@article{pmid38439188, title = {Marc van Regenmortel, personal recollections on a forward-thinking editor}, author = {Jean-Luc Pellequer and Eric Westhof}, doi = {10.1002/jmr.3080}, issn = {1099-1352}, year = {2024}, date = {2024-03-01}, urldate = {2024-03-01}, journal = {J Mol Recognit}, pages = {e3080}, abstract = {Marc van Regenmortel was the Editor-in-Chief of the Journal of Molecular Recognition for the last 25 years. Without attempting to summarize Marc's exceptional career and achievements, we would like to tell the story of the tortuous and contingent path to the unravelling of a key molecular recognition process in antigenicity. Life is indeed full of contingencies and scientific life, full of meetings and random encounters, is prone to contingencies, a key element in discovery and innovation.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{pmid38425897, title = {Peptide-Based Covalent Inhibitors Bearing Mild Electrophiles to Target a Conserved His Residue of the Bacterial Sliding Clamp}, author = {Guillaume Compain and Clément Monsarrat and Julie Blagojevic and Karl Brillet and Philippe Dumas and Philippe Hammann and Lauriane Kuhn and Isabelle Martiel and Sylvain Engilberge and Vincent Oliéric and Philippe Wolff and Dominique Y Burnouf and Jérôme Wagner and Gilles Guichard}, doi = {10.1021/jacsau.3c00572}, issn = {2691-3704}, year = {2024}, date = {2024-02-01}, urldate = {2024-02-01}, journal = {JACS Au}, volume = {4}, number = {2}, pages = {432--440}, abstract = {Peptide-based covalent inhibitors targeted to nucleophilic protein residues have recently emerged as new modalities to target protein-protein interactions (PPIs) as they may provide some benefits over more classic competitive inhibitors. Covalent inhibitors are generally targeted to cysteine, the most intrinsically reactive amino acid residue, and to lysine, which is more abundant at the surface of proteins but much less frequently to histidine. Herein, we report the structure-guided design of targeted covalent inhibitors (TCIs) able to bind covalently and selectively to the bacterial sliding clamp (SC), by reacting with a well-conserved histidine residue located on the edge of the peptide-binding pocket. SC is an essential component of the bacterial DNA replication machinery, identified as a promising target for the development of new antibacterial compounds. Thermodynamic and kinetic analyses of ligands bearing different mild electrophilic warheads confirmed the higher efficiency of the chloroacetamide compared to Michael acceptors. Two high-resolution X-ray structures of covalent inhibitor-SC adducts were obtained, revealing the canonical orientation of the ligand and details of covalent bond formation with histidine. Proteomic studies were consistent with a selective SC engagement by the chloroacetamide-based TCI. Finally, the TCI of SC was substantially more active than the parent noncovalent inhibitor in an in vitro SC-dependent DNA synthesis assay, validating the potential of the approach to design covalent inhibitors of protein-protein interactions targeted to histidine.}, keywords = {ARN-MS, ENNIFAR, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38307811, title = {Pathophysiology of human mitochondrial tRNA metabolism}, author = {Jian-Hui Zhang and Gilbert Eriani and Xiao-Long Zhou}, doi = {10.1016/j.tem.2024.01.002}, issn = {1879-3061}, year = {2024}, date = {2024-02-01}, urldate = {2024-02-01}, journal = {Trends Endocrinol Metab}, abstract = {Mitochondria play multiple critical roles in cellular activity. In particular, mitochondrial translation is pivotal in the regulation of mitochondrial and cellular homeostasis. In this forum article, we discuss human mitochondrial tRNA metabolism and highlight its tight connection with various mitochondrial diseases caused by mutations in aminoacyl-tRNA synthetases, tRNAs, and tRNA-modifying enzymes.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38408720, title = {Characterization of SLA RNA promoter from dengue virus and its interaction with the viral non-structural NS5 protein}, author = {Karl Brillet and Marta Janczuk-Richter and Amanda Poon and Joanne Laukart-Bradley and Eric Ennifar and Isabelle Lebars}, doi = {10.1016/j.biochi.2024.02.005}, issn = {1638-6183}, year = {2024}, date = {2024-02-01}, urldate = {2024-02-01}, journal = {Biochimie}, volume = {222}, pages = {87--100}, abstract = {The Dengue virus (DENV) is the most significant arthropod-borne viral pathogen in humans with 400 million infections annually. DENV comprises four distinct serotypes (DENV-1 to -4) which complicates vaccine development. Any of the four serotypes can cause clinical illness but with distinctive infection dynamics. Variations in sequences identified within the four genomes induce structural differences in crucial RNA motifs that were suggested to be correlated to the degree of pathogenicity among DENV-1 to -4. In particular, the RNA Stem-loop A (SLA) at the 5'-end of the genome, acts as a key regulator of the viral replication cycle by interacting with the viral NS5 polymerase to initiate the minus-strand viral RNA synthesis and later to methylate and cap the synthesized RNA. The molecular details of this interaction remain not fully described. Here, we report the solution secondary structures of SLA from DENV-1 to -4. Our results highlight that the four SLA exhibit structural and dynamic differences. Secondly, to determine whether SLA RNA contains serotype-specific determinants for the recognition by the viral NS5 protein, we investigated interactions between SLA from DENV -1 to -4 and DENV2 NS5 using combined biophysical approaches. Our results show that NS5 from DENV2 is able to bind SLA from other serotypes, but that other viral or host factors may be necessary to stabilize the complex and promote the catalytically active state of the NS5. By contrast, we show that a serotype-specific binding is driven by specific interactions involving conformational changes within the SLA RNA.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38070350, title = {Highly efficient and sensitive membrane-based concentration process allows quantification, surveillance, and sequencing of viruses in large volumes of wastewater}, author = {G El Soufi and L Di Jorio and Z Gerber and N Cluzel and J Van Assche and D Delafoy and R Olaso and C Daviaud and T Loustau and C Schwartz and D Trebouet and O Hernalsteens and V Marechal and S Raffestin and D Rousset and C Van Lint and J F Deleuze and M Boni and and O Rohr and M Villain-Gambier and C Wallet}, doi = {10.1016/j.watres.2023.120959}, issn = {1879-2448}, year = {2024}, date = {2024-02-01}, urldate = {2024-02-01}, journal = {Water Res}, volume = {249}, pages = {120959}, abstract = {Wastewater-based epidemiology is experiencing exponential development. Despite undeniable advantages compared to patient-centered approaches (cost, anonymity, survey of large populations without bias, detection of asymptomatic infected peoples…), major technical limitations persist. Among them is the low sensitivity of the current methods used for quantifying and sequencing viral genomes from wastewater. In situations of low viral circulation, during initial stages of viral emergences, or in areas experiencing heavy rains, the extremely low concentrations of viruses in wastewater may fall below the limit of detection of the current methods. The availability during crisis and the cost of the commercial kits, as well as the requirement of expensive materials such as high-speed centrifuge, can also present major blocks to the development of wastewater-based epidemiological survey, specifically in low-income countries. Thereby, highly sensitive, low cost and standardized methods are still needed, to increase the predictability of the viral emergences, to survey low-circulating viruses and to make the results from different labs comparable. Here, we outline and characterize new protocols for concentrating and quantifying SARS-CoV-2 from large volumes (500 mL-1 L) of untreated wastewater. In addition, we report that the methods are applicable for monitoring and sequencing. Our nucleic acid extraction technique (the routine C: 5 mL method) does not require sophisticated equipment such as automatons and is not reliant on commercial kits, making it readily available to a broader range of laboratories for routine epidemiological survey. Furthermore, we demonstrate the efficiency, the repeatability, and the high sensitivity of a new membrane-based concentration method (MBC: 500 mL method) for enveloped (SARS-CoV-2) and non-enveloped (F-specific RNA phages of genogroup II / FRNAPH GGII) viruses. We show that the MBC method allows the quantification and the monitoring of viruses in wastewater with a significantly improved sensitivity compared to the routine C method. In contexts of low viral circulation, we report quantifications of SARS-CoV-2 in wastewater at concentrations as low as 40 genome copies per liter. In highly diluted samples collected in wastewater treatment plants of French Guiana, we confirmed the accuracy of the MBC method compared to the estimations done with the routine C method. Finally, we demonstrate that both the routine C method processing 5 mL and the MBC method processing 500 mL of untreated wastewater are both compatible with SARS-CoV-2 sequencing. We show that the quality of the sequence is correlated with the concentration of the extracted viral genome. Of note, the quality of the sequences obtained with some MBC processed wastewater was improved by dilutions or enzyme substitutions suggesting the presence of specific enzyme inhibitors in some wastewater. To the best of our knowledge, our MBC method is one of the first efficient, sensitive, and repeatable method characterized for SARS-CoV-2 quantification and sequencing from large volumes of wastewater.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{CF2024, title = {Neofunctionalization driven by positive selection led to the retention of the loqs2 gene encoding an Aedes specific dsRNA binding protein}, author = {Estevez-Castro CF and Rodrigues MF and Babarit A and Ferreira FV and de Andrade EG and Marois E and Cogni R and Aguiar ERGR and Marques JT and Olmo RP}, url = {https://doi.org/10.1186/s12915-024-01821-4}, doi = {10.1186/s12915-024-01821-4}, year = {2024}, date = {2024-01-25}, urldate = {2024-01-25}, journal = {BMC Biol }, volume = {22}, number = {14}, abstract = {Background Mosquito borne viruses, such as dengue, Zika, yellow fever and Chikungunya, cause millions of infections every year. These viruses are mostly transmitted by two urban-adapted mosquito species, Aedes aegypti and Aedes albopictus. Although mechanistic understanding remains largely unknown, Aedes mosquitoes may have unique adaptations that lower the impact of viral infection. Recently, we reported the identification of an Aedes specific double-stranded RNA binding protein (dsRBP), named Loqs2, that is involved in the control of infection by dengue and Zika viruses in mosquitoes. Preliminary analyses suggested that the loqs2 gene is a paralog of loquacious (loqs) and r2d2, two co-factors of the RNA interference (RNAi) pathway, a major antiviral mechanism in insects. Results Here we analyzed the origin and evolution of loqs2. Our data suggest that loqs2 originated from two independent duplications of the first double-stranded RNA binding domain of loqs that occurred before the origin of the Aedes Stegomyia subgenus, around 31 million years ago. We show that the loqs2 gene is evolving under relaxed purifying selection at a faster pace than loqs, with evidence of neofunctionalization driven by positive selection. Accordingly, we observed that Loqs2 is localized mainly in the nucleus, different from R2D2 and both isoforms of Loqs that are cytoplasmic. In contrast to r2d2 and loqs, loqs2 expression is stage- and tissue-specific, restricted mostly to reproductive tissues in adult Ae. aegypti and Ae. albopictus. Transgenic mosquitoes engineered to express loqs2 ubiquitously undergo developmental arrest at larval stages that correlates with massive dysregulation of gene expression without major effects on microRNAs or other endogenous small RNAs, classically associated with RNA interference. Conclusions Our results uncover the peculiar origin and neofunctionalization of loqs2 driven by positive selection. This study shows an example of unique adaptations in Aedes mosquitoes that could ultimately help explain their effectiveness as virus vectors.}, keywords = {Aedes mosquitoes, Double-stranded RNA (dsRNA), dsRNA binding protein (dsRBP), loqs2, M3i, marois, Marques, Olmo, RNA interference (RNAi)}, pubstate = {published}, tppubtype = {article} } @article{Green2024, title = {A population modification gene drive targeting both Saglin and Lipophorin disables Plasmodium transmission in Anopheles mosquitoes}, author = {Emily I Green and Etienne Jaouen and Dennis Klug and Roenick Proveti Olmo and Amandine Gautier and Stéphanie Blandin and Eric Marois}, url = {https://elifesciences.org/articles/93142}, doi = {10.7554/eLife.93142}, year = {2024}, date = {2024-01-12}, journal = {Genetics and Genomics}, abstract = {Lipophorin is an essential, highly expressed lipid transport protein that is secreted and circulates in insect hemolymph. We hijacked the Anopheles coluzzii Lipophorin gene to make it co-express a single-chain version of antibody 2A10, which binds sporozoites of the malaria parasite Plasmodium falciparum. The resulting transgenic mosquitoes show a markedly decreased ability to transmit Plasmodium berghei expressing the P. falciparum circumsporozoite protein to mice. To force the spread of this antimalarial transgene in a mosquito population, we designed and tested several CRISPR/Cas9-based gene drives. One of these is installed in, and disrupts, the pro-parasitic gene Saglin and also cleaves wild-type Lipophorin, causing the anti-malarial modified Lipophorin version to replace the wild type and hitch-hike together with the Saglin drive. Although generating drive-resistant alleles and showing instability in its gRNA-encoding multiplex array, the Saglin-based gene drive reached high levels in caged mosquito populations and efficiently promoted the simultaneous spread of the antimalarial Lipophorin::Sc2A10 allele. This combination is expected to decrease parasite transmission via two different mechanisms. This work contributes to the design of novel strategies to spread antimalarial transgenes in mosquitoes, and illustrates some expected and unexpected outcomes encountered when establishing a population modification gene drive.}, keywords = {Anopheles, blandin, gene drive, Lipophorin, M3i, marois, marque, Marques, Olmo, Plasmodium, Saglin}, pubstate = {published}, tppubtype = {article} } @online{Marois2024, title = {Using the CRISPR / Cas9 system for genome editing in Anopheles mosquitoes}, author = { Eric Marois}, url = {https://hal.science/hal-04380430/document}, doi = {HAL Id: hal-04380430}, year = {2024}, date = {2024-01-08}, urldate = {2024-01-08}, journal = {HAL science}, abstract = {The advent of the CRISPR / Cas9 technology permits the targeted editing of mosquito genomes, ranging from site-directed mutagenesis of genes of interest yielding knockout mutations (which arise by insertion / deletion of a few nucleotides) to site-specific insertion of exogenous DNA sequences such as fluorescence markers or even large gene drive cassettes, themselves encoding the components of the CRISPR / Cas9 system. To obtain these heritable targeted changes, genome editing requires the delivery of Cas9 protein and its guide RNA(s) to the developing germ tissue of an embryo. Different species require adaptation of this basic principle to accommodate for their specific biology. Here, we describe a technical pipeline based on delivering the CRISPR/Cas9 components in the form of injected plasmid or as transgenes, resulting in highly efficient gene editing in Anopheles malaria vector mosquitoes. We have reliably employed these methods to mutagenize > 20 different loci of interest in Anopheles coluzzii to date. }, keywords = {Anopheles, CRISPR/Cas9, Genome editing, M3i, marois, mosquitoes}, pubstate = {published}, tppubtype = {online} } @article{pmid38507196, title = {Immunoprecipitation Methods to Isolate Messenger Ribonucleoprotein Complexes (mRNP)}, author = {Hassan Hayek and Lauriane Gross and Fatima Alghoul and Franck Martin and Gilbert Eriani and Christine Allmang}, doi = {10.1007/978-3-031-52193-5_1}, issn = {0065-2598}, year = {2024}, date = {2024-01-01}, urldate = {2024-01-01}, journal = {Adv Exp Med Biol}, volume = {3234}, pages = {1--15}, abstract = {Throughout their life cycle, messenger RNAs (mRNAs) associate with proteins to form ribonucleoproteins (mRNPs). Each mRNA is part of multiple successive mRNP complexes that participate in their biogenesis, cellular localization, translation and decay. The dynamic composition of mRNP complexes and their structural remodelling play crucial roles in the control of gene expression. Studying the endogenous composition of different mRNP complexes is a major challenge. In this chapter, we describe the variety of protein-centric immunoprecipitation methods available for the identification of mRNP complexes and the requirements for their experimental settings.}, keywords = {ERIANI, MARTIN, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38507197, title = {Purification of In Vivo or In Vitro-Assembled RNA-Protein Complexes by RNA Centric Methods}, author = {Aurélie Janvier and Hassan Hayek and Fatima Alghoul and Lauriane Gross and Christine Allmang and Franck Martin and Gilbert Eriani}, doi = {10.1007/978-3-031-52193-5_2}, issn = {0065-2598}, year = {2024}, date = {2024-01-01}, urldate = {2024-01-01}, journal = {Adv Exp Med Biol}, volume = {3234}, pages = {17--29}, abstract = {Throughout their entire life cycle, RNAs are associated with RNA-binding proteins (RBPs), forming ribonucleoprotein (RNP) complexes with highly dynamic compositions and very diverse functions in RNA metabolism, including splicing, translational regulation, ribosome assembly. Many RNPs remain poorly characterized due to the challenges inherent in their purification and subsequent biochemical characterization. Therefore, developing methods to isolate specific RNA-protein complexes is an important initial step toward understanding their function. Many elegant methodologies have been developed to isolate RNPs. This chapter describes different approaches and methods devised for RNA-specific purification of a target RNP. We focused on general methods for selecting RNPs that target a given RNA under conditions favourable for the copurification of associated factors including RNAs and protein components of the RNP.}, keywords = {ERIANI, MARTIN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38217659, title = {In-Gel Cyanoethylation for Pseudouridines Mass Spectrometry Detection of Bacterial Regulatory RNA}, author = {Antony Lechner and Philippe Wolff}, doi = {10.1007/978-1-0716-3565-0_15}, issn = {1940-6029}, year = {2024}, date = {2024-01-01}, urldate = {2024-01-01}, journal = {Methods Mol Biol}, volume = {2741}, pages = {273--287}, abstract = {Regulatory RNAs, as well as many RNA families, contain chemically modified nucleotides, including pseudouridines (ψ). To map nucleotide modifications, approaches based on enzymatic digestion of RNA followed by nano liquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) analysis were implemented several years ago. However, detection of ψ by mass spectrometry (MS) is challenging as ψ exhibits the same mass as uridine. Thus, a chemical labeling strategy using acrylonitrile was developed to detect this mass-silent modification. Acrylonitrile reacts specifically to ψ to form 1-cyanoethylpseudouridine (Ceψ), resulting in a mass shift of ψ detectable by MS. Here, a protocol detailing the steps from the purification of RNA by polyacrylamide gel electrophoresis, including in-gel labeling of ψ, to MS data interpretation to map ψ and other modifications is proposed. To demonstrate its efficiency, the protocol was applied to bacterial regulatory RNAs from E. coli: 6S RNA and transfer-messenger RNA (tmRNA, also known as 10Sa RNA). Moreover, ribonuclease P (RNase P) was also mapped using this approach. This method enabled the detection of several ψ at single nucleotide resolution.}, keywords = {ARN-MS, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38217649, title = {Ribosome Profiling Methods Adapted to the Study of RNA-Dependent Translation Regulation in Staphylococcus aureus}, author = {Maximilian P Kohl and Béatrice Chane-Woon-Ming and Roberto Bahena-Ceron and Jose Jaramillo-Ponce and Laura Antoine and Lucas Herrgott and Pascale Romby and Stefano Marzi}, doi = {10.1007/978-1-0716-3565-0_5}, issn = {1940-6029}, year = {2024}, date = {2024-01-01}, urldate = {2024-01-01}, journal = {Methods Mol Biol}, volume = {2741}, pages = {73--100}, abstract = {Noncoding RNAs, including regulatory RNAs (sRNAs), are instrumental in regulating gene expression in pathogenic bacteria, allowing them to adapt to various stresses encountered in their host environments. Staphylococcus aureus is a well-studied model for RNA-mediated regulation of virulence and pathogenicity, with sRNAs playing significant roles in shaping S. aureus interactions with human and animal hosts. By modulating the translation and/or stability of target mRNAs, sRNAs regulate the synthesis of virulence factors and regulatory proteins required for pathogenesis. Moreover, perturbation of the levels of RNA modifications in two other classes of noncoding RNAs, rRNAs, and tRNAs, has been proposed to contribute to stress adaptation. However, the study of how these various factors affect translation regulation has often been restricted to specific genes, using in vivo reporters and/or in vitro translation systems. Genome-wide sequencing approaches offer novel perspectives for studying RNA-dependent regulation. In particular, ribosome profiling methods provide a powerful resource for characterizing the overall landscape of translational regulation, contributing to a better understanding of S. aureus physiopathology. Here, we describe protocols that we have adapted to perform ribosome profiling in S. aureus.}, keywords = {MARZI, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38287188, title = {The helicase domain of human Dicer prevents RNAi-independent activation of antiviral and inflammatory pathways}, author = {Morgane Baldaccini and Léa Gaucherand and Béatrice Chane-Woon-Ming and Mélanie Messmer and Floriane Gucciardi and Sébastien Pfeffer}, doi = {10.1038/s44318-024-00035-2}, issn = {1460-2075}, year = {2024}, date = {2024-01-01}, urldate = {2024-01-01}, journal = {EMBO J}, abstract = {In mammalian somatic cells, the relative contribution of RNAi and the type I interferon response during viral infection is unclear. The apparent inefficiency of antiviral RNAi might be due to self-limiting properties and mitigating co-factors of the key enzyme Dicer. In particular, the helicase domain of human Dicer appears to be an important restriction factor of its activity. Here, we study the involvement of several helicase-truncated mutants of human Dicer in the antiviral response. All deletion mutants display a PKR-dependent antiviral phenotype against certain viruses, and one of them, Dicer N1, acts in a completely RNAi-independent manner. Transcriptomic analyses show that many genes from the interferon and inflammatory response pathways are upregulated in Dicer N1 expressing cells. We show that some of these genes are controlled by NF-kB and that blocking this pathway abrogates the antiviral phenotype of Dicer N1. Our findings highlight the crosstalk between Dicer, PKR, and the NF-kB pathway, and suggest that human Dicer may have repurposed its helicase domain to prevent basal activation of antiviral and inflammatory pathways.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38286022, title = {Exchanging and Releasing Information in Synthetic Digital Polymers Using a Strand-Displacement Strategy}, author = {Maria Nerantzaki and Claire Husser and Michael Ryckelynck and Jean-François Lutz}, doi = {10.1021/jacs.3c13953}, issn = {1520-5126}, year = {2024}, date = {2024-01-01}, urldate = {2024-01-01}, journal = {J Am Chem Soc}, abstract = {Toehold-mediated strand displacement (TMSD) was tested as a tool to edit information in synthetic digital polymers. Uniform DNA-polymer biohybrid macromolecules were first synthesized by automated phosphoramidite chemistry and characterized by HPLC, mass spectrometry, and polyacrylamide gel electrophoresis (PAGE). These precursors were diblock structures containing a synthetic poly(phosphodiester) (PPDE) segment covalently attached to a single-stranded DNA sequence. Three types of biohybrids were prepared herein: a substrate containing an accessible toehold as well as input and output macromolecules. The substrate and the input macromolecules contained noncoded PPDE homopolymers, whereas the output macromolecule contained a digitally encoded segment. After hybridization of the substrate with the output, incubation in the presence of the input led to efficient TMSD and the release of the digital segment. TMSD can therefore be used to erase or rewrite information in self-assembled biohybrid superstructures. Furthermore, it was found in this work that the conjugation of DNA single strands to synthetic segments of chosen lengths greatly facilitates the characterization and PAGE visualization of the TMSD process.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Structural Impact of the Interaction of the Influenza A Virus Nucleoprotein with Genomic RNA Segments}, author = {E. Quignon and D. Ferhadian and A. Hache and V. Vivet-Boudou and C. Isel and A. Printz-Schweigert and A. Donchet and T. Crépin and R. Marquet}, url = {https://www.mdpi.com/1999-4915/16/3/421}, isbn = {10.3390/v16030421}, year = {2024}, date = {2024-01-01}, urldate = {2024-01-01}, journal = {Viruses}, volume = {16}, number = {3}, pages = {421}, abstract = {Influenza A viruses (IAVs) possess a segmented genome consisting of eight viral RNAs (vRNAs) associated with multiple copies of viral nucleoprotein (NP) and a viral polymerase complex. Despite the crucial role of RNA structure in IAV replication, the impact of NP binding on vRNA structure is not well understood. In this study, we employed SHAPE chemical probing to compare the structure of NS and M vRNAs of WSN IAV in various states: before the addition of NP, in complex with NP, and after the removal of NP. Comparison of the RNA structures before the addition of NP and after its removal reveals that NP, while introducing limited changes, remodels local structures in both vRNAs and long-range interactions in the NS vRNA, suggesting a potentially biologically relevant RNA chaperone activity. In contrast, NP significantly alters the structure of vRNAs in vRNA/NP complexes, though incorporating experimental data into RNA secondary structure prediction proved challenging. Finally, our results suggest that NP not only binds single-stranded RNA but also helices with interruptions, such as bulges or small internal loops, with a preference for G-poor and C/U-rich regions.}, keywords = {MARQUET, MARQUET influenza A virus NP nucleoprotein vRNA RNA structure RNA chaperon chemical probing, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37876025, title = {Ascorbate deficiency increases progression of shigellosis in guinea pigs and mice infection models}, author = {Jurate Skerniskyte and Céline Mulet and Antonin C André and Mark C Anderson and Louise Injarabian and Achim Buck and Verena M Prade and Philippe J Sansonetti and Sophie Reibel-Foisset and Axel K Walch and Michel Lebel and Jens Lykkesfeldt and Benoit S Marteyn}, doi = {10.1080/19490976.2023.2271597}, issn = {1949-0984}, year = {2023}, date = {2023-12-01}, urldate = {2023-12-01}, journal = {Gut Microbes}, volume = {15}, number = {2}, pages = {2271597}, abstract = { spp. are the causative agents of bacterial dysentery and shigellosis, mainly in children living in developing countries. The study of entire life cycle and the evaluation of vaccine candidates' protective efficacy have been hampered by the lack of a suitable animal model of infection. None of the studies evaluated so far (rabbit, guinea pig, mouse) allowed the recapitulation of full shigellosis symptoms upon oral challenge. Historical reports have suggested that dysentery and scurvy are both metabolic diseases associated with ascorbate deficiency. Mammals, which are susceptible to infection (humans, non-human primates and guinea pigs) are among the few species unable to synthesize ascorbate. We optimized a low-ascorbate diet to induce moderate ascorbate deficiency, but not scurvy, in guinea pigs to investigate whether poor vitamin C status increases the progression of shigellosis. Moderate ascorbate deficiency increased shigellosis symptom severity during an extended period of time (up to 48 h) in all strains tested (, 5a, and 2a). At late time points, an important influx of neutrophils was observed both within the disrupted colonic mucosa and in the luminal compartment, although was able to disseminate deep into the organ to reach the sub-mucosal layer and the bloodstream. Moreover, we found that ascorbate deficiency also increased penetration into the colon epithelium layer in a Gulo mouse infection model. The use of these new rodent models of shigellosis opens new doors for the study of both infection strategies and immune responses to infection.}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38050960, title = {Contribution of tRNA sequence and modifications to the decoding preferences of E. coli and M. mycoides tRNAGlyUCC for synonymous glycine codons}, author = {Maria Kompatscher and Karolina Bartosik and Kevin Erharter and Raphael Plangger and Fabian Sebastian Juen and Christoph Kreutz and Ronald Micura and Eric Westhof and Matthias D Erlacher}, doi = {10.1093/nar/gkad1136}, issn = {1362-4962}, year = {2023}, date = {2023-12-01}, urldate = {2023-12-01}, journal = {Nucleic Acids Res}, abstract = {tRNA superwobbling, used by certain bacteria and organelles, is an intriguing decoding concept in which a single tRNA isoacceptor is used to decode all synonymous codons of a four-fold degenerate codon box. While Escherichia coli relies on three tRNAGly isoacceptors to decode the four glycine codons (GGN), Mycoplasma mycoides requires only a single tRNAGly. Both organisms express tRNAGly with the anticodon UCC, which are remarkably similar in sequence but different in their decoding ability. By systematically introducing mutations and altering the number and type of tRNA modifications using chemically synthesized tRNAs, we elucidated the contribution of individual nucleotides and chemical groups to decoding by the E. coli and M. mycoides tRNAGly. The tRNA sequence was identified as the key factor for superwobbling, revealing the T-arm sequence as a novel pivotal element. In addition, the presence of tRNA modifications, although not essential for providing superwobbling, was shown to delicately fine-tune and balance the decoding of synonymous codons. This emphasizes that the tRNA sequence and its modifications together form an intricate system of high complexity that is indispensable for accurate and efficient decoding.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{pmid38164596, title = {RlmQ: A Newly Discovered rRNA Modification Enzyme Bridging RNA Modification and Virulence Traits in }, author = {Roberto Bahena-Ceron and Chloe Teixeira and Jose R Jaramillo Ponce and Philippe Wolff and Florence Couzon and Pauline François and Bruno Klaholz and François Vandenesch and Pascale Romby and Karen Moreau and Stefano Marzi}, doi = {10.1261/rna.079850.123}, issn = {1469-9001}, year = {2023}, date = {2023-12-01}, urldate = {2023-12-01}, journal = {RNA}, volume = {30}, number = {3}, pages = {200-212}, abstract = {rRNA modifications play crucial roles in fine-tuning the delicate balance between translation speed and accuracy, yet the underlying mechanisms remain elusive. Comparative analyses of the ribosomal RNA modifications in taxonomically distant bacteria could help define their general, as well as species-specific, roles. In this study, we identified a new methyltransferase, RlmQ, in responsible for the Gram-positive specific mG2601, which is not modified in (G2574). We also demonstrate the absence of methylation on C1989, equivalent to C1962, which is methylated at position 5 by the Gram-negative specific RlmI methyltransferase, a paralogue of RlmQ. Both modifications ( mG2601 and mC1962) are situated within the same tRNA accommodation corridor, hinting at a potential shared function in translation. Inactivation of Q causes the loss of methylation at G2601 and significantly impacts growth, cytotoxicity, and biofilm formation. These findings unravel the intricate connections between rRNA modifications, translation, and virulence in pathogenic Gram-positive bacteria.}, keywords = {ARN-MS, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid38254646, title = {Plasmodium, the Apicomplexa Outlier When It Comes to Protein Synthesis}, author = {José R Jaramillo Ponce and Magali Frugier}, doi = {10.3390/biom14010046}, issn = {2218-273X}, year = {2023}, date = {2023-12-01}, urldate = {2023-12-01}, journal = {Biomolecules}, volume = {14}, number = {1}, pages = {46}, abstract = {Plasmodium is an obligate intracellular parasite that has numerous interactions with different hosts during its elaborate life cycle. This is also the case for the other parasites belonging to the same phylum Apicomplexa. In this study, we bioinformatically identified the components of the multi-synthetase complexes (MSCs) of several Apicomplexa parasites and modelled their assembly using AlphaFold2. It appears that none of these MSCs resemble the two MSCs that we have identified and characterized in Plasmodium. Indeed, tRip, the central protein involved in the association of the two Plasmodium MSCs is different from its homologues, suggesting also that the tRip-dependent import of exogenous tRNAs is not conserved in other apicomplexan parasites. Based on this observation, we searched for obvious differences that could explain the singularity of Plasmodium protein synthesis by comparing tRNA genes and amino acid usage in the different genomes. We noted a contradiction between the large number of asparagine residues used in Plasmodium proteomes and the single gene encoding the tRNA that inserts them into proteins. This observation remains true for all the Plasmodia strains studied, even those that do not contain long asparagine homorepeats. }, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Hinze2023, title = {Knockout of OR39 reveals redundancy in the olfactory pathway regulating the acquisition of host seeking in Anopheles coluzzii}, author = {Hinze A and Pelletier J and Ghaninia M and Marois E and Hill and S Ignell R}, url = {https://doi.org/10.1098/rspb.2023.2092}, doi = {10.1098/rspb.2023.2092}, year = {2023}, date = {2023-11-29}, urldate = {2023-11-29}, booktitle = {Proc Biol Sci}, journal = {Proc Biol Sci 290(2011):20232092}, volume = {290}, number = {2011}, abstract = {The attraction of anthropophilic mosquitoes to human host cues, such as body odour and carbon dioxide, gradually increases during adult maturation. This acquisition of host-seeking behaviour correlates with age-dependent changes in odorant receptor (OR) transcript abundance and sensitivity of olfactory sensory neurons (OSNs). One OR gene of the human malaria vector, Anopheles coluzzii, AcolOR39, is significantly downregulated in mature females, and a cognate ligand of AcolOR39, sulcatone, a major component of human emanations, mediates the observed behavioural inhibition of newly emerged (teneral) females to human body odour. Knockout of AcolOR39, using CRISPR–Cas9 mutagenesis, selectively abolished sulcatone detection in OSNs, housed in trichoid sensilla. However, knockout of AcolOR39 altered neither the response rate nor the flight behaviour of teneral females in a wind tunnel, indicating the involvement of other genes, and thus a redundancy, in regulating the acquisition of host seeking in mosquitoes.}, keywords = {CRISPR–Cas9, human odour, M3i, marois, mosquito flight, odour valence, SSR}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Efficient Sex Separation by Exploiting Differential Alternative Splicing of a Dominant Marker in Aedes aegypti}, author = {Weng SC and Antoshechkin I and Marois E and Akbari O}, url = {https://doi.org/10.1371/journal.pgen.1011065}, doi = {10.1371/journal.pgen.1011065}, year = {2023}, date = {2023-11-27}, urldate = {2023-11-27}, journal = {PLoS Genetics}, abstract = {Only female mosquitoes consume blood giving them the opportunity to transmit deadly human pathogens. Therefore, it is critical to remove females before conducting releases for genetic biocontrol interventions. Here we describe a robust sex-sorting approach termed SEPARATOR (Sexing Element Produced by Alternative RNA-splicing of A Transgenic Observable Reporter) that exploits sex-specific alternative splicing of an innocuous reporter to ensure exclusive dominant male-specific expression. Using SEPARATOR, we demonstrate reliable sex selection from early larval and pupal stages in Aedes aegypti, and use a Complex Object Parametric Analyzer and Sorter (COPAS) to demonstrate scalable high-throughput sex-selection of first instar larvae. Additionally, we use this approach to sequence the transcriptomes of early larval males and females and find several genes that are sex-specifically expressed. SEPARATOR can simplify mass production of males for release programs and is designed to be cross-species portable and should be instrumental for genetic biocontrol interventions.}, keywords = {Aedes aegypti, Introns, larvae, M3i, marois, mosquitoes, pupae, rna sequencing, transcriptome analyses}, pubstate = {published}, tppubtype = {article} } @article{pmid37941143, title = {SilkMeta: a comprehensive platform for sharing and exploiting pan-genomic and multi-omic silkworm data}, author = {Kunpeng Lu and Yifei Pan and Jianghong Shen and Lin Yang and Chengyu Zhan and Shubo Liang and Shuaishuai Tai and Linrong Wan and Tian Li and Tingcai Cheng and Bi Ma and Guoqing Pan and Ningjia He and Cheng Lu and Eric Westhof and Zhonghuai Xiang and Min-Jin Han and Xiaoling Tong and Fangyin Dai}, doi = {10.1093/nar/gkad956}, issn = {1362-4962}, year = {2023}, date = {2023-11-01}, urldate = {2023-11-01}, journal = {Nucleic Acids Res}, volume = {52}, issue = {D1}, pages = {D1024-D1032}, abstract = {The silkworm Bombyx mori is a domesticated insect that serves as an animal model for research and agriculture. The silkworm super-pan-genome dataset, which we published last year, is a unique resource for the study of global genomic diversity and phenotype-genotype association. Here we present SilkMeta (http://silkmeta.org.cn), a comprehensive database covering the available silkworm pan-genome and multi-omics data. The database contains 1082 short-read genomes, 546 long-read assembled genomes, 1168 transcriptomes, 294 phenotype characterizations (phenome), tens of millions of variations (variome), 7253 long non-coding RNAs (lncRNAs), 18 717 full length transcripts and a set of population statistics. We have compiled publications on functional genomics research and genetic stock deciphering (mutant map). A range of bioinformatics tools is also provided for data visualization and retrieval. The large batch of omics data and tools were integrated in twelve functional modules that provide useful strategies and data for comparative and functional genomics research. The interactive bioinformatics platform SilkMeta will benefit not only the silkworm but also the insect biology communities.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{pmid37945696, title = {A tRNA-modifying enzyme facilitates RNase P activity in Arabidopsis nuclei}, author = {Mathilde Arrivé and Mathieu Bruggeman and Vasileios Skaltsogiannis and Léna Coudray and Yi-Fat Quan and Cédric Schelcher and Valérie Cognat and Philippe Hammann and Johana Chicher and Philippe Wolff and Anthony Gobert and Philippe Giegé}, doi = {10.1038/s41477-023-01564-0}, issn = {2055-0278}, year = {2023}, date = {2023-11-01}, urldate = {2023-11-01}, journal = {Nat Plants}, volume = {9}, issue = {12}, pages = {2031-2041}, abstract = {RNase P is the essential activity that performs the 5' maturation of transfer RNA (tRNA) precursors. Beyond the ancestral form of RNase P containing a ribozyme, protein-only RNase P enzymes termed PRORP were identified in eukaryotes. In human mitochondria, PRORP forms a complex with two protein partners to become functional. In plants, although PRORP enzymes are active alone, we investigate their interaction network to identify potential tRNA maturation complexes. Here we investigate functional interactions involving the Arabidopsis nuclear RNase P PRORP2. We show, using an immuno-affinity strategy, that PRORP2 occurs in a complex with the tRNA methyl transferases TRM1A and TRM1B in vivo. Beyond RNase P, these enzymes can also interact with RNase Z. We show that TRM1A/TRM1B localize in the nucleus and find that their double knockout mutation results in a severe macroscopic phenotype. Using a combination of immuno-detections, mass spectrometry and a transcriptome-wide tRNA sequencing approach, we observe that TRM1A/TRM1B are responsible for the mG26 modification of 70% of cytosolic tRNAs in vivo. We use the transcriptome wide tRNAseq approach as well as RNA blot hybridizations to show that RNase P activity is impaired in TRM1A/TRM1B mutants for specific tRNAs, in particular, tRNAs containing a mG modification at position 26 that are strongly downregulated in TRM1A/TRM1B mutants. Altogether, results indicate that the mG-adding enzymes TRM1A/TRM1B functionally cooperate with nuclear RNase P in vivo for the early steps of cytosolic tRNA biogenesis.}, keywords = {ARN-MS, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37806286, title = {Escherichia coli killing by epidemiologically successful sublineages of Shigella sonnei is mediated by colicins}, author = {P Malaka De Silva and Rebecca J Bennett and Lauriane Kuhn and Patryk Ngondo and Lorine Debande and Elisabeth Njamkepo and Brian Ho and François-Xavier Weill and Benoît S Marteyn and Claire Jenkins and Kate S Baker}, doi = {10.1016/j.ebiom.2023.104822}, issn = {2352-3964}, year = {2023}, date = {2023-10-01}, urldate = {2023-10-01}, journal = {EBioMedicine}, volume = {97}, pages = {104822}, abstract = {BACKGROUND: Shigella sp. are enteric pathogens which causes >125 million cases of shigellosis annually. S. sonnei accounts for about a quarter of those cases and is increasingly prevalent in industrialising nations. Being an enteric pathogen, S. sonnei benefits from outcompeting gut commensals such as Escherichia coli to establish itself and cause disease. There are numerous mechanisms that bacterial pathogens use to outcompete its rivals including molecules called colicins. A Type 6 Secretion System (T6SS) was recently described as contributing to E. coli killing in S. sonnei.nnMETHODS: We used Bulk Phenotyping of Epidemiological Replicates (BPER) which combined bacterial Genome Wide Association Studies (bGWAS) and high throughput phenotyping on a collection of S. sonnei surveillance isolates to identify the genetic features associated with E. coli killing and explore their relationship with epidemiological behaviour. We further explored the presence of colicins and T6SS components in the isolates using genomics, laboratory experimentation, and proteomics.nnFINDINGS: Our bGWAS analysis returned known and novel colicin and colicin related genes as significantly associated with E. coli killing. In silico analyses identified key colicin clusters responsible for the killing phenotype associated with epidemiologically successful sub-lineages. The killing phenotype was not associated with the presence of a T6SS. Laboratory analyses confirmed the presence of the key colicin clusters and that killing was contact-independent.nnINTERPRETATION: Colicins are responsible for E. coli killing by S. sonnei, not a T6SS. This phenotype contributes to shaping the observed epidemiology of S. sonnei and may contribute to its increasing prevalence globally. BPER is an epidemiologically relevant approach to phenotypic testing that enables the rapid identification of genetic drivers of phenotypic changes, and assessment of their relevance to epidemiology in natural settings.nnFUNDING: Biotechnology and Biological Sciences Research Council, Biotechnology and Biological Sciences Research Council Doctoral Training Partnership studentship, Wellcome Trust, Medical Research Council (UK), French National Research Agency.}, keywords = {MARTEYN, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37876231, title = {Assessment of three-dimensional RNA structure prediction in CASP15}, author = {Rhiju Das and Rachael C Kretsch and Adam J Simpkin and Thomas Mulvaney and Phillip Pham and Ramya Rangan and Fan Bu and Ronan M Keegan and Maya Topf and Daniel J Rigden and Zhichao Miao and Eric Westhof}, doi = {10.1002/prot.26602}, issn = {1097-0134}, year = {2023}, date = {2023-10-01}, urldate = {2023-10-01}, journal = {Proteins}, volume = {91}, issue = {12}, pages = {1747-1770}, abstract = {The prediction of RNA three-dimensional structures remains an unsolved problem. Here, we report assessments of RNA structure predictions in CASP15, the first CASP exercise that involved RNA structure modeling. Forty-two predictor groups submitted models for at least one of twelve RNA-containing targets. These models were evaluated by the RNA-Puzzles organizers and, separately, by a CASP-recruited team using metrics (GDT, lDDT) and approaches (Z-score rankings) initially developed for assessment of proteins and generalized here for RNA assessment. The two assessments independently ranked the same predictor groups as first (AIchemy_RNA2), second (Chen), and third (RNAPolis and GeneSilico, tied); predictions from deep learning approaches were significantly worse than these top ranked groups, which did not use deep learning. Further analyses based on direct comparison of predicted models to cryogenic electron microscopy (cryo-EM) maps and x-ray diffraction data support these rankings. With the exception of two RNA-protein complexes, models submitted by CASP15 groups correctly predicted the global fold of the RNA targets. Comparisons of CASP15 submissions to designed RNA nanostructures as well as molecular replacement trials highlight the potential utility of current RNA modeling approaches for RNA nanotechnology and structural biology, respectively. Nevertheless, challenges remain in modeling fine details such as noncanonical pairs, in ranking among submitted models, and in prediction of multiple structures resolved by cryo-EM or crystallography.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{AI2023, title = {The sex-specific factor SOA controls dosage compensation in Anopheles mosquitoes}, author = {Kalita AI and Marois E and Kozielska M and Weissing FJ and Jaouen E and Möckel MM and, Rühle F and Butter F and Basilicata MF and Keller Valsecchi CI}, url = {https://www.nature.com/articles/s41586-023-06641-0}, doi = {10.1038/s41586-023-06641-0.}, year = {2023}, date = {2023-09-28}, journal = {Nature}, volume = {623}, pages = {175–182}, abstract = {The Anopheles mosquito is one of thousands of species in which sex differences play a central part in their biology, as only females need a blood meal to produce eggs. Sex differentiation is regulated by sex chromosomes, but their presence creates a dosage imbalance between males (XY) and females (XX). Dosage compensation (DC) can re-equilibrate the expression of sex chromosomal genes. However, because DC mechanisms have only been fully characterized in a few model organisms, key questions about its evolutionary diversity and functional necessity remain unresolved1. Here we report the discovery of a previously uncharacterized gene (sex chromosome activation (SOA)) as a master regulator of DC in the malaria mosquito Anopheles gambiae. Sex-specific alternative splicing prevents functional SOA protein expression in females. The male isoform encodes a DNA-binding protein that binds the promoters of active X chromosomal genes. Expressing male SOA is sufficient to induce DC in female cells. Male mosquitoes lacking SOA or female mosquitoes ectopically expressing the male isoform exhibit X chromosome misregulation, which is compatible with viability but causes developmental delay. Thus, our molecular analyses of a DC master regulator in a non-model organism elucidates the evolutionary steps that lead to the establishment of a chromosome-specific fine-tuning mechanism.}, keywords = {Anopheles, M3i, marois, SOA}, pubstate = {published}, tppubtype = {article} } @article{Abbo2023, title = {The virome of the invasive Asian bush mosquito Aedes japonicus in Europe}, author = {Abbo SR and de Almeida JPP and Olmo RP and Balvers C and Griep JS and Linthout C and Koenraadt CJM and Silva BM and Fros JJ and Aguiar ERGR and Marois E and Pijlman GP and Marques JT}, doi = {10.1093/ve/vead041}, year = {2023}, date = {2023-09-22}, urldate = {2023-09-22}, journal = {Virus Evol.}, volume = {9}, issue = {2}, abstract = {The Asian bush mosquito Aedes japonicus is rapidly invading North America and Europe. Due to its potential to transmit multiple pathogenic arthropod-borne (arbo)viruses including Zika virus, West Nile virus, and chikungunya virus, it is important to understand the biology of this vector mosquito in more detail. In addition to arboviruses, mosquitoes can also carry insect-specific viruses that are receiving increasing attention due to their potential effects on host physiology and arbovirus transmission. In this study, we characterized the collection of viruses, referred to as the virome, circulating in Ae. japonicus populations in the Netherlands and France. Applying a small RNA-based metagenomic approach to Ae. japonicus, we uncovered a distinct group of viruses present in samples from both the Netherlands and France. These included one known virus, Ae. japonicus narnavirus 1 (AejapNV1), and three new virus species that we named Ae. japonicus totivirus 1 (AejapTV1), Ae. japonicus anphevirus 1 (AejapAV1) and Ae. japonicus bunyavirus 1 (AejapBV1). We also discovered sequences that were presumably derived from two additional novel viruses: Ae. japonicus bunyavirus 2 (AejapBV2) and Ae. japonicus rhabdovirus 1 (AejapRV1). All six viruses induced strong RNA interference responses, including the production of twenty-one nucleotide-sized small interfering RNAs, a signature of active replication in the host. Notably, AejapBV1 and AejapBV2 belong to different viral families; however, no RNA-dependent RNA polymerase sequence has been found for AejapBV2. Intriguingly, our small RNA-based approach identified an ∼1-kb long ambigrammatic RNA that is associated with AejapNV1 as a secondary segment but showed no similarity to any sequence in public databases. We confirmed the presence of AejapNV1 primary and secondary segments, AejapTV1, AejapAV1, and AejapBV1 by reverse transcriptase polymerase chain reaction (PCR) in wild-caught Ae. japonicus mosquitoes. AejapNV1 and AejapTV1 were found at high prevalence (87-100 per cent) in adult females, adult males, and larvae. Using a small RNA-based, sequence-independent metagenomic strategy, we uncovered a conserved and prevalent virome among Ae. japonicus mosquito populations. The high prevalence of AejapNV1 and AejapTV1 across all tested mosquito life stages suggests that these viruses are intimately associated with Ae. japonicus.}, keywords = {Aedes japonicus, anphevirus, bunyavirus, M3i, marois, Marques, metagenomics, mosquito, Olmo, rhabdovirus, RNA Interference, totivirus, virome}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Extensive uORF translation from HIV-1 transcripts elicits specific T cell immune responses in infected individuals and conditions DDX3 dependency for expression of main ORFs}, author = {LABARONNE , E. and DECIMO , D. and BERTRAND , L. and GUIGUETTAZ , L. and SOHIER , T.J.M. and CLUET , D. and VIVET-BOUDOU , V. and DAHOUI , C. and FRANçOIS , P. and HATIN , I. and LAMBOTTE , O. and SAMRI , A. and AUTRAN , B. and ETIENNE , L. and GOUJON , C. and PAILLART , J.-C. and NAMY , O. and RAMIREZ , B.C. and OHLMANN , T. and MORRIS , A. and RICCI, E.P.}, doi = {10.1101/2022.04.29.489990}, year = {2023}, date = {2023-09-20}, urldate = {2023-09-20}, journal = {(bioRxiv & medRxiv)}, keywords = {PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {The virus-induced cyclic dinucleotide 2'3'-c-di-GMP mediates STING-dependent antiviral immunity in Drosophila}, author = {Cai H and Li L and Slavik KM and Huang J and Yin T and Ai X and Hédelin L and Haas G and Xiang Z and Yang Y and Li X and Chen Y and Wei Z and Deng H and Chen D and Jiao R and Martins N and Meignin C and Kranzusch PJ and Imler JL}, editor = {Elsevier Inc. }, url = {https://pubmed.ncbi.nlm.nih.gov/37659413/}, doi = {10.1016/j.immuni.2023.08.006 }, year = {2023}, date = {2023-09-12}, urldate = {2023-09-12}, journal = {Immunity}, volume = {56}, issue = {9}, pages = {1991-2005}, abstract = {In mammals, the enzyme cGAS senses the presence of cytosolic DNA and synthesizes the cyclic dinucleotide (CDN) 2'3'-cGAMP, which triggers STING-dependent immunity. In Drosophila melanogaster, two cGAS-like receptors (cGLRs) produce 3'2'-cGAMP and 2'3'-cGAMP to activate STING. We explored CDN-mediated immunity in 14 Drosophila species covering 50 million years of evolution and found that 2'3'-cGAMP and 3'2'-cGAMP failed to control infection by Drosophila C virus in D. serrata and two other species. We discovered diverse CDNs produced in a cGLR-dependent manner in response to viral infection in D. melanogaster, including 2'3'-c-di-GMP. This CDN was a more potent STING agonist than cGAMP in D. melanogaster and it also activated a strong antiviral transcriptional response in D. serrata. Our results shed light on the evolution of cGLRs in flies and provide a basis for understanding the function and regulation of this emerging family of pattern recognition receptors in animal innate immunity. }, keywords = {c-di-GMP, cGAMP, cGAS, cGLR, cyclic dinucleotide, Drosophila, Evolution, Hua, imler, M3i, meignin, pattern recognition receptor, STING, virus}, pubstate = {published}, tppubtype = {article} } @article{Marois2023, title = {Screening Mosquito Larvae Under a Fluorescence Binocular Microscope}, author = {Eric Marois}, doi = {10.1101/pdb.prot108306 }, year = {2023}, date = {2023-09-11}, urldate = {2023-09-11}, journal = {Cold Spring Harb Protoc}, keywords = {fluorescent protein, M3i, marois}, pubstate = {published}, tppubtype = {article} } @article{pmid37669377, title = {Mammalian mitochondrial translation infidelity leads to oxidative stress-induced cell cycle arrest and cardiomyopathy}, author = {Wen-Qiang Zheng and Jian-Hui Zhang and Zi-Han Li and Xiuxiu Liu and Yong Zhang and Shuo Huang and Jinsong Li and Bin Zhou and Gilbert Eriani and En-Duo Wang and Xiao-Long Zhou}, doi = {10.1073/pnas.2309714120}, issn = {1091-6490}, year = {2023}, date = {2023-09-01}, urldate = {2023-09-01}, journal = {Proc Natl Acad Sci U S A}, volume = {120}, number = {37}, pages = {e2309714120}, abstract = {Proofreading (editing) of mischarged tRNAs by cytoplasmic aminoacyl-tRNA synthetases (aaRSs), whose impairment causes neurodegeneration and cardiac diseases, is of high significance for protein homeostasis. However, whether mitochondrial translation needs fidelity and the significance of editing by mitochondrial aaRSs have been unclear. Here, we show that mammalian cells critically depended on the editing of mitochondrial threonyl-tRNA synthetase (mtThrRS, encoded by ), disruption of which accumulated Ser-tRNA and generated a large abundance of Thr-to-Ser misincorporated peptides in vivo. Such infidelity impaired mitochondrial translation and oxidative phosphorylation, causing oxidative stress and cell cycle arrest in the G0/G1 phase. Notably, reactive oxygen species (ROS) scavenging by N-acetylcysteine attenuated this abnormal cell proliferation. A mouse model of heart-specific defective mtThrRS editing was established. Increased ROS levels, blocked cardiomyocyte proliferation, contractile dysfunction, dilated cardiomyopathy, and cardiac fibrosis were observed. Our results elucidate that mitochondria critically require a high level of translational accuracy at Thr codons and highlight the cellular dysfunctions and imbalance in tissue homeostasis caused by mitochondrial mistranslation.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37704020, title = {Microfluidic Droplet Stabilization via SPAAC Promoted Antibody Conjugation at the Water/Oil Interface}, author = {Robin Dufossez and Marie-Pierre Krafft and Sylvain Ursuegui and Michel Mosser and Safae Mouftakhir and Ketty Pernod and Guilhem Chaubet and Michael Ryckelynck and Alain Wagner}, doi = {10.1021/acsami.3c10655}, issn = {1944-8252}, year = {2023}, date = {2023-09-01}, urldate = {2023-09-01}, journal = {ACS Appl Mater Interfaces}, volume = {15}, issue = {38}, pages = {45498-45505}, abstract = {Droplet-based microfluidics is leading the development of miniaturized, rapid, and sensitive version of enzyme-linked immunosorbent assays (ELISAs), a central method for protein detection. These assays involve the use of a functionalized surface able to selectively capture the desired analyte. Using the droplet's oil water interface as a capture surface requires designing custom-perfluorinated fluorosurfactants bearing azide-containing polar groups, which spontaneously react when forming the droplet with strain-alkyne-functionalized antibodies solubilized in the aqueous phase. In this article, we present our research on the influence of the structure of surfactant's hydrophilic heads on the efficiency of SPAAC functionalization and on the effect of this antibody grafting process on droplet stability. We have shown that while short linkers lead to high grafting efficiency, long linkers lead to high stability, and that an intermediate size is required to balance both parameters. In the described family of surfactants, the optimal structure proved to be a PEG linker connecting a polar di-azide head and a per-fluoropolyether tail (Krytox). We also found that grafting an increasing amount of antibody, thus increasing interface coverage, increases droplet stability. It thus appears that such a bi-partite system with a reactive fluoro-surfactant in the oil phase and reactive antibody counterpart in the aqueous phase gives access in situ to novel surfactant construct providing unexplored interface structures and droplet functionality.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37741547, title = {The interplay between cis- and trans-acting factors drives selective mRNA translation initiation in eukaryotes}, author = {Antonin Tidu and Franck Martin}, doi = {10.1016/j.biochi.2023.09.017}, issn = {1638-6183}, year = {2023}, date = {2023-09-01}, urldate = {2023-09-01}, journal = {Biochimie}, volume = {217}, pages = {20-30}, abstract = {Translation initiation consists in the assembly of the small and large ribosomal subunits on the start codon. This important step directly modulates the general proteome in living cells. Recently, genome wide studies revealed unexpected translation initiation events from unsuspected novel open reading frames resulting in the synthesis of a so-called 'dark proteome'. Indeed, the identification of the start codon by the translation machinery is a critical step that defines the translational landscape of the cell. Therefore, translation initiation is a highly regulated process in all organisms. In this review, we focus on the various cis- and trans-acting factors that rule the regulation of translation initiation in eukaryotes. Recent discoveries have shown that the guidance of the translation machinery for the choice of the start codon require sophisticated molecular mechanisms. In particular, the 5'UTR and the coding sequences contain cis-acting elements that trigger the use of AUG codons but also non-AUG codons to initiate protein synthesis. The use of these alternative start codons is also largely influenced by numerous trans-acting elements that drive selective mRNA translation in response to environmental changes.}, keywords = {MARTIN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37399950, title = {Targeting and eradicating latent CNS reservoirs of HIV-1: Original strategies and new models}, author = {Sepideh Saeb and Clémentine Wallet and Olivier Rohr and Christian Schwartz and Thomas Loustau}, doi = {10.1016/j.bcp.2023.115679}, issn = {1873-2968}, year = {2023}, date = {2023-08-01}, urldate = {2023-08-01}, journal = {Biochem Pharmacol}, volume = {214}, pages = {115679}, abstract = {Nowadays, combination antiretroviral therapy (cART) is the standard treatment for all people with human immunodeficiency virus (HIV-1). Although cART is effective in treating productive infection, it does not eliminate latent reservoirs of the virus. This leads to lifelong treatment associated with the occurrence of side effects and the development of drug-resistant HIV-1. Suppression of viral latency is therefore the major hurdle to HIV-1 eradication. Multiple mechanisms exist to regulate viral gene expression and drive the transcriptional and post-transcriptional establishment of latency. Epigenetic processes are amongst the most studied mechanisms influencing both productive and latent infection states. The central nervous system (CNS) represents a key anatomical sanctuary for HIV and is the focal point of considerable research efforts. However, limited and difficult access to CNS compartments makes understanding the HIV-1 infection state in latent brain cells such as microglial cells, astrocytes, and perivascular macrophages challenging. This review examines the latest advances on epigenetic transformations involved in CNS viral latency and targeting of brain reservoirs. Evidence from clinical studies as well as in vivo and in vitro models of HIV-1 persistence in the CNS will be discussed, with a special focus on recent 3D in vitro models such as human brain organoids. Finally, the review will address therapeutic considerations for targeting latent CNS reservoirs.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{IGNELL2023, title = {A mosquito-specific antennal protein is critical for the attraction to human odor in the malaria vector Anopheles gambiae}, author = {Julien PELLETIER AND Mengistu DAWIT AND Majid GHANINIA AND Eric MAROIS AND Rickard IGNELL}, editor = { }, url = {https://doi.org/10.1016/j.ibmb.2023.103988}, doi = {j.ibmb.2023.103988}, year = {2023}, date = {2023-07-11}, urldate = {2023-07-11}, journal = {Insect Biochemistry and Molecular Biology}, volume = {159}, issue = {August 2023}, abstract = {Mosquitoes rely mainly on the sense of smell to decipher their environment and locate suitable food sources, hosts for blood feeding and oviposition sites. The molecular bases of olfaction involve multigenic families of olfactory proteins that have evolved to interact with a narrow set of odorants that are critical for survival. Understanding the complex interplay between diversified repertoires of olfactory proteins and ecologically-relevant odorant signals, which elicit important behaviors, is fundamental for the design of novel control strategies targeting the sense of smell of disease vector mosquitoes. Previously, large multigene families of odorant receptor and ionotropic receptor proteins, as well as a subset of odorant-binding proteins have been shown to mediate the selectivity and sensitivity of the mosquito olfactory system. In this study, we identify a mosquito-specific antennal protein (MSAP) gene as a novel molecular actor of odorant reception. MSAP is highly conserved across mosquito species and is transcribed at an extremely high level in female antennae. In order to understand its role in the mosquito olfactory system, we generated knockout mutant lines in Anopheles gambiae, and performed comparative analysis of behavioral and physiological responses to human-associated odorants. We found that MSAP promotes female mosquito attraction to human odor and enhances the sensitivity of the antennae to a variety of odorants. These findings suggest that MSAP is an important component of the mosquito olfactory system, which until now has gone completely unnoticed.}, keywords = {antenna, chemoreceptor, M3i, marois, mosquitoes, olfaction}, pubstate = {published}, tppubtype = {article} } @misc{pmid37394995, title = {FEBS fellowships: supporting excellent science for over four decades}, author = {Duncan E Wright and Alain Krol}, doi = {10.1002/2211-5463.13659}, issn = {2211-5463}, year = {2023}, date = {2023-07-01}, urldate = {2023-07-01}, journal = {FEBS Open Bio}, volume = {13}, number = {7}, pages = {1138--1139}, abstract = {The Federation of European Biochemical Societies (FEBS) awarded FEBS Long-Term Fellowships from 1979 until 2020, at which time the scheme was replaced with the FEBS Excellence Award. Over four decades, FEBS awarded a huge number of Long-Term Fellowships, helping to support and promote the careers of excellent young researchers across Europe. To celebrate the exciting work performed by the FEBS Long-Term Fellows, we present here a special 'In the Limelight' issue of FEBS Open Bio, containing four Mini-reviews and four Research Protocols authored by the fellows themselves. The four Review articles provide timely updates on the respective research fields, while the Research Protocols describe how to perform challenging experimental methods in detail. We hope this issue will be a valuable resource for the community, and a celebration of the high-quality work done by young scientists.}, keywords = {KROL, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{pmid37414209, title = {RNA aptamers developed against tRip: A preliminary approach targeting tRNA entry in Plasmodium}, author = {Martina Pitolli and Marta Cela and Caroline Paulus and Joëlle Rudinger-Thirion and Magali Frugier}, doi = {10.1016/j.biochi.2023.06.011}, issn = {1638-6183}, year = {2023}, date = {2023-07-01}, urldate = {2023-07-01}, journal = {Biochimie}, volume = {217}, pages = {106-115}, abstract = {Malaria is caused by Plasmodium parasites that multiply inside host cells and can be lethal when P. falciparum is involved. We identified tRip as a membrane protein that facilitates the import of exogenous transfer RNA (tRNA) into the parasite. tRip encompasses a tRNA binding domain exposed on the parasite surface. We used the SELEX approach to isolate high-affinity and specific tRip-binding RNA motifs from a library of random 25 nucleotide-long sequences. In five rounds of combined negative and positive selections, an enriched pool of aptamers was obtained; sequencing revealed that they were all different in their primary sequence; only by comparing their structure predictions did most of the selected aptamers reveal a conserved 5-nucleotide motif sequence. We showed that the integral motif is essential for tRip-binding while the rest of the molecule can be significantly reduced or mutated as long as the motif is presented in a single-stranded region. Such RNA aptamers bind in place of the original tRNA substrate and act as an efficient competitor, suggesting that they can block tRip function and slow parasite development.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37503833b, title = {SARS-CoV-2 NSP1 induces mRNA cleavages on the ribosome}, author = {Yann Tardivat and Piotr Sosnowski and Antonin Tidu and Eric Westhof and Gilbert Eriani and Franck Martin}, doi = {10.1093/nar/gkad627}, issn = {1362-4962}, year = {2023}, date = {2023-07-01}, urldate = {2023-07-01}, journal = {Nucleic Acids Res}, volume = {51}, issue = {16}, pages = {8677-8690}, abstract = {In severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the non-structural protein NSP1 inhibits translation of host mRNAs by binding to the mRNA entry channel of the ribosome and, together with the 5'-untranslated region (UTR) of the viral mRNAs, allows the evasion of that inhibition. Here, we show that NSP1 mediates endonucleolytic cleavages of both host and viral mRNAs in the 5'UTR, but with different cleavage patterns. The first pattern is observed in host mRNAs with cleavages interspersed regularly and close to the 5' cap (6-11 nt downstream of the cap). Those cleavage positions depend more on the position relative to the 5' cap than on the sequence itself. The second cleavage pattern occurs at high NSP1 concentrations and only in SARS-CoV-2 RNAs, with the cleavages clustered at positions 45, 46 and 49. Both patterns of cleavage occur with the mRNA and NSP1 bound to the ribosome, with the SL1 hairpin at the 5' end sufficient to protect from NSP1-mediated degradation at low NSP1 concentrations. We show further that the N-terminal domain of NSP1 is necessary and sufficient for efficient cleavage. We suggest that in the ribosome-bound NSP1 protein the catalytic residues of the N-terminal domain are unmasked by the remodelling of the α1- and α2-helices of the C-terminal domain.}, keywords = {ERIANI, MARTIN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Lutrat2023, title = {Combining two Genetic Sexing Strains allows sorting of non-transgenic males for Aedes genetic control}, author = {Lutrat C and Burckbuchler M and Olmo RP and Beugnon R and Fontaine A and Akbari OS and Argiles-Herrero R and Baldet T and Bouyer J and Marois E}, url = {https://www.nature.com/articles/s42003-023-05030-7}, doi = {10.1038/s42003-023-05030-7}, year = {2023}, date = {2023-06-16}, urldate = {2023-06-16}, journal = {Commun Biol.}, volume = {6}, issue = {1}, pages = {646}, abstract = {Chemical control of disease vectoring mosquitoes Aedes albopictus and Aedes aegypti is costly, unsustainable, and increasingly ineffective due to the spread of insecticide resistance. The Sterile Insect Technique is a valuable alternative but is limited by slow, error-prone, and wasteful sex-separation methods. Here, we present four Genetic Sexing Strains (two for each Aedes species) based on fluorescence markers linked to the m and M sex loci, allowing for the isolation of transgenic males. Furthermore, we demonstrate how combining these sexing strains enables the production of non-transgenic males. In a mass-rearing facility, 100,000 first instar male larvae could be sorted in under 1.5 h with an estimated 0.01–0.1% female contamination on a single machine. Cost-efficiency analyses revealed that using these strains could result in important savings while setting up and running a mass-rearing facility. Altogether, these Genetic Sexing Strains should enable a major upscaling in control programmes against these important vectors.}, keywords = {Aedes, M3i, marois, mosquitoes, Olmo, vectoring}, pubstate = {published}, tppubtype = {article} } @article{pmid37349586, title = {Cut site preference allows influenza A virus PA-X to discriminate between host and viral mRNAs}, author = {Lea Gaucherand and Amrita Iyer and Isabel Gilabert and Chris H Rycroft and Marta M Gaglia}, doi = {10.1038/s41564-023-01409-8}, issn = {2058-5276}, year = {2023}, date = {2023-06-01}, urldate = {2023-06-01}, journal = {Nat Microbiol}, abstract = {Many viruses block host gene expression to take over the infected cell. This process, termed host shutoff, is thought to promote viral replication by preventing antiviral responses and redirecting cellular resources to viral processes. Several viruses from divergent families accomplish host shutoff through RNA degradation by endoribonucleases. However, viruses also need to ensure expression of their own genes. The influenza A virus endoribonuclease PA-X solves this problem by sparing viral mRNAs and some host RNAs necessary for viral replication. To understand how PA-X distinguishes between RNAs, we characterized PA-X cut sites transcriptome-wide using 5' rapid amplification of complementary DNA ends coupled to high-throughput sequencing. This analysis, along with RNA structure predictions and validation experiments using reporters, shows that PA-Xs from multiple influenza strains preferentially cleave RNAs at GCUG tetramers in hairpin loops. Importantly, GCUG tetramers are enriched in the human but not the influenza transcriptome. Moreover, optimal PA-X cut sites inserted in the influenza A virus genome are quickly selected against during viral replication in cells. This finding suggests that PA-X evolved these cleavage characteristics to preferentially target host over viral mRNAs in a manner reminiscent of cellular self versus non-self discrimination.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37347186, title = {Complex Regulation of Gamma-Hemolysin Expression Impacts Staphylococcus aureus Virulence}, author = {Mariane Pivard and Isabelle Caldelari and Virginie Brun and Delphine Croisier and Michel Jaquinod and Nelson Anzala and Benoît Gilquin and Chloé Teixeira and Yvonne Benito and Florence Couzon and Pascale Romby and Karen Moreau and François Vandenesch}, doi = {10.1128/spectrum.01073-23}, issn = {2165-0497}, year = {2023}, date = {2023-06-01}, urldate = {2023-06-01}, journal = {Microbiol Spectr}, pages = {e0107323}, abstract = {Staphylococcus aureus gamma-hemolysin CB (HlgCB) is a core-genome-encoded pore-forming toxin that targets the C5a receptor, similar to the phage-encoded Panton-Valentine leucocidin (PVL). Absolute quantification by mass spectrometry of HlgCB in 39 community-acquired pneumonia (CAP) isolates showed considerable variations in the HlgC and HlgB yields between isolates. Moreover, although HlgC and HlgB are encoded on a single operon, their levels were dissociated in 10% of the clinical strains studied. To decipher the molecular basis for the variation in expression and protein production among strains, different regulation levels were analyzed in representative clinical isolates and reference strains. Both the HlgCB level and the HlgC/HlgB ratio were found to depend on promoter activity and mRNA processing and translation. Strikingly, only one single nucleotide polymorphism (SNP) in the 5' untranslated region (UTR) of mRNA strongly impaired translation in the USA300 strain, leading to a strong decrease in the level of HlgC but not in HlgB. Finally, we found that high levels of HlgCB synthesis led to mortality in a rabbit model of pneumonia, correlated with the implication of the role of HlgCB in severe S. aureus CAP. Taken together, this work illustrates the complexity of virulence factor expression in clinical strains and demonstrates a butterfly effect where subtle genomic variations have a major impact on phenotype and virulence. S. aureus virulence in pneumonia results in its ability to produce several virulence factors, including the leucocidin PVL. Here, we demonstrate that HlgCB, another leucocidin, which targets the same receptors as PVL, highly contributes to S. aureus virulence in -negative strains. In addition, considerable variations in HlgCB quantities are observed among clinical isolates from patients with CAP. Biomolecular analyses have revealed that a few SNPs in the promoter sequences and only one SNP in the 5' UTR of mRNA induce the differential expression of , drastically impacting mRNA translation. This work illustrates the subtlety of regulatory mechanisms in bacteria, especially the sometimes major effects on phenotypes of single nucleotide variation in noncoding regions.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37224537, title = {Sequential disruption of SPLASH-identified vRNA-vRNA interactions challenges their role in influenza A virus genome packaging}, author = {Celia Jakob and Gabriel L Lovate and Daniel Desirò and Lara Gießler and Redmond P Smyth and Roland Marquet and Kevin Lamkiewicz and Manja Marz and Martin Schwemmle and Hardin Bolte}, doi = {10.1093/nar/gkad442}, issn = {1362-4962}, year = {2023}, date = {2023-05-01}, urldate = {2023-05-01}, journal = {Nucleic Acids Res}, volume = {51}, issue = {12}, pages = {6479-6494}, abstract = {A fundamental step in the influenza A virus (IAV) replication cycle is the coordinated packaging of eight distinct genomic RNA segments (i.e. vRNAs) into a viral particle. Although this process is thought to be controlled by specific vRNA-vRNA interactions between the genome segments, few functional interactions have been validated. Recently, a large number of potentially functional vRNA-vRNA interactions have been detected in purified virions using the RNA interactome capture method SPLASH. However, their functional significance in coordinated genome packaging remains largely unclear. Here, we show by systematic mutational analysis that mutant A/SC35M (H7N7) viruses lacking several prominent SPLASH-identified vRNA-vRNA interactions involving the HA segment package the eight genome segments as efficiently as the wild-type virus. We therefore propose that the vRNA-vRNA interactions identified by SPLASH in IAV particles are not necessarily critical for the genome packaging process, leaving the underlying molecular mechanism elusive.}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37267103, title = {The E3 ubiquitin ligase FBXL6 controls the quality of newly synthesized mitochondrial ribosomal proteins}, author = {Julie Lavie and Claude Lalou and Walid Mahfouf and Jean-William Dupuy and Aurélie Lacaule and Agata Ars Cywinska and Didier Lacombe and Anne-Marie Duchêne and Anne-Aurélie Raymond and Hamid Reza Rezvani and Richard Patryk Ngondo and Giovanni Bénard}, doi = {10.1016/j.celrep.2023.112579}, issn = {2211-1247}, year = {2023}, date = {2023-05-01}, urldate = {2023-05-01}, journal = {Cell Rep}, volume = {42}, number = {6}, pages = {112579}, abstract = {In mammals, about 99% of mitochondrial proteins are synthesized in the cytosol as precursors that are subsequently imported into the organelle. The mitochondrial health and functions rely on an accurate quality control of these imported proteins. Here, we show that the E3 ubiquitin ligase F box/leucine-rich-repeat protein 6 (FBXL6) regulates the quality of cytosolically translated mitochondrial proteins. Indeed, we found that FBXL6 substrates are newly synthesized mitochondrial ribosomal proteins. This E3 binds to chaperones involved in the folding and trafficking of newly synthesized peptide and to ribosomal-associated quality control proteins. Deletion of these interacting partners is sufficient to hamper interactions between FBXL6 and its substrate. Furthermore, we show that cells lacking FBXL6 fail to degrade specifically mistranslated mitochondrial ribosomal proteins. Finally, showing the role of FBXL6-dependent mechanism, FBXL6-knockout (KO) cells display mitochondrial ribosomal protein aggregations, altered mitochondrial metabolism, and inhibited cell cycle in oxidative conditions.}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37254812, title = {A homozygous mutation in the human selenocysteine tRNA gene impairs UGA recoding activity and selenoproteome regulation by selenium}, author = {Caroline Vindry and Olivia Guillin and Philippe Wolff and Paul Marie and Franck Mortreux and Philippe E Mangeot and Théophile Ohlmann and Laurent Chavatte}, doi = {10.1093/nar/gkad482}, issn = {1362-4962}, year = {2023}, date = {2023-05-01}, urldate = {2023-05-01}, journal = {Nucleic Acids Res}, volume = {51}, issue = {14}, pages = {7580-7601}, abstract = {The selenocysteine (Sec) tRNA (tRNA[Ser]Sec) governs Sec insertion into selenoproteins by the recoding of a UGA codon, typically used as a stop codon. A homozygous point mutation (C65G) in the human tRNA[Ser]Sec acceptor arm has been reported by two independent groups and was associated with symptoms such as thyroid dysfunction and low blood selenium levels; however, the extent of altered selenoprotein synthesis resulting from this mutation has yet to be comprehensively investigated. In this study, we used CRISPR/Cas9 technology to engineer homozygous and heterozygous mutant human cells, which we then compared with the parental cell lines. This C65G mutation affected many aspects of tRNA[Ser]Sec integrity and activity. Firstly, the expression level of tRNA[Ser]Sec was significantly reduced due to an altered recruitment of RNA polymerase III at the promoter. Secondly, selenoprotein expression was strongly altered, but, more surprisingly, it was no longer sensitive to selenium supplementation. Mass spectrometry analyses revealed a tRNA isoform with unmodified wobble nucleotide U34 in mutant cells that correlated with reduced UGA recoding activities. Overall, this study demonstrates the pleiotropic effect of a single C65G mutation on both tRNA phenotype and selenoproteome expression.}, keywords = {ARN-MS, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid37254203, title = {DNA topoisomerase 1 represses HIV-1 promoter activity through its interaction with a guanine quadruplex present in the LTR sequence}, author = {María José Lista and Anne-Caroline Jousset and Mingpan Cheng and Violaine Saint-André and Elouan Perrot and Melissa Rodrigues and Carmelo Di Primo and Danielle Gadelle and Elenia Toccafondi and Emmanuel Segeral and Clarisse Berlioz-Torrent and Stéphane Emiliani and Jean-Louis Mergny and Marc Lavigne}, doi = {10.1186/s12977-023-00627-6}, issn = {1742-4690}, year = {2023}, date = {2023-05-01}, urldate = {2023-05-01}, journal = {Retrovirology}, volume = {20}, number = {1}, pages = {10}, abstract = {BACKGROUND: Once integrated in the genome of infected cells, HIV-1 provirus is transcribed by the cellular transcription machinery. This process is regulated by both viral and cellular factors, which are necessary for an efficient viral replication as well as for the setting up of viral latency, leading to a repressed transcription of the integrated provirus.nnRESULTS: In this study, we examined the role of two parameters in HIV-1 LTR promoter activity. We identified DNA topoisomerase1 (TOP1) to be a potent repressor of this promoter and linked this repression to its catalytic domain. Additionally, we confirmed the folding of a Guanine quadruplex (G4) structure in the HIV-1 promoter and its repressive effect. We demonstrated a direct interaction between TOP1 and this G4 structure, providing evidence of a functional relationship between the two repressive elements. Mutations abolishing G4 folding affected TOP1/G4 interaction and hindered G4-dependent inhibition of TOP1 catalytic activity in vitro. As a result, HIV-1 promoter activity was reactivated in a native chromatin environment. Lastly, we noticed an enrichment of predicted G4 sequences in the promoter of TOP1-repressed cellular genes.nnCONCLUSIONS: Our results demonstrate the formation of a TOP1/G4 complex on the HIV-1 LTR promoter and its repressive effect on the promoter activity. They reveal the existence of a new mechanism of TOP1/G4-dependent transcriptional repression conserved between viral and human genes. This mechanism contrasts with the known property of TOP1 as global transcriptional activator and offers new perspectives for anti-cancer and anti-viral strategies.}, keywords = {MARQUET, NEGRONI, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36436882, title = {FluorMango, an RNA-Based Fluorogenic Biosensor for the Direct and Specific Detection of Fluoride}, author = {Claire Husser and Stéphane Vuilleumier and Michael Ryckelynck}, doi = {10.1002/smll.202205232}, issn = {1613-6829}, year = {2023}, date = {2023-03-29}, urldate = {2023-03-01}, journal = {Small}, volume = {19}, issue = {13}, pages = {e2205232}, abstract = {Nucleic acids are not only essential actors of cell life but also extremely appealing molecular objects in the development of synthetic molecules for biotechnological application, such as biosensors to report on the presence and concentration of a target ligand by emission of a measurable signal. In this work, FluorMango, a fluorogenic ribonucleic acid (RNA)-based biosensor specific for fluoride is introduced. The molecule consists of two RNA aptamer modules, a fluoride-specific sensor derived from the crcB riboswitch which changes its structure upon interaction with the target ion, and the light-up RNA Mango-III that emits fluorescence when complexed with a fluorogen. The two modules are connected by an optimized communication module identified by ultrahigh-throughput screening, which results in extremely high fluorescence of FluorMango in the presence of fluoride, and background fluorescence in its absence. The value and efficiency of this biosensor by direct monitoring of defluorinase activity in living bacterial cells is illustrated, and the use of this new tool in future screening campaigns aiming at discovering new defluorinase activities is discussed.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{deFaria2023, title = {Protocol for the analysis of double-stranded RNAs in virus-infected insect cells using anti-dsRNA antibodies}, author = {Isaque J.S. de Faria and Jean-Luc Imler and João T. Marques}, url = {https://doi.org/10.1016/j.xpro.2022.102033}, doi = {10.1016/j.xpro.2022.102033}, year = {2023}, date = {2023-03-17}, urldate = {2023-03-17}, journal = {STAR Protocols}, volume = {4}, issue = {1}, abstract = {Characterization of double-stranded (ds)RNAs is relevant to the understanding of viral replication and immune sensing. Here, we provide a protocol describing the use of anti-dsRNA antibodies for immunofluorescence and immunoblotting in virus-infected insect cells, which can also be applied to tissues and other organisms. We describe the procedures to prepare insect cells for viral infection, followed by RNA extraction and in vitro production of synthetic dsRNA controls. We then detail the steps for dsRNA detection by immunoblotting and immunofluorescence. For complete details on the use and execution of this protocol, please refer to de Faria et al. (2022).1}, keywords = {antibody, cell bioloby, imler, Immunology, M3i, Marques, microbiology, Microscopy, Molecular Biology}, pubstate = {published}, tppubtype = {article} } @article{Huang2023, title = {A Toll pathway effector protects Drosophila specifically from distinct toxins secreted by a fungus or a bacterium}, author = {Jianqiong Huang and Yanyan Lou and Jiyong Liu and Philippe Bulet and Chuping Cai and Kaiyu Ma and Renjie Jiao and Jules A Hoffmann and Samuel Liégeois and Zi Lia and Dominique Ferrandon}, editor = {Hugo Bellen, Baylor College of Medicine, Houston, TX}, url = {https://doi.org/10.1073/pnas.2205140120}, doi = {10.1073/pnas.2205140120}, year = {2023}, date = {2023-03-14}, urldate = {2023-03-14}, journal = {PNAS}, volume = {120}, number = {12}, abstract = {Major immune response pathways control the expression of hundreds of genes that represent potential effectors of the immune response. The Drosophila Toll pathway is required in the host defenses against several Gram-positive bacterial infections as well as against fungal infections. The current paradigm is that peptides secreted in the hemolymph during the systemic immune response are either bona fide antimicrobial peptides or likely ones. The finding of a dual role for one Toll pathway effector in the resilience to both Enterococcus faecalis and Metarhizium robertsii infections underscores an original concept in insect innate immunity. Evolution can select effectors tailored to protect the host from the action of microbial toxins of prokaryotic or eukaryotic origin, independently of antibodies or detoxification pathways.}, keywords = {Baramicin A, Destruxin A, disease tolerance, enterocin O16, ferrandon, hoffmann, M3i, microbial toxins, resilience}, pubstate = {published}, tppubtype = {article} } @article{Klug2023, title = {The salivary protein Saglin facilitates efficient midgut colonization of Anopheles mosquitoes by malaria parasites}, author = {Dennis Klug and Amandine Gautier and Eric Calvo and Eric Marois and Stéphanie A. Blandin}, url = {https://doi.org/10.1371/journal.ppat.1010538}, doi = {10.1371/journal.ppat.1010538}, year = {2023}, date = {2023-03-02}, urldate = {2023-03-02}, booktitle = {Plos Pathogens}, journal = {Plos Pathogens}, volume = {19}, number = {3}, issue = {3}, abstract = {Female mosquitoes rely on blood feeding to acquire sufficient nutrients for egg development. Because of the importance of this process mosquitoes evolved salivary proteins with a broad range of functions acting as blood thinners, anti-coagulants and immunosuppressants. The effect of these proteins on the blood at the bite site directly influences the size of the blood bolus a female takes up in a given time frame. Both, time of feeding and bolus size, are important parameters for fecundity and survival. Recent studies have shown that a significant proportion of salivated proteins is re-ingested during feeding and becomes part of the blood meal. Here we investigated the salivary protein Saglin which has been previously suggested as putative receptor mediating malaria parasite entry into the salivary gland. By engineering a loss-of-function mutant in An. coluzzi we could show that the absence of Saglin impairs the development of parasite stages in the blood meal of the rodent malaria parasite P. berghei and the human malaria parasite P. falciparum lowering the parasite burden of subsequent stages and preventing efficient transmission at low infection densities. Furthermore, we could show that Saglin is present in the blood meal after feeding possibly indicating a previously overlooked parasite-vector interaction.}, keywords = {blandin, BLOOD, M3i, malarial parasites, marois, mosquitoes, Oocysts, Parasitic Diseases, Plasmodium, salivary glands, sporozoites}, pubstate = {published}, tppubtype = {article} } @article{pmid36996029, title = {A snapshot on HIV-1 evolution through the identification of phylogenetic-specific properties of HIV-1 integrases M/O}, author = {Elenia Toccafondi and Marine Kanja and Flore Winter and Daniela Lener and Matteo Negroni}, doi = {10.1371/journal.ppat.1011207}, issn = {1553-7374}, year = {2023}, date = {2023-03-01}, urldate = {2023-03-01}, journal = {PLoS Pathog}, volume = {19}, number = {3}, pages = {e1011207}, abstract = {Transmissions of simian viruses to humans has originated the different groups of HIV-1. We recently identified a functional motif (CLA), in the C-terminal domain of the integrase, essential for integration in HIV-1 group M. Here, we found that the motif is instead dispensable in group O isolates, because of the presence, in the N-terminal domain of HIV-1 O of a specific sequence, Q7G27P41H44, that we define as the NOG motif. Alterations of reverse transcription and of 3' processing observed by mutating the CLA motif of IN M are fully rescued to wt levels by inserting the sequence of the NOG motif in the N-ter of the protein. These results indicate that the two motifs (CLA and NOG) functionally complement each other and a working model accounting for these observations is proposed. The establishment of these two alternative motifs seems to be due to the different phylogenetic origin and history of these two groups. Indeed, the NOG motif is already present in the ancestor of group O (SIVgor) while it is absent from SIVcpzPtt, the ancestor of group M. The CLA motif, instead, seems to have emerged after SIVcpzPtt has been transferred to humans, since no conservation is found at the same positions in these simian viruses. These results show the existence of two-group specific motifs in HIV-1 M and O integrases. In each group, only one of the motifs is functional, potentially leading the other motif to diverge from its original function and, in an evolutionary perspective, assist other functions of the protein, further increasing HIV genetic diversity.}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36617674b, title = {Proteomics-based determination of double-stranded RNA interactome reveals known and new factors involved in Sindbis virus infection}, author = {Erika Girardi and Mélanie Messmer and Paula Lopez and Aurélie Fender and Johana Chicher and Béatrice Chane-Woon-Ming and Philippe Hammann and Sébastien Pfeffer}, doi = {10.1261/rna.079270.122}, issn = {1469-9001}, year = {2023}, date = {2023-03-01}, urldate = {2023-03-01}, journal = {RNA}, volume = {29}, number = {3}, pages = {361--375}, abstract = {Viruses are obligate intracellular parasites, which depend on the host cellular machineries to replicate their genome and complete their infectious cycle. Long double-stranded (ds)RNA is a common viral by-product originating during RNA virus replication and is universally sensed as a danger signal to trigger the antiviral response. As a result, viruses hide dsRNA intermediates into viral replication factories and have evolved strategies to hijack cellular proteins for their benefit. The characterization of the host factors associated with viral dsRNA and involved in viral replication remains a major challenge to develop new antiviral drugs against RNA viruses. Here, we performed anti-dsRNA immunoprecipitation followed by mass spectrometry analysis to fully characterize the dsRNA interactome in Sindbis virus (SINV) infected human cells. Among the identified proteins, we characterized SFPQ (splicing factor, proline-glutamine rich) as a new dsRNA-associated proviral factor upon SINV infection. We showed that SFPQ depletion reduces SINV infection in human HCT116 and SK-N-BE(2) cells, suggesting that SFPQ enhances viral production. We demonstrated that the cytoplasmic fraction of SFPQ partially colocalizes with dsRNA upon SINV infection. In agreement, we proved by RNA-IP that SFPQ can bind dsRNA and viral RNA. Furthermore, we showed that overexpression of a wild-type, but not an RNA binding mutant SFPQ, increased viral infection, suggesting that RNA binding is essential for its positive effect on the virus. Overall, this study provides the community with a compendium of dsRNA-associated factors during viral infection and identifies SFPQ as a new proviral dsRNA binding protein.}, keywords = {PFEFFER, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36821722, title = {Droplet Surface Immunoassay by Relocation (D-SIRe) for High-Throughput Analysis of Cytosolic Proteins at the Single-Cell Level}, author = {Robin Dufossez and Sylvain Ursuegui and Stephanie Baudrey and Ketty Pernod and Safae Mouftakhir and Mustapha Oulad-Abdelghani and Michel Mosser and Guilhem Chaubet and Michael Ryckelynck and Alain Wagner}, doi = {10.1021/acs.analchem.2c05168}, issn = {1520-6882}, year = {2023}, date = {2023-02-01}, urldate = {2023-02-01}, journal = {Anal Chem}, volume = {95}, issue = {9}, pages = {4470-4478}, abstract = {Enzyme-linked immunosorbent assay (ELISA) is a central analytic method in biological science for the detection of proteins. Introduction of droplet-based microfluidics allowed the development of miniaturized, less-consuming, and more sensitive ELISA assays by coencapsulating the biological sample and antibody-functionalized particles. We report herein an alternative in-droplet immunoassay format, which avoids the use of particles. It exploits the oil/aqueous-phase interface as a protein capture and detection surface. This is achieved using tailored perfluorinated surfactants bearing azide-functionalized PEG-based polar headgroups, which spontaneously react when meeting at the droplet formation site, with strained alkyne-functionalized antibodies solubilized in the water phase. The resulting antibody-functionalized inner surface can then be used to capture a target protein. This surface capture process leads to concomitant relocation at the surface of a labeled detection antibody and in turn to a drastic change in the shape of the fluorescence signal from a convex shape (not captured) to a characteristic concave shape (captured). This novel droplet surface immunoassay by fluorescence relocation (D-SIRe) proved to be fast and sensitive at 2.3 attomoles of analyte per droplet. It was further demonstrated to allow detection of cytosolic proteins at the single bacteria level.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36800291, title = {Promiscuous splicing-derived hairpins are dominant substrates of tailing-mediated defense of miRNA biogenesis in mammals}, author = {Seungjae Lee and David Jee and Sid Srivastava and Acong Yang and Abhinav Ramidi and Renfu Shang and Diane Bortolamiol-Becet and Sébastien Pfeffer and Shuo Gu and Jiayu Wen and Eric C Lai}, doi = {10.1016/j.celrep.2023.112111}, issn = {2211-1247}, year = {2023}, date = {2023-02-01}, urldate = {2023-02-01}, journal = {Cell Rep}, volume = {42}, number = {2}, pages = {112111}, abstract = {Canonical microRNA (miRNA) hairpins are processed by the RNase III enzymes Drosha and Dicer into ∼22 nt RNAs loaded into an Argonaute (Ago) effector. In addition, splicing generates numerous intronic hairpins that bypass Drosha (mirtrons) to yield mature miRNAs. Here, we identify hundreds of previously unannotated, splicing-derived hairpins in intermediate-length (∼50-100 nt) but not small (20-30 nt) RNA data. Since we originally defined mirtrons from small RNA duplexes, we term this larger set as structured splicing-derived RNAs (ssdRNAs). These associate with Dicer and/or Ago complexes, but generally accumulate modestly and are poorly conserved. We propose they contaminate the canonical miRNA pathway, which consequently requires defense against the siege of splicing-derived substrates. Accordingly, ssdRNAs/mirtrons comprise dominant hairpin substrates for 3' tailing by multiple terminal nucleotidyltransferases, notably TUT4/7 and TENT2. Overall, the rampant proliferation of young mammalian mirtrons/ssdRNAs, coupled with an inhibitory molecular defense, comprises a Red Queen's race of intragenomic conflict.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36744444, title = {The tRNA identity landscape for aminoacylation and beyond}, author = {Richard Giegé and Gilbert Eriani}, doi = {10.1093/nar/gkad007}, issn = {1362-4962}, year = {2023}, date = {2023-02-01}, urldate = {2023-02-01}, journal = {Nucleic Acids Res}, volume = {51}, number = {4}, pages = {1528--1570}, abstract = {tRNAs are key partners in ribosome-dependent protein synthesis. This process is highly dependent on the fidelity of tRNA aminoacylation by aminoacyl-tRNA synthetases and relies primarily on sets of identities within tRNA molecules composed of determinants and antideterminants preventing mischarging by non-cognate synthetases. Such identity sets were discovered in the tRNAs of a few model organisms, and their properties were generalized as universal identity rules. Since then, the panel of identity elements governing the accuracy of tRNA aminoacylation has expanded considerably, but the increasing number of reported functional idiosyncrasies has led to some confusion. In parallel, the description of other processes involving tRNAs, often well beyond aminoacylation, has progressed considerably, greatly expanding their interactome and uncovering multiple novel identities on the same tRNA molecule. This review highlights key findings on the mechanistics and evolution of tRNA and tRNA-like identities. In addition, new methods and their results for searching sets of multiple identities on a single tRNA are discussed. Taken together, this knowledge shows that a comprehensive understanding of the functional role of individual and collective nucleotide identity sets in tRNA molecules is needed for medical, biotechnological and other applications.}, keywords = {ERIANI, GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Olmo.2023, title = {Mosquito vector competence for dengue is modulated by insect-specific viruses}, author = {Olmo R.P. and Todjro Y.M.H. and Aguiar E.R.G.R. and de Almeida J.P.P. and Ferreira F.V. and Armache J.N. and de Faria I.J.S. and Ferreira A.G.A. and Amadou S.C.D. and Silva A.T.S. and de Souza K.P.R. and Vilela A.P.P. and Babarit A. and Tan C.H. and Diallo M. and Gaye A. and Paupy C. and Obame-Nkoghe J. and Visser T.M. and Koenraadt C.J.M. and Wongsokarijo M.A. and Cruz A.L.C. and Prieto M.T. and Parra M.C.P. and Nogueira M.L., and Avelino-Silva V. and Mota R.N. and Borges M.A.Z. and Drumond B.P. and Kroon E.G. and Recker M. and Sedda L. and Marois E. and Imler J.L. and Marques J.T. }, url = {https://doi.org/}, doi = {10.1038/s41564-022-01289-4}, year = {2023}, date = {2023-01-05}, urldate = {2023-01-05}, journal = {Nature Microbiology}, volume = {8}, issue = {1}, abstract = {Aedes aegypti and A. albopictus mosquitoes are the main vectors for dengue virus (DENV) and other arboviruses, including Zika virus (ZIKV). Understanding the factors that affect transmission of arboviruses from mosquitoes to humans is a priority because it could inform public health and targeted interventions. Reasoning that interactions among viruses in the vector insect might affect transmission, we analysed the viromes of 815 urban Aedes mosquitoes collected from 12 countries worldwide. Two mosquito-specific viruses, Phasi Charoen-like virus (PCLV) and Humaita Tubiacanga virus (HTV), were the most abundant in A. aegypti worldwide. Spatiotemporal analyses of virus circulation in an endemic urban area revealed a 200% increase in chances of having DENV in wild A. aegypti mosquitoes when both HTV and PCLV were present. Using a mouse model in the laboratory, we showed that the presence of HTV and PCLV increased the ability of mosquitoes to transmit DENV and ZIKV to a vertebrate host. By transcriptomic analysis, we found that in DENV-infected mosquitoes, HTV and PCLV block the downregulation of histone H4, which we identify as an important proviral host factor in vivo.}, keywords = {Dengue, imler, M3i, marois, Marques, mosquito, Olmo, virus}, pubstate = {published}, tppubtype = {article} } @article{pmid36156788, title = {High-Throughput Development and Optimization of RNA-Based Fluorogenic Biosensors of Small Molecules Using Droplet-Based Microfluidics}, author = {Claire Husser and Stéphanie Baudrey and Michael Ryckelynck}, doi = {10.1007/978-1-0716-2695-5_19}, issn = {1940-6029}, year = {2023}, date = {2023-01-01}, urldate = {2023-01-01}, journal = {Methods Mol Biol}, volume = {2570}, pages = {243--269}, abstract = {Small-molecule sensing is a major issue as they can serve both in fundamental science and as makers of various diseases, contaminations, or even environment pollution. RNA aptamers are single-stranded nucleic acids that can adopt different conformations and specifically recognize a wide range of ligands, making them good candidates to develop biosensors of small molecules. Recently, light-up RNA aptamers have been introduced and used as starting building blocks of RNA-based fluorogenic biosensors. They are typically made of three domains: a reporter domain (a light-up aptamer), connected to a sensor domain (another aptamer) via a communication module. The latter is instrumental as being in charge of information transmission between the sensor and the reporting domains. Here we present an ultrahigh-throughput screening procedure to develop RNA-based fluorogenic biosensors by selecting optimized communication modules through an exhaustive functional exploration of every possible sequence permutation using droplet-based microfluidics and next-generation sequencing.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36598775, title = {Nucleos'ID: A New Search Engine Enabling the Untargeted Identification of RNA Post-transcriptional Modifications from Tandem Mass Spectrometry Analyses of Nucleosides}, author = {Clarisse Gosset-Erard and Mévie Didierjean and Jérome Pansanel and Antony Lechner and Philippe Wolff and Lauriane Kuhn and Frédéric Aubriet and Emmanuelle Leize-Wagner and Patrick Chaimbault and Yannis-Nicolas François}, doi = {10.1021/acs.analchem.2c04722}, issn = {1520-6882}, year = {2023}, date = {2023-01-01}, urldate = {2023-01-01}, journal = {Anal Chem}, volume = {95}, issue = {2}, pages = {1608-1617}, abstract = {As RNA post-transcriptional modifications are of growing interest, several methods were developed for their characterization. One of them established for their identification, at the nucleosidic level, is the hyphenation of separation methods, such as liquid chromatography or capillary electrophoresis, to tandem mass spectrometry. However, to our knowledge, no software is yet available for the untargeted identification of RNA post-transcriptional modifications from MS/MS data-dependent acquisitions. Thus, very long and tedious manual data interpretations are required. To meet the need of easier and faster data interpretation, a new user-friendly search engine, called Nucleos'ID, was developed for CE-MS/MS and LC-MS/MS users. Performances of this new software were evaluated on CE-MS/MS data from nucleoside analyses of already well-described transfer RNA and total tRNA extract. All samples showed great true positive, true negative, and false discovery rates considering the database size containing all modified and unmodified nucleosides referenced in the literature. The true positive and true negative rates obtained were above 0.94, while the false discovery rates were between 0.09 and 0.17. To increase the level of sample complexity, untargeted identification of several RNA modifications from 70S ribosome was achieved by the Nucleos'ID search following CE-MS/MS analysis.}, keywords = {ARN-MS, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36308692, title = {Exploring Protein Interactome Data with IPinquiry: Statistical Analysis and Data Visualization by Spectral Counts}, author = {Lauriane Kuhn and Timothée Vincent and Philippe Hammann and Hélène Zuber}, doi = {10.1007/978-1-0716-1967-4_11}, issn = {1940-6029}, year = {2023}, date = {2023-01-01}, journal = {Methods Mol Biol}, volume = {2426}, pages = {243--265}, abstract = {Immunoprecipitation mass spectrometry (IP-MS) is a popular method for the identification of protein-protein interactions. This approach is particularly powerful when information is collected without a priori knowledge and has been successively used as a first key step for the elucidation of many complex protein networks. IP-MS consists in the affinity purification of a protein of interest and of its interacting proteins followed by protein identification and quantification by mass spectrometry analysis. We developed an R package, named IPinquiry, dedicated to IP-MS analysis and based on the spectral count quantification method. The main purpose of this package is to provide a simple R pipeline with a limited number of processing steps to facilitate data exploration for biologists. This package allows to perform differential analysis of protein accumulation between two groups of IP experiments, to retrieve protein annotations, to export results, and to create different types of graphics. Here we describe the step-by-step procedure for an interactome analysis using IPinquiry from data loading to result export and plot production.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid36606712, title = {Solution x-ray scattering highlights discrepancies in Plasmodium multi-aminoacyl-tRNA synthetase complexes}, author = {José R Jaramillo Ponce and Anne Théobald-Dietrich and Philippe Bénas and Caroline Paulus and Claude Sauter and Magali Frugier}, doi = {10.1002/pro.4564}, issn = {1469-896X}, year = {2023}, date = {2023-01-01}, urldate = {2023-01-01}, journal = {Protein Sci}, pages = {e4564}, abstract = {tRip is a tRNA import protein specific to Plasmodium, the causative agent of malaria. In addition to its membrane localization and tRNA trafficking properties, tRip has the capacity to associate with three aminoacyl-tRNA synthetases (aaRS), the glutamyl- (ERS), glutaminyl- (QRS), and methionyl- (MRS) tRNA synthetases. In eukaryotes, such multi-aaRSs complexes (MSC) regulate the moonlighting activities of aaRSs. In Plasmodium, tRip and the three aaRSs all contain an N-terminal GST-like domain involved in the assembly of two independent complexes: the Q-complex (tRip:ERS:QRS) and the M-complex (tRip:ERS:MRS) with a 2:2:2 stoichiometry and in which the association of the GST-like domains of tRip and ERS (tRip-N:ERS-N) is central. In this study, the crystal structure of the N-terminal GST-like domain of ERS was solved and made possible further investigation of the solution architecture of the Q- and M-complexes by small-angle X-ray scattering (SAXS). This strategy relied on the engineering of a tRip-N-ERS-N chimeric protein to study the structural scaffold of both Plasmodium MSCs and confirm the unique homodimerization pattern of tRip in solution. The biological impact of these structural arrangements is discussed. This article is protected by copyright. All rights reserved.}, keywords = {FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Algorithms and programs for the shell decomposition of oscillating functions in space}, author = {Urzhumtseva L. and Lunin V. and Urzhumtsev A.}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1107/S160057672201144X}, doi = {10.1107/S160057672201144X}, year = {2023}, date = {2023-01-01}, urldate = {2023-01-01}, journal = {J Appl Cryst}, volume = {56}, number = {1}, pages = {302-311}, abstract = {Real-space refinement of atomic models in macromolecular crystallography and cryo-electron microscopy fits a model to a map obtained with experimental data. To do so, the atomic model is converted into a map of limited resolution and then this map is compared quantitatively with the experimental one. For an appropriate comparison, the atomic contributions comprising the model map should reflect the resolution of the experimental map and the atomic displacement parameter (ADP) values. Such contributions are spherically symmetric oscillating functions, different for chemically different kinds of atoms, different ADPs and different resolution values, and their derivatives with respect to atomic parameters rule the model refinement. For given parameter values, every contribution may be calculated numerically using two Fourier transforms, which is highly time consuming and makes calculation of the respective derivatives problematic. Alternatively, for an atom of each required type its contribution can be expressed in an analytical form as a sum of specially designed terms. Each term is different from zero essentially inside a spherical shell, and changing the ADP value does not change its form but rather changes the value of one of its arguments. In general, these terms become a convenient tool for the decomposition of oscillating spherically symmetric functions. This work describes the algorithms and respective software, named dec3D, to carry out such a shell decomposition for density contributions of different kinds of atoms and ions.}, keywords = {Unité ARN, Urzhumtseva}, pubstate = {published}, tppubtype = {article} } @article{pmid36846801b, title = { CspA stimulates translation in the cold of its own mRNA by promoting ribosome progression}, author = {Anna Maria Giuliodori and Riccardo Belardinelli and Melodie Duval and Raffaella Garofalo and Emma Schenckbecher and Vasili Hauryliuk and Eric Ennifar and Stefano Marzi}, doi = {10.3389/fmicb.2023.1118329}, issn = {1664-302X}, year = {2023}, date = {2023-01-01}, urldate = {2023-01-01}, journal = {Front Microbiol}, volume = {14}, pages = {1118329}, abstract = { CspA is an RNA binding protein that accumulates during cold-shock and stimulates translation of several mRNAs-including its own. Translation in the cold of mRNA involves a cis-acting thermosensor element, which enhances ribosome binding, and the trans-acting action of CspA. Using reconstituted translation systems and probing experiments we show that, at low temperature, CspA specifically promotes the translation of the mRNA folded in the conformation less accessible to the ribosome, which is formed at 37°C but is retained upon cold shock. CspA interacts with its mRNA without inducing large structural rearrangements, but allowing the progression of the ribosomes during the transition from translation initiation to translation elongation. A similar structure-dependent mechanism may be responsible for the CspA-dependent translation stimulation observed with other probed mRNAs, for which the transition to the elongation phase is progressively facilitated during cold acclimation with the accumulation of CspA.}, keywords = {ENNIFAR, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @misc{pmid37383636, title = {Editorial: Interview with the translational apparatus: stories of intriguing circuits and mechanisms to regulate translation in bacteria, volume II}, author = {Anna Maria Giuliodori and Paola Londei and Stefano Marzi}, doi = {10.3389/fmicb.2023.1195257}, issn = {1664-302X}, year = {2023}, date = {2023-01-01}, urldate = {2023-01-01}, journal = {Front Microbiol}, volume = {14}, pages = {1195257}, keywords = {MARZI, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @inbook{nokey, title = {Methods to Analyze Post-transcriptional Modifications Applied to Stable RNAs in Staphylococcus aureus}, author = {R. Bahena-Ceron and J. R. Jaramillo Ponce and H. Kanazawa and L. Antoine and P. Wolff and V. Marchand and B. Klaholz and Y Motorin and P. Romby and S. Marzi}, editor = {Springer}, url = {https://link.springer.com/chapter/10.1007/978-3-031-36390-0_11}, year = {2023}, date = {2023-01-01}, urldate = {2023-01-01}, booktitle = {RNA Structure and Function}, volume = {14}, pages = {233-258}, abstract = {RNA modifications contribute to the various functions of RNAs in all living organisms. Some of these modifications are dynamic and contribute to the regulation of gene expression. In bacteria, their roles in stress, environmental adaptation, and in infections caused by pathogens have been recently fully recognized. In this review, we describe several methodologies including mass spectrometry, next-generation RNA sequencing methods, biochemical approaches, and cryo-EM structural analysis that are used to detect and localize the modifications in tRNAs and rRNAs. We illustrate how the combination of methods was necessary to avoid technical biases for a successful mapping of the modifications in tRNAs and rRNAs in Staphylococcus aureus.}, keywords = {ROMBY, ROMBY MARZI, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{pmid36344159, title = {Data, data, burning deep, in the forests of the net}, author = {E Westhof}, doi = {10.1016/j.bbrc.2022.09.030}, issn = {1090-2104}, year = {2022}, date = {2022-12-01}, urldate = {2022-12-01}, journal = {Biochem Biophys Res Commun}, volume = {633}, pages = {42-44}, abstract = {Continuous and imaginative technological developments are leading to a massive accumulation of various types of data in all areas of biological research. As a result, the central importance of databases is increasing. Databases related to biology must not only be structured using controlled vocabularies, but also be fully integrated into the whole biological domain. To achieve this goal, they must be systematically grounded in biological evolution and exploit the available tools of evolutionary systematics to contribute to our understanding of life processes.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{pmid36458616, title = {Direct calculation of cryo-EM and crystallographic model maps for real-space refinement}, author = {Alexandre G Urzhumtsev and Ludmila M Urzhumtseva and Vladimir Y Lunin}, doi = {10.1107/S2059798322010907}, issn = {2059-7983}, year = {2022}, date = {2022-12-01}, urldate = {2022-12-01}, journal = {Acta Crystallogr D Struct Biol}, volume = {78}, number = {Pt 12}, pages = {1451--1468}, abstract = {This work addresses the problem of the calculation of limited-resolution maps from an atomic model in cryo-electron microscopy and in X-ray and neutron crystallography, including cases where the resolution varies from one molecular region to another. Such maps are necessary in real-space refinement for comparison with the experimental maps. For an appropriate numeric comparison, the calculated maps should reproduce not only the structural features contained in the experimental maps but also the principal map distortions. These model maps can be obtained with no use of Fourier transforms but, similar to density distributions, as a sum of individual atomic contributions. Such contributions, referred to as atomic density images, are atomic densities morphed to reflect distortions of the experimental map, in particular the loss of resolution. They are described by functions composed of a central peak surrounded by Fourier ripples. For practical calculations, atomic images should be cut at some distance. It is shown that to reach a reasonable accuracy such a distance should be significantly larger than the distance customarily applied when calculating density distributions. This is a consequence of the slow rate with which the amplitude of the Fourier ripples decreases. Such a large distance means that at least a few ripples should be included in calculations in order to obtain a map that is sufficiently accurate. Oscillating functions describing these atomic contributions depend, for a given atomic type, on the resolution and on the atomic displacement parameter values. To express both the central peak and the Fourier ripples of the atomic images, these functions are represented by the sums of especially designed terms, each concentrated in a spherical shell and depending analytically on the atomic parameters. In this work, the strength of the dependence of the accuracy of resulting map on the accuracy of the atomic displacement parameters and on the truncation distance, i.e. the number of ripples included in atomic density images, is analyzed. This analysis is completed by practical aspects of the calculation of maps of inhomogeneous resolution. Tests show that the calculation of limited-resolution maps from an atomic model as a sum of atomic contributions requires a large truncation radius extending beyond the central peak of an atomic image and the first Fourier ripples. The article discusses the practical details of such calculations expressing atomic contributions as analytic functions of the atomic coordinates, the atomic displacement parameters and the local resolution.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36450801, title = {LARS2 variants can present as premature ovarian insufficiency in the absence of overt hearing loss}, author = {Anne Sophie Neyroud and Joëlle Rudinger-Thirion and Magali Frugier and Lisa G Riley and Maud Bidet and Linda Akloul and Andrea Simpson and David Gilot and John Christodoulou and Célia Ravel and Andrew H Sinclair and Marc-Antoine Belaud-Rotureau and Elena J Tucker and Sylvie Jaillard}, doi = {10.1038/s41431-022-01252-1}, issn = {1476-5438}, year = {2022}, date = {2022-12-01}, urldate = {2022-12-01}, journal = {Eur J Hum Genet}, volume = {31}, issue = {4}, pages = {453-460}, abstract = {Premature ovarian insufficiency (POI) affects 1 in 100 women and is a leading cause of female infertility. There are over 80 genes in which variants can cause POI, with these explaining only a minority of cases. Whole exome sequencing (WES) can be a useful tool for POI patient management, allowing clinical care to be personalized to underlying cause. We performed WES to investigate two French sisters, whose only clinical complaint was POI. Surprisingly, they shared one known and one novel likely pathogenic variant in the Perrault syndrome gene, LARS2. Using amino-acylation studies, we established that the novel missense variant significantly impairs LARS2 function. Perrault syndrome is characterized by sensorineural hearing loss in addition to POI. This molecular diagnosis alerted the sisters to the significance of their difficulty in following conversation. Subsequent audiology assessment revealed a mild bilateral hearing loss. We describe the first cases presenting with perceived isolated POI and causative variants in a Perrault syndrome gene. Our study expands the phenotypic spectrum associated with LARS2 variants and highlights the clinical benefit of having a genetic diagnosis, with prediction of potential co-morbidity and prompt and appropriate medical care, in this case by an audiologist for early detection of hearing loss.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36454943, title = {Multiscale Modeling of Phosphate···π Contacts in RNA U-Turns Exposes Differences between Quantum-Chemical and AMBER Force Field Descriptions}, author = {Klaudia Mráziková and Holger Kruse and Vojtěch Mlýnský and Pascal Auffinger and Jiří Šponer}, doi = {10.1021/acs.jcim.2c01064}, issn = {1549-960X}, year = {2022}, date = {2022-12-01}, urldate = {2022-12-01}, journal = {J Chem Inf Model}, volume = {62}, issue = {23}, pages = {6182-6200}, abstract = {Phosphate···π, also called anion···π, contacts occur between nucleobases and anionic phosphate oxygens (OP2) in r(GNRA) and r(UNNN) U-turn motifs (N = A,G,C,U; R = A,G). These contacts were investigated using state-of-the-art quantum-chemical methods (QM) to characterize their physicochemical properties and to serve as a reference to evaluate AMBER force field (AFF) performance. We found that phosphate···π interaction energies calculated with the AFF for dimethyl phosphate···nucleobase model systems are less stabilizing in comparison with double-hybrid DFT and that minimum contact distances are larger for all nucleobases. These distance stretches are also observed in large-scale AFF vs QM/MM computations and classical molecular dynamics (MD) simulations on several r(gcGNRAgc) tetraloop hairpins when compared to experimental data extracted from X-ray/cryo-EM structures (res. ≤ 2.5 Å) using the WebFR3D bioinformatic tool. MD simulations further revealed shifted OP2/nucleobase positions. We propose that discrepancies between the QM and AFF result from a combination of missing polarization in the AFF combined with too large AFF Lennard-Jones (LJ) radii of nucleobase carbon atoms in addition to an exaggerated short-range repulsion of the LJ repulsive term. We compared these results with earlier data gathered on lone pair···π contacts in CpG Z-steps occurring in r(UNCG) tetraloops. In both instances, charge transfer calculations do not support any significant → π* donation effects. We also investigated thiophosphate···π contacts that showed reduced stabilizing interaction energies when compared to phosphate···π contacts. Thus, we challenge suggestions that the experimentally observed enhanced thermodynamic stability of phosphorothioated r(GNRA) tetraloops can be explained by larger London dispersion.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36617674, title = {Proteomics-based determination of double stranded RNA interactome reveals known and new factors involved in Sindbis virus infection}, author = {Erika Girardi and Melanie Messmer and Paula Lopez and Aurelie Fender and Johana Chicher and Beatrice Chane-Woon-Ming and Philippe Hammann and Sebastien Pfeffer}, doi = {10.1261/rna.079270.122}, issn = {1469-9001}, year = {2022}, date = {2022-12-01}, urldate = {2022-12-01}, journal = {RNA}, volume = {29}, issue = {3}, pages = {361-375}, abstract = {Viruses are obligate intracellular parasites, which depend on the host cellular machineries to replicate their genome and complete their infectious cycle. Long double stranded (ds)RNA is a common viral by-product originating during RNA virus replication and is universally sensed as a danger signal to trigger the antiviral response. As a result, viruses hide dsRNA intermediates into viral replication factories and have evolved strategies to hijack cellular proteins for their benefit. The characterization of the host factors associated with viral dsRNA and involved in viral replication remains a major challenge to develop new antiviral drugs against RNA viruses. Here, we performed anti-dsRNA immunoprecipitation followed by mass spectrometry analysis to fully characterize the dsRNA interactome in Sindbis virus (SINV) infected human cells. Among the identified proteins, we characterized SFPQ (Splicing factor, proline-glutamine rich) as a new dsRNA-associated proviral factor upon SINV infection. We showed that SFPQ depletion reduces SINV infection in human HCT116 and SK-N-BE(2) cells, suggesting that SFPQ enhances viral production. We demonstrated that the cytoplasmic fraction of SFPQ partially colocalizes with dsRNA upon SINV infection. In agreement, we proved by RNA-IP that SFPQ can bind dsRNA and viral RNA. Furthermore, we showed that overexpression of a wild type, but not an RNA binding mutant SFPQ, increased viral infection, suggesting that RNA binding is essential for its positive effect on the virus. Overall, this study provides the community with a compendium of dsRNA-associated factors during viral infection and identifies SFPQ as a new proviral dsRNA binding protein.}, keywords = {PFEFFER, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36322050, title = {The Toll pathway mediates Drosophila resilience to Aspergillus mycotoxins through specific Bomanins}, author = {Rui Xu and Yanyan Lou and Antonin Tidu and Philippe Bulet and Thorsten Heinekamp and Franck Martin and Axel Brakhage and Zi Li and Samuel Liégeois and Dominique Ferrandon}, url = {https://pubmed.ncbi.nlm.nih.gov/36322050/}, doi = {10.15252/embr.202256036}, issn = {1469-3178}, year = {2022}, date = {2022-11-01}, urldate = {2022-11-01}, journal = {EMBO Rep}, pages = {e56036}, abstract = {Host defense against infections encompasses both resistance, which targets microorganisms for neutralization or elimination, and resilience/disease tolerance, which allows the host to withstand/tolerate pathogens and repair damages. In Drosophila, the Toll signaling pathway is thought to mediate resistance against fungal infections by regulating the secretion of antimicrobial peptides, potentially including Bomanins. We find that Aspergillus fumigatus kills Drosophila Toll pathway mutants without invasion because its dissemination is blocked by melanization, suggesting a role for Toll in host defense distinct from resistance. We report that mutants affecting the Toll pathway or the 55C Bomanin locus are susceptible to the injection of two Aspergillus mycotoxins, restrictocin and verruculogen. The vulnerability of 55C deletion mutants to these mycotoxins is rescued by the overexpression of Bomanins specific to each challenge. Mechanistically, flies in which BomS6 is expressed in the nervous system exhibit an enhanced recovery from the tremors induced by injected verruculogen and display improved survival. Thus, innate immunity also protects the host against the action of microbial toxins through secreted peptides and thereby increases its resilience to infection.}, keywords = {ERIANI, ferrandon, M3i, MARTIN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36422372, title = {A Methylotrophic Bacterium Growing with the Antidiabetic Drug Metformin as Its Sole Carbon, Nitrogen and Energy Source}, author = {Pauline Chaignaud and Christelle Gruffaz and Adrien Borreca and Stéphanie Fouteau and Lauriane Kuhn and Jérémy Masbou and Zoé Rouy and Philippe Hammann and Gwenaël Imfeld and David Roche and Stéphane Vuilleumier}, doi = {10.3390/microorganisms10112302}, issn = {2076-2607}, year = {2022}, date = {2022-11-01}, journal = {Microorganisms}, volume = {10}, number = {11}, abstract = {Metformin is one of the most prescribed antidiabetic agents worldwide and is also considered for other therapeutic applications including cancer and endocrine disorders. It is largely unmetabolized by human enzymes and its presence in the environment has raised concern, with reported toxic effects on aquatic life and potentially also on humans. We report on the isolation and characterisation of strain MD1, an aerobic methylotrophic bacterium growing with metformin as its sole carbon, nitrogen and energy source. Strain MD1 degrades metformin into dimethylamine used for growth, and guanylurea as a side-product. Sequence analysis of its fully assembled genome showed its affiliation to . Differential proteomics and transcriptomics, as well as mini-transposon mutagenesis of the strain, point to genes and proteins essential for growth with metformin and potentially associated with hydrolytic C-N cleavage of metformin or with cellular transport of metformin and guanylurea. The obtained results suggest the recent evolution of the growth-supporting capacity of strain MD1 to degrade metformin. Our results identify candidate proteins of the enzymatic system for metformin transformation in strain MD1 and will inform future research on the fate of metformin and its degradation products in the environment and in humans.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid36353712b, title = { 30S Ribosomal Subunit Purification and Its Biochemical and Cryo-EM Analysis}, author = {Margarita Belinite and Iskander Khusainov and Stefano Marzi}, doi = {10.21769/BioProtoc.4532}, issn = {2331-8325}, year = {2022}, date = {2022-10-01}, urldate = {2022-10-01}, journal = {Bio Protoc}, volume = {12}, number = {20}, abstract = {The ribosome is a complex cellular machinery whose solved structure allowed for an incredible leap in structural biology research. Different ions bind to the ribosome, stabilizing inter-subunit interfaces and structurally linking rRNAs, proteins, and ligands. Besides cations such as K and Mg , polyamines are known to stabilize the folding of RNA and overall structure. The bacterial ribosome is composed of a small (30S) subunit containing the decoding center and a large (50S) subunit devoted to peptide bond formation. We have previously shown that the small ribosomal subunit of is sensitive to changes in ionic conditions and polyamines concentration. In particular, its decoding center, where mRNA codons and tRNA anticodons interact, is prone to structural deformations in the absence of spermidine. Here, we report a detailed protocol for the purification of the intact and functional 30S, achieved through specific ionic conditions and the addition of spermidine. Using this protocol, we obtained the cryo-electron microscopy (cryo-EM) structure of the 30S-mRNA complex from at 3.6 Å resolution. The 30S-mRNA complex formation was verified by a toeprinting assay. In this article, we also include a description of toeprinting and cryo-EM protocols. The described protocols can be further used to study the process of translation regulation. Graphical abstract.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36050837, title = {FRIENDLY (FMT) is an RNA binding protein associated with cytosolic ribosomes at the mitochondrial surface}, author = {Mickaele Hemono and Thalia Salinas-Giegé and Jeanne Roignant and Audrey Vingadassalon and Philippe Hammann and Elodie Ubrig and Patryk Ngondo and Anne-Marie Duchêne}, doi = {10.1111/tpj.15962}, issn = {1365-313X}, year = {2022}, date = {2022-10-01}, urldate = {2022-10-01}, journal = {Plant J}, volume = {112}, number = {2}, pages = {309--321}, abstract = {The spatial organization of protein synthesis in the eukaryotic cell is essential for maintaining the integrity of the proteome and the functioning of the cell. Translation on free polysomes or on ribosomes associated with the endoplasmic reticulum has been studied for a long time. More recent data have revealed selective translation of mRNAs in other compartments, in particular at the surface of mitochondria. Although these processes have been described in many organisms, particularky in plants, the mRNA targeting and localized translation mechanisms remain poorly understood. Here, the Arabidopsis thaliana Friendly (FMT) protein is shown to be a cytosolic RNA binding protein that associates with cytosolic ribosomes at the surface of mitochondria. FMT knockout delays seedling development and causes mitochondrial clustering. The mutation also disrupts the mitochondrial proteome, as well as the localization of nuclear transcripts encoding mitochondrial proteins at the surface of mitochondria. These data indicate that FMT participates in the localization of mRNAs and their translation at the surface of mitochondria.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid36161446, title = {The DEAD box RNA helicase DDX42 is an intrinsic inhibitor of positive-strand RNA viruses}, author = {Boris Bonaventure and Antoine Rebendenne and Ana Luiza Chaves Valadão and Mary Arnaud-Arnould and Ségolène Gracias and Francisco Garcia de Gracia and Joe McKellar and Emmanuel Labaronne and Marine Tauziet and Valérie Vivet-Boudou and Eric Bernard and Laurence Briant and Nathalie Gros and Wassila Djilli and Valérie Courgnaud and Hugues Parrinello and Stéphanie Rialle and Mickaël Blaise and Laurent Lacroix and Marc Lavigne and Jean-Christophe Paillart and Emiliano P Ricci and Reiner Schulz and Nolwenn Jouvenet and Olivier Moncorgé and Caroline Goujon}, url = {https://pubmed.ncbi.nlm.nih.gov/36161446/}, doi = {10.15252/embr.202154061}, issn = {1469-3178}, year = {2022}, date = {2022-09-01}, urldate = {2022-09-01}, journal = {EMBO Rep}, pages = {e54061}, abstract = {Genome-wide screens are powerful approaches to unravel regulators of viral infections. Here, a CRISPR screen identifies the RNA helicase DDX42 as an intrinsic antiviral inhibitor of HIV-1. Depletion of endogenous DDX42 increases HIV-1 DNA accumulation and infection in cell lines and primary cells. DDX42 overexpression inhibits HIV-1 infection, whereas expression of a dominant-negative mutant increases infection. Importantly, DDX42 also restricts LINE-1 retrotransposition and infection with other retroviruses and positive-strand RNA viruses, including CHIKV and SARS-CoV-2. However, DDX42 does not impact the replication of several negative-strand RNA viruses, arguing against an unspecific effect on target cells, which is confirmed by RNA-seq analysis. Proximity ligation assays show DDX42 in the vicinity of viral elements, and cross-linking RNA immunoprecipitation confirms a specific interaction of DDX42 with RNAs from sensitive viruses. Moreover, recombinant DDX42 inhibits HIV-1 reverse transcription in vitro. Together, our data strongly suggest a direct mode of action of DDX42 on viral ribonucleoprotein complexes. Our results identify DDX42 as an intrinsic viral inhibitor, opening new perspectives to target the life cycle of numerous RNA viruses.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36153338, title = {High-resolution silkworm pan-genome provides genetic insights into artificial selection and ecological adaptation}, author = {Xiaoling Tong and Min-Jin Han and Kunpeng Lu and Shuaishuai Tai and Shubo Liang and Yucheng Liu and Hai Hu and Jianghong Shen and Anxing Long and Chengyu Zhan and Xin Ding and Shuo Liu and Qiang Gao and Bili Zhang and Linli Zhou and Duan Tan and Yajie Yuan and Nangkuo Guo and Yan-Hong Li and Zhangyan Wu and Lulu Liu and Chunlin Li and Yaru Lu and Tingting Gai and Yahui Zhang and Renkui Yang and Heying Qian and Yanqun Liu and Jiangwen Luo and Lu Zheng and Jinghou Lou and Yunwu Peng and Weidong Zuo and Jiangbo Song and Songzhen He and Songyuan Wu and Yunlong Zou and Lei Zhou and Lan Cheng and Yuxia Tang and Guotao Cheng and Lianwei Yuan and Weiming He and Jiabao Xu and Tao Fu and Yang Xiao and Ting Lei and Anying Xu and Ye Yin and Jian Wang and Antónia Monteiro and E Westhof and Cheng Lu and Zhixi Tian and Wen Wang and Zhonghuai Xiang and Fangyin Dai}, doi = {10.1038/s41467-022-33366-x}, issn = {2041-1723}, year = {2022}, date = {2022-09-01}, urldate = {2022-09-01}, journal = {Nat Commun}, volume = {13}, number = {1}, pages = {5619}, abstract = {The silkworm Bombyx mori is an important economic insect for producing silk, the "queen of fabrics". The currently available genomes limit the understanding of its genetic diversity and the discovery of valuable alleles for breeding. Here, we deeply re-sequence 1,078 silkworms and assemble long-read genomes for 545 representatives. We construct a high-resolution pan-genome dataset representing almost the entire genomic content in the silkworm. We find that the silkworm population harbors a high density of genomic variants and identify 7308 new genes, 4260 (22%) core genes, and 3,432,266 non-redundant structure variations (SVs). We reveal hundreds of genes and SVs that may contribute to the artificial selection (domestication and breeding) of silkworm. Further, we focus on four genes responsible, respectively, for two economic (silk yield and silk fineness) and two ecologically adaptive traits (egg diapause and aposematic coloration). Taken together, our population-scale genomic resources will promote functional genomics studies and breeding improvement for silkworm.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{pmid36139107, title = {eIF3 Interacts with Selenoprotein mRNAs}, author = {Hassan Hayek and Gilbert Eriani and Christine Allmang}, doi = {10.3390/biom12091268}, issn = {2218-273X}, year = {2022}, date = {2022-09-01}, urldate = {2022-09-01}, journal = {Biomolecules}, volume = {12}, number = {9}, abstract = {The synthesis of selenoproteins requires the co-translational recoding of an in-frame UGASec codon. Interactions between the Selenocysteine Insertion Sequence (SECIS) and the SECIS binding protein 2 (SBP2) in the 3'untranslated region (3'UTR) of selenoprotein mRNAs enable the recruitment of the selenocysteine insertion machinery. Several selenoprotein mRNAs undergo unusual cap hypermethylation and are not recognized by the translation initiation factor 4E (eIF4E) but nevertheless translated. The human eukaryotic translation initiation factor 3 (eIF3), composed of 13 subunits (a-m), can selectively recruit several cellular mRNAs and plays roles in specialized translation initiation. Here, we analyzed the ability of eIF3 to interact with selenoprotein mRNAs. By combining ribonucleoprotein immunoprecipitation (RNP IP) in vivo and in vitro with cross-linking experiments, we found interactions between eIF3 and a subgroup of selenoprotein mRNAs. We showed that eIF3 preferentially interacts with hypermethylated capped selenoprotein mRNAs rather than mG-capped mRNAs. We identified direct contacts between GPx1 mRNA and eIF3 c, d, and e subunits and showed the existence of common interaction patterns for all hypermethylated capped selenoprotein mRNAs. Differential interactions of eIF3 with selenoprotein mRNAs may trigger specific translation pathways independent of eIF4E. eIF3 could represent a new player in the translation regulation and hierarchy of selenoprotein expression.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36167663, title = {CARD9 in neutrophils protects from colitis and controls mitochondrial metabolism and cell survival}, author = {Camille Danne and Chloé Michaudel and Jurate Skerniskyte and Julien Planchais and Aurélie Magniez and Allison Agus and Marie-Laure Michel and Bruno Lamas and Gregory Da Costa and Madeleine Spatz and Cyriane Oeuvray and Chloé Galbert and Maxime Poirier and Yazhou Wang and Alexia Lapière and Nathalie Rolhion and Tatiana Ledent and Cédric Pionneau and Solenne Chardonnet and Floriant Bellvert and Edern Cahoreau and Amandine Rocher and Rafael Rose Arguello and Carole Peyssonnaux and Sabine Louis and Mathias L Richard and Philippe Langella and Jamel El-Benna and Benoit Marteyn and Harry Sokol}, doi = {10.1136/gutjnl-2022-326917}, issn = {1468-3288}, year = {2022}, date = {2022-09-01}, urldate = {2022-09-01}, journal = {Gut}, volume = {72}, issue = {6}, pages = {1081-1092}, abstract = {OBJECTIVES: Inflammatory bowel disease (IBD) results from a combination of genetic predisposition, dysbiosis of the gut microbiota and environmental factors, leading to alterations in the gastrointestinal immune response and chronic inflammation. Caspase recruitment domain 9 (), one of the IBD susceptibility genes, has been shown to protect against intestinal inflammation and fungal infection. However, the cell types and mechanisms involved in the CARD9 protective role against inflammation remain unknown. DESIGN: We used dextran sulfate sodium (DSS)-induced and adoptive transfer colitis models in total and conditional CARD9 knock-out mice to uncover which cell types play a role in the CARD9 protective phenotype. The impact of deletion on neutrophil function was assessed by an in vivo model of fungal infection and various functional assays, including endpoint dilution assay, apoptosis assay by flow cytometry, proteomics and real-time bioenergetic profile analysis (Seahorse). RESULTS: Lymphocytes are not intrinsically involved in the CARD9 protective role against colitis. CARD9 expression in neutrophils, but not in epithelial or CD11c+cells, protects against DSS-induced colitis. In the absence of CARD9, mitochondrial dysfunction increases mitochondrial reactive oxygen species production leading to the premature death of neutrophilsthrough apoptosis, especially in oxidative environment. The decreased functional neutrophils in tissues might explain the impaired containment of fungi and increased susceptibility to intestinal inflammation. CONCLUSION: These results provide new insight into the role of CARD9 in neutrophil mitochondrial function and its involvement in intestinal inflammation, paving the way for new therapeutic strategies targeting neutrophils.}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36108089, title = {A tRNA-derived fragment present in E. coli OMVs regulates host cell gene expression and proliferation}, author = {Idrissa Diallo and Jeffrey Ho and Marine Lambert and Abderrahim Benmoussa and Zeinab Husseini and David Lalaouna and Eric Massé and Patrick Provost}, doi = {10.1371/journal.ppat.1010827}, issn = {1553-7374}, year = {2022}, date = {2022-09-01}, urldate = {2022-09-01}, journal = {PLoS Pathog}, volume = {18}, number = {9}, pages = {e1010827}, abstract = {RNA-sequencing has led to a spectacular increase in the repertoire of bacterial sRNAs and improved our understanding of their biological functions. Bacterial sRNAs have also been found in outer membrane vesicles (OMVs), raising questions about their potential involvement in bacteria-host relationship, but few studies have documented this issue. Recent RNA-Sequencing analyses of bacterial RNA unveiled the existence of abundant very small RNAs (vsRNAs) shorter than 16 nt. These especially include tRNA fragments (tRFs) that are selectively loaded in OMVs and are predicted to target host mRNAs. Here, in Escherichia coli (E. coli), we report the existence of an abundant vsRNA, Ile-tRF-5X, which is selectively modulated by environmental stress, while remaining unaffected by inhibition of transcription or translation. Ile-tRF-5X is released through OMVs and can be transferred to human HCT116 cells, where it promoted MAP3K4 expression. Our findings provide a novel perspective and paradigm on the existing symbiosis between bacteria and human cells.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid36140409, title = {Dynamic Regulation of NF-κB Response in Innate Immunity: The Case of the IMD Pathway in Drosophila}, author = {Alexandre Cammarata-Mouchtouris and Adrian Acker and Akira Goto and Di Chen and Nicolas Matt and Vincent Leclerc}, doi = {10.3390/biomedicines10092304}, issn = {2227-9059}, year = {2022}, date = {2022-09-01}, urldate = {2022-09-01}, journal = {Biomedicines}, volume = {10}, number = {9}, abstract = {Metazoans have developed strategies to protect themselves from pathogenic attack. These preserved mechanisms constitute the immune system, composed of innate and adaptive responses. Among the two kinds, the innate immune system involves the activation of a fast response. NF-κB signaling pathways are activated during infections and lead to the expression of timely-controlled immune response genes. However, activation of NF-κB pathways can be deleterious when uncontrolled. Their regulation is necessary to prevent the development of inflammatory diseases or cancers. The similarity of the NF-κB pathways mediating immune mechanisms in insects and mammals makes a suitable model for studying the innate immune response and learning general mechanisms that are also relevant for humans. In this review, we summarize what is known about the dynamic regulation of the central NF-κB-pathways and go into detail on the molecular level of the IMD pathway. We report on the role of the nuclear protein Akirin in the regulation of the NF-κB Relish immune response. The use of the model allows the understanding of the fine-tuned regulation of this central NF-κB pathway.}, keywords = {M3i, matt}, pubstate = {published}, tppubtype = {article} } @article{pmid35993811, title = {The influenza A virus genome packaging network - complex, flexible and yet unsolved}, author = {Celia Jakob and Rithu Paul-Stansilaus and Martin Schwemmle and Roland Marquet and Hardin Bolte}, doi = {10.1093/nar/gkac688}, issn = {1362-4962}, year = {2022}, date = {2022-08-01}, urldate = {2022-08-01}, journal = {Nucleic Acids Res}, volume = {50}, issue = {16}, pages = {9023-9038}, abstract = {The genome of influenza A virus (IAV) consists of eight unique viral RNA segments. This genome organization allows genetic reassortment between co-infecting IAV strains, whereby new IAVs with altered genome segment compositions emerge. While it is known that reassortment events can create pandemic IAVs, it remains impossible to anticipate reassortment outcomes with pandemic prospects. Recent research indicates that reassortment is promoted by a viral genome packaging mechanism that delivers the eight genome segments as a supramolecular complex into the virus particle. This finding holds promise of predicting pandemic IAVs by understanding the intermolecular interactions governing this genome packaging mechanism. Here, we critically review the prevailing mechanistic model postulating that IAV genome packaging is orchestrated by a network of intersegmental RNA-RNA interactions. Although we find supporting evidence, including segment-specific packaging signals and experimentally proposed RNA-RNA interaction networks, this mechanistic model remains debatable due to a current shortage of functionally validated intersegmental RNA-RNA interactions. We speculate that identifying such functional intersegmental RNA-RNA contacts might be hampered by limitations of the utilized probing techniques and the inherent complexity of the genome packaging mechanism. Nevertheless, we anticipate that improved probing strategies combined with a mutagenesis-based validation could facilitate their discovery.}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{dedaMarques2022, title = {Invading viral DNA triggers dsRNA synthesis by RNA polymerase II to activate antiviral RNA interference in Drosophila}, author = {Isaque J.S. de Faria and Eric R.G.R. Aguiar and Roenick P. Olmo and Juliana Alves da Silva and Laurent Daeffler and Richard W. Carthew and Jean-Luc Imler and Joao T. Marques}, doi = {10.1016/j.celrep.2022.110976}, year = {2022}, date = {2022-06-21}, urldate = {2022-06-21}, journal = {Cell Reports}, volume = {39}, pages = {110976}, abstract = {dsRNA sensing triggers antiviral responses against RNA and DNA viruses in diverse eukaryotes. In Drosophila, Invertebrate iridescent virus 6 (IIV-6), a large DNA virus, triggers production of small interfering RNAs (siRNAs) by the dsRNA sensor Dicer-2. Here, we show that host RNA polymerase II (RNAPII) bidirec- tionally transcribes specific AT-rich regions of the IIV-6 DNA genome to generate dsRNA. Both replicative and naked IIV-6 genomes trigger production of dsRNA in Drosophila cells, implying direct sensing of invading DNA. Loquacious-PD, a Dicer-2 co-factor essential for the biogenesis of endogenous siRNAs, is dispensable for processing of IIV-6-derived dsRNAs, which suggests that they are distinct. Consistent with this finding, inhibition of the RNAPII co-factor P-TEFb affects the synthesis of endogenous, but not virus-derived, dsRNA. Altogether, our results suggest that a non-canonical RNAPII complex recognizes invading viral DNA to synthesize virus-derived dsRNA, which activates the antiviral siRNA pathway in Drosophila.}, keywords = {antiviral, Drosophila, dsRNA, imler, M3i, Marques, protocol, RNA Interference}, pubstate = {published}, tppubtype = {article} } @article{pmid35687141, title = {Towards a comprehensive understanding of RNA deamination: synthesis and properties of xanthosine-modified RNA}, author = {Stefan Mair and Kevin Erharter and Eva Renard and Karl Brillet and Melanie Brunner and Alexandra Lusser and Christoph Kreutz and Eric Ennifar and Ronald Micura}, doi = {10.1093/nar/gkac477}, issn = {1362-4962}, year = {2022}, date = {2022-06-01}, urldate = {2022-06-01}, journal = {Nucleic Acids Res}, volume = {50}, issue = {11}, pages = {6038-6051}, abstract = {Nucleobase deamination, such as A-to-I editing, represents an important posttranscriptional modification of RNA. When deamination affects guanosines, a xanthosine (X) containing RNA is generated. However, the biological significance and chemical consequences on RNA are poorly understood. We present a comprehensive study on the preparation and biophysical properties of X-modified RNA. Thermodynamic analyses revealed that base pairing strength is reduced to a level similar to that observed for a G•U replacement. Applying NMR spectroscopy and X-ray crystallography, we demonstrate that X can form distinct wobble geometries with uridine depending on the sequence context. In contrast, X pairing with cytidine occurs either through wobble geometry involving protonated C or in Watson-Crick-like arrangement. This indicates that the different pairing modes are of comparable stability separated by low energetic barriers for switching. Furthermore, we demonstrate that the flexible pairing properties directly affect the recognition of X-modified RNA by reverse transcription enzymes. Primer extension assays and PCR-based sequencing analysis reveal that X is preferentially read as G or A and that the ratio depends on the type of reverse transcriptase. Taken together, our results elucidate important properties of X-modified RNA paving the way for future studies on its biological significance.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid35702737, title = {Post-transcriptional regulation of polycistronic microRNAs}, author = {Monika Vilimova and Sébastien Pfeffer}, doi = {10.1002/wrna.1749}, issn = {1757-7012}, year = {2022}, date = {2022-06-01}, urldate = {2022-06-01}, journal = {Wiley Interdiscip Rev RNA}, volume = {14}, issue = {2}, pages = {e1749}, abstract = {An important proportion of microRNA (miRNA) genes tend to lie close to each other within animal genomes. Such genomic organization is generally referred to as miRNA clusters. Even though many miRNA clusters have been greatly studied, most attention has been usually focused on functional impacts of clustered miRNA co-expression. However, there is also another compelling aspect about these miRNA clusters, their polycistronic nature. Being transcribed on a single RNA precursor, polycistronic miRNAs benefit from common transcriptional regulation allowing their coordinated expression. And yet, numerous reports have revealed striking discrepancies in the accumulation of mature miRNAs produced from the same cluster. Indeed, the larger polycistronic transcripts can act as platforms providing unforeseen post-transcriptional regulatory mechanisms controlling individual miRNA processing, thus leading to differential miRNA expression, and sometimes even challenging the general assumption that polycistronic miRNAs are co-expressed. In this review, we aim to address the current knowledge about how miRNA polycistrons are post-transcriptionally regulated. In particular, we will focus on the mechanisms occurring at the level of the primary transcript, which are highly relevant for individual miRNA processing and as such have a direct repercussion on miRNA function within the cell. This article is categorized under: RNA Processing > Processing of Small RNAs Regulatory RNAs/RNAi/Riboswitches > Biogenesis of Effector Small RNAs RNA Interactions with Proteins and Other Molecules > RNA-Protein Complexes.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid35732654, title = {RNase III CLASH in MRSA uncovers sRNA regulatory networks coupling metabolism to toxin expression}, author = {Stuart W McKellar and Ivayla Ivanova and Pedro Arede and Rachel L Zapf and Noémie Mercier and Liang-Cui Chu and Daniel G Mediati and Amy C Pickering and Paul Briaud and Robert G Foster and Grzegorz Kudla and J Ross Fitzgerald and Isabelle Caldelari and Ronan K Carroll and Jai J Tree and Sander Granneman}, doi = {10.1038/s41467-022-31173-y}, issn = {2041-1723}, year = {2022}, date = {2022-06-01}, urldate = {2022-06-01}, journal = {Nat Commun}, volume = {13}, number = {1}, pages = {3560}, abstract = {Methicillin-resistant Staphylococcus aureus (MRSA) is a bacterial pathogen responsible for significant human morbidity and mortality. Post-transcriptional regulation by small RNAs (sRNAs) has emerged as an important mechanism for controlling virulence. However, the functionality of the majority of sRNAs during infection is unknown. To address this, we performed UV cross-linking, ligation, and sequencing of hybrids (CLASH) in MRSA to identify sRNA-RNA interactions under conditions that mimic the host environment. Using a double-stranded endoribonuclease III as bait, we uncovered hundreds of novel sRNA-RNA pairs. Strikingly, our results suggest that the production of small membrane-permeabilizing toxins is under extensive sRNA-mediated regulation and that their expression is intimately connected to metabolism. Additionally, we also uncover an sRNA sponging interaction between RsaE and RsaI. Taken together, we present a comprehensive analysis of sRNA-target interactions in MRSA and provide details on how these contribute to the control of virulence in response to changes in metabolism.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid35710145b, title = {The Bacillus subtilis open reading frame ysgA encodes the SPOUT methyltransferase RlmP forming 2'-O-methylguanosine at position 2553 in the A-loop of 23S rRNA}, author = {Martine L Roovers and Geoffray Labar and Philippe Wolff and Andre Feller and Dany Van Elder and Romuald Soin and Cyril Gueydan and Veronique Kruys and Louis Droogmans}, doi = {10.1261/rna.079131.122}, issn = {1469-9001}, year = {2022}, date = {2022-06-01}, urldate = {2022-06-01}, journal = {RNA}, volume = {28}, issue = {9}, pages = {1185-1196}, abstract = {A previous bioinformatic analysis predicted that the ysgA open reading frame of Bacillus subtilis encodes an RNA methyltransferase of the SPOUT superfamily. Here we show that YsgA is the 2'-O-methyltransferase that targets position G2553 (Escherichia coli numbering) of the A-loop of 23S rRNA. This was shown by a combination of biochemical and mass spectrometry approaches using both rRNA extracted from B. subtilis wild-type or ΔysgA cells and in vitro synthesized rRNA. When the target G2553 is mutated, YsgA is able to methylate the ribose of adenosine. However it cannot methylate cytidine nor uridine. The enzyme modifies free 23S rRNA but not the fully assembled ribosome nor the 50S subunit, suggesting that the modification occurs early during ribosome biogenesis. Nevertheless ribosome subunits assembly is unaffected in a B. subtilis ΔysgA mutant strain. The crystal structure of the recombinant YsgA protein, combined with mutagenesis data, outlined in this article highlights a typical SPOUT fold preceded by an L7Ae/L30 (eL8/eL30 in a new nomenclature) N-terminal domain.}, keywords = {ARN-MS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid35500104, title = {Uptake Mechanisms and Regulatory Responses to MECAM- and DOTAM-Based Artificial Siderophores and Their Antibiotic Conjugates in }, author = {Sarah Fritsch and Véronique Gasser and Carsten Peukert and Lukas Pinkert and Lauriane Kuhn and Quentin Perraud and Vincent Normant and Mark Brönstrup and Isabelle J Schalk}, doi = {10.1021/acsinfecdis.2c00049}, issn = {2373-8227}, year = {2022}, date = {2022-06-01}, journal = {ACS Infect Dis}, volume = {8}, number = {6}, pages = {1134--1146}, abstract = {The development of new antibiotics against Gram-negative bacteria has to deal with the low permeability of the outer membrane. This obstacle can be overcome by utilizing siderophore-dependent iron uptake pathways as entrance routes for antibiotic uptake. Iron-chelating siderophores are actively imported by bacteria, and their conjugation to antibiotics allows smuggling the latter into bacterial cells. Synthetic siderophore mimetics based on MECAM (1,3,5-,',″-tris-(2,3-dihydroxybenzoyl)-triaminomethylbenzene) and DOTAM (1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane) cores, both chelating iron via catechol groups, have been recently applied as versatile carriers of functional cargo. In the present study, we show that MECAM and the MECAM-ampicillin conjugate transport iron into cells via the catechol-type outer membrane transporters PfeA and PirA and DOTAM solely via PirA. Differential proteomics and quantitative real-time polymerase chain reaction (qRT-PCR) showed that MECAM import induced the expression of , whereas led to an increase in the expression of and , a gene conferring ampicillin resistance. The presence of DOTAM did not induce the expression of but upregulated the expression of two zinc transporters ( and ), pointing out that bacteria become zinc starved in the presence of this compound. Iron uptake experiments with radioactive Fe demonstrated that import of this nutrient by MECAM and DOTAM was as efficient as with the natural siderophore enterobactin. The study provides a functional validation for DOTAM- and MECAM-based artificial siderophore mimetics as vehicles for the delivery of cargo into Gram-negative bacteria.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid35429693, title = {Novel role of UHRF1 in the epigenetic repression of the latent HIV-1}, author = {Roxane Verdikt and Maryam Bendoumou and Sophie Bouchat and Lorena Nestola and Alexander O Pasternak and Gilles Darcis and Véronique Avettand-Fenoel and Caroline Vanhulle and Amina Aït-Ammar and Marion Santangelo and Estelle Plant and Valentin Le Douce and Nadège Delacourt and Aurelija Cicilionytė and Coca Necsoi and Francis Corazza and Caroline Pereira Bittencourt Passaes and Christian Schwartz and Martin Bizet and François Fuks and Asier Sáez-Cirión and Christine Rouzioux and Stéphane De Wit and Ben Berkhout and Virginie Gautier and Olivier Rohr and Carine Van Lint}, doi = {10.1016/j.ebiom.2022.103985}, issn = {2352-3964}, year = {2022}, date = {2022-05-01}, urldate = {2022-05-01}, journal = {EBioMedicine}, volume = {79}, pages = {103985}, abstract = {BACKGROUND: The multiplicity, heterogeneity, and dynamic nature of human immunodeficiency virus type-1 (HIV-1) latency mechanisms are reflected in the current lack of functional cure for HIV-1. Accordingly, all classes of latency-reversing agents (LRAs) have been reported to present variable ex vivo potencies. Here, we investigated the molecular mechanisms underlying the potency variability of one LRA: the DNA methylation inhibitor 5-aza-2'-deoxycytidine (5-AzadC).nnMETHODS: We employed epigenetic interrogation methods (electrophoretic mobility shift assays, chromatin immunoprecipitation, Infinium array) in complementary HIV-1 infection models (latently-infected T-cell line models, primary CD4 T-cell models and ex vivo cultures of PBMCs from HIV individuals). Extracellular staining of cell surface receptors and intracellular metabolic activity were measured in drug-treated cells. HIV-1 expression in reactivation studies was explored by combining the measures of capsid p24 protein, green fluorescence protein signal, intracellular and extracellular viral RNA and viral DNA.nnFINDINGS: We uncovered specific demethylation CpG signatures induced by 5-AzadC in the HIV-1 promoter. By analyzing the binding modalities to these CpG, we revealed the recruitment of the epigenetic integrator Ubiquitin-like with PHD and RING finger domain 1 (UHRF1) to the HIV-1 promoter. We showed that UHRF1 redundantly binds to the HIV-1 promoter with different binding modalities where DNA methylation was either non-essential, essential or enhancing UHRF1 binding. We further demonstrated the role of UHRF1 in the epigenetic repression of the latent viral promoter by a concerted control of DNA and histone methylations.nnINTERPRETATION: A better understanding of the molecular mechanisms of HIV-1 latency allows for the development of innovative antiviral strategies. As a proof-of-concept, we showed that pharmacological inhibition of UHRF1 in ex vivo HIV patient cell cultures resulted in potent viral reactivation from latency. Together, we identify UHRF1 as a novel actor in HIV-1 epigenetic silencing and highlight that it constitutes a new molecular target for HIV-1 cure strategies.nnFUNDING: Funding was provided by the Belgian National Fund for Scientific Research (F.R.S.-FNRS, Belgium), the « Fondation Roi Baudouin », the NEAT (European AIDS Treatment Network) program, the Internationale Brachet Stiftung, ViiV Healthcare, the Télévie, the Walloon Region (« Fonds de Maturation »), « Les Amis des Instituts Pasteur à Bruxelles, asbl », the University of Brussels (Action de Recherche Concertée ULB grant), the Marie Skodowska Curie COFUND action, the European Union's Horizon 2020 research and innovation program under grant agreement No 691119-EU4HIVCURE-H2020-MSCA-RISE-2015, the French Agency for Research on AIDS and Viral Hepatitis (ANRS), the Sidaction and the "Alsace contre le Cancer" Foundation. This work is supported by 1UM1AI164562-01, co-funded by National Heart, Lung and Blood Institute, National Institute of Diabetes and Digestive and Kidney Diseases, National Institute of Neurological Disorders and Stroke, National Institute on Drug Abuse and the National Institute of Allergy and Infectious Diseases.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid35417704, title = {The Arabidopsis F-box protein FBW2 targets AGO1 for degradation to prevent spurious loading of illegitimate small RNA}, author = {Thibaut Hacquard and Marion Clavel and Patricia Baldrich and Esther Lechner and Imma Pérez-Salamó and Mikhail Schepetilnikov and Benoît Derrien and Marieke Dubois and Philippe Hammann and Lauriane Kuhn and Danaé Brun and Nathalie Bouteiller and Nicolas Baumberger and Hervé Vaucheret and Blake C Meyers and Pascal Genschik}, doi = {10.1016/j.celrep.2022.110671}, issn = {2211-1247}, year = {2022}, date = {2022-04-01}, journal = {Cell Rep}, volume = {39}, number = {2}, pages = {110671}, abstract = {RNA silencing is a conserved mechanism in eukaryotes involved in development and defense against viruses. In plants, ARGONAUTE1 (AGO1) protein plays a central role in both microRNA- and small interfering RNA-directed silencing, and its expression is regulated at multiple levels. Here, we report that the F-box protein FBW2 assembles an SCF complex that selectively targets for proteolysis AGO1 when it is unloaded and mutated. Although FBW2 loss of function does not lead to strong growth or developmental defects, it significantly increases RNA-silencing activity. Interestingly, under conditions in which small-RNA accumulation is affected, the failure to degrade AGO1 in fbw2 mutants becomes more deleterious for the plant. Accordingly, the non-degradable AGO1 protein assembles high-molecular-weight complexes and binds illegitimate small RNA, leading to off-target cleavage. Therefore, control of AGO1 homeostasis by FBW2 plays an important role in quality control of RNA silencing.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid34968484, title = {Suicide gene therapy in cancer and HIV-1 infection: An alternative to conventional treatments}, author = {Sepideh Saeb and Jeanne Van Assche and Thomas Loustau and Olivier Rohr and Clémentine Wallet and Christian Schwartz}, doi = {10.1016/j.bcp.2021.114893}, issn = {1873-2968}, year = {2022}, date = {2022-03-01}, journal = {Biochem Pharmacol}, volume = {197}, pages = {114893}, abstract = {Suicide Gene Therapy (SGT) aims to introduce a gene encoding either a toxin or an enzyme making the targeted cell more sensitive to chemotherapy. SGT represents an alternative approach to combat pathologies where conventional treatments fail such as pancreatic cancer or the high-grade glioblastoma which are still desperately lethal. We review the possibility to use SGT to treat these cancers which have shown promising results in vitro and in preclinical trials. However, SGT has so far failed in phase III clinical trials thus further improvements are awaited. We can now take advantages of the many advances made in SGT for treating cancer to combat other pathologies such as HIV-1 infection. In the review we also discuss the feasibility to add SGT to the therapeutic arsenal used to cure HIV-1-infected patients. Indeed, preliminary results suggest that both productive and latently infected cells are targeted by the SGT. In the last section, we address the limitations of this approach and how we might improve it.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{Imler2022, title = {cGAS-like receptor-mediated immunity: the insect perspective}, author = {H Cai and C Meignin and JL Imler}, doi = {10.1016/j.coi.2022.01.005}, year = {2022}, date = {2022-02-08}, urldate = {2022-02-08}, journal = {Current Opinion in Immunology}, volume = {74}, pages = {183-189}, abstract = {The cGAS-STING pathway plays a central role in the detection of DNA in the cytosol of mammalian cells and activation of immunity. Although the early evolutionary origin of this pathway in animals has been noted, its ancestral functions have remained elusive so far. We review here new findings in invertebrates establishing a role in sensing and signaling infection, triggering potent transcriptional responses, in addition to autophagy. Results from flies and moths/butterflies points to the importance of STING signaling in antiviral immunity in insects. The recent characterization of cGAS-like receptors in Drosophila reveals the plasticity of this family of pattern-recognition receptors, able to accommodate ligands different from DNA and to produce cyclic dinucleotides beyond 2′3′-cGAMP.}, keywords = {imler, M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{pmid34791434, title = {A NYN domain protein directly interacts with DECAPPING1 and is required for phyllotactic pattern}, author = {Marlene Schiaffini and Clara Chicois and Aude Pouclet and Tiphaine Chartier and Elodie Ubrig and Anthony Gobert and Hélène Zuber and Jérôme Mutterer and Johana Chicher and Lauriane Kuhn and Philippe Hammann and Dominique Gagliardi and Damien Garcia}, doi = {10.1093/plphys/kiab529}, issn = {1532-2548}, year = {2022}, date = {2022-02-01}, urldate = {2022-02-01}, journal = {Plant Physiol}, volume = {188}, number = {2}, pages = {1174--1188}, abstract = {In eukaryotes, general mRNA decay requires the decapping complex. The activity of this complex depends on its catalytic subunit, DECAPPING2 (DCP2), and its interaction with decapping enhancers, including its main partner DECAPPING1 (DCP1). Here, we report that in Arabidopsis thaliana, DCP1 also interacts with a NYN domain endoribonuclease, hence named DCP1-ASSOCIATED NYN ENDORIBONUCLEASE 1 (DNE1). Interestingly, we found DNE1 predominantly associated with DCP1, but not with DCP2, and reciprocally, suggesting the existence of two distinct protein complexes. We also showed that the catalytic residues of DNE1 are required to repress the expression of mRNAs in planta upon transient expression. The overexpression of DNE1 in transgenic lines led to growth defects and a similar gene deregulation signature than inactivation of the decapping complex. Finally, the combination of dne1 and dcp2 mutations revealed a functional redundancy between DNE1 and DCP2 in controlling phyllotactic pattern formation. Our work identifies DNE1, a hitherto unknown DCP1 protein partner highly conserved in the plant kingdom and identifies its importance for developmental robustness.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid34664350, title = {A role for PchHI as the ABC transporter in iron acquisition by the siderophore pyochelin in Pseudomonas aeruginosa}, author = {Béatrice Roche and Mariel A Garcia-Rivera and Vincent Normant and Lauriane Kuhn and Philippe Hammann and Mark Brönstrup and Gaëtan L A Mislin and Isabelle J Schalk}, doi = {10.1111/1462-2920.15811}, issn = {1462-2920}, year = {2022}, date = {2022-02-01}, journal = {Environ Microbiol}, volume = {24}, number = {2}, pages = {866--877}, abstract = {Iron is an essential nutrient for bacterial growth but poorly bioavailable. Bacteria scavenge ferric iron by synthesizing and secreting siderophores, small compounds with a high affinity for iron. Pyochelin (PCH) is one of the two siderophores produced by the opportunistic pathogen Pseudomonas aeruginosa. After capturing a ferric iron molecule, PCH-Fe is imported back into bacteria first by the outer membrane transporter FptA and then by the inner membrane permease FptX. Here, using molecular biology, Fe uptake assays, and LC-MS/MS quantification, we first find a role for PchHI as the heterodimeric ABC transporter involved in the siderophore-free iron uptake into the bacterial cytoplasm. We also provide the first evidence that PCH is able to reach the bacterial periplasm and cytoplasm when both FptA and FptX are expressed. Finally, we detected an interaction between PchH and FptX, linking the ABC transporter PchHI with the inner permease FptX in the PCH-Fe uptake pathway. These results pave the way for a better understanding of the PCH siderophore pathway, giving future directions to tackle P. aeruginosa infections.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid33350053, title = {Opportunistic use of catecholamine neurotransmitters as siderophores to access iron by Pseudomonas aeruginosa}, author = {Quentin Perraud and Lauriane Kuhn and Sarah Fritsch and Gwenaëlle Graulier and Véronique Gasser and Vincent Normant and Philippe Hammann and Isabelle J Schalk}, doi = {10.1111/1462-2920.15372}, issn = {1462-2920}, year = {2022}, date = {2022-02-01}, journal = {Environ Microbiol}, volume = {24}, number = {2}, pages = {878--893}, abstract = {Iron is an essential nutrient for bacterial growth and the cause of a fierce battle between the pathogen and host during infection. Bacteria have developed several strategies to access iron from the host, the most common being the production of siderophores, small iron-chelating molecules secreted into the bacterial environment. The opportunist pathogen Pseudomonas aeruginosa produces two siderophores, pyoverdine and pyochelin, and is also able to use a wide panoply of xenosiderophores, siderophores produced by other microorganisms. Here, we demonstrate that catecholamine neurotransmitters (dopamine, l-DOPA, epinephrine and norepinephrine) are able to chelate iron and efficiently bring iron into P. aeruginosa cells via TonB-dependent transporters (TBDTs). Bacterial growth assays under strong iron-restricted conditions and with numerous mutants showed that the TBDTs involved are PiuA and PirA. PiuA exhibited more pronounced specificity for dopamine uptake than for norepinephrine, epinephrine and l-DOPA, whereas PirA specificity appeared to be higher for l-DOPA and norepinephrine. Proteomic and qRT-PCR approaches showed pirA transcription and expression to be induced in the presence of all four catecholamines. Finally, the oxidative properties of catecholamines enable them to reduce iron, and we observed ferrous iron uptake via the FeoABC system in the presence of l-DOPA.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {The 3'UTR-derived sRNA RsaG coordinates redox homeostasis and metabolism adaptation in response to glucose-6-phosphate uptake in Staphylococcus aureus}, author = {E. Desgranges and L. Barrientos and L. Herrgott and S. Marzi and A. Toledo-Arana and K. Moreau and F. Vandenesch and P. Romby and I. Caldelari}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34783400}, doi = {10.1111/mmi.14845}, isbn = {34783400}, year = {2022}, date = {2022-01-01}, urldate = {2021-01-01}, journal = {Mol Microbiol}, abstract = {Staphylococcus aureus RsaG is a 3' untranslated region (3'UTR) derived sRNA from the conserved uhpT gene encoding a glucose-6-phosphate (G6P) transporter expressed in response to extracellular G6P. The transcript uhpT-RsaG undergoes degradation from 5' to 3' end by the action of the exoribonucleases J1/J2, which are blocked by a stable hairpin structure at the 5' end of RsaG, leading to its accumulation. RsaG together with uhpT are induced when bacteria are internalized into host cells or in presence of mucus-secreting cells. Using MS2 affinity purification coupled with RNA sequencing, several RNAs were identified as targets including mRNAs encoding the transcriptional factors Rex, CcpA, SarA and the sRNA RsaI. Our data suggested that RsaG contributes to the control of redox homeostasis and adjusts metabolism to changing environmental conditions. RsaG uses different molecular mechanisms to stabilize, to degrade, or to repress translation of its mRNA targets. While RsaG is conserved only in closely related species, the uhpT 3'UTR of the ape pathogen S. simiae harbors a sRNA, whose sequence is highly different, and which does not respond to G6P levels. Our results hypothesized that the 3'UTRs from UhpT transporter encoding mRNAs could have rapidly evolved to enable adaptation to host niches.}, note = {1365-2958 (Electronic) 0950-382X (Linking) Journal Article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {CD169(+) macrophages in lymph node and spleen critically depend on dual RANK and LTbetaR signaling}, author = {A. Camara and A. C. Lavanant and J. Abe and H. L. Desforges and Y. O. Alexandre and E. Girardi and Z. Igamberdieva and K. Asano and M. Tanaka and T. Hehlgans and K. Pfeffer and S. Pfeffer and S. N. Mueller and J. V. Stein and C. G. Mueller}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35031565}, isbn = {35031565}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {119}, number = {3}, pages = {e2108540119}, abstract = {CD169(+) macrophages reside in lymph node (LN) and spleen and play an important role in the immune defense against pathogens. As resident macrophages, they are responsive to environmental cues to shape their tissue-specific identity. We have previously shown that LN CD169(+) macrophages require RANKL for formation of their niche and their differentiation. Here, we demonstrate that they are also dependent on direct lymphotoxin beta (LTbeta) receptor (R) signaling. In the absence or the reduced expression of either RANK or LTbetaR, their differentiation is perturbed, generating myeloid cells expressing SIGN-R1 in LNs. Conditions of combined haploinsufficiencies of RANK and LTbetaR revealed that both receptors contribute equally to LN CD169(+) macrophage differentiation. In the spleen, the Cd169-directed ablation of either receptor results in a selective loss of marginal metallophilic macrophages (MMMs). Using a RANKL reporter mouse, we identify splenic marginal zone stromal cells as a source of RANKL and demonstrate that it participates in MMM differentiation. The loss of MMMs had no effect on the splenic B cell compartments but compromised viral capture and the expansion of virus-specific CD8(+) T cells. Taken together, the data provide evidence that CD169(+) macrophage differentiation in LN and spleen requires dual signals from LTbetaR and RANK with implications for the immune response.}, note = {1091-6490 (Electronic) 0027-8424 (Linking) Journal Article}, keywords = {PFEFFER, Team-Mueller, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Tau mRNA Metabolism in Neurodegenerative Diseases: A Tangle Journey}, author = {P. J. Da Costa and M. Hamdane and L. Buee and F. Martin}, url = {https://www.mdpi.com/2227-9059/10/2/241/htm}, doi = {10.3390/biomedicines10020241}, year = {2022}, date = {2022-01-01}, journal = {Biomedicines}, volume = {10}, number = {2}, pages = {241}, abstract = {Tau proteins are known to be mainly involved in regulation of microtubule dynamics. Besides this function, which is critical for axonal transport and signal transduction, tau proteins also have other roles in neurons. Moreover, tau proteins are turned into aggregates and consequently trigger many neurodegenerative diseases termed tauopathies, of which Alzheimerメs disease (AD) is the figurehead. Such pathological aggregation processes are critical for the onset of these diseases. Among the various causes of tau protein pathogenicity, abnormal tau mRNA metabolism, expression and dysregulation of tau post-translational modifications are critical steps. Moreover, the relevance of tau function to general mRNA metabolism has been highlighted recently in tauopathies. In this review, we mainly focus on how mRNA metabolism impacts the onset and development of tauopathies. Thus, we intend to portray how mRNA metabolism of, or mediated by, tau is associated with neurodegenerative diseases.}, keywords = {ERIANI, mRNA metabolism, Neurodegenerative Diseases, tau protein, Translation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Structural maturation of the HIV-1 RNA 5' untranslated region by Pr55(Gag) and its maturation products}, author = {O. Gilmer and E. Mailler and J. C. Paillart and A. Mouhand and C. Tisne and J. Mak and R. P. Smyth and R. Marquet and V. Vivet-Boudou}, url = {https://www.tandfonline.com/doi/full/10.1080/15476286.2021.2021677}, isbn = {35067194}, year = {2022}, date = {2022-01-01}, journal = {RNA Biol}, volume = {19}, number = {1}, pages = {191-205}, abstract = {Maturation of the HIV-1 viral particles shortly after budding is required for infectivity. During this process, the Pr55(Gag) precursor undergoes a cascade of proteolytic cleavages, and whilst the structural rearrangements of the viral proteins are well understood, the concomitant maturation of the genomic RNA (gRNA) structure is unexplored, despite evidence that it is required for infectivity. To get insight into this process, we systematically analysed the interactions between Pr55(Gag) or its maturation products (NCp15, NCp9 and NCp7) and the 5' gRNA region and their structural consequences, in vitro. We show that Pr55(Gag) and its maturation products mostly bind at different RNA sites and with different contributions of their two zinc knuckle domains. Importantly, these proteins have different transient and permanent effects on the RNA structure, the late NCp9 and NCp7 inducing dramatic structural rearrangements. Altogether, our results reveal the distinct contributions of the different Pr55(Gag) maturation products on the gRNA structural maturation.}, note = {1555-8584 (Electronic) 1547-6286 (Linking) Journal Article}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {A case of QARS1 associated epileptic encephalopathy and review of epilepsy in aminoacyl-tRNA synthetase disorders}, author = {D. L. Chan and J. Rudinger-Thirion and M. Frugier and L. G. Riley and G. Ho and K. Kothur and S. S. Mohammad}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34774383}, doi = {10.1016/j.braindev.2021.10.009}, isbn = {34774383}, year = {2022}, date = {2022-01-01}, journal = {Brain Dev}, volume = {44}, number = {2}, pages = {142-147}, abstract = {INTRODUCTION: Mutations in QARS1, which encodes human glutaminyl-tRNA synthetase, have been associated with epilepsy, developmental regression, progressive microcephaly and cerebral atrophy. Epilepsy caused by variants in QARS1 is usually drug-resistant and intractable. Childhood onset epilepsy is also reported in various aminoacyl-tRNA synthetase disorders. We describe a case with a milder neurological phenotype than previously reported with QARS1 variants and review the seizure associations with aminoacyl-tRNA synthetase disorders. CASE REPORT: The patient is a 4-year-old girl presenting at 6 weeks of age with orofacial dyskinesia and hand stereotypies. She developed focal seizures at 7 months of age. Serial electroencephalograms showed shifting focality. Her seizures were controlled after introduction of carbamazepine. Progress MRI showed very mild cortical volume loss without myelination abnormalities or cerebellar atrophy. She was found to have novel compound heterozygous variants in QARS1 (NM_005051.2): c.[1132C > T];[1574G > A], p.[(Arg378Cys)];[(Arg525Gln)] originally classified as "variants of uncertain significance" and later upgraded to "likely pathogenic" based on functional testing and updated variant database review. Functional testing showed reduced solubility of the corresponding QARS1 mutants in vitro, but only mild two-fold loss in catalytic efficiency with the c.1132C > T variant and no noted change in tRNA(Gln) aminoacylation with the c.1574G > A variant. CONCLUSION: We describe two QARS1 variants associated with overall conserved tRNA aminoacylation activity but characterized by significantly reduced QARS protein solubility, resulting in a milder clinical phenotype. 86% of previous patients reported with QARS1 had epilepsy and 79% were pharmaco-resistant. We also summarise literature regarding epilepsy in aminoacyl-tRNA synthetase disorders, which is also often early onset, severe and drug-refractory.}, note = {1872-7131 (Electronic) 0387-7604 (Linking) Case Reports}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{normant_how_2022, title = {How the Presence of Hemin Affects the Expression of the Different Iron Uptake Pathways in Pseudomonas aeruginosa Cells}, author = {Vincent Normant and Lauriane Kuhn and Mathilde Munier and Philippe Hammann and Gaëtan L. A. Mislin and Isabelle J. Schalk}, doi = {10.1021/acsinfecdis.1c00525}, issn = {2373-8227}, year = {2022}, date = {2022-01-01}, journal = {ACS infectious diseases}, volume = {8}, number = {1}, pages = {183--196}, abstract = {Iron is an essential nutriment for almost all organisms, but this metal is poorly bioavailable. During infection, bacteria access iron from the host by importing either iron or heme. Pseudomonas aeruginosa, a gram-negative pathogen, secretes two siderophores, pyoverdine (PVD) and pyochelin (PCH), to access iron and is also able to use many siderophores produced by other microorganisms (called xenosiderophores). To access heme, P. aeruginosa uses three distinct uptake pathways, named Has, Phu, and Hxu. We previously showed that P. aeruginosa expresses the Has and Phu heme uptake systems and the PVD- and PCH-dependent iron uptake pathways in iron-restricted growth conditions, using proteomic and RT-qPCR approaches. Here, using the same approaches, we show that physiological concentrations of hemin in the bacterial growth medium result in the repression of the expression of the proteins of the PVD- and PCH-dependent iron uptake pathways, leading to less production of these two siderophores. This indicates that the pathogen adapts its phenotype to use hemin as an iron source rather than produce PVD and PCH to access iron. Moreover, the presence of both hemin and a xenosiderophore resulted in (i) the strong induction of the expression of the proteins of the added xenosiderophore uptake pathway, (ii) repression of the PVD- and PCH-dependent iron uptake pathways, and (iii) no effect on the expression levels of the Has, Phu, or Hxu systems, indicating that bacteria use both xenosiderophores and heme to access iron.}, keywords = {Hemin, hemin (heme) uptake, Iron, iron homeostasis, iron uptake, outer membrane transporters, PPSE, proteomics, Pseudomonas aeruginosa, siderophore, Siderophores}, pubstate = {published}, tppubtype = {article} } @article{pmid35012549b, title = {The interactome of CLUH reveals its association to SPAG5 and its co-translational proximity to mitochondrial proteins}, author = {Mickaële Hémono and Alexandre Haller and Johana Chicher and Anne-Marie Duchêne and Richard Patryk Ngondo}, doi = {10.1186/s12915-021-01213-y}, issn = {1741-7007}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {BMC Biol}, volume = {20}, number = {1}, pages = {13}, abstract = {BACKGROUND: Mitochondria require thousands of proteins to fulfill their essential function in energy production and other fundamental biological processes. These proteins are mostly encoded by the nuclear genome, translated in the cytoplasm before being imported into the organelle. RNA binding proteins (RBPs) are central players in the regulation of this process by affecting mRNA translation, stability, or localization. CLUH is an RBP recognizing specifically mRNAs coding for mitochondrial proteins, but its precise molecular function and interacting partners remain undiscovered in mammals. RESULTS: Here we reveal for the first time CLUH interactome in mammalian cells. Using both co-IP and BioID proximity-labeling approaches, we identify novel molecular partners interacting stably or transiently with CLUH in HCT116 cells and mouse embryonic stem cells. We reveal stable RNA-independent interactions of CLUH with itself and with SPAG5 in cytosolic granular structures. More importantly, we uncover an unexpected proximity of CLUH to mitochondrial proteins and their cognate mRNAs in the cytosol. We show that this interaction occurs during the process of active translation and is dependent on CLUH TPR domain. CONCLUSIONS: Overall, through the analysis of CLUH interactome, our study sheds a new light on CLUH molecular function by revealing new partners and by highlighting its link to the translation and subcellular localization of some mRNAs coding for mitochondrial proteins.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {The battle for oxygen during bacterial and fungal infections}, author = {A. C. Andre and M. Laborde and B. S. Marteyn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35131160}, doi = {10.1016/j.tim.2022.01.002}, isbn = {35131160}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Trends Microbiol}, volume = {30}, issue = {7}, pages = {643-653}, abstract = {Bacterial and fungal pathogens face various microenvironmental conditions during infection. In addition to acidosis, nutrient consumption, and hypercapnia, pathogen infections are associated with hypoxia, which is induced by bacterial and fungal respiration during the formation of foci of infection or biofilms. Consequently, the in vivo interaction between host immune cells and pathogens is anticipated to occur mainly under low-oxygen conditions. Various infectious disease models have reported that pathogens benefit from hypoxia, which dampens the oxygen-dependent antimicrobial activities of macrophages and neutrophils, such as the production of reactive oxygen species (ROS). Due to their dual respiration capacity (aerobic and anaerobic) or phenotypical adaptation (e.g., dormancy), pathogens have the capacity to survive and disseminate in the absence of oxygen. In addition, hypoxia modulates various mechanisms of pathogen virulence, promoting the dissemination of pathogens. Further investigations are still required to evaluate the relative importance of oxygen on the capacity of pathogens to invade and colonize host organs and to better understand alternative strategies developed by immune cells to circumvent pathogen dissemination in the absence of oxygen. Addressing this important and fundamental question in various models of infection may direct the development of innovative therapeutic strategies.}, note = {1878-4380 (Electronic) 0966-842X (Linking) Journal Article Review}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Visualization of Retroviral Gag-Genomic RNA Cellular Interactions Leading to Genome Encapsidation and Viral Assembly: An Overview}, author = {S. Bernacchi}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35215917}, doi = {10.3390/v14020324}, isbn = {35215917}, year = {2022}, date = {2022-01-01}, journal = {Viruses}, volume = {14}, number = {2}, abstract = {Retroviruses must selectively recognize their unspliced RNA genome (gRNA) among abundant cellular and spliced viral RNAs to assemble into newly formed viral particles. Retroviral gRNA packaging is governed by Gag precursors that also orchestrate all the aspects of viral assembly. Retroviral life cycles, and especially the HIV-1 one, have been previously extensively analyzed by several methods, most of them based on molecular biology and biochemistry approaches. Despite these efforts, the spatio-temporal mechanisms leading to gRNA packaging and viral assembly are only partially understood. Nevertheless, in these last decades, progress in novel bioimaging microscopic approaches (as FFS, FRAP, TIRF, and wide-field microscopy) have allowed for the tracking of retroviral Gag and gRNA in living cells, thus providing important insights at high spatial and temporal resolution of the events regulating the late phases of the retroviral life cycle. Here, the implementation of these recent bioimaging tools based on highly performing strategies to label fluorescent macromolecules is described. This report also summarizes recent gains in the current understanding of the mechanisms employed by retroviral Gag polyproteins to regulate molecular mechanisms enabling gRNA packaging and the formation of retroviral particles, highlighting variations and similarities among the different retroviruses.}, note = {1999-4915 (Electronic) 1999-4915 (Linking) Journal Article Review Research Support, Non-U.S. Gov't}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Correlated sequence signatures are present within the genomic 5'UTR RNA and NSP1 protein in coronaviruses}, author = {P Sosnowski and A Tidu and G Eriani and E Westhof and F Martin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35236777}, doi = {10.1261/rna.078972.121}, isbn = {35236777}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Rna}, volume = {28}, issue = {5}, pages = {729-741}, abstract = {The 5'UTR part of coronavirus genomes plays key roles in the viral replication cycle and the translation of the viral mRNAs. The first 75-80 nucleotides, also called the leader sequence, are identical for the genomic mRNA and for the subgenomic mRNAs. Recently, it was shown that cooperative actions of a 5'UTR segment and the non-structural protein NSP1 are essential for both the inhibition of host mRNAs and for specific translation of viral mRNAs. Here, sequence analyses of both the 5'UTR RNA segment and the NSP1 protein have been done for several coronaviruses with special attention to the betacoronaviruses. The conclusions are (i) precise specific molecular signatures can be found in both the RNA and the NSP1 protein; (ii) both types of signatures strongly correlate between each other. Indeed, definite sequence motifs in the RNA correlate with sequence motifs in the protein indicating a co-evolution of 5'UTR with NSP1 in betacoronaviruses. Experimental mutational data on 5'UTR and NSP1 from SARS-CoV-2 using cell-free translation extracts support those conclusions and show that the N-terminal half of the NSP1 protein contains conserved key residues that are essential for evasion to the inhibitory effect of NSP1 on translation.}, note = {1469-9001 (Electronic) 1355-8382 (Linking) Journal Article}, keywords = {ERIANI, MARTIN, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {RACK1 Associates with RNA-Binding Proteins Vigilin and SERBP1 to Facilitate Dengue Virus Replication}, author = {A. Brugier and M. L. Hafirrassou and M. Pourcelot and M. Baldaccini and V. Kril and L. Couture and B. M. Kummerer and S. Gallois-Montbrun and L. Bonnet-Madin and P. O. Vidalain and C. Delaugerre and S. Pfeffer and L. Meertens and A. Amara}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35266803}, doi = {10.1128/jvi.01962-21}, isbn = {35266803}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {J Virol}, volume = {96}, issue = {7}, pages = {e0196221}, abstract = {Dengue virus (DENV) is a mosquito-borne flavivirus responsible for dengue disease, a major human health concern for which no effective treatment is available. DENV relies heavily on the host cellular machinery for productive infection. Here, we show that the scaffold protein RACK1, which is part of the DENV replication complex, mediates infection by binding to the 40S ribosomal subunit. Mass spectrometry analysis of RACK1 partners coupled to an RNA interference screen-identified Vigilin and SERBP1 as DENV host-dependency factors. Both are RNA-binding proteins that interact with the DENV genome. Genetic ablation of Vigilin or SERBP1 rendered cells poorly susceptible to DENV, as well as related flaviviruses, by hampering the translation and replication steps. Finally, we established that a Vigilin or SERBP1 mutant lacking RACK1 binding but still interacting with the viral RNA is unable to mediate DENV infection. We propose that RACK1 recruits Vigilin and SERBP1, linking the DENV genome to the translation machinery for efficient infection. IMPORTANCE We recently identified the scaffolding RACK1 protein as an important host-dependency factor for dengue virus (DENV), a positive-stranded RNA virus responsible for the most prevalent mosquito-borne viral disease worldwide. Here, we have performed the first RACK1 interactome in human cells and identified Vigilin and SERBP1 as DENV host-dependency factors. Both are RNA-binding proteins that interact with the DENV RNA to regulate viral replication. Importantly, Vigilin and SERBP1 interact with RACK1 and the DENV viral RNA (vRNA) to mediate viral replication. Overall, our results suggest that RACK1 acts as a binding platform at the surface of the 40S ribosomal subunit to recruit Vigilin and SERBP1, which may therefore function as linkers between the viral RNA and the translation machinery to facilitate infection.}, note = {1098-5514 (Electronic) 0022-538X (Linking) Journal Article}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Eukaryotic tRNA sequences present conserved and amino acid-specific structural signatures}, author = {E Westhof and B Thornlow and P P Chan and T M Lowe}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35380696}, doi = {10.1093/nar/gkac222}, isbn = {35380696}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Nucleic Acids Res}, volume = {50}, issue = {7}, pages = {4100-4112}, abstract = {Metazoan organisms have many tRNA genes responsible for decoding amino acids. The set of all tRNA genes can be grouped in sets of common amino acids and isoacceptor tRNAs that are aminoacylated by corresponding aminoacyl-tRNA synthetases. Analysis of tRNA alignments shows that, despite the high number of tRNA genes, specific tRNA sequence motifs are highly conserved across multicellular eukaryotes. The conservation often extends throughout the isoacceptors and isodecoders with, in some cases, two sets of conserved isodecoders. This study is focused on non-Watson-Crick base pairs in the helical stems, especially GoU pairs. Each of the four helical stems may contain one or more conserved GoU pairs. Some are amino acid specific and could represent identity elements for the cognate aminoacyl tRNA synthetases. Other GoU pairs are found in more than a single amino acid and could be critical for native folding of the tRNAs. Interestingly, some GoU pairs are anticodon-specific, and others are found in phylogenetically-specific clades. Although the distribution of conservation likely reflects a balance between accommodating isotype-specific functions as well as those shared by all tRNAs essential for ribosomal translation, such conservations may indicate the existence of specialized tRNAs for specific translation targets, cellular conditions, or alternative functions.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Viral and cellular translation during SARS-CoV-2 infection}, author = {G. Eriani and F. Martin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35429230}, doi = {10.1002/2211-5463.13413}, isbn = {35429230}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {FEBS Open Bio}, volume = {12}, issue = {9}, pages = {1584-1601}, abstract = {SARS-CoV-2 is a betacoronavirus that emerged in China in December 2019 and which is the causative agent of the Covid-19 pandemic. This enveloped virus contains a large positive-sense single-stranded RNA genome. In this review, we summarize the current knowledge on the molecular mechanisms for the translation of both viral transcripts and cellular messenger RNAs. Non-structural proteins are encoded by the genomic RNA and are produced in the early steps of infection. In contrast, the structural proteins are produced from subgenomic RNAs that are translated in the late phase of the infectious program. Non-structural protein 1 (NSP1) is a key molecule that regulates both viral and cellular translation. In addition, NSP1 interferes with multiple steps of the interferon I pathway and thereby blocks host antiviral responses. Therefore, NSP1 is a drug target of choice for the development of antiviral therapies.}, note = {2211-5463 (Electronic) 2211-5463 (Linking) Journal Article Review}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Fluorogenic RNA-Based Biosensor to Sense the Glycolytic Flux in Mammalian Cells}, author = {I. Geraci and A. Autour and G. Pietruschka and A. Shiian and M. Borisova and C. Mayer and M. Ryckelynck and G. Mayer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35427113}, doi = {10.1021/acschembio.2c00100}, isbn = {35427113}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {ACS Chem Biol}, volume = {17}, issue = {5}, pages = {1164-1173}, abstract = {The visualization of metabolic flux in real time requires sensor molecules that transduce variations of metabolite concentrations into an appropriate output signal. In this regard, fluorogenic RNA-based biosensors are promising molecular tools as they fluoresce only upon binding to another molecule. However, to date no such sensor is available that enables the direct observation of key metabolites in mammalian cells. Toward this direction, we selected and characterized an RNA light-up sensor designed to respond to fructose 1,6-bisphosphate and applied it to probe glycolytic flux variation in mammal cells.}, note = {1554-8937 (Electronic) 1554-8929 (Linking) Journal Article}, keywords = {Labex, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Rational Design of Self-Quenched Rhodamine Dimers as Fluorogenic Aptamer Probes for Live-Cell RNA Imaging}, author = {K. T. Fam and R. Pelletier and F. Bouhedda and M. Ryckelynck and M. Collot and A. S. Klymchenko}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35486532}, doi = {10.1021/acs.analchem.1c04556}, isbn = {35486532}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Anal Chem}, volume = {94}, issue = {18}, pages = {6657-6664}, abstract = {With the growing interest in the understanding of the importance of RNAs in health and disease, detection of RNAs in living cells is of high importance. Fluorogenic dyes that light up specifically selected RNA aptamers constitute an attractive direction in the design of RNA imaging probes. In this work, based on our recently proposed concept of a fluorogenic dimer, we aim to develop a robust molecular tool for intracellular RNA imaging. We rationally designed a fluorogenic self-quenched dimer (orange Gemini, o-Gemini) based on rhodamine and evaluated its capacity to light up its cognate aptamer o-Coral in solution and live cells. We found that the removal of biotin from the dimer slightly improved the fluorogenic response without losing the affinity to the cognate aptamer (o-Coral). On the other hand, replacing sulforhodamine with a carboxyrhodamine produced drastic improvement of the affinity and the turn-on response to o-Coral and, thus, a better limit of detection. In live cells expressing o-Coral-tagged RNAs, the carboxyrhodamine analogue of o-Gemini without a biotin unit displayed a higher signal as well as faster internalization into the cells. We suppose that less hydrophilic carboxyrhodamine compared to sulforhodamine can more readily penetrate through the cell plasma membrane and, together with its higher affinity to o-Coral, provide the observed improvement in the imaging experiments. The promiscuity of the o-Coral RNA aptamer to the fluorogenic dimer allowed us to tune a fluorogen chemical structure and thus drastically improve the fluorescence response of the probe to o-Coral-tagged RNAs.}, note = {1520-6882 (Electronic) 0003-2700 (Linking) Journal Article}, keywords = {Labex, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Growth-Associated Droplet Shrinkage for Bacterial Quantification, Growth Monitoring, and Separation by Ultrahigh-Throughput Microfluidics}, author = {E. Geersens and S. Vuilleumier and M. Ryckelynck}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35449964}, doi = {10.1021/acsomega.2c00248}, isbn = {35449964}, year = {2022}, date = {2022-01-01}, journal = {ACS Omega}, volume = {7}, number = {14}, pages = {12039-12047}, abstract = {Microbiology still relies on en masse cultivation for selection, isolation, and characterization of microorganisms of interest. This constrains the diversity of microbial types and metabolisms that can be investigated in the laboratory also because of intercellular competition during cultivation. Cell individualization by droplet-based microfluidics prior to experimental analysis provides an attractive alternative to access a larger fraction of the microbial biosphere, miniaturizing the required equipment and minimizing reagent use for increased and more efficient analytical throughput. Here, we show that cultivation of a model two-strain bacterial community in droplets significantly reduces representation bias in the grown culture compared to batch cultivation. Further, and based on the droplet shrinkage observed upon cell proliferation, we provide proof-of-concept for a simple strategy that allows absolute quantification of microbial cells in a sample as well as selective recovery of microorganisms of interest for subsequent experimental characterization.}, note = {2470-1343 (Electronic) 2470-1343 (Linking) Journal Article}, keywords = {Labex, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Discovery of two distinct aminoacyl-tRNA synthetase complexes anchored to the Plasmodium surface tRNA import protein}, author = {J. R. Jaramillo Ponce and D. Kapps and C. Paulus and J. Chicher and M. Frugier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35487244}, doi = {0.1016/j.jbc.2022.101987}, isbn = {35487244}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {J Biol Chem}, volume = {298}, issue = {6}, pages = {101987}, abstract = {Aminoacyl-tRNA synthetases (aaRSs) attach amino acids to their cognate transfer RNAs. In eukaryotes, a subset of cytosolic aaRSs is organized into a multi-synthetase complex (MSC), along with specialized scaffolding proteins referred to as aaRS-interacting multifunctional proteins (AIMPs). In Plasmodium, the causative agent of malaria, the tRNA import protein (tRip), is a membrane protein that has been shown to participate in tRNA trafficking; here, we show that tRip also functions as an AIMP. We identified three aaRSs, namely the glutamyl- (ERS), glutaminyl- (QRS), and methionyl- (MRS) tRNA synthetases, which were specifically co-immunoprecipitated with tRip in P. berghei blood stage parasites. All four proteins contain an N-terminal GST-like domain that was demonstrated to be involved in MSC assembly. In contrast to previous studies, further dissection of GST-like interactions identified two exclusive heterotrimeric complexes: the Q-complex (tRip:ERS:QRS) and the M-complex (tRip:ERS:MRS). Gel filtration and light scattering suggest a 2:2:2 stoichiometry for both complexes but with distinct biophysical properties, and mutational analysis further revealed that the GST-like domains of QRS and MRS use different strategies to bind ERS. Taken together our results demonstrate that neither the singular homodimerization of tRip, nor its localization in the parasite plasma membrane prevents the formation of MSCs in Plasmodium. Besides, the extracellular localization of the tRNA-binding module of tRip is compensated by the presence of additional tRNA-binding modules fused to MRS and QRS, providing each MSC with two spatially distinct functions: aminoacylation of intraparasitic tRNAs and binding of extracellular tRNAs. This unique host-pathogen interaction is discussed.}, note = {1083-351X (Electronic) 0021-9258 (Linking) Journal Article}, keywords = {FRUGIER, Labex, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Phytobeneficial traits of rhizobacteria under the control of multiple molecular dialogues}, author = {A. Laveilhe and S. Fochesato and D. Lalaouna and T. Heulin and W. Achouak}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35502577}, doi = {10.1111/1751-7915.14023}, isbn = {35502577}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Microb Biotechnol}, volume = {15}, issue = {7}, pages = {2083-2096}, abstract = {Pseudomonads play crucial roles in plant growth promotion and control of plant diseases. However, under natural conditions, other microorganisms competing for the same nutrient resources in the rhizosphere may exert negative control over their phytobeneficial characteristics. We assessed the expression of phytobeneficial genes involved in biocontrol, biostimulation and iron regulation such as, phlD, hcnA, acdS, and iron-small regulatory RNAs prrF1 and prrF2 in Pseudomonas brassicacearum co-cultivated with three phytopathogenic fungi, and two rhizobacteria in the presence or absence of Brassica napus, and in relation to iron availability. We found that the antifungal activity of P. brassicacearum depends mostly on the production of DAPG and not on HCN whose production is suppressed by fungi. We have also shown that the two-competing bacterial strains modulate the plant growth promotion activity of P. brassicacearum by modifying the expression of phlD, hcnA and acdS according to iron availability. Overall, it allows us to better understand the complexity of the multiple molecular dialogues that take place underground between microorganisms and between plants and its rhizosphere microbiota and to show that synergy in favour of phytobeneficial gene expression may exist between different bacterial species.}, note = {1751-7915 (Electronic) 1751-7915 (Linking) Journal Article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {1-Deazaguanosine-Modified RNA: The Missing Piece for Functional RNA Atomic Mutagenesis}, author = {R. Bereiter and E. Renard and K. Breuker and C. Kreutz and E. Ennifar and R. Micura}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35666572}, doi = {10.1021/jacs.2c01877}, isbn = {35666572}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {J Am Chem Soc}, volume = {144}, issue = {23}, pages = {10344-10352}, abstract = {Atomic mutagenesis is the key to advance our understanding of RNA recognition and RNA catalysis. To this end, deazanucleosides are utilized to evaluate the participation of specific atoms in these processes. One of the remaining challenges is access to RNA-containing 1-deazaguanosine (c(1)G). Here, we present the synthesis of this nucleoside and its phosphoramidite, allowing first time access to c(1)G-modified RNA. Thermodynamic analyses revealed the base pairing parameters for c(1)G-modified RNA. Furthermore, by NMR spectroscopy, a c(1)G-triggered switch of Watson-Crick into Hoogsteen pairing in HIV-2 TAR RNA was identified. Additionally, using X-ray structure analysis, a guanine-phosphate backbone interaction affecting RNA fold stability was characterized, and finally, the critical impact of an active-site guanine in twister ribozyme on the phosphodiester cleavage was revealed. Taken together, our study lays the synthetic basis for c(1)G-modified RNA and demonstrates the power of the completed deazanucleoside toolbox for RNA atomic mutagenesis needed to achieve in-depth understanding of RNA recognition and catalysis.}, note = {1520-5126 (Electronic) 0002-7863 (Linking) Journal Article}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid35761003, title = {Droplet-Based Microfluidic Chip Design, Fabrication, and Use for Ultrahigh-Throughput DNA Analysis and Quantification}, author = {Stéphanie Baudrey and Roger Cubi and Michael Ryckelynck}, doi = {10.1007/978-3-031-04039-9_18}, issn = {0065-2598}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Adv Exp Med Biol}, volume = {1379}, pages = {445--460}, abstract = {DNA is widely used as a biomarker of contamination, infection, or disease, which has stimulated the development of a wide palette of detection and quantification methods. Even though several analytical approaches based on isothermal amplification have been proposed, DNA is still mainly detected and quantified by quantitative PCR (qPCR). However, for some analyses (e.g., in cancer research) qPCR may suffer from limitations arising from competitions between highly similar template DNAs, the presence of inhibitors, or suboptimal primer design. Nevertheless, digitalizing the analysis (i.e., individualizing DNA molecules into compartments prior to amplifying them in situ) allows to address most of these issues. By its capacity to generate and manipulate millions of highly similar picoliter volume water-in-oil droplets, microfluidics offers both the required miniaturization and parallelization capacity, and led to the introduction of digital droplet PCR (ddPCR). This chapter aims at introducing the reader to the basic principles behind ddPCR while also providing the key guidelines to fabricate, set up, and use his/her own ddPCR platform. We further provide procedures to detect and quantify DNA either purified in solution or directly from individualized cells. This approach not only gives access to DNA absolute concentration with unrivaled sensitivity, but it may also be the starting point of more complex in vitro analytical pipelines discussed at the end of the chapter.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid35865816, title = {Battle for Metals: Regulatory RNAs at the Front Line}, author = {Mathilde Charbonnier and Gabriela González-Espinoza and Thomas E Kehl-Fie and David Lalaouna}, doi = {10.3389/fcimb.2022.952948}, issn = {2235-2988}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Front Cell Infect Microbiol}, volume = {12}, pages = {952948}, abstract = {Metal such as iron, zinc, manganese, and nickel are essential elements for bacteria. These nutrients are required in crucial structural and catalytic roles in biological processes, including precursor biosynthesis, DNA replication, transcription, respiration, and oxidative stress responses. While essential, in excess these nutrients can also be toxic. The immune system leverages both of these facets, to limit bacterial proliferation and combat invaders. Metal binding immune proteins reduce the bioavailability of metals at the infection sites starving intruders, while immune cells intoxicate pathogens by providing metals in excess leading to enzyme mismetallation and/or reactive oxygen species generation. In this dynamic metal environment, maintaining metal homeostasis is a critical process that must be precisely coordinated. To achieve this, bacteria utilize diverse metal uptake and efflux systems controlled by metalloregulatory proteins. Recently, small regulatory RNAs (sRNAs) have been revealed to be critical post-transcriptional regulators, working in conjunction with transcription factors to promote rapid adaptation and to fine-tune bacterial adaptation to metal abundance. In this mini review, we discuss the expanding role for sRNAs in iron homeostasis, but also in orchestrating adaptation to the availability of other metals like manganese and nickel. Furthermore, we describe the sRNA-mediated interdependency between metal homeostasis and oxidative stress responses, and how regulatory networks controlled by sRNAs contribute to survival and virulence.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Brief considerations on targeting RNA with small molecules}, author = {Q Vincens and E Westhof}, url = {https://facultyopinions.com/prime/reports/b/11/39/}, doi = {10.12703/r/11-39}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Faculty Reviews}, volume = {11}, number = {39}, abstract = {For more than three decades, RNA has been known to be a relevant and attractive macromolecule to target but figuring out which RNA should be targeted and how remains challenging. Recent years have seen the confluence of approaches for screening, drug optimization, and target validation that have led to the approval of a few RNA-targeting therapeutics for clinical applications. This focused perspective aims to highlight — but not exhaustively review — key factors accounting for these successes while pointing at crucial aspects worth considering for further breakthroughs.}, keywords = {Drug discovery RNA structure & dynamics RNA targeting SARS-CoV2, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{pmid34878758, title = {How the Presence of Hemin Affects the Expression of the Different Iron Uptake Pathways in Cells}, author = {Vincent Normant and Lauriane Kuhn and Mathilde Munier and Philippe Hammann and Gaëtan L A Mislin and Isabelle J Schalk}, doi = {10.1021/acsinfecdis.1c00525}, issn = {2373-8227}, year = {2022}, date = {2022-01-01}, journal = {ACS Infect Dis}, volume = {8}, number = {1}, pages = {183--196}, abstract = {Iron is an essential nutriment for almost all organisms, but this metal is poorly bioavailable. During infection, bacteria access iron from the host by importing either iron or heme. , a gram-negative pathogen, secretes two siderophores, pyoverdine (PVD) and pyochelin (PCH), to access iron and is also able to use many siderophores produced by other microorganisms (called xenosiderophores). To access heme, uses three distinct uptake pathways, named Has, Phu, and Hxu. We previously showed that expresses the Has and Phu heme uptake systems and the PVD- and PCH-dependent iron uptake pathways in iron-restricted growth conditions, using proteomic and RT-qPCR approaches. Here, using the same approaches, we show that physiological concentrations of hemin in the bacterial growth medium result in the repression of the expression of the proteins of the PVD- and PCH-dependent iron uptake pathways, leading to less production of these two siderophores. This indicates that the pathogen adapts its phenotype to use hemin as an iron source rather than produce PVD and PCH to access iron. Moreover, the presence of both hemin and a xenosiderophore resulted in (i) the strong induction of the expression of the proteins of the added xenosiderophore uptake pathway, (ii) repression of the PVD- and PCH-dependent iron uptake pathways, and (iii) no effect on the expression levels of the Has, Phu, or Hxu systems, indicating that bacteria use both xenosiderophores and heme to access iron.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{Libre2022, title = {A Conserved uORF Regulates APOBEC3G Translation and Is Targeted by HIV-1 Vif Protein to Repress the Antiviral Factor}, author = {C Libre and T Seissler and S Guerrero and J Batisse and C Verriez and B Stupfler and O Gilmer and R Cabrera-Rodriguez and M Weber and A Valenzuela-Fernandez and A Cimarelli and L Etienne and R Marquet and J C Paillart}, url = {https://www.mdpi.com/2227-9059/10/1/13}, doi = {10.1101/2021.01.13.426487}, year = {2022}, date = {2022-01-01}, urldate = {2022-01-01}, journal = {Biomedicines}, volume = {10}, number = {1}, pages = {13}, abstract = {The HIV-1 Vif protein is essential for viral fitness and pathogenicity. Vif decreases expression of cellular cytosine deaminases APOBEC3G (A3G), A3F, A3D and A3H, which inhibit HIV-1 replication by inducing hypermutations during reverse transcription. Vif counteracts A3G by several non-redundant mechanisms (transcription, translation and protein degradation) that concur in reducing the levels of A3G in cell and in preventing its incorporation into viral particles. How Vif affects A3G translation remains unclear. Here, we uncovered the importance of a short conserved uORF (upstream ORF) located within two critical stem-loop structures of the 5-untranslated region (5-UTR) of A3G mRNA. Extensive mutagenesis of A3G 5-UTR, combined with an analysis of their translational effect in transfected cells, indicated that the uORF represses A3G translation and that A3G mRNA is translated through a dual leaky-scanning and re-initiation mechanism. Interestingly, the uORF is also mandatory for the Vif-mediated repression of A3G translation. Furthermore, we showed that the redirection of A3G mRNA into stress granules was dependent not only on Vif, but also on the uORF. Overall, we discovered that A3G translation is regulated by a small uORF conserved in the human population and that Vif uses this specific motif to repress its translation.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Chikungunya virus envelope protein E2 provides a targeting vector for antigen delivery to human dermal CD14+ dendritic cells}, author = { Brulefert and Kraemer and Cumin and Selle and Hoste and Gad and Rühl and Madinier and Chaloin and Münz and Després and MUELLER and Flacher}, doi = {10.1016/j.jid.2021.04.027}, year = {2021}, date = {2021-12-01}, journal = {J. Invest. Dermatol}, volume = {141}, number = {12}, pages = {2985-2989}, keywords = {Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{waltz_how_2021, title = {How to build a ribosome from RNA fragments in Chlamydomonas mitochondria}, author = {Florent Waltz and Thalia Salinas-Giegé and Robert Englmeier and Herrade Meichel and Heddy Soufari and Lauriane Kuhn and Stefan Pfeffer and Friedrich Förster and Benjamin D. Engel and Philippe Giegé and Laurence Drouard and Yaser Hashem}, doi = {10.1038/s41467-021-27200-z}, issn = {2041-1723}, year = {2021}, date = {2021-12-01}, journal = {Nature Communications}, volume = {12}, number = {1}, pages = {7176}, abstract = {Mitochondria are the powerhouse of eukaryotic cells. They possess their own gene expression machineries where highly divergent and specialized ribosomes, named hereafter mitoribosomes, translate the few essential messenger RNAs still encoded by mitochondrial genomes. Here, we present a biochemical and structural characterization of the mitoribosome in the model green alga Chlamydomonas reinhardtii, as well as a functional study of some of its specific components. Single particle cryo-electron microscopy resolves how the Chlamydomonas mitoribosome is assembled from 13 rRNA fragments encoded by separate non-contiguous gene pieces. Additional proteins, mainly OPR, PPR and mTERF helical repeat proteins, are found in Chlamydomonas mitoribosome, revealing the structure of an OPR protein in complex with its RNA binding partner. Targeted amiRNA silencing indicates that these ribosomal proteins are required for mitoribosome integrity. Finally, we use cryo-electron tomography to show that Chlamydomonas mitoribosomes are attached to the inner mitochondrial membrane via two contact points mediated by Chlamydomonas-specific proteins. Our study expands our understanding of mitoribosome diversity and the various strategies these specialized molecular machines adopt for membrane tethering.}, keywords = {Chlamydomonas reinhardtii, Cryoelectron Microscopy, mitochondria, Mitochondrial Proteins, Mitochondrial Ribosomes, PPSE, Ribosomal Proteins, ribosomes, RNA}, pubstate = {published}, tppubtype = {article} } @article{pmid34853303, title = {Cross-species analysis of viral nucleic acid interacting proteins identifies TAOKs as innate immune regulators}, author = {Friederike L Pennemann and Assel Mussabekova and Christian Urban and Alexey Stukalov and Line Lykke Andersen and Vincent Grass and Teresa Maria Lavacca and Cathleen Holze and Lila Oubraham and Yasmine Benamrouche and Enrico Girardi and Rasha E Boulos and Rune Hartmann and Giulio Superti-Furga and Matthias Habjan and Jean-Luc Imler and Carine Meignin and Andreas Pichlmair}, doi = {10.1038/s41467-021-27192-w}, issn = {2041-1723}, year = {2021}, date = {2021-12-01}, urldate = {2021-12-01}, journal = {Nat Commun}, volume = {12}, number = {1}, pages = {7009}, abstract = {The cell intrinsic antiviral response of multicellular organisms developed over millions of years and critically relies on the ability to sense and eliminate viral nucleic acids. Here we use an affinity proteomics approach in evolutionary distant species (human, mouse and fly) to identify proteins that are conserved in their ability to associate with diverse viral nucleic acids. This approach shows a core of orthologous proteins targeting viral genetic material and species-specific interactions. Functional characterization of the influence of 181 candidates on replication of 6 distinct viruses in human cells and flies identifies 128 nucleic acid binding proteins with an impact on virus growth. We identify the family of TAO kinases (TAOK1, -2 and -3) as dsRNA-interacting antiviral proteins and show their requirement for type-I interferon induction. Depletion of TAO kinases in mammals or flies leads to an impaired response to virus infection characterized by a reduced induction of interferon stimulated genes in mammals and impaired expression of srg1 and diedel in flies. Overall, our study shows a larger set of proteins able to mediate the interaction between viral genetic material and host factors than anticipated so far, attesting to the ancestral roots of innate immunity and to the lineage-specific pressures exerted by viruses.}, keywords = {antiviral innate immunity, imler, M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{schiaffini_nyn_2021, title = {A NYN domain protein directly interacts with DECAPPING1 and is required for phyllotactic pattern}, author = {Marlene Schiaffini and Clara Chicois and Aude Pouclet and Tiphaine Chartier and Elodie Ubrig and Anthony Gobert and Hélène Zuber and Jérôme Mutterer and Johana Chicher and Lauriane Kuhn and Philippe Hammann and Dominique Gagliardi and Damien Garcia}, doi = {10.1093/plphys/kiab529}, issn = {1532-2548}, year = {2021}, date = {2021-11-01}, urldate = {2021-11-01}, journal = {Plant Physiology}, pages = {kiab529}, abstract = {In eukaryotes, general mRNA decay requires the decapping complex. The activity of this complex depends on its catalytic subunit, DECAPPING2 (DCP2), and its interaction with decapping enhancers, including its main partner DECAPPING1 (DCP1). Here, we report that in Arabidopsis thaliana, DCP1 also interacts with a NYN domain endoribonuclease, hence named DCP1-ASSOCIATED NYN ENDORIBONUCLEASE 1 (DNE1). Interestingly, we found DNE1 predominantly associated with DCP1, but not with DCP2, and reciprocally, suggesting the existence of two distinct protein complexes. We also showed that the catalytic residues of DNE1 are required to repress the expression of mRNAs in planta upon transient expression. The overexpression of DNE1 in transgenic lines led to growth defects and a similar gene deregulation signature than inactivation of the decapping complex. Finally, the combination of dne1 and dcp2 mutations revealed a functional redundancy between DNE1 and DCP2 in controlling phyllotactic pattern formation. Our work identifies DNE1, a hitherto unknown DCP1 protein partner highly conserved in the plant kingdom and identifies its importance for developmental robustness.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{incarbone_immunocapture_2021, title = {Immunocapture of dsRNA-bound proteins provides insight into Tobacco rattle virus replication complexes and reveals Arabidopsis DRB2 to be a wide-spectrum antiviral effector}, author = {Marco Incarbone and Marion Clavel and Baptiste Monsion and Lauriane Kuhn and Hélène Scheer and Émilie Vantard and Vianney Poignavent and Patrice Dunoyer and Pascal Genschik and Christophe Ritzenthaler}, doi = {10.1093/plcell/koab214}, issn = {1532-298X}, year = {2021}, date = {2021-11-01}, journal = {The Plant Cell}, volume = {33}, number = {11}, pages = {3402--3420}, abstract = {Plant RNA viruses form organized membrane-bound replication complexes to replicate their genomes. This process requires virus- and host-encoded proteins and leads to the production of double-stranded RNA (dsRNA) replication intermediates. Here, we describe the use of Arabidopsis thaliana expressing GFP-tagged dsRNA-binding protein (B2:GFP) to pull down dsRNA and associated proteins in planta upon infection with Tobacco rattle virus (TRV). Mass spectrometry analysis of the dsRNA-B2:GFP-bound proteins from infected plants revealed the presence of viral proteins and numerous host proteins. Among a selection of nine host candidate proteins, eight showed relocalization upon infection, and seven of these colocalized with B2-labeled TRV replication complexes. Infection of A. thaliana T-DNA mutant lines for eight such factors revealed that genetic knockout of dsRNA-BINDING PROTEIN 2 (DRB2) leads to increased TRV accumulation and DRB2 overexpression caused a decrease in the accumulation of four different plant RNA viruses, indicating that DRB2 has a potent and wide-ranging antiviral activity. We propose B2:GFP-mediated pull down of dsRNA to be a versatile method to explore virus replication complex proteomes and to discover key host virus replication factors. Given the universality of dsRNA, development of this tool holds great potential to investigate RNA viruses in other host organisms.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{roche_role_2021, title = {A role for PchHI as the ABC transporter in iron acquisition by the siderophore pyochelin in Pseudomonas aeruginosa}, author = {Béatrice Roche and Mariel A. Garcia-Rivera and Vincent Normant and Lauriane Kuhn and Philippe Hammann and Mark Brönstrup and Gaëtan L. A. Mislin and Isabelle J. Schalk}, doi = {10.1111/1462-2920.15811}, issn = {1462-2920}, year = {2021}, date = {2021-10-01}, journal = {Environmental Microbiology}, abstract = {Iron is an essential nutrient for bacterial growth but poorly bioavailable. Bacteria scavenge ferric iron by synthesizing and secreting siderophores, small compounds with a high affinity for iron. Pyochelin (PCH) is one of the two siderophores produced by the opportunistic pathogen Pseudomonas aeruginosa. After capturing a ferric iron molecule, PCH-Fe is imported back into bacteria first by the outer membrane transporter FptA and then by the inner membrane permease FptX. Here, using molecular biology, 55 Fe uptake assays, and LC-MS/MS quantification, we first find a role for PchHI as the heterodimeric ABC transporter involved in the siderophore-free iron uptake into the bacterial cytoplasm. We also provide the first evidence that PCH is able to reach the bacterial periplasm and cytoplasm when both FptA and FptX are expressed. Finally, we detected an interaction between PchH and FptX, linking the ABC transporter PchHI with the inner permease FptX in the PCH-Fe uptake pathway. These results pave the way for a better understanding of the PCH siderophore pathway, giving future directions to tackle P. aeruginosa infections.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @misc{pmid34520556, title = {Correction to 'Cis regulation within a cluster of viral microRNAs'}, author = {Monika Vilimova and Maud Contrant and Ramy Randrianjafy and Philippe Dumas and Endrit Elbasani and Päivi M Ojala and Sébastien Pfeffer and Aurélie Fender}, doi = {10.1093/nar/gkab806}, issn = {1362-4962}, year = {2021}, date = {2021-10-01}, urldate = {2021-10-01}, journal = {Nucleic Acids Res}, volume = {49}, number = {18}, pages = {10804--10805}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{arquier_brain_2021, title = {Brain adiponectin signaling controls peripheral insulin response in Drosophila}, author = {Nathalie Arquier and Marianne Bjordal and Philippe Hammann and Lauriane Kuhn and Pierre Léopold}, doi = {10.1038/s41467-021-25940-6}, issn = {2041-1723}, year = {2021}, date = {2021-09-01}, journal = {Nature Communications}, volume = {12}, number = {1}, pages = {5633}, abstract = {The brain plays a key role in energy homeostasis, detecting nutrients, metabolites and circulating hormones from peripheral organs and integrating this information to control food intake and energy expenditure. Here, we show that a group of neurons in the Drosophila larval brain expresses the adiponectin receptor (AdipoR) and controls systemic growth and metabolism through insulin signaling. We identify glucose-regulated protein 78 (Grp78) as a circulating antagonist of AdipoR function produced by fat cells in response to dietary sugar. We further show that central AdipoR signaling inhibits peripheral Juvenile Hormone (JH) response, promoting insulin signaling. In conclusion, we identify a neuroendocrine axis whereby AdipoR-positive neurons control systemic insulin response.}, keywords = {Adiponectin, Animals, Brain, Cell Line, Drosophila melanogaster, Drosophila Proteins, Energy Metabolism, Genetically Modified, Hemolymph, Homeostasis, Insulin, Juvenile Hormones, Larva, Neurons, PPSE, Receptors, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{khong_chemical_2021, title = {Chemical Modifications Induced by Phthalic Anhydride, a Respiratory Sensitizer, in Reconstructed Human Epidermis: A Combined HRMAS NMR and LC-MS/MS Proteomic Approach}, author = {Minh-Thuong Khong and Valérie Berl and Lauriane Kuhn and Philippe Hammann and Jean-Pierre Lepoittevin}, doi = {10.1021/acs.chemrestox.1c00172}, issn = {1520-5010}, year = {2021}, date = {2021-09-01}, journal = {Chemical Research in Toxicology}, volume = {34}, number = {9}, pages = {2087--2099}, abstract = {Chemical skin and respiratory allergies are becoming a major health problem. To date our knowledge on the process of protein haptenation is still limited and mainly derived from studies performed in solution using model nucleophiles. In order to better understand chemical interactions between chemical allergens and the skin, we have investigated the reactivity of phthalic anhydride 1 (PA), a chemical respiratory sensitizer, toward reconstructed human epidermis (RHE). This study was performed using a new approach combining HRMAS NMR to investigate the in situ chemical reactivity and LC-MS/MS to identify modified epidermal proteins. In RHE, the reaction of PA appeared to be quite fast and the major product formed was phthalic acid. Two amide type adducts on lysine residues were observed and after 8h of incubation, we also observed the formation of an imide type cyclized adducts with lysine. In parallel, RHE samples topically exposed to phthalic anhydride (13C)-1 were analyzed using the shotgun proteomics method. Thus, 948 different proteins were extracted and identified, 135 of which being modified by PA, i.e., 14.2% of the extracted proteome. A total of 211 amino acids were modified by PA and validated by fragmentation spectra. We thus identified 154 modified lysines, 22 modified histidines, 30 modified tyrosines, and 5 modified arginines. The rate of modified residues, as a proportion of the total number of modifiable nucleophilic residues in RHE, was rather low (1%). At the protein level, modified proteins were mainly type I and type II keratins and other proteins which are abundant in the epidermis such as protein S100A, Caspase 14, annexin A2, serpin B3, fatty-acid binding protein 5, histone H2, H3, H4, etc. However, the most modified protein, mainly on histidine residues, was filaggrin, a protein that is of low abundance (0.0266 mol %) and rich in histidine.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Current clinical and pre-clinical imaging approaches to study the cancer-associated immune system}, author = {Mueller and Gaiddon and Venkatasamy}, doi = {doi: 10.3389/fimmu.2021.716860}, year = {2021}, date = {2021-09-01}, journal = {Frontiers in Immunology}, keywords = {Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{pmid34452932, title = {Protein Phosphatase 4 Negatively Regulates the Immune Deficiency-NF-κB Pathway during the Immune Response}, author = {Layale Salem Wehbe and Dana Barakat and Adrian Acker and Rita El Khoury and Jean-Marc Reichhart and Nicolas Matt and Laure El Chamy}, doi = {10.4049/jimmunol.1901497}, issn = {1550-6606}, year = {2021}, date = {2021-08-27}, urldate = {2021-08-27}, journal = {J Immunol}, volume = {207}, number = {6}, pages = {1616--1626}, abstract = {The evolutionarily conserved immune deficiency (IMD) signaling pathway shields against bacterial infections. It regulates the expression of antimicrobial peptides encoding genes through the activation of the NF-κB transcription factor Relish. Tight regulation of the signaling cascade ensures a balanced immune response, which is otherwise highly harmful. Several phosphorylation events mediate intracellular progression of the IMD pathway. However, signal termination by dephosphorylation remains largely elusive. Here, we identify the highly conserved protein phosphatase 4 (PP4) complex as a bona fide negative regulator of the IMD pathway. RNA interference-mediated gene silencing of , , and which encode the catalytic and regulatory subunits of the phosphatase complex, respectively, caused a marked upregulation of bacterial-induced antimicrobial peptide gene expression in both S2 cells and adult flies. Deregulated IMD signaling is associated with reduced lifespan of -deficient flies in the absence of any infection. In contrast, flies overexpressing this phosphatase are highly sensitive to bacterial infections. Altogether, our results highlight an evolutionarily conserved function of PP4c in the regulation of NF-κB signaling from to mammals.}, keywords = {Drosophila, IKK complex, IMD, immune response, M3i, matt, NF-κB, PP4 complex}, pubstate = {published}, tppubtype = {article} } @article{pmid34191026, title = {Monitoring Insect Transposable Elements in Large Double-Stranded DNA Viruses Reveals Host-to-Virus and Virus-to-Virus Transposition}, author = {Vincent Loiseau and Jean Peccoud and Clémence Bouzar and Sandra Guillier and Jiangbin Fan and Gianpiero Gueli Alletti and Carine Meignin and Elisabeth A Herniou and Brian A Federici and Jörg T Wennmann and Johannes A Jehle and Richard Cordaux and Clément Gilbert}, doi = {10.1093/molbev/msab198}, issn = {1537-1719}, year = {2021}, date = {2021-08-01}, urldate = {2021-08-01}, journal = {Mol Biol Evol}, volume = {38}, number = {9}, pages = {3512--3530}, abstract = {The mechanisms by which transposable elements (TEs) can be horizontally transferred between animals are unknown, but viruses are possible candidate vectors. Here, we surveyed the presence of host-derived TEs in viral genomes in 35 deep sequencing data sets produced from 11 host-virus systems, encompassing nine arthropod host species (five lepidopterans, two dipterans, and two crustaceans) and six different double-stranded (ds) DNA viruses (four baculoviruses and two iridoviruses). We found evidence of viral-borne TEs in 14 data sets, with frequencies of viral genomes carrying a TE ranging from 0.01% to 26.33% for baculoviruses and from 0.45% to 7.36% for iridoviruses. The analysis of viral populations separated by a single replication cycle revealed that viral-borne TEs originating from an initial host species can be retrieved after viral replication in another host species, sometimes at higher frequencies. Furthermore, we detected a strong increase in the number of integrations in a viral population for a TE absent from the hosts' genomes, indicating that this TE has undergone intense transposition within the viral population. Finally, we provide evidence that many TEs found integrated in viral genomes (15/41) have been horizontally transferred in insects. Altogether, our results indicate that multiple large dsDNA viruses have the capacity to shuttle TEs in insects and they underline the potential of viruses to act as vectors of horizontal transfer of TEs. Furthermore, the finding that TEs can transpose between viral genomes of a viral species sets viruses as possible new niches in which TEs can persist and evolve.}, keywords = {M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{Goto2021, title = {Verloren negatively regulates the expression of IMD pathway dependent antimicrobial peptides in Drosophila}, author = {Pragya Prakash and Arghyashree Roychowdhury-Sinha and Akira Goto}, url = {https://www.nature.com/articles/s41598-021-94973-0}, doi = {10.1038/s41598-021-94973-0}, year = {2021}, date = {2021-07-30}, journal = {Scientific Reports}, volume = {11}, number = {15549}, abstract = {Drosophila immune deficiency (IMD) pathway is similar to the human tumor necrosis factor receptor (TNFR) signaling pathway and is preferentially activated by Gram-negative bacterial infection. Recent studies highlighted the importance of IMD pathway regulation as it is tightly controlled by numbers of negative regulators at multiple levels. Here, we report a new negative regulator of the IMD pathway, Verloren (Velo). Silencing of Velo led to constitutive expression of the IMD pathway dependent antimicrobial peptides (AMPs), and Escherichia coli stimulation further enhanced the AMP expression. Epistatic analysis indicated that Velo knock-down mediated AMP upregulation is dependent on the canonical members of the IMD pathway. The immune fluorescent study using overexpression constructs revealed that Velo resides both in the nucleus and cytoplasm, but the majority (~ 75%) is localized in the nucleus. We also observed from in vivo analysis that Velo knock-down flies exhibit significant upregulation of the AMP expression and reduced bacterial load. Survival experiments showed that Velo knock-down flies have a short lifespan and are susceptible to the infection of pathogenic Gram-negative bacteria, P. aeruginosa. Taken together, these data suggest that Velo is an additional new negative regulator of the IMD pathway, possibly acting in both the nucleus and cytoplasm.}, keywords = {bacteria, Biochemistry, DNA, Fungi, Gene Expression, gene regulation, Genetics, hoffmann, Immunochemistry, Immunology, infection, inflammation, Innate immune cells, innate immunity, M3i, microbiology, Molecular Biology, pathogens, RNA, RNAi, Signal Transduction, Transcription}, pubstate = {published}, tppubtype = {article} } @article{Hartmann2021, title = {Two cGAS-like receptors induce antiviral immunity in Drosophila}, author = {Andreas Holleufer and Kasper Grønbjerg Winther and Hans Henrik Gad and Xianlong Ai and Yuqiang Chen and Lihua Li and Ziming Wei and Huimin Deng and Jiyong Liu and Ninna Ahlmann Frederiksen and Bine Simonsen and Line Lykke Andersen and Karin Kleigrewe and Louise Dalskov and Andreas Pichlmair and Hua Cai and Jean-Luc Imler and Rune Hartmann}, editor = {Nature Publishing Group}, doi = {https://doi.org/10.1038/s41586-021-03800-z}, year = {2021}, date = {2021-07-14}, urldate = {2021-07-14}, journal = {Nature}, volume = {597}, pages = {114-118}, abstract = {In mammals, cyclic GMP-AMP (cGAMP) synthase (cGAS) produces the cyclic dinucleotide (CDN) 2'3'-cGAMP in response to cytosolic DNA and this triggers an antiviral immune response. cGAS belongs to a large family of cGAS/DncV-like nucleotidyltransferases, present in both prokaryotes1 and eukaryotes2–5. In bacteria, these enzymes synthesize a range of cyclic oligonucleotide and have recently emerged as important regulators of phage infections6–8. Here, we identify two novel cGAS-like receptors (cGLRs) in the insect Drosophila melanogaster. We show that cGLR1 and cGLR2 activate Sting and NF-κB dependent antiviral immunity in response to infection with RNA or DNA viruses. cGLR1 is activated by dsRNA to produce the novel CDN 3'2'-cGAMP whereas cGLR2 produces a combination of 2'3'-cGAMP and 3'2' cGAMP in response to a yet unidentified stimulus. Our data establish cGAS as the founding member of a family of receptors sensing different types of nucleic acids and triggering immunity through production of CDNs beyond 2'3'-cGAMP.}, keywords = {antiviral immunity, cGAS-like receptors, Drosophila, imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{pmid34261127, title = {cGAS-like receptors sense RNA and control 3'2'-cGAMP signalling in Drosophila}, author = {Kailey M Slavik and Benjamin R Morehouse and Adelyn E Ragucci and Wen Zhou and Xianlong Ai and Yuqiang Chen and Lihua Li and Ziming Wei and Heike Bähre and Martin König and Roland Seifert and Amy S Y Lee and Hua Cai and Jean-Luc Imler and Philip J Kranzusch}, doi = {10.1038/s41586-021-03743-5}, issn = {1476-4687}, year = {2021}, date = {2021-07-14}, journal = {Nature}, volume = {597}, number = {7874}, pages = {109--113}, abstract = {Cyclic GMP-AMP synthase (cGAS) is a cytosolic DNA sensor that produces the second messenger cG[2'-5']pA[3'-5']p (2'3'-cGAMP) and controls activation of innate immunity in mammalian cells. Animal genomes typically encode multiple proteins with predicted homology to cGAS, but the function of these uncharacterized enzymes is unknown. Here we show that cGAS-like receptors (cGLRs) are innate immune sensors that are capable of recognizing divergent molecular patterns and catalysing synthesis of distinct nucleotide second messenger signals. Crystal structures of human and insect cGLRs reveal a nucleotidyltransferase signalling core shared with cGAS and a diversified primary ligand-binding surface modified with notable insertions and deletions. We demonstrate that surface remodelling of cGLRs enables altered ligand specificity and used a forward biochemical screen to identify cGLR1 as a double-stranded RNA sensor in the model organism Drosophila melanogaster. We show that RNA recognition activates Drosophila cGLR1 to synthesize the novel product cG[3'-5']pA[2'-5']p (3'2'-cGAMP). A crystal structure of Drosophila stimulator of interferon genes (dSTING) in complex with 3'2'-cGAMP explains selective isomer recognition, and 3'2'-cGAMP induces an enhanced antiviral state in vivo that protects from viral infection. Similar to radiation of Toll-like receptors in pathogen immunity, our results establish cGLRs as a diverse family of metazoan pattern recognition receptors.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{nettersheim_dna_2021, title = {DNA polymerase η is a substrate for calpain: a possible mechanism for pol η retention in UV-induced replication foci}, author = {Jo-Ann Nettersheim and Régine Janel-Bintz and Lauriane Kuhn and Agnès M. Cordonnier}, doi = {10.1242/jcs.258637}, issn = {1477-9137}, year = {2021}, date = {2021-07-01}, journal = {Journal of Cell Science}, volume = {134}, number = {13}, pages = {jcs258637}, abstract = {DNA polymerase η (pol η) is specifically required for translesion DNA synthesis across UV-induced DNA lesions. Recruitment of this error-prone DNA polymerase is tightly regulated during replication to avoid mutagenesis and perturbation of fork progression. Here, we report that pol η interacts with the calpain small subunit-1 (CAPNS1) in a yeast two-hybrid screening. This interaction is functional, as demonstrated by the ability of endogenous calpain to mediate calcium-dependent cleavage of pol η in cell-free extracts and in living cells treated with a calcium ionophore. The proteolysis of pol η was found to occur at position 465, leading to a catalytically active truncated protein containing the PCNA-interacting motif PIP1. Unexpectedly, cell treatment with the specific calpain inhibitor calpeptin resulted in a decreased extent of pol η foci after UV irradiation, indicating that calpain positively regulates pol η accumulation in replication foci.}, keywords = {calpain, Calpain protease, CAPNS1, DNA Damage, DNA damage response, DNA polymerase η, DNA Repair, DNA Replication, DNA-Directed DNA Polymerase, PPSE, Replication foci, Translesion DNA synthesis}, pubstate = {published}, tppubtype = {article} } @article{mancera-martinez_phosphorylation_2021, title = {Phosphorylation of a reinitiation supporting protein, RISP, determines its function in translation reinitiation}, author = {Eder Mancera-Martínez and Yihan Dong and Joelle Makarian and Ola Srour and Odon Thiébeauld and Muhammed Jamsheer and Johana Chicher and Philippe Hammann and Mikhail Schepetilnikov and Lyubov A. Ryabova}, doi = {10.1093/nar/gkab501}, issn = {1362-4962}, year = {2021}, date = {2021-07-01}, journal = {Nucleic Acids Research}, volume = {49}, number = {12}, pages = {6908--6924}, abstract = {Reinitiation supporting protein, RISP, interacts with 60S (60S ribosomal subunit) and eIF3 (eukaryotic initiation factor 3) in plants. TOR (target-of-rapamycin) mediates RISP phosphorylation at residue Ser267, favoring its binding to eL24 (60S ribosomal protein L24). In a viral context, RISP, when phosphorylated, binds the CaMV transactivator/ viroplasmin, TAV, to assist in an exceptional mechanism of reinitiation after long ORF translation. Moreover, we show here that RISP interacts with eIF2 via eIF2β and TOR downstream target 40S ribosomal protein eS6. A RISP phosphorylation knockout, RISP-S267A, binds preferentially eIF2β, and both form a ternary complex with eIF3a in vitro. Accordingly, transient overexpression in plant protoplasts of RISP-S267A, but not a RISP phosphorylation mimic, RISP-S267D, favors translation initiation. In contrast, RISP-S267D preferentially binds eS6, and, when bound to the C-terminus of eS6, can capture 60S in a highly specific manner in vitro, suggesting that it mediates 60S loading during reinitiation. Indeed, eS6-deficient plants are highly resistant to CaMV due to their reduced reinitiation capacity. Strikingly, an eS6 phosphomimic, when stably expressed in eS6-deficient plants, can fully restore the reinitiation deficiency of these plants in cellular and viral contexts. These results suggest that RISP function in translation (re)initiation is regulated by phosphorylation at Ser267.}, keywords = {Arabidopsis, Arabidopsis Proteins, Caulimovirus, Eukaryotic, Eukaryotic Initiation Factor-2B, Eukaryotic Initiation Factor-3, Large, Peptide Chain Initiation, Phosphorylation, PPSE, Ribosomal Protein S6, Ribosome Subunits, translational}, pubstate = {published}, tppubtype = {article} } @article{Cai2021, title = {cGAS-STING: insight on the evolution of a primordial antiviral signaling cassette}, author = {Hua Cai and Jean-Luc Imler }, url = { https://doi.org/10.12703/r/10-54}, doi = {10.12703/r/10-54}, year = {2021}, date = {2021-06-08}, journal = {Faculty Reviews}, volume = {10}, pages = {54}, abstract = {Stimulator of interferon genes (STING) functions in the cytosolic DNA-sensing pathway of innate immunity in mammals. It is activated upon binding the cyclic dinucleotide 2′3′-cGAMP, a second messenger produced by the enzyme cyclic guanosine monophosphate–adenosine monophosphate synthase (cGAS), which acts as the receptor for DNA in this pathway, and triggers the expression of interferons and other viral stress-induced genes. The ancient origin of STING in the evolution of animals had been noted, but its primitive function was speculative. We review here recent advances in the remarkable history of cGAS-STING signaling, which establish that cGAS is a member of the family of cGAS/DncV-like nucleotidyltransferases (CD-NTases). In bacteria, CD-NTases synthesize a wide range of cyclic oligonucleotide second messengers in response to bacteriophage infections, which in turn activate a variety of effector proteins to abort phage infection. Among these effectors, some are related to STING, revealing an ancestral function for the cGAS-STING cassette in antiviral host defense. Study of STING signaling in invertebrate animals is consistent with an early acquisition in the history of metazoans of CD-NTase- and STING-encoding genes to counter the universal threat of viruses. In particular, STING-dependent immunity appears to play a previously unsuspected important role in some insects. These discoveries open up interesting perspectives for the use of model organisms to decipher emerging aspects of cGAS-STING biology in mammals, such as the activation of interferon-independent responses or the function and regulation of cGAS in the nucleus.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{dahlet_e2f6_2021, title = {E2F6 initiates stable epigenetic silencing of germline genes during embryonic development}, author = {Thomas Dahlet and Matthias Truss and Ute Frede and Hala Al Adhami and Anaïs F. Bardet and Michael Dumas and Judith Vallet and Johana Chicher and Philippe Hammann and Sarah Kottnik and Peter Hansen and Uschi Luz and Gonzalo Alvarez and Ghislain Auclair and Jochen Hecht and Peter N. Robinson and Christian Hagemeier and Michael Weber}, doi = {10.1038/s41467-021-23596-w}, issn = {2041-1723}, year = {2021}, date = {2021-06-01}, journal = {Nature Communications}, volume = {12}, number = {1}, pages = {3582}, abstract = {In mouse development, long-term silencing by CpG island DNA methylation is specifically targeted to germline genes; however, the molecular mechanisms of this specificity remain unclear. Here, we demonstrate that the transcription factor E2F6, a member of the polycomb repressive complex 1.6 (PRC1.6), is critical to target and initiate epigenetic silencing at germline genes in early embryogenesis. Genome-wide, E2F6 binds preferentially to CpG islands in embryonic cells. E2F6 cooperates with MGA to silence a subgroup of germline genes in mouse embryonic stem cells and in embryos, a function that critically depends on the E2F6 marked box domain. Inactivation of E2f6 leads to a failure to deposit CpG island DNA methylation at these genes during implantation. Furthermore, E2F6 is required to initiate epigenetic silencing in early embryonic cells but becomes dispensable for the maintenance in differentiated cells. Our findings elucidate the mechanisms of epigenetic targeting of germline genes and provide a paradigm for how transient repression signals by DNA-binding factors in early embryonic cells are translated into long-term epigenetic silencing during mouse development.}, keywords = {Animals, Binding Sites, Cell Differentiation, CpG Islands, CRISPR-Cas Systems, DNA Methylation, E2F6 Transcription Factor, Embryonic Development, Epigenesis, Gene Silencing, Genetic, Germ Cells, Knockout, Mice, Mouse Embryonic Stem Cells, Polycomb Repressive Complex 1, PPSE, RNA, Small Interfering}, pubstate = {published}, tppubtype = {article} } @article{Leite2021, title = {Distinct Roles of Hemocytes at Different Stages of Infection by Dengue and Zika Viruses in Aedes aegypti Mosquitoes}, author = {Thiago H J F Leite and Alvaro G A Ferreira and Jean-Luc Imler and João T Marques}, url = {https://www.frontiersin.org/articles/10.3389/fimmu.2021.660873/full}, doi = {10.3389/fimmu.2021.660873}, year = {2021}, date = {2021-05-13}, journal = {Frontiers in immunology}, volume = {12}, pages = {660873}, abstract = {Aedes aegypti mosquitoes are vectors for arboviruses of medical importance such as dengue (DENV) and Zika (ZIKV) viruses. Different innate immune pathways contribute to the control of arboviruses in the mosquito vector including RNA interference, Toll and Jak- STAT pathways. However, the role of cellular responses mediated by circulating macrophage-like cells known as hemocytes remains unclear. Here we show that hemocytes are recruited to the midgut of Ae. aegypti mosquitoes in response to DENV or ZIKV. Blockade of the phagocytic function of hemocytes using latex beads induced increased accumulation of hemocytes in the midgut and a reduction in virus infection levels in this organ. In contrast, inhibition of phagocytosis by hemocytes led to increased systemic dissemination and replication of DENV and ZIKV. Hence, our work reveals a dual role for hemocytes in Ae. aegypti mosquitoes, whereby phagocytosis is not required to control viral infection in the midgut but is essential to restrict systemic dissemination. Further understanding of the mechanism behind this duality could help the design of vector-based strategies to prevent transmission of arboviruses.}, keywords = {Aedes, Dengue, Hemocytes, imler, innate immunity, M3i, Marques, Zika}, pubstate = {published}, tppubtype = {article} } @article{pmid34059075, title = {Brain HIV-1 latently-infected reservoirs targeted by the suicide gene strategy}, author = {Sepideh Saeb and Mehrdad Ravanshad and Mahmoud Reza Pourkarim and Fadoua Daouad and Kazem Baesi and Olivier Rohr and Clémentine Wallet and Christian Schwartz}, doi = {10.1186/s12985-021-01584-2}, issn = {1743-422X}, year = {2021}, date = {2021-05-01}, urldate = {2021-05-01}, journal = {Virol J}, volume = {18}, number = {1}, pages = {107}, abstract = {Reducing the pool of HIV-1 reservoirs in patients is a must to achieve functional cure. The most prominent HIV-1 cell reservoirs are resting CD4 + T cells and brain derived microglial cells. Infected microglial cells are believed to be the source of peripheral tissues reseedings and the emergence of drug resistance. Clearing infected cells from the brain is therefore crucial. However, many characteristics of microglial cells and the central nervous system make extremely difficult their eradication from brain reservoirs. Current methods, such as the "shock and kill", the "block and lock" and gene editing strategies cannot override these difficulties. Therefore, new strategies have to be designed when considering the elimination of brain reservoirs. We set up an original gene suicide strategy using latently infected microglial cells as model cells. In this paper we provide proof of concept of this strategy.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{chen2021, title = {A time course transcriptomic analysis of host and injected oncogenic cells reveals new aspects of Drosophila immune defenses}, author = {Di Chen and Arghyashree Roychowdhury-Sinha and Pragya Prakash and Xiao Lan and Wenmin Fan and Akira Goto and Jules A Hoffmann}, url = {https://www.pnas.org/content/118/12/e2100825118}, doi = {https://doi.org/10.1073/pnas.2100825118}, year = {2021}, date = {2021-03-23}, urldate = {2021-03-23}, journal = {PNAS}, volume = {118}, number = {12}, abstract = {Oncogenic RasV12 cells [A. Simcox et al., PLoS Genet. 4, e1000142 (2008)] injected into adult males proliferated massively after a lag period of several days, and led to the demise of the flies after 2 to 3 wk. The injection induced an early massive transcriptomic response that, unexpectedly, included more than 100 genes encoding chemoreceptors of various families. The kinetics of induction and the identities of the induced genes differed markedly from the responses generated by injections of microbes. Subsequently, hundreds of genes were up-regulated, attesting to intense catabolic activities in the flies, active tracheogenesis, and cuticulogenesis, as well as stress and inflammation-type responses. At 11 d after the injections, GFP-positive oncogenic cells isolated from the host flies exhibited a markedly different transcriptomic profile from that of the host and distinct from that at the time of their injection, including in particular up-regulated expression of genes typical for cells engaged in the classical antimicrobial response of Drosophila.}, keywords = {cancer, chemoreceptor, Drosophila melanogaster, goto, hoffmann, innate immunity, M3i, RasV12}, pubstate = {published}, tppubtype = {article} } @article{gasser_esterase_2021, title = {The Esterase PfeE, the Achilles' Heel in the Battle for Iron between Pseudomonas aeruginosa and Escherichia coli}, author = {Véronique Gasser and Lauriane Kuhn and Thibaut Hubert and Laurent Aussel and Philippe Hammann and Isabelle J. Schalk}, doi = {10.3390/ijms22062814}, issn = {1422-0067}, year = {2021}, date = {2021-03-01}, urldate = {2021-03-01}, journal = {International Journal of Molecular Sciences}, volume = {22}, number = {6}, pages = {2814}, abstract = {Bacteria access iron, a key nutrient, by producing siderophores or using siderophores produced by other microorganisms. The pathogen Pseudomonas aeruginosa produces two siderophores but is also able to pirate enterobactin (ENT), the siderophore produced by Escherichia coli. ENT-Fe complexes are imported across the outer membrane of P. aeruginosa by the two outer membrane transporters PfeA and PirA. Iron is released from ENT in the P. aeruginosa periplasm by hydrolysis of ENT by the esterase PfeE. We show here that pfeE gene deletion renders P. aeruginosa unable to grow in the presence of ENT because it is unable to access iron via this siderophore. Two-species co-cultures under iron-restricted conditions show that P. aeruginosa strongly represses the growth of E. coli as long it is able to produce its own siderophores. Both strains are present in similar proportions in the culture as long as the siderophore-deficient P. aeruginosa strain is able to use ENT produced by E. coli to access iron. If pfeE is deleted, E. coli has the upper hand in the culture and P. aeruginosa growth is repressed. Overall, these data show that PfeE is the Achilles' heel of P. aeruginosa in communities with bacteria producing ENT.}, keywords = {Carrier Proteins, co-cultures, enterobactin, Escherichia coli, Escherichia coli Proteins, Esterases, Iron, iron homeostasis, iron uptake, outer membrane transporters, PPSE, Pseudomonas aeruginosa, siderophore, TonB}, pubstate = {published}, tppubtype = {article} } @article{enkler_cex1_2021, title = {Cex1 is a component of the COPI intracellular trafficking machinery}, author = {Ludovic Enkler and Bruno Rinaldi and Johan Owen Craene and Philippe Hammann and Osamu Nureki and Bruno Senger and Sylvie Friant and Hubert D. Becker}, doi = {10.1242/bio.058528}, issn = {2046-6390}, year = {2021}, date = {2021-03-01}, journal = {Biology Open}, volume = {10}, number = {3}, pages = {bio058528}, abstract = {COPI (coatomer complex I) coated vesicles are involved in Golgi-to-ER and intra-Golgi trafficking pathways, and mediate retrieval of ER resident proteins. Functions and components of the COPI-mediated trafficking pathways, beyond the canonical set of Sec/Arf proteins, are constantly increasing in number and complexity. In mammalian cells, GORAB, SCYL1 and SCYL3 proteins regulate Golgi morphology and protein glycosylation in concert with the COPI machinery. Here, we show that Cex1, homologous to the mammalian SCYL proteins, is a component of the yeast COPI machinery, by interacting with Sec27, Sec28 and Sec33 (Ret1/Cop1) proteins of the COPI coat. Cex1 was initially reported to mediate channeling of aminoacylated tRNA outside of the nucleus. Our data show that Cex1 localizes at membrane compartments, on structures positive for the Sec33 α-COP subunit. Moreover, the Wbp1 protein required for N-glycosylation and interacting via its di-lysine motif with the Sec27 β'-COP subunit is mis-targeted in cex1Δ deletion mutant cells. Our data point to the possibility of developing Cex1 yeast-based models to study neurodegenerative disorders linked to pathogenic mutations of its human homologue SCYL1.}, keywords = {Arc1, Cex1, COPI coat, PPSE, SCYL1, trafficking}, pubstate = {published}, tppubtype = {article} } @article{scheer_tutase_2021, title = {The TUTase URT1 connects decapping activators and prevents the accumulation of excessively deadenylated mRNAs to avoid siRNA biogenesis}, author = {Hélène Scheer and Caroline Almeida and Emilie Ferrier and Quentin Simonnot and Laure Poirier and David Pflieger and François M. Sement and Sandrine Koechler and Christina Piermaria and Paweł Krawczyk and Seweryn Mroczek and Johana Chicher and Lauriane Kuhn and Andrzej Dziembowski and Philippe Hammann and Hélène Zuber and Dominique Gagliardi}, doi = {10.1038/s41467-021-21382-2}, issn = {2041-1723}, year = {2021}, date = {2021-02-01}, journal = {Nature Communications}, volume = {12}, number = {1}, pages = {1298}, abstract = {Uridylation is a widespread modification destabilizing eukaryotic mRNAs. Yet, molecular mechanisms underlying TUTase-mediated mRNA degradation remain mostly unresolved. Here, we report that the Arabidopsis TUTase URT1 participates in a molecular network connecting several translational repressors/decapping activators. URT1 directly interacts with DECAPPING 5 (DCP5), the Arabidopsis ortholog of human LSM14 and yeast Scd6, and this interaction connects URT1 to additional decay factors like DDX6/Dhh1-like RNA helicases. Nanopore direct RNA sequencing reveals a global role of URT1 in shaping poly(A) tail length, notably by preventing the accumulation of excessively deadenylated mRNAs. Based on in vitro and in planta data, we propose a model that explains how URT1 could reduce the accumulation of oligo(A)-tailed mRNAs both by favoring their degradation and because 3' terminal uridines intrinsically hinder deadenylation. Importantly, preventing the accumulation of excessively deadenylated mRNAs avoids the biogenesis of illegitimate siRNAs that silence endogenous mRNAs and perturb Arabidopsis growth and development.}, keywords = {Arabidopsis, Arabidopsis Proteins, Co-Repressor Proteins, DEAD-box RNA Helicases, Gene Expression Regulation, Humans, messenger, Plant, PPSE, Proto-Oncogene Proteins, Ribonucleoproteins, RNA, RNA Nucleotidyltransferases, RNA Stability, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Small Interfering, Tobacco, transcriptome, Uridine}, pubstate = {published}, tppubtype = {article} } @article{H2021, title = {Insect decline: immediate action is needed}, author = {H Jactel and JL Imler and L Lambrechts and AB Failloux and JD Lebreton and Y Le Maho and JC Duplessy and P Cossart and P Grandcolas}, url = {https://doi.org/10.5802/crbiol.37}, doi = {10.5802/crbiol.37}, issn = {17683238}, year = {2021}, date = {2021-01-25}, journal = {Comptes Rendus. Biologies}, abstract = { Insects appeared more than 400 million years ago and they represent the richest and most diverse taxonomic group with several million species. Yet, under the combined effect of the loss of natural habitats, the intensification of agriculture with massive use of pesticides, global warming and biological invasions, insects show alarming signs of decline. Although difficult to quantify, species extinction and population reductions are confirmed for many ecosystems. This results in a loss of services such as the pollination of plants, including food crops, the recycling of organic matter, the supply of goods such as honey and the stability of food webs. It is therefore urgent to halt the decline of Insects. We recommend implementing long-term monitoring of populations, tackling the causes of insect decline by reducing the use of synthetic insecticides, preserving natural habitats, and reinventing a positive relationship between humans and insects. Apparus il y a plus de 400 millions d’années, les Insectes représentent le groupe taxonomique le plus riche et diversifié, avec plusieurs millions d’espèces. Sous l’effet de la disparition des habitats, de l’intensification de l’agriculture avec l’usage massif des pesticides, du réchauffement climatique et des invasions biologiques, les Insectes montrent des signes alarmants de déclin. Bien que difficiles à quantifier, la disparition des espèces et la réduction de leurs populations sont avérées et communes à de nombreux écosystèmes. Elles se traduisent par une perte des services rendus, comme la pollinisa- tion des plantes vivrières, le recyclage de la matière organique, la fourniture de biens comme le miel, et l’équilibre des réseaux trophiques. Il est donc urgent de freiner le déclin des Insectes. Pour cela, il faut mettre en œuvre des suivis à long terme des populations, réduire l’usage des insecticides de syn- thèse, préserver les habitats naturels, et réinventer la relation de l’Homme à l’Insecte en revalorisant son image et ses usages.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{, title = {Distinct pathogenic mechanisms of various RARS1 mutations in Pelizaeus-Merzbacher-like disease}, author = {G Li and G Eriani and E D Wang and X L Zhou}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33515434}, doi = {10.1007/s11427-020-1838-2}, isbn = {33515434}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Sci China Life Sci}, volume = {64}, number = {10}, pages = {1645-1660}, abstract = {Mutations of the genes encoding aminoacyl-tRNA synthetases are highly associated with various central nervous system disorders. Recurrent mutations, including c.5A>G, p.D2G; c.1367C>T, p.S456L; c.1535G>A, p.R512Q and c.1846_1847del, p. Y616Lfs*6 of RARS1 gene, which encodes two forms of human cytoplasmic arginyl-tRNA synthetase (hArgRS), are linked to Pelizaeus-Merzbacher-like disease (PMLD) with unclear pathogenesis. Among these mutations, c.5A>G is the most extensively reported mutation, leading to a p.D2G mutation in the N-terminal extension of the long-form hArgRS. Here, we showed the detrimental effects of R512Q substitution and DeltaC mutations on the structure and function of hArgRS, while the most frequent mutation c.5A>G, p.D2G acted in a different manner without impairing hArgRS activity. The nucleotide substitution c.5A>G reduced translation of hArgRS mRNA, and an upstream open reading frame contributed to the suppressed translation of the downstream main ORF. Taken together, our results elucidated distinct pathogenic mechanisms of various RARS1 mutations in PMLD.}, note = {1869-1889 (Electronic) 1674-7305 (Linking) Journal Article}, keywords = {aminoacyl-tRNA synthetase (aaRS), central nervous system (CNS), ERIANI, Protein Biosynthesis, translation initiation, tRNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Mercier2021, title = {MS2-Affinity Purification Coupled with RNA Sequencing in Gram-Positive Bacteria}, author = {N Mercier and K Prévost and E Massé and P Romby and I Caldelari and D Lalaouna}, url = {https://www.jove.com/t/61731/ms2-affinity-purification-coupled-with-rna-sequencing-gram-positive}, doi = {10.3791/61731}, year = {2021}, date = {2021-01-01}, journal = {J Vis Exp}, pages = {e61731}, abstract = {Although small regulatory RNAs (sRNAs) are widespread among the bacterial domain of life, the functions of many of them remain poorly characterized notably due to the difficulty of identifying their mRNA targets. Here, we described a modified protocol of the MS2-Affinity Purification coupled with RNA Sequencing (MAPS) technology, aiming to reveal all RNA partners of a specific sRNA in vivo. Broadly, the MS2 aptamer is fused to the 5’ extremity of the sRNA of interest. This construct is then expressed in vivo, allowing the MS2-sRNA to interact with its cellular partners. After bacterial harvesting, cells are mechanically lysed. The crude extract is loaded into an amylose-based chromatography column previously coated with the MS2 protein fused to the maltose binding protein. This enables the specific capture of MS2-sRNA and interacting RNAs. After elution, co-purified RNAs are identified by high-throughput RNA sequencing and subsequent bioinformatic analysis. The following protocol has been implemented in the Gram-positive human pathogen Staphylococcus aureus and is, in principle, transposable to any Gram-positive bacteria. To sum up, MAPS technology constitutes an efficient method to deeply explore the regulatory network of a particular sRNA, offering a snapshot of its whole targetome. However, it is important to keep in mind that putative targets identified by MAPS still need to be validated by complementary experimental approaches.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{muller_tridimensional_2021, title = {[Tridimensional in vitro models of nervous and immune systems in the skin]}, author = {Quentin Muller and François Berthod and Vincent Flacher}, doi = {10.1051/medsci/2020260}, issn = {1958-5381}, year = {2021}, date = {2021-01-01}, journal = {Medecine Sciences: M/S}, volume = {37}, number = {1}, pages = {68--76}, abstract = {The immune system and the sensory nervous system are responsible for perceiving danger under distinct yet complementary forms. In the last few years, neuroimmune interactions have become an important topic of dermatological research for conditions including wound healing, atopic dermatitis and psoriasis. We present here a selection of tridimensional in vitro models that reproduce skin structure and integrate an immune or a sensory function. Future evolutions of such models are expected to greatly contribute in a better understanding of reciprocal influences between sensory nervous system and immune system.}, keywords = {Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{Catalan-Moreno2021, title = {RNA thermoswitches modulate Staphylococcus aureus adaptation to ambient temperatures}, author = {A Catalan-Moreno and M Cela and P Menendez-Gil and N Irurzun and C J Caballero and I Caldelari and A Toledo-Arana}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33660769}, doi = {0.1093/nar/gkab117}, isbn = {33660769}, year = {2021}, date = {2021-01-01}, journal = {Nucleic Acids Res}, pages = {on press}, abstract = {Thermoregulation of virulence genes in bacterial pathogens is essential for environment-to-host transition. However, the mechanisms governing cold adaptation when outside the host remain poorly understood. Here, we found that the production of cold shock proteins CspB and CspC from Staphylococcus aureus is controlled by two paralogous RNA thermoswitches. Through in silico prediction, enzymatic probing and site-directed mutagenesis, we demonstrated that cspB and cspC 5'UTRs adopt alternative RNA structures that shift from one another upon temperature shifts. The open (O) conformation that facilitates mRNA translation is favoured at ambient temperatures (22 degrees C). Conversely, the alternative locked (L) conformation, where the ribosome binding site (RBS) is sequestered in a double-stranded RNA structure, is folded at host-related temperatures (37 degrees C). These structural rearrangements depend on a long RNA hairpin found in the O conformation that sequesters the anti-RBS sequence. Notably, the remaining S. aureus CSP, CspA, may interact with a UUUGUUU motif located in the loop of this long hairpin and favour the folding of the L conformation. This folding represses CspB and CspC production at 37 degrees C. Simultaneous deletion of the cspB/cspC genes or their RNA thermoswitches significantly decreases S. aureus growth rate at ambient temperatures, highlighting the importance of CspB/CspC thermoregulation when S. aureus transitions from the host to the environment.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{F2021, title = {Identification of Pr78Gag binding sites on the Mason-Pfizer monkey virus genomic RNA packaging determinants}, author = {F Pitchai and A Chameettachal and V Vivet-Boudou and L M Ali and V N Pillai and A Krishnan and S Bernacchi and F Mustafa and Marquet R and T A Rizvi}, url = {https://www.sciencedirect.com/science/article/pii/S0022283621001224?via%3Dihub}, doi = {10.1016/j.jmb.2021.166923}, year = {2021}, date = {2021-01-01}, journal = {J Mol Biol}, volume = {433}, number = {10}, pages = {166923}, abstract = {How retroviral Gag proteins recognize the packaging signals (Psi) on their genomic RNA (gRNA) is a key question that we addressed here using Mason-Pfizer monkey virus (MPMV) as a model system by combining band-shift assays and footprinting experiments. Our data show that Pr78Gag selects gRNA against spliced viral RNA by simultaneously binding to two single stranded loops on the MPMV Psi RNA: (1) a large purine loop (ssPurines), and (2) a loop which partially overlaps with a mostly base-paired purine repeat (bpPurines) and extends into a GU-rich binding motif. Importantly, this second Gag binding site is located immediately downstream of the major splice donor (mSD) and is thus absent from the spliced viral RNAs. Identifying elements crucial for MPMV gRNA packaging should help in understanding not only the mechanism of virion assembly by retroviruses, but also facilitate construction of safer retroviral vectors for human gene therapy.}, keywords = {MARQUET, Retroviruses Gag-RNA interactions Purines Footprinting hSHAPE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{C2021b, title = {Post-Translational Modifications of Retroviral HIV-1 Gag Precursors: An Overview of Their Biological Role}, author = {C Busienne and R Marquet and J C Paillart and S Bernacchi}, url = {https://www.mdpi.com/1422-0067/22/6/2871}, doi = {10.3390/ijms22062871}, year = {2021}, date = {2021-01-01}, journal = {Int. J. Mol. Sci.}, volume = {22}, number = {6}, pages = {2871}, abstract = {Protein post-translational modifications (PTMs) play key roles in eukaryotes since they finely regulate numerous mechanisms used to diversify the protein functions and to modulate their signaling networks. Besides, these chemical modifications also take part in the viral hijacking of the host, and also contribute to the cellular response to viral infections. All domains of the human immunodeficiency virus type 1 (HIV-1) Gag precursor of 55-kDa (Pr55Gag), which is the central actor for viral RNA specific recruitment and genome packaging, are post-translationally modified. In this review, we summarize the current knowledge about HIV-1 Pr55Gag PTMs such as myristoylation, phosphorylation, ubiquitination, sumoylation, methylation, and ISGylation in order to figure out how these modifications affect the precursor functions and viral replication. Indeed, in HIV-1, PTMs regulate the precursor trafficking between cell compartments and its anchoring at the plasma membrane, where viral assembly occurs. Interestingly, PTMs also allow Pr55Gag to hijack the cell machinery to achieve viral budding as they drive recognition between viral proteins or cellular components such as the ESCRT machinery. Finally, we will describe and compare PTMs of several other retroviral Gag proteins to give a global overview of their role in the retroviral life cycle.}, keywords = {HIV-1, MARQUET, PAILLART, post-translational modifications, Pr55Gag precursor, retroviral Gag precursors, retroviral life cycle, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Chameettachal2021, title = {A purine loop and the primer binding site are critical for the selective encapsidation of mouse mammary tumor virus genomic RNA by Pr77Gag}, author = {A Chameettachal and V Vivet-Boudou and F N Pitchai and Pillai V N and L M Ali and A Krishnan and S Bernacchi and F Mustafa and R Marquet and T A Rizvi}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Nucleic Acids Res}, volume = {49}, number = {8}, pages = {4668-4688}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Hayek2021, title = {eIF3 interacts with histone H4 messenger RNA to regulate its translation}, author = {H Hayek and L Gross and A Janvier and L Schaeffer and F Martin and G Eriani and C Allmang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33766559}, doi = {10.1016/j.jbc.2021.100578}, isbn = {33766559}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {J Biol Chem}, volume = {296}, pages = {100578}, abstract = {In eukaryotes, various alternative translation initiation mechanisms have been unveiled for the translation of specific mRNAs. Some do not conform to the conventional scanning-initiation model. Translation initiation of histone H4 mRNA combines both canonical (cap-dependent) and viral initiation strategies (no-scanning, internal recruitment of initiation factors). Specific H4 mRNA structures tether the translation machinery directly onto the initiation codon and allow massive production of histone H4 during the S phase of the cell cycle. The human eukaryotic translation initiation factor 3 (eIF3), composed of 13 subunits (a-m), was shown to selectively recruit and control the expression of several cellular mRNAs. Whether eIF3 mediates H4 mRNA translation remains to be elucidated. Here, we report that eIF3 binds to a stem-loop structure (eIF3-BS) located in the coding region of H4 mRNA. Combining cross-linking and ribonucleoprotein immunoprecipitation experiments in vivo and in vitro, we also found that eIF3 binds to H1, H2A, H2B and H3 histone mRNAs. We identified direct contacts between eIF3c, d, e, g subunits and histone mRNAs but observed distinct interaction patterns to each histone mRNA. Our results show that eIF3 depletion in vivo reduces histone mRNA binding and modulates histone neosynthesis, suggesting that synthesis of histones is sensitive to the levels of eIF3. Thus, we provide evidence that eIF3 acts as a regulator of histone translation.}, note = {1083-351X (Electronic) 0021-9258 (Linking) Journal Article}, keywords = {ERIANI, eukaryotic initiation factor, histone mRNA, protein synthesis, RNA protein interaction, RNA structure, translation initiation, translation regulation}, pubstate = {published}, tppubtype = {article} } @article{Westhof2021, title = {An RNA-centric historical narrative around the Protein Data Bank}, author = {E Westhof and N B Leontis}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33744291}, doi = {10.1016/j.jbc.2021.100555}, isbn = {33744291}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {J Biol Chem}, volume = {296}, pages = {100555}, abstract = {Some of the amazing contributions brought to the scientific community by the PDB are described. The focus is on nucleic acid structures with a bias towards RNA. The evolution and key roles in science of the PDB and other structural databases for nucleic acids illustrate how small initial ideas can become huge and indispensable resources with the unflinching willingness of scientists to cooperate globally. The progress in the understanding of the molecular interactions driving RNA architectures followed the rapid increase in RNA structures in the PDB. That increase was consecutive to improvements in chemical synthesis and purification of RNA molecules, as well as in biophysical methods for structure determination and computer technology. The RNA modeling efforts from the early beginnings are also described together with their links to the state of structural knowledge and technological development. Structures of RNA and of its assemblies are physical objects which, together with genomic data, allow us to integrate present-day biological functions and the historical evolution in all living species on earth.}, note = {1083-351X (Electronic) 0021-9258 (Linking) Journal Article Review}, keywords = {Computational Biology, Databases, modelling, Protein Data Bank, RNA, Structural biology, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{C2021c, title = {Structure-Switching RNAs: From Gene Expression Regulation to Small Molecule Detection}, author = {C Husser and N Dentz and M Ryckelynck}, url = {https://doi.org/10.1002/sstr.202000132}, doi = {10.1002/sstr.202000132}, year = {2021}, date = {2021-01-01}, journal = {Small Structures}, volume = {2}, number = {4}, pages = {2000132}, abstract = {RNA is instrumental to cell life in many aspects, especially gene expression regulation. Among the various known regulatory RNAs, riboswitches are particularly interesting cis‐acting molecules as they do not need cellular factor to achieve their function and are therefore highly portable from one organism to the other. These molecules usually found in the 5′ untranslated region of bacterial messenger RNAs are able to specifically sense a target ligand via an aptamer domain prior to transmitting this recognition event to an expression platform that turns on, or off, the expression of downstream genes. In addition to their obvious scientific interest, these modular molecules can also serve for the development of synthetic RNA devices with applications ranging from the control of transgene expression in gene therapy to the specific biosensing of small molecules. The engineering of such nanomachines is greatly facilitated by the proper understanding of their structure as well as the introduction of new technologies. Herein, a general overview of the current knowledge on natural riboswitches prior to explaining the main strategies used to develop new synthetic structure‐switching molecules (riboswitches or biosensors) controlled by small molecules is given.}, keywords = {biosensing, Gene Expression Regulation, riboswitches, RNA aptamers, RYCKELYNCK, Synthetic Biology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Stupfler2021, title = {Degradation-independent inhibition of APOBEC3G by HIV-1 Vif protein}, author = {B Stupfler and C Verriez and S Gallois-Montbrun and R Marquet and J C Paillart}, url = {https://www.mdpi.com/1999-4915/13/4/617}, doi = {10.3390/v13040617}, year = {2021}, date = {2021-01-01}, journal = {Viruses}, volume = {13}, number = {4}, pages = {617}, abstract = {The ubiquitin–proteasome system plays an important role in the cell under normal physiological conditions but also during viral infections. Indeed, many auxiliary proteins from the (HIV-1) divert this system to its own advantage, notably to induce the degradation of cellular restriction factors. For instance, the HIV-1 viral infectivity factor (Vif) has been shown to specifically counteract several cellular deaminases belonging to the apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC3 or A3) family (A3A to A3H) by recruiting an E3-ubiquitin ligase complex and inducing their polyubiquitination and degradation through the proteasome. Although this pathway has been extensively characterized so far, Vif has also been shown to impede A3s through degradation-independent processes, but research on this matter remains limited. In this review, we describe our current knowledge regarding the degradation-independent inhibition of A3s, and A3G in particular, by the HIV-1 Vif protein, the molecular mechanisms involved, and highlight important properties of this small viral protein.}, keywords = {APOBEC3G, deamination, encapsidation, HIV, MARQUET, PAILLART, proteasome, RNP granules, Translation, ubiquitin, Unité ARN, vif}, pubstate = {published}, tppubtype = {article} } @article{Chagot2021, title = {Application of NMR Spectroscopy to Determine the 3D Structure of Small Non-Coding RNAs}, author = {M E Chagot and M Quinternet and X Manival and I Lebars}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33792884}, doi = {10.1007/978-1-0716-1386-3_19}, isbn = {33792884}, year = {2021}, date = {2021-01-01}, journal = {Methods Mol Biol}, volume = {2300}, pages = {251-266}, abstract = {Many RNA architectures were discovered to be involved in a wide range of essential biological processes in all organisms from carrying genetic information to gene expression regulation. The remarkable ability of RNAs to adopt various architectures depending on their environment enables the achievement of their myriads of biological functions. Nuclear Magnetic Resonance (NMR) is a powerful technique to investigate both their structure and dynamics. NMR is also a key tool for studying interactions between RNAs and their numerous partners such as small molecules, ions, proteins, or other nucleic acids.In this chapter, to illustrate the use of NMR for 3D structure determination of small noncoding RNA, we describe detailed methods that we used for the yeast C/D box small nucleolar RNA U14 from sample preparation to 3D structure calculation.}, note = {1940-6029 (Electronic) 1064-3745 (Linking) Journal Article}, keywords = {3D structure, dynamics, ENNIFAR, NMR, RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Andre2021, title = {The selective advantage of facultative anaerobes relies on their unique ability to cope with changing oxygen levels during infection}, author = {A C Andre and L Debande and B S Marteyn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33813807}, doi = {10.1111/cmi.13338}, isbn = {33813807}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Cell Microbiol}, volume = {23}, number = {8}, pages = {e13338}, abstract = {Bacteria, including those that are pathogenic, have been generally classified according to their ability to survive and grow in the presence or absence of oxygen: aerobic and anaerobic bacteria, respectively. Strict aerobes require oxygen to grow (e.g. Neisseria), and strict anaerobes grow exclusively without, and do not survive oxygen exposure (e.g. Clostridia); aerotolerant bacteria (e.g. Lactobacilli) are insensitive to oxygen exposure. Facultative anaerobes (e.g. E. coli) have the unique ability to grow in the presence or in the absence of oxygen. and are thus well-adapted to these changing conditions which may constitute an underestimated selective advantage for infection. In the WHO antibiotic-resistant "priority pathogens" list, facultative anaerobes are overrepresented (8 among 12 listed pathogens), consistent with clinical studies performed in populations particularly susceptible to infectious diseases. Bacteria aerobic respiratory chain plays a central role in oxygen consumption, leading to the formation of hypoxic infectious sites (infectious hypoxia). Facultative anaerobes have developed a wide diversity of aerotolerance and anaerotolerance strategies in vivo. However, at a single cell level, the modulation of the intracellular oxygen level in host infected cells remains elusive and will be discussed in this review. In conclusion, the ability of facultative bacteria to evolve in the presence or the absence of oxygen is essential for their virulence strategy and constitute a selective advantage. This article is protected by copyright. All rights reserved.}, note = {1462-5822 (Electronic) 1462-5814 (Linking) Journal Article Review}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{deWijn2021, title = {Crystallization and Structural Determination of an Enzyme:Substrate Complex by Serial Crystallography in a Versatile Microfluidic Chip}, author = {R de Wijn and K Rollet and V Olieric and O Hennig and N Thome and C Nous and C Paulus and B Lorber and H Betat and M Morl and C Sauter}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33818565}, doi = {10.3791/61972}, isbn = {33818565}, year = {2021}, date = {2021-01-01}, journal = {J Vis Exp}, number = {169}, abstract = {The preparation of well diffracting crystals and their handling before their X-ray analysis are two critical steps of biocrystallographic studies. We describe a versatile microfluidic chip that enables the production of crystals by the efficient method of counter-diffusion. The convection-free environment provided by the microfluidic channels is ideal for crystal growth and useful to diffuse a substrate into the active site of the crystalline enzyme. Here we applied this approach to the CCA-adding enzyme of the psychrophilic bacterium Planococcus halocryophilus in the presented example. After crystallization and substrate diffusion/soaking, the crystal structure of the enzyme:substrate complex was determined at room temperature by serial crystallography and the analysis of multiple crystals directly inside the chip. The whole procedure preserves the genuine diffraction properties of the samples because it requires no crystal handling.}, note = {1940-087X (Electronic) 1940-087X (Linking) Journal Article Video-Audio Media}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Injarabian2021, title = {Reducing neutrophil exposure to oxygen allows their basal state maintenance}, author = {L Injarabian and J Skerniskyte and Q G Gianetto and V Witko-Sarsat and B S Marteyn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33811670}, doi = {10.1111/imcb.12458}, isbn = {33811670}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Immunol Cell Biol}, volume = {99}, number = {7}, pages = {782-789}, abstract = {Neutrophils are the most abundant circulating white blood cells and are the central players of the innate immune response. During their lifecycle, neutrophils mainly evolve under low oxygen conditions (0.1-4% O2), to which they are well adapted. Neutrophils are atypical cells since they are highly glycolytic, and susceptible to oxygen exposure, which induces their activation and death, through mechanisms, which remain currently elusive. Nevertheless, nearly all studies conducted on neutrophils are carried out under atmospheric oxygen (21%), corresponding to hyperoxia. Here, we investigated the impact of hyperoxia during neutrophil purification and culture on neutrophil viability, activation and cytosolic protein content. We demonstrate that neutrophil hyper-activation (CD62L shedding) is induced during culture under hyperoxic conditions (24 h), compared to neutrophils cultured under anoxic conditions. Spontaneous neutrophil extracellular trap (NET) formation is observed when neutrophils face hyperoxia during purification or culture. In addition, we show that maintaining neutrophils in autologous plasma is the preferred strategy to maintain their basal state. Our results show that manipulating neutrophils under hyperoxic conditions leads to the loss of 57 cytosolic proteins during purification, while it does not lead to an immediate impact on neutrophil activation (CD11b(high), CD54(high), CD62L(neg)) or viability (DAPI(+)). We identified two clusters of proteins belonging to the cholesterol metabolism and to the complement and coagulation cascade pathways, which are highly susceptible to neutrophil oxygen exposure during neutrophil purification. In conclusion, protecting neutrophil from oxygen during their purification and culture is recommended to avoid activation and prevent the alteration cytosolic protein composition.}, note = {1440-1711 (Electronic) 0818-9641 (Linking) Journal Article}, keywords = {Activation, anoxia, hyperoxia, MARTEYN, neutrophils, Unité ARN, viability}, pubstate = {published}, tppubtype = {article} } @article{, title = {Macromolecular interactions in vitro, comparing classical and novel approaches}, author = {C Velours and M Aumont-Nicaise and S Uebel and P England and A Velazquez-Campoy and D Stroebel and G Bec and P Soule and C Quetard and C Ebel and A Roussel and J B Charbonnier and P F Varela}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33792745}, doi = {10.1007/s00249-021-01517-5}, isbn = {33792745}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Eur Biophys J}, volume = {50}, number = {3-4}, pages = {313-330}, abstract = {Biophysical quantification of protein interactions is central to unveil the molecular mechanisms of cellular processes. Researchers can choose from a wide panel of biophysical methods that quantify molecular interactions in different ways, including both classical and more novel techniques. We report the outcome of an ARBRE-MOBIEU training school held in June 2019 in Gif-sur-Yvette, France (https://mosbio.sciencesconf.org/). Twenty European students benefited from a week's training with theoretical and practical sessions in six complementary approaches: (1) analytical ultracentrifugation with or without a fluorescence detector system (AUC-FDS), (2) isothermal titration calorimetry (ITC), (3) size exclusion chromatography coupled to multi-angle light scattering (SEC-MALS), (4) bio-layer interferometry (BLI), (5) microscale thermophoresis (MST) and, (6) switchSENSE. They implemented all these methods on two examples of macromolecular interactions with nanomolar affinity: first, a protein-protein interaction between an artificial alphaRep binder, and its target protein, also an alphaRep; second, a protein-DNA interaction between a DNA repair complex, Ku70/Ku80 (hereafter called Ku), and its cognate DNA ligand. We report the approaches used to analyze the two systems under study and thereby showcase application of each of the six techniques. The workshop provided students with improved understanding of the advantages and limitations of different methods, enabling future choices concerning approaches that are most relevant or informative for specific kinds of sample and interaction.}, note = {1432-1017 (Electronic) 0175-7571 (Linking) Journal Article}, keywords = {Artificial binders, Double-stranded DNA breaks repair factors, ENNIFAR, Macromolecular interactions, Molecular scale biophysics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{Alghoul2021, title = {RNA Secondary Structure Study by Chemical Probing Methods Using DMS and CMCT}, author = {F Alghoul and G Eriani and F Martin}, editor = {M Rederstorff}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33792883}, doi = {10.1007/978-1-0716-1386-3_18}, isbn = {978-1-0716-1385-6/ISSN}, year = {2021}, date = {2021-01-01}, booktitle = {Methods Mol Biol}, volume = {2300}, pages = {241-250}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {RNA folds into secondary structures that can serve in understanding various RNA functions (Weeks KM. Curr Opin Struct Biol 20(3):295-304, 2010). Chemical probing is a method that enables the characterization of RNA secondary structures using chemical reagents that specifically modify RNA nucleotides that are located in single-stranded areas. In our protocol, we used Dimethyl Sulfate (DMS) and Cyclohexyl-3-(2-Morpholinoethyl) Carbodiimide metho-p-Toluene sulfonate (CMCT) that are both base-specific modifying reagents (Behm-Ansmant I, et al. J Nucleic Acids 2011:408053, 2011). These modifications are mapped by primer extension arrests using 5' fluorescently labeled primers. In this protocol, we show a comprehensive method to identify RNA secondary structures in vitro using fluorescently labeled oligos. To demonstrate the efficiency of the method, we give an example of a structure we have designed which corresponds to a part of the 5'-UTR regulatory element called Translation Inhibitory Element (TIE) from Hox a3 mRNA (Xue S, et al. Nature 517(7532):33-38, 2015).}, note = {1940-6029 (Electronic) 1064-3745 (Linking) Journal Article}, keywords = {Capillary electrophoresis, chemical probing, CMCT, DMS, ERIANI, Primer extension, QuSHAPE, RNA secondary structure, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{Bouhedda2021, title = {microIVC-Seq: A Method for Ultrahigh-Throughput Development and Functional Characterization of Small RNAs}, author = {F Bouhedda and R Cubi and S Baudrey and M Ryckelynck}, editor = {M Rederstorff}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33792882}, doi = {10.1007/978-1-0716-1386-3_17}, isbn = {978-1-0716-1385-6/ISSN}, year = {2021}, date = {2021-01-01}, booktitle = {Small Non-Coding RNAs}, volume = {2300}, pages = {203-237}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, edition = {Small Non-Coding RNAs}, series = {Methods in Molecular Biology}, abstract = {For a long time, artificial RNAs have been developed by in vitro selection methodologies like Systematic Evolution of Ligands by EXponential enrichment (SELEX). Yet, even though this technology is extremely powerful to isolate specific and high-affinity binders, it is less suited for the isolation of RNAs optimized for more complex functions such as fluorescence emission or multiple-turnover catalysis. Whereas such RNAs should ideally be developed by screening approaches, conventional microtiter plate assays become rapidly cost-prohibitive. However, the advent of droplet-based microfluidics recently enabled us to devise microfluidic-assisted In Vitro Compartmentalization (muIVC), a strongly miniaturized and highly parallelized screening technology allowing to functionally screen millions of mutants in a single day while using a very low amount of reagent. Used in combination with high-throughput sequencing, the resulting muIVC-seq pipeline described in this chapter now allows rapid and semiautomated screening to be performed at low cost and in an ultrahigh-throughput regime.}, note = {1940-6029 (Electronic) 1064-3745 (Linking) Journal Article}, keywords = {Artificial RNAs, directed evolution, Droplet microfluidics, Functional screening, In vitro selection, RNA, RYCKELYNCK, RYCKELYNCK Artificial RNAs, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{Skerniskyte2021, title = {OmpA Protein-Deficient Acinetobacter baumannii Outer Membrane Vesicles Trigger Reduced Inflammatory Response}, author = {J Skerniskyte and E Karazijaite and A Luciunaite and E Suziedeliene}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33807410}, doi = {10.3390/pathogens10040407}, isbn = {33807410}, year = {2021}, date = {2021-01-01}, journal = {Pathogens}, volume = {10}, number = {4}, pages = {407}, abstract = {Multidrug resistant Acinetobacter baumannii shows a growing number of nosocomial infections worldwide during the last decade. The outer membrane vesicles (OMVs) produced by this bacterium draw increasing attention as a possible treatment target. OMVs have been implicated in the reduction of antibiotic level in the surrounding environment, transfer of virulence factors into the host cells, and induction of inflammatory response. Although the evidence on the involvement of OMVs in A. baumannii pathogenesis is currently growing, their role during inflammation is insufficiently explored. It is likely that bacteria, by secreting OMVs, can expand the area of their exposure and prepare surrounding matrix for infection. Here, we investigated the impact of A. baumannii OMVs on activation of macrophages in vitro. We show that OmpA protein present in A. baumannii OMVs substantially contributes to the proinflammatory response in J774 murine macrophages and to the cell death in both lung epithelium cells and macrophages. The loss of OmpA protein in OMVs, obtained from A. baumannii ompA mutant, resulted in the altered expression of genes coding for IL-6, NLRP3 and IL-1beta proinflammatory molecules in macrophages in vitro. These results imply that OmpA protein in bacterial OMVs could trigger a more intense proinflammatory response.}, note = {2076-0817 (Print) 2076-0817 (Linking) Journal Article}, keywords = {Acinetobacter baumannii, inflammasome, inflammation, Macrophages, MARTEYN, outer membrane vesicles, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Impact of 3-deazapurine nucleobases on RNA properties}, author = {R Bereiter and M Himmelstoss and E Renard and E Mairhofer and M Egger and K Breuker and C Kreutz and E Ennifar and R Micura}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33856457}, isbn = {33856457}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Nucleic Acids Res}, volume = {49}, number = {8}, pages = {4281-4293}, abstract = {Deazapurine nucleosides such as 3-deazaadenosine (c3A) are crucial for atomic mutagenesis studies of functional RNAs. They were the key for our current mechanistic understanding of ribosomal peptide bond formation and of phosphodiester cleavage in recently discovered small ribozymes, such as twister and pistol RNAs. Here, we present a comprehensive study on the impact of c3A and the thus far underinvestigated 3-deazaguanosine (c3G) on RNA properties. We found that these nucleosides can decrease thermodynamic stability of base pairing to a significant extent. The effects are much more pronounced for 3-deazapurine nucleosides compared to their constitutional isomers of 7-deazapurine nucleosides (c7G, c7A). We furthermore investigated base pair opening dynamics by solution NMR spectroscopy and revealed significantly enhanced imino proton exchange rates. Additionally, we solved the X-ray structure of a c3A-modified RNA and visualized the hydration pattern of the minor groove. Importantly, the characteristic water molecule that is hydrogen-bonded to the purine N3 atom and always observed in a natural double helix is lacking in the 3-deazapurine-modified counterpart. Both, the findings by NMR and X-ray crystallographic methods hence provide a rationale for the reduced pairing strength. Taken together, our comparative study is a first major step towards a comprehensive understanding of this important class of nucleoside modifications.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {CDX2 regulates ACE expression in blood development and leukemia cells}, author = {R El Omar and E Julien and K Biasch and B Guffroy and B Lioure and L Vallat and I Gross and C Domon-Dell and F Lanza and C Gachet and M Negroni and J N Freund and M Tavian}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33843985}, doi = {10.1182/bloodadvances.2020003563}, isbn = {33843985}, year = {2021}, date = {2021-01-01}, journal = {Blood Adv}, volume = {5}, number = {7}, pages = {2012-2016}, note = {2473-9537 (Electronic) 2473-9529 (Linking) Journal Article}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Biophysical Studies of the Binding of Viral RNA with the 80S Ribosome Using switchSENSE}, author = {E Schenckbecher and G Bec and T Sakamoto and B Meyer and E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33877606}, doi = {10.1007/978-1-0716-1197-5_15}, isbn = {33877606}, year = {2021}, date = {2021-01-01}, journal = {Methods Mol Biol}, volume = {2263}, pages = {341-350}, abstract = {Translation initiation, in both eukaryotes and bacteria, requires essential elements such as mRNA, ribosome, initiator tRNA, and a set of initiation factors. For each domain of life, canonical mechanisms and signals are observed to initiate protein synthesis. However, other initiation mechanism can be used, especially in viral mRNAs. Some viruses hijack cellular machinery to translate some of their mRNAs through a noncanonical initiation pathway using internal ribosome entry site (IRES), a highly structured RNAs which can directly recruit the ribosome with a restricted set of initiation factors, and in some cases even without cap and initiator tRNA. In this chapter, we describe the use of biosensors relying on electro-switchable nanolevers using the switchSENSE((R)) technology, to investigate kinetics of the intergenic (IGR) IRES of the cricket paralysis virus (CrPV) binding to 80S yeast ribosome. This study provides a proof of concept for the application of this method on large complexes.}, note = {1940-6029 (Electronic) 1064-3745 (Linking) Journal Article}, keywords = {Biophysics, ENNIFAR, Kinetics, Ribosome, RNA, switchSENSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 Capsid Core: A Bullet to the Heart of the Target Cell}, author = {E Toccafondi and D Lener and M Negroni}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33868211}, doi = {10.3389/fmicb.2021.652486}, isbn = {33868211}, year = {2021}, date = {2021-01-01}, journal = {Front Microbiol}, volume = {12}, pages = {652486}, abstract = {The first step of the intracellular phase of retroviral infection is the release of the viral capsid core in the cytoplasm. This structure contains the viral genetic material that will be reverse transcribed and integrated into the genome of infected cells. Up to recent times, the role of the capsid core was considered essentially to protect this genetic material during the earlier phases of this process. However, increasing evidence demonstrates that the permanence inside the cell of the capsid as an intact, or almost intact, structure is longer than thought. This suggests its involvement in more aspects of the infectious cycle than previously foreseen, particularly in the steps of viral genomic material translocation into the nucleus and in the phases preceding integration. During the trip across the infected cell, many host factors are brought to interact with the capsid, some possessing antiviral properties, others, serving as viral cofactors. All these interactions rely on the properties of the unique component of the capsid core, the capsid protein CA. Likely, the drawback of ensuring these multiple functions is the extreme genetic fragility that has been shown to characterize this protein. Here, we recapitulate the busy agenda of an HIV-1 capsid in the infectious process, in particular in the light of the most recent findings.}, note = {1664-302X (Print) 1664-302X (Linking) Journal Article Review}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A multi-laboratory benchmark study of isothermal titration calorimetry (ITC) using Ca(2+) and Mg(2+) binding to EDTA}, author = {A Velazquez-Campoy and B Claro and O Abian and J Horing and L Bourlon and R Claveria-Gimeno and E Ennifar and P England and J B Chaires and D Wu and G Piszczek and C Brautigam and S C Tso and H Zhao and P Schuck and S Keller and M Bastos}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33864101}, doi = {10.1007/s00249-021-01523-7}, isbn = {33864101}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Eur Biophys J}, volume = {50}, number = {3-4}, pages = {429-451}, abstract = {A small-scale ITC benchmarking study was performed involving 9 biophysics laboratories/facilities, to evaluate inter-laboratory and intra-laboratory basal levels of uncertainty. Our prime goal was to assess a number of important factors that can influence both the data gathered by this technique and the thermodynamic parameter values derived therefrom. In its first part, the study involved 5 laboratories and 13 different instruments, working with centrally prepared samples and the same experimental protocol. The second part involved 4 additional laboratories and 6 more instruments, where the users prepared their own samples according to provided instructions and did the experiments following the same protocol as in the first part. The study design comprised: (1) selecting a minimal set of laboratories; (2) providing very stable samples; (3) providing samples not requiring preparation or manipulation; and (4) providing a well-defined and detailed experimental protocol. Thus, we were able to assess: (i) the variability due to instrument and data analysis performed by each user on centrally prepared samples; (ii) the comparability of data retrieved when using 4 different software packages to analyze the same data, besides the data analysis carried out by the different users on their own experimental results; and (iii) the variability due to local sample preparation (second part of the study). Individual values, as well as averages and standard deviations for the binding parameters for EDTA-cation interaction, were used as metrics for comparing the equilibrium association constant (logK), enthalpy of interaction (DeltaH), and the so-called "stoichiometry" (n), a concentration-correction factor.}, note = {1432-1017 (Electronic) 0175-7571 (Linking) Journal Article}, keywords = {Benchmark study, Data treatment, ENNIFAR, Isothermal Titration Calorimetry (ITC), Ligand-binding, Sample preparation, Standard reaction, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Goettsch2021, title = {ITN-VIROINF: Understanding (Harmful) Virus-Host Interactions by Linking Virology and Bioinformatics}, author = {W Goettsch and N Beerenwinkel and L Deng and L Dolken and B E Dutilh and F Erhard and L Kaderali and M von Kleist and R Marquet and J Matthijnssens and S McCallin and D McMahon and T Rattei and R P Van Rij and D L Robertson and M Schwemmle and N Stern-Ginossar and M Marz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33925452}, doi = {10.3390/v13050766}, isbn = {33925452}, year = {2021}, date = {2021-01-01}, journal = {Viruses}, volume = {13}, number = {5}, abstract = {Many recent studies highlight the fundamental importance of viruses. Besides their important role as human and animal pathogens, their beneficial, commensal or harmful functions are poorly understood. By developing and applying tailored bioinformatical tools in important virological models, the Marie Sklodowska-Curie Initiative International Training Network VIROINF will provide a better understanding of viruses and the interaction with their hosts. This will open the door to validate methods of improving viral growth, morphogenesis and development, as well as to control strategies against unwanted microorganisms. The key feature of VIROINF is its interdisciplinary nature, which brings together virologists and bioinformaticians to achieve common goals.}, note = {1999-4915 (Electronic) 1999-4915 (Linking) Journal Article}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {microIVC-Useq: a microfluidic-assisted high-throughput functionnal screening in tandem with next generation sequencing and artificial neural network to rapidly characterize RNA molecules}, author = {R Cubi and F Bouhedda and M Collot and A S Klymchenko and M Ryckelynck}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33952671}, doi = {10.1261/rna.077586.120}, isbn = {33952671}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Rna}, volume = {27}, number = {7}, pages = {841-853}, abstract = {The function of an RNA is intimately linked to its three-dimensional structure. X-ray crystallography or NMR allow the fine structural characterization of small RNA (e.g., aptamers) with a precision down to atomic resolution. Yet, these technics are time consuming, laborious and do not inform on mutational robustness and the extent to which a sequence can be modified without altering RNA function, an important set of information to assist RNA engineering. On another hand, thought powerful, in silico predictions still lack the required accuracy. These limitations can be overcome by using high-throughput microfluidic-assisted functional screening technologies, as they allow exploring large mutant libraries in a rapid and cost-effective manner. Among them, we recently introduced the microfluidic-assisted In Vitro Compartmentalization (microIVC), an efficient screening strategy in which reactions are performed in picoliter droplets at rates of several thousand per second. We later improved microIVC efficiency by using in tandem with high throughput sequencing, thought a laborious bioinformatic step was still required at the end of the process. In the present work, we strongly increased the automation level of the pipeline by implementing an artificial neural network enabling unsupervised bioinformatic analysis. We demonstrate the efficiency of this "microIVC-Useq" technology by rapidly identifying a set of sequences readily accepted by a key domain of the light-up RNA aptamer SRB-2. This work not only shed some new light on the way this aptamer can be engineered, but it also allowed us to easily identify new variants with an up-to 10-fold improved performance.}, note = {1469-9001 (Electronic) 1355-8382 (Linking) Journal Article}, keywords = {Bioinformatics, droplet-based microfluidics, high-throughput screening, light-up RNA aptamer, RNA engineering, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Montavon2021, title = {Human DICER helicase domain recruits PKR and modulates its antiviral activity}, author = {T C Montavon and M Baldaccini and M Lefevre and E Girardi and B Chane-Woon-Ming and M Messmer and P Hammann and J Chicher and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=33984068}, doi = {0.1371/journal.ppat.1009549}, isbn = {33984068}, year = {2021}, date = {2021-01-01}, journal = {PLoS Pathog}, volume = {17}, number = {5}, pages = {e1009549}, abstract = {The antiviral innate immune response mainly involves type I interferon (IFN) in mammalian cells. The contribution of the RNA silencing machinery remains to be established, but several recent studies indicate that the ribonuclease DICER can generate viral siRNAs in specific conditions. It has also been proposed that type I IFN and RNA silencing could be mutually exclusive antiviral responses. In order to decipher the implication of DICER during infection of human cells with alphaviruses such as the Sindbis virus and Semliki forest virus, we determined its interactome by proteomics analysis. We show that DICER specifically interacts with several double-stranded RNA binding proteins and RNA helicases during viral infection. In particular, proteins such as DHX9, ADAR-1 and the protein kinase RNA-activated (PKR) are enriched with DICER in virus-infected cells. We demonstrate that the helicase domain of DICER is essential for this interaction and that its deletion confers antiviral properties to this protein in an RNAi-independent, PKR-dependent, manner.}, note = {1553-7374 (Electronic) 1553-7366 (Linking) Journal Article}, keywords = {PFEFFER, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Alghoul2021b, title = {Translation inhibitory elements from Hoxa3 and Hoxa11 mRNAs use uORFs for translation inhibition}, author = {F Alghoul and S Laure and G Eriani and F Martin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34076576}, doi = {10.7554/eLife.66369}, isbn = {34076576}, year = {2021}, date = {2021-01-01}, journal = {Elife}, volume = {10}, abstract = {During embryogenesis, Hox mRNA translation is tightly regulated by a sophisticated molecular mechanism that combines two RNA regulons located in their 5'UTR. First, an internal ribosome entry site (IRES) enables cap-independent translation. The second regulon is a translation inhibitory element or TIE, which ensures concomitant cap-dependent translation inhibition. In this study, we deciphered the molecular mechanisms of mouse Hoxa3 and Hoxa11 TIEs. Both TIEs possess an upstream open reading frame (uORF) that is critical to inhibit cap-dependent translation. However, the molecular mechanisms used are different. In Hoxa3 TIE, we identify an uORF which inhibits cap-dependent translation and we show the requirement of the non-canonical initiation factor eIF2D for this process. The mode of action of Hoxa11 TIE is different, it also contains an uORF but it is a minimal uORF formed by an uAUG followed immediately by a stop codon, namely a 'start-stop'. The 'start-stop' sequence is species-specific and in mice, is located upstream of a highly stable stem loop structure which stalls the 80S ribosome and thereby inhibits cap-dependent translation of Hoxa11 main ORF.}, note = {2050-084X (Electronic) 2050-084X (Linking) Journal Article}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The structure of the mouse ADAT2/ADAT3 complex reveals the molecular basis for mammalian tRNA wobble adenosine-to-inosine deamination}, author = {E Ramos-Morales and E Bayam and J Del-Pozo-Rodriguez and T Salinas-Giege and M Marek and P Tilly and P Wolff and E Troesch and E Ennifar and L Drouard and J D Godin and C Romier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34057470}, doi = {10.1093/nar/gkab436}, isbn = {34057470}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Nucleic Acids Res}, volume = {49}, number = {11}, pages = {6529-6548}, abstract = {Post-transcriptional modification of tRNA wobble adenosine into inosine is crucial for decoding multiple mRNA codons by a single tRNA. The eukaryotic wobble adenosine-to-inosine modification is catalysed by the ADAT (ADAT2/ADAT3) complex that modifies up to eight tRNAs, requiring a full tRNA for activity. Yet, ADAT catalytic mechanism and its implication in neurodevelopmental disorders remain poorly understood. Here, we have characterized mouse ADAT and provide the molecular basis for tRNAs deamination by ADAT2 as well as ADAT3 inactivation by loss of catalytic and tRNA-binding determinants. We show that tRNA binding and deamination can vary depending on the cognate tRNA but absolutely rely on the eukaryote-specific ADAT3 N-terminal domain. This domain can rotate with respect to the ADAT catalytic domain to present and position the tRNA anticodon-stem-loop correctly in ADAT2 active site. A founder mutation in the ADAT3 N-terminal domain, which causes intellectual disability, does not affect tRNA binding despite the structural changes it induces but most likely hinders optimal presentation of the tRNA anticodon-stem-loop to ADAT2.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article}, keywords = {ARN-MS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Evolutionary history expands the range of signaling interactions in hybrid multikinase networks}, author = {P Ortet and S Fochesato and A F Bitbol and D E Whitworth and D Lalaouna and C Santaella and T Heulin and W Achouak and M Barakat}, url = {https://pubmed.ncbi.nlm.nih.gov/34083699/}, doi = {10.1038/s41598-021-91260-w}, isbn = {34083699}, year = {2021}, date = {2021-01-01}, journal = {Sci Rep}, volume = {11}, number = {1}, pages = {11763}, abstract = {Two-component systems (TCSs) are ubiquitous signaling pathways, typically comprising a sensory histidine kinase (HK) and a response regulator, which communicate via intermolecular kinase-to-receiver domain phosphotransfer. Hybrid HKs constitute non-canonical TCS signaling pathways, with transmitter and receiver domains within a single protein communicating via intramolecular phosphotransfer. Here, we report how evolutionary relationships between hybrid HKs can be used as predictors of potential intermolecular and intramolecular interactions ('phylogenetic promiscuity'). We used domain-swap genes chimeras to investigate the specificity of phosphotransfer within hybrid HKs of the GacS-GacA multikinase network of Pseudomonas brassicacearum. The receiver domain of GacS was replaced with those from nine donor hybrid HKs. Three chimeras with receivers from other hybrid HKs demonstrated correct functioning through complementation of a gacS mutant, which was dependent on strains having a functional gacA. Formation of functional chimeras was predictable on the basis of evolutionary heritage, and raises the possibility that HKs sharing a common ancestor with GacS might remain components of the contemporary GacS network. The results also demonstrate that understanding the evolutionary heritage of signaling domains in sophisticated networks allows their rational rewiring by simple domain transplantation, with implications for the creation of designer networks and inference of functional interactions.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {2'-O-Trifluoromethylated RNA - a powerful modification for RNA chemistry and NMR spectroscopy}, author = {M Himmelstoss and K Erharter and E Renard and E Ennifar and C Kreutz and R Micura}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34094374}, doi = {10.1039/d0sc04520a}, isbn = {34094374}, year = {2021}, date = {2021-01-01}, journal = {Chem Sci}, volume = {11}, number = {41}, pages = {11322-11330}, abstract = {New RNA modifications are needed to advance our toolbox for targeted manipulation of RNA. In particular, the development of high-performance reporter groups facilitating spectroscopic analysis of RNA structure and dynamics, and of RNA-ligand interactions has attracted considerable interest. To this end, fluorine labeling in conjunction with (19)F-NMR spectroscopy has emerged as a powerful strategy. Appropriate probes for RNA previously focused on single fluorine atoms attached to the 5-position of pyrimidine nucleobases or at the ribose 2'-position. To increase NMR sensitivity, trifluoromethyl labeling approaches have been developed, with the ribose 2'-SCF3 modification being the most prominent one. A major drawback of the 2'-SCF3 group, however, is its strong impact on RNA base pairing stability. Interestingly, RNA containing the structurally related 2'-OCF3 modification has not yet been reported. Therefore, we set out to overcome the synthetic challenges toward 2'-OCF3 labeled RNA and to investigate the impact of this modification. We present the syntheses of 2'-OCF3 adenosine and cytidine phosphoramidites and their incorporation into oligoribonucleotides by solid-phase synthesis. Importantly, it turns out that the 2'-OCF3 group has only a slight destabilizing effect when located in double helical regions which is consistent with the preferential C3'-endo conformation of the 2'-OCF3 ribose as reflected in the (3) J (H1'-H2') coupling constants. Furthermore, we demonstrate the exceptionally high sensitivity of the new label in (19)F-NMR analysis of RNA structure equilibria and of RNA-small molecule interactions. The study is complemented by a crystal structure at 0.9 A resolution of a 27 nt hairpin RNA containing a single 2'-OCF3 group that well integrates into the minor groove. The new label carries high potential to outcompete currently applied fluorine labels for nucleic acid NMR spectroscopy because of its significantly advanced performance.}, note = {2041-6520 (Print) 2041-6520 (Linking) Journal Article}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Vitamin C levels in a Central-African mother-infant cohort: Does hypovitaminosis C increase the risk of enteric infections?}, author = {V Moya-Alvarez and J J Koyembi and L M Kaye and J R Mbecko and H Sanke-Waigana and S G Djorie and Y T Nyasenu and D Mad-Bondo and J B Kongoma and S Nakib and Y Madec and G Ulmann and N Neveux and P J Sansonetti and M Vray and B Marteyn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34137176}, doi = {10.1111/mcn.13215}, isbn = {34137176}, year = {2021}, date = {2021-01-01}, journal = {Matern Child Nutr}, pages = {e13215}, abstract = {In the MITICA (Mother-to-Infant TransmIssion of microbiota in Central-Africa) study, 48 mothers and their 50 infants were followed from delivery to 6 months between December 2017 and June 2019 in Bangui (Central-African Republic). Blood tests and stool analyses were performed in mothers at delivery, and their offspring at birth, 11 weeks and 25 weeks. Stool cultures were performed in specific growth media for Salmonella, Shigella, E. coli, Campylobacter, Enerobacter, Vibrio cholerae, Citrobacter and Klebsiella, as well as rotavirus, yeasts and parasitological exams. The median vitamin C levels in mothers at delivery were 15.3 mumol/L (inter-quartile-range [IQR] 6.2-27.8 mumol/L). In infants, the median vitamin C levels at birth were 35.2 mumol/L (IQR 16.5-63.9 mumol/L). At 11 and 25 weeks, the median vitamin C levels were 41.5 mumol/L (IQR 18.7-71.6 mumol/L) and 18.2 mumol/L (IQR 2.3-46.6 mumol/L), respectively. Hypovitaminosis C was defined as seric vitamin C levels <28 mumol/L and vitamin C deficiency was defined as vitamin C levels <11 mumol/L according to the WHO definition. In mothers, the prevalence of hypovitaminosis-C and vitamin C deficiency at delivery was 34/45 (75.6%) and 19/45 (42.2%), respectively. In infants, the prevalence of hypovitaminosis-C and vitamin C deficiency at 6 months was 18/33 (54.6%) and 11/33 (33.3%), respectively. Vitamin C levels in mothers and infants were correlated at birth (Spearman's rho = 0.5; P value = 0.002), and infants had significantly higher levels of vitamin C (median = 35.2 mumol/L; IQR 16.5-63.9 mumol/L), compared to mothers (median = 15.3 mumol/L; IQR 6.2-27.8 mumol/L; P value <0.001). The offspring of vitamin C-deficient mothers had significantly lower vitamin C levels at delivery (median = 18.7 mumol/L; IQR 13.3-30.7 mumol/L), compared to the offspring of non-deficient mothers (median = 62.2 mumol/L; IQR 34.6-89.2 mumol/L; P value <0.001). Infants with hypovitaminosis-C were at significantly higher risk of having a positive stool culture during the first 6 months of life (adjusted OR = 5.3, 95% CI 1.1; 26.1; P value = 0.038).}, note = {1740-8709 (Electronic) 1740-8695 (Linking) Journal Article}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Editorial: Interview With the Translational Apparatus: Stories of Intriguing Circuits and Mechanisms to Regulate Translation in Bacteria}, author = {A M Giuliodori and S Marzi}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34220790}, doi = {10.3389/fmicb.2021.707354}, isbn = {34220790}, year = {2021}, date = {2021-01-01}, journal = {Front Microbiol}, volume = {12}, pages = {707354}, note = {1664-302X (Print) 1664-302X (Linking) Editorial}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comparative study on tertiary contacts and folding of RNase P RNAs from a psychrophilic, a mesophilic/radiation-resistant and a thermophilic bacterium}, author = {M Marszalkowski and A Werner and R Feltens and D Helmecke and M Gossringer and E Westhof and R K Hartmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34266994}, doi = {10.1261/rna.078735.121}, isbn = {34266994}, year = {2021}, date = {2021-01-01}, journal = {Rna}, abstract = {In most bacterial type A RNase P RNAs (P RNAs), two major loop-helix tertiary contacts (L8-P4 and L18-P8) help to orient the two independently folding S- and C-domains for concerted recognition of precursor tRNA substrates. Here, we analyze the effects of mutations in these tertiary contacts in P RNAs from three different species: (i) the psychrophilic bacterium Pseudoalteromonas translucida (Ptr), (ii) the mesophilic radiation-resistant bacterium Deinococcus radiodurans (Dra), and (iii) the thermophilic bacterium Thermus thermophilus (Tth). We show by UV melting experiments that simultaneous disruption of these two interdomain contacts has a stabilizing effect on all three P RNAs. This can be inferred from reduced RNA unfolding at lower temperatures and a more concerted unfolding at higher temperatures. Thus, when the two domains tightly interact via the tertiary contacts, one domain facilitates structural transitions in the other. P RNA mutants with disrupted interdomain contacts showed severe kinetic defects that were most pronounced upon simultaneous disruption of the L8-P4 and L18-P8 contacts. At 37 degrees C, the mildest effects were observed for the thermostable Tth RNA. A third interdomain contact, L9-P1, makes only a minor contribution to P RNA tertiary folding. Furthermore, D. radiodurans RNase P RNA forms an additional pseudoknot structure between the P9 and P12 of its S-domain. This interaction was found to be particularly crucial for RNase P holoenzyme activity at near-physiological Mg2+ concentrations (2 mM). We further analyzed an exceptionally stable folding trap of the G,C-rich Tth P RNA.}, note = {1469-9001 (Electronic) 1355-8382 (Linking) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Assembling the Current Pieces: The Puzzle of RNA-Mediated Regulation in Staphylococcus aureus}, author = {L Barrientos and N Mercier and D Lalaouna and I Caldelari}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34367109}, doi = {10.3389/fmicb.2021.706690}, isbn = {34367109}, year = {2021}, date = {2021-01-01}, journal = {Front Microbiol}, volume = {12}, pages = {706690}, abstract = {The success of the major opportunistic human Staphylococcus aureus relies on the production of numerous virulence factors, which allow rapid colonization and dissemination in any tissues. Indeed, regulation of its virulence is multifactorial, and based on the production of transcriptional factors, two-component systems (TCS) and small regulatory RNAs (sRNAs). Advances in high-throughput sequencing technologies have unveiled the existence of hundreds of potential RNAs with regulatory functions, but only a fraction of which have been validated in vivo. These discoveries have modified our thinking and understanding of bacterial physiology and virulence fitness by placing sRNAs, alongside transcriptional regulators, at the center of complex and intertwined regulatory networks that allow S. aureus to rapidly adapt to the environmental cues present at infection sites. In this review, we describe the recently acquired knowledge of characterized regulatory RNAs in S. aureus that are associated with metal starvation, nutrient availability, stress responses and virulence. These findings highlight the importance of sRNAs for the comprehension of S. aureus infection processes while raising questions about the interplay between these key regulators and the pathways they control.}, note = {1664-302X (Print) 1664-302X (Linking) Journal Article Review}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Burning the Candle at Both Ends: Have Exoribonucleases Driven Divergence of Regulatory RNA Mechanisms in Bacteria?}, author = {D G Mediati and D Lalaouna and J J Tree}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34372700}, doi = {10.1128/mBio.01041-21}, isbn = {34372700}, year = {2021}, date = {2021-01-01}, journal = {mBio}, pages = {e0104121}, abstract = {Regulatory RNAs have emerged as ubiquitous gene regulators in all bacterial species studied to date. The combination of sequence-specific RNA interactions and malleable RNA structure has allowed regulatory RNA to adopt different mechanisms of gene regulation in a diversity of genetic backgrounds. In the model Gammaproteobacteria Escherichia coli and Salmonella, the regulatory RNA chaperone Hfq appears to play a global role in gene regulation, directly controlling approximately 20 to 25% of the entire transcriptome. While the model Firmicutes Bacillus subtilis and Staphylococcus aureus encode a Hfq homologue, its role has been significantly depreciated. These bacteria also have marked differences in RNA turnover. E. coli and Salmonella degrade RNA through internal endonucleolytic and 3'-->5' exonucleolytic cleavage that appears to allow transient accumulation of mRNA 3' UTR cleavage fragments that contain stabilizing 3' structures. In contrast, B. subtilis and S. aureus are able to exonucleolytically attack internally cleaved RNA from both the 5' and 3' ends, efficiently degrading mRNA 3' UTR fragments. Here, we propose that the lack of 5'-->3' exoribonuclease activity in Gammaproteobacteria has allowed the accumulation of mRNA 3' UTR ends as the "default" setting. This in turn may have provided a larger pool of unconstrained RNA sequences that has fueled the expansion of Hfq function and small RNA (sRNA) regulation in E. coli and Salmonella. Conversely, the exoribonuclease RNase J may be a significant barrier to the evolution of 3' UTR sRNAs in B. subtilis and S. aureus that has limited the pool of RNA ligands available to Hfq and other sRNA chaperones, depreciating their function in these model Firmicutes.}, note = {2150-7511 (Electronic) Journal Article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Safe and easy in vitro evaluation of tmRNA-SmpB-mediated trans-translation from ESKAPE pathogenic bacteria}, author = {M Thepaut and R Campos-Silva and E Renard and F Barloy-Hubler and E Ennifar and D Boujard and R Gillet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34353925}, doi = {10.1261/rna.078773.121}, isbn = {34353925}, year = {2021}, date = {2021-01-01}, journal = {Rna}, abstract = {In bacteria, trans-translation is the major quality control system for rescuing stalled ribosomes. It is mediated by tmRNA, a hybrid RNA with properties of both a tRNA and a mRNA, and the small protein SmpB. Because trans-translation is absent in eukaryotes but necessary for bacterial fitness or survival, it is a promising target for the development of novel antibiotics. To facilitate screening of chemical libraries, various reliable in vitro and in vivo systems have been created for assessing trans-translational activity. However, the aim of the current work was to permit the safe and easy in vitro evaluation of trans-translation from pathogenic bacteria, which are obviously the ones we should be targeting. Based on green fluorescent protein (GFP) reassembly during active trans-translation, we have created a cell-free assay adapted to the rapid evaluation of trans-translation in ESKAPE bacteria, with 24 different possible combinations. It can be used for easy high-throughput screening of chemical compounds as well as for exploring the mechanism of trans-translation in these pathogens.}, note = {1469-9001 (Electronic) 1355-8382 (Linking) Journal Article}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Bacterial translation machinery for deliberate mistranslation of the genetic code}, author = {O Vargas-Rodriguez and A H Badran and K S Hoffman and M Chen and A Crnkovic and Y Ding and J R Krieger and E Westhof and D Soll and S Melnikov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34413202}, doi = {10.1073/pnas.2110797118}, isbn = {34413202}, year = {2021}, date = {2021-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {118}, number = {35}, abstract = {Inaccurate expression of the genetic code, also known as mistranslation, is an emerging paradigm in microbial studies. Growing evidence suggests that many microbial pathogens can deliberately mistranslate their genetic code to help invade a host or evade host immune responses. However, discovering different capacities for deliberate mistranslation remains a challenge because each group of pathogens typically employs a unique mistranslation mechanism. In this study, we address this problem by studying duplicated genes of aminoacyl-transfer RNA (tRNA) synthetases. Using bacterial prolyl-tRNA synthetase (ProRS) genes as an example, we identify an anomalous ProRS isoform, ProRSx, and a corresponding tRNA, tRNA(ProA), that are predominately found in plant pathogens from Streptomyces species. We then show that tRNA(ProA) has an unusual hybrid structure that allows this tRNA to mistranslate alanine codons as proline. Finally, we provide biochemical, genetic, and mass spectrometric evidence that cells which express ProRSx and tRNA(ProA) can translate GCU alanine codons as both alanine and proline. This dual use of alanine codons creates a hidden proteome diversity due to stochastic Ala-->Pro mutations in protein sequences. Thus, we show that important plant pathogens are equipped with a tool to alter the identity of their sense codons. This finding reveals the initial example of a natural tRNA synthetase/tRNA pair for dedicated mistranslation of sense codons.}, note = {1091-6490 (Electronic) 0027-8424 (Linking) Journal Article}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Vilimova2021, title = {Cis regulation within a cluster of viral microRNAs}, author = {M Vilimova and M Contrant and R Randrianjafy and P Dumas and E Elbasani and P M Ojala and S Pfeffer and A Fender}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34417603}, doi = {10.1093/nar/gkab806}, isbn = {34417603}, year = {2021}, date = {2021-01-01}, journal = {Nucleic Acids Res}, abstract = {MicroRNAs (miRNAs) are small regulatory RNAs involved in virtually all biological processes. Although many of them are co-expressed from clusters, little is known regarding the impact of this organization on the regulation of their accumulation. In this study, we set to decipher a regulatory mechanism controlling the expression of the ten clustered pre-miRNAs from Kaposi's sarcoma associated herpesvirus (KSHV). We measured in vitro the efficiency of cleavage of each individual pre-miRNA by the Microprocessor and found that pre-miR-K1 and -K3 were the most efficiently cleaved pre-miRNAs. A mutational analysis showed that, in addition to producing mature miRNAs, they are also important for the optimal expression of the whole set of miRNAs. We showed that this feature depends on the presence of a canonical pre-miRNA at this location since we could functionally replace pre-miR-K1 by a heterologous pre-miRNA. Further in vitro processing analysis suggests that the two stem-loops act in cis and that the cluster is cleaved in a sequential manner. Finally, we exploited this characteristic of the cluster to inhibit the expression of the whole set of miRNAs by targeting the pre-miR-K1 with LNA-based antisense oligonucleotides in cells either expressing a synthetic construct or latently infected with KSHV.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA Modifications in Pathogenic Bacteria: Impact on Host Adaptation and Virulence}, author = {L Antoine and R Bahena-Ceron and H Devi Bunwaree and M Gobry and V Loegler and P Romby and S Marzi}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34440299}, doi = {10.3390/genes12081125}, isbn = {34440299}, year = {2021}, date = {2021-01-01}, journal = {Genes (Basel)}, volume = {12}, number = {8}, abstract = {RNA modifications are involved in numerous biological processes and are present in all RNA classes. These modifications can be constitutive or modulated in response to adaptive processes. RNA modifications play multiple functions since they can impact RNA base-pairings, recognition by proteins, decoding, as well as RNA structure and stability. However, their roles in stress, environmental adaptation and during infections caused by pathogenic bacteria have just started to be appreciated. With the development of modern technologies in mass spectrometry and deep sequencing, recent examples of modifications regulating host-pathogen interactions have been demonstrated. They show how RNA modifications can regulate immune responses, antibiotic resistance, expression of virulence genes, and bacterial persistence. Here, we illustrate some of these findings, and highlight the strategies used to characterize RNA modifications, and their potential for new therapeutic applications.}, note = {2073-4425 (Electronic) 2073-4425 (Linking) Journal Article Review}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Welker2021, title = {Importance of Viral Late Domains in Budding and Release of Enveloped RNA Viruses}, author = {L Welker and J C Paillart and S Bernacchi}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34452424}, doi = {10.3390/v13081559}, isbn = {34452424}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Viruses}, volume = {13}, number = {8}, pages = {1559}, abstract = {Late assembly (L) domains are conserved sequences that are necessary for the late steps of viral replication, acting like cellular adaptors to engage the ESCRT membrane fission machinery that promote virion release. These short sequences, whose mutation or deletion produce the accumulation of immature virions at the plasma membrane, were firstly identified within retroviral Gag precursors, and in a further step, also in structural proteins of many other enveloped RNA viruses including arenaviruses, filoviruses, rhabdoviruses, reoviruses, and paramyxoviruses. Three classes of L domains have been identified thus far (PT/SAP, YPXnL/LXXLF, and PPxY), even if it has recently been suggested that other motifs could act as L domains. Here, we summarize the current state of knowledge of the different types of L domains and their cellular partners in the budding events of RNA viruses, with a particular focus on retroviruses.}, note = {1999-4915 (Electronic) 1999-4915 (Linking) Journal Article Review}, keywords = {PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Antibacterial activity of a dual peptide targeting the Escherichia coli sliding clamp and the ribosome}, author = {C Andre and F Veillard and P Wolff and A M Lobstein and G Compain and C Monsarrat and J M Reichhart and D Y Burnouf and G Guichard and J E Wagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34459830}, doi = {10.1039/d1cb90020j}, isbn = {34459830}, year = {2021}, date = {2021-01-01}, journal = {RSC Chem Biol}, volume = {2}, number = {4}, pages = {1296}, abstract = {[This corrects the article DOI: 10.1039/D0CB00060D.].}, note = {2633-0679 (Electronic) 2633-0679 (Linking) Published Erratum}, keywords = {ARN-MS, ENNIFAR, reichhart, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of host tRNAs preferentially recognized by the Plasmodium surface protein tRip}, author = {M Cela and A Theobald-Dietrich and J Rudinger-Thirion and P Wolff and R Geslain and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34530443}, doi = {10.1093/nar/gkab769}, isbn = {34530443}, year = {2021}, date = {2021-01-01}, journal = {Nucleic Acids Res}, abstract = {Malaria is a life-threatening and devastating parasitic disease. Our previous work showed that parasite development requires the import of exogenous transfer RNAs (tRNAs), which represents a novel and unique form of host-pathogen interaction, as well as a potentially druggable target. This import is mediated by tRip (tRNA import protein), a membrane protein located on the parasite surface. tRip displays an extracellular domain homologous to the well-characterized OB-fold tRNA-binding domain, a structural motif known to indiscriminately interact with tRNAs. We used MIST (Microarray Identification of Shifted tRNAs), a previously established in vitro approach, to systematically assess the specificity of complexes between native Homo sapiens tRNAs and recombinant Plasmodium falciparum tRip. We demonstrate that tRip unexpectedly binds to host tRNAs with a wide range of affinities, suggesting that only a small subset of human tRNAs is preferentially imported into the parasite. In particular, we show with in vitro transcribed constructs that tRip does not bind specific tRNAs solely based on their primary sequence, hinting that post-transcriptional modifications modulate the formation of our host/parasite molecular complex. Finally, we discuss the potential utilization of the most efficient tRip ligands for the translation of the parasite's genetic information.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article}, keywords = {ARN-MS, FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Chemical and Enzymatic Probing of Viral RNAs: From Infancy to Maturity and Beyond}, author = {O Gilmer and E Quignon and A C Jousset and J C Paillart and R Marquet and V Vivet-Boudou}, url = {https://www.mdpi.com/1999-4915/13/10/1894}, doi = {v13101894}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Viruses}, volume = {13}, number = {10}, pages = {1894}, abstract = {RNA molecules are key players in a variety of biological events, and this is particularly true for viral RNAs. To better understand the replication of those pathogens and try to block them, special attention has been paid to the structure of their RNAs. Methods to probe RNA structures have been developed since the 1960s; even if they have evolved over the years, they are still in use today and provide useful information on the folding of RNA molecules, including viral RNAs. The aim of this review is to offer a historical perspective on the structural probing methods used to decipher RNA structures before the development of the selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) methodology and to show how they have influenced the current probing techniques. Actually, these technological breakthroughs, which involved advanced detection methods, were made possible thanks to the development of next-generation sequencing (NGS) but also to the previous works accumulated in the field of structural RNA biology. Finally, we will also discuss how high-throughput SHAPE (hSHAPE) paved the way for the development of sophisticated RNA structural techniques.}, keywords = {Capillary electrophoresis, chemical probe, enzymatic probe, high-throughput sequencing, MARQUET, mutational profiling, PAILLART, RNA, SHAPE, structure, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Binding of the RNA Chaperone Hfq on Target mRNAs Promotes the Small RNA RyhB-Induced Degradation in Escherichia coli}, author = {D. Lalaouna and K. Prévost and S. Park and T. Chénard and M - P. Bouchard and M - P. Caron and C. K. Vanderpool and J. Fei and E. Massé}, url = {https://www.mdpi.com/2311-553X/7/4/64}, doi = {10.3390/ncrna7040064}, year = {2021}, date = {2021-01-01}, journal = {Non-Coding RNA}, volume = {7}, number = {4}, pages = {64}, abstract = {Many RNA-RNA interactions depend on molecular chaperones to form and remain stable in living cells. A prime example is the RNA chaperone Hfq, which is a critical effector involved in regulatory interactions between small RNAs (sRNAs) and cognate target mRNAs in Enterobacteriaceae. While there is a great deal of in vitro biochemical evidence supporting the model that Hfq enhances rates or affinities of sRNA:mRNA interactions, there is little corroborating in vivo evidence. Here we used in vivo tools including reporter genes, co-purification assays, and super-resolution microscopy to analyze the role of Hfq in RyhB-mediated regulation, and we found that Hfq is often unnecessary for efficient RyhB:mRNA complex formation in vivo. Remarkably, our data suggest that a primary function of Hfq is to promote RyhB-induced cleavage of mRNA targets by RNase E. Moreover, our work indicates that Hfq plays a more limited role in dictating regulatory outcomes following sRNAs RybB and DsrA complex formation with specific target mRNAs. Our investigation helps evaluate the roles played by Hfq in some RNA-mediated regulation.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Real-time tracking of root hair nucleus morphodynamics using a microfluidic approach}, author = {G. Singh and D. Pereira and S. Baudrey and E. Hoffmann and M. Ryckelynck and A. Asnacios and M. E. Chaboute}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34562320}, doi = {10.1111/tpj.15511}, isbn = {34562320}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Plant J}, volume = {108}, issue = {2}, pages = {303-313}, abstract = {Root hairs (RHs) are tubular extensions of root epidermal cells that favour nutrient uptake and microbe interactions. RH shows a fast apical growth, constituting a unique single cell model system to analyse cellular morphodynamics. In this context, live cell imaging using microfluidics recently developed to analyze root development is appealing, but high-resolution imaging is still lacking to study accurate spatiotemporal morphodynamics of organelles. Here, we provide a powerful coverslip based microfluidic device (CMD) which enables us to capture high resolution confocal imaging of Arabidopsis RH development with real-time monitoring of nuclear movement and shape changes. To validate the setup, we confirmed the typical RH growth rates and the mean nuclear positioning previously reported with classical methods. Moreover, in order to illustrate the possibilities offered by the CMD, we have compared the real-time variations in the circularity, area, and aspect ratio of nuclei moving in growing and mature RH. Interestingly, we observed higher aspect ratios in the nuclei of mature RH, correlating with higher speeds of nuclear migration. This observation opens the way for further investigations of the effect of mechanical constraints on nuclear shape changes during RH growth and nuclear migration and its role in RH and plant development.}, note = {1365-313X (Electronic) 0960-7412 (Linking) Journal Article}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {CCA-addition in the cold: Structural characterization of the psychrophilic CCA-adding enzyme from the permafrost bacterium Planococcus halocryophilus}, author = {R. De Wijn and K. Rollet and F. G.M.Ernst and K. Wellner and H. Betat and M. Mörl and M. Sauter}, url = {https://www.sciencedirect.com/science/article/pii/S2001037021004402?via%3Dihub}, doi = {10.1016/j.csbj.2021.10.018}, isbn = {ISBN/2001-0370}, year = {2021}, date = {2021-01-01}, journal = {Computational and Structural Biotechnology Journal}, volume = {19}, pages = {5845-5855}, abstract = {CCA-adding enzymes are highly specific RNA polymerases that add and maintain the sequence C-C-A at tRNA 3ム-ends. Recently, we could reveal that cold adaptation of such a polymerase is not only achieved at the expense of enzyme stability, but also at the cost of polymerization fidelity. Enzymes from psychrophilic organisms usually show an increased structural flexibility to enable catalysis at low temperatures. Here, polymerases face a dilemma, as there is a discrepancy between the need for a tightly controlled flexibility during polymerization and an increased flexibility as strategy for cold adaptation. Based on structural and biochemical analyses, we contribute to clarify the cold adaptation strategy of the psychrophilic CCA-adding enzyme from Planococcus halocryophilus, a gram-positive bacterium thriving in the arctic permafrost at low temperatures down to −15 °C. A comparison with the closely related enzyme from the thermophilic bacterium Geobacillus stearothermophilus reveals several features of cold adaptation - a significantly reduced amount of alpha-helical elements in the C-terminal tRNA-binding region and a structural adaptation in one of the highly conserved catalytic core motifs located in the N-terminal catalytic core of the enzyme}, keywords = {CCA-adding enzyme, Cold adaptation, Psychrophilic protein, Psychrophilic RNA polymerase, SAUTER, SAXS, tRNA, Unité ARN, X-ray crystallography}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Short-Range Imbalances in the AMBER Lennard-Jones Potential for (Deoxy)Ribose.Nucleobase Lone-Pair.pi Contacts in Nucleic Acids}, author = {K. Mrazikova and J. Sponer and V. Mlynsky and P. Auffinger and H. Kruse}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34738826}, doi = {10.1021/acs.jcim.1c01047}, isbn = {34738826}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {J Chem Inf Model}, volume = {61}, number = {11}, pages = {5644-5657}, abstract = {The lone-pair.pi (lp.pi) (deoxy)ribose.nucleobase stacking is a recurring interaction in Z-DNA and RNAs that is characterized by sub-van der Waals lp.pi contacts (<3.0 A). It is a part of the structural signature of CpG Z-step motifs in Z-DNA and r(UNCG) tetraloops that are known to behave poorly in molecular dynamics (MD) simulations. Although the exact origin of the MD simulation issues remains unclear, a significant part of the problem might be due to an imbalanced description of nonbonded interactions, including the characteristic lp.pi stacking. To gain insights into the links between lp.pi stacking and MD, we present an in-depth comparison between accurate large-basis-set double-hybrid Kohn-Sham density functional theory calculations DSD-BLYP-D3/ma-def2-QZVPP (DHDF-D3) and data obtained with the nonbonded potential of the AMBER force field (AFF) for NpN Z-steps (N = G, A, C, and U). Among other differences, we found that the AFF overestimates the DHDF-D3 lp.pi distances by approximately 0.1-0.2 A, while the deviation between the DHDF-D3 and AFF descriptions sharply increases in the short-range region of the interaction. Based on atom-in-molecule polarizabilities and symmetry-adapted perturbation theory analysis, we inferred that the DHDF-D3 versus AFF differences partly originate in identical nucleobase carbon atom Lennard-Jones (LJ) parameters despite the presence/absence of connected electron-withdrawing groups that lead to different effective volumes or vdW radii. Thus, to precisely model the very short CpG lp.pi contact distances, we recommend revision of the nucleobase atom LJ parameters. Additionally, we suggest that the large discrepancy between DHDF-D3 and AFF short-range repulsive part of the interaction energy potential may significantly contribute to the poor performances of MD simulations of nucleic acid systems containing Z-steps. Understanding where, and if possible why, the point-charge-type effective potentials reach their limits is vital for developing next-generation FFs and for addressing specific issues in contemporary MD simulations.}, note = {1549-960X (Electronic) 1549-9596 (Linking) Journal Article}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {A case of QARS1 associated epileptic encephalopathy and review of epilepsy in aminoacyl-tRNA synthetase disorders}, author = {D. L. Chan and J. Rudinger-Thirion and M. Frugier and L. G. Riley and G. Ho and K. Kothur and S. S. Mohammad}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34774383}, doi = {10.1016/j.braindev.2021.10.009}, isbn = {34774383}, year = {2021}, date = {2021-01-01}, journal = {Brain Dev}, abstract = {INTRODUCTION: Mutations in QARS1, which encodes human glutaminyl-tRNA synthetase, have been associated with epilepsy, developmental regression, progressive microcephaly and cerebral atrophy. Epilepsy caused by variants in QARS1 is usually drug-resistant and intractable. Childhood onset epilepsy is also reported in various aminoacyl-tRNA synthetase disorders. We describe a case with a milder neurological phenotype than previously reported with QARS1 variants and review the seizure associations with aminoacyl-tRNA synthetase disorders. CASE REPORT: The patient is a 4-year-old girl presenting at 6 weeks of age with orofacial dyskinesia and hand stereotypies. She developed focal seizures at 7 months of age. Serial electroencephalograms showed shifting focality. Her seizures were controlled after introduction of carbamazepine. Progress MRI showed very mild cortical volume loss without myelination abnormalities or cerebellar atrophy. She was found to have novel compound heterozygous variants in QARS1 (NM_005051.2): c.[1132C > T];[1574G > A], p.[(Arg378Cys)];[(Arg525Gln)] originally classified as "variants of uncertain significance" and later upgraded to "likely pathogenic" based on functional testing and updated variant database review. Functional testing showed reduced solubility of the corresponding QARS1 mutants in vitro, but only mild two-fold loss in catalytic efficiency with the c.1132C > T variant and no noted change in tRNA(Gln) aminoacylation with the c.1574G > A variant. CONCLUSION: We describe two QARS1 variants associated with overall conserved tRNA aminoacylation activity but characterized by significantly reduced QARS protein solubility, resulting in a milder clinical phenotype. 86% of previous patients reported with QARS1 had epilepsy and 79% were pharmaco-resistant. We also summarise literature regarding epilepsy in aminoacyl-tRNA synthetase disorders, which is also often early onset, severe and drug-refractory.}, note = {1872-7131 (Electronic) 0387-7604 (Linking) Case Reports}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{meteignier_arabidopsis_2021, title = {Arabidopsis mTERF9 protein promotes chloroplast ribosomal assembly and translation by establishing ribonucleoprotein interactions in vivo}, author = {Louis-Valentin Méteignier and Rabea Ghandour and Aude Zimmerman and Lauriane Kuhn and Jörg Meurer and Reimo Zoschke and Kamel Hammani}, doi = {10.1093/nar/gkaa1244}, issn = {1362-4962}, year = {2021}, date = {2021-01-01}, journal = {Nucleic Acids Research}, volume = {49}, number = {2}, pages = {1114--1132}, abstract = {The mitochondrial transcription termination factor proteins are nuclear-encoded nucleic acid binders defined by degenerate tandem helical-repeats of ∼30 amino acids. They are found in metazoans and plants where they localize in organelles. In higher plants, the mTERF family comprises ∼30 members and several of these have been linked to plant development and response to abiotic stress. However, knowledge of the molecular basis underlying these physiological effects is scarce. We show that the Arabidopsis mTERF9 protein promotes the accumulation of the 16S and 23S rRNAs in chloroplasts, and interacts predominantly with the 16S rRNA in vivo and in vitro. Furthermore, mTERF9 is found in large complexes containing ribosomes and polysomes in chloroplasts. The comprehensive analysis of mTERF9 in vivo protein interactome identified many subunits of the 70S ribosome whose assembly is compromised in the null mterf9 mutant, putative ribosome biogenesis factors and CPN60 chaperonins. Protein interaction assays in yeast revealed that mTERF9 directly interact with these proteins. Our data demonstrate that mTERF9 integrates protein-protein and protein-RNA interactions to promote chloroplast ribosomal assembly and translation. Besides extending our knowledge of mTERF functional repertoire in plants, these findings provide an important insight into the chloroplast ribosome biogenesis.}, keywords = {16S, 23S, Arabidopsis, Arabidopsis Proteins, Chloroplast Proteins, Chloroplasts, Gene Expression Regulation, Organelle Biogenesis, Peptide Termination Factors, Plant, Polyribosomes, PPSE, Protein Biosynthesis, Recombinant Proteins, Ribonucleoproteins, Ribosomal, ribosomes, RNA, Substrate Specificity}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Evaluation of the stereochemical quality of predicted RNA 3D models in the RNA-Puzzles submissions}, author = {F Carrascoza and M Antczak and Z Miao and E Westhof and M Szachniuk}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34819324}, doi = {10.1261/rna.078685.121}, isbn = {34819324}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Rna}, abstract = {In silico prediction is a well-established approach to derive a general shape of an RNA molecule based on its sequence or secondary structure. This paper reports an analysis of the stereochemical quality of the RNA three-dimensional models predicted using dedicated computer programs. The stereochemistry of 1,052 RNA 3D structures, including 1,030 models predicted by fully automated and human-guided approaches within 22 RNA-Puzzles challenges and reference structures, is analysed. The evaluation is based on standards of RNA stereochemistry that the Protein Data Bank requires from deposited experimental structures. Deviations from standard bond lengths and angles, planarity or chirality are quantified. A reduction in the number of such deviations should help in the improvement of RNA 3D structure modelling approaches.}, note = {1469-9001 (Electronic) 1355-8382 (Linking) Journal Article}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Iterative Structure-Based Optimization of Short Peptides Targeting the Bacterial Sliding Clamp}, author = {C. Monsarrat and G. Compain and C. Andre and S. Engilberge and I. Martiel and V. Olieric and P. Wolff and K. Brillet and M. Landolfo and C. Silva da Veiga and J. Wagner and G. Guichard and D. Y. Burnouf}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34806883}, doi = {10.1021/acs.jmedchem.1c00918}, isbn = {34806883}, year = {2021}, date = {2021-01-01}, journal = {J Med Chem}, abstract = {The bacterial DNA sliding clamp (SC), or replication processivity factor, is a promising target for the development of novel antibiotics. We report a structure-activity relationship study of a new series of peptides interacting within the Escherichia coli SC ((Ec)SC) binding pocket. Various modifications were explored including N-alkylation of the peptide bonds, extension of the N-terminal moiety, and introduction of hydrophobic and constrained residues at the C-terminus. In each category, single modifications were identified that increased affinity to (Ec)SC. A combination of such modifications yielded in several cases to a substantially increased affinity compared to the parent peptides with Kd in the range of 30-80 nM. X-ray structure analysis of 11 peptide/(Ec)SC co-crystals revealed new interactions at the peptide-protein interface (i.e., stacking interactions, hydrogen bonds, and hydrophobic contacts) that can account for the improved binding. Several compounds among the best binders were also found to be more effective in inhibiting SC-dependent DNA synthesis.}, note = {1520-4804 (Electronic) 0022-2623 (Linking) Journal Article}, keywords = {ARN-MS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {The HIV-1 Nucleocapsid Regulates Its Own Condensation by Phase-Separated Activity-Enhancing Sequestration of the Viral Protease during Maturation}, author = {S. Lyonnais and S. K. Sadiq and C. Lorca-Oro and L. Dufau and S. Nieto-Marquez and T. Escriba and N. Gabrielli and X. Tan and M. Ouizougun-Oubari and J. Okoronkwo and M. Reboud-Ravaux and J. M. Gatell and R. Marquet and J. C. Paillart and A. Meyerhans and C. Tisne and R. J. Gorelick and G. Mirambeau}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34835118}, doi = {10.3390/v13112312}, isbn = {34835118}, year = {2021}, date = {2021-01-01}, journal = {Viruses}, volume = {13}, number = {11}, abstract = {A growing number of studies indicate that mRNAs and long ncRNAs can affect protein populations by assembling dynamic ribonucleoprotein (RNP) granules. These phase-separated molecular 'sponges', stabilized by quinary (transient and weak) interactions, control proteins involved in numerous biological functions. Retroviruses such as HIV-1 form by self-assembly when their genomic RNA (gRNA) traps Gag and GagPol polyprotein precursors. Infectivity requires extracellular budding of the particle followed by maturation, an ordered processing of approximately 2400 Gag and approximately 120 GagPol by the viral protease (PR). This leads to a condensed gRNA-NCp7 nucleocapsid and a CAp24-self-assembled capsid surrounding the RNP. The choreography by which all of these components dynamically interact during virus maturation is one of the missing milestones to fully depict the HIV life cycle. Here, we describe how HIV-1 has evolved a dynamic RNP granule with successive weak-strong-moderate quinary NC-gRNA networks during the sequential processing of the GagNC domain. We also reveal two palindromic RNA-binding triads on NC, KxxFxxQ and QxxFxxK, that provide quinary NC-gRNA interactions. Consequently, the nucleocapsid complex appears properly aggregated for capsid reassembly and reverse transcription, mandatory processes for viral infectivity. We show that PR is sequestered within this RNP and drives its maturation/condensation within minutes, this process being most effective at the end of budding. We anticipate such findings will stimulate further investigations of quinary interactions and emergent mechanisms in crowded environments throughout the wide and growing array of RNP granules.}, note = {1999-4915 (Electronic) 1999-4915 (Linking) Journal Article}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {The HIV-1 Integrase C-Terminal domain induces TAR RNA structural changes promoting Tat binding}, author = {C. Rocchi and C. Louvat and A. Miele and J. Batisse and C. Guillon and L. Ballut and D. Lener and M. Negroni and M. Ruff and P. Gouet and F. Fiorini}, url = {https://www.biorxiv.org/content/10.1101/2021.10.21.465253v1.full.pdf+html}, doi = {10.1101/2021.10.21.465253}, year = {2021}, date = {2021-01-01}, abstract = {Recent evidence indicated that HIV-1 Integrase (IN) binds genomic viral RNA (gRNA) playing a critical role in viral particle morphogenesis and gRNA stability in host cells. Combining biophysical and biochemical approaches we show that the C-terminal flexible 18-residues tail of IN acts as a sensor of the peculiar apical structure of trans-activation response element RNA (TAR), directly interacting with its hexaloop. We highlighted how the whole IN C-terminal domain, once bound to TAR, can change its structure assisting the binding of Tat, the HIV trans-activator protein, which finally displaces IN from TAR. Our results are consistent with the emerging role of IN in early stage of proviral transcription and suggest new steps of HIV-1 life cycle that can be considered as therapeutic targets.}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Untangling the roles of RNA helicases in antiviral innate immunity}, author = {M. Baldaccini and S. Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34882751}, doi = {10.1371/journal.ppat.1010072}, isbn = {34882751}, year = {2021}, date = {2021-01-01}, journal = {PLoS Pathog}, volume = {17}, number = {12}, pages = {e1010072}, abstract = {One of the first layers of protection that metazoans put in place to defend themselves against viruses rely on the use of proteins containing DExD/H-box helicase domains. These members of the duplex RNA-activated ATPase (DRA) family act as sensors of double-stranded RNA (dsRNA) molecules, a universal marker of viral infections. DRAs can be classified into 2 subgroups based on their mode of action: They can either act directly on the dsRNA, or they can trigger a signaling cascade. In the first group, the type III ribonuclease Dicer plays a key role to activate the antiviral RNA interference (RNAi) pathway by cleaving the viral dsRNA into small interfering RNAs (siRNAs). This represents the main innate antiviral immune mechanism in arthropods and nematodes. Even though Dicer is present and functional in mammals, the second group of DRAs, containing the RIG-I-like RNA helicases, appears to have functionally replaced RNAi and activate type I interferon (IFN) response upon dsRNA sensing. However, recent findings tend to blur the frontier between these 2 mechanisms, thereby highlighting the crucial and diverse roles played by RNA helicases in antiviral innate immunity. Here, we will review our current knowledge of the importance of these key proteins in viral infection, with a special focus on the interplay between the 2 main types of response that are activated by dsRNA.}, note = {1553-7374 (Electronic) 1553-7366 (Linking) Journal Article Review}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {Stabilization of Ribosomal RNA of the Small Subunit by Spermidine in Staphylococcus aureus}, author = {M. Belinite and I. Khusainov and H. Soufari and S. Marzi and P. Romby and M. Yusupov and Y. Hashem}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=34869582}, doi = {10.3389/fmolb.2021.738752}, isbn = {34869582}, year = {2021}, date = {2021-01-01}, journal = {Front Mol Biosci}, volume = {8}, pages = {738752}, abstract = {Cryo-electron microscopy is now used as a method of choice in structural biology for studying protein synthesis, a process mediated by the ribosome machinery. In order to achieve high-resolution structures using this approach, one needs to obtain homogeneous and stable samples, which requires optimization of ribosome purification in a species-dependent manner. This is especially critical for the bacterial small ribosomal subunit that tends to be unstable in the absence of ligands. Here, we report a protocol for purification of stable 30 S from the Gram-positive bacterium Staphylococcus aureus and its cryo-EM structures: in presence of spermidine at a resolution ranging between 3.4 and 3.6 A and in its absence at 5.3 A. Using biochemical characterization and cryo-EM, we demonstrate the importance of spermidine for stabilization of the 30 S via preserving favorable conformation of the helix 44.}, note = {2296-889X (Print) 2296-889X (Linking) Journal Article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{madel:hal-03064977, title = {Osteoclasts contribute to early development of chronic inflammation by promoting dysregulated hematopoiesis and myeloid skewing}, author = {Maria-Bernadette Madel and Lidia Ibanez and Thomas Ciucci and Julia Halper and Majlinda Topi and Henri-Jean Garchon and Matthieu Rouleau and Christopher Georges Mueller and Laurent Peyrin Biroulet and David Moulin and Claudine Blin-Wakkach and Abdelilah Wakkach}, url = {https://hal.archives-ouvertes.fr/hal-03064977}, doi = {10.1101/2020.12.09.418137}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, note = {working paper or preprint}, keywords = {Team-Mueller}, pubstate = {published}, tppubtype = {article} } @inbook{nokey, title = {Compartmentalization-based technologies for in vitro selection and evolution of ribozymes and light-up RNA aptamers}, author = {F. Bouhedda and M. Ryckelynck}, editor = {W. Winkler & B. Masquida S. Müller}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/9783527814527.ch28}, doi = {10.1002/9783527814527.ch28}, year = {2021}, date = {2021-01-01}, booktitle = {Ribozymes}, volume = {28}, pages = {721-738}, publisher = {John Wiley & Sons, Ltd}, abstract = {Summary Catalytic RNAs, also known as ribozymes, are naturally found in every living cell where they can occupy functions as important as peptide bond formation catalysis or intron splicing just as two examples. Besides, ribozymes are thought to be very ancient molecules that might have been the key actors of the so-called RNA world, but they also hold great promise for plenty of modern applications. These features have stimulated the development of in vitro evolution methodologies aiming at characterizing existing but also isolate new artificial ribozymes. Whereas bulk approaches in which all the RNA sequences of library are assayed in a single reaction mixture may be efficient to select fast, single-turn-over and/or self-modifying catalysts, this format is less adapted to the isolation of multiple turnover trans-acting molecules. Instead, a compartmentalization approach in which each variant is isolated and assayed into an individual compartment is better suited. In this chapter, we review the different strategies available to perform such compartmentalization and that range from hand-made water-in-oil emulsion to more advanced microfluidic-assisted ultrahigh-throughput screening. We finally extend the applications scope of these technologies to other RNAs (i.e., light-up RNA aptamers) for which a functional screening may also reveal more efficient than more conventional bulk in vitro selections.}, keywords = {catalytic RNAs, Droplet microfluidics, fluorescence, in vitro evolution, In vitro selection, light-up RNA aptamers, ribozymes, RYCKELYNCK, screening, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{Bec2021, title = {switchSENSE Technology for Analysis of DNA Polymerase Kinetics }, author = {G Bec and E Ennifar }, editor = {M Rederstorff}, year = {2021}, date = {2021-01-01}, journal = {Methods in Molecular Biology }, volume = {2247}, pages = {145-153}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, edition = {Small Non-Coding RNAs}, series = {Methods in Molecular Biology}, abstract = {The switchSENSE technology is a recent approach based on surface sensor chips for the analysis of interactions of macromolecules. The technology relies on electro-switchable DNA nanolevers tethered at one end on a gold surface via a sulfur linker and labeled with a Cy3 dye on the other end. The switchSENSE approach is effective in the determination of a large panel of biophysical parameters such as binding kinetics, dissociation constant, hydrodynamic radius, or melting temperature. In addition, it can also give access to some enzymatic data such as nuclease or polymerase activity. Here we describe a DNA polymerase assay that allows retrieving, in a single experimental set, association and dissociation rates, as well as the catalytic rate of the enzyme. }, keywords = {Biosensor, DNA polymerase, ENNIFAR, HIV reverse transcriptase, Kinetics, switchSENSE technology, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{Vicens2021, title = {Interaction Networks of Ribosomal Expansion Segments in Kinetoplastids }, author = {Q Vicens and A Bochler and A Jobe and J Frank and Y Hashem }, url = {https://pubmed.ncbi.nlm.nih.gov/33252739/}, doi = {10.1007/978-3-030-58971-4_13 }, year = {2021}, date = {2021-01-01}, volume = {96}, pages = {433-450}, publisher = {Subcellular Biochemistry}, abstract = {Expansion segments (ES) are insertions of a few to hundreds of nucleotides at discrete locations on eukaryotic ribosomal RNA (rRNA) chains. Some cluster around ‘hot spots’ involved in translation regulation and some may participate in biogenesis. Whether ES play the same roles in different organisms is currently unclear, especially since their size may vary dramatically from one species to another and very little is known about their functions. Most likely, ES variation is linked to adaptation to a particular environment. In this chapter, we compare the interaction networks of ES from four kinetoplastid parasites, which have evolved in diverse insect vectors and mammalian hosts: Trypanosoma cruzi, Trypanosoma brucei, Leishmania donovani and Leishmania major. Here, we comparatively analyze ribosome structures from these representative kinetoplastids and ascertain meaningful structural differences from mammalian ribosomes. We base our analysis on sequence alignments and three-dimensional structures of 80S ribosomes solved by cryo-electron microscopy (cryo-EM). Striking differences in size are observed between ribosomes of different parasites, indicating that not all ES are expanded equally. Larger ES are not always matched by large surrounding ES or proteins extensions in their vicinity, a particularity that may lead to clues about their biological function. ES display different species-specific patterns of conservation, which underscore the density of their interaction network at the surface of the ribosome. Making sense of the conservation patterns of ES is part of a global effort to lay the basis for functional studies aimed at discovering unique kinetoplastid-specific sites suitable for therapeutic applications against these human and often animal pathogens.}, keywords = {Eukaryotic translation, Expansion segment, Kinetoplastid parasite, Ribosomal RNA, Ribosome structure, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Neocles B. Leontis (1955 - 2020)}, author = {P B Moore and A Petrov and E Westhof and C L Zirbel}, url = {https://ibmc.cnrs.fr/wp-content/uploads/2021/01/Neocles-B_RNA.pdf}, doi = {10.1261/rna.078673.121}, isbn = {33452229}, year = {2021}, date = {2021-01-01}, journal = {Rna}, abstract = {None.}, note = {1469-9001 (Electronic) 1355-8382 (Linking) Journal Article}, keywords = {WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Amplifying and Fine-Tuning Rsm sRNAs Expression and Stability to Optimize the Survival of Pseudomonas brassicacerum in Nutrient-Poor Environments}, author = {D Lalaouna and S Fochesato and M Harir and P Ortet and P Schmitt-Kopplin and T Heulin and W Achouak}, url = {https://www.mdpi.com/2076-2607/9/2/250}, doi = {10.3390/microorganisms9020250}, year = {2021}, date = {2021-01-01}, journal = {Microorganisms}, volume = {9}, number = {2}, pages = {250}, abstract = {In the beneficial plant root-associated Pseudomonas brassicacearum strain NFM421, the GacS/GacA two-component system positively controls biofilm formation and the production of secondary metabolites through the synthesis of rsmX, rsmY and rsmZ. Here, we evidenced the genetic amplification of Rsm sRNAs by the discovery of a novel 110-nt long sRNA encoding gene, rsmX-2, generated by the duplication of rsmX-1 (formerly rsmX). Like the others rsm genes, its overexpression overrides the gacA mutation. We explored the expression and the stability of rsmX-1, rsmX-2, rsmY and rsmZ encoding genes under rich or nutrient-poor conditions, and showed that their amount is fine-tuned at the transcriptional and more interestingly at the post-transcriptional level. Unlike rsmY and rsmZ, we noticed that the expression of rsmX-1 and rsmX-2 genes was exclusively GacA-dependent. The highest expression level and longest half-life for each sRNA were correlated with the highest ppGpp and cyclic-di-GMP levels and were recorded under nutrient-poor conditions. Together, these data support the view that the Rsm system in P. brassicacearum is likely linked to the stringent response, and seems to be required for bacterial adaptation to nutritional stress.}, keywords = {GacA-dependent, nutritional stress, Pseudomonas brassicacearum, ROMBY, Rsm sRNAs, sRNAs stability, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid33514850, title = {Inhibition of HIV-1 gene transcription by KAP1 in myeloid lineage}, author = {Amina Ait-Ammar and Maxime Bellefroid and Fadoua Daouad and Valérie Martinelli and Jeanne Van Assche and Clémentine Wallet and Anthony Rodari and Marco De Rovere and Birthe Fahrenkrog and Christian Schwartz and Carine Van Lint and Virginie Gautier and Olivier Rohr}, doi = {10.1038/s41598-021-82164-w}, issn = {2045-2322}, year = {2021}, date = {2021-01-01}, urldate = {2021-01-01}, journal = {Sci Rep}, volume = {11}, number = {1}, pages = {2692}, abstract = {HIV-1 latency generates reservoirs that prevent viral eradication by the current therapies. To find strategies toward an HIV cure, detailed understandings of the molecular mechanisms underlying establishment and persistence of the reservoirs are needed. The cellular transcription factor KAP1 is known as a potent repressor of gene transcription. Here we report that KAP1 represses HIV-1 gene expression in myeloid cells including microglial cells, the major reservoir of the central nervous system. Mechanistically, KAP1 interacts and colocalizes with the viral transactivator Tat to promote its degradation via the proteasome pathway and repress HIV-1 gene expression. In myeloid models of latent HIV-1 infection, the depletion of KAP1 increased viral gene elongation and reactivated HIV-1 expression. Bound to the latent HIV-1 promoter, KAP1 associates and cooperates with CTIP2, a key epigenetic silencer of HIV-1 expression in microglial cells. In addition, Tat and CTIP2 compete for KAP1 binding suggesting a dynamic modulation of the KAP1 cellular partners upon HIV-1 infection. Altogether, our results suggest that KAP1 contributes to the establishment and the persistence of HIV-1 latency in myeloid cells.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Schneider2020, title = {Sensing and signalling viral infection in drosophila}, author = {J Schneider and JL Imler}, url = {https://doi.org/10.1016/j.dci.2020.103985 }, doi = {10.1016/j.dci.2020.103985}, year = {2020}, date = {2020-12-23}, journal = {Developmental and Comparative Immunology}, volume = {117}, abstract = {The fruitfly Drosophila melanogaster is a valuable model to unravel mechanisms of innate immunity, in particular in the context of viral infections. RNA interference, and more specifically the small interfering RNA pathway, is a major component of antiviral immunity in drosophila. In addition, the contribution of inducible transcriptional responses to the control of viruses in drosophila and other invertebrates is increasingly recognized. In particular, the recent discovery of a STING-IKKβ-Relish signalling cassette in drosophila has confirmed that NF-κB tran- scription factors play an important role in the control of viral infections, in addition to bacterial and fungal infections. Here, we review recent developments in the field, which begin to shed light on the mechanisms involved in sensing of viral infections and in signalling leading to production of antiviral effectors.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{Thomès2020, title = {Mosaic evolution of the phosphopantothenate biosynthesis pathway in bacteria and archaea}, author = { L Thomès and A Lescure}, url = {https://academic.oup.com/gbe/advance-article/doi/10.1093/gbe/evaa262/6035135}, doi = {10.1093/gbe/evaa262}, year = {2020}, date = {2020-12-15}, journal = {Genome Biology and Evolution}, volume = {13}, number = {2}, pages = {evaa262}, abstract = {Phosphopantothenate is a precursor to synthesis of Coenzyme A (CoA), a molecule essential to many metabolic pathways. Organisms of the archaeal phyla were shown to utilize a different phosphopantothenate biosynthetic pathway from the eukaryotic and bacterial one. In this study, we report that symbiotic bacteria from the group Candidatus poribacteria present enzymes of the archaeal pathway, namely pantoate kinase (PoK) and phosphopantothenate synthetase (PPS), mirroring what was demonstrated for Picrophilus torridus, an archaea partially utilizing the bacterial pathway. Our results support the ancient origin of the CoA pathway in the three domains of life, but also highlight its complex and dynamic evolution. Importantly, this study helps to improve protein annotation for this pathway in the Candidatus poribacteria group and other related organisms.}, keywords = {Candidatus poribacteria, Coenzyme A, LESCURE, phosphopantothenate pathway, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Abaeva2020, title = {The Halastavi árva Virus Intergenic Region IRES Promotes Translation by the Simplest Possible Initiation Mechanism }, author = {I S Abaeva and Q Vicens and A Bochler and H Soufari and A Simonetti and T V Pestova and Y Hashem and C U T Hellen}, url = {https://pubmed.ncbi.nlm.nih.gov/33296660/}, doi = {10.1016/j.celrep.2020.108476 }, year = {2020}, date = {2020-12-08}, journal = {Cell Reports}, volume = {33}, number = {10}, pages = {108476}, abstract = {Dicistrovirus intergenic region internal ribosomal entry sites (IGR IRESs) do not require initiator tRNA, an AUG codon, or initiation factors and jumpstart translation from the middle of the elongation cycle via formation of IRES/80S complexes resembling the pre-translocation state. eEF2 then translocates the [codon-anticodon]-mimicking pseudoknot I (PKI) from ribosomal A sites to P sites, bringing the first sense codon into the decoding center. Halastavi árva virus (HalV) contains an IGR that is related to previously described IGR IRESs but lacks domain 2, which enables these IRESs to bind to individual 40S ribosomal subunits. By using in vitro reconstitution and cryoelectron microscopy (cryo-EM), we now report that the HalV IGR IRES functions by the simplest initiation mechanism that involves binding to 80S ribosomes such that PKI is placed in the P site, so that the A site contains the first codon that is directly accessible for decoding without prior eEF2-mediated translocation of PKI. }, keywords = {Cricket paralysis virus, Dicistrovirus, ENNIFAR, Halastavi árva virus; IRES, intergenic region, internal ribosomal entry site, pseudoknot, Ribosome, SERBP1, SERPINE1 mRNA binding protein 1, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{A.2020, title = {The viral protein NSP1 acts as a ribosome gatekeeper for shutting down host translation and fostering SARS-CoV-2 translation}, author = {A Tidu and A Janvier and L Schaeffer and P Sosnowski and L Kuhn and P Hammann and E Westhof and G Eriani and F Martin }, url = {https://rnajournal.cshlp.org/content/early/2020/12/02/rna.078121.120}, doi = {10.1261/rna.078121.120 }, year = {2020}, date = {2020-12-02}, journal = {RNA}, volume = {27}, number = {3}, pages = {253-264}, abstract = {SARS-CoV-2 coronavirus is responsible for Covid-19 pandemic. In the early phase of infection, the single-strand positive RNA genome is translated into non-structural proteins (NSP). One of the first proteins produced during viral infection, NSP1, binds to the host ribosome and blocks the mRNA entry channel. This triggers translation inhibition of cellular translation. In spite of the presence of NSP1 on the ribosome, viral translation proceeds however. The molecular mechanism of the so-called viral evasion to NSP1 inhibition remains elusive. Here, we confirm that viral translation is maintained in the presence of NSP1. The evasion to NSP1-inhibition is mediated by the cis-acting RNA hairpin SL1 in the 5'UTR of SARS-CoV-2. NSP1-evasion can be transferred on a reporter transcript by SL1 transplantation. The apical part of SL1 is only required for viral translation. We show that NSP1 remains bound on the ribosome during viral translation. We suggest that the interaction between NSP1 and SL1 frees the mRNA accommodation channel while maintaining NSP1 bound to the ribosome. Thus, NSP1 acts as a ribosome gatekeeper, shutting down host translation or fostering SARS-CoV-2 translation depending on the presence of the SL1 5'UTR hairpin. SL1 is also present and necessary for translation of sub-genomic RNAs in the late phase of the infectious program. Consequently, therapeutic strategies targeting SL1 should affect viral translation at early and late stages of infection. Therefore, SL1 might be seen as a genuine 'Achille heel' of the virus. }, keywords = {ERIANI, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{P.2020, title = {Deflating the RNA Mg 2+ bubble. Stereochemistry to the rescue! }, author = {P Auffinger and E Ennifar and L D'Ascenzo}, url = {https://doi.org/10.1261/rna.076067.120}, doi = {10.1261/rna.076067.120 }, year = {2020}, date = {2020-12-02}, journal = {RNA}, volume = {27}, number = {3}, pages = {243-252}, abstract = {Proper evaluation of the ionic structure of biomolecular systems remains challenging in X ray and cryo-EM techniques but is essential for advancing our understanding of complex structure/activity/solvent relationships. However, numerous studies overestimate the number of MgProper evaluation of the ionic structure of biomolecular systems remains challenging in X ray and cryo-EM techniques but is essential for advancing our understanding of complex structure/activity/solvent relationships. However, numerous studies overestimate the number of Mg2+ in the deposited structures and underrate the importance of stereochemical rules to correctly assign these ions. Herein, we re-evaluate the PDBid 6QNR and 6SJ6 models of the ribosome ionic structure and establish that stereochemical principles need consideration when evaluating ion binding features, even when K+ anomalous signals are available as it is the case for 6QNR. Assignment errors can result in misleading conceptions of the solvent structure of ribosomes and other RNA systems and should therefore be avoided. Our analysis resulted in a significant decrease of the number of bound Mg2+ in the 6QNR structure, suggesting that K+ and not Mg2+ is the prevalent ion in the ribosome 1st solvation shell. We stress that the use of proper stereochemical guidelines is critical for deflating the current Mg2+ bubble witnessed in many ribosome and other RNA structures. Herewith, we would like to draw the attention of the researchers interested in the ionic structure of biomolecular systems on the importance and complementarity of stereochemistry and other ion identification techniques such as those pertaining to the detection of anomalous signals of transition metals and K+ We also stress that for the identification of lighter ions such as Mg2+, Na+, , stereochemistry coupled with high resolution structures remain the best currently available option. in the deposited structures and underrate the importance of stereochemical rules to correctly assign these ions. Herein, we re-evaluate the PDBid 6QNR and 6SJ6 models of the ribosome ionic structure and establish that stereochemical principles need consideration when evaluating ion binding features, even when K+ anomalous signals are available as it is the case for 6QNR. Assignment errors can result in misleading conceptions of the solvent structure of ribosomes and other RNA systems and should therefore be avoided. Our analysis resulted in a significant decrease of the number of bound Mg2+ in the 6QNR structure, suggesting that K+ and not Mg2+ is the prevalent ion in the ribosome 1st solvation shell. We stress that the use of proper stereochemical guidelines is critical for deflating the current Mg2+ bubble witnessed in many ribosome and other RNA structures. Herewith, we would like to draw the attention of the researchers interested in the ionic structure of biomolecular systems on the importance and complementarity of stereochemistry and other ion identification techniques such as those pertaining to the detection of anomalous signals of transition metals and K+ We also stress that for the identification of lighter ions such as Mg2+, Na+, , stereochemistry coupled with high resolution structures remain the best currently available option. }, keywords = {cryo-EM, ENNIFAR, ionic atmosphere, solvent, Unité ARN, validation, X-Ray}, pubstate = {published}, tppubtype = {article} } @article{pmid33441291, title = {[Viral sensing by RNA helicases]}, author = {C Rousseau and C Meignin}, url = {https://pubmed.ncbi.nlm.nih.gov/33441291/}, doi = {10.1684/vir.2020.0871 }, isbn = {1267-8694}, year = {2020}, date = {2020-12-01}, journal = {Virologie (Montrouge)}, volume = {24}, number = {6}, pages = {419-436}, keywords = {antiviral innate immunity, M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{pmid33441288, title = {Viral sensing by RNA helicases}, author = {C Rousseau and C Meignin}, url = {https://pubmed.ncbi.nlm.nih.gov/33441288/}, doi = {10.1684/vir.2020.0872}, issn = {1267-8694}, year = {2020}, date = {2020-12-01}, journal = {Virologie (Montrouge)}, volume = {24}, number = {6}, pages = {36-52}, keywords = {antiviral innate immunity, M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{perraud_opportunistic_2020, title = {Opportunistic use of catecholamine neurotransmitters as siderophores to access iron by Pseudomonas aeruginosa}, author = {Quentin Perraud and Lauriane Kuhn and Sarah Fritsch and Gwenaëlle Graulier and Véronique Gasser and Vincent Normant and Philippe Hammann and Isabelle J. Schalk}, doi = {10.1111/1462-2920.15372}, issn = {1462-2920}, year = {2020}, date = {2020-12-01}, journal = {Environmental Microbiology}, abstract = {Iron is an essential nutrient for bacterial growth and the cause of a fierce battle between the pathogen and host during infection. Bacteria have developed several strategies to access iron from the host, the most common being the production of siderophores, small iron-chelating molecules secreted into the bacterial environment. The opportunist pathogen Pseudomonas aeruginosa produces two siderophores, pyoverdine and pyochelin, and is also able to use a wide panoply of xenosiderophores, siderophores produced by other microorganisms. Here, we demonstrate that catecholamine neurotransmitters (dopamine, l-DOPA, epinephrine and norepinephrine) are able to chelate iron and efficiently bring iron into P. aeruginosa cells via TonB-dependent transporters (TBDTs). Bacterial growth assays under strong iron-restricted conditions and with numerous mutants showed that the TBDTs involved are PiuA and PirA. PiuA exhibited more pronounced specificity for dopamine uptake than for norepinephrine, epinephrine and l-DOPA, whereas PirA specificity appeared to be higher for l-DOPA and norepinephrine. Proteomic and qRT-PCR approaches showed pirA transcription and expression to be induced in the presence of all four catecholamines. Finally, the oxidative properties of catecholamines enable them to reduce iron, and we observed ferrous iron uptake via the FeoABC system in the presence of l-DOPA.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{ruiz_toxicological_2020, title = {Toxicological evaluation of highly water dispersible few-layer graphene in vivo}, author = {Amalia Ruiz and Matteo Andrea Lucherelli and Diane Murera and Delphine Lamon and Cécilia Ménard-Moyon and Alberto Bianco}, url = {https://linkinghub.elsevier.com/retrieve/pii/S0008622320307855}, doi = {10.1016/j.carbon.2020.08.023}, issn = {00086223}, year = {2020}, date = {2020-12-01}, urldate = {2020-11-20}, journal = {Carbon}, volume = {170}, pages = {347--360}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{Cai_2020, title = {2'3'-cGAMP triggers a STING- and NF-κB-dependent broad antiviral response in Drosophila}, author = {H Cai and A Holleufer and B Simonsen and J Schneider and A Lemoine and HH Gad and J Huang and J Huang and D Chen and T Peng and JT Marques and R Hartmann and N Martins and JL Imler}, url = {https://pubmed.ncbi.nlm.nih.gov/33262294/}, doi = {10.1126/scisignal.abc4537 }, year = {2020}, date = {2020-12-01}, journal = {Science Signaling}, volume = {13}, number = {660}, pages = {eabc4537}, abstract = {We previously reported that an ortholog of STING regulates infection by picorna-like viruses in Drosophila In mammals, STING is activated by the cyclic dinucleotide 2'3'-cGAMP produced by cGAS, which acts as a receptor for cytosolic DNA. Here, we showed that injection of flies with 2'3'-cGAMP induced the expression of dSTING-regulated genes. Coinjection of 2'3'-cGAMP with a panel of RNA or DNA viruses resulted in substantially reduced viral replication. This 2'3'-cGAMP-mediated protection was still observed in flies with mutations in Atg7 and AGO2, genes that encode key components of the autophagy and small interfering RNA pathways, respectively. By contrast, this protection was abrogated in flies with mutations in the gene encoding the NF-κB transcription factor Relish. Transcriptomic analysis of 2'3'-cGAMP-injected flies revealed a complex response pattern in which genes were rapidly induced, induced after a delay, or induced in a sustained manner. Our results reveal that dSTING regulates an NF-κB-dependent antiviral program that predates the emergence of interferons in vertebrates. }, keywords = {imler, M3i, Marques, STING}, pubstate = {published}, tppubtype = {article} } @inbook{Imler_2020, title = {L'immunité innée}, author = {JL Imler and JA Hofmann}, editor = {P Cramer and A Meignien}, isbn = {9791090119895}, year = {2020}, date = {2020-11-30}, volume = {Le défi des maladies infectieuses}, pages = {191-203}, publisher = {Les éditions du Palais}, edition = {Docis}, keywords = {hoffmann, imler, M3i}, pubstate = {published}, tppubtype = {inbook} } @article{12020, title = {Adaptation of the Romanomermis culicivorax CCA-Adding Enzyme to Miniaturized Armless tRNA Substrates }, author = {O Hennig and S Philipp and S Bonin and K Rollet and T Kolberg and T Jühling and H Betat and C Sauter and M Mörl }, url = {https://www.mdpi.com/1422-0067/21/23/9047}, doi = {10.3390/ijms21239047 }, year = {2020}, date = {2020-11-28}, journal = {International Journal of Molecular Sciences}, volume = {21}, number = {23}, pages = {E9047}, abstract = {The mitochondrial genome of the nematode Romanomermis culicivorax encodes for miniaturized hairpin-like tRNA molecules that lack D- as well as T-arms, strongly deviating from the consensus cloverleaf. The single tRNA nucleotidyltransferase of this organism is fully active on armless tRNAs, while the human counterpart is not able to add a complete CCA-end. Transplanting single regions of the Romanomermis enzyme into the human counterpart, we identified a beta-turn element of the catalytic core that-when inserted into the human enzyme-confers full CCA-adding activity on armless tRNAs. This region, originally identified to position the 3'-end of the tRNA primer in the catalytic core, dramatically increases the enzyme's substrate affinity. While conventional tRNA substrates bind to the enzyme by interactions with the T-arm, this is not possible in the case of armless tRNAs, and the strong contribution of the beta-turn compensates for an otherwise too weak interaction required for the addition of a complete CCA-terminus. This compensation demonstrates the remarkable evolutionary plasticity of the catalytic core elements of this enzyme to adapt to unconventional tRNA substrates. }, keywords = {CCA-adding enzyme, co-evolution, evolutionary plasticity, minimalized armless tRNAs, SAUTER, tRNA nucleotidyltransferase, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Hayoun2020, title = {Dichloromethane Degradation Pathway from Unsequenced Hyphomicrobium sp. MC8b Rapidly Explored by Pan-Proteomics }, author = {K Hayoun and E Geersens and C C Laczny and R Halder and C Lázaro Sánchez and A Manna and F Bringel and M Ryckelynck and P Wilmesand E E L Muller and B Alpha-Bazin and J Armengaud and S Vuilleumier }, url = {https://www.mdpi.com/2076-2607/8/12/1876}, doi = { 10.3390/microorganisms8121876 }, year = {2020}, date = {2020-11-27}, journal = {Microorganisms}, volume = {8}, number = {12}, pages = {E1876}, abstract = {Several bacteria are able to degrade the major industrial solvent dichloromethane (DCM) by using the conserved dehalogenase DcmA, the only system for DCM degradation characterised at the sequence level so far. Using differential proteomics, we rapidly identified key determinants of DCM degradation for Hyphomicrobium sp. MC8b, an unsequenced facultative methylotrophic DCM-degrading strain. For this, we designed a pan-proteomics database comprising the annotated genome sequences of 13 distinct Hyphomicrobium strains. Compared to growth with methanol, growth with DCM induces drastic changes in the proteome of strain MC8b. Dichloromethane dehalogenase DcmA was detected by differential pan-proteomics, but only with poor sequence coverage, suggesting atypical characteristics of the DCM dehalogenation system in this strain. More peptides were assigned to DcmA by error-tolerant search, warranting subsequent sequencing of the genome of strain MC8b, which revealed a highly divergent set of dcm genes in this strain. This suggests that the dcm enzymatic system is less strongly conserved than previously believed, and that substantial molecular evolution of dcm genes has occurred beyond their horizontal transfer in the bacterial domain. Our study showed the power of pan-proteomics for quick characterization of new strains belonging to branches of the Tree of Life that are densely genome-sequenced. }, keywords = {dcm genes, dehalogenation, dichloromethane, differential proteomics, genome sequencing, Hyphomicrobium, Nanopore, pan-proteomics, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{S.2020, title = {Special Issue "Function and Structure of Viral Ribonucleoproteins Complexes"}, author = {S Bernacchi}, url = {https://www.mdpi.com/1999-4915/12/12/1355}, doi = { 10.3390/v12121355 }, year = {2020}, date = {2020-11-26}, journal = {Viruses}, volume = {12}, number = {12}, pages = {E1355}, abstract = {RNA viruses are extraordinary evolution machines that efficiently ensure their replication by taking advantage of the association with viral and cellular components to form ribonucleic complexes (vRNPs) [...]. }, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{B2020b, title = {Advances in RNA 3D Structure Modeling Using Experimental Data}, author = {B Li and Y Cao and E Westhof and Z Miao}, doi = {https://doi.org/10.3389/fgene.2020.574485}, year = {2020}, date = {2020-11-23}, journal = {Front Genet .}, volume = {11}, number = {574485}, abstract = {RNA is a unique bio-macromolecule that can both record genetic information and perform biological functions in a variety of molecular processes, including transcription, splicing, translation, and even regulating protein function. RNAs adopt specific three-dimensional conformations to enable their functions. Experimental determination of high-resolution RNA structures using x-ray crystallography is both laborious and demands expertise, thus, hindering our comprehension of RNA structural biology. The computational modeling of RNA structure was a milestone in the birth of bioinformatics. Although computational modeling has been greatly improved over the last decade showing many successful cases, the accuracy of such computational modeling is not only length-dependent but also varies according to the complexity of the structure. To increase credibility, various experimental data were integrated into computational modeling. In this review, we summarize the experiments that can be integrated into RNA structure modeling as well as the computational methods based on these experimental data. We also demonstrate how computational modeling can help the experimental determination of RNA structure. We highlight the recent advances in computational modeling which can offer reliable structure models using high-throughput experimental data.}, keywords = {3D shape, chemical probing, RNA structure, RNA-puzzles, structure prediction, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{Duvergé2020, title = {Pseudotyping Lentiviral Vectors: When the Clothes Make the Virus }, author = {A Duvergé and M Negroni}, url = {https://doi.org/10.3390/v12111311}, doi = {10.3390/v12111311}, year = {2020}, date = {2020-11-11}, journal = {Viruses}, volume = {12}, number = {11}, pages = {1311}, abstract = {Delivering transgenes to human cells through transduction with viral vectors constitutes one of the most encouraging approaches in gene therapy. Lentivirus-derived vectors are among the most promising vectors for these approaches. When the genetic modification of the cell must be performed in vivo, efficient specific transduction of the cell targetsof the therapy in the absence of off-targeting constitutes the Holy Grail of gene therapy. For viral therapy, this is largely determined by the characteristics of the surface proteins carried by the vector. In this regard, an important property of lentiviral vectors is the possibility of being pseudotyped by envelopes of other viruses, widening the panel of proteins with which they can bearmed. Here, we discuss how this is achieved at the molecular level and what the properties and the potentialities of the different envelope proteins that can be used for pseudotyping these vectorsare.}, keywords = {envelope proteins, gene therapy, lentiviral vectors, NEGRONI, pseudotyping, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{L.2020, title = {Role of Purine-Rich Regions in Mason-Pfizer Monkey Virus (MPMV) Genomic RNA Packaging and Propagation }, author = {L M Ali and F N Pitchai and V Vivet-Boudou and A Chameettachal and A Jabeen and V N Pillai and F Mustafa and R Marquet and T A Rizvi }, url = {https://www.frontiersin.org/articles/10.3389/fmicb.2020.595410/full}, doi = { 10.3389/fmicb.2020.595410 }, year = {2020}, date = {2020-11-05}, journal = {Frontiers in Microbiology}, number = {11}, pages = {595410}, abstract = {A distinguishing feature of the Mason-Pfizer monkey virus (MPMV) packaging signal RNA secondary structure is a single-stranded purine-rich sequence (ssPurines) in close vicinity to a palindromic stem loop (Pal SL) that functions as MPMV dimerization initiation site (DIS). However, unlike other retroviruses, MPMV contains a partially base-paired repeat sequence of ssPurines (bpPurines) in the adjacent region. Both purine-rich sequences have earlier been proposed to act as potentially redundant Gag binding sites to initiate the process of MPMV genomic RNA (gRNA) packaging. The objective of this study was to investigate the biological significance of ssPurines and bpPurines in MPMV gRNA packaging by systematic mutational and biochemical probing analyses. Deletion of either ssPurines or bpPurines individually had no significant effect on MPMV gRNA packaging, but it was severely compromised when both sequences were deleted simultaneously. Selective 2' hydroxyl acylation analyzed by primer extension (SHAPE) analysis of the mutant RNAs revealed only mild effects on structure by deletion of either ssPurines or bpPurines, while the structure was dramatically affected by the two simultaneous deletions. This suggests that ssPurines and bpPurines play a redundant role in MPMV gRNA packaging, probably as Gag binding sites to facilitate gRNA capture and encapsidation. Interestingly, the deletion of bpPurines revealed an additional severe defect on RNA propagation that was independent of the presence or absence of ssPurines or the gRNA structure of the region. These findings further suggest that the bpPurines play an additional role in the early steps of MPMV replication cycle that is yet to be identified. }, keywords = {base paired purines, MARQUET, Mason-Pfizer monkey virus, PAILLART, retroviruses, RNA packaging, RNA secondary structure, RNA-Gag interaction, SHAPE, single-stranded purines, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{soufari_structure_2020, title = {Structure of the mature kinetoplastids mitoribosome and insights into its large subunit biogenesis.}, author = {Heddy Soufari and Florent Waltz and Camila Parrot and Stéphanie Durrieu-Gaillard and Anthony Bochler and Lauriane Kuhn and Marie Sissler and Yaser Hashem}, doi = {10.1073/pnas.2011301117}, issn = {1091-6490 0027-8424}, year = {2020}, date = {2020-11-01}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {117}, number = {47}, pages = {29851--29861}, abstract = {Kinetoplastids are unicellular eukaryotic parasites responsible for such human pathologies as Chagas disease, sleeping sickness, and leishmaniasis. They have a single large mitochondrion, essential for the parasite survival. In kinetoplastid mitochondria, most of the molecular machineries and gene expression processes have significantly diverged and specialized, with an extreme example being their mitochondrial ribosomes. These large complexes are in charge of translating the few essential mRNAs encoded by mitochondrial genomes. Structural studies performed in Trypanosoma brucei already highlighted the numerous peculiarities of these mitoribosomes and the maturation of their small subunit. However, several important aspects mainly related to the large subunit (LSU) remain elusive, such as the structure and maturation of its ribosomal RNA. Here we present a cryo-electron microscopy study of the protozoans Leishmania tarentolae and Trypanosoma cruzi mitoribosomes. For both species, we obtained the structure of their mature mitoribosomes, complete rRNA of the LSU, as well as previously unidentified ribosomal proteins. In addition, we introduce the structure of an LSU assembly intermediate in the presence of 16 identified maturation factors. These maturation factors act on both the intersubunit and the solvent sides of the LSU, where they refold and chemically modify the rRNA and prevent early translation before full maturation of the LSU.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{Abel2020, title = {Structural insights into the membrane receptor ShuA in DDM micelles and in a model of gram-negative bacteria outer membrane as seen by SAXS and MD simulations }, author = {S Abel and M Marchi and J Solier and S Finet and K Brillet and F Bonneté }, url = {https://pubmed.ncbi.nlm.nih.gov/33157097/}, doi = {10.1016/j.bbamem.2020.183504 }, year = {2020}, date = {2020-11-01}, journal = {Biochim Biophys Acta Biomembr}, volume = {in press}, abstract = {Successful crystallization of membrane proteins in detergent micelles depends on key factors such as conformational stability of the protein in micellar assemblies, the protein-detergent complex (PDC) monodispersity and favorable protein crystal contacts by suitable shielding of the protein hydrophobic surface by the detergent belt. With the aim of studying the influence of amphiphilic environment on membrane protein structure, stability and crystallizability, we combine molecular dynamics (MD) simulations with SEC-MALLS and SEC-SAXS (Size Exclusion Chromatography in line with Multi Angle Laser Light Scattering or Small Angle X-ray Scattering) experiments to describe the protein-detergent interactions that could help to rationalize PDC crystallization. In this context, we compare the protein-detergent interactions of ShuA from Shigella dysenteriae in n-Dodecyl-β-D-Maltopyranoside (DDM) with ShuA inserted in a realistic model of gram-negative bacteria outer membrane (OM) containing a mixture of bacterial lipopolysaccharide and phospholipids. To evaluate the quality of the PDC models, we compute the corresponding SAXS curves from the MD trajectories and compare with the experimental ones. We show that computed SAXS curves obtained from the MD trajectories reproduce better the SAXS obtained from the SEC-SAXS experiments for ShuA surrounded by 268 DDM molecules. The MD results show that the DDM molecules form around ShuA a closed belt whose the hydrophobic thickness appears slightly smaller (~22 Å) than the hydrophobic transmembrane domain of the protein (24.6 Å) suggested by Orientations of Proteins in Membranes (OPM) database. The simulations also show that ShuA transmembrane domain is remarkably stable in all the systems except for the extracellular and periplasmic loops that exhibit larger movements due to specific molecular interactions with lipopolysaccharides (LPS). We finally point out that this detergent behavior may lead to the occlusion of the periplasmic hydrophilic surface and poor crystal contacts leading to difficulties in crystallization of ShuA in DDM. }, keywords = {DDM, ENNIFAR, Membrane transporter ShuA, Molecular dynamics simulations, Outer membrane model, SEC-MALLS, SEC-SAXS, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Petitjean2020, title = {Genome-Wide CRISPR-Cas9 Screen Reveals the Importance of the Heparan Sulfate Pathway and the Conserved Oligomeric Golgi Complex for Synthetic Double-Stranded RNA Uptake and Sindbis Virus Infection }, author = {O Petitjean and E Girardi and RP Ngondon and V Lupashin and S Pfeffer }, url = {https://pubmed.ncbi.nlm.nih.gov/33177215/}, doi = {10.1128/mSphere.00914-20 }, year = {2020}, date = {2020-11-01}, journal = { mSphere}, volume = {5}, number = {6}, pages = {e00914-20}, abstract = {Double-stranded RNA (dsRNA) is the hallmark of many viral infections. dsRNA is produced either by RNA viruses during replication or by DNA viruses upon convergent transcription. Synthetic dsRNA is also able to mimic viral-induced activation of innate immune response and cell death. In this study, we employed a genome-wide CRISPR-Cas9 loss-of-function screen based on cell survival in order to identify genes implicated in the host response to dsRNA. By challenging HCT116 human cells with either synthetic dsRNA or Sindbis virus (SINV), we identified the heparan sulfate (HS) pathway as a crucial factor for dsRNA entry, and we validated SINV dependency on HS. Interestingly, we uncovered a novel role for COG4, a component of the conserved oligomeric Golgi (COG) complex, as a factor involved in cell survival to both dsRNA and SINV in human cells. We showed that COG4 knockout led to a decrease of extracellular HS that specifically affected dsRNA transfection efficiency and reduced viral production, which explains the increased cell survival of these mutants.IMPORTANCE When facing a viral infection, the organism has to put in place a number of defense mechanisms in order to clear the pathogen from the cell. At the early phase of this preparation for fighting against the invader, the innate immune response is triggered by the sensing of danger signals. Among those molecular cues, double-stranded RNA (dsRNA) is a very potent inducer of different reactions at the cellular level that can ultimately lead to cell death. Using a genome-wide screening approach, we set to identify genes involved in dsRNA entry, sensing, and apoptosis induction in human cells. This allowed us to determine that the heparan sulfate pathway and the conserved oligomeric Golgi complex are key determinants allowing entry of both dsRNA and viral nucleic acid leading to cell death. }, keywords = {complex oligomeric Golgi complex, CRISPR-Cas9 screen, double-stranded RNA, heparan-sulfate, PFEFFER, Transfection, Unité ARN, virus}, pubstate = {published}, tppubtype = {article} } @article{Despons2020, title = {How Many Messenger RNAs Can Be Translated by the START Mechanism? }, author = {L Despons and F Martin }, url = {https://pubmed.ncbi.nlm.nih.gov/33171614/}, doi = {10.3390/ijms21218373 }, year = {2020}, date = {2020-11-01}, journal = {International Journal of Molecular Sciences}, volume = {21}, number = {21}, pages = {8373}, abstract = {Translation initiation is a key step in the protein synthesis stage of the gene expression pathway of all living cells. In this important process, ribosomes have to accurately find the AUG start codon in order to ensure the integrity of the proteome. "Structure Assisted RNA Translation", or "START", has been proposed to use stable secondary structures located in the coding sequence to augment start site selection by steric hindrance of the progression of pre-initiation complex on messenger RNA. This implies that such structures have to be located downstream and at on optimal distance from the AUG start codon (i.e., downstream nucleotide +16). In order to assess the importance of the START mechanism in the overall mRNA translation process, we developed a bioinformatic tool to screen coding sequences for such stable structures in a 50 nucleotide-long window spanning the nucleotides from +16 to +65. We screened eight bacterial genomes and six eukaryotic genomes. We found stable structures in 0.6-2.5% of eukaryotic coding sequences. Among these, approximately half of them were structures predicted to form G-quadruplex structures. In humans, we selected 747 structures. In bacteria, the coding sequences from Gram-positive bacteria contained 2.6-4.2% stable structures, whereas the structures were less abundant in Gram-negative bacteria (0.2-2.7%). In contrast to eukaryotes, putative G-quadruplex structures are very rare in the coding sequence of bacteria. Altogether, our study reveals that the START mechanism seems to be an ancient strategy to facilitate the start codon recognition that is used in different kingdoms of life. }, keywords = {ERIANI, LESCURE, mRNA, Ribosome, secondary structures, START, translation initiation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{soufari_specific_2020, title = {Specific features and assembly of the plant mitochondrial complex I revealed by cryo-EM.}, author = {Heddy Soufari and Camila Parrot and Lauriane Kuhn and Florent Waltz and Yaser Hashem}, doi = {10.1038/s41467-020-18814-w}, issn = {2041-1723 2041-1723}, year = {2020}, date = {2020-10-01}, journal = {Nature communications}, volume = {11}, number = {1}, pages = {5195}, abstract = {Mitochondria are the powerhouses of eukaryotic cells and the site of essential metabolic reactions. Complex I or NADH:ubiquinone oxidoreductase is the main entry site for electrons into the mitochondrial respiratory chain and constitutes the largest of the respiratory complexes. Its structure and composition vary across eukaryote species. However, high resolution structures are available only for one group of eukaryotes, opisthokonts. In plants, only biochemical studies were carried out, already hinting at the peculiar composition of complex I in the green lineage. Here, we report several cryo-electron microscopy structures of the plant mitochondrial complex I. We describe the structure and composition of the plant respiratory complex I, including the ancestral mitochondrial domain composed of the carbonic anhydrase. We show that the carbonic anhydrase is a heterotrimeric complex with only one conserved active site. This domain is crucial for the overall stability of complex I as well as a peculiar lipid complex composed of cardiolipin and phosphatidylinositols. Moreover, we also describe the structure of one of the plant-specific complex I assembly intermediates, lacking the whole P(D) module, in presence of the maturation factor GLDH. GLDH prevents the binding of the plant specific P1 protein, responsible for the linkage of the P(P) to the P(D) module.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{incarbone_characterization_2020, title = {Characterization of a DCL2-Insensitive Tomato Bushy Stunt Virus Isolate Infecting Arabidopsis thaliana.}, author = {Marco Incarbone and Hélene Scheer and Jean-Michel Hily and Lauriane Kuhn and Mathieu Erhardt and Patrice Dunoyer and Denise Altenbach and Christophe Ritzenthaler}, doi = {10.3390/v12101121}, issn = {1999-4915 1999-4915}, year = {2020}, date = {2020-10-01}, journal = {Viruses}, volume = {12}, number = {10}, abstract = {Tomato bushy stunt virus (TBSV), the type member of the genus Tombusvirus in the family Tombusviridae is one of the best studied plant viruses. The TBSV natural and experimental host range covers a wide spectrum of plants including agricultural crops, ornamentals, vegetables and Nicotiana benthamiana. However, Arabidopsis thaliana, the well-established model organism in plant biology, genetics and plant-microbe interactions is absent from the list of known TBSV host plant species. Most of our recent knowledge of the virus life cycle has emanated from studies in Saccharomyces cerevisiae, a surrogate host for TBSV that lacks crucial plant antiviral mechanisms such as RNA interference (RNAi). Here, we identified and characterized a TBSV isolate able to infect Arabidopsis with high efficiency. We demonstrated by confocal and 3D electron microscopy that in Arabidopsis TBSV-BS3Ng replicates in association with clustered peroxisomes in which numerous spherules are induced. A dsRNA-centered immunoprecipitation analysis allowed the identification of TBSV-associated host components including DRB2 and DRB4, which perfectly localized to replication sites, and NFD2 that accumulated in larger viral factories in which peroxisomes cluster. By challenging knock-out mutants for key RNAi factors, we showed that TBSV-BS3Ng undergoes a non-canonical RNAi defensive reaction. In fact, unlike other RNA viruses described, no 22nt TBSV-derived small RNA are detected in the absence of DCL4, indicating that this virus is DCL2-insensitive. The new Arabidopsis-TBSV-BS3Ng pathosystem should provide a valuable new model for dissecting plant-virus interactions in complement to Saccharomyces cerevisiae.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{normant_nocardamine-dependent_2020, title = {Nocardamine-Dependent Iron Uptake in Pseudomonas aeruginosa: Exclusive Involvement of the FoxA Outer Membrane Transporter.}, author = {Vincent Normant and Inokentijs Josts and Lauriane Kuhn and Quentin Perraud and Sarah Fritsch and Philippe Hammann and Gaëtan L A Mislin and Henning Tidow and Isabelle J Schalk}, doi = {10.1021/acschembio.0c00535}, issn = {1554-8937 1554-8929}, year = {2020}, date = {2020-10-01}, journal = {ACS chemical biology}, volume = {15}, number = {10}, pages = {2741--2751}, abstract = {Iron is a key nutrient for almost all living organisms. Paradoxically, it is poorly soluble and consequently poorly bioavailable. Bacteria have thus developed multiple strategies to access this metal. One of the most common consists of the use of siderophores, small compounds that chelate ferric iron with very high affinity. Many bacteria are able to produce their own siderophores or use those produced by other microorganisms (exosiderophores) in a piracy strategy. Pseudomonas aeruginosa produces two siderophores, pyoverdine and pyochelin, and is also able to use a large panel of exosiderophores. We investigated the ability of P. aeruginosa to use nocardamine (NOCA) and ferrioxamine B (DFOB) as exosiderophores under iron-limited planktonic growth conditions. Proteomic and RT-qPCR approaches showed induction of the transcription and expression of the outer membrane transporter FoxA in the presence of NOCA or DFOB in the bacterial environment. Expression of the proteins of the heme- or pyoverdine- and pyochelin-dependent iron uptake pathways was not affected by the presence of these two tris-hydroxamate siderophores. (55)Fe uptake assays using foxA mutants showed ferri-NOCA to be exclusively transported by FoxA, whereas ferri-DFOB was transported by FoxA and at least one other unidentified transporter. The crystal structure of FoxA complexed with NOCA-Fe revealed very similar siderophore binding sites between NOCA-Fe and DFOB-Fe. We discuss iron uptake by hydroxamate exosiderophores in P. aeruginosa cells in light of these results.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{pmid32619426, title = {Evolution of a concept: From accessory protein to key virulence factor, the case of HIV-1 Vpr}, author = {Clémentine Wallet and Olivier Rohr and Christian Schwartz}, doi = {10.1016/j.bcp.2020.114128}, issn = {1873-2968}, year = {2020}, date = {2020-10-01}, urldate = {2020-10-01}, journal = {Biochem Pharmacol}, volume = {180}, pages = {114128}, abstract = {Back in 1989 some studies have shown that the viral protein Vpr was dispensable for HIV-1 replication in vitro. From then the concept of accessory or auxiliary protein for Vpr has emerged and it is still used to date. However, Vpr soon appeared to be very important for in vivo virus spread and pathogenesis. Vpr has been involved in many biological functions including regulation of reverse transcriptase activity, the nuclear import of the pre-integration complex (PIC), HIV-1 transcription, gene splicing, apoptosis and in cell cycle arrest. Thus, we might rather consider Vpr as a true virulence factor instead of just an accessory factor. At present, Vpr can be regarded as a potential and promising target in different strategies aiming to fight infected cells including latently infected cells.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comparative patterns of modified nucleotides in individual tRNA species from a mesophilic and two thermophilic archaea}, author = {P Wolff and C Villette and J Zumsteg and D Heintz and L Antoine and B Chane-Woon-Ming and L Droogmans and H Grosjean and E Westhof}, url = {https://pubmed.ncbi.nlm.nih.gov/32994183/}, doi = {10.1261/rna.077537.120}, year = {2020}, date = {2020-09-29}, journal = {RNA}, volume = {26}, number = {12}, pages = {1957-1975}, abstract = {To improve and complete our knowledge of archaeal tRNA modification patterns, we have identified and compared the modification pattern (type and location) in tRNAs of three very different archaeal species, Methanococcus maripaludis (a mesophilic methanogen), Pyrococcus furiosus (a hyperthermophile thermococcale) and Sulfolobus acidocaldarius (an acidophilic thermophilic sulfolobale). Most abundant isoacceptor tRNAs (79 in total) for each of the 20 amino acids were isolated by two-dimensional gel electrophoresis followed by in-gel RNase digestions. The resulting oligonucleotide fragments were separated by nanoLC and their nucleotide content analyzed by mass spectrometry (MS/MS). Analysis of total modified nucleosides obtained from complete digestion of bulk tRNAs was also performed. Distinct base- and/or ribose-methylations, cytidine acetylations and thiolated pyrimidines were identified, some at new positions in tRNAs. Novel, some tentatively identified, modifications were also found. The least diversified modification landscape is observed in the mesophilic Methanococcus maripaludis and the most complex one in Sulfolobus acidocaldarius. Notable observations are the frequent occurrence of ac4C nucleotides in thermophilic archaeal tRNAs, the presence of m7G at positions 1 and 10 in Pyrococcus furiosus tRNAs, and the use of wyosine derivatives at position 37 of tRNAs especially those decoding U1- and C1-starting codons. These results complete those already obtained by others with sets of archaeal tRNAs from Methanocaldococcus jannaschii and Haloferax volcanii.}, keywords = {(hyper)thermophiles Archaea mass spectrometry modifications tRNA, ARN-MS, ENNIFAR, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{volohonsky_plasmodium_2020, title = {Kinetics of Plasmodium midgut invasion in Anopheles mosquitoes}, author = {Gloria Volohonsky and Perrine Paul-Gilloteaux and Jitka Stafkova and Julien Soichot and Jean Salamero and Elena A. Levashina }, editor = {Kenneth D. Vernick, Institut Pasteur, FRANCE}, url = {https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1008739}, doi = {10.1371/journal.ppat.1008739}, issn = {1553-7374}, year = {2020}, date = {2020-09-18}, journal = {PLoS Pathog}, volume = {16}, number = {9}, abstract = {The traversal of the mosquito midgut cells is one of the critical stages in the life cycle of malaria parasites. Motile parasite forms, called ookinetes, traverse the midgut epithelium in a dynamic process which is not fully understood. Here, we harnessed transgenic reporters to track invasion of Plasmodium parasites in African and Indian mosquito species. We found important differences in parasite dynam- ics between the two Anopheles species and demonstrated a role of the mosquito comple- ment-like system in regulation of parasite invasion of the midgut cells.}, keywords = {Anopheles, M3i, Malaria, marois, midgut, Plasmodium}, pubstate = {published}, tppubtype = {article} } @article{spenle_tenascin-c_2020, title = {Tenascin-C Orchestrates an Immune-Suppressive Tumor Microenvironment in Oral Squamous Cell Carcinoma}, author = {Caroline Spenlé and Thomas Loustau and Devadarssen Murdamoothoo and William Erne and Stephanie Beghelli-de la Forest Divonne and Romain Veber and Luciana Petti and Pierre Bourdely and Matthias Mörgelin and Eva-Maria Brauchle and Gérard Cremel and Vony Randrianarisoa and Abdouramane Camara and Samah Rekima and Sebastian Schaub and Kelly Nouhen and Thomas Imhof and Uwe Hansen and Nicodème Paul and Raphael Carapito and Nicolas Pythoud and Aurélie Hirschler and Christine Carapito and Hélène Dumortier and Christopher G Mueller and Manuel Koch and Katja Schenke-Layland and Shigeyuki Kon and Anne Sudaka and Fabienne Anjuère and Ellen Van Obberghen-Schilling and Gertraud Orend}, doi = {10.1158/2326-6066.CIR-20-0074}, issn = {2326-6074}, year = {2020}, date = {2020-09-01}, journal = {Cancer Immunology Research}, volume = {8}, number = {9}, pages = {1122--1138}, abstract = {Inherent immune suppression represents a major challenge in the treatment of human cancer. The extracellular matrix molecule tenascin-C promotes cancer by multiple mechanisms, yet the roles of tenascin-C in tumor immunity are incompletely understood. Using a 4NQO-induced oral squamous cell carcinoma (OSCC) model with abundant and absent tenascin-C, we demonstrated that tenascin-C enforced an immune-suppressive lymphoid stroma via CCL21/CCR7 signaling, leading to increased metastatic tumors. Through TLR4, tenascin-C increased expression of CCR7 in CD11c+ myeloid cells. By inducing CCL21 in lymphatic endothelial cells via integrin α9β1 and binding to CCL21, tenascin-C immobilized CD11c+ cells in the stroma. Inversion of the lymph node-to-tumor CCL21 gradient, recruitment of T regulatory cells, high expression of anti-inflammatory cytokines, and matrisomal components were hallmarks of the tenascin-C-instructed lymphoid stroma. Ablation of tenascin-C or CCR7 blockade inhibited the lymphoid immune-suppressive stromal properties, reducing tumor growth, progression, and metastasis. Thus, targeting CCR7 could be relevant in human head and neck tumors, as high tenascin-C expression and an immune-suppressive stroma correlate to poor patient survival.}, keywords = {Dumortier, I2CT, Team-Dumortier, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{lin_comparative_2020, title = {Comparative Effects of Graphene and Molybdenum Disulfide on Human Macrophage Toxicity}, author = {Hazel Lin and Ding‐Kun Ji and Matteo Andrea Lucherelli and Giacomo Reina and Stefano Ippolito and Paolo Samorì and Alberto Bianco}, url = {https://onlinelibrary.wiley.com/doi/10.1002/smll.202002194}, doi = {10.1002/smll.202002194}, issn = {1613-6810, 1613-6829}, year = {2020}, date = {2020-09-01}, urldate = {2020-11-20}, journal = {Small}, volume = {16}, number = {35}, pages = {2002194}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{newman_nose--brain_2020, title = {Nose-to-Brain Translocation and Cerebral Biodegradation of Thin Graphene Oxide Nanosheets}, author = {Leon Newman and Artur Filipe Rodrigues and Dhifaf A Jasim and Isabella Anna Vacchi and Cécilia Ménard-Moyon and Alberto Bianco and Cyrill Bussy and Kostas Kostarelos}, url = {https://linkinghub.elsevier.com/retrieve/pii/S2666386420301879}, doi = {10.1016/j.xcrp.2020.100176}, issn = {26663864}, year = {2020}, date = {2020-09-01}, urldate = {2020-11-20}, journal = {Cell Reports Physical Science}, volume = {1}, number = {9}, pages = {100176}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{reina_hard_2020, title = {Hard Nanomaterials in Time of Viral Pandemics}, author = {Giacomo Reina and Shiyuan Peng and Lucas Jacquemin and Andrés Felipe Andrade and Alberto Bianco}, url = {https://pubs.acs.org/doi/10.1021/acsnano.0c04117}, doi = {10.1021/acsnano.0c04117}, issn = {1936-0851, 1936-086X}, year = {2020}, date = {2020-08-01}, urldate = {2020-11-20}, journal = {ACS Nano}, volume = {14}, number = {8}, pages = {9364--9388}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pelin_partial_2020, title = {Partial Reversibility of the Cytotoxic Effect Induced by Graphene-Based Materials in Skin Keratinocytes}, author = {Marco Pelin and Hazel Lin and Arianna Gazzi and Silvio Sosa and Cristina Ponti and Amaya Ortega and Amaia Zurutuza and Ester Vázquez and Maurizio Prato and Aurelia Tubaro and Alberto Bianco}, url = {https://www.mdpi.com/2079-4991/10/8/1602}, doi = {10.3390/nano10081602}, issn = {2079-4991}, year = {2020}, date = {2020-08-01}, urldate = {2020-11-20}, journal = {Nanomaterials}, volume = {10}, number = {8}, pages = {1602}, abstract = {In the frame of graphene-based material (GBM) hazard characterization, particular attention should be given to the cutaneous effects. Hence, this study investigates if HaCaT skin keratinocytes exposed to high concentrations of few-layer graphene (FLG) or partially dehydrated graphene oxide (d-GO) for a short time can recover from the cytotoxic insult, measured by means of cell viability, mitochondrial damage and oxidative stress, after GBM removal from the cell medium. When compared to 24 or 72 h continuous exposure, recovery experiments suggest that the cytotoxicity induced by 24 h exposure to GBM is only partially recovered after 48 h culture in GBM-free medium. This partial recovery, higher for FLG as compared to GO, is not mediated by autophagy and could be the consequence of GBM internalization into cells. The ability of GBMs to be internalized inside keratinocytes together with the partial reversibility of the cellular damage is important in assessing the risk associated with skin exposure to GBM-containing devices.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Unusual tertiary pairs in eukaryotic tRNA-Ala}, author = {E Westhof and S Liang and X Tong and X Ding and L Zheng and F Dai}, url = {https://pubmed.ncbi.nlm.nih.gov/32737189/}, doi = {10.1261/rna.076299.120}, year = {2020}, date = {2020-07-31}, journal = {RNA}, volume = {26}, number = {11}, pages = {1519-1529}, abstract = {tRNA molecules have well-defined sequence conservations that reflect the conserved tertiary pairs maintaining their architecture and functions during the translation processes. An analysis of aligned tRNA sequences present in the GtRNAdb data base (The Lowe Lab, University of California, Santa Cruz) led to surprising conservations on some cytosolic tRNAs specific for Alanine compared to other tRNA species, including tRNAs specific for Glycine. First, besides the well-known G3oU70 base pair in the amino acid stem, there is the frequent occurrence of a second wobble pair at G30oU40, a pair overwhelmingly observed as a Watson-Crick pair throughout phylogeny. Secondly, the tertiary pair R15/Y48 occurs as a purine-purine R15/A48 pair. Finally, the conserved T54/A58 pair maintaining the fold of the T-loop is observed as a purine-purine A54/A58 pair. The R15/A48 and A54/A58 pairs always occur together. The G30oU40 pair occurs alone or together with these other two pairs. The pairing variations are observed to variable extent depending on phylogeny. Among eukaryotes, insects display simultaneously all variations, while mammals present either the G30oU40 pair or both R15/A48 and A54/A58. tRNAs with the anticodon 34A(I)GC36 are the most prone to display all those pair variations in mammals and insects. tRNAs with anticodon Y34GC36 have preferentially G30oU40 only. These unusual pairs are not observed in bacterial, or archaeal tRNAs, probably because of the avoidance of A34-containing anticodons in 4-codon boxes. Among eukaryotes, these unusual pairing features were not observed in fungi and nematodes. These unusual structural features may affect transcription rates (e.g. 54/58) or ribosomal translocation (30/40).}, keywords = {Ala Gly insects mammals tRNA, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{Ryckelynck2020, title = {Development and Applications of Fluorogen/Light-Up RNA Aptamer Pairs for RNA Detection and More}, author = {M Ryckelynck}, url = {https://link.springer.com/protocol/10.1007%2F978-1-0716-0712-1_5}, doi = {10.1007/978-1-0716-0712-1_5 }, year = {2020}, date = {2020-07-25}, journal = {RNA Tagging}, volume = {2166}, pages = {73-102}, publisher = {Methods in Molecular Biology}, abstract = {The central role of RNA in living systems made it highly desirable to have noninvasive and sensitive technologies allowing for imaging the synthesis and the location of these molecules in living cells. This need motivated the development of small pro-fluorescent molecules called “fluorogens” that become fluorescent upon binding to genetically encodable RNAs called “light-up aptamers.” Yet, the development of these fluorogen/light-up RNA pairs is a long and thorough process starting with the careful design of the fluorogen and pursued by the selection of a specific and efficient synthetic aptamer. This chapter summarizes the main design and the selection strategies used up to now prior to introducing the main pairs. Then, the vast application potential of these molecules for live-cell RNA imaging and other applications is presented and discussed.}, keywords = {aptamer, biosensing, Engineering, fluorogen, Functional screening, Live-cell imaging, RNA, RYCKELYNCK, SELEX, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{veillard2020, title = {Antibacterial activity of a dual peptide targeting the Escherichia coli sliding clamp and the ribosome}, author = {Christophe André and Florian Veillard and Philippe Wolff and Anne-marie Lobstein and Guillaume Compain and Clément Monsarrat and Jean-marc Reichhart and Camille Noûs and Dominique Burnouf and Gilles Guichard and Jerome Wagner }, url = {https://pubs.rsc.org/en/content/articlelanding/2020/CB/D0CB00060D#!divAbstract}, doi = {10.1039/D0CB00060D}, year = {2020}, date = {2020-07-16}, journal = {RSC Chemical Biology}, volume = {1}, number = {3}, pages = {137-147}, abstract = {The bacterial processivity factor, or sliding clamp (SC), is a target of choice for new antibacterial drugs development. We have previously developed peptides that target Escherichia coli SC and block its interaction with DNA polymerases in vitro. Here, one such SC binding peptide was fused to a Proline-rich AntiMicrobial Peptide (PrAMP) to allow its internalization into E. coli cells. Co-immunoprecipitation assays with a N-terminally modified bifunctional peptide that still enters the bacteria but fails to interact with the bacterial ribosome, the major target of PrAMPs, demonstrate that it actually interacts with the bacterial SC. Moreover, when compared to SC non-binding controls, this peptide induces a ten-fold higher antibacterial activity against E. coli, showing that the observed antimicrobial activity is linked to SC binding. Finally, an unmodified bifunctional compound significantly increases the survival of Drosophila melanogaster flies challenged by an E. coli infection. Our study demonstrates the potential of PrAMPs to transport antibiotics into the bacterial cytoplasm and validates the development of drugs targeting the bacterial processivity factor of Gram-negative bacteria as a promising new class of antibiotics.}, keywords = {Antibacterial, Antimicrobial peptides, Bacterial, clamp, E. coli, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{kroll-hermi_proteasome_2020, title = {Proteasome subunit PSMC3 variants cause neurosensory syndrome combining deafness and cataract due to proteotoxic stress.}, author = {Ariane Kröll-Hermi and Frédéric Ebstein and Corinne Stoetzel and Véronique Geoffroy and Elise Schaefer and Sophie Scheidecker and Séverine Bär and Masanari Takamiya and Koichi Kawakami and Barbara A Zieba and Fouzia Studer and Valerie Pelletier and Carine Eyermann and Claude Speeg-Schatz and Vincent Laugel and Dan Lipsker and Florian Sandron and Steven McGinn and Anne Boland and Jean-François Deleuze and Lauriane Kuhn and Johana Chicher and Philippe Hammann and Sylvie Friant and Christelle Etard and Elke Krüger and Jean Muller and Uwe Strähle and Hélène Dollfus}, doi = {10.15252/emmm.201911861}, issn = {1757-4684 1757-4676 1757-4676}, year = {2020}, date = {2020-07-01}, journal = {EMBO molecular medicine}, volume = {12}, number = {7}, pages = {e11861}, abstract = {The ubiquitin-proteasome system degrades ubiquitin-modified proteins to maintain protein homeostasis and to control signalling. Whole-genome sequencing of patients with severe deafness and early-onset cataracts as part of a neurological, sensorial and cutaneous novel syndrome identified a unique deep intronic homozygous variant in the PSMC3 gene, encoding the proteasome ATPase subunit Rpt5, which lead to the transcription of a cryptic exon. The proteasome content and activity in patient's fibroblasts was however unaffected. Nevertheless, patient's cells exhibited impaired protein homeostasis characterized by accumulation of ubiquitinated proteins suggesting severe proteotoxic stress. Indeed, the TCF11/Nrf1 transcriptional pathway allowing proteasome recovery after proteasome inhibition is permanently activated in the patient's fibroblasts. Upon chemical proteasome inhibition, this pathway was however impaired in patient's cells, which were unable to compensate for proteotoxic stress although a higher proteasome content and activity. Zebrafish modelling for knockout in PSMC3 remarkably reproduced the human phenotype with inner ear development anomalies as well as cataracts, suggesting that Rpt5 plays a major role in inner ear, lens and central nervous system development.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{Lalouna2020, title = {Manganese: the battle of the two armies}, author = {D Lalaouna and P Romby}, url = {https://univoak.eu/islandora/object/islandora:105630}, year = {2020}, date = {2020-07-01}, journal = {Project Repository Journal}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{jacoberger-foissac_liposomes_2020, title = {Liposomes as tunable platform to decipher the antitumor immune response triggered by TLR and NLR agonists}, author = {Célia Jacoberger-Foissac and Hanadi Saliba and May Wantz and Cendrine Seguin and Vincent Flacher and Benoît Frisch and Béatrice Heurtault and Sylvie Fournel}, doi = {10.1016/j.ejpb.2020.05.026}, issn = {1873-3441}, year = {2020}, date = {2020-07-01}, journal = {European Journal of Pharmaceutics and Biopharmaceutics: Official Journal of Arbeitsgemeinschaft Fur Pharmazeutische Verfahrenstechnik e.V}, volume = {152}, pages = {348--357}, abstract = {Liposomes are powerful tools for the optimization of peptides and adjuvant composition in cancer vaccines. Here, we take advantage of a liposomal platform versatility to develop three vaccine candidates associating a peptide from HA influenza virus protein as CD4 epitope, a peptide from HPV16 E7 oncoprotein as CD8 epitope and TLR4, TLR2/6 or NOD1 agonists as adjuvant. Liposomal vaccine containing MPLA (TLR4 liposomes), are the most effective treatment against the HPV-transformed orthotopic lung tumor mouse model, TC-1. This vaccine induces a potent Th1-oriented antitumor immunity, which leads to a significant reduction in tumor growth and a prolonged survival of mice, even when injected after tumor appearance. This efficacy is dependent on CD8+ T cells. Subcutaneous injection of this treatment induces the migration of skin DCs to draining lymph nodes. Interestingly, TLR2/6 liposomes trigger a weaker Th1-immune response which is not sufficient for the induction of a prolonged antitumor activity. Although NOD1 liposome treatment results in the control of early tumor growth, it does not extend mice survival. Surprisingly, the antitumor activity of NOD1 vaccine is not associated with a specific adaptive immune response. This study shows that our modulable platform can be used for the strategical development of vaccines.}, keywords = {Delivery system, HPV-transformed pulmonary tumors, Liposomal nanoparticles, Team-Mueller, Therapeutic vaccines, Toll-like and nod-like receptor agonists}, pubstate = {published}, tppubtype = {article} } @article{Liégeois2020b, title = {An atlas for hemocytes in an insect}, author = {Samuel Liégeois and Dominique Ferrandon}, editor = {Elife}, url = {https://elifesciences.org/articles/59113}, doi = {10.7554/eLife.59113}, year = {2020}, date = {2020-06-30}, journal = {Elife}, abstract = {Single-cell RNA sequencing has revealed distinct subpopulations of hemocytes in fruit fly larvae}, keywords = {Drosophila, ferrandon, Hemocytes, M3i, single-cell}, pubstate = {published}, tppubtype = {article} } @article{rodrigues_sizedependent_2020, title = {Size‐Dependent Pulmonary Impact of Thin Graphene Oxide Sheets in Mice: Toward Safe‐by‐Design}, author = {Artur Filipe Rodrigues and Leon Newman and Dhifaf Jasim and Sourav P Mukherjee and Jun Wang and Isabella A Vacchi and Cécilia Ménard‐Moyon and Alberto Bianco and Bengt Fadeel and Kostas Kostarelos and Cyrill Bussy}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/advs.201903200}, doi = {10.1002/advs.201903200}, issn = {2198-3844, 2198-3844}, year = {2020}, date = {2020-06-01}, urldate = {2020-11-20}, journal = {Advanced Science}, volume = {7}, number = {12}, pages = {1903200}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{orecchioni_toward_2020, title = {Toward High‐Dimensional Single‐Cell Analysis of Graphene Oxide Biological Impact: Tracking on Immune Cells by Single‐Cell Mass Cytometry}, author = {Marco Orecchioni and Valentina Bordoni and Claudia Fuoco and Giacomo Reina and Hazel Lin and Martina Zoccheddu and Acelya Yilmazer and Barbara Zavan and Gianni Cesareni and Davide Bedognetti and Alberto Bianco and Lucia Gemma Delogu}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/smll.202000123}, doi = {10.1002/smll.202000123}, issn = {1613-6810, 1613-6829}, year = {2020}, date = {2020-05-01}, urldate = {2020-11-20}, journal = {Small}, volume = {16}, number = {21}, pages = {2000123}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{vacchi_strategies_2020, title = {Strategies for the Controlled Covalent Double Functionalization of Graphene Oxide}, author = {Isabella A Vacchi and Shi Guo and Jésus Raya and Alberto Bianco and Cécilia Ménard‐Moyon}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/chem.201905785}, doi = {10.1002/chem.201905785}, issn = {0947-6539, 1521-3765}, year = {2020}, date = {2020-05-01}, urldate = {2020-11-20}, journal = {Chemistry – A European Journal}, volume = {26}, number = {29}, pages = {6591--6598}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{perraud_phenotypic_2020, title = {Phenotypic Adaption of Pseudomonas aeruginosa by Hacking Siderophores Produced by Other Microorganisms.}, author = {Quentin Perraud and Paola Cantero and Béatrice Roche and Véronique Gasser and Vincent P Normant and Lauriane Kuhn and Philippe Hammann and Gaëtan L A Mislin and Laurence Ehret-Sabatier and Isabelle J Schalk}, doi = {10.1074/mcp.RA119.001829}, issn = {1535-9484 1535-9476 1535-9476}, year = {2020}, date = {2020-04-01}, journal = {Molecular & cellular proteomics : MCP}, volume = {19}, number = {4}, pages = {589--607}, abstract = {Bacteria secrete siderophores to access iron, a key nutrient poorly bioavailable and the source of strong competition between microorganisms in most biotopes. Many bacteria also use siderophores produced by other microorganisms (exosiderophores) in a piracy strategy. Pseudomonas aeruginosa, an opportunistic pathogen, produces two siderophores, pyoverdine and pyochelin, and is also able to use a panel of exosiderophores. We first investigated expression of the various iron-uptake pathways of P. aeruginosa in three different growth media using proteomic and RT-qPCR approaches and observed three different phenotypic patterns, indicating complex phenotypic plasticity in the expression of the various iron-uptake pathways. We then investigated the phenotypic plasticity of iron-uptake pathway expression in the presence of various exosiderophores (present individually or as a mixture) under planktonic growth conditions, as well as in an epithelial cell infection assay. In all growth conditions tested, catechol-type exosiderophores were clearly more efficient in inducing the expression of their corresponding transporters than the others, showing that bacteria opt for the use of catechol siderophores to access iron when they are present in the environment. In parallel, expression of the proteins of the pyochelin pathway was significantly repressed under most conditions tested, as well as that of proteins of the pyoverdine pathway, but to a lesser extent. There was no effect on the expression of the heme and ferrous uptake pathways. Overall, these data provide precise insights on how P. aeruginosa adjusts the expression of its various iron-uptake pathways (phenotypic plasticity and switching) to match varying levels of iron and competition.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{janel-bintz_proteomic_2020, title = {Proteomic Analysis of DNA Synthesis on a Structured DNA Template in Human Cellular Extracts: Interplay Between NHEJ and Replication-Associated Proteins.}, author = {Régine Janel-Bintz and Lauriane Kuhn and Philippe Frit and Johana Chicher and Jérôme Wagner and Lajos Haracska and Philippe Hammann and Agnès M Cordonnier}, doi = {10.1002/pmic.201900184}, issn = {1615-9861 1615-9853}, year = {2020}, date = {2020-02-01}, journal = {Proteomics}, volume = {20}, number = {3-4}, pages = {e1900184}, abstract = {It is established that short inverted repeats trigger base substitution mutagenesis in human cells. However, how the replication machinery deals with structured DNA is unknown. It has been previously reported that in human cell-free extracts, DNA primer extension using a structured single-stranded template is transiently blocked at DNA hairpins. Here, the proteomic analysis of proteins bound to the DNA template is reported and evidence that the DNA-PK complex (DNA-PKcs and the Ku heterodimer) recognizes, and is activated by, structured single-stranded DNA is provided. Hijacking the DNA-PK complex by double-stranded oligonucleotides results in a large removal of the pausing sites and an elevated DNA extension efficiency. Conversely, DNA-PKcs inhibition results in its stabilization on the template, along with other proteins acting downstream in the Non-Homologous End-Joining (NHEJ) pathway, especially the XRCC4-DNA ligase 4 complex and the cofactor PAXX. Retention of NHEJ factors to the DNA in the absence of DNA-PKcs activity correlates with additional halts of primer extension, suggesting that these proteins hinder the progression of the DNA synthesis at these sites. Overall these results raise the possibility that, upon binding to hairpins formed onto ssDNA during fork progression, the DNA-PK complex interferes with replication fork dynamics in vivo.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{Aguiar_2020, title = {A single unidirectional piRNA cluster similar to the flamenco locus is the major source of EVE-derived transcription and small RNAs in Aedes aegypti mosquitoes }, author = {ERGR Aguiar and JPP de Almeida and LR Queiroz and LS Oliveira and RP Olmo and IJDS de Faria and JL Imler and A Gruber and BJ Matthews and JT Marques}, url = {https://rnajournal.cshlp.org/content/26/5/581.long}, doi = {10.1261/rna.073965.119}, year = {2020}, date = {2020-01-29}, journal = {RNA}, volume = {26}, number = {5}, pages = {581-594}, abstract = {Endogenous viral elements (EVEs) are found in many eukaryotic genomes. Despite considerable knowledge about genomic elements such as transposons (TEs) and retroviruses, we still lack information about nonretroviral EVEs. Aedes aegypti mosquitoes have a highly repetitive genome that is covered with EVEs. Here, we identified 129 nonretroviral EVEs in the AaegL5 version of the A. aegypti genome. These EVEs were significantly associated with TEs and preferentially located in repeat-rich clusters within intergenic regions. Genome-wide transcriptome analysis showed that most EVEs generated transcripts although only around 1.4% were sense RNAs. The majority of EVE transcription was antisense and correlated with the generation of EVE-derived small RNAs. A single genomic cluster of EVEs located in a 143 kb repetitive region in chromosome 2 contributed with 42% of antisense transcription and 45% of small RNAs derived from viral elements. This region was enriched for TE-EVE hybrids organized in the same coding strand. These generated a single long antisense transcript that correlated with the generation of phased primary PIWI-interacting RNAs (piRNAs). The putative promoter of this region had a conserved binding site for the transcription factor Cubitus interruptus, a key regulator of the flamenco locus in Drosophila melanogaster Here, we have identified a single unidirectional piRNA cluster in the A. aegypti genome that is the major source of EVE transcription fueling the generation of antisense small RNAs in mosquitoes. We propose that this region is a flamenco-like locus in A. aegypti due to its relatedness to the major unidirectional piRNA cluster in Drosophila melanogaster. }, keywords = {A. aegypti, Aedes aegypti, endogenous viral elements, EVE, flamenco locus, imler, M3i, Marques, piRNA, piRNAs, RNA Interference}, pubstate = {published}, tppubtype = {article} } @inbook{Liégeois2020, title = {Methods to Quantify In Vivo Phagocytic Uptake and Opsonization of Live or Killed Microbes in Drosophila melanogaster}, author = {Samuel Liégeois and Wenhui Wang and Dominique Ferrandon}, editor = {Springer Protocols Handbooks}, url = {https://link.springer.com/protocol/10.1007%2F978-1-0716-0259-1_5}, doi = {10.1007/978-1-0716-0259-1_5}, isbn = {9781071602584}, year = {2020}, date = {2020-01-28}, pages = {79}, publisher = {Springer Protocols Handbooks}, chapter = {5}, series = {Immunity in Insects}, abstract = {Here we describe different phagocytosis assays in Drosophila, using various killed or live microbes (bacteria and fungi). Different ex vivo and in vivo approaches are shown, to quantify larval and adult phagocytosis of microorganisms by hemocytes. We also explain how to perform an in vivo opsonization assay. Altogether, these protocols represent a useful range of tools to the researcher interested in the detailed analysis of phagocytosis in the context of the study of host-pathogen relationships.}, keywords = {Drosophila, ferrandon, M3i, opsonization, Phagocytosis, protocol}, pubstate = {published}, tppubtype = {inbook} } @article{goto2020, title = {The Kinase IKKβ Regulates a STING-and NF-κB-Dependent Antiviral Response Pathway in Drosophila}, author = {Akira Goto and Kiyoshi Okado and Nelson Martins and Hua Cai and Vincent Barbier and Olivier Lamiable and Laurent Troxler and Estelle Santiago and Lauriane Kuhn and Donggi Paik and Neal Silverman and Andreas Holleufer and Rune Hartmann and Jiyong Liu and Tao Peng and Jules A Hoffmann and Carine Meignin and Laurent Daeffler and Jean-Luc Imler }, url = {https://www-sciencedirect-com.insb.bib.cnrs.fr/science/article/pii/S107476131930528X}, doi = {10.1016/j.immuni.2019.12.009 }, year = {2020}, date = {2020-01-14}, journal = {Immunity}, volume = {52}, number = {1}, pages = {200}, abstract = {Antiviral immunity inDrosophilainvolves RNA inter-ference and poorly characterized inducible re-sponses. Here, we showed that two components ofthe IMD pathway, the kinase dIKKband the tran-scription factor Relish, were required to controlinfection by two picorna-like viruses. We identifieda set of genes induced by viral infection and regu-lated by dIKKband Relish, which included an ortho-log of STING. We showed that dSTING participatedin the control of infection by picorna-like viruses,acting upstream of dIKKbto regulate expression ofNazo, an antiviral factor. Our data reveal an antiviralfunction for STING in an animal model devoid of inter-ferons and suggest an evolutionarily ancient role forthis molecule in antiviral immunity.}, keywords = {antiviral, Drosophila, hoffmann, imler, Kinase, M3i, meignin, STING}, pubstate = {published}, tppubtype = {article} } @article{pmid32843367, title = {In vitro studies provide insight into effects of Đicer-2 helicase mutations in Đrosophila melanogaster}, author = {H M Donelick and L Talide and M Bellet and P J Aruscavage and E Lauret and E R G R Aguiar and J T Marques and C Meignin and B L Bass}, year = {2020}, date = {2020-01-01}, journal = {RNA}, volume = {26}, number = {12}, pages = {1847--1861}, note = {[PubMed Central:hrefhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC7668257PMC7668257] [DOI:hrefhttps://dx.doi.org/10.1261/rna.077289.12010.1261/rna.077289.120] [PubMed:hrefhttps://www.ncbi.nlm.nih.gov/pubmed/2136255421362554]}, keywords = {antiviral innate immunity, Dicer-2, Drosophila melanogaster, M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{pmid32002191, title = {Wide spectrum and high frequency of genomic structural variation, including transposable elements, in large double-stranded ĐNA viruses}, author = {V Loiseau and E A Herniou and Y Moreau and N Lévêque and C Meignin and L Daeffler and B Federici and R Cordaux and C Gilbert}, year = {2020}, date = {2020-01-01}, journal = {Virus Evol}, volume = {6}, number = {1}, pages = {vez060}, note = {[PubMed Central:hrefhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC6983493PMC6983493] [DOI:hrefhttps://dx.doi.org/10.1093/ve/vez06010.1093/ve/vez060] [PubMed:hrefhttps://www.ncbi.nlm.nih.gov/pubmed/2739260627392606]}, keywords = {M3i, meignin, virus}, pubstate = {published}, tppubtype = {article} } @article{brulefert_vitamin_2020, title = {Vitamin D3-elicited CD14+ human skin dendritic cells promote thymic stromal lymphopoietin-independent type 2 T-helper responses}, author = {Adrien Brulefert and Astrid Hoste and Quentin Muller and Jean-Daniel Fauny and Christopher G Mueller and Vincent Flacher}, doi = {10.1111/all.14718}, issn = {1398-9995}, year = {2020}, date = {2020-01-01}, journal = {Allergy}, abstract = {BACKGROUND: Immune modulation by vitamin D3 through dendritic cells (DCs) remains controversial. Human DCs exposed in vitro counteract type-1 T-helper (Th1) differentiation and induce regulatory T cells. However, cutaneous application on mice promotes Th2-driven inflammation resembling atopic dermatitis and relying on thymic stromal lymphopoietin (TSLP) from keratinocytes and T-cell orientation by TSLP-stimulated skin DCs. We studied the effects of vitamin D3 in human skin, focusing on TSLP production and the role of skin DCs in T-cell differentiation. METHODS: Human healthy skin explants were exposed in vitro to vitamin D3 analogs. Migrating DCs were analyzed and TSLP quantified in the supernatant. Allogeneic naïve CD4+ T cells were cocultured with DCs to assess their proliferation and cytokine production. RESULTS: Vitamin D3 induced skin DCs to differentiate Th2 cells producing IL-4 and IL-13. Vitamin D3 triggered TSLP release in textasciitilde30% of skin explants, correlating with IL-13 detection in Th2 cells. In these donors, blocking TSLP receptor during skin explant cultures abrogated IL-13 production, yet IL-4+ Th2 cells were unaffected. Among skin DCs emerged CD14+ cells that had responded directly to vitamin D3 and differed from classical CD14+ dermal emigrants. Vitamin D3-elicited CD14+ DCs sufficed to promote IL-4+ Th2 cells in a TSLP-independent manner. CONCLUSION: Vitamin D3, despite inducing TSLP in some donors, had a direct influence on skin DCs, affecting their phenotype and ability to drive Th2 responses independently of TSLP. Our findings pave the way toward in vitro systems that accurately model human cutaneous Th2 responses, notably involved in atopic dermatitis.}, keywords = {atopic dermatitis, Dendritic cell, T helper 2, Team-Mueller, thymic stromal lymphopoietin, vitamin D3}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Native Electrospray Ionization Mass Spectrometry of RNA-Ligand Complexes}, author = {P Wolff and E Ennifar}, editor = {V Arluison and F Wien}, url = {https://pubmed.ncbi.nlm.nih.gov/32006311}, doi = {10.1007/978-1-0716-0278-2_9}, isbn = {32006311}, year = {2020}, date = {2020-01-01}, booktitle = {RNA Spectroscopy: Methods and Protocols}, volume = {2113}, pages = {111-118}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Native electrospray ionization mass spectrometry (native ESI-MS) is a powerful tool to investigate non-covalent biomolecular interactions. It has been widely used to study protein complexes, but only few examples are described for the analysis of complexes involving RNA-RNA interactions. Here, we provide a detailed protocol for native ESI-MS analysis of RNA complexes. As an example, we present the analysis of the HIV-1 genomic RNA dimerization initiation site (DIS) extended duplex dimer bound to the aminoglycoside antibiotic lividomycin.}, keywords = {ARN-MS, ENNIFAR, Native mass spectrometry RNA, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Machine Learning of Reverse Transcription Signatures of Variegated Polymerases Allows Mapping and Discrimination of Methylated Purines in Limited Transcriptomes}, author = {S Werner and L Schmidt and V Marchand and T Kemmer and C Falschlunger and M V Sednev and G Bec and E Ennifar and C Höbartner and R Micura and Y Motorin and A Hildebrandt and M Helm}, url = {https://pubmed.ncbi.nlm.nih.gov/32095818}, doi = {10.1093/nar/gkaa113}, isbn = {32095818}, year = {2020}, date = {2020-01-01}, journal = {Nucleic Acids Res}, volume = {48}, number = {7}, pages = {3734-3746}, abstract = {Reverse transcription (RT) of RNA templates containing RNA modifications leads to synthesis of cDNA containing information on the modification in the form of misincorporation, arrest, or nucleotide skipping events. A compilation of such events from multiple cDNAs represents an RT-signature that is typical for a given modification, but, as we show here, depends also on the reverse transcriptase enzyme. A comparison of 13 different enzymes revealed a range of RT-signatures, with individual enzymes exhibiting average arrest rates between 20 and 75%, as well as average misincorporation rates between 30 and 75% in the read-through cDNA. Using RT-signatures from individual enzymes to train a random forest model as a machine learning regimen for prediction of modifications, we found strongly variegated success rates for the prediction of methylated purines, as exemplified with N1-methyladenosine (m1A). Among the 13 enzymes, a correlation was found between read length, misincorporation, and prediction success. Inversely, low average read length was correlated to high arrest rate and lower prediction success. The three most successful polymerases were then applied to the characterization of RT-signatures of other methylated purines. Guanosines featuring methyl groups on the Watson-Crick face were identified with high confidence, but discrimination between m1G and m22G was only partially successful. In summary, the results suggest that, given sufficient coverage and a set of specifically optimized reaction conditions for reverse transcription, all RNA modifications that impede Watson-Crick bonds can be distinguished by their RT-signature.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Site-Specific Spin Labeling of RNA for NMR and EPR Structural Studies}, author = {B Vileno and I Lebars}, editor = {V Arluison and F Wien}, url = {https://pubmed.ncbi.nlm.nih.gov/32006317}, doi = {10.1007/978-1-0716-0278-2_15}, isbn = {32006317}, year = {2020}, date = {2020-01-01}, booktitle = {RNA Spectroscopy: Methods and Protocols}, volume = {2113}, pages = {217-235}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Many RNA architectures were discovered to be involved in essential biological pathways acting as catalysts and/or regulators of gene expression, transcription, translation, splicing, or viral infection. The key to understand their diverse biological functions is to investigate their structure and dynamic. Nuclear Magnetic Resonance (NMR) is a powerful method to gain insight into these properties. However, the study of high-molecular-weight RNAs by NMR remains challenging. Advances in biochemical and NMR methods over the recent years allow to overcome the limitation of NMR. In particular, the incorporation of paramagnetic probes, coupled to the measurement of the induced effects on nuclear spins, has become an efficient tool providing long-range distance restraints and information on dynamic in solution. At the same time, the use of spin label enabled the application of Electron Paramagnetic Resonance (EPR) to study biological macromolecules. Combining NMR and EPR is emerging as a new approach to investigate the architecture of biological systems. Here, we describe an efficient protocol to introduce a paramagnetic probe into a RNA at a specific position. This method enables various combinations of isotopic labeling for NMR and is also of interest for EPR studies.}, keywords = {ENNIFAR, RNA NMR EPR Paramagnetic Spin-label, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {A unique ferrous iron binding mode is associated to large conformational changes for the transport protein FpvC of Pseudomonas aeruginosa}, author = {A Vigouroux and M Aumont-Nicaise and A Boussac and L Marty and L Lo Bello and P Legrand and K Brillet and I J Schalk and S Moréra}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31318478}, doi = {10.1111/febs.15004}, isbn = {31318478}, year = {2020}, date = {2020-01-01}, journal = {FEBS J}, volume = {287}, number = {2}, pages = {295-309}, abstract = {Pseudomonas aeruginosa secretes pyoverdine, a major siderophore to get access to iron, an essential nutrient. Pyoverdine scavenges ferric iron in the bacterial environment with the resulting complex internalized by bacteria. Iron release from pyoverdine in the periplasm involves an iron reduction by an inner membrane reductase and two solute-binding proteins (SBPs) FpvC and FpvF in association with their ABC transporter. FpvC and FpvF belong to two different subgroups of SBPs within the structural cluster A: FpvC and FpvF were proposed to be a metal-binding protein and a ferrisiderophore binding protein, respectively. Here, we report the redox state and the binding mode of iron to FpvC. We first solved the crystal structure of FpvC bound to a fortuitous Ni2+ by single anomalous dispersion method. Using a different protein purification strategy, we determined the structure of FpvC with manganese and iron, which binds to FpvC in a ferrous state as demonstrated by electron paramagnetic resonance. FpvC is the first example of a hexahistidine metal site among SBPs in which the Fe2+ redox state is stabilized under aerobic conditions. Using biophysics methods, we showed that FpvC reversibly bind a broad range of divalent ions. The structure of a mutant mimicking the apo FpvC reveals a protein in an open state with large conformational changes when compared with the metal-bound FpvC. These results highlight that the canonical metal site in FpvC is distinct from those yet described in SBPs and they provide new insights into the mechanism of PVD-Fe dissociation in P. aeruginosa. This article is protected by copyright. All rights reserved.}, keywords = {ENNIFAR, Pseudomonas aeruginosa iron pyoverdine siderophore solute-binding protein, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {2,6-Diaminopurine as a Highly Potent Corrector of UGA Nonsense Mutations}, author = {C Trzaska and S Amand and C Bailly and C Leroy and V Marchand and E Duvernois-Berthet and J M Saliou and H Benhabiles and E Werkmeister and T Chassat and R Guilbert and D Hannebique and A Mouray and M C Copin and P A Moreau and E Adriaenssens and A Kulozik and E Westhof and D Tulasne and Y Motorin and S Rebuffat and F Lejeune}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32198346?dopt=Abstract}, doi = {10.1038/s41467-020-15140-z}, isbn = {32198346}, year = {2020}, date = {2020-01-01}, journal = {Nat Commun}, volume = {11}, number = {1}, pages = {1509}, abstract = {Nonsense mutations cause about 10% of genetic disease cases, and no treatments are available. Nonsense mutations can be corrected by molecules with nonsense mutation readthrough activity. An extract of the mushroom Lepista inversa has recently shown high-efficiency correction of UGA and UAA nonsense mutations. One active constituent of this extract is 2,6-diaminopurine (DAP). In Calu-6 cancer cells, in which TP53 gene has a UGA nonsense mutation, DAP treatment increases p53 level. It also decreases the growth of tumors arising from Calu-6 cells injected into immunodeficient nude mice. DAP acts by interfering with the activity of a tRNA-specific 2'-O-methyltransferase (FTSJ1) responsible for cytosine 34 modification in tRNATrp. Low-toxicity and high-efficiency UGA nonsense mutation correction make DAP a good candidate for the development of treatments for genetic diseases caused by nonsense mutations.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure-Guided Engineering of the Homodimeric Mango-IV Fluorescence Turn-on Aptamer Yields an RNA FRET Pair}, author = {3rd R J Trachman and R Cojocaru and D Wu and G Piszczek and M Ryckelynck and P J Unrau and A R Ferré-D'Amaré}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32386573?dopt=Abstract}, doi = {10.1016/j.str.2020.04.007}, isbn = {32386573}, year = {2020}, date = {2020-01-01}, journal = {Structure}, volume = {28}, number = {7}, pages = {776-785.e773}, abstract = {Fluorescent RNA aptamers have been used in cells as biosensor reporters and tags for tracking transcripts. Recently, combined SELEX and microfluidic fluorescence sorting yielded three aptamers that activate fluorescence of TO1-Biotin: Mango-II, Mango-III, and Mango-IV. Of these, Mango-IV was best at imaging RNAs in both fixed and live mammalian cells. To understand how Mango-IV achieves activity in cells, we determined its crystal structure complexed with TO1-Biotin. The structure reveals a domain-swapped homodimer with two independent G-quadruplex fluorophore binding pockets. Structure-based analyses indicate that the Mango-IV core has relaxed fluorophore specificity, and a tendency to reorganize binding pocket residues. These molecular properties may endow it with robustness in the cellular milieu. Based on the domain-swapped structure, heterodimers between Mango-IV and the fluorescent aptamer iSpinach, joined by Watson-Crick base pairing, were constructed. These exhibited FRET between their respective aptamer-activated fluorophores, advancing fluorescent aptamer technology toward multi-color, RNA-based imaging of RNA coexpression and colocalization.}, keywords = {RNA aptamer RNA fluorescence RNA structure fluorescent dye structure-guided engineering, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Structural Analysis of RNA by Small-Angle X-ray Scattering}, author = {A Théobald-Dietrich and R de Wijn and K Rollet and A Bluhm and J Rudinger-Thirion and C Paulus and B Lorber and A Thureau and M Frugier and C Sauter}, editor = {V Arluison and F Wien}, url = {https://pubmed.ncbi.nlm.nih.gov/32006316}, doi = {10.1007/978-1-0716-0278-2_14}, isbn = {32006316}, year = {2020}, date = {2020-01-01}, booktitle = {RNA Spectroscopy: Methods and Protocols}, volume = {2113}, pages = {189-215}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Over the past two decades small-angle X-ray scattering (SAXS) has become a popular method to characterize solutions of biomolecules including ribonucleic acid (RNA). In an integrative structural approach, SAXS is complementary to crystallography, NMR, and electron microscopy and provides information about RNA architecture and dynamics. This chapter highlights the practical advantages of combining size-exclusion chromatography and SAXS at synchrotron facilities. It is illustrated by practical case studies of samples ranging from single hairpins and tRNA to a large IRES. The emphasis is also put on sample preparation which is a critical step of SAXS analysis and on optimized protocols for in vitro RNA synthesis ensuring the production of mg amount of pure and homogeneous molecules.}, keywords = {FRUGIER, IRES Integrative structural biology RNA SEC-SAXS Structure tRNA, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Structural Insights Into the Mammalian Late-Stage Initiation Complexes}, author = {A Simonetti and E Guca and A Bochler and L Kuhn and Y Hashem}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32268096?dopt=Abstract}, doi = {10.1016/j.celrep.2020.03.061}, isbn = {32268096}, year = {2020}, date = {2020-01-01}, journal = {Cell Rep}, volume = {31}, number = {1}, pages = {107497}, abstract = {In higher eukaryotes, the mRNA sequence in the direct vicinity of the start codon, called the Kozak sequence (CRCCaugG, where R is a purine), is known to influence the rate of the initiation process. However, the molecular basis underlying its role remains poorly understood. Here, we present the cryoelectron microscopy (cryo-EM) structures of mammalian late-stage 48S initiation complexes (LS48S ICs) in the presence of two different native mRNA sequences, β-globin and histone 4, at overall resolution of 3 and 3.5 Å, respectively. Our high-resolution structures unravel key interactions from the mRNA to eukaryotic initiation factors (eIFs): 1A, 2, 3, 18S rRNA, and several 40S ribosomal proteins. In addition, we are able to study the structural role of ABCE1 in the formation of native 48S ICs. Our results reveal a comprehensive map of ribosome/eIF-mRNA and ribosome/eIF-tRNA interactions and suggest the impact of mRNA sequence on the structure of the LS48S IC.}, keywords = {ENNIFAR, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Grad-cryo-EM: Tool to Isolate Translation Initiation Complexes From Rabbit Reticulocyte Lysate Suitable for Structural Studies}, author = {J Rol-Moreno and L Kuhn and S Marzi and A Simonetti}, editor = {V Arluison and F Wien}, url = {https://pubmed.ncbi.nlm.nih.gov/32006323}, doi = {10.1007/978-1-0716-0278-2_21}, isbn = {32006323}, year = {2020}, date = {2020-01-01}, urldate = {2020-01-01}, booktitle = {RNA Spectroscopy: Methods and Protocols}, volume = {2113}, pages = {329-339}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Since its development, single-particle cryogenic electron microscopy (cryo-EM) has played a central role in the study at medium resolution of both bacterial and eukaryotic ribosomal complexes. With the advent of the direct electron detectors and new processing software which allow obtaining structures at atomic resolution, formerly obtained only by X-ray crystallography, cryo-EM has become the method of choice for the structural analysis of the translation machinery. In most of the cases, the ribosomal complexes at different stages of the translation process are assembled in vitro from purified components, which limit the analysis to previously well-characterized complexes with known factors composition. The initiation phase of the protein synthesis is a very dynamic process during which several proteins interact with the translation apparatus leading to the formation of a chronological series of initiation complexes (ICs). Here we describe a method to isolate ICs assembled on natural in vitro transcribed mRNA directly from rabbit reticulocyte lysate (RRL) by sucrose density gradient centrifugation. The Grad-cryo-EM approach allows investigating structures and composition of intermediate ribosomal complexes prepared in near-native condition by cryo-EM and mass spectrometry analyses. This is a powerful approach, which could be used to study translation initiation of any mRNAs, including IRES containing ones, and which could be adapted to different cell extracts.}, keywords = {ENNIFAR, PPSE, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {The Expanding LARS2 Phenotypic Spectrum: HLASA, Perrault Syndrome With Leukodystrophy, and Mitochondrial Myopathy}, author = {L G Riley and J Rudinger-Thirion and M Frugier and M Wilson and M Luig and T I Alahakoon and C Y Nixon and E P Kirk and T Roscioli and S Lunke and Z Stark and K J Wierenga and S Palle and M Walsh and E Higgs and S Arbuckle and S Thirukeswaran and A G Compton and D R Thorburn and J Christodoulou}, url = {https://pubmed.ncbi.nlm.nih.gov/32442335/?dopt=Abstract}, doi = {doi: 10.1002/humu.24050}, isbn = {32442335}, year = {2020}, date = {2020-01-01}, journal = {Hum Mutat}, volume = {41}, number = {8}, pages = {1425-1434}, abstract = {Perrault syndrome is a rare autosomal recessive disorder characterized by sensorineural hearing loss (SNHL) in both sexes and primary ovarian insufficiency in 46, XX karyotype females. Biallelic variants in five genes are reported to be causative: HSD17B4, HARS2, LARS2, CLPP and C10orf2. Here we present eight families affected by Perrault syndrome. In five families we identified novel or previously reported variants in HSD17B4, LARS2, CLPP and C10orf2. The proband from each family was whole exome sequenced and variants confirmed by Sanger sequencing. A female was compound heterozygous for a known, p.(Gly16Ser) and novel, p.(Val82Phe) variant in D-bifunctional protein (HSD17B4). A family was homozygous for mitochondrial leucyl aminocyl tRNA synthetase (mtLeuRS) (LARS2) p.(Thr522Asn), previously associated with Perrault syndrome. A further family was compound heterozygous for mtLeuRS, p.(Thr522Asn) and a novel variant, p.(Met117Ile). Affected individuals with LARS2 variants had low frequency SNHL, a feature previously described in Perrault syndrome. A female with significant neurological disability was compound heterozygous for p.(Arg323Gln) and p.(Asn399Ser) variants in Twinkle (C10orf2). A male was homozygous for a novel variant in CLPP, p.(Cys144Arg). In three families there were no putative pathogenic variants in these genes confirming additional disease-causing genes remain unidentified. We have expanded the spectrum of disease-causing variants associated with Perrault syndrome.}, keywords = {FRUGIER, Perrault syndrome low frequency hearing loss primary ovarian insufficiency sensorineural hearing loss, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Nature of the Purine at Position 34 in tRNAs of 4-codon Boxes Is Correlated With Nucleotides at Positions 32 and 38 to Maintain Decoding Fidelity}, author = {K Pernod and L Schaeffer and J Chicher and E Hok and C Rick and R Geslain and G Eriani and E Westhof and M Ryckelynck and F Martin}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32266934?dopt=Abstract}, doi = {10.1093/nar/gkaa221}, isbn = {32266934}, year = {2020}, date = {2020-01-01}, journal = {Nucleic Acids Res}, volume = {48}, number = {11}, pages = {6170-6183}, abstract = {Translation fidelity relies essentially on the ability of ribosomes to accurately recognize triplet interactions between codons on mRNAs and anticodons of tRNAs. To determine the codon-anticodon pairs that are efficiently accepted by the eukaryotic ribosome, we took advantage of the IRES from the intergenic region (IGR) of the Cricket Paralysis Virus. It contains an essential pseudoknot PKI that structurally and functionally mimics a codon-anticodon helix. We screened the entire set of 4096 possible combinations using ultrahigh-throughput screenings combining coupled transcription/translation and droplet-based microfluidics. Only 97 combinations are efficiently accepted and accommodated for translocation and further elongation: 38 combinations involve cognate recognition with Watson-Crick pairs and 59 involve near-cognate recognition pairs with at least one mismatch. More than half of the near-cognate combinations (36/59) contain a G at the first position of the anticodon (numbered 34 of tRNA). G34-containing tRNAs decoding 4-codon boxes are almost absent from eukaryotic genomes in contrast to bacterial genomes. We reconstructed these missing tRNAs and could demonstrate that these tRNAs are toxic to cells due to their miscoding capacity in eukaryotic translation systems. We also show that the nature of the purine at position 34 is correlated with the nucleotides present at 32 and 38.}, keywords = {ERIANI, PPSE, RYCKELYNCK, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural Basis of Nanobody Recognition of Grapevine Fanleaf Virus and of Virus Resistance Loss}, author = {I Orlov and C Hemmer and L Ackerer and B Lorber and A Ghannam and V Poignavent and K Hleibieh and C Sauter and C Schmitt-Keichinger and L Belval and J M Hily and A Marmonier and V Komar and S Gersch and P Schellenberger and P Bron and E Vigne and S Muyldermans and O Lemaire and G Demangeat and C Ritzenthaler and B P Klaholz}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32371486?dopt=Abstract}, doi = {10.1073/pnas.1913681117}, isbn = {32371486}, year = {2020}, date = {2020-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {117}, number = {20}, pages = {10848-10855}, abstract = {Grapevine fanleaf virus (GFLV) is a picorna-like plant virus transmitted by nematodes that affects vineyards worldwide. Nanobody (Nb)-mediated resistance against GFLV has been created recently, and shown to be highly effective in plants, including grapevine, but the underlying mechanism is unknown. Here we present the high-resolution cryo electron microscopy structure of the GFLV-Nb23 complex, which provides the basis for molecular recognition by the Nb. The structure reveals a composite binding site bridging over three domains of one capsid protein (CP) monomer. The structure provides a precise mapping of the Nb23 epitope on the GFLV capsid in which the antigen loop is accommodated through an induced-fit mechanism. Moreover, we uncover and characterize several resistance-breaking GFLV isolates with amino acids mapping within this epitope, including C-terminal extensions of the CP, which would sterically interfere with Nb binding. Escape variants with such extended CP fail to be transmitted by nematodes linking Nb-mediated resistance to vector transmission. Together, these data provide insights into the molecular mechanism of Nb23-mediated recognition of GFLV and of virus resistance loss.}, keywords = {GFLV nanobody structural biology virus, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dynamic nanopore long-read sequencing analysis of HIV-1 splicing events during the early steps of infection}, author = {N Nguyen Quang and S Goudey and E Ségéral and A Mohammad and S Lemoine and C Blugeon and M Versapuech and J C Paillart and C Berlioz-Torrent and S Emiliani and S Gallois-Montbrun}, url = {https://pubmed.ncbi.nlm.nih.gov/32807178/}, doi = {10.1186/s12977-020-00533-1}, isbn = {32807178}, year = {2020}, date = {2020-01-01}, journal = {Retrovirology}, volume = {17}, number = {1}, pages = {25}, abstract = {Background Alternative splicing is a key step in Human Immunodeficiency Virus type 1 (HIV-1) replication that is tightly regulated both temporally and spatially. More than 50 different transcripts can be generated from a single HIV-1 unspliced pre-messenger RNA (pre-mRNA) and a balanced proportion of unspliced and spliced transcripts is critical for the production of infectious virions. Understanding the mechanisms involved in the regulation of viral RNA is therefore of potential therapeutic interest. However, monitoring the regulation of alternative splicing events at a transcriptome-wide level during cell infection is challenging. Here we used the long-read cDNA sequencing developed by Oxford Nanopore Technologies (ONT) to explore in a quantitative manner the complexity of the HIV-1 transcriptome regulation in infected primary CD4+ T cells. Results ONT reads mapping to the viral genome proved sufficiently long to span all possible splice junctions, even distant ones, and to be assigned to a total of 150 exon combinations. Fifty-three viral RNA isoforms, including 14 new ones were further considered for quantification. Relative levels of viral RNAs determined by ONT sequencing showed a high degree of reproducibility, compared favourably to those produced in previous reports and highly correlated with quantitative PCR (qPCR) data. To get further insights into alternative splicing regulation, we then compiled quantifications of splice site (SS) usage and transcript levels to build モsplice treesヤ, a quantitative representation of the cascade of events leading to the different viral isoforms. This approach allowed visualizing the complete rewiring of SS usages upon perturbation of SS D2 and its impact on viral isoform levels. Furthermore, we produced the first dynamic picture of the cascade of events occurring between 12 and 24 h of viral infection. In particular, our data highlighted the importance of non-coding exons in viral RNA transcriptome regulation. Conclusion ONT sequencing is a convenient and reliable strategy that enabled us to grasp the dynamic of the early splicing events modulating the viral RNA landscape in HIV-1 infected cells. Background}, keywords = {HIV RNA Alternative splicing Viral transcriptome ONT long-read sequencing, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Secondary structure of the SARS-CoV-2 5'-UTR}, author = {Z Miao and A Tidu and G Eriani and F Martin}, url = {https://pubmed.ncbi.nlm.nih.gov/32965173/}, doi = {10.1080/15476286.2020.1814556}, isbn = {32965173}, year = {2020}, date = {2020-01-01}, journal = {RNA Biol}, volume = {18}, number = {4}, pages = {447-456}, abstract = {The SARS-CoV-2, a positive-sense single-stranded RNA Coronavirus, is a global threat to human health. Thus, understanding its life cycle mechanistically would be important to facilitate the design of antiviral drugs. A key aspect of viral progression is the synthesis of viral proteins by the ribosome of the human host. In Coronaviruses, this process is regulated by the viral 5' and 3' untranslated regions (UTRs), but the precise regulatory mechanism has not yet been well understood. In particular, the 5'-UTR of the viral genome is most likely involved in translation initiation of viral proteins. Here, we performed inline probing and RNase V1 probing to establish a model of the secondary structure of SARS-CoV-2 5'-UTR. We found that the 5'-UTR contains stable structures including a very stable four-way junction close to the AUG start codon. Sequence alignment analysis of SARS-CoV-2 variants 5'-UTRs revealed a highly conserved structure with few co-variations that confirmed our secondary structure model based on probing experiments.}, keywords = {5ʹ-UTR SARS-CoV-2 probing secondary structure, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA-Puzzles Round IV: 3D Structure Predictions of Four Ribozymes and Two Aptamers}, author = {Z Miao and R W Adamiak and M Antczak and M J Boniecki and J M Bujnicki and S J Chen and C Y Cheng and Y Cheng and F C Chou and R Das and N V Dokholyan and F Ding and C Geniesse and Y Jiang and A Joshi and A Krokhotin and M Magnus and O Mailhot and F Major and T H Mann and P Piatkowski and R Pluta and M Popenda and J Sarzynska and L Sun and M Szachniuk and S Tian and J Wang and J Wang and A M Watkins and J Wiedemann and Y Xiao and X Xu and J D Yesselman and D Zhang and Y Zhang and Z Zhang and C Zhao and P Zhao and Y Zhou and T Zok and A Zyla and A Ren and R T Batey and B L Golden and L Huang and D M Lilley and Y Liu and D J Patel and E Westhof}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32371455?dopt=Abstract}, doi = {10.1261/rna.075341.120}, isbn = {32371455}, year = {2020}, date = {2020-01-01}, journal = {RNA}, volume = {26}, number = {8}, pages = {982-995}, abstract = {RNA-Puzzles is a collective endeavor dedicated to the advancement and improvement of RNA 3D structure prediction. With agreement from crystallographers, the RNA structures are predicted by various groups before the publication of the crystal structures. We now report the prediction of six RNA sequences: four structures of nucleolytic ribozymes and two of riboswitches. Systematic protocols for comparing models and crystal structures are described and analyzed. In these six puzzles, we discuss a) the comparison between the automated web server and human experts; b) the prediction of coaxial stacking; c) the prediction of structural details and ligand binding; d) the development of novel prediction methods; and e) the potential improvements to be made. It is illustrated that correct coaxial stacking and tertiary contacts are key for the prediction of RNA architecture, while ligand binding modes can be only predicted with low resolution and accurate ligand binding prediction still remains out of reach. All the predicted models are available for the future development of force field parameters and the improvement of comparison and assessment tools.}, keywords = {RNA structure aptamer modeing prediction ribozyme, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Differential evolution in 3'UTRs leads to specific gene expression in Staphylococcus}, author = {P Menendez-Gil and C J Caballero and A Catalan-Moreno and N Irurzun and I Barrio-Hernandez and I Caldelari and A Toledo-Arana}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32016395}, doi = {10.1093/nar/gkaa047}, isbn = {32016395}, year = {2020}, date = {2020-01-01}, journal = {Nucleic Acids Res}, volume = {48}, number = {5}, pages = {2544-2563}, abstract = {The evolution of gene expression regulation has contributed to species differentiation. The 3' untranslated regions (3'UTRs) of mRNAs include regulatory elements that modulate gene expression; however, our knowledge of their implications in the divergence of bacterial species is currently limited. In this study, we performed genome-wide comparative analyses of mRNAs encoding orthologous proteins from the genus Staphylococcus and found that mRNA conservation was lost mostly downstream of the coding sequence (CDS), indicating the presence of high sequence diversity in the 3'UTRs of orthologous genes. Transcriptomic mapping of different staphylococcal species confirmed that 3'UTRs were also variable in length. We constructed chimeric mRNAs carrying the 3'UTR of orthologous genes and demonstrated that 3'UTR sequence variations affect protein production. This suggested that species-specific functional 3'UTRs might be specifically selected during evolution. 3'UTR variations may occur through different processes, including gene rearrangements, local nucleotide changes, and the transposition of insertion sequences. By extending the conservation analyses to specific 3'UTRs, as well as the entire set of Escherichia coli and Bacillus subtilis mRNAs, we showed that 3'UTR variability is widespread in bacteria. In summary, our work unveils an evolutionary bias within 3'UTRs that results in species-specific non-coding sequences that may contribute to bacterial diversity.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA-Puzzles toolkit: a computational resource of RNA 3D structure benchmark datasets, structure manipulation, and evaluation tools}, author = {M Magnus and M Antczak and T Zok and J Wiedemann and P Lukasiak and Y Cao and J M Bujnicki and E Westhof and M Szachniuk and Z Miao}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31799609}, doi = {10.1093/nar/gkz1108.}, isbn = {31799609}, year = {2020}, date = {2020-01-01}, journal = {Nucleic Acids Res}, volume = {48}, number = {2}, pages = {576-588}, abstract = {Significant improvements have been made in the efficiency and accuracy of RNA 3D structure prediction methods during the succeeding challenges of RNA-Puzzles, a community-wide effort on the assessment of blind prediction of RNA tertiary structures. The RNA-Puzzles contest has shown, among others, that the development and validation of computational methods for RNA fold prediction strongly depend on the benchmark datasets and the structure comparison algorithms. Yet, there has been no systematic benchmark set or decoy structures available for the 3D structure prediction of RNA, hindering the standardization of comparative tests in the modeling of RNA structure. Furthermore, there has not been a unified set of tools that allows deep and complete RNA structure analysis, and at the same time, that is easy to use. Here, we present RNA-Puzzles toolkit, a computational resource including (i) decoy sets generated by different RNA 3D structure prediction methods (raw, for-evaluation and standardized datasets), (ii) 3D structure normalization, analysis, manipulation, visualization tools (RNA_format, RNA_normalizer, rna-tools) and (iii) 3D structure comparison metric tools (RNAQUA, MCQ4Structures). This resource provides a full list of computational tools as well as a standard RNA 3D structure prediction assessment protocol for the community.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {High-throughput Fluorescence-Based Screen Identifies the Neuronal microRNA miR-124 as a Positive Regulator of Alphavirus Infection}, author = {P Lopez and E Girardi and B C Mounce and A Weiss and B Chane-Woon-Ming and M Messmer and P Kaukinen and A Kopp and D Bortolamiol-Becet and A Fendri and M Vignuzzi and L Brino and S Pfeffer}, url = {https://pubmed.ncbi.nlm.nih.gov/32102877}, doi = {10.1128/JVI.02145-19}, isbn = {32102877}, year = {2020}, date = {2020-01-01}, journal = {J Virol}, volume = {94}, number = {9}, pages = {e02145-02119}, abstract = {Micro (mi)RNAs are small regulatory RNAs, which act by modulating the expression of target genes. In addition to their role in maintaining essential physiological functions in the cell, miRNAs can also regulate viral infections. They can do so directly by targeting RNAs of viral origin or indirectly by targeting host mRNAs and this can result in a positive or negative outcome for the virus. Here, we performed a fluorescence-based miRNA genome-wide screen in order to identify cellular miRNAs involved in the regulation of arbovirus infection in human cells. We identified sixteen miRNAs showing a positive effect on Sindbis virus (SINV) expressing GFP, among which a number of neuron-specific ones such as miR-124. We confirmed that overexpression of miR-124 increases both SINV structural protein translation and viral production and that this effect is mediated by its seed sequence. We further demonstrated that the SINV genome possesses a binding site for miR-124. Both inhibition of miR-124 or silent mutations to disrupt this binding site in the viral RNA abolished the positive regulation. We also proved that miR-124 inhibition reduces SINV infection in human differentiated neuronal cells. Finally, we showed that the proviral effect of miR-124 is conserved for other alphaviruses as its inhibition reduces chikungunya virus (CHIKV) viral production in human cells. Altogether, our work expands the panel of positive regulation of the viral cycle by direct binding of host miRNAs to the viral RNA and provides new insights into the role of cellular miRNAs as regulators of alphavirus infection.IMPORTANCEArthropod-borne (arbo) viruses are part of a class of pathogens that are transmitted to their final hosts by insects. Because of climate change, the habitat of some of these insects, such as mosquitoes, is shifting, thereby facilitating the emergence of viral epidemics. Among the pathologies associated with arboviruses infection, neurological diseases like meningitis or encephalitis represent a significant health burden. Using a genome-wide miRNA screen, we identified the neuronal miR-124 as a positive regulator of the Sindbis and chikungunya alphaviruses. We also showed that this effect was in part direct, thereby opening novel avenues to treat alphaviruses infection.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of Post-Transcriptional RNA Modifications by Sheathless Capillary Electrophoresis-High Resolution Mass Spectrometry}, author = {A Lechner and P Wolff and E Leize-Wagner and Y N François}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32343557?dopt=Abstract}, doi = {10.1021/acs.analchem.0c01345}, isbn = {32343557}, year = {2020}, date = {2020-01-01}, journal = {Anal Chem}, volume = {92}, number = {10}, pages = {7363-7370}, abstract = {Over the past decade there has been a growing interest in RNA modification analysis. High performance liquid chromatography-tandem mass spectrometry coupling (HPLC-MS/MS) is classically used to characterize post-transcriptional modifications of ribonucleic acids (RNAs). Here we propose a novel and simple workflow based on capillary zone electrophoresis-tandem mass spectrometry (CE-MS/MS), in positive mode, to characterize RNA modifications at nucleoside and oligonucleotide levels. By first totally digesting the purified RNA, prior to CE-MS/MS analysis, we were able to identify the nucleoside modifications. Then, using a bottom-up approach, sequencing of the RNAs and mapping of the modifications were performed. Sequence coverages from 68% to 97% were obtained for four tRNAs. Furthermore, unambiguous identification and mapping of several modifications were achieved.}, keywords = {ARN-MS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Short but Weak! The Z-DNA lone-pair···π Conundrum Challenges Standard Carbon Van Der Waals Radii}, author = {H Kruse and K Mrazikova and L D'Ascenzo and J Sponer and P Auffinger}, url = {https://pubmed.ncbi.nlm.nih.gov/32516461/}, doi = {10.1002/anie.202004201}, isbn = {32516461}, year = {2020}, date = {2020-01-01}, journal = {Angew Chem Int Ed Engl}, pages = {in press}, abstract = {Current interest for lone-pair···π (lp···π) non-covalent interactions is gaining momentum in biochemistry and (supramolecular)chemistry. However, the physico-chemical origin of the exceptionally short (≈2.8 Å) oxygen to nucleobase plane distances observed in prototypical Z-DNA CpG steps remains unclear. Accurate quantum mechanical calculations including SAPT2+3 interaction energy decompositions demonstrate that lp···π contacts do not result from n→π* orbital overlaps but from weak dispersion and electrostatics interactions combined with stereochemical effects imposed by the structural context. They also suggest that the carbon van der Waals (vdW) radii originally derived for sp3carbons should not be used for the smaller sp2 atoms. Using a more adapted carbon vdW radius results in lp···π contacts being no longer of the sub-vdW type. Overall, our findings challenge the whole lp···π concept that refers to elusive orbital interactions that cannot explain short contact distances.}, keywords = {ENNIFAR, lp···pi interactions molecular recognition non-covalent interactions van der Waals radii Z-DNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {NKNK: a New Essential Motif in the C-Terminal Domain of HIV-1 Group M Integrases}, author = {M Kanja and P Cappy and N Levy and O Oladosu and S Schmidt and P Rossolillo and F Winter and R Gasser and C Moog and M Ruff and M Negroni and D Lener}, url = {https://pubmed.ncbi.nlm.nih.gov/32727879/}, doi = {10.1128/JVI.01035-20}, isbn = {32727879}, year = {2020}, date = {2020-01-01}, journal = {J Virol}, volume = {94}, number = {20}, pages = {e01035-01020}, abstract = {Using coevolution network interference based on comparison of two phylogenetically distantly related isolates, one from the main group M and the other from the minor group O of HIV-1, we identify, in the C-terminal domain (CTD) of integrase, a new functional motif constituted by four noncontiguous amino acids (N222K240N254K273). Mutating the lysines abolishes integration through decreased 3' processing and inefficient nuclear import of reverse-transcribed genomes. Solution of the crystal structures of wild-type (wt) and mutated CTDs shows that the motif generates a positive surface potential that is important for integration. The number of charges in the motif appears more crucial than their position within the motif. Indeed, the positions of the K's could be permutated or additional K's could be inserted in the motif, generally without affecting integration per se Despite this potential genetic flexibility, the NKNK arrangement is strictly conserved in natural sequences, indicative of an effective purifying selection exerted at steps other than integration. Accordingly, reverse transcription was reduced even in the mutants that retained wt integration levels, indicating that specifically the wt sequence is optimal for carrying out the multiple functions that integrase exerts. We propose that the existence of several amino acid arrangements within the motif, with comparable efficiencies of integration per se, might have constituted an asset for the acquisition of additional functions during viral evolution.IMPORTANCE Intensive studies of HIV-1 have revealed its extraordinary ability to adapt to environmental and immunological challenges, an ability that is also at the basis of antiviral treatment escape. Here, by deconvoluting the different roles of the viral integrase in the various steps of the infectious cycle, we report how the existence of alternative equally efficient structural arrangements for carrying out one function opens up the possibility of adapting to the optimization of further functionalities exerted by the same protein. Such a property provides an asset to increase the efficiency of the infectious process. On the other hand, though, the identification of this new motif provides a potential target for interfering simultaneously with multiple functions of the protein.}, keywords = {evolution human immunodeficiency virus integrase phylogenetic groups, NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ascorbate maintains a low plasma oxygen level}, author = {L Injarabian and M Scherlinger and A Devin and S Ransac and J Lykkesfeldt and B S Marteyn}, url = {https://pubmed.ncbi.nlm.nih.gov/32606354/}, doi = {10.1038/s41598-020-67778-w}, isbn = {32606354}, year = {2020}, date = {2020-01-01}, journal = {Sci Rep}, volume = {10}, number = {1}, pages = {10659}, abstract = {In human blood, oxygen is mainly transported by red blood cells. Accordingly, the dissolved oxygen level in plasma is expected to be limited, although it has not been quantified yet. Here, by developing dedicated methods and tools, we determined that human plasma pO2 = 8.4 mmHg (1.1% O2). Oxygen solubility in plasma was believed to be similar to water. Here we reveal that plasma has an additional ascorbate-dependent oxygen-reduction activity. Plasma experimental oxygenation oxidizes ascorbate (49.5 μM in fresh plasma vs < 2 μM in oxidized plasma) and abolishes this capacity, which is restored by ascorbate supplementation. We confirmed these results in vivo, showing that the plasma pO2 is significantly higher in ascorbate-deficient guinea pigs (Ascorbateplasma < 2 μM), compared to control (Ascorbateplasma > 15 μM). Plasma low oxygen level preserves the integrity of oxidation-sensitive components such as ubiquinol. Circulating leucocytes are well adapted to these conditions, since the abundance of their mitochondrial network is limited. These results shed a new light on the importance of oxygen exposure on leucocyte biological study, in regards with the reducing conditions they encounter in vivo; but also, on the manipulation of blood products to improve their integrity and potentially improve transfusions' efficacy.}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Functional microRNA screen uncovers O-linked N-acetylglucosamine transferase as a host factor modulating hepatitis C virus morphogenesis and infectivity}, author = {K Herzog and S Bandiera and S Pernot and C Fauvelle and F Jühling and A Weiss and A Bull and S C Durand and B Chane-Woon-Ming and S Pfeffer and M Mercey and H Lerat and J C Meunier and W Raffelsberger and L Brino and T F Baumert and M B Zeisel}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31076402?dopt=Abstract}, doi = {10.1136/gutjnl-2018-317423}, isbn = {31076402}, year = {2020}, date = {2020-01-01}, journal = {Gut}, volume = {69}, number = {2}, pages = {380-392}, abstract = {OBJECTIVE: Infection of human hepatocytes by the hepatitis C virus (HCV) is a multistep process involving both viral and host factors. microRNAs (miRNAs) are small non-coding RNAs that post-transcriptionally regulate gene expression. Given that miRNAs were indicated to regulate between 30% and 75% of all human genes, we aimed to investigate the functional and regulatory role of miRNAs for the HCV life cycle. DESIGN: To systematically reveal human miRNAs affecting the HCV life cycle, we performed a two-step functional high-throughput miRNA mimic screen in Huh7.5.1 cells infected with recombinant cell culture-derived HCV. miRNA targeting was then assessed using a combination of computational and functional approaches. RESULTS: We uncovered miR-501-3p and miR-619-3p as novel modulators of HCV assembly/release. We discovered that these miRNAs regulate O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) protein expression and identified OGT and O-GlcNAcylation as regulators of HCV morphogenesis and infectivity. Furthermore, increased OGT expression in patient-derived liver tissue was associated with HCV-induced liver disease and cancer. CONCLUSION: miR-501-3p and miR-619-3p and their target OGT are previously undiscovered regulatory host factors for HCV assembly and infectivity. In addition to its effect on HCV morphogenesis, OGT may play a role in HCV-induced liver disease and hepatocarcinogenesis.}, keywords = {HCV hepatitis C hepatocyte molecular mechanisms, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Roadblocks and fast tracks: How RNA binding proteins affect the viral RNA journey in the cell}, author = {E Girardi and S Pfeffer and T F Baumert and K Majzoub}, url = {https://pubmed.ncbi.nlm.nih.gov/32847707/}, doi = {10.1016/j.semcdb.2020.08.006}, isbn = {32847707}, year = {2020}, date = {2020-01-01}, journal = {Semin Cell Dev Biol}, volume = {S1084-9521}, number = {20}, pages = {30091-4}, abstract = {As obligate intracellular parasites with limited coding capacity, RNA viruses rely on host cells to complete their multiplication cycle. Viral RNAs (vRNAs) are central to infection. They carry all the necessary information for a virus to synthesize its proteins, replicate and spread and could also play essential non-coding roles. Regardless of its origin or tropism, vRNA has by definition evolved in the presence of host RNA Binding Proteins (RBPs), which resulted in intricate and complicated interactions with these factors. While on one hand some host RBPs recognize vRNA as non-self and mobilize host antiviral defenses, vRNA must also co-opt other host RBPs to promote viral infection. Focusing on pathogenic RNA viruses, we will review important scenarios of RBP-vRNA interactions during which host RBPs recognize, modify or degrade vRNAs. We will then focus on how vRNA hijacks the largest ribonucleoprotein complex (RNP) in the cell, the ribosome, to selectively promote the synthesis of its proteins. We will finally reflect on how novel technologies are helping in deepening our understanding of vRNA-host RBPs interactions, which can be ultimately leveraged to combat everlasting viral threats.}, keywords = {Innate Immunity RNA biology RNA viruses Technology Viral RNA sensing Viral Translation Virology., PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Prevention of dsRNA-induced interferon signaling by AGO1x is linked to breast cancer cell proliferation}, author = {S Ghosh and J C Guimaraes and M Lanzafame and A Schmidt and A P Syed and B Dimitriades and A Börsch and S Ghosh and N Mittal and T Montavon and A L Correia and J Danner and G Meister and L M Terracciano and S Pfeffer and S Piscuoglio and M Zavolan}, url = {https://pubmed.ncbi.nlm.nih.gov/32812257/}, doi = {10.15252/embj.2019103922}, isbn = {32812257}, year = {2020}, date = {2020-01-01}, journal = {EMBO J}, pages = {in press}, abstract = {Translational readthrough, i.e., elongation of polypeptide chains beyond the stop codon, was initially reported for viral RNA, but later found also on eukaryotic transcripts, resulting in proteome diversification and protein-level modulation. Here, we report that AGO1x, an evolutionarily conserved translational readthrough isoform of Argonaute 1, is generated in highly proliferative breast cancer cells, where it curbs accumulation of double-stranded RNAs (dsRNAs) and consequent induction of interferon responses and apoptosis. In contrast to other mammalian Argonaute protein family members with primarily cytoplasmic functions, AGO1x exhibits nuclear localization in the vicinity of nucleoli. We identify AGO1x interaction with the polyribonucleotide nucleotidyltransferase 1 (PNPT1) and show that the depletion of this protein further augments dsRNA accumulation. Our study thus uncovers a novel function of an Argonaute protein in buffering the endogenous dsRNA-induced interferon responses, different than the canonical function of AGO proteins in the miRNA effector pathway. As AGO1x expression is tightly linked to breast cancer cell proliferation, our study thus suggests a new direction for limiting tumor growth.}, keywords = {Argonaute 1 breast cancer endogenous dsRNA interferon response translation readthrough, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The power of cooperation: Experimental and computational approaches in the functional characterization of bacterial sRNAs}, author = {J Georg and D Lalaouna and S Hou and S C Lott and I Caldelari and S Marzi and W R Hess and P Romby}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31705780}, doi = {10.1111/mmi.14420}, isbn = {31705780}, year = {2020}, date = {2020-01-01}, journal = {Mol Microbiol}, volume = {113}, number = {3}, pages = {603-612}, abstract = {Trans-acting small regulatory RNAs (sRNAs) are key players in the regulation of gene expression in bacteria. There are hundreds of different sRNAs in a typical bacterium, which in contrast to eukaryotic miRNAs are more heterogeneous in length, sequence composition, and secondary structure. The vast majority of sRNAs function post-transcriptionally by binding to other RNAs (mRNAs, sRNAs) through rather short regions of imperfect sequence complementarity. Besides, every single sRNA may interact with dozens of different target RNAs and impact gene expression either negatively or positively. These facts contributed to the view that the entirety of the regulatory targets of a given sRNA, its targetome, is challenging to identify. However, recent developments show that a more comprehensive sRNA's targetome can be achieved through the combination of experimental and computational approaches. Here, we give a short introduction into these methods followed by a description of two sRNAs, RyhB and RsaA, to illustrate the particular strengths and weaknesses of these approaches in more detail. RyhB is an sRNA involved in iron homeostasis in Enterobacteriaceae, while RsaA is a modulator of virulence in Staphylococcus aureus. Using such a combined strategy, a better appreciation of the sRNA-dependent regulatory networks is now attainable.}, keywords = {ROMBY, Staphylococcus aureus CopraRNA MAPS post-transcriptional regulation sRNAs, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Thioguanosine Conversion Enables mRNA-Lifetime Evaluation by RNA Sequencing Using Double Metabolic Labeling (TUC-seq DUAL)}, author = {C Gasser and I Delazer and E Neuner and K Pascher and K Brillet and S Klotz and L Trixl and M Himmelstoß and E Ennifar and D Rieder and A Lusser and R Micura}, url = {https://pubmed.ncbi.nlm.nih.gov/31999864/}, doi = {10.1002/anie.201916272}, isbn = {31999864}, year = {2020}, date = {2020-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {59}, number = {17}, pages = {6881-6886}, abstract = {Temporal information about cellular RNA populations is essential to understand the functional roles of RNA. We have developed the hydrazine/NH4 Cl/OsO4 -based conversion of 6-thioguanosine (6sG) into A', where A' constitutes a 6-hydrazino purine derivative. A' retains the Watson-Crick base-pair mode and is efficiently decoded as adenosine in primer extension assays and in RNA sequencing. Because 6sG is applicable to metabolic labeling of freshly synthesized RNA and because the conversion chemistry is fully compatible with the conversion of the frequently used metabolic label 4-thiouridine (4sU) into C, the combination of both modified nucleosides in dual-labeling setups enables high accuracy measurements of RNA decay. This approach, termed TUC-seq DUAL, uses the two modified nucleosides in subsequent pulses and their simultaneous detection, enabling mRNA-lifetime evaluation with unprecedented precision.}, keywords = {ENNIFAR, RNA sequencing RNA structures gene expression nucleoside modifications oligonucleotides, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Navigation through the twists and turns of RNA sequencing technologies: Application to bacterial regulatory RNAs}, author = {E Desgranges and I Caldelari and S Marzi and D Lalaouna}, url = {https://www.ncbi.nlm.nih.gov/pubmed/32068131}, doi = {10.1016/j.bbagrm.2020.194506}, isbn = {32068131}, year = {2020}, date = {2020-01-01}, journal = {Biochim Biophys Acta Gene Regul Mech}, volume = {1863}, number = {3}, pages = {194506}, abstract = {Discovered in the 1980s, small regulatory RNAs (sRNAs) are now considered key actors in virtually all aspects of bacterial physiology and virulence. Together with transcriptional and translational regulatory proteins, they integrate and often are hubs of complex regulatory networks, responsible for bacterial response/adaptation to various perceived stimuli. The recent development of powerful RNA sequencing technologies has facilitated the identification and characterization of sRNAs (length, structure and expression conditions) and their RNA targets in several bacteria. Nevertheless, it could be very difficult for non-experts to understand the advantages and drawbacks related to each offered option and, consequently, to make an informed choice. Therefore, the main goal of this review is to provide a guide to navigate through the twists and turns of high-throughput RNA sequencing technologies, with a specific focus on those applied to the study of sRNAs. This article is part of a Special Issue entitled: RNA and gene control in bacteria edited by Dr. M. Guillier and F. Repoila.}, keywords = {Bacteria Post-transcriptional regulation RNA sequencing Regulatory network Small regulatory RNA Targetome, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Monitoring the Production of High Diffraction-Quality Crystals of Two Enzymes in Real Time Using In Situ Dynamic Light Scattering}, author = {R de Wijn and K Rollet and S Engilberge and A G McEwen and O Hennig and H Betat and M Mörl and F Riobé and O Maury and E Girard and P Bénas and B Lorber and C Sauter}, url = {https://www.mdpi.com/2073-4352/10/2/65}, doi = {10.3390/cryst10020065}, year = {2020}, date = {2020-01-01}, journal = {Crystals}, volume = {10}, number = {2}, pages = {65}, abstract = {The reproducible preparation of well-diffracting crystals is a prerequisite for every structural study based on crystallography. An instrument called XtalController has recently been designed that allows the monitoring of crystallization assays using dynamic light scattering and microscopy, and integrates piezo pumps to alter the composition of the mother liquor during the experiment. We have applied this technology to study the crystallization of two enzymes, the CCA-adding enzyme of the psychrophilic bacterium Planococcus halocryophilus, and the lysozyme from hen egg white in the presence of a synthetic chemical nucleant. We were able to (i) detect early nucleation events and (ii) drive the crystallization system (through cycles of dissolution/crystallization) toward growth conditions yielding crystals with excellent diffraction properties. This technology opens a way to the rational production of samples for crystallography, ranging from nanocrystals for electron diffraction, microcrystals for serial or conventional X-ray diffraction, to larger crystals for neutron diffraction.}, keywords = {enzyme crystallization dynamic light scattering nucleation nucleant Tb-Xo4 crystallophore microcrystals nanocrystals X-ray diffraction XtalController, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selenoprotein N is an endoplasmic reticulum calcium sensor that links luminal calcium levels to a redox activity}, author = {A Chernorudskiy and E Varone and S F Colombo and S Fumagalli and A Cagnotto and A Cattaneo and M Briens and M Baltzinger and L Kuhn and A Bachi and A Berardi and M Salmona and G Musco and N Borgese and A Lescure and E Zito}, url = {https://pubmed.ncbi.nlm.nih.gov/32817544/}, doi = {10.1073/pnas.2003847117}, isbn = {32817544}, year = {2020}, date = {2020-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {117}, number = {35}, pages = {21288-21298}, abstract = {The endoplasmic reticulum (ER) is the reservoir for calcium in cells. Luminal calcium levels are determined by calcium-sensing proteins that trigger calcium dynamics in response to calcium fluctuations. Here we report that Selenoprotein N (SEPN1) is a type II transmembrane protein that senses ER calcium fluctuations by binding this ion through a luminal EF-hand domain. In vitro and in vivo experiments show that via this domain, SEPN1 responds to diminished luminal calcium levels, dynamically changing its oligomeric state and enhancing its redox-dependent interaction with cellular partners, including the ER calcium pump sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA). Importantly, single amino acid substitutions in the EF-hand domain of SEPN1 identified as clinical variations are shown to impair its calcium-binding and calcium-dependent structural changes, suggesting a key role of the EF-hand domain in SEPN1 function. In conclusion, SEPN1 is a ER calcium sensor that responds to luminal calcium depletion, changing its oligomeric state and acting as a reductase to refill ER calcium stores.}, keywords = {LESCURE, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Different views of the dynamic landscape covered by the 5'-hairpin of the 7SK small nuclear RNA}, author = {K Brillet and D Martinez-Zapien and G Bec and E Ennifar and A C Dock-Bregeon and I Lebars}, url = {https://pubmed.ncbi.nlm.nih.gov/32430362/}, doi = {10.1261/rna.074955.120}, isbn = {32430362}, year = {2020}, date = {2020-01-01}, journal = {RNA}, volume = {26}, number = {9}, pages = {1184-1197}, abstract = {The 7SK small nuclear RNA (7SKsnRNA) plays a key role in the regulation of RNA polymerase II by sequestrating and inhibiting the positive transcription elongation factor b (P-TEFb) in the 7SK ribonucleoprotein complex (7SKsnRNP), a process mediated by interaction with the protein HEXIM. P-TEFb is also an essential cellular factor recruited by the viral protein Tat to ensure the replication of the viral RNA in the infection cycle of the human immunodeficiency virus (HIV-1). Tat promotes the release of P-TEFb from the 7SKsnRNP and subsequent activation of transcription, by displacing HEXIM from the 5'-hairpin of the 7SKsnRNA. This hairpin (HP1), comprising the signature sequence of the 7SKsnRNA, has been the subject of three independent structural studies aiming at identifying the structural features that could drive the recognition by the two proteins, both depending on Arginine Rich Motifs (ARM). Interestingly, four distinct structures were determined. In an attempt to provide a comprehensive view of the structure-function relationship of this versatile RNA, we present here a structural analysis of the models, highlighting how HP1 is able to adopt distinct conformations with significant impact on the compactness of the molecule. Since these models are solved under different conditions by NMR and crystallography, the impact of the buffer composition on the conformational variation was investigated by complementary biophysical approaches. Finally, using Isothermal Titration Calorimetry, we determined the thermodynamic signatures of the Tat-ARM and HEXIM-ARM peptide interactions with the RNA, showing that they are associated with distinct binding}, keywords = {7SK HEXIM RNA Tat structure, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Zinc Fingers in HIV-1 Gag Precursor Are Not Equivalent for gRNA Recruitment at the Plasma Membrane}, author = {E Boutant and J Bonzi and H Anton and M Bin Nasim and R Cathagne and E Real and D Dujardin and P Carl and P Didier and J C Paillart and R Marquet and Y Mely and H de Rocquigny and S Bernacchi}, url = {https://pubmed.ncbi.nlm.nih.gov/32574557/}, doi = {10.1016/j.bpj.2020.05.035}, isbn = {32574557}, year = {2020}, date = {2020-01-01}, journal = {Biophys J}, volume = {119}, number = {2}, pages = {419-433}, abstract = {The human immunodeficiency virus type 1 Gag precursor specifically selects the unspliced viral genomic RNA (gRNA) from the bulk of cellular and spliced viral RNAs via its nucleocapsid (NC) domain and drives gRNA encapsidation at the plasma membrane (PM). To further identify the determinants governing the intracellular trafficking of Gag-gRNA complexes and their accumulation at the PM, we compared, in living and fixed cells, the interactions between gRNA and wild-type Gag or Gag mutants carrying deletions in NC zinc fingers (ZFs) or a nonmyristoylated version of Gag. Our data showed that the deletion of both ZFs simultaneously or the complete NC domain completely abolished intracytoplasmic Gag-gRNA interactions. Deletion of either ZF delayed the delivery of gRNA to the PM but did not prevent Gag-gRNA interactions in the cytoplasm, indicating that the two ZFs display redundant roles in this respect. However, ZF2 played a more prominent role than ZF1 in the accumulation of the ribonucleoprotein complexes at the PM. Finally, the myristate group, which is mandatory for anchoring the complexes at the PM, was found to be dispensable for the association of Gag with the gRNA in the cytosol.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Analysis of the HIV-1 Genomic RNA Dimerization Initiation Site Binding to Aminoglycoside Antibiotics Using Isothermal Titration Calorimetry}, author = {S Bernacchi and E Ennifar}, editor = {V Arluison and F Wien}, url = {https://pubmed.ncbi.nlm.nih.gov/32006318}, doi = {10.1007/978-1-0716-0278-2_16}, isbn = {32006318}, year = {2020}, date = {2020-01-01}, booktitle = {RNA Spectroscopy: Methods and Protocols}, volume = {2113}, pages = {237-250}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Isothermal titration calorimetry (ITC) provides a sensitive, powerful, and accurate tool to suitably analyze the thermodynamic of RNA binding events. This approach does not require any modification or labeling of the system under analysis and is performed in solution. ITC is a very convenient technique that provides an accurate determination of binding parameters, as well as a complete thermodynamic profile of the molecular interactions. Here we show how this approach can be used to characterize the interactions between the dimerization initiation site (DIS) RNA localized within the HIV-1 viral genome and aminoglycoside antibiotics. Our ITC study showed that the 4,5-disubstituted 2-desoxystreptamine (2-DOS) aminoglycosides can bind the DIS with a nanomolar affinity and a high specificity.}, keywords = {Aminoglycosides, dimerization, drug interaction, ENNIFAR, HIV-1, initiation site, ITC, MARQUET, PAILLART, RNA, Thermodynamics RNA, Unité ARN, Viral}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Dynamic Light Scattering Analysis on RNA Associated to Proteins}, author = {S Bernacchi}, editor = {V Arluison and F Wien}, url = {https://pubmed.ncbi.nlm.nih.gov/32006306}, doi = {10.1007/978-1-0716-0278-2_4}, isbn = {32006306}, year = {2020}, date = {2020-01-01}, booktitle = {RNA Spectroscopy: Methods and Protocols}, volume = {2113}, pages = {31-39}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Dynamic light scattering represents an accurate, robust, and reliable technique to analyze molecule size in solution and monitor their interactions in real time. Here, we describe how to analyze by DLS an RNA-protein interaction. In our frame, we studied complexes formed between RNA fragments derived from the genome of HIV-1 in association with the viral precursor Pr55Gag. These interactions are crucial for the specific selection of the viral genomic RNA (gRNA) from the bulk of the viral spliced and cellular RNAs. This chapter displays how DLS allows to characterize the interactions that regulate the early steps of viral assembly.}, keywords = {Dynamic light scattering, Hydrodynamic radius Diffusion coefficient, MARQUET, PAILLART, Pr55Gag, precursor, Protein-RNA interactions, Unité ARN, Viral assembly, Viral genomic RNA Viral spliced RNA}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Mapping of Posttranscriptional tRNA Modifications by Two-Dimensional Gel Electrophoresis Mass Spectrometry}, author = {L Antoine and P Wolff}, editor = {V Arluison and F Wien}, url = {https://pubmed.ncbi.nlm.nih.gov/32006310}, doi = {10.1007/978-1-0716-0278-2_8}, isbn = {32006310}, year = {2020}, date = {2020-01-01}, booktitle = {RNA Spectroscopy: Methods and Protocols}, volume = {2113}, pages = {101-110}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {RNA modification mapping by mass spectrometry (MS) is based on the use of specific ribonucleases (RNases) that generate short oligonucleotide digestion products which are further separated by nano-liquid chromatography and analyzed by MS and MS/MS. Recent developments in MS instrumentation allow the possibility to deeply explore posttranscriptional modifications. Notably, development of nano-liquid chromatography and nano-electrospray drastically increases the detection sensitivity and allows the identification and sequencing of RNA digested fragments separated and extracted from two-dimensional polyacrylamide gels, as long as the mapping and characterization of ribonucleotide modifications.}, keywords = {2D Gel isolation Nano-LC-MS/MS Posttranscriptional tRNA modifications, ARN-MS, ENNIFAR, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{malanagahalli_few_2020, title = {Few Layer Graphene Does Not Affect Cellular Homeostasis of Mouse Macrophages}, author = {Sowmya Malanagahalli and Diane Murera and Cristina Martín and Hazel Lin and Nadége Wadier and Hélène Dumortier and Ester Vázquez and Alberto Bianco}, doi = {10.3390/nano10020228}, issn = {2079-4991}, year = {2020}, date = {2020-01-01}, journal = {Nanomaterials (Basel, Switzerland)}, volume = {10}, number = {2}, abstract = {: Graphene-related materials (GRMs) are widely used in various applications due to their unique properties. A growing number of reports describe the impact of different carbon nanomaterials, including graphene oxide (GO), reduced GO (rGO), and carbon nanotubes (CNT), on immune cells, but there is still a very limited number of studies on graphene. In this work, we investigated the biological responses of few layer graphene (FLG) on mouse macrophages (bone marrow derived macrophages, BMDMs), which are part of the first line of defense in innate immunity. In particular, our paper describes our findings of short-term FLG treatment in BMDMs with a focus on observing material internalization and changes in general cell morphology. Subsequent investigation of cytotoxicity parameters showed that increasing doses of FLG did not hamper the viability of cells and did not trigger inflammatory responses. Basal level induced autophagic activity sufficed to maintain the cellular homeostasis of FLG treated cells. Our results shed light on the impact of FLG on primary macrophages and show that FLG does not elicit immunological responses leading to cell death.}, keywords = {Autophagy, bone marrow derived macrophages, carbon nanomaterials, Dumortier, graphene, I2CT, primary immune cells, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{summer_ybey_2020, title = {YBEY is an essential biogenesis factor for mitochondrial ribosomes.}, author = {Sabrina Summer and Anna Smirnova and Alessandro Gabriele and Ursula Toth and Akinyemi Mandela Fasemore and Konrad U Förstner and Lauriane Kuhn and Johana Chicher and Philippe Hammann and Goran Mitulović and Nina Entelis and Ivan Tarassov and Walter Rossmanith and Alexandre Smirnov}, doi = {10.1093/nar/gkaa148}, issn = {1362-4962 0305-1048 0305-1048}, year = {2020}, date = {2020-01-01}, journal = {Nucleic acids research}, volume = {48}, number = {17}, pages = {9762--9786}, abstract = {Ribosome biogenesis requires numerous trans-acting factors, some of which are deeply conserved. In Bacteria, the endoribonuclease YbeY is believed to be involved in 16S rRNA 3'-end processing and its loss was associated with ribosomal abnormalities. In Eukarya, YBEY appears to generally localize to mitochondria (or chloroplasts). Here we show that the deletion of human YBEY results in a severe respiratory deficiency and morphologically abnormal mitochondria as an apparent consequence of impaired mitochondrial translation. Reduced stability of 12S rRNA and the deficiency of several proteins of the small ribosomal subunit in YBEY knockout cells pointed towards a defect in mitochondrial ribosome biogenesis. The specific interaction of mitoribosomal protein uS11m with YBEY suggests that the latter helps to properly incorporate uS11m into the nascent small subunit in its late assembly stage. This scenario shows similarities with final stages of cytosolic ribosome biogenesis, and may represent a late checkpoint before the mitoribosome engages in translation.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{fujii_tailoring_2020, title = {Tailoring the interaction between graphene oxide and antibacterial pyridinium salts by terminal functional groups}, author = {R Fujii and K Okubo and S Takashiba and A Bianco and Y Nishina}, url = {https://linkinghub.elsevier.com/retrieve/pii/S0008622319311972}, doi = {10.1016/j.carbon.2019.11.094}, issn = {00086223}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Carbon}, volume = {160}, pages = {204--210}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{guo_flexible_2020, title = {A Flexible Method for Covalent Double Functionalization of Graphene Oxide}, author = {Shi Guo and Yuta Nishina and Alberto Bianco and Cécilia Ménard‐Moyon}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/anie.201913461}, doi = {10.1002/anie.201913461}, issn = {1433-7851, 1521-3773}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Angewandte Chemie International Edition}, volume = {59}, number = {4}, pages = {1542--1547}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{wang_neutron_2020, title = {Neutron Activated 153Sm Sealed in Carbon Nanocapsules for textitin Vivo Imaging and Tumor Radiotherapy}, author = {Julie T -W Wang and Rebecca Klippstein and Markus Martincic and Elzbieta Pach and Robert Feldman and Martin Šefl and Yves Michel and Daniel Asker and Jane K Sosabowski and Martin Kalbac and Tatiana Da Ros and Cécilia Ménard-Moyon and Alberto Bianco and Ioanna Kyriakou and Dimitris Emfietzoglou and Jean-Claude Saccavini and Belén Ballesteros and Khuloud T Al-Jamal and Gerard Tobias}, url = {https://pubs.acs.org/doi/10.1021/acsnano.9b04898}, doi = {10.1021/acsnano.9b04898}, issn = {1936-0851, 1936-086X}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {ACS Nano}, volume = {14}, number = {1}, pages = {129--141}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bajczyk_serrate_2020, title = {SERRATE interacts with the nuclear exosome targeting (NEXT) complex to degrade primary miRNA precursors in Arabidopsis.}, author = {Mateusz Bajczyk and Heike Lange and Dawid Bielewicz and Lukasz Szewc and Susheel S Bhat and Jakub Dolata and Lauriane Kuhn and Zofia Szweykowska-Kulinska and Dominique Gagliardi and Artur Jarmolowski}, doi = {10.1093/nar/gkaa373}, issn = {1362-4962 0305-1048 0305-1048}, year = {2020}, date = {2020-01-01}, journal = {Nucleic acids research}, volume = {48}, number = {12}, pages = {6839--6854}, abstract = {SERRATE/ARS2 is a conserved RNA effector protein involved in transcription, processing and export of different types of RNAs. In Arabidopsis, the best-studied function of SERRATE (SE) is to promote miRNA processing. Here, we report that SE interacts with the nuclear exosome targeting (NEXT) complex, comprising the RNA helicase HEN2, the RNA binding protein RBM7 and one of the two zinc-knuckle proteins ZCCHC8A/ZCCHC8B. The identification of common targets of SE and HEN2 by RNA-seq supports the idea that SE cooperates with NEXT for RNA surveillance by the nuclear exosome. Among the RNA targets accumulating in absence of SE or NEXT are miRNA precursors. Loss of NEXT components results in the accumulation of pri-miRNAs without affecting levels of miRNAs, indicating that NEXT is, unlike SE, not required for miRNA processing. As compared to se-2, se-2 hen2-2 double mutants showed increased accumulation of pri-miRNAs, but partially restored levels of mature miRNAs and attenuated developmental defects. We propose that the slow degradation of pri-miRNAs caused by loss of HEN2 compensates for the poor miRNA processing efficiency in se-2 mutants, and that SE regulates miRNA biogenesis through its double contribution in promoting miRNA processing but also pri-miRNA degradation through the recruitment of the NEXT complex.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{backes_production_2020, title = {Production and processing of graphene and related materials}, author = {Claudia Backes and Amr M Abdelkader and Concepción Alonso and Amandine Andrieux-Ledier and Raul Arenal and Jon Azpeitia and Nilanthy Balakrishnan and Luca Banszerus and Julien Barjon and Ruben Bartali and Sebastiano Bellani and Claire Berger and Reinhard Berger and Bernal M M Ortega and Carlo Bernard and Peter H Beton and André Beyer and Alberto Bianco and Peter Bøggild and Francesco Bonaccorso and Gabriela Borin Barin and Cristina Botas and Rebeca A Bueno and Daniel Carriazo and Andres Castellanos-Gomez and Meganne Christian and Artur Ciesielski and Tymoteusz Ciuk and Matthew T Cole and Jonathan Coleman and Camilla Coletti and Luigi Crema and Huanyao Cun and Daniela Dasler and Domenico De Fazio and Noel Díez and Simon Drieschner and Georg S Duesberg and Roman Fasel and Xinliang Feng and Alberto Fina and Stiven Forti and Costas Galiotis and Giovanni Garberoglio and Jorge M García and Jose Antonio Garrido and Marco Gibertini and Armin Gölzhäuser and Julio Gómez and Thomas Greber and Frank Hauke and Adrian Hemmi and Irene Hernandez-Rodriguez and Andreas Hirsch and Stephen A Hodge and Yves Huttel and Peter U Jepsen and Ignacio Jimenez and Ute Kaiser and Tommi Kaplas and HoKwon Kim and Andras Kis and Konstantinos Papagelis and Kostas Kostarelos and Aleksandra Krajewska and Kangho Lee and Changfeng Li and Harri Lipsanen and Andrea Liscio and Martin R Lohe and Annick Loiseau and Lucia Lombardi and Maria Francisca López and Oliver Martin and Cristina Martín and Lidia Martínez and Jose Angel Martin-Gago and José Ignacio Martínez and Nicola Marzari and Álvaro Mayoral and John McManus and Manuela Melucci and Javier Méndez and Cesar Merino and Pablo Merino and Andreas P Meyer and Elisa Miniussi and Vaidotas Miseikis and Neeraj Mishra and Vittorio Morandi and Carmen Munuera and Roberto Muñoz and Hugo Nolan and Luca Ortolani and Anna K Ott and Irene Palacio and Vincenzo Palermo and John Parthenios and Iwona Pasternak and Amalia Patane and Maurizio Prato and Henri Prevost and Vladimir Prudkovskiy and Nicola Pugno and Teófilo Rojo and Antonio Rossi and Pascal Ruffieux and Paolo Samorì and Léonard Schué and Eki Setijadi and Thomas Seyller and Giorgio Speranza and Christoph Stampfer and Ingrid Stenger and Wlodek Strupinski and Yuri Svirko and Simone Taioli and Kenneth B K Teo and Matteo Testi and Flavia Tomarchio and Mauro Tortello and Emanuele Treossi and Andrey Turchanin and Ester Vazquez and Elvira Villaro and Patrick R Whelan and Zhenyuan Xia and Rositza Yakimova and Sheng Yang and Reza G Yazdi and Chanyoung Yim and Duhee Yoon and Xianghui Zhang and Xiaodong Zhuang and Luigi Colombo and Andrea C Ferrari and Mar Garcia-Hernandez}, url = {https://iopscience.iop.org/article/10.1088/2053-1583/ab1e0a}, doi = {10.1088/2053-1583/ab1e0a}, issn = {2053-1583}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {2D Materials}, volume = {7}, number = {2}, pages = {022001}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{dinesh_hybrid_2020, title = {Hybrid Interfaces Made of Nanotubes and Backbone-Altered Dipeptides Tune Neuronal Network Architecture}, author = {Bhimareddy Dinesh and Manuela Medelin and Denis Scaini and Federica Lareno Faccini and Federica Quici and Laura Ballerini and Alberto Bianco}, url = {https://pubs.acs.org/doi/10.1021/acschemneuro.9b00522}, doi = {10.1021/acschemneuro.9b00522}, issn = {1948-7193, 1948-7193}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {ACS Chemical Neuroscience}, volume = {11}, number = {2}, pages = {162--172}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{aloisi_synthesis_2020, title = {Synthesis and Characterization of Adamantane-Containing Heteropeptides with a Chirality Switch: Synthesis and Characterization of Adamantane-Containing Heteropeptides with a Chirality Switch}, author = {Adriano Aloisi and Niels Johan Christensen and Kasper K Sørensen and Chloé Guilbaud-Chéreau and Knud J Jensen and Alberto Bianco}, url = {http://doi.wiley.com/10.1002/ejoc.201901666}, doi = {10.1002/ejoc.201901666}, issn = {1434193X}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {European Journal of Organic Chemistry}, volume = {2020}, number = {7}, pages = {815--820}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{wang_neutron-irradiated_2020, title = {Neutron-irradiated antibody-functionalised carbon nanocapsules for targeted cancer radiotherapy}, author = {Julie Tzu-Wen Wang and Cinzia Spinato and Rebecca Klippstein and Pedro Miguel Costa and Markus Martincic and Elzbieta Pach and Aritz Perez Ruiz de Garibay and Cécilia Ménard-Moyon and Robert Feldman and Yves Michel and Martin Šefl and Ioanna Kyriakou and Dimitris Emfietzoglou and Jean-Claude Saccavini and Belén Ballesteros and Gerard Tobias and Alberto Bianco and Khuloud T Al-Jamal}, url = {https://linkinghub.elsevier.com/retrieve/pii/S0008622320302062}, doi = {10.1016/j.carbon.2020.02.060}, issn = {00086223}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Carbon}, volume = {162}, pages = {410--422}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bianco_carbon_2020, title = {Carbon science perspective in 2020: Current research and future challenges}, author = {Alberto Bianco and Yuan Chen and Elzbieta Frackowiak and Michael Holzinger and Nikhil Koratkar and Vincent Meunier and Sergey Mikhailovsky and Michael Strano and Juan M D Tascon and Mauricio Terrones}, url = {https://linkinghub.elsevier.com/retrieve/pii/S0008622320300622}, doi = {10.1016/j.carbon.2020.01.055}, issn = {00086223}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Carbon}, volume = {161}, pages = {373--391}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{lucherelli_rational_2020, title = {Rational Chemical Multifunctionalization of Graphene Interface Enhances Targeted Cancer Therapy}, author = {Matteo Andrea Lucherelli and Yue Yu and Giacomo Reina and Gonzalo Abellán and Eijiro Miyako and Alberto Bianco}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/anie.201916112}, doi = {10.1002/anie.201916112}, issn = {1433-7851, 1521-3773}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Angewandte Chemie International Edition}, volume = {59}, number = {33}, pages = {14034--14039}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{soltani_carbon_2020, title = {Carbon Nanomaterials Applied for the Treatment of Inflammatory Diseases: Preclinical Evidence}, author = {Rym Soltani and Shi Guo and Alberto Bianco and Cécilia Ménard‐Moyon}, url = {https://onlinelibrary.wiley.com/doi/10.1002/adtp.202000051}, doi = {10.1002/adtp.202000051}, issn = {2366-3987, 2366-3987}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Advanced Therapeutics}, volume = {3}, number = {9}, pages = {2000051}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{gazzi_graphene_2020, title = {Graphene, other carbon nanomaterials and the immune system: toward nanoimmunity-by-design}, author = {Arianna Gazzi and Laura Fusco and Marco Orecchioni and Silvia Ferrari and Giulia Franzoni and Stephen J Yan and Matthias Rieckher and Guotao Peng and Matteo Andrea Lucherelli and Isabella Anna Vacchi and Ngoc Do Quyen Chau and Alejandro Criado and Akcan Istif and Donato Mancino and Antonio Dominguez and Hagen Eckert and Ester Vázquez and Tatiana Da Ros and Paola Nicolussi and Vincenzo Palermo and Björn Schumacher and Gianaurelio Cuniberti and Yiyong Mai and Cecilia Clementi and Matteo Pasquali and Xinliang Feng and Kostas Kostarelos and Acelya Yilmazer and Davide Bedognetti and Bengt Fadeel and Maurizio Prato and Alberto Bianco and Lucia Gemma Delogu}, url = {https://iopscience.iop.org/article/10.1088/2515-7639/ab9317}, doi = {10.1088/2515-7639/ab9317}, issn = {2515-7639}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Journal of Physics: Materials}, volume = {3}, number = {3}, pages = {034009}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{raya_kinetics_2020, title = {Kinetics of 1H–13C multiple-contact multiple-contact cross-polarization as a powerful tool to determine the structure and dynamics of complex materials: application to graphene oxide}, author = {Jésus Raya and Alberto Bianco and Jérôme Hirschinger}, url = {http://xlink.rsc.org/?DOI=D0CP00454E}, doi = {10.1039/D0CP00454E}, issn = {1463-9076, 1463-9084}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Physical Chemistry Chemical Physics}, volume = {22}, number = {21}, pages = {12209--12227}, abstract = {Structural and dynamical details are probed by kinetics of 1 H– 13 C single- and multiple-contact cross-polarization in graphene oxide. , Hartmann–Hahn cross-polarization (HHCP) is the most widely used solid-state NMR technique to enhance the magnetization of dilute spins from abundant spins. Furthermore, as the kinetics of CP depends on dipolar interactions, it contains valuable information on molecular structure and dynamics. In this work, analytical solutions are derived for the kinetics of HHCP and multiple-contact CP (MC-CP) using both classical and non-classical spin-coupling models including the effects of molecular dynamics and several 1 H, 13 C relaxation and 1 H– 13 C CP experiments are performed in graphene oxide (GO). HHCP is found to be inefficient in our GO sample due to very fast 1 H T 1ρ relaxation. By contrast, the MC-CP technique which alleviates most of the magnetization loss by 1 H T 1ρ relaxation leads to a much larger polarization transfer efficiency reducing the measuring time by an order of magnitude. A detailed analysis of the HHCP and MC-CP kinetics indicates the existence of at least two different kinds of hydroxyl (C–OH) functional groups in GO, the major fraction (∼90%) of these groups being in the unusual “slow CP regime” in which the rate of 1 H T 1ρ relaxation is fast compared to the rate of cross-polarization. This 13 C signal component is attributed to mobile C–OH groups interacting preferentially with fast-relaxing water molecules while the remaining carbons (∼10%) in the usual “fast CP regime” are assigned to C–OH groups involved in hydrogen bonding with neighboring hydroxyl and/or epoxy groups.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ji_controlled_2020, title = {Controlled functionalization of carbon nanodots for targeted intracellular production of reactive oxygen species}, author = {Ding-Kun Ji and Giacomo Reina and Shi Guo and Matilde Eredia and Paolo Samorì and Cécilia Ménard-Moyon and Alberto Bianco}, url = {http://xlink.rsc.org/?DOI=D0NH00300J}, doi = {10.1039/D0NH00300J}, issn = {2055-6756, 2055-6764}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Nanoscale Horizons}, volume = {5}, number = {8}, pages = {1240--1249}, abstract = {Multifunctional carbon nanodots with deep-red emission were prepared. These carbon nanodots are suitable for targeted intracellular production of reactive oxygen species by laser irradiation leading to efficient cancer cell death. , Controlled intracellular release of exogenous reactive oxygen species (ROS) is an innovative and efficient strategy for cancer treatment. Well-designed materials, which can produce ROS in targeted cells, minimizing side effects, still need to be explored as new generation nanomedicines. Here, red-emissive carbon nanodots (CNDs) with intrinsic theranostic properties are devised, and further modified with folic acid (FA) ligand through a controlled covalent functionalization for targeted cell imaging and intracellular production of ROS. We demonstrated that covalent functionalization is an effective strategy to prevent the aggregation of the dots, leading to superior colloidal stability, enhanced luminescence and ROS generation. Indeed, the functional nanodots possess a deep-red emission and good dispersibility under physiological conditions. Importantly, they show excellent targeting properties and generation of high levels of ROS under 660 nm laser irradiation, leading to efficient cell death. These unique properties enable FA-modified carbon nanodots to act as a multifunctional nanoplatform for simultaneous targeted imaging and efficient photodynamic therapy to induce cancer cell death.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{guo_is_2020, title = {Is carboxylation an efficient method for graphene oxide functionalization?}, author = {Shi Guo and Jésus Raya and Dingkun Ji and Yuta Nishina and Cécilia Ménard-Moyon and Alberto Bianco}, url = {http://xlink.rsc.org/?DOI=D0NA00561D}, doi = {10.1039/D0NA00561D}, issn = {2516-0230}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Nanoscale Advances}, volume = {2}, number = {9}, pages = {4085--4092}, abstract = {We investigated the carboxylation of graphene oxide using chloroacetic acid and different amounts of NaOH. Increase of carboxyl groups was observed only at high amounts of NaOH, but partial reduction attenuates the yield of further functionalization. , Graphene oxide (GO) is one of the most popular materials applied in different research areas thanks to its unique properties. The application of GO requires well-designed protocols to introduce different functionalities on its surface, exploiting the oxygenated groups already present. Due to the complex and unstable chemical environment on the GO surface, it is recommended to perform the functionalization under mild conditions. The carboxylation of GO is a widely used method to introduce additional carboxylic acids, which could be further modified through amidation or esterification reactions. The strategy already reported in the literature requires harsh conditions (excess amount of sodium hydroxide). GO is readily reduced under basic conditions, but the reduction of GO during the carboxylation is barely studied. In this work, we performed the carboxylation using chloroacetic acid with different amounts of sodium hydroxide and characterized the functionalized GO with various techniques. The carboxylated GO was exploited to develop a double functionalization approach combining an epoxide ring opening reaction and an amidation. The results showed that strong basic conditions were necessary to derivatize GO. Nevertheless, these conditions resulted in a partial reduction of GO and some functionalities on GO were removed during the reaction, thus reducing the total efficiency of the functionalization in comparison to an epoxide ring opening reaction, indicating that carboxylation is not an efficient approach for the functionalization of GO.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ma_degradation-by-design_2020, title = {Degradation-by-design: how chemical functionalization enhances the biodegradability and safety of 2D materials}, author = {Baojin Ma and Cristina Martín and Rajendra Kurapati and Alberto Bianco}, url = {http://xlink.rsc.org/?DOI=C9CS00822E}, doi = {10.1039/C9CS00822E}, issn = {0306-0012, 1460-4744}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Chemical Society Reviews}, volume = {49}, number = {17}, pages = {6224--6247}, abstract = {A large number of graphene and other 2D materials are currently explored for the development of new technologies. The assessment of their biodegradability is one of the fundamental aspects for their safe application. , A large number of graphene and other 2D materials are currently used for the development of new technologies, increasingly entering different industrial sectors. Interrogating the impact of such 2D materials on health and environment is crucial for both modulating their potential toxicity in living organisms and eliminating them from the environment. In this context, understanding if 2D materials are bio-persistent is mandatory. In this review we describe the importance of biodegradability and decomposition of 2D materials. We initially cover the biodegradation of graphene family materials, followed by other emerging classes of 2D materials including transition metal dichalcogenides and oxides, Xenes, Mxenes and other non-metallic 2D materials. We explain the role of defects and functional groups, introduced onto the surface of the materials during their preparation, and the consequences of chemical functionalization on biodegradability. In strong relation to the chemistry on 2D materials, we describe the concept of “degradation-by-design” that we contributed to develop, and which concerns the covalent modification with appropriate molecules to enhance the biodegradability of 2D materials. Finally, we cover the importance of designing new biodegradable 2D conjugates and devices for biomedical applications as drug delivery carriers, in bioelectronics, and tissue engineering. We would like to highlight that the biodegradation of 2D materials mainly depends on the type of material, the chemical functionalization, the aqueous dispersibility and the redox potentials of the different oxidative environments. Biodegradation is one of the necessary conditions for the safe application of 2D materials. Therefore, we hope that this review will help to better understand their biodegradation processes, and will stimulate the chemists to explore new chemical strategies to design safer products, composites and devices containing 2D materials.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{luan_degradation_2020, title = {Degradation of Structurally Defined Graphene Nanoribbons by Myeloperoxidase and the Photo‐Fenton Reaction}, author = {Xiangfeng Luan and Cristina Martín and Pengfei Zhang and Qian Li and Isabella Anna Vacchi and Lucia Gemma Delogu and Yiyong Mai and Alberto Bianco}, url = {https://onlinelibrary.wiley.com/doi/10.1002/anie.202008925}, doi = {10.1002/anie.202008925}, issn = {1433-7851, 1521-3773}, year = {2020}, date = {2020-01-01}, urldate = {2020-11-20}, journal = {Angewandte Chemie International Edition}, volume = {59}, number = {42}, pages = {18515--18521}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{Verriez2020, title = {Les APOBEC : histoire d’une famille de protéines antivirales et mutagènes.}, author = {C Verriez, R Marquet, J-C Paillart and B Stupfler}, year = {2020}, date = {2020-01-01}, journal = {Virologie}, volume = {24}, number = {6}, pages = {381-418}, abstract = {La réponse immunitaire innée est une réponse non spécifique qui constitue la première ligne de défense en cas d’infection, notamment en permettant l’élimination des pathogènes par phagocytose ou apoptose. Au sein des cellules immunitaires, cette réponse innée se caractérise entre autres par la synthèse de protéines nommées facteurs de restriction dont le rôle est d’inhiber la réplication virale. Parmi ces facteurs, les protéines de la famille APOBEC3 (Apolipoprotein B mRNA-editing Enzyme Catalytic polypeptide-like 3 ou A3) constituent des facteurs antiviraux majeurs qui ciblent de nombreux types de virus. L’une des cibles des A3 est le virus de l’immunodéficience humaine de type 1 (VIH-1) : l’activité désaminase de certaines A3 convertit une fraction des cytidines du génome viral en uridines et perturbe son expression. Néanmoins, le VIH-1 contrecarre les A3 en exprimant la protéine Vif qui les inhibe en détournant divers mécanismes cellulaires. Par ailleurs, les APOBEC3 participent au maintien de l’intégrité génétique par l’inhibition des rétroéléments mais contribuent également à la cancérogenèse, à l’image d’A3A et A3B, deux facteurs majeurs dans ce processus. L’éventail de leurs activités, combiné aux récentes études montrant leur implication dans la régulation de virus émergents (Zika, SARS-CoV-2), permettent d’envisager les A3 ainsi que leurs partenaires viraux comme axes thérapeutiques.}, keywords = {APOBEC3, cancer, Facteurs de restriction, MARQUET, PAILLART, Unité ARN, vif, VIH-1}, pubstate = {published}, tppubtype = {article} } @article{meteignier_arabidopsis_2020, title = {The Arabidopsis mTERF-repeat MDA1 protein plays a dual function in transcription and stabilization of specific chloroplast transcripts within the psbE and ndhH operons.}, author = {Louis-Valentin Méteignier and Rabea Ghandour and Karin Meierhoff and Aude Zimmerman and Johana Chicher and Nicolas Baumberger and Abdelmalek Alioua and Jörg Meurer and Reimo Zoschke and Kamel Hammani}, doi = {10.1111/nph.16625}, issn = {1469-8137 0028-646X 0028-646X}, year = {2020}, date = {2020-01-01}, journal = {The New phytologist}, volume = {227}, number = {5}, pages = {1376--1391}, abstract = {The mTERF gene family encodes for nucleic acid binding proteins that are predicted to regulate organellar gene expression in eukaryotes. Despite the implication of this gene family in plant development and response to abiotic stresses, a precise molecular function was assigned to only a handful number of its c. 30 members in plants. Using a reverse genetics approach in Arabidopsis thaliana and combining molecular and biochemical techniques, we revealed new functions for the chloroplast mTERF protein, MDA1. We demonstrated that MDA1 associates in vivo with components of the plastid-encoded RNA polymerase and transcriptional active chromosome complexes. MDA1 protein binds in vivo and in vitro with specificity to 27-bp DNA sequences near the 5'-end of psbE and ndhA chloroplast genes to stimulate their transcription, and additionally promotes the stabilization of the 5'-ends of processed psbE and ndhA messenger (m)RNAs. Finally, we provided evidence that MDA1 function in gene transcription likely coordinates RNA folding and the action of chloroplast RNA-binding proteins on mRNA stabilization. Our results provide examples for the unexpected implication of DNA binding proteins and gene transcription in the regulation of mRNA stability in chloroplasts, blurring the boundaries between DNA and RNA metabolism in this organelle.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{chico_cul3bpm_2020, title = {CUL3(BPM) E3 ubiquitin ligases regulate MYC2, MYC3, and MYC4 stability and JA responses.}, author = {Jose Manuel Chico and Esther Lechner and Gemma Fernandez-Barbero and Esther Canibano and Gloria García-Casado and Jose Manuel Franco-Zorrilla and Philippe Hammann and Angel M Zamarreño and Jose M García-Mina and Vicente Rubio and Pascal Genschik and Roberto Solano}, doi = {10.1073/pnas.1912199117}, issn = {1091-6490 0027-8424 0027-8424}, year = {2020}, date = {2020-01-01}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {117}, number = {11}, pages = {6205--6215}, abstract = {The jasmonate (JA)-pathway regulators MYC2, MYC3, and MYC4 are central nodes in plant signaling networks integrating environmental and developmental signals to fine-tune JA defenses and plant growth. Continuous activation of MYC activity is potentially lethal. Hence, MYCs need to be tightly regulated in order to optimize plant fitness. Among the increasing number of mechanisms regulating MYC activity, protein stability is arising as a major player. However, how the levels of MYC proteins are modulated is still poorly understood. Here, we report that MYC2, MYC3, and MYC4 are targets of BPM (BTB/POZ-MATH) proteins, which act as substrate adaptors of CUL3-based E3 ubiquitin ligases. Reduction of function of CUL3(BPM) in amiR-bpm lines, bpm235 triple mutants, and cul3ab double mutants enhances MYC2 and MYC3 stability and accumulation and potentiates plant responses to JA such as root-growth inhibition and MYC-regulated gene expression. Moreover, MYC3 polyubiquitination levels are reduced in amiR-bpm lines. BPM3 protein is stabilized by JA, suggesting a negative feedback regulatory mechanism to control MYC activity, avoiding harmful runaway responses. Our results uncover a layer for JA-pathway regulation by CUL3(BPM)-mediated degradation of MYC transcription factors.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{mueller_sinusoidal_2020, title = {[The sinusoidal microenvironment regulates the niche and the differentiation of lymph node macrophages]}, author = {Christopher G Mueller and Abdouramane Camara and Vincent Flacher}, doi = {10.1051/medsci/2020148}, issn = {1958-5381}, year = {2020}, date = {2020-01-01}, journal = {Medecine Sciences: M/S}, volume = {36}, number = {10}, pages = {835--838}, keywords = {Animals, Capillaries, Cell Differentiation, Cellular, Humans, Immunity, Lymph Nodes, Lymphatic Vessels, Macrophages, Stem Cell Niche, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural Differences in Translation Initiation between Pathogenic Trypanosomatids and Their Mammalian Hosts}, author = {A Bochler and J B Querido and T Prilepskaja and H Soufari and A Simonetti and M L Del Cistia and L Kuhn and A R Ribeiro and L S Valášek and Y Hashem}, url = {https://pubmed.ncbi.nlm.nih.gov/33357443/}, doi = {10.1016/j.celrep.2020.108534}, isbn = {33357443}, year = {2020}, date = {2020-01-01}, urldate = {2020-01-01}, journal = {Cell Rep}, volume = {33}, number = {12}, pages = {108534}, abstract = {Canonical mRNA translation in eukaryotes begins with the formation of the 43S pre-initiation complex (PIC). Its assembly requires binding of initiator Met-tRNAiMet and several eukaryotic initiation factors (eIFs) to the small ribosomal subunit (40S). Compared to their mammalian hosts, trypanosomatids present significant structural differences in their 40S, suggesting substantial variability in translation initiation. Here, we determine the structure of the 43S PIC from Trypanosoma cruzi, the parasite causing Chagas disease. Our structure shows numerous specific features, such as the variant eIF3 structure and its unique interactions with the large rRNA expansion segments (ESs) 9S, 7S, and 6S, and the association of a kinetoplastid-specific DDX60-like helicase. It also reveals the 40S-binding site of the eIF5 C-terminal domain and structures of key terminal tails of several conserved eIFs underlying their activities within the PIC. Our results are corroborated by glutathione S-transferase (GST) pull-down assays in both human and T. cruzi and mass spectrometry data.}, keywords = {ENNIFAR, ES6(S) ES7(S) ES9(S) Trypanosoma cruzi cryo-EM eIF1 eIF2 eIF3 eIF5-CTD k-DDX60 the 43S pre-initiation complex translation initiation, HASCHEM, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {La Société de Biologie de Strasbourg: 100 ans au service de la science et de la société}, author = {P Antony and S Fournel and J Zoll and J M Mantz and K Befort and D Massotte and P Giégé and J Céraline and D Metzger and H Becker and L Drouard and C Florentz and R Martin and C Nébigil and S Potier and A Schaefer and E Schaeffer and C Schuster and A Bresson and E Quéméneur and L Choulier and N Matt and L Monassier and C Lugnier and L Freysz and J A Hoffmann and H Dreyfus and C Romier}, url = {https://pubmed.ncbi.nlm.nih.gov/33357372/}, isbn = {33357372}, year = {2020}, date = {2020-01-01}, journal = {Biol Aujourdhui}, volume = {214}, number = {3-4}, pages = {137-148}, abstract = {Founded in 1919, the Society of Biology of Strasbourg (SBS) is a learned society whose purpose is the dissemination and promotion of scientific knowledge in biology. Subsidiary of the Society of Biology, the SBS celebrated its Centenary on Wednesday, the 16th of October 2019 on the Strasbourg University campus and at the Strasbourg City Hall. This day allowed retracing the various milestones of the SBS, through its main strengths, its difficulties and its permanent goal to meet scientific and societal challenges. The common thread of this day was the transmission of knowledge related to the past, the present, but also the future. At the start of the 21st century, the SBS must continue to reinvent itself to pursue its objective of transmitting scientific knowledge in biology and beyond. Scientific talks performed by senior scientists and former SBS thesis prizes awardees, a round table, and informal discussions reflected the history and the dynamism of the SBS association. All SBS Centennial participants have set the first milestone for the SBS Bicentennial.}, keywords = {Historique History achievements and milestones futur de la recherche en biologie future of research in biology interdisciplinarity interdisciplinarité réalisations transmission des savoirs/connaissances transmission of knowledge, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A dimerization-based fluorogenic dye-aptamer module for RNA imaging in live cells}, author = {F Bouhedda and K T Fam and M Collot and A Autour and S Marzi and A Klymchenko and M Ryckelynck}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31636432}, doi = {https://doi.org/10.1038/s41589-019-0381-8}, isbn = {31636432}, year = {2020}, date = {2020-01-01}, journal = {Nat Chem Biol}, volume = {16}, number = {1}, pages = {69-76}, abstract = {Live-cell imaging of RNA has remained a challenge because of the lack of naturally fluorescent RNAs. Recently developed RNA aptamers that can light-up small fluorogenic dyes could overcome this limitation, but they still suffer from poor brightness and photostability. Here, we propose the concept of a cell-permeable fluorogenic dimer of self-quenched sulforhodamine B dyes (Gemini-561) and the corresponding dimerized aptamer (o-Coral) that can drastically enhance performance of the current RNA imaging method. The improved brightness and photostability, together with high affinity of this complex, allowed direct fluorescence imaging in live mammalian cells of RNA polymerase III transcription products as well as messenger RNAs labeled with a single copy of the aptamer; that is, without tag multimerization. The developed fluorogenic module enables fast and sensitive detection of RNA inside live cells, while the proposed design concept opens the route to new generation of ultrabright RNA probes.}, keywords = {ROMBY, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid32105815, title = {Analysis of RNA binding properties of human Ku protein reveals its interactions with 7SK snRNA and protein components of 7SK snRNP complex}, author = {Olga Shadrina and Irina Garanina and Sergey Korolev and Timofei Zatsepin and Jeanne Van Assche and Fadoua Daouad and Clementine Wallet and Olivier Rohr and Marina Gottikh}, doi = {10.1016/j.biochi.2020.02.016}, issn = {1638-6183}, year = {2020}, date = {2020-01-01}, urldate = {2020-01-01}, journal = {Biochimie}, volume = {171-172}, pages = {110--123}, abstract = {Human Ku heterodimeric protein composed of Ku70 and Ku80 subunits plays an important role in the non-homologous end-joining DNA repair pathway as a sensor of double strand DNA breaks. Ku is also involved in numerous cellular processes, and in some of them it acts in an RNA-dependent manner. However, RNA binding properties of the human Ku have not been well studied. Here we have analyzed interactions of a recombinant Ku heterodimer with a set of RNAs of various structure as well as eCLIP (enhanced crosslinking and immunoprecipitation) data for human Ku70. As a result, we have proposed a consensus RNA structure preferable for the Ku binding that is a hairpin possessing a bulge just near GpG sequence-containing terminal loop. 7SK snRNA is a scaffold for a ribonucleoprotein complex (7SK snRNP), which is known to participate in transcription regulation. We have shown that the recombinant Ku specifically binds a G-rich loop of hairpin 1 within 7SK snRNA. Moreover, Ku protein has been co-precipitated from HEK 293T cells with endogenous 7SK snRNA and such proteins included in 7SK snRNP as HEXIM1, Cdk9 and CTIP2. Ku and Cdk9 binding is found to be RNA-independent, meanwhile HEXIM1 and Ku co-precipitation depended on the presence of intact 7SK snRNA. The latter result has been confirmed using recombinant HEXIM1 and Ku proteins. Colocalization of Ku and CTIP2 was additionally confirmed by confocal microscopy. These results allow us to propose human Ku as a new component of the 7SK snRNP complex.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{gurcan_closer_2019, title = {A closer look at the genotoxicity of graphene based materials}, author = {Cansu Gurcan and Hadiseh Taheri and Alberto Bianco and Lucia Gemma Delogu and Acelya Yilmazer}, url = {https://doi.org/10.1088%2F2515-7639%2Fab5844}, doi = {10.1088/2515-7639/ab5844}, issn = {2515-7639}, year = {2019}, date = {2019-12-01}, urldate = {2020-04-01}, journal = {Journal of Physics: Materials}, volume = {3}, number = {1}, pages = {014007}, abstract = {Graphene-based materials (GBMs) have attracted many scientists because of their optical, thermal, mechanical and electronic properties. Their good dispersibility in different type of solvents including water, the possibility to formulate them according to desired function, and the wide surface area, which can allow various chemical modifications, expanded the use of these materials in biological systems. For these reasons, GBMs have been extensively studied in vitro and in vivo in the biomedical field. However, the toxicity and genotoxicity of GBMs must be thoroughly investigated before they can be translated into clinical settings. The main mechanism of graphene toxicity is thought to be caused by reactive oxygen species produced in cells, which in turn interact with various biomolecules including DNA. In this review we aimed to discuss different genotoxicity studies performed with GBMs with specific focus on the different cell types and conditions. By comparing and discussing such reports, scientists will be able to engineer non toxic GBMs for future preclinical and/or clinical studies. In order to allow a safer and faster transition to clinic, future studies should involve state-of-the-art technologies such as systems biology approaches or three-dimensional microfluidic systems, which can better predict the normal physiological scenario.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bouchoucha_determination_2019, title = {Determination of protein-only RNase P interactome in Arabidopsis mitochondria and chloroplasts identifies a complex between PRORP1 and another NYN domain nuclease.}, author = {Ayoub Bouchoucha and Florent Waltz and Géraldine Bonnard and Mathilde Arrivé and Philippe Hammann and Lauriane Kuhn and Cédric Schelcher and Hélène Zuber and Anthony Gobert and Philippe Giegé}, doi = {10.1111/tpj.14458}, issn = {1365-313X 0960-7412}, year = {2019}, date = {2019-11-01}, journal = {The Plant journal : for cell and molecular biology}, volume = {100}, number = {3}, pages = {549--561}, abstract = {The essential type of endonuclease that removes 5' leader sequences from transfer RNA precursors is called RNase P. While ribonucleoprotein RNase P enzymes containing a ribozyme are found in all domains of life, another type of RNase P called 'PRORP', for 'PROtein-only RNase P', is composed of protein that occurs only in a wide variety of eukaryotes, in organelles and in the nucleus. Here, to find how PRORP functions integrate with other cell processes, we explored the protein interaction network of PRORP1 in Arabidopsis mitochondria and chloroplasts. Although PRORP proteins function as single subunit enzymes in vitro, we found that PRORP1 occurs in protein complexes and is present in high-molecular-weight fractions that contain mitochondrial ribosomes. The analysis of immunoprecipitated protein complexes identified proteins involved in organellar gene expression processes. In particular, direct interaction was established between PRORP1 and MNU2 a mitochondrial nuclease. A specific domain of MNU2 and a conserved signature of PRORP1 were found to be directly accountable for this protein interaction. Altogether, results revealed the existence of an RNA maturation complex in Arabidopsis mitochondria and suggested that PRORP proteins cooperated with other gene expression factors for RNA maturation in vivo.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{bordoni_stimulation_2019, title = {Stimulation of bone formation by monocyte-activator functionalized graphene oxide in vivo}, author = {Valentina Bordoni and Giacomo Reina and Marco Orecchioni and Giulia Furesi and Stefanie Thiele and Chiara Gardin and Barbara Zavan and Gianaurelio Cuniberti and Alberto Bianco and Martina Rauner and Lucia G Delogu}, doi = {10.1039/c9nr03975a}, issn = {2040-3372}, year = {2019}, date = {2019-11-01}, journal = {Nanoscale}, volume = {11}, number = {41}, pages = {19408--19421}, abstract = {Nanosystems are able to enhance bone regeneration, a complex process requiring the mutual interplay between immune and skeletal cells. Activated monocytes can communicate pro-osteogenic signals to mesenchymal stem cells and promote osteogenesis. Thus, the activation of monocytes is a promising strategy to improve bone regeneration. Nanomaterials specifically selected to provoke immune-mediated bone formation are still missing. As a proof of concept, we apply here the intrinsic immune-characteristics of graphene oxide (GO) with the well-recognized osteoinductive capacity of calcium phosphate (CaP) in a biocompatible nanomaterial called maGO-CaP (monocytes activator GO complexed with CaP). In the presence of monocytes, the alkaline phosphatase activity and the expression of osteogenic markers increased. Studying the mechanisms of action, we detected an up-regulation of Wnt and BMP signaling, two key osteogenic pathways. The role of the immune activation was evidenced by the over-production of oncostatin M, a pro-osteogenic factor produced by monocytes. Finally, we tested the pro-osteogenic effects of maGO-CaP in vivo. maGO-CaP injected into the tibia of mice enhanced local bone mass and the bone formation rate. Our study suggests that maGO-CaP can activate monocytes to enhance osteogenesis ex vivo and in vivo.}, keywords = {Animals, Biocompatible Materials, Bone Morphogenetic Protein 2, Calcium Phosphates, Cell Differentiation, Cell Survival, Coculture Techniques, Graphite, Humans, I2CT, Inbred C57BL, Male, Mesenchymal Stem Cells, Mice, Monocytes, Oncostatin M, Osteoblasts, Osteogenesis, Signal Transduction, Team-Bianco, Tibia, Wnt Proteins}, pubstate = {published}, tppubtype = {article} } @article{Brown2019, title = {Large expert-curated database for benchmarking document similarity detection in biomedical literature search}, author = {P Brown, RELISH Consortium and Y Zhou}, doi = {https://doi.org/10.1093/database/baz085}, year = {2019}, date = {2019-10-29}, journal = {Database}, volume = {2019}, number = {2019}, pages = {baz085}, abstract = {Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency–Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{vargas-franco_genetically-achieved_2019, title = {Genetically-achieved disturbances to the expression levels of TNFSF11 receptors modulate the effects of zoledronic acid on growing mouse skeletons}, author = {Jorge William Vargas-Franco and Beatriz Castaneda and Andrea Gama and Christopher G Mueller and Dominique Heymann and Françoise Rédini and Frédéric Lézot}, doi = {10.1016/j.bcp.2019.06.027}, issn = {1873-2968}, year = {2019}, date = {2019-10-01}, journal = {Biochemical Pharmacology}, volume = {168}, pages = {133--148}, abstract = {Zoledronic acid (ZOL), a nitrogen bisphosphonate (N-BP), is currently used to treat and control pediatric osteolytic diseases. Variations in the intensity of the effects and side effects of N-BPs have been reported with no clear explanations regarding their origins. We wonder if such variations could be associated with different levels of RANKL signaling activity in growing bone during and after the treatment with N-BPs. To answer this question, ZOL was injected into neonate C57BL/6J mice with different genetically-determined RANKL signaling activity levels (Opg+/+textbackslashRankTg-, Opg+/+textbackslashRankTg+, Opg+/-textbackslashRankTg-, Opg+/-textbackslashRankTg+, Opg-/-textbackslashRankTg- and Opg-/-textbackslashRankTg+ mice) following a protocol (4 injections from post-natal day 1 to 7 at the dose of 50 μg/kg) that mimics those used in onco-pediatric patients. At the end of pediatric growth (1 and half months) and at an adult age (10 months), the bone morphometric and mineral parameters were measured using μCT in the tibia and skull for the different mice. A histologic analysis of the dental and periodontal tissues was also performed. At the end of pediatric growth, a delay in long bone and skull bone growth, a blockage of tooth eruption, some molar root alterations and a neoplasia-like structure associated with incisor development were found. Interestingly, the magnitude of these side effects was reduced by Opg deficiency (Opg-/-) but increased by Rank overexpression (RankTg). Analysis of the skeletal phenotype at ten months confirmed respectively the beneficial and harmful effects of Opg deficiency and Rank overexpression. These results validated the hypothesis that the RANKL signaling activity level in the bone microenvironment is implicated in the modulation of the response to ZOL. Further studies will be necessary to understand the underlying molecular mechanisms, which will help decipher the variability in the effects of N-BPs reported in the human population. SIGNIFICANT STATEMENTS: The present study establishes that in mice the RANKL signaling activity level is a major modulator of the effects and side-effects of bisphosphonates on the individual skeleton during growth. However, the modulatory actions are dependent on the ways in which this level of activity is increased. A decrease in OPG expression is beneficial to the skeletal phenotype observed at the end of growth, while RANK overexpression deteriorates it. Far removed from pediatric treatment, in adults, the skeletal phenotypes initially observed at the end of growth for the different levels of RANKL signaling activity were maintained, although significant improvement was associated only with reductions in OPG expression.}, keywords = {Animals, Bone Density Conservation Agents, Bone Development, Craniofacial bone, Gene Knockout Techniques, Growth, Inbred C57BL, Knockout, Long bone, Mice, Newborn, Osteoprotegerin, RANK ligand, RANKL/RANK/OPG, Skull, Team-Mueller, Tibia, Tooth, X-Ray Microtomography, Zoledronic acid}, pubstate = {published}, tppubtype = {article} } @article{lange_rst1_2019, title = {RST1 and RIPR connect the cytosolic RNA exosome to the Ski complex in Arabidopsis.}, author = {Heike Lange and Simon Y A Ndecky and Carlos Gomez-Diaz and David Pflieger and Nicolas Butel and Julie Zumsteg and Lauriane Kuhn and Christina Piermaria and Johana Chicher and Michael Christie and Ezgi S Karaaslan and Patricia L M Lang and Detlef Weigel and Hervé Vaucheret and Philippe Hammann and Dominique Gagliardi}, doi = {10.1038/s41467-019-11807-4}, issn = {2041-1723 2041-1723}, year = {2019}, date = {2019-08-01}, journal = {Nature communications}, volume = {10}, number = {1}, pages = {3871}, abstract = {The RNA exosome is a key 3'-5' exoribonuclease with an evolutionarily conserved structure and function. Its cytosolic functions require the co-factors SKI7 and the Ski complex. Here we demonstrate by co-purification experiments that the ARM-repeat protein RESURRECTION1 (RST1) and RST1 INTERACTING PROTEIN (RIPR) connect the cytosolic Arabidopsis RNA exosome to the Ski complex. rst1 and ripr mutants accumulate RNA quality control siRNAs (rqc-siRNAs) produced by the post-transcriptional gene silencing (PTGS) machinery when mRNA degradation is compromised. The small RNA populations observed in rst1 and ripr mutants are also detected in mutants lacking the RRP45B/CER7 core exosome subunit. Thus, molecular and genetic evidence supports a physical and functional link between RST1, RIPR and the RNA exosome. Our data reveal the existence of additional cytosolic exosome co-factors besides the known Ski subunits. RST1 is not restricted to plants, as homologues with a similar domain architecture but unknown function exist in animals, including humans.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{mueller_skin_2019, title = {Of skin and bone: did Langerhans cells and osteoclasts evolve from a common ancestor?}, author = {Christopher G Mueller and Benjamin Voisin}, doi = {10.1111/joa.12543}, issn = {1469-7580}, year = {2019}, date = {2019-08-01}, journal = {Journal of Anatomy}, volume = {235}, number = {2}, pages = {412--417}, abstract = {Skin Langerhans cells are antigen-presenting cells of the interfollicular epidermis and the upper part of the hair follicle, whereas osteoclasts are specialized bone-resorbing macrophages. Although at first view these two cell types appear to have little in common, a closer analysis reveals shared features, and when taking into account their surrounding environment, a hypothesis can be developed that Langerhans cells and osteoclasts have evolved from a common ancestral cell type. In this mini-review, we have compared the ontogenetic features of Langerhans cells and osteoclasts from a genetic and a functional point of view, an issue that so far has been overlooked. The gene programs that control cell differentiation, and the body parts where they reside, present surprising similarities. Whereas the function of osteoclasts in bone degradation has been established since the first vertebrates, Langerhans cells may have undergone a stepwise adaptation from aquatic to terrestrial life. Their cell function co-evolved with the imperatives of the skin to protect against physical impact, heat, water loss and pathogens, which implied the capacity of Langerhans cells to associate with skin appendages and to develop immunostimulatory functions. For the highly versatile and efficient immune system of modern vertebrates, Langerhans cells may be a memory of the past.}, keywords = {Animals, Biological Evolution, Dendritic cell, Evolution, hair follicle, Humans, Langerhans cell, Langerhans Cells, Macrophage, OSTEOCLAST, Osteoclasts, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{Talide_2020, title = {Sensing Viral Infections in Insects: A Dearth of Pathway Receptors}, author = {L Talide and JL Imler and C Meignin}, url = {https://www.caister.com/insectvirology2}, doi = {10.21775/cimb.034.031}, isbn = {9781912530090}, year = {2019}, date = {2019-06-06}, journal = {Curr Issues Mol Biol}, volume = {34}, pages = {31-60}, abstract = {Insects, the most diverse group of animals, can be infected by an extraordinary diversity of viruses. Among them, arthropod-borne viruses can be transmitted to humans, while bee and silkworm viruses cause important economic losses. Like all invertebrates, insects rely solely on innate immunity to counter viral infections. Protein-based mechanisms, involving restriction factors and evolutionarily conserved signaling pathways regulating transcription factors of the NF-kB and STAT families, participate in the control of viral infections in insects. In addition, RNA-based responses play a major role in the silencing of viral RNAs. We review here our current state of knowledge on insect antiviral defense mechanisms, which include conserved as well as adaptive, insect-specific strategies. Identification of the innate immunity receptors that sense viral infection in insects remains a major challenge for the field. }, keywords = {imler, innate immunity, M3i, meignin, STAT, viral Infection}, pubstate = {published}, tppubtype = {article} } @article{Olmo_2019, title = {The insect reservoir of biodiversity for viruses and for antiviral mechanisms}, author = {RP Olmo and NE Martins and ERGR Aguiar and JT Marques and JL Imler}, url = {https://www.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652019000600604&lng=en&nrm=iso&tlng=en}, doi = {10.1590/0001-3765201920190122}, year = {2019}, date = {2019-06-03}, journal = { An Acad Bras Cienc }, volume = {91}, number = {Suppl 3}, pages = {e20190122}, abstract = {Insects are the most diverse group of animals. They can be infected by an extraordinary diversity of viruses. Among them, arthropod-borne viruses (arboviruses) can be transmitted to humans. High-throughput sequencing of small RNAs from insects provides insight into their virome, which may help understand the dynamics of vector borne infectious diseases. Furthermore, investigating the mechanisms that restrict viral infections in insects points to genetic innovations that may inspire novel antiviral strategies. }, keywords = {antiviral immunity, imler, insects, M3i, Marques, metagenomics, restriction factors, RNA Interference, virome}, pubstate = {published}, tppubtype = {article} } @inbook{Einhorn_2019, title = {Insect Immunity: From Systemic to Chemosensory Organs Protection}, author = {E. Einhorn and JL Imler}, editor = {JF Picimbon}, url = {https://link.springer.com/chapter/10.1007%2F978-3-030-05165-5_9}, isbn = {9783030051648}, year = {2019}, date = {2019-05-17}, volume = {2}, pages = {205-229}, publisher = {Springer}, abstract = {Insects are confronted to a wide range of infectious microorganisms. Tissues in direct contact with the environment, such as olfactory organs, are particularly exposed to pathogens. We review here the immune mechanisms operating in insects to control infections. Experiments conducted on the model organism Drosophila melanogaster (fruit fly) have provided genetic evidence that insects rely on both cellular and humoral mechanisms to control infections. Once epithelial barriers have been breached, circulating or membrane-associated innate immunity receptors trigger signaling in the fat body and lead to secretion of high concentrations of antimicrobial peptides active on fungi and bacteria in the hemolymph. This induced response involves the evolutionarily conserved Toll and immune deficiency (IMD) signaling pathways, which promote nuclear translocation of transcription factors of the NF-κB family. In addition, different subsets of differentiated blood cells or hemocytes can neutralize bacteria, fungi or parasites by phagocytosis, production of microbicidal compounds, or encapsulation. An alternative to mount costly immune responses is to sense pathogens through chemosensory cues and avoid them. Interestingly, some families of molecules, including the Toll receptors, participate in both olfaction and immunity.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {inbook} } @article{schaeffer_sp2-iminosugar_2019, title = {sp2-Iminosugar glycolipids as inhibitors of lipopolysaccharide-mediated human dendritic cell activation in vitro and of acute inflammation in mice in vivo}, author = {Evelyne Schaeffer and Elena M Sánchez-Fernández and Rita Gonçalves-Pereira and Vincent Flacher and Delphine Lamon and Monique Duval and Jean-Daniel Fauny and José M García Fernández and Christopher G Mueller and Carmen Ortiz Mellet}, doi = {10.1016/j.ejmech.2019.02.078}, issn = {1768-3254}, year = {2019}, date = {2019-05-01}, journal = {European Journal of Medicinal Chemistry}, volume = {169}, pages = {111--120}, abstract = {Glycolipid mimetics consisting of a bicyclic polyhydroxypiperidine-cyclic carbamate core and a pseudoanomeric hydrophobic tail, termed sp2-iminosugar glycolipids (sp2-IGLs), target microglia during neuroinflammatory processes. Here we have synthesized and investigated new variants of sp2-IGLs for their ability to suppress the activation of human monocyte-derived dendritic cells (DCs) by lipopolysaccharide (LPS) signaling through Toll-like receptor 4. We report that the best lead was (1R)-1-dodecylsulfonyl-5N,6O-oxomethylidenenojirimycin (DSO2-ONJ), able to inhibit LPS-induced TNFα production and maturation of DCs. Immunovisualization experiments, using a mannoside glycolipid conjugate (MGC) that also suppress LPS-mediated DC activation as control, evidenced a distinct mode of action for the sp2-IGLs: unlike MGCs, DSO2-ONJ did not elicit internalization of the LPS co-receptor CD14 or induce its co-localization with the Toll-like receptor 4. In a mouse model of LPS-induced acute inflammation, DSO2-ONJ demonstrated anti-inflammatory activity by inhibiting the production of the pro-inflammatory interleukin-6. The ensemble of the data highlights sp2-IGLs as a promising new class of molecules against inflammation by interfering in Toll-like receptor intracellular signaling.}, keywords = {Activation, Acute Disease, Animals, antagonists & inhibitors, CD14, Cells, chemical synthesis, Chemistry, CO-RECEPTOR, Cultured, Dendritic cell, Dendritic Cells, Dose-Response Relationship, Drug, drug effects, drug therapy, Glycolipid, Glycolipids, Human, Humans, Iminosugar, immunopathology, IN VITRO, In vivo, Inbred C57BL, inflammation, Interleukin-6, lipopolysaccharide, Lipopolysaccharides, LPS, Male, Maturation, metabolism, Mice, MICROGLIA, Molecular Structure, mouse, pathology, Pharmacology, PRODUCTION, Receptor, signaling, Structure-Activity Relationship, Sulfone, Sulfoxide, Tail, target, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{Martins_2019jbio, title = {Les insectes : un fantastique réservoir de virus et de gènes antiviraux}, author = {NE Martins and RP Olmo and ERGR Aguiar and JT Marques and JL Imler}, url = {https://www.biologie-journal.org/articles/jbio/abs/2018/02/jbio190008/jbio190008.html}, doi = {10.1051/jbio/2019008}, year = {2019}, date = {2019-04-11}, journal = {Biologie Aujourd'hui}, volume = {212}, number = {3-4}, pages = {101-106}, abstract = {Les insectes forment le groupe d’animaux qui présente la plus grande diversité. Des travaux récents de métagénomique montrent qu’ils peuvent être infectés par une diversité extraordinaire de virus. Parmi eux, les arbovirus (arthropod-borne viruses) peuvent être transmis à l’Homme par les insectes hématophages, notamment les moustiques. Le séquençage à haut débit des petits ARN des insectes fournit des informations sur leur virome, un paramètre qui pourrait contribuer à expliquer la dynamique de la transmission des maladies infectieuses par des insectes vecteurs. D’autre part, la caractérisation des mécanismes qui restreignent les infections virales chez les insectes révèle des innovations génétiques qui pourraient à terme inspirer de nouvelles stratégies antivirales.}, keywords = {antiviral immunity, ARN interference, imler, Insect, M3i, Marques, metagenomic, virome}, pubstate = {published}, tppubtype = {article} } @article{Martins_2019i, title = {A Transgenic Flock House Virus Replicon Reveals an RNAi Independent Antiviral Mechanism Acting in Drosophila Follicular Somatic Cells }, author = {N Martins and A Lemoine and E Santiago and S Paro and JL Imler and C Meignin}, url = {https://www.g3journal.org/content/9/2/403.long}, doi = {10.1534/g3.118.200872}, year = {2019}, date = {2019-02-07}, journal = { G3 (Bethesda)}, volume = {9}, number = {2}, pages = {403-412}, keywords = {antiviral immunity, Drosophila melanogaster, Follicular somatic cells, Genetics of Immunity, imler, M3i, meignin, replicon, Viral replicon}, pubstate = {published}, tppubtype = {article} } @article{Franchet2019, title = {Phosphatidic acid as a limiting host metabolite for the proliferation of the microsporidium Tubulinosema ratisbonensis in Drosophila flies}, author = {Adrien Franchet and Sebastian Niehus and Gaetan Caravello and Dominique Ferrandon}, editor = {Nature Publishing Group}, url = {https://www.nature.com/articles/s41564-018-0344-y}, doi = {10.1038/s41564-018-0344-y}, isbn = {2058-5276}, year = {2019}, date = {2019-01-28}, journal = {Nature Microbiology}, volume = {4}, number = {4}, pages = {645}, abstract = {A Drosophila melanogaster systemic infection model for the microsporidian Tubulinosema ratisbonensis reveals that the parasite hijacks host phosphatidic acid, which is a limiting precursor for synthesis of parasite membranes and therefore proliferation.}, keywords = {Drosophila, ferrandon, Host-Parasite Interactions, Lipids, M3i, metabolism, microsporidia}, pubstate = {published}, tppubtype = {article} } @article{Achee2019, title = {Alternative strategies for mosquito-borne arbovirus control}, author = { Nicole L. Achee and John P. Grieco and Hassan Vatandoost and Gonçalo Seixas and Joao Pinto and Lee Ching-NG and Ademir J. Martins and Waraporn Juntarajumnong and Vincent Corbel and Clement Gouagna and Jean-Philippe David and James G. Logan and James Orsborne and Eric Marois and Gregor J. Devine and John Vontas }, url = {https://doi.org/10.1371/journal.pntd.0006822}, doi = {10.1371/journal.pntd.0006822}, year = {2019}, date = {2019-01-03}, journal = {PLoS Neglected Tropical Diseases}, abstract = {International public health workers are challenged by the burden of arthropod-borne viral diseases, to include mosquito-borne arboviruses transmitted by Aedes aegypti and A. albopictus due in part to lack of sustainable vector control and insecticide resistance (IR), as well as the inability to scale up and sustain existing interventions for prevention of urban epidemics. As a consequence, there has been increasing interest to advance the development of alternative methods. This review provides a general overview of alternative vector-control strategies under development for the control of arbovirus mosquito vectors and highlights how each could offer innovative public health value. Considerations to regulations, acceptance, and sustainability are also provided.}, keywords = {arbovirus, M3i, marois, mosquito}, pubstate = {published}, tppubtype = {article} } @article{fillatre_teads_2019, title = {TEADs, Yap, Taz, Vgll4s transcription factors control the establishment of Left-Right asymmetry in zebrafish}, author = {Jonathan Fillatre and Jean-Daniel Fauny and Jasmine Alexandra Fels and Cheng Li and Mary Goll and Christine Thisse and Bernard Thisse}, doi = {10.7554/eLife.45241}, issn = {2050-084X}, year = {2019}, date = {2019-01-01}, journal = {eLife}, volume = {8}, abstract = {In many vertebrates, establishment of Left-Right (LR) asymmetry results from the activity of a ciliated organ functioning as the LR Organizer (LRO). While regulation of the formation of this structure by major signaling pathways has been described, the transcriptional control of LRO formation is poorly understood. Using the zebrafish model, we show that the transcription factors and cofactors mediating or regulating the transcriptional outcome of the Hippo signaling pathway play a pivotal role in controlling the expression of genes essential to the formation of the LRO including ligands and receptors of signaling pathways involved in this process and most genes required for motile ciliogenesis. Moreover, the transcription cofactor, Vgll4l regulates epigenetic programming in LRO progenitors by controlling the expression of writers and readers of DNA methylation marks. Altogether, our study uncovers a novel and essential role for the transcriptional effectors and regulators of the Hippo pathway in establishing LR asymmetry.}, keywords = {Animals, Body Patterning, Developmental, developmental biology, Gene Expression Regulation, Hippo pathway, I2CT, Imagerie, Left-Right asymmetry, Left-Right Organizer, Signal Transduction, Taz, Transcription Factors, Vgll4, Yap, Zebrafish}, pubstate = {published}, tppubtype = {article} } @article{murera_few_2019, title = {Few layer graphene does not affect the function and the autophagic activity of primary lymphocytes}, author = {Diane Murera and Sowmya Malaganahalli and Cristina Martín and Giacomo Reina and Jean-Daniel Fauny and Hélène Dumortier and Ester Vázquez and Alberto Bianco}, doi = {10.1039/c9nr00846b}, issn = {2040-3372}, year = {2019}, date = {2019-01-01}, journal = {Nanoscale}, volume = {11}, number = {21}, pages = {10493--10503}, abstract = {Carbon-based nanomaterials represent a new tool in future medical applications. Thus, focusing on the evaluation of the degree of their safety has been growing in the last years. In this study we were particularly interested in understanding the impact of few layer graphene (FLG) on primary murine lymphocytes. These B and T cells, that are the second, but specialized, line of defense of the immune system, rely on various mechanisms to ensure their efficient function and maintenance. One of these mechanisms is autophagy that can be triggered by various nanomaterials in some types of cells. For these reasons, we were interested in evaluating the way FLG could affect this process in lymphocytes. Our results point out that FLG neither impacts the viability and activation of T and B cells nor their autophagic activity. Using confocal microscopy, we were also able to see that FLG does not appear to cause any membrane damage and does not penetrate inside of these cells. Overall, our data do not show any effect of this material on lymphocyte homeostasis, which is one more argument in favor of the continuation of studies investigating the potential of FLG for therapeutic applications.}, keywords = {Animals, Autophagy, B-Lymphocytes, Dumortier, Graphite, I2CT, Inbred BALB C, Mice, Nanostructures, T-Lymphocytes, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{, title = {The multiple flavors of GoU pairs in RNA}, author = {E Westhof and M Yusupov and G Yusupova}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31033092?dopt=Abstract}, doi = {10.1002/jmr.2782}, isbn = {31033092}, year = {2019}, date = {2019-01-01}, journal = {J Mol Recognit}, pages = {e2782}, abstract = {Wobble GU pairs (or GoU) occur frequently within double-stranded RNA helices interspersed within the standard G═C and A─U Watson-Crick pairs. However, other types of GoU pairs interacting on their Watson-Crick edges have been observed. The structural and functional roles of such alternative GoU pairs are surprisingly diverse and reflect the various pairings G and U can form by exploiting all the subtleties of their electronic configurations. Here, the structural characteristics of the GoU pairs are updated following the recent crystallographic structures of functional ribosomal complexes and the development in our understanding of ribosomal translation.}, keywords = {GoU pair anticodon codon mRNA miscoding ribosome tRNA tautomer, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pseudouridines or how to draw on weak energy differences}, author = {E Westhof}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31761086}, doi = {10.1016/j.bbrc.2019.10.009}, isbn = {31761086}, year = {2019}, date = {2019-01-01}, journal = {Biochem Biophys Res Commun}, volume = {520}, number = {4}, pages = {702-704}, abstract = {In many RNA molecules, pseudouridines occur at conserved positions in functional sites. A great diversity of pseudouridine synthases guarantees the insertion of the modified base at precise locations. The accepted structural role of pseudouridines is a reduction of the RNA flexibility around the modification site. However, experiments rarely yield clear-cut evidence. The article "Dynamic stacking of an expected branch point adenosine in duplexes containing pseudouridine-modified or unmodified U2 snRNA sites" published in 2019 in Biochemical and Biophysical Research Communication by Kennedy et al. constitute a provocative case [1]. This example illustrates how a definite conformational state can be selected through small energy differences in a constrained environment.}, keywords = {RNA, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{Vicens2019, title = {A forum where french-speaking faculty can exchange research on teaching}, author = {Q Vicens and X Coumoul and J L Souciet}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31141270?dopt=Abstract}, doi = {10.1002/bmb.21258}, isbn = {31141270}, year = {2019}, date = {2019-01-01}, journal = {Biochem Mol Biol Educ}, volume = {47}, number = {5}, pages = {599-606}, abstract = {In order to help promote instructional change at French-speaking universities in Europe, we initiated a series of 1-day events centered on learning innovations. Since 2015, these events have been taking place every 6 months at the Université Paris Descartes, with the moral support of three learned scientific societies, the French Academy of Sciences, and sponsoring by leaders in textbook editing and classroom technologies. Each event gathers ~ 40 participants (faculty members, postdocs, and educational specialists) from four countries (Belgium, France, Luxemburg, Switzerland) and invitees, who share their active learning practices, flipped classroom variations representing the most popular strategy. Their experience revealed that faculty who invest themselves in revamping teaching are still isolated at their institutions, although institutional and national support have now been gaining momentum. In particular, the role of educational specialists (known as ingénieurs pédagogiques in France) is key to help faculty move away from lecturing. Overall, our event series illustrates that a hands-off approach is effective to foster a cross-border community of committed academics in a context where the process of changing the way we teach at universities is still in its infancy.}, keywords = {ducational specialist faculty development instructional change, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Biallelic variants in \textit{LARS2} and \textit{KARS} cause deafness and (ovario)leukodystrophy}, author = {M S van der Knaap and M Bugiani and M I Mendes and L G Riley and D E C Smith and J Rudinger-Thirion and M Frugier and M Breur and J Crawford and J van Gaalen and M Schouten and M Willems and Q Waisfisz and F T Mau-Them and R J Rodenburg and R J Taft and B Keren and J Christodoulou and C Depienne and C Simons and G S Salomons and F Mochel}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30737337}, doi = {10.1212/WNL.0000000000007098}, isbn = {30737337}, year = {2019}, date = {2019-01-01}, journal = {Neurology}, volume = {92}, number = {11}, pages = {e1225-e1237}, abstract = {OBJECTIVE: To describe the leukodystrophy caused by pathogenic variants in LARS2 and KARS, encoding mitochondrial leucyl transfer RNA (tRNA) synthase and mitochondrial and cytoplasmic lysyl tRNA synthase, respectively. METHODS: We composed a group of 5 patients with leukodystrophy, in whom whole-genome or whole-exome sequencing revealed pathogenic variants in LARS2 or KARS. Clinical information, brain MRIs, and postmortem brain autopsy data were collected. We assessed aminoacylation activities of purified mutant recombinant mitochondrial leucyl tRNA synthase and performed aminoacylation assays on patients' lymphoblasts and fibroblasts. RESULTS: Patients had a combination of early-onset deafness and later-onset neurologic deterioration caused by progressive brain white matter abnormalities on MRI. Female patients with LARS2 pathogenic variants had premature ovarian failure. In 2 patients, MRI showed additional signs of early-onset vascular abnormalities. In 2 other patients with LARS2 and KARS pathogenic variants, magnetic resonance spectroscopy revealed elevated white matter lactate, suggesting mitochondrial disease. Pathology in one patient with LARS2 pathogenic variants displayed evidence of primary disease of oligodendrocytes and astrocytes with lack of myelin and deficient astrogliosis. Aminoacylation activities of purified recombinant mutant leucyl tRNA synthase showed a 3-fold loss of catalytic efficiency. Aminoacylation assays on patients' lymphoblasts and fibroblasts showed about 50% reduction of enzyme activity. CONCLUSION: This study adds LARS2 and KARS pathogenic variants as gene defects that may underlie deafness, ovarian failure, and leukodystrophy with mitochondrial signature. We discuss the specific MRI characteristics shared by leukodystrophies caused by mitochondrial tRNA synthase defects. We propose to add aminoacylation assays as biochemical diagnostic tools for leukodystrophies.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {py_convrot: rotation conventions, to understand and to apply}, author = {L Urzhumtseva and A Urzhumtsev}, url = {https://doi.org/10.1107/S1600576719007313}, doi = {10.1107/S1600576719007313}, year = {2019}, date = {2019-01-01}, journal = {J Appl Cryst}, volume = {52}, number = {4}, pages = {869-881}, abstract = {Rotation is a core crystallographic operation. Two sets of Cartesian coordinates of each point of a rotated object, those before and after rotation, are linearly related, and the coefficients of these linear combinations can be represented in matrix form. This 3 × 3 matrix is unique for all points and thus describes unambiguously a particular rotation. However, its nine elements are mutually dependent and are not interpretable in a straightforward way. To describe rotations by independent and comprehensible parameters, crystallographic software usually refers to Euler or to polar angles. In crystallography and cryo-electron microscopy, there exists a large choice of conventions, making direct comparison of rotation parameters difficult and sometimes confusing. The program py_convrot, written in Python, is a converter of parameters describing rotations. In particular, it deals with all possible choices of polar angles and with all kinds of Euler angles, including all choices of rotation axes and rotation directions. Using a menu, a user can build their own rotation parameterization; its action can be viewed with an interactive graphical tool, Demo. The tables in this article and the extended help pages of the program describe details of these parameterizations and the decomposition of rotation matrices into all types of parameters. The program allows orthogonalization conventions and symmetry operations to be taken into account. This makes the program and its supporting materials both an illustrative teaching material, especially for non-specialists in mathematics and computing, and a tool for practical use.}, keywords = {rigid-body rotation rotation matrices rotation conventions teaching tools interactive illustrations Euler angles polar angles direction cosines, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure and functional reselection of the Mango-III fluorogenic RNA aptamer}, author = {3rd R J Trachman and A Autour and S C Y Jeng and A Abdolahzadeh and A Andreoni and R Cojocaru and R Garipov and E V Dolgosheina and J R Knutson and M Ryckelynck and P J Unrau and A R Ferré-D'Amaré}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30992561?dopt=Abstract}, doi = {10.1038/s41589-019-0267-9}, isbn = {30992561}, year = {2019}, date = {2019-01-01}, journal = {Nat Chem Biol}, volume = {15}, number = {5}, pages = {472-479}, abstract = {Several turn-on RNA aptamers that activate small-molecule fluorophores have been selected in vitro. Among these, the ~30 nucleotide Mango-III is notable because it binds the thiazole orange derivative TO1-Biotin with high affinity and fluoresces brightly (quantum yield 0.55). Uniquely among related aptamers, Mango-III exhibits biphasic thermal melting, characteristic of molecules with tertiary structure. We report crystal structures of TO1-Biotin complexes of Mango-III, a structure-guided mutant Mango-III(A10U), and a functionally reselected mutant iMango-III. The structures reveal a globular architecture arising from an unprecedented pseudoknot-like connectivity between a G-quadruplex and an embedded non-canonical duplex. The fluorophore is restrained into a planar conformation by the G-quadruplex, a lone, long-range trans Watson-Crick pair (whose A10U mutation increases quantum yield to 0.66), and a pyrimidine perpendicular to the nucleobase planes of those motifs. The improved iMango-III and Mango-III(A10U) fluoresce ~50% brighter than enhanced green fluorescent protein, making them suitable tags for live cell RNA visualization.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HDAC4 Levels Control Sensibility toward Cisplatin in Gastric Cancer via the p53-p73/BIK Pathway}, author = {M E Spaety and A Gries and A Badie and A Venkatasamy and B Romain and C Orvain and K Yanagihara and K Okamoto and A C Jung and G Mellitzer and S Pfeffer and C Gaiddon}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31703394}, doi = {10.3390/cancers11111747}, isbn = {31703394}, year = {2019}, date = {2019-01-01}, journal = {Cancers (Basel)}, volume = {11}, number = {11}, pages = {1747}, abstract = {Gastric cancer (GC) remains a health issue due to the low efficiency of therapies, such as cisplatin. This unsatisfactory situation highlights the necessity of finding factors impacting GC sensibility to therapies. We analyzed the cisplatin pangenomic response in cancer cells and found HDAC4 as a major epigenetic regulator being inhibited. HDAC4 mRNA repression was partly mediated by the cisplatin-induced expression of miR-140. At a functional level, HDAC4 inhibition favored cisplatin cytotoxicity and reduced tumor growth. Inversely, overexpression of HDAC4 inhibits cisplatin cytotoxicity. Importantly, HDAC4 expression was found to be elevated in gastric tumors compared to healthy tissues, and in particular in specific molecular subgroups. Furthermore, mutations in HDAC4 correlate with good prognosis. Pathway analysis of genes whose expression in patients correlated strongly with HDAC4 highlighted DNA damage, p53 stabilization, and apoptosis as processes downregulated by HDAC4. This was further confirmed by silencing of HDAC4, which favored cisplatin-induced apoptosis characterized by cleavage of caspase 3 and induction of proapoptotic genes, such as BIK, in part via a p53-dependent mechanism. Altogether, these results reveal HDAC4 as a resistance factor for cisplatin in GC cells that impacts on patients' survival.}, keywords = {BIK HDAC4 cisplatin gastric cancer miR-140 p53 p73, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ribosomal mistranslation leads to silencing of the unfolded protein response and increased mitochondrial biogenesis}, author = {D Shcherbakov and Y Teo and H Boukari and A Cortes-Sanchon and M Mantovani and I Osinnii and J Moore and R Juskeviciene and M Brilkova and S Duscha and H S Kumar and E Laczko and H Rehrauer and E Westhof and R Akbergenov and E C Böttger}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31637312}, doi = {10.1038/s42003-019-0626-9}, isbn = {31637312}, year = {2019}, date = {2019-01-01}, journal = {Commun Biol}, volume = {2}, pages = {381}, abstract = {Translation fidelity is the limiting factor in the accuracy of gene expression. With an estimated frequency of 10-4, errors in mRNA decoding occur in a mostly stochastic manner. Little is known about the response of higher eukaryotes to chronic loss of ribosomal accuracy as per an increase in the random error rate of mRNA decoding. Here, we present a global and comprehensive picture of the cellular changes in response to translational accuracy in mammalian ribosomes impaired by genetic manipulation. In addition to affecting established protein quality control pathways, such as elevated transcript levels for cytosolic chaperones, activation of the ubiquitin-proteasome system, and translational slowdown, ribosomal mistranslation led to unexpected responses. In particular, we observed increased mitochondrial biogenesis associated with import of misfolded proteins into the mitochondria and silencing of the unfolded protein response in the endoplasmic reticulum.}, keywords = {Protein folding Protein transport Transcriptomics, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {ITC Studies of Ribosome/Antibiotics Interactions}, author = {E Schenckbecher and B Meyer and E Ennifar}, editor = {E Ennifar}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30929237}, doi = {10.1007/978-1-4939-9179-2_7}, isbn = {30929237}, year = {2019}, date = {2019-01-01}, booktitle = {Microcalorimetry of Biological Molecules: Methods and Protocols}, volume = {1964}, pages = {89-98}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {The fight against multiresistant bacteria responsible for nosocomial diseases has recently been classified as an absolute priority by the World Health Organization. For some organisms, priority status has even been assessed as critical, as almost all currently available antibiotics are now inefficient against these "super-bacteria." Ribosome is a major target of several antibiotics, and extensive biochemical and structural studies led to a better understanding of the mechanism of action of drugs targeting translation (Blair et al., Nat Rev Microbiol 13:42-51, 2015; Lin et al., Annu Rev Biochem 87:451-478, 2018; Wilson, Nat Rev Microbiol 12:35-48, 2014; Yonath, Annu Rev Biochem 74:649-79, 2005). However, our knowledge regarding thermodynamic data of compounds targeting the ribosome, which are yet essential for a complete understanding of translation inhibition mechanisms by drugs, is still very poor.In this chapter we describe the use of ITC microcalorimetry to investigate the binding of bacterial ribosome to two antibiotics targeting the peptide tunnel: macrolides and proline-rich antimicrobial peptides (PrAMPs). This strategy yields reliable and artifact-free binding parameters for antibiotics and provides an original view on ribosome/antibiotics interactions.}, keywords = {Antibiotics Antimicrobial peptide Competition ITC Macrolide Ribosome, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Molecular basis for the differential interaction of plant mitochondrial VDAC proteins with tRNAs}, author = {T Salinas and S E Farouk-Ameqrane and E Ubrig and C Sauter and A M Duchêne and L Maréchal-Drouard}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30517754?dopt=Abstract}, doi = {10.1093/nar/gky1247}, isbn = {30519697}, year = {2019}, date = {2019-01-01}, journal = {Nucleic Acids Res}, volume = {47}, number = {2}, pages = {1048-1049}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Development and engineering of artificial RNAs}, author = {M Ryckelynck}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31195094?dopt=Abstract}, doi = {10.1016/j.ymeth.2019.06.009}, isbn = {31195094}, year = {2019}, date = {2019-01-01}, journal = {Methods}, volume = {161}, pages = {1-2}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Importance of potassium ions for ribosome structure and function revealed by long-wavelength X-ray diffraction}, author = {A Rozov and I Khusainov and K El Omari and R Duman and V Mykhaylyk and M Yusupov and E Westhof and A Wagner and G Yusupova}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31175275?dopt=Abstract}, doi = {10.1038/s41467-019-10409-4}, isbn = {31175275}, year = {2019}, date = {2019-01-01}, journal = {Nat Commun}, volume = {10}, number = {1}, pages = {2519}, abstract = {The ribosome, the largest RNA-containing macromolecular machinery in cells, requires metal ions not only to maintain its three-dimensional fold but also to perform protein synthesis. Despite the vast biochemical data regarding the importance of metal ions for efficient protein synthesis and the increasing number of ribosome structures solved by X-ray crystallography or cryo-electron microscopy, the assignment of metal ions within the ribosome remains elusive due to methodological limitations. Here we present extensive experimental data on the potassium composition and environment in two structures of functional ribosome complexes obtained by measurement of the potassium anomalous signal at the K-edge, derived from long-wavelength X-ray diffraction data. We elucidate the role of potassium ions in protein synthesis at the three-dimensional level, most notably, in the environment of the ribosome functional decoding and peptidyl transferase centers. Our data expand the fundamental knowledge of the mechanism of ribosome function and structural integrity.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {AFFINImeter: A software to analyze molecular recognition processes from experimental data}, author = {A Pineiro and E Munoz and J Sabin and M Costas and M Bastos and A Velazquez-Campoy and P F Garrido and P Dumas and E Ennifar and L Garcia-Rio and J Rial and D Pérez and P Fraga and A Rodriguez and C Cotelo}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30849378?dopt=Abstract}, doi = {10.1016/j.ab.2019.02.031}, isbn = {30849378}, year = {2019}, date = {2019-01-01}, journal = {Anal Biochem}, volume = {5}, number = {577}, pages = {117-134}, abstract = {The comprehension of molecular recognition phenomena demands the understanding of the energetic and kinetic processes involved. General equations valid for the thermodynamic analysis of any observable that is assessed as a function of the concentration of the involved compounds are described, together with their implementation in the AFFINImeter software. Here, a maximum of three different molecular species that can interact with each other to form an enormous variety of surpramolecular complexes are considered. The corrections currently employed to take into account the effects of dilution, volume displacement, concentration errors and those due to external factors, especially in the case of ITC measurements, are included. The methods used to fit the model parameters to the experimental data, and to generate the uncertainties are described in detail. A simulation tool and the so called kinITC analysis to get kinetic information from calorimetric experiments are also presented. An example of how to take advantage of the AFFINImeter software for the global multi-temperature analysis of a system exhibiting cooperative 1:2 interactions is presented and the results are compared with data previously published. Some useful recommendations for the analysis of experiments aimed at studying molecular interactions are provided.}, keywords = {Affinity Analysis software Isothermal titration calorimetry Kinetics Molecular recognition Thermodynamics, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mapping of Shigella flexneri's tissue distribution and type III secretion apparatus activity during infection of the large intestine of guinea pigs}, author = {G Nigro and E T Arena and M Sachse and M Moya-Nilges and B S Marteyn and P J Sansonetti and F X Campbell-Valois}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31578543}, doi = {10.1093/femspd/ftz054}, isbn = {31578543}, year = {2019}, date = {2019-01-01}, journal = {Pathog Dis}, volume = {77}, number = {7}, pages = {ftz054}, abstract = {Shigella spp. are bacterial pathogens that invade the human colonic mucosa using a type III secretion apparatus (T3SA), a proteinaceous device activated upon contact with host cells. Active T3SAs translocate proteins that carve the intracellular niche of Shigella spp. Nevertheless, the activation state of the T3SA has not been addressed in vivo. Here, we used a green fluorescent protein transcription-based secretion activity reporter (TSAR) to provide a spatio-temporal description of S. flexneri T3SAs activity in the colon of Guinea pigs. First, we observed that early mucus release is triggered in the vicinity of luminal bacteria with inactive T3SA. Subsequent mucosal invasion showed bacteria with active T3SA associated with the brush border, eventually penetrating into epithelial cells. From 2 to 8 h post-challenge, the infection foci expanded, and these intracellular bacteria displayed homogeneously high-secreting activity, while extracellular foci within the lamina propria featured bacteria with low secretion activity. We also found evidence that within lamina propria macrophages, bacteria reside in vacuoles instead of accessing the cytosol. Finally, bacteria were cleared from tissues between 8 and 24 h post-challenge, highlighting the hit-and-run colonization strategy of Shigella. This study demonstrates how genetically encoded reporters can contribute to deciphering pathogenesis in vivo.}, keywords = {MARTEYN, Shigella in vivo infection fluorescence microscopy genetically encoded reporters large intestine type III secretion system, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Thermodynamic and Kinetic Analysis of Isothermal Titration Calorimetry Experiments by Using KinITC in AFFINImeter}, author = {E Munoz and J Sabin and J Rial and D Pérez and E Ennifar and P Dumas and A Pineiro}, editor = {E Ennifar}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30929246}, doi = {10.1007/978-1-4939-9179-2_16}, isbn = {30929246}, year = {2019}, date = {2019-01-01}, booktitle = {Microcalorimetry of Biological Molecules: Methods and Protocols}, volume = {1964}, pages = {225-239}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Standard molecular binding isothermal titration calorimetric (ITC) experiments are designed to get thermodynamic information: changes in Gibbs energy, enthalpy, and entropy associated to the studied process. Traditionally, the kinetic information contained in the ITC raw signal has been ignored. For a usual one-step process, this corresponds to the rate constants for the association and the dissociation of the complex (kon and koff). The availability of highly sensitive ITC instruments with low response time, together with the development of theoretical methods and of public software for the proper analysis of the signal, cancels any reason for not retrieving this kinetic information. Here we describe how to further exploit ITC experiments of simple one-step interactions by using the software AFFINImeter.The method is exemplified using a standard reference system for thermodynamic and kinetic molecular binding analysis: the interaction of carbonic anhydrase (CA) with its inhibitor 4-carboxybenzenesulfonamide (4-CBS) at several temperatures. It is to be emphasized that old experiments initially designed and executed just for thermodynamic analysis can be readily recycled by using AFFINImeter to retrieve the previously ignored kinetic information.}, keywords = {AFFINImeter Carbonic anhydrase Data analysis ITC KinITC Kinetics Molecular binding Thermodynamics, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Thermodynamics of Molecular Machines Using Incremental ITC}, author = {B Meyer and C da Veiga and P Dumas and E Ennifar}, editor = {E Ennifar}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30929240}, doi = {10.1007/978-1-4939-9179-2_10}, isbn = {30929240}, year = {2019}, date = {2019-01-01}, booktitle = {Microcalorimetry of Biological Molecules: Methods and Protocols}, volume = {1964}, pages = {129-140}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {Molecular biomachines, such as DNA and RNA polymerases or the ribosome, are fascinating biological assemblies able to swiftly perform repeated and highly regulated tasks, with a remarkable accuracy. Significant advances in structural studies during the past 20 years provided a wealth of information regarding their architecture and considerably contributed to a better understanding of their mechanism of action. However, the three-dimensional structure of a biological nanomachine alone does not provide access to its detailed mechanism of action, even when obtained at atomic resolution. When combined with other biophysical approaches, thermodynamic data, together with kinetic data, are essential for a complete description of any binding interaction, revealing forces driving complex formation and providing insights into mechanisms of action. We have developed an incremental ITC approach that is well-suitable for analysis of biomolecular machines. This strategy allows a dissection of molecular biomachine reactions through successive additions in the ITC cell of consecutive substrates.}, keywords = {ENNIFAR, HIV reverse transcriptase Incremental ITC Mechanism of action Molecular biomachines Thermodynamics, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {The evolution of RNA structural probing methods: From gels to next-generation sequencing}, author = {E Mailler and J C Paillart and R Marquet and R P Smyth and V Vivet-Boudou}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30485688?dopt=Abstract}, doi = {10.1002/wrna.1518}, isbn = {30485688}, year = {2019}, date = {2019-01-01}, journal = {Wiley Interdiscip Rev RNA}, volume = {10}, number = {2}, pages = {e1518}, abstract = {RNA molecules are important players in all domains of life and the study of the relationship between their multiple flexible states and the associated biological roles has increased in recent years. For several decades, chemical and enzymatic structural probing experiments have been used to determine RNA structure. During this time, there has been a steady improvement in probing reagents and experimental methods, and today the structural biologist community has a large range of tools at its disposal to probe the secondary structure of RNAs in vitro and in cells. Early experiments used radioactive labeling and polyacrylamide gel electrophoresis as read-out methods. This was superseded by capillary electrophoresis, and more recently by next-generation sequencing. Today, powerful structural probing methods can characterize RNA structure on a genome-wide scale. In this review, we will provide an overview of RNA structural probing methodologies from a historical and technical perspective. This article is categorized under: RNA Structure and Dynamics > RNA Structure, Dynamics, and Chemistry RNA Methods > RNA Analyses in vitro and In Silico RNA Methods > RNA Analyses in Cells.}, keywords = {MARQUET, NGS RNA probing structure, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Importance of cellular microRNAs in the regulation of viral infections]}, author = {P Lopez and E Girardi and S Pfeffer}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31532379}, doi = {10.1051/medsci/2019130}, isbn = {31532379}, year = {2019}, date = {2019-01-01}, journal = {Med Sci (Paris)}, volume = {35}, number = {8-9}, pages = {667-673}, abstract = {Viruses are obligatory intracellular parasites that rely on a wide range of cellular factors to successfully accomplish their infectious cycle. Among those, micro (mi)RNAs have recently emerged as important modulators of viral infections. These small regulatory molecules act as repressors of gene expression. During infection, miRNAs can function by targeting either cellular or viral RNAs. In this review, we will recapitulate what has been reported to date on this interplay between cellular miRNAs and viruses and the effect on the infection. Furthermore, we will briefly discuss the possibilities of interfering with the infection through the modulation of this pathway to develop novel antiviral therapies.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleobase carbonyl groups are poor Mg^{2+} inner-sphere binders but excellent monovalent ion binders - A critical PDB survey}, author = {F Leonarski and L D'Ascenzo and P Auffinger}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30409785?dopt=Abstract}, doi = {10.1261/rna.068437.118}, isbn = {30409785}, year = {2019}, date = {2019-01-01}, journal = {RNA}, volume = {25}, number = {2}, pages = {173-192}, abstract = {Precise knowledge of Mg2+ inner-sphere binding site characteristics is vital for understanding the structure and function of nucleic acid systems. Unfortunately, the PDB, that represents the main source of Mg2+ binding sites, contains a significant number of ion assignment issues that significantly blur our understanding of the functions of these ions. Here, following a preceding study devoted to Mg2+ binding to nucleobase nitrogens, we surveyed PDB nucleic acid crystallographic structures with resolutions < 2.9 Å to classify the Mg2+ inner-sphere binding patterns to nucleotide carbonyl, ribose hydroxyl, cyclic ether and phosphodiester oxygen atoms to derive a set of "prior-knowledge" nucleobase Mg2+ binding sites. We report that crystallographic examples of trustworthy nucleobase Mg2+ binding sites are fewer than expected given that many of those bind misidentified Na+ or K+ We also emphasize that binding of Na+ and K+ to nucleic acids is much more frequent than anticipated. Overall, we provide crystallographic evidence that nucleobases are poor inner-sphere Mg2+ binders but good binders for monovalent ions. Based on strict stereochemical criteria, we propose an extended set of guidelines designed to help in the assignment and validation of ions directly contacting nucleobase and ribose atoms. These guidelines should help in the interpretation of X-ray and cryo-EM solvent density maps. When borderline metal ion stereochemistry is observed, alternative placement of Na+, K+, or Ca2+ should be considered. We also critically examine the use of lanthanides (Yb3+, Tb3+) as Mg2+ substitutes in crystallography experiments.}, keywords = {DNA lanthanides magnesium ribosome ribozyme, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Lalaouna2019, title = {RsaC sRNA modulates the oxidative stress response of \textit{Staphylococcus aureus} during manganese starvation}, author = {D Lalaouna and J Baude and Z Wu and A Tomasini and J Chicher and S Marzi and F Vandenesch and P Romby and I Caldelari and K Moreau}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31504767?dopt=Abstract}, doi = {10.1093/nar/gkz728}, isbn = {31504767}, year = {2019}, date = {2019-01-01}, journal = {Nucleic Acids Res}, volume = {47}, number = {18}, pages = {9871-9887}, abstract = {The human opportunistic pathogen Staphylococcus aureus produces numerous small regulatory RNAs (sRNAs) for which functions are still poorly understood. Here, we focused on an atypical and large sRNA called RsaC. Its length varies between different isolates due to the presence of repeated sequences at the 5′ end while its 3′ part is structurally independent and highly conserved. Using MS2-affinity purification coupled with RNA sequencing (MAPS) and quantitative differential proteomics, sodA mRNA was identified as a primary target of RsaC sRNA. SodA is a Mn-dependent superoxide dismutase involved in oxidative stress response. Remarkably, rsaC gene is co-transcribed with the major manganese ABC transporter MntABC and, consequently, RsaC is mainly produced in response to Mn starvation. This 3′UTR-derived sRNA is released from mntABC-RsaC precursor after cleavage by RNase III. The mature and stable form of RsaC inhibits the synthesis of the Mn-containing enzyme SodA synthesis and favors the oxidative stress response mediated by SodM, an alternative SOD enzyme using either Mn or Fe as co-factor. In addition, other putative targets of RsaC are involved in oxidative stress (ROS and NOS) and metal homeostasis (Fe and Zn). Consequently, RsaC may balance two interconnected defensive responses, i.e. oxidative stress and metal-dependent nutritional immunity.}, keywords = {PPSE, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comment on "Evaluating Unexpectedly Short Non-covalent Distances in X-ray Crystal Structures of Proteins with Electronic Structure Analysis"}, author = {H Kruse and J Sponer and P Auffinger}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31424937?dopt=Abstract}, doi = {10.1021/acs.jcim.9b00473}, isbn = {31424937}, year = {2019}, date = {2019-01-01}, journal = {J Chem Inf Model}, volume = {59}, number = {9}, pages = {3605-3608}, abstract = {1 Institute of Biophysics of the Czech Academy of Sciences, Kralovopolska 135, CZ-61265 Brno, Czech Republic. 2 Architecture et Réactivité de l'ARN, Université de Strasbourg, Institut de Biologie Moléculaire et Cellulaire du CNRS, Strasbourg 67084, France.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Purification and Functional Characterization of a Biologically Active Full-Length Feline Immunodeficiency Virus (FIV) Pr50^{Gag}.}, author = {A Krishnan and V Pillai and A Chameettachal and L M Ali and F Nuzra Nagoor Pitchai and S Tariq and F Mustafa and R Marquet and T A Rizvi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31357656?report=&dispmax=200&tool=PubCrawler_2.23}, doi = {10.3390/v11080689}, isbn = {31357656}, year = {2019}, date = {2019-01-01}, journal = {Viruses}, volume = {11}, number = {8}, pages = {689}, abstract = {The feline immunodeficiency virus (FIV) full-length Pr50Gag precursor is a key player in the assembly of new viral particles. It is also a critical component of the efficient selection and packaging of two copies of genomic RNA (gRNA) into the newly formed virus particles from a wide pool of cellular and spliced viral RNA. To understand the molecular mechanisms involved during FIV gRNA packaging, we expressed the His6-tagged and untagged recombinant FIV Pr50Gag protein both in eukaryotic and prokaryotic cells. The recombinant Pr50Gag-His6-tag fusion protein was purified from soluble fractions of prokaryotic cultures using immobilized metal affinity chromatography (IMAC). This purified protein was able to assemble in vitro into virus-like particles (VLPs), indicating that it preserved its ability to oligomerize/multimerize. Furthermore, VLPs formed in eukaryotic cells by the FIV full-length Pr50Gag both in the presence and absence of His6-tag could package FIV sub-genomic RNA to similar levels, suggesting that the biological activity of the recombinant full-length Pr50Gag fusion protein was retained in the presence of His6-tag at the carboxy terminus. Successful expression and purification of a biologically active, recombinant full-length Pr50Gag-His6-tag fusion protein will allow study of the intricate RNA-protein interactions involved during FIV gRNA encapsidation.}, keywords = {Gag protein purification His-tag fusion protein Pr50Gag protein expression feline immunodeficiency virus (FIV) retroviral RNA packaging viral assembly, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Stabilizing role of structural elements within the 5´ Untranslated Region (UTR) and gag sequences in Mason-Pfizer monkey virus (MPMV) genomic RNA packaging}, author = {R M Kalloush and V Vivet-Boudou and L M Ali and V Pillai and F Mustafa and R Marquet and T A Rizvi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30773097?dopt=Abstract}, doi = {10.1080/15476286.2019.1572424}, isbn = {30773097}, year = {2019}, date = {2019-01-01}, journal = {RNA Biol}, volume = {16}, number = {5}, pages = {612-625}, abstract = {The Mason-Pfizer monkey virus (MPMV) genomic RNA (gRNA) packaging signal is a highly-structured element with several stem-loops held together by two phylogenetically conserved long-range interactions (LRIs) between U5 and gag complementary sequences. These LRIs play a critical role in maintaining the structure of the 5´ end of the MPMV gRNA. Thus, one could hypothesize that the overall RNA secondary structure of this region is further architecturally held together by three other stem loops (SL3, Gag SL1, and Gag SL2) comprising of sequences from the distal parts of the 5´untranslated region (5' UTR) to ~ 120 nucleotides into gag, excluding gag sequences involved in forming the U5-Gag LRIs. To provide functional evidence for the biological significance of these stem loops during gRNA encapsidation, these structural motifs were mutated and their effects on MPMV RNA packaging and propagation were tested in a single round trans-complementation assay. The mutant RNA structures were further studied by high throughput SHAPE (hSHAPE) assay. Our results reveal that sequences involved in forming these three stem loops do not play crucial roles at an individual level during MPMV gRNA packaging or propagation. Further structure-function analysis indicates that the U5-Gag LRIs have a more important architectural role in stabilizing the higher order structure of the 5´ UTR than the three stem loops which have a more secondary and perhaps indirect role in stabilizing the overall RNA secondary structure of the region. Our work provides a better understanding of the molecular interactions that take place during MPMV gRNA packaging.}, keywords = {MARQUET, MARQUET PAILLART Mason-Pfizer monkey virus (MPMV) RNA packaging RNA secondary structure Retroviruses U5/Gag LRIs gag, Mason-Pfizer monkey virus (MPMV) RNA packaging RNA secondary structure Retroviruses U5/Gag LRIs gag, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A tRNA-mimic Strategy to Explore the Role of G34 of tRNA^{Gly} in Translation and Codon Frameshifting}, author = {A Janvier and L Despons and L Schaeffer and A Tidu and F Martin and G Eriani}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31405256?dopt=Abstract}, doi = {10.3390/ijms20163911}, isbn = {31405256}, year = {2019}, date = {2019-01-01}, journal = {Int J Mol Sci}, volume = {20}, number = {16}, pages = {3911}, abstract = {Decoding of the 61 sense codons of the genetic code requires a variable number of tRNAs that establish codon-anticodon interactions. Thanks to the wobble base pairing at the third codon position, less than 61 different tRNA isoacceptors are needed to decode the whole set of codons. On the tRNA, a subtle distribution of nucleoside modifications shapes the anticodon loop structure and participates to accurate decoding and reading frame maintenance. Interestingly, although the 61 anticodons should exist in tRNAs, a strict absence of some tRNAs decoders is found in several codon families. For instance, in Eukaryotes, G34-containing tRNAs translating 3-, 4- and 6-codon boxes are absent. This includes tRNA specific for Ala, Arg, Ile, Leu, Pro, Ser, Thr, and Val. tRNAGly is the only exception for which in the three kingdoms, a G34-containing tRNA exists to decode C3 and U3-ending codons. To understand why G34-tRNAGly exists, we analysed at the genome wide level the codon distribution in codon +1 relative to the four GGN Gly codons. When considering codon GGU, a bias was found towards an unusual high usage of codons starting with a G whatever the amino acid at +1 codon. It is expected that GGU codons are decoded by G34-containing tRNAGly, decoding also GGC codons. Translation studies revealed that the presence of a G at the first position of the downstream codon reduces the +1 frameshift by stabilizing the G34•U3 wobble interaction. This result partially explains why G34-containing tRNAGly exists in Eukaryotes whereas all the other G34-containing tRNAs for multiple codon boxes are absent.}, keywords = {ERIANI, IRES element frameshifting genetic code glycine codon tRNA translation, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A variant in MRPS14 (uS14m) causes perinatal hypertrophic cardiomyopathy with neonatal lactic acidosis, growth retardation, dysmorphic features and neurological involvement}, author = {C B Jackson and M Huemer and R Bolognini and F Martin and G Szinnai and B C Donner and U Richter and B J Battersby and J M Nuoffer and A Suomalainen and A Schaller}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30358850}, doi = {10.1093/hmg/ddy374}, isbn = {30358850}, year = {2019}, date = {2019-01-01}, journal = {Hum Mol Genet}, volume = {28}, number = {4}, pages = {639-649}, abstract = {Dysfunction of mitochondrial translation is increasingly an important molecular cause of human disease, but structural defects of mitochondrial ribosomal subunits are rare. We used next-generation sequencing to identify a homozygous variant in the mitochondrial small ribosomal protein 14 (MRPS14, uS14m) in a patient manifesting with perinatal hypertrophic cardiomyopathy, growth retardation, muscle hypotonia, elevated lactate, dysmorphy and mental retardation. In skeletal muscle and fibroblasts from the patient there was biochemical deficiency in complex IV of the respiratory chain. In fibroblasts mitochondrial translation was impaired and ectopic expression of a wild type MRPS14 cDNA functionally complemented this defect. Surprisingly, the mutant uS14m was stable and did not affect assembly of the small ribosomal subunit. Instead, structural modeling of the uS14m mutation predicted a disruption to the ribosomal mRNA channel. Collectively, our data demonstrates pathogenic mutations in MRPS14 can manifest as a perinatal-onset mitochondrial hypertrophic cardiomyopathy with a novel molecular pathogenic mechanism that impairs the function of mitochondrial ribosomes during translation elongation or mitochondrial mRNA recruitment rather than assembly.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Neutrophil Metabolic Shift during their Lifecycle: Impact on their Survival and Activation}, author = {L Injarabian and A Devin and S Ransac and B S Marteyn}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31906243?dopt=Abstract}, doi = {10.3390/ijms21010287}, isbn = {31906243}, year = {2019}, date = {2019-01-01}, journal = {Int J Mol Sci}, volume = {21}, number = {1}, pages = {287}, abstract = {Polymorphonuclear neutrophils (PMNs) are innate immune cells, which represent 50% to 70% of the total circulating leukocytes. How PMNs adapt to various microenvironments encountered during their life cycle, from the bone marrow, to the blood plasma fraction, and to inflamed or infected tissues remains largely unexplored. Metabolic shifts have been reported in other immune cells such as macrophages or lymphocytes, in response to local changes in their microenvironment, and in association with a modulation of their pro-inflammatory or anti-inflammatory functions. The potential contribution of metabolic shifts in the modulation of neutrophil activation or survival is anticipated even though it is not yet fully described. If neutrophils are considered to be mainly glycolytic, the relative importance of alternative metabolic pathways, such as the pentose phosphate pathway, glutaminolysis, or the mitochondrial oxidative metabolism, has not been fully considered during activation. This statement may be explained by the lack of knowledge regarding the local availability of key metabolites such as glucose, glutamine, and substrates, such as oxygen from the bone marrow to inflamed tissues. As highlighted in this review, the link between specific metabolic pathways and neutrophil activation has been outlined in many reports. However, the impact of neutrophil activation on metabolic shifts' induction has not yet been explored. Beyond its importance in neutrophil survival capacity in response to available metabolites, metabolic shifts may also contribute to neutrophil population heterogeneity reported in cancer (tumor-associated neutrophil) or auto-immune diseases (Low/High Density Neutrophils). This represents an active field of research. In conclusion, the characterization of neutrophil metabolic shifts is an emerging field that may provide important knowledge on neutrophil physiology and activation modulation. The related question of microenvironmental changes occurring during inflammation, to which neutrophils will respond to, will have to be addressed to fully appreciate the importance of neutrophil metabolic shifts in inflammatory diseases.}, keywords = {energetic metabolism infection inflammation neutrophils nutrient availability oxygen sensing, MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{reina_chemical_2019, title = {Chemical Functionalization of Nanodiamonds: Opportunities and Challenges Ahead}, author = {Giacomo Reina and Li Zhao and Alberto Bianco and Naoki Komatsu}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/anie.201905997}, doi = {10.1002/anie.201905997}, issn = {1521-3773}, year = {2019}, date = {2019-01-01}, urldate = {2020-04-01}, journal = {Angewandte Chemie International Edition}, volume = {58}, number = {50}, pages = {17918--17929}, abstract = {Nanodiamond(ND)-based technologies are flourishing in a wide variety of fields spanning from electronics and optics to biomedicine. NDs are considered a family of nanomaterials with an sp3 carbon core and a variety of sizes, shapes, and surfaces. They show interesting physicochemical properties such as hardness, stiffness, and chemical stability. Additionally, they can undergo ad-hoc core and surface functionalization, which tailors them for the desired applications. Noteworthy, the properties of NDs and their surface chemistry are highly dependent on the synthetic method used to prepare them. In this Minireview, we describe the preparation of NDs from the materials-chemistry viewpoint. The different methodologies of synthesis, purification, and surface functionalization as well as biomedical applications are critically discussed. New synthetic approaches as well as limits and obstacles of NDs are presented and analyzed.}, keywords = {carbon materials, diamond, Functionalization, I2CT, imaging, Team-Bianco, therapy}, pubstate = {published}, tppubtype = {article} } @article{, title = {Erratum for: Bioengineered Human Organ-on-Chip Reveals Intestinal Microenvironment and Mechanical Forces Impacting Shigella Infection}, author = {A Grassart and V Malardé and S Gobaa and A Sartori-Rupp and J Kerns and K Karalis and B Marteyn and P Sansonetti and N Sauvonnet}, url = {https://pubmed.ncbi.nlm.nih.gov/31600505-bioengineered-human-organ-on-chip-reveals-intestinal-microenvironment-and-mechanical-forces-impacting-shigella-infection/}, doi = {10.1016/j.chom.2019.09.007}, isbn = {31600505}, year = {2019}, date = {2019-01-01}, journal = {Cell Host Microbe}, volume = {26}, number = {4}, pages = {565}, keywords = {MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Bioengineered Human Organ-on-Chip Reveals Intestinal Microenvironment and Mechanical Forces Impacting Shigella Infection}, author = {A Grassart and V Malardé and S Gobaa and A Sartori-Rupp and J Kerns and K Karalis and B Marteyn and P Sansonetti and N Sauvonnet}, url = {https://pubmed.ncbi.nlm.nih.gov/31492657-bioengineered-human-organ-on-chip-reveals-intestinal-microenvironment-and-mechanical-forces-impacting-shigella-infection/?from_term=Grassart+marteyn&from_pos=1}, doi = {10.1016/j.chom.2019.08.007}, isbn = {31492657}, year = {2019}, date = {2019-01-01}, journal = {Cell Host Microbe}, volume = {26}, number = {3}, pages = {435-444.e434}, abstract = {Intestinal epithelial cells are constantly exposed to pathogens and mechanical forces. However, the impact of mechanical forces on infections leading to diarrheal diseases remains largely unknown. Here, we addressed whether flow and peristalsis impact the infectivity of the human pathogen Shigella within a 3D colonic epithelium using Intestine-Chip technology. Strikingly, infection is significantly increased and minimal bacterial loads are sufficient to invade enterocytes from the apical side and trigger loss of barrier integrity, thereby shifting the paradigm about early stage Shigella invasion. Shigella quickly colonizes epithelial crypt-like invaginations and demonstrates the essential role of the microenvironment. Furthermore, by modulating the mechanical forces of the microenvironment, we find that peristalsis impacts Shigella invasion. Collectively, our results reveal that Shigella leverages the intestinal microenvironment by taking advantage of the microarchitecture and mechanical forces to efficiently invade the intestine. This approach will enable molecular and mechanistic interrogation of human-restricted enteric pathogens.}, keywords = {Gut-on-Chip Intestine-Chip Organ-on-Chip enterocyte host-pathogen interactions intestine microengineering peristalsis shear stress stretching, MARTEYN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{arbogast_atg5_2019, title = {ATG5 is required for B cell polarization and presentation of particulate antigens.}, author = {Florent Arbogast and Johan Arnold and Philippe Hammann and Lauriane Kuhn and Johana Chicher and Diane Murera and Justine Weishaar and Sylviane Muller and Jean-Daniel Fauny and Frédéric Gros}, doi = {10.1080/15548627.2018.1516327}, issn = {1554-8635 1554-8627 1554-8627}, year = {2019}, date = {2019-01-01}, journal = {Autophagy}, volume = {15}, number = {2}, pages = {280--294}, abstract = {The involvement of macroautophagy/autophagy proteins in B-cell receptor (BCR) trafficking, although suspected, is not well understood. We show that ATG5 (autophagy related 5) contributes to BCR polarization after stimulation and internalization into LAMP1 (lysosomal-associated membrane protein 1)(+) and major histocompatibility complex class II (MHC-II)(+) compartments. BCR polarization is crucial in the context of immobilized antigen processing. Moreover, antigen presentation to cognate T cells is decreased in the absence of ATG5 when the model antigen OVAL/ovalbumin is provided in an immobilized form in contrast to the normal presentation of soluble OVAL. We further show that ATG5 is required for centrosome polarization and actin nucleation in the immune synapse area. This event is accompanied by an increased interaction between ATG16L1 (autophagy related 16-like 1 [S. cerevisiae]) and the microtubule-organizing center-associated protein PCM1 (pericentriolar material 1). In the human B cell line BJAB, PCM1 is required for BCR polarization after stimulation. We thus propose that the ATG12 (autophagy related 12)-ATG5-ATG16L1 complex under BCR stimulation allows its interaction with PCM1 and consequently facilitates centrosome relocalization to the immune synapse, optimizing the presentation of particulate antigens. Abbreviations: ACTB: actin beta; ACTR2/3: ARP2/3 actin-related protein 2/3; APC: antigen-presenting cells; ATG: autophagy-related; BCR: B cell receptor; BECN1/Beclin 1: beclin 1, autophagy related; CDC42: cell division cycle 42; Cr2: complement receptor 2; CSFE: carboxyfluorescein succinimidyl ester; DAPI: 4',6-diamidino-2-phenylindole dihydrochloride; EEA1: early endosome antigen 1; ELISA: enzyme-linked immunosorbent assay; FITC: fluorescein isothyocyanate; GC: germinal center; GJA1/CX3: gap junction protein, alpha 1; Ig: immunoglobulin; LAMP1: lysosomal-associated membrane protein 1; LAP: LC3-associated phagocytosis; LM: littermate; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MAPK/ERK: mitogen activated protein kinase; MHC-II: major histocompatibility complex class II; MIIC: MHC class II compartment; OVAL: ovalbumin; PBS: phosphate-buffered saline; PCM1: pericentriolar material 1; PtdIns3K: phosphatidylinositol 3-kinase; PTPRC/CD45RB/B220; Protein tyrosine phosphatase, receptor type, C; SYK: spleen tyrosine kinase; TBS: Tris-buffered saline; TCR: T cell receptor; ULK1: unc-51 like kinase 1.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {When a common biological role does not imply common disease outcomes: Disparate pathology linked to human mitochondrial aminoacyl-tRNA synthetases}, author = {L E González-Serrano and J W Chihade and M Sissler}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30647134?dopt=Abstract}, doi = {10.1074/jbc.REV118.002953}, isbn = {30647134}, year = {2019}, date = {2019-01-01}, journal = {J Biol Chem}, volume = {294}, number = {14}, pages = {5309-5320}, abstract = {Mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs) are essential components of the mitochondrial translation machinery. The correlation of mitochondrial disorders with mutations in these enzymes has raised the interest of the scientific community over the past several years. Most surprising has been the wide-ranging presentation of clinical manifestations in patients with mt-aaRS mutations, despite the enzymes' common biochemical role. Even among cases where a common physiological system is affected, phenotypes, severity, and age of onset varies depending on which mt-aaRS is mutated. Here we review work done thus far and propose a categorization of diseases based on tissue specificity that highlights emerging patterns. We further discuss multiple in vitro and in cellulo efforts to characterize the behavior of wildtype and mutant mt-aaRSs that have shaped hypotheses about the molecular causes of these pathologies. Much remains to do in order to complete our understanding of these proteins. We expect that futher work is likely to result in the discovery of new roles for the mt-aaRSs in addition to their fundamental function in mitochondrial translation, informing the development of treatment strategies and diagnoses.}, keywords = {aminoacyl tRNA synthetase central nervous system (CNS) genetic disease mitochondria mutant, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{waltz_small_2019, title = {Small is big in Arabidopsis mitochondrial ribosome.}, author = {Florent Waltz and Tan-Trung Nguyen and Mathilde Arrivé and Anthony Bochler and Johana Chicher and Philippe Hammann and Lauriane Kuhn and Martine Quadrado and Hakim Mireau and Yaser Hashem and Philippe Giegé}, doi = {10.1038/s41477-018-0339-y}, issn = {2055-0278 2055-0278}, year = {2019}, date = {2019-01-01}, journal = {Nature plants}, volume = {5}, number = {1}, pages = {106--117}, abstract = {Mitochondria are responsible for energy production through aerobic respiration, and represent the powerhouse of eukaryotic cells. Their metabolism and gene expression processes combine bacterial-like features and traits that evolved in eukaryotes. Among mitochondrial gene expression processes, translation remains the most elusive. In plants, while numerous pentatricopeptide repeat (PPR) proteins are involved in all steps of gene expression, their function in mitochondrial translation remains unclear. Here we present the biochemical characterization of Arabidopsis mitochondrial ribosomes and identify their protein subunit composition. Complementary biochemical approaches identified 19 plant-specific mitoribosome proteins, of which ten are PPR proteins. The knockout mutations of ribosomal PPR (rPPR) genes result in distinct macroscopic phenotypes, including lethality and severe growth delay. The molecular analysis of rppr1 mutants using ribosome profiling, as well as the analysis of mitochondrial protein levels, demonstrate rPPR1 to be a generic translation factor that is a novel function for PPR proteins. Finally, single-particle cryo-electron microscopy (cryo-EM) reveals the unique structural architecture of Arabidopsis mitoribosomes, characterized by a very large small ribosomal subunit, larger than the large subunit, bearing an additional RNA domain grafted onto the head. Overall, our results show that Arabidopsis mitoribosomes are substantially divergent from bacterial and other eukaryote mitoribosomes, in terms of both structure and protein content.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {History of tRNA research in Strasbourg}, author = {C Florentz and R Giege}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31185141?dopt=Abstract}, doi = {10.1002/iub.2079}, isbn = {31185141}, year = {2019}, date = {2019-01-01}, journal = {IUBMB Life}, volume = {71}, number = {8}, pages = {1066-1087}, abstract = {The tRNA molecules, in addition to translating the genetic code into protein and defining the second genetic code via their aminoacylation by aminoacyl-tRNA synthetases, act in many other cellular functions and dysfunctions. This article, illustrated by personal souvenirs, covers the history of ~60 years tRNA research in Strasbourg. Typical examples point up how the work in Strasbourg was a two-way street, influenced by and at the same time influencing investigators outside of France. All along, research in Strasbourg has nurtured the structural and functional diversity of tRNA. It produced massive sequence and crystallographic data on tRNA and its partners, thereby leading to a deeper physicochemical understanding of tRNA architecture, dynamics, and identity. Moreover, it emphasized the role of nucleoside modifications and in the last two decades, highlighted tRNA idiosyncrasies in plants and organelles, together with cellular and health-focused aspects. The tRNA field benefited from a rich local academic heritage and a strong support by both university and CNRS. Its broad interlinks to the worldwide community of tRNA researchers opens to an exciting future.}, keywords = {FLORENTZ, GIEGE, Strasbourg epistemology evolution genetic code history structural biology transfer RNA translation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @book{, title = {Microcalorimetry of Biological Molecules: Methods and Protocols}, editor = {E Ennifar}, year = {2019}, date = {2019-01-01}, volume = {1964}, publisher = {Springer Protocols, Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {This volume provides methods on microcalorimetry approaches to investigate complex biological molecular systems. Chapters guide readers through Differential Scanning Calorimetry (DSC), Isothermal Titration Calorimetry (ITC), and advanced data processing. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Microcalorimetry of Biological Molecules: Methods and Protocols aims to ensure successful results in the further study of this vital field.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {book} } @article{, title = {Noncoding RNA}, author = {E Desgranges and S Marzi and K Moreau and P Romby and I Caldelari}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31004423?dopt=Abstract}, doi = {10.1128/microbiolspec.GPP3-0038-2018}, isbn = {31004423}, year = {2019}, date = {2019-01-01}, journal = {Microbiol Spectr}, volume = {7}, number = {2}, pages = {1-2}, abstract = {Regulatory RNAs, present in many bacterial genomes and particularly in pathogenic bacteria such as Staphylococcus aureus, control the expression of genes encoding virulence factors or metabolic proteins. They are extremely diverse and include noncoding RNAs (sRNA), antisense RNAs, and some 5' or 3' untranslated regions of messenger RNAs that act as sensors for metabolites, tRNAs, or environmental conditions (e.g., temperature, pH). In this review we focus on specific examples of sRNAs of S. aureus that illustrate how numerous sRNAs and associated proteins are embedded in complex networks of regulation. In addition, we discuss the CRISPR-Cas systems defined as an RNA-interference-like mechanism, which also exist in staphylococcal strains.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[RsaI, a multifaceted regulatory RNA, modulates the metabolism of the opportunistic pathogen Staphylococcus aureus]}, author = {E Desgranges and D Bronesky and A Corvaglia and P François and C Caballero and L Prado and A Toledo-Arana and I Lasa and K Moreau and F Vandenesch and S Marzi and P Romby and I Caldelari}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31903946?dopt=Abstract}, doi = {10.1051/medsci/2019235}, isbn = {31903946}, year = {2019}, date = {2019-01-01}, journal = {Med Sci (Paris)}, volume = {35}, number = {12}, pages = {1221-1223}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{trinquier_trna_2019, title = {tRNA Maturation Defects Lead to Inhibition of rRNA Processing via Synthesis of pppGpp.}, author = {Aude Trinquier and Jonathan E Ulmer and Laetitia Gilet and Sabine Figaro and Philippe Hammann and Lauriane Kuhn and Frédérique Braun and Ciarán Condon}, doi = {10.1016/j.molcel.2019.03.030}, issn = {1097-4164 1097-2765}, year = {2019}, date = {2019-01-01}, journal = {Molecular cell}, volume = {74}, number = {6}, pages = {1227--1238.e3}, abstract = {rRNAs and tRNAs universally require processing from longer primary transcripts to become functional for translation. Here, we describe an unsuspected link between tRNA maturation and the 3' processing of 16S rRNA, a key step in preparing the small ribosomal subunit for interaction with the Shine-Dalgarno sequence in prokaryotic translation initiation. We show that an accumulation of either 5' or 3' immature tRNAs triggers RelA-dependent production of the stringent response alarmone (p)ppGpp in the Gram-positive model organism Bacillus subtilis. The accumulation of (p)ppGpp and accompanying decrease in GTP levels specifically inhibit 16S rRNA 3' maturation. We suggest that cells can exploit this mechanism to sense potential slowdowns in tRNA maturation and adjust rRNA processing accordingly to maintain the appropriate functional balance between these two major components of the translation apparatus.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {A simple and versatile microfluidic device for efficient biomacromolecule crystallization and structural analysis by serial crystallography}, author = {R de Wijn and O Hennig and J Roche and S Engilberge and K Rollet and P Fernandez-Millan and K Brillet and H Betat and M Mörl and A Roussel and E Girard and C Mueller-Dieckmann and G C Fox and V Olieric and J A Gavira and B Lorber and C Sauter}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31098026?dopt=Abstract}, doi = {10.1107/S2052252519003622}, isbn = {31098026}, year = {2019}, date = {2019-01-01}, journal = {IUCrJ}, volume = {6}, number = {Pt 3}, pages = {454-464}, abstract = {Determining optimal conditions for the production of well diffracting crystals is a key step in every biocrystallography project. Here, a microfluidic device is described that enables the production of crystals by counter-diffusion and their direct on-chip analysis by serial crystallography at room temperature. Nine 'non-model' and diverse biomacromolecules, including seven soluble proteins, a membrane protein and an RNA duplex, were crystallized and treated on-chip with a variety of standard techniques including micro-seeding, crystal soaking with ligands and crystal detection by fluorescence. Furthermore, the crystal structures of four proteins and an RNA were determined based on serial data collected on four synchrotron beamlines, demonstrating the general applicability of this multipurpose chip concept.}, keywords = {ChipX3 counter-diffusion crystallization ligand soaking macromolecule microfluidics protein structure room temperature seeding serial crystallography trace fluorescent labeling, ENNIFAR, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The yeast C/D box snoRNA U14 adopts a }, author = {M E Chagot and M Quinternet and B Rothé and B Charpentier and J Coutant and X Manival and I Lebars}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30914254?dopt=Abstract}, doi = {10.1016/j.biochi.2019.03.014}, isbn = {30914254}, year = {2019}, date = {2019-01-01}, journal = {Biochimie}, volume = {164}, pages = {70-82}, abstract = {Non-coding RNAs associate with proteins to form ribonucleoproteins (RNPs), such as ribosome, box C/D snoRNPs, H/ACA snoRNPs, ribonuclease P, telomerase and spliceosome to ensure cell viability. The assembly of these RNA-protein complexes relies on the ability of the RNA to adopt the correct bound conformation. K-turn motifs represent ubiquitous binding platform for proteins found in several cellular environment. This structural motif has an internal three-nucleotide bulge flanked on its 3' side by a G●A/A●G tandem pairs followed by one or two non-Watson-Crick pairs, and on its 5' side by a classical RNA helix. This peculiar arrangement induces a strong curvature of the phosphodiester backbone, which makes it conducive to multiple tertiary interactions. SNU13/Snu13p (Human/Yeast) binds specifically the U14 C/D box snoRNA K-turn sequence motif. This event is the prerequisite to promote the assembly of the RNP, which contains NOP58/Nop58 and NOP56/Nop56 core proteins and the 2'-O-methyl-transferase, Fibrillarin/Nop1p. The U14 small nucleolar RNA is a conserved non-coding RNA found in yeast and vertebrates required for the pre-rRNA maturation and ribose methylation. Here, we report the solution structure of the free U14 snoRNA K-turn motif (kt-U14) as determined by Nuclear Magnetic Resonance. We demonstrate that a major fraction of free kt-U14 adopts a pre-folded conformation similar to protein bound K-turn, even in the absence of divalent ions. In contrast to the kt-U4 or tyrS RNA, kt-U14 displays a sharp bent in the phosphodiester backbone. The U●U and G●A tandem base pairs are formed through weak hydrogen bonds. Finally, we show that the structure of kt-U14 is stabilized upon Snu13p binding. The structure of the free U14 RNA is the first reference example for the canonical motifs of the C/D box snoRNA family.}, keywords = {ENNIFAR, K-turn NMR Snu13 protein snoRNA U14, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A multifaceted small RNA modulates gene expression upon glucose limitation in \textit{Staphylococcus aureus}}, author = {D Bronesky and E Desgranges and A Corvaglia and P François and C J Caballero and L Prado and A Toledo-Arana and I Lasa and K Moreau and F Vandenesch and S Marzi and P Romby and I Caldelari}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30760492}, doi = {10.15252/embj.201899363}, isbn = {30760492}, year = {2019}, date = {2019-01-01}, journal = {EMBO J}, volume = {38}, number = {6}, pages = {e99363}, abstract = {Pathogenic bacteria must rapidly adapt to ever-changing environmental signals resulting in metabolism remodeling. The carbon catabolite repression, mediated by the catabolite control protein A (CcpA), is used to express genes involved in utilization and metabolism of the preferred carbon source. Here, we have identified RsaI as a CcpA-repressed small non-coding RNA that is inhibited by high glucose concentrations. When glucose is consumed, RsaI represses translation initiation of mRNAs encoding a permease of glucose uptake and the FN3K enzyme that protects proteins against damage caused by high glucose concentrations. RsaI also binds to the 3' untranslated region of icaR mRNA encoding the transcriptional repressor of exopolysaccharide production and to sRNAs induced by the uptake of glucose-6 phosphate or nitric oxide. Furthermore, RsaI expression is accompanied by a decreased transcription of genes involved in carbon catabolism pathway and an activation of genes involved in energy production, fermentation, and nitric oxide detoxification. This multifaceted RNA can be considered as a metabolic signature when glucose becomes scarce and growth is arrested.}, keywords = {ROMBY, sRNA carbon metabolism catabolite control protein A pathogenic bacteria regulatory RNAs translational regulation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Optimization of fluorogenic RNA-based biosensors using droplet-based microfluidic ultrahigh-throughput screening}, author = {A Autour and F Bouhedda and R Cubi and M Ryckelynck}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30902664?dopt=Abstract}, doi = {10.1016/j.ymeth.2019.03.015}, isbn = {30902664}, year = {2019}, date = {2019-01-01}, journal = {Methods}, volume = {161}, pages = {46-53}, abstract = {Biosensors are biological molecules able to detect and report the presence of a target molecule by the emission of a signal. Nucleic acids are particularly appealing for the design of such molecule since their great structural plasticity makes them able to specifically interact with a wide range of ligands and their structure can rearrange upon recognition to trigger a reporting event. A biosensor is typically made of three main domains: a sensing domain that is connected to a reporting domain via a communication module in charge of transmitting the sensing event through the molecule. The communication module is therefore an instrumental element of the sensor. This module is usually empirically developed through a trial-and-error strategy with the testing of only a few combinations judged relevant by the experimenter. In this work, we introduce a novel method combining the use of droplet-based microfluidics and next generation sequencing. This method allows to functionally characterize up to a million of different sequences in a single set of experiments and, by doing so, to exhaustively test every possible sequence permutations of the communication module. Here, we demonstrate the efficiency of the approach by isolating a set of optimized RNA biosensors able to sense theophylline and to convert this recognition into fluorescence emission.}, keywords = {Aptasensors Fluorogenic biosensors High-throughput screening Light-up aptamer Next generation sequencing RNA, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mapping post-transcriptional modifications in Staphylococcus aureus tRNAs by nanoLC/MSMS}, author = {L Antoine and P Wolff and E Westhof and P Romby and S Marzi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/31295507?dopt=Abstract}, doi = {10.1016/j.biochi.2019.07.003}, isbn = {31295507}, year = {2019}, date = {2019-01-01}, journal = {Biochimie}, volume = {164}, pages = {60-69}, abstract = {RNA modifications are involved in numerous biological processes. These modifications are constitutive or modulated in response to adaptive processes and can impact RNA base pairing formation, protein recognition, RNA structure and stability. tRNAs are the most abundantly modified RNA molecules. Analysis of the roles of their modifications in response to stress, environmental changes, and infections caused by pathogens, has fueled new research areas. Nevertheless, the detection of modified nucleotides in RNAs is still a challenging task. We present here a reliable method to identify and localize tRNA modifications, which was applied to the human pathogenic bacteria, Staphyloccocus aureus. The method is based on a separation of tRNA species on a two-dimensional polyacrylamide gel electrophoresis followed by nano liquid chromatography-mass spectrometry. We provided a list of modifications mapped on 25 out of the 40 tRNA species (one isoacceptor for each amino acid). This method can be easily used to monitor the dynamics of tRNA modifications in S. aureus in response to stress adaptation and during infection of the host, a relatively unexplored field.}, keywords = {2D gel isolation Staphylococcus aureus nanoLC/MSMS post-transcriptional tRNA modifications, ARN-MS, ENNIFAR, ROMBY, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interaction of a Model Peptide on Gram Negative and Gram Positive Bacterial Sliding Clamps}, author = {C André and I Martiel and P Wolff and M Landolfo and B Lorber and C Silva da Veiga and A Dejaegere and P Dumas and G Guichard and V Olieric and J G Wagner and D Y Burnouf}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30912430?dopt=Abstract}, doi = {10.1021/acsinfecdis.9b00089}, isbn = {30912430}, year = {2019}, date = {2019-01-01}, urldate = {2019-01-01}, journal = {ACS Infect Dis}, volume = {5}, number = {6}, pages = {1022-1034}, abstract = {Bacterial sliding clamps control the access of DNA polymerases to the replication fork and are appealing targets for antibacterial drugs development. It is therefore essential to decipher the polymerase-clamp binding mode across various bacterial species. Here, two residues of the E. coli clamp binding pocket, EcS346 and EcM362, and their cognate residues in M. tuberculosis and B. subtilis clamps, were mutated. The effects of these mutations on the interaction of a model peptide with these variant clamps were evaluated by thermodynamic, molecular dynamics, X-rays crystallography and biochemical analyses. EcM362 and corresponding residues in Gram positive clamps occupy a strategic position where a mobile residue is essential for an efficient peptide interaction. EcS346 has a more subtle function that modulates the pocket folding dynamics, while the equivalent residue in B. subtilis is essential for polymerase activity and might therefore be a Gram positive specific molecular marker. Finally, the peptide binds through an induced-fit process to Gram negative and positive pockets but the complex stability varies according to a pocket specific network of interactions.}, keywords = {ARN-MS, ENNIFAR, FRUGIER, ITC ligand−target interaction new antibacterials development sliding clamp, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{camara_lymph_2019, title = {Lymph Node Mesenchymal and Endothelial Stromal Cells Cooperate via the RANK-RANKL Cytokine Axis to Shape the Sinusoidal Macrophage Niche}, author = {Abdouramane Camara and Olga G Cordeiro and Farouk Alloush and Janina Sponsel and Mélanie Chypre and Lucas Onder and Kenichi Asano and Masato Tanaka and Hideo Yagita and Burkhard Ludewig and Vincent Flacher and Christopher G Mueller}, doi = {10.1016/j.immuni.2019.05.008}, issn = {1097-4180}, year = {2019}, date = {2019-01-01}, journal = {Immunity}, volume = {50}, number = {6}, pages = {1467--1481.e6}, abstract = {Tissue-resident macrophages are receptive to specific signals concentrated in cellular niches that direct their cell differentiation and maintenance genetic programs. Here, we found that deficiency of the cytokine RANKL in lymphoid tissue organizers and marginal reticular stromal cells of lymph nodes resulted in the loss of the CD169+ sinusoidal macrophages (SMs) comprising the subcapsular and the medullary subtypes. Subcapsular SM differentiation was impaired in mice with targeted RANK deficiency in SMs. Temporally controlled RANK removal in lymphatic endothelial cells (LECs) revealed that lymphatic RANK activation during embryogenesis and shortly after birth was required for the differentiation of both SM subtypes. Moreover, RANK expression by LECs was necessary for SM restoration after inflammation-induced cell loss. Thus, cooperation between mesenchymal cells and LECs shapes a niche environment that supports SM differentiation and reconstitution after inflammation.}, keywords = {Activation, Animals, Biomarkers, Cell Differentiation, Cells, Cellular, Cellular Microenvironment, cytokine, Cytokines, deficiency, Differentiation, Endothelial Cells, ENDOTHELIAL-CELLS, environment, Expression, immune regulation, Immunology, Immunophenotyping, inflammation, LYMPH, LYMPH NODE, Lymph Nodes, lymphatic endothelial cells, Lymphoid Tissue, Macrophage, Macrophages, Mesenchymal Stem Cells, mesenchymal stromal cells, Mice, rank, RANK ligand, Receptor Activator of Nuclear Factor-kappa B, Regulation, Signal Transduction, Stromal Cells, Team-Mueller, transgenic}, pubstate = {published}, tppubtype = {article} } @article{rive_improved_2019, title = {Improved Biocompatibility of Amino-Functionalized Graphene Oxide in Caenorhabditis elegans}, author = {Corvin Rive and Giacomo Reina and Prerana Wagle and Emanuele Treossi and Vincenzo Palermo and Alberto Bianco and Lucia Gemma Delogu and Matthias Rieckher and Björn Schumacher}, doi = {10.1002/smll.201902699}, issn = {1613-6829}, year = {2019}, date = {2019-01-01}, journal = {Small (Weinheim an Der Bergstrasse, Germany)}, volume = {15}, number = {45}, pages = {e1902699}, abstract = {Graphene oxide (GO) holds high promise for diagnostic and therapeutic applications in nanomedicine but reportedly displays immunotoxicity, underlining the need for developing functionalized GO with improved biocompatibility. This study describes adverse effects of GO and amino-functionalized GO (GONH2 ) during Caenorhabditis elegans development and ageing upon acute or chronic exposure. Chronic GO treatment throughout the C. elegans development causes decreased fecundity and a reduction of animal size, while acute treatment does not lead to any measurable physiological decline. However, RNA-Sequencing data reveal that acute GO exposure induces innate immune gene expression. The p38 MAP kinase, PMK-1, which is a well-established master regulator of innate immunity, protects C. elegans from chronic GO toxicity, as pmk-1 mutants show reduced tissue-functionality and facultative vivipary. In a direct comparison, GONH2 exposure does not cause detrimental effects in the wild type or in pmk-1 mutants, and the innate immune response is considerably less pronounced. This work establishes enhanced biocompatibility of amino-functionalized GO in a whole-organism, emphasizing its potential as a biomedical nanomaterial.}, keywords = {amino-functionalized graphene oxide, Caenorhabditis elegans, graphene oxide, I2CT, innate immunity, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{lucherelli_straightforward_2019, title = {A Straightforward Approach to Multifunctional Graphene}, author = {Matteo Andrea Lucherelli and Jésus Raya and Konstantin F Edelthalhammer and Frank Hauke and Andreas Hirsch and Gonzalo Abellán and Alberto Bianco}, doi = {10.1002/chem.201903165}, issn = {1521-3765}, year = {2019}, date = {2019-01-01}, journal = {Chemistry (Weinheim an Der Bergstrasse, Germany)}, volume = {25}, number = {57}, pages = {13218--13223}, abstract = {Graphene has been covalently functionalized through a one-pot reductive pathway using graphite intercalation compounds (GICs), in particular KC8 , with three different orthogonally protected derivatives of 4-aminobenzylamine. This novel multifunctional platform exhibits excellent bulk functionalization homogeneity (Hbulk ) and degree of addition while preserving the chemical functionalities of the organic addends through different protecting groups, namely: tert-butyloxycarbonyl (Boc), benzyloxycarbonyl (Cbz) and phthalimide (Pht). We have employed (temperature-dependent) statistical Raman spectroscopy (SRS), X-ray photoelectron spectroscopy (XPS), magic angle spinning solid state 13 C NMR (MAS-NMR), and a characterization tool consisting of thermogravimetric analysis coupled with gas chromatography and mass spectrometry (TG-GC-MS) to unambiguously demonstrate the covalent binding and the chemical nature of the different molecular linkers. This work paves the way for the development of smart graphene-based materials of great interest in biomedicine or electronics, to name a few, and will serve as a guide in the design of new 2D multifunctional materials.}, keywords = {carbon materials, diazonium salts, Functionalization, Graphite, I2CT, orthogonal protection, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{rauti_graphene_2019, title = {Graphene Oxide Flakes Tune Excitatory Neurotransmission in Vivo by Targeting Hippocampal Synapses}, author = {Rossana Rauti and Manuela Medelin and Leon Newman and Sandra Vranic and Giacomo Reina and Alberto Bianco and Maurizio Prato and Kostas Kostarelos and Laura Ballerini}, doi = {10.1021/acs.nanolett.8b04903}, issn = {1530-6992}, year = {2019}, date = {2019-01-01}, journal = {Nano Letters}, volume = {19}, number = {5}, pages = {2858--2870}, abstract = {Synapses compute and transmit information to connect neural circuits and are at the basis of brain operations. Alterations in their function contribute to a vast range of neuropsychiatric and neurodegenerative disorders and synapse-based therapeutic intervention, such as selective inhibition of synaptic transmission, may significantly help against serious pathologies. Graphene is a two-dimensional nanomaterial largely exploited in multiple domains of science and technology, including biomedical applications. In hippocampal neurons in culture, small graphene oxide nanosheets (s-GO) selectively depress glutamatergic activity without altering cell viability. Glutamate is the main excitatory neurotransmitter in the central nervous system and growing evidence suggests its involvement in neuropsychiatric disorders. Here we demonstrate that s-GO directly targets the release of presynaptic vesicle. We propose that s-GO flakes reduce the availability of transmitter, via promoting its fast release and subsequent depletion, leading to a decline ofglutamatergic neurotransmission. We injected s-GO in the hippocampus in vivo, and 48 h after surgery ex vivo patch-clamp recordings from brain slices show a significant reduction in glutamatergic synaptic activity in respect to saline injections.}, keywords = {Animals, Excitatory Amino Acid Agents, glutamate, Glutamic Acid, graphene, Graphite, hippocampal network, Hippocampus, Humans, I2CT, Nanostructures, Neurodegenerative Diseases, Neurons, Newborn, Primary Cell Culture, quantum dots, Rats, synapses, Synaptic Transmission, Team-Bianco, Wistar}, pubstate = {published}, tppubtype = {article} } @article{ji_physically-triggered_2019, title = {Physically-triggered nanosystems based on two-dimensional materials for cancer theranostics}, author = {Ding-Kun Ji and Cécilia Ménard-Moyon and Alberto Bianco}, doi = {10.1016/j.addr.2018.08.010}, issn = {1872-8294}, year = {2019}, date = {2019-01-01}, journal = {Advanced Drug Delivery Reviews}, volume = {138}, pages = {211--232}, abstract = {There is an increasing demand to develop effective methods for treating malignant diseases to improve healthcare in our society. Stimuli-responsive nanosystems, which can respond to internal or external stimuli are promising in cancer therapy and diagnosis due to their functionality and versatility. As a newly emerging class of nanomaterials, two-dimensional (2D) nanomaterials have attracted huge interest in many different fields including biomedicine due to their unique physical and chemical properties. In the past decade, stimuli-responsive nanosystems based on 2D nanomaterials have been widely studied, showing promising applications in cancer therapy and diagnosis, including phototherapies, magnetic therapy, drug and gene delivery, and non-invasive imaging. Here, we will focus our attention on the state-of-the-art of physically-triggered nanosystems based on graphene and two-dimensional nanomaterials for cancer therapy and diagnosis. The physical triggers include light, temperature, magnetic and electric fields.}, keywords = {2D Materials, Animals, Diagnosis, graphene, Graphite, Humans, I2CT, Light, Magnetic Fields, Nanomaterials, Nanostructures, Neoplasms, Team-Bianco, Theragnosis, Theranostic Nanomedicine, therapy}, pubstate = {published}, tppubtype = {article} } @article{pmid32038533, title = {Current Status of Latency Reversing Agents Facing the Heterogeneity of HIV-1 Cellular and Tissue Reservoirs}, author = {Amina Ait-Ammar and Anna Kula and Gilles Darcis and Roxane Verdikt and Stephane De Wit and Virginie Gautier and Patrick W G Mallon and Alessandro Marcello and Olivier Rohr and Carine Van Lint}, doi = {10.3389/fmicb.2019.03060}, issn = {1664-302X}, year = {2019}, date = {2019-01-01}, urldate = {2019-01-01}, journal = {Front Microbiol}, volume = {10}, pages = {3060}, abstract = {One of the most explored therapeutic approaches aimed at eradicating HIV-1 reservoirs is the "shock and kill" strategy which is based on HIV-1 reactivation in latently-infected cells ("shock" phase) while maintaining antiretroviral therapy (ART) in order to prevent spreading of the infection by the neosynthesized virus. This kind of strategy allows for the "kill" phase, during which latently-infected cells die from viral cytopathic effects or from host cytolytic effector mechanisms following viral reactivation. Several latency reversing agents (LRAs) with distinct mechanistic classes have been characterized to reactivate HIV-1 viral gene expression. Some LRAs have been tested in terms of their potential to purge latent HIV-1 in clinical trials, showing that reversing HIV-1 latency is possible. However, LRAs alone have failed to reduce the size of the viral reservoirs. Together with the inability of the immune system to clear the LRA-activated reservoirs and the lack of specificity of these LRAs, the heterogeneity of the reservoirs largely contributes to the limited success of clinical trials using LRAs. Indeed, HIV-1 latency is established in numerous cell types that are characterized by distinct phenotypes and metabolic properties, and these are influenced by patient history. Hence, the silencing mechanisms of HIV-1 gene expression in these cellular and tissue reservoirs need to be better understood to rationally improve this cure strategy and hopefully reach clinical success.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid31709195, title = {Microglial Cells: The Main HIV-1 Reservoir in the Brain}, author = {Clementine Wallet and Marco De Rovere and Jeanne Van Assche and Fadoua Daouad and Stéphane De Wit and Virginie Gautier and Patrick W G Mallon and Alessandro Marcello and Carine Van Lint and Olivier Rohr and Christian Schwartz}, doi = {10.3389/fcimb.2019.00362}, issn = {2235-2988}, year = {2019}, date = {2019-01-01}, urldate = {2019-01-01}, journal = {Front Cell Infect Microbiol}, volume = {9}, pages = {362}, abstract = {Despite efficient combination of the antiretroviral therapy (cART), which significantly decreased mortality and morbidity of HIV-1 infection, a definitive HIV cure has not been achieved. Hidden HIV-1 in cellular and anatomic reservoirs is the major hurdle toward a functional cure. Microglial cells, the Central Nervous system (CNS) resident macrophages, are one of the major cellular reservoirs of latent HIV-1. These cells are believed to be involved in the emergence of drugs resistance and reseeding peripheral tissues. Moreover, these long-life reservoirs are also involved in the development of HIV-1-associated neurocognitive diseases (HAND). Clearing these infected cells from the brain is therefore crucial to achieve a cure. However, many characteristics of microglial cells and the CNS hinder the eradication of these brain reservoirs. Better understandings of the specific molecular mechanisms of HIV-1 latency in microglial cells should help to design new molecules and new strategies preventing HAND and achieving HIV cure. Moreover, new strategies are needed to circumvent the limitations associated to anatomical sanctuaries with barriers such as the blood brain barrier (BBB) that reduce the access of drugs.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{olmo_control_2018, title = {Control of dengue virus in the midgut of Aedes aegypti by ectopic expression of the dsRNA-binding protein Loqs2.}, author = {Roenick P Olmo and Alvaro G A Ferreira and Tatiane C Izidoro-Toledo and Eric R G R Aguiar and Isaque J S de Faria and Kátia P R de Souza and Kátia P Osório and Lauriane Kuhn and Philippe Hammann and Elisa G de Andrade and Yaovi Mathias Todjro and Marcele N Rocha and Thiago H J F Leite and Siad C G Amadou and Juliana N Armache and Simona Paro and Caroline D de Oliveira and Fabiano D Carvalho and Luciano A Moreira and Eric Marois and Jean-Luc Imler and João T Marques}, doi = {10.1038/s41564-018-0268-6}, issn = {2058-5276 2058-5276}, year = {2018}, date = {2018-12-01}, journal = {Nature microbiology}, volume = {3}, number = {12}, pages = {1385--1393}, abstract = {Dengue virus (DENV) is an arbovirus transmitted to humans by Aedes mosquitoes(1). In the insect vector, the small interfering RNA (siRNA) pathway is an important antiviral mechanism against DENV(2-5). However, it remains unclear when and where the siRNA pathway acts during the virus cycle. Here, we show that the siRNA pathway fails to efficiently silence DENV in the midgut of Aedes aegypti although it is essential to restrict systemic replication. Accumulation of DENV-derived siRNAs in the midgut reveals that impaired silencing results from a defect downstream of small RNA biogenesis. Notably, silencing triggered by endogenous and exogenous dsRNAs remained effective in the midgut where known components of the siRNA pathway, including the double-stranded RNA (dsRNA)-binding proteins Loquacious and r2d2, had normal expression levels. We identified an Aedes-specific paralogue of loquacious and r2d2, hereafter named loqs2, which is not expressed in the midgut. Loqs2 interacts with Loquacious and r2d2 and is required to control systemic replication of DENV and also Zika virus. Furthermore, ectopic expression of Loqs2 in the midgut of transgenic mosquitoes is sufficient to restrict DENV replication and dissemination. Together, our data reveal a mechanism of tissue-specific regulation of the mosquito siRNA pathway controlled by Loqs2.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{navet_maternal_2018, title = {Maternal RANKL Reduces the Osteopetrotic Phenotype of Null Mutant Mouse Pups}, author = {Benjamin Navet and Jorge William Vargas-Franco and Andrea Gama and Jérome Amiaud and Yongwon Choi and Hideo Yagita and Christopher G Mueller and Françoise Rédini and Dominique Heymann and Beatriz Castaneda and Frédéric Lézot}, doi = {10.3390/jcm7110426}, issn = {2077-0383}, year = {2018}, date = {2018-11-01}, journal = {Journal of Clinical Medicine}, volume = {7}, number = {11}, abstract = {RANKL signalization is implicated in the morphogenesis of various organs, including the skeleton. Mice invalidated for Rankl present an osteopetrotic phenotype that was less severe than anticipated, depending on RANKL's implication in morphogenesis. The hypothesis of an attenuated phenotype, as a result of compensation during gestation by RANKL of maternal origin, was thus brought into question. In order to answer this question, Rankl null mutant pups from null mutant parents were generated, and the phenotype analyzed. The results validated the presence of a more severe osteopetrotic phenotype in the second-generation null mutant with perinatal lethality. The experiments also confirmed that RANKL signalization plays a part in the morphogenesis of skeletal elements through its involvement in cell-to-cell communication, such as in control of osteoclast differentiation. To conclude, we have demonstrated that the phenotype associated with Rankl invalidation is attenuated through compensation by RANKL of maternal origin.}, keywords = {bone, mandible, Morphogenesis, OSTEOCLAST, RANKL, skeletal growth, Team-Mueller, Tooth}, pubstate = {published}, tppubtype = {article} } @article{reynard_mannoside_2018, title = {Mannoside Glycolipid Conjugates Display Antiviral Activity Against Ebola Virus}, author = {Olivier Reynard and Evelyne Schaeffer and Valentina A Volchkova and Andrea Cimarelli and Christopher G Mueller and Viktor E Volchkov}, doi = {10.1093/infdis/jiy464}, issn = {1537-6613}, year = {2018}, date = {2018-11-01}, journal = {The Journal of Infectious Diseases}, volume = {218}, number = {suppl_5}, pages = {S666--S671}, abstract = {The West African outbreak of Ebola virus (EBOV) infection during 2013-2016 highlighted the need for development of field-applicable therapeutic drugs for this infection. Here we report that mannoside glycolipid conjugates (MGCs) consisting of a trimannose head and a lipophilic chain assembled by a linker inhibit EBOV infection not only of human monocyte-derived dendritic cells and macrophages, but also of a number of susceptible cells. Analysis of the mode of action leads us to conclude that MGCs act directly on cells, notably by preventing virus endocytosis.}, keywords = {Animals, Antiviral Agents, Chlorocebus aethiops, Ebolavirus, Glycolipids, Humans, Mannosides, Team-Mueller, Vero Cells, Virus Internalization}, pubstate = {published}, tppubtype = {article} } @article{Olmo_2018, title = {Control of dengue virus in the midgut of Aedes aegypti by ectopic expression of the dsRNA-binding protein Loqs2 }, author = {RP Olmo and AGA Ferreira and TC Izidoro-Toledo and ERGR Aguiar and IJS de Faria and KPR de Souza and KP Osório and L Kuhn and P Hammann and EG de Andrade and YM Todjro and MN Rocha and THJF Leite and SCG Amadou and JN Armache and S Paro and CD de Oliveira and FD Carvalho and LA Moreira and E Marois and JL Imler and JT Marques}, url = {https://www.nature.com/articles/s41564-018-0268-6}, doi = {10.1038/s41564-018-0268-6}, year = {2018}, date = {2018-10-29}, journal = {Nature Microbiology}, volume = {3}, number = {12}, pages = {1385-1393}, abstract = {Dengue virus (DENV) is an arbovirus transmitted to humans by Aedes mosquitoes. In the insect vector, the small interfering RNA (siRNA) pathway is an important antiviral mechanism against DENV. However, it remains unclear when and where the siRNA pathway acts during the virus cycle. Here, we show that the siRNA pathway fails to efficiently silence DENV in the midgut of Aedes aegypti although it is essential to restrict systemic replication. Accumulation of DENV-derived siRNAs in the midgut reveals that impaired silencing results from a defect downstream of small RNA biogenesis. Notably, silencing triggered by endogenous and exogenous dsRNAs remained effective in the midgut where known components of the siRNA pathway, including the double-stranded RNA (dsRNA)-binding proteins Loquacious and r2d2, had normal expression levels. We identified an Aedes-specific paralogue of loquacious and r2d2, hereafter named loqs2, which is not expressed in the midgut. Loqs2 interacts with Loquacious and r2d2 and is required to control systemic replication of DENV and also Zika virus. Furthermore, ectopic expression of Loqs2 in the midgut of transgenic mosquitoes is sufficient to restrict DENV replication and dissemination. Together, our data reveal a mechanism of tissue-specific regulation of the mosquito siRNA pathway controlled by Loqs2. }, keywords = {Aedes aegypti, Dengue, imler, M3i, marois, Marques, Zika}, pubstate = {published}, tppubtype = {article} } @article{Petitjean2018, title = {To have and have not, RNA interference as an antiviral defense system in mammals }, author = {Olivier Petitjean, Thomas Montavon and Sébastien Pfeffer}, url = {https://pubmed.ncbi.nlm.nih.gov/33111686/}, doi = { 10.1684/vir.2018.0748 }, issn = {33111686 }, year = {2018}, date = {2018-10-01}, journal = {Virologie (Montrouge)}, volume = {22}, number = {5}, pages = {251-260}, abstract = {RNA silencing is a small RNA based mechanism regulating gene expression and involved in many biological processes in most eukaryotes. In plants, nematodes and arthropods, this mechanism participates to antiviral defense. In mammals, although the RNA silencing machinery is present and needed for the microRNA pathway, its importance as an antiviral defense is still debated. In recent years, several studies have attempted to answer to the question of whether RNA silencing as an antiviral pathway is retained in mammals. However, these studies did not provide a clear answer yet. In this review, we will present the arguments for and against a relevant antiviral role of RNA interference (RNAi) in mammals, by discussing examples of active and functional mammalian antiviral RNAi in specific cell types and/or in specific conditions. }, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{chicois_upf1_2018, title = {The UPF1 interactome reveals interaction networks between RNA degradation and translation repression factors in Arabidopsis.}, author = {Clara Chicois and Hélène Scheer and Shahïnez Garcia and Hélène Zuber and Jérôme Mutterer and Johana Chicher and Philippe Hammann and Dominique Gagliardi and Damien Garcia}, doi = {10.1111/tpj.14022}, issn = {1365-313X 0960-7412}, year = {2018}, date = {2018-10-01}, journal = {The Plant journal : for cell and molecular biology}, volume = {96}, number = {1}, pages = {119--132}, abstract = {The RNA helicase UP-FRAMESHIFT (UPF1) is a key factor of nonsense-mediated decay (NMD), a mRNA decay pathway involved in RNA quality control and in the fine-tuning of gene expression. UPF1 recruits UPF2 and UPF3 to constitute the NMD core complex, which is conserved across eukaryotes. No other components of UPF1-containing ribonucleoproteins (RNPs) are known in plants, despite its key role in regulating gene expression. Here, we report the identification of a large set of proteins that co-purify with the Arabidopsis UPF1, either in an RNA-dependent or RNA-independent manner. We found that like UPF1, several of its co-purifying proteins have a dual localization in the cytosol and in P-bodies, which are dynamic structures formed by the condensation of translationally repressed mRNPs. Interestingly, more than half of the proteins of the UPF1 interactome also co-purify with DCP5, a conserved translation repressor also involved in P-body formation. We identified a terminal nucleotidyltransferase, ribonucleases and several RNA helicases among the most significantly enriched proteins co-purifying with both UPF1 and DCP5. Among these, RNA helicases are the homologs of DDX6/Dhh1, known as translation repressors in humans and yeast, respectively. Overall, this study reports a large set of proteins associated with the Arabidopsis UPF1 and DCP5, two components of P-bodies, and reveals an extensive interaction network between RNA degradation and translation repression factors. Using this resource, we identified five hitherto unknown components of P-bodies in plants, pointing out the value of this dataset for the identification of proteins potentially involved in translation repression and/or RNA degradation.}, note = {Place: England}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{navet_intrinsic_2018, title = {The Intrinsic and Extrinsic Implications of RANKL/RANK Signaling in Osteosarcoma: From Tumor Initiation to Lung Metastases}, author = {Benjamin Navet and Kosei Ando and Jorge William Vargas-Franco and Régis Brion and Jérome Amiaud and Kanji Mori and Hideo Yagita and Christopher G Mueller and Franck Verrecchia and Clotilde Dumars and Marie-Françoise Heymann and Dominique Heymann and Frédéric Lézot}, doi = {10.3390/cancers10110398}, issn = {2072-6694}, year = {2018}, date = {2018-10-01}, journal = {Cancers}, volume = {10}, number = {11}, abstract = {Background: Osteosarcoma is the most frequent form of malignant pediatric bone tumor. Despite the current therapeutic arsenal, patient life-expectancy remains low if metastases are detected at the time of diagnosis, justifying research into better knowledge at all stages of osteosarcoma ontogenesis and identification of new therapeutic targets. Receptor Activator of Nuclear factor κB (RANK)expression has been reported in osteosarcoma cells, raising the question of Receptor Activator of Nuclear factor κB Ligand (RANKL)/RANK signaling implications in these tumor cells (intrinsic), in addition to previously reported implications through osteoclast activation in the tumor microenvironment (extrinsic). Methods: Based on in vitro and in vivo experimentations using human and mouse osteosarcoma cell lines, the consequences on the main cellular processes of RANK expression in osteosarcoma cells were analyzed. Results: The results revealed that RANK expression had no impact on cell proliferation and tumor growth, but stimulated cellular differentiation and, in an immune-compromised environment, increased the number of lung metastases. The analysis of RANKL, RANK and osteoprotegerin (OPG) expressions in biopsies of a cohort of patients revealed that while RANK expression in osteosarcoma cells was not significantly different between patients with or without metastases at the time of diagnosis, the OPG/RANK ratio decreased significantly. Conclusion: Altogether, these results are in favor of RANKL-RANK signaling inhibition as an adjuvant for the treatment of osteosarcoma.}, keywords = {bone, metastases, osteosarcoma, RANKL/RANK, T-Lymphocyte, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @book{Gros2018, title = {Atlas d'immunologie}, author = {Frédéric Gros and Sylvie Fournel and Samuel Liégeois and Daniel Richard and Pauline Soulas-Sprauel}, editor = {Editions Dunod }, isbn = {9782100767052}, year = {2018}, date = {2018-09-01}, edition = {Editions Dunod}, abstract = {Cet ouvrage présente de façon synthétique et illustrée, les nombreux mécanismes de la réponse immunitaire et les différents types de cellules intervenant dans le système immunitaire. Les notions de base en immunologie, les modes d'explorations de l'immunité et les applications médicales de l'immunologie et les pathologies sont présentés sous forme de fiches illustrées par de nombreux schémas et photos en couleur.}, keywords = {atlas, cours, ferrandon, immunologie, M3i}, pubstate = {published}, tppubtype = {book} } @article{Goto2018, title = {The Kinase IKKβ Regulates a STING- and NF-κB-Dependent Antiviral Response Pathway in Drosophila}, author = {Akira Goto and Kiyoshi Okado and Nelson Martins and Hua Cai and Vincent Barbier and Olivier Lamiable and Laurent Troxler and Estelle Santiago and Lauriane Kuhn and Donggi Paik and Neal Silverman and Andreas Holleufer and Rune Hartmann and Jiyong Liu and Tao Peng and Jules A Hoffmann and Carine Meignin and Laurent Daeffler and Jean-Luc Imler}, editor = {Elsevier Inc.}, url = {https://doi.org/10.1016/j.immuni.2018.07.013}, doi = {j.immuni.2018.07.013}, year = {2018}, date = {2018-08-21}, journal = {Immunity}, number = {49}, pages = {225-234}, abstract = {Antiviral immunity in Drosophila involves RNA interference and poorly characterized inducible responses. Here, we showed that two components of the IMD pathway, the kinase dIKKβ and the transcription factor Relish, were required to control infection by two picorna-like viruses. We identified a set of genes induced by viral infection and regulated by dIKKβ and Relish, which included an ortholog of STING. We showed that dSTING participated in the control of infection by picorna-like viruses, acting upstream of dIKKβ to regulate expression of Nazo, an antiviral factor. Our data reveal an antiviral function for STING in an animal model devoid of interferons and suggest an evolutionarily ancient role for this molecule in antiviral immunity.}, keywords = {hoffmann, imler, M3i, meignin, PPSE}, pubstate = {published}, tppubtype = {article} } @inbook{Bouyer2018, title = {Pests and vector-borne diseases in the livestock industry}, author = {Bouyer J. and Marois E.}, editor = {Wageningen Academic}, url = {https://doi.org/10.3920/978-90-8686-863-6_14 }, doi = {10.3920/978-90-8686-863-6_14}, isbn = {9789086868636 }, year = {2018}, date = {2018-08-15}, volume = {5}, pages = {435–451}, chapter = {Genetic control of vectors}, series = {Ecology and Control of Vector-borne Diseases}, abstract = {In a context of tighter regulations on approved insecticide molecules, the spread of insecticide resistance in insect vectors of human and animal diseases and the introduction of exotic vectors to new territories call for the development of new pest control methods and strategies. New genetic control methods, related to the ancestral sterile insect technique (SIT), show particular promise and are being developed in response to increasing health and agricultural challenges. These include the use of symbionts like Wolbachia and the use of transgenic insect strains, some of which incorporate gene editing techniques that can lead to transgene spread (gene drive). Here we present the principles, associated opportunities and risks, as well as the degree of advancement of these various techniques for a subset of livestock pests and disease vectors including screwworms, tsetse, mosquitoes and stomoxes. We then present some case studies on recent improvements in the use of the SIT in tsetse and the release of insects carrying a dominant lethal gene, symbiont-based approaches and gene drive in mosquitoes. Finally, we call to speed up the development of genetic control, within a rigorous benefit-risk analysis framework including international public consultation.}, keywords = {insect resistance, M3i, marois}, pubstate = {published}, tppubtype = {inbook} } @article{bianco_carbon_2018, title = {A carbon science perspective in 2018: Current achievements and future challenges}, author = {Alberto Bianco and Yongsheng Chen and Yuan Chen and Debjit Ghoshal and Robert H Hurt and Yoong Ahm Kim and Nikhil Koratkar and Vincent Meunier and Mauricio Terrones}, url = {http://www.sciencedirect.com/science/article/pii/S0008622318301829}, doi = {10.1016/j.carbon.2018.02.058}, issn = {0008-6223}, year = {2018}, date = {2018-06-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {132}, pages = {785--801}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{vacchi_controlled_2018, title = {Controlled derivatization of hydroxyl groups of graphene oxide in mild conditions}, author = {Isabella A Vacchi and Jésus Raya and Alberto Bianco and Cécilia Ménard-Moyon}, url = {https://doi.org/10.1088%2F2053-1583%2Faac8a9}, doi = {10.1088/2053-1583/aac8a9}, issn = {2053-1583}, year = {2018}, date = {2018-06-01}, urldate = {2020-04-01}, journal = {2D Materials}, volume = {5}, number = {3}, pages = {035037}, abstract = {Graphene oxide (GO) is constituted of various oxygen-containing functionalities, primarily epoxides and hydroxyl groups on the basal plane, with a very low amount of carbonyl, quinone, carboxylic acid, phenol, and lactone functions at the edges. The high chemical reactivity of these oxygenated groups makes functionalization difficult to control as different reactions can occur concomitantly. In this study we have investigated the reactivity of GO towards orthogonal reactions to selectively functionalize the hydroxyl groups, which are present in a high amount. We explored both the esterification and the Williamson reaction. Our strategies present the main advantage to occur in mild conditions, thus preserving the intrinsic properties of GO, whereas most reactions reported in literature require relatively harsh conditions, leading to (partial) reduction, and/or are not chemoselective. We have also extended our study to the ketones and examined their derivatization by the Wittig reaction. This work has allowed developing two facile methods for the covalent derivatization of the hydroxyl groups in mild conditions, while GO was not reactive towards the Wittig reaction, probably due to the low amount of ketones. Overall, this work leads to a better understanding of the reactivity of GO for controlled derivatization. This opens promising perspectives for multi-functionalization of GO in order to design graphene-based nanomaterials endowed of multiple properties.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{S2018, title = {Quorum-sensing regulator RhlR but not its autoinducer RhlI enables Pseudomonas to evade opsonization}, author = {Samantha Haller and Adrien Franchet and A Hakkim and J Chen and E Drenkard and S Yu and Steffi Schirmeier and Zi Li and Nelson Martins and FM Ausubel and Samuel Liégeois and Dominique Ferrandon}, url = {http://embor.embopress.org/content/early/2018/03/09/embr.201744880}, doi = {10.15252/embr.201744880}, year = {2018}, date = {2018-03-09}, journal = {EMBO Reports}, keywords = {Drosophila, ferrandon, M3i, Opsonin Proteins, Phagocytosis, Pseudomonas aeruginosa, Quorum Sensing}, pubstate = {published}, tppubtype = {article} } @article{Dong2018, title = {CRISPR/Cas9 -mediated gene knockout of Anopheles gambiae FREP1 suppresses malaria parasite infection}, author = { Yuemei Dong and Maria L. Simões and Eric Marois and George Dimopoulos }, url = {https://doi.org/10.1371/journal.ppat.1006898}, doi = {10.1371/journal.ppat.1006898}, year = {2018}, date = {2018-03-08}, urldate = {2018-03-08}, journal = {PLoS Pathog}, volume = {14}, number = {3}, abstract = {Plasmodium relies on numerous agonists during its journey through the mosquito vector, and these agonists represent potent targets for transmission-blocking by either inhibiting or interfering with them pre- or post-transcriptionally. The recently developed CRISPR/Cas9-based genome editing tools for Anopheles mosquitoes provide new and promising opportunities for the study of agonist function and for developing malaria control strategies through gene deletion to achieve complete agonist inactivation. Here we have established a modified CRISPR/Cas9 gene editing procedure for the malaria vector Anopheles gambiae, and studied the effect of inactivating the fibrinogen-related protein 1 (FREP1) gene on the mosquito’s susceptibility to Plasmodium and on mosquito fitness. FREP1 knockout mutants developed into adult mosquitoes that showed profound suppression of infection with both human and rodent malaria parasites at the oocyst and sporozoite stages. FREP1 inactivation, however, resulted in fitness costs including a significantly lower blood-feeding propensity, fecundity and egg hatching rate, a retarded pupation time, and reduced longevity after a blood meal.}, keywords = {Anopheles gambiae, CRISPR/Cas9, Knockout, M3i, Malaria, marois}, pubstate = {published}, tppubtype = {article} } @article{Issa2018, title = {The circulating protease Persephone is an immune sensor for microbial proteolytic activities upstream of the Drosophila Toll pathway.}, author = {Najwa Issa and Nina Guillaumot and Emilie Lauret and Nicolas Matt and Christine Scaeffer-Reiss and Alain Van Dorsselaer and Jean-Marc Reichhart and Florian Veillard}, url = {http://www.cell.com/molecular-cell/fulltext/S1097-2765(18)30058-3?_returnURL=https%3A%2F%2Flinkinghub.elsevier.com%2Fretrieve%2Fpii%2FS1097276518300583%3Fshowall%3Dtrue}, doi = {10.1016/j.molcel.2018.01.029}, issn = {1097-2765}, year = {2018}, date = {2018-02-15}, journal = {Molecular Cell}, volume = {69}, number = {4}, pages = {539-550}, abstract = {Microbial or endogenous molecular patterns as well as pathogen functional features can activate innate immune systems. Whereas detection of infection by pattern recognition receptors has been investigated in details, sensing of virulence factors activities remains less characterized. In Drosophila, genetic evidences indicate that the serine protease Persephone belongs to a danger pathway activated by abnormal proteolytic activities to induce Toll signaling. However, neither the activation mechanism of this pathway nor its specificity has been determined. Here, we identify a unique region in the pro-domain of Persephone that functions as bait for exogenous proteases independently of their origin, type, or specificity. Cleavage in this bait region constitutes the first step of a sequential activation and licenses the subsequent maturation of Persephone to the endogenous cysteine cathepsin 26-29-p. Our results establish Persephone itself as an immune receptor able to sense a broad range of microbes through virulence factor activities rather than molecular patterns.}, keywords = {circulating protease, immune sensor, M3i, matt, persephone, Protease, proteolitic activities, reichhart, toll pathway}, pubstate = {published}, tppubtype = {article} } @article{schaeffer_inhibition_2018, title = {Inhibition of dengue virus infection by mannoside glycolipid conjugates}, author = {Evelyne Schaeffer and Vincent Flacher and Patrick Neuberg and Astrid Hoste and Adrien Brulefert and Jean-Daniel Fauny and Alain Wagner and Christopher G Mueller}, doi = {10.1016/j.antiviral.2018.04.005}, issn = {1872-9096}, year = {2018}, date = {2018-01-01}, journal = {Antiviral Research}, volume = {154}, pages = {116--123}, abstract = {Dengue virus (DENV), a mosquito-borne flavivirus, causes severe and potentially fatal symptoms in millions of infected individuals each year. Although dengue fever represents a major global public health problem, the vaccines or antiviral drugs proposed so far have not shown sufficient efficacy and safety, calling for new antiviral developments. Here we have shown that a mannoside glycolipid conjugate (MGC) bearing a trimannose head with a saturated lipid chain inhibited DENV productive infection. It showed remarkable cell promiscuity, being active in human skin dendritic cells, hepatoma cell lines and Vero cells, and was active against all four DENV serotypes, with an IC50 in the low micromolar range. Time-of-addition experiments and structure-activity analyses revealed the importance of the lipid chain to interfere with an early viral infection step. This, together with a correlation between antiviral activity and membrane polarization by the lipid moiety indicated that the inhibitor functions by blocking viral envelope fusion with the endosome membrane. These finding establish MGCs as a novel class of antivirals against the DENV.}, keywords = {Cell Membrane, Dendritic Cells, Dengue virus, I2CT, Imagerie, inhibitors, Macrophages, Skin, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{sawaf_defective_2018, title = {Defective BTLA functionality is rescued by restoring lipid metabolism in lupus CD4+ Ŧ cells}, author = {Matthieu Sawaf and Jean-Daniel Fauny and Renaud Felten and Flora Sagez and Jacques-Eric Gottenberg and Hélène Dumortier and Fanny Monneaux}, doi = {10.1172/jci.insight.99711}, issn = {2379-3708}, year = {2018}, date = {2018-01-01}, journal = {JCI insight}, volume = {3}, number = {13}, abstract = {Coinhibitory receptors play an important role in the prevention of autoimmune diseases, such as systemic lupus erythematosus (SLE), by limiting T cell activation. B and T lymphocyte attenuator (BTLA) is an inhibitory receptor, similar to cytotoxic T lymphocyte-associated protein 4 (CTLA-4) and programmed death 1 (PD1), that negatively regulates the immune response. The role of BTLA in the pathogenesis of autoimmune diseases in humans and, more specifically, in SLE is largely unknown. We investigated BTLA expression on various T cell subsets, and we did not observe significant variations of BTLA expression between lupus patients and healthy controls. However, the enhancement of BTLA expression after activation was significantly lower in SLE patients compared with that in healthy controls. Furthermore, we found an impaired capacity of BTLA to inhibit T cell activation in SLE due to a poor BTLA recruitment to the immunological synapse following T cell stimulation. Finally, we demonstrated that defective BTLA function can be corrected by restoring intracellular trafficking and by normalizing the lipid metabolism in lupus CD4+ T cells. Collectively, our results evidence that the BTLA signaling pathway is altered in SLE T cells and highlight the potential of targeting this pathway for the development of new therapeutic strategies in lupus.}, keywords = {80 and over, Adolescent, Adult, Aged, Autoimmune Diseases, Autoimmunity, CD4-Positive T-Lymphocytes, Cell Proliferation, CTLA-4 Antigen, Dumortier, Female, France, Humans, I2CT, Imagerie, Immunologic, Immunology, Lipid Metabolism, lupus, Lupus Erythematosus, Lymphocyte Activation, Male, Middle Aged, Monneaux, Programmed Cell Death 1 Receptor, Receptors, Signal Transduction, Systemic, Team-Dumortier, Young Adult}, pubstate = {published}, tppubtype = {article} } @article{rodrigues_immunological_2018, title = {Immunological impact of graphene oxide sheets in the abdominal cavity is governed by surface reactivity}, author = {Artur Filipe Rodrigues and Leon Newman and Dhifaf A Jasim and Isabella A Vacchi and Cécilia Ménard-Moyon and Livia E Crica and Alberto Bianco and Kostas Kostarelos and Cyrill Bussy}, doi = {10.1007/s00204-018-2303-z}, issn = {1432-0738}, year = {2018}, date = {2018-01-01}, journal = {Archives of Toxicology}, volume = {92}, number = {11}, pages = {3359--3379}, abstract = {Graphene oxide (GO) is an oxidised form of graphene that has attracted commercial interest in multiple applications, including inks, printed electronics and spray coatings, which all raise health concerns due to potential creation of inhalable aerosols. Although a number of studies have discussed the toxicity of GO sheets, the in vivo impact of their lateral dimensions is still not clear. Here, we compared the effects of large GO sheets (l-GO, 1-20 µm) with those of small GO sheets (s-GO, textbackslashtextless 1 µm) in terms of mesothelial damage and peritoneal inflammation, after intraperitoneal (i.p.) injection in mice. To benchmark the outcomes, long and rigid multi-walled carbon nanotubes (MWCNTs) that were shown to be associated with asbestos-like pathogenicity on the mesothelium were also tested. Our aim was to assess whether lateral dimensions can be a predictor of inflammogenicity for GO sheets in a similar fashion as length is for MWCNTs. While long MWCNTs dispersed in 0.5% BSA induced a granulomatous response on the diaphragmatic mesothelium and immune cell recruitment to the peritoneal cavity, GO sheets dispersed under similar conditions did not cause any response, regardless of their lateral dimensions. We further interrogated whether tuning the surface reactivity of GO by testing different dispersions (5% dextrose instead of 0.5% BSA) may change the biological outcome. Although the change of dispersion did not alter the impact of GO on the mesothelium (i.e. no granuloma), we observed that, when dispersed in protein-free 5% dextrose solution, s-GO elicited a greater recruitment of monocytic cells to the peritoneal cavity than l-GO, or when dispersed in protein-containing solution. Such recruitment coincided with the greater ability of s-GO to interact in vivo with peritoneal macrophages and was associated with a greater surface reactivity in comparison to l-GO. In conclusion, large dimension was not a determining factor of the immunological impact of GO sheets after i.p. administration. For an equal dose, GO sheets with lateral dimensions similar to the length of long MWCNTs were less pathogenic than the MWCNTs. On the other hand, surface reactivity and the ability of some smaller GO sheets to interact more readily with immune cells seem to be key parameters that can be tuned to improve the safety profile of GO. In particular, the choice of dispersion modality, which affected these two parameters, was found to be of crucial importance in the assessment of GO impact in this model. Overall, these findings are essential for a better understanding of the parameters governing GO toxicity and inflammation, and the rational design of safe GO-based formulations for various applications, including biomedicine.}, keywords = {2D Materials, Animals, carbon, Epithelium, Female, graphene oxide, Graphite, I2CT, In vivo, Inbred C57BL, inflammation, Intraperitoneal, Macrophages, Mesothelium, Mice, Nanotubes, Peritoneal, Peritoneal Cavity, Protein coating, Team-Bianco, Tissue Distribution, Toxicity}, pubstate = {published}, tppubtype = {article} } @article{wang_blocking_2018, title = {Blocking nuclear export of HSPA8 after heat shock stress severely alters cell survival.}, author = {Fengjuan Wang and Srinivasa Reddy Bonam and Nicolas Schall and Lauriane Kuhn and Philippe Hammann and Olivier Chaloin and Jean-Baptiste Madinier and Jean-Paul Briand and Nicolas Page and Sylviane Muller}, doi = {10.1038/s41598-018-34887-6}, issn = {2045-2322 2045-2322}, year = {2018}, date = {2018-01-01}, journal = {Scientific reports}, volume = {8}, number = {1}, pages = {16820}, abstract = {The nuclear translocation of endogenous heat shock cognate protein HSPA8 is a requisite for cell survival during oxidative and heat shock stress. Upon these events, cytoplasmic HSPA8 is thought to concentrate within the nucleus and nucleolus. When the situation returns to normal, HSPA8 is released from its nuclear/nucleolar anchors and redistributes into the cytoplasm. By using different stress conditions and a 21-mer phosphopeptide tool called P140, which binds HSPA8 and hampers its chaperone properties, we deciphered the cellular and molecular effects arising during this vital cytoplasmic-nuclear-cytoplasmic shuttling process. Using the non-metastatic fibroblastoid cell line MRL/N-1 derived from a MRL/MpTn-gld/gld lupus-prone mouse, we discovered that P140 treatment neutralized the egress of HSPA8 from nucleus to cytoplasm in the cell recovery phase. This lack of relocation of HSPA8 into the cytoplasm of heat-shocked MRL/N-1 cells altered the ability of these cells to survive when a second mild oxidative stress mimicking inflammatory conditions was applied. Crosslinking experiments followed by proteomics studies showed that P140 binds regions close to nuclear import and export signal sequences encompassed within the HSPA8 structure. These data are consistent with HSPA8 having a crucial cell protective role against reactive oxygen species (ROS) production by mitochondria during inflammatory conditions.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {In cell mutational interference mapping experiment (in cell MIME) identifies the 5' polyadenylation signal as a dual regulator of HIV-1 genomic RNA production and packaging}, author = {R. P. Smyth and M. R. Smith and A. C. Jousset and L. Despons and G. Laumond and T. Decoville and P. Cattenoz and C. Moog and F. Jossinet and M. Mougel and J. C. Paillart and M. Kleist and R. Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=29514260}, doi = {10.1093/nar/gky152}, isbn = {29514260}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {9}, pages = {e57}, abstract = {Non-coding RNA regulatory elements are important for viral replication, making them promising targets for therapeutic intervention. However, regulatory RNA is challenging to detect and characterise using classical structure-function assays. Here, we present in cell Mutational Interference Mapping Experiment (in cell MIME) as a way to define RNA regulatory landscapes at single nucleotide resolution under native conditions. In cell MIME is based on (i) random mutation of an RNA target, (ii) expression of mutated RNA in cells, (iii) physical separation of RNA into functional and non-functional populations, and (iv) high-throughput sequencing to identify mutations affecting function. We used in cell MIME to define RNA elements within the 5' region of the HIV-1 genomic RNA (gRNA) that are important for viral replication in cells. We identified three distinct RNA motifs controlling intracellular gRNA production, and two distinct motifs required for gRNA packaging into virions. Our analysis reveals the 73AAUAAA78 polyadenylation motif within the 5' PolyA domain as a dual regulator of gRNA production and gRNA packaging, and demonstrates that a functional polyadenylation signal is required for viral packaging even though it negatively affects gRNA production.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {5' Untranslated Regions Genome, LESCURE, Ribonucleic Acid *Virus Assembly Virus Replication, Unité ARN, Viral HEK293 Cells HIV-1/*genetics/physiology Humans Mutation Nucleotide Motifs Poly A/metabolism RNA, Viral/*biosynthesis/*chemistry *Regulatory Sequences}, pubstate = {published}, tppubtype = {article} } @article{arosio_conjugation_2018, title = {Conjugation of a GM3 lactone mimetic on carbon nanotubes enhances the related inhibition of melanoma-associated metastatic events}, author = {Paolo Arosio and Giuseppina Comito and Francesco Orsini and Alessandro Lascialfari and Paola Chiarugi and Cécilia Ménard-Moyon and Cristina Nativi and Barbara Richichi}, doi = {10.1039/c8ob01817k}, issn = {1477-0539}, year = {2018}, date = {2018-01-01}, journal = {Organic & Biomolecular Chemistry}, volume = {16}, number = {33}, pages = {6086--6095}, abstract = {GM3-ganglioside is known to be involved in melanoma proliferation. In order to modulate metastatic-related events, we have functionalized multi-walled carbon nanotubes (MWCNTs) with multiple copies of a GM3-lactone mimetic. The MWCNTs proved to guarantee the appropriate spatial arrangement of the mimetic allowing a stronger inhibition of migration and invasiveness of human melanoma (A375) cells compared to other multivalent constructs reported before. In addition, the effect of the multivalent tubular conjugate on the inhibition of specific tyrosine kinases, which are associated with the ganglioside complexes within the membrane domains, was demonstrated. Finally, the short-term fate of the conjugate was assessed, for the first time, by means of the 1H NMR relaxometry technique by exploiting the signal arising from the CNTs.}, keywords = {Antineoplastic Agents, Biomimetic Materials, carbon, Cell Line, G(M3) Ganglioside, Humans, I2CT, Melanoma, Models, Molecular, Molecular Conformation, Nanotubes, Neoplasm Metastasis, Team-Bianco, tumor}, pubstate = {published}, tppubtype = {article} } @article{poirier_toxicological_2018, title = {Toxicological effects of CdSe nanocrystals on the marine diatom Phaeodactylum tricornutum: The first mass spectrometry-based proteomic approach.}, author = {Isabelle Poirier and Marie Pallud and Lauriane Kuhn and Philippe Hammann and Arnaud Demortière and Arash Jamali and Johana Chicher and Christelle Caplat and Régis Kevin Gallon and Martine Bertrand}, doi = {10.1016/j.ecoenv.2018.01.043}, issn = {1090-2414 0147-6513}, year = {2018}, date = {2018-01-01}, journal = {Ecotoxicology and environmental safety}, volume = {152}, pages = {78--90}, abstract = {In the marine environment, benthic diatoms from estuarine and coastal sediments are among the first targets of nanoparticle pollution whose potential toxicity on marine organisms is still largely unknown. It is therefore relevant to improve our knowledge of interactions between these new pollutants and microalgae, the key players in the control of marine resources. In this study, the response of P. tricornutum to CdSe nanocrystals (CdSe NPs) of 5 nm (NP5) and 12 nm (NP12) in diameter was evaluated through microscopic, physiological, biochemical and proteomic approaches. NP5 and NP12 affected cell growth but oxygen production was only slightly decreased by NP5 after 1-d incubation time. In our experimental conditions, a high CdSe NP dissolution was observed during the first day of culture, leading to Cd bioaccumulation and oxidative stress, particularly with NP12. However, after a 7-day incubation time, proteomic analysis highlighted that P. tricornutum responded to CdSe NP toxicity by regulating numerous proteins involved in protection against oxidative stress, cellular redox homeostasis, Ca(2+) regulation and signalling, S-nitrosylation and S-glutathionylation processes and cell damage repair. These proteome changes allowed algae cells to regulate their intracellular ROS level in contaminated cultures. P. tricornutum was also capable to control its intracellular Cd concentration at a sufficiently low level to preserve its growth. To our knowledge, this is the first work allowing the identification of proteins differentially expressed by P. tricornutum subjected to NPs and thus the understanding of some molecular pathways involved in its cellular response to nanoparticles. SIGNIFICANCE: The microalgae play a key role in the control of marine resources. Moreover, they produce 50% of the atmospheric oxygen. CdSe NPs are extensively used in the industry of renewable energies and it is regrettably expected that these pollutants will sometime soon appear in the marine environment through surface runoff, urban effluents and rivers. Since estuarine and coastal sediments concentrate pollutants, benthic microalgae which live in superficial sediments will be among the first targets of nanoparticle pollution. Thus, it is relevant to improve our knowledge of interactions between diatoms and nanoparticles. Proteomics is a powerful tool for understanding the molecular mechanisms triggered by nanoparticle exposure, and our study is the first one to use this tool to identify proteins differentially expressed by P. tricornutum subjected to CdSe nanocrystals. This work is fundamental to improve our knowledge about the defence mechanisms developed by algae cells to counteract damage caused by CdSe NPs.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal Structures of the Mango-II RNA Aptamer Reveal Heterogeneous Fluorophore Binding and Guide Engineering of Variants with Improved Selectivity and Brightness}, author = {3rd R J Trachman and A Abdolahzadeh and A Andreoni and R Cojocaru and J R Knutson and M Ryckelynck and P J Unrau and A R Ferré-D'Amaré}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29768001?dopt=Abstract}, doi = {10.1021/acs.biochem.8b00399}, isbn = {29768001}, year = {2018}, date = {2018-01-01}, journal = {Biochemistry}, volume = {57}, number = {26}, pages = {3544-3548}, abstract = {Several RNA aptamers that bind small molecules and enhance their fluorescence have been successfully used to tag and track RNAs in vivo, but these genetically encodable tags have not yet achieved single-fluorophore resolution. Recently, Mango-II, an RNA that binds TO1-Biotin with ~1 nM affinity and enhances its fluorescence by >1,500-fold was isolated by fluorescence selection from the pool that yielded the original RNA Mango. We determined the crystal structures of Mango-II in complex with two fluorophores, TO1-Biotin and TO3-Biotin, and found that despite their high affinity, the ligands adopt multiple distinct conformations, indicative of a binding pocket with modest stereoselectivity. Mutational analysis of the binding site led to Mango-II(A22U), which retains high affinity for TO1-Biotin but now discriminates over 5-fold against TO3-biotin. Moreover, fluorescence enhancement of TO1-Biotin increases 18% while that of TO3-Biotin decreases by 25%. Crystallographic, spectroscopic, and analog studies show that the A22U mutation improves conformational homogeneity and shape complementarity of the fluorophore-RNA interface. Our work demonstrates that even after extensive functional selection, aptamer RNAs can be further improved through structure-guided engineering.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dimerization confers increased stability to nucleases in 5' halves from glycine and glutamic acid tRNAs}, author = {J P Tosar and F Gámbaro and L Darré and S Pantano and E Westhof and A Cayota}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29893896?dopt=Abstract}, doi = {10.1093/nar/gky495}, isbn = {29893896}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {17}, pages = {9081-9093}, abstract = {We have previously shown that 5' halves from tRNAGlyGCC and tRNAGluCUC are the most enriched small RNAs in the extracellular space of human cell lines, and especially in the non-vesicular fraction. Extracellular RNAs are believed to require protection by either encapsulation in vesicles or ribonucleoprotein complex formation. However, deproteinization of non-vesicular tRNA halves does not affect their retention in size-exclusion chromatography. Thus, we considered alternative explanations for their extracellular stability. In-silico analysis of the sequence of these tRNA-derived fragments showed that tRNAGly 5' halves can form homodimers or heterodimers with tRNAGlu 5' halves. This capacity is virtually unique to glycine tRNAs. By analyzing synthetic oligonucleotides by size exclusion chromatography, we provide evidence that dimerization is possible in vitro. tRNA halves with single point substitutions preventing dimerization are degraded faster both in controlled nuclease digestion assays and after transfection in cells, showing that dimerization can stabilize tRNA halves against the action of cellular nucleases. Finally, we give evidence supporting dimerization of endogenous tRNAGlyGCC 5' halves inside cells. Considering recent reports have shown that 5' tRNA halves from Ala and Cys can form tetramers, our results highlight RNA intermolecular structures as a new layer of complexity in the biology of tRNA-derived fragments.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {CUG initiation and frameshifting enable production of dipeptide repeat proteins from ALS/FTD C9ORF72 transcript}, author = {R Tabet and L Schaeffer and F Freyermuth and M Jambeau and M Workman and C Z Lee and C C Lin and J Jiang and K Jansen-West and H Abou-Hamdan and L Désaubry and T Gendron and L Petrucelli and F Martin and C Lagier-Tourenne}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29323119?dopt=Abstract}, doi = {10.1038/s41467-017-02643-5}, isbn = {29323119}, year = {2018}, date = {2018-01-01}, journal = {Nat Commun}, volume = {9}, number = {1}, pages = {152}, abstract = {Expansion of G4C2 repeats in the C9ORF72 gene is the most prevalent inherited form of amyotrophic lateral sclerosis and frontotemporal dementia. Expanded transcripts undergo repeat-associated non-AUG (RAN) translation producing dipeptide repeat proteins from all reading frames. We determined cis-factors and trans-factors influencing translation of the human C9ORF72 transcripts. G4C2 translation operates through a 5'-3' cap-dependent scanning mechanism, requiring a CUG codon located upstream of the repeats and an initiator Met-tRNAMeti. Production of poly-GA, poly-GP, and poly-GR proteins from the three frames is influenced by mutation of the same CUG start codon supporting a frameshifting mechanism. RAN translation is also regulated by an upstream open reading frame (uORF) present in mis-spliced C9ORF72 transcripts. Inhibitors of the pre-initiation ribosomal complex and RNA antisense oligonucleotides selectively targeting the 5'-flanking G4C2 sequence block ribosomal scanning and prevent translation. Finally, we identified an unexpected affinity of expanded transcripts for the ribosomal subunits independently from translation.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of Escherichia coli Arginyl-tRNA Synthetase in Complex with tRNA^{Arg}: Pivotal Role of the D-loop}, author = {P Stephen and S Ye and M Zhou and J Song and R Zhang and E D Wang and R Giege and S X Lin}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29678554?dopt=Abstract}, doi = {10.1016/j.jmb.2018.04.011}, isbn = {29678554}, year = {2018}, date = {2018-01-01}, journal = {J Mol Biol}, volume = {430}, number = {11}, pages = {1590-1606}, abstract = {Aminoacyl-tRNA synthetases are essential components in protein biosynthesis. Arginyl-tRNA synthetase (ArgRS) belongs to the small group of aminoacyl-tRNA synthetases requiring cognate tRNA for amino acid activation. The crystal structure of Escherichia coli (Eco) ArgRS has been solved in complex with tRNAArg at 3.0-Å resolution. With this first bacterial tRNA complex, we are attempting to bridge the gap existing in structure-function understanding in prokaryotic tRNAArg recognition. The structure shows a tight binding of tRNA on the synthetase through the identity determinant A20 from the D-loop, a tRNA recognition snapshot never elucidated structurally. This interaction of A20 involves 5 amino acids from the synthetase. Additional contacts via U20a and U16 from the D-loop reinforce the interaction. The importance of D-loop recognition in EcoArgRS functioning is supported by a mutagenesis analysis of critical amino acids that anchor tRNAArg on the synthetase; in particular, mutations at amino acids interacting with A20 affect binding affinity to the tRNA and specificity of arginylation. Altogether the structural and functional data indicate that the unprecedented ArgRS crystal structure represents a snapshot during functioning and suggest that the recognition of the D-loop by ArgRS is an important trigger that anchors tRNAArg on the synthetase. In this process, A20 plays a major role, together with prominent conformational changes in several ArgRS domains that may eventually lead to the mature ArgRS:tRNA complex and the arginine activation. Functional implications that could be idiosyncratic to the arginine identity of bacterial ArgRSs are discussed.}, keywords = {GIEGE arginine identity conformational adaptation tRNA arginylation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {In cell mutational interference mapping experiment (in cell MIME) identifies the 5′ polyadenylation signal as a dual regulator of HIV-1 genomic RNA production and packaging}, author = {R P Smyth and M R Smith and A C Jousset and L Despons and G Laumond and T Decoville and P Cattenoz and C Moog and F Jossinet and M Mougel and J C Paillart and M von Kleist and R Marquet}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29514260?dopt=Abstract}, doi = {10.1093/nar/gky152}, isbn = {29514260}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {9}, pages = {e57}, abstract = {Non-coding RNA regulatory elements are important for viral replication, making them promising targets for therapeutic intervention. However, regulatory RNA is challenging to detect and characterise using classical structure-function assays. Here, we present in cell Mutational Interference Mapping Experiment (in cell MIME) as a way to define RNA regulatory landscapes at single nucleotide resolution under native conditions. In cell MIME is based on (i) random mutation of an RNA target, (ii) expression of mutated RNA in cells, (iii) physical separation of RNA into functional and non-functional populations, and (iv) high-throughput sequencing to identify mutations affecting function. We used in cell MIME to define RNA elements within the 5′ region of the HIV-1 genomic RNA (gRNA) that are important for viral replication in cells. We identified three distinct RNA motifs controlling intracellular gRNA production, and two distinct motifs required for gRNA packaging into virions. Our analysis reveals the 73AAUAAA78 polyadenylation motif within the 5′ PolyA domain as a dual regulator of gRNA production and gRNA packaging, and demonstrates that a functional polyadenylation signal is required for viral packaging even though it negatively affects gRNA production.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA Structure-A Neglected Puppet Master for the Evolution of Virus and Host Immunity}, author = {R P Smyth and M Negroni and A M Lever and J Mak and J C Kenyon}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30283444?dopt=Abstract}, doi = {10.3389/fimmu.2018.02097}, isbn = {30283444}, year = {2018}, date = {2018-01-01}, journal = {Front Immunol}, volume = {9}, pages = {2097}, abstract = {The central dogma of molecular biology describes the flow of genetic information from DNA to protein via an RNA intermediate. For many years, RNA has been considered simply as a messenger relaying information between DNA and proteins. Recent advances in next generation sequencing technology, bioinformatics, and non-coding RNA biology have highlighted the many important roles of RNA in virtually every biological process. Our understanding of RNA biology has been further enriched by a number of significant advances in probing RNA structures. It is now appreciated that many cellular and viral biological processes are highly dependent on specific RNA structures and/or sequences, and such reliance will undoubtedly impact on the evolution of both hosts and viruses. As a contribution to this special issue on host immunity and virus evolution, it is timely to consider how RNA sequences and structures could directly influence the co-evolution between hosts and viruses. In this manuscript, we begin by stating some of the basic principles of RNA structures, followed by describing some of the critical RNA structures in both viruses and hosts. More importantly, we highlight a number of available new tools to predict and to evaluate novel RNA structures, pointing out some of the limitations readers should be aware of in their own analyses.}, keywords = {NEGRONI, RNA structure immune evasion secondary structure viral RNA viral evolution, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Thermodynamic study of aptamers binding to their target proteins}, author = {T Sakamoto and E Ennifar and Y Nakamura}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29054802?dopt=Abstract}, doi = {10.1016/j.biochi.2017.10.010}, isbn = {29054802}, year = {2018}, date = {2018-01-01}, journal = {Biochimie}, volume = {145}, pages = {91-97}, abstract = {Aptamers are nucleic acids that bind to a target molecule with high affinity and specificity, which are selected from systematic evolution of ligands by exponential enrichment (SELEX). Aptamers feature high affinity and specificity to their target molecule and a large structural diversity; biophysical tools, together with structural studies, are essential to reveal the mechanism of aptamers recognition. Furthermore, understanding the mechanism of action would also contribute to their development for therapeutic applications. Isothermal titration calorimetry (ITC) is a fast and robust method to study the physical basis of molecular interactions. In a single experiment, it provides all thermodynamic parameters of a molecular interaction, including dissociation constant, Kd; Gibbs free energy change, ΔG; enthalpy change, ΔH; entropy change, ΔS; and stoichiometry, N. The development of modern microcalorimeters significantly contributed to the expansion of the ITC use in biological systems. Therefore, ITC has been applied to the development of small therapeutic agents that bind to target proteins and is increasingly being used to study aptamer-target protein interactions. This review focuses on thermodynamic approaches for understanding the molecular principles of aptamer-target interactions.}, keywords = {Aptamer Conformation Interaction Isothermal titration calorimetry Thermodynamics, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Tautomeric G*U pairs within the molecular ribosomal grip and fidelity of decoding in bacteria}, author = {A Rozov and P Wolff and H Grosjean and M Yusupov and G Yusupova and E Westhof}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29931292?dopt=Abstract}, doi = {10.1093/nar/gky547}, isbn = {29931292}, year = {2018}, date = {2018-01-01}, urldate = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {14}, pages = {7425-7435}, abstract = {We report new crystallographic structures of Thermus thermophilus ribosomes complexed with long mRNAs and native Escherichia coli tRNAs. They complete the full set of combinations of Watson-Crick G•C and miscoding G•U pairs at the first two positions of the codon-anticodon duplex in ribosome functional complexes. Within the tight decoding center, miscoding G•U pairs occur, in all combinations, with a non-wobble geometry structurally indistinguishable from classical coding Watson-Crick pairs at the same first two positions. The contacts with the ribosomal grip surrounding the decoding center are all quasi-identical, except in the crowded environment of the amino group of a guanosine at the second position; in which case a G in the codons may be preferred. In vivo experimental data show that the translational errors due to miscoding by G•U pairs at the first two positions are the most frequently encountered ones, especially at the second position and with a G on the codon. Such preferred miscodings involve a switch from an A-U to a G•U pair in the tRNA/mRNA complex and very rarely from a G = C to a G•U pair. It is concluded that the frequencies of such occurrences are only weakly affected by the codon/anticodon structures but depend mainly on the stability and lifetime of the complex, the modifications present in the anticodon loop, especially those at positions 34 and 37, in addition to the relative concentration of cognate/near-cognate tRNA species present in the cellular tRNA pool.}, keywords = {ARN-MS, ENNIFAR, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The phenotypic spectrum of germline YARS2 variants: from isolated sideroblastic anemia to mitochondrial myopathy, lactic acidosis and sideroblastic anemia 2}, author = {L G Riley and M M Heeney and J Rudinger-Thirion and M Frugier and D R Campagna and R Zhou and G A Hale and L M Hilliard and J A Kaplan and J L Kwiatkowski and C A Sieff and D P Steensma and A J Rennings and A Simons and N Schaap and R J Roodenburg and T Kleefstra and L Arenillas and J Fita-Torró and R Ahmed and M Abboud and E Bechara and R Farah and R Y Tamminga and S S Bottomley and M Sanchez and D W Swinkels and J Christodoulou and M D Fleming}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30026338}, doi = {10.3324/haematol.2017.182659}, isbn = {30026338}, year = {2018}, date = {2018-01-01}, journal = {Haematologica}, volume = {103}, number = {12}, pages = {2008-2015}, abstract = {YARS2 variants have previously been described in patients with myopathy, lactic acidosis and sideroblastic anemia 2 (MLASA2). YARS2 encodes the mitochondrial tyrosyl-tRNA synthetase, which is responsible for conjugating tyrosine to its cognate mt-tRNA for mitochondrial protein synthesis. Here we describe 14 individuals from 11 families presenting with sideroblastic anemia and with YARS2 variants that we identified using a sideroblastic anemia gene panel or exome sequencing. The phenotype of these patients ranged from MLASA to isolated congenital sideroblastic anemia. As in previous cases, inter- and intra-familial phenotypic variability was observed, however this report includes the first cases with isolated sideroblastic anemia and patients with biallelic YARS2 variants that have no clinically ascertainable phenotype. We identified ten novel YARS2 variants and three previously reported variants. In vitro amino-acylation assays of three five novel missense variants showed they that three had less effect on the catalytic activity of YARS2 than the most commonly reported variant, p.(Phe52Leu), associated with MLASA2, which may explain the milder phenotypes in patients with these variants. However, the other two missense variants had a more severe effect on YARS2 catalytic efficiency. Several patients carried the common YARS2 c.572 G>T, p.(Gly191Val) variant (minor allele frequency = 0.1259) in trans with a rare deleterious YARS2 variant. We have previously shown that the p.(Gly191Val) variant reduces YARS2 catalytic activity. Consequently, we suggest that biallelic YARS2 variants, including severe loss-of-function alleles in trans of the common p.(Gly191Val) variant, should be considered as a cause of isolated congenital sideroblastic anemia, as well as the MLASA syndromic phenotype.}, keywords = {Congenital sideroblastic anemia MLASA YARS2 mitochondrial myopathy sideroblastic anemia, FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mining for recurrent long-range interactions in RNA structures reveals embedded hierarchies in network families}, author = {V Reinharz and A Soulé and E Westhof and J Waldispühl and A Denise}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29608773?dopt=Abstract}, doi = {10.1093/nar/gky197}, isbn = {29608773}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {8}, pages = {3841-3851}, abstract = {The wealth of the combinatorics of nucleotide base pairs enables RNA molecules to assemble into sophisticated interaction networks, which are used to create complex 3D substructures. These interaction networks are essential to shape the 3D architecture of the molecule, and also to provide the key elements to carry molecular functions such as protein or ligand binding. They are made of organised sets of long-range tertiary interactions which connect distinct secondary structure elements in 3D structures. Here, we present a de novo data-driven approach to extract automatically from large data sets of full RNA 3D structures the recurrent interaction networks (RINs). Our methodology enables us for the first time to detect the interaction networks connecting distinct components of the RNA structure, highlighting their diversity and conservation through non-related functional RNAs. We use a graphical model to perform pairwise comparisons of all RNA structures available and to extract RINs and modules. Our analysis yields a complete catalog of RNA 3D structures available in the Protein Data Bank and reveals the intricate hierarchical organization of the RNA interaction networks and modules. We assembled our results in an online database (http://carnaval.lri.fr) which will be regularly updated. Within the site, a tool allows users with a novel RNA structure to detect automatically whether the novel structure contains previously observed RINs.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Expression, purification, and characterization of biologically active full-length Mason-Pfizer monkey virus (MPMV) Pr78^{Gag}}, author = {F N N Pitchai and L Ali and V N Pillai and A Chameettachal and S S Ashraf and F Mustafa and R Marquet and T A Rizvi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30087395}, doi = {10.1038/s41598-018-30142-0}, isbn = {30087395}, year = {2018}, date = {2018-01-01}, journal = {Sci Rep}, volume = {8}, number = {1}, pages = {11793}, abstract = {MPMV precursor polypeptide Pr78Gag orchestrates assembly and packaging of genomic RNA (gRNA) into virus particles. Therefore, we have expressed recombinant full-length Pr78Gag either with or without His6-tag in bacterial as well as eukaryotic cultures and purified the recombinant protein from soluble fractions of the bacterial cultures. The recombinant Pr78Gag protein has the intrinsic ability to assemble in vitro to form virus like particles (VLPs). Consistent with this observation, the recombinant protein could form VLPs in both prokaryotes and eukaryotes. VLPs formed in eukaryotic cells by recombinant Pr78Gag with or without His6-tag can encapsidate MPMV transfer vector RNA, suggesting that the inclusion of the His6-tag to the full-length Pr78Gag did not interfere with its expression or biological function. This study demonstrates the expression and purification of a biologically active, recombinant Pr78Gag, which should pave the way to study RNA-protein interactions involved in the MPMV gRNA packaging process.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural insights into the role of diphthamide on elongation factor 2 in messenger RNA reading frame maintenance}, author = {S Pellegrino and N Demeshkina and E Mancera-Martinez and S Melnikov and A Simonetti and A Myasnikov and M Yusupov and G Yusupova and Y Hashem}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29886014?dopt=Abstract}, doi = {10.1016/j.jmb.2018.06.006}, isbn = {29886014}, year = {2018}, date = {2018-01-01}, journal = {J Mol Biol}, volume = {430}, number = {17}, pages = {2677-2687}, abstract = {One of the most critical steps of protein biosynthesis is the coupled movement of messenger RNA (mRNA), that encodes genetic information, with transfer RNAs (tRNAs) on the ribosome. In eukaryotes this process is catalyzed by a conserved G-protein, the elongation factor 2 (eEF2), which carries a unique post-translational modification, called diphthamide, found in all eukaryotic species. Here we present near-atomic resolution cryo-EM structures of yeast 80S ribosome complexes containing mRNA, tRNA and eEF2 trapped in different GTP-hydrolysis states which provide further structural insights on the role of diphthamide in the mechanism of translation fidelity in eukaryotes.}, keywords = {ENNIFAR, Ribosome translocation cryo-EM diphthamide eEF2 reading-frame maintenance, Unité ARN}, pubstate = {published}, tppubtype = {article} } @misc{, title = {The role of Selenoprotein N in the differentiation of erythroid progenitors during stress erythropoiesis}, author = {R F Paulson and S Nettleford and C Liao and Y Chen and S Hao and M Baltzinger and M Thami-Braye and A Lescure and K S Prabhu}, year = {2018}, date = {2018-01-01}, number = {Dec 25}, publisher = {BioRxiv 528638 [Preprint]}, abstract = {Low serum Se is independently associated with anemia in elderly population, dialysis patients, sickle-cells patients, and hypothyroidism patients. Previous work from our laboratory showed that dietary Se deficiency in mice showed mild anemia indicating activation of stress-erythropoietic mechanisms. Unlike steady state erythropoiesis that is primarily responsible for homeostasis to produce new erythrocytes at a constant rate, stress erythropoiesis predominates when the bone marrow cannot generate sufficient erythrocytes. During such a process, short-term reconstituting hematopoietic stem cells (CD34+Kit+Sca1+Linneg) migrate to the spleen leading to the proliferation and differentiation of stress-erythroid progenitors (SEPs). These cells lead to stress burst forming unit-erythroid cells (BFU-E) followed by terminal differentiation to erythrocytes. Recent studies demonstrate deficits in selenoproteins block the expansion and development of stress BFU-E with defects in terminal differentiation. Analysis of selenoprotein expression showed that selenoprotein W (SelenoW) was highly expressed in developing SEPs. CRISPR-Cas9 knockout of SelenoW blocked the proliferation of immature SEPs in murine and human stress erythropoiesis cultures demonstrating a central role for SelenoW in stress erythropoiesis. Using the two-culture system to generate SEPs, selenoprotein N (SelenoN) expression increased as the progenitors transition from self-renewing モstem cell likeヤ progenitors to form committed erythroid progenitors. SelenoN−/− mice showed significantly slower erythroid recovery following phenylhydrazine (PHZ)-induced acute hemolytic anemia. As in the muscle satellite cells where SelenoN regulates cellular Ca2+ signaling, SelenoN may also regulate Ca2+ signaling in SEPs to modulate pathways of differentiation. In summary, these data suggest that multiple selenoproteins, including SelenoN and SelenoW, coordinately regulate stress erythropoiesis.}, keywords = {LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Editorial overview: Protein nucleic acid interactions: order, ambiguities and disorder in recognition and complex formation between proteins and nucleic acids}, author = {D Patel and E Westhof}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30553297?dopt=Abstract}, doi = {10.1016/j.sbi.2018.11.009}, isbn = {30553297}, year = {2018}, date = {2018-01-01}, journal = {Curr Opin Struct Biol}, volume = {53}, pages = {vi-viii}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The bifurcated stem loop 4 (SL4) is crucial for efficient packaging of mouse mammary tumor virus (MMTV) genomic RNA}, author = {F Mustafa and V Vivet-Boudou and A Jabeen and L M Ali and R M Kalloush and R Marquet and T A Rizvi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29929424?dopt=Abstract}, doi = {10.1080/15476286.2018.1486661}, isbn = {29929424}, year = {2018}, date = {2018-01-01}, journal = {RNA Biol}, volume = {15}, number = {8}, pages = {1047-1059}, abstract = {Packaging the mouse mammary tumor virus (MMTV) genomic RNA (gRNA) requires the entire 5' untranslated region (UTR) in conjunction with the first 120 nucleotides of the gag gene. This region includes several palindromic (pal) sequence(s) and stable stem loops (SLs). Among these, stem loop 4 (SL4) adopts a bifurcated structure consisting of three stems, two apical loops, and an internal loop. Pal II, located in one of the apical loops, mediates gRNA dimerization, a process intricately linked to packaging. We thus hypothesized that the bifurcated SL4 structure could constitute the major gRNA packaging determinant. To test this hypothesis, the two apical loops and the flanking sequences forming the bifurcated SL4 were individually mutated. These mutations all had deleterious effects on gRNA packaging and propagation. Next, single and compensatory mutants were designed to destabilize then recreate the bifurcated SL4 structure. A structure-function analysis using bioinformatics predictions and RNA chemical probing revealed that mutations that led to the loss of the SL4 bifurcated structure abrogated RNA packaging and propagation, while compensatory mutations that recreated the native SL4 structure restored RNA packaging and propagation to wild type levels. Altogether, our results demonstrate that SL4 constitutes the principal packaging determinant of MMTV gRNA. Our findings further suggest that SL4 acts as a structural switch that can not only differentiate between RNA for translation versus packaging/dimerization, but its location also allows differentiation between spliced and unspliced RNAs during gRNA encapsidation.}, keywords = {MARQUET, Mouse mammary tumor virus RNA packaging and dimerization RNA secondary structure Retroviruses palindrome, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of histone deacetylase 8 (HDAC8) selective inhibition reveals specific active site structural and functional determinants}, author = {M Marek and T B Shaik and T Heimburg and A Chakrabarti and J Lancelot and E Ramos Morales and C Da Veiga and D V Kalinin and J Melesina and D Robaa and K Schmidtkunz and T Suzuki and R Holl and E Ennifar and R Pierce and M Jung and W Sippl and C Romier}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30347148}, doi = {10.1021/acs.jmedchem.8b01087}, isbn = {30347148}, year = {2018}, date = {2018-01-01}, journal = {J Med Chem}, volume = {61}, number = {22}, pages = {10000-10016}, abstract = {Metal-dependent histone deacetylases (HDACs) are key epigenetic regulators that represent promising therapeutic targets for the treatment of numerous human diseases. Yet, the currently FDA-approved HDAC inhibitors non-specifically target at least several of the eleven structurally similar but functionally different HDAC isozymes, which hampers their broad usage in clinical settings. Selective inhibitors targeting single HDAC isozymes are being developed, but precise understanding in molecular terms of their selectivity remains sparse. Here, we show that HDAC8-selective inhibitors adopt a L-shaped conformation required for their binding to a HDAC8-specific pocket formed by HDAC8 catalytic tyrosine and HDAC8 L1 and L6 loops. In other HDAC isozymes, a L1-L6 lock sterically prevents L-shaped inhibitor binding. Shielding of the HDAC8-specific pocket by protein engineering decreases potency of HDAC8-selective inhibitors and affects catalytic activity. Collectively, our results unravel key HDAC8 active site structural and functional determinants important for the design of next-generation chemical probes and epigenetic drugs.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Viral internal ribosomal entry sites: four classes for one goal}, author = {J Mailliot and F Martin}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29193740?dopt=Abstract}, doi = {10.1002/wrna.1458}, isbn = {29193740}, year = {2018}, date = {2018-01-01}, journal = {Wiley Interdiscip Rev RNA}, volume = {9}, number = {2}, pages = {none}, abstract = {To ensure efficient propagation, viruses need to rapidly produce viral proteins after cell entrance. Since viral genomes do not encode any components of the protein biosynthesis machinery, viral proteins must be produced by the host cell. To hi-jack the host cellular translation, viruses use a great variety of distinct strategies. Many single-stranded positive-sensed RNA viruses contain so-called internal ribosome entry sites (IRESs). IRESs are structural RNA motifs that have evolved to specific folds that recruit the host ribosomes on the viral coding sequences in order to synthesize viral proteins. In host canonical translation, recruitment of the translation machinery components is essentially guided by the 5' cap (m7 G) of mRNA. In contrast, IRESs are able to promote efficient ribosome assembly internally and in cap-independent manner. IRESs have been categorized into four classes, based on their length, nucleotide sequence, secondary and tertiary structures, as well as their mode of action. Classes I and II require the assistance of cellular auxiliary factors, the eukaryotic intiation factors (eIF), for efficient ribosome assembly. Class III IRESs require only a subset of eIFs whereas Class IV, which are the more compact, can promote translation without any eIFs. Extensive functional and structural investigations of IRESs over the past decades have allowed a better understanding of their mode of action for viral translation. Because viral translation has a pivotal role in the infectious program, IRESs are therefore attractive targets for therapeutic purposes. For further resources related to this article, please visit the WIREs website.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Uncovering the Importance of Selenium in Muscle Disease}, author = {A Lescure and M Baltzinger and E Zito}, editor = {B Michalke}, url = {https://link.springer.com/chapter/10.1007/978-3-319-95390-8_18}, doi = {978-3-319-95390-8_18}, year = {2018}, date = {2018-01-01}, booktitle = {Selenium: Series Molecular and Integrative Toxicology}, pages = {345-362}, publisher = {Springer}, address = {New-York}, series = {Molecular and Integrative Toxicology}, abstract = {A connection between selenium bioavailability and development of muscular disorders both in humans and livestock has been established for a long time. With the development of genomics, the function of several selenoproteins was shown to be involved in muscle activity, including SELENON, which was linked to an inherited form of myopathy. Development of animal models has helped to dissect the physiological dysfunction due to mutation in the SELENON gene; however the molecular activity remains elusive and only recent analysis using both in vivo and in vitro experiment provided hints toward its function in oxidative stress defence and calcium transport control. This review sets out to summarise most recent findings for the importance of selenium in muscle function and the contribution of this information to the design of strategies to cure the diseases.}, keywords = {elenium Selenoproteins Muscle disease Oxidative stress Calcium transport RYR SERCA, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {MS2-Affinity Purification Coupled With RNA Sequencing Approach in the Human Pathogen Staphylococcus aureus}, author = {D Lalaouna and E Desgranges and I Caldelari and S Marzi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30502950?dopt=Abstract}, doi = {10.1016/bs.mie.2018.08.022}, isbn = {30502950}, year = {2018}, date = {2018-01-01}, journal = {Methods Enzymol}, volume = {612}, pages = {393-411}, abstract = {Staphylococcus aureus is a Gram-positive major human pathogen involved in a wide range of human infectious diseases (from minor skin infections to septicemia, endocarditis or toxic shock syndrome). The treatment of S. aureus infections is very challenging due to the emergence of multiple antibiotic-resistant isolates. The high diversity of clinical symptoms caused by S. aureus depends on the precise expression of numerous virulence factors and stress response pathways, which are tightly regulated at every level (transcriptional, posttranscriptional, translational, and posttranslational). During the last two decades, it has become evident that small regulatory RNAs (sRNAs) play a major role in fast adaptive responses, mainly by targeting mRNA translation. sRNAs act as antisense RNAs by forming noncontiguous pairings with their target mRNAs and their mechanisms of action vary according to the interaction site. To obtain a global and detailed view of the regulatory networks involved in the adaptive processes of S. aureus, we have adapted the MAPS approach to get individual sRNA targetomes. We also set up different strategies to validate MAPS results and establish sRNA regulatory activities. As this method has been first developed in Gram-negative bacteria, we provide here a protocol for its application in S. aureus and highlight underlying differences. Finally, we discuss several points that have been and could be further improved and provide a workflow file for the automatic analysis of the sequencing in Galaxy.}, keywords = {Galaxy workflow Interactome MS2-affinity purification coupled with RNA sequencing RNA:RNA interaction Small regulatory RNA Staphylococcus aureus Targetome, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Specific Chemical Approaches for Studying Mammalian Ribosomes Complexed with Ligands Involved in Selenoprotein Synthesis}, author = {O Kossinova and A Malygin and A Krol and G Karpova}, editor = {L Chavatte}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28917038?dopt=Abstract}, doi = {10.1007/978-1-4939-7258-6_6}, isbn = {28917038}, year = {2018}, date = {2018-01-01}, booktitle = {Selenoproteins: Methods and Protocols}, volume = {1661}, pages = {73-92}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Chemical approaches are very powerful tools for investigating the molecular structure and architecture of large ribonucleoprotein complexes involving ribosomes and other components of the translation system. Application of RNA nucleotide-specific and cross-linking reagents of a broad specificity range allows the researcher to obtain information on the sites of ligand binding to the ribosome and to each other as well as on the RNA rearrangements caused by the binding. Here, we describe specific chemical approaches including chemical probing and site-directed or bifunctional reagent-mediated cross-linking, which have been used for exploring the mechanism of selenocysteine insertion into a polypeptide chain by mammalian ribosomes.}, keywords = {Chemical modification Chemical probing Cross-linking RNA Ribosome Selenocysteine Selenoprotein, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Double methylation of tRNA-U54 to 2'-O-methylthymidine (Tm) synergistically decreases immune response by Toll-like receptor 7}, author = {P Keller and I Freund and V Marchand and G Bec and R Huang and Y Motorin and T Eigenbrod and A Dalpke and M Helm}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30102387?dopt=Abstract}, doi = {10.1093/nar/gky644}, isbn = {30105811}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {18}, pages = {9764-9775}, abstract = {Sensing of nucleic acids for molecular discrimination between self and non-self is a challenging task for the innate immune system. RNA acts as a potent stimulus for pattern recognition receptors including in particular human Toll-like receptor 7 (TLR7). Certain RNA modifications limit potentially harmful self-recognition of endogenous RNA. Previous studies had identified the 2'-O-methylation of guanosine 18 (Gm18) within tRNAs as an antagonist of TLR7 leading to an impaired immune response. However, human tRNALys3 was non-stimulatory despite lacking Gm18. To identify the underlying molecular principle, interferon responses of human peripheral blood mononuclear cells to differentially modified tRNALys3 were determined. The investigation of synthetic modivariants allowed attributing a significant part of the immunosilencing effect to the 2'-O-methylthymidine (m5Um) modification at position 54. The effect was contingent upon the synergistic presence of both methyl groups at positions C5 and 2'O, as shown by the fact that neither Um54 nor m5U54 produced any effect alone. Testing permutations of the nucleobase at ribose-methylated position 54 suggested that the extent of silencing and antagonism of the TLR7 response was governed by hydrogen patterns and lipophilic interactions of the nucleobase. The results identify a new immune-modulatory endogenous RNA modification that limits TLR7 activation by RNA.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Small but large enough: structural properties of armless mitochondrial tRNAs from the nematode Romanomermis culicivorax}, author = {T Jühling and E Duchardt-Ferner and S Bonin and J Wöhnert and J Pütz and C Florentz and H Betat and C Sauter and M Mörl}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29986062?dopt=Abstract}, doi = {10.1093/nar/gky593}, isbn = {29986062}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {17}, pages = {9170-9180}, abstract = {As adapter molecules to convert the nucleic acid information into the amino acid sequence, tRNAs play a central role in protein synthesis. To fulfill this function in a reliable way, tRNAs exhibit highly conserved structural features common in all organisms and in all cellular compartments active in translation. However, in mitochondria of metazoans, certain dramatic deviations from the consensus tRNA structure are described, where some tRNAs lack the D- or T-arm without losing their function. In Enoplea, this miniaturization comes to an extreme, and functional mitochondrial tRNAs can lack both arms, leading to a considerable size reduction. Here, we investigate the secondary and tertiary structure of two such armless tRNAs from Romanomermis culicivorax. Despite their high AU content, the transcripts fold into a single and surprisingly stable hairpin structure, deviating from standard tRNAs. The three-dimensional form is boomerang-like and diverges from the standard L-shape. These results indicate that such unconventional miniaturized tRNAs can still fold into a tRNA-like shape, although their length and secondary structure are very unusual. They highlight the remarkable flexibility of the protein synthesis apparatus and suggest that the translational machinery of Enoplea mitochondria may show compensatory adaptations to accommodate these armless tRNAs for efficient translation.}, keywords = {FLORENTZ, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {How to fold and protect mitochondrial ribosomal RNA with fewer guanines}, author = {M Hosseini and P Roy and M Sissler and C L Zirbel and E Westhof and N Leontis}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30215760?dopt=Abstract}, doi = {10.1093/nar/gky762}, isbn = {30215760}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {20}, pages = {10946-10968}, abstract = {Mammalian mitochondrial ribosomes evolved from bacterial ribosomes by reduction of ribosomal RNAs, increase of ribosomal protein content, and loss of guanine nucleotides. Guanine is the base most sensitive to oxidative damage. By systematically comparing high-quality, small ribosomal subunit RNA sequence alignments and solved 3D ribosome structures from mammalian mitochondria and bacteria, we deduce rules for folding a complex RNA with the remaining guanines shielded from solvent. Almost all conserved guanines in both bacterial and mammalian mitochondrial ribosomal RNA form guanine-specific, local or long-range, RNA-RNA or RNA-protein interactions. Many solvent-exposed guanines conserved in bacteria are replaced in mammalian mitochondria by bases less sensitive to oxidation. New guanines, conserved only in the mitochondrial alignment, are strategically positioned at solvent inaccessible sites to stabilize the ribosomal RNA structure. New mitochondrial proteins substitute for truncated RNA helices, maintain mutual spatial orientations of helices, compensate for lost RNA-RNA interactions, reduce solvent accessibility of bases, and replace guanines conserved in bacteria by forming specific amino acid-RNA interactions.}, keywords = {SISSLER, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Translation of non-standard codon nucleotides reveals minimal requirements for codon-anticodon interactions}, author = {T P Hoernes and K Faserl and M A Juen and J Kremser and C Gasser and E Fuchs and X Shi and A Siewert and H Lindner and C Kreutz and R Micura and S Joseph and C Höbartner and E Westhof and A Hüttenhofer and M D Erlacher}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30451861?dopt=Abstract}, doi = {10.1038/s41467-018-07321-8}, isbn = {30451861}, year = {2018}, date = {2018-01-01}, journal = {Nat Commun}, volume = {9}, number = {1}, pages = {4865}, abstract = {The precise interplay between the mRNA codon and the tRNA anticodon is crucial for ensuring efficient and accurate translation by the ribosome. The insertion of RNA nucleobase derivatives in the mRNA allowed us to modulate the stability of the codon-anticodon interaction in the decoding site of bacterial and eukaryotic ribosomes, allowing an in-depth analysis of codon recognition. We found the hydrogen bond between the N1 of purines and the N3 of pyrimidines to be sufficient for decoding of the first two codon nucleotides, whereas adequate stacking between the RNA bases is critical at the wobble position. Inosine, found in eukaryotic mRNAs, is an important example of destabilization of the codon-anticodon interaction. Whereas single inosines are efficiently translated, multiple inosines, e.g., in the serotonin receptor 5-HT2C mRNA, inhibit translation. Thus, our results indicate that despite the robustness of the decoding process, its tolerance toward the weakening of codon-anticodon interactions is limited.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nanobody-mediated resistance to Grapevine fanleaf virus in plants.}, author = {C Hemmer and S Djennane and L Ackerer and K Hleibieh and A Marmonier and S Gersch and S Garcia and E Vigne and V Komar and M Perrin and C Gertz and L Belval and F Berthold and B Monsion and C Schmitt-Keichinger and O Lemaire and B Lorber and C Gutiérrez and S Muyldermans and G Demangeat and C Ritzenthaler}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28796912?dopt=Abstract}, doi = {10.1111/pbi.12819}, isbn = {28796912}, year = {2018}, date = {2018-01-01}, journal = {Plant Biotechnol J}, volume = {16}, number = {2}, pages = {660-671}, abstract = {Since their discovery, single-domain antigen-binding fragments of camelid-derived heavy chain-only antibodies, also known as Nanobodies (Nbs), have proven to be of outstanding interest as therapeutics against human diseases and pathogens including viruses, but their use against phytopathogens remains limited. Many plant viruses including Grapevine fanleaf virus (GFLV), a nematode-transmitted icosahedral virus and causal agent of fanleaf degenerative disease, have worldwide distribution and huge burden on crop yields representing billions of US dollars of losses annually, yet solutions to combat these viruses are often limited or inefficient. Here we identified a Nb specific to GFLV that confers strong resistance to GFLV upon stable expression in the model plant Nicotiana benthamiana and also in grapevine rootstock, the natural host of the virus. We showed that resistance was effective against a broad range of GFLV isolates independently of the inoculation method including upon nematode transmission but not against its close relative, Arabis mosaic virus. We also demonstrated that virus neutralization occurs at an early step of the virus life cycle, prior to cell-to-cell movement. Our findings will not only be instrumental to confer resistance to GFLV in grapevine but more generally they pave the way for the generation of novel antiviral strategies in plants based on Nbs. This article is protected by copyright. All rights reserved.}, keywords = {FRUGIER, GMO Nanobodies Single chain antibodies grapevine nepovirus plant virus transgenic plant, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Tracking the m^{7}G-cap during translation initiation by crosslinking methods}, author = {L Gross and L Schaeffer and F Alghoul and H Hayek and C Allmang and G Eriani and F Martin}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29307728?dopt=Abstract}, doi = {10.1016/j.ymeth.2017.12.019}, isbn = {29307728}, year = {2018}, date = {2018-01-01}, journal = {Methods}, volume = {137}, pages = {3-10}, abstract = {In eukaryotes, cap-dependent translation initiation is a sophisticated process that requires numerous trans-acting factors, the eukaryotic Initiation Factors (eIFs). Their main function is to assist the ribosome for accurate AUG start codon recognition. The whole process requires a 5'-3' scanning step and is therefore highly dynamic. Therefore translation requires a complex interplay between eIFs through assembly/release cycles. Here, we describe an original approach to assess the dynamic features of translation initiation. The principle is to use the m7Gcap located at the 5' extremity of mRNAs as a tracker to monitor RNA and protein components that are in its vicinity. Cap-binding molecules are trapped by chemical and UV crosslinking. The combination of cap crosslinking methods in cell-free translation systems with the use of specific translation inhibitors for different steps such as edeine, GMP-PNP or cycloheximide allowed assessing the cap fate during eukaryotic translation. Here, we followed the position of the cap in the histone H4 mRNA and the cap binding proteins during H4 mRNA translation.}, keywords = {Cap-dependent translation Chemical crosslinking Histone H4 mRNA Ribosome UV crosslinking eIF4E, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Modification of Selenoprotein mRNAs by Cap Tri-methylation}, author = {A S Gribling-Burrer and G Eriani and C Allmang}, editor = {L Chavatte}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28917041?dopt=Abstract}, doi = {1007/978-1-4939-7258-6_9}, isbn = {28917041}, year = {2018}, date = {2018-01-01}, booktitle = {Selenoproteins: Methods and Protocols}, volume = {1661}, pages = {125-141}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Several selenoprotein mRNAs undergo 5' cap maturation events whereby their classical monomethylated m7G cap becomes trimethylated (m32,2,7G) by the trimethylguanosine synthase 1 (Tgs1). Here, we describe immunoprecipitation methods for the detection of endogenous m32,2,7G-capped selenoprotein mRNAs from total cell extracts or after polysome fractionation of cytoplasmic extracts. We have also developed a method for the in vitro cap hypermethylation of selenoprotein mRNA transcripts using purified Tgs1 enzyme.}, keywords = {2, 7G cap, Cap hypermethylation Cap immunoprecipitation Selenoprotein mRNAs TMG cap Tgs1 m3 2, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Three human aminoacyl-tRNA synthetases have distinct sub-mitochondrial localizations that are unaffected by disease-associated mutations}, author = {L E González-Serrano and L Karim and F Pierre and H Schwenzer and A Rotig and A Munnich and M Sissler}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30006346?dopt=Abstract}, doi = {10.1074/jbc.RA118.003400}, isbn = {30006346}, year = {2018}, date = {2018-01-01}, journal = {J Biol Chem}, volume = {293}, number = {35}, pages = {13604-13615}, abstract = {Human mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs) are key enzymes in the mitochondrial protein translation system and catalyze the charging of amino acids on their cognate tRNAs. Mutations in their nuclear genes are associated with pathologies having a broad spectrum of clinical phenotypes, but with no clear molecular mechanism(s). For example, mutations in the nuclear genes encoding mt-AspRS and mt-ArgRS are correlated with the moderate neurodegenerative disorder leukoencephalopathy with brainstem and spinal cord involvement and lactate elevation (LBSL) and with the severe neurodevelopmental disorder pontocerebellar hypoplasia type 6 (PCH6), respectively. Previous studies have shown no or only minor impacts of these mutations on the canonical properties of these enzymes, indicating that the role of the mt-aaRSs in protein synthesis is mostly not affected by these mutations, but their effects on the mitochondrial localizations of aaRSs remain unclear. Here, we demonstrate that three human aaRSs, mt-AspRS, mt-ArgRS, and LysRS, each have a specific sub-mitochondrial distribution, with mt-ArgRS being exclusively localized in the membrane, LysRS exclusively in the soluble fraction, and mt-AspRS being present in both. Chemical treatments revealed that mt-AspRs is anchored in the mitochondrial membrane through electrostatic interactions, whereas mt-ArgRS uses hydrophobic interactions. We also report that novel mutations in mt-AspRS and mt-ArgRS genes from individuals with LBSL and PCH6, respectively, had no significant impact on the mitochondrial localizations of mt-AspRS and mt-ArgRS. The variable sub-mitochondrial locations for these three mt-aaRSs strongly suggest the existence of additional enzyme properties, requiring further investigation to unravel the mechanisms underlying the two neurodegenerative disorders.}, keywords = {aminoacyl tRNA synthetase dual-localization human leukodystrophy membrane-anchored mitochondria mitochondrial disorder neurodegenerative disease pontocerebellar hypoplasia translation, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {On the Importance of Host MicroRNAs During Viral Infection}, author = {E Girardi and P López and S Pfeffer}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30333857?dopt=Abstract}, doi = {10.3389/fgene.2018.00439}, isbn = {30333857}, year = {2018}, date = {2018-01-01}, journal = {Front Genet}, volume = {9}, pages = {439}, abstract = {Every living organism has to constantly face threats from the environment and deal with a large number of pathogens against which it has to defend itself to survive. Among those, viruses represent a large class of obligatory intracellular parasites, which rely on their host machinery to multiply and propagate. As a result, viruses and their hosts have engaged in an ever-evolving arms race to be able to maintain their existence. The role played by micro (mi)RNAs in this ongoing battle has been extensively studied in the past 15 years and will be the subject of this review article. We will mainly focus on cellular miRNAs and their implication during viral infection in mammals. Thus, we will describe current techniques that can be used to identify miRNAs involved in the modulation of viral infection and to characterize their targets and mode of action. We will also present different reported examples of miRNA-mediated regulation of viruses, which can have a positive outcome either for the host or for the virus. In addition, the mode of action is also of a dual nature, depending on the target of the miRNA. Indeed, the regulatory small RNA can either directly guide an Argonaute protein on a viral transcript, or target a cellular mRNA involved in the host antiviral response. We will then see whether and how viruses respond to miRNA-mediated targeting. Finally, we will discuss how our knowledge of viral targeting by miRNA can be exploited for developing new antiviral therapeutic approaches.}, keywords = {defense mechanism host–pathogen interaction microRNA post-transcriptional regulation virus, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural and functional motifs in influenza virus RNAs}, author = {D Ferhadian and M Contrant and A Printz-Schweigert and R P Smyth and J C Paillart and R Marquet}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29651275?dopt=Abstract}, doi = {10.3389/fmicb.2018.00559}, isbn = {29651275}, year = {2018}, date = {2018-01-01}, journal = {Front Microbiol}, volume = {9}, pages = {599}, abstract = {Influenza A viruses (IAV) are responsible for recurrent influenza epidemics and occasional devastating pandemics in humans and animals. They belong to the Orthomyxoviridae family and their genome consists of eight (-) sense viral RNA (vRNA) segments of different lengths coding for at least 11 viral proteins. A heterotrimeric polymerase complex is bound to the promoter consisting of the 13 5′-terminal and 12 3′-terminal nucleotides of each vRNA, while internal parts of the vRNAs are associated with multiple copies of the viral nucleoprotein (NP), thus forming ribonucleoproteins (vRNP). Transcription and replication of vRNAs result in viral mRNAs (vmRNAs) and complementary RNAs (cRNAs), respectively. Complementary RNAs are the exact positive copies of vRNAs; they also form ribonucleoproteins (cRNPs) and are intermediate templates in the vRNA amplification process. On the contrary, vmRNAs have a 5′ cap snatched from cellular mRNAs and a 3′ polyA tail, both gained by the viral polymerase complex. Hence, unlike vRNAs and cRNAs, vmRNAs do not have a terminal promoter able to recruit the viral polymerase. Furthermore, synthesis of at least two viral proteins requires vmRNA splicing. Except for extensive analysis of the viral promoter structure and function and a few, mostly bioinformatics, studies addressing the vRNA and vmRNA structure, structural studies of the influenza A vRNAs, cRNAs, and vmRNAs are still in their infancy. The recent crystal structures of the influenza polymerase heterotrimeric complex drastically improved our understanding of the replication and transcription processes. The vRNA structure has been mainly studied in vitro using RNA probing, but its structure has been very recently studied within native vRNPs using crosslinking and RNA probing coupled to next generation RNA sequencing. Concerning vmRNAs, most studies focused on the segment M and NS splice sites and several structures initially predicted by bioinformatics analysis have now been validated experimentally and their role in the viral life cycle demonstrated. This review aims to compile the structural motifs found in the different RNA classes (vRNA, cRNA, and vmRNA) of influenza viruses and their function in the viral replication cycle.}, keywords = {CRNA RNA RNA structure influenza influenza A virus promoter vRN, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {START: STructure-Assisted RNA Translation}, author = {G Eriani and F Martin}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30176155?dopt=Abstract}, doi = {10.1080/15476286.2018.1518855}, isbn = {30176155}, year = {2018}, date = {2018-01-01}, journal = {RNA Biol}, volume = {15}, number = {9}, pages = {1250-1253}, abstract = {Cap-dependent translation initiation begins by assembly of a pre-initiation ribosomal complex that scans the 5' Untranslated Region in order to localise the start codon. During this process, RNA secondary structures are melted by RNA helicases. Guenther et al reported that the yeast helicase Ded1, an orthologue of the mammalian DDX3 helicase, is responsible for this activity. When Ded1 is non-functional, RNA structures in the 5'UTR promote translation initiation on Alternative Translation Initiation Sites (ATIS) lead to uORF translation and consequently down-regulation of the main ORF. This mechanism is driven by the sole presence of RNA secondary structures located downstream of ATIS. Translation initiation mediated by RNA structures is found in other messenger RNAs. We propose to name this novel mechanism STructure-Assisted-RNA-Translation or START.}, keywords = {Ded1/DDX3 RNA helicase Ribosome Scanning Translation initiation messenger RNA, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The NC domain of HIV-1 Gag contributes to the interaction of Gag with TSG101}, author = {S E El Meshri and E Boutant and A Mouhand and A Thomas and V Larue and L Richert and V Vivet-Boudou and Y Mély and C Tisné and D Muriaux and H de Rocquigny}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29571744?report=&dispmax=200&tool=PubCrawler_2.39}, doi = {10.1016/j.bbagen.2018.03.020}, isbn = {29571744}, year = {2018}, date = {2018-01-01}, journal = {Biochim Biophys Acta-Gen Subj}, volume = {1862}, number = {6}, pages = {1421-1431}, abstract = {BACKGROUND: HIV-1 Gag polyprotein orchestrates the assembly of viral particles. Its C-terminus consists of the nucleocapsid (NC) domain that interacts with RNA, and the p6 domain containing the PTAP motif that binds the cellular ESCRT factor TSG101 and ALIX. Deletion of the NC domain of Gag (GagNC) results in defective Gag assembly, a decrease in virus production and, thus probably affects recruitment of the ESCRT machinery. To investigate the role of GagNC in this recruitment, we analysed its impact on TSG101 and ALIX localisations and interactions in cells expressing Gag. METHODS: Cells expressing mCherry-Gag or derivatives, alone or together with eGFP-TSG101 or eGFP-ALIX, were analysed by confocal microscopy and FLIM-FRET. Chemical shift mapping between TSG101-UEV motif and Gag C-terminus was performed by NMR. RESULTS: We show that deletion of NC or of its two zinc fingers decreases the amount of Gag-TSG101 interacting complexes in cells. These findings are supported by NMR data showing chemical shift perturbations in the NC domain in- and outside - of the zinc finger elements upon TSG101 binding. The NMR data further identify a large stretch of amino acids within the p6 domain directly interacting with TSG101. CONCLUSION: The NC zinc fingers and p6 domain of Gag participate in the formation of the Gag-TSG101 complex and in its cellular localisation. GENERAL SIGNIFICANCE: This study illustrates that the NC and p6 domains cooperate in the interaction with TSG101 during HIV-1 budding. In addition, details on the Gag-TSG101 complex were obtained by combining two high resolution biophysical techniques.}, keywords = {FRET Gag HIV NMR Nucleocapsid TSG101, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Retroviral RNA dimerization: from structure to functions}, author = {N Dubois and R Marquet and J C Paillart and S Bernacchi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29623074?dopt=Abstract}, doi = {doi.org/10.3389/fmicb.2018.00527}, isbn = {29623074}, year = {2018}, date = {2018-01-01}, journal = {Front Microbiol}, volume = {9}, pages = {527}, abstract = {The genome of the retroviruses is a dimer composed by two homologous copies of genomic RNA (gRNA) molecules of positive polarity. The dimerization process allows two gRNA molecules to be non-covalently linked together through intermolecular base-pairing. This step is critical for the viral life cycle and is highly conserved among retroviruses with the exception of spumaretroviruses. Furthermore, packaging of two gRNA copies into viral particles presents an important evolutionary advantage for immune system evasion and drug resistance. Recent studies reported RNA switches models regulating not only gRNA dimerization, but also translation and packaging, and a spatio-temporal characterization of viral gRNA dimerization within cells are now at hand. This review summarizes our current understanding on the structural features of the dimerization signals for a variety of retroviruses (HIVs, MLV, RSV, BLV, MMTV, MPMV…), the mechanisms of RNA dimer formation and functional implications in the retroviral cycle.}, keywords = {HIV MuLV RNA dimerization function retrovirus structure, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The C-terminal p6 domain of the HIV-1 Pr55^{Gag} precursor is required for specific binding to the genomic RNA.}, author = {N Dubois and K K Khoo and S Ghossein and T Seissler and P Wolff and W J McKinstry and J Mak and J C Paillart and R Marquet and S Bernacchi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29954247?dopt=Abstract}, doi = {10.1080/15476286.2018.1481696}, isbn = {29954247}, year = {2018}, date = {2018-01-01}, journal = {RNA Biol}, volume = {15}, number = {7}, pages = {923-936}, abstract = {The Pr55Gag precursor specifically selects the HIV-1 genomic RNA (gRNA) from a large excess of cellular and partially or fully spliced viral RNAs and drives the virus assembly at the plasma membrane. During these processes, the NC domain of Pr55Gag interacts with the gRNA, while its C-terminal p6 domain binds cellular and viral factors and orchestrates viral particle release. Gag∆p6 is a truncated form of Pr55Gag lacking the p6 domain usually used as a default surrogate for wild type Pr55Gag for in vitro analysis. With recent advance in production of full-length recombinant Pr55Gag, here, we tested whether the p6 domain also contributes to the RNA binding specificity of Pr55Gag by systematically comparing binding of Pr55Gag and Gag∆p6 to a panel of viral and cellular RNAs. Unexpectedly, our fluorescence data reveal that the p6 domain is absolutely required for specific binding of Pr55Gag to the HIV-1 gRNA. Its deletion resulted not only in a decreased affinity for gRNA, but also in an increased affinity for spliced viral and cellular RNAs. In contrast Gag∆p6 displayed a similar affinity for all tested RNAs. Removal of the C-terminal His-tag from Pr55Gag and Gag∆p6 uniformly increased the Kd values of the RNA-protein complexes by ~2.5 fold but did not affect the binding specificities of these proteins. Altogether, our results demonstrate a novel role of the p6 domain in the specificity of Pr55Gag-RNA interactions, and strongly suggest that the p6 domain contributes to the discrimination of HIV-1 gRNA from cellular and spliced viral mRNAs, which is necessary for its selective encapsidation.}, keywords = {ENNIFAR, HIV-1 Pr55Gag RNA-protein binding specificity fluorescence spectroscopy genomic RNA p6 domain, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Combining crystallogenesis methods to produce diffraction-quality crystals of a psychrophilic tRNA-maturation enzyme}, author = {R de Wijn and O Hennig and F G M Ernst and B Lorber and H Betat and M Mörl and C Sauter}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30387781?dopt=Abstract}, doi = {10.1107/S2053230X18014590}, isbn = {30387781}, year = {2018}, date = {2018-01-01}, journal = {Acta Crystallogr F Struct Biol Commun}, volume = {74}, number = {Pt 11}, pages = {747-753}, abstract = {The determination of conditions for the reproducible growth of well diffracting crystals is a critical step in every biocrystallographic study. On the occasion of a new structural biology project, several advanced crystallogenesis approaches were tested in order to increase the success rate of crystallization. These methods included screening by microseed matrix screening, optimization by counter-diffusion and crystal detection by trace fluorescent labeling, and are easily accessible to any laboratory. Their combination proved to be particularly efficient in the case of the target, a 48 kDa CCA-adding enzyme from the psychrophilic bacterium Planococcus halocryophilus. A workflow summarizes the overall strategy, which led to the production of crystals that diffracted to better than 2 Å resolution and may be of general interest for a variety of applications.}, keywords = {CCA-adding enzyme Planococcus halocryophilus counter-diffusion crystallogenesis microseeding optimization tRNA maturation trace fluorescent labeling, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of receptors for UNCG and GNRA Z-turns and their occurrence in rRNA}, author = {L D'Ascenzo and Q Vicens and P Auffinger}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29986118?dopt=Abstract}, doi = {10.1093/nar/gky578}, isbn = {29986118}, year = {2018}, date = {2018-01-01}, journal = {Nucleic Acids Res}, volume = {46}, number = {15}, pages = {7989-7997}, abstract = {In contrast to GNRA tetraloop receptors that are common in RNA, receptors for the more thermostable UNCG loops have remained elusive for almost three decades. An analysis of all RNA structures with resolution ≤3.0 Å from the PDB allowed us to identify three previously unnoticed receptors for UNCG and GNRA tetraloops that adopt a common UNCG fold, named 'Z-turn' in agreement with our previously published nomenclature. These receptors recognize the solvent accessible second Z-turn nucleotide in different but specific ways. Two receptors participating in a complex network of tertiary interactions are associated with the rRNA UUCG and GAAA Z-turns capping helices H62 and H35a in rRNA large subunits. Structural comparison of fully assembled ribosomes and comparative sequence analysis of >6500 rRNA sequences helped us recognize that these motifs are almost universally conserved in rRNA, where they may contribute to organize the large subunit around the subdomain-IV four-way junction. The third UCCG receptor was identified in a rRNA/protein construct crystallized at acidic pH. These three non-redundant Z-turn receptors are relevant for our understanding of the assembly of rRNA and other long-non-coding RNAs, as well as for the design of novel folding motifs for synthetic biology.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Regulation of primary-microRNA processing}, author = {A Creugny and A Fender and S Pfeffer}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29683487?dopt=Abstract}, doi = {10.1002/1873-3468.13067}, isbn = {29683487}, year = {2018}, date = {2018-01-01}, journal = {FEBS Lett}, volume = {592}, number = {12}, pages = {1980-1996}, abstract = {MicroRNAs (miRNAs) are evolutionarily conserved small regulatory RNAs that participate in the fine-tuning of many, if not all, fundamental biological processes. Molecular mechanisms involved in miRNA biogenesis and mode of action have been elucidated in the past two decades. Similar to many cellular pathways, miRNA processing and function can be globally or specifically regulated at several levels and by numerous proteins and RNAs. Given their role as fine-tuning molecules, it is essential for miRNA expression to be tightly regulated in order to maintain cellular homeostasis. Here, we review our current knowledge of the first step of their maturation occurring in the nucleus and how it can be specifically and dynamically modulated. This article is protected by copyright. All rights reserved.}, keywords = {Drosha biogenesis microRNA regulation, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Biochemical and Functional Characterization of Mouse Mammary Tumor Virus Full-Length Pr77^{Gag} Expressed in Prokaryotic and Eukaryotic Cells}, author = {A Chameettachal and V N Pillai and L M Ali and F N N Pitchai and M T Ardah and F Mustafa and R Marquet and T A Rizvi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29912170?dopt=Abstract}, doi = {10.3390/v10060334}, isbn = {29912170}, year = {2018}, date = {2018-01-01}, journal = {Viruses}, volume = {10}, number = {6}, pages = {334}, abstract = {The mouse mammary tumor virus (MMTV) Pr77Gag polypeptide is an essential retroviral structural protein without which infectious viral particles cannot be formed. This process requires specific recognition and packaging of dimerized genomic RNA (gRNA) by Gag during virus assembly. Most of the previous work on retroviral assembly has used either the nucleocapsid portion of Gag, or other truncated Gag derivatives—not the natural substrate for virus assembly. In order to understand the molecular mechanism of MMTV gRNA packaging process, we expressed and purified full-length recombinant Pr77Gag-His₆-tag fusion protein from soluble fractions of bacterial cultures. We show that the purified Pr77Gag-His₆-tag protein retained the ability to assemble virus-like particles (VLPs) in vitro with morphologically similar immature intracellular particles. The recombinant proteins (with and without His₆-tag) could both be expressed in prokaryotic and eukaryotic cells and had the ability to form VLPs in vivo. Most importantly, the recombinant Pr77Gag-His₆-tag fusion proteins capable of making VLPs in eukaryotic cells were competent for packaging sub-genomic MMTV RNAs. The successful expression and purification of a biologically active, full-length MMTV Pr77Gag should lay down the foundation towards performing RNA⁻protein interaction(s), especially for structure-function studies and towards understanding molecular intricacies during MMTV gRNA packaging and assembly processes.}, keywords = {MARQUET, PAILLART, Pr77Gag RNA packaging RNA–Gag interactions RNA–protein interaction mouse mammary tumor virus (MMTV) protein assembly protein expression protein purification retrovirus, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Plasmodium apicoplast tyrosyl-tRNA synthetase recognizes an unusual, simplified identity set in cognate tRNATyr}, author = {M Cela and C Paulus and M A S Santos and G R Moura and M Frugier and J Rudinger-Thirion}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30592748?dopt=Abstract}, doi = {10.1371/journal.pone.0209805}, isbn = {30592748}, year = {2018}, date = {2018-01-01}, journal = {PLoS One}, volume = {13}, number = {12}, pages = {e0209805}, abstract = {The life cycle of Plasmodium falciparum, the agent responsible for malaria, depends on both cytosolic and apicoplast translation fidelity. Apicoplast aminoacyl-tRNA synthetases (aaRS) are bacterial-like enzymes devoted to organellar tRNA aminoacylation. They are all encoded by the nuclear genome and are translocated into the apicoplast only after cytosolic biosynthesis. Apicoplast aaRSs contain numerous idiosyncratic sequence insertions: An understanding of the roles of these insertions has remained elusive and they hinder efforts to heterologously overexpress these proteins. Moreover, the A/T rich content of the Plasmodium genome leads to A/U rich apicoplast tRNA substrates that display structural plasticity. Here, we focus on the P. falciparum apicoplast tyrosyl-tRNA synthetase (Pf-apiTyrRS) and its cognate tRNATyr substrate (Pf-apitRNATyr). Cloning and expression strategies used to obtain an active and functional recombinant Pf-apiTyrRS are reported. Functional analyses established that only three weak identity elements in the apitRNATyr promote specific recognition by the cognate Pf-apiTyrRS and that positive identity elements usually found in the tRNATyr acceptor stem are excluded from this set. This finding brings to light an unusual behavior for a tRNATyr aminoacylation system and suggests that Pf-apiTyrRS uses primarily negative recognition elements to direct tyrosylation specificity.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Light-Up RNA Aptamers and Their Cognate Fluorogens: From Their Development to Their Applications}, author = {F Bouhedda and A Autour and M Ryckelynck}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29295531?dopt=Abstract}, doi = {10.3390/ijms19010044}, isbn = {29295531}, year = {2018}, date = {2018-01-01}, journal = {Int J Mol Sci}, volume = {19}, number = {1}, pages = {44}, abstract = {An RNA-based fluorogenic module consists of a light-up RNA aptamer able to specifically interact with a fluorogen to form a fluorescent complex. Over the past decade, significant efforts have been devoted to the development of such modules, which now cover the whole visible spectrum, as well as to their engineering to serve in a wide range of applications. In this review, we summarize the different strategies used to develop each partner (the fluorogen and the light-up RNA aptamer) prior to giving an overview of their applications that range from live-cell RNA imaging to the set-up of high-throughput drug screening pipelines. We then conclude with a critical discussion on the current limitations of these modules and how combining in vitro selection with screening approaches may help develop even better molecules.}, keywords = {RNA biosensing fluorescence fluorogen fluorogenic dye gene expression monitoring in vitro evolution light-up aptamer live-cell imaging, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Phloem-Triggered Virus-Induced Gene Silencing Using a Recombinant Polerovirus}, author = {D Bortolamiol-Bécet and B Monsion and S Chapuis and K Hleibieh and D Scheidecker and A Alioua and F Bogaert and F Revers and V Brault and V Ziegler-Graff}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30405546?dopt=Abstract}, doi = {10.3389/fmicb.2018.02449}, isbn = {30405546}, year = {2018}, date = {2018-01-01}, journal = {Front Microbiol}, volume = {9}, pages = {2449}, abstract = {The phloem-limited poleroviruses infect Arabidopsis thaliana without causing noticeable disease symptoms. In order to facilitate visual infection identification, we developed virus-induced gene silencing (VIGS) vectors derived from Turnip yellows virus (TuYV). Short sequences from the host gene AtCHLI1 required for chlorophyll biosynthesis [42 nucleotides in sense or antisense orientation or as an inverted-repeat (IR), or an 81 nucleotide sense fragment] were inserted into the 3' non-coding region of the TuYV genome to screen for the most efficient and robust silencing vector. All recombinant viruses produced a clear vein chlorosis phenotype on infected Arabidopsis plants due to the expression inhibition of the AtCHLI1 gene. The introduction of a sense-oriented sequence into TuYV genome resulted in a virus exhibiting a more sustainable chlorosis than the virus containing an IR of the same length. This observation was correlated with a higher stability of the sense sequence insertion in the viral genome. In order to evaluate the impact of the TuYV silencing suppressor P0 in the VIGS mechanism a P0 knock-out mutation was introduced into the recombinant TuYV viruses. They induced a similar but milder vein clearing phenotype due to lower viral accumulation. This indicates that P0 does not hinder the performances of the TuYV silencing effect and confirms that in the viral infection context, P0 has no major impact on the production, propagation and action of the short distance silencing signal in phloem cells. Finally, we showed that TuYV can be used to strongly silence the phloem specific AtRTM1 gene. The TuYV-derived VIGS vectors therefore represent powerful tools to easily detect and monitor TuYV in infected plants and conduct functional analysis of phloem-restricted genes. Moreover this example indicates the potential of poleroviruses for use in functional genomic studies of agronomic plants.}, keywords = {PFEFFER, Unité ARN, VIGS phloem polerovirus suppressor of RNA silencing visual monitoring of infection}, pubstate = {published}, tppubtype = {article} } @article{, title = {Fluorogenic RNA Mango aptamers for imaging small non-coding RNAs in mammalian cells}, author = {A Autour and S C Y Jeng and A D Cawte and A Abdolahzadeh and A Galli and SSS. Panchapakesan and D Rueda and M Ryckelynck and P J Unrau}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29440634?dopt=Abstract}, doi = {10.1038/s41467-018-02993-8}, isbn = {29440634}, year = {2018}, date = {2018-01-01}, journal = {Nat Commun}, volume = {9}, number = {1}, pages = {656}, abstract = {Despite having many key roles in cellular biology, directly imaging biologically important RNAs has been hindered by a lack of fluorescent tools equivalent to the fluorescent proteins available to study cellular proteins. Ideal RNA labelling systems must preserve biological function, have photophysical properties similar to existing fluorescent proteins, and be compatible with established live and fixed cell protein labelling strategies. Here, we report a microfluidics-based selection of three new high-affinity RNA Mango fluorogenic aptamers. Two of these are as bright or brighter than enhanced GFP when bound to TO1-Biotin. Furthermore, we show that the new Mangos can accurately image the subcellular localization of three small non-coding RNAs (5S, U6, and a box C/D scaRNA) in fixed and live mammalian cells. These new aptamers have many potential applications to study RNA function and dynamics both in vitro and in mammalian cells.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutant MRPS5 affects mitoribosomal accuracy and confers stress-related behavioral alterations}, author = {R Akbergenov and S Duscha and A K Fritz and R Juskeviciene and N Oishi and K Schmitt and D Shcherbakov and Y Teo and H Boukari and P Freihofer and P Isnard-Petit and B Oettinghaus and S Frank and K Thiam and H Rehrauer and E Westhof and J Schacht and A Eckert and D Wolfer and E C Böttger}, url = {https://www.ncbi.nlm.nih.gov/pubmed/30237157?dopt=Abstract}, doi = {10.15252/embr.201846193}, isbn = {30237157}, year = {2018}, date = {2018-01-01}, journal = {EMBO Rep}, volume = {19}, number = {11}, pages = {e46193}, abstract = {The 1555 A to G substitution in mitochondrial 12S A-site rRNA is associated with maternally transmitted deafness of variable penetrance in the absence of otherwise overt disease. Here, we recapitulate the suggested A1555G-mediated pathomechanism in an experimental model of mitoribosomal mistranslation by directed mutagenesis of mitoribosomal protein MRPS5. We first establish that the ratio of cysteine/methionine incorporation and read-through of mtDNA-encoded MT-CO1 protein constitute reliable measures of mitoribosomal misreading. Next, we demonstrate that human HEK293 cells expressing mutant V336Y MRPS5 show increased mitoribosomal mistranslation. As for immortalized lymphocytes of individuals with the pathogenic A1555G mutation, we find little changes in the transcriptome of mutant V336Y MRPS5 HEK cells, except for a coordinated upregulation of transcripts for cytoplasmic ribosomal proteins. Homozygous knock-in mutant Mrps5 V338Y mice show impaired mitochondrial function and a phenotype composed of enhanced susceptibility to noise-induced hearing damage and anxiety-related behavioral alterations. The experimental data in V338Y mutant mice point to a key role of mitochondrial translation and function in stress-related behavioral and physiological adaptations.}, keywords = {aging disease misreading mitochondria protein synthesis, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{schaeffer_inhibition_2018b, title = {Inhibition of dengue virus infection by mannoside glycolipid conjugates}, author = {Evelyne Schaeffer and Vincent Flacher and Patrick Neuberg and Astrid Hoste and Adrien Brulefert and Jean-Daniel Fauny and Alain Wagner and Christopher G Mueller}, doi = {10.1016/j.antiviral.2018.04.005}, issn = {1872-9096}, year = {2018}, date = {2018-01-01}, journal = {Antiviral Research}, volume = {154}, pages = {116--123}, abstract = {Dengue virus (DENV), a mosquito-borne flavivirus, causes severe and potentially fatal symptoms in millions of infected individuals each year. Although dengue fever represents a major global public health problem, the vaccines or antiviral drugs proposed so far have not shown sufficient efficacy and safety, calling for new antiviral developments. Here we have shown that a mannoside glycolipid conjugate (MGC) bearing a trimannose head with a saturated lipid chain inhibited DENV productive infection. It showed remarkable cell promiscuity, being active in human skin dendritic cells, hepatoma cell lines and Vero cells, and was active against all four DENV serotypes, with an IC50 in the low micromolar range. Time-of-addition experiments and structure-activity analyses revealed the importance of the lipid chain to interfere with an early viral infection step. This, together with a correlation between antiviral activity and membrane polarization by the lipid moiety indicated that the inhibitor functions by blocking viral envelope fusion with the endosome membrane. These finding establish MGCs as a novel class of antivirals against the DENV.}, keywords = {Animals, Antiviral Agents, Cell Line, Cell Membrane, Chemistry, Chlorocebus aethiops, Dendritic Cells, Dengue, Dengue virus, development, Drug, Drug Discovery, Flavivirus, function, Fusion, Glycolipids, Health, Hep G2 Cells, Human, Humans, immunopathology, infection, inhibition, inhibitors, Inhibitory Concentration 50, lipid, Macrophages, Mannosides, Membrane, Serogroup, Skin, Team-Mueller, vaccine, Vaccines, Vero Cells, viral Infection, virus, Virus Replication}, pubstate = {published}, tppubtype = {article} } @article{muller_development_2018, title = {Development of an innervated tissue-engineered skin with human sensory neurons and Schwann cells differentiated from iPS cells}, author = {Quentin Muller and Marie-Josée Beaudet and Thiéry De Serres-Bérard and Sabrina Bellenfant and Vincent Flacher and François Berthod}, doi = {10.1016/j.actbio.2018.10.011}, issn = {1878-7568}, year = {2018}, date = {2018-01-01}, journal = {Acta Biomaterialia}, volume = {82}, pages = {93--101}, abstract = {Cutaneous innervation is increasingly recognized as a major element of skin physiopathology through the neurogenic inflammation driven by neuropeptides that are sensed by endothelial cells and the immune system. To investigate this process in vitro, models of innervated tissue-engineered skin (TES) were developed, yet exclusively with murine sensory neurons extracted from dorsal root ganglions. In order to build a fully human model of innervated TES, we used induced pluripotent stem cells (iPSC) generated from human skin fibroblasts. Nearly 100% of the iPSC differentiated into sensory neurons were shown to express the neuronal markers BRN3A and β3-tubulin after 19 days of maturation. In addition, these cells were also positive to TRPV1 and neurofilament M, and some of them expressed Substance P, TrkA and TRPA1. When stimulated with molecules inducing neuropeptide release, iPSC-derived neurons released Substance P and CGRP, both in conventional monolayer culture and after seeding in a 3D fibroblast-populated collagen sponge model. Schwann cells, the essential partners of neurons for function and axonal migration, were also successfully differentiated from human iPSC as shown by their expression of the markers S100, GFAP, p75 and SOX10. When cultured for one additional month in the TES model, iPSC-derived neurons seeded at the bottom of the sponge formed a network of neurites spanning the whole TES up to the epidermis, but only when combined with mouse or iPSC-derived Schwann cells. This unique model of human innervated TES should be highly useful for the study of cutaneous neuroinflammation. STATEMENT OF SIGNIFICANCE: The purpose of this work was to develop in vitro an innovative fully human tissue-engineered skin enabling the investigation of the influence of cutaneous innervation on skin pathophysiology. To reach that aim, neurons were differentiated from human induced pluripotent stem cells (iPSCs) generated from normal human skin fibroblasts. This innervated tissue-engineered skin model will be the first one to show iPSC-derived neurons can be successfully used to build a 3D nerve network in vitro. Since innervation has been recently recognized to play a central role in many human skin diseases, such as psoriasis and atopic dermatitis, this construct promises to be at the forefront to model these diseases while using patient-derived cells.}, keywords = {atopic dermatitis, Axonal migration, Biological, Canada, Cells, CGRP, Chemistry, COLLAGEN, Culture, Dermatitis, development, disease, Endothelial Cells, ENDOTHELIAL-CELLS, Epidermis, Expression, Fibroblast, Fibroblasts, function, Human, Humans, Immune System, Immunology, immunopathology, IN VITRO, Induced Pluripotent Stem Cells, inflammation, INNERVATION, Maturation, migration, Models, mouse, murine, Nerve, Neurites, Neurogenic Inflammation, Neurons, NEUROPEPTIDE, Neuropeptides, physiopathology, Pluripotent Stem Cells, Psoriasis, SCHWANN CELLS, Sensory Receptor Cells, Skin, skin disease, Skin Diseases, stem, Stem Cells, SUBSTANCE, SUBSTANCE P, Team-Mueller, Tissue Engineering, TRPV1}, pubstate = {published}, tppubtype = {article} } @article{montavon_characterization_2018, title = {Characterization of DCL4 missense alleles provides insights into its ability to process distinct classes of dsRNA substrates.}, author = {Thomas Montavon and Yerim Kwon and Aude Zimmermann and Philippe Hammann and Timothée Vincent and Valérie Cognat and Marc Bergdoll and Fabrice Michel and Patrice Dunoyer}, doi = {10.1111/tpj.13941}, issn = {1365-313X 0960-7412}, year = {2018}, date = {2018-01-01}, journal = {The Plant journal : for cell and molecular biology}, volume = {95}, number = {2}, pages = {204--218}, abstract = {In the model plant Arabidopsis thaliana, four Dicer-like proteins (DCL1-4) mediate the production of various classes of small RNAs (sRNAs). Among these four proteins, DCL4 is by far the most versatile RNaseIII-like enzyme, and previously identified dcl4 missense alleles were shown to uncouple the production of the various classes of DCL4-dependent sRNAs. Yet little is known about the molecular mechanism behind this uncoupling. Here, by studying the subcellular localization, interactome and binding to the sRNA precursors of three distinct dcl4 missense alleles, we simultaneously highlight the absolute requirement of a specific residue in the helicase domain for the efficient production of all DCL4-dependent sRNAs, and identify, within the PAZ domain, an important determinant of DCL4 versatility that is mandatory for the efficient processing of intramolecular fold-back double-stranded RNA (dsRNA) precursors, but that is dispensable for the production of small interfering RNAs (siRNAs) from RDR-dependent dsRNA susbtrates. This study not only provides insights into the DCL4 mode of action, but also delineates interesting tools to further study the complexity of RNA silencing pathways in plants, and possibly other organisms.}, note = {Place: England}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{mueller_molecular_2018, title = {Molecular and Cellular Requirements for the Assembly of Tertiary Lymphoid Structures}, author = {C G Mueller and S Nayar and J Campos and F Barone}, doi = {10.1007/978-3-319-78127-3_4}, issn = {0065-2598}, year = {2018}, date = {2018-01-01}, journal = {Advances in Experimental Medicine and Biology}, volume = {1060}, pages = {55--72}, abstract = {At sites of chronic inflammation, recruited immune cells form structures that resemble secondary lymphoid organs (SLOs). Those are characterized by segregated areas of prevalent T- or B-cell aggregation, differentiation of high endothelial venules (HEVs) and local activation of resident stromal cells. B-cell proliferation and affinity maturation towards locally displayed autoantigens have been demonstrated at those sites, known as tertiary lymphoid structures (TLSs). TLS formation has been associated with local disease persistence and progression as well as increased systemic manifestations. While bearing a similar histological structure to SLO, the signals that regulate TLS and SLO formation can diverge, and a series of pro-inflammatory cytokines has been ascribed as responsible for TLS formation at different anatomical sites. Here we review the structural elements as well as the signals responsible for TLS aggregation, aiming to provide an overview to this complex immunological phenomenon.}, keywords = {Animals, CCL21, CXCL13, Endothelial and stromal cells, Humans, Lymphotoxin, Molecular Targeted Therapy, RANKL, Sjögren’s syndrome, Team-Mueller, Tertiary lymphoid structures, TNF}, pubstate = {published}, tppubtype = {article} } @article{mueller_cellular_2018, title = {Cellular and Vascular Components of Tertiary Lymphoid Structures}, author = {Christopher George Mueller and Saba Nayar and David Gardner and Francesca Barone}, doi = {10.1007/978-1-4939-8709-2_2}, issn = {1940-6029}, year = {2018}, date = {2018-01-01}, journal = {Methods in Molecular Biology (Clifton, N.J.)}, volume = {1845}, pages = {17--30}, abstract = {Inflammatory immune cells recruited at the site of chronic inflammation form structures that resemble secondary lymphoid organs (SLO). These are characterized by segregated areas of prevalent T- or B-cell aggregation, differentiation of high endothelial venules, and local activation of resident stromal cells, including lymphatic endothelial cells. B-cell proliferation and affinity maturation toward locally displayed autoantigens have been demonstrated at these sites, known as tertiary lymphoid structures (TLS). TLS formation during chronic inflammation has been associated with local disease persistence and progression, as well as increased systemic manifestations. While bearing a similar histological structure to SLO, the signals that regulate TLS and SLO formation can diverge and a series of pro-inflammatory cytokines have been ascribed as responsible for TLS formation at different anatomical sites. Moreover, for a long time the structural compartment that regulates TLS homeostasis, including survival and recirculation of leucocytes has been neglected. In this chapter, we summarize the novel data available on TLS formation, structural organization, and the functional and anatomical links connecting TLS and SLOs.}, keywords = {Animals, Biomarkers, CCL21, Cell Survival, Cellular Microenvironment, CXCL13, Cytokines, Humans, Immunity, inflammation, Innate, LYMPHATIC VESSEL, Lymphocyte, Lymphocyte Subsets, Lymphotoxin, Multigene Family, Neovascularization, Pathologic, Receptors, Signal Transduction, Sjögren’s syndrome, Stromal cell, Team-Mueller, Tertiary lymphoid organ, Tertiary lymphoid structures, TNF-α, Tumor Necrosis Factor}, pubstate = {published}, tppubtype = {article} } @article{fadeel_safety_2018, title = {Safety Assessment of Graphene-Based Materials: Focus on Human Health and the Environment}, author = {Bengt Fadeel and Cyrill Bussy and Sonia Merino and Ester Vázquez and Emmanuel Flahaut and Florence Mouchet and Lauris Evariste and Laury Gauthier and Antti J Koivisto and Ulla Vogel and Cristina Martín and Lucia G Delogu and Tina Buerki-Thurnherr and Peter Wick and Didier Beloin-Saint-Pierre and Roland Hischier and Marco Pelin and Fabio Candotto Carniel and Mauro Tretiach and Fabrizia Cesca and Fabio Benfenati and Denis Scaini and Laura Ballerini and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, doi = {10.1021/acsnano.8b04758}, issn = {1936-086X}, year = {2018}, date = {2018-01-01}, journal = {ACS nano}, volume = {12}, number = {11}, pages = {10582--10620}, abstract = {Graphene and its derivatives are heralded as "miracle" materials with manifold applications in different sectors of society from electronics to energy storage to medicine. The increasing exploitation of graphene-based materials (GBMs) necessitates a comprehensive evaluation of the potential impact of these materials on human health and the environment. Here, we discuss synthesis and characterization of GBMs as well as human and environmental hazard assessment of GBMs using in vitro and in vivo model systems with the aim to understand the properties that underlie the biological effects of these materials; not all GBMs are alike, and it is essential that we disentangle the structure-activity relationships for this class of materials.}, keywords = {Animals, carbon nanomaterials, environment, Environmental Monitoring, Exposure, graphene, Graphite, hazard, Health, Humans, I2CT, life cycle assessment, Materials Testing, Nanostructures, Risk Assessment, safety, Structure-Activity Relationship, Team-Bianco, Toxicity}, pubstate = {published}, tppubtype = {article} } @article{ferreira_small_2018, title = {The small non-coding RNA response to virus infection in the Leishmania vector Lutzomyia longipalpis}, author = {Flávia Viana Ferreira and Eric Roberto Guimarães Rocha Aguiar and Roenick Proveti Olmo and Karla Pollyanna Vieira de Oliveira and Emanuele Guimarães Silva and Maurício Roberto Viana Sant'Anna and Nelder Figueiredo de Gontijo and Erna Geessien Kroon and Jean-Luc Imler and João Trindade Marques}, doi = {10.1371/journal.pntd.0006569}, issn = {1935-2735}, year = {2018}, date = {2018-01-01}, journal = {PLoS Negl Trop Dis}, volume = {12}, number = {6}, pages = {e0006569}, abstract = {Sandflies are well known vectors for Leishmania but also transmit a number of arthropod-borne viruses (arboviruses). Few studies have addressed the interaction between sandflies and arboviruses. RNA interference (RNAi) mechanisms utilize small non-coding RNAs to regulate different aspects of host-pathogen interactions. The small interfering RNA (siRNA) pathway is a broad antiviral mechanism in insects. In addition, at least in mosquitoes, another RNAi mechanism mediated by PIWI interacting RNAs (piRNAs) is activated by viral infection. Finally, endogenous microRNAs (miRNA) may also regulate host immune responses. Here, we analyzed the small non-coding RNA response to Vesicular stomatitis virus (VSV) infection in the sandfly Lutzoymia longipalpis. We detected abundant production of virus-derived siRNAs after VSV infection in adult sandflies. However, there was no production of virus-derived piRNAs and only mild changes in the expression of vector miRNAs in response to infection. We also observed abundant production of virus-derived siRNAs against two other viruses in Lutzomyia Lulo cells. Together, our results suggest that the siRNA but not the piRNA pathway mediates an antiviral response in sandflies. In agreement with this hypothesis, pre-treatment of cells with dsRNA against VSV was able to inhibit viral replication while knock-down of the central siRNA component, Argonaute-2, led to increased virus levels. Our work begins to elucidate the role of RNAi mechanisms in the interaction between L. longipalpis and viruses and should also open the way for studies with other sandfly-borne pathogens.}, keywords = {Animals, Host-Pathogen Interactions, imler, Insect Vectors, Leishmania, M3i, ncRNA, Psychodidae, RNA, RNA Interference, Small Interfering, Untranslated, Vesicular stomatitis Indiana virus, Viral}, pubstate = {published}, tppubtype = {article} } @article{kuhn_definition_2017, title = {Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS}, author = {Lauriane Kuhn and Karim Majzoub and Evelyne Einhorn and Johana Chicher and Julien Pompon and Jean-Luc Imler and Philippe Hammann and Carine Meignin}, doi = {10.1534/g3.117.042564}, issn = {2160-1836}, year = {2017}, date = {2017-12-31}, journal = {G3 (Bethesda)}, abstract = {Receptor for Activated C kinase 1 (RACK1) is a scaffold protein that has been found in association with several signaling complexes, and with the 40S subunit of the ribosome. Using the model organism Drosophila melanogaster, we recently showed that RACK1 is required at the ribosome for IRES-mediated translation of viruses. Here, we report a proteomic characterization of the interactome of RACK1 in Drosophila S2 cells. We carried out Label-Free quantitation using both Data-Dependent and Data-Independent Acquisition and observed a significant advantage for the Sequential Window Acquisition of all THeoretical fragment-ion spectra (SWATH) method both in terms of identification of interactants and quantification of low abundance proteins. These data represent the first SWATH spectral library available for Drosophila and will be a useful resource for the community. A total of 52 interacting proteins were identified, including several molecules involved in translation such as structural components of the ribosome, factors regulating translation initiation or elongation and RNA binding proteins. Among these 52 proteins, 15 were identified as partners by the SWATH strategy only. Interestingly, these 15 proteins are significantly enriched for the functions translation and nucleic acid binding. This enrichment reflects the engagement of RACK1 at the ribosome and highlights the added value of SWATH analysis. A functional screen did not reveal any protein sharing the interesting properties of RACK1, which is required for IRES-dependent translation and not essential for cell viability. Intriguingly however, 10 of the RACK1 partners identified restrict replication of Cricket paralysis virus, an IRES-containing virus.}, keywords = {imler, M3i, meignin, PPSE}, pubstate = {published}, tppubtype = {article} } @article{Triller2017, title = {Natural parasite exposure induces protective human anti-malarial antibodies}, author = {Triller G and Scally SW and Costa G and Pissarev M and Kreschel C and Bosch A and Marois E and Sack BK and Murugan R and Salman AM and Janse CJ and Khan SM and Kappe SH and Adegnika AA and Mordmüller B and Levashina EA and Julien JP and Wardemann H}, url = {https://doi.org/10.1016/j.immuni.2017.11.007}, doi = {10.1016/j.immuni.2017.11.007}, year = {2017}, date = {2017-12-19}, journal = {Immunity }, volume = {47}, issue = {6}, pages = {1197-1209}, abstract = {Antibodies against the NANP repeat of circumsporozoite protein (CSP), the major surface antigen of Plasmodium falciparum (Pf) sporozoites, can protect from malaria in animal models but protective humoral immunity is difficult to induce in humans. Here we cloned and characterized rare affinity-matured human NANP-reactive memory B cell antibodies elicited by natural Pf exposure that potently inhibited parasite transmission and development in vivo. We unveiled the molecular details of antibody binding to two distinct protective epitopes within the NANP repeat. NANP repeat recognition was largely mediated by germline encoded and immunoglobulin (Ig) heavy-chain complementarity determining region 3 (HCDR3) residues, whereas affinity maturation contributed predominantly to stabilizing the antigen-binding site conformation. Combined, our findings illustrate the power of exploring human anti-CSP antibody responses to develop tools for malaria control in the mammalian and the mosquito vector and provide a molecular basis for the structure-based design of next-generation CSP malaria vaccines. }, keywords = {anti-malarial antibodie, M3i, marois}, pubstate = {published}, tppubtype = {article} } @article{Jelena2017, title = {The serine protease homolog spheroide is involved in sensing of pathogenic Gram-positive bacteria}, author = {Jelena Patrnogic and Vincent Leclerc}, url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5718610/}, doi = {10.1371/journal.pone.0188339}, year = {2017}, date = {2017-12-06}, journal = {PLoS One}, volume = {12}, number = {12}, abstract = {In Drosophila, recognition of pathogens such as Gram-positive bacteria and fungi triggers the activation of proteolytic cascades and the subsequent activation of the Toll pathway. This response can be achieved by either detection of pathogen associated molecular patterns or by sensing microbial proteolytic activities ("danger signals"). Previous data suggested that certain serine protease homologs (serine protease folds that lack an active catalytic triad) could be involved in the pathway. We generated a null mutant of the serine protease homolog spheroide (sphe). These mutant flies are susceptible to Enterococcus faecalis infection and unable to fully activate the Toll pathway. Sphe is required to activate the Toll pathway after challenge with pathogenic Gram-Positive bacteria. Sphe functions in the danger signal pathway, downstream or at the level of Persephone.}, keywords = {Fungi, Gram-Positive Bacteria, serine protease, spheroide, toll pathway}, pubstate = {published}, tppubtype = {article} } @article{pmid28687465, title = {On the way to find a cure: Purging latent HIV-1 reservoirs}, author = {Christian Schwartz and Sophie Bouchat and Céline Marban and Virginie Gautier and Carine Van Lint and Olivier Rohr and Valentin Le Douce}, doi = {10.1016/j.bcp.2017.07.001}, issn = {1873-2968}, year = {2017}, date = {2017-12-01}, urldate = {2017-12-01}, journal = {Biochem Pharmacol}, volume = {146}, pages = {10--22}, abstract = {Introduction of cART in 1996 has drastically increased the life expectancy of people living with HIV-1. However, this treatment has not allowed cure as cessation of cART is associated with a rapid viral rebound. The main barrier to the eradication of the virus is related to the persistence of latent HIV reservoirs. Evidence is now accumulating that purging the HIV-1 reservoir might lead to a cure or a remission. The most studied strategy is the so called "shock and kill" therapy. This strategy is based on reactivation of dormant viruses from the latently-infected reservoirs (the shock) followed by the eradication of the reservoirs (the kill). This review focuses mainly on the recent advances made in the "shock and kill" therapy. We believe that a cure or a remission will come from combinatorial approaches i.e. combination of drugs to reactivate the dormant virus from all the reservoirs including the one located in sanctuaries, and combination of strategies boosting the immune system. Alternative strategies based on cell and gene therapy or based in inducing deep latency, which are evoked in this review reinforce the idea that at least a remission is attainable.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{kurapati_covalent_2017, title = {Covalent chemical functionalization enhances the biodegradation of graphene oxide}, author = {Rajendra Kurapati and Fanny Bonachera and Julie Russier and Adukamparai Rajukrishnan Sureshbabu and Cécilia Ménard-Moyon and Kostas Kostarelos and Alberto Bianco}, url = {https://doi.org/10.1088%2F2053-1583%2Faa8f0a}, doi = {10.1088/2053-1583/aa8f0a}, issn = {2053-1583}, year = {2017}, date = {2017-11-01}, urldate = {2020-04-01}, journal = {2D Materials}, volume = {5}, number = {1}, pages = {015020}, abstract = {Biodegradation of the graphene-based materials is an emerging issue due to their estimated widespread usage in different industries. Indeed, a few concerns have been raised about their biopersistence. Here, we propose the design of surface-functionalized graphene oxide (GO) with the capacity to degrade more effectively compared to unmodified GO using horseradish peroxidase (HRP). For this purpose, we have functionalized the surface of GO with two well-known substrates of HRP namely coumarin and catechol. The biodegradation of all conjugates has been followed by Raman, dynamic light scattering and electron microscopy. Molecular docking and gel electrophoresis have been carried out to gain more insights into the interaction between GO conjugates and HRP. Our studies have revealed better binding when GO is functionalized with coumarin or catechol compared to control GOs. All results prove that GO functionalized with coumarin and catechol moieties display a faster and more efficient biodegradation over GO.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{chau_elucidation_2017, title = {Elucidation of siRNA complexation efficiency by graphene oxide and reduced graphene oxide}, author = {Ngoc Do Quyen Chau and Giacomo Reina and Jésus Raya and Isabella Anna Vacchi and Cécilia Ménard-Moyon and Yuta Nishina and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S0008622317306942}, doi = {10.1016/j.carbon.2017.07.016}, issn = {0008-6223}, year = {2017}, date = {2017-10-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {122}, pages = {643--652}, abstract = {Gene therapy has attracted tremendous attention as a promising method for the treatment of many diseases. Graphene oxide (GO), the oxidized form of graphene, is showing lot of potential in this field. Indeed, the polar GO oxygenated functional groups make this material highly hydrophilic, leading to a good dispersibility in water. Until now, the interaction of small interference RNA (siRNA) with graphene materials has not been elucidated. The main goal of this work was to develop a novel platform to complex siRNA molecules and to rationalize the supramolecular interactions between GO surface and the double strand RNA. Our study focused first on green and facile methods such as hydrothermal or vitamin C treatments to prepare graphene oxides at various percentage of oxygen. Epoxidation was also explored to reintroduce epoxide groups on reduced GO for further functionalization. Subsequently, we performed the covalent functionalization of GO with triethyleneglycoldiamine (TEG) and with low molecular weight polyethyleneimine (PEI) via the epoxy ring opening reaction. Finally, by gel electrophoresis, we were able to correlate the GO complexation ability with the graphene surface chemistry. In addition, ozonated GO functionalized with PEI showed a high complexing capacity for siRNA, proving to be a promising candidate for gene silencing.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{chypre_porphyrin_2017, title = {Porphyrin Derivatives Inhibit the Interaction between Receptor Activator of NF-κB and Its Ligand}, author = {Mélanie Chypre and Maria-Bernadette Madel and Olivier Chaloin and Claudine Blin-Wakkach and Christophe Morice and Christopher G Mueller}, doi = {10.1002/cmdc.201700462}, issn = {1860-7187}, year = {2017}, date = {2017-10-01}, journal = {ChemMedChem}, volume = {12}, number = {20}, pages = {1697--1702}, abstract = {Receptor activator of NF-κB (RANK), a member of the TNF-receptor superfamily, plays an important role in bone resorption and stimulates immune and epithelial cell activation. Denosumab, a human monoclonal antibody that blocks the RANK ligand (RANKL), is approved for the treatment of osteoporosis and bone metastasis. However, a small molecule that inhibits the RANK-RANKL interaction would be beneficial to decrease cost and to facilitate treatments with orally available therapeutic agents. Herein we report the discovery of the first nonpeptidic inhibitors of RANK-RANKL interactions. In screening a chemical library by competitive ELISA, the porphyrin verteporfin was identified as a hit. Derivatives were screened, and the chlorin-macrocycle-containing pheophorbide A and purpurin 18 were found to bind recombinant RANKL, to inhibit RANK-RANKL interactions in the ELISA, and to suppress the RANKL-dependent activation of model cells and the differentiation of RANK-expressing precursors into osteoclasts. This discovery of a family of small molecules that inhibit RANK activation presents an initial basis for further development of nonpeptidic therapeutic agents targeting the interaction between RANK and RANKL.}, keywords = {Animals, Cell Survival, cell-based assays, ELISA, Humans, Jurkat Cells, Mice, Molecular Structure, Osteoclasts, Osteogenesis, porphyrins, Protein Binding, RANK ligand, receptor activator of NF-κB, Receptor Activator of Nuclear Factor-kappa B, Structure-Activity Relationship, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{Gross_2017, title = {The IRES5'UTR of the dicistrovirus cricket paralysis virus is a type III IRES containing an essential pseudoknot structure}, author = {E Gross and Q Vincens and E Einhorn and A Noireterre and L Schaeffer and L Kuhn and JL Imler and Eriani and C Meignin and F Martin}, url = {https://academic.oup.com/nar/article/45/15/8993/3978035}, doi = {10.1093/nar/gkx622 }, year = {2017}, date = {2017-09-06}, journal = {Nucleic Acids Research}, volume = {45}, number = {15}, pages = {8993-9004}, abstract = {Cricket paralysis virus (CrPV) is a dicistrovirus. Its positive-sense single-stranded RNA genome contains two internal ribosomal entry sites (IRESs). The 5' untranslated region (5'UTR) IRES5'UTR mediates translation of non-structural proteins encoded by ORF1 whereas the well-known intergenic region (IGR) IRESIGR is required for translation of structural proteins from open reading frame 2 in the late phase of infection. Concerted action of both IRES is essential for host translation shut-off and viral translation. IRESIGR has been extensively studied, in contrast the IRES5'UTR remains largely unexplored. Here, we define the minimal IRES element required for efficient translation initiation in drosophila S2 cell-free extracts. We show that IRES5'UTR promotes direct recruitment of the ribosome on the cognate viral AUG start codon without any scanning step, using a Hepatitis-C virus-related translation initiation mechanism. Mass spectrometry analysis revealed that IRES5'UTR recruits eukaryotic initiation factor 3, confirming that it belongs to type III class of IRES elements. Using Selective 2'-hydroxyl acylation analyzed by primer extension and DMS probing, we established a secondary structure model of 5'UTR and of the minimal IRES5'UTR. The IRES5'UTR contains a pseudoknot structure that is essential for proper folding and ribosome recruitment. Overall, our results pave the way for studies addressing the synergy and interplay between the two IRES from CrPV. }, keywords = {imler, IRES, M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{onder_lymphatic_2017, title = {Lymphatic Endothelial Cells Control Initiation of Lymph Node Organogenesis}, author = {Lucas Onder and Urs Mörbe and Natalia Pikor and Mario Novkovic and Hung-Wei Cheng and Thomas Hehlgans and Klaus Pfeffer and Burkhard Becher and Ari Waisman and Thomas Rülicke and Jennifer Gommerman and Christopher G Mueller and Shinichiro Sawa and Elke Scandella and Burkhard Ludewig}, doi = {10.1016/j.immuni.2017.05.008}, issn = {1097-4180}, year = {2017}, date = {2017-07-01}, journal = {Immunity}, volume = {47}, number = {1}, pages = {80--92.e4}, abstract = {Lymph nodes (LNs) are strategically situated throughout the body at junctures of the blood vascular and lymphatic systems to direct immune responses against antigens draining from peripheral tissues. The current paradigm describes LN development as a programmed process that is governed through the interaction between mesenchymal lymphoid tissue organizer (LTo) cells and hematopoietic lymphoid tissue inducer (LTi) cells. Using cell-type-specific ablation of key molecules involved in lymphoid organogenesis, we found that initiation of LN development is dependent on LTi-cell-mediated activation of lymphatic endothelial cells (LECs) and that engagement of mesenchymal stromal cells is a succeeding event. LEC activation was mediated mainly by signaling through receptor activator of NF-κB (RANK) and the non-canonical NF-κB pathway and was steered by sphingosine-1-phosphate-receptor-dependent retention of LTi cells in the LN anlage. Finally, the finding that pharmacologically enforced interaction between LTi cells and LECs promotes ectopic LN formation underscores the central LTo function of LECs.}, keywords = {Animals, Cell Differentiation, Cells, Choristoma, Cultured, Embryo, Endothelial Cells, fibroblastic reticular cells, Inbred C57BL, lymph node organogenesis, Lymph Nodes, lymphatic and blood endothelial cells, lymphoid stromal cells, lymphoid tissue organizer cells, Lymphotoxin beta Receptor, Lysosphingolipid, Mammalian, Mesenchymal Stem Cells, mesenchymal stromal cells, Mice, NF-kappa B, Organogenesis, Receptor Activator of Nuclear Factor-kappa B, Receptors, Signal Transduction, Team-Mueller, transgenic}, pubstate = {published}, tppubtype = {article} } @article{FM2017, title = {Complex Coding and Regulatory Polymorphisms in a Restriction Factor Determine the Susceptibility of Drosophila to Viral Infection}, author = {Cao C Cogni R Barbier V Jiggins FM}, url = {https://academic.oup.com/genetics/article/206/4/2159/6072650}, doi = {10.1534/genetics.117.201970}, year = {2017}, date = {2017-06-19}, journal = {Genetics}, volume = {206}, pages = {2159-2173}, abstract = {It is common to find that major-effect genes are an important cause of variation in susceptibility to infection. Here we have characterized natural variation in a gene called pastrel that explains over half of the genetic variance in susceptibility to the Drosophila C virus (DCV) in populations of Drosophila melanogaster. We found extensive allelic heterogeneity, with a sample of seven alleles of pastrel from around the world conferring four phenotypically distinct levels of resistance. By modifying candidate SNPs in transgenic flies, we show that the largest effect is caused by an amino acid polymorphism that arose when an ancestral threonine was mutated to alanine, greatly increasing resistance to DCV. Overexpression of the ancestral, susceptible allele provides strong protection against DCV; indicating that this mutation acted to improve an existing restriction factor. The pastrel locus also contains complex structural variation and cis-regulatory polymorphisms altering gene expression. We find that higher expression of pastrel is associated with increased survival after DCV infection. To understand why this variation is maintained in populations, we investigated genetic variation surrounding the amino acid variant that is causing flies to be resistant. We found no evidence of natural selection causing either recent changes in allele frequency or geographical variation in frequency, suggesting that this is an old polymorphism that has been maintained at a stable frequency. Overall, our data demonstrate how complex genetic variation at a single locus can control susceptibility to a virulent natural pathogen.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{Autour2017, title = {Ultrahigh-Throughput Improvement and Discovery of Enzymes Using Droplet-Based Microfluidic Screening}, author = {A Autour and M Ryckelynck}, url = {https://doi.org/10.3390/mi8040128}, doi = {10.3390/mi8040128}, year = {2017}, date = {2017-04-18}, journal = {Micromachines}, volume = {8}, number = {4}, pages = {128}, abstract = {Enzymes are extremely valuable tools for industrial, environmental, and biotechnological applications and there is a constant need for improving existing biological catalysts and for discovering new ones. Screening microbe or gene libraries is an efficient way of identifying new enzymes. In this view, droplet-based microfluidics appears to be one of the most powerful approaches as it allows inexpensive screenings in well-controlled conditions and an ultrahigh-throughput regime. This review aims to introduce the main microfluidic devices and concepts to be considered for such screening before presenting and discussing the latest successful applications of the technology for enzyme discovery. }, keywords = {directed evolution, droplet-based microfluidics, enzyme improvement, high-throughput screening, RYCKELYNCK, single-cell, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{gies_identification_2017, title = {Identification of autoreactive B cells with labeled nucleosomes}, author = {Vincent Gies and Alain Wagner and Cécile Seifert and Aurélien Guffroy and Jean-D. Fauny and Anne-M. Knapp and Jean-L. Pasquali and Thierry Martin and Hélène Dumortier and Anne-S. Korganow and Pauline Soulas-Sprauel}, doi = {10.1038/s41598-017-00664-0}, issn = {2045-2322}, year = {2017}, date = {2017-04-01}, journal = {Scientific Reports}, volume = {7}, number = {1}, pages = {602}, abstract = {The pathogenesis of autoimmune diseases has not been completely elucidated yet, and only a few specific treatments have been developed so far. In autoimmune diseases mediated by pathogenic autoantibodies, such as systemic lupus erythematosus, the specific detection and analysis of autoreactive B cells is crucial for a better understanding of the physiopathology. Biological characterization of these cells may help to define new therapeutic targets. Very few techniques allowing the precise detection of autoreactive B cells have been described so far. Herein we propose a new flow cytometry technique for specific detection of anti-nucleosome B cells, which secrete autoantibodies in systemic lupus erythematosus, using labeled nucleosomes. We produced different fluorochrome-labeled nucleosomes, characterized them, and finally tested them in flow cytometry. Nucleosomes labeled via the cysteines present in H3 histone specifically bind to autoreactive B cells in the anti-DNA transgenic B6.56R mice model. The present work validates the use of fluorochrome-labeled nucleosomes via cysteines to identify anti-nucleosome B cells and offers new opportunities for the description of autoreactive B cell phenotype.}, keywords = {Dumortier, I2CT, Imagerie, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{pmid28256531, title = {Repression of Human T-lymphotropic virus type 1 Long Terminal Repeat sense transcription by Sp1 recruitment to novel Sp1 binding sites}, author = {Sylvain Fauquenoy and Gwenaëlle Robette and Anna Kula and Caroline Vanhulle and Sophie Bouchat and Nadège Delacourt and Anthony Rodari and Céline Marban and Christian Schwartz and Arsène Burny and Olivier Rohr and Benoit Van Driessche and Carine Van Lint}, doi = {10.1038/srep43221}, issn = {2045-2322}, year = {2017}, date = {2017-03-01}, urldate = {2017-03-01}, journal = {Sci Rep}, volume = {7}, pages = {43221}, abstract = {Human T-lymphotropic Virus type 1 (HTLV-1) infection is characterized by viral latency in the majority of infected cells and by the absence of viremia. These features are thought to be due to the repression of viral sense transcription in vivo. Here, our in silico analysis of the HTLV-1 Long Terminal Repeat (LTR) promoter nucleotide sequence revealed, in addition to the four Sp1 binding sites previously identified, the presence of two additional potential Sp1 sites within the R region. We demonstrated that the Sp1 and Sp3 transcription factors bound in vitro to these two sites and compared the binding affinity for Sp1 of all six different HTLV-1 Sp1 sites. By chromatin immunoprecipitation experiments, we showed Sp1 recruitment in vivo to the newly identified Sp1 sites. We demonstrated in the nucleosomal context of an episomal reporter vector that the Sp1 sites interfered with both the sense and antisense LTR promoter activities. Interestingly, the Sp1 sites exhibited together a repressor effect on the LTR sense transcriptional activity but had no effect on the LTR antisense activity. Thus, our results demonstrate the presence of two new functional Sp1 binding sites in the HTLV-1 LTR, which act as negative cis-regulatory elements of sense viral transcription.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{gies_b_2017, title = {B cells differentiate in human thymus and express AIRE}, author = {Vincent Gies and Aurélien Guffroy and François Danion and Philippe Billaud and Céline Keime and Jean-Daniel Fauny and Sandrine Susini and Anne Soley and Thierry Martin and Jean-Louis Pasquali and Frédéric Gros and Isabelle André-Schmutz and Pauline Soulas-Sprauel and Anne-Sophie Korganow}, doi = {10.1016/j.jaci.2016.09.044}, issn = {1097-6825}, year = {2017}, date = {2017-03-01}, journal = {The Journal of Allergy and Clinical Immunology}, volume = {139}, number = {3}, pages = {1049--1052.e12}, keywords = {I2CT, Imagerie}, pubstate = {published}, tppubtype = {article} } @article{gies_identification_2017b, title = {Identification of autoreactive B cells with labeled nucleosomes}, author = {Vincent Gies and Alain Wagner and Cécile Seifert and Aurélien Guffroy and Jean-D. Fauny and Anne-M. Knapp and Jean-L. Pasquali and Thierry Martin and Hélène Dumortier and Anne-S. Korganow and Pauline Soulas-Sprauel}, doi = {10.1038/s41598-017-00664-0}, issn = {2045-2322}, year = {2017}, date = {2017-01-01}, journal = {Scientific Reports}, volume = {7}, number = {1}, pages = {602}, abstract = {The pathogenesis of autoimmune diseases has not been completely elucidated yet, and only a few specific treatments have been developed so far. In autoimmune diseases mediated by pathogenic autoantibodies, such as systemic lupus erythematosus, the specific detection and analysis of autoreactive B cells is crucial for a better understanding of the physiopathology. Biological characterization of these cells may help to define new therapeutic targets. Very few techniques allowing the precise detection of autoreactive B cells have been described so far. Herein we propose a new flow cytometry technique for specific detection of anti-nucleosome B cells, which secrete autoantibodies in systemic lupus erythematosus, using labeled nucleosomes. We produced different fluorochrome-labeled nucleosomes, characterized them, and finally tested them in flow cytometry. Nucleosomes labeled via the cysteines present in H3 histone specifically bind to autoreactive B cells in the anti-DNA transgenic B6.56R mice model. The present work validates the use of fluorochrome-labeled nucleosomes via cysteines to identify anti-nucleosome B cells and offers new opportunities for the description of autoreactive B cell phenotype.}, keywords = {Animals, Autoantibodies, Autoantigens, Autoimmunity, B-Lymphocytes, Biomarkers, Cell Line, Dumortier, Female, Flow Cytometry, Humans, I2CT, Lupus Erythematosus, Mice, Nucleosomes, Staining and Labeling, Systemic, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{kurapati_enzymatic_2017, title = {Enzymatic Biodegradability of Pristine and Functionalized Transition Metal Dichalcogenide MoS2 Nanosheets}, author = {Rajendra Kurapati and Laura Muzi and Aritz Perez Ruiz de Garibay and Julie Russier and Damien Voiry and Isabella A Vacchi and Manish Chhowalla and Alberto Bianco}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adfm.201605176}, doi = {10.1002/adfm.201605176}, issn = {1616-3028}, year = {2017}, date = {2017-01-01}, urldate = {2020-04-01}, journal = {Advanced Functional Materials}, volume = {27}, number = {7}, pages = {1605176}, abstract = {2D transition metal dichalcogenide MoS2 nanosheets are increasingly attracting interests due to their promising applications in materials science and biomedicine. However, their biocompatibility and their biodegradability have not been thoroughly studied yet. Here, the biodegradability of exfoliated pristine and covalently functionalized MoS2 (f-MoS2) is investigated. First, biodegradability of these nanomaterials is evaluated using plant horseradish peroxidase and human myeloperoxidase. The results reveal that the enzymatic degradability rate of MoS2 and f-MoS2 is slower than in the case of the simple treatment with H2O2 alone. In parallel, high biocompatibility of both pristine and f-MoS2 nanosheets is found up to 100 µg mL−1 in both cell lines (HeLa and Raw264.7) and primary immune cells. In addition, no immune cell activation and minimal pro-inflammatory cytokine release are observed in RAW264.7 and human monocyte-derived macrophages, suggesting a negligible cellular impact of such materials. Furthermore, the effects of degraded MoS2 and partially degraded f-MoS2 products on cell viability and activation are studied in cancer and immune cells. A certain cytotoxicity is measured at the highest concentrations. Finally, to prove that the cellular impact is due to cell uptake, the internalization of both pristine and functionalized MoS2 in cancer and primary immune cells is assessed.}, keywords = {Cytotoxicity, degradation, graphene-related materials, I2CT, molybdenum disulfide, peroxidases, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{kuhn_definition_2017b, title = {Definition of a RACK1 Interaction Network in Drosophila melanogaster Using SWATH-MS}, author = {Lauriane Kuhn and Karim Majzoub and Evelyne Einhorn and Johana Chicher and Julien Pompon and Jean-Luc Imler and Philippe Hammann and Carine Meignin}, doi = {10.1534/g3.117.042564}, issn = {2160-1836}, year = {2017}, date = {2017-01-01}, urldate = {2017-01-01}, journal = {G3 (Bethesda)}, abstract = {Receptor for Activated C kinase 1 (RACK1) is a scaffold protein that has been found in association with several signaling complexes, and with the 40S subunit of the ribosome. Using the model organism Drosophila melanogaster, we recently showed that RACK1 is required at the ribosome for IRES-mediated translation of viruses. Here, we report a proteomic characterization of the interactome of RACK1 in Drosophila S2 cells. We carried out Label-Free quantitation using both Data-Dependent and Data-Independent Acquisition and observed a significant advantage for the Sequential Window Acquisition of all THeoretical fragment-ion spectra (SWATH) method both in terms of identification of interactants and quantification of low abundance proteins. These data represent the first SWATH spectral library available for Drosophila and will be a useful resource for the community. A total of 52 interacting proteins were identified, including several molecules involved in translation such as structural components of the ribosome, factors regulating translation initiation or elongation and RNA binding proteins. Among these 52 proteins, 15 were identified as partners by the SWATH strategy only. Interestingly, these 15 proteins are significantly enriched for the functions translation and nucleic acid binding. This enrichment reflects the engagement of RACK1 at the ribosome and highlights the added value of SWATH analysis. A functional screen did not reveal any protein sharing the interesting properties of RACK1, which is required for IRES-dependent translation and not essential for cell viability. Intriguingly however, 10 of the RACK1 partners identified restrict replication of Cricket paralysis virus, an IRES-containing virus.}, keywords = {meignin, PPSE}, pubstate = {published}, tppubtype = {article} } @article{gross_ires5utr_2017, title = {The IRES5'UTR of the dicistrovirus cricket paralysis virus is a type III IRES containing an essential pseudoknot structure}, author = {Lauriane Gross and Quentin Vicens and Evelyne Einhorn and Audrey Noireterre and Laure Schaeffer and Lauriane Kuhn and Jean-Luc Imler and Gilbert Eriani and Carine Meignin and Franck Martin}, doi = {10.1093/nar/gkx622}, issn = {1362-4962}, year = {2017}, date = {2017-01-01}, urldate = {2017-01-01}, journal = {Nucleic Acids Research}, volume = {45}, number = {15}, pages = {8993--9004}, abstract = {Cricket paralysis virus (CrPV) is a dicistrovirus. Its positive-sense single-stranded RNA genome contains two internal ribosomal entry sites (IRESs). The 5' untranslated region (5'UTR) IRES5'UTR mediates translation of non-structural proteins encoded by ORF1 whereas the well-known intergenic region (IGR) IRESIGR is required for translation of structural proteins from open reading frame 2 in the late phase of infection. Concerted action of both IRES is essential for host translation shut-off and viral translation. IRESIGR has been extensively studied, in contrast the IRES5'UTR remains largely unexplored. Here, we define the minimal IRES element required for efficient translation initiation in drosophila S2 cell-free extracts. We show that IRES5'UTR promotes direct recruitment of the ribosome on the cognate viral AUG start codon without any scanning step, using a Hepatitis-C virus-related translation initiation mechanism. Mass spectrometry analysis revealed that IRES5'UTR recruits eukaryotic initiation factor 3, confirming that it belongs to type III class of IRES elements. Using Selective 2'-hydroxyl acylation analyzed by primer extension and DMS probing, we established a secondary structure model of 5'UTR and of the minimal IRES5'UTR. The IRES5'UTR contains a pseudoknot structure that is essential for proper folding and ribosome recruitment. Overall, our results pave the way for studies addressing the synergy and interplay between the two IRES from CrPV.}, keywords = {meignin, PPSE}, pubstate = {published}, tppubtype = {article} } @article{nokey, title = {RNAcentral: a comprehensive database of non-coding RNA sequences}, author = {A. I. Petrov and S. J. E. Kay and I. Kalvari and K. L. Howe and K. A. Gray and E. A. Bruford and P. J. Kersey and G. Cochrane and R. D. Finn and A. Bateman and A. Kozomara and S. Griffiths-Jones and A. Frankish and C. W. Zwieb and B. Y. Lau and K. P. Williams and P. P. Chan and T. M. Lowe and J. J. Cannone and R. Gutell and M. A. Machnicka and J. M. Bujnicki and M. Yoshihama and N. Kenmochi and B. Chai and J. R. Cole and M. Szymanski and W. M. Karlowski and V. Wood and E. Huala and T. Z. Berardini and Y. Zhao and R. Chen and W. Zhu and M. D. Paraskevopoulou and I. S. Vlachos and A. G. Hatzigeorgiou and L. Ma and Z. Zhang and J. Puetz and P. F. Stadler and D. McDonald and S. Basu and P. Fey and S. R. Engel and J. M. Cherry and P. J. Volders and P. Mestdagh and J. Wower and M. B. Clark and X. C. Quek and M. E. Dinger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=27794554}, doi = {10.1093/nar/gkw1008}, isbn = {27794554}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {D1}, pages = {D128-D134}, abstract = {RNAcentral is a database of non-coding RNA (ncRNA) sequences that aggregates data from specialised ncRNA resources and provides a single entry point for accessing ncRNA sequences of all ncRNA types from all organisms. Since its launch in 2014, RNAcentral has integrated twelve new resources, taking the total number of collaborating database to 22, and began importing new types of data, such as modified nucleotides from MODOMICS and PDB. We created new species-specific identifiers that refer to unique RNA sequences within a context of single species. The website has been subject to continuous improvements focusing on text and sequence similarity searches as well as genome browsing functionality. All RNAcentral data is provided for free and is available for browsing, bulk downloads, and programmatic access at http://rnacentral.org/.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Animals *Databases, Nucleic Acid Genomics Humans Nucleotides/chemistry RNA, RNA Species Specificity, Unité ARN, Untranslated/*chemistry Sequence Analysis}, pubstate = {published}, tppubtype = {article} } @article{carapito_two_2017, title = {Two proteomic methodologies for defining N-termini of mature human mitochondrial aminoacyl-tRNA synthetases.}, author = {Christine Carapito and Lauriane Kuhn and Loukmane Karim and Magali Rompais and Thierry Rabilloud and Hagen Schwenzer and Marie Sissler}, doi = {10.1016/j.ymeth.2016.10.012}, issn = {1095-9130 1046-2023}, year = {2017}, date = {2017-01-01}, journal = {Methods (San Diego, Calif.)}, volume = {113}, pages = {111--119}, abstract = {Human mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs) are encoded in the nucleus, synthesized in the cytosol and targeted for importation into mitochondria by a N-terminal mitochondrial targeting sequence. This targeting sequence is presumably cleaved upon entry into the mitochondria, following a process still not fully deciphered in human, despite essential roles for the mitochondrial biogenesis. Maturation processes are indeed essential both for the release of a functional enzyme and to route correctly the protein within mitochondria. The absence of consensus sequences for cleavage sites and the discovery of possible multiple proteolytic steps render predictions of N-termini difficult. Further, the knowledge of the cleavages is key for the design of protein constructions compatible with efficient production in bacterial strains. Finally, full comprehension becomes essential because a growing number of mutations are found in genes coding for mt-aaRS. In the present study, we take advantage of proteomic methodological developments and identified, in mitochondria, three N-termini for the human mitochondrial aspartyl-tRNA synthetase. This first description of the co-existence of different forms opens new perspectives in the biological understanding of this enzyme. Those methods are extended to the whole set of human mt-aaRSs and methodological advice are provided for further investigations.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {Native ESI Mass Spectrometry Can Help to Avoid Wrong Interpretations from Isothermal Titration Calorimetry in Difficult Situations.}, author = {P Wolff and C Da Veiga and E Ennifar and G Bec and G Guichard and D Burnouf and P Dumas}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27957716?dopt=Abstract}, doi = {10.1007/s13361-016-1534-6}, isbn = {27957716}, year = {2017}, date = {2017-01-01}, journal = {J Am Soc Mass Spectrom}, volume = {28}, number = {2}, pages = {347-357}, abstract = {We studied by native ESI-MS the binding of various DNA-polymerase-derived peptides onto DNA-polymerase processivity rings from Escherichia coli, Pseudomonas aeruginosa, and Mycobacterium tuberculosis. These homodimeric rings present two equivalent specific binding sites, which leads to successive formation during a titration experiment of singly- and doubly occupied rings. By using the ESI-MS free-ring spectrum as a ruler, we derived by robust linear regression the fractions of the different ring species at each step of a titration experiment. These results led to accurate Kd values (from 0.03 to 0.5 μM) along with the probability of peptide loss due to gas phase dissociation (GPD). We show that this good quality is due to the increased information content of a titration experiment with a homodimer. Isothermal titration calorimetry (ITC) led with the same binding model to Kd(ITC) values systematically higher than their ESI-MS counterparts and, often, to poor fit of the ITC curves. A processing with two competing modes of binding on the same site requiring determination of two (Kd, ΔH) pairs greatly improved the fits and yielded a second Kd(ITC) close to Kd(ESI-MS). The striking features are: (1) ITC detected a minor binding mode (~20%) of 'low-affinity' that did not appear with ESI-MS; (2) the simplest processing of ITC data with only one (Kd, ΔH) pair led wrongly to the Kd of the low-affinity binding mode but to the ΔH of the high-affinity binding mode. Analogous misleading results might well exist in published data based on ITC experiments. Graphical Abstract ᅟ.}, keywords = {ARN-MS, DNA-polymerase processivity rings ESI-MS ITC Kd determination, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Embraced by eIF3: structural and functional insights into the roles of eIF3 across the translation cycle}, author = {L S Valasek and J Zeman and S Wagner and P Beznoskova and Z Pavlíkova and M P Mohammad and V Hronova and A Herrmannova and Y Hashem and S Gunisova}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28981723?dopt=Abstract}, doi = {10.1093/nar/gkx805}, isbn = {28981723}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {19}, pages = {10948-10968}, abstract = {Protein synthesis is mediated via numerous molecules including the ribosome, mRNA, tRNAs, as well as translation initiation, elongation and release factors. Some of these factors play several roles throughout the entire process to ensure proper assembly of the preinitiation complex on the right mRNA, accurate selection of the initiation codon, errorless production of the encoded polypeptide and its proper termination. Perhaps, the most intriguing of these multitasking factors is the eukaryotic initiation factor eIF3. Recent evidence strongly suggests that this factor, which coordinates the progress of most of the initiation steps, does not come off the initiation complex upon subunit joining, but instead it remains bound to 80S ribosomes and gradually falls off during the first few elongation cycles to: (1) promote resumption of scanning on the same mRNA molecule for reinitiation downstream-in case of translation of upstream ORFs short enough to preserve eIF3 bound; or (2) come back during termination on long ORFs to fine tune its fidelity or, if signaled, promote programmed stop codon readthrough. Here, we unite recent structural views of the eIF3-40S complex and discus all known eIF3 roles to provide a broad picture of the eIF3's impact on translational control in eukaryotic cells.}, keywords = {HASHEM, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The RNA targetome of Staphylococcus aureus non-coding RNA RsaA: impact on cell surface properties and defense mechanisms.}, author = {A Tomasini and K Moreau and J Chicher and T Geissmann and F Vandenesch and P Romby and S Marzi and I Caldelari}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28379505}, doi = {10.1093/nar/gkx219}, isbn = {28379505}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {11}, pages = {6746-6760}, abstract = {The virulon of Staphyloccocus aureus is controlled by intricate connections between transcriptional and post-transcriptional regulators including proteins and small non-coding RNAs (sRNAs). Many of the sRNAs regulate gene expression through base-pairings with mRNAs. However, characterization of the direct sRNA targets in Gram-positive bacteria remained a difficult challenge. Here, we have applied and adapted the MS2-affinity purification approach coupled to RNA sequencing (MAPS) to determine the targetome of RsaA sRNA of S. aureus, known to repress the synthesis of the transcriptional regulator MgrA. Several mRNAs were enriched with RsaA expanding its regulatory network. Besides mgrA, several of these mRNAs encode a family of SsaA-like enzymes involved in peptidoglycan metabolism and the secreted anti-inflammatory FLIPr protein. Using a combination of in vivo and in vitro approaches, these mRNAs were validated as direct RsaA targets. Quantitative differential proteomics of wild-type and mutant strains corroborated the MAPS results. Additionally, it revealed that RsaA indirectly activated the synthesis of surface proteins supporting previous data that RsaA stimulated biofilm formation and favoured chronic infections. All together, this study shows that MAPS could also be easily applied in Gram-positive bacteria for identification of sRNA targetome.}, keywords = {PPSE, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Recent Advances in Mitochondrial Aminoacyl-tRNA Synthetases and Disease}, author = {M Sissler and L E González-Serrano and E Westhof}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28716624?dopt=Abstract}, doi = {10.1016/j.molmed.2017.06.002}, isbn = {28716624}, year = {2017}, date = {2017-01-01}, journal = {Trends Mol Med}, volume = {23}, number = {8}, pages = {693-708}, abstract = {Dysfunctions in mitochondria - the powerhouses of the cell - lead to several human pathologies. Because mitochondria integrate nuclear and mitochondrial genetic systems, they are richly intertwined with cellular activities. The nucleus-encoded mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs) are key components of the mitochondrial translation apparatus. Mutations in these enzymes predominantly affect the central nervous system (CNS) but also target other organs. Comparable mutations in mt-aaRSs can lead to vastly diverse diseases, occurring at different stages in life, and within different tissues; this represents a confounding issue. With newer information available, we propose that the pleiotropy and tissue-specificity of mt-aaRS-associated diseases result from the molecular integration of mitochondrial translation events within the cell; namely, through specific crosstalk between the cellular program and the energy demands of the cell. We place particular focus on neuronal cells.}, keywords = {aminoacyl-tRNA synthetase central nervous system mitochondrial disease mitochondrial translation moonlighting proteins unfolded protein response, SISSLER, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Hijacking of the Ubiquitin/Proteasome Pathway by the HIV Auxiliary Proteins}, author = {T Seissler and R Marquet and J C Paillart}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29088112?dopt=Abstract}, doi = {10.3390/v9110322}, isbn = {29088112}, year = {2017}, date = {2017-01-01}, journal = {Viruses}, volume = {9}, number = {11}, pages = {322}, abstract = {The ubiquitin-proteasome system (UPS) ensures regulation of the protein pool in the cell by ubiquitination of proteins followed by their degradation by the proteasome. It plays a central role in the cell under normal physiological conditions as well as during viral infections. On the one hand, the UPS can be used by the cell to degrade viral proteins, thereby restricting the viral infection. On the other hand, it can also be subverted by the virus to its own advantage, notably to induce degradation of cellular restriction factors. This makes the UPS a central player in viral restriction and counter-restriction. In this respect, the human immunodeficiency viruses (HIV-1 and 2) represent excellent examples. Indeed, many steps of the HIV life cycle are restricted by cellular proteins, some of which are themselves components of the UPS. However, HIV itself hijacks the UPS to mediate defense against several cellular restriction factors. For example, the HIV auxiliary proteins Vif, Vpx and Vpu counteract specific restriction factors by the recruitment of cellular UPS components. In this review, we describe the interplay between HIV and the UPS to illustrate its role in the restriction of viral infections and its hijacking by viral proteins for counter-restriction.}, keywords = {APOBEC BST2/Tetherin HIV March8 SAMHD1 TRIM5α proteasome restriction factors ubiquitin, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Correction: Structural Insights into Viral Determinants of Nematode Mediated Grapevine fanleaf virus Transmission.}, author = {P Schellenberger and C Sauter and B Lorber and P Bron and S Trapani and M Bergdoll and A Marmonier and C Schmitt-Kelchinger and O Lemaire and G Demangeat and C Ritzenthaler}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28296962?dopt=Abstract}, doi = {10.1371/journal.ppat.1006268}, isbn = {28296962}, year = {2017}, date = {2017-01-01}, journal = {PLoS Pathog}, volume = {13}, number = {3}, pages = {e1006268}, abstract = {[This corrects the article DOI: 10.1371/journal.ppat.1002034.]. Erratum for Structural insights into viral determinants of nematode mediated Grapevine fanleaf virus transmission. [PLoS Pathog. 2011]}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Synthesis, Thermodynamic Properties, and Crystal Structure of RNA Oligonucleotides Containing 5-Hydroxymethylcytosine.}, author = {C Riml and A Lusser and E Ennifar and R Micura}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28707898?dopt=Abstract}, doi = {10.1021/acs.joc.7b01171}, isbn = {28707898}, year = {2017}, date = {2017-01-01}, journal = {J Org Chem}, volume = {82}, number = {15}, pages = {7939-7945}, abstract = {5-Hydroxymethylcytosine (hm5C) is an RNA modification that has attracted significant interest because of the finding that RNA hydroxymethylation can favor mRNA translation. For insight into the mechanistic details of hm5C function to be obtained, the availability of RNAs containing this modification at defined positions that can be used for in vitro studies is highly desirable. In this work, we present an eight-step route to 5-hydroxymethylcytidine (hm5rC) phosphoramidite for solid-phase synthesis of modified RNA oligonucleotides. Furthermore, we examined the effects of hm5rC on RNA duplex stability and its impact on structure formation using the sarcin-ricin loop (SRL) motif. Thermal denaturation experiments revealed that hm5rC increases RNA duplex stability. By contrast, when cytosine within an UNCG tetraloop motif was replaced by hm5rC, the thermodynamic stability of the corresponding hairpin fold was attenuated. Importantly, incorporation of hm5rC into the SRL motif resulted in an RNA crystal structure at 0.85 Å resolution. Besides changes in the hydration pattern at the site of modification, a slight opening of the hm5rC-G pair compared to the unmodified C-G in the native structure was revealed.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Biophysical analysis of Arabidopsis protein-only RNase P alone and in complex with tRNA provides a refined model of tRNA binding.}, author = {F Pinker and C Schelcher and P Fernández-Millán and A Gobert and C Birck and A Thureau and P Roblin and P Giege and C Sauter}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28696260?dopt=Abstract}, doi = {10.1074/jbc.M117.782078}, isbn = {28696260}, year = {2017}, date = {2017-01-01}, journal = {J Biol Chem}, volume = {292}, number = {34}, pages = {13904-13913}, abstract = {RNase P is a universal enzyme that removes 5' leader sequences from tRNA precursors. The enzyme is therefore essential for maturation of functional tRNAs and mRNA translation. RNase P represents a unique example of an enzyme that can occur either as ribonucleoprotein or as protein alone. The latter form of the enzyme called PRORP (PRotein-Only RNase P) is widespread in eukaryotes, in which it can provide organellar or nuclear RNase P activities. Here, we have focused on Arabidopsis nuclear PRORP2 and its interaction with tRNA substrates. Affinity measurements helped assess the respective importance of individual pentatricopeptide repeat motifs in PRORP2 for RNA binding. We characterized the PRORP2 structure by X-ray crystallography and by small-angle X-ray scattering (SAXS) in solution, as well as that of its complex with a tRNA precursor by SAXS. Of note, our study reports the first structural data of a PRORP-tRNA complex. Combined with complementary biochemical and biophysical analyses, our structural data suggest that PRORP2 undergoes conformational changes to accommodate its substrate. In particular, the catalytic domain and the RNA binding domain can move around a central hinge. Altogether, this work provides a refined model of the PRORP-tRNA complex that illustrates how protein-only RNase P enzymes specifically bind tRNA and highlights the contribution of protein dynamics to achieve this specific interaction.}, keywords = {PRORP RNA processing X-ray crystallography pentatricopeptide repeat (PPR) precursor tRNA (pre-tRNA) ribonuclease P (RNase P) small-angle X-ray scattering (SAXS) tRNA maturation, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {MiSynPat: an integrated knowledge base linking clinical, genetic, and structural data for disease-causing mutations in human mitochondrial aminoacyl-tRNA synthetases.}, author = {L Moulinier and R Ripp and G Castillo and O Poch and M Sissler}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28608363}, doi = {10.1002/humu.23277}, isbn = {28608363}, year = {2017}, date = {2017-01-01}, journal = {Hum Mutat}, volume = {38}, number = {10}, pages = {1316-1324}, abstract = {Numerous mutations in each of the mitochondrial aminoacyl-tRNA synthetases have been implicated in human diseases. The mutations are autosomal and recessive and lead mainly to neurological disorders, although with pleiotropic effects. The processes and interactions that drive the etiology of the disorders associated with mitochondrial aminoacyl-tRNA synthetases are far from understood. The complexity of the clinical, genetic and structural data requires concerted, interdisciplinary efforts to understand the molecular biology of these disorders. Towards this goal, we designed MiSynPat, a comprehensive knowledge base together with an ergonomic web server designed to organize and access all pertinent information (sequences, multiple sequence alignments, structures, disease descriptions, mutation characteristics, original literature) on the disease-linked human mitochondrial aminoacyl-tRNA synthetases. With MiSynPat, a user can also evaluate the impact of a possible mutation on sequence-conservation-structure in order to foster the links between basic and clinical researchers and to facilitate future diagnosis. The proposed integrated view, coupled with research on disease-related mitochondrial aminoacyl-tRNA synthetases, will help to reveal new functions for these enzymes and to open new vistas in the molecular biology of the cell. The purpose of MiSynPat, freely available at http://misynpat.org, is to constitute a reference and a converging resource for scientists and clinicians. This article is protected by copyright. All rights reserved.}, keywords = {3D structures aminoacyl-tRNA synthetases disease-causing mutations knowledge base mitochondrial disorders sequence alignments, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA Structure: Advances and Assessment of 3D Structure Prediction.}, author = {Z Miao and E Westhof}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28375730}, doi = {10.1146/annurev-biophys-070816-034125}, isbn = {28375730}, year = {2017}, date = {2017-01-01}, journal = {Annu Rev Biophys}, volume = {46}, pages = {483-503}, abstract = {Biological functions of RNA molecules are dependent upon sustained specific three-dimensional (3D) structures of RNA, with or without the help of proteins. Understanding of RNA structure is frequently based on 2D structures, which describe only the Watson-Crick (WC) base pairs. Here, we hierarchically review the structural elements of RNA and how they contribute to RNA 3D structure. We focus our analysis on the non-WC base pairs and on RNA modules. Several computer programs have now been designed to predict RNA modules. We describe the RNA-Puzzles initiative, which is a community-wide, blind assessment of RNA 3D structure prediction programs to determine the capabilities and bottlenecks of current predictions. The assessment metrics used in RNA-Puzzles are briefly described. The detection of RNA 3D modules from sequence data and their automatic implementation belong to the current challenges in RNA 3D structure prediction. Expected final online publication date for the Annual Review of Biophysics Volume 46 is May 20, 2017. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA-Puzzles Round III: 3D RNA structure prediction of five riboswitches and one ribozyme.}, author = {Z Miao and R W Adamiak and M Antczak and R T Batey and A J Becka and M Biesiada and M J Boniecki and J Bujnicki and S J Chen and C Y Cheng and F C Chou and A R Ferré-D'Amaré and R Das and W K Dawson and D Feng and N V Dokholyan and S Dunin-Horkawicz and C Geniesse and K Kappel and W Kladwang and A Krokhotin and G E Lach and F Major and T H Mann and M Magnus and K Pachulska-Wieczorek and D J Patel and J A Piccirilli and M Popenda and K J Purzycka and A Ren and G M Rice and Jr. J Santalucia and J Sarzynska and M Szachniuk and A Tandon and J J Trausch and S Tian and J Wang and K M Weeks and 2nd. B Williams and Y Xiao and X Xu and D Zhang and T Zok and E Westhof}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28138060?dopt=Abstract}, doi = {10.1261/rna.060368.116}, isbn = {28138060}, year = {2017}, date = {2017-01-01}, journal = {RNA}, volume = {23}, number = {5}, pages = {655-672}, abstract = {RNA-Puzzles is a collective experiment in blind 3D RNA structure prediction. We report here a third round of RNA-Puzzles. Five puzzles, 4, 8, 12, 13, 14, all structures of riboswitch aptamers and puzzle 7, a ribozyme structure, are included in this round of the experiment. The riboswitch structures include biological binding sites for small molecules (S-adenosyl methionine, cyclic diadenosine monophosphate, 5-amino 4-imidazole carboxamide riboside 5'-triphosphate, glutamine) and proteins (YbxF) and one set describes large conformational changes between ligand-free and ligand-bound states; the Varkud satellite ribozyme is the most recently solved structure of a known large ribozyme. All the puzzles have established biological functions and require structural understanding to appreciate their molecular mechanisms. Through the use of fast-track experimental data, including multidimensional chemical mapping, and accurate prediction of RNA secondary structure, a large portion of the contacts in 3D have been predicted correctly leading to similar topologies for the top ranking predictions. Template-based and homology-derived predictions could predict structures to particularly high accuracies. However, achieving biological insights from de novo prediction of RNA 3D structures still depends on the size and complexity of the RNA. Blind computational predictions of RNA structures already appear to provide useful structural information in many cases. Similar to the previous RNA-Puzzles Round II experiment, the prediction of non-Watson-Crick interactions and the observed high atomic clash scores reveal notable need for algorithm of improvement. All prediction models and assessment results are available at http://ahsoka.u-strasbg.fr/rnapuzzles/.}, keywords = {3D prediction X-ray structures bioinformatics force fields models structure quality, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {ABCE1: A special factor that orchestrates translation at the crossroad between recycling and initiation.}, author = {E Mancera-Martinez and J Brito Querido and L S Valasek and A Simonetti and Y Hashem}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28498001?dopt=Abstract}, doi = {10.1080/15476286.2016.1269993}, isbn = {28498001}, year = {2017}, date = {2017-01-01}, journal = {RNA Biol}, volume = {14}, number = {10}, pages = {1279-1285}, abstract = {For many years initiation and termination of mRNA translation have been studied separately. However, a direct link between these two isolated stages has been suggested by the fact that some initiation factors also control termination and can even promote ribosome recycling; i.e. the last stage where post-terminating 80 S ribosomes are split to start a new round of initiation. Notably, it is now firmly established that, among other factors, ribosomal recycling critically requires the NTPase ABCE1. However, several earlier reports have proposed that ABCE1 also somehow participates in the initiation complex assembly. Based on an extended analysis of our recently published late-stage 48 S initiation complex from rabbit, here we provide new mechanistic insights into this putative role of ABCE1 in initiation. This point of view represents the first structural evidence in which the regulatory role of the recycling factor ABCE1 in initiation is discussed and establishes a corner stone for elucidating the interplay between ABCE1 and several initiation factors during the transit from ribosomal recycling to formation of the elongation competent 80 S initiation complex.}, keywords = {ABCE1 cryo-EM recycling ribosome translation, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Erratum}, author = {E Lioliou and P Fechter and I Caldelari and B C Jester and S Dubrac and A C Helfer and S Boisset and F Vandenesch and P Romby and T Geissmann}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29243975?dopt=Abstract}, doi = {10.1080/15476286.2016.1184436}, isbn = {29243975}, year = {2017}, date = {2017-01-01}, journal = {RNA Biol}, volume = {14}, number = {12}, pages = {1827}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mg2+ ions: do they bind to nucleobase nitrogens?}, author = {F Leonarski and L D'Ascenzo and P Auffinger}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27923930?dopt=Abstract}, doi = {10.1093/nar/gkw1175}, isbn = {27923930}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {2}, pages = {987-1004}, abstract = {Given the many roles proposed for Mg2+ in nucleic acids, it is essential to accurately determine their binding modes. Here, we surveyed the PDB to classify Mg2+ inner-sphere binding patterns to nucleobase imine N1/N3/N7 atoms. Among those, purine N7 atoms are considered to be the best nucleobase binding sites for divalent metals. Further, Mg2+ coordination to N7 has been implied in several ribozyme catalytic mechanisms. We report that Mg2+ assigned near imine nitrogens derive mostly from poor interpretations of electron density patterns and are most often misidentified Na+, K+, NH4+ ions, water molecules or spurious density peaks. Consequently, apart from few documented exceptions, Mg2+ ions do not bind to N7 atoms. Without much of a surprise, Mn2+, Zn2+ and Cd2+, which have a higher affinity for nitrogens, may contact N7 atoms when present in crystallization buffers. In this respect, we describe for the first time a potential Zn2+ ribosomal binding site involving two purine N7 atoms. Further, we provide a set of guidelines to help in the assignment of Mg2+ in crystallographic, cryo-EM, NMR and model building practices and discuss implications of our findings related to ion substitution experiments.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A novel double kink-turn module in euryarchaeal RNase P RNAs.}, author = {L B Lai and A Tanimoto and S M Lai and W Y Chen and I A Marathe and E Westhof and V H Wysocki and V Gopalan}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28525600?dopt=Abstract}, doi = {10.1093/nar/gkx388}, isbn = {28525600}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {12}, pages = {7432-7440}, abstract = {RNase P is primarily responsible for the 5΄ maturation of transfer RNAs (tRNAs) in all domains of life. Archaeal RNase P is a ribonucleoprotein made up of one catalytic RNA and five protein cofactors including L7Ae, which is known to bind the kink-turn (K-turn), an RNA structural element that causes axial bending. However, the number and location of K-turns in archaeal RNase P RNAs (RPRs) are unclear. As part of an integrated approach, we used native mass spectrometry to assess the number of L7Ae copies that bound the RPR and site-specific hydroxyl radical-mediated footprinting to localize the K-turns. Mutagenesis of each of the putative K-turns singly or in combination decreased the number of bound L7Ae copies, and either eliminated or changed the L7Ae footprint on the mutant RPRs. In addition, our results support an unprecedented 'double K-turn' module in type A and type M archaeal RPR variants.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Acute Exposure to Cigarette Smoking Followed by Myocardial Infarction Aggravates Renal Damage in an \textit{In Vivo} Mouse Model.}, author = {F Kobeissy and A Shaito and A Kaplan and L Baki and H Hayek and C Dagher-Hamalian and A Nehme and R Ghali and E Abidi and A Husari and A Zeidan and F A Zouein and K Zibara}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29177025?dopt=Abstract}, doi = {10.1155/2017/5135241}, isbn = {29177025}, year = {2017}, date = {2017-01-01}, journal = {Oxid Med Cell Longev}, volume = {2017}, pages = {5135241}, abstract = {Cigarette smoking (S) is a risk factor for progressive chronic kidney disease, renal dysfunction, and renal failure. In this study, the effect of smoking on kidney function was investigated in a mouse model of myocardial infarction (MI) using 4 groups: control (C), smoking (S), MI, and S+MI. Histological analysis of S+MI group showed alterations in kidney structure including swelling of the proximal convoluted tubules (PCTs), thinning of the epithelial lining, focal loss of the brush border of PCTs, and patchy glomerular retraction. Molecular analysis revealed that nephrin expression was significantly reduced in the S+MI group, whereas sodium-hydrogen exchanger-1 (NHE-1) was significantly increased, suggesting altered glomerular filtration and kidney functions. Moreover, S+MI group, but not S alone, showed a significant increase in the expression of connective tissue growth factor (CTGF) and fibrotic proteins fibronectin (FN) and α-smooth muscle actin (SMA), in comparison to controls, in addition to a significant increase in mRNA levels of IL-6 and TNF-α inflammatory markers. Finally, reactive oxygen species (ROS) production was significantly accentuated in S+MI group concomitant with a significant increase in NOX-4 protein levels. In conclusion, smoking aggravates murine acute renal damage caused by MI at the structural and molecular levels by exacerbating renal dysfunction.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of the 70S ribosome from human pathogen Staphylococcus aureus.}, author = {I Khusainov and Q Vicens and A Bochler and F Grosse and A Myasnikov and J F Ménétret and J Chicher and S Marzi and P Romby and G Yusupova and M Yusupov and Y Hashem}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28123039?dopt=Abstract}, doi = {10.1093/nar/gkw1126}, isbn = {28123039}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {2}, pages = {1026}, abstract = {Erratum for Structure of the 70S ribosome from human pathogen Staphylococcus aureus.}, keywords = {PPSE, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structures and dynamics of hibernating ribosomes from \textit{Staphylococcus aureus} mediated by intermolecular interactions of HPF.}, author = {I Khusainov and Q Vicens and R Ayupov and K Usachev and A Myasnikov and A Simonetti and S Validov and B Kieffer and G Yusupova and M Yusupov and Y Hashem}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28645916?dopt=Abstract}, doi = {10.15252/embj.201696105}, isbn = {28645916}, year = {2017}, date = {2017-01-01}, journal = {EMBO J}, volume = {36}, number = {14}, pages = {2073-2087}, abstract = {In bacteria, ribosomal hibernation shuts down translation as a response to stress, through reversible binding of stress-induced proteins to ribosomes. This process typically involves the formation of 100S ribosome dimers. Here, we present the structures of hibernating ribosomes from human pathogen Staphylococcus aureus containing a long variant of the hibernation-promoting factor (SaHPF) that we solved using cryo-electron microscopy. Our reconstructions reveal that the N-terminal domain (NTD) of SaHPF binds to the 30S subunit as observed for shorter variants of HPF in other species. The C-terminal domain (CTD) of SaHPF protrudes out of each ribosome in order to mediate dimerization. Using NMR, we characterized the interactions at the CTD-dimer interface. Secondary interactions are provided by helix 26 of the 16S ribosomal RNA We also show that ribosomes in the 100S particle adopt both rotated and unrotated conformations. Overall, our work illustrates a specific mode of ribosome dimerization by long HPF, a finding that may help improve the selectivity of antimicrobials.}, keywords = {cryo-electron microscopy hibernation pathogen ribosome, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Efficient and Accurate Translation Initiation Directed by TISU Involves RPS3 and RPS10e Binding and Differential Eukaryotic Initiation Factor 1A Regulation}, author = {O Haimov and H Sinvani and F Martin and I Ulitsky and R Emmanuel and A Tamarkin-Ben-Harush and A Vardy and R Dikstein}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28584194?dopt=Abstract}, doi = {10.1128/MCB.00150-17}, isbn = {28584194}, year = {2017}, date = {2017-01-01}, journal = {Mol Cell Biol}, volume = {37}, number = {15}, pages = {e00150-00117}, abstract = {Canonical translation initiation involves ribosomal scanning but short 5' UTR mRNAs are translated in a scanning-independent manner. The extent and mechanism of scanning independent translation is not fully understood. Here we report that short 5' UTR mRNAs constitute a substantial fraction of the translatome. Short 5' UTR mRNAs are enriched with TISU, a 12-nucleotide element directing efficient scanning-independent translation. Comprehensive mutagenesis revealed that each AUG flanking nucleotides of TISU contributes to translational strength but only few are important for accuracy. Using site-specific UV crosslinking of ribosomal complexes assembled on TISU mRNA we demonstrate specific binding of TISU to ribosomal proteins at the E and the A sites. We identified RPS3 as the major TISU-binding protein in the 48S complex A-site. Upon 80S formation RPS3 interaction is weakened and switched to RPS10e (former name RPS10). We further demonstrate that TISU is particularly dependent on eIF1A which interacts with both RPS3 and RPS10e. Our findings suggest that the cap-recruited ribosome specifically binds the TISU nucleotides at the A and the E sites in cooperation with eIF1A to promote scanning arrest.}, keywords = {ERIANI, RPS10 RPS10e RPS3 TISU eIF1A short 5′UTR translation initiation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The IRES5'UTR of the dicistrovirus cricket paralysis virus is a type III IRES containing an essential pseudoknot structure}, author = {L Gross and Q Vicens and E Einhorn and A Noireterre and L Schaeffer and L Kuhn and JL Imler and G Eriani and C Meignin and F Martin}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28911115?dopt=Abstract}, doi = {10.1093/nar/gkx622}, isbn = {28911115}, year = {2017}, date = {2017-01-01}, urldate = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {15}, pages = {8993-9004}, abstract = {Cricket paralysis virus (CrPV) is a dicistrovirus. Its positive-sense single-stranded RNA genome contains two internal ribosomal entry sites (IRESs). The 5' untranslated region (5'UTR) IRES5'UTR mediates translation of non-structural proteins encoded by ORF1 whereas the well-known intergenic region (IGR) IRESIGR is required for translation of structural proteins from open reading frame 2 in the late phase of infection. Concerted action of both IRES is essential for host translation shut-off and viral translation. IRESIGR has been extensively studied, in contrast the IRES5'UTR remains largely unexplored. Here, we define the minimal IRES element required for efficient translation initiation in drosophila S2 cell-free extracts. We show that IRES5'UTR promotes direct recruitment of the ribosome on the cognate viral AUG start codon without any scanning step, using a Hepatitis-C virus-related translation initiation mechanism. Mass spectrometry analysis revealed that IRES5'UTR recruits eukaryotic initiation factor 3, confirming that it belongs to type III class of IRES elements. Using Selective 2'-hydroxyl acylation analyzed by primer extension and DMS probing, we established a secondary structure model of 5'UTR and of the minimal IRES5'UTR. The IRES5'UTR contains a pseudoknot structure that is essential for proper folding and ribosome recruitment. Overall, our results pave the way for studies addressing the synergy and interplay between the two IRES from CrPV.}, keywords = {ERIANI, meignin, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {SECIS-binding protein 2 interacts with the SMN complex and the methylosome for selenoprotein mRNP assembly and translation.}, author = {A S Gribling-Burrer and M Leichter and L Wurth and A Huttin and F Schlotter and N Troffer-Charlier and V Cura and M Barkats and J Cavarelli and S Massenet and C Allmang}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28115638?dopt=Abstract}, doi = {10.1093/nar/gkx031}, isbn = {28115638}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {9}, pages = {5399-5413}, abstract = {Selenoprotein synthesis requires the co-translational recoding of a UGASec codon. This process involves an RNA structural element, called Selenocysteine Insertion Sequence (SECIS) and the SECIS binding protein 2 (SBP2). Several selenoprotein mRNAs undergo unusual cap hypermethylation by the trimethylguanosine synthase 1 (Tgs1), which is recruited by the ubiquitous Survival of MotoNeurons (SMN) protein. SMN, the protein involved in spinal muscular atrophy, is part of a chaperone complex that collaborates with the methylosome for RNP assembly. Here, we analyze the role of individual SMN and methylosome components in selenoprotein mRNP assembly and translation. We show that SBP2 interacts directly with four proteins of the SMN complex and the methylosome core proteins. Nevertheless, SBP2 is not a methylation substrate of the methylosome. We found that both SMN and methylosome complexes are required for efficient translation of the selenoprotein GPx1 in vivo We establish that the steady-state level of several selenoprotein mRNAs, major regulators of oxidative stress damage in neurons, is specifically reduced in the spinal cord of SMN-deficient mice and that cap hypermethylation of GPx1 mRNA is affected. Altogether we identified a new function of the SMN complex and the methylosome in selenoprotein mRNP assembly and expression.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{nehmar_therapeutic_2017, title = {Therapeutic Modulation of Plasmacytoid Dendritic Cells in Experimental Arthritis}, author = {Ramzi Nehmar and Ghada Alsaleh and Benjamin Voisin and Vincent Flacher and Alexandre Mariotte and Victoria Saferding and Antonia Puchner and Birgit Niederreiter and Thierry Vandamme and Gernot Schabbauer and Philippe Kastner and Susan Chan and Peggy Kirstetter and Martin Holcmann and Christopher Mueller and Jean Sibilia and Seiamak Bahram and Stephan Blüml and Philippe Georgel}, doi = {10.1002/art.40225}, issn = {2326-5205}, year = {2017}, date = {2017-01-01}, journal = {Arthritis & Rheumatology (Hoboken, N.J.)}, volume = {69}, number = {11}, pages = {2124--2135}, abstract = {OBJECTIVE: The role of plasmacytoid dendritic cells (PDCs) and type I interferons (IFNs) in rheumatoid arthritis (RA) remains a subject of controversy. This study was undertaken to explore the contribution of PDCs and type I IFNs to RA pathogenesis using various animal models of PDC depletion and to monitor the effect of localized PDC recruitment and activation on joint inflammation and bone damage. METHODS: Mice with K/BxN serum-induced arthritis, collagen-induced arthritis, and human tumor necrosis factor transgene insertion were studied. Symptoms were evaluated by visual scoring, quantification of paw swelling, determination of cytokine levels by enzyme-linked immunosorbent assay, and histologic analysis. Imiquimod-dependent therapeutic effects were monitored by transcriptome analysis (using quantitative reverse transcriptase-polymerase chain reaction) and flow cytometric analysis of the periarticular tissue. RESULTS: PDC-deficient mice showed exacerbation of inflammatory and arthritis symptoms after arthritogenic serum transfer. In contrast, enhancing PDC recruitment and activation to arthritic joints by topical application of the Toll-like receptor 7 (TLR-7) agonist imiquimod significantly ameliorated arthritis in various mouse models. Imiquimod induced an IFN signature and led to reduced infiltration of inflammatory cells. CONCLUSION: The therapeutic effects of imiquimod on joint inflammation and bone destruction are dependent on TLR-7 sensing by PDCs and type I IFN signaling. Our findings indicate that local recruitment and activation of PDCs represents an attractive therapeutic opportunity for RA patients.}, keywords = {Activation, Adjuvants, Aminoquinolines, Analysis, Animal, Animals, arthritis, Assay, cancer, Cells, cytokine, Cytokines, Dendritic Cells, DEPLETION, Disease Models, drug effects, Enzyme-Linked Immunosorbent Assay, Experimental, Flow Cytometry, Gene Expression Profiling, Genetics, GLYCOPROTEIN, Glycoproteins, Human, Humans, IFN, IKAROS, Ikaros Transcription Factor, imiquimod, Immunologic, Immunology, immunopathology, inflammation, interferon, Interferon Type I, interferons, Knockout, Membrane, Membrane Glycoproteins, METHOD, methods, Mice, MODULATION, mouse, Necrosis, NECROSIS-FACTOR-ALPHA, pathogenesis, Patients, Pharmacology, physiology, plasmacytoid dendritic cells, Protein, Receptor, Reverse Transcriptase Polymerase Chain Reaction, rheumatoid, rheumatoid arthritis, Serum, signaling, Team-Mueller, TLR7, Toll-Like Receptor 7, TOPICAL APPLICATION, Transcription, TRANSCRIPTION FACTOR, transcriptome, transgenic, tumor, Tumor Necrosis Factor, Tumor Necrosis Factor-alpha}, pubstate = {published}, tppubtype = {article} } @article{, title = {What macromolecular crystallogenesis tells us - what is needed in the future}, author = {R Giege}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28875021?report=&dispmax=200&tool=PubCrawler_2.38}, doi = {10.1107/S2052252517006595}, isbn = {28875021}, year = {2017}, date = {2017-01-01}, journal = {IUCrJ}, volume = {4}, number = {Pt 4}, pages = {340-349}, abstract = {Crystallogenesis is a longstanding topic that has transformed into a discipline that is mainly focused on the preparation of crystals for practising crystallo-graphers. Although the idiosyncratic features of proteins have to be taken into account, the crystallization of proteins is governed by the same physics as the crystallization of inorganic materials. At present, a diversified panel of crystallization methods adapted to proteins has been validated, and although only a few methods are in current practice, the success rate of crystallization has increased constantly, leading to the determination of ∼105 X-ray structures. These structures reveal a huge repertoire of protein folds, but they only cover a restricted part of macromolecular diversity across the tree of life. In the future, crystals representative of missing structures or that will better document the structural dynamics and functional steps underlying biological processes need to be grown. For the pertinent choice of biologically relevant targets, computer-guided analysis of structural databases is needed. From another perspective, crystallization is a self-assembly process that can occur in the bulk of crowded fluids, with crystals being supramolecular assemblies. Life also uses self-assembly and supramolecular processes leading to transient, or less often stable, complexes. An integrated view of supramolecularity implies that proteins crystallizing either in vitro or in vivo or participating in cellular processes share common attributes, notably determinants and antideterminants that favour or disfavour their correct or incorrect associations. As a result, under in vivo conditions proteins show a balance between features that favour or disfavour association. If this balance is broken, disorders/diseases occur. Understanding crystallization under in vivo conditions is a challenge for the future. In this quest, the analysis of packing contacts and contacts within oligomers will be crucial in order to decipher the rules governing protein self-assembly and will guide the engineering of novel biomaterials. In a wider perspective, understanding such contacts will open the route towards supramolecular biology and generalized crystallogenesis.}, keywords = {crowding crystal engineering crystallizability crystallization predictors crystallogenesis determinant and antideterminant evolution packing self-assembly rules supramolecularity surface patches symmetry and asymmetry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure and fluorescence properties of the iSpinach aptamer in complex with DFHBI}, author = {P Fernandez-Millan and A Autour and E Ennifar and E Westhof and M Ryckelynck}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28939697?dopt=Abstract}, doi = {10.1261/rna.063008}, isbn = {28939697}, year = {2017}, date = {2017-01-01}, journal = {RNA}, volume = {23}, number = {12}, pages = {1788-1795}, abstract = {Fluorogenic RNA aptamers are short nucleic acids able to specifically interact with small molecules and strongly enhance their fluorescence upon complex formation. Among the different systems recently introduced, Spinach, an aptamer forming a fluorescent complex with the 3,5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI), is one of the most promising. Using random mutagenesis and ultrahigh-throughput screening, we recently developed iSpinach, an improved version of the aptamer, endowed with an increased folding efficiency and thermal stability. iSpinach is a shorter version of Spinach comprising five mutations whom the exact role was not deciphered yet. In this work, we co-crystallized a re-engineered version of iSpinach in complex with the DFHBI and solved the x-ray structure of the complex at 2 Å resolution. Only a few mutations were required to optimize iSpinach production and crystallization, underlying the good folding capacity of the molecule. The measured fluorescence half-lives in the crystal were 60% higher than in solution. Comparisons with structures previously reported for Spinach allows shedding some light on the possible function of the different beneficial mutations carried by iSpinach.}, keywords = {DFHBI Spinach crystal structure fluorescence fluorogenic RNA aptamer, ENNIFAR, RYCKELYNCK, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{saliba_enhancing_2017, title = {Enhancing tumor specific immune responses by transcutaneous vaccination}, author = {Hanadi Saliba and Béatrice Heurtault and Hasnaa Bouharoun-Tayoun and Vincent Flacher and Benoît Frisch and Sylvie Fournel and Soulaima Chamat}, doi = {10.1080/14760584.2017.1382357}, issn = {1744-8395}, year = {2017}, date = {2017-01-01}, journal = {Expert Review of Vaccines}, volume = {16}, number = {11}, pages = {1079--1094}, abstract = {INTRODUCTION: Our understanding of the involvement of the immune system in cancer control has increased over recent years. However, the development of cancer vaccines intended to reverse tumor-induced immune tolerance remains slow as most current vaccine candidates exhibit limited clinical efficacy. The skin is particularly rich with multiple subsets of dendritic cells (DCs) that are involved to varying degrees in the induction of robust immune responses. Transcutaneous administration of cancer vaccines may therefore harness the immune potential of these DCs, however, this approach is hampered by the impermeability of the stratum corneum. Innovative vaccine formulations including various nanoparticles, such as liposomes, are therefore needed to properly deliver cancer vaccine components to skin DCs. Areas covered: The recent insights into skin DC subsets and their functional specialization, the potential of nanoparticle-based vaccines in transcutaneous cancer vaccination and, finally, the most relevant clinical trial advances in liposomal and in cutaneous cancer vaccines will be discussed. Expert commentary: To define the optimal conditions for mounting protective skin DC-induced anti-tumor immune responses, investigation of the cellular and molecular interplay that controls tumor progression should be pursued in parallel with clinical development. The resulting knowledge will then be translated into improved cancer vaccines that better target the most appropriate immune players.}, keywords = {Administration, Cancer vaccine, Cancer Vaccines, Clinical Trials as Topic, Cutaneous, Dendritic Cells, Humans, liposome, Liposomes, nanoparticle, Nanoparticles, Neoplasms, Skin, skin dendritic cell, Team-Mueller, transcutaneous vaccination, Treatment Outcome, Vaccination}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular architecture and dynamics of ASH1-mRNA recognition by its mRNA-transport complex}, author = {F T Edelmann and A Schlundt and R G Heym and A Jenner and A Niedner-Boblenz and M I Syed and J C Paillart and R Stehle and R Janowski and M Sattler and R P Jansen and D Niessing}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28092367?dopt=Abstract}, doi = {10.1038/nsmb.3351}, isbn = {28092367}, year = {2017}, date = {2017-01-01}, journal = {Nat Struct Mol Biol}, volume = {24}, number = {2}, pages = {152-161}, abstract = {mRNA localization is an essential mechanism of gene regulation and is required for processes such as stem-cell division, embryogenesis and neuronal plasticity. It is not known which features in the cis-acting mRNA localization elements (LEs) are specifically recognized by motor-containing transport complexes. To the best of our knowledge, no high-resolution structure is available for any LE in complex with its cognate protein complex. Using X-ray crystallography and complementary techniques, we carried out a detailed assessment of an LE of the ASH1 mRNA from yeast, its complex with its shuttling RNA-binding protein She2p, and its highly specific, cytoplasmic complex with She3p. Although the RNA alone formed a flexible stem loop, She2p binding induced marked conformational changes. However, only joining by the unstructured She3p resulted in specific RNA recognition. The notable RNA rearrangements and joint action of a globular and an unfolded RNA-binding protein offer unprecedented insights into the step-wise maturation of an mRNA-transport complex.}, keywords = {Cell polarity RNA transport X-ray crystallography, MARQUET, MARQUET PAILLART Cell polarity RNA transport X-ray crystallography, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{debard_nonconventional_2017, title = {Nonconventional localizations of cytosolic aminoacyl-tRNA synthetases in yeast and human cells.}, author = {Sylvain Debard and Gaétan Bader and Johan-Owen De Craene and Ludovic Enkler and Séverine Bär and Daphné Laporte and Philippe Hammann and Evelyne Myslinski and Bruno Senger and Sylvie Friant and Hubert Dominique Becker}, doi = {10.1016/j.ymeth.2016.09.017}, issn = {1095-9130 1046-2023}, year = {2017}, date = {2017-01-01}, journal = {Methods (San Diego, Calif.)}, volume = {113}, pages = {91--104}, abstract = {By definition, cytosolic aminoacyl-tRNA synthetases (aaRSs) should be restricted to the cytosol of eukaryotic cells where they supply translating ribosomes with their aminoacyl-tRNA substrates. However, it has been shown that other translationally-active compartments like mitochondria and plastids can simultaneously contain the cytosolic aaRS and its corresponding organellar ortholog suggesting that both forms do not share the same organellar function. In addition, a fair number of cytosolic aaRSs have also been found in the nucleus of cells from several species. Hence, these supposedly cytosolic-restricted enzymes have instead the potential to be multi-localized. As expected, in all examples that were studied so far, when the cytosolic aaRS is imported inside an organelle that already contains its bona fide corresponding organellar-restricted aaRSs, the cytosolic form was proven to exert a nonconventional and essential function. Some of these essential functions include regulating homeostasis and protecting against various stresses. It thus becomes critical to assess meticulously the subcellular localization of each of these cytosolic aaRSs to unravel their additional roles. With this objective in mind, we provide here a review on what is currently known about cytosolic aaRSs multi-compartmentalization and we describe all commonly used protocols and procedures for identifying the compartments in which cytosolic aaRSs relocalize in yeast and human cells.}, note = {Place: United States}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {Argonaute proteins regulate HIV-1 multiply spliced RNA and viral production in a Dicer independent manner.}, author = {A Eckenfelder and E Ségéral and N Pinzon and D Ulveling and C Amadori and M Charpentier and S Nidelet and J P Concordet and J F Zagury and J C Paillart and C Berlioz-Torrent and H Seitz and S Emiliani and S Gallois-Montbrun}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28003477}, doi = {10.1093/nar/gkw1289}, isbn = {28003477}, year = {2017}, date = {2017-01-01}, journal = {Nucleic Acids Res}, volume = {45}, number = {7}, pages = {4158-4173}, abstract = {Argonaute (Ago) proteins associate with microRNAs (miRNAs) to form the core of the RNA-induced silencing complex (RISC) that mediates post-transcriptional gene silencing of target mRNAs. As key players in anti-viral defense, Ago proteins are thought to have the ability to interact with human immunodeficiency virus type 1 (HIV-1) RNA. However, the role of this interaction in regulating HIV-1 replication has been debated. Here, we used high throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP) to explore the interaction between Ago2 and HIV-1 RNA in infected cells. By only considering reads of 50 nucleotides length in our analysis, we identified more than 30 distinct binding sites for Ago2 along the viral RNA genome. Using reporter assays, we found four binding sites, located near splice donor sites, capable of repressing Luciferase gene expression in an Ago-dependent manner. Furthermore, inhibition of Ago1 and Ago2 levels in cells expressing HIV-1 led to an increase of viral multiply spliced transcripts and to a strong reduction in the extracellular CAp24 level. Depletion of Dicer did not affect these activities. Our results highlight a new role of Ago proteins in the control of multiply spliced HIV-1 transcript levels and viral production, independently of the miRNA pathway.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Site-Directed Chemical Probing to map transient RNA/protein interactions.}, author = {M Duval and A Marenna and C Chevalier and S Marzi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28027957}, doi = {10.1016/j.ymeth.2016.12.011}, isbn = {28027957}, year = {2017}, date = {2017-01-01}, journal = {Methods}, volume = {117}, pages = {48-58}, abstract = {RNA-protein interactions are at the bases of many biological processes, forming either tight and stable functional ribonucleoprotein (RNP) complexes (i.e. the ribosome) or transitory ones, such as the complexes involving RNA chaperone proteins. To localize the sites where a protein interacts on an RNA molecule, a common simple and inexpensive biochemical method is the footprinting technique. The protein leaves its footprint on the RNA acting as a shield to protect the regions of interaction from chemical modification or cleavages obtained with chemical or enzymatic nucleases. This method has proven its efficiency to study in vitro the organization of stable RNA-protein complexes. Nevertheless, when the protein binds the RNA very dynamically, with high off-rates, protections are very often difficult to observe. For the analysis of these transient complexes, we describe an alternative strategy adapted from the Site Directed Chemical Probing (SDCP) approach and we compare it with classical footprinting. SDCP relies on the modification of the RNA binding protein to tether an RNA probe (usually Fe-EDTA) to specific protein positions. Local cleavages on the regions of interaction can be used to localize the protein and position its domains on the RNA molecule. This method has been used in the past to monitor stable complexes; we provide here a detailed protocol and a practical example of its application to the study of Escherichia coli RNA chaperone protein S1 and its transitory complexes with mRNAs.}, keywords = {BABE RNA chaperone proteins RNA/protein interaction hydroxyl radicals ribosomal protein S1 rpsO mRNA site directed chemical probing transient ribonucleoprotein complexes, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{incarbone_neutralization_2017, title = {Neutralization of mobile antiviral small RNA through peroxisomal import.}, author = {M Incarbone and A Zimmermann and P Hammann and M Erhardt and F Michel and P Dunoyer}, doi = {10.1038/nplants.2017.94}, issn = {2055-0278 2055-0278}, year = {2017}, date = {2017-01-01}, journal = {Nature plants}, volume = {3}, pages = {17094}, abstract = {In animals, certain viral proteins are targeted to peroxisomes to dampen the antiviral immune response mediated by these organelles(1-3). In plants, RNA interference (RNAi) mediated by small interfering (si)RNA is the main antiviral defence mechanism. To protect themselves against the cell- and non-cell autonomous effects of RNAi, viruses produce viral suppressors of RNA silencing (VSR)(4), whose study is crucial to properly understand the biological cycle of plant viruses and potentially find new solutions to control these pathogens. By combining biochemical approaches, cell-specific inhibition of RNAi movement and peroxisome isolation, we show here that one such VSR, the peanut clump virus (PCV)-encoded P15, isolates siRNA from the symplasm by delivering them into the peroxisomal matrix. Infection with PCV lacking this ability reveals that piggybacking of these VSR-bound nucleic acids into peroxisomes potentiates viral systemic movement by preventing the spread of antiviral siRNA. Collectively, these results highlight organellar confinement of antiviral molecules as a novel pathogenic strategy that may have its direct counterpart in other plant and animal viruses.}, note = {Place: England}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {sRNA-mediated activation of gene expression by inhibition of 5'-3' exonucleolytic mRNA degradation.}, author = {S Durand and F Braun and A C Helfer and P Romby and C Condon}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28436820?dopt=Abstract}, doi = {10.7554/eLife.23602}, isbn = {28436820}, year = {2017}, date = {2017-01-01}, journal = {Elife}, volume = {6}, pages = {e23602}, abstract = {Post-transcriptional control by small regulatory RNA (sRNA) is critical for rapid adaptive processes. sRNAs can directly modulate mRNA degradation in Proteobacteria without interfering with translation. However, Firmicutes have a fundamentally different set of ribonucleases for mRNA degradation and whether sRNAs can regulate the activity of these enzymes is an open question. We show that Bacillus subtilis RoxS, a major trans-acting sRNA shared with Staphylococus aureus, prevents degradation of the yflS mRNA, encoding a malate transporter. In the presence of malate, RoxS transiently escapes from repression by the NADH-sensitive transcription factor Rex and binds to the extreme 5'-end of yflS mRNA. This impairs the 5'-3' exoribonuclease activity of RNase J1, increasing the half-life of the primary transcript and concomitantly enhances ribosome binding to increase expression of the transporter. Globally, the different targets regulated by RoxS suggest that it helps readjust the cellular NAD+/NADH balance when perturbed by different stimuli.}, keywords = {b. subtilis chromosomes genes infectious disease microbiology, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{montavon_specific_2017, title = {A specific dsRNA-binding protein complex selectively sequesters endogenous inverted-repeat siRNA precursors and inhibits their processing.}, author = {Thomas Montavon and Yerim Kwon and Aude Zimmermann and Philippe Hammann and Timothée Vincent and Valérie Cognat and Fabrice Michel and Patrice Dunoyer}, doi = {10.1093/nar/gkw1264}, issn = {1362-4962 0305-1048 0305-1048}, year = {2017}, date = {2017-01-01}, journal = {Nucleic acids research}, volume = {45}, number = {3}, pages = {1330--1344}, abstract = {In plants, several dsRNA-binding proteins (DRBs) have been shown to play important roles in various RNA silencing pathways, mostly by promoting the efficiency and/or accuracy of Dicer-like proteins (DCL)-mediated small RNA production. Among the DRBs encoded by the Arabidopsis genome, we recently identified DRB7.2 whose function in RNA silencing was unknown. Here, we show that DRB7.2 is specifically involved in siRNA production from endogenous inverted-repeat (endoIR) loci. This function requires its interacting partner DRB4, the main cofactor of DCL4 and is achieved through specific sequestration of endoIR dsRNA precursors, thereby repressing their access and processing by the siRNA-generating DCLs. The present study also provides multiple lines of evidence showing that DRB4 is partitioned into, at least, two distinct cellular pools fulfilling different functions, through mutually exclusive binding with either DCL4 or DRB7.2. Collectively, these findings revealed that plants have evolved a specific DRB complex that modulates selectively the production of endoIR-siRNAs. The existence of such a complex and its implication regarding the still elusive biological function of plant endoIR-siRNA will be discussed.}, keywords = {Inverted Repeat Sequences, PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nonconventional localizations of cytosolic aminoacyl-tRNA synthetases in yeast and human cells.}, author = {S Debard and G Bader and J O De Craene and L Enkler and S Bär and D Laporte and E Myslinski and B Senger and S Friant and H D Becker}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27725303?dopt=Abstract}, doi = {10.1016/j.ymeth.2016.09.017}, isbn = {27725303}, year = {2017}, date = {2017-01-01}, journal = {Methods}, volume = {113}, pages = {91-104}, abstract = {By definition, cytosolic aminoacyl-tRNA synthetases (aaRSs) should be restricted to the cytosol of eukaryotic cells where they supply translating ribosomes with their aminoacyl-tRNA substrates. However, it has been shown that other translationally-active compartments like mitochondria and plastids can simultaneously contain the cytosolic aaRS and its corresponding organellar ortholog suggesting that both forms do not share the same organellar function. In addition, a fair number of cytosolic aaRSs have also been found in the nucleus of cells from several species. Hence, these supposedly cytosolic-restricted enzymes have instead the potential to be multi-localized. As expected, in all examples that were studied, when the cytosolic aaRS is imported inside an organelle that already contains the bona fide corresponding organellar-restricted aaRSs, the cytosolic form was proven to exert a nonconventional and essential function. Some of these essential functions include homeostasis and protection against various stresses. It thus becomes critical to assess meticulously the subcellular localization of each of these cytosolic aaRSs to unravel their additional roles. With this objective in mind, we provide here a review on what is currently known about cytosolic aaRSs multi-compartmentalization and we describe all commonly used protocols and procedures for identifying the compartment in which cytosolic aaRSs relocalize in yeast and human cells.}, keywords = {JOSSINET Fractionation Human MTS Microscopy NLS Yeast aaRS tRNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Revisiting GNRA and UNCG folds: U-turns versus Z-turns in RNA hairpin loops.}, author = {L D'Ascenzo and F Leonarski and Q Vicens and P Auffinger}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27999116?dopt=Abstract}, doi = {10.1261/rna.059097.116}, isbn = {27999116}, year = {2017}, date = {2017-01-01}, journal = {RNA}, volume = {23}, number = {3}, pages = {259-269}, abstract = {When thinking about RNA three-dimensional structures, coming across GNRA and UNCG tetraloops is perceived as a boon since their folds have been extensively described. Nevertheless, analyzing loop conformations within RNA and RNP structures led us to uncover several instances of GNRA and UNCG loops that do not fold as expected. We noticed that when a GNRA does not assume its "natural" fold, it adopts the one we typically associate with a UNCG sequence. The same folding interconversion may occur for loops with UNCG sequences, for instance within tRNA anticodon loops. Hence, we show that some structured tetranucleotide sequences starting with G or U can adopt either of these folds. The underlying structural basis that defines these two fold types is the mutually exclusive stacking of a backbone oxygen on either the first (in GNRA) or the last nucleobase (in UNCG), generating an oxygen-π contact. We thereby propose to refrain from using sequences to distinguish between loop conformations. Instead, we suggest using descriptors such as U-turn (for "GNRA-type" folds) and a newly described Z-turn (for "UNCG-type" folds). Because tetraloops adopt for the largest part only two (inter)convertible turns, we are better able to interpret from a structural perspective loop interchangeability occurring in ribosomes and viral RNA. In this respect, we propose a general view on the inclination for a given sequence to adopt (or not) a specific fold. We also suggest how long-noncoding RNAs may adopt discrete but transient structures, which are therefore hard to predict.}, keywords = {ENNIFAR, RNA folding RNA motif structure prediction tRNA anticodon tetraloop, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutations in signal recognition particle SRP54 cause syndromic neutropenia with Shwachman-Diamond-like features}, author = {R Carapito and M Konantz and J C Paillart and Z Miao and A Pichot and M S Leduc and Y Yang and K L Bergstrom and D H Mahoney and D L Shardy and G Alsaleh and L Naegely and A Kolmer and N Paul and A Hanauer and V Rolli and J S Müller and E Alghisi and L Sauteur and C Macquin and A Morlon and C S Sancho and P Amati-Bonneau and V Procaccio and A L Mosca-Boidron and N Marle and N Osmani and O Lefebvre and J G Goetz and S Unal and N A Akarsu and M Radosavljevic and M P Chenard and F Rialland and A Grain and M C Béné and M Eveillard and M Vincent and J Guy and L Faivre and C Thauvin-Robinet and J Thevenon and K Myers and M D Fleming and A Shimamura and E Bottollier-Lemallaz and E Westhof and C Lengerke and B Isidor and S Bahram}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28972538?dopt=Abstract}, doi = {10.1172/JCI92876}, isbn = {28972538}, year = {2017}, date = {2017-01-01}, journal = {J Clin Invest}, volume = {127}, number = {11}, pages = {4090-4103}, abstract = {Shwachman-Diamond syndrome (SDS) (OMIM #260400) is a rare inherited bone marrow failure syndrome (IBMFS) that is primarily characterized by neutropenia and exocrine pancreatic insufficiency. Seventy-five to ninety percent of patients have compound heterozygous loss-of-function mutations in the Shwachman-Bodian-Diamond syndrome (sbds) gene. Using trio whole-exome sequencing (WES) in an sbds-negative SDS family and candidate gene sequencing in additional SBDS-negative SDS cases or molecularly undiagnosed IBMFS cases, we identified 3 independent patients, each of whom carried a de novo missense variant in srp54 (encoding signal recognition particle 54 kDa). These 3 patients shared congenital neutropenia linked with various other SDS phenotypes. 3D protein modeling revealed that the 3 variants affect highly conserved amino acids within the GTPase domain of the protein that are critical for GTP and receptor binding. Indeed, we observed that the GTPase activity of the mutated proteins was impaired. The level of SRP54 mRNA in the bone marrow was 3.6-fold lower in patients with SRP54-mutations than in healthy controls. Profound reductions in neutrophil counts and chemotaxis as well as a diminished exocrine pancreas size in a SRP54-knockdown zebrafish model faithfully recapitulated the human phenotype. In conclusion, autosomal dominant mutations in SRP54, a key member of the cotranslation protein-targeting pathway, lead to syndromic neutropenia with a Shwachman-Diamond-like phenotype.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Two proteomic methodologies for defining N-termini of mature human mitochondrial aminoacyl-tRNA synthetases.}, author = {C Carapito and L Kuhn and L Karim and M Rompais and T Rabilloud and H Schwenzer and M Sissler}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27793688?dopt=Abstract}, doi = {10.1016/j.ymeth.2016.10.012}, isbn = {27793688}, year = {2017}, date = {2017-01-01}, journal = {Methods}, volume = {113}, pages = {111-119}, abstract = {Human mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs) are encoded in the nucleus, synthesized in the cytosol and targeted for importation into mitochondria by a N-terminal mitochondrial targeting sequence. This targeting sequence is presumably cleaved upon entry into the mitochondria, following a process still not fully deciphered in human, despite essential roles for the mitochondrial biogenesis. Maturation processes are indeed essential both for the release of a functional enzyme and to route correctly the protein within mitochondria. The absence of consensus sequences for cleavage sites and the discovery of possible multiple proteolytic steps render predictions of N-termini difficult. Further, the knowledge of the cleavages is key for the design of protein constructions compatible with efficient production in bacterial strains. Finally, full comprehension becomes essential because a growing number of mutations are found in genes coding for mt-aaRS. In the present study, we take advantage of proteomic methodological developments and identified, in mitochondria, three N-termini for the human mitochondrial aspartyl-tRNA synthetase. This first description of the co-existence of different forms opens new perspectives in the biological understanding of this enzyme. Those methods are extended to the whole set of human mt-aaRSs and methodological advice are provided for further investigations.}, keywords = {PPSE, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 sequences in the epidemic suggest an alternative pathway for the generation of the Long Terminal Repeats}, author = {P Cappy and A Moisan and F De Oliveira and J C Plantier and M Negroni}, url = {https://www.nature.com/articles/s41598-017-14135-z}, doi = {10.1038/s41598-017-14135-z}, isbn = {29057964}, year = {2017}, date = {2017-01-01}, journal = {Sci Rep}, volume = {7}, number = {1}, pages = {13715}, abstract = {To generate the long-terminal repeats (LTR) that border the integrated viral genome, two-strand transfer steps must occur during reverse transcription. Analysis of the genetic polymorphisms that are present in the LTR of HIV-1 heterozygous virions in single infection cycle studies has revealed which of the two copies of genomic RNAs is used for each transfer event. Thus, the first event of strand transfer has been described to be either intra- or intermolecular, while the second event is generally intramolecular. Here, we repeated these analyses using sequences from HIV databases and extended the study to the regions surrounding the LTR. We observed a striking correlation between the pattern of recombination in the LTR and the phylogenetic origin of the surrounding sequences. This correlation suggests that the second-strand transfer can be either intra- or intermolecular and, interestingly, could reflect an effect of proximity between nucleic acids that would guide this transfer. This factor could be particularly relevant for heterozygous viruses containing highly divergent genomic RNAs, such as those considered in the present study.}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Complete genome sequence and annotation of the Staphylococcus aureus strain HG001}, author = {I Caldelari and B Chane-Woon-Ming and C Noirot and K Moreau and P Romby and C Gaspin and S Marzi}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28798184?dopt=Abstract}, doi = {10.1128/genomeA.00783-17}, isbn = {28798184}, year = {2017}, date = {2017-01-01}, journal = {Genome Announc}, volume = {5}, number = {32}, pages = {e00783-00717}, abstract = {Staphylococcus aureus is an opportunistic Gram-positive pathogen responsible for a wide range of infections from minor skin abscesses to life-threatening diseases. Here, we report the draft genome assembly and current annotation of the HG001 strain, a derivative of the RN1 (NCT8325) strain with restored rbsU (a positive activator of SigB).}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The cryo-EM Structure of a Novel 40S Kinetoplastid-Specific Ribosomal Protein}, author = {J Brito Querido and E Mancera-Martinez and Q Vicens and A Bochler and J Chicher and A Simonetti and Y Hashem}, url = {https://www.ncbi.nlm.nih.gov/pubmed/29107485?dopt=Abstract}, doi = {10.1016/j.str.2017.09.014}, isbn = {29107485}, year = {2017}, date = {2017-01-01}, journal = {Structure}, volume = {25}, number = {12}, pages = {1785-1794}, abstract = {Kinetoplastids are potentially lethal protozoan pathogens affecting more than 20 million people worldwide. There is a critical need for more specific targets for the development of safer anti-kinetoplastid therapeutic molecules that can replace the scarce and highly cytotoxic current drugs. The kinetoplastid ribosome represents a potential therapeutic target due to its relative structural divergence when compared with its human counterpart. However, several kinetoplastid-specific ribosomal features remain uncharacterized. Here, we present the near-atomic cryoelectron microscopy structure of a novel bona fide kinetoplastid-specific ribosomal (r-) protein (KSRP) bound to the ribosome. KSRP is an essential protein located at the solvent face of the 40S subunit, where it binds and stabilizes kinetoplastid-specific domains of rRNA, suggesting its role in ribosome integrity. KSRP also interacts with the r-protein eS6 at a region that is only conserved in kinetoplastids. The kinetoplastid-specific ribosomal environment of KSRP provides a promising target for the design of safer anti-kinetoplastidian drugs.}, keywords = {PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 Pr55Gag binds genomic and spliced RNAs with different affinity and stoechiometry.}, author = {S Bernacchi and E W Abd El-Wahab and N Dubois and M Hijnen and R P Smyth and J Mak and R Marquet and J C Paillart}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27841704?dopt=Abstract}, doi = {10.1080/15476286.2016.1256533}, isbn = {27841704}, year = {2017}, date = {2017-01-01}, journal = {RNA Biol}, volume = {14}, number = {90}, pages = {103}, abstract = {The HIV-1 Pr55Gag precursor specifically selects genomic RNA (gRNA) from a large variety of cellular and spliced viral RNAs (svRNAs), however the molecular mechanisms of this selective recognition remains poorly understood. To gain better understanding of this process, we analyzed the interactions between Pr55Gag and a large panel of viral RNA (vRNA) fragments encompassing the main packaging signal (Psi) and its flanking regions by fluorescence spectroscopy. We showed that the gRNA harbors a high affinity binding site which is absent from svRNA species, suggesting that this site might be crucial for selecting the HIV-1 genome. Our stoichiometry analysis of protein/RNA complexes revealed that few copies of Pr55Gag specifically associate with the 5メ region of the gRNA. Besides, we found that gRNA dimerization significantly impacts Pr55Gag binding, and we confirmed that the internal loop of stem-loop 1 (SL1) in Psi is crucial for specific interaction with Pr55Gag. Our analysis of gRNA fragments of different length supports the existence of a long-range tertiary interaction involving sequences upstream and downstream of the Psi region. This long-range interaction might promote optimal exposure of SL1 for efficient Pr55Gag recognition. Altogether, our results shed light on the molecular mechanisms allowing the specific selection of gRNA by Pr55Gag amongst a variety of svRNAs, all harboring SL1 in their first common exon.}, keywords = {HIV-1 Pr55Gag fluorescence spectroscopy genomic RNA selection high affinity binding site protein-RNA interaction stoichiometry, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleic acid nanomaterials: Silver-wired DNA}, author = {P Auffinger and E Ennifar}, url = {https://www.ncbi.nlm.nih.gov/pubmed/28937666?dopt=Abstract}, doi = {10.1038/nchem.2869}, isbn = {28937666}, year = {2017}, date = {2017-01-01}, journal = {Nat Chem}, volume = {9}, number = {10}, pages = {932-934}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{sojod_rankranklopg_2017, title = {RANK/RANKL/OPG Signalization Implication in Periodontitis: New Evidence from a RANK Transgenic Mouse Model}, author = {Bouchra Sojod and Danielle Chateau and Christopher G Mueller and Sylvie Babajko and Ariane Berdal and Frédéric Lézot and Beatriz Castaneda}, doi = {10.3389/fphys.2017.00338}, issn = {1664-042X}, year = {2017}, date = {2017-01-01}, journal = {Frontiers in Physiology}, volume = {8}, pages = {338}, abstract = {Periodontitis is based on a complex inflammatory over-response combined with possible genetic predisposition factors. The RANKL/RANK/OPG signaling pathway is implicated in bone resorption through its key function in osteoclast differentiation and activation, as well as in the inflammatory response. This central element of osteo-immunology has been suggested to be perturbed in several diseases, including periodontitis, as it is a predisposing factor for this disease. The aim of the present study was to validate this hypothesis using a transgenic mouse line, which over-expresses RANK (RTg) and develops a periodontitis-like phenotype at 5 months of age. RTg mice exhibited severe alveolar bone loss, an increased number of TRAP positive cells, and disorganization of periodontal ligaments. This phenotype was more pronounced in females. We also observed dental root resorption lacunas. Hyperplasia of the gingival epithelium, including Malassez epithelial rests, was visible as early as 25 days, preceding any other symptoms. These results demonstrate that perturbations of the RANKL/RANK/OPG system constitute a core element of periodontitis, and more globally, osteo-immune diseases.}, keywords = {alveolar bone, gingival epithelium, malassez epithelial rests (MER), Osteoclasts, periodontitis, rank, root resorption, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{russier_few-layer_2017, title = {Few-Layer Graphene Kills Selectively Tumor Cells from Myelomonocytic Leukemia Patients}, author = {Julie Russier and Verónica León and Marco Orecchioni and Eri Hirata and Patrizia Virdis and Claudio Fozza and Francesco Sgarrella and Gianaurelio Cuniberti and Maurizio Prato and Ester Vázquez and Alberto Bianco and Lucia G Delogu}, doi = {10.1002/anie.201700078}, issn = {1521-3773}, year = {2017}, date = {2017-01-01}, journal = {Angewandte Chemie (International Ed. in English)}, volume = {56}, number = {11}, pages = {3014--3019}, abstract = {In the cure of cancer, a major cause of today's mortality, chemotherapy is the most common treatment, though serious frequent challenges are encountered by current anticancer drugs. We discovered that few-layer graphene (FLG) dispersions have a specific killer action on monocytes, showing neither toxic nor activation effects on other immune cells. We confirmed the therapeutic application of graphene on an aggressive type of cancer that is myelomonocytic leukemia, where the monocytes are in their malignant form. We demonstrated that graphene has the unique ability to target and boost specifically the necrosis of monocytic cancer cells. Moreover, the comparison between FLG and a common chemotherapeutic drug, etoposide, confirmed the higher specificity and toxicity of FLG. Since current chemotherapy treatments of leukemia still cause serious problems, these findings open the way to new and safer therapeutic approaches.}, keywords = {Acute, cancer therapy, Chronic, Cultured, graphene, Graphite, Humans, I2CT, Immune System, leukemia, Leukocytes, Mononuclear, Myeloid, Myelomonocytic, myelomonocytic leukemia, Nanomaterials, Particle Size, Surface Properties, Team-Bianco, Tumor Cells}, pubstate = {published}, tppubtype = {article} } @article{volohonsky_transgenic_2017, title = {Transgenic Expression of the Anti-parasitic Factor TEP1 in the Malaria Mosquito Anopheles gambiae}, author = {Gloria Volohonsky and Ann-Katrin Hopp and Mélanie Saenger and Julien Soichot and Heidi Scholze and Jens Boch and Stéphanie A Blandin and Eric Marois}, editor = {Kenneth D Vernick}, url = {http://dx.plos.org/10.1371/journal.ppat.1006113}, doi = {10.1371/journal.ppat.1006113}, issn = {1553-7374}, year = {2017}, date = {2017-01-01}, urldate = {2017-02-01}, journal = {PLOS Pathogens}, volume = {13}, number = {1}, pages = {e1006113}, keywords = {Anopheles gambiae, anti-parasitic factor, blandin, M3i, Malaria, marois, TEP1, transgenic}, pubstate = {published}, tppubtype = {article} } @article{mussabekova_innate_2017, title = {Innate and intrinsic antiviral immunity in Drosophila}, author = {Assel Mussabekova and Laurent Daeffler and Jean-Luc Imler}, doi = {10.1007/s00018-017-2453-9}, issn = {1420-9071}, year = {2017}, date = {2017-01-01}, journal = {Cell. Mol. Life Sci.}, abstract = {The fruit fly Drosophila melanogaster has been a valuable model to investigate the genetic mechanisms of innate immunity. Initially focused on the resistance to bacteria and fungi, these studies have been extended to include antiviral immunity over the last decade. Like all living organisms, insects are continually exposed to viruses and have developed efficient defense mechanisms. We review here our current understanding on antiviral host defense in fruit flies. A major antiviral defense in Drosophila is RNA interference, in particular the small interfering (si) RNA pathway. In addition, complex inducible responses and restriction factors contribute to the control of infections. Some of the genes involved in these pathways have been conserved through evolution, highlighting loci that may account for susceptibility to viral infections in humans. Other genes are not conserved and represent species-specific innovations.}, keywords = {Argonaute 2, Dicer-2, IMD pathway, imler, Jak/STAT pathway, M3i, NF-κB}, pubstate = {published}, tppubtype = {article} } @article{koltun_sumo-targeted_2017, title = {The SUMO-targeted ubiquitin ligase, Dgrn, is essential for Drosophila innate immunity}, author = {Bella Koltun and Eliza Shackelford and François Bonnay and Nicolas Matt and Jean-Marc Reichhart and Amir Orian}, url = {http://www.intjdevbiol.com/paper.php?doi=160250ao}, doi = {10.1387/ijdb.160250ao}, issn = {0214-6282}, year = {2017}, date = {2017-01-01}, urldate = {2017-07-12}, journal = {The International Journal of Developmental Biology}, volume = {61}, number = {3-4-5}, pages = {319--327}, keywords = {Dgrn, Drosophila, innate immunity, Ligase, M3i, matt, reichhart, SUMO, target, ubiquitin}, pubstate = {published}, tppubtype = {article} } @article{Chamy2017, title = {Advances in Myeloid-Like Cell Origins and Functions in the Model Organism Drosophila melanogaster}, author = {Laure El Chamy and Nicolas Matt and Jean-Marc Reichhart}, url = {http://www.asmscience.org/content/journal/microbiolspec/10.1128/microbiolspec.MCHD-0038-2016}, doi = {10.1128/microbiolspec.MCHD-0038-2016}, issn = {2165-0497}, year = {2017}, date = {2017-01-01}, urldate = {2017-07-12}, journal = {Microbiology Spectrum}, volume = {5}, number = {1}, keywords = {Drosophila melanogaster, M3i, matt, Myeloid-Like Cell Origins, reichhart}, pubstate = {published}, tppubtype = {article} } @article{pmid27755105, title = {Reactivation capacity by latency-reversing agents ex vivo correlates with the size of the HIV-1 reservoir}, author = {Gilles Darcis and Sophie Bouchat and Anna Kula and Benoit Van Driessche and Nadège Delacourt and Caroline Vanhulle and Véronique Avettand-Fenoel and Stéphane De Wit and Olivier Rohr and Christine Rouzioux and Carine Van Lint}, doi = {10.1097/QAD.0000000000001290}, issn = {1473-5571}, year = {2017}, date = {2017-01-01}, urldate = {2017-01-01}, journal = {AIDS}, volume = {31}, number = {2}, pages = {181--189}, abstract = {OBJECTIVE: HIV-1 reservoirs are the major hurdle to virus clearance in combination antiretroviral therapy (cART)-treated patients. An approach to eradicating HIV-1 involves reversing latency in cART-treated patients to make latent cells visible to the host immune system. Stimulation of patient cell cultures with latency-reversing agents (LRAs) ex vivo results in heterogeneous responses among HIV-infected patients. Determinants of this heterogeneity are unknown and consequently important to determine.nnDESIGN AND METHODS: Here, we grouped and retrospectively analyzed the data from our two recent HIV-1 reactivation studies to investigate the role of the HIV-1 reservoir size in the reactivation capacity by LRAs in ex vivo cultures of CD8-depleted peripheral blood mononuclear cells (PBMCs) isolated from 54 cART-treated patients and of resting CD4 T cells isolated from 30 cART-treated patients.nnRESULTS: Our results established a statistically relevant positive correlation between the HIV-1 reservoir size measured by total cell-associated HIV-1 DNA and the frequency of positive HIV-1 recovery measurements in response to various LRAs in ex vivo cultures of cells isolated from cART-treated HIV aviremic patients. HIV-1 reservoir size also correlated with the extracellular HIV-1 RNA median level measured in supernatants of cell cultures following LRA treatments. However, we identified HIV patients whose positive measurements frequency and median level of extracellular HIV-1 RNA deviated from linearity relative to their corresponding HIV reservoir size.nnCONCLUSION: We demonstrated that the reservoir size is one predictive marker of LRA effectiveness but this parameter alone is not sufficient. The identification of other predictive markers is necessary to predict the success of HIV anti-latency approaches.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Lee2016, title = {Enterocyte Purge and Rapid Recovery Is a Resilience Reaction of the Gut Epithelium to Pore-Forming Toxin Attack}, author = {Kwang-Zin Lee and Matthieu Lestradet and Catherine Socha and Stefanie Schirmeier and Antonin Schmitz and Caroline Spenlé and Olivier Lefebvre and Céline Keime and Wennida M. Yamba and Richard Bou Aoun and Samuel Liegeois and Yannick Schwab and Patricia Simon-Assmann and Frédéric Dalle and Dominique Ferrandon}, editor = {L Abate}, url = {http://www.sciencedirect.com/science/article/pii/S193131281630436X}, doi = {10.1016/j.chom.2016.10.010}, issn = {1931-3128}, year = {2016}, date = {2016-11-23}, urldate = {2016-11-25}, journal = {Cell Host & Microbe}, abstract = {Summary Besides digesting nutrients, the gut protects the host against invasion by pathogens. Enterocytes may be subjected to damage by both microbial and host defensive responses, causing their death. Here, we report a rapid epithelial response that alleviates infection stress and protects the enterocytes from the action of microbial virulence factors. Intestinal epithelia exposed to hemolysin, a pore-forming toxin secreted by Serratia marcescens, undergo an evolutionarily conserved process of thinning followed by the recovery of their initial thickness within a few hours. In response to hemolysin attack, Drosophila melanogaster enterocytes extrude most of their apical cytoplasm, including damaged organelles such as mitochondria, yet do not lyse. We identify two secreted peptides, the expression of which requires CyclinJ, that mediate the recovery phase in which enterocytes regain their original shape and volume. Epithelial thinning and recovery constitute a fast and efficient response to intestinal infections, with pore-forming toxins acting as alarm signals.}, keywords = {Epithelium, ferrandon, gut, M3i, resilience}, pubstate = {published}, tppubtype = {article} } @article{pmid27725726, title = {HIC1 controls cellular- and HIV-1- gene transcription via interactions with CTIP2 and HMGA1}, author = {Valentin Le Douce and Faezeh Forouzanfar and Sebastian Eilebrecht and Benoit Van Driessche and Amina Ait-Ammar and Roxane Verdikt and Yoshihito Kurashige and Céline Marban and Virginie Gautier and Ermanno Candolfi and Arndt G Benecke and Carine Van Lint and Olivier Rohr and Christian Schwartz}, doi = {10.1038/srep34920}, issn = {2045-2322}, year = {2016}, date = {2016-10-01}, urldate = {2016-10-01}, journal = {Sci Rep}, volume = {6}, pages = {34920}, abstract = {Among many cellular transcriptional regulators, Bcl11b/CTIP2 and HGMA1 have been described to control the establishment and the persistence of HIV-1 latency in microglial cells, the main viral reservoir in the brain. In this present work, we identify and characterize a transcription factor i.e. HIC1, which physically interacts with both Bcl11b/CTIP2 and HMGA1 to co-regulate specific subsets of cellular genes and the viral HIV-1 gene. Our results suggest that HIC1 represses Tat dependent HIV-1 transcription. Interestingly, this repression of Tat function is linked to HIC1 K314 acetylation status and to SIRT1 deacetylase activity. Finally, we show that HIC1 interacts and cooperates with HGMA1 to regulate Tat dependent HIV-1 transcription. Our results also suggest that HIC1 repression of Tat function happens in a TAR dependent manner and that this TAR element may serve as HIC1 reservoir at the viral promoter to facilitate HIC1/TAT interaction.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{poirier_ability_2016, title = {Ability of the marine bacterium Pseudomonas fluorescens BA3SM1 to counteract the toxicity of CdSe nanoparticles.}, author = {Isabelle Poirier and Lauriane Kuhn and Arnaud Demortière and Boris Mirvaux and Philippe Hammann and Johana Chicher and Christelle Caplat and Marie Pallud and Martine Bertrand}, doi = {10.1016/j.jprot.2016.07.021}, issn = {1876-7737 1874-3919}, year = {2016}, date = {2016-10-01}, journal = {Journal of proteomics}, volume = {148}, pages = {213--227}, abstract = {In the marine environment, bacteria from estuarine and coastal sediments are among the first targets of nanoparticle pollution; it is therefore relevant to improve the knowledge of interactions between bacteria and nanoparticles. In this work, the response of the marine bacterium Pseudomonas fluorescens BA3SM1 to CdSe nanocrystals (CdSe NPs) of 3nm (NP3) and 8nm (NP8) in diameter was evaluated through microscopic, physiological, biochemical and proteomic approaches. Transmission electron microscopy images showed that NP3 were able to penetrate the bacteria, while NP8 were highly concentrated around the cells, embedded in large exopolysaccharides. In our experimental conditions, both CdSe NP sizes induced a decrease in respiration during the stationary growth phase, while only NP8 caused growth retardation and a decrease in pyoverdine production. Proteomic analyses highlighted that the strain responded to CdSe NP toxicity by inducing various defence mechanisms such as cell aggregation, extracellular CdSe NP sequestration, effective protection against oxidative stress, modifications of envelope organization and properties, and cadmium export. In addition, BA3SM1 presented a biosorption capacity of 1.6×10(16)NP3/g dry weight and 1.7×10(15)NP8/g dry weight. This strain therefore appears as a promising agent for NP bioremediation processes. Proteomic data are available via ProteomeXchange with identifier PXD004012. BIOLOGICAL SIGNIFICANCE: To the best of our knowledge, this is the first report focussing on the effects of CdSe colloidal nanocrystals (CdSe NPs) on a marine strain of Pseudomonas fluorescens. CdSe NPs are extensively used in the industry of renewable energies and it is regrettably expected that these pollutants will sometime soon appear in the marine environment through surface runoff, urban effluents and rivers. Bacteria living in estuarine and coastal sediments will be among the first targets of these new pollutants. The pseudomonads are frequently found in these ecosystems. They are involved in several biogeochemical cycles and are known for their high resistance to pollutants. Consequently, this study focussing on the effects of CdSe NPs on the marine strain P. fluorescens BA3SM1 is highly relevant for several reasons. First, it aims at improving knowledge about the interactions between bacteria and NPs. This is fundamental to effectively use NPs against pathogenic bacteria. Secondly, in spite of CdSe NP interactions with the bacterial cells, the strain BA3SM1 can develop various strategies to counteract CdSe NP toxicity and ensure its growth. It exhibits interesting properties to sequester CdSe NPs and it retains its ability to form biofilm. The strain therefore appears as a promising agent for NP bioremediation thanks to biofiltration processes. Finally, this study shows that CdSe NPs of 8nm in diameter cause a decrease in the secretion of siderophore pyoverdine, a secondary metabolite playing a key role in microbial ecology since it drives bacterial survival and competitiveness in ecosystems. Bacteria producing effective siderophores survive better in a Fe-deficient environment where they antagonize the growth of other microbes thought iron deprivation. Furthermore, siderophores are also employed as virulence factors in human pathogenic strains such as P. aeruginosa. Consequently, this study highlights that NPs can impact the secondary metabolism of bacteria with environmental and medical implications. In addition, in this work, Data-Dependant Acquisition (DDA) provided state of the art Mass Spectrometry data by Spectral Counting and MS1 Label-Free. The combination of these two well-known proteomic techniques including manual validations strengthened the identification and quantification of regulated proteins. Moreover, numerous correlations between proteomic analyses and other observations (physiological, biochemical, microscopic) consolidated our interpretations.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{jasim_thickness_2016, title = {Thickness of functionalized graphene oxide sheets plays critical role in tissue accumulation and urinary excretion: A pilot PET/CT study}, author = {Dhifaf A Jasim and Herve Boutin and Michael Fairclough and Cécilia Ménard-Moyon and Christian Prenant and Alberto Bianco and Kostas Kostarelos}, url = {http://www.sciencedirect.com/science/article/pii/S2352940716300099}, doi = {10.1016/j.apmt.2016.04.003}, issn = {2352-9407}, year = {2016}, date = {2016-09-01}, urldate = {2020-04-01}, journal = {Applied Materials Today}, volume = {4}, pages = {24--30}, abstract = {We have recently reported that administration of thin graphene oxide (GO) sheets in the systemic circulation of rodents leads to rapid urinary excretion for the majority of injected dose and accumulation by the reticuloendothelial system organs for the remaining dose. In this study, graphene oxide was functionalized with a chelating moiety (DOTA, (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid)) and labeled with [64Cu] for positron emission computed tomography (PET/CT) imaging. The thin functionalized graphene oxide material (f-GO-thin) consisted of a few layers (∼5nm) in thickness. Aging of the f-GO-thin material led to re-stacking of the flakes that resulted in materials of increased thickness (f-GO-thick) without altering their lateral dimension. These two types of f-GOs were comparatively studied pharmacologically to reveal the previously unexplored in vivo role of graphene oxide sheet thickness. Our results showed that a significantly larger fraction of the thicker GO sheets (47.5% of injected dose) remained within the body of living animals 24h after intravenous administration, residing mainly in the spleen and liver. The thinner GO sheets were predominantly (76.9% of injected dose) excreted through the glomerular filter into the urine. This pilot study provides an initial correlation between graphene-based material structure and pharmacological profile that is imperative towards understanding of how 2D structures behave in vivo to give information on potential biomedical applications.}, keywords = {carbon, I2CT, imaging, Nanomedicine, Pharmacokinetics, Pharmacology, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ehrhardt_redox-cycler_2016, title = {The redox-cycler plasmodione is a fast acting antimalarial lead compound with pronounced activity against sexual and early asexual blood-stage parasites}, author = {Katharina Ehrhardt and Christiane Deregnaucourt and Alice-Anne Goetz and Tzvetomira Tzanova and Valentina Gallo and Paolo Arese and Bruno Pradines and Sophie H Adjalley and Denyse Bagrel and Stephanie A Blandin and Michael Lanzer and Elisabeth Davioud-Charvet}, url = {http://aac.asm.org/content/60/9/5146}, doi = {10.1128/AAC.02975-15}, issn = {1098-6596}, year = {2016}, date = {2016-09-01}, journal = {Antimicrob. Agents Chemother.}, volume = {60}, number = {9}, pages = {5146-5158}, abstract = {Previously, we presented the chemical design of a promising series of antimalarial agents, 3-[substituted-benzyl]-menadiones, with potent in vitro and in vivo activities. Ongoing studies on the mode of action of antimalarial 3-[substituted-benzyl]-menadiones revealed that these agents disturb the redox balance of the parasitized erythrocyte by acting as redox-cyclers - a strategy that is broadly recognized for the development of new antimalarial agents. Here, we report a detailed parasitological characterization of the in vitro activity profile of the lead compound 3-[4-(trifluoromethyl)benzyl]-menadione 1c (henceforth called plasmodione) against intra-erythrocytic stages of the human malaria parasite Plasmodium falciparum We show that plasmodione acts rapidly against asexual blood stages, thereby disrupting the clinically relevant intra-erythrocytic life cycle of the parasite, and furthermore has potent activity against early gametocytes. The lead's antiplasmodial activity was unaffected by the most common resistance mechanisms to clinically used antimalarials. Moreover, plasmodione has a low potential to induce drug resistance and a fast killing speed as observed by culturing parasites under continuous drug pressure. Drug interactions with licensed antimalarial drugs were also established using the fixed-ratio isobologram method. Initial toxicological profiling suggests that it is a safe agent for possible human use. Our studies identify plasmodione as a promising antimalarial lead compound and strongly support the future development of redox-active benzylmenadiones as antimalarial agents.}, keywords = {antimalarial, blandin, M3i, parasites, Plasmodium, redox-cycler}, pubstate = {published}, tppubtype = {article} } @inbook{Paro0000, title = {Encyclopedia of Immunobiology}, author = {Simona Paro and Jean-Luc Imler}, editor = {M Ratcliffe}, year = {2016}, date = {2016-08-01}, volume = {1}, pages = {454-461}, publisher = {Elsevier}, chapter = {“Immunity in insects”}, keywords = {imler, Immunity, Insect, M3i}, pubstate = {published}, tppubtype = {inbook} } @book{leclerc2016, title = {Biologie du développement}, author = {Daniel Boujard and Vincent Leclerc and Stéphane Vincent}, editor = {Editions Dunod}, url = {https://www.dunod.com/sciences-techniques/biologie-du-developpement}, isbn = {978-2-10-072085-9}, year = {2016}, date = {2016-06-01}, booktitle = {Biologie du développement}, pages = {304}, edition = {Editions Dunod}, crossref = {EAN: 9782100720859}, abstract = {La biologie du développement étudie les phénomènes qui permettent le passage d’un œuf à un organisme adulte. Cette discipline a beaucoup évolué ces dernières années avec l'essor des nouvelles techniques d’imagerie in vivo, des « omiques » et des techniques de transgénèse. Aux progrès technologiques s’ajoutent les avancées importantes en génétique et épigénétique, tout cela permettant d’avoir aujourd’hui une vision beaucoup plus intégrée des mécanismes fondamentaux de la biologie du développement. L’objectif de l’ouvrage est de fournir une vision synthétique et moderne de la discipline tout en montrant son lien avec la biologie cellulaire, la génétique, la physiologie et la médecine. Les principaux modèles, historiques ou émergents, sont présentés puis les grandes étapes du développement sont détaillées en s’appuyant sur les données les plus récentes. Un chapitre aborde l’importance de l’apport de la biologie du développement dans la compréhension de nombreuses pathologies, dont le cancer.}, keywords = {biologie, CAPES, développement, Licence, Master, Médecine}, pubstate = {published}, tppubtype = {book} } @article{arnold_autophagy_2016, title = {Autophagy is dispensable for B-cell development but essential for humoral autoimmune responses}, author = {J Arnold and D Murera and F Arbogast and J -D Fauny and S Muller and F Gros}, doi = {10.1038/cdd.2015.149}, issn = {1476-5403}, year = {2016}, date = {2016-05-01}, journal = {Cell Death and Differentiation}, volume = {23}, number = {5}, pages = {853--864}, abstract = {To gain new insight into the role of B-cell autophagy, we generated two novel mouse models deficient for the autophagy-related gene (Atg)5, one from the outset pro-B cell stage (Atg5(f/-) Mb1 cre) and the other in mature B cells only (Atg5(f/-) CD21 cre). We show that autophagy is dispensable for pro- to pre-B cell transition, but necessary at a basal level to maintain normal numbers of peripheral B cells. It appears non-essential for B-cell activation under B-cell receptor stimulation but required for their survival after lipopolysaccharide stimulation that drives plasmablast differentiation and for specific IgM production after immunization. Results obtained using Atg5(f/-) CD21 cre × C57BL/6(lpr/lpr) autoimmune-prone mice show that B-cell autophagy is involved in the maintenance of anti-nuclear antibody secretion, elevated number of long-lived plasma cells, and sustains IgG deposits in the kidneys. Thus, treatments specifically targeting autophagy might be beneficial in systemic autoimmune diseases.}, keywords = {I2CT, Imagerie}, pubstate = {published}, tppubtype = {article} } @article{modugno_comparative_2016, title = {A comparative study on the enzymatic biodegradability of covalently functionalized double- and multi-walled carbon nanotubes}, author = {Gloria Modugno and Fayçal Ksar and Alessia Battigelli and Julie Russier and Pierre Lonchambon and Edelma Eleto da Silva and Cécilia Ménard-Moyon and Brigitte Soula and Anne-Marie Galibert and Mathieu Pinault and Emmanuel Flahaut and Martine Mayne-L'Hermite and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S0008622316300239}, doi = {10.1016/j.carbon.2016.01.023}, issn = {0008-6223}, year = {2016}, date = {2016-04-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {100}, pages = {367--374}, abstract = {The assessment of the biodegradability potential of carbon nanotubes (CNTs) is a fundamental point towards their applications in materials science and biomedicine. Due to the continuous concerns about the fate of such type of nanomaterials, it is very important to understand if they can undergo degradation under certain conditions and if the morphology and structure of the nanotubes play a role in this process. For this purpose we have decided to undertake a comparative study on the enzymatic degradation of CNTs with concentric multilayers. Double-walled (DW) and multi-walled (MW) CNTs of various lengths, degrees of oxidation and functionalizations using different methods were treated with horseradish peroxidase (HRP). While all tested DWCNTs resulted resistant to the biodegradation, some of the MWCNTs were partially degraded by the enzyme. We have found that short oxidized multi-walled CNTs functionalized by amidation were reduced in length and presented a high amount of defects at the end of the period of treatment with HRP. This comparative study holds its importance in the understanding of the structural changes of different types of nanotubes towards the catalytic enzymatic degradation and will help to design safer CNTs for future applications.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{muzi_examining_2016, title = {Examining the impact of multi-layer graphene using cellular and amphibian models}, author = {Laura Muzi and Florence Mouchet and Stéphanie Cadarsi and Izabela Janowska and Julie Russier and Cécilia Ménard-Moyon and Gianfranco Risuleo and Brigitte Soula and Anne-Marie Galibert and Emmanuel Flahaut and Eric Pinelli and Laury Gauthier and Alberto Bianco}, url = {https://doi.org/10.1088%2F2053-1583%2F3%2F2%2F025009}, doi = {10.1088/2053-1583/3/2/025009}, issn = {2053-1583}, year = {2016}, date = {2016-04-01}, urldate = {2020-04-01}, journal = {2D Materials}, volume = {3}, number = {2}, pages = {025009}, abstract = {In the last few years, graphene has been defined as the revolutionary material showing an incredible expansion in industrial applications. Different graphene forms have been applied in several contexts, spreading from energy technologies and electronics to food and agriculture technologies. Graphene showed promises also in the biomedical field. Hopeful results have been already obtained in diagnostic, drug delivery, tissue regeneration and photothermal cancer ablation. In view of the enormous development of graphene-based technologies, a careful assessment of its impact on health and environment is demanded. It is evident how investigating the graphene toxicity is of fundamental importance in the context of medical purposes. On the other hand, the nanomaterial present in the environment, likely to be generated all along the industrial life-cycle, may have harmful effects on living organisms. In the present work, an important contribution on the impact of multi-layer graphene (MLG) on health and environment is given by using a multifaceted approach. For the first purpose, the effect of the material on two mammalian cell models was assessed. Key cytotoxicity parameters were considered such as cell viability and inflammatory response induction. This was combined with an evaluation of MLG toxicity towards Xenopus laevis, used as both in vivo and environmental model organism.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ibrahim_improvement_2016, title = {Improvement of Mitochondria Extract from Saccharomyces cerevisiae Characterization in Shotgun Proteomics Using Sheathless Capillary Electrophoresis Coupled to Tandem Mass Spectrometry.}, author = {Marianne Ibrahim and Rabah Gahoual and Ludovic Enkler and Hubert Dominique Becker and Johana Chicher and Philippe Hammann and Yannis-Nicolas François and Lauriane Kuhn and Emmanuelle Leize-Wagner}, doi = {10.1093/chromsci/bmw005}, issn = {1945-239X 0021-9665 0021-9665}, year = {2016}, date = {2016-04-01}, journal = {Journal of chromatographic science}, volume = {54}, number = {4}, pages = {653--663}, abstract = {In this work, we describe the characterization of a quantity-limited sample (100 ng) of yeast mitochondria by shotgun bottom-up proteomics. Sample characterization was carried out by sheathless capillary electrophoresis, equipped with a high sensitivity porous tip and coupled to tandem mass spectrometry (CESI-MS-MS) and concomitantly with a state-of-art nano flow liquid chromatography coupled to a similar mass spectrometry (MS) system (nanoLC-MS-MS). With single injections, both nanoLC-MS-MS and CESI-MS-MS 60 min-long separation experiments allowed us to identify 271 proteins (976 unique peptides) and 300 proteins (1,765 unique peptides) respectively, demonstrating a significant specificity and complementarity in identification depending on the physicochemical separation employed. Such complementary, maximizing the number of analytes detected, presents a powerful tool to deepen a biological sample's proteomic characterization. A comprehensive study of the specificity provided by each separating technique was also performed using the different properties of the identified peptides: molecular weight, mass-to-charge ratio (m/z), isoelectric point (pI), sequence coverage or MS-MS spectral quality enabled to determine the contribution of each separation. For example, CESI-MS-MS enables to identify larger peptides and eases the detection of those having extreme pI without impairing spectral quality. The addition of peptides, and therefore proteins identified by both techniques allowed us to increase significantly the sequence coverages and then the confidence of characterization. In this study, we also demonstrated that the two yeast enolase isoenzymes were both characterized in the CESI-MS-MS data set. The observation of discriminant proteotypic peptides is facilitated when a high number of precursors with high-quality MS-MS spectra are generated.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{said_structural_2016, title = {Structural characterization of antibody drug conjugate by a combination of intact, middle-up and bottom-up techniques using sheathless capillary electrophoresis - Tandem mass spectrometry as nanoESI infusion platform and separation method.}, author = {Nassur Said and Rabah Gahoual and Lauriane Kuhn and Alain Beck and Yannis-Nicolas François and Emmanuelle Leize-Wagner}, doi = {10.1016/j.aca.2016.03.006}, issn = {1873-4324 0003-2670}, year = {2016}, date = {2016-04-01}, journal = {Analytica chimica acta}, volume = {918}, pages = {50--59}, abstract = {Antibody-drug conjugates (ADCs) represent a fast growing class of biotherapeutic products. Their production leads to a distribution of species exhibiting different number of conjugated drugs overlaying the inherent complexity resulting from the monoclonal antibody format, such as glycoforms. ADCs require an additional level of characterization compared to first generation of biotherapeutics obtained through multiple analytical techniques for complete structure assessment. We report the development of complementary approaches implementing sheathless capillary electrophoresis-mass spectrometry (sheathless CE-MS) to characterize the different aspects defining the structure of brentuximab vedotin. Native MS using sheathless CE-MS instrument as a nanoESI infusion platform enabled accurate mass measurements and estimation of the average drug to antibody ratio alongside to drug load distribution. Middle-up analysis performed after limited IdeS proteolysis allowed to study independently the light chain, Fab and F(ab')2 subunits incorporating 1, 0 to 4 and 0 to 8 payloads respectively. Finally, a CZE-ESI-MS/MS methodology was developed in order to be compatible with hydrophobic drug composing ADCs. From a single injection, complete sequence coverage could be achieved. Using the same dataset, glycosylation and drug-loaded peptides could be simultaneously identified revealing robust information regarding their respective localization and abundance. Drug-loaded peptide fragmentation mass spectra study demonstrated drug specific fragments reinforcing identification confidence, undescribed so far. Results reveal the method ability to characterize ADCs primary structure in a comprehensive manner while reducing tremendously the number of experiments required. Data generated showed that sheathless CZE-ESI-MS/MS characteristics position the methodology developed as a relevant alternative for comprehensive multilevel characterization of these complex biomolecules.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{zhang_carbon_2016, title = {Carbon science in 2016: Status, challenges and perspectives}, author = {Jin Zhang and Mauricio Terrones and Chong Rae Park and Rahul Mukherjee and Marc Monthioux and Nikhil Koratkar and Yern Seung Kim and Robert Hurt and Elzbieta Frackowiak and Toshiaki Enoki and Yuan Chen and Yongsheng Chen and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S0008622315006818}, doi = {10.1016/j.carbon.2015.11.060}, issn = {0008-6223}, year = {2016}, date = {2016-03-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {98}, pages = {708--732}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{chypre_characterization_2016, title = {Characterization and application of two RANK-specific antibodies with different biological activities}, author = {M Chypre and J Seaman and O G Cordeiro and L Willen and K A Knoop and A Buchanan and R C Sainson and I R Williams and H Yagita and P Schneider and C G Mueller}, doi = {10.1016/j.imlet.2016.01.003}, year = {2016}, date = {2016-03-01}, journal = {Immunol.Lett.}, volume = {171}, number = {1879-0542 (Electronic)}, pages = {5--14}, abstract = {Antibodies play an important role in therapy and investigative biomedical research. The TNF-family member Receptor Activator of NF-kappaB (RANK) is known for its role in bone homeostasis and is increasingly recognized as a central player in immune regulation and epithelial cell activation. However, the study of RANK biology has been hampered by missing or insufficient characterization of high affinity tools that recognize RANK. Here, we present a careful description and comparison of two antibodies, RANK-02 obtained by phage display (Newa, 2014 [1]) and R12-31 generated by immunization (Kamijo, 2006 [2]). We found that both antibodies recognized mouse RANK with high affinity, while RANK-02 and R12-31 recognized human RANK with high and lower affinities, respectively. Using a cell apoptosis assay based on stimulation of a RANK:Fas fusion protein, and a cellular NF-kappaB signaling assay, we showed that R12-31 was agonist for both species. R12-31 interfered little or not at all with the binding of RANKL to RANK, in contrast to RANK-02 that efficiently prevented this interaction. Depending on the assay and species, RANK-02 was either a weak agonist or a partial antagonist of RANK. Both antibodies recognized human Langerhans cells, previously shown to express RANK, while dermal dendritic cells were poorly labeled. In vivo R12-31 agonist activity was demonstrated by its ability to induce the formation of intestinal villous microfold cells in mice. This characterization of two monoclonal antibodies should now allow better evaluation of their application as therapeutic reagents and investigative tools}, keywords = {Activation, Animals, ANTAGONIST, Antibodies, antibody, Antibody Affinity, Apoptosis, Assay, Cell Differentiation, Cell Surface Display Techniques, Cellular, Chemistry, comparison, Dendritic Cells, DERMAL DENDRITIC CELLS, Epithelial Cells, Epithelial microfold cell, Epitopes, Fusion, FUSION PROTEIN, HEK293 Cells, Homeostasis, Human, Humans, immune regulation, Immunization, Immunology, Immunomodulation, immunopathology, In vivo, Inbred C57BL, Intestines, Jurkat Cells, Langerhans cell, Langerhans Cells, Mice, Monoclonal, monoclonal antibody, MONOCLONAL-ANTIBODY, mouse, NF-kappa B, NF-kappaB, pathology, Protein, rank, RANK (TNFRSF11a), Receptor, Receptor Activator of Nuclear Factor-kappa B, Regulation, Secondary, Signal Transduction, signaling, Team-Mueller, therapy}, pubstate = {published}, tppubtype = {article} } @article{marangon_synergic_2016, title = {Synergic mechanisms of photothermal and photodynamic therapies mediated by photosensitizer/carbon nanotube complexes}, author = {Iris Marangon and Cécilia Ménard-Moyon and Amanda K A Silva and Alberto Bianco and Nathalie Luciani and Florence Gazeau}, url = {http://www.sciencedirect.com/science/article/pii/S0008622315301421}, doi = {10.1016/j.carbon.2015.08.023}, issn = {0008-6223}, year = {2016}, date = {2016-02-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {97}, pages = {110--123}, series = {BIOMEDICAL APPLICATIONS OF CARBON NANOMATERIALS}, abstract = {We report the construct of a nanosystem based on multi-walled carbon nanotubes (MWCNT) and the photosensitizer m-tetrahydroxyphenylchlorin (mTHPC) for cancer treatment by combination of photodynamic (PDT) and photothermal (PTT) therapy. The photoactivity of the complex mTHPC/MWCNT was studied revealing quenching of the photosensitizer associated to CNT and almost total release of the photosensitizer into the cell cytoplasm after uptake by SKOV3 ovarian cancer cells. The photothermal and photodynamic cytotoxicity of these mTHPC/MWCNT on/off complexes was assessed at the cell level by viability test, imaging flow cytometry, confocal microscopy and transmission electron microscopy and at the molecular level by a proteomic analysis of apoptosis-related proteins and genomic analysis of 84 genes involved in oxidative stress. Cytotoxicity correlated at the cell level to the uptake of mTHPC/MWCNT, while PDT and PTT treatment induced different signaling pathways leading to cell apoptosis. For the first time, the mechanisms of PDT/PTT synergy in eradication of cancer cells were studied revealing that distinct cell responses to PDT and PTT defeat the cell defense to oxidative stress.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bianco_introduction_2016, title = {Introduction to the special issue on biomedical applications of carbon nanomaterials}, author = {Alberto Bianco and Zhuang Liu}, url = {http://www.sciencedirect.com/science/article/pii/S0008622315006314}, doi = {10.1016/j.carbon.2015.09.043}, issn = {0008-6223}, year = {2016}, date = {2016-02-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {97}, pages = {124--125}, series = {BIOMEDICAL APPLICATIONS OF CARBON NANOMATERIALS}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{lamiable_cytokine_2016, title = {Cytokine Diedel and a viral homologue suppress the IMD pathway in Drosophila}, author = {Olivier Lamiable and Christine Kellenberger and Cordula Kemp and Laurent Troxler and Nadège Pelte and Michael Boutros and Joao Trindade Marques and Laurent Daeffler and Jules A Hoffmann and Alain Roussel and Jean-Luc Imler}, url = {http://www.pnas.org/content/113/3/698.abstract}, doi = {10.1073/pnas.1516122113}, issn = {0027-8424, 1091-6490}, year = {2016}, date = {2016-01-19}, urldate = {2016-01-07}, journal = {PNAS}, volume = {113}, number = {3}, pages = {698–703}, abstract = {Viruses are obligatory intracellular parasites that suffer strong evolutionary pressure from the host immune system. Rapidly evolving viral genomes can adapt to this pressure by acquiring genes that counteract host defense mechanisms. For example, many vertebrate DNA viruses have hijacked cellular genes encoding cytokines or cytokine receptors to disrupt host cell communication. Insect viruses express suppressors of RNA interference or apoptosis, highlighting the importance of these cell intrinsic antiviral mechanisms in invertebrates. Here, we report the identification and characterization of a family of proteins encoded by insect DNA viruses that are homologous to a 12-kDa circulating protein encoded by the virus-induced Drosophila gene diedel (die). We show that die mutant flies have shortened lifespan and succumb more rapidly than controls when infected with Sindbis virus. This reduced viability is associated with deregulated activation of the immune deficiency (IMD) pathway of host defense and can be rescued by mutations in the genes encoding the homolog of IKKγ or IMD itself. Our results reveal an endogenous pathway that is exploited by insect viruses to modulate NF-κB signaling and promote fly survival during the antiviral response.}, keywords = {antiviral immunity, bioinformatic, cytokine, Edin, hoffmann, imler, M3i, Sindbis Virus, virokine}, pubstate = {published}, tppubtype = {article} } @article{, title = {iSpinach: a fluorogenic RNA aptamer optimized for in vitro applications.}, author = {A Autour and E Westhof and M Ryckelynck}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26932363?dopt=Abstract}, doi = {10.1093/nar/gkw083}, isbn = {26932363}, year = {2016}, date = {2016-01-01}, journal = {Nucleic Acids Res}, volume = {44}, number = {6}, pages = {2491-2500}, abstract = {Using random mutagenesis and high throughput screening by microfluidic-assisted In Vitro Compartmentalization, we report the isolation of an order of magnitude times brighter mutants of the light-up RNA aptamers Spinach that are far less salt-sensitive and with a much higher thermal stability than the parent molecule. Further engineering gave iSpinach, a molecule with folding and fluorescence properties surpassing those of all currently known aptamer based on the fluorogenic co-factor 3,5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI). We illustrate the potential of iSpinach in a new sensitive and high throughput-compatible fluorogenic assay that measures co-transcriptionally the catalytic constant (kcat) of a model ribozyme.}, keywords = {RYCKELYNCK, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Sodium and Potassium Interactions with Nucleic Acids}, author = {P Auffinger and L D'Ascenzo and E Ennifar}, editor = {A Sigel and H Sigel and R Sigel}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26860302?dopt=Abstract}, doi = {10.1007/978-3-319-21756-7_6}, isbn = {26860302}, year = {2016}, date = {2016-01-01}, booktitle = {The Alkali Metal Ions: Their Role for Life}, volume = {16}, pages = {167-201}, publisher = {Springer}, series = {Metal Ions in Life Sciences}, abstract = {Metal ions are essential cofactors for the structure and functions of nucleic acids. Yet, the early discovery in the 70s of the crucial role of Mg(2+) in stabilizing tRNA structures has occulted for a long time the importance of monovalent cations. Renewed interest in these ions was brought in the late 90s by the discovery of specific potassium metal ions in the core of a group I intron. Their importance in nucleic acid folding and catalytic activity is now well established. However, detection of K(+) and Na(+) ions is notoriously problematic and the question about their specificity is recurrent. Here we review the different methods that can be used to detect K(+) and Na(+) ions in nucleic acid structures such as X-ray crystallography, nuclear magnetic resonance or molecular dynamics simulations. We also discuss specific versus non-specific binding to different structures through various examples.}, keywords = {DNA Hydration K+ Metal binding Molecular dynamics simulations Monovalent ions Na+·NMR Potassium RNA Sodium Solvation X-ray crystallography, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Reduced DICER1 expression bestows rheumatoid arthritis synoviocytes proinflammatory properties and resistance to apoptotic stimuli.}, author = {G Alsaleh and R Nehmar and S Blüml and C Schleiss and E Ostermann and J P Dillenseger and A Sayeh and P Choquet and D Dembele and A Francois and J H Salmon and N Paul and G Schabbauer and G Bierry and A Meyer and J E Gottenberg and G Haas and S Pfeffer and L Vallat and J Sibilia and S Bahram and P Georgel}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26882526}, doi = {10.1002/art.39641}, isbn = {26882526}, year = {2016}, date = {2016-01-01}, journal = {Arthritis Rheumatol}, volume = {68}, number = {8}, pages = {1839-1848}, abstract = {Objectives While the regulatory role of individual microRNAs in rheumatoid arthritis is well established, the role of DICER1 in the pathogenesis of the disease has not yet been investigated. Here, we analyze the expression of factors involved in miRNA biogenesis in synoviocytes (FLS) from RA patients and monitored arthritis triggered by K/BxN serum transfer in Dicer-deficient mice. Methods Genes and precursor miRNAs expression was quantified by RT-qPCR. miRNAs macroarray profiling was monitored by RT-qPCR. Cytokines were quantified by ELISA. Experimental arthritis in mice was achieved by serum transfer from K/BxN donors. Apoptosis was quantified using an ELISA assay. Results Here we report decreased DICER1 and mature miRNA expression in synovial fibroblasts isolated from rheumatoid arthritis (RA) patients. These cells are hyperresponsive to LPS, as evidenced by increased IL-6 secretion upon stimulation. Experimental serum transfer arthritis in Dicer mouse mutants confirmed that unbalanced miRNAs biogenesis correlates with enhanced inflammatory response. Finally, synoviocytes from both RA patients and from Dicer mutant mouse exhibit increased resistance to apoptotic stimuli. Conclusion Our work further substantiates the important role of DICER1 in the maintenance of homeostasis and the regulation of inflammatory responses. This article is protected by copyright. All rights reserved.}, keywords = {DICER1 inflammation microRNA rheumatoid arthritis, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @incollection{, title = {Synthetic Capped mRNAs for Cap-Specific Photo-Cross-Linking Experiments.}, author = {J Kowalska and F Martin and J Jemielity}, editor = {R Rhoads}, year = {2016}, date = {2016-01-01}, booktitle = {Synthetic mRNA: Production, Introduction Into Cells, and Physiological Consequences}, volume = {1428}, pages = {31-43}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {he 7-methylguanosine triphosphate cap present at the 5' ends of eukaryotic mRNAs plays numerous roles in mRNA expression and metabolism. The identification and studies on cap-binding partners can be significantly advanced using tailored chemical tools such as synthetic cap analogues or RNAs carrying modified cap structures. Here we provide protocols for the production of mRNAs specifically labeled within the 5' cap with a nucleoside capable of being photo-activated, either 6-thioguanosine or 7-methyl-6-thioguanosine, which can be used in photo-cross-linking experiments to identify or characterize cap-binding biomolecules. We also describe a protocol for the cross-linking experiments with capped RNAs to map histone H4 cap-binding pocket.}, keywords = {ERIANI 5′ cap analogs 6-thioguanosine Cap binding Histone mRNA Photo-cross-linking Transcription mRNA stability}, pubstate = {published}, tppubtype = {incollection} } @article{belval_display_2016, title = {Display of whole proteins on inner and outer surfaces of Grapevine fanleaf virus-like particles}, author = {Lorène Belval and Caroline Hemmer and Claude Sauter and Catherine Reinbold and Jean-Daniel Fauny and François Berthold and Léa Ackerer and Corinne Schmitt-Keichinger and Olivier Lemaire and Gérard Demangeat and Christophe Ritzenthaler}, doi = {10.1111/pbi.12582}, issn = {1467-7652}, year = {2016}, date = {2016-01-01}, journal = {Plant Biotechnology Journal}, abstract = {Virus-like particles (VLPs) derived from non-enveloped viruses result from the self-assembly of capsid proteins (CPs). They generally display similar structural features to viral particles but are non-infectious and their inner cavity and outer surface can potentially be adapted to serve as nanocarriers of great biotechnological interest. While a VLP outer surface is generally amenable to chemical or genetic modifications, encaging a cargo within particles can be more complex and is often limited to small molecules or peptides. Examples where both inner cavity and outer surface have been used to simultaneously encapsulate and expose entire proteins remain scarce. Here we describe the production of spherical VLPs exposing fluorescent proteins at either their outer surface or inner cavity as a result of the self-assembly of a single genetically modified viral structural protein, the CP of grapevine fanleaf virus (GFLV). We found that the N- and C-terminal ends of the GFLV CP allow the genetic fusion of proteins as large as 27 kDa and the plant-based production of nucleic-acid free VLPs. Remarkably, expression of N- or C-terminal CP fusions, resulted in the production of VLPs with recombinant proteins exposed either to the inner cavity or the outer surface, respectively, while coexpression of both fusion proteins led to the formation hybrid VLP, although rather inefficiently. Such properties are rather unique for a single viral structural protein and open new potential avenues for the design of safe and versatile nanocarriers, particularly for the targeted delivery of bioactive molecules. This article is protected by copyright. All rights reserved.}, keywords = {I2CT, Imagerie, SAUTER}, pubstate = {published}, tppubtype = {article} } @article{sawaf_follicular_2016, title = {Follicular Helper Ŧ Cells in Systemic Lupus Erythematosus: Why Should They Be Considered as Interesting Therapeutic Targets?}, author = {Matthieu Sawaf and Hélène Dumortier and Fanny Monneaux}, doi = {10.1155/2016/5767106}, issn = {2314-7156}, year = {2016}, date = {2016-01-01}, journal = {Journal of Immunology Research}, volume = {2016}, pages = {5767106}, abstract = {Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by B cell hyperactivity leading to the production of autoantibodies, some of which having a deleterious effect. Reducing autoantibody production thus represents a way of controlling lupus pathogenesis, and a better understanding of the molecular and cellular factors involved in the differentiation of B cells into plasma cells could allow identifying new therapeutic targets. Follicular helper T cells (TFH) represent a distinct subset of CD4(+) T cells specialized in providing help to B cells. They are required for the formation of germinal centers and the generation of long-lived serological memory and, as such, are suspected to play a central role in SLE. Recent advances in the field of TFH biology have allowed the identification of important molecular factors involved in TFH differentiation, regulation, and function. Interestingly, some of these TFH-related molecules have been described to be dysregulated in lupus patients. In the present review, we give an overview of the aberrant expression and/or function of such key players in lupus, and we highlight their potential as therapeutic targets.}, keywords = {Adult, Autoantibodies, B-Lymphocytes, Cell Differentiation, Dumortier, Germinal Center, Helper-Inducer, Humans, I2CT, Lupus Erythematosus, Molecular Targeted Therapy, Monneaux, Plasma Cells, Systemic, T-Lymphocytes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{spinato_different_2016, title = {Different chemical strategies to aminate oxidised multi-walled carbon nanotubes for siRNA complexation and delivery}, author = {Cinzia Spinato and Davide Giust and Isabella Anna Vacchi and Cécilia Ménard-Moyon and Kostas Kostarelos and Alberto Bianco}, url = {https://pubs.rsc.org/en/content/articlelanding/2016/tb/c5tb02088c}, doi = {10.1039/C5TB02088C}, issn = {2050-7518}, year = {2016}, date = {2016-01-01}, urldate = {2020-04-01}, journal = {Journal of Materials Chemistry B}, volume = {4}, number = {3}, pages = {431--441}, abstract = {In this work, we have investigated the preparation of amino-functionalised multi-walled carbon nanotubes (MWCNTs) as potential carriers for the delivery of siRNA. Several studies have shown promising results exploiting functionalised CNTs for the delivery of genetic material in vitro and in vivo. Our groups have previously observed that the type of surface functionalisation used to modify oxidised MWCNTs (oxMWCNTs) can lead to significant differences in nanotube cellular uptake and delivery capability. In those studies, amino-functionalised CNTs were obtained by cycloaddition reactions. Here, we focused on the direct conversion of the carboxylic groups present on oxMWCNTs into amines, and we attempted different synthetic strategies in order to directly tether the amines onto the CNTs, without extending the lateral chain. The functionalised material was characterised by X-ray photoelectron spectroscopy, Fourier transform infra-red spectroscopy and transmission electron microscopy, and the most water-dispersible CNTs were selected for siRNA complexation and cellular uptake studies. The aminated conjugates are demonstrated to be promising vectors to achieve intracellular transport of genetic information.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{rottloff_proteome_2016, title = {Proteome analysis of digestive fluids in Nepenthes pitchers.}, author = {Sandy Rottloff and Sissi Miguel and Flore Biteau and Estelle Nisse and Philippe Hammann and Lauriane Kuhn and Johana Chicher and Vincent Bazile and Laurence Gaume and Benoit Mignard and Alain Hehn and Frédéric Bourgaud}, doi = {10.1093/aob/mcw001}, issn = {1095-8290 0305-7364 0305-7364}, year = {2016}, date = {2016-01-01}, journal = {Annals of botany}, volume = {117}, number = {3}, pages = {479--495}, abstract = {BACKGROUND AND AIMS: Carnivorous plants have developed strategies to enable growth in nutrient-poor soils. For the genus Nepenthes, this strategy represents producing pitcher-modified leaves that can trap and digest various prey. These pitchers produce a digestive fluid composed of proteins, including hydrolytic enzymes. The focus of this study was on the identification of these proteins. METHODS: In order to better characterize and have an overview of these proteins, digestive fluid was sampled from pitchers at different stages of maturity from five species of Nepenthes (N. mirabilis, N. alata, N. sanguinea, N. bicalcarata and N. albomarginata) that vary in their ecological niches and grew under different conditions. Three complementary approaches based on transcriptomic resources, mass spectrometry and in silico analysis were used. KEY RESULTS: This study permitted the identification of 29 proteins excreted in the pitchers. Twenty of these proteins were never reported in Nepenthes previously and included serine carboxypeptidases, α- and β-galactosidases, lipid transfer proteins and esterases/lipases. These 20 proteins display sequence signals allowing their secretion into the pitcher fluid. CONCLUSIONS: Nepenthes pitcher plants have evolved an arsenal of enzymes to digest prey caught in their traps. The panel of new proteins identified in this study provides new insights into the digestive process of these carnivorous plants.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{dietrich_interleukin-36_2016, title = {Interleukin-36 potently stimulates human M2 macrophages, Langerhans cells and keratinocytes to produce pro-inflammatory cytokines}, author = {Damien Dietrich and Praxedis Martin and Vincent Flacher and Yu Sun and David Jarrossay and Nicolo Brembilla and Christopher Mueller and Heather A Arnett and Gaby Palmer and Jennifer Towne and Cem Gabay}, doi = {10.1016/j.cyto.2016.05.012}, issn = {1096-0023}, year = {2016}, date = {2016-01-01}, journal = {Cytokine}, volume = {84}, pages = {88--98}, abstract = {Interleukin (IL)-36 cytokines belong to the IL-1 family and include three agonists, IL-36 α, β and γ and one inhibitor, IL-36 receptor antagonist (IL-36Ra). IL-36 and IL-1 (α and β) activate similar intracellular pathways via their related heterodimeric receptors, IL-36R/IL-1RAcP and IL-1R1/IL-1RAcP, respectively. However, excessive IL-36 versus IL-1 signaling induces different phenotypes in humans, which may be related to differential expression of their respective receptors. We examined the expression of IL-36R, IL-1R1 and IL-1RAcP mRNA in human peripheral blood, tonsil and skin immune cells by RT-qPCR. Monocyte-derived dendritic cells (MDDC), M0, M1 or M2-polarized macrophages, primary keratinocytes, dermal macrophages and Langerhans cells (LC) were stimulated with IL-1β or IL-36β. Cytokine production was assessed by RT-qPCR and immunoassays. The highest levels of IL-36R mRNA were found in skin-derived keratinocytes, LC, dermal macrophages and dermal CD1a(+) DC. In the blood and in tonsils, IL-36R mRNA was predominantly found in myeloid cells. By contrast, IL-1R1 mRNA was detected in almost all cell types with higher levels in tonsil and skin compared to peripheral blood immune cells. IL-36β was as potent as IL-1β in stimulating M2 macrophages, keratinocytes and LC, less potent than IL-1β in stimulating M0 macrophages and MDDC, and exerted no effects in M1 and dermal macrophages. Levels of IL-1Ra diminished the ability of M2 macrophages to respond to IL-1. Taken together, these data are consistent with the association of excessive IL-36 signaling with an inflammatory skin phenotype and identify human LC and M2 macrophages as new IL-36 target cells.}, keywords = {agonists, ANTAGONIST, BLOOD, Cells, Cellular, Chemistry, Cultured, cytokine, CYTOKINE PRODUCTION, Cytokines, Dendritic Cells, DERMATOLOGY, Expression, Human, Humans, IL-1, IL-1R1, IL-1ra, IL-36, IL-36R, Immunoassay, Immunology, immunopathology, inflammation, Interleukin, Interleukin-1 Receptor Accessory Protein, Interleukin-1 Type I, KERATINOCYTES, Langerhans Cells, Macrophage, Macrophages, messenger, Molecular Biology, Monocytes, mRNA, Myeloid Cells, pathology, Phenotype, PRODUCTION, PROINFLAMMATORY CYTOKINES, Receptor, receptor antagonist, Receptors, RNA, signaling, Skin, target, Team-Mueller, TONSIL}, pubstate = {published}, tppubtype = {article} } @article{stoetzel_mutation_2016, title = {A mutation in VPS15 (PIK3R4) causes a ciliopathy and affects IFT20 release from the cis-Golgi.}, author = {Corinne Stoetzel and Séverine Bär and Johan-Owen De Craene and Sophie Scheidecker and Christelle Etard and Johana Chicher and Jennifer R Reck and Isabelle Perrault and Véronique Geoffroy and Kirsley Chennen and Uwe Strähle and Philippe Hammann and Sylvie Friant and Hélène Dollfus}, doi = {10.1038/ncomms13586}, issn = {2041-1723 2041-1723}, year = {2016}, date = {2016-01-01}, journal = {Nature communications}, volume = {7}, pages = {13586}, abstract = {Ciliopathies are a group of diseases that affect kidney and retina among other organs. Here, we identify a missense mutation in PIK3R4 (phosphoinositide 3-kinase regulatory subunit 4, named VPS15) in a family with a ciliopathy phenotype. Besides being required for trafficking and autophagy, we show that VPS15 regulates primary cilium length in human fibroblasts, as well as ciliary processes in zebrafish. Furthermore, we demonstrate its interaction with the golgin GM130 and its localization to the Golgi. The VPS15-R998Q patient mutation impairs Golgi trafficking functions in humanized yeast cells. Moreover, in VPS15-R998Q patient fibroblasts, the intraflagellar transport protein IFT20 is not localized to vesicles trafficking to the cilium but is restricted to the Golgi. Our findings suggest that at the Golgi, VPS15 and GM130 form a protein complex devoid of VPS34 to ensure the IFT20-dependent sorting and transport of membrane proteins from the cis-Golgi to the primary cilium.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{dobrenel_arabidopsis_2016, title = {The Arabidopsis TOR Kinase Specifically Regulates the Expression of Nuclear Genes Coding for Plastidic Ribosomal Proteins and the Phosphorylation of the Cytosolic Ribosomal Protein S6.}, author = {Thomas Dobrenel and Eder Mancera-Martínez and Céline Forzani and Marianne Azzopardi and Marlène Davanture and Manon Moreau and Mikhail Schepetilnikov and Johana Chicher and Olivier Langella and Michel Zivy and Christophe Robaglia and Lyubov A Ryabova and Johannes Hanson and Christian Meyer}, doi = {10.3389/fpls.2016.01611}, issn = {1664-462X 1664-462X 1664-462X}, year = {2016}, date = {2016-01-01}, journal = {Frontiers in plant science}, volume = {7}, pages = {1611}, abstract = {Protein translation is an energy consuming process that has to be fine-tuned at both the cell and organism levels to match the availability of resources. The target of rapamycin kinase (TOR) is a key regulator of a large range of biological processes in response to environmental cues. In this study, we have investigated the effects of TOR inactivation on the expression and regulation of Arabidopsis ribosomal proteins at different levels of analysis, namely from transcriptomic to phosphoproteomic. TOR inactivation resulted in a coordinated down-regulation of the transcription and translation of nuclear-encoded mRNAs coding for plastidic ribosomal proteins, which could explain the chlorotic phenotype of the TOR silenced plants. We have identified in the 5' untranslated regions (UTRs) of this set of genes a conserved sequence related to the 5' terminal oligopyrimidine motif, which is known to confer translational regulation by the TOR kinase in other eukaryotes. Furthermore, the phosphoproteomic analysis of the ribosomal fraction following TOR inactivation revealed a lower phosphorylation of the conserved Ser240 residue in the C-terminal region of the 40S ribosomal protein S6 (RPS6). These results were confirmed by Western blot analysis using an antibody that specifically recognizes phosphorylated Ser240 in RPS6. Finally, this antibody was used to follow TOR activity in plants. Our results thus uncover a multi-level regulation of plant ribosomal genes and proteins by the TOR kinase.}, keywords = {Phosphorylation, plastid, PPSE, proteomic, Ribosome, RPS6, TOR kinase, transcriptomic, translatomic}, pubstate = {published}, tppubtype = {article} } @article{harashima_modulation_2016, title = {Modulation of plant growth in vivo and identification of kinase substrates using an analog-sensitive variant of CYCLIN-DEPENDENT KINASE A;1.}, author = {Hirofumi Harashima and Nico Dissmeyer and Philippe Hammann and Yuko Nomura and Katharina Kramer and Hirofumi Nakagami and Arp Schnittger}, doi = {10.1186/s12870-016-0900-7}, issn = {1471-2229 1471-2229}, year = {2016}, date = {2016-01-01}, journal = {BMC plant biology}, volume = {16}, number = {1}, pages = {209}, abstract = {BACKGROUND: Modulation of protein activity by phosphorylation through kinases and subsequent de-phosphorylation by phosphatases is one of the most prominent cellular control mechanisms. Thus, identification of kinase substrates is pivotal for the understanding of many - if not all - molecular biological processes. Equally, the possibility to deliberately tune kinase activity is of great value to analyze the biological process controlled by a particular kinase. RESULTS: Here we have applied a chemical genetic approach and generated an analog-sensitive version of CDKA;1, the central cell-cycle regulator in Arabidopsis and homolog of the yeast Cdc2/CDC28 kinases. This variant could largely rescue a cdka;1 mutant and is biochemically active, albeit less than the wild type. Applying bulky kinase inhibitors allowed the reduction of kinase activity in an organismic context in vivo and the modulation of plant growth. To isolate CDK substrates, we have adopted a two-dimensional differential gel electrophoresis strategy, and searched for proteins that showed mobility changes in fluorescently labeled extracts from plants expressing the analog-sensitive version of CDKA;1 with and without adding a bulky ATP variant. A pilot set of five proteins involved in a range of different processes could be confirmed in independent kinase assays to be phosphorylated by CDKA;1 approving the applicability of the here-developed method to identify substrates. CONCLUSION: The here presented generation of an analog-sensitive CDKA;1 version is functional and represent a novel tool to modulate kinase activity in vivo and identify kinase substrates. Our here performed pilot screen led to the identification of CDK targets that link cell proliferation control to sugar metabolism, proline proteolysis, and glucosinolate production providing a hint how cell proliferation and growth are integrated with plant development and physiology.}, keywords = {Arabidopsis, Cell cycle, Kinase, Mitosis, Phosphorylation, PPSE, Substrate}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Perspectives and Pitfalls in Nucleic Acids Crystallography}, author = {E Westhof}, editor = {E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26227033?dopt=Abstract}, doi = {10.1007/978-1-4939-2763-0_1}, isbn = {26227033}, year = {2016}, date = {2016-01-01}, booktitle = {Nucleic Acid Crystallography: Methods and Protocols}, volume = {1320}, pages = {3-8}, publisher = {Humana Press}, address = {NY}, series = {Methods in Molecular Biology}, abstract = {X-ray crystallography offers precious and striking knowledge on biomolecular architectures. Although safeguards do exist to guarantee the accuracy of the structures deposited in databases, they are not always applied, leading to the spread of inaccurate data. The importance of validation reports in the publication process is emphasized.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {A Human Disease-causing Point Mutation in Mitochondrial Threonyl-tRNA Synthetase Induces Both Structural and Functional Defects.}, author = {Y Wang and X L Zhou and Z R Ruan and R J Liu and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26811336?dopt=Abstract}, doi = {10.1074/jbc.M115.700849}, isbn = {26811336}, year = {2016}, date = {2016-01-01}, journal = {J Biol Chem}, volume = {291}, number = {12}, pages = {6507-6520}, abstract = {Mitochondria require all translational components, including aminoacyl-tRNA synthetases (aaRSs), to complete organelle protein synthesis. Some aaRS mutations cause mitochondrial disorders, including human mitochondrial threonyl-tRNA synthetase (hmtThrRS) (encoded by TARS2), the P282L mutation of which causes mitochondrial encephalomyopathies. However, its catalytic and structural consequences remain unclear. Herein, we cloned TARS2 and purified the wild-type and P282L mutant hmtThrRS. hmtThrRS misactivates non-cognate Ser and uses post-transfer editing to clear erroneously synthesized products. In vitro and in vivo analyses revealed that the mutation induces a decrease in Thr activation, aminoacylation, and proofreading activities and a change in the protein structure and/or stability, which might cause reduced catalytic efficiency. We also identified a splicing variant of TARS2 mRNA lacking exons 8 and 9, the protein product of which is targeted into mitochondria. In HEK293T cells, the variant does not dimerize and cannot complement the ThrRS knock-out strain in yeast, suggesting that the truncated protein is inactive and might have a non-canonical function, as observed for other aaRS fragments. The present study describes the aminoacylation and editing properties of hmtThrRS, clarifies the molecular consequences of the P282L mutation, and shows that the yeast ThrRS-deletion model is suitable to test pathology-associated point mutations or alternative splicing variants of mammalian aaRS mRNAs.}, keywords = {alternative splicing aminoacyl-tRNA synthetase enzyme kinetics mitochondria mitochondrial disease threonyl-tRNA synthetase, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{cordeiro_integrin-alpha_2016, title = {Integrin-Alpha IIb Identifies Murine Lymph Node Lymphatic Endothelial Cells Responsive to RANKL}, author = {Olga G Cordeiro and Mélanie Chypre and Nathalie Brouard and Simon Rauber and Farouk Alloush and Monica Romera-Hernandez and Cécile Bénézech and Zhi Li and Anita Eckly and Mark C Coles and Antal Rot and Hideo Yagita and Catherine Léon and Burkhard Ludewig and Tom Cupedo and François Lanza and Christopher G Mueller}, doi = {10.1371/journal.pone.0151848}, issn = {1932-6203}, year = {2016}, date = {2016-01-01}, journal = {PloS One}, volume = {11}, number = {3}, pages = {e0151848}, abstract = {Microenvironment and activation signals likely imprint heterogeneity in the lymphatic endothelial cell (LEC) population. Particularly LECs of secondary lymphoid organs are exposed to different cell types and immune stimuli. However, our understanding of the nature of LEC activation signals and their cell source within the secondary lymphoid organ in the steady state remains incomplete. Here we show that integrin alpha 2b (ITGA2b), known to be carried by platelets, megakaryocytes and hematopoietic progenitors, is expressed by a lymph node subset of LECs, residing in medullary, cortical and subcapsular sinuses. In the subcapsular sinus, the floor but not the ceiling layer expresses the integrin, being excluded from ACKR4+ LECs but overlapping with MAdCAM-1 expression. ITGA2b expression increases in response to immunization, raising the possibility that heterogeneous ITGA2b levels reflect variation in exposure to activation signals. We show that alterations of the level of receptor activator of NF-κB ligand (RANKL), by overexpression, neutralization or deletion from stromal marginal reticular cells, affected the proportion of ITGA2b+ LECs. Lymph node LECs but not peripheral LECs express RANK. In addition, we found that lymphotoxin-β receptor signaling likewise regulated the proportion of ITGA2b+ LECs. These findings demonstrate that stromal reticular cells activate LECs via RANKL and support the action of hematopoietic cell-derived lymphotoxin.}, keywords = {Activation, Animals, Cells, Cultured, Endothelial Cells, ENDOTHELIAL-CELLS, Expression, Fibronectins, Immunization, Immunology, immunopathology, Inbred C57BL, infection, ligand, LYMPH, LYMPH NODE, Lymph Nodes, lymphoid organs, Lymphotoxin, Lymphotoxin-beta, Mice, murine, NF-kappaB, Platelet Membrane Glycoprotein IIb, PLATELETS, PROGENITORS, rank, RANK ligand, Receptor, Secondary, Signal Transduction, signaling, SINUS, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{, title = {A deafness and diabetes associated tRNA mutation caused the deficient pseudouridinylation at position 55 in tRNAGlu and mitochondrial dysfunction.}, author = {M Wang and H Liu and J Zheng and B Chen and M Zhou and W Fan and H Wang and X Liang and X Zhou and G Eriani and P Jiang and M X Guan}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27519417?dopt=Abstract}, doi = {10.1074/jbc.M116.739482}, isbn = {27519417}, year = {2016}, date = {2016-01-01}, journal = {J Biol Chem}, volume = {291}, number = {40}, pages = {21029-21041}, abstract = {Several mitochondrial tRNA mutations have been associated with maternally inherited diabetes and deafness (MIDD). However, the pathophysiology of these tRNA mutations remains poorly understood. In this report, we identified the novel homoplasmic 14692A>G mutation in the mitochondrial tRNAGlu gene among three Han Chinese families with maternally inherited diabetes and deafness. The m.14692A>G mutation affected a highly conserved uridine at position 55 of TΨC loop of tRNAGlu. The uridine is modified to pseudouridine (Ψ55), which plays an important role in the structure and function of this tRNA. Using lymphoblastoid cell lines derived from a Chinese family, we demonstrated that the m.14692A>G mutation caused the loss of Ψ55 modification and increased the angiogenin-mediated endonucleolytic cleavage in mutant tRNAGlu. The destabilization of base-pairing (18A-Ψ55) caused by the m.14692A>G mutation perturbed the conformation and stability of tRNAGlu. Approximately 65% decrease in the steady-state level of tRNAGlu was observed in mutant cells, compared to control cells. A failure in tRNAGlu metabolism impaired mitochondrial translation, especially for polypeptides with high proportion of glutamic acid codons such as MT-ND1, MT-ND6 and MT-CO2 in mutant cells. An impairment of mitochondrial translation caused the defective respiratory capacity, especially reducing activities of complexes I and IV. Furthermore, marked decreases in the levels of mitochondrial ATP and membrane potential were observed in mutant cells. These mitochondrial dysfunctions caused an increasing production of reactive oxygen species in the mutant cells. Our findings may provide new insights into pathophysiology of MIDD, which was primarily manifested by the deficient nucleotide modification of mitochondrial tRNAGlu.}, keywords = {diabetes hearing mitochondrial DNA (mtDNA) mitochondrial disease mutant pathogenesis post-translational modification (PTM) transfer RNA (tRNA), ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {COMPaRS: a stand-alone program for map comparison using quantile rank scaling}, author = {L Urzhumtseva and A Urzhumtsev}, url = {journals.iucr.org/j/issues/2016/06/00/jo5026/index.html}, doi = {S1600576716015752}, year = {2016}, date = {2016-01-01}, journal = {J Appl Cryst}, volume = {49}, number = {6}, pages = {2270-2275}, abstract = {The usual metrics for comparison of two crystallographic or cryoEM maps, for example the overall map correlation coefficient, measure the similarity of two sets of values with no consideration of their position in space. In contrast, when analyzing the maps visually it is the positions of sets of points with map values equal to or greater than some cutoff level that is of interest. An intrinsic and scale-invariant characteristic of such a set is the quantile rank defining the fraction of grid nodes (or of the unit-cell volume) with values less than this cutoff level. Comparison of the quantile ranks associated with the same point in the two maps is very similar to a comparison of the isosurfaces. The program COMPaRS uses new metrics for map comparison based on this idea: this gives quantitative results that agree with the qualitative results obtained from a visual analysis.}, keywords = {map comparison map correlation coefficient quantile rank scaling discrepancy function peak comparison computer programs., Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Helical Symmetry of Nucleic Acids: Obstacle or Help in Structure Solution?}, author = {A Urzhumtsev and L Urzhumtseva and U Baumann}, editor = {E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26227048?dopt=Abstract}, doi = {10.1007/978-1-4939-2763-0_16}, isbn = {26227048}, year = {2016}, date = {2016-01-01}, urldate = {2016-01-01}, booktitle = {Nucleic Acid Crystallography: Methods and Protocols}, volume = {1320}, pages = {259-267}, publisher = {Humana Press}, address = {NY}, series = {Methods in Molecular Biology}, abstract = {Crystallographic molecular replacement method is the key tool to define an atomic structure of nucleic acids. Frequently nucleic acids are packed forming continuous helices in the crystal. This arrangement of individual molecules in "infinite" pseudo helical structures in crystal may be the reason why the molecular replacement fails to find a unique position of the search atomic model as the method requires. The Patterson function, calculated as a Fourier series with diffraction intensities, has auxiliary peaks for such a molecular packing. Those near the origin peak indicate the orientation of the helices. The coordinates of other peaks are related to the molecular position and the rotation angle between two such "infinite" helices. Thus, the peak analysis allows getting molecular position even without a search model. An intelligent selecting and averaging of the phase sets corresponding to multiple probable positions of the search model again result in a unique solution but in the form of a Fourier synthesis and not a model. This synthesis can be used then to build an atomic model as it is the case for usual phasing methods.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Two novel members of the LhrC family of small RNAs in Listeria monocytogenes with overlapping regulatory functions but distinctive expression profiles.}, author = {M Storm Mollerup and J A Ross and A C Helfer and K Meistrup and P Romby and B H Kallipolitis}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27400116?dopt=Abstract}, doi = {10.1080/15476286.2016.1208332}, isbn = {27400116}, year = {2016}, date = {2016-01-01}, journal = {RNA Biol}, volume = {13}, number = {9}, pages = {895-915}, abstract = {Multicopy small RNAs (sRNAs) have gained recognition as an important feature of bacterial gene regulation. In the human pathogen Listeria monocytogenes, five homologous sRNAs, called LhrC1-5, control gene expression by base pairing to target mRNAs though three conserved UCCC motifs common to all five LhrCs. We show here that the sRNAs Rli22 and Rli33-1 are structurally and functionally related to LhrC1-5, expanding the LhrC family to seven members, which makes it the largest multicopy sRNA family reported so far. Rli22 and Rli33-1 both contain two UCCC motifs important for post-transcriptional repression of three LhrC target genes. One such target, oppA, encodes a virulence-associated oligo-peptide binding protein. Like LhrC1-5, Rli22 and Rli33-1 employ their UCCC motifs to recognize the Shine-Dalgarno region of oppA mRNA and prevent formation of the ribosomal complex, demonstrating that the seven sRNAs act in a functionally redundant manner. However, differential expression profiles of the sRNAs under infection-relevant conditions suggest that they might also possess non-overlapping functions. Collectively, this makes the LhrC family a unique case for studying the purpose of sRNA multiplicity in the context of bacterial virulence.}, keywords = {Listeria monocytogenes antisense multiplicity post-transcriptional regulation sRNAs, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interplay between Catalysts and Substrates for Activity of Class Ib Aminoacyl-tRNA Synthetases and Implications for Pharmacology.}, author = {P Stephen and S X Lin and R Giegè}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26286212}, doi = {10.2174/1568026615666150819110018}, isbn = {26286212}, year = {2016}, date = {2016-01-01}, journal = {Curr Top Med Chem}, volume = {16}, number = {6}, pages = {616-633}, abstract = {Aminoacyl-tRNA synthetase:transfer RNA (aaRS:tRNA) systems became recently essential targets in molecular medicine, because perturbed recognition of cognate tRNAs by aaRSs and poor precision in tRNA aminoacylation do not guarantee accurate protein biosynthesis, thus leading to diseases. Sets of identity determinants situated at particular zones of tRNA are responsible for functional accuracy. Recent work in X-ray crystallography has revealed various snapshots of aaRS:ligand complexes which represent the stages required for aminoacylation. Here we focus on a small group of class I aaRSs conserved in evolution, the ArgRSs, GluRSs, GlnRSs, and atypical LysRSs found mostly in Archaea and in a few Bacteria, that catalyze amino acid activation only in the presence of their cognate tRNAs. Structural and functional features of these aaRSs, ranked in subclass Ib, together with their peculiar mode of tRNA recognition and identity expression are reviewed and compared. Strategies to inhibit class Ib aaRS:tRNA aminoacylation systems, their dysfunction leading to human diseases, and the implications for pharmacology are outlined.}, keywords = {GIEGE Aminoacylation tRNA Aminoacyl-tRNA synthetase and diseases Aminoacyl-tRNA synthetase as drug target Protein-biomolecule interactions tRNA identity determinants, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A step forward understanding HIV-1 diversity}, author = {R P Smyth and M Negroni}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27093884?dopt=Abstract}, doi = {10.1186/s12977-016-0259-8}, isbn = {27093884}, year = {2016}, date = {2016-01-01}, journal = {Retrovirology}, volume = {13}, number = {1}, pages = {27}, abstract = {Human immunodeficiency virus (HIV) populations are characterized by extensive genetic diversity. Antigenic diversification is essential for escape from immune selection and therapy, and remains one of the major obstacles for the development of an efficient vaccine strategy. Even if intensive efforts have been made for understanding the molecular mechanisms responsible for genetic diversity in HIV, conclusive data in vivo is still lacking. Recent works have addressed this issue, focusing on the identification of the sources of genetic diversity during in vivo infections and on the estimate of the pervasiveness of genetic recombination during replication in vivo. Surprisingly, it appears that despite the error-prone nature of the viral polymerase, the bulk of mutations found in patients are indeed due to the effect of a cellular restriction factor. This factor tends to hypermutate the viral genome abolishing viral infectivity. When hypermutation is incomplete, the virus retains infectivity and converts the effect of the cellular factor to its advantage by exploiting it to generate genetic diversity that is beneficial for viral propagation. This view contrasts the long-standing dogma that viral diversity is due to the intrinsic error-prone nature of the viral replication cycle. Besides hypermutations and mutations, recombination is also a pervasive source of genetic diversity. The estimate of the frequency at which this process takes place in vivo has remained elusive, despite extensive efforts in this sense. Now, using single genome amplification, and starting from publically available datasets, it has been obtained a confirmation of the estimates previously made using tissue culture studies. These recent findings are presented here and their implications for the development of future researches are discussed.}, keywords = {alternative splicing aminoacyl-tRNA synthetase enzyme kinetics mitochondria mitochondrial disease threonyl-tRNA synthetase, MARQUET, NEGRONI, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {MIMEAnTo - Profiling functional RNA in Mutational Interference Mapping Experiments.}, author = {M R Smith and R P Smyth and R Marquet and M von Kleist}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27402903}, doi = {10.1093/bioinformatics/btw479}, isbn = {27402903}, year = {2016}, date = {2016-01-01}, journal = {Bioinformatics}, volume = {32}, number = {21}, pages = {3369-3370}, abstract = {The mutational interference mapping experiment (MIME) is a powerful method that, coupled to a bioinformatics analysis pipeline, allows the identification of domains and structures in RNA that are important for its function. In MIME, target RNAs are randomly mutated, selected by function, physically separated and sequenced using next-generation sequencing (NGS). Quantitative effects of each mutation at each position in the RNA can be recovered with statistical certainty using the herein developed user-friendly, cross-platform software MIMEAnTo (MIME Analysis Tool). AVAILABILITY AND IMPLEMENTATION: MIMEAnTo is implemented in C++ using the boost library as well as Qt for the graphical user interface and is distributed under GPL (http://www.gnu.org/licences/gpl). The libraries are statically linked in a stand alone executable and are not required on the system. The plots are generated with gnuplot. Gnuplot-iostream (https://github.com/dstahlke/gnuplot-iostream) serves as gnuplot interface. Standalone executables including examples and source code can be downloaded from https://github.com/maureensmith/MIMEAnTo SUPPLEMENTARY INFORMATION: Supplementary data is available at Bioinformatics online.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {What is so special about neuronal translation? (comment on DOI 10.1002/bies.201600052).}, author = {M Sissler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27427507?dopt=Abstract}, doi = {10.1002/bies.201600131}, isbn = {27427507}, year = {2016}, date = {2016-01-01}, journal = {Bioessays}, volume = {38}, number = {9}, pages = {816}, keywords = {SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {eIF3 Peripheral Subunits Rearrangement after mRNA Binding and Start-Codon Recognition.}, author = {A Simonetti and J Brito Querido and A Myasnikov and E Mancera-Martinez and A Renaud and L Kuhn and Y Hashem}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27373335}, doi = {10.1016/j.molcel.2016.05.033.}, isbn = {27373335}, year = {2016}, date = {2016-01-01}, journal = {Mol Cell}, volume = {63}, number = {2}, pages = {206-217}, abstract = {mRNA translation initiation in eukaryotes requires the cooperation of a dozen eukaryotic initiation factors (eIFs) forming several complexes, which leads to mRNA attachment to the small ribosomal 40S subunit, mRNA scanning for start codon, and accommodation of initiator tRNA at the 40S P site. eIF3, composed of 13 subunits, 8 core (a, c, e, f, h, l, k, and m) and 5 peripheral (b, d, g, i, and j), plays a central role during this process. Here we report a cryo-electron microscopy structure of a mammalian 48S initiation complex at 5.8 Å resolution. It shows the relocation of subunits eIF3i and eIF3g to the 40S intersubunit face on the GTPase binding site, at a late stage in initiation. On the basis of a previous study, we demonstrate the relocation of eIF3b to the 40S intersubunit face, binding below the eIF2-Met-tRNAiMet ternary complex upon mRNA attachment. Our analysis reveals the deep rearrangement of eIF3 and unravels the molecular mechanism underlying eIF3 function in mRNA scanning and timing of ribosomal subunit joining.}, keywords = {PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @incollection{, title = {Synthetic Capped mRNAs for Cap-Specific Photo-Cross-Linking Experiments.}, author = { J. Kowalska and F. Martin and J. Jemielity}, editor = { R. Rhoads}, year = {2016}, date = {2016-01-01}, booktitle = {Synthetic mRNA: Production, Introduction Into Cells, and Physiological Consequences}, volume = {1428}, pages = {31-43}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {he 7-methylguanosine triphosphate cap present at the 5' ends of eukaryotic mRNAs plays numerous roles in mRNA expression and metabolism. The identification and studies on cap-binding partners can be significantly advanced using tailored chemical tools such as synthetic cap analogues or RNAs carrying modified cap structures. Here we provide protocols for the production of mRNAs specifically labeled within the 5' cap with a nucleoside capable of being photo-activated, either 6-thioguanosine or 7-methyl-6-thioguanosine, which can be used in photo-cross-linking experiments to identify or characterize cap-binding biomolecules. We also describe a protocol for the cross-linking experiments with capped RNAs to map histone H4 cap-binding pocket.}, keywords = {5′, 6-thioguanosine, analogs, Binding, Cap, ERIANI, Histone, mRNA, Photo-cross-linking, stability, Transcription}, pubstate = {published}, tppubtype = {incollection} } @article{, title = {Mechanistic and Structural Studies of Protein-Only RNase P Compared to Ribonucleoproteins Reveal the Two Faces of the Same Enzymatic Activity.}, author = {C Schelcher and C Sauter and P Giege}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27348014?dopt=Abstract}, doi = {10.3390/biom6030030}, isbn = {27348014}, year = {2016}, date = {2016-01-01}, journal = {Biomolecules}, volume = {6}, number = {3}, pages = {30}, abstract = {RNase P, the essential activity that performs the 5' maturation of tRNA precursors, can be achieved either by ribonucleoproteins containing a ribozyme present in the three domains of life or by protein-only enzymes called protein-only RNase P (PRORP) that occur in eukaryote nuclei and organelles. A fast growing list of studies has investigated three-dimensional structures and mode of action of PRORP proteins. Results suggest that similar to ribozymes, PRORP proteins have two main domains. A clear functional analogy can be drawn between the specificity domain of the RNase P ribozyme and PRORP pentatricopeptide repeat domain, and between the ribozyme catalytic domain and PRORP N4BP1, YacP-like Nuclease domain. Moreover, both types of enzymes appear to dock with the acceptor arm of tRNA precursors and make specific contacts with the corner of pre-tRNAs. While some clear differences can still be delineated between PRORP and ribonucleoprotein (RNP) RNase P, the two types of enzymes seem to use, fundamentally, the same catalytic mechanism involving two metal ions. The occurrence of PRORP and RNP RNase P represents a remarkable example of convergent evolution. It might be the unique witness of an ongoing replacement of catalytic RNAs by proteins for enzymatic activities.}, keywords = {PRORP RNase P crystal structures kinetic analyses tRNA biogenesis, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The ribosome prohibits the G•U wobble geometry at the first position of the codon-anticodon helix.}, author = {A Rozov and E Westhof and M Yusupov and G Yusupova}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27174928?dopt=Abstract}, doi = {10.1093/nar/gkw431}, isbn = {27174928}, year = {2016}, date = {2016-01-01}, journal = {Nucleic Acids Res}, volume = {44}, number = {13}, pages = {6434-6441}, abstract = {Precise conversion of genetic information into proteins is essential to cellular health. However, a margin of error exists and is at its highest on the stage of translation of mRNA by the ribosome. Here we present three crystal structures of 70S ribosome complexes with messenger RNA and transfer RNAs and show that when a G•U base pair is at the first position of the codon-anticodon helix a conventional wobble pair cannot form because of inescapable steric clash between the guanosine of the A codon and the key nucleotide of decoding center adenosine 1493 of 16S rRNA. In our structure the rigid ribosomal decoding center, which is identically shaped for cognate or near-cognate tRNAs, forces this pair to adopt a geometry close to that of a canonical G•C pair. We further strengthen our hypothesis that spatial mimicry due either to base tautomerism or ionization dominates the translation infidelity mechanism.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {New Structural Insights into Translational Miscoding.}, author = {A Rozov and N Demeshkina and E Westhof and M Yusupov and G Yusupova}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27372401}, doi = {10.1016/j.tibs.2016.06.001}, isbn = {27372401}, year = {2016}, date = {2016-01-01}, journal = {Trends Biochem Sci}, volume = {41}, number = {9}, pages = {798-814}, abstract = {The fidelity of translation depends strongly on the selection of the correct aminoacyl-tRNA that is complementary to the mRNA codon present in the ribosomal decoding center. The ribosome occasionally makes mistakes by selecting the wrong substrate from the pool of aminoacyl-tRNAs. Here, we summarize recent structural advances that may help to clarify the origin of missense errors that occur during decoding. These developments suggest that discrimination between tRNAs is based primarily on steric complementarity and shape acceptance rather than on the number of hydrogen bonds between the molding of the decoding center and the codon-anticodon duplex. They strengthen the hypothesis that spatial mimicry, due either to base tautomerism or ionization, drives infidelity in ribosomal translation.}, keywords = {crystallography decoding ribosome structure translation, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Novel base-pairing interactions at the tRNA wobble position crucial for accurate reading of the genetic code.}, author = {A Rozov and N Demeshkina and I Khusainov and E Westhof and M Yusupov and G Yusupova}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26791911?dopt=Abstract}, doi = {10.1038/ncomms10457}, isbn = {26791911}, year = {2016}, date = {2016-01-01}, journal = {Nat Commun}, volume = {21}, number = {7}, pages = {10457}, abstract = {Posttranscriptional modifications at the wobble position of transfer RNAs play a substantial role in deciphering the degenerate genetic code on the ribosome. The number and variety of modifications suggest different mechanisms of action during messenger RNA decoding, of which only a few were described so far. Here, on the basis of several 70S ribosome complex X-ray structures, we demonstrate how Escherichia coli tRNA(Lys)UUU with hypermodified 5-methylaminomethyl-2-thiouridine (mnm(5)s(2)U) at the wobble position discriminates between cognate codons AAA and AAG, and near-cognate stop codon UAA or isoleucine codon AUA, with which it forms pyrimidine-pyrimidine mismatches. We show that mnm(5)s(2)U forms an unusual pair with guanosine at the wobble position that expands general knowledge on the degeneracy of the genetic code and specifies a powerful role of tRNA modifications in translation. Our models consolidate the translational fidelity mechanism proposed previously where the steric complementarity and shape acceptance dominate the decoding mechanism.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {LARS2 Variants Associated with Hydrops, Lactic Acidosis, Sideroblastic Anemia, and Multisystem Failure.}, author = {L G Riley and J Rudinger-Thirion and K Schmitz-Abe and D R Thorburn and R L Davis and J Teo and S Arbuckle and S T Cooper and D R Campagna and M Frugier and K Markianos and C M Sue and M D Fleming and J Christodoulou}, editor = {E Morava}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26537577?dopt=Abstract}, doi = {10.1007/8904_2015_515}, isbn = {26537577}, year = {2016}, date = {2016-01-01}, booktitle = {JIMD Rep}, volume = {28}, pages = {49-57}, publisher = {Springer}, edition = {SSIEM}, abstract = {Pathogenic variants in mitochondrial aminoacyl-tRNA synthetases result in a broad range of mitochondrial respiratory chain disorders despite their shared role in mitochondrial protein synthesis. LARS2 encodes the mitochondrial leucyl-tRNA synthetase, which attaches leucine to its cognate tRNA. Sequence variants in LARS2 have previously been associated with Perrault syndrome, characterized by premature ovarian failure and hearing loss (OMIM #615300). In this study, we report variants in LARS2 that are associated with a severe multisystem metabolic disorder. The proband was born prematurely with severe lactic acidosis, hydrops, and sideroblastic anemia. She had multisystem complications with hyaline membrane disease, impaired cardiac function, a coagulopathy, pulmonary hypertension, and progressive renal disease and succumbed at 5 days of age. Whole exome sequencing of patient DNA revealed compound heterozygous variants in LARS2 (c.1289C>T; p.Ala430Val and c.1565C>A; p.Thr522Asn). The c.1565C>A (p.Thr522Asn) LARS2 variant has previously been associated with Perrault syndrome and both identified variants are predicted to be damaging (SIFT, PolyPhen). Muscle and liver samples from the proband did not display marked mitochondrial respiratory chain enzyme deficiency. Immunoblotting of patient muscle and liver showed LARS2 levels were reduced in liver and complex I protein levels were reduced in patient muscle and liver. Aminoacylation assays revealed p.Ala430Val LARS2 had an 18-fold loss of catalytic efficiency and p.Thr522Asn a 9-fold loss compared to wild-type LARS2. We suggest that the identified LARS2 variants are responsible for the severe multisystem clinical phenotype seen in this baby and that mutations in LARS2 can result in variable phenotypes.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Requirements for nucleocapsid-mediated regulation of reverse transcription during the late steps of HIV-1 assembly.}, author = {P J Racine and C Chamontin and H de Rocquigny and S Bernacchi and J C Paillart and M Mougel}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27273064?dopt=Abstract}, doi = {10.1038/srep27536}, isbn = {27273064}, year = {2016}, date = {2016-01-01}, journal = {Sci Rep}, volume = {6}, pages = {27536}, abstract = {HIV-1 is a retrovirus replicating within cells by reverse transcribing its genomic RNA (gRNA) into DNA. Within cells, virus assembly requires the structural Gag proteins with few accessory proteins, notably the viral infectivity factor (Vif) and two copies of gRNA as well as cellular factors to converge to the plasma membrane. In this process, the nucleocapsid (NC) domain of Gag binds to the packaging signal of gRNA which consists of a series of stem-loops (SL1-SL3) ensuring gRNA selection and packaging into virions. Interestingly, mutating NC activates a late-occurring reverse transcription (RT) step in producer cells, leading to the release of DNA-containing HIV-1 particles. In order to decipher the molecular mechanism regulating this late RT, we explored the role of several key partners of NC, such as Vif, gRNA and the cellular cytidine deaminase APOBEC3G that restricts HIV-1 infection by targeting the RT. By studying combinations of deletions of these putative players, we revealed that NC, SL1-SL3 and in lesser extent Vif, but not APOBEC3G, interplay regulates the late RT.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleolin promotes heat shock-associated translation of VEGF-D to promote tumor lymphangiogenesis.}, author = {F Morfoisse and F Tatin and F Hantelys and A Adoue and A C Helfer and S Cassant-Sourdy and F Pujol and A Gomez-Brouchet and L Ligat and F Lopez and S Pyronnet and J Courty and J Guillermet-Guibert and S Marzi and R J Schneider and A C Prats and B H Garmy-Susini}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27280395?dopt=Abstract}, doi = {10.1158/0008-5472.CAN-15-3140}, isbn = {27280395}, year = {2016}, date = {2016-01-01}, journal = {Cancer Res}, volume = {76}, number = {15}, pages = {4394-4405}, abstract = {The vascular endothelial growth factor VEGF-D promotes metastasis by inducing lymphangiogenesis and dilatation of the lymphatic vasculature, facilitating tumor cell extravasion. Here we report a novel level of control for VEGF-D expression at the level of protein translation. In human tumor cells, VEGF-D colocalized with eIF4GI and 4E-BP1, which can program increased initiation at IRES motifs on mRNA by the translational initiation complex. In murine tumors, the steady-state level of VEGF-D protein was increased despite the overexpression and dephosphorylation of 4E-BP1, which downregulates protein synthesis, suggesting the presence of an IRES in the 5' UTR of VEGF-D mRNA. We found that nucleolin, a nucleolar protein involved in ribosomal maturation, bound directly to the 5'UTR of VEGF-D mRNA, thereby improving its translation following heat shock stress via IRES activation. Nucleolin blockade by RNAi-mediated silencing or pharmacological inhibition reduced VEGF-D translation along with a subsequent constriction of lymphatic vessels in tumors. Our results identify nucleolin as a key regulator of VEGF-D expression, deepening understanding of lymphangiogenesis control during tumor formation.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RBscore&NBench: a high-level web server for nucleic acid binding residues prediction with a large-scale benchmarking database.}, author = {Z Miao and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27084939?dopt=Abstract}, doi = {10.1093/nar/gkw251}, isbn = {27084949}, year = {2016}, date = {2016-01-01}, journal = {Nucleic Acids Res}, volume = {44}, number = {W1}, pages = {W562-W567}, abstract = {RBscore&NBench combines a web server, RBscore and a database, NBench. RBscore predicts RNA-/DNA-binding residues in proteins and visualizes the prediction scores and features on protein structures. The scoring scheme of RBscore directly links feature values to nucleic acid binding probabilities and illustrates the nucleic acid binding energy funnel on the protein surface. To avoid dataset, binding site definition and assessment metric biases, we compared RBscore with 18 web servers and 3 stand-alone programs on 41 datasets, which demonstrated the high and stable accuracy of RBscore. A comprehensive comparison led us to develop a benchmark database named NBench. The web server is available on:http://ahsoka.u-strasbg.fr/rbscorenbench/.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Quantifying side-chain conformational variations in protein structure.}, author = {Z Miao and Y Cao}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27845406?dopt=Abstract}, doi = {10.1038/srep37024}, isbn = {27845406}, year = {2016}, date = {2016-01-01}, journal = {Sci Rep}, volume = {6}, pages = {37024}, abstract = {Protein side-chain conformation is closely related to their biological functions. The side-chain prediction is a key step in protein design, protein docking and structure optimization. However, side-chain polymorphism comprehensively exists in protein as various types and has been long overlooked by side-chain prediction. But such conformational variations have not been quantitatively studied and the correlations between these variations and residue features are vague. Here, we performed statistical analyses on large scale data sets and found that the side-chain conformational flexibility is closely related to the exposure to solvent, degree of freedom and hydrophilicity. These analyses allowed us to quantify different types of side-chain variabilities in PDB. The results underscore that protein side-chain conformation prediction is not a single-answer problem, leading us to reconsider the assessment approaches of side-chain prediction programs.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of the interaction between the HIV-1 Gag structural polyprotein and the cellular ribosomal protein L7 and its implication in viral nucleic acid remodeling.}, author = {H E Mekdad and E Boutant and H Karnib and M E Biedma and K K Sharma and I Malytska and G Laumond and M Roy and E Réal and J C Paillart and C Moog and J L Darlix and Y Mély and H de Rocquigny}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27515235}, doi = {10.1186/s12977-016-0287-4}, isbn = {27515235}, year = {2016}, date = {2016-01-01}, journal = {Retrovirology}, volume = {13}, number = {1}, pages = {54}, abstract = {Background In HIV-1 infected cells, the integrated viral DNA is transcribed by the host cell machinery to generate the full length HIV-1 RNA (FL RNA) that serves as mRNA encoding for the Gag and GagPol precursors. Virion formation is orchestrated by Gag, and the current view is that a specific interaction between newly made Gag molecules and FL RNA initiates the process. This in turn would cause FL RNA dimerization by the NC domain of Gag (GagNC). However the RNA chaperoning activity of unprocessed Gag is low as compared to the mature NC protein. This prompted us to search for GagNC co-factors. Results Here we report that RPL7, a major ribosomal protein involved in translation regulation, is a partner of Gag via its interaction with the NC domain. This interaction is mediated by the NC zinc fingers and the N- and C-termini of RPL7, respectively, but seems independent of RNA binding, Gag oligomerization and its interaction with the plasma membrane. Interestingly, RPL7 is shown for the first time to exhibit a potent DNA/RNA chaperone activity higher than that of Gag. In addition, Gag and RPL7 can function in concert to drive rapid nucleic acid hybridization. Conclusions Our results show that GagNC interacts with the ribosomal protein RPL7, favoring the notion that RPL7 could be a Gag helper chaperoning factor possibly contributing to the start of Gag assembly.}, keywords = {HIV Gag RPL7 Interaction Chaperone activity Nucleocapsid, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Enzymatic Paradox of Yeast Arginyl-tRNA Synthetase: Exclusive Arginine Transfer Controlled by a Flexible Mechanism of tRNA Recognition.}, author = {A McShane and E Hok and J Tomberlin and G Eriani and R Geslain}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26844776?dopt=Abstract}, doi = {10.1371/journal.pone.0148460}, isbn = {26844776}, year = {2016}, date = {2016-01-01}, journal = {PLoS One}, volume = {11}, number = {2}, pages = {e0148460}, abstract = {Identity determinants are essential for the accurate recognition of transfer RNAs by aminoacyl-tRNA synthetases. To date, arginine determinants in the yeast Saccharomyces cerevisiae have been identified exclusively in vitro and only on a limited number of tRNA Arginine isoacceptors. In the current study, we favor a full cellular approach and expand the investigation of arginine determinants to all four tRNA Arg isoacceptors. More precisely, this work scrutinizes the relevance of the tRNA nucleotides at position 20, 35 and 36 in the yeast arginylation reaction. We built 21 mutants by site-directed mutagenesis and tested their functionality in YAL5, a previously engineered yeast knockout deficient for the expression of tRNA Arg CCG. Arginylation levels were also monitored using Northern blot. Our data collected in vivo correlate with previous observations. C35 is the prominent arginine determinant followed by G36 or U36 (G/U36). In addition, although there is no major arginine determinant in the D loop, the recognition of tRNA Arg ICG relies to some extent on the nucleotide at position 20. This work refines the existing model for tRNA Arg recognition. Our observations indicate that yeast Arginyl-tRNA synthetase (yArgRS) relies on distinct mechanisms to aminoacylate the four isoacceptors. Finally, according to our refined model, yArgRS is able to accommodate tRNA Arg scaffolds presenting N34, C/G35 and G/A/U36 anticodons while maintaining specificity. We discuss the mechanistic and potential physiological implications of these findings.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transient compartmentalization of RNA replicators prevents extinction due to parasites.}, author = {S Matsumura and A Kun and M Ryckelynck and F Coldren and A Szilagyi and F Jossinet and C Rick and P Nghe and E Szathmary and A D Griffiths}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27940874?dopt=Abstract}, doi = {10.1126/science.aag1582}, isbn = {27940874}, year = {2016}, date = {2016-01-01}, journal = {Science}, volume = {354}, number = {6317}, pages = {1293-1296}, abstract = {The appearance of molecular replicators (molecules that can be copied) was probably a critical step in the origin of life. However, parasitic replicators would take over and would have prevented life from taking off unless the replicators were compartmentalized in reproducing protocells. Paradoxically, control of protocell reproduction would seem to require evolved replicators. We show here that a simpler population structure, based on cycles of transient compartmentalization (TC) and mixing of RNA replicators, is sufficient to prevent takeover by parasitic mutants. TC tends to select for ensembles of replicators that replicate at a similar rate, including a diversity of parasites that could serve as a source of opportunistic functionality. Thus, TC in natural, abiological compartments could have allowed life to take hold.}, keywords = {RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ribosomal 18S rRNA base pairs with mRNA during eukaryotic translation initiation.}, author = {F Martin and J F Ménétret and A Simonetti and A G Myasnikov and Q Vicens and L Prongidi-Fix and S K Natchiar and B P Klaholz and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27554013?dopt=Abstract}, doi = {10.1038/ncomms12622}, isbn = {27554013}, year = {2016}, date = {2016-01-01}, journal = {Nat Commun}, volume = {7}, pages = {12622}, abstract = {Eukaryotic mRNAs often contain a Kozak sequence that helps tether the ribosome to the AUG start codon. The mRNA of histone H4 (h4) does not undergo classical ribosome scanning but has evolved a specific tethering mechanism. The cryo-EM structure of the rabbit ribosome complex with mouse h4 shows that the mRNA forms a folded, repressive structure at the mRNA entry site on the 40S subunit next to the tip of helix 16 of 18S ribosomal RNA (rRNA). Toe-printing and mutational assays reveal that an interaction exists between a purine-rich sequence in h4 mRNA and a complementary UUUC sequence of helix h16. Together the present data establish that the h4 mRNA harbours a sequence complementary to an 18S rRNA sequence which tethers the mRNA to the ribosome to promote proper start codon positioning, complementing the interactions of the 40S subunit with the Kozak sequence that flanks the AUG start codon.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The life-cycle of the HIV-1 Gag-RNA complex.}, author = {E Mailler and S Bernacchi and R Marquet and J C Paillart and V Vivet-Boudou and R P Smyth}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27626439?dopt=Abstract}, doi = {10.3390/v8090248}, isbn = {27626439}, year = {2016}, date = {2016-01-01}, journal = {Viruses}, volume = {8}, number = {9}, pages = {248}, abstract = {Human immunodeficiency virus type 1 (HIV-1) replication is a highly regulated process requiring the recruitment of viral and cellular components to the plasma membrane for assembly into infectious particles. This review highlights the recent process of understanding the selection of the genomic RNA (gRNA) by the viral Pr55Gag precursor polyprotein, and the processes leading to its incorporation into viral particles.}, keywords = {HIV-1 packaging Gag genomic RNA assembly, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Various checkpoints prevent the synthesis of Staphylococcus aureus peptidoglycan hydrolase LytM in the stationary growth phase.}, author = {E Lioliou and P Fechter and I Caldelari and B C Jester and S Dubrac and A C Helfer and S Boisset and F Vandenesch and P Romby and T Geissmann}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26901414}, doi = {10.1080/15476286.2016.1153209}, isbn = {26901414}, year = {2016}, date = {2016-01-01}, journal = {RNA Biol}, volume = {13}, number = {4}, pages = {427-440}, abstract = {In Staphylococcus aureus, peptidoglycan metabolism plays a role in the host inflammatory response and pathogenesis. Transcription of the peptidoglycan hydrolases is activated by the essential two-component system WalKR at low cell density. During stationary growth phase, WalKR is not active and transcription of the peptidoglycan hydrolase genes is repressed. In this work, we studied regulation of expression of the glycylglycine endopeptidase LytM. We show that, in addition to the transcriptional regulation mediated by WalKR, the synthesis of LytM is negatively controlled by a unique mechanism at the stationary growth phase. We have identified two different mRNAs encoding lytM, which vary in the length of their 5' untranslated (5'UTR) regions. LytM is predominantly produced from the WalKR-regulated mRNA transcript carrying a short 5'UTR. The lytM mRNA is also transcribed as part of a polycistronic operon with the upstream SA0264 gene and is constitutively expressed. Although SA0264 protein can be synthesized from the longer operon transcript, lytM cannot be translated because its ribosome-binding site is sequestered into a translationally inactive secondary structure. In addition, the effector of the agr system, RNAIII, can inhibit translation of lytM present on the operon without altering the transcript level but does not have an effect on the translation of the upstream gene. We propose that this dual regulation of lytM expression, at the transcriptional and post-transcriptional levels, contributes to prevent cell wall damage during the stationary phase of growth.}, keywords = {LytM RNAIII Staphylococcus aureus autolysin cell wall peptidoglycan peptidoglycan hydrolase post-transcriptional regulation, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {What do we know about selenium contribution to muscle physiology?}, author = {A Lescure and M Briens and A Ferreiro}, editor = {D L Hatfield and U Schweizer and P A Tsuji and V Gladyshev}, url = {http://link.springer.com/chapter/10.1007/978-3-319-41283-2_40}, doi = {10.1007/978-3-319-41283-2_40}, year = {2016}, date = {2016-01-01}, booktitle = {Selenium: Its molecular biology and role in human health}, pages = {475-486}, publisher = {Springer}, edition = {4}, abstract = {Early observations of selenium (Se) deficiencies in human and livestock revealed the importance of this trace element for normal muscle function. However, the molecular and cellular dysfunction connecting low selenium diets to muscular diseases remain elusive. Importantly, mutations in the SEPN1 gene encoding selenoprotein N (SEPN1) were shown to cause an inherited muscular disease in humans. Therefore, it is expected that understanding the role of SEPN1 and the related pathophysiology will unveil the participation of Se in molecular processes essential for muscle function and pave the way for targeted therapeutics. However, the functional characterization of SEPN1 is still lacking. Analysis of its activity in cellular and animal models pointed to its involvement in oxidative stress defense and in control of calcium (Ca2+) handling. A link between the activities of SEPN1 and of the Ca2+ transporters, RyR1 and SERCA, was shown, but the enzymatic reaction catalyzed by SEPN1 has not been characterized.}, keywords = {Intracellular calcium handling Muscle Muscular dystrophies Oxidative stress Selenium Selenoprotein N, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Molecular basis and specificity of H2A.Z-H2B recognition and deposition by the histone chaperone YL1.}, author = {C M Latrick and M Marek and K Ouararhni and C Papin and I Stoll and M Ignatyeva and A Obri and E Ennifar and S Dimitrov and C Romier and A Hamiche}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26974126?dopt=Abstract}, doi = {10.1038/nsmb.3189}, isbn = {26974126}, year = {2016}, date = {2016-01-01}, journal = {Nat Struct Mol Biol}, volume = {23}, number = {4}, pages = {309-316}, abstract = {H2A.Z, a widely conserved histone variant, is evicted from chromatin by the histone chaperone ANP32E. However, to date, no deposition chaperone for H2A.Z is known in metazoans. Here, we identify YL1 as a specific H2A.Z-deposition chaperone. The 2.7-Å-resolution crystal structure of the human YL1-H2A.Z-H2B complex shows that YL1 binding, similarly to ANP32E binding, triggers an extension of the H2A.Z αC helix. The interaction with YL1 is, however, more extensive and includes both the extended acidic patch and the entire DNA-binding surface of H2A.Z-H2B. Substitution of only four amino acid residues of H2A is sufficient for the formation of an H2A.Z-like interface specifically recognized by YL1. Collectively, our data reveal the molecular basis of H2A.Z-specific recognition by YL1 and shed light on the mechanism of H2A.Z transfer to the nucleosome by the ATP-dependent chromatin-remodeling complexes SRCAP and P400-TIP60.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Synthetic Capped mRNAs for Cap-Specific Photo-Cross-Linking Experiments.}, author = {J Kowalska and F Martin and J Jemielity}, editor = {R Rhoads}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27236790?dopt=Abstract}, doi = {10.1007/978-1-4939-3625-0_2}, isbn = {27236790}, year = {2016}, date = {2016-01-01}, booktitle = {Synthetic mRNA: Production, Introduction Into Cells, and Physiological Consequences}, volume = {1428}, pages = {31-43}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {he 7-methylguanosine triphosphate cap present at the 5' ends of eukaryotic mRNAs plays numerous roles in mRNA expression and metabolism. The identification and studies on cap-binding partners can be significantly advanced using tailored chemical tools such as synthetic cap analogues or RNAs carrying modified cap structures. Here we provide protocols for the production of mRNAs specifically labeled within the 5' cap with a nucleoside capable of being photo-activated, either 6-thioguanosine or 7-methyl-6-thioguanosine, which can be used in photo-cross-linking experiments to identify or characterize cap-binding biomolecules. We also describe a protocol for the cross-linking experiments with capped RNAs to map histone H4 cap-binding pocket.}, keywords = {cap analogs 6-thioguanosine Cap binding Histone mRNA Photo-cross-linking Transcription mRNA stability, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {[A glimpse on Staphylococcus aureus translation machinery and its control].}, author = {I Khusainov and A Marenna and M Cerciat and P Fechter and Y Hashem and S Marzi and P Romby and G Yusupova and M Yusupov}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27668596?dopt=Abstract}, doi = {10.7868/S0026898416040042}, isbn = {27668596}, year = {2016}, date = {2016-01-01}, journal = {Mol Biol (Mosk)}, volume = {50}, number = {4}, pages = {549-557. Russian}, abstract = {Staphylococcus aureus is a major opportunistic and versatile pathogen. Because the bacteria rapidly evolve multi-resistances towards antibiotics, there is an urgent need to find novel targets and alternative strategies to cure bacterial infections. Here, we provide a brief overview on the knowledge acquired on S. aureus ribosomes, which is one of the major antibiotic targets. We will show that subtle differences exist between the translation at the initiation step of Gram-negative and Gram-positive bacteria although their ribosomes display a remarkable degree of resemblance. In addition, we will illustrate using specific examples the diversity of mechanisms controlling translation initiation in S. aureus that contribute to shape the expression of the virulence factors in a temporal and dynamic manner.}, keywords = {ROMBY, Staphylococcus aureus post-transcriptional regulation quorum sensing regulatory RNAs, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {OB or Not OB: Idiosyncratic utilization of the tRNA-binding OB-fold domain in unicellular, pathogenic eukaryotes.}, author = {D Kapps and M Cela and A Théobald-Dietrich and T Hendrickson and M Frugier}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27714804?dopt=Abstract}, doi = {10.1002/1873-3468.12441}, isbn = {27714804}, year = {2016}, date = {2016-01-01}, journal = {FEBS Lett}, volume = {590}, number = {23}, pages = {4180-4191}, abstract = {In this Review, we examine the so-called 'OB-fold', a tRNA-binding domain homologous to the bacterial tRNA-binding protein Trbp111. We highlight the ability of OB-fold homologs to bind tRNAs and summarize their distribution in evolution. Nature has capitalized on the advantageous effects acquired when an OB-fold domain binds to tRNA by evolutionarily selecting this domain for fusion to different enzymes. Here, we review our current understanding of how the complexity of OB-fold containing proteins and enzymes developed to expand their functions, especially in unicellular, pathogenic eukaryotes. This article is protected by copyright. All rights reserved.}, keywords = {FRUGIER, Trbp111 parasites pathogenic eukaryotes tRNA binding protein, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Packaging of Mason-Pfizer monkey virus (MPMV) genomic RNA depends upon conserved long-range interactions (LRIs) between U5 and gag sequences.}, author = {R M Kalloush and V Vivet-Boudou and L M Ali and F Mustafa and R Marquet and T A Rizvi}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27095024?dopt=Abstract}, doi = {10.1261/rna.055731.115}, isbn = {27095024}, year = {2016}, date = {2016-01-01}, journal = {RNA}, volume = {22}, number = {6}, pages = {905-919}, abstract = {MPMV has great potential for development as a vector for gene therapy. In this respect, precisely defining the sequences and structural motifs that are important for dimerization and packaging of its genomic RNA (gRNA) are of utmost importance. A distinguishing feature of the MPMV gRNA packaging signal is two phylogenetically conserved long-range interactions (LRIs) between U5 andgagcomplementary sequences, LRI-I and LRI-II. To test their biological significance in the MPMV life cycle, we introduced mutations into these structural motifs and tested their effects on MPMV gRNA packaging and propagation. Furthermore, we probed the structure of key mutants using SHAPE (selective 2'hydroxyl acylation analyzed by primer extension). Disrupting base-pairing of the LRIs affected gRNA packaging and propagation, demonstrating their significance to the MPMV life cycle. A double mutant restoring a heterologous LRI-I was fully functional, whereas a similar LRI-II mutant failed to restore gRNA packaging and propagation. These results demonstrate that while LRI-I acts at the structural level, maintaining base-pairing is not sufficient for LRI-II function. In addition, in vitro RNA dimerization assays indicated that the loss of RNA packaging in LRI mutants could not be attributed to the defects in dimerization. Our findings suggest that U5-gagLRIs play an important architectural role in maintaining the structure of the 5' region of the MPMV gRNA, expanding the crucial role of LRIs to the nonlentiviral group of retroviruses.}, keywords = {MARQUET, Mason-Pfizer monkey virus (MPMV) RNA packaging and dimerization RNA secondary structure SHAPE (selective 2′hydroxyl acylation analyzed by primer extension) long-range interactions (LRI) retroviruses, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Experimental Approaches to Study Genome Packaging of Influenza A Viruses.}, author = {C Isel and S Munier and N Naffakh}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27517951?dopt=Abstract}, doi = {10.3390/v8080218}, isbn = {27517951}, year = {2016}, date = {2016-01-01}, journal = {Viruses}, volume = {8}, number = {8}, pages = {218}, abstract = {The genome of influenza A viruses (IAV) consists of eight single-stranded negative sense viral RNAs (vRNAs) encapsidated into viral ribonucleoproteins (vRNPs). It is now well established that genome packaging (i.e., the incorporation of a set of eight distinct vRNPs into budding viral particles), follows a specific pathway guided by segment-specific cis-acting packaging signals on each vRNA. However, the precise nature and function of the packaging signals, and the mechanisms underlying the assembly of vRNPs into sub-bundles in the cytoplasm and their selective packaging at the viral budding site, remain largely unknown. Here, we review the diverse and complementary methods currently being used to elucidate these aspects of the viral cycle. They range from conventional and competitive reverse genetics, single molecule imaging of vRNPs by fluorescence in situ hybridization (FISH) and high-resolution electron microscopy and tomography of budding viral particles, to solely in vitro approaches to investigate vRNA-vRNA interactions at the molecular level.}, keywords = {MARQUET, PAILLART, RNA-RNA interaction competitive reverse genetics influenza virus packaging assay packaging signal single-molecule FISH, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of factors involved in target RNA-directed microRNA degradation.}, author = {G Haas and S Cetin and M Messmer and B Chane-Woon-Ming and O Terenzi and J Chicher and L Kuhn and P Hammann and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26809675?dopt=Abstract}, doi = {10.1093/nar/gkw040}, isbn = {26809675}, year = {2016}, date = {2016-01-01}, journal = {Nucleic Acids Res}, volume = {44}, number = {6}, pages = {2873-2887}, abstract = {The mechanism by which micro (mi)RNAs control their target gene expression is now well understood. It is however less clear how the level of miRNAs themselves is regulated. Under specific conditions, abundant and highly complementary target RNA can trigger miRNA degradation by a mechanism involving nucleotide addition and exonucleolytic degradation. One such mechanism has been previously observed to occur naturally during viral infection. To date, the molecular details of this phenomenon are not known. We report here that both the degree of complementarity and the ratio of miRNA/target abundance are crucial for the efficient decay of the small RNA. Using a proteomic approach based on the transfection of biotinylated antimiRNA oligonucleotides, we set to identify the factors involved in target-mediated miRNA degradation. Among the retrieved proteins, we identified members of the RNA-induced silencing complex, but also RNA modifying and degradation enzymes. We further validate and characterize the importance of one of these, the Perlman Syndrome 3'-5' exonuclease DIS3L2. We show that this protein interacts with Argonaute 2 and functionally validate its role in target-directed miRNA degradation both by artificial targets and in the context of mouse cytomegalovirus infection}, keywords = {PFEFFER, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The expression of a viral microRNA is regulated by clustering to allow optimal B cell transformation.}, author = {J Haar and M Contrant and K Bernhardt and R Feederle and S Diederichs and S Pfeffer and H J Delecluse}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26635399?dopt=Abstract}, doi = {10.1093/nar/gkv1330}, isbn = {26635399}, year = {2016}, date = {2016-01-01}, journal = {Nucleic Acids Res}, volume = {44}, number = {3}, pages = {1326-1341}, abstract = {The Epstein-Barr virus (EBV) transforms B cells by expressing latent proteins and the BHRF1 microRNA cluster. MiR-BHRF1-3, its most transforming member, belongs to the recently identified group of weakly expressed microRNAs. We show here that miR-BHRF1-3 displays an unusually low propensity to form a stem-loop structure, an effect potentiated by miR-BHRF1-3's proximity to the BHRF1 polyA site. Cloning miR-BHRF1-2 or a cellular microRNA, but not a ribozyme, 5' of miR-BHRF1-3 markedly enhanced its expression. However, a virus carrying mutated miR-BHRF1-2 seed regions expressed miR-BHRF1-3 at normal levels and was fully transforming. Therefore, miR-BHRF1-2's role during transformation is independent of its seed regions, revealing a new microRNA function. Increasing the distance between miR-BHRF1-2 and miR-BHRF1-3 in EBV enhanced miR-BHRF1-3's expression but decreased its transforming potential. Thus, the expression of some microRNAs must be restricted to a narrow range, as achieved by placing miR-BHRF1-3 under the control of miR-BHRF1-2.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Translational regulation of APOBEC3G mRNA by Vif requires its 5′UTR and contributes to restoring HIV-1 infectivity}, author = {S X Guerrero and C Libre and J Batisse and G Mercenne and D Richer and G Laumond and T Decoville and C Moog and R Marquet and J C Paillart}, url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5171582/}, doi = {10.1038/srep39507}, year = {2016}, date = {2016-01-01}, journal = {Sci Rep}, volume = {6}, pages = {39507}, abstract = {The essential HIV-1 viral infectivity factor (Vif) allows productive infection of non-permissive cells expressing cytidine deaminases APOBEC3G (A3G) and A3F by decreasing their cellular level, and preventing their incorporation into virions. Unlike the Vif-induced degradation of A3G, the functional role of the inhibition of A3G translation by Vif remained unclear. Here, we show that two stem-loop structures within the 5メ-untranslated region of A3G mRNA are crucial for translation inhibition by Vif in cells, and most Vif alleles neutralize A3G translation efficiently. Interestingly, K26R mutation in Vif abolishes degradation of A3G by the proteasome but has no effect at the translational level, indicating these two pathways are independent. These two mechanisms, proteasomal degradation and translational inhibition, similarly contribute to decrease the cellular level of A3G by Vif and to prevent its incorporation into virions. Importantly, inhibition of A3G translation is sufficient to partially restore viral infectivity in the absence of proteosomal degradation. These findings demonstrate that HIV-1 has evolved redundant mechanisms to specifically inhibit the potent antiviral activity of A3G.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Quantitative and predictive model of kinetic regulation by E. coli TPP riboswitches.}, author = {S Guedich and B Puffer-Enders and M Baltzinger and G Hoffmann and C Da Veiga and F Jossinet and S Thore and G Bec and E Ennifar and D Burnouf and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26932506?dopt=Abstract}, doi = {10.1080/15476286.2016.1142040}, isbn = {26932506}, year = {2016}, date = {2016-01-01}, journal = {RNA Biol}, volume = {13}, number = {4}, pages = {373-390}, abstract = {Riboswitches are non-coding elements upstream or downstream of mRNAs that, upon binding of a specific ligand, regulate transcription and/or translation initiation in bacteria, or alternative splicing in plants and fungi. We have studied thiamine pyrophosphate (TPP) riboswitches regulating translation of thiM operon and transcription and translation of thiC operon in E. coli, and that of THIC in the plant A. thaliana. For all, we ascertained an induced-fit mechanism involving initial binding of the TPP followed by a conformational change leading to a higher-affinity complex. The experimental values obtained for all kinetic and thermodynamic parameters of TPP binding imply that the regulation by A. thaliana riboswitch is governed by mass-action law, whereas it is of kinetic nature for the two bacterial riboswitches. Kinetic regulation requires that the RNA polymerase pauses after synthesis of each riboswitch aptamer to leave time for TPP binding, but only when its concentration is sufficient. A quantitative model of regulation highlighted how the pausing time has to be linked to the kinetic rates of initial TPP binding to obtain an ON/OFF switch in the correct concentration range of TPP. We verified the existence of these pauses and the model prediction on their duration. Our analysis also led to quantitative estimates of the respective efficiency of kinetic and thermodynamic regulations, which shows that kinetically regulated riboswitches react more sharply to concentration variation of their ligand than thermodynamically regulated riboswitches. This rationalizes the interest of kinetic regulation and confirms empirical observations that were obtained by numerical simulations.}, keywords = {ENNIFAR, Kinetic Regulation kinITC pyrophosphate riboswitches thiamine, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {An integrated, structure- and energy-based view of the genetic code.}, author = {H Grosjean and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27448410?dopt=Abstract}, doi = {10.1093/nar/gkw608}, isbn = {27448410}, year = {2016}, date = {2016-01-01}, journal = {Nucleic Acids Res}, volume = {44}, number = {17}, pages = {8020-8040}, abstract = {The principles of mRNA decoding are conserved among all extant life forms. We present an integrative view of all the interaction networks between mRNA, tRNA and rRNA: the intrinsic stability of codon-anticodon duplex, the conformation of the anticodon hairpin, the presence of modified nucleotides, the occurrence of non-Watson-Crick pairs in the codon-anticodon helix and the interactions with bases of rRNA at the A-site decoding site. We derive a more information-rich, alternative representation of the genetic code, that is circular with an unsymmetrical distribution of codons leading to a clear segregation between GC-rich 4-codon boxes and AU-rich 2:2-codon and 3:1-codon boxes. All tRNA sequence variations can be visualized, within an internal structural and energy framework, for each organism, and each anticodon of the sense codons. The multiplicity and complexity of nucleotide modifications at positions 34 and 37 of the anticodon loop segregate meaningfully, and correlate well with the necessity to stabilize AU-rich codon-anticodon pairs and to avoid miscoding in split codon boxes. The evolution and expansion of the genetic code is viewed as being originally based on GC content with progressive introduction of A/U together with tRNA modifications. The representation we present should help the engineering of the genetic code to include non-natural amino acids.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selenoprotein Gene Nomenclature.}, author = {V N Gladyshev and E S Arnér and M J Berry and R Brigelius-Flohé and E A Bruford and R F Burk and B A Carlson and S Castellano and L Chavatte and M Conrad and P R Copeland and A M Diamond and D M Driscoll and A Ferreiro and L Flohé and F R Green and R Guigó and D E Handy and D L Hatfield and J Hesketh and P R Hoffmann and A Holmgren and R J Hondal and M T Howard and K Huang and H Y Kim and I Y Kim and J Köhrle and A Krol and G V Kryukov and B J Lee and B C Lee and X G Lei and Q Liu and A Lescure and A V Lobanov and J Loscalzo and M Maiorino and M Mariotti and K S Prabhu and M P Rayman and S Rozovsky and G Salinas and L Schomburg and U Schweizer and M Simonović and R A Sunde and P A Tsuji and S Tweedie and F Ursini and Y Zhang}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27645994?dopt=Abstract}, doi = {10.1074/jbc.M116.756155}, isbn = {27645994}, year = {2016}, date = {2016-01-01}, journal = {J Biol Chem}, volume = {291}, number = {46}, pages = {24036-24040}, abstract = {The human genome contains 25 genes coding for selenocysteine-containing proteins (selenoproteins). These proteins are involved in a variety of functions, most notably redox homeostasis. Selenoprotein enzymes with known functions are designated according to these functions: TXNRD1, TXNRD2, and TXNRD3 (thioredoxin reductases), GPX1, GPX2, GPX3, GPX4 and GPX6 (glutathione peroxidases), DIO1, DIO2, and DIO3 (iodothyronine deiodinases), MSRB1 (methionine-R-sulfoxide reductase 1) and SEPHS2 (selenophosphate synthetase 2). Selenoproteins without known functions have traditionally been denoted by SEL or SEP symbols. However, these symbols are sometimes ambiguous and conflict with the approved nomenclature for several other genes. Therefore, there is a need to implement a rational and coherent nomenclature system for selenoprotein-encoding genes. Our solution is to use the root symbol SELENO followed by a letter. This nomenclature applies to SELENOF (selenoprotein F, the 15 kDa selenoprotein, SEP15), SELENOH (selenoprotein H, SELH, C11orf31), SELENOI (selenoprotein I, SELI, EPT1), SELENOK (selenoprotein K, SELK), SELENOM (selenoprotein M, SELM), SELENON (selenoprotein N, SEPN1, SELN), SELENOO (selenoprotein O, SELO), SELENOP (selenoprotein P, SeP, SEPP1, SELP), SELENOS (selenoprotein S, SELS, SEPS1, VIMP), SELENOT (selenoprotein T, SELT), SELENOV (selenoprotein V, SELV) and SELENOW (selenoprotein W, SELW, SEPW1). This system, approved by the HUGO Gene Nomenclature Committee, also resolves conflicting, missing and ambiguous designations for selenoprotein genes and is applicable to selenoproteins across vertebrates.}, keywords = {function gene name genomics nomenclature selenium selenocysteine selenoprotein structure-function, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Influenza NA and PB1 Gene Segments Interact during the Formation of Viral Progeny: Localization of the Binding Region within the PB1 Gene.}, author = {B Gilbertson and T Zheng and M Gerber and A Printz-Schweigert and C Ong and R Marquet and C Isel and S Rockman and L Bown}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27556479?dopt=Abstract}, doi = {10.3390/v8080238}, isbn = {27556479}, year = {2016}, date = {2016-01-01}, journal = {Viruses}, volume = {8}, number = {8}, pages = {238}, abstract = {he influenza A virus genome comprises eight negative-sense viral RNAs (vRNAs) that form individual ribonucleoprotein (RNP) complexes. In order to incorporate a complete set of each of these vRNAs, the virus uses a selective packaging mechanism that facilitates co-packaging of specific gene segments but whose molecular basis is still not fully understood. Recently, we used a competitive transfection model where plasmids encoding the A/Puerto Rico/8/34 (PR8) and A/Udorn/307/72 (Udorn) PB1 gene segments were competed to show that the Udorn PB1 gene segment is preferentially co-packaged into progeny virions with the Udorn NA gene segment. Here we created chimeric PB1 genes combining both Udorn and PR8 PB1 sequences to further define the location within the Udorn PB1 gene that drives co-segregation of these genes and show that nucleotides 1776-2070 of the PB1 gene are crucial for preferential selection. In vitro assays examining specific interactions between Udorn NA vRNA and purified vRNAs transcribed from chimeric PB1 genes also supported the importance of this region in the PB1-NA interaction. Hence, this work identifies an association between viral genes that are co-selected during packaging. It also reveals a region potentially important in the RNP-RNP interactions within the supramolecular complex that is predicted to form prior to budding to allow one of each segment to be packaged in the viral progeny. Our study lays the foundation to understand the co-selection of specific genes, which may be critical to the emergence of new viruses with pandemic potential.}, keywords = {MARQUET, PAILLART, RNA-RNA interaction competitive transfection gene segments influenza virus packaging reassortment viral polymerase, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoacyl-tRNA Synthetases in the Bacterial World.}, author = {R Giege and M Springer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27223819?dopt=Abstract}, doi = {10.1128/ecosalplus.ESP-0002-2016}, isbn = {27223819}, year = {2016}, date = {2016-01-01}, journal = {EcoSal Plus}, volume = {7}, number = {1}, pages = {none}, abstract = {Aminoacyl-tRNA synthetases (aaRSs) are modular enzymes globally conserved in the three kingdoms of life. All catalyze the same two-step reaction, i.e., the attachment of a proteinogenic amino acid on their cognate tRNAs, thereby mediating the correct expression of the genetic code. In addition, some aaRSs acquired other functions beyond this key role in translation. Genomics and X-ray crystallography have revealed great structural diversity in aaRSs (e.g., in oligomery and modularity, in ranking into two distinct groups each subdivided in 3 subgroups, by additional domains appended on the catalytic modules). AaRSs show huge structural plasticity related to function and limited idiosyncrasies that are kingdom or even species specific (e.g., the presence in many Bacteria of non discriminating aaRSs compensating for the absence of one or two specific aaRSs, notably AsnRS and/or GlnRS). Diversity, as well, occurs in the mechanisms of aaRS gene regulation that are not conserved in evolution, notably between distant groups such as Gram-positive and Gram-negative Bacteria. The review focuses on bacterial aaRSs (and their paralogs) and covers their structure, function, regulation, and evolution. Structure/function relationships are emphasized, notably the enzymology of tRNA aminoacylation and the editing mechanisms for correction of activation and charging errors. The huge amount of genomic and structural data that accumulated in last two decades is reviewed, showing how the field moved from essentially reductionist biology towards more global and integrated approaches. Likewise, the alternative functions of aaRSs and those of aaRS paralogs (e.g., during cell wall biogenesis and other metabolic processes in or outside protein synthesis) are reviewed. Since aaRS phylogenies present promiscuous bacterial, archaeal, and eukaryal features, similarities and differences in the properties of aaRSs from the three kingdoms of life are pinpointed throughout the review and distinctive characteristics of bacterium-like synthetases from organelles are outlined}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Buffering deleterious polymorphisms in highly constrained parts of HIV-1 envelope by flexible regions.}, author = {R Gasser and M Hamoudi and M Pellicciotta and Z Zhou and C Visdeloup and P Colin and M Braibant and B Lagane and M Negroni}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27473399?dopt=Abstract}, doi = {10.1186/s12977-016-0285-6}, year = {2016}, date = {2016-01-01}, urldate = {2016-01-01}, journal = {Retrovirology}, volume = {13}, number = {1}, pages = {50}, abstract = {BACKGROUND: Covariation is an essential process that leads to coevolution of parts of proteins and genomes. In organisms subject to strong selective pressure, coevolution is central to keep the balance between the opposite requirements of antigenic variation and retention of functionality. Being the viral component most exposed to the external environment, the HIV-1 glycoprotein gp120 constitutes the main target of the immune response. Accordingly its more external portions are characterised by extensive sequence heterogeneity fostering constant antigenic variation. RESULTS: We report that a single polymorphism, present at the level of the viral population in the conserved internal region C2, was sufficient to totally abolish Env functionality when introduced in an exogenous genetic context. The prominent defect of the non-functional protein is a block occurring after recognition of the co-receptor CCR5, likely due to an interference with the subsequent conformational changes that lead to membrane fusion. We also report that the presence of compensatory polymorphisms at the level of the external and hypervariable region V3 fully restored the functionality of the protein. The functional revertant presents different antigenic profiles and sensitivity to the entry inhibitor TAK 779. CONCLUSIONS: Our data suggest that variable regions, besides harbouring intrinsic extensive antigenic diversity, can also contribute to sequence diversification in more structurally constrained parts of the gp120 by buffering the deleterious effect of polymorphisms, further increasing the genetic flexibility of the protein and the antigenic repertoire of the viral population.}, keywords = {Antigenic variation Coevolution Envelope HIV Viral entry, NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transfer RNA: from pioneering crystallographic studies to contemporary tRNA biology.}, author = {P Fernandez-Millan and C Schelcher and J Chihade and B Masquida and P Giege and C Sauter}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26968773?dopt=Abstract}, doi = {10.1016/j.abb.2016.03.005}, isbn = {26968773}, year = {2016}, date = {2016-01-01}, journal = {Arch Biochem Biophys}, volume = {602}, pages = {95-105}, abstract = {Transfer RNAs (tRNAs) play a key role in protein synthesis as adaptor molecules between messenger RNA and protein sequences on the ribosome. Their discovery in the early sixties provoked a worldwide infatuation with the study of their architecture and their function in the decoding of genetic information. tRNAs are also emblematic molecules in crystallography: the determination of the first tRNA crystal structures represented a milestone in structural biology and tRNAs were for a long period the sole source of information on RNA folding, architecture, and post-transcriptional modifications. Crystallographic data on tRNAs in complex with aminoacyl-tRNA synthetases (aaRSs) also provided the first insight into protein:RNA interactions. Beyond the translation process and the history of structural investigations on tRNA, this review also illustrates the renewal of tRNA biology with the discovery of a growing number of tRNA partners in the cell, the involvement of tRNAs in a variety of regulatory and metabolic pathways, and emerging applications in biotechnology and synthetic biology.}, keywords = {RNA:protein recognition crystallography genetic code protein synthesis transfer RNA translation, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Traditional Chemical Mapping of RNA Structure In Vitro and In Vivo.}, author = {P Fechter and D Parmentier and Z Wu and O Fuchsbauer and P Romby and S Marzi}, editor = {D Turner and D Mathews}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27665595?dopt=Abstract}, doi = {10.1007/978-1-4939-6433-8_7}, isbn = {27665595}, year = {2016}, date = {2016-01-01}, booktitle = {RNA Structure Determination: Methods and Protocols}, volume = {1490}, pages = {83-103}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Chemical probing is often used to gain knowledge on the secondary and tertiary structures of RNA molecules either free or engaged in complexes with ligands. The method monitors the reactivity of each nucleotide towards chemicals of various specificities reflecting the hydrogen bonding environment of each nucleotide within the RNA molecule. In addition, information can be obtained on the binding site of a ligand (noncoding RNAs, protein, metabolites), and on RNA conformational changes that accompanied ligand binding or perturbation of the environmental cues. The detection of the modifications can be obtained either by using end-labeled RNA molecules or by primer extension using reverse transcriptase. The goal of this chapter is to provide the reader with an experimental guide to probe the structure of RNA in vitro and in vivo with the most suitable chemical probes.}, keywords = {Chemical mapping Lead(II) induced cleavages RNA structure probing, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @book{, title = {Nucleic Acid Crystallography: Methods and Protocols}, editor = {E Ennifar}, year = {2016}, date = {2016-01-01}, volume = {1320}, publisher = {Humana Press}, address = {New York, NY}, series = {Methods in Molecular Biology}, abstract = {This volume includes comprehensive and up-to-date coverage of all nucleic acid-specific steps used in X-ray crystallography, from macromolecule production to structure determination. Chapters dedicated to RNA preparation and crystallogenesis will be of interest to beginners, while chapters focused on data collection, phasing and refinement will be particularly useful to researchers with a higher level of expertise. Several functional case studies are also presented in the last part of the book. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and thorough, Nucleic Acid Crystallography: Methods and Protocols presents protocols that are aimed at both researchers and students who are interested in the structural biology of DNA or RNA, alone or in complex with proteins or ligands.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {book} } @article{, title = {Genome engineering in the yeast pathogen Candida glabrata using the CRISPR-Cas9 system.}, author = {L Enkler and D Richer and A L Marchand and D Ferrandon and F Jossinet}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27767081?dopt=Abstract}, doi = {10.1038/srep35766}, isbn = {27767081}, year = {2016}, date = {2016-01-01}, journal = {Sci Rep}, volume = {6}, pages = {35766}, abstract = {Among Candida species, the opportunistic fungal pathogen Candida glabrata has become the second most common causative agent of candidiasis in the world and a major public health concern. Yet, few molecular tools and resources are available to explore the biology of C. glabrata and to better understand its virulence during infection. In this study, we describe a robust experimental strategy to generate loss-of-function mutants in C. glabrata. The procedure is based on the development of three main tools: (i) a recombinant strain of C. glabrata constitutively expressing the CRISPR-Cas9 system, (ii) an online program facilitating the selection of the most efficient guide RNAs for a given C. glabrata gene, and (iii) the identification of mutant strains by the Surveyor technique and sequencing. As a proof-of-concept, we have tested the virulence of some mutants in vivo in a Drosophila melanogaster infection model. Our results suggest that yps11 and a previously uncharacterized serine/threonine kinase are involved, directly or indirectly, in the ability of the pathogenic yeast to infect this model host organism.}, keywords = {JOSSINET, M3i, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Extending ITC to Kinetics with kinITC.}, author = {P Dumas and E Ennifar and C Da Veiga and G Bec and W Palau and C Di Primo and A Piñeiro and J Sabin and E Muñoz and J Rial}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26794354?dopt=Abstract}, doi = {10.1016/bs.mie.2015.08.026}, isbn = {26794354}, year = {2016}, date = {2016-01-01}, journal = {Methods Enzymol}, volume = {567}, pages = {157-180}, abstract = {Isothermal titration calorimetry (ITC) has long been used for kinetic studies in chemistry, but this remained confined to enzymatic studies in the biological field. In fact, the biological community has long had the tendency of ignoring the kinetic possibilities of ITC considering it solely as a thermodynamic technique, whereas surface plasmon resonance is seen as the kinetic technique par excellence. However, the primary signal recorded by ITC is a heat power which is directly related to the kinetics of the reaction. Here, it is shown how this kinetic signal can be recovered by using kinITC, the kinetic extension of ITC. The theoretical basis of kinITC is detailed for the most common situation of a second-order reaction A+B Ω C characterized by kinetic parameters kon, koff. A simplified kinITC-ETC method based upon the determination of an "Equilibration Time Curve" (ETC) is presented. The ETC is obtained by automatic determination of the "effective end" of each injection. The method is illustrated with experimental results with a comparison to Surface Plasmon Resonance (SPR) data. kon values were obtained in a wide range, from 10(3) to 0.5×10(6)M(-1)s(-1). All procedures were implemented in the program AFFINImeter (https://www.affinimeter.com/).}, keywords = {Biological calorimetry Equilibration time curve ITC Kinetics Microcalorimetry kinITC, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {'Z-DNA like' fragments in RNA: a recurring structural motif with implications for folding, RNA/protein recognition and immune response.}, author = {L D'Ascenzo and F Leonarski and Q Vicens and P Auffinger}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27151194?dopt=Abstract}, doi = {10.1093/nar/gkw388}, isbn = {27151194}, year = {2016}, date = {2016-01-01}, journal = {Nucleic Acids Res}, volume = {44}, number = {12}, pages = {5944-5956}, abstract = {Since the work of Alexander Rich, who solved the first Z-DNA crystal structure, we have known that d(CpG) steps can adopt a particular structure that leads to forming left-handed helices. However, it is still largely unrecognized that other sequences can adopt 'left-handed' conformations in DNA and RNA, in double as well as single stranded contexts. These 'Z-like' steps involve the coexistence of several rare structural features: a C2'-endo puckering, a syn nucleotide and a lone pair-π stacking between a ribose O4' atom and a nucleobase. This particular arrangement induces a conformational stress in the RNA backbone, which limits the occurrence of Z-like steps to ≈0.1% of all dinucleotide steps in the PDB. Here, we report over 600 instances of Z-like steps, which are located within r(UNCG) tetraloops but also in small and large RNAs including riboswitches, ribozymes and ribosomes. Given their complexity, Z-like steps are probably associated with slow folding kinetics and once formed could lock a fold through the formation of unique long-range contacts. Proteins involved in immunologic response also specifically recognize/induce these peculiar folds. Thus, characterizing the conformational features of these motifs could be a key to understanding the immune response at a structural level.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Anions in Nucleic Acid Crystallography}, author = {L D'Ascenzo and P Auffinger}, editor = {E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26227054?dopt=Abstract}, doi = {10.1007/978-1-4939-2763-0_22}, isbn = {26227054}, year = {2016}, date = {2016-01-01}, booktitle = {Nucleic Acid Crystallography: Methods and Protocols}, volume = {1320}, pages = {337-351}, publisher = {Humana Press}, address = {NY}, series = {Methods in Molecular Biology}, abstract = {Nucleic acid crystallization buffers contain a large variety of chemicals fitting specific needs. Among them, anions are often solely considered for pH-regulating purposes and as cationic co-salts while their ability to directly bind to nucleic acid structures is rarely taken into account. Here we review current knowledge related to the use of anions in crystallization buffers along with data on their biological prevalence. Chloride ions are frequently identified in crystal structures but display low cytosolic concentrations. Hence, they are thought to be distant from nucleic acid structures in the cell. Sulfate ions are also frequently identified in crystal structures but their localization in the cell remains elusive. Nevertheless, the characterization of the binding properties of these ions is essential for better interpreting the solvent structure in crystals and consequently, avoiding mislabeling of electron densities. Furthermore, understanding the binding properties of these anions should help to get clues related to their potential effects in crowded cellular environments.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Isothermal Titration Calorimetry: Assisted Crystallization of RNA-Ligand Complexes}, author = {C Da Veiga and J Mezher and P Dumas and E Ennifar}, editor = {E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26227041?dopt=Abstract}, doi = {10.1007/978-1-4939-2763-0_9}, isbn = {26227041}, year = {2016}, date = {2016-01-01}, booktitle = {Nucleic Acid Crystallography: Methods and Protocols}, volume = {1320}, pages = {127-143}, publisher = {Humana Press}, address = {NY}, series = {Methods in Molecular Biology}, abstract = {The success rate of nucleic acids/ligands co-crystallization can be significantly improved by performing preliminary biophysical analyses. Among suitable biophysical approaches, isothermal titration calorimetry (ITC) is certainly a method of choice. ITC can be used in a wide range of experimental conditions to monitor in real time the formation of the RNA- or DNA-ligand complex, with the advantage of providing in addition the complete binding profile of the interaction. Following the ITC experiment, the complex is ready to be concentrated for crystallization trials. This chapter describes a detailed experimental protocol for using ITC as a tool for monitoring RNA/small molecule binding, followed by co-crystallization.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{lamiable_analysis_2016, title = {Analysis of the Contribution of Hemocytes and Autophagy to Drosophila Antiviral Immunity}, author = {Olivier Lamiable and Johan Arnold and Isaque Joao Silva da de Faria and Roenick Proveti Olmo and Francesco Bergami and Carine Meignin and Jules A Hoffmann and Joao Trindade Marques and Jean-Luc Imler}, url = {http://jvi.asm.org/content/90/11/5415}, doi = {10.1128/JVI.00238-16}, issn = {0022-538X, 1098-5514}, year = {2016}, date = {2016-01-01}, urldate = {2016-06-05}, journal = {J. Virol.}, volume = {90}, number = {11}, pages = {5415--5426}, keywords = {antiviral immunity, Autophagy, Hemocytes, hoffmann, imler, M3i, meignin}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 mutation and recombination rates are different in macrophages and T-cells}, author = {D Cromer and T E Schlub and R P Smyth and A J Grimm and A Chopra and S Mallal and M P Davenport and J Mak}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27110814?dopt=Abstract}, doi = {10.3390/v8040118}, year = {2016}, date = {2016-01-01}, journal = {Viruses}, volume = {8}, number = {4}, pages = {118}, abstract = {High rates of mutation and recombination help human immunodeficiency virus (HIV) to evade the immune system and develop resistance to antiretroviral therapy. Macrophages and T-cells are the natural target cells of HIV-1 infection. A consensus has not been reached as to whether HIV replication results in differential recombination between primary T-cells and macrophages. Here, we used HIV with silent mutation markers along with next generation sequencing to compare the mutation and the recombination rates of HIV directly in T lymphocytes and macrophages. We observed a more than four-fold higher recombination rate of HIV in macrophages compared to T-cells (p < 0.001) and demonstrated that this difference is not due to different reliance on C-X-C chemokine receptor type 4 (CXCR4) and C-C chemokine receptor type 5 (CCR5) co-receptors between T-cells and macrophages. We also found that the pattern of recombination across the HIV genome (hot and cold spots) remains constant between T-cells and macrophages despite a three-fold increase in the overall recombination rate. This indicates that the difference in rates is a general feature of HIV DNA synthesis during macrophage infection. In contrast to HIV recombination, we found that T-cells have a 30% higher mutation rate than macrophages (p < 0.001) and that the mutational profile is similar between these cell types. Unexpectedly, we found no association between mutation and recombination in macrophages, in contrast to T-cells. Our data highlights some of the fundamental difference of HIV recombination and mutation amongst these two major target cells of infection. Understanding these differences will provide invaluable insights toward HIV evolution and how the virus evades immune surveillance and anti-retroviral therapeutics.}, keywords = {HIV mutation recombination evolution, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{marques_diversity_2016, title = {The diversity of insect antiviral immunity: insights from viruses}, author = {João T Marques and Jean-Luc Imler}, url = {http://www.sciencedirect.com/science/article/pii/S1369527416300571}, doi = {10.1016/j.mib.2016.05.002}, issn = {1369-5274}, year = {2016}, date = {2016-01-01}, urldate = {2016-06-05}, journal = {Current Opinion in Microbiology}, volume = {32}, pages = {71--76}, abstract = {Insects represent over 70% of all animal species. Recent virome analyses reveal unprecedented genetic diversity of insect viruses, which appears to match that of their hosts. Thus, insect-virus interactions may provide information on a vast repertoire of antiviral immune mechanisms. Tapping into this diversity is challenging because of several constraints imposed by the uniqueness of each insect model. Nevertheless, it is clear that many conserved and divergent pathways participate in the control of viral infection in insects. Co-evolution between hosts and viruses favors the development of immune evasion mechanisms by the pathogen. Viral suppressors can offer unique perspective on host pathways and emphasize the importance of RNA interference, apoptosis, but also NF-κB pathways and translation control in insect antiviral immunity.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structures of a group II intron lariat primed for reverse splicing.}, author = {M Costa and H Walbott and D Monachello and E Westhof and F Michel}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27934709?dopt=Abstract}, doi = {10.1126/science.aaf9258}, isbn = {27934709}, year = {2016}, date = {2016-01-01}, journal = {Science}, volume = {354}, number = {6316}, pages = {aaf9258}, abstract = {The 2'-5' branch of nuclear premessenger introns is believed to have been inherited from self-splicing group II introns, which are retrotransposons of bacterial origin. Our crystal structures at 3.4 and 3.5 angstrom of an excised group II intron in branched ("lariat") form show that the 2'-5' branch organizes a network of active-site tertiary interactions that position the intron terminal 3'-hydroxyl group into a configuration poised to initiate reverse splicing, the first step in retrotransposition. Moreover, the branchpoint and flanking helices must undergo a base-pairing switch after branch formation. A group II-based model of the active site of the nuclear splicing machinery (the spliceosome) is proposed. The crucial role of the lariat conformation in active-site assembly and catalysis explains its prevalence in modern splicing.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{martins_discovery_2016, title = {Discovery of novel targets for antivirals: learning from flies}, author = {Nelson Martins and Jean-Luc Imler and Carine Meignin}, url = {http://www.sciencedirect.com/science/article/pii/S1879625716301274}, doi = {10.1016/j.coviro.2016.09.005}, issn = {1879-6265}, year = {2016}, date = {2016-01-01}, journal = {Curr Opin Virol}, volume = {20}, pages = {64--70}, abstract = {Developing antiviral drugs is challenging due to the small number of targets in viruses, and the rapid evolution of viral genes. Animals have evolved a number of efficient antiviral defence mechanisms, which can serve as a source of inspiration for novel therapies. The genetically tractable insect Drosophila belongs to the most diverse group of animals. Genetic and transcriptomic analyses have recently identified Drosophila genes encoding viral restriction factors. Some of them represent evolutionary novelties and their characterization may provide hints for the design of directly acting antivirals. In addition, functional screens revealed conserved host factors required for efficient viral translation, such as the ribosomal protein RACK1 and the release factor Pelo. These proteins are promising candidates for host-targeted antivirals.}, keywords = {antiviral, imler, M3i, meignin, target}, pubstate = {published}, tppubtype = {article} } @article{, title = {A pseudouridylation switch in rRNA is implicated in ribosome function during the life cycle of Trypanosoma brucei.}, author = {V Chikne and T Doniger and K S Rajan and O Bartok and D Eliaz and S Cohen-Chalamish and C Tschudi and R Unger and Y Hashem and S Kadener and S Michaeli}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27142987?dopt=Abstract}, doi = {10.1038/srep25296}, isbn = {27142987}, year = {2016}, date = {2016-01-01}, journal = {Sci Rep}, volume = {6}, pages = {25296}, abstract = {The protozoan parasite Trypanosoma brucei, which causes devastating diseases in humans and animals in sub-Saharan Africa, undergoes a complex life cycle between the mammalian host and the blood-feeding tsetse fly vector. However, little is known about how the parasite performs most molecular functions in such different environments. Here, we provide evidence for the intriguing possibility that pseudouridylation of rRNA plays an important role in the capacity of the parasite to transit between the insect midgut and the mammalian bloodstream. Briefly, we mapped pseudouridines (Ψ) on rRNA by Ψ-seq in procyclic form (PCF) and bloodstream form (BSF) trypanosomes. We detected 68 Ψs on rRNA, which are guided by H/ACA small nucleolar RNAs (snoRNA). The small RNome of both life cycle stages was determined by HiSeq and 83 H/ACAs were identified. We observed an elevation of 21 Ψs modifications in BSF as a result of increased levels of the guiding snoRNAs. Overexpression of snoRNAs guiding modification on H69 provided a slight growth advantage to PCF parasites at 30 °C. Interestingly, these modifications are predicted to significantly alter the secondary structure of the large subunit (LSU) rRNA suggesting that hypermodified positions may contribute to the adaption of ribosome function during cycling between the two hosts.}, keywords = {HASHEM, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{lamiable_wntd_2016, title = {WntD and Diedel: Two immunomodulatory cytokines in Drosophila immunity}, author = {Olivier Lamiable and Carine Meignin and Jean-Luc Imler}, url = {http://www.tandfonline.com/doi/abs/10.1080/19336934.2016.1202387?journalCode=kfly20}, doi = {10.1080/19336934.2016.1202387}, issn = {1933-6942}, year = {2016}, date = {2016-01-01}, journal = {Fly (Austin)}, volume = {10}, number = {4}, pages = {187--194}, abstract = {Remarkable progress has been made on the understanding of the basic mechanisms of innate immunity in flies, from sensing infection to production of effector molecules. However, how the immune response is orchestrated at the level of the organism remains poorly understood. While cytokines activating immune responses, such as Spaetzle or Unpaired-3, have been identified and characterized in Drosophila, much less is known regarding immunosuppressor cytokines. In a recent publication, we reported the identification of a novel cytokine, Diedel, which acts as systemic negative regulator of the IMD pathway. Here, we discuss the similarities between Diedel and WntD, another immunomodulatory cytokine and present evidence that the 2 molecules act independently from one another.}, keywords = {Cytokines, Drosophila, IMD pathway, imler, innate immunity, M3i, meignin, virus}, pubstate = {published}, tppubtype = {article} } @article{, title = {Multianalytical Study of the Binding between a Small Chiral Molecule and a DNA Aptamer: Evidence for Asymmetric Steric Effect upon 3'- versus 5'-End Sequence Modification.}, author = {L Challier and R Miranda-Castro and B Barbe and C Fave and B Limoges and E Peyrin and C Ravelet and E Fiore and P Labbé and L Coche-Guérente and E Ennifar and G Bec and P Dumas and F Mavré and V Noël}, url = {https://www.ncbi.nlm.nih.gov/pubmed/27934108?dopt=Abstract}, doi = {10.1021/acs.analchem.6b04046}, isbn = {27934108}, year = {2016}, date = {2016-01-01}, journal = {Anal Chem}, volume = {88}, number = {23}, pages = {11963-11971}, abstract = {Nucleic acid aptamers are involved in a broad field of applications ranging from therapeutics to analytics. Deciphering the binding mechanisms between aptamers and small ligands is therefore crucial to improve and optimize existing applications and to develop new ones. Particularly interesting is the enantiospecific binding mechanism involving small molecules with nonprestructured aptamers. One archetypal example is the chiral binding between l-tyrosinamide and its 49-mer aptamer for which neither structural nor mechanistic information is available. In the present work, we have taken advantage of a multiple analytical characterization strategy (i.e., using electroanalytical techniques such as kinetic rotating droplet electrochemistry, fluorescence polarization, isothermal titration calorimetry, and quartz crystal microbalance) for interpreting the nature of binding process. Screening of the binding thermodynamics and kinetics with a wide range of aptamer sequences revealed the lack of symmetry between the two ends of the 23-mer minimal binding sequence, showing an unprecedented influence of the 5' aptamer modification on the bimolecular binding rate constant kon and no significant effect on the dissociation rate constant koff. The results we have obtained lead us to conclude that the enantiospecific binding reaction occurs through an induced-fit mechanism, wherein the ligand promotes a primary nucleation binding step near the 5'-end of the aptamer followed by a directional folding of the aptamer around its target from 5'-end to 3'-end. Functionalization of the 5'-end position by a chemical label, a polydA tail, a protein, or a surface influences the kinetic/thermodynamic constants up to 2 orders of magnitude in the extreme case of a surface immobilized aptamer, while significantly weaker effect is observed for a 3'-end modification. The reason is that steric hindrance must be overcome to nucleate the binding complex in the presence of a modification near the nucleation site.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{baron_lbp/bpi_2016, title = {The LBP/BPI multigenic family in invertebrates: Evolutionary history and evidences of specialization in mollusks}, author = {Olga Lucia Baron and Emeline Deleury and Jean-Marc Reichhart and Christine Coustau}, url = {http://linkinghub.elsevier.com/retrieve/pii/S0145305X15300756}, doi = {10.1016/j.dci.2015.11.006}, issn = {0145305X}, year = {2016}, date = {2016-01-01}, urldate = {2017-07-12}, journal = {Developmental & Comparative Immunology}, volume = {57}, pages = {20--30}, keywords = {BPI, Evolution, Invertebrate, LBP, M3i, mollusks, multigenic family, reichhart}, pubstate = {published}, tppubtype = {article} } @article{, title = {Staphylococcus aureus RNAIII and Its Regulon Link Quorum Sensing, Stress Responses, Metabolic Adaptation, and Regulation of Virulence Gene Expression.}, author = {D Bronesky and Z Wu and S Marzi and P Walter and T Geissmann and K Moreau and F Vandenesch and I Caldelari and P Romby}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27482744}, doi = {10.1146/annurev-micro-102215-095708}, isbn = {27482744}, year = {2016}, date = {2016-01-01}, journal = {Annu Rev Microbiol}, volume = {70}, pages = {299-316}, abstract = {Staphylococcus aureus RNAIII is one of the main intracellular effectors of the quorum-sensing system. It is a multifunctional RNA that encodes a small peptide, and its noncoding parts act as antisense RNAs to regulate the translation and/or the stability of mRNAs encoding transcriptional regulators, major virulence factors, and cell wall metabolism enzymes. In this review, we explain how regulatory proteins and RNAIII are embedded in complex regulatory circuits to express virulence factors in a dynamic and timely manner in response to stress and environmental and metabolic changes. Expected final online publication date for the Annual Review of Microbiology Volume 70 is September 08, 2016. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Apicomplexa-specific tRip facilitates import of exogenous tRNAs into malaria parasites}, author = {T Bour and N Mahmoudi and D Kapps and S Thiberge and D Bargieri and R Ménard and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27071116}, doi = {10.1073/pnas.1600476113}, isbn = {27071116}, year = {2016}, date = {2016-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {113}, number = {17}, pages = {4717-4722}, abstract = {The malaria-causingPlasmodiumparasites are transmitted to vertebrates by mosquitoes. To support their growth and replication, these intracellular parasites, which belong to the phylumApicomplexa,have developed mechanisms to exploit their hosts. These mechanisms include expropriation of small metabolites from infected host cells, such as purine nucleotides and amino acids. Heretofore, no evidence suggested that transfer RNAs (tRNAs) could also be exploited. We identified an unusual gene inApicomplexawith a coding sequence for membrane-docking and structure-specific tRNA binding. ThisApicomplexaprotein-designated tRip (tRNA import protein)-is anchored to the parasite plasma membrane and directs import of exogenous tRNAs. In the absence of tRip, the fitness of the parasite stage that multiplies in the blood is significantly reduced, indicating that the parasite may need host tRNAs to sustain its own translation and/or as regulatory RNAs.Plasmodiumis thus the first example, to our knowledge, of a cell importing exogenous tRNAs, suggesting a remarkable adaptation of this parasite to extend its reach into host cell biology.}, keywords = {FRUGIER, Plasmodium tRNA trafficking, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Display of whole proteins on inner and outer surfaces of Grapevine fanleaf virus-like particles.}, author = {L Belval and C Hemmer and C Sauter and C Reinbold and J D Fauny and F Berthold and L Ackerer and C Schmitt-Keichinger and O Lemaire and G Demangeat and C Ritzenthaler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/27178344?dopt=Abstract}, doi = {10.1111/pbi.12582}, isbn = {27178344}, year = {2016}, date = {2016-01-01}, journal = {Plant Biotechnol J}, volume = {14}, number = {12}, pages = {2288-2299}, abstract = {Virus-like particles (VLPs) derived from non-enveloped viruses result from the self-assembly of capsid proteins (CPs). They generally display similar structural features to viral particles but are non-infectious and their inner cavity and outer surface can potentially be adapted to serve as nanocarriers of great biotechnological interest. While a VLP outer surface is generally amenable to chemical or genetic modifications, encaging a cargo within particles can be more complex and is often limited to small molecules or peptides. Examples where both inner cavity and outer surface have been used to simultaneously encapsulate and expose entire proteins remain scarce. Here we describe the production of spherical VLPs exposing fluorescent proteins at either their outer surface or inner cavity as a result of the self-assembly of a single genetically modified viral structural protein, the CP of grapevine fanleaf virus (GFLV). We found that the N- and C-terminal ends of the GFLV CP allow the genetic fusion of proteins as large as 27 kDa and the plant-based production of nucleic-acid free VLPs. Remarkably, expression of N- or C-terminal CP fusions, resulted in the production of VLPs with recombinant proteins exposed either to the inner cavity or the outer surface, respectively, while coexpression of both fusion proteins led to the formation hybrid VLP, although rather inefficiently. Such properties are rather unique for a single viral structural protein and open new potential avenues for the design of safe and versatile nanocarriers, particularly for the targeted delivery of bioactive molecules.}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{flacher_mannoside_2015, title = {Mannoside Glycolipid Conjugates Display Anti-inflammatory Activity by Inhibition of Toll-like Receptor-4 Mediated Cell Activation}, author = {Vincent Flacher and Patrick Neuberg and Floriane Point and François Daubeuf and Quentin Muller and David Sigwalt and Jean-Daniel Fauny and Jean-Serge Remy and Nelly Frossard and Alain Wagner and Christopher G Mueller and Evelyne Schaeffer}, doi = {10.1021/acschembio.5b00552}, issn = {1554-8937}, year = {2015}, date = {2015-12-01}, journal = {ACS chemical biology}, volume = {10}, number = {12}, pages = {2697--2705}, abstract = {Inhibition of excessive Toll-like receptor 4 (TLR4) signaling is a therapeutic approach pursued for many inflammatory diseases. We report that Mannoside Glycolipid Conjugates (MGCs) selectively blocked TLR4-mediated activation of human monocytes and monocyte-derived dendritic cells (DCs) by lipopolysaccharide (LPS). They potently suppressed pro-inflammatory cytokine secretion and maturation of DCs exposed to LPS, leading to impaired T cell stimulation. MGCs did not interfere with LPS and could act in a delayed manner, hours after LPS stimulation. Their inhibitory action required both the sugar heads and the lipid chain, although the nature of the sugar and the structure of the lipid tail could be modified. They blocked early signaling events at the cell membrane, enhanced internalization of CD14 receptors, and prevented colocalization of CD14 and TLR4, thereby abolishing NF-κB nuclear translocation. When the best lead conjugate was tested in a mouse model of LPS-induced acute lung inflammation, it displayed an anti-inflammatory action by suppressing the recruitment of neutrophils. Thus, MGCs could serve as promising leads for the development of selective TLR4 antagonistic agents for inflammatory diseases.}, keywords = {I2CT, Imagerie, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{flacher_mannoside_2015b, title = {Mannoside Glycolipid Conjugates Display Anti-inflammatory Activity by Inhibition of Toll-like Receptor-4 Mediated Cell Activation}, author = {Vincent Flacher and Patrick Neuberg and Floriane Point and François Daubeuf and Quentin Muller and David Sigwalt and Jean-Daniel Fauny and Jean-Serge Remy and Nelly Frossard and Alain Wagner and Christopher G Mueller and Evelyne Schaeffer}, doi = {10.1021/acschembio.5b00552}, issn = {1554-8937}, year = {2015}, date = {2015-12-01}, journal = {ACS chemical biology}, volume = {10}, number = {12}, pages = {2697--2705}, abstract = {Inhibition of excessive Toll-like receptor 4 (TLR4) signaling is a therapeutic approach pursued for many inflammatory diseases. We report that Mannoside Glycolipid Conjugates (MGCs) selectively blocked TLR4-mediated activation of human monocytes and monocyte-derived dendritic cells (DCs) by lipopolysaccharide (LPS). They potently suppressed pro-inflammatory cytokine secretion and maturation of DCs exposed to LPS, leading to impaired T cell stimulation. MGCs did not interfere with LPS and could act in a delayed manner, hours after LPS stimulation. Their inhibitory action required both the sugar heads and the lipid chain, although the nature of the sugar and the structure of the lipid tail could be modified. They blocked early signaling events at the cell membrane, enhanced internalization of CD14 receptors, and prevented colocalization of CD14 and TLR4, thereby abolishing NF-κB nuclear translocation. When the best lead conjugate was tested in a mouse model of LPS-induced acute lung inflammation, it displayed an anti-inflammatory action by suppressing the recruitment of neutrophils. Thus, MGCs could serve as promising leads for the development of selective TLR4 antagonistic agents for inflammatory diseases.}, keywords = {Activation, Animals, Anti-Inflammatory Agents, Carbohydrate Sequence, CD14, Cell Membrane, Cells, Chemistry, Cultured, cytokine, Dendritic Cells, development, disease, Glycolipids, Human, Humans, immunopathology, Inbred BALB C, inflammation, inhibition, lipid, lipopolysaccharide, Lipopolysaccharides, LPS, LUNG, Mannosides, Maturation, Membrane, Mice, monocyte, Monocytes, mouse, neutrophils, NF-kappaB, Pneumonia, Protein-Serine-Threonine Kinases, Receptor, secretion, signaling, Structure-Activity Relationship, Tail, Team-Mueller, TLR4, Toll-Like Receptor 4}, pubstate = {published}, tppubtype = {article} } @article{mairhofer_impaired_2015, title = {Impaired gp100-Specific CD8(+) Ŧ-Cell Responses in the Presence of Myeloid-Derived Suppressor Cells in a Spontaneous Mouse Melanoma Model}, author = {David G Mairhofer and Daniela Ortner and Christoph H Tripp and Sandra Schaffenrath and Viktor Fleming and Lukas Heger and Kerstin Komenda and Daniela Reider and Diana Dudziak and Suzie Chen and Jürgen C Becker and Vincent Flacher and Patrizia Stoitzner}, doi = {10.1038/jid.2015.241}, issn = {1523-1747}, year = {2015}, date = {2015-11-01}, journal = {The Journal of Investigative Dermatology}, volume = {135}, number = {11}, pages = {2785--2793}, abstract = {Murine tumor models that closely reflect human diseases are important tools to investigate carcinogenesis and tumor immunity. The transgenic (tg) mouse strain tg(Grm1)EPv develops spontaneous melanoma due to ectopic overexpression of the metabotropic glutamate receptor 1 (Grm1) in melanocytes. In the present study, we characterized the immune status and functional properties of immune cells in tumor-bearing mice. Melanoma development was accompanied by a reduction in the percentages of CD4(+) T cells including regulatory T cells (Tregs) in CD45(+) leukocytes present in tumor tissue and draining lymph nodes (LNs). In contrast, the percentages of CD8(+) T cells were unchanged, and these cells showed an activated phenotype in tumor mice. Endogenous melanoma-associated antigen glycoprotein 100 (gp100)-specific CD8(+) T cells were not deleted during tumor development, as revealed by pentamer staining in the skin and draining LNs. They, however, were unresponsive to ex vivo gp100-peptide stimulation in late-stage tumor mice. Interestingly, immunosuppressive myeloid-derived suppressor cells (MDSCs) were recruited to tumor tissue with a preferential accumulation of granulocytic MDSC (grMDSCs) over monocytic MDSC (moMDSCs). Both subsets produced Arginase-1, inducible nitric oxide synthase (iNOS), and transforming growth factor-β and suppressed T-cell proliferation in vitro. In this work, we describe the immune status of a spontaneous melanoma mouse model that provides an interesting tool to develop future immunotherapeutical strategies.}, keywords = {Analysis of Variance, Animal, Animals, Antigen, cancer, CARCINOGENESIS, CD8-Positive T-Lymphocytes, Cell Proliferation, Cultured, DERMATOLOGY, development, disease, Disease Models, Experimental, GLYCOPROTEIN, gp100 Melanoma Antigen, Growth, Human, Humans, Immunity, Immunologic, IN VITRO, Inbred C57BL, iNOS, Leukocytes, LYMPH, LYMPH NODE, Lymph Nodes, Lymphocyte Activation, MELANOCYTES, Melanoma, Mice, mouse, murine, NITRIC OXIDE, nitric oxide synthase, Phenotype, Proliferation, Random Allocation, Receptor, Regulatory, RESPONSES, Skin, SUBSETS, Suppressor Factors, T CELLS, T-CELLS, T-Lymphocytes, Team-Mueller, Transforming Growth Factor beta, transgenic, tumor, Tumor Cells, tumor immunity}, pubstate = {published}, tppubtype = {article} } @article{haid_langerhans_2015, title = {Langerhans cells in the sebaceous gland of the murine skin}, author = {Bernhard Haid and David E Schlögl and Martin Hermann and Christoph H Tripp and Patrizia Stoitzner and Nikolaus Romani and Vincent Flacher}, doi = {10.1111/exd.12803}, issn = {1600-0625}, year = {2015}, date = {2015-11-01}, journal = {Experimental Dermatology}, volume = {24}, number = {11}, pages = {899--901}, keywords = {Animals, Dendritic Cells, DERMATOLOGY, DERMIS, Epidermis, Inbred BALB C, Inbred C57BL, Langerhans Cells, Langerin, Letter, Mice, murine, pilosebaceous unit, sebaceous gland, Sebaceous Glands, Skin, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{veillard_drosophila_2015, title = {Drosophila melanogaster clip-domain serine proteases: Structure, function and regulation}, author = {Florian Veillard and Laurent Troxler and Jean-Marc Reichhart}, url = {http://www.sciencedirect.com/science/article/pii/S030090841500317X}, doi = {10.1016/j.biochi.2015.10.007}, isbn = {0300-9084}, year = {2015}, date = {2015-10-08}, journal = {Biochimie}, volume = {122}, pages = {255-269}, abstract = {Mammalian chymotrypsin-like serine proteases (SPs) are one of the best-studied family of enzymes with roles in a wide range of physiological processes, including digestion, blood coagulation, fibrinolysis and humoral immunity. Extracellular SPs can form cascades, in which one protease activates the zymogen of the next protease in the chain, to amplify physiological or pathological signals. These extracellular SPs are generally multi-domain proteins, with pro-domains that are involved in protein–protein interactions critical for the sequential organization of the cascades, the control of their intensity and their proper localization. Far less is known about invertebrate SPs than their mammalian counterparts. In insect genomes, SPs and their proteolytically inactive homologs (SPHs) constitute large protein families. In addition to the chymotrypsin fold, many of these proteins contain additional structural domains, often with conserved mammalian orthologues. However, the largest group of arthropod SP regulatory modules is the clip domains family, which has only been identified in arthropods. The clip-domain SPs are extracellular and have roles in the immune response and embryonic development. The powerful reverse-genetics tools in Drosophila melanogaster have been essential to identify the functions of clip-SPs and their organization in sequential cascades. This review focuses on the current knowledge of Drosophila clip-SPs and presents, when necessary, data obtained in other insect models. We will first cover the biochemical and structural features of clip domain SPs and SPHs. Clip-SPs are implicated in three main biological processes: the control of the dorso-ventral patterning during embryonic development; the activation of the Toll-mediated response to microbial infections and the prophenoloxydase cascade, which triggers melanization. Finally, we review the regulation of SPs and SPHs, from specificity of activation to inhibition by endogenous or pathogen-encoded inhibitors.}, keywords = {bioinformatic, Chymotrypsin, clip domain, Immunity, Insect, M3i, Melanization, reichhart, Serine Proteases, Serpin, Toll}, pubstate = {published}, tppubtype = {article} } @article{J2015, title = {A New Role of the Mosquito Complement-like Cascade in Male Fertility in Anopheles gambiae}, author = {Julien Pompon and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26394016}, year = {2015}, date = {2015-09-22}, journal = {PLoS Biology}, volume = {13}, number = {9}, pages = {e1002255}, abstract = {Thioester-containing protein 1 (TEP1) is a key immune factor that determines mosquito resistance to a wide range of pathogens, including malaria parasites. Here we report a new allele-specific function of TEP1 in male fertility. We demonstrate that during spermatogenesis TEP1 binds to and removes damaged cells through the same complement-like cascade that kills malaria parasites in the mosquito midgut. Further, higher fertility rates are mediated by an allele that renders the mosquito susceptible to Plasmodium. By elucidating the molecular and genetic mechanisms underlying TEP1 function in spermatogenesis, our study suggests that pleiotropic antagonism between reproduction and immunity may shape resistance of mosquito populations to malaria parasites.}, keywords = {Anopheles gambiae, TEP1}, pubstate = {published}, tppubtype = {article} } @article{el_chamy_multilayered_2015, title = {The multilayered innate immune defense of the gut}, author = {Laure El Chamy and Nicolas Matt and Monde Ntwasa and Jean-Marc Reichhart}, url = {http://www.biomedj.org/text.asp?2015/38/4/276/158621}, doi = {10.4103/2319-4170.158621}, issn = {2320-2890}, year = {2015}, date = {2015-08-01}, journal = {Biomed J}, volume = {38}, number = {4}, pages = {276--284}, abstract = {In the wild, the fruit fly Drosophila melanogaster thrives on rotten fruit. The digestive tract maintains a powerful gut immune barrier to regulate the ingested microbiota, including entomopathogenic bacteria. This gut immune barrier includes a chitinous peritrophic matrix that isolates the gut contents from the epithelial cells. In addition, the epithelial cells are tightly sealed by septate junctions and can mount an inducible immune response. This local response can be activated by invasive bacteria, or triggered by commensal bacteria in the gut lumen. As with chronic inflammation in mammals, constitutive activation of the gut innate immune response is detrimental to the health of flies. Accordingly, the Drosophila gut innate immune response is tightly regulated to maintain the endogenous microbiota, while preventing infections by pathogenic microorganisms.}, keywords = {fly, gut, innate immunity, M3i, matt, reichhart}, pubstate = {published}, tppubtype = {article} } @article{aguiar_sequence-independent_2015, title = {Sequence-independent characterization of viruses based on the pattern of viral small RNAs produced by the host}, author = {Eric Roberto Guimarães Rocha Aguiar and Roenick Proveti Olmo and Simona Paro and Flavia Viana Ferreira and Isaque João Silva da de Faria and Yaovi Mathias Honore Todjro and Francisco Pereira Lobo and Erna Geessien Kroon and Carine Meignin and Derek Gatherer and Jean-Luc Imler and João Trindade Marques}, url = {http://nar.oxfordjournals.org/lookup/doi/10.1093/nar/gkv587}, doi = {10.1093/nar/gkv587}, issn = {1362-4962}, year = {2015}, date = {2015-07-01}, journal = {Nucleic Acids Research}, volume = {43}, number = {13}, pages = {6191--6206}, abstract = {Virus surveillance in vector insects is potentially of great benefit to public health. Large-scale sequencing of small and long RNAs has previously been used to detect viruses, but without any formal comparison of different strategies. Furthermore, the identification of viral sequences largely depends on similarity searches against reference databases. Here, we developed a sequence-independent strategy based on virus-derived small RNAs produced by the host response, such as the RNA interference pathway. In insects, we compared sequences of small and long RNAs, demonstrating that viral sequences are enriched in the small RNA fraction. We also noted that the small RNA size profile is a unique signature for each virus and can be used to identify novel viral sequences without known relatives in reference databases. Using this strategy, we characterized six novel viruses in the viromes of laboratory fruit flies and wild populations of two insect vectors: mosquitoes and sandflies. We also show that the small RNA profile could be used to infer viral tropism for ovaries among other aspects of virus biology. Additionally, our results suggest that virus detection utilizing small RNAs can also be applied to vertebrates, although not as efficiently as to plants and insects.}, keywords = {Animals, Contig Mapping, Female, imler, insects, M3i, meignin, Ovary, Plants, RNA, Sequence Analysis, Small Untranslated, Vertebrates, Viral, Viral Tropism, viruses}, pubstate = {published}, tppubtype = {article} } @article{pmid26225566, title = {An In-Depth Comparison of Latency-Reversing Agent Combinations in Various In Vitro and Ex Vivo HIV-1 Latency Models Identified Bryostatin-1+JQ1 and Ingenol-B+JQ1 to Potently Reactivate Viral Gene Expression}, author = {Gilles Darcis and Anna Kula and Sophie Bouchat and Koh Fujinaga and Francis Corazza and Amina Ait-Ammar and Nadège Delacourt and Adeline Melard and Kabamba Kabeya and Caroline Vanhulle and Benoit Van Driessche and Jean-Stéphane Gatot and Thomas Cherrier and Luiz F Pianowski and Lucio Gama and Christian Schwartz and Jorge Vila and Arsène Burny and Nathan Clumeck and Michel Moutschen and Stéphane De Wit and B Matija Peterlin and Christine Rouzioux and Olivier Rohr and Carine Van Lint}, doi = {10.1371/journal.ppat.1005063}, issn = {1553-7374}, year = {2015}, date = {2015-07-01}, urldate = {2015-07-01}, journal = {PLoS Pathog}, volume = {11}, number = {7}, pages = {e1005063}, abstract = {The persistence of latently infected cells in patients under combinatory antiretroviral therapy (cART) is a major hurdle to HIV-1 eradication. Strategies to purge these reservoirs are needed and activation of viral gene expression in latently infected cells is one promising strategy. Bromodomain and Extraterminal (BET) bromodomain inhibitors (BETi) are compounds able to reactivate latent proviruses in a positive transcription elongation factor b (P-TEFb)-dependent manner. In this study, we tested the reactivation potential of protein kinase C (PKC) agonists (prostratin, bryostatin-1 and ingenol-B), which are known to activate NF-κB signaling pathway as well as P-TEFb, used alone or in combination with P-TEFb-releasing agents (HMBA and BETi (JQ1, I-BET, I-BET151)). Using in vitro HIV-1 post-integration latency model cell lines of T-lymphoid and myeloid lineages, we demonstrated that PKC agonists and P-TEFb-releasing agents alone acted as potent latency-reversing agents (LRAs) and that their combinations led to synergistic activation of HIV-1 expression at the viral mRNA and protein levels. Mechanistically, combined treatments led to higher activations of P-TEFb and NF-κB than the corresponding individual drug treatments. Importantly, we observed in ex vivo cultures of CD8+-depleted PBMCs from 35 cART-treated HIV-1+ aviremic patients that the percentage of reactivated cultures following combinatory bryostatin-1+JQ1 treatment was identical to the percentage observed with anti-CD3+anti-CD28 antibodies positive control stimulation. Remarkably, in ex vivo cultures of resting CD4+ T cells isolated from 15 HIV-1+ cART-treated aviremic patients, the combinations bryostatin-1+JQ1 and ingenol-B+JQ1 released infectious viruses to levels similar to that obtained with the positive control stimulation. The potent effects of these two combination treatments were already detected 24 hours post-stimulation. These results constitute the first demonstration of LRA combinations exhibiting such a potent effect and represent a proof-of-concept for the co-administration of two different types of LRAs as a potential strategy to reduce the size of the latent HIV-1 reservoirs.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{schaeffer_dermal_2015b, title = {Dermal CD14(+) Dendritic Cell and Macrophage Infection by Dengue Virus Is Stimulated by Interleukin-4}, author = {Evelyne Schaeffer and Vincent Flacher and Vasiliki Papageorgiou and Marion Decossas and Jean-Daniel Fauny and Melanie Krämer and Christopher G Mueller}, doi = {10.1038/jid.2014.525}, issn = {1523-1747}, year = {2015}, date = {2015-07-01}, journal = {The Journal of Investigative Dermatology}, volume = {135}, number = {7}, pages = {1743--1751}, abstract = {Dengue virus (DENV) is responsible for the most prevalent arthropod-borne viral infection in humans. Events decisive for disease development occur in the skin after virus inoculation by the mosquito. Yet, the role of human dermis-resident immune cells in dengue infection and disease remains elusive. Here we investigated how dermal dendritic cells (dDCs) and macrophages (dMs) react to DENV and impact on immunopathology. We show that both CD1c(+) and CD14(+) dDC subsets were infected, but viral load greatly increased in CD14(+) dDCs upon IL-4 stimulation, which correlated with upregulation of virus-binding lectins Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Nonintegrin (DC-SIGN/CD209) and mannose receptor (CD206). IL-4 also enhanced T-cell activation by dDCs, which was further increased upon dengue infection. dMs purified from digested dermis were initially poorly infected but actively replicated the virus and produced TNF-α upon lectin upregulation in response to IL-4. DC-SIGN(+) cells are abundant in inflammatory skin with scabies infection or Th2-type dermatitis, suggesting that skin reactions to mosquito bites heighten the risk of infection and subsequent immunopathology. Our data identify dDCs and dMs as primary arbovirus target cells in humans and suggest that dDCs initiate a potent virus-directed T-cell response, whereas dMs fuel the inflammatory cascade characteristic of dengue fever.}, keywords = {Abdominal Wall, Activation, Adhesion, adhesion molecules, Antigen-Presenting Cells, arbovirus, C-Type, Cell Adhesion, Cell Adhesion Molecules, Cell Surface, Cells, Chemistry, Confocal, Cultured, cytokine, Cytokines, cytology, Dendritic Cells, Dengue, Dengue virus, DERMAL DENDRITIC CELLS, Dermatitis, DERMIS, development, disease, Enzyme-Linked Immunosorbent Assay, Epidermal Cells, Epidermis, Human, Humans, ICAM-3, IL-4, Immunology, immunopathology, infection, Interleukin-4, Langerhans Cells, LECTIN, Lectins, Lymphocyte Activation, Macrophage, Macrophages, metabolism, Microscopy, pathogenicity, physiopathology, Receptor, Receptors, Scabies, Sensitivity and Specificity, Skin, Skin Diseases, SUBSETS, T CELL ACTIVATION, target, Team-Mueller, TNF ALPHA, Viral, viral Infection, Viral Load, virology, virus}, pubstate = {published}, tppubtype = {article} } @article{bianco_safety_2015, title = {Safety concerns on graphene and 2D materials: a Flagship perspective}, author = {Alberto Bianco and Maurizio Prato}, url = {https://doi.org/10.1088%2F2053-1583%2F2%2F3%2F030201}, doi = {10.1088/2053-1583/2/3/030201}, issn = {2053-1583}, year = {2015}, date = {2015-06-01}, urldate = {2020-04-01}, journal = {2D Materials}, volume = {2}, number = {3}, pages = {030201}, abstract = {This is a specially commissioned editorial from the Graphene Flagship Work Package on Health and Environment. This editorial is part of the 2D Materials focus collection on ‘Progress on the science and applications of two-dimensional materials’, published in association with the Graphene Flagship. It provides an overview of key, recent advances from the ‘Health and Environment’ work package and is not intended as a comprehensive review of this field.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @inbook{Majzoub2015, title = {Encyclopedia of Molecular Cell Biology and Molecular Medicine}, author = {Karim Majzoub and Jean-Luc Imler}, editor = {Wiley-VCH Verlag}, doi = {10.1002/3527600906.mcb.201500003}, year = {2015}, date = {2015-04-28}, volume = {1}, pages = {192-228}, publisher = {GmbH & Co. KGaA}, chapter = {« RNAi to treat virus infections »}, abstract = {In spite of its young age, the field of RNA interference has already yielded major advances in the laboratory. This sequence-specific mechanism of gene regulation also holds strong promise for the development of a new generation of drugs, in particular to control the everlasting threat of viral infections. Here, the mechanisms and pathways of RNA interference are reviewed, with emphasis placed on how RNA silencing forms a potent antiviral immune mechanism in plants and invertebrates. The approaches developed to use RNA interference to control viral infections in mammals are then described. Finally, the problems encountered while translating this revolutionary technology into the clinic are presented, and the advances currently developed to overcome these limitations are discussed.}, keywords = {antiviral, Argonaute, Delivery, imler, Immunity, lipofection, M3i, microRNA (miRNA), RNA Virus Infections, RNAi, small hairpin RNA (shRNA), small interfering RNA (siRNA)}, pubstate = {published}, tppubtype = {inbook} } @article{G2015, title = {Tools for Anopheles gambiae Transgenesis}, author = {Gloria Volohonsky and Olivier Terenzi and Julien Soichot and Daniel A Naujoks and Tony Nolan and Nikolai Windbichler and Delphine Kapps and Andie L Smidler and Anaïs Vittu and Giulia Costa and Stefanie Steinert and Elena A Levashina and Stéphanie A Blandin and Eric Marois}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25869647}, year = {2015}, date = {2015-04-13}, journal = {G3 (Bethesda)}, volume = {5}, number = {6}, pages = {1151-63}, abstract = {Transgenesis is an essential tool to investigate gene function and to introduce desired characters in laboratory organisms. Setting-up transgenesis in non-model organisms is challenging due to the diversity of biological life traits and due to knowledge gaps in genomic information. Some procedures will be broadly applicable to many organisms, and others have to be specifically developed for the target species. Transgenesis in disease vector mosquitoes has existed since the 2000s but has remained limited by the delicate biology of these insects. Here, we report a compilation of the transgenesis tools that we have designed for the malaria vector Anopheles gambiae, including new docking strains, convenient transgenesis plasmids, a puromycin resistance selection marker, mosquitoes expressing cre recombinase, and various reporter lines defining the activity of cloned promoters. This toolbox contributed to rendering transgenesis routine in this species and is now enabling the development of increasingly refined genetic manipulations such as targeted mutagenesis. Some of the reagents and procedures reported here are easily transferable to other nonmodel species, including other disease vector or agricultural pest insects.}, keywords = {Anopheles gambiae, bioinformatic, blandin, M3i, marois, transgenesis}, pubstate = {published}, tppubtype = {article} } @article{paro_sensing_2015, title = {Sensing viral RNAs by Dicer/RIG-I like ATPases across species}, author = {Simona Paro and Jean-Luc Imler and Carine Meignin}, url = {http://linkinghub.elsevier.com/retrieve/pii/S0952791515000102}, doi = {10.1016/j.coi.2015.01.009}, issn = {1879-0372}, year = {2015}, date = {2015-02-01}, journal = {Current Opinion in Immunology}, volume = {32}, pages = {106--113}, abstract = {Induction of antiviral immunity in vertebrates and invertebrates relies on members of the RIG-I-like receptor and Dicer families, respectively. Although these proteins have different size and domain composition, members of both families share a conserved DECH-box helicase domain. This helicase, also known as a duplex RNA activated ATPase, or DRA domain, plays an important role in viral RNA sensing. Crystallographic and electron microscopy studies of the RIG-I and Dicer DRA domains indicate a common structure and that similar conformational changes are induced by dsRNA binding. Genetic and biochemical studies on the function and regulation of DRAs reveal similarities, but also some differences, between viral RNA sensing mechanisms in nematodes, flies and mammals.}, keywords = {Adenosine Triphosphatases, Animals, DEAD-box RNA Helicases, Humans, imler, M3i, meignin, Protein Binding, Protein Interaction Domains and Motifs, Ribonuclease III, RNA, Viral, Virus Diseases, viruses}, pubstate = {published}, tppubtype = {article} } @article{DE2015, title = {Highly evolvable malaria vectors: The genomes of 16 Anopheles mosquitoes.}, author = {D E Neafsey and R M Waterhouse and M R Abai and S S Aganezov and M A Alekseyev and J E Allen and J Amon and B Arcà and P Arensburger and G Artemov and L A Assour and H Basseri and A Berlin and B W Birren and Stéphanie A Blandin and A I Brockman and T R Burkot and A Burt and C S Chan and C Chauve and J C Chiu and M Christensen and C Costantini and V L M Davidson and E Deligianni and T Dottorini and V Dritsou and S B Gabriel and W M Guelbeogo and A B Hall and M W Han and T Hlaing and D S T Hughes and A M Jenkins and X Jiang and I Jungreis and E G Kakani and M Kamali and P Kemppainen and R C Kennedy and I K Kirmitzoglou and L L Koekemoer and N Laban and N Langridge and M K N Lawniczak and M Lirakis and N F Lobo and E Lowy and R M MacCallum and C Mao and G Maslen and C Mbogo and J McCarthy and K Michel and S N Mitchell and W Moore and K A Murphy and A N Naumenko and Tony Nolan and E M Novoa and S O’Loughlin and C Oringanje and M A Oshaghi and N Pakpour and P A Papathanos and A N Peery and Michael Povelones and A Prakash and D A Price and A Rajaraman and L J Reimer and D C Rinker and A Rokas and T L Russell and N F Sagnon and M V Sharakhova and T Shea and F A Simão and F Simard and M A Slotman and P Somboon and V Stegniy and C J Struchiner and G W C Thomas and M Tojo and P Topalis and J M C Tubio and M F Unger and J Vontas and C Walton and C S Wilding and J H Willis and Y-C Wu and G Yan and E M Zdobnov and X Zhou and Flaminia Catteruccia and Georges K Christophides and F H Collins and R S Cornman and Andrea Crisanti and M J Donnelly and S J Emrich and M C Fontaine and W Gelbart and M W Hahn and I A Hansen and P I Howell and Fotis C Kafatos and M Kellis and D Lawson and C Louis and S Luckhart and M A T Muskavitch and J M Ribeiro and M A Riehle and I V Sharakhov and Z Tu and L J Zwiebel and N J Besansky}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25554792}, year = {2015}, date = {2015-01-02}, journal = {Science}, volume = {347}, number = {6217}, pages = {1258522}, abstract = {Variation in vectorial capacity for human malaria among Anopheles mosquito species is determined by many factors, including behavior, immunity, and life history. To investigate the genomic basis of vectorial capacity and explore new avenues for vector control, we sequenced the genomes of 16 anopheline mosquito species from diverse locations spanning ~100 million years of evolution. Comparative analyses show faster rates of gene gain and loss, elevated gene shuffling on the X chromosome, and more intron losses, relative to Drosophila. Some determinants of vectorial capacity, such as chemosensory genes, do not show elevated turnover but instead diversify through protein-sequence changes. This dynamism of anopheline genes and genomes may contribute to their flexible capacity to take advantage of new ecological niches, including adapting to humans as primary hosts.}, keywords = {Anopheles, blandin, genomes, M3i}, pubstate = {published}, tppubtype = {article} } @article{, title = {Modulation of aminoacylation and editing properties of leucyl-tRNA synthetase by a conserved structural module.}, author = {W Yan and Q Ye and M Tan and X Chen and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25817995?dopt=Abstract}, doi = {10.1074/jbc.M115.639492}, isbn = {25817995}, year = {2015}, date = {2015-01-01}, journal = {J Biol Chem}, volume = {290}, number = {19}, pages = {12256-12267}, abstract = {A conserved structural module following the KMSKS catalytic loop exhibits α-α-β-α topology in class Ia and Ib aminoacyl-tRNA synthetases. However, the function of this domain has received little attention. Here, we describe the effect this module has on the aminoacylation and editing capacities of leucyl-tRNA synthetases (LeuRSs) by characterizing the key residues from various species. Mutation of highly conserved basic residues on the third α-helix of this domain impairs the affinity of LeuRS for the anticodon stem of tRNALeu, which decreases both aminoacylation and editing activities. Two glycine residues on this α-helix contribute to flexibility, leucine activation and editing of LeuRS from Escherichia coli (EcLeuRS). Acidic residues on the β-strand enhance the editing activity of EcLeuRS and sense the size of the tRNALeu D-loop. Incorporation of these residues stimulates the tRNA-dependent editing activity of the chimeric minimalist enzyme MmLeuRS fused with the connective polypeptide 1 (CP1) editing domain and leucine-specific domain of (LSD) from EcLeuRS. Together, these results reveal the stem contact (SC)-fold to be functional as well as a structural linker between the catalytic site and tRNA binding domain. Sequence comparison of the EcLeuRS SC-fold domain with editing-deficient enzymes suggests that key residues of this module have evolved an adaptive strategy to follow the editing functions of LeuRSs.}, keywords = {aminoacyl tRNA synthetase aminoacylation editing enzyme evolution protein synthesis stem contact fold transfer RNA (tRNA), ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Activity-Fed Translation (AFT) Assay: A New High-Throughput Screening Strategy for Enzymes in Droplets.}, author = {G Woronoff and M Ryckelynck and J Wessel and O Schicke and A Griffiths and Soumillion P.}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25914325?dopt=Abstract}, doi = {10.1002/cbic.201500087}, year = {2015}, date = {2015-01-01}, journal = {Chembiochem}, volume = {16}, number = {9}, pages = {1343-1349}, abstract = {There is an increasing demand for the development of sensitive enzymatic assays compatible with droplet-based microfluidics. Here we describe an original strategy, activity-fed translation (AFT), based on the coupling of enzymatic activity to in vitro translation of a fluorescent protein. We show that methionine release upon the hydrolysis of phenylacetylmethionine by penicillin acylase enabled in vitro expression of green fluorescent protein. An autocatalytic setup where both proteins are expressed makes the assay highly sensitive, as fluorescence was detected in droplets containing single PAC genes. Adding a PCR step in the droplets prior to the assay increased the sensitivity further. The strategy is potentially applicable for any activity that can be coupled to the production of an amino acid, and as the microdroplet volume is small the use of costly reagents such as in vitro expression mixtures is not limiting for high-throughput screening projects}, keywords = {droplet-based microfluidics enzyme catalysis in vitro translation penicillin acylase protein expression, RYCKELYNCK, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA structure from deep sequencing.}, author = {E Westhof}, url = {http://www.nature.com/nbt/journal/v33/n9/full/nbt.3338.html}, doi = {10.1038/nbt.3338}, isbn = {26348961}, year = {2015}, date = {2015-01-01}, journal = {Nat Biotechnol}, volume = {33}, number = {9}, pages = {928-929}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Twenty years of RNA crystallography.}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25780106?dopt=Abstract}, doi = {10.1261/rna.049726.115}, isbn = {25780106}, year = {2015}, date = {2015-01-01}, journal = {RNA}, volume = {21}, number = {4}, pages = {486-487}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{brunke_mice_2015b, title = {Of mice, flies--and men? Comparing fungal infection models for large-scale screening efforts}, author = {Sascha Brunke and Jessica Quintin and Lydia Kasper and Ilse D Jacobsen and Martin E Richter and Ekkehard Hiller and Tobias Schwarzmüller and Christophe d'Enfert and Karl Kuchler and Steffen Rupp and Bernhard Hube and Dominique Ferrandon}, url = {http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4415897/}, doi = {10.1242/dmm.019901}, issn = {1754-8411}, year = {2015}, date = {2015-01-01}, journal = {Dis Model Mech}, volume = {8}, number = {5}, pages = {473--486}, abstract = {Studying infectious diseases requires suitable hosts for experimental in vivo infections. Recent years have seen the advent of many alternatives to murine infection models. However, the use of non-mammalian models is still controversial because it is often unclear how well findings from these systems predict virulence potential in humans or other mammals. Here, we compare the commonly used models, fruit fly and mouse (representing invertebrate and mammalian hosts), for their similarities and degree of correlation upon infection with a library of mutants of an important fungal pathogen, the yeast Candida glabrata. Using two indices, for fly survival time and for mouse fungal burden in specific organs, we show a good agreement between the models. We provide a suitable predictive model for estimating the virulence potential of C. glabrata mutants in the mouse from fly survival data. As examples, we found cell wall integrity mutants attenuated in flies, and mutants of a MAP kinase pathway had defective virulence in flies and reduced relative pathogen fitness in mice. In addition, mutants with strongly reduced in vitro growth generally, but not always, had reduced virulence in flies. Overall, we demonstrate that surveying Drosophila survival after infection is a suitable model to predict the outcome of murine infections, especially for severely attenuated C. glabrata mutants. Pre-screening of mutants in an invertebrate Drosophila model can, thus, provide a good estimate of the probability of finding a strain with reduced microbial burden in the mouse host.}, keywords = {Alternative infection models, Candida glabrata, ferrandon, Fungal virulence factors, M3i, Mutant library, Signature-tagged mutagenesis}, pubstate = {published}, tppubtype = {article} } @article{, title = {Small RNAs in Bacteria and Archaea: Who They Are, What They Do, and How They Do It.}, author = {E G Wagner and P Romby}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26296935?dopt=Abstract}, doi = {10.1016/bs.adgen.2015.05.001}, isbn = {26296935}, year = {2015}, date = {2015-01-01}, journal = {Adv Genet}, volume = {90}, pages = {133-208}, abstract = {Small RNAs are ubiquitously present regulators in all kingdoms of life. Most bacterial and archaeal small RNAs (sRNAs) act by antisense mechanisms on multiple target mRNAs, thereby globally affecting essentially any conceivable trait-stress responses, adaptive metabolic changes, virulence etc. The sRNAs display many distinct mechanisms of action, most of them through effects on target mRNA translation and/or stability, and helper proteins like Hfq often play key roles. Recent data highlight the interplay between posttranscriptional control by sRNAs and transcription factor-mediated transcriptional control, and cross talk through mutual regulation of regulators. Based on the properties that distinguish sRNA-type from transcription factors-type control, we begin to glimpse why sRNAs have evolved as a second, essential layer of gene regulation. This review will discuss the prevalence of sRNAs, who they are, what biological roles they play, and how they carry out their functions.}, keywords = {Antisense RNA Hfq Interconnected networks Posttranscriptional regulation RNA sponges Regulatory RNA, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{hafirassou_[fly_2015b, title = {[From fly to man: RACK1, an essential actor of the dependent viral translation of IRES].}, author = {Lamine Mohamed Hafirassou and Carine Meignin and Thomas Baumert and Catherine Schuster}, year = {2015}, date = {2015-01-01}, journal = {Médecine sciences : M/S}, volume = {31}, number = {5}, pages = {469--472}, keywords = {meignin}, pubstate = {published}, tppubtype = {article} } @article{, title = {Evaluation of anti-HIV-1 mutagenic nucleoside analogues.}, author = {V Vivet-Boudou and C Isel and Y El Safadi and R P Smyth and G Laumond and C Moog and J C Paillart and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25398876?dopt=Abstract}, isbn = {25398876}, year = {2015}, date = {2015-01-01}, journal = {J Biol Chem}, volume = {290}, number = {1}, pages = {371-383}, abstract = {Because of their high mutation rates, RNA viruses and retroviruses replicate close to the threshold of viability. Their existence as quasi-species has pioneered the concept of "lethal mutagenesis" that prompted us to synthesize pyrimidine nucleoside analogues with antiviral activity in cell culture consistent with an accumulation of deleterious mutations in the HIV-1 genome. However, testing all potentially mutagenic compounds in cell-based assays is tedious and costly. Here, we describe two simple in vitro biophysical/biochemical assays that allow prediction of the mutagenic potential of deoxyribonucleoside analogues. The first assay compares the thermal stabilities of matched and mismatched base pairs in DNA duplexes containing or not the nucleoside analogues as follows. A promising candidate should display a small destabilization of the matched base pair compared with the natural nucleoside and the smallest gap possible between the stabilities of the matched and mismatched base pairs. From this assay, we predicted that two of our compounds, 5-hydroxymethyl-2'-deoxyuridine and 5-hydroxymethyl-2'-deoxycytidine, should be mutagenic. The second in vitro reverse transcription assay assesses DNA synthesis opposite nucleoside analogues inserted into a template strand and subsequent extension of the newly synthesized base pairs. Once again, only 5-hydroxymethyl-2'-deoxyuridine and 5-hydroxymethyl-2'-deoxycytidine are predicted to be efficient mutagens. The predictive potential of our fast and easy first line screens was confirmed by detailed analysis of the mutation spectrum induced by the compounds in cell culture because only compounds 5-hydroxymethyl-2'-deoxyuridine and 5-hydroxymethyl-2'-deoxycytidine were found to increase the mutation frequency by 3.1- and 3.4-fold, respectively.}, keywords = {antiviral agent human immunodeficiency virus (HIV) lethal mutagenesis nucleoside/nucleotide analogue reverse transcription, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{hammond_crispr-cas9_2015, title = {A CRISPR-Cas9 gene drive system targeting female reproduction in the malaria mosquito vector Anopheles gambiae}, author = {Andrew Hammond and Roberto Galizi and Kyros Kyrou and Alekos Simoni and Carla Siniscalchi and Dimitris Katsanos and Matthew Gribble and Dean Baker and Eric Marois and Steven Russell and Austin Burt and Nikolai Windbichler and Andrea Crisanti and Tony Nolan}, url = {http://www.nature.com/doifinder/10.1038/nbt.3439}, doi = {10.1038/nbt.3439}, issn = {1087-0156, 1546-1696}, year = {2015}, date = {2015-01-01}, urldate = {2016-01-26}, journal = {Nature Biotechnology}, volume = {34}, number = {1}, pages = {78--83}, keywords = {M3i, marois}, pubstate = {published}, tppubtype = {article} } @article{, title = {Spot peptide arrays and SPR measurements: throughput and quantification in antibody selectivity studies.}, author = {T Vernet and L Choulier and Y Nominé and L Bellard and M Baltzinger and G Travé and D Altschuh}, url = {https://www.ncbi.nlm.nih.gov/pubmed/25960426}, doi = {10.1002/jmr.2477}, isbn = {25960426}, year = {2015}, date = {2015-01-01}, journal = {J Mol Recognit}, volume = {28}, number = {10}, pages = {635-644}, abstract = {Antibody selectivity represents a major issue in the development of efficient immuno-therapeutics and detection assays. Its description requires a comparison of the affinities of the antibody for a significant number of antigen variants. In the case of peptide antigens, this task can now be addressed to a significant level of details owing to improvements in spot peptide array technologies. They allow the high-throughput mutational analysis of peptides with, depending on assay design, an evaluation of binding stabilities. Here, we examine the cross-reactive capacity of an antibody fragment using the PEPperCHIP(®) technology platform (PEPperPRINT GmbH, Heidelberg, Germany; >8800 peptides per microarray) combined with the surface plasmon resonance characterization (Biacore(®) technology; GE-Healthcare Biacore, Uppsala, Sweden) of a subset of interactions. ScFv1F4 recognizes the N-terminal end of oncoprotein E6 of human papilloma virus 16. The spot permutation analysis (i.e. each position substituted by all amino acids except cysteine) of the wild type decapeptide (sequence (6)TAMFQDPQER(15)) and of 15 variants thereof defined the optimal epitope and provided a ranking for variant recognition. The SPR affinity measurements mostly validated the ranking of complex stabilities deduced from array data and defined the sensitivity of spot fluorescence intensities, bringing further insight into the conditions for cross-reactivity. Our data demonstrate the importance of throughput and quantification in the assessment of antibody selectivity.}, keywords = {DUMAS antibody selectivity antigen-antibody recognition cross-reactivity peptide array spot peptide surface plasmon resonance, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The HDAC6/APOBEC3G complex regulates HIV-1 infectiveness by inducing Vif autophagic degradation.}, author = {M S Valera and L de Armas Rillo and J Barroso-González and S Ziglio and J Batisse and N Dubois and S Marrero-Hernández and S Borel and L García-Expósito and M Biard-Piechaczyk and J C Paillart and A Valenzuela-Fernández}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26105074?dopt=Abstract}, doi = {10.1186/s12977-015-0181-5}, isbn = {26105074}, year = {2015}, date = {2015-01-01}, journal = {Retrovirology}, volume = {12}, pages = {53}, abstract = {BACKGROUND: Human immunodeficiency virus type 1 (HIV-1) has evolved a complex strategy to overcome the immune barriers it encounters throughout an organism thanks to its viral infectivity factor (Vif), a key protein for HIV-1 infectivity and in vivo pathogenesis. Vif interacts with and promotes "apolipoprotein B mRNA-editing enzyme-catalytic, polypeptide-like 3G" (A3G) ubiquitination and subsequent degradation by the proteasome, thus eluding A3G restriction activity against HIV-1. RESULTS: We found that cellular histone deacetylase 6 (HDAC6) directly interacts with A3G through its C-terminal BUZ domain (residues 841-1,215) to undergo a cellular co-distribution along microtubules and cytoplasm. The HDAC6/A3G complex occurs in the absence or presence of Vif, competes for Vif-mediated A3G degradation, and accounts for A3G steady-state expression level. In fact, HDAC6 directly interacts with and promotes Vif autophagic clearance, thanks to its C-terminal BUZ domain, a process requiring the deacetylase activity of HDAC6. HDAC6 degrades Vif without affecting the core binding factor β (CBF-β), a Vif-associated partner reported to be key for Vif- mediated A3G degradation. Thus HDAC6 antagonizes the proviral activity of Vif/CBF-β-associated complex by targeting Vif and stabilizing A3G. Finally, in cells producing virions, we observed a clear-cut correlation between the ability of HDAC6 to degrade Vif and to restore A3G expression, suggesting that HDAC6 controls the amount of Vif incorporated into nascent virions and the ability of HIV-1 particles of being infectious. This effect seems independent on the presence of A3G inside virions and on viral tropism. CONCLUSIONS: Our study identifies for the first time a new cellular complex, HDAC6/A3G, involved in the autophagic degradation of Vif, and suggests that HDAC6 represents a new antiviral factor capable of controlling HIV-1 infectiveness by counteracting Vif and its functions.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {EFRESOL: effective resolution of a diffraction data set}, author = {L Urzhumtseva and A Urzhumtsev}, url = {http://scripts.iucr.org/cgi-bin/paper?S1600576715001648}, doi = {10.1107/S1600576715001648}, year = {2015}, date = {2015-01-01}, journal = {J Appl Cryst}, volume = {48}, number = {2}, pages = {589-597}, abstract = {The resolution of a diffraction data set conveys the details that one expects to distinguish in the Fourier maps calculated with these data. For example, individual atoms in a macromolecular chain cannot be resolved in the maps calculated with 2 Å resolution data sets, while they can be resolved in accurate maps calculated with 1 Å resolution data. However, if a data set is incomplete its high-resolution cutoff becomes less straightforward to interpret. For instance, a Fourier map calculated using a 1 Å resolution data set with many high-resolution reflections missing may rather resemble a map corresponding to 2 Å resolution data. The authors have proposed a method that redefines the traditional notion of data resolution, making it more formal and general. This manuscript presents the corresponding tool, the program EFRESOL. For a data set of an arbitrary completeness, the program calculates its mean, highest and lowest effective resolutions. These values are established through the minimum distance between two point scatterers when their images are still resolved as separate peaks in the Fourier maps calculated with the given data set. Additionally, the program calculates the optical resolution, which is defined as the minimum distance for typical atoms of the structure when they are resolved in a hypothetical synthesis obtained with the given amplitudes and the exact phases if they are known. Both effective and optical resolutions show the `resolving power' of the diffraction data set.}, keywords = {high resolution resolution cutoff incomplete data sets anisotropic data sets point scatterers effective resolution optical resolution computer programs, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {105 Stabilization effects induced by modified nucleotides in tRNA T-loop motifs.}, author = {D Sydow and F Leonarski and L D'Ascenzo and P Auffinger}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26103316?dopt=Abstract}, doi = {10.1080/07391102.2015.1032667}, isbn = {26103316}, year = {2015}, date = {2015-01-01}, journal = {J Biomol Struct Dyn}, volume = {33}, number = {Suppl 1}, pages = {66}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dynamical features of the Plasmodium falciparum ribosome during translation.}, author = {M Sun and W Li and K Blomqvist and S Das and Y Hashem and J D Dvorin and J Frank}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26432834?dopt=Abstract}, doi = {10.1093/nar/gkv991}, isbn = {26432834}, year = {2015}, date = {2015-01-01}, journal = {Nucleic Acids Res}, volume = {43}, number = {21}, pages = {10515-10524}, abstract = {Plasmodium falciparum, the mosquito-transmitted Apicomplexan parasite, causes the most severe form of human malaria. In the asexual blood-stage, the parasite resides within erythrocytes where it proliferates, multiplies and finally spreads to new erythrocytes. Development of drugs targeting the ribosome, the site of protein synthesis, requires specific knowledge of its structure and work cycle, and, critically, the ways they differ from those in the human host. Here, we present five cryo-electron microscopy (cryo-EM) reconstructions of ribosomes purified from P. falciparum blood-stage schizonts at sub-nanometer resolution. Atomic models were built from these density maps by flexible fitting. Significantly, our study has taken advantage of new capabilities of cryo-EM, in visualizing several structures co-existing in the sample at once, at a resolution sufficient for building atomic models. We have discovered structural and dynamic features that differentiate the ribosomes of P. falciparum from those of mammalian system. Prompted by the absence of RACK1 on the ribosome in our and an earlier study we confirmed that RACK1 does not specifically co-purify with the 80S fraction in schizonts. More extensive studies, using cryo-EM methodology, of translation in the parasite will provide structural knowledge that may lead to development of novel anti-malarials.}, keywords = {HASHEM, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interplay between Catalysts and Substrates for Activity of Class Ib Aminoacyl-tRNA Synthetases and Implications for Pharmacology.}, author = {P Stephen and S X Lin and R Giege}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26286212?dopt=Abstract}, isbn = {26286212}, year = {2015}, date = {2015-01-01}, journal = {Curr Top Med Chem}, pages = {in print}, abstract = {Aminoacyl-tRNA synthetase:transfer RNA (aaRS:tRNA) systems became recently essential targets in molecular medicine, because perturbed recognition of cognate tRNAs by aaRSs and poor precision in tRNA aminoacylation do not guarantee accurate protein biosynthesis,thus leading to diseases. Sets of identity determinants situated at particular zones of tRNA are responsible for functional accuracy and recent work in X-ray crystallography has revealed various snapshots of aaRS:ligand complexes which represent the stages required for aminoacylation. Here we focus on a small group of class I aaRSs conserved in evolution, the ArgRSs, GluRSs, GlnRSs, and atypical LysRSs, found mostly in Archaea and in a few Bacteria, that catalyze amino acid activation only in the presence of their cognate tRNAs. Structural and functional features of these aaRSs, ranked in subclass Ib, together with their peculiar mode of tRNA recognition and identity expression are reviewed and compared. Strategies to inhibit class Ib aaRS:tRNA aminoacylation systems, their dysfunction leading to human diseases, and the implications for pharmacology are outlined.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Small RNA deep sequencing identifies viral microRNAs during malignant catarrhal fever induced by alcelaphine herpesvirus 1.}, author = {O Sorel and L Tuddenham and F Myster and L Palmeira and P Kerkhofs and S Pfeffer and A Vanderplasschen and B G Dewals}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26329753}, doi = {10.1099/jgv.0.000272}, isbn = {26329753}, year = {2015}, date = {2015-01-01}, journal = {J Gen Virol}, volume = {96}, number = {11}, pages = {3360-3372}, abstract = {Alcelaphine herpesvirus 1 (AlHV-1) is a γ-herpesvirus (γ-HV) carried asymptomatically by wildebeest. Upon cross-species transmission, AlHV-1 induces a fatal lymphoproliferative disease named malignant catarrhal fever (MCF) in many ruminants, including cattle and the rabbit model. Latency has been shown to be essential for MCF induction. However, the mechanisms causing the activation and proliferation of infected CD8+ T cells are unknown. Many γ-HVs express microRNAs (miRNAs). These small noncoding RNAs can regulate expression of host or viral target genes involved in various pathways and are thought to facilitate viral infection and/or mediate activation and proliferation of infected lymphocytes. AlHV-1 genome has been predicted to encode a large number of miRNAs. However, their precise contribution in viral infection and pathogenesis in vivo remains unknown. Here, using small RNAs cloning and sequencing we identified 36 potential miRNAs expressed in a lymphoblastoid cell line propagated from a calf infected with AlHV-1 and developing MCF. Among the sequenced candidate miRNAs, 32 were expressed on the reverse strand of the genome in two main clusters. The expression of these 32 viral miRNAs was further validated using northern blot and qRT-PCR in lymphoid organs of MCF-developing calves or rabbits. To determine the concerted contribution in MCF of 28 viral miRNAs clustered in the non-protein-coding region of the AlHV-1 genome, a recombinant virus was produced. The absence of these 28 miRNAs did not affect viral growth in vitro or MCF induction in rabbits, indicating that the AlHV-1 miRNAs clustered in this non-protein-coding genomic region are dispensable MCF induction.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutational interference mapping experiment (MIME) for studying RNA structure and function.}, author = {R P Smyth and L Despons and G Huili and S Bernacchi and M Hijnen and J Mak and F Jossinet and L Weixi and J C Paillart and M von Kleist and R Marquet}, url = {http://www.nature.com/nmeth/journal/v12/n9/full/nmeth.3490.html}, doi = {10.1038/nmeth.3490}, isbn = {26237229}, year = {2015}, date = {2015-01-01}, journal = {Nat Methods}, volume = {12}, pages = {866-872}, abstract = {RNA regulates many biological processes; however, identifying functional RNA sequences and structures is complex and time-consuming. We introduce a method, mutational interference mapping experiment (MIME), to identify, at single-nucleotide resolution, the primary sequence and secondary structures of an RNA molecule that are crucial for its function. MIME is based on random mutagenesis of the RNA target followed by functional selection and next-generation sequencing. Our analytical approach allows the recovery of quantitative binding parameters and permits the identification of base-pairing partners directly from the sequencing data. We used this method to map the binding site of the human immunodeficiency virus-1 (HIV-1) Pr55Gag protein on the viral genomic RNA in vitro, and showed that, by analyzing permitted base-pairing patterns, we could model RNA structure motifs that are crucial for protein binding.}, keywords = {MARQUET, PAILLART, Protein folding Molecular biophysics X-ray crystallography Cryoelectron microscopy Non-coding RNAs High-throughput screening RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutations of Human NARS2, Encoding the Mitochondrial Asparaginyl-tRNA Synthetase, Cause Nonsyndromic Deafness and Leigh Syndrome.}, author = {M Simon and E M Richard and X Wang and M Shahzad and V H Huang and T A Qaiser and P Potluri and S E Mahl and A Davila and S Nazli and S Hancock and M Yu and J Gargus and R Chang and N Al-Sheqaih and W G Newman and J Abdenur and A Starr and R Hegde and T Dorn and A Busch and E Park and J F Wu and H Schwenzer and A Flierl and C Florentz and M Sissler and S N Khan and R Li and M X Guan and T B Friedman and D K Wu and V Procaccio and S Riazuddin and D C Wallace and Z M Ahmed and T Huang and S Riazuddin}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25807530?dopt=Abstract}, doi = {10.1371/journal.pgen.1005097. eCollection 2015}, isbn = {25807530}, year = {2015}, date = {2015-01-01}, journal = {PLoS Genet}, volume = {11}, number = {3}, pages = {e1005097}, abstract = {Here we demonstrate association of variants in the mitochondrial asparaginyl-tRNA synthetase NARS2 with human hearing loss and Leigh syndrome. A homozygous missense mutation ([c.637G>T; p.Val213Phe]) is the underlying cause of nonsyndromic hearing loss (DFNB94) and compound heterozygous mutations ([c.969T>A; p.Tyr323*] + [c.1142A>G; p.Asn381Ser]) result in mitochondrial respiratory chain deficiency and Leigh syndrome, which is a neurodegenerative disease characterized by symmetric, bilateral lesions in the basal ganglia, thalamus, and brain stem. The severity of the genetic lesions and their effects on NARS2 protein structure cosegregate with the phenotype. A hypothetical truncated NARS2 protein, secondary to the Leigh syndrome mutation p.Tyr323* is not detectable and p.Asn381Ser further decreases NARS2 protein levels in patient fibroblasts. p.Asn381Ser also disrupts dimerization of NARS2, while the hearing loss p.Val213Phe variant has no effect on NARS2 oligomerization. Additionally we demonstrate decreased steady-state levels of mt-tRNAAsn in fibroblasts from the Leigh syndrome patients. In these cells we show that a decrease in oxygen consumption rates (OCR) and electron transport chain (ETC) activity can be rescued by overexpression of wild type NARS2. However, overexpression of the hearing loss associated p.Val213Phe mutant protein in these fibroblasts cannot complement the OCR and ETC defects. Our findings establish lesions in NARS2 as a new cause for nonsyndromic hearing loss and Leigh syndrome.}, keywords = {FLORENTZ, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Neurodegenerative disease-associated mutants of a human mitochondrial aminoacyl-tRNA synthetase present individual molecular signatures.}, author = {C Sauter and B Lorber and A Gaudry and L Karim and H Schwenzer and F Wien and P Roblin and C Florentz and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26620921?dopt=Abstract}, doi = {10.1038/srep17332}, isbn = {26620921}, year = {2015}, date = {2015-01-01}, journal = {Sci Rep}, volume = {5}, pages = {17332}, abstract = {Mutations in human mitochondrial aminoacyl-tRNA synthetases are associated with a variety of neurodegenerative disorders. The effects of these mutations on the structure and function of the enzymes remain to be established. Here, we investigate six mutants of the aspartyl-tRNA synthetase correlated with leukoencephalopathies. Our integrated strategy, combining an ensemble of biochemical and biophysical approaches, reveals that mutants are diversely affected with respect to their solubility in cellular extracts and stability in solution, but not in architecture. Mutations with mild effects on solubility occur in patients as allelic combinations whereas those with strong effects on solubility or on aminoacylation are necessarily associated with a partially functional allele. The fact that all mutations show individual molecular and cellular signatures and affect amino acids only conserved in mammals, points towards an alternative function besides aminoacylation.}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNA Biology in Mitochondria.}, author = {T Salinas-Giege and R Giege and P Giege}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25734984?dopt=Abstract}, isbn = {25734984}, year = {2015}, date = {2015-01-01}, journal = {Int J Mol Sci}, volume = {16}, number = {3}, pages = {4518-4559}, abstract = {Mitochondria are the powerhouses of eukaryotic cells. They are considered as semi-autonomous because they have retained genomes inherited from their prokaryotic ancestor and host fully functional gene expression machineries. These organelles have attracted considerable attention because they combine bacterial-like traits with novel features that evolved in the host cell. Among them, mitochondria use many specific pathways to obtain complete and functional sets of tRNAs as required for translation. In some instances, tRNA genes have been partially or entirely transferred to the nucleus and mitochondria require precise import systems to attain their pool of tRNAs. Still, tRNA genes have also often been maintained in mitochondria. Their genetic arrangement is more diverse than previously envisaged. The expression and maturation of mitochondrial tRNAs often use specific enzymes that evolved during eukaryote history. For instance many mitochondria use a eukaryote-specific RNase P enzyme devoid of RNA. The structure itself of mitochondrial encoded tRNAs is also very diverse, as e.g., in Metazoan, where tRNAs often show non canonical or truncated structures. As a result, the translational machinery in mitochondria evolved adapted strategies to accommodate the peculiarities of these tRNAs, in particular simplified identity rules for their aminoacylation. Here, we review the specific features of tRNA biology in mitochondria from model species representing the major eukaryotic groups, with an emphasis on recent research on tRNA import, maturation and aminoacylation.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Using droplet-based microfluidics to improve the catalytic properties of RNA under multiple-turnover conditions.}, author = {M Ryckelynck and S Baudrey and C Rick and A Marin and F Coldren and E Westhof and A D Griffiths}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25605963?dopt=Abstract}, doi = {10.1261/rna.048033.114}, isbn = {25605963}, year = {2015}, date = {2015-01-01}, journal = {RNA}, volume = {21}, number = {3}, pages = {458-469}, abstract = {In vitro evolution methodologies are powerful approaches to identify RNA with new functionalities. While Systematic Evolution of Ligands by Exponential enrichment (SELEX) is an efficient approach to generate new RNA aptamers, it is less suited for the isolation of efficient ribozymes as it does not select directly for the catalysis. In vitro compartmentalization (IVC) in aqueous droplets in emulsions allows catalytic RNAs to be selected under multiple-turnover conditions but suffers severe limitations that can be overcome using the droplet-based microfluidics workflow described in this paper. Using microfluidics, millions of genes in a library can be individually compartmentalized in highly monodisperse aqueous droplets and serial operations performed on them. This allows the different steps of the evolution process (gene amplification, transcription, and phenotypic assay) to be uncoupled, making the method highly flexible, applicable to the selection and evolution of a variety of RNAs, and easily adaptable for evolution of DNA or proteins. To demonstrate the method, we performed cycles of random mutagenesis and selection to evolve the X-motif, a ribozyme which, like many ribozymes selected using SELEX, has limited multiple-turnover activity. This led to the selection of variants, likely to be the optimal ribozymes that can be generated using point mutagenesis alone, with a turnover number under multiple-turnover conditions, kss cat, ∼28-fold higher than the original X-motif, primarily due to an increase in the rate of product release, the rate-limiting step in the multiple-turnover reaction.}, keywords = {RNA droplet-based microfluidics high-throughput screening in vitro evolution ribozymes, RYCKELYNCK, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of Lethal Mutations in Yeast Threonyl-tRNA Synthetase Revealing Critical Residues in Its Human Homolog.}, author = {Z R Ruan and Z P Fang and Q Ye and H Y Lei and G Eriani and X L Zhou and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25416776?dopt=Abstract}, doi = {10.1074/jbc.M114.599886}, isbn = {25416776}, year = {2015}, date = {2015-01-01}, journal = {J Biol Chem}, volume = {290}, number = {3}, pages = {1664-1678}, abstract = {Aminoacyl-tRNA synthetases (aaRSs) are a group of ancient enzymes catalyzing aminoacylation and editing reactions for protein biosynthesis. Increasing evidence suggests that these critical enzymes are often associated with mammalian disorders. Therefore, complete determination of the enzymes functions is essential for informed diagnosis and treatment. Here, we show that a yeast knockout strain for the threonyl-tRNA synthetase (ThrRS) gene is an excellent platform for such an investigation. Saccharomyces cerevisiae ThrRS (ScThrRS) has a unique modular structure containing four structural domains and a eukaryotic-specific N-terminal extension. Using randomly mutated libraries of the ThrRS gene (thrS) and a genetic screen, a set of loss-of-function mutants were identified. The mutations affected the synthetic and editing activities and influenced the dimer interface. The results also highlighted the role of the N-terminal extension for enzymatic activity and protein stability. To gain insights into the pathological mechanisms induced by mutated aaRSs, we systematically introduced the loss-of-function mutations into the human cytoplasmic ThrRS gene. All mutations induced similar detrimental effects, showing that the yeast model could be used to study pathology-associated point mutations in mammalian aaRSs.}, keywords = {aminoacyl tRNA synthetase aminoacylation editing mutagenesis protein structure transfer RNA (tRNA) translation, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural insights into the translational infidelity mechanism.}, author = {A Rozov and N Demeshkina and E Westhof and M Yusupov and G Yusupova}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26037619?dopt=Abstract}, doi = {10.1038/ncomms8251}, isbn = {26037619}, year = {2015}, date = {2015-01-01}, journal = {Nat Commun}, volume = {6}, pages = {7251}, abstract = {The decoding of mRNA on the ribosome is the least accurate process during genetic information transfer. Here we propose a unified decoding mechanism based on 11 high-resolution X-ray structures of the 70S ribosome that explains the occurrence of missense errors during translation. We determined ribosome structures in rare states where incorrect tRNAs were incorporated into the peptidyl-tRNA-binding site. These structures show that in the codon-anticodon duplex, a G·U mismatch adopts the Watson-Crick geometry, indicating a shift in the tautomeric equilibrium or ionization of the nucleobase. Additional structures with mismatches in the 70S decoding centre show that the binding of any tRNA induces identical rearrangements in the centre, which favours either isosteric or close to the Watson-Crick geometry codon-anticodon pairs. Overall, the results suggest that a mismatch escapes discrimination by preserving the shape of a Watson-Crick pair and indicate that geometric selection via tautomerism or ionization dominates the translational infidelity mechanism.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {A glimpse at long regulatory RNAs in various organisms.}, author = {P Romby and O Dontsova}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26365222?dopt=Abstract}, doi = {10.1016/j.biochi.2015.08.011}, isbn = {26365222}, year = {2015}, date = {2015-01-01}, journal = {Biochimie}, volume = {117}, pages = {1-2}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Disorder-to-order transition of MAGI-1 PDZ1 C-terminal extension upon peptide binding: thermodynamic and dynamic insights.}, author = {J Ramirez and R Recht and S Charbonnier and E Ennifar and R A Atkinson and G Travé and Y Nominé and B Kieffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25590897?dopt=Abstract}, doi = {10.1021/bi500845j}, isbn = {25590897}, year = {2015}, date = {2015-01-01}, journal = {Biochemistry}, volume = {54}, number = {6}, pages = {1327-1337}, abstract = {PDZ domains are highly abundant protein-protein interaction modules commonly found in multidomain scaffold proteins. The PDZ1 domain of MAGI-1, a protein present at cellular tight junctions that contains 6 PDZ domains, is targeted by the E6 oncoprotein of the high-risk Human Papilloma Virus. Thermodynamic and dynamic studies using complementary ITC and NMR 15N heteronuclear relaxation measurements were conducted at different temperatures to decipher the molecular mechanism of this interaction. The binding of E6 peptides to the MAGI-1 PDZ1 is accompanied by an unusually large and negative change in heat capacity ΔCp that is attributed to a disorder-to-order transition of the C-terminal extension of the PDZ1 domain, observed upon E6 binding. Analysis of the temperature dependent thermodynamic parameters and 15N NMR relaxation data of a PDZ1 mutant for which this disorder-to-order transition was abolished allows the unusual thermodynamic signature of the E6 binding to be correlated to the local folding of PDZ1 C-terminal extension. The comparison of exchange contributions observed for wild-type and mutant proteins explains how the variation in solvent exposed area may compensate for the loss of conformational entropy and further designates a distinct set of few residues that mediate this local folding phenomenon.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization and crystallographic analysis of an Arabidopsis nuclear proteinaceous RNase P}, author = {F Pinker and P Giege and C Sauter}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26527263?dopt=Abstract}, doi = {10.1107/S2053230X15017033}, isbn = {26527263}, year = {2015}, date = {2015-01-01}, journal = {Acta Crystallogr F Struct Biol Commun}, volume = {71}, number = {Pt 11}, pages = {1372-1377}, abstract = {RNase P activity is ubiquitous and involves the 5' maturation of precursor tRNAs. For a long time, it was thought that all RNases P were ribonucleoproteic enzymes. However, the characterization of RNase P in human mitochondria and in plants revealed a novel kind of RNase P composed of protein only, called PRORP for `proteinaceous RNase P'. Whereas in human mitochondria PRORP has two partners that are required for RNase P activity, PRORP proteins are active as single-subunit enzymes in plants. Three paralogues of PRORP are found in Arabidopsis thaliana. PRORP1 is responsible for RNase P in mitochondria and chloroplasts, while PRORP2 and PRORP3 are nuclear enzymes. Here, the purification and crystallization of the Arabidopsis PRORP2 protein are reported. Optimization of the initial crystallization conditions led to crystals that diffracted to 3 Å resolution.}, keywords = {RNase P nuclear PRORP PPR tRNA maturation enzyme, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Modulation of the microRNA cluster miR-183-96-182 expression by the Epstein-Barr virus latent membrane protein 1.}, author = {L Oussaief and A Fendri and B Chane-Woon-Ming and R Poirey and H J Delecluse and I Joab and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26401047?dopt=Abstract}, doi = {10.1128/JVI.01757-15}, isbn = {26401047}, year = {2015}, date = {2015-01-01}, journal = {J Virol}, volume = {89}, number = {23}, pages = {12178-12188}, abstract = {Epstein-Barr virus (EBV) is an oncogenic human herpesvirus involved in the pathogenesis of Burkitt's lymphoma (BL) and various other lymphoproliferative disorders. In BL, EBV protein expression is restricted to the EBNA1, but small non-coding RNAs such as the EBERs and micro (mi)RNAs can also be detected. miRNAs play major roles in crucial processes such as proliferation, differentiation and cell death. It has recently become clear that alteration in the expression profile of miRNAs contribute to the pathogenesis of a number of malignancies. During latent infection, EBV expresses 25 viral pre-miRNAs and modulates the expression of specific cellular miRNAs, such as miR-155 and miR-146, which potentially play a role in oncogenesis. Here, we established the small RNA expression profiles of three BL cell lines. Using large-scale sequencing coupled to northern blot and real-time RT-PCR analysis validation, we demonstrated the differential expression of some cellular and viral miRNAs. High-level expression of the miR-183-96-182 cluster and EBV miR-BART cluster was significantly associated with EBV type I latency. This expression was not affected by viral reactivation since TGF-β1 stimulation did not significantly change the miRNA profiles. However, using several approaches, including de novo infection with a mutant virus, we present evidence that the expression of latent membrane protein (LMP) 1 triggered down-regulation of the expression of the miR-183-96-182 cluster. We further show that this effect involves the Akt signaling pathway. IMPORTANCE: In addition to expressing their own miRNAs, herpesviruses also impact the expression levels of cellular miRNAs. This regulation can be either positive or negative and usually results in the perturbation of pathways to create a cellular environment that is more "virus-friendly". For example, EBV induces the expression of miR-155, a well-characterized oncomiR, which leads to increased cell proliferation and decreased cell death. Here, we show that EBV-encoded LMP-1 protein is also involved in the down-regulation of a cluster of three miRNAs, miR-183, -96 and -182, which are known to be also repressed in several cancers. We therefore identify yet another potential player in EBV-induced oncogenesis.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A defense-offense multi-layered regulatory switch in a pathogenic bacterium.}, author = {M Nitzan and P Fechter and A Peer and Y Altuvia and D Bronesky and F Vandenesch and P Romby and O Biham and H Margalit}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25628364?dopt=Abstract}, doi = {10.1093/nar/gkv001}, isbn = {25628364}, year = {2015}, date = {2015-01-01}, journal = {Nucleic Acids Res}, volume = {43}, number = {3}, pages = {1357-1369}, abstract = {Cells adapt to environmental changes by efficiently adjusting gene expression programs. Staphylococcus aureus, an opportunistic pathogenic bacterium, switches between defensive and offensive modes in response to quorum sensing signal. We identified and studied the structural characteristics and dynamic properties of the core regulatory circuit governing this switch by deterministic and stochastic computational methods, as well as experimentally. This module, termed here Double Selector Switch (DSS), comprises the RNA regulator RNAIII and the transcription factor Rot, defining a double-layered switch involving both transcriptional and post-transcriptional regulations. It coordinates the inverse expression of two sets of target genes, immuno-modulators and exotoxins, expressed during the defensive and offensive modes, respectively. Our computational and experimental analyses show that the DSS guarantees fine-tuned coordination of the inverse expression of its two gene sets, tight regulation, and filtering of noisy signals. We also identified variants of this circuit in other bacterial systems, suggesting it is used as a molecular switch in various cellular contexts and offering its use as a template for an effective switching device in synthetic biology studies.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A Large-Scale Assessment of Nucleic Acids Binding Site Prediction Programs.}, author = {Z Miao and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26681179?dopt=Abstract}, doi = {10.1371/journal.pcbi.1004639}, isbn = {26681179}, year = {2015}, date = {2015-01-01}, journal = {PLoS Comput Biol}, volume = {11}, number = {12}, pages = {e1004639}, abstract = {Computational prediction of nucleic acid binding sites in proteins are necessary to disentangle functional mechanisms in most biological processes and to explore the binding mechanisms. Several strategies have been proposed, but the state-of-the-art approaches display a great diversity in i) the definition of nucleic acid binding sites; ii) the training and test datasets; iii) the algorithmic methods for the prediction strategies; iv) the performance measures and v) the distribution and availability of the prediction programs. Here we report a large-scale assessment of 19 web servers and 3 stand-alone programs on 41 datasets including more than 5000 proteins derived from 3D structures of protein-nucleic acid complexes. Well-defined binary assessment criteria (specificity, sensitivity, precision, accuracy…) are applied. We found that i) the tools have been greatly improved over the years; ii) some of the approaches suffer from theoretical defects and there is still room for sorting out the essential mechanisms of binding; iii) RNA binding and DNA binding appear to follow similar driving forces and iv) dataset bias may exist in some methods.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Prediction of nucleic acid binding probability in proteins: a neighboring residue network based score.}, author = {Z Miao and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25940624?dopt=Abstract}, doi = {10.1093/nar/gkv446}, isbn = {25940624}, year = {2015}, date = {2015-01-01}, journal = {Nucleic Acids Res}, volume = {43}, number = {11}, pages = {5340-5351}, abstract = {We describe a general binding score for predicting the nucleic acid binding probability in proteins. The score is directly derived from physicochemical and evolutionary features and integrates a residue neighboring network approach. Our process achieves stable and high accuracies on both DNA- and RNA-binding proteins and illustrates how the main driving forces for nucleic acid binding are common. Because of the effective integration of the synergetic effects of the network of neighboring residues and the fact that the prediction yields a hierarchical scoring on the protein surface, energy funnels for nucleic acid binding appear on protein surfaces, pointing to the dynamic process occurring in the binding of nucleic acids to proteins.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA-Puzzles Round II: assessment of RNA structure prediction programs applied to three large RNA structures.}, author = {Z Miao and R W Adamiak and M F Blanchet and M Boniecki and J M Bujnicki and S J Chen and C Cheng and G Chojnowski and F C Chou and P Cordero and J A Cruz and A R Ferré-D'amaré and R Das and F Ding and N V Dokholyan and S Dunin-Horkawicz and W Kladwang and A Krokhotin and G Lach and M Magnus and F Major and T H Mann and B Masquida and D Matelska and M Meyer and A Peselis and M Popenda and K J Purzycka and A Serganov and J Stasiewicz and M Szachniuk and A Tandon and S Tian and J Wang and Y Xiao and X Xu and J Zhang and P Zhao and T Zok and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25883046}, doi = {10.1261/rna.049502.114}, isbn = {25883046}, year = {2015}, date = {2015-01-01}, journal = {RNA}, volume = {21}, number = {6}, pages = {1066-1084}, abstract = {This paper is a report of a second round of RNA-Puzzles, a collective and blind experiment in three-dimensional (3D) RNA structure prediction. Three puzzles, Puzzles 5, 6, and 10, represented sequences of three large RNA structures with limited or no homology with previously solved RNA molecules. A lariat-capping ribozyme, as well as riboswitches complexed to adenosylcobalamin and tRNA, were predicted by seven groups using RNAComposer, ModeRNA/SimRNA, Vfold, Rosetta, DMD, MC-Fold, 3dRNA, and AMBER refinement. Some groups derived models using data from state-of-the-art chemical-mapping methods (SHAPE, DMS, CMCT, and mutate-and-map). The comparisons between the predictions and the three subsequently released crystallographic structures, solved at diffraction resolutions of 2.5-3.2 Å, were carried out automatically using various sets of quality indicators. The comparisons clearly demonstrate the state of present-day de novo prediction abilities as well as the limitations of these state-of-the-art methods. All of the best prediction models have similar topologies to the native structures, which suggests that computational methods for RNA structure prediction can already provide useful structural information for biological problems. However, the prediction accuracy for non-Watson-Crick interactions, key to proper folding of RNAs, is low and some predicted models had high Clash Scores. These two difficulties point to some of the continuing bottlenecks in RNA structure prediction. All submitted models are available for download at http://ahsoka.u-strasbg.fr/rnapuzzles/.}, keywords = {3D prediction X-ray structures bioinformatics force fields models structure quality, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Clearance of persistent hepatitis C virus infection in humanized mice using a claudin-1-targeting monoclonal antibody.}, author = {L Mailly and F Xiao and J Lupberger and G K Wilson and P Aubert and F H Duong and D Calabrese and C Leboeuf and I Fofana and C Thumann and S Bandiera and M Lütgehetmann and T Volz and C Davis and H J Harris and C J Mee and E Girardi and B Chane-Woon-Ming and M Ericsson and N Fletcher and R Bartenschlager and P Pessaux and K Vercauteren and P Meuleman and P Villa and L Kaderali and S Pfeffer and M H Heim and M Neunlist and M B Zeisel and M Dandri and J A McKeating and E Robinet and T F Baumert}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25798937?dopt=Abstract}, doi = {10.1038/nbt.3179}, isbn = {25798937}, year = {2015}, date = {2015-01-01}, journal = {Nat Biotechnol}, volume = {33}, number = {5}, pages = {549-554}, abstract = {Hepatitis C virus (HCV) infection is a leading cause of liver cirrhosis and cancer. Cell entry of HCV and other pathogens is mediated by tight junction (TJ) proteins, but successful therapeutic targeting of TJ proteins has not been reported yet. Using a human liver-chimeric mouse model, we show that a monoclonal antibody specific for the TJ protein claudin-1 (ref. 7) eliminates chronic HCV infection without detectable toxicity. This antibody inhibits HCV entry, cell-cell transmission and virus-induced signaling events. Antibody treatment reduces the number of HCV-infected hepatocytes in vivo, highlighting the need for de novo infection by means of host entry factors to maintain chronic infection. In summary, we demonstrate that an antibody targeting a virus receptor can cure chronic viral infection and uncover TJ proteins as targets for antiviral therapy.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {APOBEC3F/G and Vif: action and counteractions.}, author = {C Libre and J Batisse and S Guerrero and R Marquet and J C Paillart}, editor = {T Hope and M Stevenson and D Richman}, url = {http://link.springer.com/referenceworkentry/10.1007/978-1-4614-9610-6_376-1}, doi = {10.1007/978-1-4614-9610-6_376-1}, year = {2015}, date = {2015-01-01}, booktitle = {Encyclopedia of AIDS}, pages = {1-12}, publisher = {Springer New York}, abstract = {The primary targets of HIV-1 (T lymphocytes, macrophages, and monocytes) are able to limit HIV-1 replication by expressing the restriction factors APOBEC3F and APOBEC3G (A3F/G). These two proteins have cytidine deaminase activity and induce mutations during reverse transcription of the genomic RNA that could be lethal for the virus. A3F/G also interfere with the reverse transcription and integration processes independently from this deaminase activity. To counteract this restriction, HIV-1 has evolved the viral infectivity factor (Vif), a multifunctional protein that is able to reduce the cellular A3F/G expression level through two major mechanisms: (1) Vif induces degradation of A3F/G by the proteasome by recruiting an E3 ubiquitin ligase complex, and (2) Vif inhibits A3F/G translation by interacting with the 5′ untranslated region (UTR) of their mRNAs. These two mechanisms ultimately reduce the packaging of A3F/G into virions. The intimate relationship between Vif and A3F/G.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Efficient Estimation of Three-Dimensional Covariance and its Application in the Analysis of Heterogeneous Samples in Cryo-Electron Microscopy.}, author = {H Y Liao and Y Hashem and J Frank}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25982529?dopt=Abstract}, doi = {10.1016/j.str.2015.04.004}, isbn = {25982529}, year = {2015}, date = {2015-01-01}, journal = {Structure}, volume = {23}, number = {6}, pages = {1129-1137}, abstract = {Single-particle cryogenic electron microscopy (cryo-EM) is a powerful tool for the study of macromolecular structures at high resolution. Classification allows multiple structural states to be extracted and reconstructed from the same sample. One classification approach is via the covariance matrix, which captures the correlation between every pair of voxels. Earlier approaches employ computing-intensive resampling and estimate only the eigenvectors of the matrix, which are then used in a separate fast classification step. We propose an iterative scheme to explicitly estimate the covariance matrix in its entirety. In our approach, the flexibility in choosing the solution domain allows us to examine a part of the molecule in greater detail. Three-dimensional covariance maps obtained in this way from experimental data (cryo-EM images of the eukaryotic pre-initiation complex) prove to be in excellent agreement with conclusions derived by using traditional approaches, revealing in addition the interdependencies of ligand bindings and structural changes.}, keywords = {HASHEM, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Calpain Cleaves Most Components in the Multiple Aminoacyl-tRNA Synthetase Complex and Affects Their Functions.}, author = {H Y Lei and X L Zhou and Z R Ruan and W C Sun and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26324710?dopt=Abstract}, doi = {10.1074/jbc.M115.681999}, isbn = {26324710}, year = {2015}, date = {2015-01-01}, journal = {J Biol Chem}, volume = {290}, number = {43}, pages = {26314-26327}, abstract = {Nine aminoacyl-tRNA synthetases (aaRSs) and three scaffold proteins form a super multiple aminoacyl-tRNA synthetase complex (MSC) in the human cytoplasm. Domains that have been added progressively to MSC components during evolution are linked by unstructured flexible peptides, producing an elongated and multiarmed MSC structure that is easily attacked by proteases in vivo. A yeast two-hybrid screen for proteins interacting with LeuRS, a representative MSC member, identified calpain 2, a calcium-activated neutral cysteine protease. Calpain 2 and calpain 1 could partially hydrolyze most MSC components to generate specific fragments that resembled those reported previously. The cleavage sites of calpain in ArgRS, GlnRS, and p43 were precisely mapped. After cleavage, their N-terminal regions were removed. Sixty-three amino acid residues were removed from the N terminus of ArgRS to form ArgRSΔN63; GlnRS formed GlnRSΔN198, and p43 formed p43ΔN106. GlnRSΔN198 had a much weaker affinity for its substrates, tRNA(Gln) and glutamine. p43ΔN106 was the same as the previously reported p43-derived apoptosis-released factor. The formation of p43ΔN106 by calpain depended on Ca(2+) and could be specifically inhibited by calpeptin and by RNAi of the regulatory subunit of calpain in vivo. These results showed, for the first time, that calpain plays an essential role in dissociating the MSC and might regulate the canonical and non-canonical functions of certain components of the MSC.}, keywords = {aminoacyl tRNA synthetase aminoacylation calpain protein fragments protein secretion proteolysis stress, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of a ribosomal RNA phosphate oxygen during the EF-G-triggered GTP hydrolysis.}, author = {M Koch and S Flür and C Kreutz and E Ennifar and R Micura and N Polacek}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25941362?dopt=Abstract}, doi = {10.1073/pnas.1505231112}, isbn = {25941362}, year = {2015}, date = {2015-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {112}, number = {20}, pages = {E2561-2568}, abstract = {Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases.}, keywords = {ENNIFAR, RNA solid-phase synthesis methylphosphonate modification protein biosynthesis tRNA translocation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Preface.}, author = {F Jossinet and Y Ponty and J Waldispühl}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25768234?dopt=Abstract}, doi = {10.1089/cmb.2015.010P}, isbn = {25768234}, year = {2015}, date = {2015-01-01}, journal = {J Comput Biol}, volume = {22}, number = {3}, pages = {189}, keywords = {JOSSINET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 replication and the cellular eukaryotic translation apparatus}, author = {S X Guerrero and J Batisse and C Libre and S Bernacchi and R Marquet and J C Paillart}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25606970?dopt=Abstract}, isbn = {25606970}, year = {2015}, date = {2015-01-01}, journal = {Viruses}, volume = {7}, number = {1}, pages = {199-218}, abstract = {Eukaryotic translation is a complex process composed of three main steps: initiation, elongation, and termination. During infections by RNA- and DNA-viruses, the eukaryotic translation machinery is used to assure optimal viral protein synthesis. Human immunodeficiency virus type I (HIV-1) uses several non-canonical pathways to translate its own proteins, such as leaky scanning, frameshifting, shunt, and cap-independent mechanisms. Moreover, HIV-1 modulates the host translation machinery by targeting key translation factors and overcomes different cellular obstacles that affect protein translation. In this review, we describe how HIV-1 proteins target several components of the eukaryotic translation machinery, which consequently improves viral translation and replication.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cross-species comparative analysis of Dicer proteins during Sindbis virus infection.}, author = {E Girardi and M Lefèvre and B Chane-Woon-Ming and S Paro and B Claydon and JL Imler and C Meignin and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26024431?dopt=Abstract}, doi = {10.1038/srep10693}, isbn = {26024431}, year = {2015}, date = {2015-01-01}, urldate = {2015-01-01}, journal = {Sci Rep}, volume = {5}, pages = {10693}, abstract = {In plants and invertebrates RNA silencing is a major defense mechanism against virus infections. The first event in RNA silencing is dicing of long double stranded RNAs into small interfering RNAs (siRNAs). The Dicer proteins involved in this process are phylogenetically conserved and have the same domain organization. Accordingly, the production of viral derived siRNAs has also been observed in the mouse, but only in restricted cell types. To gain insight on this restriction, we compare the dicing activity of human Dicer and fly Dicer-2 in the context of Sindbis virus (SINV) infection. Expression of human Dicer in flies inefficiently rescues the production of viral siRNAs but confers some protection against SINV. Conversely, expression of Dicer-2 in human cells allows the production of viral 21 nt small RNAs. However, this does not confer resistance to viral infection, but on the contrary results in stronger accumulation of viral RNA. We further show that Dicer-2 expression in human cells perturbs interferon (IFN) signaling pathways and antagonizes protein kinase R (PKR)-mediated antiviral immunity. Overall, our data suggest that a functional incompatibility between the Dicer and IFN pathways explains the predominance of the IFN response in mammalian somatic cells.}, keywords = {meignin, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The external domains of the HIV-1 envelope are a mutational cold spot.}, author = {R Geller and P Domingo-Calap and J M Cuevas and P Rossolillo and M Negroni and R Sanjuán}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26450412?dopt=Abstract}, doi = {10.1038/ncomms9571}, isbn = {26450412}, year = {2015}, date = {2015-01-01}, journal = {Nat Commun}, volume = {6}, pages = {8571}, abstract = {In RNA viruses, mutations occur fast and have large fitness effects. While this affords remarkable adaptability, it can also endanger viral survival due to the accumulation of deleterious mutations. How RNA viruses reconcile these two opposed facets of mutation is still unknown. Here we show that, in human immunodeficiency virus (HIV-1), spontaneous mutations are not randomly located along the viral genome. We find that the viral mutation rate experiences a threefold reduction in the region encoding the most external domains of the viral envelope, which are strongly targeted by neutralizing antibodies. This contrasts with the hypermutation mechanisms deployed by other, more slowly mutating pathogens such as DNA viruses and bacteria, in response to immune pressure. We show that downregulation of the mutation rate in HIV-1 is exerted by the template RNA through changes in sequence context and secondary structure, which control the activity of apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (A3)-mediated cytidine deamination and the fidelity of the viral reverse transcriptase.}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {MIST, a Novel Approach to Reveal Hidden Substrate Specificity in Aminoacyl-tRNA Synthetases.}, author = {G Eriani and J Karam and J Jacinto and E Morris Richard and R Geslain}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26067673?dopt=Abstract}, doi = {10.1371/journal.pone.0130042}, isbn = {26067673}, year = {2015}, date = {2015-01-01}, journal = {PLoS One}, volume = {10}, number = {6}, pages = {e0130042}, abstract = {Aminoacyl-tRNA synthetases (AARSs) constitute a family of RNA-binding proteins, that participate in the translation of the genetic code, by covalently linking amino acids to appropriate tRNAs. Due to their fundamental importance for cell life, AARSs are likely to be one of the most ancient families of enzymes and have therefore been characterized extensively. Paradoxically, little is known about their capacity to discriminate tRNAs mainly because of the practical challenges that represent precise and systematic tRNA identification. This work describes a new technical and conceptual approach named MIST (Microarray Identification of Shifted tRNAs) designed to study the formation of tRNA/AARS complexes independently from the aminoacylation reaction. MIST combines electrophoretic mobility shift assays with microarray analyses. Although MIST is a non-cellular assay, it fully integrates the notion of tRNA competition. In this study we focus on yeast cytoplasmic Arginyl-tRNA synthetase (yArgRS) and investigate in depth its ability to discriminate cellular tRNAs. We report that yArgRS in submicromolar concentrations binds cognate and non-cognate tRNAs with a wide range of apparent affinities. In particular, we demonstrate that yArgRS binds preferentially to type II tRNAs but does not support their misaminoacylation. Our results reveal important new trends in tRNA/AARS complex formation and potential deep physiological implications.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Multiple ways to regulate translation initiation in bacteria: mechanisms, regulatory circuits, dynamics.}, author = {M Duval and A Simonetti and I Caldelari and S Marzi}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25792421?dopt=Abstract}, doi = {10.1016/j.biochi.2015.03.007}, isbn = {25792421}, year = {2015}, date = {2015-01-01}, journal = {Biochimie}, volume = {114}, pages = {18-29}, abstract = {To adapt their metabolism rapidly and constantly in response to environmental variations, bacteria often target the translation initiation process, during which the ribosome assembles on the mRNA. Here, we review different mechanisms of regulation mediated by cis-acting elements, sRNAs and proteins, showing, when possible, their intimate connection with the translational apparatus. Indeed the ribosome itself could play a direct role in several regulatory mechanisms. Different features of the regulatory signals (sequences, structures and their positions on the mRNA) are contributing to the large variety of regulatory mechanisms. Ribosome heterogeneity, variation of individual cells responses and the spatial and temporal organization of the translation process add more layers of complexity. This hampers to define manageable set of rules for bacterial translation initiation control.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {sRNA and mRNA turnover in Gram-positive bacteria.}, author = {S Durand and A Tomasini and F Braun and C Condon and P Romby}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25934118?dopt=Abstract}, isbn = {25934118}, year = {2015}, date = {2015-01-01}, journal = {FEMS Microbiol Rev}, volume = {39}, number = {3}, pages = {316-330}, abstract = {t is widely recognized that RNA degradation plays a critical role in gene regulation when fast adaptation of cell growth is required to respond to stress and changing environmental conditions. Bacterial ribonucleases acting alone or in concert with various trans-acting regulatory factors are important mediators of RNA degradation. Here, we will give an overview of what is known about ribonucleases in several Gram-positive bacteria, their specificities and mechanisms of action. In addition, we will illustrate how sRNAs act in a coordinated manner with ribonucleases to regulate the turnover of particular mRNA targets, and the complex interplay existing between the ribosome, the ribonucleases and RNAs.}, keywords = {Gram-positive bacteria mRNA stability ribonucleases sRNA, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{schaeffer_dermal_2015, title = {Dermal CD14(+) Dendritic Cell and Macrophage Infection by Dengue Virus Is Stimulated by Interleukin-4}, author = {Evelyne Schaeffer and Vincent Flacher and Vasiliki Papageorgiou and Marion Decossas and Jean-Daniel Fauny and Melanie Krämer and Christopher G Mueller}, doi = {10.1038/jid.2014.525}, issn = {1523-1747}, year = {2015}, date = {2015-01-01}, journal = {The Journal of Investigative Dermatology}, volume = {135}, number = {7}, pages = {1743--1751}, abstract = {Dengue virus (DENV) is responsible for the most prevalent arthropod-borne viral infection in humans. Events decisive for disease development occur in the skin after virus inoculation by the mosquito. Yet, the role of human dermis-resident immune cells in dengue infection and disease remains elusive. Here we investigated how dermal dendritic cells (dDCs) and macrophages (dMs) react to DENV and impact on immunopathology. We show that both CD1c(+) and CD14(+) dDC subsets were infected, but viral load greatly increased in CD14(+) dDCs upon IL-4 stimulation, which correlated with upregulation of virus-binding lectins Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Nonintegrin (DC-SIGN/CD209) and mannose receptor (CD206). IL-4 also enhanced T-cell activation by dDCs, which was further increased upon dengue infection. dMs purified from digested dermis were initially poorly infected but actively replicated the virus and produced TNF-α upon lectin upregulation in response to IL-4. DC-SIGN(+) cells are abundant in inflammatory skin with scabies infection or Th2-type dermatitis, suggesting that skin reactions to mosquito bites heighten the risk of infection and subsequent immunopathology. Our data identify dDCs and dMs as primary arbovirus target cells in humans and suggest that dDCs initiate a potent virus-directed T-cell response, whereas dMs fuel the inflammatory cascade characteristic of dengue fever.}, keywords = {Abdominal Wall, Antigen-Presenting Cells, C-Type, Cell Adhesion Molecules, Cell Surface, Cells, Confocal, Cultured, Cytokines, Dengue, Dengue virus, Enzyme-Linked Immunosorbent Assay, Epidermis, Humans, I2CT, Imagerie, Interleukin-4, Langerhans Cells, Lectins, Lymphocyte Activation, Macrophages, Microscopy, Receptors, Sensitivity and Specificity, Skin Diseases, Team-Mueller, Viral}, pubstate = {published}, tppubtype = {article} } @article{, title = {A Nitric Oxide Regulated Small RNA Controls Expression of Genes Involved in Redox Homeostasis in Bacillus subtilis.}, author = {S Durand and F Braun and E Lioliou and C Romilly and A C Helfer and L Kuhn and N Quittot and P Nicolas and P Romby and C Condon}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25643072?dopt=Abstract}, doi = {10.1371/journal.pgen.1004957}, isbn = {25643072}, year = {2015}, date = {2015-01-01}, journal = {PLoS Genet}, volume = {11}, number = {2}, pages = {e1004957}, abstract = {RsaE is the only known trans-acting small regulatory RNA (sRNA) besides the ubiquitous 6S RNA that is conserved between the human pathogen Staphylococcus aureus and the soil-dwelling Firmicute Bacillus subtilis. Although a number of RsaE targets are known in S. aureus, neither the environmental signals that lead to its expression nor its physiological role are known. Here we show that expression of the B. subtilis homolog of RsaE is regulated by the presence of nitric oxide (NO) in the cellular milieu. Control of expression by NO is dependent on the ResDE two-component system in B. subtilis and we determined that the same is true in S. aureus. Transcriptome and proteome analyses revealed that many genes with functions related to oxidative stress and oxidation-reduction reactions were up-regulated in a B. subtilis strain lacking this sRNA. We have thus renamed it RoxS. The prediction of RoxS-dependent mRNA targets also suggested a significant enrichment for mRNAs related to respiration and electron transfer. Among the potential direct mRNA targets, we have validated the ppnKB mRNA, encoding an NAD+/NADH kinase, both in vivo and in vitro. RoxS controls both translation initiation and the stability of this transcript, in the latter case via two independent pathways implicating RNase Y and RNase III. Furthermore, RNase Y intervenes at an additional level by processing the 5' end of the RoxS sRNA removing about 20 nucleotides. Processing of RoxS allows it to interact more efficiently with a second target, the sucCD mRNA, encoding succinyl-CoA synthase, thus expanding the repertoire of targets recognized by this sRNA.}, keywords = {PPSE, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of mammalian eIF3 in the context of the 43S preinitiation complex.}, author = {A des Georges and V Dhote and L Kuhn and C U Hellen and T V Pestova and J Frank and Y Hashem}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26344199}, doi = {10.1038/nature14891}, isbn = {26344199}, year = {2015}, date = {2015-01-01}, journal = {Nature}, volume = {525}, number = {7570}, pages = {491-495}, abstract = {During eukaryotic translation initiation, 43S complexes, comprising a 40S ribosomal subunit, initiator transfer RNA and initiation factors (eIF) 2, 3, 1 and 1A, attach to the 5'-terminal region of messenger RNA and scan along it to the initiation codon. Scanning on structured mRNAs also requires the DExH-box protein DHX29. Mammalian eIF3 contains 13 subunits and participates in nearly all steps of translation initiation. Eight subunits having PCI (proteasome, COP9 signalosome, eIF3) or MPN (Mpr1, Pad1, amino-terminal) domains constitute the structural core of eIF3, to which five peripheral subunits are flexibly linked. Here we present a cryo-electron microscopy structure of eIF3 in the context of the DHX29-bound 43S complex, showing the PCI/MPN core at ∼6 Å resolution. It reveals the organization of the individual subunits and their interactions with components of the 43S complex. We were able to build near-complete polyalanine-level models of the eIF3 PCI/MPN core and of two peripheral subunits. The implications for understanding mRNA ribosomal attachment and scanning are discussed.}, keywords = {HASHEM, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {106 Ion-π interactions in biomolecular systems.}, author = {L D'Ascenzo and P Auffinger}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26103317?dopt=Abstract}, doi = {10.1080/07391102.2015.1032668.}, isbn = {26103317}, year = {2015}, date = {2015-01-01}, journal = {J Biomol Struct Dyn}, volume = {33}, number = {Suppl 1}, pages = {67}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {A comprehensive classification and nomenclature of carboxyl-carboxyl(ate) supramolecular motifs and related catemers: implications for biomolecular systems.}, author = {L D'Ascenzo and P Auffinger}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25827369?dopt=Abstract}, doi = {10.1107/S205252061500270X}, isbn = {25827369}, year = {2015}, date = {2015-01-01}, journal = {Acta Crystallogr B Struct Sci Cryst Eng Mater}, volume = {71}, number = {Pt 2}, pages = {164-175}, abstract = {Carboxyl and carboxylate groups form important supramolecular motifs (synthons). Besides carboxyl cyclic dimers, carboxyl and carboxylate groups can associate through a single hydrogen bond. Carboxylic groups can further form polymeric-like catemer chains within crystals. To date, no exhaustive classification of these motifs has been established. In this work, 17 association types were identified (13 carboxyl-carboxyl and 4 carboxyl-carboxylate motifs) by taking into account the syn and anti carboxyl conformers, as well as the syn and anti lone pairs of the O atoms. From these data, a simple rule was derived stating that only eight distinct catemer motifs involving repetitive combinations of syn and anti carboxyl groups can be formed. Examples extracted from the Cambridge Structural Database (CSD) for all identified dimers and catemers are presented, as well as statistical data related to their occurrence and conformational preferences. The inter-carboxyl(ate) and carboxyl(ate)-water hydrogen-bond properties are described, stressing the occurrence of very short (strong) hydrogen bonds. The precise characterization and classification of these supramolecular motifs should be of interest in crystal engineering, pharmaceutical and also biomolecular sciences, where similar motifs occur in the form of pairs of Asp/Glu amino acids or motifs involving ligands bearing carboxyl(ate) groups. Hence, we present data emphasizing how the analysis of hydrogen-containing small molecules of high resolution can help understand structural aspects of larger and more complex biomolecular systems of lower resolution.}, keywords = {biomolecular systems crystal engineering pharmaceuticals supramolecular motifs, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Potent Sensitisation of Cancer Cells to Anticancer Drugs by a Quadruple Mutant of the Human Deoxycytidine Kinase.}, author = {S T Coulibaly and P Rossolillo and F Winter and F K Kretzschmar and M Brayé and D P Martin and D Lener and M Negroni}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26485161?dopt=Abstract}, doi = {10.1371/journal.pone.0140741}, isbn = {26485161}, year = {2015}, date = {2015-01-01}, journal = {PLoS One}, volume = {10}, number = {10}, pages = {e0140741}, abstract = {Identifying enzymes that, once introduced in cancer cells, lead to an increased efficiency of treatment constitutes an important goal for biomedical applications. Using an original procedure whereby mutant genes are generated based on the use of conditional lentivector genome mobilisation, we recently described, for the first time, the identification of a human deoxycytidine kinase (dCK) mutant (G12) that sensitises a panel of cancer cell lines to treatment with the dCK analogue gemcitabine. Here, starting from the G12 variant itself, we generated a new library and identified a mutant (M36) that triggers even greater sensitisation to gemcitabine than G12. With respect to G12, M36 presents an additional mutation located in the region that constitutes the interface of the dCK dimer. The simple presence of this mutation halves both the IC50 and the proportion of residual cells resistant to the treatment. Furthermore, the use of vectors with self-inactivating LTRs leads to an increased sensitivity to treatment, a result compatible with a relief of the transcriptional interference exerted by the U3 promoter on the internal promoter that drives the expression of M36. Importantly, a remarkable effect is also observed in treatments with the anticancer compound cytarabine (AraC), for which a 10,000 fold decrease in IC50 occurred. By triggering the sensitisation of various cancer cell types with poor prognosis to two commonly used anticancer compounds M36 is a promising candidate for suicide gene approaches.}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Mutant human deoxycytidine kinase.}, author = {S Coulibaly and P Rossolillo and M Negroni}, isbn = {EP 15305545.4}, year = {2015}, date = {2015-01-01}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Purification of mRNA-programmed translation initiation complexes suitable for mass spectrometry analysis.}, author = {J Chicher and A Simonetti and L Kuhn and L Schaeffer and P Hammann and G Eriani and F Martin}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25914180?dopt=Abstract}, doi = {10.1002/pmic.201400628}, isbn = {25914180}, year = {2015}, date = {2015-01-01}, journal = {Proteomics}, volume = {15}, number = {14}, pages = {2417-2425}, abstract = {Liquid Chromatography coupled to tandem Mass Spectrometry (nanoLC-MS/MS) is a powerful analytical technique for the identification and mass analysis of complex protein mixtures. Here we present a combination of methods developed for the extensive/deep proteomic analysis of purified ribosome/mRNA particles assembled in rabbit reticulocyte lysate (RRL). Ribosomes are assembled on chimeric biotinylated mRNA-DNA molecules immobilized on streptavidin-coated beads and incubated with RRL to form initiation complexes. After washing steps, the complexes are trypsin-digested directly on the beads in semi-native condition or after their elution from the beads in denaturing Laemmli buffer. The nanoLC-MS/MS analysis performed on complexes assembled on β-globin, viral HCV and histone H4 mRNAs revealed significant differences in initiation factors composition in agreement with models of translation initiation used by these different types of mRNAs. Using Laemmli-denaturing condition induces release of deeply buried peptides from the ribosome and eukaryotic initiation factor 3 (eIF3) allowing the identification of the nearly complete set of ribosomal proteins. This article is protected by copyright. All rights reserved.}, keywords = {ERIANI, PPSE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 viral infectivity factor interacts with microtubule-associated protein light chain 3 and inhibits autophagy}, author = {S Borel and V Robert-Hebmann and J Alfaisal and A Jain and M Faure and L Espert and L Chaloin and J C Paillart and T Johansen and M Biard-Piechaczyk}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25490467?dopt=Abstract}, isbn = {25490467}, year = {2015}, date = {2015-01-01}, journal = {AIDS}, volume = {29}, number = {3}, pages = {275-286}, abstract = {Objective: Autophagy, an important anti-viral process triggered during HIV-1 entry by gp41-dependent membrane fusion, is repressed in infected CD4 T cells by an unknown mechanism. The aim of this study was to identify the role of Vif in the autophagy blockade. Design/methods: To determine the role of Vif in autophagy inhibition, we used cell lines that express CD4 and CXCR4 and primary CD4 T cells. Pull-down experiments, immunoprecipitation assays and computational analyses were performed to analyze the interaction between Vif and LC3B, a major autophagy component, in presence or absence of the anti-viral host factor APOBEC3G, after HIV-1 infection or ectopic expression of Vif. Autophagy was analyzed after infection by viruses expressing Vif (NL4.3) or not (NL4.3ΔVif), or after exogenous Vif expression. Results: We demonstrate that the C-terminal part of Vif interacts directly with LC3B, independently of the presence of APOBEC3G. Vif binds to pro-LC3 and Atg4-cleaved LC3 forms, and glycine 120, the amino acid conjugated to phosphatidylethanolamine on autophagosomes, is required. Importantly, we evidence that Vif inhibits autophagy during HIV-1 infection. Indeed, autophagy is detected in target cells infected by NL4.3ΔVif, but prevented in cells infected by NL4.3. Furthermore, autophagy triggered in NL4.3ΔVif-infected cells is inhibited when Vif is expressed in trans but is still active when target cells express a mutant of Vif that binds weakly to LC3B. Conclusions: Our study unveils that Vif inhibits autophagy independently of its action on APOBEC3G and therefore suggest a new function of this viral protein in restricting innate anti-viral mechanisms.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {miR-122 - a key factor and therapeutic target in liver disease.}, author = {S Bandiera and S Pfeffer and T F Baumert and M B Zeisel}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25308172?dopt=Abstract}, doi = {10.1016/j.jhep.2014.10.004}, isbn = {25308172}, year = {2015}, date = {2015-01-01}, journal = {J Hepatol}, volume = {62}, number = {2}, pages = {448-457}, abstract = {Being the largest internal organ of the human body with the unique ability of self-regeneration, the liver is involved in a wide variety of vital functions that require highly orchestrated and controlled biochemical processes. Increasing evidence suggests that microRNA (miRNA) are essential for the regulation of liver development, regeneration and metabolic functions. Hence, alterations in intrahepatic miRNA networks have been associated with liver disease including hepatitis, steatosis, cirrhosis and hepatocellular carcinoma (HCC). miR-122 is the most frequent miRNA in the adult liver, and a central player in liver biology and disease. Furthermore, miR-122 has been shown to be an essential host factor for hepatitis C virus (HCV) infection and an antiviral target complementary to standard of care using direct-acting antivirals or interferon-based treatment. This review summarizes our current understanding of the key role of miR-122 in liver physiology and disease highlighting its role in HCC and viral hepatitis. We also discuss the perspectives of miRNA-based therapeutic approaches for viral hepatitis and liver disease.}, keywords = {HBV HCC HCV Hepatitis Liver disease pathogenesis miR-122, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Elaborate uORF/IRES features control expression and localization of human glycyl-tRNA synthetase.}, author = {J Alexandrova and C Paulus and J Rudinger-Thirion and F Jossinet and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26327585?dopt=Abstract}, doi = {10.1080/15476286.2015.1086866}, isbn = {26327585}, year = {2015}, date = {2015-01-01}, journal = {RNA Biol}, volume = {12}, number = {12}, pages = {1301-1313}, abstract = {The canonical activity of glycyl-tRNA synthetase (GARS) is to charge glycine onto its cognate tRNAs. However, outside translation, GARS also participates in many other functions. A single gene encodes both the cytosolic and mitochondrial forms of GARS but two mRNA isoforms were identified. Using immunolocalization assays, in vitro translation assays and bicistronic constructs we provide experimental evidence that one of these mRNAs tightly controls expression and localization of human GARS. An intricate regulatory domain was found in its 5'-UTR which displays a functional Internal Ribosome Entry Site and an upstream Open Reading Frame. Together, these elements hinder the synthesis of the mitochondrial GARS and target the translation of the cytosolic enzyme to ER-bound ribosomes. This finding reveals a complex picture of GARS translation and localization in mammals. In this context, we discuss how human GARS expression could influence its moonlighting activities and its involvement in diseases.}, keywords = {Aminoacyl-tRNA synthetase IRES post-transcriptional control uORF, FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{jacquemin_ox40_2015, title = {OX40 Ligand Contributes to Human Lupus Pathogenesis by Promoting Ŧ Follicular Helper Response}, author = {Clément Jacquemin and Nathalie Schmitt and Cécile Contin-Bordes and Yang Liu and Priya Narayanan and Julien Seneschal and Typhanie Maurouard and David Dougall and Emily Spence Davizon and Hélène Dumortier and Isabelle Douchet and Loïc Raffray and Christophe Richez and Estibaliz Lazaro and Pierre Duffau and Marie-Elise Truchetet and Liliane Khoryati and Patrick Mercié and Lionel Couzi and Pierre Merville and Thierry Schaeverbeke and Jean-François Viallard and Jean-Luc Pellegrin and Jean-François Moreau and Sylviane Muller and Sandy Zurawski and Robert L Coffman and Virginia Pascual and Hideki Ueno and Patrick Blanco}, doi = {10.1016/j.immuni.2015.05.012}, issn = {1097-4180}, year = {2015}, date = {2015-01-01}, journal = {Immunity}, volume = {42}, number = {6}, pages = {1159--1170}, abstract = {Increased activity of T follicular helper (Tfh) cells plays a major pathogenic role in systemic lupus erythematosus (SLE). However, the mechanisms that cause aberrant Tfh cell responses in SLE remain elusive. Here we showed the OX40 ligand (OX40L)-OX40 axis contributes to the aberrant Tfh response in SLE. OX40L was expressed by myeloid antigen-presenting cells (APCs), but not B cells, in blood and in inflamed tissues in adult and pediatric SLE patients. The frequency of circulating OX40L-expressing myeloid APCs positively correlated with disease activity and the frequency of ICOS(+) blood Tfh cells in SLE. OX40 signals promoted naive and memory CD4(+) T cells to express multiple Tfh cell molecules and were sufficient to induce them to become functional B cell helpers. Immune complexes containing RNA induced OX40L expression on myeloid APCs via TLR7 activation. Our study provides a rationale to target the OX40L-OX40 axis as a therapeutic modality for SLE.}, keywords = {Adolescent, Adult, Aged, Antigen Presentation, B-Lymphocytes, Cell Differentiation, Cells, Cultured, Cytokines, Disease Progression, Dumortier, Female, Helper-Inducer, Humans, I2CT, Immunologic Memory, Inducible T-Cell Co-Stimulator Protein, Lupus Erythematosus, Lymphocyte Activation, Male, Middle Aged, Molecular Targeted Therapy, Myeloid Cells, OX40, OX40 Ligand, Receptors, RNA, Signal Transduction, Systemic, T-Lymphocytes, Team-Dumortier, Toll-Like Receptor 7, Young Adult}, pubstate = {published}, tppubtype = {article} } @article{marchesan_winding_2015, title = {The winding road for carbon nanotubes in nanomedicine}, author = {Silvia Marchesan and Kostas Kostarelos and Alberto Bianco and Maurizio Prato}, url = {http://www.sciencedirect.com/science/article/pii/S1369702114002594}, doi = {10.1016/j.mattod.2014.07.009}, issn = {1369-7021}, year = {2015}, date = {2015-01-01}, urldate = {2020-04-01}, journal = {Materials Today}, volume = {18}, number = {1}, pages = {12--19}, abstract = {Carbon nanotubes (CNTs) are recognized as promising nanomaterials for technological advancement. However, the stigma of structural similarity with asbestos fibers has slowed down progress of CNTs in nanomedicine. Nevertheless, it also prompted thorough studies that have revealed that functionalized CNTs (fCNTs) can biologically behave in a very different and safer manner. Here we review pristine and fCNT fate in biological settings, focusing on the importance of protein interaction, formation of the protein corona, and modulation of immune response. The emerging consensus on the desirable fCNT properties to achieve immunological neutrality, and even biodegradation, shows great promise for CNT adoption in medicine.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{karttunen_oxygen-dependent_2015, title = {Oxygen-dependent hydroxylation by FIH regulates the TRPV3 ion channel.}, author = {Sarah Karttunen and Michael Duffield and Nathan R Scrimgeour and Lauren Squires and Wai Li Lim and Mark L Dallas and Jason L Scragg and Johana Chicher and Keyur A Dave and Murray L Whitelaw and Chris Peers and Jeffrey J Gorman and Jonathan M Gleadle and Grigori Y Rychkov and Daniel J Peet}, doi = {10.1242/jcs.158451}, issn = {1477-9137 0021-9533}, year = {2015}, date = {2015-01-01}, journal = {Journal of cell science}, volume = {128}, number = {2}, pages = {225--231}, abstract = {Factor inhibiting HIF (FIH, also known as HIF1AN) is an oxygen-dependent asparaginyl hydroxylase that regulates the hypoxia-inducible factors (HIFs). Several proteins containing ankyrin repeat domains (ARDs) have been characterised as substrates of FIH, although there is little evidence for a functional consequence of hydroxylation on these substrates. This study demonstrates that the transient receptor potential vanilloid 3 (TRPV3) channel is hydroxylated by FIH on asparagine 242 within the cytoplasmic ARD. Hypoxia, FIH inhibitors and mutation of asparagine 242 all potentiated TRPV3-mediated current, without altering TRPV3 protein levels, indicating that oxygen-dependent hydroxylation inhibits TRPV3 activity. This novel mechanism of channel regulation by oxygen-dependent asparaginyl hydroxylation is likely to extend to other ion channels.}, note = {Place: England}, keywords = {alpha Subunit/genetics/*metabolism, Amino Acid Sequence, Ankyrin Repeat/genetics, Cell Hypoxia/*genetics, FIH, HEK293 Cells, Humans, Hydroxylation, Hydroxylation/genetics, Hypoxia, Hypoxia-Inducible Factor 1, Mixed Function Oxygenases/antagonists & inhibitors/genetics/*metabolism, Mutation, Oxygen/metabolism, PPSE, Protein Binding, Repressor Proteins/antagonists & inhibitors/genetics/*metabolism, TRPV Cation Channels/genetics/*metabolism, TRPV3}, pubstate = {published}, tppubtype = {article} } @article{weber-lotfi_nucleic_2015, title = {Nucleic acid import into mitochondria: New insights into the translocation pathways.}, author = {Frédérique Weber-Lotfi and Milana V Koulintchenko and Noha Ibrahim and Philippe Hammann and Daria V Mileshina and Yuri M Konstantinov and André Dietrich}, doi = {10.1016/j.bbamcr.2015.09.011}, issn = {0006-3002 0006-3002}, year = {2015}, date = {2015-01-01}, journal = {Biochimica et biophysica acta}, volume = {1853}, number = {12}, pages = {3165--3181}, abstract = {Mitochondria have retained indispensable but limited genetic information and they import both proteins and nucleic acids from the cytosol. RNA import is essential for gene expression and regulation, whereas competence for DNA uptake is likely to contribute to organellar genome dynamics and evolution. Contrary to protein import mechanisms, the way nucleic acids cross the mitochondrial membranes remains poorly understood. Using proteomic, genetic and biochemical approaches with both plant and yeast organelles, we develop here a model for DNA uptake into mitochondria. The first step includes the voltage-dependent anion channel and an outer membrane-located precursor fraction of a protein normally located in the inner membrane. To proceed, the DNA is then potentially recruited in the intermembrane space by an accessible subunit of one of the respiratory chain complexes. Final translocation through the inner membrane remains the most versatile but points to the components considered to make the mitochondrial permeability transition pore. Depending on the size, DNA and RNA cooperate or compete for mitochondrial uptake, which shows that they share import mechanisms. On the other hand, our results imply the existence of more than one route for nucleic acid translocation into mitochondria.}, note = {Place: Netherlands}, keywords = {Arabidopsis/metabolism, Biological Transport, Import factor, mitochondria, Mitochondria/*metabolism, Nucleic acid transport, Nucleic Acids/*metabolism, Permeability transition pore, Plant, PPSE, Saccharomyces cerevisiae/metabolism, Yeast}, pubstate = {published}, tppubtype = {article} } @article{lezot_skeletal_2015, title = {Skeletal consequences of RANKL-blocking antibody (IK22-5) injections during growth: mouse strain disparities and synergic effect with zoledronic acid}, author = {Frédéric Lézot and Julie Chesneau and Benjamin Navet and Bérengère Gobin and Jérome Amiaud and YongWon Choi and Hideo Yagita and Beatriz Castaneda and Ariane Berdal and Christopher G Mueller and Françoise Rédini and Dominique Heymann}, doi = {10.1016/j.bone.2014.12.011}, issn = {1873-2763}, year = {2015}, date = {2015-01-01}, journal = {Bone}, volume = {73}, pages = {51--59}, abstract = {High doses of bone resorption inhibitors are currently under evaluation in pediatric oncology. Previous works have evidenced transient arrest in long bone and skull bone growth and tooth eruption blockage when mice were treated with zoledronic acid (ZOL). The question of potential similar effects with a RANKL-blocking antibody (IK22.5) was raised. Sensitivity disparities in these inhibitors between mouse strains and synergic effects of zoledronic acid and a RANKL-blocking antibody were subsidiary questions. In order to answer these questions, newborn C57BL/6J and CD1 mice were injected every two or three days (4 injections in total so 7 or 10 days of treatment length) with high doses of a RANKL-blocking antibody. The consequences on the tibia, craniofacial bones and teeth were analyzed by μCT and histology at the end of the treatment and one, two and three months later. The results obtained showed that RANKL-blocking antibody injections induced a transient arrest of tibia and skull bone growth and an irreversible blockage of tooth eruption in C57BL/6J mice. In CD1 mice, tooth eruption defects were also present but only at much higher doses. Similar mouse strain differences were obtained with zoledronic acid. Finally, a synergic effect of the two inhibitors was evidenced. In conclusion as previously observed for bisphosphonates (ZOL), a RANKL-blocking antibody induced a transient arrest in long bone and skull bone growth and a blockage of tooth eruption with however disparities between mouse strains with regard to this last effect. A synergic effect of both bone resorption inhibitors was also demonstrated.}, keywords = {Animals, Antibodies, Bone Density Conservation Agents, Bone Development, Bone resorption, Diphosphonates, Female, Imidazoles, Inbred C57BL, Mice, Newborn, Pregnancy, RANK ligand, RANKL, Side effect, Skeleton growth, Team-Mueller, Tooth Eruption, Zoledronic acid}, pubstate = {published}, tppubtype = {article} } @article{chtarbanova_drosophila_2014, title = {Drosophila C virus systemic infection leads to intestinal obstruction}, author = {Stanislava Chtarbanova and Olivier Lamiable and Kwang-Zin Lee and Delphine Galiana and Laurent Troxler and Carine Meignin and Charles Hetru and Jules A Hoffmann and Laurent Daeffler and Jean-Luc Imler}, url = {http://jvi.asm.org/content/88/24/14057}, doi = {10.1128/JVI.02320-14}, issn = {1098-5514}, year = {2014}, date = {2014-12-01}, journal = {Journal of Virology}, volume = {88}, number = {24}, pages = {14057--14069}, abstract = {Drosophila C virus (DCV) is a positive-sense RNA virus belonging to the Dicistroviridae family. This natural pathogen of the model organism Drosophila melanogaster is commonly used to investigate antiviral host defense in flies, which involves both RNA interference and inducible responses. Although lethality is used routinely as a readout for the efficiency of the antiviral immune response in these studies, virus-induced pathologies in flies still are poorly understood. Here, we characterize the pathogenesis associated with systemic DCV infection. Comparison of the transcriptome of flies infected with DCV or two other positive-sense RNA viruses, Flock House virus and Sindbis virus, reveals that DCV infection, unlike those of the other two viruses, represses the expression of a large number of genes. Several of these genes are expressed specifically in the midgut and also are repressed by starvation. We show that systemic DCV infection triggers a nutritional stress in Drosophila which results from intestinal obstruction with the accumulation of peritrophic matrix at the entry of the midgut and the accumulation of the food ingested in the crop, a blind muscular food storage organ. The related virus cricket paralysis virus (CrPV), which efficiently grows in Drosophila, does not trigger this pathology. We show that DCV, but not CrPV, infects the smooth muscles surrounding the crop, causing extensive cytopathology and strongly reducing the rate of contractions. We conclude that the pathogenesis associated with systemic DCV infection results from the tropism of the virus for an important organ within the foregut of dipteran insects, the crop. IMPORTANCE: DCV is one of the few identified natural viral pathogens affecting the model organism Drosophila melanogaster. As such, it is an important virus for the deciphering of host-virus interactions in insects. We characterize here the pathogenesis associated with DCV infection in flies and show that it results from the tropism of the virus for an essential but poorly characterized organ in the digestive tract, the crop. Our results may have relevance for other members of the Dicistroviridae, some of which are pathogenic to beneficial or pest insect species.}, keywords = {Animals, bioinformatic, Dicistroviridae, Female, Gastrointestinal Tract, Gene Expression Profiling, hoffmann, imler, Intestinal Obstruction, M3i, meignin, Muscle, Nodaviridae, Sindbis Virus, Smooth, Viral Tropism}, pubstate = {published}, tppubtype = {article} } @article{poirier_effect_2014, title = {The effect of cold stress on the proteome of the marine bacterium Pseudomonas fluorescens BA3SM1 and its ability to cope with metal excess.}, author = {Isabelle Poirier and Lauriane Kuhn and Christelle Caplat and Philippe Hammann and Martine Bertrand}, doi = {10.1016/j.aquatox.2014.10.002}, issn = {1879-1514 0166-445X}, year = {2014}, date = {2014-12-01}, journal = {Aquatic toxicology (Amsterdam, Netherlands)}, volume = {157}, pages = {120--133}, abstract = {This study examined the effect of cold stress on the proteome and metal tolerance of Pseudomonas fluorescens BA3SM1, a marine strain isolated from tidal flat sediments. When cold stress (+10 °C for 36 h) was applied before moderate metal stress (0.4 mM Cd, 0.6 mM Cd, 1.5 mM Zn, and 1.5 mM Cu), growth disturbances induced by metal, in comparison with respective controls, were reduced for Cd and Zn while they were pronounced for Cu. This marine strain was able to respond to cold stress through a number of changes in protein regulation. Analysis of the predicted differentially expressed protein functions demonstrated that some mechanisms developed under cold stress were similar to those developed in response to Cd, Zn, and Cu. Therefore, pre-cold stress could help this strain to better counteract toxicity of moderate concentrations of some metals. P. fluorescens BA3SM1 was able to remove up to 404.3 mg Cd/g dry weight, 172.5 mg Zn/g dry weight, and 11.3 mg Cu/g dry weight and its metal biosorption ability seemed to be related to the bacterial growth phase. Thus, P. fluorescens BA3SM1 appears as a promising agent for bioremediation processes, even at low temperatures.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{frechin_expression_2014, title = {Expression of nuclear and mitochondrial genes encoding ATP synthase is synchronized by disassembly of a multisynthetase complex.}, author = {Mathieu Frechin and Ludovic Enkler and Emmanuel Tetaud and Daphné Laporte and Bruno Senger and Corinne Blancard and Philippe Hammann and Gaétan Bader and Sandra Clauder-Münster and Lars M Steinmetz and Robert Pierre Martin and Jean-Paul di Rago and Hubert Dominique Becker}, doi = {10.1016/j.molcel.2014.10.015}, issn = {1097-4164 1097-2765}, year = {2014}, date = {2014-12-01}, journal = {Molecular cell}, volume = {56}, number = {6}, pages = {763--776}, abstract = {In eukaryotic cells, oxidative phosphorylation involves multisubunit complexes of mixed genetic origin. Assembling these complexes requires an organelle-independent synchronizing system for the proper expression of nuclear and mitochondrial genes. Here we show that proper expression of the F1FO ATP synthase (complex V) depends on a cytosolic complex (AME) made of two aminoacyl-tRNA synthetases (cERS and cMRS) attached to an anchor protein, Arc1p. When yeast cells adapt to respiration the Snf1/4 glucose-sensing pathway inhibits ARC1 expression triggering simultaneous release of cERS and cMRS. Free cMRS and cERS relocate to the nucleus and mitochondria, respectively, to synchronize nuclear transcription and mitochondrial translation of ATP synthase genes. Strains releasing asynchronously the two aminoacyl-tRNA synthetases display aberrant expression of nuclear and mitochondrial genes encoding subunits of complex V resulting in severe defects of the oxidative phosphorylation mechanism. This work shows that the AME complex coordinates expression of enzymes that require intergenomic control.}, note = {Place: United States}, keywords = {Cell Nucleus/genetics, Fungal, Gene Expression, Gene Expression Regulation, Mitochondria/genetics, Multienzyme Complexes, PPSE, Protein Multimerization, Proton-Translocating ATPases/*genetics/metabolism, RNA-Binding Proteins/physiology, Saccharomyces cerevisiae Proteins/physiology, Saccharomyces cerevisiae/enzymology/*genetics}, pubstate = {published}, tppubtype = {article} } @article{bonnay_akirin_2014, title = {Akirin specifies NF-κB selectivity of Drosophila innate immune response via chromatin remodeling}, author = {François Bonnay and Xuan-Hung Nguyen and Eva Cohen-Berros and Laurent Troxler and Eric Batsche and Jacques Camonis and Osamu Takeuchi and Jean-Marc Reichhart and Nicolas Matt}, doi = {10.15252/embj.201488456}, issn = {1460-2075}, year = {2014}, date = {2014-10-01}, journal = {EMBO J.}, volume = {33}, number = {20}, pages = {2349--2362}, abstract = {The network of NF-κB-dependent transcription that activates both pro- and anti-inflammatory genes in mammals is still unclear. As NF-κB factors are evolutionarily conserved, we used Drosophila to understand this network. The NF-κB transcription factor Relish activates effector gene expression following Gram-negative bacterial immune challenge. Here, we show, using a genome-wide approach, that the conserved nuclear protein Akirin is a NF-κB co-factor required for the activation of a subset of Relish-dependent genes correlating with the presence of H3K4ac epigenetic marks. A large-scale unbiased proteomic analysis revealed that Akirin orchestrates NF-κB transcriptional selectivity through the recruitment of the Osa-containing-SWI/SNF-like Brahma complex (BAP). Immune challenge in Drosophila shows that Akirin is required for the transcription of a subset of effector genes, but dispensable for the transcription of genes that are negative regulators of the innate immune response. Therefore, Akirins act as molecular selectors specifying the choice between subsets of NF-κB target genes. The discovery of this mechanism, conserved in mammals, paves the way for the establishment of more specific and less toxic anti-inflammatory drugs targeting pro-inflammatory genes.}, keywords = {Animals, bioinformatic, Cell Cycle Proteins, Chromatin Assembly and Disassembly, chromatin remodeling, DNA-Binding Proteins, Female, Genetic, Immunity, Innate, Innate immune response, M3i, Male, matt, Mutation, NF-kappa B, NF‐κB, Promoter Regions, proteomics, reichhart, Trans-Activators, Transcription Factors, Transcriptional Activation, Two-Hybrid System Techniques}, pubstate = {published}, tppubtype = {article} } @article{tartey_akirin2_2014, title = {Akirin2 is critical for inducing inflammatory genes by bridging IκB-ζ and the SWI/SNF complex}, author = {Sarang Tartey and Kazufumi Matsushita and Alexis Vandenbon and Daisuke Ori and Tomoko Imamura and Takashi Mino and Daron M Standley and Jules A Hoffmann and Jean-Marc Reichhart and Shizuo Akira and Osamu Takeuchi}, doi = {10.15252/embj.201488447}, issn = {1460-2075}, year = {2014}, date = {2014-10-01}, journal = {EMBO J.}, volume = {33}, number = {20}, pages = {2332--2348}, abstract = {Transcription of inflammatory genes in innate immune cells is coordinately regulated by transcription factors, including NF-κB, and chromatin modifiers. However, it remains unclear how microbial sensing initiates chromatin remodeling. Here, we show that Akirin2, an evolutionarily conserved nuclear protein, bridges NF-κB and the chromatin remodeling SWI/SNF complex by interacting with BRG1-Associated Factor 60 (BAF60) proteins as well as IκB-ζ, which forms a complex with the NF-κB p50 subunit. These interactions are essential for Toll-like receptor-, RIG-I-, and Listeria-mediated expression of proinflammatory genes including Il6 and Il12b in macrophages. Consistently, effective clearance of Listeria infection required Akirin2. Furthermore, Akirin2 and IκB-ζ recruitment to the Il6 promoter depend upon the presence of IκB-ζ and Akirin2, respectively, for regulation of chromatin remodeling. BAF60 proteins were also essential for the induction of Il6 in response to LPS stimulation. Collectively, the IκB-ζ-Akirin2-BAF60 complex physically links the NF-κB and SWI/SNF complexes in innate immune cell activation. By recruiting SWI/SNF chromatin remodellers to IκB-ζ, transcriptional coactivator for NF-κB, the conserved nuclear protein Akirin2 stimulates pro-inflammatory gene promoters in mouse macrophages during innate immune responses to viral or bacterial infection.}, keywords = {Adaptor Proteins, Animals, Cell Nucleus, Chromatin Assembly and Disassembly, chromatin remodeling, Chromosomal Proteins, cytokine, Cytokines, Female, Gene Expression Regulation, gene regulation, Genetic, hoffmann, Humans, Immunity, Innate, innate immunity, Knockout, Listeria monocytogenes, M3i, Macrophages, Male, Mice, Multiprotein Complexes, Non-Histone, Nuclear Proteins, Promoter Regions, Protein Binding, reichhart, Repressor Proteins, Sequence Deletion, Signal Transducing, Transcriptional Activation}, pubstate = {published}, tppubtype = {article} } @article{amcheslavsky_enteroendocrine_2014b, title = {Enteroendocrine cells support intestinal stem-cell-mediated homeostasis in Drosophila}, author = {Alla Amcheslavsky and Wei Song and Qi Li and Yingchao Nie and Ivan Bragatto and Dominique Ferrandon and Norbert Perrimon and Tony Y Ip}, doi = {10.1016/j.celrep.2014.08.052}, issn = {2211-1247}, year = {2014}, date = {2014-10-01}, journal = {Cell Rep}, volume = {9}, number = {1}, pages = {32--39}, abstract = {Intestinal stem cells in the adult Drosophila midgut are regulated by growth factors produced from the surrounding niche cells including enterocytes and visceral muscle. The role of the other major cell type, the secretory enteroendocrine cells, in regulating intestinal stem cells remains unclear. We show here that newly eclosed scute loss-of-function mutant flies are completely devoid of enteroendocrine cells. These enteroendocrine cell-less flies have normal ingestion and fecundity but shorter lifespan. Moreover, in these newly eclosed mutant flies, the diet-stimulated midgut growth that depends on the insulin-like peptide 3 expression in the surrounding muscle is defective. The depletion of Tachykinin-producing enteroendocrine cells or knockdown of Tachykinin leads to a similar although less severe phenotype. These results establish that enteroendocrine cells serve as an important link between diet and visceral muscle expression of an insulin-like growth factor to stimulate intestinal stem cell proliferation and tissue growth.}, keywords = {Animals, Cell Differentiation, Enterocytes, Enteroendocrine Cells, Female, ferrandon, Homeostasis, Intestines, M3i, Male, Stem Cells, Tachykinins}, pubstate = {published}, tppubtype = {article} } @inbook{P2014, title = {Transgenic insects: techniques and applications}, author = {P Gabrieli and Eric Marois and Flaminia Catteruccia}, editor = {MQ Benedicts}, year = {2014}, date = {2014-10-01}, pages = {188-207}, publisher = {CABI}, chapter = {Sexual sterilization of mosquitoes}, keywords = {insects, M3i, marois, sterile}, pubstate = {published}, tppubtype = {inbook} } @article{flacher_murine_2014, title = {Murine Langerin+ dermal dendritic cells prime CD8+ Ŧ cells while Langerhans cells induce cross-tolerance}, author = {Vincent Flacher and Christoph H Tripp and David G Mairhofer and Ralph M Steinman and Patrizia Stoitzner and Juliana Idoyaga and Nikolaus Romani}, doi = {10.15252/emmm.201303283}, issn = {1757-4684}, year = {2014}, date = {2014-09-01}, journal = {EMBO molecular medicine}, volume = {6}, number = {9}, pages = {1191--1204}, abstract = {Skin dendritic cells (DCs) control the immunogenicity of cutaneously administered vaccines. Antigens targeted to DCs via the C-type lectin Langerin/CD207 are cross-presented to CD8(+) T cells in vivo. We investigated the relative roles of Langerhans cells (LCs) and Langerin(+) dermal DCs (dDCs) in different vaccination settings. Poly(I:C) and anti-CD40 agonist antibody promoted cytotoxic responses upon intradermal immunization with ovalbumin (OVA)-coupled anti-Langerin antibodies (Langerin/OVA). This correlated with CD70 upregulation in Langerin(+) dDCs, but not LCs. In chimeric mice where Langerin targeting was restricted to dDCs, CD8(+) T-cell memory was enhanced. Conversely, providing Langerin/OVA exclusively to LCs failed to prime cytotoxicity, despite initial antigen cross-presentation to CD8(+) T cells. Langerin/OVA combined with imiquimod could not prime CD8(+) T cells and resulted in poor cytotoxicity in subsequent responses. This tolerance induction required targeting and maturation of LCs. Altogether, Langerin(+) dDCs prime long-lasting cytotoxic responses, while cross-presentation by LCs negatively influences CD8(+) T-cell priming. Moreover, this highlights that DCs exposed to TLR agonists can still induce tolerance and supports the existence of qualitatively different DC maturation programs.}, keywords = {agonists, Animals, Antibodies, antibody, Antigen, Antigen Presentation, Antigens, C-Type, C-type lectin, cancer, CD70, CD8-Positive T-Lymphocytes, CD8+ T cells, CD8+ T‐cell responses, Cellular, CROSS-PRESENTATION, Cross-Priming, Cytotoxicity, Dendritic Cells, DERMAL DENDRITIC CELLS, DERMATOLOGY, disease, imiquimod, Immunization, IMMUNOGENICITY, Immunologic Memory, Immunological, Immunology, In vivo, Inbred C57BL, INDUCTION, Intradermal, Langerhans Cells, LECTIN, Lectins, Mannose-Binding Lectins, Maturation, Mice, Models, murine, OVALBUMIN, physiology, priming, RESPONSES, Skin, Surface, T CELLS, T-CELLS, Team-Mueller, tolerance, Vaccination, vaccine, Vaccines}, pubstate = {published}, tppubtype = {article} } @article{lamiable_induced_2014, title = {Induced antiviral innate immunity in Drosophila}, author = {Olivier Lamiable and Jean-Luc Imler}, doi = {10.1016/j.mib.2014.05.006}, issn = {1879-0364}, year = {2014}, date = {2014-08-01}, journal = {Current Opinion in Microbiology}, volume = {20}, pages = {62--68}, abstract = {Immunity to viral infections in the model organism Drosophila melanogaster involves both RNA interference and additional induced responses. The latter include not only cellular mechanisms such as programmed cell death and autophagy, but also the induction of a large set of genes, some of which contribute to the control of viral replication and resistance to infection. This induced response to infection is complex and involves both virus-specific and cell-type specific mechanisms. We review here recent developments, from the sensing of viral infection to the induction of signaling pathways and production of antiviral effector molecules. Our current understanding, although still partial, validates the Drosophila model of antiviral induced immunity for insect pests and disease vectors, as well as for mammals.}, keywords = {Animals, Gene Expression Regulation, Host-Pathogen Interactions, imler, Immunity, Innate, M3i, RNA Viruses, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{lange_rna_2014, title = {The RNA helicases AtMTR4 and HEN2 target specific subsets of nuclear transcripts for degradation by the nuclear exosome in Arabidopsis thaliana.}, author = {Heike Lange and Hélène Zuber and François M Sement and Johana Chicher and Lauriane Kuhn and Philippe Hammann and Véronique Brunaud and Caroline Bérard and Nathalie Bouteiller and Sandrine Balzergue and Sébastien Aubourg and Marie-Laure Martin-Magniette and Hervé Vaucheret and Dominique Gagliardi}, doi = {10.1371/journal.pgen.1004564}, issn = {1553-7404 1553-7390 1553-7390}, year = {2014}, date = {2014-08-01}, journal = {PLoS genetics}, volume = {10}, number = {8}, pages = {e1004564}, abstract = {The RNA exosome is the major 3'-5' RNA degradation machine of eukaryotic cells and participates in processing, surveillance and turnover of both nuclear and cytoplasmic RNA. In both yeast and human, all nuclear functions of the exosome require the RNA helicase MTR4. We show that the Arabidopsis core exosome can associate with two related RNA helicases, AtMTR4 and HEN2. Reciprocal co-immunoprecipitation shows that each of the RNA helicases co-purifies with the exosome core complex and with distinct sets of specific proteins. While AtMTR4 is a predominantly nucleolar protein, HEN2 is located in the nucleoplasm and appears to be excluded from nucleoli. We have previously shown that the major role of AtMTR4 is the degradation of rRNA precursors and rRNA maturation by-products. Here, we demonstrate that HEN2 is involved in the degradation of a large number of polyadenylated nuclear exosome substrates such as snoRNA and miRNA precursors, incompletely spliced mRNAs, and spurious transcripts produced from pseudogenes and intergenic regions. Only a weak accumulation of these exosome substrate targets is observed in mtr4 mutants, suggesting that MTR4 can contribute, but plays rather a minor role for the degradation of non-ribosomal RNAs and cryptic transcripts in Arabidopsis. Consistently, transgene post-transcriptional gene silencing (PTGS) is marginally affected in mtr4 mutants, but increased in hen2 mutants, suggesting that it is mostly the nucleoplasmic exosome that degrades aberrant transgene RNAs to limit their entry in the PTGS pathway. Interestingly, HEN2 is conserved throughout green algae, mosses and land plants but absent from metazoans and other eukaryotic lineages. Our data indicate that, in contrast to human and yeast, plants have two functionally specialized RNA helicases that assist the exosome in the degradation of specific nucleolar and nucleoplasmic RNA populations, respectively.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{goto_chromatin_2014, title = {The chromatin regulator DMAP1 modulates activity of the nuclear factor B (NF-B) transcription factor Relish in the Drosophila innate immune response}, author = {Akira Goto and Hidehiro Fukuyama and Jean-Luc Imler and Jules A Hoffmann}, doi = {10.1074/jbc.C114.553719}, issn = {1083-351X}, year = {2014}, date = {2014-07-01}, journal = {The Journal of Biological Chemistry}, volume = {289}, number = {30}, pages = {20470--20476}, abstract = {The host defense of the model organism Drosophila is under the control of two major signaling cascades controlling transcription factors of the NF-B family, the Toll and the immune deficiency (IMD) pathways. The latter shares extensive similarities with the mammalian TNF-R pathway and was initially discovered for its role in anti-Gram-negative bacterial reactions. A previous interactome study from this laboratory reported that an unexpectedly large number of proteins are binding to the canonical components of the IMD pathway. Here, we focus on DNA methyltransferase-associated protein 1 (DMAP1), which this study identified as an interactant of Relish, a Drosophila transcription factor reminiscent of the mammalian p105 NF-B protein. We show that silencing of DMAP1 expression both in S2 cells and in flies results in a significant reduction of Escherichia coli-induced expression of antimicrobial peptides. Epistatic analysis indicates that DMAP1 acts in parallel or downstream of Relish. Co-immunoprecipitation experiments further reveal that, in addition to Relish, DMAP1 also interacts with Akirin and the Brahma-associated protein 55 kDa (BAP55). Taken together, these results reveal that DMAP1 is a novel nuclear modulator of the IMD pathway, possibly acting at the level of chromatin remodeling.}, keywords = {Animals, Cell Line, Chromatin Assembly and Disassembly, Epistasis, Escherichia coli, Escherichia coli Infections, Genetic, hoffmann, imler, Immunity, Innate, M3i, NF-kappa B, Repressor Proteins, Signal Transduction, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{F2014, title = {Evidence of natural Wolbachia infections in field populations of Anopheles gambiae}, author = {Francesco Baldini and N Segata and Julien Pompon and P Marcenac and W R Shaw and R Dabire and A Diabate and Elena A Levashina and Flaminia Catteruccia}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24905191}, year = {2014}, date = {2014-06-06}, journal = {Nature Commun.}, volume = {5}, pages = {3985}, abstract = {Wolbachia are maternally transmitted intracellular bacteria that invade insect populations by manipulating their reproduction and immunity and thus limiting the spread of numerous human pathogens. Experimental Wolbachia infections can reduce Plasmodium numbers in Anopheles mosquitoes in the laboratory, however, natural Wolbachia infections in field anophelines have never been reported. Here we show evidence of Wolbachia infections in Anopheles gambiae in Burkina Faso, West Africa. Sequencing of the 16S rRNA gene identified Wolbachia sequences in both female and male germlines across two seasons, and determined that these sequences are vertically transmitted from mother to offspring. Whole-genome sequencing of positive samples suggests that the genetic material identified in An. gambiae belongs to a novel Wolbachia strain, related to but distinct from strains infecting other arthropods. The evidence of Wolbachia infections in natural Anopheles populations promotes further investigations on the possible use of natural Wolbachia-Anopheles associations to limit malaria transmission.}, keywords = {Anopheles gambiae, field, Wolbachia}, pubstate = {published}, tppubtype = {article} } @inbook{Imler2014, title = {Le défi des maladies infectieuses}, author = {Jean-Luc Imler and Jules A Hoffmann}, editor = {Ph. Cramer collection Guérir et prévenir demain}, isbn = {978-2-85525-390-9}, year = {2014}, date = {2014-06-01}, pages = {167-174}, publisher = {Editions DOCIS}, chapter = {« L’immunité innée »}, abstract = {La lèpre fait des ravages dès l’Antiquité, les épidémies de peste tuent au Moyen Âge et celles de choléra dévastent l’Inde. La tuberculose émerge véritablement au XIXème siècle, la grippe espagnole a fait vingt millions de morts en 1918 et de nouvelles maladies apparaissent : les infections hospitalières, les hépatites, le SIDA, les fièvres hémorragiques, la légionellose,… Toutes ces maladies ont un point commun, ce sont des maladies infectieuses qui sont pour la médecine un vrai défi, tant elles sont dévastatrices. L’innovation a eu et continue à avoir un rôle essentiel dans la caractérisation de ces maladies, la découverte de l’agent responsable, leur traitement et leur prévention. Ce livre, dont les auteurs font partie des plus éminents spécialistes français et européens des maladies infectieuses décrit, de façon abordable par tous, aussi bien les découvertes et les inventions essentielles à ce domaine que les avancées médicales.}, keywords = {hoffmann, imler, innate immunity, M3i, maladies infectieuses}, pubstate = {published}, tppubtype = {inbook} } @article{F2014b, title = {Site-specific genetic engineering of the Anopheles gambiae Y chromosome}, author = {F Bernardini and R Galizi and M Menichelli and P A Papathanos and V Dritsou and Eric Marois and Andrea Crisanti and Nikolai Windbichler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24821795}, year = {2014}, date = {2014-05-27}, journal = {Proc Natl Acad Sci U S A.}, volume = {111}, number = {21}, pages = {7600-5}, abstract = {Despite its function in sex determination and its role in driving genome evolution, the Y chromosome remains poorly understood in most species. Y chromosomes are gene-poor, repeat-rich and largely heterochromatic and therefore represent a difficult target for genetic engineering. The Y chromosome of the human malaria vector Anopheles gambiae appears to be involved in sex determination although very little is known about both its structure and function. Here, we characterize a transgenic strain of this mosquito species, obtained by transposon-mediated integration of a transgene construct onto the Y chromosome. Using meganuclease-induced homologous repair we introduce a site-specific recombination signal onto the Y chromosome and show that the resulting docking line can be used for secondary integration. To demonstrate its utility, we study the activity of a germ-line-specific promoter when located on the Y chromosome. We also show that Y-linked fluorescent transgenes allow automated sex separation of this important vector species, providing the means to generate large single-sex populations. Our findings will aid studies of sex chromosome function and enable the development of male-exclusive genetic traits for vector control.}, keywords = {Anopheles gambiae, Biotechnology, M3i, marois, SIT, transgenesis}, pubstate = {published}, tppubtype = {article} } @article{pmid24122342, title = {The many lives of CTIP2: from AIDS to cancer and cardiac hypertrophy}, author = {Valentin Le Douce and Thomas Cherrier and Raphael Riclet and Olivier Rohr and Christian Schwartz}, doi = {10.1002/jcp.24490}, issn = {1097-4652}, year = {2014}, date = {2014-05-01}, urldate = {2014-05-01}, journal = {J Cell Physiol}, volume = {229}, number = {5}, pages = {533--537}, abstract = {CTIP2 is a key transcriptional regulator involved in numerous physiological functions. Initial works have shown the importance of CTIP2 in the establishment and persistence of HIV latency in microglial cells, the main latent/quiescent viral reservoir in the brain. Recent studies have highlighted the importance of CTIP2 in several other pathologies, such as cardiac hypertrophy and various types of human malignancies. Targeting CTIP2 may therefore constitute a new approach in the treatment of these pathologies.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid24623795, title = {HMGA1 recruits CTIP2-repressed P-TEFb to the HIV-1 and cellular target promoters}, author = {Sebastian Eilebrecht and Valentin Le Douce and Raphael Riclet and Brice Targat and Houda Hallay and Benot Van Driessche and Christian Schwartz and Gwenaëlle Robette and Carine Van Lint and Olivier Rohr and Arndt G Benecke}, doi = {10.1093/nar/gku168}, issn = {1362-4962}, year = {2014}, date = {2014-04-01}, urldate = {2014-04-01}, journal = {Nucleic Acids Res}, volume = {42}, number = {8}, pages = {4962--4971}, abstract = {Active positive transcription elongation factor b (P-TEFb) is essential for cellular and human immunodeficiency virus type 1 (HIV-1) transcription elongation. CTIP2 represses P-TEFb activity in a complex containing 7SK RNA and HEXIM1. Recently, the inactive 7SK/P-TEFb small nuclear RNP (snRNP) has been detected at the HIV-1 core promoter as well as at the promoters of cellular genes, but a recruiting mechanism still remains unknown to date. Here we show global synergy between CTIP2 and the 7SK-binding chromatin master-regulator HMGA1 in terms of P-TEFb-dependent endogenous and HIV-1 gene expression regulation. While CTIP2 and HMGA1 concordingly repress the expression of cellular 7SK-dependent P-TEFb targets, the simultaneous knock-down of CTIP2 and HMGA1 also results in a boost in Tat-dependent and independent HIV-1 promoter activity. Chromatin immunoprecipitation experiments reveal a significant loss of CTIP2/7SK/P-TEFb snRNP recruitment to cellular gene promoters and the HIV-1 promoter on HMGA1 knock-down. Our findings not only provide insights into a recruiting mechanism for the inactive 7SK/P-TEFb snRNP, but may also contribute to a better understanding of viral latency.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{imler_overview_2014, title = {Overview of Drosophila immunity: a historical perspective}, author = {Jean-Luc Imler}, doi = {10.1016/j.dci.2013.08.018}, issn = {1879-0089}, year = {2014}, date = {2014-01-01}, journal = {Developmental and Comparative Immunology}, volume = {42}, number = {1}, pages = {3--15}, abstract = {The functional analysis of genes from the model organism Drosophila melanogaster has provided invaluable information for many cellular and developmental or physiological processes, including immunity. The best-understood aspect of Drosophila immunity is the inducible humoral response, first recognized in 1972. This pioneering work led to a remarkable series of findings over the next 30 years, ranging from the identification and characterization of the antimicrobial peptides produced, to the deciphering of the signalling pathways activating the genes that encode them and, ultimately, to the discovery of the receptors sensing infection. These studies on an insect model coincided with a revival of the field of innate immunity, and had an unanticipated impact on the biomedical field.}, keywords = {Allergy and Immunology, Animal, Animals, Antimicrobial Cationic Peptides, Antimicrobial peptides, history, Humans, IMD pathway, imler, Immunity, Innate, innate immunity, M3i, Models, Pattern recognition receptors, Signal Transduction, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{leclerc_drosophila_2014, title = {Drosophila melanogaster model in innate immunity.}, author = {Vincent Leclerc and Laure El Chamy and Monde Ntwasa and Jean-Marc Reichhart}, year = {2014}, date = {2014-01-01}, journal = {Trends in Entomology}, volume = {10}, pages = {73--86}, abstract = {Due relative simplicity of the fly and availability of large amounts of genetic tools, Drosophila melanogaster has proven to be an excellent model to study the basic principles of innate immunity. This is illustrated by the discovery of the Toll-like receptor functions in pathogen sensing, recognised by the 2011 Nobel Prize in Physiology and Medicine awarded to Jules Hoffmann. Drosophila can also be used as an in vivo, genetically tractable model, to analyse various aspects of host-pathogen interactions including virulence factor mechanisms of action.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{haller_assessing_2014b, title = {Assessing Pseudomonas virulence with a nonmammalian host: Drosophila melanogaster}, author = {Samantha Haller and Stefanie Limmer and Dominique Ferrandon}, doi = {10.1007/978-1-4939-0473-0_56}, issn = {1940-6029}, year = {2014}, date = {2014-01-01}, journal = {Methods Mol. Biol.}, volume = {1149}, pages = {723--740}, abstract = {Drosophila melanogaster flies represent an interesting model to study host-pathogen interactions as: (1) they are cheap and easy to raise rapidly and do not bring up ethical issues, (2) available genetic tools are highly sophisticated, for instance allowing tissue-specific alteration of gene expression, e.g., of immune genes, (3) they have a relatively complex organization, with distinct digestive tract and body cavity in which local or systemic infections, respectively, take place, (4) a medium throughput can be achieved in genetic screens, for instance looking for Pseudomonas aeruginosa mutants with altered virulence. We present here the techniques used to investigate host-pathogen relationships, namely the two major models of infections as well as the relevant parameters used to monitor the infection (survival, bacterial titer, induction of host immune response).}, keywords = {Animal, Animals, Antimicrobial Cationic Peptides, Biological Assay, Colony Count, Disease Models, ferrandon, Hemolymph, Host-Pathogen Interactions, M3i, Mammals, Microbial, Pseudomonas aeruginosa, Pseudomonas Infections, Reverse Transcriptase Polymerase Chain Reaction, Virulence}, pubstate = {published}, tppubtype = {article} } @article{lestradet_drosophila_2014b, title = {Drosophila as a model for intestinal infections}, author = {Matthieu Lestradet and Kwan Zin Lee and Dominique Ferrandon}, doi = {10.1007/978-1-4939-1261-2_2}, issn = {1940-6029 (Electronic) 1064-3745 (Linking)}, year = {2014}, date = {2014-01-01}, journal = {Methods Mol Biol}, volume = {1197}, pages = {11--40}, abstract = {Drosophila melanogaster is a powerful model to study infections thanks to the power of its genetics and knowledge on its biology accumulated for over a century. While the systemic humoral immune response against invading microbes has been intensively studied in the past two decades, the study of intestinal infections is more recent. Here, we present the methods that are currently in use to probe various aspects of the host-pathogen interactions between Drosophila and ingested microbes, with an emphasis on the study of the midgut epithelium, which constitutes the major interface between the organism and the microbe-rich ingested food.}, keywords = {Animal, Animals, Bacterial Physiological Phenomena, Disease Models, ferrandon, Gastrointestinal Tract, Host-Pathogen Interactions, M3i}, pubstate = {published}, tppubtype = {article} } @article{schwarzmuller_systematic_2014b, title = {Systematic phenotyping of a large-scale Candida glabrata deletion collection reveals novel antifungal tolerance genes}, author = {Tobias Schwarzmüller and Biao Ma and Ekkehard Hiller and Fabian Istel and Michael Tscherner and Sascha Brunke and Lauren Ames and Arnaud Firon and Brian Green and Vitor Cabral and Marina Marcet-Houben and Ilse D Jacobsen and Jessica Quintin and Katja Seider and Ingrid Frohner and Walter Glaser and Helmut Jungwirth and Sophie Bachellier-Bassi and Murielle Chauvel and Ute Zeidler and Dominique Ferrandon and Toni Gabaldón and Bernhard Hube and Christophe d'Enfert and Steffen Rupp and Brendan Cormack and Ken Haynes and Karl Kuchler}, doi = {10.1371/journal.ppat.1004211}, issn = {1553-7374}, year = {2014}, date = {2014-01-01}, journal = {PLoS Pathog.}, volume = {10}, number = {6}, pages = {e1004211}, abstract = {The opportunistic fungal pathogen Candida glabrata is a frequent cause of candidiasis, causing infections ranging from superficial to life-threatening disseminated disease. The inherent tolerance of C. glabrata to azole drugs makes this pathogen a serious clinical threat. To identify novel genes implicated in antifungal drug tolerance, we have constructed a large-scale C. glabrata deletion library consisting of 619 unique, individually bar-coded mutant strains, each lacking one specific gene, all together representing almost 12% of the genome. Functional analysis of this library in a series of phenotypic and fitness assays identified numerous genes required for growth of C. glabrata under normal or specific stress conditions, as well as a number of novel genes involved in tolerance to clinically important antifungal drugs such as azoles and echinocandins. We identified 38 deletion strains displaying strongly increased susceptibility to caspofungin, 28 of which encoding proteins that have not previously been linked to echinocandin tolerance. Our results demonstrate the potential of the C. glabrata mutant collection as a valuable resource in functional genomics studies of this important fungal pathogen of humans, and to facilitate the identification of putative novel antifungal drug target and virulence genes.}, keywords = {Antifungal Agents, Azoles, Biofilms, Candida glabrata, Candidiasis, Cell Wall, Drug Resistance, Echinocandins, ferrandon, Fungal, Fungal Proteins, Gene Deletion, Gene Knockout Techniques, Gene Library, M3i, Microbial Sensitivity Tests, Osmotic Pressure, Phenotype}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Design of RNA Binders: Targeting the HIV Replication Cycle as a Case Study.}, author = {A Blond and E Ennifar and C Tisné and L Micouin}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25100137?dopt=Abstract}, doi = {10.1002/cmdc.201402259}, isbn = {25100137}, year = {2014}, date = {2014-01-01}, journal = {ChemMedChem}, volume = {9}, number = {9}, pages = {1982-96}, abstract = {The human immunodeficiency virus 1 (HIV-1) replication cycle is finely tuned with many important steps involving RNA-RNA or protein-RNA interactions, all of them being potential targets for the development of new antiviral compounds. This cycle can also be considered as a good benchmark for the evaluation of early-stage strategies aiming at designing drugs that bind to RNA, with the possibility to correlate in vitro activities with antiviral properties. In this review, we highlight different approaches developed to interfere with four important steps of the HIV-1 replication cycle: the early stage of reverse transcription, the transactivation of viral transcription, the nuclear export of partially spliced transcripts and the dimerization step.}, keywords = {DUMAS, ENNIFAR, RNA recognition antiviral agents drug design human immunodeficiency virus new targets, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Surprising base pairing and structural properties of 2'-trifluoromethylthio-modified ribonucleic acids.}, author = {M Košutić and L Jud and C Da Veiga and M Frener and K Fauster and C Kreutz and E Ennifar and R Micura}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24766131?dopt=Abstract}, doi = {10.1021/ja5005637}, isbn = {24766131}, year = {2014}, date = {2014-01-01}, journal = {J Am Chem Soc}, volume = {136}, number = {18}, pages = {6656-63}, abstract = {The chemical synthesis of ribonucleic acids (RNA) with novel chemical modifications is largely driven by the motivation to identify eligible functional probes for the various applications in life sciences. To this end, we have a strong focus on the development of novel fluorinated RNA derivatives that are powerful in NMR spectroscopic analysis of RNA folding and RNA ligand interactions. Here, we report on the synthesis of 2'-SCF3 pyrimidine nucleoside containing oligoribonucleotides and the comprehensive investigation of their structure and base pairing properties. While this modification has a modest impact on thermodynamic stability when it resides in single-stranded regions, it was found to be destabilizing to a surprisingly high extent when located in double helical regions. Our NMR spectroscopic investigations on short single-stranded RNA revealed a strong preference for C2'-endo conformation of the 2'-SCF3 ribose unit. Together with a recent computational study (L. Li, J. W. Szostak, J. Am. Chem. Soc. 2014, 136, 2858-2865) that estimated the extent of destabilization caused by a single C2'-endo nucleotide within a native RNA duplex to amount to 6 kcal mol(-1) because of disruption of the planar base pair structure, these findings support the notion that the intrinsic preference for C2'-endo conformation of 2'-SCF3 nucleosides is most likely responsible for the pronounced destabilization of double helices. Importantly, we were able to crystallize 2'-SCF3 modified RNAs and solved their X-ray structures at atomic resolution. Interestingly, the 2'-SCF3 containing nucleosides that were engaged in distinct mismatch arrangements, but also in a standard Watson-Crick base pair, adopted the same C3'-endo ribose conformations as observed in the structure of the unmodified RNA. Likely, strong crystal packing interactions account for this observation. In all structures, the fluorine atoms made surprisingly close contacts to the oxygen atoms of the corresponding pyrimidine nucleobase (O2), and the 2'-SCF3 moieties participated in defined water-bridged hydrogen-bonding networks in the minor groove. All these features allow a rationalization of the structural determinants of the 2'-SCF3 nucleoside modification and correlate them to base pairing properties.}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A minimalist mitochondrial threonyl-tRNA synthetase exhibits tRNA-isoacceptor specificity during proofreading.}, author = {X L Zhou and Z R Ruan and M Wang and Z P Fang and Y Wang and Y Chen and R J Liu and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25414329?dopt=Abstract}, doi = {10.1093/nar/gku1218}, isbn = {25414329}, year = {2014}, date = {2014-01-01}, journal = {Nucleic Acids Res}, volume = {42}, number = {22}, pages = {13873-13886}, abstract = {Yeast mitochondria contain a minimalist threonyl-tRNA synthetase (ThrRS) composed only of the catalytic core and tRNA binding domain but lacking the entire editing domain. Besides the usual tRNAThr2, some budding yeasts, such as Saccharomyces cerevisiae, also contain a non-canonical tRNAThr1 with an enlarged 8-nucleotide anticodon loop, reprograming the usual leucine CUN codons to threonine. This raises interesting questions about the aminoacylation fidelity of such ThrRSs and the possible contribution of the two tRNAThrs during editing. Here, we found that, despite the absence of the editing domain, S. cerevisiae mitochondrial ThrRS (ScmtThrRS) harbors a tRNA-dependent pre-transfer editing activity. Remarkably, only the usual tRNAThr2 stimulated pre-transfer editing, thus, establishing the first example of a synthetase exhibiting tRNA-isoacceptor specificity during pre-transfer editing. We also showed that the failure of tRNAThr1 to stimulate tRNA-dependent pre-transfer editing was due to the lack of an editing domain. Using assays of the complementation of a ScmtThrRS gene knockout strain, we showed that the catalytic core and tRNA binding domain of ScmtThrRS co-evolved to recognize the unusual tRNAThr1. In combination, the results provide insights into the tRNA-dependent editing process and suggest that tRNA-dependent pre-transfer editing takes place in the aminoacylation catalytic core.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Hypermethylated-capped selenoprotein mRNAs in mammals.}, author = {L Wurth and A S Gribling-Burrer and C Verheggen and M Leichter and A Takeuchi and S Baudrey and F Martin and A Krol and E Bertrand and C Allmang}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25013170}, doi = {10.1093/nar/gku580}, isbn = {25013170}, year = {2014}, date = {2014-01-01}, journal = {Nucleic Acids Res}, volume = {42}, number = {13}, pages = {8663-8677}, abstract = {Mammalian mRNAs are generated by complex and coordinated biogenesis pathways and acquire 5'-end m7G caps that play fundamental roles in processing and translation. Here we show that several selenoprotein mRNAs are not recognized efficiently by translation initiation factor eIF4E because they bear a hypermethylated cap. This cap modification is acquired via a 5'-end maturation pathway similar to that of the small nucle(ol)ar RNAs (sn- and snoRNAs). Our findings also establish that the trimethylguanosine synthase 1 (Tgs1) interacts with selenoprotein mRNAs for cap hypermethylation and that assembly chaperones and core proteins devoted to sn- and snoRNP maturation contribute to recruiting Tgs1 to selenoprotein mRNPs. We further demonstrate that the hypermethylated-capped selenoprotein mRNAs localize to the cytoplasm, are associated with polysomes and thus translated. Moreover, we found that the activity of Tgs1, but not of eIF4E, is required for the synthesis of the GPx1 selenoprotein in vivo.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Differential Modes of Peptide Binding onto Replicative Sliding Clamps from Various Bacterial Origins.}, author = {P Wolff and I Amal and V Oliéric and O Chaloin and G Gygli and E Ennifar and B Lorber and G Guichard and J E Wagner and A Dejaegere and D Y Burnouf}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25170813?dopt=Abstract}, doi = {10.1021/jm500467a}, isbn = {25170813}, year = {2014}, date = {2014-01-01}, journal = {J Med Chem}, volume = {57}, number = {18}, pages = {7565-7576}, abstract = {Bacterial sliding clamps are molecular hubs that interact with many proteins involved in DNA metabolism through their binding, via a conserved peptidic sequence, into a universally conserved pocket. This interacting pocket is acknowledged as a potential molecular target for the development of new antibiotics. We previously designed short peptides with an improved affinity for the Escherichia coli binding pocket. Here we show that these peptides differentially interact with other bacterial clamps, despite the fact that all pockets are structurally similar. Thermodynamic and modeling analyses of the interactions differentiate between two categories of clamps: group I clamps interacts efficiently with our designed peptides and assembles the Escherichia coli and related orthologs clamps, while group II poorly interact with the same peptides and includes Bacillus subtilis and other Gram+ clamps. These studies also suggest that the peptide binding process could occur via different mechanisms depending on which type of clamp it binds to.}, keywords = {ENNIFAR, FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Recognition of Watson-Crick base pairs: constraints and limits due to geometric selection and tautomerism}, author = {E Westhof and M Yusupov and G Yusupova}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24765524}, doi = {10.12703/P6-19}, isbn = {24765524}, year = {2014}, date = {2014-01-01}, journal = {F1000Prime Rep}, volume = {6}, pages = {19}, abstract = {The natural bases of nucleic acids have a strong preference for one tautomer form, guaranteeing fidelity in their hydrogen bonding potential. However, base pairs observed in recent crystal structures of polymerases and ribosomes are best explained by an alternative base tautomer, leading to the formation of base pairs with Watson-Crick-like geometries. These observations set limits to geometric selection in molecular recognition of complementary Watson-Crick pairs for fidelity in replication and translation processes.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Isostericity and tautomerism of base pairs in nucleic acids.}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24950426?dopt=Abstract}, doi = {10.1016/j.febslet.2014.06.031.}, isbn = {24950426}, year = {2014}, date = {2014-01-01}, journal = {FEBS Lett}, volume = {588}, number = {15}, pages = {2464-2469}, abstract = {The natural bases of nucleic acids form a great variety of base pairs with at least two hydrogen bonds between them. They are classified in twelve main families, with the Watson-Crick family being one of them. In a given family, some of the base pairs are isosteric between them, meaning that the positions and the distances between the C1' carbon atoms are very similar. The isostericity of Watson-Crick pairs between the complementary bases forms the basis of RNA helices and of the resulting RNA secondary structure. Several defined suites of non-Watson-Crick base pairs assemble into RNA modules that form recurrent, rather regular, building blocks of the tertiary architecture of folded RNAs. RNA modules are intrinsic to RNA architecture are therefore disconnected from a biological function specifically attached to a RNA sequence. RNA modules occur in all kingdoms of life and in structured RNAs with diverse functions. Because of chemical and geometrical constraints, isostericity between non-Watson-Crick pairs is restricted and this leads to higher sequence conservation in RNA modules with, consequently, greater difficulties in extracting 3D information from sequence analysis. Nucleic acid helices have to be recognised in several biological processes like replication or translational decoding. In polymerases and the ribosomal decoding site, the recognition occurs on the minor groove sides of the helical fragments. With the use of alternative conformations, protonated or tautomeric forms of the bases, some base pairs with Watson-Crick-like geometries can form and be stabilized. Several of these pairs with Watson-Crick-like geometries extend the concept of isostericity beyond the number of isosteric pairs formed between complementary bases. These observations set therefore limits and constraints to geometric selection in molecular recognition of complementary Watson-Crick pairs for fidelity in replication and translation processes.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Biological evidence for the world's smallest tRNAs.}, author = {S Wende and E G Platzer and F Jühling and J Pütz and C Florentz and P F Stadler and M Mörl}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23958440?dopt=Abstract}, doi = {10.1016/j.biochi.2013.07.034}, isbn = {23958440}, year = {2014}, date = {2014-01-01}, journal = {Biochimie}, volume = {100}, pages = {151-158}, abstract = {Due to their function as adapters in translation, tRNA molecules share a common structural organization in all kingdoms and organelles with ribosomal protein biosynthesis. A typical tRNA has a cloverleaf-like secondary structure, consisting of acceptor stem, D-arm, anticodon arm, a variable region, and T-arm, with an average length of 73 nucleotides. In several mitochondrial genomes, however, tRNA genes encode transcripts that show a considerable deviation of this standard, having reduced D- or T-arms or even completely lack one of these elements, resulting in tRNAs as small as 66 nts. An extreme case of such truncations is found in the mitochondria of Enoplea. Here, several tRNA genes are annotated that lack both the D- and the T-arm, suggesting even shorter transcripts with a length of only 42 nts. However, direct evidence for these exceptional tRNAs, which were predicted by purely computational means, has been lacking so far. Here, we demonstrate that several of these miniaturized armless tRNAs consisting only of acceptor- and anticodon-arms are indeed transcribed and correctly processed by non-encoded CCA addition in the mermithid Romanomermis culicivorax. This is the first direct evidence for the existence and functionality of the smallest tRNAs ever identified so far. It opens new possibilities towards exploration/assessment of minimal structural motifs defining a functional tRNA and their evolution.}, keywords = {FLORENTZ SISSLER CCA-addition Enoplea armless tRNA mitochondria processing tRNA structure, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Quantifying resolution limiting factors in subtomogram averaged cryo-electron tomography using simulations.}, author = {L M Voortman and M Vulović and M Maletta and A Voigt and E M Franken and A Simonetti and P J Peters and L J van Vliet and B Rieger}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24998892}, doi = {10.1016/j.jsb.2014.06.007}, isbn = {24998892}, year = {2014}, date = {2014-01-01}, journal = {J Struct Biol}, volume = {187}, number = {2}, pages = {103-111}, abstract = {Cryo-electron tomography (CET) is the only available technique capable of characterizing the structure of biological macromolecules in conditions close to the native state. With the advent of subtomogram averaging, as a post-processing step to CET, resolutions in the (sub-) nanometer range have become within reach. In addition to advances in instrumentation and experiments, the reconstruction scheme has improved by inclusion of more accurate contrast transfer function (CTF) correction methods, better defocus estimation, and better alignments of the tilt-series and subtomograms. To quantify the importance of each contribution, we have split the full process from data collection to reconstruction into different steps. For the purpose of evaluation we have acquired tilt-series of ribosomes in such a way that we could precisely determine the defocus of each macromolecule. Then, we simulated tilt-series using the InSilicoTEM package and applied tomogram reconstruction and subtomogram averaging. Through large scale simulations under different conditions and parameter settings we find that tilt-series alignment is the resolution limiting factor for our experimental data. Using simulations, we find that when this alignment inaccuracy is alleviated, tilted CTF correction improves the final resolution, or equivalently, the same resolution can be achieved using less particles. Furthermore, we predict from which resolution onwards better CTF correction and defocus estimation methods are required. We obtain a final average using 3198 ribosomes with a resolution of 2.2nm on the experimental data. Our simulations suggest that with the same number of particles a resolution of 1.2nm could be achieved by improving the tilt-series alignment.}, keywords = {Acquisition protocol Cryo-EM Ribosome Subtomogram averaging TEM image simulation Tilted CTF correction Tomography, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Biogenesis of Circular RNAs.}, author = {Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25259915?dopt=Abstract}, doi = {10.1016/j.cell.2014.09.005}, isbn = {25259915}, year = {2014}, date = {2014-01-01}, journal = {Cell}, volume = {159}, number = {1}, pages = {13-14}, abstract = {Circular RNAs are generated during splicing through various mechanisms. Ashwal-Fluss et al. demonstrate that exon circularization and linear splicing compete with each other in a tissue-specific fashion, and Zhang et al. show that exon circularization depends on flanking intronic complementary sequences. Both papers show that several types of circular RNA transcripts can be produced from a single gene.}, keywords = {ERIANI, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The importance of regulatory RNAs in Staphylococcus aureus.}, author = {A Tomasini and P François and B P Howden and P Fechter and P Romby and I Caldelari}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24291227}, doi = {10.1016/j.meegid.2013.11.016}, isbn = {24291227}, year = {2014}, date = {2014-01-01}, journal = {Infect Genet Evol}, volume = {21}, number = {C}, pages = {616-626}, abstract = {RNA molecules with regulatory functions in pathogenic bacteria have benefited from a renewed interest these two last decades. In Staphylococcus aureus, recent genome-wide approaches have led to the discovery that almost 10-20% of genes code for RNAs with critical regulatory roles in adaptive processes. These RNAs include trans-acting RNAs, which mostly act through binding to target mRNAs, and cis-acting RNAs, which include regulatory regions of mRNAs responding to various metabolic signals. Besides recent analysis of S. aureus transcriptome has revealed an unprecedented existence of pervasive transcription generating a high number of weakly expressed antisense RNAs along the genome as well as numerous mRNAs with overlapped regions. Here, we will illustrate the diversity of trans-acting RNAs and illustrate how they are integrated into complex regulatory circuits, which link metabolism, stress response and virulence.}, keywords = {Regulatory RNA Staphylococcus Virulence, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Small RNA cloning and sequencing strategy affects host and viral microRNA expression signatures}, author = {G Stik and B Muylkens and D Coupeau and S Laurent and G Dambrine and M Messmer and B Chane-Woon-Ming and S Pfeffer and D Rasschaert}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24746587}, doi = {10.1016/j.jbiotec.2014.04.005}, isbn = {24746587}, year = {2014}, date = {2014-01-01}, journal = {J Biotechnol}, volume = {181}, pages = {35-44}, abstract = {The establishment of the microRNA (miRNA) expression signatures is the basic element to investigate the role played by these regulatory molecules in the biology of an organism. Marek's disease virus 1 (MDV-1) is an avian herpesvirus that naturally infects chicken and induces T cells lymphomas. During latency, MDV-1, like other herpesviruses, expresses a limited subset of transcripts. These include three miRNA clusters. Several studies identified the expression of virus and host encoded miRNAs from MDV-1 infected cell cultures and chickens. But a high discrepancy was observed when miRNA cloning frequencies obtained from different cloning and sequencing protocols were compared. Thus, we analyzed the effect of small RNA library preparation and sequencing on the miRNA frequencies obtained from the same RNA samples collected during MDV-1 infection of chicken at different steps of the oncoviral pathogenesis. Qualitative and quantitative variations were found in the data, depending on the strategy used. One of the mature miRNA derived from the latency-associated-transcript (LAT), mdv1-miR-M7-5p, showed the highest variation. Its cloning frequency was 50% of the viral miRNA counts when a small scale sequencing approach was used. Its frequency was 100 times less abundant when determined through the deep sequencing approach. Northern blot analysis showed a better correlation with the miRNA frequencies found by the small scale sequencing approach. By analyzing the cellular miRNA repertoire, we also found a gap between the two sequencing approaches. Collectively, our study indicates that next-generation sequencing data considered alone are limited for assessing the absolute copy number of transcripts. Thus, the quantification of small RNA should be addressed by compiling data obtained by using different techniques such as microarrays, qRT-PCR and NB analysis in support of high throughput sequencing data. These observations should be considered when miRNA variations are studied prior addressing functional studies.}, keywords = {Marek's disease virus MicroRNA Small RNA cloning and sequencing Viral infection, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identifying recombination hotspots in the HIV-1 genome.}, author = {R P Smyth and T E Schlub and A J Grimm and C Waugh and P Ellenberg and A Chopra and S Mallal and D Cromer and J Mak and M P Davenport}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24371048?dopt=Abstract}, doi = {10.1128/JVI.03014-13}, isbn = {24371048}, year = {2014}, date = {2014-01-01}, journal = {J Virol}, volume = {88}, number = {5}, pages = {2891-2902}, abstract = {HIV-1 infection is characterised by the rapid generation of genetic diversity that facilitates viral escape from immune selection and antiretroviral therapy. Despite recombination's crucial role in viral diversity and evolution, little is known about the genomic factors that influence recombination between highly similar genomes. In this study, we use a minimally modified full length HIV-1 genome and high throughput sequence analysis to study recombination in gag and pol in T cells. We find that recombination is favoured at a number of recombination hotspots, where recombination occurs six times more frequently than at corresponding coldspots. Interestingly, these hotspots occur near important features of the HIV-1 genome, but do not occur at sites immediately around protease inhibitor or reverse transcriptase inhibitor drug resistance mutations. We show that the recombination hot and cold spots are consistent across five blood donors and are independent of co-receptor mediated entry. Finally, we check common experimental confounders and find that these are not driving the location of recombination hotspots. This is the first study to identify the location of recombination hotspots, between two similar viral genomes with great statistical power and under conditions that closely reflect natural recombination events amongst HIV-1 quasispecies.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of RNA binding and chaperoning activities of HIV-1 Vif protein: Importance of the C-terminal unstructured tail.}, author = {D Sleiman and S Bernacchi and S X Guerrero and F Brachet and V Larue and J C Paillart and C Tisne}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25144404?dopt=Abstract}, doi = {10.4161/rna.29546}, isbn = {25144404}, year = {2014}, date = {2014-01-01}, journal = {RNA Biol}, volume = {11}, number = {7}, pages = {906-920}, abstract = {The viral infectivity factor (Vif) is essential for the productive infection and dissemination of HIV-1 in non-permissive cells, containing the cellular anti-HIV defense cytosine deaminases APOBEC3 (A3G and A3F). Vif neutralizes the antiviral activities of the APOBEC3G/F by diverse mechanisms including their degradation through the ubiquitin/proteasome pathway and their translational inhibition. In addition, Vif appears to be an active partner of the late steps of viral replication by interacting with Pr55Gag, reverse transcriptase and genomic RNA. Here, we expressed and purified full-length and truncated Vif proteins, and analyzed their RNA binding and chaperone properties. First, we showed by CD and NMR spectroscopies that the N-terminal domain of Vif is highly structured in solution, whereas the C-terminal domain remains mainly unfolded. Both domains exhibited substantial RNA binding capacities with dissociation constants in the nanomolar range, whereas the basic unfolded C-terminal domain of Vif was responsible in part for its RNA chaperone activity. Second, we showed by NMR chemical shift mapping that Vif and NCp7 share the same binding sites on tRNALys3, the primer of HIV-1 reverse transcriptase. Finally, our results indicate that Vif has potent RNA chaperone activity and provide direct evidence for an important role of the unstructured C-terminal domain of Vif in this capacity.}, keywords = {HIV RNA chaperone Vif nucleocapsid unstructured domain, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A PNPase Dependent CRISPR System in Listeria.}, author = {N Sesto and M Touchon and J M Andrade and J Kondo and E P Rocha and C M Arraiano and C Archambaud and E Westhof and P Romby and P Cossart}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24415952?dopt=Abstract}, doi = {10.1371/journal.pgen.1004065}, isbn = {24415952}, year = {2014}, date = {2014-01-01}, journal = {PLoS Genet}, volume = {10}, number = {1}, pages = {e1004065}, abstract = {The human bacterial pathogen Listeria monocytogenes is emerging as a model organism to study RNA-mediated regulation in pathogenic bacteria. A class of non-coding RNAs called CRISPRs (clustered regularly interspaced short palindromic repeats) has been described to confer bacterial resistance against invading bacteriophages and conjugative plasmids. CRISPR function relies on the activity of CRISPR associated (cas) genes that encode a large family of proteins with nuclease or helicase activities and DNA and RNA binding domains. Here, we characterized a CRISPR element (RliB) that is expressed and processed in the L. monocytogenes strain EGD-e, which is completely devoid of cas genes. Structural probing revealed that RliB has an unexpected secondary structure comprising basepair interactions between the repeats and the adjacent spacers in place of canonical hairpins formed by the palindromic repeats. Moreover, in contrast to other CRISPR-Cas systems identified in Listeria, RliB-CRISPR is ubiquitously present among Listeria genomes at the same genomic locus and is never associated with the cas genes. We showed that RliB-CRISPR is a substrate for the endogenously encoded polynucleotide phosphorylase (PNPase) enzyme. The spacers of the different Listeria RliB-CRISPRs share many sequences with temperate and virulent phages. Furthermore, we show that a cas-less RliB-CRISPR lowers the acquisition frequency of a plasmid carrying the matching protospacer, provided that trans encoded cas genes of a second CRISPR-Cas system are present in the genome. Importantly, we show that PNPase is required for RliB-CRISPR mediated DNA interference. Altogether, our data reveal a yet undescribed CRISPR system whose both processing and activity depend on PNPase, highlighting a new and unexpected function for PNPase in "CRISPRology".}, keywords = {ROMBY, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pathogenic Implications of Human Mitochondrial Aminoacyl-tRNA Synthetases.}, author = {H Schwenzer and J Zoll and C Florentz and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23824528}, doi = {10.1007/128_2013_457}, isbn = {23824528}, year = {2014}, date = {2014-01-01}, journal = {Top Curr Chem}, volume = {344}, pages = {247-292}, abstract = {Mitochondria are considered as the powerhouse of eukaryotic cells. They host several central metabolic processes fueling the oxidative phosphorylation pathway (OXPHOS) that produces ATP from its precursors ADP and inorganic phosphate Pi (PPi). The respiratory chain complexes responsible for the OXPHOS pathway are formed from complementary sets of protein subunits encoded by the nuclear genome and the mitochondrial genome, respectively. The expression of the mitochondrial genome requires a specific and fully active translation machinery from which aminoacyl-tRNA synthetases (aaRSs) are key actors. Whilst the macromolecules involved in mammalian mitochondrial translation have been under investigation for many years, there has been an explosion of interest in human mitochondrial aaRSs (mt-aaRSs) since the discovery of a large (and growing) number of mutations in these genes that are linked to a variety of neurodegenerative disorders. Herein we will review the present knowledge on mt-aaRSs in terms of their biogenesis, their connection to mitochondrial respiration, i.e., the respiratory chain (RC) complexes, and to the mitochondrial translation machinery. The pathology-related mutations detected so far are described, with special attention given to their impact on mt-aaRSs biogenesis, functioning, and/or subsequent activities. The collected data to date shed light on the diverse routes that are linking primary molecular possible impact of a mutation to its phenotypic expression. It is envisioned that a variety of mechanisms, inside and outside the translation machinery, would play a role on the heterogeneous manifestations of mitochondrial disorders.}, keywords = {Aminoacyl-tRNA synthetase Human mitochondrial disorders Pathology-related mutations Respiratory chain defects, FLORENTZ, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Released selective pressure on a structural domain gives new insights on the functional relaxation of mitochondrial aspartyl-tRNA synthetase.}, author = {H Schwenzer and G C Scheper and N Zorn and L Moulinier and A Gaudry and E Leize and F Martin and C Florentz and O Poch and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24120687}, doi = {10.1016/j.biochi.2013.09.027}, isbn = {24120687}, year = {2014}, date = {2014-01-01}, journal = {Biochimie}, volume = {100}, pages = {18-26}, abstract = {Mammalian mitochondrial aminoacyl-tRNA synthetases are nuclear-encoded enzymes that are essential for mitochondrial protein synthesis. Due to an endosymbiotic origin of the mitochondria, many of them share structural domains with homologous bacterial enzymes of same specificity. This is also the case for human mitochondrial aspartyl-tRNA synthetase (AspRS) that shares the so-called bacterial insertion domain with bacterial homologs. The function of this domain in the mitochondrial proteins is unclear. Here, we show by bioinformatic analyses that the sequences coding for the bacterial insertion domain are less conserved in opisthokont and protist than in bacteria and viridiplantae. The divergence suggests a loss of evolutionary pressure on this domain for non-plant mitochondrial AspRSs. This discovery is further connected with the herein described occurrence of alternatively spliced transcripts of the mRNAs coding for some mammalian mitochondrial AspRSs. Interestingly, the spliced transcripts alternately lack one of the four exons that code for the bacterial insertion domain. Although we showed that the human alternative transcript is present in all tested tissues; co-exists with the full-length form, possesses 5'- and 3'-UTRs, a poly-A tail and is bound to polysomes, we were unable to detect the corresponding protein. The relaxed selective pressure combined with the occurrence of alternative splicing, involving a single structural sub-domain, favors the hypothesis of the loss of function of this domain for AspRSs of mitochondrial location. This evolutionary divergence is in line with other characteristics, established for the human mt-AspRS, that indicate a functional relaxation of non-viridiplantae mt-AspRSs when compared to bacterial and plant ones, despite their common ancestry.}, keywords = {Aminoacyl-tRNA synthetase Bioinformatics MTS Mitochondria Molecular biology Translation aaRS aminoacyl-tRNA synthetase (specificity is indicated by the name of the amino acid abbreviated in a three-letter code, e.g. AspRS stands for aspartyl-tRNA synthetase) mitochondrial mitochondrial targeting sequence mt, ERIANI, FLORENTZ, SISSLER, transferred to the cognate tRNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {15-20% of HIV substitution mutations are associated with recombination.}, author = {T E Schlub and A J Grimm and R P Smyth and D Cromer and A Chopra and S Mallal and V Venturi and C Waugh and J Mak and M P Davenport}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24453357?dopt=Abstract}, doi = {10.1128/JVI.03136-13}, isbn = {24453357}, year = {2014}, date = {2014-01-01}, journal = {J Virol}, volume = {88}, number = {7}, pages = {3837-3849}, abstract = {HIV undergoes a high rate of mutation and recombination during reverse transcription, but it is not known whether these events occur independently or are linked mechanistically. Here we use a system of silent marker mutations in HIV and a single round of infection in primary T-lymphocytes, combined with a high-throughput sequencing and mathematical modelling approach to directly estimate the viral recombination and mutation rates. From >7 million nt of sequences from HIV infection, we observe 4801 recombination events and 859 substitution mutations (≈1.51 and 0.12 events per 1000 nt respectively). We use experimental controls to account for PCR-induced and transfection-induced recombination and sequencing error. We find the single cycle virus-induced mutation rate is 4.6 × 10-5 mutations per nt after correction. By sorting our data into recombined and non-recombined sequences, we find a significantly higher mutation rate in recombined regions (p=0.003, Fisher's exact). We use a permutation approach to eliminate a number of potential confounding factors and confirm that mutation occurs around the site of recombination, and is not simply co-located in the genome. By comparing mutation rates in recombined and non-recombined regions we find that recombination-associated mutations account for 15-20% of all mutations occurring during reverse transcription.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Plongée au cœur des molécules du vivant (Diving into the heart of the molecules of life)}, author = {D Sauter}, year = {2014}, date = {2014-01-01}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Molecular basis for the differential interaction of plant mitochondrial VDAC proteins with tRNAs.}, author = {T Salinas and S El Farouk-Ameqrane and E Ubrig and C Sauter and A M Duchêne and L Maréchal-Drouard}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25114051?dopt=Abstract}, doi = {10.1093/nar/gku728}, isbn = {25114051}, year = {2014}, date = {2014-01-01}, journal = {Nucleic Acids Res}, volume = {42}, number = {15}, pages = {9937-9948}, abstract = {In plants, the voltage-dependent anion-selective channel (VDAC) is a major component of a pathway involved in transfer RNA (tRNA) translocation through the mitochondrial outer membrane. However, the way in which VDAC proteins interact with tRNAs is still unknown. Potato mitochondria contain two major mitochondrial VDAC proteins, VDAC34 and VDAC36. These two proteins, composed of a N-terminal α-helix and of 19 β-strands forming a β-barrel structure, share 75% sequence identity. Here, using both northwestern and gel shift experiments, we report that these two proteins interact differentially with nucleic acids. VDAC34 binds more efficiently with tRNAs or other nucleic acids than VDAC36. To further identify specific features and critical amino acids required for tRNA binding, 21 VDAC34 mutants were constructed and analyzed by northwestern. This allowed us to show that the β-barrel structure of VDAC34 and the first 50 amino acids that contain the α-helix are essential for RNA binding. Altogether the work shows that during evolution, plant mitochondrial VDAC proteins have diverged so as to interact differentially with nucleic acids, and this may reflect their involvement in various specialized biological functions.}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A Non-Coding RNA Promotes Bacterial Persistence and Decreases Virulence by Regulating a Regulator in Staphylococcus aureus.}, author = {C Romilly and C Lays and A Tomasini and I Caldelari and Y Benito and P Hammann and T Geissmann and S Boisset and P Romby and F Vandenesch}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24651379?dopt=Abstract}, doi = {10.1371/journal.ppat.1003979}, isbn = {24651379}, year = {2014}, date = {2014-01-01}, journal = {PLoS Pathog}, volume = {10}, number = {3}, pages = {e1003979}, abstract = {Staphylococcus aureus produces a high number of RNAs for which the functions are poorly understood. Several non-coding RNAs carry a C-rich sequence suggesting that they regulate mRNAs at the post-transcriptional level. We demonstrate that the Sigma B-dependent RsaA RNA represses the synthesis of the global transcriptional regulator MgrA by forming an imperfect duplex with the Shine and Dalgarno sequence and a loop-loop interaction within the coding region of the target mRNA. These two recognition sites are required for translation repression. Consequently, RsaA causes enhanced production of biofilm and a decreased synthesis of capsule formation in several strain backgrounds. These phenotypes led to a decreased protection of S. aureus against opsonophagocytic killing by polymorphonuclear leukocytes compared to the mutant strains lacking RsaA. Mice animal models showed that RsaA attenuates the severity of acute systemic infections and enhances chronic catheter infection. RsaA takes part in a regulatory network that contributes to the complex interactions of S. aureus with the host immune system to moderate invasiveness and favour chronic infections. It is the first example of a conserved small RNA in S. aureus functioning as a virulence suppressor of acute infections. Because S. aureus is essentially a human commensal, we propose that RsaA has been positively selected through evolution to support commensalism and saprophytic interactions with the host.}, keywords = {PPSE, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {In-line alignment and Mg(2+) coordination at the cleavage site of the env22 twister ribozyme.}, author = {A Ren and M Košutić and K R Rajashankar and M Frener and T Santner and E Westhof and R Micura and D J Patel}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25410397?dopt=Abstract}, doi = {10.1038/ncomms6534}, isbn = {25410397}, year = {2014}, date = {2014-01-01}, journal = {Nat Commun}, volume = {5}, pages = {5534}, abstract = {Small self-cleaving nucleolytic ribozymes contain catalytic domains that accelerate site-specific cleavage/ligation of phosphodiester backbones. We report on the 2.9-Å crystal structure of the env22 twister ribozyme, which adopts a compact tertiary fold stabilized by co-helical stacking, double-pseudoknot formation and long-range pairing interactions. The U-A cleavage site adopts a splayed-apart conformation with the modelled 2'-O of U positioned for in-line attack on the adjacent to-be-cleaved P-O5' bond. Both an invariant guanosine and a Mg(2+) are directly coordinated to the non-bridging phosphate oxygens at the U-A cleavage step, with the former positioned to contribute to catalysis and the latter to structural integrity. The impact of key mutations on cleavage activity identified an invariant guanosine that contributes to catalysis. Our structure of the in-line aligned env22 twister ribozyme is compared with two recently reported twister ribozymes structures, which adopt similar global folds, but differ in conformational features around the cleavage site.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Fusion with Anticodon Binding Domain of GluRS is Not Sufficient to Alter the Substrate Specificity of a Chimeric Glu-Q-RS.}, author = {S Ray and M Blaise and B Roy and S Ghosh and D Kern and R Banerjee}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24374508}, doi = {10.1007/s10930-013-9537-7}, isbn = {24374508}, year = {2014}, date = {2014-01-01}, journal = {Protein J}, volume = {33}, number = {1}, pages = {48-60}, abstract = {Glutamyl-queuosine-tRNAAsp synthetase (Glu-Q-RS) is a paralog of glutamyl-tRNA synthetase (GluRS) and is found in more than forty species of proteobacteria, cyanobacteria, and actinobacteria. Glu-Q-RS shows striking structural similarity with N-terminal catalytic domain of GluRS (NGluRS) but it lacks the C-terminal anticodon binding domain (CGluRS). In spite of structural similarities, Glu-Q-RS and NGluRS differ in their functional properties. Glu-Q-RS glutamylates the Q34 nucleotide of the anticodon of tRNAAsp whereas NGluRS constitutes the catalytic domain of GluRS catalyzing the transfer of Glu on the acceptor end of tRNAGlu. Since NGluRS is able to catalyze aminoacylation of only tRNAGlu the glutamylation capacity of tRNAAsp by Glu-Q-RS is surprising. To understand the substrate specificity of Glu-Q-RS we undertook a systemic approach by investigating the biophysical and biochemical properties of the NGluRS (1-301), CGluRS (314-471) and Glu-Q-RS-CGluRS, (1-298 of Glu-Q-RS fused to 314-471 from GluRS). Circular dichroism, fluorescence spectroscopy and differential scanning calorimetry analyses revealed absence of N-terminal domain (1-298 of Glu-Q-RS) and C-terminal domain (314-471 from GluRS) communication in chimera, in contrast to the native full length GluRS. The chimeric Glu-Q-RS is still able to aminoacylate tRNAAsp but has also the capacity to bind tRNAGlu. However the chimeric protein is unable to aminoacylate tRNAGlu probably as a consequence of the lack of domain-domain communication.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Critical Role of Zinc Ion on E. coli Glutamyl-Queuosine-tRNA^{Asp} Synthetase (Glu-Q-RS) Structure and Function.}, author = {S Ray and V Banerjee and M Blaise and B Banerjee and K P Das and D Kern and R Banerjee}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24505021?dopt=Abstract}, doi = {10.1007/s10930-014-9546-1}, isbn = {24505021}, year = {2014}, date = {2014-01-01}, journal = {Protein J}, volume = {33}, number = {2}, pages = {143-149}, abstract = {Glutamyl-queuosine-tRNAAsp synthetase (Glu-Q-RS) and glutamyl-tRNA synthetase (GluRS), differ widely by their function although they share close structural resemblance within their catalytic core of GluRS. In particular both Escherichia coli GluRS and Glu-Q-RS contain a single zinc-binding site in their putative tRNA acceptor stem-binding domain. It has been shown that the zinc is crucial for correct positioning of the tRNAGlu acceptor-end in the active site of E. coli GluRS. To address the role of zinc ion in Glu-Q-RS, the C101S/C103S Glu-Q-RS variant is constructed. Energy dispersive X-ray fluorescence show that the zinc ion still remained coordinated but the variant became structurally labile and acquired aggregation capacity. The extent of aggregation of the protein is significantly decreased in presence of the small substrates and more particularly by adenosine triphosphate. Addition of zinc increased significantly the solubility of the variant. The aminoacylation assay reveals a decrease in activity of the variant even after addition of zinc as compared to the wild-type, although the secondary structure of the protein is not altered as shown by the Fourier transform infrared spectroscopy study.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Herpesviruses Encode Their Own MicroRNAs.}, author = {S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24778302?dopt=Abstract}, doi = {10.1373/clinchem.2013.216408}, isbn = {24778302}, year = {2014}, date = {2014-01-01}, journal = {Clin Chem}, volume = {60}, number = {5}, pages = {791-792}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cap analogs containing 6-thioguanosine - reagents for the synthesis of mRNAs selectively photo-crosslinkable with cap-binding biomolecules.}, author = {M Nowakowska and J Kowalska and F Martin and A d'Orchymont and J Zuberek and M Lukaszewicz and E Darzynkiewicz and J Jemielity}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24763507}, doi = {10.1039/c4ob00059e}, isbn = {24763507}, year = {2014}, date = {2014-01-01}, journal = {Org Biomol Chem}, volume = {12}, number = {27}, pages = {4841-4847}, abstract = {Numerous biomolecules recognize the 7-methylguanosine cap structure present at the 5' ends of eukaryotic mRNAs. Photo-crosslinking is a valuable technique to study these interactions. We report three anti-reverse cap analogs containing a photo-activable nucleoside, 6-thioguanosine (6SG), that enable the synthesis of capped RNAs with 6SG positioned exclusively as the first transcribed nucleotide. The effect of the 6-thioguanosine moiety on binding to the translation factor eIF4E and the efficiency of mRNA translation was determined. The utility of mRNAs with a 6SG-modified cap in crosslinking experiments is shown by mapping the histone H4 cap-binding pocket.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {ZNF143 is regulated through alternative 3'UTR isoforms.}, author = {R P Ngondo and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24952353?dopt=Abstract}, doi = {10.1016/j.biochi.2014.06.008}, isbn = {24952353}, year = {2014}, date = {2014-01-01}, journal = {Biochimie}, volume = {104}, pages = {137-146}, abstract = {ZNF143 is a ubiquitously expressed transcription factor conserved in all vertebrates, regulating genes involved in primary metabolism and cell growth. It is therefore crucial to tightly maintain the adequate level of this factor in the cell. Although ZNF143 expression is auto-regulated at the transcriptional level, nothing is known about the post-transcriptional events influencing its expression. In this work, performed in mammalian cells, we show that ZNF143 expresses different 3'-untranslated regions (3'-UTR) as a result of alternative polyadenylation. These 3'UTR isoforms have a diverse impact on the ZNF143 transcript fate. Indeed, we show that the longest isoform, unlike the short one, contains a destabilizing AU-Rich element and is targeted by the miRNA 590-3p. Additionally we observed a correlation between ZNF143 downregulation and miR-590-3p up-regulation in retinoic acid treated teratocarcinoma cells. This strongly suggests that ZNF143 post-transcriptional regulation depends on the long 3'UTR isoform during teratocarcinoma cells differentiation. Finally we evidenced that the alternative polyadenylation site usage is independent of the previously identified ZNF143 transcriptional auto-regulation.}, keywords = {3'UTR ARE element NT2D1 ZNF143 alternative polyadenylation miR-590-3p, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 DIS stem loop forms an obligatory bent kissing intermediate in the dimerization pathway.}, author = {H Mundigala and J B Michaux and A L Feig and E Ennifar and D Rueda}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24813449?dopt=Abstract}, doi = {10.1093/nar/gku332}, isbn = {24813449}, year = {2014}, date = {2014-01-01}, journal = {Nucleic Acids Res}, volume = {42}, number = {11}, pages = {7281-7289}, abstract = {The HIV-1 dimerization initiation sequence (DIS) is a conserved palindrome in the apical loop of a conserved hairpin motif in the 5'-untranslated region of its RNA genome. DIS hairpin plays an important role in genome dimerization by forming a 'kissing complex' between two complementary hairpins. Understanding the kinetics of this interaction is key to exploiting DIS as a possible human immunodeficiency virus (HIV) drug target. Here, we present a single-molecule Förster resonance energy transfer (smFRET) study of the dimerization reaction kinetics. Our data show the real-time formation and dissociation dynamics of individual kissing complexes, as well as the formation of the mature extended duplex complex that is ultimately required for virion packaging. Interestingly, the single-molecule trajectories reveal the presence of a previously unobserved bent intermediate required for extended duplex formation. The universally conserved A272 is essential for the formation of this intermediate, which is stabilized by Mg2+, but not by K+ cations. We propose a 3D model of a possible bent intermediate and a minimal dimerization pathway consisting of three steps with two obligatory intermediates (kissing complex and bent intermediate) and driven by Mg2+ ions.}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Hypoxia Induces VEGF-C Expression in Metastatic Tumor Cells via a HIF-1α-Independent Translation-Mediated Mechanism.}, author = {F Morfoisse and A Kuchnio and C Frainay and A Gomez-Brouchet and M B Delisle and S Marzi and A C Helfer and F Hantelys and F Pujol and J Guillermet-Guibert and C Bousquet and M Dewerchin and S Pyronnet and A C Prats and P Carmeliet and B Garmy-Susini}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24388748?dopt=Abstract}, doi = {10.1016/j.celrep.2013.12.011}, isbn = {24388748}, year = {2014}, date = {2014-01-01}, journal = {Cell Rep}, volume = {6}, number = {1}, pages = {155-167}, abstract = {Various tumors metastasize via lymph vessels and lymph nodes to distant organs. Even though tumors are hypoxic, the mechanisms of how hypoxia regulates lymphangiogenesis remain poorly characterized. Here, we show that hypoxia reduced vascular endothelial growth factor C (VEGF-C) transcription and cap-dependent translation via the upregulation of hypophosphorylated 4E-binding protein 1 (4E-BP1). However, initiation of VEGF-C translation was induced by hypoxia through an internal ribosome entry site (IRES)-dependent mechanism. IRES-dependent VEGF-C translation was independent of hypoxia-inducible factor 1α (HIF-1α) signaling. Notably, the VEGF-C IRES activity was higher in metastasizing tumor cells in lymph nodes than in primary tumors, most likely because lymph vessels in these lymph nodes were severely hypoxic. Overall, this transcription-independent but translation-dependent upregulation of VEGF-C in hypoxia stimulates lymphangiogenesis in tumors and lymph nodes and may contribute to lymphatic metastasis.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Speciation of a group I intron into a lariat capping ribozyme.}, author = {M Meyer and H Nielsen and V Oliéric and P Roblin and S D Johansen and E Westhof and B Masquida}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24821772?dopt=Abstract}, doi = {10.1073/pnas.1322248111}, isbn = {24821772}, year = {2014}, date = {2014-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {111}, number = {21}, pages = {7659-7664}, abstract = {The lariat-capping (LC) ribozyme is a natural ribozyme isolated from eukaryotic microorganisms. Despite apparent structural similarity to group I introns, the LC ribozyme catalyzes cleavage by a 2',5' branching reaction, leaving the 3' product with a 3-nt lariat cap that functionally substitutes for a conventional mRNA cap in the downstream pre-mRNA encoding a homing endonuclease. We describe the crystal structures of the precleavage and postcleavage LC ribozymes, which suggest that structural features inherited from group I ribozymes have undergone speciation due to profound changes in molecular selection pressure, ultimately giving rise to an original branching ribozyme family. The structures elucidate the role of key elements that regulate the activity of the LC ribozyme by conformational switching and suggest a mechanism by which the signal for branching is transmitted to the catalytic core. The structures also show how conserved interactions twist residues, forming the lariat to join chemical groups involved in branching.}, keywords = {GIR1 RNA catalysis RNA structure SAXS crystallography, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @book{, title = {Mitochondria: An organelle for life.}, editor = {L Maréchal-Drouard and M Sissler and I Tarassov}, year = {2014}, date = {2014-01-01}, volume = {100}, publisher = {ELSEVIER}, series = {Biochimie}, keywords = {SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {book} } @article{, title = {RACK1 Controls IRES-Mediated Translation of Viruses.}, author = {K Majzoub and M L Hafirassou and C Meignin and A Goto and S Marzi and A Fedorova and Y Verdier and J Vinh and J A Hoffmann and F Martin and T F Baumert and C Schuster and JL Imler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/25416947}, doi = {10.1016/j.cell.2014.10.041}, isbn = {25416947}, year = {2014}, date = {2014-01-01}, journal = {Cell}, volume = {159}, number = {5}, pages = {1086-1095}, abstract = {Fighting viral infections is hampered by the scarcity of viral targets and their variability, resulting in development of resistance. Viruses depend on cellular molecules-which are attractive alternative targets-for their life cycle, provided that they are dispensable for normal cell functions. Using the model organism Drosophila melanogaster, we identify the ribosomal protein RACK1 as a cellular factor required for infection by internal ribosome entry site (IRES)-containing viruses. We further show that RACK1 is an essential determinant for hepatitis C virus translation and infection, indicating that its function is conserved for distantly related human and fly viruses. Inhibition of RACK1 does not affect Drosophila or human cell viability and proliferation, and RACK1-silenced adult flies are viable, indicating that this protein is not essential for general translation. Our findings demonstrate a specific function for RACK1 in selective mRNA translation and uncover a target for the development of broad antiviral intervention.}, keywords = {ERIANI, hoffmann, imler, M3i, meignin, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @book{, title = {Handbook of RNA Biochemistry}, editor = {R K Hartmann and A Bindereif and A Schon and E Westhof}, year = {2014}, date = {2014-01-01}, volume = {1}, publisher = {Wiley-Vch-Verlag}, edition = {OUVRAGE}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {book} } @article{marangon_covalent_2014, title = {Covalent Functionalization of Multi-walled Carbon Nanotubes with a Gadolinium Chelate for Efficient T1-Weighted Magnetic Resonance Imaging}, author = {Iris Marangon and Cécilia Ménard‐Moyon and Jelena Kolosnjaj‐Tabi and Marie Lys Béoutis and Lénaic Lartigue and Damien Alloyeau and Elzbieta Pach and Belén Ballesteros and Gwennhael Autret and Tsedev Ninjbadgar and Dermot F Brougham and Alberto Bianco and Florence Gazeau}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adfm.201402234}, doi = {10.1002/adfm.201402234}, issn = {1616-3028}, year = {2014}, date = {2014-01-01}, urldate = {2020-04-02}, journal = {Advanced Functional Materials}, volume = {24}, number = {45}, pages = {7173--7186}, abstract = {Given the promise of carbon nanotubes (CNTs) for photothermal therapy, drug delivery, tissue engineering, and gene therapy, there is a need for non-invasive imaging methods to monitor CNT distribution and fate in the body. In this study, non-ionizing whole-body high field magnetic resonance imaging (MRI) is used to follow the distribution of water-dispersible non-toxic functionalized CNTs administrated intravenously to mice. Oxidized CNTs are endowed with positive MRI contrast properties by covalent functionalization with the chelating ligand diethylenetriaminepentaacetic dianhydride (DTPA), followed by chelation to Gd3+. The structural and magnetic properties, MR relaxivities, cellular uptake, and application for MRI cell imaging of Gd-CNTs in comparison to the precursor oxidized CNTs are evaluated. Despite the intrinsic T2 contrast of oxidized CNTs internalized in macrophages, the anchoring of paramagnetic gadolinium onto the nanotube sidewall allows efficient T1 contrast and MR signal enhancement, which is preserved after CNT internalization by cells. Hence, due to their high dispersibility, Gd-CNTs have the potential to produce positive contrast in vivo following injection into the bloodstream. The uptake of Gd-CNTs in the liver and spleen is assessed using MRI, while rapid renal clearance of extracellular Gd-CNTs is observed, confirming the evidences of other studies using different imaging modalities.}, keywords = {Carbon nanotubes, contrast agents, I2CT, magnetic resonance imaging, Nanomedicine, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{gahoual_monoclonal_2014, title = {Monoclonal antibodies biosimilarity assessment using transient isotachophoresis capillary zone electrophoresis-tandem mass spectrometry.}, author = {Rabah Gahoual and Michaël Biacchi and Johana Chicher and Lauriane Kuhn and Philippe Hammann and Alain Beck and Emmanuelle Leize-Wagner and Yannis N François}, doi = {10.4161/mabs.36305}, issn = {1942-0870 1942-0862 1942-0862}, year = {2014}, date = {2014-01-01}, journal = {mAbs}, volume = {6}, number = {6}, pages = {1464--1473}, abstract = {Out of all categories, monoclonal antibody (mAb) therapeutics attract the most interest due to their strong therapeutic potency and specificity. Six of the 10 top-selling drugs are antibody-based therapeutics that will lose patent protection soon. The European Medicines Agency has pioneered the regulatory framework for approval of biosimilar products and approved the first biosimilar antibodies by the end of 2013. As highly complex glycoproteins with a wide range of micro-variants, mAbs require extensive characterization through multiple analytical methods for structure assessment rendering manufacturing control and biosimilarity studies particularly product and time-consuming. Here, capillary zone electrophoresis coupled to mass spectrometry by a sheathless interface (CESI-MS) was used to characterize marketed reference mAbs and their respective biosimilar candidate simultaneously over different facets of their primary structure. CESI-MS/MS data were compared between approved mAbs and their biosimilar candidates to prove/disconfirm biosimilarity regarding recent regulation directives. Using only a single sample injection of 200 fmol, CESI-MS/MS data enabled 100% amino acids (AA) sequence characterization, which allows a difference of even one AA between 2 samples to be distinguished precisely. Simultaneously glycoforms were characterized regarding their structures and position through fragmentation spectra and glycoforms semiquantitative analysis was established, showing the capacity of the developed methodology to detect up to 16 different glycans. Other posttranslational modifications hotspots were characterized while their relative occurrence levels were estimated and compared to biosimilars. These results proved the value of using CESI-MS because the separation selectivity and ionization efficiency provided by the system allowed substantial improvement in the characterization workflow robustness and accuracy. Biosimilarity assessment could be performed routinely with a single injection of each candidate enabling improvements in the biosimilar development pipeline.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{hardre_selective_2014, title = {The selective biotin tagging and thermolysin proteolysis of chloroplast outer envelope proteins reveals information on protein topology and association into complexes.}, author = {Hélène Hardré and Lauriane Kuhn and Catherine Albrieux and Juliette Jouhet and Morgane Michaud and Daphné Seigneurin-Berny and Denis Falconet and Maryse A Block and Eric Maréchal}, doi = {10.3389/fpls.2014.00203}, issn = {1664-462X 1664-462X 1664-462X}, year = {2014}, date = {2014-01-01}, journal = {Frontiers in plant science}, volume = {5}, pages = {203}, abstract = {The understanding of chloroplast function requires the precise localization of proteins in each of its sub-compartments. High-sensitivity mass spectrometry has allowed the inventory of proteins in thylakoid, stroma, and envelope fractions. Concerning membrane association, proteins can be either integral or peripheral or even soluble proteins bound transiently to a membrane complex. We sought a method providing information at the surface of the outer envelope membrane (OEM), based on specific tagging with biotin or proteolysis using thermolysin, a non-membrane permeable protease. To evaluate this method, envelope, thylakoid, and stroma proteins were separated by two-dimensional electrophoresis and analyzed by immunostaining and mass spectrometry. A short selection of proteins associated to the chloroplast envelope fraction was checked after superficial treatments of intact chloroplasts. We showed that this method could allow the characterization of OEM embedded proteins facing the cytosol, as well as peripheral and soluble proteins associated via tight or lose interactions. Some stromal proteins were associated with biotinylated spots and analyzes are still needed to determine whether polypeptides were tagged prior import or if they co-migrated with OEM proteins. This method also suggests that some proteins associated with the inner envelope membrane (IEM) might need the integrity of a trans-envelope (IEM-OEM) protein complex (e.g., division ring-forming components) or at least an intact OEM partner. Following this evaluation, proteomic analyzes should be refined and the putative role of inter-membrane space components stabilizing trans-envelope complexes demonstrated. For future comprehensive studies, perspectives include the dynamic analyses of OEM proteins and IEM-OEM complexes in various physiological contexts and using virtually any other purified membrane organelle.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{voisin_anatomical_2014, title = {Anatomical distribution analysis reveals lack of Langerin+ dermal dendritic cells in footpads and tail of C57BL/6 mice}, author = {Benjamin Voisin and David Gabriel Mairhofer and Suzie Chen and Patrizia Stoitzner and Christopher George Mueller and Vincent Flacher}, doi = {10.1111/exd.12373}, issn = {1600-0625}, year = {2014}, date = {2014-01-01}, journal = {Experimental Dermatology}, volume = {23}, number = {5}, pages = {354--356}, abstract = {Epidermal Langerhans cells (LCs) and dermal dendritic cells (dDCs) capture cutaneous antigens and present them to T-cells in lymph nodes (LNs). The function of LCs and Langerin+ dDCs was extensively studied in the mouse, but their anatomical repartition is unknown. Here, we found LCs in back skin, footpads and tail skin of C57BL/6, BALB/c, 129/Sv and CBA/J mice. Langerin+ dDCs were readily observed in back skin of all strains, but only in footpads and tail of BALB/c and CBA/J mice. Similarly, while LCs were equally present in all LNs and strains, Langerin+ dDCs were found in popliteal LNs (draining footpads) only in BALB/c and CBA/J mice. The sciatic LNs, which we identified as the major tail-draining lymphoid organ, were devoid of Langerin+ dDCs in all strains. Thus, functionally different DCs reside in different skin areas, with variations among mouse strains, implying a potential impact on the cutaneous immune reaction.}, keywords = {Analysis, Animals, Antigen, Antigens, C-Type, CD, CD11c Antigen, Cell Adhesion Molecules, Dendritic Cells, DERMAL DENDRITIC CELLS, Epithelial Cell Adhesion Molecule, footpad skin, function, Hindlimb, immunopathology, Inbred BALB C, Inbred C57BL, Inbred CBA, inflammation, Integrin alpha Chains, Langerhans Cells, Lectins, Letter, Leukocyte Common Antigens, LYMPH, LYMPH NODE, Lymph Nodes, Mannose-Binding Lectins, Mice, mouse, Neoplasm, Skin, skin-draining lymph nodes, Surface, T CELLS, T-CELLS, Tail, tail skin, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{hemmerlin_profiling_2014, title = {Profiling of defense responses in Escherichia coli treated with fosmidomycin.}, author = {Andréa Hemmerlin and Denis Tritsch and Philippe Hammann and Michel Rohmer and Thomas J Bach}, doi = {10.1016/j.biochi.2013.11.008}, issn = {1638-6183 0300-9084}, year = {2014}, date = {2014-01-01}, journal = {Biochimie}, volume = {99}, pages = {54--62}, abstract = {The mevalonate-independent isoprenoid biosynthesis pathway has been recognized as a promising target for designing new antibiotics. But pathogens treated with compounds such as fosmidomycin, a slow binding inhibitor of 1-deoxy-D-xylulose 5-phosphate reducto-isomerase, the second enzyme in this pathway, develop rapid drug resistance. In Escherichia coli, acquired resistance results mostly from inactivating the cAMP-dependent glpT transporter, thereby preventing import of the inhibitor. Such mutant strains are characterized by cross-resistance to fosfomycin, by susceptibility to efflux pump inhibitors, by disability to use glycerol 3-phosphate as a carbon source or by increased activity of the promoter controlling the expression of the glpABC regulon when grown in presence of fosmidomycin. The quite challenging task consists in conceiving new and efficient inhibitors avoiding resistance acquisition. They should be efficient in blocking the target enzyme, but should also be durably taken up by the organism. To address this issue, it is essential to characterize the mechanisms the pathogen exploits to defeat the antibiotic before resistance is acquired. Having this in mind, a 2-D Fluorescence Difference Gel Electrophoresis proteomic approach has been applied to identify defense responses in E. coli cells being shortly exposed to fosmidomycin (3 h). It seems that combined strategies are promptly induced. The major one consists in preventing toxic effects of the compound either by adapting metabolism and/or by getting rid of the molecule. The strategy adopted by the bacteria is to eliminate the drug from the cell or to increase the tolerance to oxidative stress. The design of new, but still efficient drugs, needs consideration of such rapid modulations required to adapt cell growth in contact of the inhibitor.}, note = {Place: France}, keywords = {1-Deoxy-d-xylulose 5-phosphate reducto-isomerase, Anti-Bacterial Agents/*pharmacology, Antibiotics, Bacterial, Disk Diffusion Antimicrobial Tests, Drug Resistance, Escherichia coli Proteins/*metabolism, Escherichia coli/drug effects/growth & development/*metabolism, Fosfomycin/*analogs & derivatives/pharmacology, Fosmidomycin, Isoprenoid, Oxidative Stress, Phenotype, PPSE, Proteome/metabolism, Resistance acquisition}, pubstate = {published}, tppubtype = {article} } @article{hammann_method_2014, title = {A method to map changes in bacterial surface composition induced by regulatory RNAs in Escherichia coli and Staphylococcus aureus.}, author = {P Hammann and D Parmentier and M Cerciat and J Reimegård and A-C Helfer and S Boisset and M Guillier and F Vandenesch and G E H Wagner and P Romby and P Fechter}, doi = {10.1016/j.biochi.2014.07.011}, issn = {1638-6183 0300-9084}, year = {2014}, date = {2014-01-01}, journal = {Biochimie}, volume = {106}, pages = {175--179}, abstract = {We have adapted a method to map cell surface proteins and to monitor the effect of specific regulatory RNAs on the surface composition of the bacteria. This method involves direct labeling of surface proteins of living bacteria using fluorescent dyes and a subsequent separation of the crude extract by 2D gel electrophoresis. The strategy yields a substantial enrichment in surface proteins over cytoplasmic proteins. We validated this method by monitoring the effect of the regulatory RNA MicA in Escherichia coli, which regulates the synthesis of several outer membrane proteins, and highlighted the role of Staphylococcus aureus RNAIII for the maintenance of cell wall integrity.}, note = {Place: France}, keywords = {Bacterial Outer Membrane Proteins/metabolism, Bacterial Proteins/*metabolism, Bacterial/genetics/*metabolism, Base Sequence, Carbocyanines/metabolism, Cell Membrane/*metabolism, Cell Wall/metabolism, Confocal, DIGE, Electrophoresis, Escherichia coli/genetics/*metabolism, Gel, Mass, Matrix-Assisted Laser Desorption-Ionization, Microscopy, Molecular Sequence Data, Non-coding RNAs, Post-transcriptional regulation, PPSE, Reproducibility of Results, RNA, Spectrometry, Staining and Labeling/methods, Staphylococcus aureus/genetics/*metabolism, Surface proteins, Two-Dimensional/methods}, pubstate = {published}, tppubtype = {article} } @article{mueller_emerging_2014, title = {Emerging immune functions of non-hematopoietic stromal cells}, author = {Christopher G Mueller and Mark Christopher Coles}, doi = {10.3389/fimmu.2014.00437}, issn = {1664-3224}, year = {2014}, date = {2014-01-01}, journal = {Frontiers in Immunology}, volume = {5}, pages = {437}, keywords = {development, immune cells, inflammation, Lymphoid Tissue, Stromal Cells, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{, title = {Specific recognition of the HIV-1 genomic RNA by the Gag precursor.}, author = {E W Abd El-Wahab and R P Smyth and E Mailler and S Bernacchi and V Vivet-Boudou and M Hijnen and F Jossinet and J Mak and J C Paillart and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24986025?dopt=Abstract}, doi = {10.1038/ncomms5304}, isbn = {24986025}, year = {2014}, date = {2014-01-01}, journal = {Nat Commun}, volume = {5}, pages = {4304}, abstract = {During assembly of HIV-1 particles in infected cells, the viral Pr55(Gag) protein (or Gag precursor) must select the viral genomic RNA (gRNA) from a variety of cellular and viral spliced RNAs. However, there is no consensus on how Pr55(Gag) achieves this selection. Here, by using RNA binding and footprinting assays, we demonstrate that the primary Pr55(Gag) binding site on the gRNA consists of the internal loop and the lower part of stem-loop 1 (SL1), the upper part of which initiates gRNA dimerization. A double regulation ensures specific binding of Pr55(Gag) to the gRNA despite the fact that SL1 is also present in spliced viral RNAs. The region upstream of SL1, which is present in all HIV-1 RNAs, prevents binding to SL1, but this negative effect is counteracted by sequences downstream of SL4, which are unique to the gRNA.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{lamanna_adamantane-based_2014, title = {Adamantane-based dendrons for trimerization of the therapeutic P140 peptide}, author = {Giuseppe Lamanna and Maxime Grillaud and Christophe Macri and Olivier Chaloin and Sylviane Muller and Alberto Bianco}, doi = {10.1016/j.biomaterials.2014.05.017}, issn = {1878-5905}, year = {2014}, date = {2014-01-01}, journal = {Biomaterials}, volume = {35}, number = {26}, pages = {7553--7561}, abstract = {Dendrons constituted of an adamantane core, a focal point and three arms, were synthetized starting from a multifunctional adamantane derivative. Maleimido groups at the periphery of the scaffold were used to covalently attach the peptide called P140, a therapeutic phosphopeptide controlling disease activity in systemic lupus, both in mice and patients. Biotinylation of the trimers at the focal point was performed using click chemistry and the conjugates were studied in terms of solubility, binding affinity to its receptor, the HSPA8/HSC70 chaperone protein, effect on HSPA8 folding property and in vivo activity. The results showed that the trimerization of P140 peptide does not trigger aggregation or steric hindrances during the interaction with HSPA8 protein. Compared to the monomeric cognate peptide, the trivalent P140 peptide displayed the same capacity, in vitro, to down-regulate HSPA8 activity and, in vivo in MRL/lpr lupus-prone mice, to reduce abnormal blood hypercellularity. The control trimer synthesized with the same scaffold and a scrambled sequence of P140 showed no effect in vivo. This work reveals that adamantane-based scaffolds with a well-defined spatial conformation are promising trivalent systems for molecular recognition and for biomedical applications.}, keywords = {Adamantane, Animals, Biotin, C3-symmetry, Dendrimers, Dendrons, Drug Carriers, Female, HSC70 Heat-Shock Proteins, HSPA8 protein, Humans, I2CT, Inbred MRL lpr, Lupus Erythematosus, Mice, P140 peptide, Peptide Fragments, Systemic, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{servant_graphene_2014, title = {Graphene for multi-functional synthetic biology: the last 'zeitgeist' in nanomedicine}, author = {A Servant and A Bianco and M Prato and K Kostarelos}, doi = {10.1016/j.bmcl.2014.01.051}, issn = {1464-3405}, year = {2014}, date = {2014-01-01}, journal = {Bioorganic & Medicinal Chemistry Letters}, volume = {24}, number = {7}, pages = {1638--1649}, abstract = {The high versatility of graphene has attracted significant attention in many areas of scientific research from electronics to physics and mechanics. One of the most intriguing utilisation of graphene remains however in nanomedicine and synthetic biology. In particular, the last decade has witnessed an exponential growth in the generation of novel candidate therapeutics of multiple biological activities based on graphene constructs with small molecules, such as anti-cancer drugs. In this Digest, we summarise the different synthetic strategies and routes available to fabricate these promising graphene conjugates and the opportunities for the design of multi-functional tools for synthetic biology that they offer.}, keywords = {Antineoplastic Agents, carbon, Drug delivery, Drug Design, Graphite, I2CT, Nanomaterials, Nanomedicine, nanotechnology, Synthetic Biology, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{petrillo_cytoplasmic_2013, title = {Cytoplasmic granule formation and translational inhibition of nodaviral RNAs in the absence of the double-stranded RNA binding protein B2}, author = {Jessica E Petrillo and Arno P Venter and James R Short and Radhika Gopal and Safia Deddouche and Olivier Lamiable and Jean-Luc Imler and Anette Schneemann}, doi = {10.1128/JVI.02362-13}, issn = {1098-5514}, year = {2013}, date = {2013-12-01}, journal = {Journal of Virology}, volume = {87}, number = {24}, pages = {13409--13421}, abstract = {Flock House virus (FHV) is a positive-sense RNA insect virus with a bipartite genome. RNA1 encodes the RNA-dependent RNA polymerase, and RNA2 encodes the capsid protein. A third protein, B2, is translated from a subgenomic RNA3 derived from the 3' end of RNA1. B2 is a double-stranded RNA (dsRNA) binding protein that inhibits RNA silencing, a major antiviral defense pathway in insects. FHV is conveniently propagated in Drosophila melanogaster cells but can also be grown in mammalian cells. It was previously reported that B2 is dispensable for FHV RNA replication in BHK21 cells; therefore, we chose this cell line to generate a viral mutant that lacked the ability to produce B2. Consistent with published results, we found that RNA replication was indeed vigorous but the yield of progeny virus was negligible. Closer inspection revealed that infected cells contained very small amounts of coat protein despite an abundance of RNA2. B2 mutants that had reduced affinity for dsRNA produced analogous results, suggesting that the dsRNA binding capacity of B2 somehow played a role in coat protein synthesis. Using fluorescence in situ hybridization of FHV RNAs, we discovered that RNA2 is recruited into large cytoplasmic granules in the absence of B2, whereas the distribution of RNA1 remains largely unaffected. We conclude that B2, by binding to double-stranded regions in progeny RNA2, prevents recruitment of RNA2 into cellular structures, where it is translationally silenced. This represents a novel function of B2 that further contributes to successful completion of the nodaviral life cycle.}, keywords = {Animals, Capsid Proteins, Cell Line, Cricetinae, Cytoplasmic Granules, Double-Stranded, imler, M3i, Nodaviridae, Protein Biosynthesis, RNA, RNA Virus Infections, RNA-Binding Proteins, Viral, Viral Proteins}, pubstate = {published}, tppubtype = {article} } @article{bianco_all_2013, title = {All in the graphene family – A recommended nomenclature for two-dimensional carbon materials}, author = {Alberto Bianco and Hui-Ming Cheng and Toshiaki Enoki and Yury Gogotsi and Robert H Hurt and Nikhil Koratkar and Takashi Kyotani and Marc Monthioux and Chong Rae Park and Juan M D Tascon and Jin Zhang}, url = {http://www.sciencedirect.com/science/article/pii/S0008622313008002}, doi = {10.1016/j.carbon.2013.08.038}, issn = {0008-6223}, year = {2013}, date = {2013-12-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {65}, pages = {1--6}, abstract = {Interest in two-dimensional, sheet-like or flake-like carbon forms has expanded beyond monolayer graphene to include related materials with significant variations in layer number, lateral dimension, rotational faulting, and chemical modification. Describing this family of “graphene materials” has been causing confusion in the Carbon journal and in the scientific literature as a whole. The international editorial team for Carbon believes that the time has come for a discussion on a rational naming system for two-dimensional carbon forms. We propose here a first nomenclature for two-dimensional carbons that could guide authors toward a more precise description of their subject materials, and could allow the field to move forward with a higher degree of common understanding.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{kabbage_tropomyosin-4_2013, title = {Tropomyosin-4 correlates with higher SBR grades and tubular differentiation in infiltrating ductal breast carcinomas: an immunohistochemical and proteomics-based study.}, author = {Maria Kabbage and Mounir Trimeche and Hela Ben Nasr and Philippe Hammann and Lauriane Kuhn and Bechr Hamrita and Karim Chahed}, doi = {10.1007/s13277-013-0939-0}, issn = {1423-0380 1010-4283}, year = {2013}, date = {2013-12-01}, journal = {Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine}, volume = {34}, number = {6}, pages = {3593--3602}, abstract = {The aim of this study is to evaluate tropomyosin-4 (TM4) expression in infiltrating ductal breast carcinomas (IDCAs), as well as its prognostic significance. Using a 2-DE/MALDI-TOF mass spectrometry investigation coupled with an immunohistochemical approach, we have assessed the expression of TM4 in IDCAs, as well as in other types of breast tumors. Proteomic analyses revealed an increased expression of tropomyosin-4 in IDCA tumors. Using immunohistochemistry, overexpression of tropomyosin-4 was confirmed in 51 additional tumor specimens. Statistical analyses revealed, however, no significant correlations between tropomyosin-4 expression and clinicopathological parameters of the disease including tumor stage, patient age, estrogen and progesterone receptor status, and lymph node metastasis occurrence. A significant association was found, however, with a high Scarf-Bloom-Richardson (SBR) grade, a known marker of tumor severity. Additionally, the SBR component showing a correlation with TM4 expression was the tubular differentiation status. This study demonstrates the upregulation of tropomyosin-4 in IDCA tissues, which may highlight its involvement in breast cancer development. Our findings also support a link between tropomyosin-4 expression and aggressiveness of IDCA tumors.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{schaeffer_dynamic_2013, title = {Dynamic micelles of mannoside glycolipids are more efficient than polymers for inhibiting HIV-1 trans-infection}, author = {Evelyne Schaeffer and Laure Dehuyser and David Sigwalt and Vincent Flacher and Serena Bernacchi and Olivier Chaloin and Jean-Serge Remy and Christopher G Mueller and Rachid Baati and Alain Wagner}, doi = {10.1021/bc4000806}, issn = {1520-4812}, year = {2013}, date = {2013-11-01}, journal = {Bioconjugate Chemistry}, volume = {24}, number = {11}, pages = {1813--1823}, abstract = {Mannoside glycolipid conjugates are able to inhibit human immunodeficiency virus type 1 (HIV-1) trans-infection mediated by human dendritic cells (DCs). The conjugates are formed by three building blocks: a linear or branched mannose head, a hydrophilic linker, and a 24-carbon lipid chain. We have shown that, even as single molecules, these compounds efficiently target mannose-binding lectins, such as DC-specific ICAM-3-grabbing nonintegrin (DC-SIGN) important for HIV-1 transmission. With the goal to optimize their inhibitory activity by supramolecular structure formation, we have compared saturated and unsaturated conjugates, as single molecules, self-assemblies of dynamic micelles, and photopolymerized cross-linked polymers. Surface plasmon resonance showed that, unexpectedly, polymers of trivalent conjugates did not display a higher binding affinity for DC-SIGN than single molecules. Interactions on a chip or in solution were independent of calcium; however, binding to DCs was inhibited by a calcium chelator. Moreover, HIV-1 trans-infection was mostly inhibited by dynamic micelles and not by rigid polymers. The inhibition data revealed a clear correlation between the structure and molecular assembly of a conjugate and its biological antiviral activity. We present an interaction model between DC-SIGN and conjugates-either single molecules, micelles, or polymers-that highlights that the most effective interactions by dynamic micelles involve both mannose heads and lipid chains. Our data reveal that trivalent glycolipid conjugates display the highest microbicide potential for HIV prophylaxis, as dynamic micelles conjugates and not as rigid polymers.}, keywords = {Anti-HIV Agents, Calcium, Cells, Chemistry, Cultured, Dendritic Cells, Dose-Response Relationship, Drug, Electron, fluorescence, Glycolipids, HIV, HIV Infections, HIV-1, Human, Humans, immunodeficiency, immunopathology, inhibition, LECTIN, Lectins, lipid, Mannosides, Micelles, Microbial Sensitivity Tests, Microscopy, Models, Molecular, Molecular Structure, Polymers, prophylaxis, Spectrometry, Structure-Activity Relationship, Surface Plasmon Resonance, target, Team-Mueller, Thermodynamics, Transmission, virus}, pubstate = {published}, tppubtype = {article} } @article{AL2013, title = {Targeted Mutagenesis in the Malaria Mosquito Using TALE Nucleases}, author = {Andie L Smidler and Olivier Terenzi and Julien Soichot and Elena A Levashina and Eric Marois}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23977401}, year = {2013}, date = {2013-08-15}, journal = {PLoS One}, volume = {8}, number = {8}, pages = {e74511}, abstract = {Anopheles gambiae, the main mosquito vector of human malaria, is a challenging organism to manipulate genetically. As a consequence, reverse genetics studies in this disease vector have been largely limited to RNA interference experiments. Here, we report the targeted disruption of the immunity gene TEP1 using transgenic expression of Transcription-Activator Like Effector Nucleases (TALENs), and the isolation of several TEP1 mutant A. gambiae lines. These mutations inhibited protein production and rendered TEP1 mutants hypersusceptible to Plasmodium berghei. The TALEN technology opens up new avenues for genetic analysis in this disease vector and may offer novel biotechnology-based approaches for malaria control.}, keywords = {M3i, marois, Mutagenesis, TALEN}, pubstate = {published}, tppubtype = {article} } @article{pmid23660570, title = {Non-coding RNAs: novel players in chromatin-regulation during viral latency}, author = {Sebastian Eilebrecht and Christian Schwartz and Olivier Rohr}, doi = {10.1016/j.coviro.2013.04.001}, issn = {1879-6265}, year = {2013}, date = {2013-08-01}, urldate = {2013-08-01}, journal = {Curr Opin Virol}, volume = {3}, number = {4}, pages = {387--393}, abstract = {Chromatin structure plays an essential role during gene expression regulation not only in the case of the host cellular genome, but also during the viral life cycle. Epigenetic chromatin marks thereby define, whether a gene promoter is accessible for the transcription machinery or whether a repressive heterochromatin state is established. The heterochromatin-mediated repression of lytic viral genes results in viral latency, enabling the virus to persist dormant without being recognized by the host immune system, but keeping the potential for reactivation. Arising new systems biology approaches are starting to uncover an unexpected multiplicity and variety of non-coding (nc)RNAs playing important roles during chromatin structure control, likely constituting a novel layer in epigenetic regulation. In this review we give an overview of chromatin-regulatory viral and host cellular ncRNAs and their links to viral latency.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid23852730, title = {CTIP2 is a negative regulator of P-TEFb}, author = {Thomas Cherrier and Valentin Le Douce and Sebastian Eilebrecht and Raphael Riclet and Céline Marban and Franck Dequiedt and Yannick Goumon and Jean-Christophe Paillart and Mathias Mericskay and Ara Parlakian and Pedro Bausero and Wasim Abbas and Georges Herbein and Siavash K Kurdistani and Xavier Grana and Benoit Van Driessche and Christian Schwartz and Ermanno Candolfi and Arndt G Benecke and Carine Van Lint and Olivier Rohr}, doi = {10.1073/pnas.1220136110}, issn = {1091-6490}, year = {2013}, date = {2013-07-01}, urldate = {2013-07-01}, journal = {Proc Natl Acad Sci U S A}, volume = {110}, number = {31}, pages = {12655--12660}, abstract = {The positive transcription elongation factor b (P-TEFb) is involved in physiological and pathological events including inflammation, cancer, AIDS, and cardiac hypertrophy. The balance between its active and inactive form is tightly controlled to ensure cellular integrity. We report that the transcriptional repressor CTIP2 is a major modulator of P-TEFb activity. CTIP2 copurifies and interacts with an inactive P-TEFb complex containing the 7SK snRNA and HEXIM1. CTIP2 associates directly with HEXIM1 and, via the loop 2 of the 7SK snRNA, with P-TEFb. In this nucleoprotein complex, CTIP2 significantly represses the Cdk9 kinase activity of P-TEFb. Accordingly, we show that CTIP2 inhibits large sets of P-TEFb- and 7SK snRNA-sensitive genes. In hearts of hypertrophic cardiomyopathic mice, CTIP2 controls P-TEFb-sensitive pathways involved in the establishment of this pathology. Overexpression of the β-myosin heavy chain protein contributes to the pathological cardiac wall thickening. The inactive P-TEFb complex associates with CTIP2 at the MYH7 gene promoter to repress its activity. Taken together, our results strongly suggest that CTIP2 controls P-TEFb function in physiological and pathological conditions.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid23855931, title = {Pseudomonas DING proteins as human transcriptional regulators and HIV-1 antagonists}, author = {Andrew Suh and Valentin Le Douce and Olivier Rohr and Christian Schwartz and Ken Scott}, doi = {10.1186/1743-422X-10-234}, issn = {1743-422X}, year = {2013}, date = {2013-07-01}, urldate = {2013-07-01}, journal = {Virol J}, volume = {10}, pages = {234}, abstract = {BACKGROUND: Anti-HIV-1 therapy depends upon multiple agents that target different phases of the viral replication cycle. Recent reports indicate that plant and human DING proteins are unique in targeting viral gene transcription as the basis of their anti-HIV-1 therapy.nnMETHODS: Two cloned DING genes from Pseudomonas were transiently expressed in human cells, and effects on NFκB-mediated transcription, HIV-1 transcription, and HIV-1 production were measured.nnRESULTS: Both DING proteins elevated NFκB-mediated transcription. In microglial cells, one protein, from P. aeruginosa PA14, suppressed HIV-1 transcription; the other protein, from P. fluorescens SBW25, was inactive. The PA14DING protein also reduces HIV-1 production in microglial cells.nnCONCLUSIONS: Structural differences between the two DING proteins highlight regions of the PA14DING protein essential to the anti-HIV-1 activity, and may guide the design of therapeutic agents.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{Batisse2013, title = {APOBEC3G impairs the multimerization of the HIV-1 Vif protein in living cells}, author = {J Batisse and S X Guerrero and S Bernacchi and L Richert and J Godet and V Goldschmidt and Y Mély and R Marquet and H de Rocquigny and J C Paillart }, url = {https://pubmed.ncbi.nlm.nih.gov/23576497/}, doi = {10.1128/JVI.03494-12}, year = {2013}, date = {2013-07-01}, journal = {Journal of Virology}, volume = {87}, number = {11}, pages = {6492-506}, abstract = {The HIV-1 viral infectivity factor (Vif) is a small basic protein essential for viral fitness and pathogenicity. Vif allows productive infection in nonpermissive cells, including most natural HIV-1 target cells, by counteracting the cellular cytosine deaminases APOBEC3G (apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3G [A3G]) and A3F. Vif is also associated with the viral assembly complex and packaged into viral particles through interactions with the viral genomic RNA and the nucleocapsid domain of Pr55(Gag). Recently, we showed that oligomerization of Vif into high-molecular-mass complexes induces Vif folding and influences its binding to high-affinity RNA binding sites present in the HIV genomic RNA. To get further insight into the role of Vif multimerization in viral assembly and A3G repression, we used fluorescence lifetime imaging microscopy (FLIM)- and fluorescence resonance energy transfer (FRET)-based assays to investigate Vif-Vif interactions in living cells. By using two N-terminally tagged Vif proteins, we show that Vif-Vif interactions occur in living cells. This oligomerization is strongly reduced when the putative Vif multimerization domain ((161)PPLP(164)) is mutated, indicating that this domain is crucial, but that regions outside this motif also participate in Vif oligomerization. When coexpressed together with Pr55(Gag), Vif is largely relocated to the cell membrane, where Vif oligomerization also occurs. Interestingly, wild-type A3G strongly interferes with Vif multimerization, contrary to an A3G mutant that does not bind to Vif. These findings confirm that Vif oligomerization occurs in living cells partly through its C-terminal motif and suggest that A3G may target and perturb the Vif oligomerization state to limit its functions in the cell. }, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{M2013, title = {Silencing of Genes and Alleles by RNAi in Anopheles gambiae}, author = {Marina Lamacchia and John Randy Clayton and R Wang-Sattler and Lars M Steinmetz and Elena A Levashina and Stéphanie A Blandin}, year = {2013}, date = {2013-06-13}, journal = {Methods Mol Biol.}, volume = {923}, pages = {161-76}, abstract = {Anopheles gambiae mosquitoes are the major vectors of human malaria parasites. However, mosquitoes are not passive hosts for parasites, actively limiting their development in vivo. Our current understanding of the mosquito antiparasitic response is mostly based on the phenotypic analysis of gene knockdowns obtained by RNA interference (RNAi), through the injection or transfection of long dsRNAs in adult mosquitoes or cultured cells, respectively. Recently, RNAi has been extended to silence specifically one allele of a given gene in a heterozygous context, thus allowing to compare the contribution of different alleles to a phenotype in the same genetic background.}, keywords = {blandin, dsRNA, M3i, RNAi}, pubstate = {published}, tppubtype = {article} } @article{fukuyama_landscape_2013, title = {Landscape of protein-protein interactions in Drosophila immune deficiency signaling during bacterial challenge}, author = {Hidehiro Fukuyama and Yann Verdier and Yongsheng Guan and Chieko Makino-Okamura and Victoria Shilova and Xi Liu and Elie Maksoud and Jun Matsubayashi and Iman Haddad and Kerstin Spirohn and Kenichiro Ono and Charles Hetru and Jean Rossier and Trey Ideker and Michael Boutros and Joëlle Vinh and Jules A Hoffmann}, doi = {10.1073/pnas.1304380110}, issn = {1091-6490}, year = {2013}, date = {2013-06-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {110}, number = {26}, pages = {10717--10722}, abstract = {The Drosophila defense against pathogens largely relies on the activation of two signaling pathways: immune deficiency (IMD) and Toll. The IMD pathway is triggered mainly by Gram-negative bacteria, whereas the Toll pathway responds predominantly to Gram-positive bacteria and fungi. The activation of these pathways leads to the rapid induction of numerous NF-κB-induced immune response genes, including antimicrobial peptide genes. The IMD pathway shows significant similarities with the TNF receptor pathway. Recent evidence indicates that the IMD pathway is also activated in response to various noninfectious stimuli (i.e., inflammatory-like reactions). To gain a better understanding of the molecular machinery underlying the pleiotropic functions of this pathway, we first performed a comprehensive proteomics analysis to identify the proteins interacting with the 11 canonical members of the pathway initially identified by genetic studies. We identified 369 interacting proteins (corresponding to 291 genes) in heat-killed Escherichia coli-stimulated Drosophila S2 cells, 92% of which have human orthologs. A comparative analysis of gene ontology from fly or human gene annotation databases points to four significant common categories: (i) the NuA4, nucleosome acetyltransferase of H4, histone acetyltransferase complex, (ii) the switching defective/sucrose nonfermenting-type chromatin remodeling complex, (iii) transcription coactivator activity, and (iv) translation factor activity. Here we demonstrate that sumoylation of the IκB kinase homolog immune response-deficient 5 plays an important role in the induction of antimicrobial peptide genes through a highly conserved sumoylation consensus site during bacterial challenge. Taken together, the proteomics data presented here provide a unique avenue for a comparative functional analysis of proteins involved in innate immune reactions in flies and mammals.}, keywords = {Amino Acid, Animals, Chromatin Assembly and Disassembly, Escherichia coli, functional proteomics, Genes, Genetically Modified, Histone Acetyltransferases, hoffmann, Host-Pathogen Interactions, Humans, IMD interactome, Insect, M3i, Models, Molecular, Protein Interaction Maps, Sequence Homology, Signal Transduction, small ubiquitin-like modifier}, pubstate = {published}, tppubtype = {article} } @article{gahoual_rapid_2013, title = {Rapid and multi-level characterization of trastuzumab using sheathless capillary electrophoresis-tandem mass spectrometry.}, author = {Rabah Gahoual and Alicia Burr and Jean-Marc Busnel and Lauriane Kuhn and Phillipe Hammann and Alain Beck and Yannis-Nicolas François and Emmanuelle Leize-Wagner}, doi = {10.4161/mabs.23995}, issn = {1942-0870 1942-0862 1942-0862}, year = {2013}, date = {2013-06-01}, journal = {mAbs}, volume = {5}, number = {3}, pages = {479--490}, abstract = {Monoclonal antibodies (mAbs) are highly complex proteins that display a wide range of microheterogeneity that requires multiple analytical methods for full structure assessment and quality control. As a consequence, the characterization of mAbs on different levels is particularly product - and time - consuming. This work presents the characterization of trastuzumab sequence using sheathless capillary electrophoresis (referred as CESI) - tandem mass spectrometry (CESI-MS/MS). Using this bottom-up proteomic-like approach, CESI-MS/MS provided 100% sequence coverage for both heavy and light chain via peptide fragment fingerprinting (PFF) identification. The result was accomplished in a single shot, corresponding to the analysis of 100 fmoles of digest. The same analysis also enabled precise characterization of the post-translational hot spots of trastuzumab, used as a representative widely marketed therapeutic mAb, including the structural confirmation of the five major N-glycoforms.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{SE2013, title = {AP-1/Fos-TGase2 axis mediates wounding-induced Plasmodium falciparum killing in Anopheles gambiae}, author = {Sandrine E Nsango and Julien Pompon and Ting Xie and Annika Rademacher and Malou Fraiture and Martine Thoma and P H Awono-Ambene and R S Moyou and Isabelle Morlais and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23592781}, year = {2013}, date = {2013-05-31}, journal = {J Biol Chem.}, volume = {288}, number = {22}, pages = {16145-54}, abstract = {Anopheline mosquitoes are the only vectors of human malaria worldwide. It is now widely accepted that mosquito immune responses play a crucial role in restricting Plasmodium development within the vector; therefore, further dissection of the molecular mechanisms underlying these processes should inform new vector control strategies urgently needed to roll back the disease. Here, using genome-wide transcriptional profiling, bioinformatics, and functional gene analysis, we identify a new axis of mosquito resistance to monoclonal Plasmodium falciparum infections that includes the AP-1 transcription factor Fos and the transglutaminase 2 (TGase2), a cross-linking enzyme with known roles in wound responses. We demonstrate that Fos regulates induction of TGase2 expression after wounding but does not affect expression of the components of the well characterized complement-like system. Silencing of Fos or of TGase2 aborts the wounding-induced mosquito killing of P. falciparum. These results reveal multiple signaling pathways that are required for efficient Plasmodium killing in Anopheles gambiae.}, keywords = {AP-1, Fos-TGase2, wounding}, pubstate = {published}, tppubtype = {article} } @article{kobayashi_shigella_2013, title = {The Shigella OspC3 effector inhibits caspase-4, antagonizes inflammatory cell death, and promotes epithelial infection}, author = {Taira Kobayashi and Michinaga Ogawa and Takahito Sanada and Hitomi Mimuro and Minsoo Kim and Hiroshi Ashida and Reiko Akakura and Mitsutaka Yoshida and Magdalena Kawalec and Jean-Marc Reichhart and Tsunehiro Mizushima and Chihiro Sasakawa}, doi = {10.1016/j.chom.2013.04.012}, issn = {1934-6069}, year = {2013}, date = {2013-05-01}, journal = {Cell Host Microbe}, volume = {13}, number = {5}, pages = {570--583}, abstract = {Caspase-mediated inflammatory cell death acts as an intrinsic defense mechanism against infection. Bacterial pathogens deploy countermeasures against inflammatory cell death, but the mechanisms by which they do this remain largely unclear. In a screen for Shigella flexneri effectors that regulate cell death during infection, we discovered that Shigella infection induced acute inflammatory, caspase-4-dependent epithelial cell death, which is counteracted by the bacterial OspC3 effector. OspC3 interacts with the caspase-4-p19 subunit and inhibits its activation by preventing caspase-4-p19 and caspase-4-p10 heterodimerization by depositing the conserved OspC3 X1-Y-X₂-D-X₃ motif at the putative catalytic pocket of caspase-4. Infection of guinea pigs with a Shigella ospC3-deficient mutant resulted in enhanced inflammatory cell death and associated symptoms, correlating with decreased bacterial burdens. Salmonella Typhimurium and enteropathogenic Escherichia coli infection also induced caspase-4-dependent epithelial death. These findings highlight the importance of caspase-4-dependent innate immune responses and demonstrate that Shigella delivers a caspase-4-specific inhibitor to delay epithelial cell death and promote infection.}, keywords = {Animal, Animals, Bacillary, Bacterial, Bacterial Proteins, Caspases, Cell Death, Cell Line, Disease Models, DNA, Dysentery, Enzyme Inhibitors, Epithelial Cells, Escherichia coli, Gene Knockout Techniques, Guinea Pigs, Host-Pathogen Interactions, Humans, Initiator, M3i, Protein Binding, Protein Interaction Mapping, reichhart, Salmonella typhimurium, Sequence Analysis, Shigella flexneri, Virulence Factors}, pubstate = {published}, tppubtype = {article} } @article{kabbage_calreticulin_2013, title = {Calreticulin expression in infiltrating ductal breast carcinomas: relationships with disease progression and humoral immune responses.}, author = {Maria Kabbage and Mounir Trimeche and Sarra Bergaoui and Philippe Hammann and Lauriane Kuhn and Bechr Hamrita and Hela ben Nasr and Anouar Chaieb and Lotfi Chouchane and Karim Chahed}, doi = {10.1007/s13277-013-0661-y}, issn = {1423-0380 1010-4283}, year = {2013}, date = {2013-04-01}, journal = {Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine}, volume = {34}, number = {2}, pages = {1177--1188}, abstract = {The aim of this study was to evaluate calreticulin expression in infiltrating ductal breast carcinomas (IDCAs), as well as its relationships with clinicopathological parameters of the disease. Using a two-dimensional gel electrophoresis/matrix-assisted laser desorption ionization time of flight mass spectrometry investigation coupled to an immunohistochemical approach, we have assessed the expression of calreticulin in IDCAs, as well as in other types of breast tumors. The humoral immune response against calreticulin was estimated using a serological proteomics-based strategy. Proteomic analyses revealed an increased expression of calreticulin in IDCA tumors. Using immunohistochemistry, overexpression of calreticulin was confirmed in 51 additional tumor specimens. Statistical analyses revealed, however, no significant correlations between calreticulin expression and clinicopathological parameters of the disease including tumor stage, patient age, SBR grade, and lymph node metastasis occurrence. A significant association was found, however, with estrogen receptor status. This study demonstrates the upregulation of calreticulin in IDCA tissues which may highlight its involvement in breast cancer development. Our findings also support a link between calreticulin expression and estrogen transduction pathways. Our results do not, however, support the involvement of calreticulin in the development of a humoral immune response in IDCAs.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{vanBerge2013, title = {Pathogenic mutations causing LBSL affect mitochondrial aspartyl-tRNA synthetase in diverse ways}, author = { L van Berge and J Kevenaar and E Polder and A Gaudry and C Florentz and M Sissler and M S van der Knaap and G C Scheper }, url = {https://pubmed.ncbi.nlm.nih.gov/23216004/}, doi = {10.1042/BJ20121564 }, year = {2013}, date = {2013-03-04}, journal = {Biochemical Journal}, volume = {450}, number = {2}, pages = {345-50}, abstract = {The autosomal recessive white matter disorder LBSL (leukoencephalopathy with brain stem and spinal cord involvement and lactate elevation) is caused by mutations in DARS2, coding for mtAspRS (mitochondrial aspartyl-tRNA synthetase). Generally, patients are compound heterozygous for mutations in DARS2. Many different mutations have been identified in patients, including several missense mutations. In the present study, we have examined the effects of missense mutations found in LBSL patients on the expression, enzyme activity, localization and dimerization of mtAspRS, which is important for understanding the cellular defect underlying the pathogenesis of the disease. Nine different missense mutations were analysed and were shown to have various effects on mtAspRS properties. Several mutations have a direct effect on the catalytic activity of the enzyme; others have an effect on protein expression or dimerization. Most mutations have a clear impact on at least one of the properties of mtAspRS studied, probably resulting in a small contribution of the missense variants to the mitochondrial aspartylation activity in the cell. }, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{quintin_drosophila_2013b, title = {The Drosophila Toll pathway controls but does not clear Candida glabrata infections}, author = {Jessica Quintin and Joelle Asmar and Alexey A Matskevich and Marie-Céline Lafarge and Dominique Ferrandon}, doi = {10.4049/jimmunol.1201861}, issn = {1550-6606}, year = {2013}, date = {2013-03-01}, journal = {J. Immunol.}, volume = {190}, number = {6}, pages = {2818--2827}, abstract = {The pathogenicity of Candida glabrata to patients remains poorly understood for lack of convenient animal models to screen large numbers of mutants for altered virulence. In this study, we explore the minihost model Drosophila melanogaster from the dual perspective of host and pathogen. As in vertebrates, wild-type flies contain C. glabrata systemic infections yet are unable to kill the injected yeasts. As for other fungal infections in Drosophila, the Toll pathway restrains C. glabrata proliferation. Persistent C. glabrata yeasts in wild-type flies do not appear to be able to take shelter in hemocytes from the action of the Toll pathway, the effectors of which remain to be identified. Toll pathway mutant flies succumb to injected C. glabrata. In this immunosuppressed background, cellular defenses provide a residual level of protection. Although both the Gram-negative binding protein 3 pattern recognition receptor and the Persephone protease-dependent detection pathway are required for Toll pathway activation by C. glabrata, only GNBP3, and not psh mutants, are susceptible to the infection. Both Candida albicans and C. glabrata are restrained by the Toll pathway, yet the comparative study of phenoloxidase activation reveals a differential activity of the Toll pathway against these two fungal pathogens. Finally, we establish that the high-osmolarity glycerol pathway and yapsins are required for virulence of C. glabrata in this model. Unexpectedly, yapsins do not appear to be required to counteract the cellular immune response but are needed for the colonization of the wild-type host.}, keywords = {Adaptor Proteins, Animal, Animals, Antigens, Candida glabrata, Candidiasis, Cells, Cultured, Differentiation, Disease Models, ferrandon, Immunologic, M3i, Phagocytosis, Receptors, Signal Transducing, Signal Transduction, Toll-Like Receptors, Virulence}, pubstate = {published}, tppubtype = {article} } @article{serag_nanobiotechnology_2013, title = {Nanobiotechnology meets plant cell biology: carbon nanotubes as organelle targeting nanocarriers}, author = {Maged F Serag and Noritada Kaji and Satoshi Habuchi and Alberto Bianco and Yoshinobu Baba}, url = {https://pubs.rsc.org/en/content/articlelanding/2013/ra/c2ra22766e}, doi = {10.1039/C2RA22766E}, issn = {2046-2069}, year = {2013}, date = {2013-03-01}, urldate = {2020-04-01}, journal = {RSC Advances}, volume = {3}, number = {15}, pages = {4856--4862}, abstract = {For years, nanotechnology has shown great promise in the fields of biomedical and biotechnological sciences and medical research. In this review, we demonstrate its versatility and applicability in plant cell biology studies. Specifically, we discuss the ability of functionalized carbon nanotubes to penetrate the plant cell wall, target specific organelles, probe protein-carrier activity and induce organelle recycling in plant cells. We also, shed light on prospective applications of carbon nanomaterials in cell biology and plant cell transformation.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{poirier_strategies_2013, title = {Strategies developed by the marine bacterium Pseudomonas fluorescens BA3SM1 to resist metals: A proteome analysis.}, author = {Isabelle Poirier and Philippe Hammann and Lauriane Kuhn and Martine Bertrand}, doi = {10.1016/j.aquatox.2012.12.006}, issn = {1879-1514 0166-445X}, year = {2013}, date = {2013-03-01}, journal = {Aquatic toxicology (Amsterdam, Netherlands)}, volume = {128-129}, pages = {215--232}, abstract = {A global proteomic evaluation of the response of the marine bacterium Pseudomonas fluorescens BA3SM1 to Cd, Zn and Cu was performed by two dimensional gel electrophoresis followed by mass spectrometry. When stressed with Cd, the most toxic metal for P. fluorescens BA3SM1, cell growth is rapidly affected and the number of proteins up-regulated (sixteen for 0.4 mM Cd) remains low in comparison with results obtained for Zn and Cu (twenty eight for 1.5mM Zn and forty four for 1.5 mM Cu). The changes in protein expression indicate that the cell adapts to metals by inducing essentially seven defense mechanisms: cell aggregation/biofilm formation (Zn=CutextbackslashtextgreaterCd); modification of envelope properties to increase the extracellular metal biosorption and/or control the uptake of metal (CutextbackslashtextgreaterZn); metal export (Cd=Zn and probably Cu); responses to oxidative stress (CutextbackslashtextgreaterZntextbackslashtextgreaterCd); intracellular metal sequestration (Zn=Cu and probably Cd); hydrolysis of abnormally folded proteins (Cd=Cu), and the over-synthesis of proteins inhibited by metal (CdtextbackslashtextgreaterCutextbackslashtextgreaterZn). To the best of our knowledge, this is the first report showing that a marine P. fluorescens is able to acquire a metal-resistant phenotype, making the strain BA3SM1 a promising agent for bioremediation processes.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{Neuenfeldt2013, title = {Thermodynamic properties distinguish human mitochondrial aspartyl-tRNA synthetase from bacterial homolog with same 3D architecture}, author = {A Neuenfeldt and B Lorber and E Ennifar and A Gaudry and C Sauter and M Sissler and C Florentz }, url = {https://pubmed.ncbi.nlm.nih.gov/23275545/}, doi = { 10.1093/nar/gks1322 }, year = {2013}, date = {2013-02-04}, journal = {Nucleic Acids Research}, volume = {41}, number = {4}, pages = {2698-708}, abstract = {In the mammalian mitochondrial translation apparatus, the proteins and their partner RNAs are coded by two genomes. The proteins are nuclear-encoded and resemble their homologs, whereas the RNAs coming from the rapidly evolving mitochondrial genome have lost critical structural information. This raises the question of molecular adaptation of these proteins to their peculiar partner RNAs. The crystal structure of the homodimeric bacterial-type human mitochondrial aspartyl-tRNA synthetase (DRS) confirmed a 3D architecture close to that of Escherichia coli DRS. However, the mitochondrial enzyme distinguishes by an enlarged catalytic groove, a more electropositive surface potential and an alternate interaction network at the subunits interface. It also presented a thermal stability reduced by as much as 12°C. Isothermal titration calorimetry analyses revealed that the affinity of the mitochondrial enzyme for cognate and non-cognate tRNAs is one order of magnitude higher, but with different enthalpy and entropy contributions. They further indicated that both enzymes bind an adenylate analog by a cooperative allosteric mechanism with different thermodynamic contributions. The larger flexibility of the mitochondrial synthetase with respect to the bacterial enzyme, in combination with a preserved architecture, may represent an evolutionary process, allowing nuclear-encoded proteins to cooperate with degenerated organelle RNAs. }, keywords = {ENNIFAR, FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{bonnay_big_2013, title = {Big bang gene modulates gut immune tolerance in Drosophila}, author = {François Bonnay and Eva Cohen-Berros and Martine Hoffmann and Sabrina Y Kim and Gabrielle L Boulianne and Jules A Hoffmann and Nicolas Matt and Jean-Marc Reichhart}, doi = {10.1073/pnas.1221910110}, issn = {1091-6490}, year = {2013}, date = {2013-02-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {110}, number = {8}, pages = {2957--2962}, abstract = {Chronic inflammation of the intestine is detrimental to mammals. Similarly, constant activation of the immune response in the gut by the endogenous flora is suspected to be harmful to Drosophila. Therefore, the innate immune response in the gut of Drosophila melanogaster is tightly balanced to simultaneously prevent infections by pathogenic microorganisms and tolerate the endogenous flora. Here we describe the role of the big bang (bbg) gene, encoding multiple membrane-associated PDZ (PSD-95, Discs-large, ZO-1) domain-containing protein isoforms, in the modulation of the gut immune response. We show that in the adult Drosophila midgut, BBG is present at the level of the septate junctions, on the apical side of the enterocytes. In the absence of BBG, these junctions become loose, enabling the intestinal flora to trigger a constitutive activation of the anterior midgut immune response. This chronic epithelial inflammation leads to a reduced lifespan of bbg mutant flies. Clearing the commensal flora by antibiotics prevents the abnormal activation of the gut immune response and restores a normal lifespan. We now provide genetic evidence that Drosophila septate junctions are part of the gut immune barrier, a function that is evolutionarily conserved in mammals. Collectively, our data suggest that septate junctions are required to maintain the subtle balance between immune tolerance and immune response in the Drosophila gut, which represents a powerful model to study inflammatory bowel diseases.}, keywords = {Animals, hoffmann, Immune Tolerance, Longevity, M3i, matt, Membrane Proteins, reichhart}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_complementary_2013b, title = {The complementary facets of epithelial host defenses in the genetic model organism Drosophila melanogaster: from resistance to resilience}, author = {Dominique Ferrandon}, doi = {10.1016/j.coi.2012.11.008}, issn = {1879-0372}, year = {2013}, date = {2013-02-01}, journal = {Curr. Opin. Immunol.}, volume = {25}, number = {1}, pages = {59--70}, abstract = {Significant advances have been made in our understanding of the host defense against microbial infections taking place at frontier epithelia of Drosophila flies. Immune deficiency (IMD), the major NF-κB immune response pathway induced in these epithelia, displays remarkable adaptations in its activation and regulation in the respiratory and digestive tract. The host defense against ingested pathogens is not limited to resistance, that is, the immune response. It also involves resilience, the capacity of the host to endure and repair damages inflicted by pathogens or the host's own immune response. For instance, enterocytes damaged by pathogens, the microbiota of aging flies, or host-derived reactive oxygen species (ROS), are replaced under the control of multiple pathways by the compensatory proliferation of intestinal stem cells (ISCs).}, keywords = {Adult Stem Cells, aging, Animal, Animals, Cell Proliferation, Disease Models, Enterocytes, ferrandon, Humans, Immunity, Intestinal Mucosa, M3i, Metagenome, Stem Cell Niche, Wound Healing}, pubstate = {published}, tppubtype = {article} } @article{boscolo_sandwich_2013, title = {Sandwich ELISA Assay for the Quantitation of Palytoxin and Its Analogs in Natural Samples}, author = {S Boscolo and M Pelin and M De Bortoli and G Fontanive and A Barreras and F Berti and S Sosa and O Chaloin and A Bianco and T Yasumoto and M Prato and M Poli and A Tubaro}, url = {https://doi.org/10.1021/es304222t}, doi = {10.1021/es304222t}, issn = {0013-936X}, year = {2013}, date = {2013-02-01}, urldate = {2020-04-01}, journal = {Environmental Science & Technology}, volume = {47}, number = {4}, pages = {2034--2042}, abstract = {Palytoxins are potent marine biotoxins that have recently become endemic to the Mediterranean Sea, and are becoming more frequently associated with seafood. Due to their high toxicity, suitable methods to quantify palytoxins are needed. Thus, we developed an indirect sandwich ELISA for palytoxin and 42-hydroxy-palytoxin. An intralaboratory study demonstrated sensitivity (limit of detection}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{baron_parental_2013, title = {Parental transfer of the antimicrobial protein LBP/BPI protects Biomphalaria glabrata eggs against oomycete infections}, author = {Olga Lucia Baron and Pieter van West and Benoit Industri and Michel Ponchet and Géraldine Dubreuil and Benjamin Gourbal and Jean-Marc Reichhart and Christine Coustau}, doi = {10.1371/journal.ppat.1003792}, issn = {1553-7374}, year = {2013}, date = {2013-01-01}, journal = {PLoS Pathog.}, volume = {9}, number = {12}, pages = {e1003792}, abstract = {Vertebrate females transfer antibodies via the placenta, colostrum and milk or via the egg yolk to protect their immunologically immature offspring against pathogens. This evolutionarily important transfer of immunity is poorly documented in invertebrates and basic questions remain regarding the nature and extent of parental protection of offspring. In this study, we show that a lipopolysaccharide binding protein/bactericidal permeability increasing protein family member from the invertebrate Biomphalaria glabrata (BgLBP/BPI1) is massively loaded into the eggs of this freshwater snail. Native and recombinant proteins displayed conserved LPS-binding, antibacterial and membrane permeabilizing activities. A broad screening of various pathogens revealed a previously unknown biocidal activity of the protein against pathogenic water molds (oomycetes), which is conserved in human BPI. RNAi-dependent silencing of LBP/BPI in the parent snails resulted in a significant reduction of reproductive success and extensive death of eggs through oomycete infections. This work provides the first functional evidence that a LBP/BPI is involved in the parental immune protection of invertebrate offspring and reveals a novel and conserved biocidal activity for LBP/BPI family members.}, keywords = {Acute-Phase Proteins, Animals, Antimicrobial Cationic Peptides, Biomphalaria, Blood Proteins, Carrier Proteins, Cell Membrane, Cell Membrane Permeability, Cloning, Escherichia coli, Female, Immunity, infection, M3i, Maternally-Acquired, Membrane Glycoproteins, Microbial Sensitivity Tests, Molecular, Oomycetes, Recombinant Proteins, reichhart, Zygote}, pubstate = {published}, tppubtype = {article} } @article{ayyaz_negative_2013b, title = {A negative role for MyD88 in the resistance to starvation as revealed in an intestinal infection of Drosophila melanogaster with the Gram-positive bacterium Staphylococcus xylosus}, author = {Arshad Ayyaz and Philippe Giammarinaro and Samuel Liégeois and Matthieu Lestradet and Dominique Ferrandon}, doi = {10.1016/j.imbio.2012.07.027}, issn = {1878-3279}, year = {2013}, date = {2013-01-01}, journal = {Immunobiology}, volume = {218}, number = {4}, pages = {635--644}, abstract = {Drosophila melanogaster is a useful model to investigate mucosal immunity. The immune response to intestinal infections is mediated partly by the Immune deficiency (IMD) pathway, which only gets activated by a type of peptidoglycan lacking in several medically important Gram-positive bacterial species such as Staphylococcus. Thus, the intestinal host defense against such bacterial strains remains poorly known. Here, we have used Staphylococcus xylosus to develop a model of intestinal infections by Gram-positive bacteria. S. xylosus behaves as an opportunistic pathogen in a septic injury model, being able to kill only flies immunodeficient either for the Toll pathway or the cellular response. When ingested, it is controlled by IMD-independent host intestinal defenses, yet flies eventually die. Having excluded an overreaction of the immune response and the action of toxins, we find that flies actually succumb to starvation, likely as a result of a competition for sucrose between the bacteria and the flies. Fat stores of wild-type flies are severely reduced within a day, a period when sucrose is not yet exhausted in the feeding solution. Interestingly, the Toll pathway mutant MyD88 is more resistant to the ingestion of S. xylosus and to starvation than wild-type flies. MyD88 flies do not rapidly deplete their fat stores when starved, in contrast to wild-type flies. Thus, we have uncovered a novel function of MyD88 in the regulation of metabolism that appears to be independent of its known roles in immunity and development.}, keywords = {Adaptor Proteins, Animal, Animals, Antigens, Differentiation, Disease Models, ferrandon, Immunity, Immunologic, Innate, Intestinal Diseases, M3i, Mucosal, Mutation, Receptors, Signal Transducing, Staphylococcal Infections, Staphylococcus, Starvation, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{, title = {Thermodynamics of HIV-1 Reverse Transcriptase in action elucidates the mechanism of action of non-nucleoside inhibitors.}, author = {G Bec and B Meyer and M A Gerard and J Steger and K Fauster and P Wolff and D Burnouf and R Micura and P Dumas and E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23742167}, isbn = {23742167}, year = {2013}, date = {2013-01-01}, journal = {J Am Chem Soc}, volume = {135}, number = {26}, pages = {9743-9752}, abstract = {HIV-1 reverse transcriptase (RT) is a heterodimeric enzyme that converts the genomic viral RNA into proviral DNA. Despite intensive biochemical and structural studies, direct thermodynamic data regarding RT interactions with its substrates are still lacking. Here we addressed the mechanism of action of RT and of non-nucleoside RT inhibitors (NNRTIs) by isothermal titration calorimetry (ITC). Using a new incremental-ITC approach, a step-by-step thermody-namic dissection of the RT polymerization activity showed that most of the driving force for DNA synthesis is provided by initial dNTP binding. Surprisingly, thermodynamic and kinetic data led to a re-interpretation of the mechanism of inhibition of NNRTIs. Binding of NNRTIs to preformed RT/DNA complexes is hindered by a kinetic barrier and NNRTIs mostly interact with free RT. Once formed, RT/NNRTI complexes bind DNA either in a seemingly polymerase-competent orientation, or form high-affinity dead-end complexes, both RT/NNRTI/DNA complexes being unable to bind the incoming nucleotide substrate.}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {X-ray crystallography as a tool for mechanism-of-action studies and drug discovery}, author = {E Ennifar}, url = {http://www.ingentaconnect.com/content/ben/cpb/2013/00000014/00000005/art00007?crawler=true}, isbn = {ISBN/1389-2010 (Print) 1873-4316 (Online)}, year = {2013}, date = {2013-01-01}, journal = {Curr Pharm Biotechnol}, volume = {14}, number = {5}, pages = {537-550}, abstract = {Knowledge of three-dimensional structures of biological macromolecules is essential for a complete understanding of many biological processes. X-ray crystallography is the most widely used technique in structural biology and can provide highly detailed structures of proteins, nucleic acids or macromolecular complexes without any size limit. In the past decade, several recent advances in biological crystallography and automation of data collection and structure solution allowed extraordinary progresses and now more than 58 000 crystal structures have been deposited into the Protein Data Bank. This wealth of structural data significantly helped the elucidation of many biological processes and led to the development of new drugs. In this review we will show how of X-ray crystallography can provide insights into the mechanism of action of biological processes and can contribute to the rationale development of ligands through structure-based drug design.}, keywords = {Drug design Ligand Molecular mechanism Protein RNA Structural biology X-ray crystallography, DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure-guided Discovery of a Novel Aminoglycoside Conjugate Targeting HIV-1 RNA Viral Genome.}, author = {E Ennifar and M W Aslam and P Strasser and G Hoffmann and P Dumas and F L van Delft}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24015986?dopt=Abstract}, doi = {10.1021/cb400498n}, isbn = {24015986}, year = {2013}, date = {2013-01-01}, journal = {ACS Chem Biol}, volume = {8}, number = {11}, pages = {2509-17}, abstract = {The Dimerization Initiation Site (DIS) of the HIV-1 genomic RNA is a conserved stem-loop that promotes viral genome dimerization by forming a loop-loop complex. The DIS constitutes a potentially interesting target since it is crucial for several key steps of the viral replication. In this work we describe the synthesis of a rationally designed aminoglycoside conjugate that binds the HIV-1 DIS viral RNA with high specificity, as shown by an extensive in vitro binding characterization. We propose a three-dimensional model of the drug/RNA interaction that perfectly fits with binding data. Our results show the feasibility of targeting the HIV DIS viral RNA dimer and open the way to the rationale design of a new class of antiviral drugs. In addition, due to similarities between the HIV-1 DIS RNA and the bacterial aminoacyl decoding site (A site) RNA, we show that this novel aminoglycoside conjugate also binds the bacterial A site with a similar affinity than natural aminoglycoside antibiotics.}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Controlling Helix Formation in the γ-Peptide Superfamily: Heterogeneous Foldamers with Urea/Amide and Urea/Carbamate Backbones}, author = {N Pendem and Y R Nelli and C Douat and L Fischer and M Laguerre and E Ennifar and B Kauffmann and G Guichard}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23460200}, doi = {10.1002/anie.201209838}, isbn = {23460200}, year = {2013}, date = {2013-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {52}, number = {15}, pages = {4147-51}, abstract = {One fold to rule them all: New heterogeneous aliphatic backbone foldamers belonging to the γ-peptide superfamily and containing various combinations of urea/amide (U/A) and urea/carbamate (U/C) units are reported. Structural studies at atomic resolution reveal hydrogen-bonded helical structures akin to that formed by cognate Un homooligomers.}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Helix-forming propensity of aliphatic urea oligomers incorporating noncanonical residue substitution patterns}, author = {N Pendem and C Douat and P Claudon and M Laguerre and S Castano and B Desbat and D Cavagnat and E Ennifar and B Kauffmann and G Guichard}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=23445529}, doi = {10.1021/ja401151v}, isbn = {23445529}, year = {2013}, date = {2013-01-01}, journal = {J Am Chem Soc}, volume = {135}, number = {12}, pages = {4884-92}, abstract = {Aliphatic N,N'-linked oligoureas are peptidomimetic foldamers that adopt a well-defined helical secondary structure stabilized by a collection of remote three-center H-bonds closing 12- and 14-membered pseudorings. Delineating the rules that govern helix formation depending on the nature of constituent units is of practical utility if one aims to utilize this helical fold to place side chains in a given arrangement and elaborate functional helices. In this work, we tested whether the helix geometry is compatible with alternative substitution patterns. The central -NH-CH(R)-CH2-NH-CO- residue in a model oligourea pentamer sequence was replaced by guest units bearing various substitution patterns [e.g., -NH-CH2-CH2-NH-CO-, -NH-CH2-CH(R)-NH-CO-, and -NH-CH(R(1))-CH(R(2))-NH-CO-], levels of preorganization (cyclic vs acyclic residues), and stereochemistries, and the helix formation was systematically assessed. The extent of helix perturbation or stabilization was primarily monitored in solution by Fourier transform IR, NMR, and electronic circular dichroism spectroscopies. Our results indicate that although three new substitution patterns were accommodated in the 2.5-helix, the helical urea backbone in short oligomers is particularly sensitive to variations in the residue substitution pattern (position and stereochemistry). For example, the trans-1,2-diaminocyclohexane unit was experimentally found to break the helix nucleation, but the corresponding cis unit did not. Theoretical calculations helped to rationalize these results. The conformational preferences in this series of oligoureas were also studied at high resolution by X-ray structure analyses of a representative set of modified oligomers.}, note = {1520-5126 (Electronic) 0002-7863 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Biomolecular Peptidomimetics/*chemistry Protein Structure, Circular Dichroism Crystallography, ENNIFAR, Fourier Transform Infrared Urea/*analogs & derivatives, Molecular Molecular Conformation Nuclear Magnetic Resonance, Secondary Spectroscopy, Unité ARN, X-Ray Cyclohexylamines/chemistry Models}, pubstate = {published}, tppubtype = {article} } @article{le_coz_circulating_2013, title = {Circulating TFH subset distribution is strongly affected in lupus patients with an active disease}, author = {Carole Le Coz and Aurélie Joublin and Jean-Louis Pasquali and Anne-Sophie Korganow and Hélène Dumortier and Fanny Monneaux}, doi = {10.1371/journal.pone.0075319}, issn = {1932-6203}, year = {2013}, date = {2013-01-01}, journal = {PloS One}, volume = {8}, number = {9}, pages = {e75319}, abstract = {Follicular helper T cells (TFH) represent a distinct subset of CD4(+) T cells specialized in providing help to B lymphocytes, which may play a central role in autoimmune diseases having a major B cell component such as systemic lupus erythematosus. Recently, TFH subsets that share common phenotypic and functional characteristics with TFH cells from germinal centers, have been described in the peripheral blood from healthy individuals. The aim of this study was to analyze the distribution of such populations in lupus patients. Circulating TFH cell subsets were defined by multicolor flow cytometry as TFH17 (CXCR3(-)CCR6(+)), TFH1 (CXCR3 (+) CCR6(-)) or TFH2 (CXCR3(-)CCR6(-)) cells among CXCR5 (+) CD45RA(-)CD4(+) T cells in the peripheral blood of 23 SLE patients and 23 sex and age-matched healthy controls. IL-21 receptor expression by B cells was analyzed by flow cytometry and the serum levels of IL-21 and Igs were determined by ELISA tests. We found that the TFH2 cell subset frequency is strongly and significantly increased in lupus patients with an active disease (SLEDAI scoretextgreater8), while the TFH1 cell subset percentage is greatly decreased. The TFH2 and TFH1 cell subset frequency alteration is associated with the presence of high Ig levels and autoantibodies in patient's sera. Moreover, the TFH2 cell subset enhancement correlates with an increased frequency of double negative memory B cells (CD27(-)IgD(-)CD19(+) cells) expressing the IL-21R. Finally, we found that IgE levels in lupus patients' sera correlate with disease activity and seem to be associated with high TFH2 cell subset frequency. In conclusion, our study describes for the first time the distribution of circulating TFH cell subsets in lupus patients. Interestingly, we found an increased frequency of TFH2 cells, which correlates with disease activity. Our results suggest that this subset might play a key role in lupus pathogenesis.}, keywords = {Adult, Aged, B-Lymphocytes, Case-Control Studies, CD4 Lymphocyte Count, CD5 Antigens, CXCR5, Cytokines, Dumortier, Female, Flow Cytometry, Helper-Inducer, Humans, I2CT, Immunoglobulin E, Immunologic Memory, Immunophenotyping, Interleukin-21, Lupus Erythematosus, Male, Middle Aged, Monneaux, Phenotype, Receptors, Systemic, T-Lymphocytes, Team-Dumortier, Th2 Cells, Young Adult}, pubstate = {published}, tppubtype = {article} } @article{dumortier_when_2013, title = {When carbon nanotubes encounter the immune system: desirable and undesirable effects}, author = {Hélène Dumortier}, doi = {10.1016/j.addr.2013.09.005}, issn = {1872-8294}, year = {2013}, date = {2013-01-01}, journal = {Advanced Drug Delivery Reviews}, volume = {65}, number = {15}, pages = {2120--2126}, abstract = {The role of our immune system is to bring efficient protection against invasion by foreign elements, not only pathogens but also any material it may be in contact with. Nanoparticles may enter the body and encounter the immune system either intentionally (e.g. administration for biomedical application) or not (e.g. respiratory occupational exposure). Therefore, it is of fundamental importance to get a thorough knowledge of the way they interact with immune cells and all related consequences. Among nanomaterials, carbon nanotubes (CNTs) are of special interest because of their tremendous field of applications. Consequently, their increasing production, processing and eventual incorporation into new types of composites and/or into biological systems have raised fundamental issues regarding their potential impact on health. This review aims at giving an overview of the known desirable and undesirable effects of CNTs on the immune system, i.e. beneficial modulation of immune cells by CNTs engineered for biomedical applications versus toxicity, inflammation and unwanted immune reactions triggered by CNTs themselves.}, keywords = {Animals, Biomedical application, carbon, Carbon nanotubes, Dumortier, Environmental Exposure, Functionalization, Humans, I2CT, Immune cell activation, Immune System, inflammation, Inhalation Exposure, Lymphocyte, Macrophage, Nanotubes, Occupational Exposure, Team-Dumortier, Toxicity}, pubstate = {published}, tppubtype = {article} } @article{russier_evidencing_2013, title = {Evidencing the mask effect of graphene oxide: a comparative study on primary human and murine phagocytic cells}, author = {Julie Russier and Emanuele Treossi and Alessia Scarsi and Francesco Perrozzi and Hélène Dumortier and Luca Ottaviano and Moreno Meneghetti and Vincenzo Palermo and Alberto Bianco}, doi = {10.1039/c3nr03543c}, issn = {2040-3372}, year = {2013}, date = {2013-01-01}, journal = {Nanoscale}, volume = {5}, number = {22}, pages = {11234--11247}, abstract = {Graphene oxide (GO) is attracting an ever-growing interest in different fields and applications. Not much is known about the possible impact of GO sheet lateral dimensions on their effects in vitro, especially on human primary cells. In an attempt to address this issue, we present a study to evaluate, how highly soluble 2-dimensional GO constituted of large or small flakes affects human monocyte derived macrophages (hMDM). For this purpose, the lateral size of GO was tuned using sonication and three samples were obtained. The non sonicated one presented large flakes (textasciitilde1.32 μm) while sonication for 2 and 26 hours generated small (textasciitilde0.27 μm) and very small (textasciitilde0.13 μm) sheets of GO, respectively. Cell studies were then conducted to evaluate the cytotoxicity, the oxidative stress induction, the activation potential and the pro-inflammatory effects of these different types of GO at increasing concentrations. In comparison, the same experiments were run on murine intraperitoneal macrophages (mIPM). The interaction between GO and cells was further examined by TEM and Raman spectroscopy. Our data revealed that the GO sheet size had a significant impact on different cellular parameters (i.e. cellular viability, ROS generation, and cellular activation). Indeed, the more the lateral dimensions of GO were reduced, the higher were the cellular internalization and the effects on cellular functionality. Our data also revealed a particular interaction of GO flakes with the cellular membrane. In fact, a GO mask due to the parallel arrangement of the graphene sheets on the cellular surface was observed. Considering the mask effect, we have hypothesized that this particular contact between GO sheets and the cell membrane could either promote their internalization or isolate cells from their environment, thus possibly accounting for the following impact on cellular parameters.}, keywords = {Animals, Cell Survival, Cells, Cultured, Cytokines, Dumortier, Graphite, Humans, I2CT, Macrophages, Mice, Monocytes, Oxidative Stress, Oxides, Reactive Oxygen Species, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{lacotte_early_2013, title = {Early differentiated CD138(high) MHCII+ IgG+ plasma cells express CXCR3 and localize into inflamed kidneys of lupus mice}, author = {Stéphanie Lacotte and Marion Decossas and Carole Le Coz and Susana Brun and Sylviane Muller and Hélène Dumortier}, doi = {10.1371/journal.pone.0058140}, issn = {1932-6203}, year = {2013}, date = {2013-01-01}, journal = {PloS One}, volume = {8}, number = {3}, pages = {e58140}, abstract = {Humoral responses are central to the development of chronic autoimmune diseases such as systemic lupus erythematosus. Indeed, autoantibody deposition is responsible for tissue damage, the kidneys being one of the main target organs. As the source of pathogenic antibodies, plasma cells are therefore critical players in this harmful scenario, both at systemic and local levels. The aim of the present study was to analyze plasma cells in NZB/W lupus mice and to get a better understanding of the mechanisms underlying their involvement in the renal inflammation process. Using various techniques (i.e. flow cytometry, quantitative PCR, ELISpot), we identified and extensively characterized three plasma cell intermediates, according to their B220/CD138/MHCII expression levels. Each of these cell subsets displays specific proliferation and antibody secretion capacities. Moreover, we evidenced that the inflammation-related CXCR3 chemokine receptor is uniquely expressed by CD138(high)MHCII(+) plasma cells, which encompass both short- and long-lived cells and mostly produce IgG (auto)antibodies. Expression of CXCR3 allows efficient chemotactic responsiveness of these cells to cognate chemokines, which production is up-regulated in the kidneys of diseased NZB/W mice. Finally, using fluorescence and electron microscopy, we demonstrated the presence of CD138(+)CXCR3(+)IgG(+) cells in inflammatory areas in the kidneys, where they are very likely involved in the injury process. Thus, early differentiated CD138(high)MHCII(+) rather than terminally differentiated CD138(high)MHCII(low) plasma cells may be involved in the renal inflammatory injury in lupus, due to CXCR3 expression and IgG secretion.}, keywords = {Animals, Autoantibodies, Cell Differentiation, CXCR3, Dumortier, Gene Expression Regulation, Histocompatibility Antigens Class II, I2CT, Immunoglobulin G, Inbred BALB C, Kidney, Leukocyte Common Antigens, Lupus Nephritis, Mice, Plasma Cells, Receptors, Syndecan-1, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{habif_phosphoproteome_2013, title = {Phosphoproteome analyses reveal specific implications of Hcls1, p21-activated kinase 1 and Ezrin in proliferation of a myeloid progenitor cell line downstream of wild-type and ITD mutant Fms-like tyrosine kinase 3 receptors.}, author = {Guillaume Habif and Marie-France Grasset and Sylvie Kieffer-Jaquinod and Lauriane Kuhn and Guy Mouchiroud and Stéphanie Gobert-Gosse}, doi = {10.1016/j.jprot.2012.09.009}, issn = {1876-7737 1874-3919}, year = {2013}, date = {2013-01-01}, journal = {Journal of proteomics}, volume = {78}, pages = {231--244}, abstract = {The tyrosine kinase receptor Flt3 (Fms-like tyrosine kinase 3) is almost always expressed in AML (acute myeloid leukemia) cells, and constitutive activation of Flt3 by ITD (internal tandem duplication) mutations is one of the most common molecular alterations known in AML, especially monocytic AML. Furthermore, Flt3-ligand (FL) was shown as an in vitro growth factor for monocytic precursors, pointing to the important role of Flt3 in the regulation of monocyte/macrophage production. To get a relevant model for studying the molecular mechanisms underlying the physiopathological role of Flt3 on monocytic lineage development, we used the IL-3 dependent murine myeloid progenitors FDC-P1 cell line to generate cells stably co-expressing murine Fms (M-CSF receptor) and human Flt3. Wild type (WT)-Flt3 expressing cells could proliferate in an FL-dependent manner, whereas those expressing Flt3-ITD all survived IL-3 deprivation and showed autonomous proliferation, whereas both types of cells could differentiate to monocytic cells in response to M-CSF. Next, by combining phosphoprotein detection or purification, comparative 2D-PAGE and mass spectrometry sequencing, we sought for downstream mediators of Flt3-WT or Flt3-ITD in FD/Fms cell proliferation. Amongst the differentially expressed and/or phosphorylated proteins, 3 showed a specific implication in FD/Fms cell proliferation: Hcls1 and the Pak1/2 in FL-dependent proliferation of Flt3-WT expressing cells and Ezrin in autonomous proliferation of Flt3-ITD expressing cells.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {CTIP2 is a negative regulator of P-TEFb}, author = {T Cherrier and V Le Douce and S Eilebrecht and R Riclet and C Marban and F Dequiedt and Y Goumon and J C Paillart and M Mericskay and A Parlakian and P Bausero and W Abbas and G Herbein and S K Kurdistani and X Grana and B Van Driessche and C Schwartz and E Candolfi and A G Benecke and C Van Lint and O Rohr}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23852730?dopt=Abstract http://www.pnas.org/content/110/31/12655.full}, isbn = {23852730}, year = {2013}, date = {2013-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {110}, number = {31}, pages = {12655-12660}, abstract = {The positive transcription elongation factor b (P-TEFb) is involved in physiological and pathological events including inflammation, cancer, AIDS, and cardiac hypertrophy. The balance between its active and inactive form is tightly controlled to ensure cellular integrity. We report that the transcriptional repressor CTIP2 is a major modulator of P-TEFb activity. CTIP2 copurifies and interacts with an inactive P-TEFb complex containing the 7SK snRNA and HEXIM1. CTIP2 associates directly with HEXIM1 and, via the loop 2 of the 7SK snRNA, with P-TEFb. In this nucleoprotein complex, CTIP2 significantly represses the Cdk9 kinase activity of P-TEFb. Accordingly, we show that CTIP2 inhibits large sets of P-TEFb- and 7SK snRNA-sensitive genes. In hearts of hypertrophic cardiomyopathic mice, CTIP2 controls P-TEFb-sensitive pathways involved in the establishment of this pathology. Overexpression of the β-myosin heavy chain protein contributes to the pathological cardiac wall thickening. The inactive P-TEFb complex associates with CTIP2 at the MYH7 gene promoter to repress its activity. Taken together, our results strongly suggest that CTIP2 controls P-TEFb function in physiological and pathological conditions.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Escherichia coli Ribosomal Protein S1 Unfolds Structured mRNAs Onto the Ribosome for Active Translation Initiation.}, author = {M Duval and A Korepanov and O Fuchsbauer and P Fechter and A Haller and A Fabbretti and L Choulier and R Micura and B P Klaholz and P Romby and M Springer and S Marzi}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24339747?dopt=Abstract}, doi = {10.1371/journal.pbio.1001731}, isbn = {24339747}, year = {2013}, date = {2013-01-01}, journal = {PLoS Biol}, volume = {11}, number = {12}, pages = {e1001731}, abstract = {Regulation of translation initiation is well appropriate to adapt cell growth in response to stress and environmental changes. Many bacterial mRNAs adopt structures in their 5' untranslated regions that modulate the accessibility of the 30S ribosomal subunit. Structured mRNAs interact with the 30S in a two-step process where the docking of a folded mRNA precedes an accommodation step. Here, we used a combination of experimental approaches in vitro (kinetic of mRNA unfolding and binding experiments to analyze mRNA-protein or mRNA-ribosome complexes, toeprinting assays to follow the formation of ribosomal initiation complexes) and in vivo (genetic) to monitor the action of ribosomal protein S1 on the initiation of structured and regulated mRNAs. We demonstrate that r-protein S1 endows the 30S with an RNA chaperone activity that is essential for the docking and the unfolding of structured mRNAs, and for the correct positioning of the initiation codon inside the decoding channel. The first three OB-fold domains of S1 retain all its activities (mRNA and 30S binding, RNA melting activity) on the 30S subunit. S1 is not required for all mRNAs and acts differently on mRNAs according to the signals present at their 5' ends. This work shows that S1 confers to the ribosome dynamic properties to initiate translation of a large set of mRNAs with diverse structural features.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @incollection{, title = {Footprinting methods for mapping RNA-protein and RNA-RNA interactions.}, author = {M Duval and C Romilly and A C Helfer and O Fuchsbauer and P Romby and S Marzi}, editor = {D Klostermeier and C Hammann}, url = {http://www.degruyter.com/view/books/9783110284959/9783110284959.29/9783110284959.29.xml?format=EBOK}, doi = {10.1515/9783110284959}, isbn = {9783110284959/ISSN}, year = {2013}, date = {2013-01-01}, booktitle = {RNA Structure and Folding}, pages = {29-50}, publisher = {Verlag Walter de Gruyter}, address = {Berlin}, abstract = {Footprinting is currently used to study protein-nucleic acid complexes and more specifically protein-RNA interactions. The method is based on the accessibility of each nucleotide of RNA bound to a ligand against chemical modifications or enzymatic cleavages. It provides information on the regions of interaction on the RNA sequence, and on the RNA conformational changes, which undergo upon ligand binding. Although these approaches are well appropriate to analyze stable RNA-protein or RNA-RNA complexes, it is however difficult to gain knowledge on transient and dynamic complexes. Here, we provide a brief guide on RNA footprinting, discuss the mechanism of action of the most currently used chemical and enzymatic probes, and illustrate the advantages and limitations of the approaches with several examples.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {incollection} } @article{, title = {Involvement of protein IF2 N domain in ribosomal subunit joining revealed from architecture and function of the full-length initiation factor.}, author = {A Simonetti and S Marzi and I M Billas and A Tsai and A Fabbretti and A G Myasnikov and P Roblin and A C Vaiana and I Hazemann and D Eiler and T A Steitz and J D Puglisi and C O Gualerzi and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/pubmed/24029017?dopt=Abstract}, doi = {10.1073/pnas.1309578110}, isbn = {24029017}, year = {2013}, date = {2013-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {110}, number = {39}, pages = {15656-61}, abstract = {Translation initiation factor 2 (IF2) promotes 30S initiation complex (IC) formation and 50S subunit joining, which produces the 70S IC. The architecture of full-length IF2, determined by small angle X-ray diffraction and cryo electron microscopy, reveals a more extended conformation of IF2 in solution and on the ribosome than in the crystal. The N-terminal domain is only partially visible in the 30S IC, but in the 70S IC, it stabilizes interactions between IF2 and the L7/L12 stalk of the 50S, and on its deletion, proper N-formyl-methionyl(fMet)-tRNAfMet positioning and efficient transpeptidation are affected. Accordingly, fast kinetics and single-molecule fluorescence data indicate that the N terminus promotes 70S IC formation by stabilizing the productive sampling of the 50S subunit during 30S IC joining. Together, our data highlight the dynamics of IF2-dependent ribosomal subunit joining and the role played by the N terminus of IF2 in this process.}, keywords = {ERIANI, integrated structural biology protein synthesis, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of the protein core of translation initiation factor 2 in apo, GTP-bound and GDP-bound forms.}, author = {A Simonetti and S Marzi and A Fabbretti and I Hazemann and L Jenner and A Urzhumtsev and C O Gualerzi and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23695237?dopt=Abstract}, doi = {10.1107/S0907444913006422}, isbn = {23695237}, year = {2013}, date = {2013-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {69}, number = {Pt 6}, pages = {925-33}, abstract = {Translation initiation factor 2 (IF2) is involved in the early steps of bacterial protein synthesis. It promotes the stabilization of the initiator tRNA on the 30S initiation complex (IC) and triggers GTP hydrolysis upon ribosomal subunit joining. While the structure of an archaeal homologue (a/eIF5B) is known, there are significant sequence and functional differences in eubacterial IF2, while the trimeric eukaryotic IF2 is completely unrelated. Here, the crystal structure of the apo IF2 protein core from Thermus thermophilus has been determined by MAD phasing and the structures of GTP and GDP complexes were also obtained. The IF2-GTP complex was trapped by soaking with GTP in the cryoprotectant. The structures revealed conformational changes of the protein upon nucleotide binding, in particular in the P-loop region, which extend to the functionally relevant switch II region. The latter carries a catalytically important and conserved histidine residue which is observed in different conformations in the GTP and GDP complexes. Overall, this work provides the first crystal structure of a eubacterial IF2 and suggests that activation of GTP hydrolysis may occur by a conformational repositioning of the histidine residue.}, keywords = {ERIANI, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{kabbage_expression_2012, title = {Expression of the molecular chaperone αB-crystallin in infiltrating ductal breast carcinomas and the significance thereof: an immunohistochemical and proteomics-based strategy.}, author = {Maria Kabbage and Mounir Trimeche and Hela Ben Nasr and Philippe Hammann and Lauriane Kuhn and Bechr Hamrita and Anouar Chaieb and Lotfi Chouchane and Karim Chahed}, doi = {10.1007/s13277-012-0490-4}, issn = {1423-0380 1010-4283}, year = {2012}, date = {2012-12-01}, journal = {Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine}, volume = {33}, number = {6}, pages = {2279--2288}, abstract = {This study aims to evaluate αB-crystallin expression in infiltrating ductal breast carcinomas (IDCAs), as well as, its prognostic significance. Using a two-dimensional electrophoresis matrix-assisted laser desorption/ionisation-time of flight mass spectrometry investigation coupled to an immunohistochemical approach, we have assessed the expression of αB-crystallin in IDCAs, as well as, in other types of breast tumors (invasive lobular carcinomas, medullary carcinomas, and in situ ductal carcinomas). Correlation between αB-crystallin expression and clinicopathological parameters of breast cancer has also been investigated. Proteomic analyses revealed an increased expression of αB-crystallin in IDCA tumors compared to adjacent nontumor tissues. Overexpression of this molecular chaperone was further confirmed in 51 tumor specimens. Statistical analyses revealed, however, no significant correlations between αB-crystallin expression and clinicopathological parameters of the disease (tumor stage, patient age, hormone receptors, SBR grade, and lymph node metastases). This study demonstrates the upregulation of αB-crystallin in IDCA tissues which may highlight its possible involvement in breast cancer development. Our findings do not, however, support the involvement of this molecular chaperone in the progression of this disease.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{anes_assessment_2012, title = {Assessment of the clinical significance of antigenic and functional levels of α1-proteinase inhibitor (α1-Pi) in infiltrating ductal breast carcinomas.}, author = {Amel Ben Anes and Hela Ben Nasr and Philippe Hammann and Lauriane Kuhn and Mounir Trimeche and Bechr Hamrita and Iheb Bougmiza and Anouar Chaieb and Hedi Khairi and Karim Chahed}, doi = {10.1016/j.clinbiochem.2012.07.099}, issn = {1873-2933 0009-9120}, year = {2012}, date = {2012-11-01}, journal = {Clinical biochemistry}, volume = {45}, number = {16-17}, pages = {1421--1431}, abstract = {OBJECTIVES: To determine the clinical significance of α1-proteinase inhibitor (α1-Pi) in infiltrating ductal breast carcinoma patients. DESIGN AND METHODS: Serum levels of α1-Pi, tryptic specific inhibitory capacity and α1-Pi circulating immune complexes were determined using radial immunodiffusion, BAPNA assays and ELISA, respectively. 2-DE-MS and immunohistochemistry were performed to examine α1-Pi protein expression. RESULTS: A decreased serum level of α1-Pi was found among breast cancer patients in comparison to controls. In addition, we found a significantly decreased mean level of α1-Pi in the node metastatic group when compared to node negative patients. However, the functional activity of the inhibitor did not decrease proportionately. Through 2-DE analyses, a differential expression of α1-Pi isoforms according to tumor stage and node metastatic development was found. CONCLUSIONS: Both α1-Pi levels and specific activity could be a source of complementary clinical information and may provide useful information for a better understanding of the mechanisms of metastasis.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{E2012, title = {High-throughput sorting of mosquito larvae for laboratory studies and for future vector control interventions}, author = {Eric Marois and C Scali and Julien Soichot and Christine Kappler and Elena A Levashina and Flaminia Catteruccia}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22929810}, year = {2012}, date = {2012-08-28}, journal = {Malaria J.}, volume = {11}, pages = {302}, abstract = {BACKGROUND: Mosquito transgenesis offers new promises for the genetic control of vector-borne infectious diseases such as malaria and dengue fever. Genetic control strategies require the release of large number of male mosquitoes into field populations, whether they are based on the use of sterile males (sterile insect technique, SIT) or on introducing genetic traits conferring refractoriness to disease transmission (population replacement). However, the current absence of high-throughput techniques for sorting different mosquito populations impairs the application of these control measures. METHODS: A method was developed to generate large mosquito populations of the desired sex and genotype. This method combines flow cytometry and the use of Anopheles gambiae transgenic lines that differentially express fluorescent markers in males and females. RESULTS: Fluorescence-assisted sorting allowed single-step isolation of homozygous transgenic mosquitoes from a mixed population. This method was also used to select wild-type males only with high efficiency and accuracy, a highly desirable tool for genetic control strategies where the release of transgenic individuals may be problematic. Importantly, sorted males showed normal mating ability compared to their unsorted brothers. CONCLUSIONS: The developed method will greatly facilitate both laboratory studies of mosquito vectorial capacity requiring high-throughput approaches and future field interventions in the fight against infectious disease vectors.}, keywords = {COPAS, M3i, marois, Sorting, transgenesis}, pubstate = {published}, tppubtype = {article} } @article{singh_nucleobase-pairing_2012, title = {Nucleobase-pairing triggers the self-assembly of uracil-ferrocene on adenine functionalized multi-walled carbon nanotubes}, author = {Prabhpreet Singh and Cécilia Ménard-Moyon and Jitendra Kumar and Bruno Fabre and Sandeep Verma and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S0008622311008670}, doi = {10.1016/j.carbon.2011.10.037}, issn = {0008-6223}, year = {2012}, date = {2012-08-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {50}, number = {9}, pages = {3170--3177}, series = {Festschrift dedicated to Peter A. Thrower, Editor-in-Chief, 1972 - 2012}, abstract = {Shortened and oxidized multi-walled carbon nanotubes (MWCNTs) were functionalized with adenine using the amidation strategy. The adenine functionalized MWCNTs (Ad-MWCNTs) were complexed with a uracil substituted ferrocene and characterized by transmission electron microscopy (TEM), high resolution TEM (HRTEM), electron diffraction X-ray spectroscopy (EDX), and atomic force microscopy (AFM). The electrochemical properties of these novel nanohybrids were studied by cyclic voltammetry. The favorable supramolecular interaction of the electroactive species with the functionalized nanotubes through the efficient adenine–uracil base-pairing can be exploited for the design of new electronic devices.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pmid22555163, title = {Histone methyltransferase inhibitors induce HIV-1 recovery in resting CD4(+) T cells from HIV-1-infected HAART-treated patients}, author = {Sophie Bouchat and Jean-Stéphane Gatot and Kabamba Kabeya and Christelle Cardona and Laurence Colin and Georges Herbein and Stéphane De Wit and Nathan Clumeck and Olivier Lambotte and Christine Rouzioux and Olivier Rohr and Carine Van Lint}, doi = {10.1097/QAD.0b013e32835535f5}, issn = {1473-5571}, year = {2012}, date = {2012-07-01}, urldate = {2012-07-01}, journal = {AIDS}, volume = {26}, number = {12}, pages = {1473--1482}, abstract = {OBJECTIVE: Reactivation of HIV-1 expression in persistent reservoirs together with an efficient HAART has been proposed as an adjuvant therapy aimed at reaching a functional cure for HIV. Previously, H3K9 methylation was shown to play a major role in chromatin-mediated repression of the HIV-1 promoter. Here, we evaluated the therapeutic potential of histone methyltransferase inhibitors (HMTIs) in reactivating HIV-1 from latency.nnDESIGN: We evaluated the reactivation potential of two specific HMTIs (chaetocin and BIX-01294, two specific inhibitors of Suv39H1 and G9a, respectively) in ex-vivo cultures of resting CD4 T cells isolated from HIV-1-infected HAART-treated individuals.nnMETHODS: We measured HIV-1 recovery in ex-vivo cultures treated with an HMTI alone or in combination with other HIV-1 inducers (in absence of IL-2 and of allogenic stimulation) of CD8-depleted peripheral blood mononuclear cells (PBMCs) or of resting CD4 T cells isolated from 67 HIV-infected, HAART-treated patients with undetectable viral load.nnRESULTS: We demonstrated, for the first time, that chaetocin induced HIV-1 recovery in 50% of CD8-depleted PBMCs cultures and in 86% of resting CD4 T-cell cultures isolated from HIV-1-infected, HAART-treated patients, whereas BIX-01294 reactivated HIV-1 expression in 80% of resting CD4 T-cell cultures isolated from similar patients. Moreover, we showed that combinatory treatments including one HMTI and either the histone deacetylase inhibitor suberoylanilide hydroxamic acid or the non-tumor-promoting NF-κB inducer prostratin had a higher reactivation potential than these compounds alone.nnCONCLUSION: Our results constitute a proof-of-concept for the therapeutic potential of HMTIs in strategies aiming at reducing the pool of latent reservoirs in HIV-infected, HAART-treated patient.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{ezekowitz_lawrences_2012, title = {Lawrence's book review unfair to Hoffmann}, author = {Alan R B Ezekowitz and Jean-Luc Dimarcq and Fotis Kafatos and Elena A Levashina and Dominique Ferrandon and Charles Hetru and Jean-Luc Imler and Jean-Marc Reichhart}, doi = {10.1016/j.cub.2012.05.015}, issn = {1879-0445}, year = {2012}, date = {2012-06-01}, journal = {Curr. Biol.}, volume = {22}, number = {12}, pages = {R482}, keywords = {Book Reviews as Topic, Ethics, ferrandon, imler, M3i, Professional, reichhart}, pubstate = {published}, tppubtype = {article} } @article{lemaitre_pillars_2012, title = {Pillars article: the dorsoventral regulatory gene cassette spätzle/Toll/cactus controls the potent antifungal response in Drosophila adults. Cell. 1996. 86: 973-983}, author = {Bruno Lemaitre and Emmanuelle Nicolas and Lydia Michaut and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {1550-6606}, year = {2012}, date = {2012-06-01}, journal = {J. Immunol.}, volume = {188}, number = {11}, pages = {5210--5220}, abstract = {The cytokine-induced activation cascade of NF-kappaB in mammals and the activation of the morphogen dorsal in Drosophila embryos show striking structural and functional similarities (Toll/IL-1, Cactus/I-kappaB, and dorsal/NF-kappaB). Here we demonstrate that these parallels extend to the immune response of Drosophila. In particular, the intracellular components of the dorsoventral signaling pathway (except for dorsal) and the extracellular Toll ligand, spätzle regulatory gene cassette, control expression of the antifungal peptide gene drosomycin in adults. We also show that mutations in the Toll signaling pathway dramatically reduce survival after fungal infection. Antibacterial genes are induced either by a distinct pathway involving the immune deficiency gene (imd) or by combined activation of both imd and dorsoventral pathways.}, keywords = {Animals, Antifungal Agents, Developmental, DNA-Binding Proteins, Gene Expression Regulation, history, hoffmann, M3i, Multigene Family, Mycoses, Phosphoproteins, reichhart, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @inbook{Imler2012, title = {Nucleic Acid Sensors and Antiviral Immunity}, author = {Jean-Luc Imler and Jules A Hoffmann}, editor = {eds T. Fujita & P. Sambhara}, year = {2012}, date = {2012-06-01}, pages = {1-18}, publisher = {Landes Bioscience}, chapter = {1 : Antiviral responses in invertebrates}, keywords = {antiviral immunity, hoffmann, imler, Invertebrate, M3i}, pubstate = {published}, tppubtype = {inbook} } @article{A2012, title = {Midgut Microbiota of the Malaria Mosquito Vector Anopheles gambiae and Interactions with Plasmodium falciparum Infection}, author = {A Boissière and M T Tchioffo and D Bachar and L Abate and A Marie and Sandrine E Nsango and H R Rhahbazkia and P H Awono-Ambene and Elena A Levashina and R Christen and Isabelle Morlais}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22693451}, year = {2012}, date = {2012-05-31}, journal = {PLoS Pathog.}, volume = {8}, number = {5}, pages = {e1002742}, abstract = {The susceptibility of Anopheles mosquitoes to Plasmodium infections relies on complex interactions between the insect vector and the malaria parasite. A number of studies have shown that the mosquito innate immune responses play an important role in controlling the malaria infection and that the strength of parasite clearance is under genetic control, but little is known about the influence of environmental factors on the transmission success. We present here evidence that the composition of the vector gut microbiota is one of the major components that determine the outcome of mosquito infections. A. gambiae mosquitoes collected in natural breeding sites from Cameroon were experimentally challenged with a wild P. falciparum isolate, and their gut bacterial content was submitted for pyrosequencing analysis. The meta-taxogenomic approach revealed a broader richness of the midgut bacterial flora than previously described. Unexpectedly, the majority of bacterial species were found in only a small proportion of mosquitoes, and only 20 genera were shared by 80% of individuals. We show that observed differences in gut bacterial flora of adult mosquitoes is a result of breeding in distinct sites, suggesting that the native aquatic source where larvae were grown determines the composition of the midgut microbiota. Importantly, the abundance of Enterobacteriaceae in the mosquito midgut correlates significantly with the Plasmodium infection status. This striking relationship highlights the role of natural gut environment in parasite transmission. Deciphering microbe-pathogen interactions offers new perspectives to control disease transmission.}, keywords = {microbiota, Plasmodium falciparum}, pubstate = {published}, tppubtype = {article} } @article{C2012, title = {Salivary gland-specific P. berghei reporter lines enable rapid evaluation of tissue-specific sporozoite loads in mosquitoes}, author = {Chandra Ramakrishnan and Annika Rademacher and Julien Soichot and Giulia Costa and Andy P Waters and Chris J Janse and J Ramesar and Blandine M Franke-Fayard and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22574152}, year = {2012}, date = {2012-05-04}, journal = {PLoS One}, volume = {7}, number = {5}, pages = {e36376}, abstract = {Malaria is a life-threatening human infectious disease transmitted by mosquitoes. Levels of the salivary gland sporozoites (sgs), the only mosquito stage infectious to a mammalian host, represent an important cumulative index of Plasmodium development within a mosquito. However, current techniques of sgs quantification are laborious and imprecise. Here, transgenic P. berghei reporter lines that produce the green fluorescent protein fused to luciferase (GFP-LUC) specifically in sgs were generated, verified and characterised. Fluorescence microscopy confirmed the sgs stage specificity of expression of the reporter gene. The luciferase activity of the reporter lines was then exploited to establish a simple and fast biochemical assay to evaluate sgs loads in whole mosquitoes. Using this assay we successfully identified differences in sgs loads in mosquitoes silenced for genes that display opposing effects on P. berghei ookinete/oocyst development. It offers a new powerful tool to study infectivity of P. berghei to the mosquito, including analysis of vector-parasite interactions and evaluation of transmission-blocking vaccines.}, keywords = {luciferase, M3i, Plasmodium berghei, salivary gland}, pubstate = {published}, tppubtype = {article} } @article{thomann_novel_2012, title = {Novel glycolipid TLR2 ligands of the type Pam2Cys-α-Gal: synthesis and biological properties}, author = {Jean-Sébastien Thomann and Fanny Monneaux and Gaëlle Creusat and Maria Vittoria Spanedda and Béatrice Heurtault and Chloé Habermacher and Francis Schuber and Line Bourel-Bonnet and Benoît Frisch}, doi = {10.1016/j.ejmech.2012.02.039}, issn = {1768-3254}, year = {2012}, date = {2012-05-01}, journal = {European Journal of Medicinal Chemistry}, volume = {51}, pages = {174--183}, abstract = {A more complete understanding of the mechanism of action of TLR agonists has fueled the investigation of new synthetic immunoadjuvants. In this context, we designed and synthesized glycolipids of the type Pam(2)Cys-α-Galactose as novel immunoadjuvants. Their synthesis required modifying a hydrophobic tBoc-[2,3-bispalmitoyloxy-(2R)-propyl]-R-cysteinyl moiety, i.e. the minimal structure required for TLR2 agonist activity, by addition of a hydrophilic head, either an α-Galactosylpyranose or an α-Galactosylfuranose to gain respectively Pam(2)CGalp and Pam(2)CGalf. While preparing a carbohydrate building block, an unexpected stereoselectivity was observed during a halide ion-catalytic process on a protected galactofuranose: the alpha anomer was obtained with surprisingly high selectivity (α/β ratiotextgreater9) and with good isolated yield (51%). The TLR2 binding properties of Pam(2)CGalp and Pam(2)CGalf were then fully evaluated. Their efficiency in triggering the proliferation of BALB/c mouse splenocytes was also compared to that of Pam(2)CAG and Pam(3)CAG, two well-established ligands of TLRs. Moreover, the maturation state of murine dendritic cells previously incubated with either Pam(2)CGalp or Pam(2)CGalf was monitored by flow cytometry and compared to that induced by lipopolysaccharide. Pam(2)CGalp and Pam(2)CGalf were found to be equivalent TLR2 agonists, and induced splenocyte proliferation and DC maturation. With very similar activity, Pam(2)CGalp and Pam(2)CGalf were also 10-fold to 100-fold better than Pam(2)CAG and Pam(3)CAG at inducing B cell proliferation. This represents the first time a glucidic head has been added to the tBoc-[2,3-bispalmitoyloxy-(2R)-propyl]-R-cysteinyl moiety whilst maintaining the immunomodulating activity. This should greatly enrich the data available on Pam(2)C structure/activity relationships.}, keywords = {Adjuvants, Animals, Cell Line, Chemistry Techniques, Female, Galactose, Glycolipids, Humans, I2CT, Immunologic, ligands, Mice, Monneaux, Structure-Activity Relationship, Synthetic, Team-Dumortier, Toll-Like Receptor 2}, pubstate = {published}, tppubtype = {article} } @article{SE2012, title = {Genetic clonality of Plasmodium falciparum affects the outcome of infection in Anopheles gambiae}, author = {Sandrine E Nsango and L Abate and Martine Thoma and Julien Pompon and Malou Fraiture and Annika Rademacher and A Berry and P H Awono-Ambene and Elena A Levashina and Isabelle Morlais}, year = {2012}, date = {2012-04-24}, journal = {Int J Parasitol.}, volume = {42}, number = {6}, pages = {589-95}, abstract = {Mosquito infections with natural isolates of Plasmodium falciparum are notoriously variable and pose a problem for reliable evaluation of efficiency of transmission-blocking agents for malaria control interventions. Here, we show that monoclonal P. falciparum isolates produce higher parasite loads than mixed ones. Induction of the mosquito immune responses by wounding efficiently decreases Plasmodium numbers in monoclonal infections but fails to do so in infections with two or more parasite genotypes. Our results point to the parasites genetic complexity as a potentially crucial component of mosquito-parasite interactions.}, keywords = {monoclonal infection, Plasmodium falciparum}, pubstate = {published}, tppubtype = {article} } @article{goto_toll_2012, title = {Toll signaling in flies and mammals: two sorts of MyD88}, author = {Akira Goto and Jean-Luc Imler}, doi = {10.1016/j.immuni.2012.04.001}, issn = {1097-4180}, year = {2012}, date = {2012-04-01}, journal = {Immunity}, volume = {36}, number = {4}, pages = {555--557}, abstract = {The mammalian MyD88 signaling molecule participates in Toll receptor signaling within the cytoplasm. In this issue of Immunity, Marek and Kagan (2012) report that Drosophila (d)MyD88 acts instead at the plasma membrane to sort the signaling adaptor Tube.}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{pmid22067449, title = {LSD1 cooperates with CTIP2 to promote HIV-1 transcriptional silencing}, author = {Valentin Le Douce and Laurence Colin and Laetitia Redel and Thomas Cherrier and Georges Herbein and Dominique Aunis and Olivier Rohr and Carine Van Lint and Christian Schwartz}, doi = {10.1093/nar/gkr857}, issn = {1362-4962}, year = {2012}, date = {2012-03-01}, urldate = {2012-03-01}, journal = {Nucleic Acids Res}, volume = {40}, number = {5}, pages = {1904--1915}, abstract = {Microglial cells are the main HIV-1 targets in the central nervous system (CNS) and constitute an important reservoir of latently infected cells. Establishment and persistence of these reservoirs rely on the chromatin structure of the integrated proviruses. We have previously demonstrated that the cellular cofactor CTIP2 forces heterochromatin formation and HIV-1 gene silencing by recruiting HDAC and HMT activities at the integrated viral promoter. In the present work, we report that the histone demethylase LSD1 represses HIV-1 transcription and viral expression in a synergistic manner with CTIP2. We show that recruitment of LSD1 at the HIV-1 proximal promoter is associated with both H3K4me3 and H3K9me3 epigenetic marks. Finally, our data suggest that LSD1-induced H3K4 trimethylation is linked to hSET1 recruitment at the integrated provirus.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{flacher_skin_2012, title = {Skin langerin+ dendritic cells transport intradermally injected anti-DEC-205 antibodies but are not essential for subsequent cytotoxic CD8+ Ŧ cell responses}, author = {V Flacher and C H Tripp and B Haid and A Kissenpfennig and B Malissen and P Stoitzner and J Idoyaga and N Romani}, year = {2012}, date = {2012-03-01}, journal = {Journal of Immunology}, volume = {188}, number = {1550-6606 (Electronic)}, pages = {2146--2155}, abstract = {Incorporation of Ags by dendritic cells (DCs) increases when Ags are targeted to endocytic receptors by mAbs. We have previously demonstrated in the mouse that mAbs against C-type lectins administered intradermally are taken up by epidermal Langerhans cells (LCs), dermal Langerin(neg) DCs, and dermal Langerin(+) DCs in situ. However, the relative contribution of these skin DC subsets to the induction of immune responses after Ag targeting has not been addressed in vivo. We show in this study that murine epidermal LCs and dermal DCs transport intradermally injected mAbs against the lectin receptor DEC-205/CD205 in vivo. Skin DCs targeted in situ with mAbs migrated through lymphatic vessels in steady state and inflammation. In the skin-draining lymph nodes, targeting mAbs were found in resident CD8alpha(+) DCs and in migrating skin DCs. More than 70% of targeted DCs expressed Langerin, including dermal Langerin(+) DCs and LCs. Numbers of targeted skin DCs in the nodes increased 2-3-fold when skin was topically inflamed by the TLR7 agonist imiquimod. Complete removal of the site where OVA-coupled anti-DEC-205 had been injected decreased endogenous cytotoxic responses against OVA peptide-loaded target cells by 40-50%. Surprisingly, selective ablation of all Langerin(+) skin DCs in Langerin-DTR knock-in mice did not affect such responses independently of the adjuvant chosen. Thus, in cutaneous immunization strategies where Ag is targeted to DCs, Langerin(+) skin DCs play a major role in transport of anti-DEC-205 mAb, although Langerin(neg) dermal DCs and CD8alpha(+) DCs are sufficient to subsequent CD8(+) T cell responses}, keywords = {administration & dosage, Animals, Antibodies, antibody, Antigen, Antigens, Biosynthesis, C-Type, C-type lectin, CD, Cell Surface, Comparative Study, Cytotoxic, Dendritic Cells, DERMATOLOGY, Gene Knock-In Techniques, Genetics, imiquimod, immune response, IMMUNE-RESPONSES, Immunization, Immunology, in situ, In vivo, Inbred BALB C, Inbred C57BL, INDUCTION, inflammation, Inflammation Mediators, Injections, Intradermal, knock-in, Langerhans Cells, LECTIN, Lectins, LYMPH, LYMPH NODE, Lymph Nodes, LYMPHATIC VESSEL, Lymphatic Vessels, mAb, Mannose-Binding Lectins, MEDIATOR, metabolism, Mice, Minor Histocompatibility Antigens, mouse, murine, Organ Culture Techniques, Ovum, pathology, physiology, Protein, Protein Transport, Rats, Receptor, Receptors, RESPONSES, Skin, SUBSETS, Surface, T-Lymphocytes, target, Team-Mueller, TLR7, transgenic}, pubstate = {published}, tppubtype = {article} } @article{serag_plant_2012, title = {The plant cell uses carbon nanotubes to build tracheary elements}, author = {Maged F Serag and Noritada Kaji and Manabu Tokeshi and Alberto Bianco and Yoshinobu Baba}, doi = {10.1039/c2ib00135g}, issn = {1757-9708}, year = {2012}, date = {2012-02-01}, journal = {Integrative Biology: Quantitative Biosciences from Nano to Macro}, volume = {4}, number = {2}, pages = {127--131}, abstract = {Since their discovery, carbon nanotubes (CNTs) have been eminent members of the nanomaterial family. Because of their unique physical, chemical and mechanical properties, they are regarded as new potential materials to bring enormous benefits in cell biology studies. Undoubtedly, the first step to prove the advantages of CNTs is to understand the basic behavior of CNTs inside the cells. In a number of studies, CNTs have been demonstrated as new carrier systems for the delivery of DNA, proteins and therapeutic molecules into living cells. However, post-uptake behavior of CNTs inside the cells has not received much consideration. Utilizing the plant cell model, we have shown in this study that the plant cells, differentiating into tracheary elements, incorporate cup-stacked carbon nanotubes (CSCNTs) into cell structure via oxidative cross-linking of monolignols to the nanotubes surface during lignin biosynthesis. This finding highlights the fate of CNTs inside plant cells and provides an example on how the plant cell can handle internalized carbon nanomaterials.}, keywords = {Arabidopsis, Atomic Force, carbon, Cell Differentiation, Confocal, Endocytosis, I2CT, Lignin, Microscopy, Nanotubes, Plant Cells, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{delogu_ex_2012, title = {Ex vivo impact of functionalized carbon nanotubes on human immune cells}, author = {Lucia Gemma Delogu and Enrica Venturelli and Roberto Manetti and Gérard Aimé Pinna and Ciriaco Carru and Roberto Madeddu and Luciano Murgia and Francesco Sgarrella and Hélène Dumortier and Alberto Bianco}, doi = {10.2217/nnm.11.101}, issn = {1748-6963}, year = {2012}, date = {2012-02-01}, journal = {Nanomedicine (London, England)}, volume = {7}, number = {2}, pages = {231--243}, abstract = {AIM: Different studies, carried out by us and others, have investigated the impact of carbon nanotubes (CNTs) in vitro and in animal models. To date, only a few studies have been performed on human cells ex vivo. There is also a lack of comparison between CNTs with varied functionalization and structural properties and their impact on different cell types. MATERIALS & METHODS: The present ex vivo human study focuses on the impact of a series of functionalized multiwalled CNTs on human T and B lymphocytes, natural killer (NK) cells and monocytes. RESULTS: Smaller diameter nanotubes are internalized more efficiently. Viability assays displayed the absence of cytotoxicity of all multiwalled CNTs used. Activation assay demonstrated a strong effect on monocytes and NK cells. CONCLUSION: Our results, on human cells ex vivo, confirmed previous studies demonstrating appropriately functionalized CNTs are nontoxic. The effects on cell functionality were significant for the monocytes and NK cells. These findings encourage the possible use of CNTs for biomedical applications either as carriers of therapeutic molecules or as immune modulator systems.}, keywords = {carbon, Cells, Cultured, Cytokines, Dumortier, Humans, I2CT, Immunity, Innate, Materials Testing, Nanotubes, T-Lymphocytes, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{pmid22427948, title = {The level of DING proteins is increased in HIV-infected patients: in vitro and in vivo studies}, author = {Ahmed Djeghader and Gerard Aragonès and Nune Darbinian and Mikael Elias and Daniel Gonzalez and Anabel García-Heredia and Raúl Beltrán-Debón and Rafal Kaminski and Guillaume Gotthard and Julien Hiblot and Anna Rull and Olivier Rohr and Christian Schwartz and Carlos Alonso-Villaverde and Jorge Joven and Jordi Camps and Eric Chabriere}, doi = {10.1371/journal.pone.0033062}, issn = {1932-6203}, year = {2012}, date = {2012-01-01}, urldate = {2012-01-01}, journal = {PLoS One}, volume = {7}, number = {3}, pages = {e33062}, abstract = {DING proteins constitute an interesting family, owing to their intriguing and important activities. However, after a decade of research, little is known about these proteins. In humans, at least five different DING proteins have been identified, which were implicated in important biological processes and diseases, including HIV. Indeed, recent data from different research groups have highlighted the anti-HIV activity of some DING representatives. These proteins share the ability to inhibit the transcriptional step of HIV-1, a key step of the viral cycle that is not yet targeted by the current therapies. Since such proteins have been isolated from humans, we undertook a comprehensive study that focuses on the relationship between these proteins and HIV-infection in an infectious context. Hence, we developed a home-made ELISA for the quantification of the concentration of DING proteins in human serum. Using this method, we were able to determine the concentration of DING proteins in healthy and HIV-infected patients. Interestingly, we observed a significant increase of the concentration of DING proteins in non treated and treated HIV-infected patients compared to controls. In addition, cell cultures infected with HIV also show an increased expression of DING proteins, ruling out the possible role of antiretroviral treatment in the increase of the expression of DING proteins. In conclusion, results from this study show that the organism reacts to HIV-infection by an overexpression of DING proteins.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{schickel_carabin_2012, title = {Carabin deficiency in B cells increases BCR-TLR9 costimulation-induced autoimmunity}, author = {Jean-Nicolas Schickel and Jean-Louis Pasquali and Anne Soley and Anne-Marie Knapp and Marion Decossas and Aurélie Kern and Jean-Daniel Fauny and Luc Marcellin and Anne-Sophie Korganow and Thierry Martin and Pauline Soulas-Sprauel}, doi = {10.1002/emmm.201201595}, issn = {1757-4684}, year = {2012}, date = {2012-01-01}, journal = {EMBO molecular medicine}, volume = {4}, number = {12}, pages = {1261--1275}, abstract = {The mechanisms behind flares of human autoimmune diseases in general, and of systemic lupus in particular, are poorly understood. The present scenario proposes that predisposing gene defects favour clinical flares under the influence of external stimuli. Here, we show that Carabin is low in B cells of (NZB × NZW) F1 mice (murine SLE model) long before the disease onset, and is low in B cells of lupus patients during the inactive phases of the disease. Using knock-out and B-cell-conditional knock-out murine models, we identify Carabin as a new negative regulator of B-cell function, whose deficiency in B cells speeds up early B-cell responses and makes the mice more susceptible to anti-dsDNA production and renal lupus flare after stimulation with a Toll-like Receptor 9 agonist, CpG-DNA. Finally, in vitro analysis of NFκB activation and Erk phosphorylation in TLR9- and B-cell receptor (BCR)-stimulated Carabin-deficient B cells strongly suggests how the internal defect synergizes with the external stimulus and proposes Carabin as a natural inhibitor of the potentially dangerous crosstalk between BCR and TLR9 pathways in self-reactive B cells.}, keywords = {Adaptor Proteins, Animals, Antigen, Autoimmunity, B-Cell, B-Lymphocytes, Carrier Proteins, Cohort Studies, DNA, Humans, I2CT, Imagerie, Inbred NZB, Inbred Strains, Mice, Phosphorylation, Prospective Studies, Receptors, Signal Transducing, Toll-Like Receptor 9, Transfection}, pubstate = {published}, tppubtype = {article} } @article{coste_crystal_2012, title = {Crystal structure of Diedel, a marker of the immune response of Drosophila melanogaster}, author = {Franck Coste and Cordula Kemp and Vanessa Bobezeau and Charles Hetru and Christine Kellenberger and Jean-Luc Imler and Alain Roussel}, doi = {10.1371/journal.pone.0033416}, issn = {1932-6203}, year = {2012}, date = {2012-01-01}, journal = {PloS One}, volume = {7}, number = {3}, pages = {e33416}, abstract = {BACKGROUND: The Drosophila melanogaster gene CG11501 is up regulated after a septic injury and was proposed to act as a negative regulator of the JAK/STAT signaling pathway. Diedel, the CG11501 gene product, is a small protein of 115 residues with 10 cysteines. METHODOLOGY/PRINCIPAL FINDINGS: We have produced Diedel in Drosophila S2 cells as an extra cellular protein thanks to its own signal peptide and solved its crystal structure at 1.15 Å resolution by SIRAS using an iodo derivative. Diedel is composed of two sub domains SD1 and SD2. SD1 is made of an antiparallel β-sheet covered by an α-helix and displays a ferredoxin-like fold. SD2 reveals a new protein fold made of loops connected by four disulfide bridges. Further structural analysis identified conserved hydrophobic residues on the surface of Diedel that may constitute a potential binding site. The existence of two conformations, cis and trans, for the proline 52 may be of interest as prolyl peptidyl isomerisation has been shown to play a role in several physiological mechanisms. The genome of D. melanogaster contains two other genes coding for proteins homologous to Diedel, namely CG43228 and CG34329. Strikingly, apart from Drosophila and the pea aphid Acyrthosiphon pisum, Diedel-related sequences were exclusively identified in a few insect DNA viruses of the Baculoviridae and Ascoviridae families. CONCLUSION/SIGNIFICANCE: Diedel, a marker of the Drosophila antimicrobial/antiviral response, is a member of a small family of proteins present in drosophilids, aphids and DNA viruses infecting lepidopterans. Diedel is an extracellular protein composed of two sub-domains. Two special structural features (hydrophobic surface patch and cis/trans conformation for proline 52) may indicate a putative interaction site, and support an extra cellular signaling function for Diedel, which is in accordance with its proposed role as negative regulator of the JAK/STAT signaling pathway.}, keywords = {Animals, Aphids, Crystallography, imler, Janus Kinases, M3i, Protein Folding, Protein Structure, Signal Transduction, STAT Transcription Factors, Tertiary, Transcription Factors, X-Ray}, pubstate = {published}, tppubtype = {article} } @article{deleury_specific_2012, title = {Specific versus non-specific immune responses in an invertebrate species evidenced by a comparative de novo sequencing study}, author = {Emeline Deleury and Géraldine Dubreuil and Namasivayam Elangovan and Eric Wajnberg and Jean-Marc Reichhart and Benjamin Gourbal and David Duval and Olga Lucia Baron and Jérôme Gouzy and Christine Coustau}, doi = {10.1371/journal.pone.0032512}, issn = {1932-6203}, year = {2012}, date = {2012-01-01}, journal = {PLoS ONE}, volume = {7}, number = {3}, pages = {e32512}, abstract = {Our present understanding of the functioning and evolutionary history of invertebrate innate immunity derives mostly from studies on a few model species belonging to ecdysozoa. In particular, the characterization of signaling pathways dedicated to specific responses towards fungi and Gram-positive or Gram-negative bacteria in Drosophila melanogaster challenged our original view of a non-specific immunity in invertebrates. However, much remains to be elucidated from lophotrochozoan species. To investigate the global specificity of the immune response in the fresh-water snail Biomphalaria glabrata, we used massive Illumina sequencing of 5'-end cDNAs to compare expression profiles after challenge by Gram-positive or Gram-negative bacteria or after a yeast challenge. 5'-end cDNA sequencing of the libraries yielded over 12 millions high quality reads. To link these short reads to expressed genes, we prepared a reference transcriptomic database through automatic assembly and annotation of the 758,510 redundant sequences (ESTs, mRNAs) of B. glabrata available in public databases. Computational analysis of Illumina reads followed by multivariate analyses allowed identification of 1685 candidate transcripts differentially expressed after an immune challenge, with a two fold ratio between transcripts showing a challenge-specific expression versus a lower or non-specific differential expression. Differential expression has been validated using quantitative PCR for a subset of randomly selected candidates. Predicted functions of annotated candidates (approx. 700 unisequences) belonged to a large extend to similar functional categories or protein types. This work significantly expands upon previous gene discovery and expression studies on B. glabrata and suggests that responses to various pathogens may involve similar immune processes or signaling pathways but different genes belonging to multigenic families. These results raise the question of the importance of gene duplication and acquisition of paralog functional diversity in the evolution of specific invertebrate immune responses.}, keywords = {Animals, Biomphalaria, Calmodulin, Cluster Analysis, Complementary, DNA, Expressed Sequence Tags, Ferritins, Gene Expression Profiling, Gene Expression Regulation, High-Throughput Nucleotide Sequencing, Immunity, Innate, M3i, messenger, Pattern Recognition, Phylogeny, Receptors, reichhart, RNA, Signal Transduction, Zinc Fingers}, pubstate = {published}, tppubtype = {article} } @article{liu_drosophila_2012, title = {Drosophila EYA regulates the immune response against DNA through an evolutionarily conserved threonine phosphatase motif}, author = {Xi Liu and Teruyuki Sano and Yongsheng Guan and Shigekazu Nagata and Jules A Hoffmann and Hidehiro Fukuyama}, doi = {10.1371/journal.pone.0042725}, issn = {1932-6203}, year = {2012}, date = {2012-01-01}, journal = {PLoS ONE}, volume = {7}, number = {8}, pages = {e42725}, abstract = {Innate immune responses against DNA are essential to counter both pathogen infections and tissue damages. Mammalian EYAs were recently shown to play a role in regulating the innate immune responses against DNA. Here, we demonstrate that the unique Drosophila eya gene is also involved in the response specific to DNA. Haploinsufficiency of eya in mutants deficient for lysosomal DNase activity (DNaseII) reduces antimicrobial peptide gene expression, a hallmark for immune responses in flies. Like the mammalian orthologues, Drosophila EYA features a N-terminal threonine and C-terminal tyrosine phosphatase domain. Through the generation of a series of mutant EYA fly strains, we show that the threonine phosphatase domain, but not the tyrosine phosphatase domain, is responsible for the innate immune response against DNA. A similar role for the threonine phosphatase domain in mammalian EYA4 had been surmised on the basis of in vitro studies. Furthermore EYA associates with IKKβ and full-length RELISH, and the induction of the IMD pathway-dependent antimicrobial peptide gene is independent of SO. Our data provide the first in vivo demonstration for the immune function of EYA and point to their conserved immune function in response to endogenous DNA, throughout evolution.}, keywords = {Amino Acid, Animals, Blotting, Conserved Sequence, Endodeoxyribonucleases, Eye Proteins, hoffmann, Immunoprecipitation, M3i, Phosphoprotein Phosphatases, Sequence Homology, Transcription Factors, Western}, pubstate = {published}, tppubtype = {article} } @article{niehus_fly_2012b, title = {Fly culture collapse disorder: detection, prophylaxis and eradication of the microsporidian parasite Tubulinosema ratisbonensis infecting Drosophila melanogaster}, author = {Sebastian Niehus and Philippe Giammarinaro and Samuel Liégeois and Jessica Quintin and Dominique Ferrandon}, doi = {10.4161/fly.20896}, issn = {1933-6942}, year = {2012}, date = {2012-01-01}, journal = {Fly (Austin)}, volume = {6}, number = {3}, pages = {193--204}, abstract = {Drosophila melanogaster is a robust model to investigate many biological problems. It is however prone to some infections, which may endanger fly stocks if left unchecked for. One such infection is caused by an obligate fungal intracellular parasite, Tubulinosema ratisbonensis, which can be found in laboratory stocks. Here, we identify and briefly characterize a T. ratisbonensis strain that was infesting our Drosophila cultures and that required intensive measures to contain and eradicate the infection. We describe the phenotypes of infested stocks. We also report PCR-based techniques that allow the detection of infested stocks with a high sensitivity. We have developed a high-throughput qPCR assay that allows the efficient parallel screening of a large number of potentially-infested stocks. We also have investigated several prophylactic measures to prevent the further contamination of stocks, namely UV-exposure, ethanol treatment, bleaching, and desiccation. Bleaching was found to kill all spores. Other treatments were less effective but were found to be sufficient to prevent further contamination of noninfested stocks. Two treatments were efficacious in curing infested stocks (1) bleaching of eggs and subsequent raising of the larvae in clean vials; (2) fumagillin treatment. These cures only work on stocks that have not become too weak to withstand the procedures.}, keywords = {Animals, Apansporoblastina, Apansporoblastina/*genetics/physiology, Base Sequence, cure, Disinfection, Disinfection/methods, DNA, DNA Primers, Drosophila melanogaster/*microbiology, ferrandon, fumagillin, Fungal, Fungal/chemistry, M3i, microsporidia, obligate intracellular parasitism, PCR detection, Phylogeny, Polymerase Chain Reaction, Polymerase Chain Reaction/methods, prophylaxis, Ribosomal, Ribosomal/chemistry, Sequence Alignment, Tubulinosema ratisbonensis}, pubstate = {published}, tppubtype = {article} } @article{meister_immune_2012, title = {Immune cell transdifferentiation: a complex crosstalk between circulating immune cells and the haematopoietic niche}, author = {Marie Meister and Dominique Ferrandon}, doi = {10.1038/embor.2011.238}, issn = {1469-3178}, year = {2012}, date = {2012-01-01}, journal = {EMBO Rep.}, volume = {13}, number = {1}, pages = {3--4}, keywords = {Animals, Cell Communication, Cell Transdifferentiation, ferrandon, Hematopoietic Stem Cells, Humans, Immune System, M3i, Signal Transduction, Stem Cell Niche}, pubstate = {published}, tppubtype = {article} } @article{zhang_uap56_2012, title = {UAP56 Couples piRNA Clusters to the Perinuclear Transposon Silencing Machinery.}, author = {Fan Zhang and Jie Wang and Jia Xu and Zhao Zhang and Birgit S Koppetsch and Nadine Schultz and Thom Vreven and Carine Meignin and Ilan Davis and Phillip D Zamore and Zhiping Weng and William E Theurkauf}, year = {2012}, date = {2012-01-01}, journal = {Cell}, volume = {151}, number = {4}, pages = {871--884}, keywords = {meignin}, pubstate = {published}, tppubtype = {article} } @article{keravis_disease_2012, title = {Disease progression in MRL/lpr lupus-prone mice is reduced by NCS 613, a specific cyclic nucleotide phosphodiesterase type 4 (PDE4) inhibitor}, author = {Thérèse Keravis and Fanny Monneaux and Issaka Yougbaré and Lucien Gazi and Jean-Jacques Bourguignon and Sylviane Muller and Claire Lugnier}, doi = {10.1371/journal.pone.0028899}, issn = {1932-6203}, year = {2012}, date = {2012-01-01}, journal = {PloS One}, volume = {7}, number = {1}, pages = {e28899}, abstract = {Systemic lupus erythematosus is a polymorphic and multigenic inflammatory autoimmune disease. Cyclic AMP (cAMP) modulates inflammation and the inhibition of cyclic nucleotide phosphodiesterase type 4 (PDE4), which specifically hydrolyzes cAMP, inhibits TNFα secretion. This study was aimed at investigating the evolution of PDE activity and expression levels during the course of the disease in MRL/lpr lupus-prone mice, and to evaluate in these mice the biological and clinical effects of treatments with pentoxifylline, denbufylline and NCS 613 PDE inhibitors. This study reveals that compared to CBA/J control mice, kidney PDE4 activity of MRL/lpr mice increases with the disease progression. Furthermore, it showed that the most potent and selective PDE4 inhibitor NCS 613 is also the most effective molecule in decreasing proteinuria and increasing survival rate of MRL/lpr mice. NCS 613 is a potent inhibitor, which is more selective for the PDE4C subtype (IC₅₀= 1.4 nM) than the other subtypes (PDE4A, IC₅₀= 44 nM; PDE4B, IC₅₀= 48 nM; and PDE4D, IC₅₀= 14 nM). Interestingly, its affinity for the High Affinity Rolipram Binding Site is relatively low (K(i) = 148 nM) in comparison to rolipram (K(i) = 3 nM). Finally, as also observed using MRL/lpr peripheral blood lymphocytes (PBLs), NCS 613 inhibits basal and LPS-induced TNFα secretion from PBLs of lupus patients, suggesting a therapeutic potential of NCS 613 in systemic lupus. This study reveals that PDE4 represent a potential therapeutic target in lupus disease.}, keywords = {Adenine, Animals, Cyclic AMP, Cyclic Nucleotide Phosphodiesterases, Disease Progression, Female, Humans, I2CT, Inbred CBA, Inbred MRL lpr, Isoenzymes, Kidney, Lipopolysaccharides, Lupus Erythematosus, Mice, Monneaux, Pentoxifylline, Phosphodiesterase 4 Inhibitors, Proteinuria, Survival Rate, Systemic, Team-Dumortier, Tumor Necrosis Factor-alpha, Type 4, Xanthines}, pubstate = {published}, tppubtype = {article} } @article{lamanna_enhancement_2012, title = {Enhancement of anti-inflammatory drug activity by multivalent adamantane-based dendrons}, author = {Giuseppe Lamanna and Julie Russier and Hélène Dumortier and Alberto Bianco}, doi = {10.1016/j.biomaterials.2012.03.072}, issn = {1878-5905}, year = {2012}, date = {2012-01-01}, journal = {Biomaterials}, volume = {33}, number = {22}, pages = {5610--5617}, abstract = {We have developed a straightforward method to prepare 1(st) and 2(nd) generation adamantane-based dendrons, previously called HYDRAmers, bearing at the periphery the anti-inflammatory drug, ibuprofen. The multivalency effect on the drug activity was studied, demonstrating that our multivalent ibuprofen-dendron conjugates exert an enhanced anti-inflammatory activity compared to free ibuprofen, in vitro. These results provide insights into the effect of HYDRAmer multivalency on biological interactions for therapeutic applications.}, keywords = {Animals, Anti-Inflammatory Agents, Cell Line, Cell Survival, Dendrimers, Drug Synergism, Dumortier, I2CT, Ibuprofen, Macrophages, Mice, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Transfer RNA structure}, author = {E Westhof and P Auffinger}, url = {http://www.els.net/WileyCDA/ElsArticle/refId-a0000527.html}, doi = {10.1002/9780470015902.a0000527.pub2}, year = {2012}, date = {2012-01-01}, booktitle = {Encyclopedia of Life Sciences}, publisher = {John Wiley & Sons}, abstract = {Transfer ribonucleic acid (tRNA) molecules that participate in the elongation step of protein synthesis on the ribosome have a conserved secondary structure, known as the cloverleaf, and fold into a common three-dimensional architecture. The conservation of the global L-shaped 3D fold is assessed by the more than 100 available crystal structures showing tRNAs in native states or in complexes where tRNAs are bound to various interacting systems such as cognate synthetases, editing, modification and processing enzymes or full ribosomes. These tRNA crystal structures display awhole range of structural adaptability features encoded in their sequence and underlying their various functions. Thus, as the number of available structural data expands, the concept of a unique tRNA structure fades out for that of an ensemble of interconnected and environmentally dependant tRNA structures.}, keywords = {transfer ribonucleic acid tRNA cloverleaf structure wobble hypothesis anticodon Watson–Crick pairs non-Watson–Crick pairs hydrogen bond Hoogsteen pairs dynamics solvation magnesium, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {RNA modeling, naturally.}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22308479}, doi = {10.1073/pnas.1121363109}, isbn = {22308479}, year = {2012}, date = {2012-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {109}, number = {8}, pages = {2691-2692}, note = {Published online: February 3, 2012}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Ribozymes, catalytically active RNA molecules. Introduction.}, author = {E Westhof}, editor = {J S Hartig}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22315059}, doi = {10.1007/978-1-61779-545-9_1}, isbn = {22315059}, year = {2012}, date = {2012-01-01}, booktitle = {Ribozymes: Methods and Protocols}, volume = {848}, pages = {1-4}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {It is now about 30 years since ribozymes, catalytically active RNA molecules, were discovered. Although the chemical versatility of RNA does not come close to that of proteins, the chemical properties of nucleic acid systems are nevertheless thoroughly exploited in biological systems, leading to diverse ways of accelerating chemical reactions. Ribozymes are truly fascinating biological molecules. After all, is catalytic RNA an accident of life or, instead, is life an accident of catalytic RNA?}, note = {Methods in molecular biology (Clifton, N.J.)}, keywords = {RNA catalysis Ribozyme Hammerhead, Unité ARN, WESTHOF, WESTHOF RNA catalysis Ribozyme Hammerhead Overview}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {HIV-1 polymerase inhibition by nucleoside analogs: cellular- and kinetic parameters of efficacy, susceptibility and resistance selection.}, author = {M von Kleist and P Metzner and R Marquet and C Schütte}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22275860}, doi = {10.1371/journal.pcbi.1002359}, isbn = {22275860}, year = {2012}, date = {2012-01-01}, journal = {PLoS Comput Biol}, volume = {8}, number = {1}, pages = {e1002359}, abstract = {Nucleoside analogs (NAs) are used to treat numerous viral infections and cancer. They compete with endogenous nucleotides (dNTP/NTP) for incorporation into nascent DNA/RNA and inhibit replication by preventing subsequent primer extension. To date, an integrated mathematical model that could allow the analysis of their mechanism of action, of the various resistance mechanisms, and their effect on viral fitness is still lacking. We present the first mechanistic mathematical model of polymerase inhibition by NAs that takes into account the reversibility of polymerase inhibition. Analytical solutions for the model point out the cellular- and kinetic aspects of inhibition. Our model correctly predicts for HIV-1 that resistance against nucleoside analog reverse transcriptase inhibitors (NRTIs) can be conferred by decreasing their incorporation rate, increasing their excision rate, or decreasing their affinity for the polymerase enzyme. For all analyzed NRTIs and their combinations, model-predicted macroscopic parameters (efficacy, fitness and toxicity) were consistent with observations. NRTI efficacy was found to greatly vary between distinct target cells. Surprisingly, target cells with low dNTP/NTP levels may not confer hyper-susceptibility to inhibition, whereas cells with high dNTP/NTP contents are likely to confer natural resistance. Our model also allows quantification of the selective advantage of mutations by integrating their effects on viral fitness and drug susceptibility. For zidovudine triphosphate (AZT-TP), we predict that this selective advantage, as well as the minimal concentration required to select thymidine-associated mutations (TAMs) are highly cell-dependent. The developed model allows studying various resistance mechanisms, inherent fitness effects, selection forces and epistasis based on microscopic kinetic data. It can readily be embedded in extended models of the complete HIV-1 reverse transcription process, or analogous processes in other viruses and help to guide drug development and improve our understanding of the mechanisms of resistance development during treatment.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Small RNA deep-sequencing identifies microRNAs and other small non-coding RNAs from human herpesvirus 6 (HHV-6B).}, author = {L Tuddenham and J S Jung and B Chane-Woon-Ming and L Dölken and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22114334}, doi = {10.1128/​ JVI.05911-11}, isbn = {22114334}, year = {2012}, date = {2012-01-01}, journal = {J Virol}, volume = {86}, pages = {1638-1649}, abstract = {Roseolovirus, or human herpesvirus 6 (HHV-6) is a ubiquitous human pathogen infecting over 95% of the population by the age of two years. As with other herpesviruses, reactivation of HHV-6 can present with severe complications in immunocompromised individuals. Recent studies have highlighted the importance of herpesvirus-derived micro (mi)RNAs in modulating both cellular and viral gene expression. An initial report, which computed the likelihood of various viruses to encode for miRNAs, did not predict HHV-6 miRNAs. To experimentally screen for small HHV-6 encoded RNAs, we conducted large-scale sequencing of Sup-T-1 cells lytically infected with a laboratory strain of HHV-6B. This revealed an abundant 60-65 nucleotide RNA of unknown function derived from the lytic origin of replication (OriLyt) that gave rise to smaller RNA species of 18-19 nucleotides in length. In addition, we identified four pre-miRNAs, whose mature forms accumulated in Argonaute 2. In contrast to other beta-herpesviruses, HHV-6B miRNAs are expressed from direct repeat regions (DR(L) and DR(R)) located at either side of the genome. All miRNAs are conserved in the closely related HHV-6A variant, and one of them is a seed ortholog of the human miR-582-5p. Similar to alpha-herpesvirus miRNAs, they are expressed antisense to immediate early ORFs and thus have the potential to regulate key viral regulators.}, note = {Published ahead of print 23 November 2011}, keywords = {MicroRNA Roseolovirus HHV-6 Sup-T-1 OriLyt, PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Expression of Small Regulatory RNAs in Clinical Samples Reflects the Different Life Styles of Staphylococcus aureus in Colonization vs. Infection.}, author = {J Song and C Lays and F Vandenesch and Y Benito and M Bes and Y Chu and G Lina and P Romby and T Geissmann and S Boisset}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22629378?dopt=Abstract http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0037294}, isbn = {22629378}, year = {2012}, date = {2012-01-01}, journal = {PLoS One}, volume = {7}, number = {5}, pages = {e37294}, abstract = {Small RNAs (sRNAs) are involved in the post-transcriptional regulation of metabolic pathways and in responses to stress and virulence. We analyzed the expression levels of five sRNAs of Staphylococcus aureus during human colonization or infection. Total RNA was isolated from nasal carriers, abscesses and cystic fibrosis patients (20 subjects per condition). The expression levels of the sRNAs were measured in the clinical samples and compared with those of the corresponding strains grown in vitro. Five sRNAs were encoded and expressed in all clinical strains in vitro. In vivo, the global expression of the five sRNAs was extremely variable in the abscessed patients, more homogeneous in the cystic fibrosis patients, and highly uniform in the nasal carrier samples. The expression levels of the sRNAs in vivo resembled those obtained at exponential phase or late exponential phase of growth in vitro, for three and one sRNA respectively; while for one sRNA, the expression was always higher in vivo as compared to in vitro growth. The in vitro conditions do not uniformly mimic the in vivo conditions for sRNA expression. Nasal colonization is associated with a unique expression pattern of sRNA that might reflect the commensalism of S. aureus in this niche.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The origin of genetic diversity in HIV-1.}, author = {R P Smyth and M P Davenport and J Mak}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22728444?dopt=Abstract}, doi = {22728444}, year = {2012}, date = {2012-01-01}, journal = {Virus Res}, volume = {169}, number = {2}, pages = {415-429}, abstract = {One of the hallmarks of HIV infection is the rapid development of a genetically complex population (quasispecies) from an initially limited number of infectious particles. Genetic diversity remains one of the major obstacles to eradication of HIV. The viral quasispecies can respond rapidly to selective pressures, such as that imposed by the immune system and antiretroviral therapy, and frustrates vaccine design efforts. Two unique features of retroviral replication are responsible for the unprecedented variation generated during infection. First, mutations are frequently introduced into the viral genome by the error prone viral reverse transcriptase and through the actions of host cellular factors, such as the APOBEC family of nucleic acid editing enzymes. Second, the HIV reverse transcriptase can utilize both copies of the co-packaged viral genome in a process termed retroviral recombination. When the co-packaged viral genomes are genetically different, retroviral recombination can lead to the shuffling of mutations between viral genomes in the quasispecies. This review outlines the stages of the retroviral life cycle where genetic variation is introduced, focusing on the principal mechanisms of mutation and recombination. Understanding the mechanistic origin of genetic diversity is essential to combating HIV.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Initiation of HIV-1 reverse transcription and functional role of nucleocapsid-mediated tRNA/viral genome interactions.}, author = {D Sleiman and V Goldschmidt and P Barraud and R Marquet and J C Paillart and C Tisné}, url = {http://www.sciencedirect.com/science/article/pii/S0168170212002031?v=s5}, doi = {10.1016/j.virusres.2012.06.006}, isbn = {22721779}, year = {2012}, date = {2012-01-01}, journal = {Virus Res}, volume = {169}, number = {2}, pages = {324-339}, abstract = {HIV-1 reverse transcription is initiated from a tRNALys3 molecule annealed to the viral RNA at the primer binding site (PBS). The annealing of tRNALys3 requires the opening of its three-dimensional structure and RNA rearrangements to form an efficient initiation complex recognized by the reverse transcriptase. This annealing is mediated by the nucleocapsid protein (NC). In this paper, we first review the actual knowledge about HIV-1 viral RNA and tRNALys3 structures. Then, we summarize the studies explaining how NC chaperones the formation of the tRNALys3/PBS binary complex. Additional NMR data that investigated the NC interaction with tRNALys3 D-loop are presented. Lastly, we focused on the additional interactions occurring between tRNALys3 and the viral RNA and showed that they are dependent on HIV-1 isolates, ie the sequence and the structure of the viral RNA.}, keywords = {MARQUET, PAILLART, Reverse transcription HIV tRNALys3 Nucleocapsid, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{lacerda_translocation_2012, title = {Translocation mechanisms of chemically functionalised carbon nanotubes across plasma membranes}, author = {Lara Lacerda and Julie Russier and Giorgia Pastorin and Antonia M Herrero and Enrica Venturelli and Hélène Dumortier and Khuloud T Al-Jamal and Maurizio Prato and Kostas Kostarelos and Alberto Bianco}, doi = {10.1016/j.biomaterials.2012.01.024}, issn = {1878-5905}, year = {2012}, date = {2012-01-01}, journal = {Biomaterials}, volume = {33}, number = {11}, pages = {3334--3343}, abstract = {Understanding the mechanisms responsible for carbon nanotube (CNT) internalisation into live cells is considered critical both from a fundamental point of view and for further engineering of CNT-based delivery systems to intracellular targets. While several studies are focused on the development of such CNT-based delivery systems, attempts to systematically elucidate the cellular uptake mechanisms of CNTs are still rather limited. The aim of the present study is to evaluate the cellular internalisation of chemically functionalised multi-walled carbon nanotubes (f-MWCNTs) in the presence of different well-known cellular uptake inhibitors. Our data reveal how f-MWCNTs are able to translocate across cell membranes of both phagocytic and non-phagocytic cell lines. We have evidenced that at least 30-50% of f-MWCNTs are taken up by cells through an energy-independent mechanism. This characteristic makes nanotubes loaded with therapeutic or diagnostic cargos extremely interesting as the release of active molecules directly into the cytoplasm increase their biological activity and therapeutic efficacy.}, keywords = {Animals, carbon, Cell Line, Cell Membrane, Dumortier, I2CT, Macrophages, Mice, Nanotubes, Phagocytosis, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure-activity relationships among the kanamycin aminoglycosides: role of ring I hydroxy and amino groups.}, author = {S Salian and T Matt and R Akbergenov and S Harish and M Meyer and S Duscha and D Shcherbakov and B B Bernet and A Vasella and E Westhof and E C Bottger}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22948879?dopt=Abstract}, doi = {10.1128/AAC.01326-12}, isbn = {22948879}, year = {2012}, date = {2012-01-01}, journal = {Antimicrob Agents Chemother}, volume = {56}, number = {12}, pages = {6104-6108}, abstract = {The kanamycins form an important subgroup of the 4,6-disubstituted 2-deoxystreptamine aminoglycoside antibiotics, comprising kanamycin A, kanamycin B, tobramycin and dibekacin. These compounds interfere with protein synthesis by targeting the ribosomal decoding A site and they differ in the number and location of amino and hydroxy groups of the glucopyranosyl moiety (ring I). We have synthesized kanamycin analogues characterized by subtle variations of the 2' and 6' substituents of ring I. The functional activities of the kanamycins and the synthesized analogues were investigated i) in cell-free translation assays on wild-type and mutant bacterial ribosomes to study drug-target interaction, ii) in MIC assays to assess antibacterial activity, and iii) in rabbit reticulocyte translation assays to determine activity on eukaryotic ribosomes. Position 2' forms an intramolecular H-bond with O5 of ring II, helping the relative orientations of the two rings with respect to each other. This bond becomes critical for drug activity when a 6' OH substituent is present. Our results point to complex synergistic interactions crucial for aminoglycoside binding and activity.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Riboswitch (T-Box)-mediated control of tRNA-dependent amidation in clostridium acetobutylicum rationalizes gene and pathway redundancy for asparagine and asparaginyl-tRNAAsn synthesis.}, author = {N Y Saad and B Schiel and M Braye and J T Heap and N P Minton and P Duerre and H D Becker}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22505715?dopt=Abstract}, doi = {10.1074/jbc.M111.332304}, isbn = {22505715}, year = {2012}, date = {2012-01-01}, journal = {J Biol Chem}, volume = {287}, number = {24}, pages = {20382-20394}, abstract = {Analysis of the Gram-positive Clostridium acetobutylicum genome reveals an inexplicable level of redundancy for the genes putatively involved in asparagine (Asn) and Asn-tRNAAsn synthesis. Besides a duplicated set of gatCAB tRNA-dependent amidotransferase genes, there is a triplication of aspartyl-tRNA synthetase (AspRS) genes and a duplication of asparagine synthetase B (AsnB) genes. This genomic landscape leads to the suspicion of the incoherent simultaneous use of the direct and indirect pathways of Asn and Asn-tRNAAsn formation. Through a combination of biochemical and genetic approaches, we show that C. acetobutylicum forms Asn and Asn-tRNAAsn by tRNA-dependent amidation. We demonstrate that an entire transamidation pathway composed of AspRS and one set of GatCAB genes is organized as an operon under the control of a tRNAAsn-dependent T-Box riboswitch. Finally, our results suggest that this exceptional gene redundancy might be interconnected to control tRNA-dependent Asn synthesis, which in turn might be involved in controlling the metabolic switch from acidogenesis to solventogenesis in C. acetobutylicum.}, note = {First Published on April 13, 2012}, keywords = {BECKER KERN Aminoacyl tRNA synthesis Bacterial metabolism Riboswitch Transcription regulation Transfer RNA (tRNA) T-Box Antitermination tRNA-dependent synthesis of Asn, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Retrovolution: HIV-Driven Evolution of Cellular Genes and Improvement of Anticancer Drug Activation}, author = {P Rossolillo and F Winter and E Simon-Loriere and S Gallois-Montbrun and M Negroni}, url = {http://www.plosgenetics.org/article/info%3Adoi%2F10.1371%2Fjournal.pgen.1002904}, doi = {10.1371/journal.pgen.1002904}, isbn = {22927829}, year = {2012}, date = {2012-01-01}, journal = {PLoS Genet}, volume = {8}, number = {8}, pages = {e1002904}, abstract = {In evolution strategies aimed at isolating molecules with new functions, screening for the desired phenotype is generally performed in vitro or in bacteria. When the final goal of the strategy is the modification of the human cell, the mutants selected with these preliminary screenings may fail to confer the desired phenotype, due to the complex networks that regulate gene expression in higher eukaryotes. We developed a system where, by mimicking successive infection cycles with HIV-1 derived vectors containing the gene target of the evolution in their genome, libraries of gene mutants are generated in the human cell, where they can be directly screened. As a proof of concept we created a library of mutants of the human deoxycytidine kinase (dCK) gene, involved in the activation of nucleoside analogues used in cancer treatment, with the aim of isolating a variant sensitizing cancer cells to the chemotherapy compound Gemcitabine, to be used in gene therapy for anti-cancer approaches or as a poorly immunogenic negative selection marker for cell transplantation approaches. We describe the isolation of a dCK mutant, G12, inducing a 300-fold sensitization to Gemcitabine in cells originally resistant to the prodrug (Messa 10K), an effect 60 times stronger than the one induced by the wt enzyme. The phenotype is observed in different tumour cell lines irrespective of the insertion site of the transgene and is due to a change in specificity of the mutated kinase in favour of the nucleoside analogue. The mutations characterizing G12 are distant from the active site of the enzyme and are unpredictable on a rational basis, fully validating the pragmatic approach followed. Besides the potential interest of the G12 dCK variant for therapeutic purposes, the methodology developed is of interest for a large panel of applications in biotechnology and basic research.}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{lamanna_multifunctionalized_2012, title = {Multifunctionalized carbon nanotubes as advanced multimodal nanomaterials for biomedical applications}, author = {Giuseppe Lamanna and Alessia Battigelli and Cécilia Ménard-Moyon and Alberto Bianco}, url = {https://www.degruyter.com/view/journals/ntrev/1/1/article-p17.xml}, doi = {10.1515/ntrev-2011-0002}, issn = {2191-9089, 2191-9097}, year = {2012}, date = {2012-01-01}, urldate = {2020-04-01}, journal = {Nanotechnology Reviews}, volume = {1}, number = {1}, pages = {17--29}, abstract = {textbackslashtextlesssection class="abstract"textbackslashtextgreatertextbackslashtextlessh2 class="abstractTitle text-title my-1" id="d739e2"textbackslashtextgreaterAbstracttextbackslashtextless/h2textbackslashtextgreatertextbackslashtextlessptextbackslashtextgreaterThe increasing importance of nanotechnology in the field of biomedical applications has encouraged the development of new nanomaterials endowed with multiple functions. Novel nanoscale drug delivery systems with diagnostic, imaging and therapeutic properties hold many promises for the treatment of different types of diseases, including cancer, infection and neurodegenerative syndromes. Functionalized carbon nanotubes (CNTs) are one of the most recent type of nanomaterial developed in biomedicine as they can be designed and imparted with multimodal capabilities. Indeed, the possibility of inserting different functionalities on CNTs is opening the possibility to exploit them on new strategies that combine diagnosis with improved therapeutic efficacies. In this review, we describe the different approaches that have been recently developed to generate multifunctionalized CNTs for biomedical applications. In particular, covalent and non-covalent double and triple functionalization methods are discussed, putting in evidence their use textbackslashtextlessemtextbackslashtextgreaterin vitrotextbackslashtextless/emtextbackslashtextgreater and textbackslashtextlessemtextbackslashtextgreaterin vivotextbackslashtextless/emtextbackslashtextgreater and highlighting the advantages and the drawbacks of these new systems. Preclinical studies have demonstrated that multifunctional CNTs are highly promising when combining diagnostic, imaging and therapeutic modalities.textbackslashtextless/ptextbackslashtextgreatertextbackslashtextless/sectiontextbackslashtextgreater}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Loop-loop interactions involved in antisense regulation are processed by the endoribonuclease III in Staphylococcus aureus.}, author = {C Romilly and C Chevalier and S Marzi and B Masquida and T Geissmann and F Vandenesch and E Westhof and P Romby}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23134978?dopt=Abstract}, doi = {10.4161/rna.22710}, isbn = {23134978}, year = {2012}, date = {2012-01-01}, journal = {RNA Biol}, volume = {9}, number = {12}, pages = {1461-1472}, abstract = {The endoribonuclease III (RNase III) belongs to the enzyme family known to process double-stranded RNAs. Staphylococcus aureus RNase III was shown to regulate, in concert with the quorum sensing induced RNAIII, the degradation of several mRNAs encoding virulence factors and the transcriptional repressor of toxins Rot. Two of the mRNA-RNAIII complexes involve fully base paired loop-loop interactions with similar sequences that are cleaved by RNase III at a unique position. We show here that the sequence of the base pairs within the loop-loop interaction was not critical for RNase III cleavage, but that the co-axial stacking of three consecutive helices provides an ideal topology for RNase III recognition. In contrast, RNase III induces several strong cleavages in a regular helix, which carries a sequence similar to the loop-loop interaction. The introduction of a bulged loop that interrupts the regular helix restrains the number of cleavages. This work shows that S. aureus RNase III is able to bind and cleave a variety of RNA-mRNA substrates, and that specific structure elements direct the action of RNase III.}, keywords = {ROMBY, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{fabre_immobilization_2012, title = {Immobilization of double functionalized carbon nanotubes on glassy carbon electrodes for the electrochemical sensing of the biotin–avidin affinity}, author = {Bruno Fabre and Cristian Samorì and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S1572665711005893}, doi = {10.1016/j.jelechem.2011.11.029}, issn = {1572-6657}, year = {2012}, date = {2012-01-01}, urldate = {2020-04-01}, journal = {Journal of Electroanalytical Chemistry}, volume = {665}, pages = {90--94}, abstract = {Multi-walled carbon nanotubes (MWCNTs) double functionalized with redox-active ferrocene and biotin (Fc–Biot-MWCNTs) were synthesized and used for the electrochemical detection of avidin. After dispersion in perfluorosulfonated polymer Nafion and immobilization on the electrode surfaces, the cyclic voltammetry response of the modified electrodes showed in aqueous medium a quasi-reversible one-electron system at 0.46V vs. SCE, assigned to the bound ferrocene/ferrocenium redox couple. Upon the addition of avidin in the range 0.9–20nM, a stepwise decrease of both anodic and cathodic peak currents ascribed to the ferrocene was observed. These electrochemical changes are specifically due to the formation of biotin–avidin complex and are explained by complexation-induced modifications in the environment of covalently bound ferrocene.}, keywords = {Avidin, Biotin, Electrochemical detection, Ferrocene, Functionalized carbon nanotubes, I2CT, Nafion, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Current knowledge on regulatory RNAs and their machineries in Staphylococcus aureus}, author = {C Romilly and I Caldelari and D Parmentier and E Lioliou and P Romby and P Fechter}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22546940}, doi = {0.4161/rna.20103}, isbn = {22546940}, year = {2012}, date = {2012-01-01}, journal = {RNA Biol}, volume = {9}, number = {4}, pages = {402-413}, abstract = {Staphylococcus aureus is one of the major human pathogens, which causes numerous community-associated and hospital-acquired infections. The regulation of the expression of numerous virulence factors is coordinated by complex interplays between two component systems, transcriptional regulatory proteins, and regulatory RNAs. Recent studies have identified numerous novel RNAs comprising cis-acting regulatory RNAs, antisense RNAs, small non coding RNAs and small mRNAs encoding peptides. We present here several examples of RNAs regulating S. aureus pathogenicity and describe various aspects of antisense regulation.}, keywords = {ROMBY, Staphylococcus aureus Gene regulation Peptides Regulatory RNAs RNA-binding proteins Virulence, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {TLR2 Expression Is Regulated by MicroRNA miR-19 in Rheumatoid Fibroblast-like Synoviocytes.}, author = {L Philippe and G Alsaleh and G Suffert and A Meyer and P Georgel and J Sibilia and D Wachsmann and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22105995}, doi = {10.4049/​ jimmunol.1102348}, isbn = {22105995}, year = {2012}, date = {2012-01-01}, journal = {J Immunol}, volume = {188}, number = {1}, pages = {454-461}, abstract = {Resident cells, such as fibroblast-like synoviocytes (FLS), play a crucial role in rheumatoid arthritis (RA). They are implicated in the inflammatory response and play a key role in osteoarticular destruction. Moreover, RA FLS spread RA to unaffected joints. Pathogen-associated molecular patterns and damage-associated molecular patterns have been found to activate RA FLS by interacting with pattern recognition receptors, such as TLR. RA FLS express a large number of TLR, and TLR2 was demonstrated to be involved in RA inflammation. Because microRNA have emerged as important controllers of TLR expression and signaling, the aim of this study was to evaluate their potential involvement in the control of TLR2 expression by RA FLS. We first showed that Tlr2 expression is strongly upregulated in RA FLS in response to TLR2 ligands. Using a microRNA microarray analysis, we identified one miRNA in activated RA FLS, miR-19b, which was downregulated and predicted to target Tlr2 mRNA. Downregulation of miR-19b and miR-19a, which belongs to the same cluster, was confirmed by real-time quantitative PCR. Transfection of RA FLS with miR-19a/b mimics decreased TLR2 protein expression. In parallel, we found that both IL-6 and matrix metalloproteinase 3 secretion was significantly downregulated in activated FLS transfected with either mimic. Moreover, using a luciferase assay, we showed that miR-19a/b directly target Tlr2 mRNA. Taken together, our data point toward an important role for miR-19a/b in the regulation of IL-6 and matrix metalloproteinase 3 release by controlling TLR2 expression, as well as provide evidence that miR-19a/b can act as negative regulators of inflammation in humans.}, note = {Published online: November 21, 2011}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Deregulation of Type I IFN-Dependent Genes Correlates with Increased Susceptibility to Cytomegalovirus Acute Infection of Dicer Mutant Mice.}, author = {E Ostermann and L Tuddenham and C Macquin and G Alsaleh and J Schreiber-Becker and M Tanguy and S Bahram and S Pfeffer and P Georgel}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22916300?dopt=Abstract}, doi = {10.1371/journal.pone.0043744}, isbn = {22916300}, year = {2012}, date = {2012-01-01}, journal = {PLoS One}, volume = {7}, number = {8}, pages = {e43744}, abstract = {Regulation of gene expression by microRNAs (miRNAs) is now considered as an essential mechanism for cell development and homeostasis. Indeed, numerous studies have reported that modulating their expression, maturation, or activity can affect cell survival, identity or activation. In particular, miRNAs are key players in the tight regulation of signaling cascades, and as such, they appear as perfectly suited immunomodulators. Several immune-related processes, including inflammation, have recently been demonstrated to require specific miRNAs. In addition, the discovery of herpesvirus-encoded miRNAs has reinforced this assumption. To decipher the potential roles of miRNAs in innate antiviral immune response, we developed an in vivo model based on the inoculation of mouse cytomegalovirus (MCMV) in mice. Furthermore, we exploited a mouse line carrying a hypomorphic mutation in the Dicer gene to visualize the impact of impaired miRNA biogenesis upon the anti-MCMV response. Our data indicate that miRNAs are important actors in mounting an efficient response against herpesviruses. We suggest that a rapid and transient interferon response following viral infection requires miRNA-dependent repressor release. In addition, our in vivo efforts identified several miRNA targets, thus providing a conceptual framework for future analyzes on the regulation of specific actors involved in the Type I interferon pathway.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A structural module in RNase P expands the variety of RNA kinks.}, author = {M Meyer and E Westhof and B Masquida}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22336704}, isbn = {22336704}, year = {2012}, date = {2012-01-01}, journal = {RNA Biol}, volume = {9}, number = {3}, pages = {254-260}, abstract = {RNA structures are built from recurrent modules that can be identified by structural and comparative sequence analysis. In order to assemble sets of helices in compact architectures, modules that introduce bends and kinks are necessary. Among such modules, kink-turns form an important family that presents sequence and structural characteristics. Here, we describe an internal loop in the bacterial type A RNase P RNA that sets helices bound at the junctions exactly in the same relative positions as in kink-turns but without the structural signatures typical of kink-turns. Our work suggests that identifying a structural module in a subset of RNA sequences constitutes a strategy to identify distinct sequential motifs sharing common structural characteristics.}, keywords = {Kink-turn RNA module RNA motif RNA structure RNase P Structural alignment, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Protein capsules assembled via isobutyramide grafts: sequential growth, biofunctionalization, and cellular uptake.}, author = {D Mertz and J Cui and Y Yan and G Devlin and C Chaubaroux and A Dochter and R Alles and P Lavalle and J C Voegel and A Blencowe and P Auffinger and F Caruso}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22950440?dopt=Abstract http://pubs.acs.org/doi/full/10.1021/nn302024t}, doi = {10.1021/nn302024t}, isbn = {22950440}, year = {2012}, date = {2012-01-01}, journal = {ACS Nano}, volume = {6}, number = {9}, pages = {7584-7594}, abstract = {We report the sequential assembly of proteins via the alternating physical adsorption of human serum albumin (HSA) and chemical grafting with isobutyramide (IBAM) or bromoisobutyramide (BrIBAM) groups. This approach, performed on silica template particles, leads to the formation of noncovalent protein films with controlled growth at the nanometer scale. Further, after template removal, hollow protein capsules with tunable wall thicknesses and high mechanical stability are obtained. The use of BrIBAM, compared to IBAM grafts, leads to significantly thicker capsule walls, highlighting the influence of the bromine atoms in the assembly process, which is discussed in terms of a theoretical model of noncovalent interactions. Another feature of the process is the possibility to functionalize the HSA capsules with other biologically active macromolecules, including enzymes, polysaccharides, or DNA plasmids, demonstrating the versatility of this approach. We also report that BrIBAM-HSA and IBAM-HSA capsules display negligible cytotoxicity in vitro with HeLa cells and that their cellular uptake is dependent on the thickness of the capsule walls. These findings support the potential use of these protein capsules in tailored biological applications such as drug delivery.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA mimicry, a decoy for regulatory proteins.}, author = {S Marzi and P Romby}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22098101}, doi = {10.1111/j.1365-2958.2011.07911.x}, isbn = {22098101}, year = {2012}, date = {2012-01-01}, journal = {Mol Microbiol}, volume = {83}, number = {1}, pages = {1-6}, abstract = {Small non-coding RNA molecules (sRNA) are key regulators participating in complex networks, which adapt metabolism in response to environmental changes. In this issue of Molecular Microbiology, and in a related paper in Proc. Natl. Acad. Sci. USA, Moreno and colleagues and Sonnleitner and colleagues report on novel sRNAs, which act as decoys to inhibit the activity of the master post-transcriptional regulatory protein Crc. Crc is a key protein involved in carbon catabolite repression that optimizes metabolism improving the adaptation of the bacteria to their diverse habitats. Crc is a novel RNA-binding protein that regulates translation of multiple target mRNAs. Two regulatory sRNAs in Pseudomonas putida mimic the natural mRNA targets of Crc and counteract the action of Crc by sequestrating the protein when catabolite repression is absent. Crc trapping by a sRNA is a mechanism reminiscent to the regulation of the repressor of secondary metabolites (RsmA) in Pseudomonas, and highlights the suitability of RNA-dependent regulation to rapidly adjust cell growth in response to environmental changes.}, note = {Article first published online: 21 NOV 2011}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Fifteen years of the yeast three-hybrid system: RNA-protein interactions under investigation.}, author = {F Martin}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22841566?dopt=Abstract}, doi = {10.1016/j.ymeth.2012.07.016}, isbn = {2284156}, year = {2012}, date = {2012-01-01}, journal = {Methods}, volume = {58}, number = {4}, pages = {367-375}, abstract = {In 1996, the Wickens and the Kuhl labs developed the yeast three-hybrid system independently. By expressing two chimeric proteins and one chimeric RNA molecule in Saccharomyces cerevisiae, this method allows in vivo monitoring of RNA-protein interactions by measuring the expression levels of HIS3 and LacZ reporter genes. Specific RNA targets have been used to characterize unknown RNA binding proteins. Previously described RNA binding proteins have also been used as bait to select new RNA targets. Finally, this method has been widely used to investigate or confirm previously suspected RNA-protein interactions. However, this method falls short in some aspects, such as RNA display and selection of false positive molecules. This review will summarize the results obtained with this method from the past 15years, as well as on recent efforts to improve its specificity.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Degradation of cellular mir-27 by a novel, highly abundant viral transcript is important for efficient virus replication in vivo.}, author = {L Marcinowski and M Tanguy and A Krmpotic and B Rädle and V J Lisnić and L Tuddenham and B Chane-Woon-Ming and Z Ruzsics and F Erhard and C Benkartek and M Babic and R Zimmer and J Trgovcich and U H Koszinowski and S Jonjic and S Pfeffer and L Dölken}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22346748}, isbn = {22346748}, year = {2012}, date = {2012-01-01}, journal = {PLoS Pathog}, volume = {8}, number = {2}, pages = {e1002510}, abstract = {Cytomegaloviruses express large amounts of viral miRNAs during lytic infection, yet, they only modestly alter the cellular miRNA profile. The most prominent alteration upon lytic murine cytomegalovirus (MCMV) infection is the rapid degradation of the cellular miR-27a and miR-27b. Here, we report that this regulation is mediated by the ∼1.7 kb spliced and highly abundant MCMV m169 transcript. Specificity to miR-27a/b is mediated by a single, apparently optimized, miRNA binding site located in its 3'-UTR. This site is easily and efficiently retargeted to other cellular and viral miRNAs by target site replacement. Expression of the 3'-UTR of m169 by an adenoviral vector was sufficient to mediate its function, indicating that no other viral factors are essential in this process. Degradation of miR-27a/b was found to be accompanied by 3'-tailing and -trimming. Despite its dramatic effect on miRNA stability, we found this interaction to be mutual, indicating potential regulation of m169 by miR-27a/b. Most interestingly, three mutant viruses no longer able to target miR-27a/b, either due to miRNA target site disruption or target site replacement, showed significant attenuation in multiple organs as early as 4 days post infection, indicating that degradation of miR-27a/b is important for efficient MCMV replication in vivo.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pichia sorbitophila, an Interspecies Yeast Hybrid, Reveals Early Steps of Genome Resolution After Polyploidization.}, author = {V L Louis and L Despons and A Friedrich and T Martin and P Durrens and S Casarégola and C Neuvéglise and C Fairhead and C Marck and J A Cruz and M L Straub and V Kugler and C Sacerdot and Z Uzunov and A Thierry and S Weiss and C Bleykasten and J De Montigny and N Jacques and P Jung and M Lemaire and S Mallet and G Morel and G F Richard and A Sarkar and G Savel and J Schacherer and M L Seret and E Talla and G Samson and C Jubin and J Poulain and B Vacherie and V Barbe and E Pelletier and D J Sherman and E Westhof and J Weissenbach and P V Baret and P Wincker and C Gaillardin and B Dujon and J L Souciet}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22384408?dopt=Abstract}, isbn = {22384408}, year = {2012}, date = {2012-01-01}, journal = {G3 (Bethesda)}, volume = {2}, number = {2}, pages = {299-311}, abstract = {Polyploidization is an important process in the evolution of eukaryotic genomes, but ensuing molecular mechanisms remain to be clarified. Autopolyploidization or whole-genome duplication events frequently are resolved in resulting lineages by the loss of single genes from most duplicated pairs, causing transient gene dosage imbalance and accelerating speciation through meiotic infertility. Allopolyploidization or formation of interspecies hybrids raises the problem of genetic incompatibility (Bateson-Dobzhansky-Muller effect) and may be resolved by the accumulation of mutational changes in resulting lineages. In this article, we show that an osmotolerant yeast species, Pichia sorbitophila, recently isolated in a concentrated sorbitol solution in industry, illustrates this last situation. Its genome is a mosaic of homologous and homeologous chromosomes, or parts thereof, that corresponds to a recently formed hybrid in the process of evolution. The respective parental contributions to this genome were characterized using existing variations in GC content. The genomic changes that occurred during the short period since hybrid formation were identified (e.g., loss of heterozygosity, unilateral loss of rDNA, reciprocal exchange) and distinguished from those undergone by the two parental genomes after separation from their common ancestor (i.e., NUMT (NUclear sequences of MiTochondrial origin) insertions, gene acquisitions, gene location movements, reciprocal translocation). We found that the physiological characteristics of this new yeast species are determined by specific but unequal contributions of its two parents, one of which could be identified as very closely related to an extant Pichia farinosa strain.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Protein analysis by dynamic light scattering: Methods and techniques for students.}, author = {B Lorber and F Fischer and M Bailly and H Roy and D Kern}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23166025}, doi = {10.1002/bmb.20644}, isbn = {23166025}, year = {2012}, date = {2012-01-01}, journal = {Biochem Mol Biol Educ}, volume = {40}, number = {6}, pages = {372-382}, abstract = {Dynamic light scattering (DLS) analyses are routinely used in biology laboratories to detect aggregates in macromolecular solutions, to determine the size of proteins, nucleic acids, and complexes or to monitor the binding of ligands. This article is written for graduate and undergraduate students with access to DLS and for faculty members who wish to incorporate DLS into a lab activity, a practical course or research. It reviews the basic concepts of light scattering measurements and addresses four critical aspects of the analysis and interpretation of DLS results. To ensure reproducible quantitative data, attention should be paid to controlling the preparation and handling of proteins or assemblies because variations in the state of aggregation, induced by minor changes in experimental condition or technique, might compromise DLS results and affect protein activity. Variables like temperature, solvent viscosity, and inter-particle interactions may also influence particle size determination. Every point is illustrated by case studies, including a commercially available albumin, a small RNA virus isolated from plants, as well as four soluble proteins and a ribonucleoprotein assembly purified and characterized by students in the frame of their master degree. © 2012 by The International Union of Biochemistry and Molecular Biology.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Global Regulatory Functions of the Staphylococcus aureus Endoribonuclease III in Gene Expression.}, author = {E Lioliou and C M Sharma and I Caldelari and A C Helfer and P Fechter and F Vandenesch and J Vogel and P Romby}, url = {http://www.plosgenetics.org/article/info%3Adoi%2F10.1371%2Fjournal.pgen.1002782 http://www.ncbi.nlm.nih.gov/pubmed/22761586?dopt=Abstract}, isbn = {22761586}, year = {2012}, date = {2012-01-01}, journal = {PLoS Genet}, volume = {8}, number = {6}, pages = {e1002782}, abstract = {RNA turnover plays an important role in both virulence and adaptation to stress in the Gram-positive human pathogen Staphylococcus aureus. However, the molecular players and mechanisms involved in these processes are poorly understood. Here, we explored the functions of S. aureus endoribonuclease III (RNase III), a member of the ubiquitous family of double-strand-specific endoribonucleases. To define genomic transcripts that are bound and processed by RNase III, we performed deep sequencing on cDNA libraries generated from RNAs that were co-immunoprecipitated with wild-type RNase III or two different cleavage-defective mutant variants in vivo. Several newly identified RNase III targets were validated by independent experimental methods. We identified various classes of structured RNAs as RNase III substrates and demonstrated that this enzyme is involved in the maturation of rRNAs and tRNAs, regulates the turnover of mRNAs and non-coding RNAs, and autoregulates its synthesis by cleaving within the coding region of its own mRNA. Moreover, we identified a positive effect of RNase III on protein synthesis based on novel mechanisms. RNase III-mediated cleavage in the 5' untranslated region (5'UTR) enhanced the stability and translation of cspA mRNA, which encodes the major cold-shock protein. Furthermore, RNase III cleaved overlapping 5'UTRs of divergently transcribed genes to generate leaderless mRNAs, which constitutes a novel way to co-regulate neighboring genes. In agreement with recent findings, low abundance antisense RNAs covering 44% of the annotated genes were captured by co-immunoprecipitation with RNase III mutant proteins. Thus, in addition to gene regulation, RNase III is associated with RNA quality control of pervasive transcription. Overall, this study illustrates the complexity of post-transcriptional regulation mediated by RNase III.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {RNA-mediated regulation of virulence gene expression: another layer of complexity}, author = {E Lioliou and C Romilly and T Geissman and F Vandenesch and P Romby}, editor = {A Filloux}, url = {http://www.horizonpress.com/regulatorynetworks}, year = {2012}, date = {2012-01-01}, booktitle = {Bacterial Regulatory Networks}, pages = {143-166}, publisher = {Caister Academic Press}, address = {Norwich, UK}, abstract = {Many pathogenic bacteria cause serious diseases in humans, animals, and plants. Due to the appearance of resistance to multiple antibiotics, it has become important to fully understand the regulatory networks that lead to the production of virulence factors that help the bacteria combat the host defense machinery, acquire nutrients, and survive and/or proliferate within the host. In recent years, complex interplays between transcriptional regulatory proteins, two-component systems, and regulatory RNAs have been described, establishing the gene expression patterns in pathogenic bacteria. In this review, several examples will illustrate the diversity of regulatory RNAs and how they are integrated into the regulatory circuits required for virulence gene expression, with special emphasis on the mechanisms of regulation at the molecular level.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Selenoprotein N: Its role in disease.}, author = {A Lescure and P Castets and D Grunwald and V Allamand and M Howard}, editor = {D Hatfield and M Berry and V Gladyshev}, url = {http://www.springerlink.com/content/h457r0g274533522/}, doi = {10.1007/978-1-4614-1025-6_22}, year = {2012}, date = {2012-01-01}, booktitle = {Selenium: Its molecular biology and role in human health}, pages = {283-294}, publisher = {Springer}, edition = {2}, abstract = {Selenoprotein N is among the newly identified selenoproteins, initially discovered in silico with no known molecular function. It has become the focus of attention because mutations in the selenoprotein N gene are linked to a group of muscle disorders, now referred as SEPN1-related myopathies. An emerging view arising from recent findings is that the loss of selenoprotein N leads to cellular sensitivity to oxidative stress and loss of calcium homeostasis. Studies of animal models for SEPN1-Related Myopathies revealed the fate of sensitized muscle may depend on stresses to which it is subjected, and defects in the function of selenoprotein N-deficient muscle progenitor cells during development in zebrafish embryos or during muscle regeneration in fully developed mouse muscle. Dysfunction of these different processes raises significant questions regarding which of the phenotypic manifestations of SEPN1-Related Myopathies are initiated by events during development and which are progressive in nature arising from dysfunction of mature muscle.}, keywords = {KROL, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Modeling RNA Molecules}, author = {N Leontis and E Westhof}, editor = {N Leontis and E Westhof}, url = {http://link.springer.com/chapter/10.1007/978-3-642-25740-7_2}, doi = {10.1007/978-3-642-25740-7_2}, year = {2012}, date = {2012-01-01}, booktitle = {RNA 3D Structure Analysis and Predication}, pages = {5-17}, publisher = {Springer}, series = {Nucleic Acids and Molecular Biology}, abstract = {A primary activity of scientific work is the construction of models to represent the nature and workings of phenomena we observe in the world around us. Models that represent the molecular components of living system in three dimensions (3D) and at atomic resolution are highly valued in molecular and structural biology. For example, the decipherment of the 3D structures of ribosomes, the complex protein-synthesizing nanomachines of the cell, represents a tremendous achievement, recently recognized with the Nobel Prize in Chemistry (http://nobelprize.org/nobel_prizes/chemistry/laureates/2009/). Nonetheless, this phenomenal success is tempered by the realization that even now, over 10 years after the first ribosome structures were solved, we still do not understand fully several aspects of their functioning. For all who have grappled with the complexities of ribosome structures, Richard Feynmannメs pithy statement, モWhat I cannot create, I do not understand,ヤ rings especially true (Hawking 2001). This physics-based realization contrasts with another point of view of modeling. To paraphrase R. W. Hamming, who said, モThe purpose of computing is insight, not numbersヤ (Hamming 1971), we should remember that the purpose of molecular modeling is functional insight, not detailed atomic models per se. Therefore, as we seek to improve our abilities to construct 3D models for molecules for which we do not yet have experimental atomic-resolution structures, we should bear in mind that it may not be necessary to achieve some arbitrary precision in the atomic coordinates to provide insight into biological function. Rather, we should think carefully to identify those predicted features that yield important insights}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @book{, title = {RNA 3D Structure Analysis and Prediction}, editor = {N Leontis and E Westhof}, year = {2012}, date = {2012-01-01}, publisher = {Springer, Berlin}, series = {Nucleic Acids and Molecular Biology}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {book} } @article{, title = {Automated prediction of three-way junction topological families in RNA secondary structures.}, author = {A Lamiable and D Barth and A Denise and F Quessette and S Vial and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22326420}, doi = {10.1016/j.compbiolchem.2011.11.001}, isbn = {22326420}, year = {2012}, date = {2012-01-01}, journal = {Comput Biol Chem}, volume = {37}, pages = {1-5}, abstract = {We present an algorithm for automatically predicting the topological family of any RNA three-way junction, given only the information from the secondary structure: the sequence and the Watson-Crick pairings. The parameters of the algorithm have been determined on a data set of 33 three-way junctions whose 3D conformation is known. We applied the algorithm on 53 other junctions and compared the predictions to the real shape of those junctions. We show that the correct answer is selected out of nine possible configurations 64% of the time. Additionally, these results are noticeably improved if homology information is used. The resulting software, Cartaj, is available online and downloadable (with source) at: http://cartaj.lri.fr.}, note = {Available online 11 January 2012.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Armless mitochondrial tRNAs in enoplea (nematoda).}, author = {F Juhling and J Putz and C Florentz and P F Stadler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23018779?dopt=Abstract}, doi = {10.4161/rna.21630}, isbn = {23018779}, year = {2012}, date = {2012-01-01}, journal = {RNA Biol}, volume = {9}, number = {9}, pages = {1161 - 1166}, abstract = {The mitochondrial genome of metazoan animal typically encodes 22 tRNAs. Nematode mt-tRNAs normally lack the T-stem and instead feature a replacement loop. In the class Enoplea, putative mt-tRNAs that are even further reduced have been predicted to lack both the T- and the D-arm. Here we investigate these tRNA candidates in detail. Three lines of computational evidence support that they are indeed minimal functional mt-tRNAs: (1) the high level of conservation of both sequence and secondary structure, (2) the perfect preservation of the anticodons, and (3) the persistence of these sequence elements throughout several genome rearrangements that place them between different flanking genes.}, keywords = {Enoplea Mermithidae Mitochondrial tRNA D-loop T-loop, FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Improved systematic tRNA gene annotation allows new insights into the evolution of mitochondrial tRNA structures and into the mechanisms of mitochondrial genome rearrangements.}, author = {F Juhling and J Putz and M Bernt and A Donath and M Middendorf and C Florentz and P F Stadler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22139921}, doi = {10.1093/nar/gkr1131}, isbn = {22139921}, year = {2012}, date = {2012-01-01}, journal = {Nucleic Acids Res}, volume = {40}, number = {7}, pages = {2833-2845}, abstract = {Transfer RNAs (tRNAs) are present in all types of cells as well as in organelles. tRNAs of animal mitochondria show a low level of primary sequence conservation and exhibit 'bizarre' secondary structures, lacking complete domains of the common cloverleaf. Such sequences are hard to detect and hence frequently missed in computational analyses and mitochondrial genome annotation. Here, we introduce an automatic annotation procedure for mitochondrial tRNA genes in Metazoa based on sequence and structural information in manually curated covariance models. The method, applied to re-annotate 1876 available metazoan mitochondrial RefSeq genomes, allows to distinguish between remaining functional genes and degrading 'pseudogenes', even at early stages of divergence. The subsequent analysis of a comprehensive set of mitochondrial tRNA genes gives new insights into the evolution of structures of mitochondrial tRNA sequences as well as into the mechanisms of genome rearrangements. We find frequent losses of tRNA genes concentrated in basal Metazoa, frequent independent losses of individual parts of tRNA genes, particularly in Arthropoda, and wide-spread conserved overlaps of tRNAs in opposite reading direction. Direct evidence for several recent Tandem Duplication-Random Loss events is gained, demonstrating that this mechanism has an impact on the appearance of new mitochondrial gene orders.}, note = {First published online: December 1, 2011}, keywords = {FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {When ribonucleases come into play in pathogens: a survey of gram-positive bacteria.}, author = {B C Jester and P Romby and E Lioliou}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22550495}, doi = {10.1155/2012/592196}, issn = {ISSN: 1687-918X (Print) ISSN: 1687-9198 (Online)}, year = {2012}, date = {2012-01-01}, journal = {Int J Microbiol}, volume = {2012}, pages = {592196}, abstract = {It is widely acknowledged that RNA stability plays critical roles in bacterial adaptation and survival in different environments like those encountered when bacteria infect a host. Bacterial ribonucleases acting alone or in concert with regulatory RNAs or RNA binding proteins are the mediators of the regulatory outcome on RNA stability. We will give a current update of what is known about ribonucleases in the model Gram-positive organism Bacillus subtilis and will describe their established roles in virulence in several Gram-positive pathogenic bacteria that are imposing major health concerns worldwide. Implications on bacterial evolution through stabilization/transfer of genetic material (phage or plasmid DNA) as a result of ribonucleases' functions will be covered. The role of ribonucleases in emergence of antibiotic resistance and new concepts in drug design will additionally be discussed.}, note = {Epub 2012 Mar 13.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vivo identification of essential nucleotides in tRNALeu to its functions by using a constructed yeast tRNALeu knockout strain.}, author = {Q Huang and P Yao and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22917587?dopt=Abstract}, doi = {10.1093/nar/gks783}, isbn = {22917587}, year = {2012}, date = {2012-01-01}, journal = {Nucleic Acids Res}, volume = {40}, number = {20}, pages = {10463-10477}, abstract = {The fidelity of protein biosynthesis requires the aminoacylation of tRNA with its cognate amino acid catalyzed by aminoacyl-tRNA synthetase with high levels of accuracy and efficiency. Crucial bases in tRNA(Leu) to aminoacylation or editing functions of leucyl-tRNA synthetase have been extensively studied mainly by in vitro methods. In the present study, we constructed two Saccharomyces cerevisiae tRNA(Leu) knockout strains carrying deletions of the genes for tRNA(Leu)(GAG) and tRNA(Leu)(UAG). Disrupting the single gene encoding tRNA(Leu)(GAG) had no phenotypic consequence when compared to the wild-type strain. While disrupting the three genes for tRNA(Leu)(UAG) had a lethal effect on the yeast strain, indicating that tRNA(Leu)(UAG) decoding capacity could not be compensated by another tRNA(Leu) isoacceptor. Using the triple tRNA knockout strain and a randomly mutated library of tRNA(Leu)(UAG), a selection to identify critical tRNA(Leu) elements was performed. In this way, mutations inducing in vivo decreases of tRNA levels or aminoacylation or editing ability by leucyl-tRNA synthetase were identified. Overall, the data showed that the triple tRNA knockout strain is a suitable tool for in vivo studies and identification of essential nucleotides of the tRNA.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoacyl-tRNA Synthetases in the Bacterial World.}, author = {R Giege and M Springer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/26442825}, doi = {10.1128/ecosalplus.4.2.1}, isbn = {26442825}, year = {2012}, date = {2012-01-01}, journal = {EcoSal Plus}, volume = {5}, number = {1}, pages = {none}, abstract = {Aminoacyl-tRNAsynthetases (aaRSs) are modular enzymesglobally conserved in the three kingdoms of life. All catalyze the same two-step reaction, i.e., the attachment of a proteinogenic amino acid on their cognate tRNAs, thereby mediating the correct expression of the genetic code. In addition, some aaRSs acquired other functions beyond this key role in translation.Genomics and X-ray crystallography have revealed great structural diversity in aaRSs (e.g.,in oligomery and modularity, in ranking into two distinct groups each subdivided in 3 subgroups, by additional domains appended on the catalytic modules). AaRSs show hugestructural plasticity related to function andlimited idiosyncrasies that are kingdom or even speciesspecific (e.g.,the presence in many Bacteria of non discriminating aaRSs compensating for the absence of one or two specific aaRSs, notably AsnRS and/or GlnRS).Diversity, as well, occurs in the mechanisms of aaRS gene regulation that are not conserved in evolution, notably betweendistant groups such as Gram-positive and Gram-negative Bacteria.Thereview focuses on bacterial aaRSs (and their paralogs) and covers their structure, function, regulation,and evolution. Structure/function relationships are emphasized, notably the enzymology of tRNA aminoacylation and the editing mechanisms for correction of activation and charging errors. The huge amount of genomic and structural data that accumulatedin last two decades is reviewed,showing how thefield moved from essentially reductionist biologytowards more global and integrated approaches. Likewise, the alternative functions of aaRSs and those of aaRSparalogs (e.g., during cellwall biogenesis and other metabolic processes in or outside protein synthesis) are reviewed. Since aaRS phylogenies present promiscuous bacterial, archaeal, and eukaryal features, similarities and differences in the properties of aaRSs from the three kingdoms of life are pinpointedthroughout the reviewand distinctive characteristics of bacterium-like synthetases from organelles are outlined.}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of transfer RNAs: similarity and variability}, author = {R Giege and F Juhling and J Putz and P Stadler and C Sauter and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21957054?dopt=Abstrac}, doi = {10.1002/wrna.103}, isbn = {21957054}, year = {2012}, date = {2012-01-01}, journal = {Wiley Interdiscip Rev RNA}, volume = {3}, number = {1}, pages = {37-61}, abstract = {Transfer RNAs (tRNAs) are ancient molecules whose origin goes back to the beginning of life on Earth. Key partners in the ribosome-translation machinery, tRNAs read genetic information on messenger RNA and deliver codon specified amino acids attached to their distal 3′-extremity for peptide bond synthesis on the ribosome. In addition to this universal function, tRNAs participate in a wealth of other biological processes and undergo intricate maturation events. Our understanding of tRNA biology has been mainly phenomenological, but ongoing progress in structural biology is giving a robust physico-chemical basis that explains many facets of tRNA functions. Advanced sequence analysis of tRNA genes and their RNA transcripts have uncovered rules that underly tRNA 2D folding and 3D L-shaped architecture, as well as provided clues about their evolution. The increasing number of X-ray structures of free, protein- and ribosome-bound tRNA, reveal structural details accounting for the identity of the 22 tRNA families (one for each proteinogenic amino acid) and for the multifunctionality of a given family. Importantly, the structural role of post-transcriptional tRNA modifications is being deciphered. On the other hand, the plasticity of tRNA structure during function has been illustrated using a variety of technical approaches that allow dynamical insights. The large range of structural properties not only allows tRNAs to be the key actors of translation, but also sustain a diversity of unrelated functions from which only a few have already been pinpointed. Many surprises can still be expected. WIREs RNA 2012, 3:37–61. doi: 10.1002/wrna.103}, keywords = {FLORENTZ, FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Re-designed N-terminus enhances expression, solubility and crystallizability of mitochondrial protein.}, author = {A Gaudry and B Lorber and A Neuenfeldt and C Sauter and C Florentz and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22871419?report=&dispmax=200&tool=PubCrawler_2.23}, isbn = {22871419}, year = {2012}, date = {2012-01-01}, journal = {Protein Eng Des Sel}, volume = {25}, number = {9}, pages = {473-481}, abstract = {Mitochondrial aminoacyl-tRNA synthetases are key enzymes in translation. They are encoded by the nuclear genome, synthesized as precursors in the cytosol and imported. Most are matured by cleavage of their N-terminal targeting sequence. The poor expression of mature proteins in prokaryotic systems, along with their low solubility and stability after purification are major obstacles for biophysical and crystallographic studies. The purpose of the present work was to analyze the influence of additives on a slightly soluble aspartyl-tRNA synthetase and of the N-terminal sequence of the protein on its expression and solubility. On the one hand, the solubility of the enzyme was augmented to some extent in the presence of a chemical analog of the intermediary product aspartyl-adenylate, 5'-O-[N-(L aspartyl) sulfamoyl] adenosine. On the other hand, expression was enhanced by extending the N-terminus by seven natural amino acids from the predicted targeting sequence. The re-designed enzyme was active, monodisperse, more soluble and yielded crystals that are suitable for structure determination. This result underlines the importance of the N-terminal residue sequence for solubility. It suggests that additional criteria should be taken into account for the prediction of cleavage sites in mitochondrial targeting sequences.}, keywords = {FLORENTZ, FRUGIER, SAUTER, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence and structure requirements for specific recognition of HIV-1 TAR and DIS RNA by the HIV-1 Vif protein.}, author = {S Freisz and J Mezher and L Hafirassou and P Wolff and Y Nomine and C Romier and P Dumas and E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22767258?dopt=Abstract}, isbn = {22767258}, year = {2012}, date = {2012-01-01}, journal = {RNA Biol}, volume = {9}, number = {7}, pages = {966 - 977}, abstract = {The HIV-1 Vif protein plays an essential role in the regulation of the infectivity of HIV-1 virion and in vivo pathogenesis. Vif neutralizes the human DNA-editing enzyme APOBEC3 protein, an antiretroviral cellular factor from the innate immune system, allowing the virus to escape the host defence system. It was shown that Vif is packaged into viral particles through specific interactions with the viral genomic RNA. Conserved and structured sequences from the 5'-noncoding region, such as the Tat-responsive element (TAR) or the genomic RNA dimerization initiation site (DIS), are primary binding sites for Vif. In the present study we used isothermal titration calorimetry to investigate sequence and structure determinants important for Vif binding to short viral RNA corresponding to TAR and DIS stem-loops. We showed that Vif specifically binds TAR and DIS in the low nanomolar range. In addition, Vif primarily binds the TAR UCU bulge, but not the apical loop. Determinants for Vif binding to the DIS loop-loop complex are likely more complex and involve the self-complementary loop together with the upper part of the stem. These results suggest that Tat-TAR inhibitors or DIS small molecule binders might be also effective to disturb Vif-TAR and Vif-DIS binding in order to reduce Vif packaging into virions.}, keywords = {ENNIFAR, HIV Biophysics RNA-protein interactions Microcalorimetry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A supramolecular assembly formed by influenza A virus genomic RNA segments.}, author = {E Fournier and V Moules and B Essere and J C Paillart and J D Sirbat and C Isel and A Cavalier and J P Rolland and D Thomas and B Lina and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22075989}, doi = {10.1093/nar/gkr985}, isbn = {22075989}, year = {2012}, date = {2012-01-01}, journal = {Nucleic Acids Res}, volume = {40}, number = {5}, pages = {2197-2209}, abstract = {The influenza A virus genome consists of eight viral RNAs (vRNAs) that form viral ribonucleoproteins (vRNPs). Even though evidence supporting segment-specific packaging of vRNAs is accumulating, the mechanism ensuring selective packaging of one copy of each vRNA into the viral particles remains largely unknown. We used electron tomography to show that the eight vRNPs emerge from a common 'transition zone' located underneath the matrix layer at the budding tip of the virions, where they appear to be interconnected and often form a star-like structure. This zone appears as a platform in 3D surface rendering and is thick enough to contain all known packaging signals. In vitro, all vRNA segments are involved in a single network of intermolecular interactions. The regions involved in the strongest interactions were identified and correspond to known packaging signals. A limited set of nucleotides in the 5' region of vRNA 7 was shown to interact with vRNA 6 and to be crucial for packaging of the former vRNA. Collectively, our findings support a model in which the eight genomic RNA segments are selected and packaged as an organized supramolecular complex held together by direct base pairing of the packaging signals.}, note = {First published online: November 10, 2011}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interaction network linking the human H3N2 influenza A virus genomic RNA segments.}, author = {E Fournier and V Moules and B Essere and J C Paillart and J D Sirbat and A Cavalier and J P Rolland and D Thomas and B Lina and C Isel and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23063835?dopt=Abstract}, doi = {10.1016/j.vaccine.2012.09.079}, issn = {ISSN: 0264-410X}, year = {2012}, date = {2012-01-01}, journal = {Vaccine}, volume = {30}, number = {51}, pages = {7359-7367}, abstract = {he genome of influenza A viruses is comprised of eight negative-sense viral RNAs (vRNAs) that form viral ribonucleoproteins (vRNPs). In order to be infectious, an influenza A viral particle must encapsidate at least one copy of each of the vRNAs. Thus, even though genome segmentation is evolutionary advantageous, it undeniably complicates viral assembly, which is believed to occur through a selective mechanism that still remains to be understood. Using electron tomography 3D-reconstructions, we show that the eight vRNPs of an influenza A Moscow/10/99 (H3N2) virus are interconnected within a star-like structure as they emerge from a unique "transition zone" at the budding tip of the virions. Notably, this "transition zone" is thick enough to accommodate all described packaging signals. We also report that, in vitro, each vRNA segment is involved in a direct contact with at least one other vRNA partner, in a single network of intermolecular interactions. We show that in several cases, the regions involved in vRNA/vRNA interactions overlap with previously identified packaging signals. Our results thus provide support for the involvement of RNA/RNA interactions in the selection and specific packaging of influenza A genomic RNAs, which appear embedded into an organised supramolecular complex likely held together by direct base-pairings between packaging signals.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{hamrita_towards_2012, title = {[Towards a better knowledge of breast cancer physiopathology: the proteomics approach].}, author = {Bechr Hamrita and Hela Ben Nasr and Philippe Hammann and Lauriane Kuhn and Amel Ben Anes and Saloua Dimassi and Anouar Chaieb and Hedi Khairi and Karim Chahed}, doi = {10.1684/abc.2012.0741}, issn = {1950-6112 0003-3898}, year = {2012}, date = {2012-01-01}, journal = {Annales de biologie clinique}, volume = {70}, number = {5}, pages = {553--565}, abstract = {Breast cancer represents a major public health problem. Approximately one woman in ten is likely to develop a malignant tumor of the breast in their lifetime. The frequency of breast cancer is rising steadily for 20 years and the practical benefits in the diagnosis, prognosis and treatment of this disease are still too limited. Actually, there is no tumor marker with a specificity and sensitivity sufficient to have an utility in clinical and early diagnosis of breast cancer, although, carcinoembryonic antigen (CEA), MUC-1 and CA 15-3 were reported to be useful as markers for monitoring this disease. Thus, proteomics approaches are needed for the discovery and the identification of new protein biomarkers that may allow a better understanding of biological mechanisms of breast tumor development and serve as potential therapeutic targets. This article reviews advances in this field, as well as, the major contribution of these markers in breast pathology, with a focus on their biological characteristics and their clinical and therapeutic involvement.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {The asparagine-transamidosome from Helicobacter pylori: a dual-kinetic mode in non-discriminating aspartyl-tRNA synthetase safeguards the genetic code.}, author = {F Fischer and J L Huot and B Lorber and G Diss and T L Hendrickson and H D Becker and J Lapointe and D Kern}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22362756?dopt=Abstract}, doi = {10.1093/nar/gks167}, isbn = {22362756}, year = {2012}, date = {2012-01-01}, journal = {Nucleic Acids Res}, volume = {40}, number = {11}, pages = {4965-4976}, abstract = {Helicobacter pylori catalyzes Asn-tRNA(Asn) formation by use of the indirect pathway that involves charging of Asp onto tRNA(Asn) by a non-discriminating aspartyl-tRNA synthetase (ND-AspRS), followed by conversion of the mischarged Asp into Asn by the GatCAB amidotransferase. We show that the partners of asparaginylation assemble into a dynamic Asn-transamidosome, which uses a different strategy than the Gln-transamidosome to prevent the release of the mischarged aminoacyl-tRNA intermediate. The complex is described by gel-filtration, dynamic light scattering and kinetic measurements. Two strategies for asparaginylation are shown: (i) tRNA(Asn) binds GatCAB first, allowing aminoacylation and immediate transamidation once ND-AspRS joins the complex; (ii) tRNA(Asn) is bound by ND-AspRS which releases the Asp-tRNA(Asn) product much slower than the cognate Asp-tRNA(Asp); this kinetic peculiarity allows GatCAB to bind and transamidate Asp-tRNA(Asn) before its release by the ND-AspRS. These results are discussed in the context of the interrelation between the Asn and Gln-transamidosomes which use the same GatCAB in H. pylori, and shed light on a kinetic mechanism that ensures faithful codon reassignment for Asn.}, note = {First published online: February 22, 2012}, keywords = {BECKER KERN FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Adaptation of aminoacylation identity rules to mammalian mitochondria.}, author = {A Fender and A Gaudry and F Jühling and M Sissler and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22402012?dopt=Abstract}, doi = {10.1016/j.biochi.2012.02.030}, isbn = {22402012}, year = {2012}, date = {2012-01-01}, journal = {Biochimie}, volume = {94}, number = {5}, pages = {1090-1097}, abstract = {Many mammalian mitochondrial aminoacyl-tRNA synthetases are of bacterial-type and share structural domains with homologous bacterial enzymes of the same specificity. Despite this high similarity, synthetases from bacteria are known for their inability to aminoacylate mitochondrial tRNAs, while mitochondrial enzymes do aminoacylate bacterial tRNAs. Here, the reasons for non-aminoacylation by a bacterial enzyme of a mitochondrial tRNA have been explored. A mutagenic analysis performed on in vitro transcribed human mitochondrial tRNA(Asp) variants tested for their ability to become aspartylated by Escherichia coli aspartyl-tRNA synthetase, reveals that full conversion cannot be achieved on the basis of the currently established tRNA/synthetase recognition rules. Integration of the full set of aspartylation identity elements and stabilization of the structural tRNA scaffold by restoration of D- and T-loop interactions, enable only a partial gain in aspartylation efficiency. The sequence context and high structural instability of the mitochondrial tRNA are additional features hindering optimal adaptation of the tRNA to the bacterial enzyme. Our data support the hypothesis that non-aminoacylation of mitochondrial tRNAs by bacterial synthetases is linked to the large sequence and structural relaxation of the organelle encoded tRNAs, itself a consequence of the high rate of mitochondrial genome divergence.}, note = {Available online 1 March 2012}, keywords = {FLORENTZ, Identity elements Mitochondrial tRNA Cross-aminoacylation Structural plasticity, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {2'-Azido RNA, a versatile tool for chemical biology: synthesis, X-ray structure, siRNA applications, click labeling.}, author = {K Fauster and M Hartl and T Santner and M Aigner and C Kreutz and K Bister and E Ennifar and R Micura}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22273279}, doi = {10.1021/cb200510k}, isbn = {22273279}, year = {2012}, date = {2012-01-01}, journal = {ACS Chem Biol}, volume = {7}, number = {3}, pages = {581-589}, abstract = {Chemical modification can significantly enrich the structural and functional repertoire of ribonucleic acids and endow them with new outstanding properties. Here, we report the syntheses of novel 2'-azido cytidine and 2'-azido guanosine building blocks and demonstrate their efficient site-specific incorporation into RNA by mastering the synthetic challenge of using phosphoramidite chemistry in the presence of azido groups. Our study includes the detailed characterization of 2'-azido nucleoside containing RNA using UV-melting profile analysis and CD and NMR spectroscopy. Importantly, the X-ray crystallographic analysis of 2'-azido uridine and 2'-azido adenosine modified RNAs reveals crucial structural details of this modification within an A-form double helical environment. The 2'-azido group supports the C3'-endo ribose conformation and shows distinct water-bridged hydrogen bonding patterns in the minor groove. Additionally, siRNA induced silencing of the brain acid soluble protein (BASP1) encoding gene in chicken fibroblasts demonstrated that 2'-azido modifications are well tolerated in the guide strand, even directly at the cleavage site. Furthermore, the 2'-azido modifications are compatible with 2'-fluoro and/or 2'-O-methyl modifications to achieve siRNAs of rich modification patterns and tunable properties, such as increased nuclease resistance or additional chemical reactivity. The latter was demonstrated by the utilization of the 2'-azido groups for bioorthogonal Click reactions that allows efficient fluorescent labeling of the RNA. In summary, the present comprehensive investigation on site-specifically modified 2'-azido RNA including all four nucleosides provides a basic rationale behind the physico- and biochemical properties of this flexible and thus far neglected type of RNA modification.}, note = {Online: January 24, 2012}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A pyrophosphatase activity associated with purified HIV-1 particles.}, author = {C Ducloux and M Mougel and V Goldschmidt and L Didierlaurent and R Marquet and C Isel}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22766015?dopt=Abstract}, doi = {10.1016/j.biochi.2012.06.025}, isbn = {22766015}, year = {2012}, date = {2012-01-01}, journal = {Biochimie}, volume = {94}, number = {12}, pages = {2498-2507}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A new understanding of the decoding principle on the ribosome.}, author = {N Demeshkina and L Jenner and E Westhof and M Yusupov and G Yusupova}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22437501?dopt=Abstract}, doi = {10.1038/nature10913}, isbn = {22437501}, year = {2012}, date = {2012-01-01}, journal = {Nature}, volume = {484}, number = {7393}, pages = {256-259}, abstract = {During protein synthesis, the ribosome accurately selects transfer RNAs (tRNAs) in accordance with the messenger RNA (mRNA) triplet in the decoding centre. tRNA selection is initiated by elongation factor Tu, which delivers tRNA to the aminoacyl tRNA-binding site (A site) and hydrolyses GTP upon establishing codon-anticodon interactions in the decoding centre. At the following proofreading step the ribosome re-examines the tRNA and rejects it if it does not match the A codon. It was suggested that universally conserved G530, A1492 and A1493 of 16S ribosomal RNA, critical for tRNA binding in the A site, actively monitor cognate tRNA, and that recognition of the correct codon-anticodon duplex induces an overall ribosome conformational change (domain closure). Here we propose an integrated mechanism for decoding based on six X-ray structures of the 70S ribosome determined at 3.1-3.4 Å resolution, modelling cognate or near-cognate states of the decoding centre at the proofreading step. We show that the 30S subunit undergoes an identical domain closure upon binding of either cognate or near-cognate tRNA. This conformational change of the 30S subunit forms a decoding centre that constrains the mRNA in such a way that the first two nucleotides of the A codon are limited to form Watson-Crick base pairs. When U·G and G·U mismatches, generally considered to form wobble base pairs, are at the first or second codon-anticodon position, the decoding centre forces this pair to adopt the geometry close to that of a canonical C·G pair. This by itself, or with distortions in the codon-anticodon mini-helix and the anticodon loop, causes the near-cognate tRNA to dissociate from the ribosome.}, note = {Published online 21 March 2012}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nonrandom distribution of cryptic repeating triplets of purines and pyrimidines (RNY)n in gp120 of HIV-1.}, author = {E De Crignis and S Guglietta and B T Foley and M Negroni and A F Di Narzo and V Waelti Da Costa and M Cavassini and P A Bart and G Pantaleo and C Graziosi}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21902591}, doi = {10.1089/AID.2011.0208}, isbn = {21902591}, year = {2012}, date = {2012-01-01}, journal = {AIDS Res Hum Retroviruses}, volume = {28}, number = {5}, pages = {:493-504}, abstract = {We have analyzed purine (R) and pyrimidine (Y) codon patterns in variable and constant regions of HIV-1 gp120 in seven patients infected with different HIV-1 subtypes and naïve to antiretroviral therapy. We have calculated the relative frequency of each in-frame codon RNY, YNR, RNR, and YNY (N=any nucleotide) in variable and constant regions of gp120, in the sequence comprised within indels and at indels' flanking sites. Our data show that hypervariable regions V1, V2, V4, and V5 are characterized by the presence of long stretches of RNY codons constituting the majority of the sequence portion comprised within insertions/deletions. In full length gp120 and within inserted/deleted fragments the number of AVT (V=A, C, G) codons did not exceed 50% of the total RNY codons. RNY strings in variable regions spanned up to 21 codons and were always in-frame. In contrast, RNY strings in constant regions were mostly out-of-frame and their length was limited to 5 codons. The frequency of the codon RNY was found to be significantly higher in variable regions (p<0.0001; t-test), within indels, and at indels' flanking sites (p<0.0001; χ² test). Analysis of the distribution of RNY strings equal to or longer than 5 codons in the full genome of HXB2 also shows that these sequences are mostly out-of-frame, unless they contain a potential N-glycosylation site or an Asparagine. These data suggest that cryptic repeats of RNY may play a role in the genesis of multiple base insertions and deletions in hypervariable regions of gp120.}, note = {NEGRONI}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reconstitution of the entire hepatitis C virus life cycle in non-hepatic cells.}, author = {D Da Costa and M Turek and D J Felmlee and E Girardi and S Pfeffer and G Long and R Bartenschlager and M B Zeisel and T F Baumert}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22896615?dopt=Abstract}, doi = {10.1128/JVI.01066-12}, isbn = {22896615}, year = {2012}, date = {2012-01-01}, journal = {J Virol}, volume = {86}, number = {21}, pages = {11919-11925}, abstract = {Hepatitis C virus (HCV) is a human hepatotropic virus, yet the relevant host factors restricting HCV infection to hepatocytes are only partially understood. We demonstrate that exogenous expression of defined host factors reconstituted the entire HCV life cycle in human non-hepatic 293T cells. This study shows robust HCV entry, RNA replication, and production of infectious virus in human non-hepatic cells, and highlights key host factors required for liver tropism of HCV.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA-Puzzles: A CASP-like evaluation of RNA three-dimensional structure prediction.}, author = {J A Cruz and M F Blanchet and M Boniecki and J M Bujnicki and S J Chen and S Cao and R Das and F Ding and N V Dokholyan and S C Flores and L Huang and C A Lavender and V Lisi and F Major and K Mikolajczak and D J Patel and A Philips and T Puton and J Santalucia and F Sijenyi and T Hermann and K Rother and M Rother and A Serganov and M Skorupski and T Soltysinski and P Sripakdeevong and I Tuszynska and K M Weeks and C Waldsich and M Wildauer and N B Leontis and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22361291?dopt=Abstract}, doi = {10.1261/rna.031054.111}, isbn = {2236129}, year = {2012}, date = {2012-01-01}, journal = {RNA}, volume = {18}, number = {4}, pages = {610-625}, abstract = {We report the results of a first, collective, blind experiment in RNA three-dimensional (3D) structure prediction, encompassing three prediction puzzles. The goals are to assess the leading edge of RNA structure prediction techniques; compare existing methods and tools; and evaluate their relative strengths, weaknesses, and limitations in terms of sequence length and structural complexity. The results should give potential users insight into the suitability of available methods for different applications and facilitate efforts in the RNA structure prediction community in ongoing efforts to improve prediction tools. We also report the creation of an automated evaluation pipeline to facilitate the analysis of future RNA structure prediction exercises.}, note = {Published in Advance February 23, 2012}, keywords = {3D prediction Bioinformatics Force fields Structure, Unité ARN, WESTHOF, WESTHOF 3D prediction Bioinformatics Force fields Structure}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutation in a primate-conserved retrotransposon reveals a noncoding RNA as a mediator of infantile encephalopathy.}, author = {F Cartault and P Munier and E Benko and I Desguerre and S Hanein and N Boddaert and S Bandiera and J Vellayoudom and P Krejbich-Trotot and M Bintner and J J Hoarau and M Girard and E Génin and P de Lonlay and A Fourmaintraux and M Naville and D Rodriguez and J Feingold and M Renouil and A Munnich and E Westhof and M Fähling and S Lyonnet and A Henrion-Caude}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22411793?dopt=Abstract}, doi = {10.1073/pnas.1111596109}, isbn = {22411793}, year = {2012}, date = {2012-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {109}, number = {13}, pages = {4980-4985}, abstract = {The human genome is densely populated with transposons and transposon-like repetitive elements. Although the impact of these transposons and elements on human genome evolution is recognized, the significance of subtle variations in their sequence remains mostly unexplored. Here we report homozygosity mapping of an infantile neurodegenerative disease locus in a genetic isolate. Complete DNA sequencing of the 400-kb linkage locus revealed a point mutation in a primate-specific retrotransposon that was transcribed as part of a unique noncoding RNA, which was expressed in the brain. In vitro knockdown of this RNA increased neuronal apoptosis, consistent with the inappropriate dosage of this RNA in vivo and with the phenotype. Moreover, structural analysis of the sequence revealed a small RNA-like hairpin that was consistent with the putative gain of a functional site when mutated. We show here that a mutation in a unique transposable element-containing RNA is associated with lethal encephalopathy, and we suggest that RNAs that harbor evolutionarily recent repetitive elements may play important roles in human brain development.}, note = {Published online: March 12, 2012}, keywords = {Genetic disease Long noncoding RNA Long interspersed element 1 Medulla oblongata Pediatrics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {A current overview of regulatory RNAs in Staphylococcus aureus}, author = {I Caldelari and P Fechter and E Lioliou and C Chevalier and C Gaspin and P Romby}, editor = {A Marchfelder and W R Hess}, url = {http://link.springer.com/content/pdf/10.1007%2F978-3-7091-0218-3_3}, doi = {10.1007/978-3-7091-0218-3_3}, year = {2012}, date = {2012-01-01}, booktitle = {Regulatory RNAs in prokaryotes}, pages = {51-75}, publisher = {SpringerWienNewYork}, abstract = {Staphylococcus aureus is a common commensal bacterial species that is usually found in the nose and on the skin of 30 % of healthy adults. However, the bacterium is a remarkably versatile pathogen that is one of the main causes of community as well as hospital-acquired infections (Cheung et al., 2004; Novick, 2003). S. aureus is responsible for systemic infections such as sepsis and endocarditis, which can be difficult to treat due to the acquisition of resistance towards numerous antibiotics in clinical use. S. aureus causes a wide spectrum of human diseases in part due to its ability to produce an array of virulence factors, which are mostly encoded by laterally acquired genomic regions, the so-called pathogenicity islands. These factors include surface proteins responsible for the adhesion and invasion of the host, exoproteins required for host immune evasion, and toxins involved in dissemination in host tissues and acquisition of nutrients (Novick, 2003). Redundancies exist to ensure that a productive infection still occurs even though one factor may be lost. In recent decades, many studies have been carried out to understand how S. aureus is able to coordinate the expression of a large panel of virulence factors at the appropriate time in order to facilitate successful infections (Novick and Geisinger, 2008). These works offer the possibility for developing anti-virulence therapies as alternative strategies for affecting the bacteria viability, i. e. by inhibiting the expression of the virulence factors that cause host damage or the interaction between the pathogen and the host (Clatworthy et al., 2007). Inhibiting virulence instead of viability may have little impact on human flora and result in weaker selective pressure for the development of antibiotic resistance. Hence, determining the regulatory networks and the dynamics involved in virulence and in fast adaptive responses are of prime importance to combating S. aureus infections.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {kinITC: a new method for obtaining joint thermodynamic and kinetic data by Isothermal Titration Calorimetry.}, author = {D Y Burnouf and E Ennifar and S Guedich and B Puffer and G Hoffmann and G Bec and F Disdier and M Baltzinger and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22126339}, doi = {10.1021/ja209057d}, isbn = {22126339}, year = {2012}, date = {2012-01-01}, journal = {J Am Chem Soc}, volume = {134}, number = {1}, pages = {559-565}, abstract = {Isothermal Titration Calorimetry is the method of choice for obtaining thermodynamic data on a great variety of systems. Here we show that modern ITC apparatus and new processing methods also allow obtaining a complete kinetic description on more diverse systems than usually thought, ranging from simple ligand binding to complex RNA folding. We illustrate these new features with a simple case (HIV-1 reverse transcriptase/inhibitor interaction), but also with the more complex case of the folding of a riboswitch triggered by Thiamine PyrophosPhate binding. The originality of the new kinITC method appears clearly by its ability of dissecting, both thermodynamically and kinetically, the two components: primary ligand binding and subsequent RNA folding. We are not aware of another single method yielding in a simple way such a deep insight into a composite process. Our study also rationalizes common observations from daily ITC use.}, note = {Publication Date (Web): November 29, 2011}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The role of Vif oligomerization and RNA chaperone activity in HIV-1 replication.}, author = {J Batisse and S X Guerrero and S Bernacchi and D Sleiman and C Gabus and J L Darlix and R Marquet and C Tisné and J C Paillart}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22728817?dopt=Abstract}, doi = {22728817}, year = {2012}, date = {2012-01-01}, journal = {Virus Res}, volume = {169}, number = {2}, pages = {361-376}, abstract = {The viral infectivity factor (Vif) is essential for the productive infection and dissemination of HIV-1 in non-permissive cells that involve most natural HIV-1 target cells. Vif counteracts the packaging of two cellular cytidine deaminases named APOBEC3G (A3G) and A3F by diverse mechanisms including the recruitment of an E3 ubiquitin ligase complex and the proteasomal degradation of A3G/A3F, the inhibition of A3G mRNA translation or by a direct competition mechanism. In addition, Vif appears to be an active partner of the late steps of viral replication by participating in virus assembly and Gag processing, thus regulating the final stage of virion formation notably genomic RNA dimerization and by inhibiting the initiation of reverse transcription. Vif is a small pleiotropic protein with multiple domains, and recent studies highlighted the importance of Vif conformation and flexibility in counteracting A3G and in binding RNA. In this review, we will focus on the oligomerization and RNA chaperone properties of Vif and show that the intrinsic disordered nature of some Vif domains could play an important role in virus assembly and replication. Experimental evidence demonstrating the RNA chaperone activity of Vif will be presented.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Ions in molecular dynamics simulations of RNA systems.}, author = {P Auffinger}, editor = {N Leontis and E Westhof}, url = {http://www.springerlink.com/content/r770wvrn0m200t04}, doi = {10.1007/978-3-642-25740-7_14}, year = {2012}, date = {2012-01-01}, booktitle = {RNA 3D Structure Analysis and Prediction (Nucleic Acids and Molecular Biology)}, volume = {27}, pages = {299-318}, publisher = {Springer, Berlin}, abstract = {Ions and water molecules are intricately associated with biomolecular systems and play important structural and functional roles that are still not well understood. For RNA systems, the functions of these ions are not limited to the neutralization of the charges carried by the polyanionic backbone, since they also bind to very specific locations of the RNA 3D fold. Hence, it is essential to include them with the greatest possible accuracy in 3D structural models and especially in molecular dynamics (MD) simulations. This review aims at describing some of the successes achieved in the modeling of monovalent and divalent ions in RNA systems, as well as to highlight important deficiencies of current force fields and MD techniques that represent important challenges for future development.}, keywords = {Molecular dynamics simulation Crystallography RNA DNA Solvation Hydration Monovalent cation Divalent cation Sodium Potassium Magnesium Na+ K+ Mg2+, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Seleneoprotein biosynthesis}, author = {C Allmang and A Krol}, editor = {J Liu and G Luo and Y Mu}, url = {http://www.springerlink.com/content/q4h42729317k1478/}, doi = {10.1007/978-3-642-22236-8_8}, year = {2012}, date = {2012-01-01}, booktitle = {Selenoproteins and Mimics}, pages = {107-124}, publisher = {Springer}, series = {Advanced Topics and Technology in China}, abstract = {The amino acid selenocysteine (Sec) is the major biological form of the trace element selenium. Sec is co-translationally incorporated in selenoproteins and is found in the active site of those that have already been assigned a function. In eukaryotes, Sec biosynthesis from serine on the selenocysteine transfer RNA (tRNASec) requires four enzymes. The synthesis of selenoproteins follows a remarkable mechanism which involves translational recoding of a UGA codon, normally used as a stop signal, into a Sec codon. A surprisingly high number of molecular partners have been identified in this machinery but their mechanism of action is still largely unknown. In this chapter, we will provide a detailed description of the knowledge of the field.}, keywords = {ERIANI, KROL, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{dehuyser_synthesis_2012, title = {Synthesis of Novel Mannoside Glycolipid Conjugates for Inhibition of HIV-1 Trans-Infection}, author = {L Dehuyser and E Schaeffer and O Chaloin and C G Mueller and R Baati and A Wagner}, year = {2012}, date = {2012-01-01}, journal = {Bioconjug.Chem.}, number = {1520-4812 (Electronic)}, abstract = {Mannose-binding lectins, such as dendritic cell-specific ICAM-3-grabbing non-integrin (DC-SIGN), are expressed at the surface of human dendritic cells (DCs) that capture and transmit human immunodeficiency virus type-1 (HIV-1) to CD4(+) cells. With the goal of reducing viral trans-infection by targeting DC-SIGN, we have designed a new class of mannoside glycolipid conjugates. We report the synthesis of amphiphiles composed of a mannose head, a hydrophilic linker essential for solubility in aqueous media, and a lipid chain of variable length. These conjugates presented unusual properties based on a cooperation between the mannoside head and the lipid chain, which enhanced the affinity and decreased the need for multivalency. With an optimal lipid length, they exhibited strong binding affinity for DC-SIGN (K(d) in the micromolar range) as assessed by surface plasmon resonance. The most active molecules were branched trimannoside conjugates, able to inhibit the interaction of the HIV-1 envelope with DCs, and to drastically reduce trans-infection of HIV-1 mediated by DCs (IC(50s) in the low micromolar range). This new class of compounds may be of potential use for prevention of HIV-1 dissemination, and also of infection by other DC-SIGN-binding human pathogens}, keywords = {Dendritic Cells, HIV-1, Human, immunodeficiency, infection, inhibition, LECTIN, Lectins, lipid, Mannose-Binding Lectins, prevention, Solubility, Surface Plasmon Resonance, synthesis, Team-Mueller, virus}, pubstate = {published}, tppubtype = {article} } @article{hess_rankl_2012, title = {RANKL induces organized lymph node growth by stromal cell proliferation}, author = {E Hess and V Duheron and M Decossas and F Lezot and A Berdal and S Chea and R Golub and M R Bosisio and S L Bridal and Y Choi and H Yagita and C G Mueller}, doi = {10.4049/jimmunol.1101513}, year = {2012}, date = {2012-01-01}, journal = {Journal of Immunology}, volume = {188}, number = {1550-6606 (Electronic)}, pages = {1245--1254}, abstract = {RANK and its ligand RANKL play important roles in the development and regulation of the immune system. We show that mice transgenic for Rank in hair follicles display massive postnatal growth of skin-draining lymph nodes. The proportions of hematopoietic and nonhematopoietic stromal cells and their organization are maintained, with the exception of an increase in B cell follicles. The hematopoietic cells are not activated and respond to immunization by foreign Ag and adjuvant. We demonstrate that soluble RANKL is overproduced from the transgenic hair follicles and that its neutralization normalizes lymph node size, inclusive area, and numbers of B cell follicles. Reticular fibroblastic and vascular stromal cells, important for secondary lymphoid organ formation and organization, express RANK and undergo hyperproliferation, which is abrogated by RANKL neutralization. In addition, they express higher levels of CXCL13 and CCL19 chemokines, as well as MAdCAM-1 and VCAM-1 cell-adhesion molecules. These findings highlight the importance of tissue-derived cues for secondary lymphoid organ homeostasis and identify RANKL as a key molecule for controlling the plasticity of the immune system}, keywords = {Animals, Cell Adhesion, Cell Adhesion Molecules, Cell Proliferation, Chemokine CCL19, Chemokine CXCL13, chemokines, CXCL13, cytology, development, Growth, growth & development, Hair, hair follicle, Homeostasis, Human, Immune System, Immunization, ligand, LYMPH, LYMPH NODE, Lymph Nodes, Mice, mouse, physiology, plasticity, Proliferation, Protein, rank, RANK ligand, Regulation, Secondary, Stromal Cells, Team-Mueller, transgenic, VCAM1}, pubstate = {published}, tppubtype = {article} } @article{romani_targeting_2012, title = {Targeting skin dendritic cells to improve intradermal vaccination}, author = {N Romani and V Flacher and C H Tripp and F Sparber and S Ebner and P Stoitzner}, doi = {10.1007/82_2010_118}, issn = {0070-217X}, year = {2012}, date = {2012-01-01}, journal = {Current Topics in Microbiology and Immunology}, volume = {351}, pages = {113--138}, abstract = {Vaccinations in medicine are typically administered into the muscle beneath the skin or into the subcutaneous fat. As a consequence, the vaccine is immunologically processed by antigen-presenting cells of the skin or the muscle. Recent evidence suggests that the clinically seldom used intradermal route is effective and possibly even superior to the conventional subcutaneous or intramuscular route. Several types of professional antigen-presenting cells inhabit the healthy skin. Epidermal Langerhans cells (CD207/langerin(+)), dermal langerin(neg), and dermal langerin(+) dendritic cells (DC) have been described, the latter subset so far only in mouse skin. In human skin langerin(neg) dermal DC can be further classified based on their reciprocal expression of CD1a and CD14. The relative contributions of these subsets to the generation of immunity or tolerance are still unclear. Yet, specializations of these different populations have become apparent. Langerhans cells in human skin appear to be specialized for induction of cytotoxic T lymphocytes; human CD14(+) dermal DC can promote antibody production by B cells. It is currently attempted to rationally devise and improve vaccines by harnessing such specific properties of skin DC. This could be achieved by specifically targeting functionally diverse skin DC subsets. We discuss here advances in our knowledge on the immunological properties of skin DC and strategies to significantly improve the outcome of vaccinations by applying this knowledge.}, keywords = {Adaptive Immunity, administration & dosage, Analysis, Animals, Antibodies, antibody, Antigen, ANTIGEN PRESENTING CELLS, Antigen-Presenting Cells, Antigens, B CELLS, B-Lymphocytes, Bacterial Infections, Biosynthesis, C-Type, CD, CD14, CD1a, Cell Lineage, cytokine, Cytokines, cytology, Cytotoxic, Dendritic Cells, DERMATOLOGY, DERMIS, Drug Delivery Systems, Expression, Human, Humans, Immunity, Immunology, INDUCTION, Injections, Innate, Intradermal, Langerhans Cells, LECTIN, Lectins, Lymphocyte Activation, Lymphocytes, Mannose-Binding Lectins, methods, Mice, mouse, Muscle, prevention & control, PRODUCTION, Protein, review, Skin, SUBSETS, T-Lymphocytes, Team-Mueller, tolerance, Vaccination, vaccine, Vaccines, Virus Diseases}, pubstate = {published}, tppubtype = {article} } @article{peltier_beet_2012, title = {Beet necrotic yellow vein virus subgenomic RNA3 is a cleavage product leading to stable non-coding RNA required for long-distance movement.}, author = {Claire Peltier and Elodie Klein and Kamal Hleibieh and Massimiliano D'Alonzo and Philippe Hammann and Salah Bouzoubaa and Claudio Ratti and David Gilmer}, doi = {10.1099/vir.0.039685-0}, issn = {1465-2099 0022-1317}, year = {2012}, date = {2012-01-01}, journal = {The Journal of general virology}, volume = {93}, number = {Pt 5}, pages = {1093--1102}, abstract = {Beet necrotic yellow vein virus (BNYVV) is a multipartite RNA virus. BNYVV RNA3 does not accumulate in non-host transgenic Arabidopsis thaliana plants when expressed using a 35S promoter. However, a 3'-derivative species has been detected in transgenic plants and in transient expression assays conducted in Nicotiana benthamiana and Beta macrocarpa. The 3'-derivative species is similar to the previously reported subgenomic RNA3 produced during virus infection. 5' RACE revealed that the truncated forms had identical 5' ends. The 5' termini carried the coremin motif also present on BNYVV RNA5, beet soil-borne mosaic virus RNA3 and 4, and cucumber mosaic virus group 2 RNAs. This RNA3 species lacks a m(7)Gppp at the 5' end of the cleavage products, whether expressed transiently or virally. Mutagenesis revealed the importance of the coremin sequence for both long-distance movement and stabilization of the cleavage product in vivo and in vitro. The isolation of various RNA3 5'-end products suggests the existence of a cleavage between nt 212 and 1234 and subsequent exonucleolytic degradation, leading to the accumulation of a non-coding RNA. When RNA3 was incubated in wheatgerm extracts, truncated forms appeared rapidly and their appearance was protein- and divalent ion-dependent.}, note = {Place: England}, keywords = {*Genome, Arabidopsis/virology, Beta vulgaris/virology, Plant Viruses/genetics/*pathogenicity, PPSE, RNA, RNA Stability, Tobacco/virology, Untranslated/*metabolism, Viral, Viral/genetics/*metabolism}, pubstate = {published}, tppubtype = {article} } @article{mueller_emerging_2012, title = {Emerging Functions of RANKL in Lymphoid Tissues}, author = {Christopher G Mueller and Estelle Hess}, doi = {10.3389/fimmu.2012.00261}, issn = {1664-3224}, year = {2012}, date = {2012-01-01}, journal = {Frontiers in Immunology}, volume = {3}, pages = {261}, abstract = {The tumor necrosis factor superfamily (TNFSF) members play pivotal roles in embryonic development of lymphoid tissue and their homeostasis. RANKL (Receptor activator of NF-κB ligand, also called TRANCE, TNFSF11) is recognized as an important player in bone homeostasis and lymphoid tissue formation. In its absence bone mass control is deregulated and lymph nodes fail to develop. While its function in bone is well described, there is still little functional insight into the action of RANKL in lymphoid tissue development and homeostasis. Here we provide an overview of the known functions of RANKL, its signaling receptor RANK and its decoy receptor OPG from the perspective of lymphoid tissue development and immune activation in the mouse. Expressed by the hematopoietic lymphoid tissue inducing (LTi) cells and the mesenchymal lymphoid tissue organizer (LTo) cells, RANKL was shown to stimulate Lymphotoxin (LT) expression and to be implicated in LTi cell accumulation. Our recent finding that RANKL also triggers proliferation of adult lymph node stroma suggests that RANKL may furthermore directly activate LTo cells. Beyond bone, the RANKL-RANK-OPG triad plays important roles in immunobiology that are waiting to be unraveled.}, keywords = {LTi, LTo, LYMPH NODE, lymphoid organs, OPG, stroma, Team-Mueller, TNFSF11, TRANCE}, pubstate = {published}, tppubtype = {article} } @article{marquet2012, title = {A transcription factor acts as a HIV-1 accomplice to destroy the cellular defences.}, author = {R Marquet and S Guerrero and S Bernacchi and S Pernot and J Batisse and J C Paillart}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22549857?dopt=AbstractPlus}, doi = {10.1051/medsci/2012284007}, year = {2012}, date = {2012-01-01}, journal = {Med Sci (Paris)}, volume = {28}, number = {4}, pages = {356-358}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @incollection{, title = {Probing RNA Structure In Vitro with Enzymes and Chemicals.}, author = {A C Helfer and C Romilly and C Chevalier and E Lioliou and S Marzi and P Romby}, editor = {R K Hartmann and A Bindereif and A Schön and E Westhof}, url = {http://onlinelibrary.wiley.com/doi/10.1002/9783527647064.ch10/summary}, doi = {10.1002/9783527647064.ch10}, isbn = {9783527327645 9783527647064/ISSN}, year = {2012}, date = {2012-01-01}, booktitle = {Handbook of RNA Biochemistry: Second, Completely Revised and Enlarged Edition}, pages = {205-230}, publisher = {Wiley-VCH}, abstract = {Introduction Enzymatic and Chemical Probes In Vivo DMS Modification Commentary Troubleshooting References}, keywords = {RNA structure probing in vitro in vivo chemical probes enzymatic probing ribonucleases RNases T1, ROMBY, T2, Unité ARN, V1 lead(II)-induced cleavages dimethylsulfate (DMS) diethylpyrocarbonate (DEPC) 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide metho-p-toluenesulfonate (CMCT) N-ethyl-N-nitrosourea (ENU) N-methylisatoic anhydride (NMIA) 1-methyl-7-nitroisatoic anhydride (1M7) benzoyl cyanide (BzCN)}, pubstate = {published}, tppubtype = {incollection} } @article{serag_functional_2011, title = {Functional platform for controlled subcellular distribution of carbon nanotubes}, author = {Maged F Serag and Noritada Kaji and Enrica Venturelli and Yukihiro Okamoto and Kazuyoshi Terasaka and Manabu Tokeshi and Hajime Mizukami and Kevin Braeckmans and Alberto Bianco and Yoshinobu Baba}, doi = {10.1021/nn2035654}, issn = {1936-086X}, year = {2011}, date = {2011-11-01}, journal = {ACS nano}, volume = {5}, number = {11}, pages = {9264--9270}, abstract = {As nanoparticles can cross different cellular barriers and access different tissues, control of their uptake and cellular fate presents a functional approach that will be broadly applicable to nanoscale technologies in cell biology. Here we show that the trafficking of single-walled carbon nanotubes (SWCNTs) through various subcellular membranes of the plant cell is facilitated or inhibited by attaching a suitable functional tag and controlling medium components. This enables a unique control over the uptake and the subcellular distribution of SWCNTs and provides a key strategy to promote their cellular elimination to minimize toxicity. Our results also demonstrate that SWCNTs are involved in a carrier-mediated transport (CMT) inside cells; this is a phenomenon that scientists could use to obtain novel molecular insights into CMT, with the potential translation to advances in subcellular nanobiology.}, keywords = {Biological Transport, carbon, Catharanthus, Exocytosis, Fluorescence Recovery After Photobleaching, Fluorescent Dyes, I2CT, Intracellular Space, Nanotubes, Surface Properties, Team-Bianco, Vacuoles}, pubstate = {published}, tppubtype = {article} } @article{ali-boucetta_cellular_2011, title = {Cellular uptake and cytotoxic impact of chemically functionalized and polymer-coated carbon nanotubes}, author = {Hanene Ali-Boucetta and Khuloud T Al-Jamal and Karin H Müller and Shouping Li and Alexandra E Porter and Ayad Eddaoudi and Maurizio Prato and Alberto Bianco and Kostas Kostarelos}, doi = {10.1002/smll.201101004}, issn = {1613-6829}, year = {2011}, date = {2011-11-01}, journal = {Small (Weinheim an Der Bergstrasse, Germany)}, volume = {7}, number = {22}, pages = {3230--3238}, abstract = {The impact of nanomaterials such as carbon nanotubes on biological matter is a topic of increasing interest and concern and requires a multifaceted approach to be resolved. A modified cytotoxic (lactate dehydrogenase (LDH)) assay is developed in an attempt to offer a valid and reliable methodology for screening carbon nanotube toxicity in vitro. Two of the most widely used types of surface-modified multiwalled carbon nanotubes (MWNTs) are tested: ammonium-functionalized MWNTs (MWNT-NH3+ ) and Pluronic F127 coated MWNTs (MWNT:F127). Chemically functionalized MWNTs show significantly greater cellular uptake into lung epithelial A549 cells compared to the non-covalently Pluronic F127-coated MWNTs. In spite of this, MWNT:F127 exhibit enhanced cytotoxicity according to the modified LDH assay. The validity of the modified LDH assay is further validated by direct comparison with other less reliable or accurate cytotoxicity assays. These findings indicate the reliability of the modified LDH assay as a screening tool to assess carbon nanotube cytotoxicity and illustrate that high levels of carbon nanotube cellular internalization do not necessarily lead to adverse responses.}, keywords = {Annexin A5, carbon, Cell Death, Cell Line, Endocytosis, Flow Cytometry, Fluorescein-5-isothiocyanate, Humans, I2CT, L-Lactate Dehydrogenase, mitochondria, Nanotubes, Polymers, Propidium, Surface Properties, Team-Bianco, tumor, water}, pubstate = {published}, tppubtype = {article} } @article{imler_[innate_2011b, title = {[Innate immunity crowned 2011 Nobel Prize winner]}, author = {Jean-Luc Imler and Dominique Ferrandon}, url = {http://dx.doi.org.gate1.inist.fr/10.1051/medsci/20112711020}, doi = {10.1051/medsci/20112711020}, issn = {0767-0974 (Print) 0767-0974 (Linking)}, year = {2011}, date = {2011-11-01}, journal = {Med Sci (Paris)}, volume = {27}, pages = {1019--24}, keywords = {*Immunity, *Nobel Prize, Biological, ferrandon, Genetic Association Studies, Humans, imler, Immunotherapy/methods/trends, Innate/genetics, M3i, Models, Molecular Targeted Therapy/trends, Seasons, Structure-Activity Relationship, Toll-Like Receptors/chemistry/genetics/isolation &amp;amp; purification/physiology}, pubstate = {published}, tppubtype = {article} } @article{montellano_fullerene_2011, title = {Fullerene C₆₀ as a multifunctional system for drug and gene delivery}, author = {Alejandro Montellano and Tatiana Da Ros and Alberto Bianco and Maurizio Prato}, doi = {10.1039/c1nr10783f}, issn = {2040-3372}, year = {2011}, date = {2011-10-01}, journal = {Nanoscale}, volume = {3}, number = {10}, pages = {4035--4041}, abstract = {The fullerene family, and especially C(60), has delighted the scientific community during the last 25 years with perspective applications in a wide variety of fields, including the biological and the biomedical domains. Several biomedical uses have been explored using water-soluble C(60)-derivatives. However, the employment of fullerenes for drug delivery is still at an early stage of development. The design and synthesis of multifunctionalized and multimodal C(60) systems able to cross the cell membranes and efficiently deliver active molecules is an attracting challenge that involves multidisciplinary strategies. Promising results have emerged in the last years, bringing fullerenes again to the front of interest. Herein, the state of the art of this emerging field is presented and illustrated with some of the most representative examples.}, keywords = {DNA, Drug Carriers, Fullerenes, Gene Transfer Techniques, I2CT, Immunoconjugates, Plasmids, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bianco_making_2011, title = {Making carbon nanotubes biocompatible and biodegradable}, author = {Alberto Bianco and Kostas Kostarelos and Maurizio Prato}, doi = {10.1039/c1cc13011k}, issn = {1364-548X}, year = {2011}, date = {2011-10-01}, journal = {Chemical Communications (Cambridge, England)}, volume = {47}, number = {37}, pages = {10182--10188}, abstract = {Carbon nanotubes are promising nanomaterials with great potential in the field of nanomedicine for both therapeutic and diagnostic applications. Different approaches have been developed to render this material biocompatible and to modulate any ensuing toxic effects. In the context of medical use, although chemically functionalised carbon nanotubes display reduced toxicity, they are still considered with scepticism due to their perceived non-biodegradability. Recently, it has been demonstrated that functionalised carbon nanotubes can be degraded by oxidative enzymes. This finding is offering a new perspective for the development of carbon nanotubes in medicine. This article highlights recent advances that can act as paradigm-shifts towards the design of biocompatible and biodegradable functionalised carbon nanotubes and allow their translation into the clinic.}, keywords = {Animals, Biocompatible, carbon, Coated Materials, HeLa Cells, Humans, I2CT, nanotechnology, Nanotubes, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{boyer_pathogen-derived_2011, title = {Pathogen-derived effectors trigger protective immunity via activation of the Rac2 enzyme and the IMD or Rip kinase signaling pathway}, author = {Laurent Boyer and Lorin Magoc and Stephanie Dejardin and Michael Cappillino and Nicholas Paquette and Charlotte Hinault and Guillaume M Charriere and Eddie W K Ip and Shannon Fracchia and Elizabeth Hennessy and Deniz Erturk-Hasdemir and Jean-Marc Reichhart and Neal Silverman and Adam Lacy-Hulbert and Lynda M Stuart}, doi = {10.1016/j.immuni.2011.08.015}, issn = {1097-4180}, year = {2011}, date = {2011-10-01}, journal = {Immunity}, volume = {35}, number = {4}, pages = {536--549}, abstract = {Although infections with virulent pathogens often induce a strong inflammatory reaction, what drives the increased immune response to pathogens compared to nonpathogenic microbes is poorly understood. One possibility is that the immune system senses the level of threat from a microorganism and augments the response accordingly. Here, focusing on cytotoxic necrotizing factor 1 (CNF1), an Escherichia coli-derived effector molecule, we showed the host indirectly sensed the pathogen by monitoring for the effector that modified RhoGTPases. CNF1 modified Rac2, which then interacted with the innate immune adaptors IMD and Rip1-Rip2 in flies and mammalian cells, respectively, to drive an immune response. This response was protective and increased the ability of the host to restrict pathogen growth, thus defining a mechanism of effector-triggered immunity that contributes to how metazoans defend against microbes with pathogenic potential.}, keywords = {Adaptor Proteins, Enzyme Activation, HEK293 Cells, Humans, M3i, rac GTP-Binding Proteins, Receptor-Interacting Protein Serine-Threonine Kinases, reichhart, Signal Transducing, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{limmer_pseudomonas_2011b, title = {Pseudomonas aeruginosa RhlR is required to neutralize the cellular immune response in a Drosophila melanogaster oral infection model}, author = {Stefanie Limmer and Samantha Haller and Eliana Drenkard and Janice Lee and Shen Yu and Christine Kocks and Frederick M Ausubel and Dominique Ferrandon}, doi = {10.1073/pnas.1114907108}, issn = {1091-6490}, year = {2011}, date = {2011-10-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {108}, number = {42}, pages = {17378--17383}, abstract = {An in-depth mechanistic understanding of microbial infection necessitates a molecular dissection of host-pathogen relationships. Both Drosophila melanogaster and Pseudomonas aeruginosa have been intensively studied. Here, we analyze the infection of D. melanogaster by P. aeruginosa by using mutants in both host and pathogen. We show that orally ingested P. aeruginosa crosses the intestinal barrier and then proliferates in the hemolymph, thereby causing the infected flies to die of bacteremia. Host defenses against ingested P. aeruginosa included an immune deficiency (IMD) response in the intestinal epithelium, systemic Toll and IMD pathway responses, and a cellular immune response controlling bacteria in the hemocoel. Although the observed cellular and intestinal immune responses appeared to act throughout the course of the infection, there was a late onset of the systemic IMD and Toll responses. In this oral infection model, P. aeruginosa PA14 did not require its type III secretion system or other well-studied virulence factors such as the two-component response regulator GacA or the protease AprA for virulence. In contrast, the quorum-sensing transcription factor RhlR, but surprisingly not LasR, played a key role in counteracting the cellular immune response against PA14, possibly at an early stage when only a few bacteria are present in the hemocoel. These results illustrate the power of studying infection from the dual perspective of host and pathogen by revealing that RhlR plays a more complex role during pathogenesis than previously appreciated.}, keywords = {Animal, Animals, Bacteremia, Bacterial Proteins, Cellular, Disease Models, ferrandon, Genes, Genetically Modified, Hemolymph, Host-Pathogen Interactions, Immunity, Insect, M3i, Mutation, Oral, Pseudomonas aeruginosa, Pseudomonas Infections, Quorum Sensing, Trans-Activators, Viral, Virulence}, pubstate = {published}, tppubtype = {article} } @article{Chtarbanova2011, title = {Immunité innée antivirale chez la drosophile}, author = {Stanislava Chtarbanova and Jean-Luc Imler}, url = {http://www.jle.com/fr/revues/vir/e-docs/immunite_innee_antivirale_chez_la_drosophile_290366/article.phtml?tab=texte}, doi = {10.1684/vir.2011.0417}, issn = {1267-8694}, year = {2011}, date = {2011-10-01}, journal = {Virologie}, volume = {15}, number = {5}, pages = {296-306}, abstract = {La drosophile est utilisée depuis une quinzaine d’années comme modèle pour l’étude des mécanismes de l’immunité innée contre les infections bactériennes et fongiques. Les mécanismes de défense de cet insecte contre les infections virales sont maintenant abordés. L’interférence ARN joue un rôle essentiel dans la détection et le contrôle des virus. Ce mécanisme implique la reconnaissance des ARN double brins viraux par Dicer-2 et leur coupure pour former des petits ARN interférants (ARNsi) de 21 nucléotides. Ces ARNsi sont ensuite chargés sur l’enzyme Argonaute-2, qui inhibera spécifiquement les ARN viraux contenant une séquence complémentaire. Une réponse immunitaire inductible contribue également au contrôle des infections virales chez la drosophile. Cette réponse implique les voies de signalisation Toll et IMD, d’une part, (régulant des facteurs de transcription de la famille NF-κB) et JAK/STAT, d’autre part. On ne sait encore que peu de choses sur la nature des molécules effectrices régulées par ces voies de signalisation et sur les récepteurs qui les activent. Même si de nombreux points d’interrogation demeurent, l’intérêt du modèle drosophile pour identifier des mécanismes de défense antivirale conservés au cours de l’évolution, et impliqués dans la transmission des arbovirus par les moustiques, est aujourd’hui bien établi.}, keywords = {*RNA Interference, antiviral immunity, Argonaute, dicer, helicase, imler, M3i, viruses}, pubstate = {published}, tppubtype = {article} } @article{singh_formation_2011, title = {Formation of efficient catalytic silver nanoparticles on carbon nanotubes by adenine functionalization}, author = {Prabhpreet Singh and Giuseppe Lamanna and Cécilia Ménard-Moyon and Francesca Maria Toma and Elena Magnano and Federica Bondino and Maurizio Prato and Sandeep Verma and Alberto Bianco}, doi = {10.1002/anie.201102976}, issn = {1521-3773}, year = {2011}, date = {2011-10-01}, journal = {Angewandte Chemie (International Ed. in English)}, volume = {50}, number = {42}, pages = {9893--9897}, abstract = {Stuck together: adenine/carbon nanotube hybrids trigger the formation of controlled-size catalytic silver nanoparticles on the nanotube surface. The catalytic efficiency of the resulting species was assessed in the oxidation of 2-methylhydroquinone to its corresponding benzoquinone, with complete recovery and without loss of activity of the catalyst.}, keywords = {Adenine, carbon, Catalysis, I2CT, Metal Nanoparticles, Molecular Structure, Nanotubes, Silver, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{hamrita_elongation_2011, title = {An elongation factor-like protein (EF-Tu) elicits a humoral response in infiltrating ductal breast carcinomas: an immunoproteomics investigation.}, author = {Bechr Hamrita and Hela Ben Nasr and Philippe Hammann and Lauriane Kuhn and Christelle-Lemaitre Guillier and Anouar Chaieb and Hedi Khairi and Karim Chahed}, doi = {10.1016/j.clinbiochem.2011.06.005}, issn = {1873-2933 0009-9120}, year = {2011}, date = {2011-09-01}, journal = {Clinical biochemistry}, volume = {44}, number = {13}, pages = {1097--1104}, abstract = {OBJECTIVES: In the current study, we have used an immunoproteomics approach to identify proteins that commonly elicit a humoral response in patients with infiltrating ductal carcinomas of the breast. DESIGN AND METHODS: Sera obtained at the time of diagnosis from 40 patients with invasive breast cancer and 42 healthy controls were screened for the presence of IgG antibodies to MCF-7 cell line proteins using a serological proteomics-based approach. RESULTS: An immunoreactive protein detected in sera from 21 of 40 patients was isolated and subsequently identified as elongation factor-Tu. CONCLUSIONS: The immunoproteomic approach implemented here offers a powerful tool for determining novel tumor antigens that induce a humoral immune response in cancer patients. From our findings, the immunoreactive EF-Tu protein and/or the related circulating antibodies may display clinical usefulness as potential diagnostic markers and provide a means for a better understanding of the molecular mechanisms underlying breast cancer development.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{venturelli_antibody_2011, title = {Antibody covalent immobilization on carbon nanotubes and assessment of antigen binding}, author = {Enrica Venturelli and Chiara Fabbro and Olivier Chaloin and Cécilia Ménard-Moyon and Cristian R Smulski and Tatiana Da Ros and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, doi = {10.1002/smll.201100137}, issn = {1613-6829}, year = {2011}, date = {2011-08-01}, journal = {Small (Weinheim an Der Bergstrasse, Germany)}, volume = {7}, number = {15}, pages = {2179--2187}, abstract = {Controlling the covalent bonding of antibodies onto functionalized carbon nanotubes is a key step in the design and preparation of nanotube-based conjugates for targeting cancer cells. For this purpose, an anti-MUC1 antibody (Ab) is linked to both multi-walled (MWCNTs) and double-walled carbon nanotubes (DWCNTs) using different synthetic strategies. The presence of the Ab attached to the nanotubes is confirmed by gel electrophoresis and thermogravimetric analysis. Most importantly, molecular recognition of the antigen by surface plasmon resonance is able to determine similar Ab binding capacities for both Ab-DWCNTs and Ab-MWCNTs. These results are very relevant for the design of future receptor-targeting strategies using chemically functionalized carbon nanotubes.}, keywords = {Antibodies, Antigens, carbon, I2CT, Immobilized, Mucin-1, nanotechnology, Nanotubes, Protein Binding, Team-Bianco, Thermogravimetry}, pubstate = {published}, tppubtype = {article} } @article{chtarbanova_microbial_2011, title = {Microbial sensing by Toll receptors: a historical perspective}, author = {Stanislava Chtarbanova and Jean-Luc Imler}, doi = {10.1161/ATVBAHA.108.179523}, issn = {1524-4636}, year = {2011}, date = {2011-08-01}, journal = {Arteriosclerosis, Thrombosis, and Vascular Biology}, volume = {31}, number = {8}, pages = {1734--1738}, abstract = {The family of Toll-like receptors plays an essential role in the induction of the immune response. These receptors sense the presence of microbial ligands and activate the nuclear factor-κB transcription factor. We review the key studies that led from the formulation of the concept of pattern recognition receptors to the characterization of Toll-like receptors, insisting on the important role played by the model organism Drosophila melanogaster and on the increasing evidence connecting these receptors to cardiovascular disease.}, keywords = {Animals, Cardiovascular Diseases, history, Host-Pathogen Interactions, Humans, imler, Immunity, Innate, M3i, Macrophages, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{guillaumond_chromatin_2011, title = {Chromatin remodeling as a mechanism for circadian prolactin transcription: rhythmic NONO and SFPQ recruitment to HLTF.}, author = {Fabienne Guillaumond and Benedicte Boyer and Denis Becquet and Severine Guillen and Lauriane Kuhn and Jerome Garin and Maya Belghazi and Olivier Bosler and Jean-Louis Franc and Anne-Marie François-Bellan}, doi = {10.1096/fj.10-178616}, issn = {1530-6860 0892-6638}, year = {2011}, date = {2011-08-01}, journal = {FASEB journal : official publication of the Federation of American Societies for Experimental Biology}, volume = {25}, number = {8}, pages = {2740--2756}, abstract = {Most clock-controlled genes (CCGs) lack the specific E-box response element necessary for direct circadian regulation. This is the case for the prolactin (Prl) gene, the expression of which oscillates in individual lactotrope pituitary cells. To characterize the processes underlying this oscillation, we used a lactotrope cell line (GH4C1 cells). In these cells, Prl gene expression fluctuated significantly during 24 h (P=0.0418). Circadian Prl transcription depended on an interaction between the pituitary-specific transcription factor, PIT-1, and the helicase-like transcription factor (HLTF), a SWI/SNF chromatin remodeler, shown here to bind the Prl promoter on an E-box that differs from the specific E-box preferentially bound by clock proteins. Circadian Prl transcription was further accompanied by marked daily chromatin transitions. While neither HLTF nor PIT-1 was rhythmically expressed, NONO and SFPQ, identified as HLTF-associated proteins by mass spectrometry, displayed a circadian pattern and bound rhythmically to the Prl promoter. Furthermore, NONO and SFPQ were functionally involved in circadian Prl transcription since overexpression of both proteins greatly reduced Prl promoter activity (Ptextbackslashtextless0.001) and disrupted its circadian pattern. A mechanism involving a rhythm in paraspeckle protein recruitment is proposed to explain how the core oscillator can generate a circadian pattern of CCGs lacking the specific E-box response element.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{lamanna_hydramers_2011, title = {HYDRAmers: design, synthesis and characterization of different generation novel Hydra-like dendrons based on multifunctionalized adamantane}, author = {Giuseppe Lamanna and Julie Russier and Cécilia Ménard-Moyon and Alberto Bianco}, doi = {10.1039/c1cc11689d}, issn = {1364-548X}, year = {2011}, date = {2011-08-01}, journal = {Chemical Communications (Cambridge, England)}, volume = {47}, number = {31}, pages = {8955--8957}, abstract = {In this communication we present a new synthetic strategy to different generation Hydra-like dendrons based on tetrafunctionalized adamantane as a building block. The novel dendrons, which we termed HYDRAmers, possess at the periphery and at the central core orthogonal protections that can be exploited for conjugation of targeting ligands, drugs and/or imaging probes.}, keywords = {Adamantane, Animals, Cell Line, Dendrimers, Drug Design, Humans, I2CT, L-Lactate Dehydrogenase, Magnetic Resonance Spectroscopy, Mice, Team-Bianco, tumor}, pubstate = {published}, tppubtype = {article} } @article{E2011, title = {The multifaceted mosquito anti-Plasmodium response}, author = {Eric Marois}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21802348}, year = {2011}, date = {2011-07-27}, journal = {Curr Opin Microbiol.}, volume = {14}, number = {4}, pages = {429-35}, abstract = {Plasmodium development within its mosquito vector is an essential step in malaria transmission, as illustrated in world regions where malaria was successfully eradicated via vector control. The innate immune system of most mosquitoes is able to completely clear a Plasmodium infection, preventing parasite transmission to humans. Understanding the biological basis of this phenomenon is expected to inspire new strategies to curb malaria incidence in countries where vector control via insecticides is unpractical, or inefficient because insecticide resistance genes have spread across mosquito populations. Several aspects of mosquito biology that condition the success of the parasite in colonizing its vector begin to be understood at the molecular level, and a wealth of recently published data highlights the multifaceted nature of the mosquito response against parasite invasion. In this brief review, we attempt to provide an integrated view of the challenges faced by the parasite to successfully invade its mosquito host, and discuss the possible intervention strategies that could exploit this knowledge for the fight against human malaria.}, keywords = {anti-Plasmodium response, M3i, marois}, pubstate = {published}, tppubtype = {article} } @article{pmid21762475, title = {Human-Phosphate-Binding-Protein inhibits HIV-1 gene transcription and replication}, author = {Thomas Cherrier and Mikael Elias and Alicia Jeudy and Guillaume Gotthard and Valentin Le Douce and Houda Hallay and Patrick Masson and Andrea Janossy and Ermanno Candolfi and Olivier Rohr and Eric Chabrière and Christian Schwartz}, doi = {10.1186/1743-422X-8-352}, issn = {1743-422X}, year = {2011}, date = {2011-07-01}, urldate = {2011-07-01}, journal = {Virol J}, volume = {8}, pages = {352}, abstract = {The Human Phosphate-Binding protein (HPBP) is a serendipitously discovered lipoprotein that binds phosphate with high affinity. HPBP belongs to the DING protein family, involved in various biological processes like cell cycle regulation. We report that HPBP inhibits HIV-1 gene transcription and replication in T cell line, primary peripherical blood lymphocytes and primary macrophages. We show that HPBP is efficient in naïve and HIV-1 AZT-resistant strains. Our results revealed HPBP as a new and potent anti HIV molecule that inhibits transcription of the virus, which has not yet been targeted by HAART and therefore opens new strategies in the treatment of HIV infection.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{al-jamal_functional_2011, title = {Functional motor recovery from brain ischemic insult by carbon nanotube-mediated siRNA silencing}, author = {Khuloud T Al-Jamal and Lisa Gherardini and Giuseppe Bardi and Antonio Nunes and Chang Guo and Cyrill Bussy and Antonia M Herrero and Alberto Bianco and Maurizio Prato and Kostas Kostarelos and Tommaso Pizzorusso}, doi = {10.1073/pnas.1100930108}, issn = {1091-6490}, year = {2011}, date = {2011-07-01}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, number = {27}, pages = {10952--10957}, abstract = {Stroke is the second cause of death worldwide with ischemic stroke accounting for 80% of all stroke insults. Caspase-3 activation contributes to brain tissue loss and downstream biochemical events that lead to programmed cell death after traumatic brain injury. Alleviation of symptoms following ischemic neuronal injury can be potentially achieved by either genetic disruption or pharmacological inhibition of caspases. Here, we studied whether silencing of Caspase-3 using carbon nanotube-mediated in vivo RNA interference (RNAi) could offer a therapeutic opportunity against stroke. Effective delivery of siRNA directly to the CNS has been shown to normalize phenotypes in animal models of several neurological diseases. It is shown here that peri-lesional stereotactic administration of a Caspase-3 siRNA (siCas 3) delivered by functionalized carbon nanotubes (f-CNT) reduced neurodegeneration and promoted functional preservation before and after focal ischemic damage of the rodent motor cortex using an endothelin-1 induced stroke model. These observations illustrate the opportunity offered by carbon nanotube-mediated siRNA delivery and gene silencing of neuronal tissue applicable to a variety of different neuropathological conditions where intervention at well localized brain foci may offer therapeutic and functional benefits.}, keywords = {Animals, Apoptosis, Base Sequence, Brain Ischemia, carbon, Caspase 3, Caspase Inhibitors, Cell Line, Cells, Cultured, Electron, Endothelin-1, Female, Genetic Therapy, I2CT, Inbred C57BL, Mice, Microscopy, Nanomedicine, Nanotubes, Neurons, Psychomotor Performance, Rats, RNA, RNA Interference, Small Interfering, Sprague-Dawley, Team-Bianco, Transmission}, pubstate = {published}, tppubtype = {article} } @article{eleftherianos_atp-sensitive_2011, title = {ATP-sensitive potassium channel (K(ATP))-dependent regulation of cardiotropic viral infections}, author = {Ioannis Eleftherianos and Sungyong Won and Stanislava Chtarbanova and Barbara Squiban and Karen Ocorr and Rolf Bodmer and Bruce Beutler and Jules A Hoffmann and Jean-Luc Imler}, doi = {10.1073/pnas.1108926108}, issn = {1091-6490}, year = {2011}, date = {2011-07-01}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, number = {29}, pages = {12024--12029}, abstract = {The effects of the cellular environment on innate immunity remain poorly characterized. Here, we show that in Drosophila ATP-sensitive potassium channels (K(ATP)) mediate resistance to a cardiotropic RNA virus, Flock House virus (FHV). FHV viral load in the heart rapidly increases in K(ATP) mutant flies, leading to increased viremia and accelerated death. The effect of K(ATP) channels is dependent on the RNA interference genes Dcr-2, AGO2, and r2d2, indicating that an activity associated with this potassium channel participates in this antiviral pathway in Drosophila. Flies treated with the K(ATP) agonist drug pinacidil are protected against FHV infection, thus demonstrating the importance of this regulation of innate immunity by the cellular environment in the heart. In mice, the Coxsackievirus B3 replicates to higher titers in the hearts of mayday mutant animals, which are deficient in the Kir6.1 subunit of K(ATP) channels, than in controls. Together, our data suggest that K(ATP) channel deregulation can have a critical impact on innate antiviral immunity in the heart.}, keywords = {Animals, Heart, HeLa Cells, hoffmann, Humans, imler, Immunity, Immunoblotting, Inbred C57BL, Innate, KATP Channels, M3i, Mice, Nodaviridae, Pinacidil, Reverse Transcriptase Polymerase Chain Reaction, RNA Interference, Tolbutamide, Viral Load, Viremia}, pubstate = {published}, tppubtype = {article} } @article{ciobanu_selection_2011, title = {Selection of a synthetic glycan oligomer from a library of DNA-templated fragments against DC-SIGN and inhibition of HIV gp120 binding to dendritic cells}, author = {M Ciobanu and K T Huang and J P Daguer and S Barluenga and O Chaloin and E Schaeffer and C G Mueller and D A Mitchell and N Winssinger}, year = {2011}, date = {2011-07-01}, journal = {Chem.Commun.(Camb.)}, number = {1364-548X (Electronic)}, abstract = {We report the synthesis of a nucleic acid-encoded carbohydrate library, its combinatorial self-assembly into 37 485 pairs and a screen against DC-SIGN leading to the identification of consensus ligand motifs. A prototypical example from the selected pairs was shown to have enhanced binding. A dendrimer incorporating the selected motifs inhibited gp120's binding to dendritic cells with higher efficiency than mannan}, keywords = {Dendritic Cells, GP120, HIV, IDENTIFICATION, inhibition, ligand, mannan, synthesis, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{V2011, title = {Functional genomics of tick thioester-containing proteins reveal the ancient origin of the complement system}, author = {V Buresova and Ondrej Hajdusek and Z Franta and G Loosova and L Grunclova and Elena A Levashina and P Kopacek}, year = {2011}, date = {2011-06-30}, journal = {J Innate Immun.}, volume = {3}, number = {6}, pages = {623-30}, abstract = {Ticks are important ectoparasites and vectors of multiple human and animal diseases. The obligatory hemophagy of ticks provides a formidable route for parasite transmission from one host to another. Parasite survival inside the tick relies on the ability of a pathogen to escape or inhibit tick immune defenses, but the molecular interactions between the tick and its pathogens remain poorly understood. Here we report that tick genomes are unique in that they contain all known classes of the α(2)-macroglobulin family (α(2)M-F) proteins: α(2)-macroglobulin pan-protease inhibitors, C3 complement components, and insect thioester-containing and macroglobulin-related proteins. By using RNA interference-mediated gene silencing in the hard tick Ixodes ricinus we demonstrated the central role of a C3-like molecule in the phagocytosis of bacteria and revealed nonredundant functions for α(2)M-F proteins. Assessment of α(2)M-F functions in a single organism should significantly contribute to the general knowledge on the evolution and function of the complement system. Importantly, understanding the tick immune mechanisms should provide new concepts for efficient transmission blocking of tick-borne diseases.}, keywords = {TEP1}, pubstate = {published}, tppubtype = {article} } @article{murphy_length-dependent_2011, title = {Length-dependent retention of carbon nanotubes in the pleural space of mice initiates sustained inflammation and progressive fibrosis on the parietal pleura}, author = {Fiona A Murphy and Craig A Poland and Rodger Duffin and Khuloud T Al-Jamal and Hanene Ali-Boucetta and Antonio Nunes and Fiona Byrne and Adriele Prina-Mello and Yuri Volkov and Shouping Li and Stephen J Mather and Alberto Bianco and Maurizio Prato and William Macnee and William A Wallace and Kostas Kostarelos and Ken Donaldson}, doi = {10.1016/j.ajpath.2011.02.040}, issn = {1525-2191}, year = {2011}, date = {2011-06-01}, journal = {The American Journal of Pathology}, volume = {178}, number = {6}, pages = {2587--2600}, abstract = {The fibrous shape of carbon nanotubes (CNTs) raises concern that they may pose an asbestos-like inhalation hazard, leading to the development of diseases, especially mesothelioma. Direct instillation of long and short CNTs into the pleural cavity, the site of mesothelioma development, produced asbestos-like length-dependent responses. The response to long CNTs and long asbestos was characterized by acute inflammation, leading to progressive fibrosis on the parietal pleura, where stomata of strictly defined size limit the egress of long, but not short, fibers. This was confirmed by demonstrating clearance of short, but not long, CNT and nickel nanowires and by visualizing the migration of short CNTs from the pleural space by single-photon emission computed tomographic imaging. Our data confirm the hypothesis that, although a proportion of all deposited particles passes through the pleura, the pathogenicity of long CNTs and other fibers arises as a result of length-dependent retention at the stomata on the parietal pleura.}, keywords = {Animals, carbon, Cell Proliferation, Disease Progression, Emission-Computed, Epithelium, Fibrosis, I2CT, inflammation, Lymph Nodes, Mediastinum, Mice, Nanotubes, Nanowires, Particle Size, Pleura, Pleural Cavity, Single-Photon, Team-Bianco, Time Factors, Tomography, X-Ray Computed}, pubstate = {published}, tppubtype = {article} } @article{al-jamal_cellular_2011, title = {Cellular uptake mechanisms of functionalised multi-walled carbon nanotubes by 3D electron tomography imaging}, author = {Khuloud T Al-Jamal and Hannah Nerl and Karin H Müller and Hanene Ali-Boucetta and Shouping Li and Peter D Haynes and Joerg R Jinschek and Maurizio Prato and Alberto Bianco and Kostas Kostarelos and Alexandra E Porter}, doi = {10.1039/c1nr10080g}, issn = {2040-3372}, year = {2011}, date = {2011-06-01}, journal = {Nanoscale}, volume = {3}, number = {6}, pages = {2627--2635}, abstract = {Carbon nanotubes (CNTs) are being investigated for a variety of biomedical applications. Despite numerous studies, the pathways by which carbon nanotubes enter cells and their subsequent intracellular trafficking and distribution remain poorly determined. Here, we use 3-D electron tomography techniques that offer optimum enhancement of contrast between carbon nanotubes and the plasma membrane to investigate the mechanisms involved in the cellular uptake of shortened, functionalised multi-walled carbon nanotubes (MWNT-NH(3)(+)). Both human lung epithelial (A549) cells, that are almost incapable of phagocytosis and primary macrophages, capable of extremely efficient phagocytosis, were used. We observed that MWNT-NH(3)(+) were internalised in both phagocytic and non-phagocytic cells by any one of three mechanisms: (a) individually via membrane wrapping; (b) individually by direct membrane translocation; and (c) in clusters within vesicular compartments. At early time points following intracellular translocation, we noticed accumulation of nanotube material within various intracellular compartments, while a long-term (14-day) study using primary human macrophages revealed that MWNT-NH(3)(+) were able to escape vesicular (phagosome) entrapment by translocating directly into the cytoplasm.}, keywords = {carbon, Cell Line, Cell Membrane, Cytoplasm, Electron Microscope Tomography, Humans, I2CT, imaging, Macrophages, Nanotubes, Phagocytosis, Phagosomes, Team-Bianco, Three-Dimensional, tumor}, pubstate = {published}, tppubtype = {article} } @article{singh_carbon_2011, title = {Carbon nanotube-nucleobase hybrids: nanorings from uracil-modified single-walled carbon nanotubes}, author = {Prabhpreet Singh and Francesca Maria Toma and Jitendra Kumar and V Venkatesh and Jesus Raya and Maurizio Prato and Sandeep Verma and Alberto Bianco}, doi = {10.1002/chem.201100312}, issn = {1521-3765}, year = {2011}, date = {2011-06-01}, journal = {Chemistry (Weinheim an Der Bergstrasse, Germany)}, volume = {17}, number = {24}, pages = {6772--6780}, abstract = {Single-walled carbon nanotubes (SWCNTs) have been covalently functionalized with uracil nucleobase. The hybrids have been characterized by using complementary spectroscopic and microscopic techniques including solid-state NMR spectroscopy. The uracil-functionalized SWCNTs are able to self-assemble into regular nanorings with a diameter of 50-70 nm, as observed by AFM and TEM. AFM shows that the rings do not have a consistent height and thickness, which indicates that they may be formed by separate bundles of CNTs. The simplest model for the nanoring formation likely involves two bundles of CNTs interacting with each other via uracil-uracil base-pairing at both CNT ends. These nanorings can be envisaged for the development of advanced electronic circuits.}, keywords = {carbon, I2CT, Magnetic Resonance Spectroscopy, Molecular Structure, Nanotubes, Team-Bianco, Uracil}, pubstate = {published}, tppubtype = {article} } @article{limmer_virulence_2011b, title = {Virulence on the fly: Drosophila melanogaster as a model genetic organism to decipher host-pathogen interactions}, author = {Stefanie Limmer and Jessica Quintin and Charles Hetru and Dominique Ferrandon}, issn = {1873-5592}, year = {2011}, date = {2011-06-01}, journal = {Curr Drug Targets}, volume = {12}, number = {7}, pages = {978--999}, abstract = {To gain an in-depth grasp of infectious processes one has to know the specific interactions between the virulence factors of the pathogen and the host defense mechanisms. A thorough understanding is crucial for identifying potential new drug targets and designing drugs against which the pathogens might not develop resistance easily. Model organisms are a useful tool for this endeavor, thanks to the power of their genetics. Drosophila melanogaster is widely used to study host-pathogen interactions. Its basal immune response is well understood and is briefly reviewed here. Considerations relevant to choosing an adequate infection model are discussed. This review then focuses mainly on infections with two categories of pathogens, the well-studied Gram-negative bacterium Pseudomonas aeruginosa and infections by fungi of medical interest. These examples provide an overview over the current knowledge on Drosophila-pathogen interactions and illustrate the approaches that can be used to study those interactions. We also discuss the usefulness and limits of Drosophila infection models for studying specific host-pathogen interactions and high-throughput drug screening.}, keywords = {Animal, Animals, Anti-Infective Agents, Disease Models, Drug Delivery Systems, Drug Design, Drug Resistance, ferrandon, Fungi, High-Throughput Screening Assays, Host-Pathogen Interactions, Humans, M3i, Microbial, Pseudomonas aeruginosa}, pubstate = {published}, tppubtype = {article} } @article{ogawa_tecpr1-dependent_2011, title = {A Tecpr1-dependent selective autophagy pathway targets bacterial pathogens}, author = {Michinaga Ogawa and Yuko Yoshikawa and Taira Kobayashi and Hitomi Mimuro and Makoto Fukumatsu and Kotaro Kiga and Zhenzi Piao and Hiroshi Ashida and Mitsutaka Yoshida and Shigeru Kakuta and Tomohiro Koyama and Yoshiyuki Goto and Takahiro Nagatake and Shinya Nagai and Hiroshi Kiyono and Magdalena Kawalec and Jean-Marc Reichhart and Chihiro Sasakawa}, doi = {10.1016/j.chom.2011.04.010}, issn = {1934-6069}, year = {2011}, date = {2011-05-01}, journal = {Cell Host Microbe}, volume = {9}, number = {5}, pages = {376--389}, abstract = {Selective autophagy of bacterial pathogens represents a host innate immune mechanism. Selective autophagy has been characterized on the basis of distinct cargo receptors but the mechanisms by which different cargo receptors are targeted for autophagic degradation remain unclear. In this study we identified a highly conserved Tectonin domain-containing protein, Tecpr1, as an Atg5 binding partner that colocalized with Atg5 at Shigella-containing phagophores. Tecpr1 activity is necessary for efficient autophagic targeting of bacteria, but has no effect on rapamycin- or starvation-induced canonical autophagy. Tecpr1 interacts with WIPI-2, a yeast Atg18 homolog and PI(3)P-interacting protein required for phagophore formation, and they colocalize to phagophores. Although Tecpr1-deficient mice appear normal, Tecpr1-deficient MEFs were defective for selective autophagy and supported increased intracellular multiplication of Shigella. Further, depolarized mitochondria and misfolded protein aggregates accumulated in the Tecpr1-knockout MEFs. Thus, we identify a Tecpr1-dependent pathway as important in targeting bacterial pathogens for selective autophagy.}, keywords = {Animals, Autophagy, Biological, Cells, Cultured, M3i, Membrane Proteins, Mice, Microtubule-Associated Proteins, Models, Phagosomes, Protein Interaction Mapping, reichhart, Shigella, Two-Hybrid System Techniques}, pubstate = {published}, tppubtype = {article} } @article{kellenberger_structure-function_2011, title = {Structure-function analysis of grass clip serine protease involved in Drosophila Toll pathway activation}, author = {Christine Kellenberger and Philippe Leone and Laurent Coquet and Thierry Jouenne and Jean-Marc Reichhart and Alain Roussel}, doi = {10.1074/jbc.M110.182741}, issn = {1083-351X}, year = {2011}, date = {2011-04-01}, journal = {J. Biol. Chem.}, volume = {286}, number = {14}, pages = {12300--12307}, abstract = {Grass is a clip domain serine protease (SP) involved in a proteolytic cascade triggering the Toll pathway activation of Drosophila during an immune response. Epistasic studies position it downstream of the apical protease ModSP and upstream of the terminal protease Spaetzle-processing enzyme. Here, we report the crystal structure of Grass zymogen. We found that Grass displays a rather deep active site cleft comparable with that of proteases of coagulation and complement cascades. A key distinctive feature is the presence of an additional loop (75-loop) in the proximity of the activation site localized on a protruding loop. All biochemical attempts to hydrolyze the activation site of Grass failed, strongly suggesting restricted access to this region. The 75-loop is thus proposed to constitute an original mechanism to prevent spontaneous activation. A comparison of Grass with clip serine proteases of known function involved in analogous proteolytic cascades allowed us to define two groups, according to the presence of the 75-loop and the conformation of the clip domain. One group (devoid of the 75-loop) contains penultimate proteases whereas the other contains terminal proteases. Using this classification, Grass appears to be a terminal protease. This result is evaluated according to the genetic data documenting Grass function.}, keywords = {Animals, Catalytic Domain, Cell Line, M3i, reichhart, Serine Proteases, Signal Transduction, Structure-Activity Relationship, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{banchet-cadeddu_use_2011, title = {Use of the NEO strategy (Nucleophilic addition/Epoxide Opening) for the synthesis of a new C-galactoside ester analogue of KRN 7000}, author = {Aline Banchet-Cadeddu and Agathe Martinez and Stéphane Guillarme and Véronique Parietti and Fanny Monneaux and Eric Hénon and Jean-Hugues Renault and Jean-Marc Nuzillard and Arnaud Haudrechy}, doi = {10.1016/j.bmcl.2011.02.044}, issn = {1464-3405}, year = {2011}, date = {2011-04-01}, journal = {Bioorganic & Medicinal Chemistry Letters}, volume = {21}, number = {8}, pages = {2510--2514}, abstract = {Our goal in the search for potentially bioactive analogues of KRN 7000 was to design an easy synthetic approach to a library of analogues using a strategy recently developed in our laboratory based on a Nucleophilic addition followed by an Epoxide Opening (the NEO strategy). Through the use of a common pivotal structure, a new C-galactoside ester analogue (23) was synthesized which showed an encouraging T(H)2 biased response during preliminary biological tests.}, keywords = {Animals, Cell Proliferation, Cells, Cultured, Esters, Galactosides, Galactosylceramides, Glycolipids, I2CT, Interferon-gamma, Interleukin-4, Mice, Monneaux, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{russier_oxidative_2011, title = {Oxidative biodegradation of single- and multi-walled carbon nanotubes}, author = {Julie Russier and Cécilia Ménard-Moyon and Enrica Venturelli and Edmond Gravel and Gabriele Marcolongo and Moreno Meneghetti and Eric Doris and Alberto Bianco}, doi = {10.1039/c0nr00779j}, issn = {2040-3372}, year = {2011}, date = {2011-03-01}, journal = {Nanoscale}, volume = {3}, number = {3}, pages = {893--896}, abstract = {In this study we compare the biodegradation of both single-walled (SWCNTs) and multi-walled carbon nanotubes (MWCNTs) using two different oxidative conditions. In particular, we demonstrate that oxidized multi-walled carbon nanotubes are highly degraded, although not to completeness when treated with horseradish peroxidase (HRP) in the presence of hydrogen peroxide.}, keywords = {Absorbable Implants, Biocompatible Materials, Body Fluids, carbon, Horseradish Peroxidase, Hydrogen Peroxide, I2CT, Macromolecular Substances, Materials Testing, Molecular Conformation, Nanotubes, Oxidation-Reduction, Particle Size, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{menard-moyon_one-pot_2011, title = {One-pot triple functionalization of carbon nanotubes}, author = {Cécilia Ménard-Moyon and Chiara Fabbro and Maurizio Prato and Alberto Bianco}, doi = {10.1002/chem.201003050}, issn = {1521-3765}, year = {2011}, date = {2011-03-01}, journal = {Chemistry (Weinheim an Der Bergstrasse, Germany)}, volume = {17}, number = {11}, pages = {3222--3227}, abstract = {Carbon nanotubes (CNTs) are very promising as carriers for the delivery of bioactive molecules. The multifunctionalization of CNTs is necessary to impart multimodalities for the development of future CNT-based multipotent therapeutic constructs. In this context, we report the first example of covalent trifunctionalization of different types of CNTs. Our strategy is a simple and efficient methodology based on the simultaneous functionalization of the nanotube surface with three different active groups. The reaction is performed in one step by arylation with diazonium salts generated in situ. The CNTs are functionalized with benzylamine moieties blocked with three different protecting groups that can be selectively removed under specific conditions. The trifunctionalized CNTs were characterized by TEM, thermogravimetric analysis, and Raman and UV/Vis/NIR spectroscopy, while the amine loading was determined by using the Kaiser test. The sequential removal of the protecting groups of the amine functions allows the grafting of the molecules of interest on the nanotube surface to be controlled.}, keywords = {Aniline Compounds, Azo Compounds, Benzylamines, carbon, I2CT, Nanotubes, Raman, Spectrophotometry, Spectrum Analysis, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{S2011, title = {Intracellular immune responses of dipteran insects}, author = {Stefanie Steinert and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21349091}, year = {2011}, date = {2011-03-01}, journal = {Immunol Rev.}, volume = {240}, number = {1}, pages = {129-40}, abstract = {Vector-borne diseases, transmitted by bloodsucking arthropods, pose worldwide socio-medical and economical problems. Some of the major human infectious diseases, such as malaria, Dengue fever, and yellow fever, are transmitted by mosquitoes. While the majority of pathogens enjoy extracellular life styles in insects, viruses and some endosymbionts are strictly intracellular. Here, we summarize our knowledge on defense reactions against intracellular microorganisms in dipteran insects and discuss the potential of insects as models to study human pathogens.}, keywords = {intracellular immune responses}, pubstate = {published}, tppubtype = {article} } @article{duheron_receptor_2011, title = {Receptor activator of NF-kappaB (RANK) stimulates the proliferation of epithelial cells of the epidermo-pilosebaceous unit}, author = {V Duheron and E Hess and M Duval and M Decossas and B Castaneda and J E Klopper and L Amoasii and J B Barbaroux and I R Williams and H Yagita and J Penninger and Y Choi and F Lezot and R Groves and R Paus and C G Mueller}, doi = {10.1073/pnas.1013054108}, year = {2011}, date = {2011-03-01}, journal = {Proc.Natl.Acad.Sci.U.S.A}, volume = {108}, number = {1091-6490 (Electronic)}, pages = {5342--5347}, abstract = {Receptor activator of NF-kappaB (RANK), known for controlling bone mass, has been recognized for its role in epithelial cell activation of the mammary gland. Because bone and the epidermo-pilosebaceous unit of the skin share a lifelong renewal activity where similar molecular players operate, and because mammary glands and hair follicles are both skin appendages, we have addressed the function of RANK in the hair follicle and the epidermis. Here, we show that mice deficient in RANK ligand (RANKL) are unable to initiate a new growth phase of the hair cycle and display arrested epidermal homeostasis. However, transgenic mice overexpressing RANK in the hair follicle or administration of recombinant RANKL both activate the hair cycle and epidermal growth. RANK is expressed by the hair follicle germ and bulge stem cells and the epidermal basal cells, cell types implicated in the renewal of the epidermo-pilosebaceous unit. RANK signaling is dispensable for the formation of the stem cell compartment and the inductive hair follicle mesenchyme, and the hair cycle can be rescued by Rankl knockout skin transplantation onto nude mice. RANKL is actively transcribed by the hair follicle at initiation of its growth phase, providing a mechanism for stem cell RANK engagement and hair-cycle entry. Thus, RANK-RANKL regulates hair renewal and epidermal homeostasis and provides a link between these two activities}, keywords = {Activation, Animals, Cell Proliferation, Chemistry, cytology, Epidermis, Epithelial Cells, function, Genetics, Growth, Hair, hair follicle, Homeostasis, Immunology, Inbred C57BL, ligand, metabolism, Mice, NF-kappa B, NF-kappaB, Nude, Osteoprotegerin, physiology, Proliferation, rank, RANK ligand, Receptor, Receptor Activator of Nuclear Factor-kappa B, signaling, Skin, Skin Transplantation, stem, Stem Cells, Team-Mueller, transgenic, TRANSGENIC MICE, TRANSPLANTATION}, pubstate = {published}, tppubtype = {article} } @article{gaillard_carbon_2011, title = {Carbon nanotube-coupled cell adhesion peptides are non-immunogenic: a promising step toward new biomedical devices}, author = {Claire Gaillard and Monique Duval and Hélène Dumortier and Alberto Bianco}, doi = {10.1002/psc.1290}, issn = {1099-1387}, year = {2011}, date = {2011-02-01}, journal = {Journal of Peptide Science: An Official Publication of the European Peptide Society}, volume = {17}, number = {2}, pages = {139--142}, abstract = {Carbon nanotubes functionalized with cell adhesion peptides can be considered as novel, promising candidates for the development of advanced drug delivery systems or for designing new generation of self-assembling nerve 'bridges'. An important step toward the integration of these types of conjugates in living bodies is the assessment of their impact on the immune system. In this direction, an integrin-derived peptide has been covalently conjugated to carbon nanotubes. Following intraperitoneal administration, peptide-carbon nanotubes do not trigger an anti-peptide antibody production. Demonstration of the immune neutrality of peptide-carbon nanotubes reinforces their potential use as substrates for neuronal regeneration in vivo.}, keywords = {carbon, Dumortier, Enzyme-Linked Immunosorbent Assay, I2CT, Nanotubes, Peptides, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{pmid22073215, title = {Genome-wide binding map of the HIV-1 Tat protein to the human genome}, author = {Céline Marban and Trent Su and Roberto Ferrari and Bing Li and Dimitrios Vatakis and Matteo Pellegrini and Jerome A Zack and Olivier Rohr and Siavash K Kurdistani}, doi = {10.1371/journal.pone.0026894}, issn = {1932-6203}, year = {2011}, date = {2011-01-01}, urldate = {2011-01-01}, journal = {PLoS One}, volume = {6}, number = {11}, pages = {e26894}, abstract = {The HIV-1 Trans-Activator of Transcription (Tat) protein binds to multiple host cellular factors and greatly enhances the level of transcription of the HIV genome. While Tat's control of viral transcription is well-studied, much less is known about the interaction of Tat with the human genome. Here, we report the genome-wide binding map of Tat to the human genome in Jurkat T cells using chromatin immunoprecipitation combined with next-generation sequencing. Surprisingly, we found that ~53% of the Tat target regions are within DNA repeat elements, greater than half of which are Alu sequences. The remaining target regions are located in introns and distal intergenic regions; only ~7% of Tat-bound regions are near transcription start sites (TSS) at gene promoters. Interestingly, Tat binds to promoters of genes that, in Jurkat cells, are bound by the ETS1 transcription factor, the CBP histone acetyltransferase and/or are enriched for histone H3 lysine 4 tri-methylation (H3K4me3) and H3K27me3. Tat binding is associated with genes enriched with functions in T cell biology and immune response. Our data reveal that Tat's interaction with the host genome is more extensive than previously thought, with potentially important implications for the viral life cycle.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{serag_trafficking_2011, title = {Trafficking and subcellular localization of multiwalled carbon nanotubes in plant cells}, author = {Maged F Serag and Noritada Kaji and Claire Gaillard and Yukihiro Okamoto and Kazuyoshi Terasaka and Mohammad Jabasini and Manabu Tokeshi and Hajime Mizukami and Alberto Bianco and Yoshinobu Baba}, doi = {10.1021/nn102344t}, issn = {1936-086X}, year = {2011}, date = {2011-01-01}, journal = {ACS nano}, volume = {5}, number = {1}, pages = {493--499}, abstract = {Major barriers to delivery of biomolecules are crossing the cellular membranes and achieving a high cytoplasmic concentration by circumventing entrapment into endosomes and other lytic organelles. Motivated by such aim, we have investigated the capability of multiwalled carbon nanotubes (MWCNTs) to penetrate the cell membrane of plant protoplasts (plant cells made devoid of their cell walls via enzymatic treatment) and studied their internalization mechanism via confocal imaging and TEM techniques. Our results indentified an endosome-escaping uptake mode of MWCNTs by plant protoplasts. Moreover, short MWCNTs (textbackslashtextless100 nm) were observed to target specific cellular substructures including the nucleus, plastids, and vacuoles. These findings are expected to have a significant impact on plant cell biology and transformation technologies.}, keywords = {Biological Transport, carbon, Catharanthus, Cell Membrane, Endosomes, I2CT, Intracellular Space, Nanotubes, Protoplasts, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{banchet-cadeddu_stimulating_2011, title = {The stimulating adventure of KRN 7000}, author = {Aline Banchet-Cadeddu and Eric Hénon and Manuel Dauchez and Jean-Hugues Renault and Fanny Monneaux and Arnaud Haudrechy}, doi = {10.1039/c0ob00975j}, issn = {1477-0539}, year = {2011}, date = {2011-01-01}, journal = {Organic & Biomolecular Chemistry}, volume = {9}, number = {9}, pages = {3080--3104}, abstract = {Associated with the CD1d protein, KRN 7000, a potent synthetic α-galactosylceramide, is known to activate the invariant NKT immune cells. This stimulation then leads to the production of different cytokines modulating a T(H)1/T(H)2 immune response balance involved in protection against several pathologies such as autoimmune diseases and cancers. Various efforts have been made toward the synthesis of simple and more functionalized analogues in order to selectively induce T(H)1 or T(H)2-type cytokine production. Since the discovery of KRN 7000, structure-activity relationships, crystallographic and modelling studies have pointed to the potential of several GalCer analogues in term of selective bioactivity, and have highlighted interesting elements in order to better understand the recognition and activation mechanisms of immune iNKT cells. By presenting an up-to-date library of analogues, collecting recent breakthroughs done in crystallography and molecular modelling, and relating them to the available biological results, we hope that this review will highlight and help the scientific community in their KRN research.}, keywords = {Adjuvants, Animals, Antigen, Antigens, CD1d, Galactosylceramides, Helper-Inducer, Humans, I2CT, Immunologic, Monneaux, Receptors, T-Cell, T-Lymphocytes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular insights into the ligand-controlled organization of the SAM-I riboswitch}, author = {B Heppell and S Blouin and A M Dussault and J Mulhbacher and E Ennifar and J C Penedo and D A Lafontaine}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21532599}, doi = {10.1038/nchembio.563}, isbn = {ISBN/1552-4469 (Electronic) 1552-4450 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {Nat Chem Biol}, volume = {7}, number = {6}, pages = {384-92}, abstract = {S-adenosylmethionine (SAM) riboswitches are widespread in bacteria, and up to five different SAM riboswitch families have been reported, highlighting the relevance of SAM regulation. On the basis of crystallographic and biochemical data, it has been postulated, but never demonstrated, that ligand recognition by SAM riboswitches involves key conformational changes in the RNA architecture. We show here that the aptamer follows a two-step hierarchical folding selectively induced by metal ions and ligand binding, each of them leading to the formation of one of the two helical stacks observed in the crystal structure. Moreover, we find that the anti-antiterminator P1 stem is rotated along its helical axis upon ligand binding, a mechanistic feature that could be common to other riboswitches. We also show that the nonconserved P4 helical domain is used as an auxiliary element to enhance the ligand-binding affinity. This work provides the first comprehensive characterization, to our knowledge, of a ligand-controlled riboswitch folding pathway.}, note = {Published online 01 May 2011}, keywords = {Aptamers, Bacterial/*chemistry *Riboswitch S-Adenosylmethionine/*chemistry, DUMAS, ENNIFAR, Nucleotide/chemistry Bacillus subtilis/genetics Binding Sites Crystallography, Unité ARN, X-Ray Ligands Metals Nucleic Acid Conformation RNA}, pubstate = {published}, tppubtype = {article} } @article{benincasa_antifungal_2011, title = {Antifungal activity of amphotericin B conjugated to carbon nanotubes}, author = {Monica Benincasa and Sabrina Pacor and Wei Wu and Maurizio Prato and Alberto Bianco and Renato Gennaro}, doi = {10.1021/nn1023522}, issn = {1936-086X}, year = {2011}, date = {2011-01-01}, journal = {ACS nano}, volume = {5}, number = {1}, pages = {199--208}, abstract = {Amphotericin B (AMB) has long been considered the most effective drug in the treatment of serious invasive fungal infections. There are, however, major limitations to its use, due to several adverse effects, including acute infusional reactions and, most relevant, a dose-dependent nephrotoxicity. At least some of these effects are attributed to the aggregation of AMB as a result of its poor water solubility. To overcome this problem, reformulated versions of the drug have been developed, including a micellar dispersion of AMB with sodium deoxycholate (AMBD), its encapsulation into liposomes, or its incorporation into lipidic complexes. The development of nanobiotechnologies provides novel potential drug delivery systems that make use of nanomaterials such as functionalized carbon nanotubes (f-CNTs), which are emerging as an innovative and efficient tool for the transport and cellular translocation of therapeutic molecules. In this study, we prepared two conjugates between f-CNTs and AMB. The antifungal activity of these conjugates was tested against a collection of reference and clinical fungal strains, in comparison to that of AMB alone or AMBD. Measured minimum inhibition concentration (MIC) values for f-CNT-AMB conjugates were either comparable to or better than those displayed by AMB and AMBD. Furthermore, AMBD-resistant Candida strains were found to be susceptible to f-CNT-AMB 1. Additional studies, aimed at understanding the mechanism of action of the conjugates, suggest a nonlytic mechanism, since the compounds show a major permeabilizing effect on the tested fungal strains only after extended incubation. Interestingly, the f-CNT-AMB 1 does not show any significant toxic effect on Jurkat cells at antifungal concentrations.}, keywords = {Amphotericin B, Antifungal Agents, Candida, carbon, Cell Membrane, Deoxycholic Acid, Drug Design, Drug Resistance, Fungal, Humans, I2CT, Jurkat Cells, Kinetics, Membrane Potentials, Nanotubes, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bou_aoun_analysis_2011, title = {Analysis of thioester-containing proteins during the innate immune response of Drosophila melanogaster}, author = {Richard Bou Aoun and Charles Hetru and Laurent Troxler and Daniel Doucet and Dominique Ferrandon and Nicolas Matt}, doi = {10.1159/000321554}, issn = {1662-8128}, year = {2011}, date = {2011-01-01}, journal = {J Innate Immun}, volume = {3}, number = {1}, pages = {52--64}, abstract = {Thioester-containing proteins (TEPs) are conserved proteins among insects that are thought to be involved in innate immunity. In Drosophila, the Tep family is composed of 6 genes named Tep1-Tep6. In this study, we investigated the phylogeny, expression pattern and roles of these genes in the host defense of Drosophila. Protostomian Tep genes are clustered in 3 distinct branches, 1 of which is specific to mosquitoes. Most D. melanogaster Tep genes are expressed in hemocytes, can be induced in the fat body, and are expressed in specific regions of the hypodermis. This expression pattern is consistent with a role in innate immunity. However, we find that TEP1, TEP2, and TEP4 are not strictly required in the body cavity to fight several bacterial and fungal infections. One possibility is that Drosophila TEPs act redundantly or that their absence can be compensated by other components of the immune response. TEPs may thus provide a subtle selective advantage during evolution. Alternatively, they may be required in host defense against specific as yet unidentified natural pathogens of Drosophila.}, keywords = {Animals, bioinformatic, DNA, Evolution, ferrandon, Gene Expression Regulation, Hemocytes, Immunity, In Situ Hybridization, Innate, M3i, matt, Molecular, Mutation, Phylogeny, Sequence Analysis}, pubstate = {published}, tppubtype = {article} } @article{reichhart_drosophila_2011, title = {The Drosophila serpins: multiple functions in immunity and morphogenesis}, author = {Jean-Marc Reichhart and David Gubb and Vincent Leclerc}, doi = {10.1016/B978-0-12-386471-0.00011-0}, issn = {1557-7988}, year = {2011}, date = {2011-01-01}, journal = {Meth. Enzymol.}, volume = {499}, pages = {205--225}, abstract = {Members of the serpin superfamily of proteins have been found in all living organisms, although rarely in bacteria or fungi. They have been extensively studied in mammals, where many rapid physiological responses are regulated by inhibitory serpins. In addition to the inhibitory serpins, a large group of noninhibitory proteins with a conserved serpin fold have also been identified in mammals. These noninhibitory proteins have a wide range of functions, from storage proteins to molecular chaperones, hormone transporters, and tumor suppressors. In contrast, until recently, very little was known about insect serpins in general, or Drosophila serpins in particular. In the last decade, however, there has been an increasing interest in the serpin biology of insects. It is becoming clear that, like in mammals, a similar wide range of physiological responses are regulated in insects and that noninhibitory serpin-fold proteins also play key roles in insect biology. Drosophila is also an important model organism that can be used to study human pathologies (among which serpinopathies or other protein conformational diseases) and mechanisms of regulation of proteolytic cascades in health or to develop strategies for control of insect pests and disease vectors. As most of our knowledge on insect serpins comes from studies on the Drosophila immune response, we survey here the Drosophila serpin literature and describe the laboratory techniques that have been developed to study serpin-regulated responses in this model genetic organism.}, keywords = {Animals, Immunity, Innate, M3i, Morphogenesis, reichhart, Serpins, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{meister_immune_2011, title = {Immune cell transdifferentiation: a complex crosstalk between circulating immune cells and the haematopoietic niche}, author = {Marie Meister and Dominique Ferrandon}, doi = {embor2011238 [pii] 10.1038/embor.2011.238}, issn = {1469-3178 (Electronic) 1469-221X (Linking)}, year = {2011}, date = {2011-01-01}, journal = {EMBO Rep}, volume = {13}, pages = {3--4}, keywords = {ferrandon, M3i}, pubstate = {published}, tppubtype = {article} } @article{lee_negative_2011b, title = {Negative regulation of immune responses on the fly}, author = {Kwang-Zin Lee and Dominique Ferrandon}, doi = {10.1038/emboj.2011.47}, issn = {1460-2075}, year = {2011}, date = {2011-01-01}, journal = {EMBO J.}, volume = {30}, number = {6}, pages = {988--990}, keywords = {*Gene Expression Regulation, *Homeostasis, Animals, bacteria, Bacteria/*immunology, Biological, Drosophila melanogaster/*immunology, Drosophila Proteins/biosynthesis/metabolism, ferrandon, Gene Expression Regulation, Homeostasis, M3i, Mitogen-Activated Protein Kinases, Mitogen-Activated Protein Kinases/metabolism, Models, NF-kappa B, NF-kappa B/metabolism, ras Proteins, ras Proteins/metabolism, Receptor Protein-Tyrosine Kinases, Receptor Protein-Tyrosine Kinases/metabolism}, pubstate = {published}, tppubtype = {article} } @article{nehme_relative_2011b, title = {Relative roles of the cellular and humoral responses in the Drosophila host defense against three gram-positive bacterial infections}, author = {Nadine T Nehme and Jessica Quintin and Ju Hyun Cho and Janice Lee and Marie-Céline Lafarge and Christine Kocks and Dominique Ferrandon}, doi = {10.1371/journal.pone.0014743}, issn = {1932-6203}, year = {2011}, date = {2011-01-01}, journal = {PLoS ONE}, volume = {6}, number = {3}, pages = {e14743}, abstract = {BACKGROUND: Two NF-kappaB signaling pathways, Toll and immune deficiency (imd), are required for survival to bacterial infections in Drosophila. In response to septic injury, these pathways mediate rapid transcriptional activation of distinct sets of effector molecules, including antimicrobial peptides, which are important components of a humoral defense response. However, it is less clear to what extent macrophage-like hemocytes contribute to host defense. METHODOLOGY/PRINCIPAL FINDINGS: In order to dissect the relative importance of humoral and cellular defenses after septic injury with three different gram-positive bacteria (Micrococcus luteus, Enterococcus faecalis, Staphylococcus aureus), we used latex bead pre-injection to ablate macrophage function in flies wildtype or mutant for various Toll and imd pathway components. We found that in all three infection models a compromised phagocytic system impaired fly survival--independently of concomitant Toll or imd pathway activation. Our data failed to confirm a role of the PGRP-SA and GNBP1 Pattern Recognition Receptors for phagocytosis of S. aureus. The Drosophila scavenger receptor Eater mediates the phagocytosis by hemocytes or S2 cells of E. faecalis and S. aureus, but not of M. luteus. In the case of M. luteus and E. faecalis, but not S. aureus, decreased survival due to defective phagocytosis could be compensated for by genetically enhancing the humoral immune response. CONCLUSIONS/SIGNIFICANCE: Our results underscore the fundamental importance of both cellular and humoral mechanisms in Drosophila immunity and shed light on the balance between these two arms of host defense depending on the invading pathogen.}, keywords = {Animals, Antimicrobial Cationic Peptides, Carrier Proteins, Cell Surface, Cellular, Enterococcus faecalis, ferrandon, Gram-Positive Bacteria, Gram-Positive Bacterial Infections, Host-Pathogen Interactions, Humoral, Immunity, Innate, M3i, Micrococcus luteus, Opsonin Proteins, Phagocytosis, Receptors, Signal Transduction, Solubility, Staphylococcus aureus}, pubstate = {published}, tppubtype = {article} } @article{herrero_functionalised_2011, title = {Functionalised carbon nanotubes: high biocompatibility with lack of toxicity}, author = {Antonia M Herrero and Lara Lacerda and Alberto Bianco and Kostas Kostarelos and Maurizio Prato}, url = {http://www.inderscience.com/link.php?id=44433}, doi = {10.1504/IJNT.2011.044433}, issn = {1475-7435, 1741-8151}, year = {2011}, date = {2011-01-01}, urldate = {2020-04-01}, journal = {International Journal of Nanotechnology}, volume = {8}, number = {10/11/12}, pages = {885}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of tRNA amino acid-accepting end in aminoacylation and its quality control}, author = {X L Zhou and D H Du and M Tan and H Y Lei and L L Ruan and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21775341}, doi = {10.1093/nar/gkr595}, issn = {1362-4962 (Electronic) 0305-1048 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {Nucleic Acids Res}, volume = {39}, number = {20}, pages = {8857-8868}, abstract = {Aminoacyl-tRNA synthetases (aaRSs) are remarkable enzymes that are in charge of the accurate recognition and ligation of amino acids and tRNA molecules. The greatest difficulty in accurate aminoacylation appears to be in discriminating between highly similar amino acids. To reduce mischarging of tRNAs by non-cognate amino acids, aaRSs have evolved an editing activity in a second active site to cleave the incorrect aminoacyl-tRNAs. Editing occurs after translocation of the aminoacyl-CCA(76) end to the editing site, switching between a hairpin and a helical conformation for aminoacylation and editing. Here, we studied the consequence of nucleotide changes in the CCA(76) accepting end of tRNA(Leu) during the aminoacylation and editing reactions. The analysis showed that the terminal A(76) is essential for both reactions, suggesting that critical interactions occur in the two catalytic sites. Substitutions of C(74) and C(75) selectively decreased aminoacylation keeping nearly unaffected editing. These mutations might favor the regular helical conformation required to reach the editing site. Mutating the editing domain residues that contribute to CCA(76) binding reduced the aminoacylation fidelity leading to cell-toxicity in the presence of non-cognate amino acids. Collectively, the data show how protein synthesis quality is controlled by the CCA(76) homogeneity of tRNAs.}, note = {DOI: 10.1093/nar/gkr595}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A quantitative RNA code for mRNA target selection by the germline fate determinant GLD-1}, author = {J E Wright and D Gaidatzis and M Senften and B M Farley and E Westhof and S P Ryder and R Ciosk}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21169991}, doi = {10.1038/emboj.2010.334}, issn = {1460-2075 (Electronic) 0261-4189 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {EMBO J}, volume = {30}, number = {3}, pages = {533-545}, abstract = {RNA-binding proteins (RBPs) are critical regulators of gene expression. To understand and predict the outcome of RBP-mediated regulation a comprehensive analysis of their interaction with RNA is necessary. The signal transduction and activation of RNA (STAR) family of RBPs includes developmental regulators and tumour suppressors such as Caenorhabditis elegans GLD-1, which is a key regulator of germ cell development. To obtain a comprehensive picture of GLD-1 interactions with the transcriptome, we identified GLD-1-associated mRNAs by RNA immunoprecipitation followed by microarray detection. Based on the computational analysis of these mRNAs we generated a predictive model, where GLD-1 association with mRNA is determined by the strength and number of 7-mer GLD-1-binding motifs (GBMs) within UTRs. We verified this quantitative model both in vitro, by competition GLD-1/GBM-binding experiments to determine relative affinity, and in vivo, by 'transplantation' experiments, where 'weak' and 'strong' GBMs imposed translational repression of increasing strength on a non-target mRNA. This study demonstrates that transcriptome-wide identification of RBP mRNA targets combined with quantitative computational analysis can generate highly predictive models of post-transcriptional regulatory networks.}, note = {Wright, Jane E Gaidatzis, Dimos Senften, Mathias Farley, Brian M Westhof, Eric Ryder, Sean P Ciosk, Rafal GM081422/GM/NIGMS NIH HHS/United States R01 GM081422-01A1/GM/NIGMS NIH HHS/United States R01 GM081422-02/GM/NIGMS NIH HHS/United States R01 GM081422-03/GM/NIGMS NIH HHS/United States R01 GM081422-04/GM/NIGMS NIH HHS/United States Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't England The EMBO journal EMBO J. 2011 Feb 2;30(3):533-45. Epub 2010 Dec 17.}, keywords = {Animals Binding, Biological Protein Interaction Domains and Motifs/*genetics RNA, Competitive Caenorhabditis elegans/*genetics/metabolism Caenorhabditis elegans Proteins/*metabolism Computational Biology/methods Gene Regulatory Networks/*genetics Immunoprecipitation Microarray Analysis *Models, Messenger/*metabolism RNA-Binding Proteins/*metabolism, Unité ARN, WESTHOF, WESTHOF Animals Binding}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure-based design of short peptide ligands binding onto the E. coli processivity ring}, author = {P Wolff and V Olieric and J P Briand and O Chaloin and A Dejaegere and P Dumas and E Ennifar and G Guichard and J Wagner and D Y Burnouf}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21619076}, doi = {10.1021/jm200311m}, issn = {1520-4804 (Electronic) 0022-2623 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {J Med Chem}, volume = {54}, number = {13}, pages = {4627-4637}, abstract = {The multimeric DNA sliding clamps confer high processivity to replicative DNA polymerases and are also binding platforms for various enzymes involved in DNA metabolism. These enzymes interact with the clamp through a small peptide that binds into a hydrophobic pocket which is a potential target for the development of new antibacterial compounds. Starting from a generic heptapeptide, we used a structure-based strategy to improve the design of new peptide ligands. Chemical modifications at specific residues result in a dramatic increase of the interaction as measured by SPR and ITC. The affinity of our best hits was improved by 2 orders of magnitude as compared to the natural ligand, reaching 10(-8) M range. The molecular basis of the interactions was analyzed by solving the co-crystal structures of the most relevant peptides bound to the clamp and reveals how chemical modifications establish new contacts and contributes to an increased affinity of the ligand.}, note = {Wolff, Philippe Olieric, Vincent Briand, Jean Paul Chaloin, Olivier Dejaegere, Annick Dumas, Philippe Ennifar, Eric Guichard, Gilles Wagner, Jerome Burnouf, Dominique Y J Med Chem. 2011 Jul 14;54(13):4627-37. Epub 2011 Jun 9.}, keywords = {Crystallography, ENNIFAR, Molecular Oligopeptides/*chemical synthesis/chemistry Protein Binding Structure-Activity Relationship Thermodynamics, Unité ARN, X-Ray DNA Polymerase III/*chemistry DNA Polymerase beta/*chemistry Drug Design Escherichia coli/*chemistry Escherichia coli Proteins/*chemistry Ligands Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Predicting and modeling RNA architecture}, author = {E Westhof and B Masquida and F Jossinet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20504963}, doi = {10.1101/cshperspect.a003632}, issn = {1943-0264 (Electronic)}, year = {2011}, date = {2011-01-01}, journal = {Cold Spring Harb Perspect Biol}, volume = {3}, number = {2}, pages = {309-320}, abstract = {A general approach for modeling the architecture of large and structured RNA molecules is described. The method exploits the modularity and the hierarchical folding of RNA architecture that is viewed as the assembly of preformed double-stranded helices defined by Watson-Crick base pairs and RNA modules maintained by non-Watson-Crick base pairs. Despite the extensive molecular neutrality observed in RNA structures, specificity in RNA folding is achieved through global constraints like lengths of helices, coaxiality of helical stacks, and structures adopted at the junctions of helices. The Assemble integrated suite of computer tools allows for sequence and structure analysis as well as interactive modeling by homology or ab initio assembly with possibilities for fitting within electronic density maps. The local key role of non-Watson-Crick pairs guides RNA architecture formation and offers metrics for assessing the accuracy of three-dimensional models in a more useful way than usual root mean square deviation (RMSD) values.}, note = {Westhof, Eric Masquida, Benoit Jossinet, Fabrice United States Cold Spring Harbor perspectives in biology Cold Spring Harb Perspect Biol. 2011 Feb 1;3(2). pii: a003632. doi: 10.1101/cshperspect.a003632.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Endless Subtleties of RNA-Protein Complexes.}, author = {E Westhof and V Fritsch}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21742256}, issn = {1878-4186}, year = {2011}, date = {2011-01-01}, journal = {Structure}, volume = {19}, number = {7}, pages = {902-903}, abstract = {New NMR structures of complexes between double-strand RNA binding domains and their RNA hairpin substrates reported by Wang et al. in this issue of Structure reveal striking adaptations in both the protein recognition element and in the RNA.}, note = {DOI:10.1016/j.str.2011.06.006}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {8-Modified-2'-Deoxyadenosine Analogues Induce Delayed Polymerization Arrest during HIV-1 Reverse Transcription.}, author = {V Vivet-Boudou and C Isel and M Sleiman and R Smyth and N Ben Gaied and P Barhoum and G Laumond and G Bec and M Götte and J Mak and A M Aubertin and A Burger and R Marquet}, url = {http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0027456}, doi = {doi: 10.1371/journal.pone.0027456}, isbn = {PMC3210175}, year = {2011}, date = {2011-01-01}, journal = {PLoS One}, volume = {6}, number = {11}, pages = {e27456}, abstract = {The occurrence of resistant viruses to any of the anti-HIV-1 compounds used in the current therapies against AIDS underlies the urge for the development of new drug targets and/or new drugs acting through novel mechanisms. While all anti-HIV-1 nucleoside analogues in clinical use and in clinical trials rely on ribose modifications for activity, we designed nucleosides with a natural deoxyribose moiety and modifications of position 8 of the adenine base. Such modifications might induce a steric clash with helix αH in the thumb domain of the p66 subunit of HIV-1 RT at a distance from the catalytic site, causing delayed chain termination. Eleven new 2'-deoxyadenosine analogues modified on position 8 of the purine base were synthesized and tested in vitro and in cell-based assays. In this paper we demonstrate for the first time that chemical modifications on position 8 of 2'-deoxyadenosine induce delayed chain termination in vitro, and also inhibit DNA synthesis when incorporated in a DNA template strand. Furthermore, one of them had moderate anti-HIV-1 activity in cell-culture. Our results constitute a proof of concept indicating that modification on the base moiety of nucleosides can induce delayed polymerization arrest and inhibit HIV-1 replication.}, note = {Published online 2011 November 7}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A program to analyze the distributions of unmeasured reflections}, author = {L Urzhumtseva and A Urzhumtsev}, url = {http://scripts.iucr.org/cgi-bin/paper?gm5016}, year = {2011}, date = {2011-01-01}, journal = {J Appl Cryst}, volume = {44}, number = {4}, pages = {865-872}, abstract = {Crystallographic Fourier maps may contain barely interpretable or non-interpretable regions if these maps are calculated with an incomplete set of diffraction data. Even a small percentage of missing data may be crucial if these data are distributed non-uniformly and form connected regions of reciprocal space. Significant time and effort can be lost trying to interpret poor maps, in improving them by phase refinement or in fighting against artefacts, whilst the problem could in fact be solved by completing the data set. To characterize the distribution of missing reflections, several types of diagrams have been suggested in addition to the usual plots of completeness in resolution shells and cumulative data completeness. A computer program, FOBSCOM, has been developed to analyze the spatial distribution of unmeasured diffraction data, to search for connected regions of unmeasured reflections and to obtain numeric characteristics of these regions. By performing this analysis, the program could help to save time during structure solution for a number of projects. It can also provide information about a possible overestimation of the map quality and model-biased features when calculated values are used to replace unmeasured data.}, note = {10.1107/S0021889811019546}, keywords = {Unité ARN, Unmeasured data Connected regions Fourier maps Computer programs}, pubstate = {published}, tppubtype = {article} } @article{, title = {Roles and regulation of microRNAs in cytomegalovirus infection}, author = {L Tuddenham and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21545855}, doi = {10.1016/j.bbagrm.2011.04.002}, issn = {0006-3002 (Print) 0006-3002 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {Biochim Biophys Acta-Gene Regul Mech}, volume = {1809}, number = {11-12}, pages = {613-622}, abstract = {MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression post-transcriptionally via binding to complementary sites typically located in the 3' untranslated regions (UTRs) of their target mRNAs. This ancient regulatory system has been conserved in eukaryotes throughout evolution, and it is therefore unsurprising that certain viruses have evolved to express their own miRNAs. Since the initial discovery of Epstein-Barr virus (EBV) derived miRNAs in 2004, over 230 viral miRNAs have been identified, the majority arising from herpesviruses. Although the functions of most viral miRNAs remain to be elucidated, an increasing number of their cellular and viral targets have been experimentally validated. Due to their non-immunogenic nature, viral miRNAs represent an elegant tool for the virus to evade the host immune system, and likely play a key role in the latent/lytic switch during the viral lifecycle. In this review, we will focus on the interactions of cytomegaloviruses with cellular and viral miRNAs during infection. This article is part of a Special Issue entitled: MicroRNAs in viral gene regulation.}, note = {Journal article Biochimica et biophysica acta Biochim Biophys Acta. 2011 Apr 21.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {New dimensions of RNA in molecular recognition and catalysis.}, author = {E C Theil and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22181677}, isbn = {22181677}, year = {2011}, date = {2011-01-01}, journal = {Acc Chem Res}, volume = {44}, number = {12}, pages = {1255-1256}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Inhibition of Aminoglycoside-Deactivating Enzymes APH(3')-IIIa and AAC(6')-Ii by Amphiphilic Paromomycin O2''-Ether Analogues.}, author = {J Szychowski and J Kondo and O Zahr and K Auclair and E Westhof and S Hanessian and J W Keillor}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21905229}, doi = {10.1002/cmdc.201100346}, isbn = {21905229}, year = {2011}, date = {2011-01-01}, journal = {ChemMedChem}, volume = {6}, number = {11}, pages = {1961-1966}, abstract = {Paromomycin analogue activity: Novel amphiphilic aminoglycosides are shown to inhibit clinically relevant deactivating enzymes, without undergoing significant deactivation themselves.}, note = {Article first published online: 8 SEP 2011}, keywords = {Aminoglycosides Antibacterial agents Antibiotics APH(3′)-IIIa Drug discovery Paromomycin, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Kaposi's sarcoma herpesvirus microRNAs target caspase 3 and regulate apoptosis.}, author = {G Suffert and G Malterer and J Hausser and J Viiliäinen and A Fender and M Contrant and T Ivacevic and V Benes and F Gros and O Voinnet and M Zavolan and P M Ojala and J G Haas and S Pfeffer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/22174674}, isbn = {22174674}, year = {2011}, date = {2011-01-01}, journal = {PLoS Pathog}, volume = {7}, number = {12}, pages = {e1002405}, abstract = {Kaposi's sarcoma herpesvirus (KSHV) encodes a cluster of twelve micro (mi)RNAs, which are abundantly expressed during both latent and lytic infection. Previous studies reported that KSHV is able to inhibit apoptosis during latent infection; we thus tested the involvement of viral miRNAs in this process. We found that both HEK293 epithelial cells and DG75 cells stably expressing KSHV miRNAs were protected from apoptosis. Potential cellular targets that were significantly down-regulated upon KSHV miRNAs expression were identified by microarray profiling. Among them, we validated by luciferase reporter assays, quantitative PCR and western blotting caspase 3 (Casp3), a critical factor for the control of apoptosis. Using site-directed mutagenesis, we found that three KSHV miRNAs, miR-K12-1, 3 and 4-3p, were responsible for the targeting of Casp3. Specific inhibition of these miRNAs in KSHV-infected cells resulted in increased expression levels of endogenous Casp3 and enhanced apoptosis. Altogether, our results suggest that KSHV miRNAs directly participate in the previously reported inhibition of apoptosis by the virus, and are thus likely to play a role in KSHV-induced oncogenesis.}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA structures, genomic organization and selection of recombinant HIV}, author = {E Simon-Loriere and P Rossolillo and M Negroni}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21422815}, doi = {10.4161/rna.8.2.15193}, issn = {1555-8584 (Electronic) 1547-6286 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {RNA Biol}, volume = {8}, number = {2}, pages = {280-286}, abstract = {Recombination is an evolutionary mechanism intrinsic to the evolution of many RNA viruses. In retroviruses and notably in the case of HIV, recombination is so frequent that it can be considered as part of its mode of replication. This process not only plays a central role in shaping HIV genetic diversity worldwide, but has also been involved in immune escape and development of resistance to antiviral treatments. Recombination does not create new mutations in the existing genetic repertoire of the virus, but creates new combinations of pre-existing polymorphisms. The simultaneous insertion of multiple substitutions in a single replication cycle leaves little room for the progressive coevolution of regions of proteins, RNA or, more in general, genomes, to accommodate these drastic sequence changes. Therefore, recombination, while allowing the virus to rapidly explore larger sequence space than the slow accumulation of point mutations, also runs the risk of generating non functional viruses. Recombination is the consequence of a switch in the template used during reverse transcription and is promoted by the presence of structured regions in the genomic RNA template. In this review, we discuss new observations suggesting that the distribution of RNA structures along the HIV genome may enhance recombination rates in regions where the resultant progeny is less likely to be impaired, and could therefore maximize the evolutionary value of this source of genetic diversity.}, note = {Journal article RNA biology RNA Biol. 2011 Mar 1;8(2).}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{bechetoille_new_2011, title = {A new organotypic model containing dermal-type macrophages}, author = {N Bechetoille and H Vachon and A Gaydon and A Boher and T Fontaine and E Schaeffer and M Decossas and V Andre-Frei and C G Mueller}, year = {2011}, date = {2011-01-01}, journal = {Experimental Dermatology}, volume = {20}, number = {1600-0625 (Electronic)}, pages = {1035--1037}, abstract = {Human skin equivalents (SEs) are popular three-dimensional (D) cell culture systems in fundamental and applied dermatology. They have been made to contain dendritic cells, but so far no study on the incorporation of potentially anti-inflammatory dermal macrophages has been performed. Here, we show that monocyte-derived dermal-type macrophages can be introduced into a rigid scaffold with dermal fibroblasts. They maintain their cell surface markers CD163, DC-SIGN/CD209 and HLA-DR, which discriminate them from monocytes and dendritic cells. They retain the ability to produce the anti-inflammatory cytokine IL-10 in response to lipopolysaccharide (LPS) and to phagocytose latex beads. We thus demonstrate the feasibility of creating macrophage-fibroblast 3D cultures as a first step towards generating SEs with dermal macrophages}, keywords = {CELL CULTURE, Chemistry, Culture, cytokine, Dendritic Cells, DERMATOLOGY, Fibroblast, Fibroblasts, HLA-DR, Human, IL-10, IL10, Immunology, Latex, Letter, lipopolysaccharide, LPS, Macrophage, Macrophages, monocyte, Monocytes, Skin, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Insights into translation initiation and termination complexes and into the polysome architecture}, author = {A Simonetti and S Marzi and A Myasnikov and J F Menetret and B P Klaholz}, editor = {M Rodnina and W Wintermeyer and R Green}, url = {http://www.springerlink.com/content/j5022n7218036w14}, doi = {10.1007/978-3-7091-0215-2_10}, year = {2011}, date = {2011-01-01}, booktitle = {Ribosomes Structures, Function, and Dynamics}, pages = {113-128}, publisher = {Springer}, address = {Wien}, abstract = {Translation initiation is the most strongly regulated phase of protein synthesis during which the synthesis of a given protein is decided on. Initiation is the least conserved step of translation, since bacteria, archaea and eukarya have distinct and very different ways to initiate translation, and many different trans-acting factors are involved in the process. In bacteria, translation initiation comprises the consecutive formation of three major intermediary initiation complexes that are assembled via a multi-step process and that differ in composition and in conformation. At the end of the initiation process, an active 70S ribosomal initiation complex (70S IC) has formed which can enter peptide bond formation.}, keywords = {ENNIFAR, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {miR-346 controls release of TNF-alpha protein and stability of its mRNA in rheumatoid arthritis via tristetraprolin stabilization}, author = {N Semaan and L Frenzel and G Alsaleh and G Suffert and J E Gottenberg and J Sibilia and S Pfeffer and D Wachsmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21611196}, doi = {10.1371/journal.pone.0019827}, issn = {1932-6203 (Electronic) 1932-6203 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {PLoS One}, volume = {6}, number = {5}, pages = {e19827}, abstract = {TNF-alpha is a major cytokine implicated in rheumatoid arthritis. Its expression is regulated both at the transcriptional and posttranscriptional levels and recent data demonstrated that miRNAs are implicated in TNF-alpha response in macrophages. LPS-activated FLS isolated from RA patients express TNF-alpha mRNA but not the mature protein. This prompted us to look for miRNAs which could be implicated in this anti-inflammatory effect. Using a microarray, we found two miRNAs, miR-125b and miR-939 predicted to target the 3'-UTR of TNF-alpha mRNA, to be up-regulated in RA FLS in response to LPS, but their repression did not restore mature TNF-alpha expression in FLS. We showed previously that miR-346, which is upregulated in LPS-activated FLS, inhibited Btk expression that stabilized TNF-alpha mRNA. Blocking miR-346 reestablished TNF-alpha expression in activated FLS. Interestingly, transfection of miR-346 in LPS-activated THP-1 cells inhibited TNF-alpha secretion. We also demonstrated that TTP, a RNA binding protein which inhibited TNF-alpha synthesis, is overexpressed in activated FLS and that inhibition of miR-346 decreases its expression. Conversely, transfection of miR-346 in LPS-activated THP-1 cells increased TTP mRNA expression and inhibited TNF-alpha release. These results indicate that miR-346 controls TNF-alpha synthesis by regulating TTP expression.}, note = {Semaan, Noha Frenzel, Laurent Alsaleh, Ghada Suffert, Guillaume Gottenberg, Jacques-Eric Sibilia, Jean Pfeffer, Sebastien Wachsmann, Dominique Research Support, Non-U.S. Gov't United States PloS one PLoS One. 2011;6(5):e19827. Epub 2011 May 17. DOI: 10.1371/journal.pone.0019827}, keywords = {Antisense/metabolism RNA, arthritis, Biological Protein Stability/drug effects Protein-Tyrosine Kinases/metabolism *RNA Stability/drug effects RNA, Messenger/genetics/metabolism Reverse Transcriptase Polymerase Chain Reaction Synovial Fluid/cytology Transfection Tristetraprolin/*metabolism Tumor Necrosis Factor-alpha/biosynthesis/genetics/*secretion, PFEFFER, Rheumatoid/*genetics/pathology Cell Line Fibroblasts/drug effects/metabolism/pathology Gene Expression Regulation Humans Lipopolysaccharides/pharmacology MicroRNAs/genetics/*metabolism Models, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{canard_generation_2011, title = {Generation of anti-DC-SIGN monoclonal antibodies capable of blocking HIV-1 gp120 binding and reactive on formalin-fixed tissue}, author = {B Canard and H Vachon and T Fontaine and J J Pin and S Paul and C Genin and C G Mueller}, year = {2011}, date = {2011-01-01}, journal = {Immunol.Lett.}, volume = {135}, number = {1879-0542 (Electronic)}, pages = {165--172}, abstract = {DC-SIGN is a C-type lectin of recognized importance in immunology and in the pathogenicity human pathogens. Monoclonal antibodies directed against DC-SIGN have been generated, but their systemic characterization for interfering with binding of the HIV-1 glycoprotein 120 has often been omitted. Moreover, so far, no anti-DC-SIGN monoclonal antibody has been described that recognizes its antigen after formalin fixation and paraffin embedding. In this study, we have generated new anti-DC-SIGN monoclonal antibodies using HeLa cells stably expressing DC-SIGN as immunogen. We have obtained 11 hybridoma clones producing antibodies that recognized DC-SIGN on monocyte-derived dendritic cells and on dermal-type macrophages. Seven monoclonal antibodies displayed a capacity to interfere with DC-SIGN binding to HIV-1 gp120. One recognized DC-SIGN on formalin-fixed dendritic cells and macrophages. Using this antibody we have obtained specific labelling of DC-SIGN and colocalisation with the dermal macrophage marker CD163 on human skin. The described monoclonal anti-human DC-SIGN antibodies will be of use to the scientific community to address fundamental immunology issues, in particular concerning macrophages and dendritic cells, and help elucidate infection events of pathogen targeting DC-SIGN as recognition receptor}, keywords = {Adhesion, adhesion molecules, Animals, Antibodies, antibody, Antigen, Antigens, Blocking, C-Type, C-type lectin, CD, Cell Adhesion, Cell Adhesion Molecules, Cell Surface, Chemistry, clones, Dendritic Cells, DERMIS, Differentiation, Fixatives, Formaldehyde, formalin-fixed tissue, Genetics, GLYCOPROTEIN, GP120, HeLa Cells, HIV, HIV Envelope Protein gp120, HIV-1, Human, Humans, hybridoma, ICAM-3, immunodeficiency, Immunology, Inbred BALB C, infection, LECTIN, Lectins, Macrophage, Macrophages, Mice, Monoclonal, monoclonal antibody, MONOCLONAL-ANTIBODY, Monocytes, Murine-Derived, Myelomonocytic, Nih 3T3 Cells, Paraffin Embedding, pathogenicity, Protein, Receptor, Receptors, recognition, Skin, Team-Mueller, virus}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural insights into viral determinants of nematode mediated Grapevine fanleaf virus transmission.}, author = {P Schellenberger and C Sauter and B Lorber and P Bron and S Trapani and M Bergdoll and A Marmonier and C Schmitt-Kelchinger and O Lemaire and G Demangeat and C Ritzenthaler}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21625570}, doi = {10.1371/journal.ppat.1002034}, isbn = {21625570}, year = {2011}, date = {2011-01-01}, journal = {PLoS Pathog}, volume = {7}, number = {5}, pages = {e1002034}, abstract = {Many animal and plant viruses rely on vectors for their transmission from host to host. Grapevine fanleaf virus (GFLV), a picorna-like virus from plants, is transmitted specifically by the ectoparasitic nematode Xiphinema index. The icosahedral capsid of GFLV, which consists of 60 identical coat protein subunits (CP), carries the determinants of this specificity. Here, we provide novel insight into GFLV transmission by nematodes through a comparative structural and functional analysis of two GFLV variants. We isolated a mutant GFLV strain (GFLV-TD) poorly transmissible by nematodes, and showed that the transmission defect is due to a glycine to aspartate mutation at position 297 (Gly297Asp) in the CP. We next determined the crystal structures of the wild-type GFLV strain F13 at 3.0 Å and of GFLV-TD at 2.7 Å resolution. The Gly297Asp mutation mapped to an exposed loop at the outer surface of the capsid and did not affect the conformation of the assembled capsid, nor of individual CP molecules. The loop is part of a positively charged pocket that includes a previously identified determinant of transmission. We propose that this pocket is a ligand-binding site with essential function in GFLV transmission by X. index. Our data suggest that perturbation of the electrostatic landscape of this pocket affects the interaction of the virion with specific receptors of the nematode's feeding apparatus, and thereby severely diminishes its transmission efficiency. These data provide a first structural insight into the interactions between a plant virus and a nematode vector.}, keywords = {FLORENTZ, FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Strategies for the crystallization of viruses: Using phase diagrams and gels to produce 3D crystals of Grapevine fanleaf virus}, author = {P Schellenberger and G Demangeat and O Lemaire and C Ritzenthaler and M Bergdoll and V Olieric and C Sauter and B Lorber}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21352920}, doi = {10.1016/j.jsb.2011.02.007}, issn = {1095-8657 (Electronic) 1047-8477 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {J Struct Biol}, volume = {174}, number = {2}, pages = {344-351}, abstract = {The small icosahedral plant RNA nepovirus Grapevine fanleaf virus (GFLV) is specifically transmitted by a nematode and causes major damage to vineyards worldwide. To elucidate the molecular mechanisms underlying the recognition between the surface of its protein capsid and cellular components of its vector, host and viral proteins synthesized upon infection, the wild type GFLV strain F13 and a natural mutant (GFLV-TD) carrying a Gly(297)Asp mutation were purified, characterized and crystallized. Subsequently, the geometry and volume of their crystals was optimized by establishing phase diagrams. GFLV-TD was twice as soluble as the parent virus in the crystallization solution and its crystals diffracted X-rays to a resolution of 2.7A. The diffraction limit of GFLV-F13 crystals was extended from 5.5 to 3A by growth in agarose gel. Preliminary crystallographic analyses indicate that both types of crystals are suitable for structure determination. Keys for the successful production of GFLV crystals include the rigorous quality control of virus preparations, crystal quality improvement using phase diagrams, and crystal lattice reinforcement by growth in agarose gel. These strategies are applicable to the production of well-diffracting crystals of other viruses and macromolecular assemblies.}, note = {Schellenberger, Pascale Demangeat, Gerard Lemaire, Olivier Ritzenthaler, Christophe Bergdoll, Marc Olieric, Vincent Sauter, Claude Lorber, Bernard United States Journal of structural biology J Struct Biol. 2011 May;174(2):344-51. Epub 2011 Feb 23.}, keywords = {FLORENTZ, FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Misfolded human tRNA isodecoder binds and neutralizes a 3' UTR-embedded Alu element}, author = {J Rudinger-Thirion and A Lescure and C Paulus and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21896722}, doi = {10.1073/pnas.1103698108}, issn = {1091-6490 (Electronic) 0027-8424 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {108}, number = {40}, pages = {E794-802}, abstract = {Several classes of small noncoding RNAs are key players in cellular metabolism including mRNA decoding, RNA processing, and mRNA stability. Here we show that a tRNA(Asp) isodecoder, corresponding to a human tRNA-derived sequence, binds to an embedded Alu RNA element contained in the 3' UTR of the human aspartyl-tRNA synthetase mRNA. This interaction between two well-known classes of RNA molecules, tRNA and Alu RNA, is driven by an unexpected structural motif and induces a global rearrangement of the 3' UTR. Besides, this 3' UTR contains two functional polyadenylation signals. We propose a model where the tRNA/Alu interaction would modulate the accessibility of the two alternative polyadenylation sites and regulate the stability of the mRNA. This unique regulation mechanism would link gene expression to RNA polymerase III transcription and may have implications in a primate-specific signal pathway.}, note = {Rudinger-Thirion, Joelle Lescure, Alain Paulus, Caroline Frugier, Magali United States Proceedings of the National Academy of Sciences of the United States of America Proc Natl Acad Sci U S A. 2011 Oct 4;108(40):E794-802. Epub 2011 Sep 6.}, keywords = {FRUGIER, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Increased muscle stress-sensitivity induced by selenoprotein N inactivation in mouse: a mammalian model for SEPN1-related myopathy.}, author = {M Rederstorff and P Castets and S Arbogast and J Lainé and S Vassilopoulos and M Beuvin and O Dubourg and A Vignaud and A Ferry and A Krol and V Allamand and P Guicheney and A Ferreiro and A Lescure}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21858002}, doi = {10.1371/journal.pone.0023094}, isbn = {21858002}, year = {2011}, date = {2011-01-01}, journal = {PLoS One}, volume = {6}, number = {8}, pages = {e23094}, abstract = {Selenium is an essential trace element and selenoprotein N (SelN) was the first selenium-containing protein shown to be directly involved in human inherited diseases. Mutations in the SEPN1 gene, encoding SelN, cause a group of muscular disorders characterized by predominant affection of axial muscles. SelN has been shown to participate in calcium and redox homeostasis, but its pathophysiological role in skeletal muscle remains largely unknown. To address SelN function in vivo, we generated a Sepn1-null mouse model by gene targeting. The Sepn1(-/-) mice had normal growth and lifespan, and were macroscopically indistinguishable from wild-type littermates. Only minor defects were observed in muscle morphology and contractile properties in SelN-deficient mice in basal conditions. However, when subjected to challenging physical exercise and stress conditions (forced swimming test), Sepn1(-/-) mice developed an obvious phenotype, characterized by limited motility and body rigidity during the swimming session, as well as a progressive curvature of the spine and predominant alteration of paravertebral muscles. This induced phenotype recapitulates the distribution of muscle involvement in patients with SEPN1-Related Myopathy, hence positioning this new animal model as a valuable tool to dissect the role of SelN in muscle function and to characterize the pathophysiological process}, keywords = {KROL, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Labeling of multiple HIV-1 proteins with the biarsenical-tetracysteine system}, author = {C F Pereira and P C Ellenberg and K L Jones and T L Fernandez and R P Smyth and D J Hawkes and M Hijnen and V Vivet-Boudou and R Marquet and I Johnson and J Mak}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21347302}, doi = {10.1371/journal.pone.0017016}, issn = {1932-6203 (Electronic) 1932-6203 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {PLoS One}, volume = {6}, number = {2}, pages = {e17016}, abstract = {Due to its small size and versatility, the biarsenical-tetracysteine system is an attractive way to label viral proteins for live cell imaging. This study describes the genetic labeling of the human immunodeficiency virus type 1 (HIV-1) structural proteins (matrix, capsid and nucleocapsid), enzymes (protease, reverse transcriptase, RNAse H and integrase) and envelope glycoprotein 120 with a tetracysteine tag in the context of a full-length virus. We measure the impact of these modifications on the natural virus infection and, most importantly, present the first infectious HIV-1 construct containing a fluorescently-labeled nucleocapsid protein. Furthermore, due to the high background levels normally associated with the labeling of tetracysteine-tagged proteins we have also optimized a metabolic labeling system that produces infectious virus containing the natural envelope glycoproteins and specifically labeled tetracysteine-tagged proteins that can easily be detected after virus infection of T-lymphocytes. This approach can be adapted to other viral systems for the visualization of the interplay between virus and host cell during infection.}, note = {Pereira, Candida F Ellenberg, Paula C Jones, Kate L Fernandez, Tara L Smyth, Redmond P Hawkes, David J Hijnen, Marcel Vivet-Boudou, Valerie Marquet, Roland Johnson, Iain Mak, Johnson Research Support, Non-U.S. Gov't United States PloS one PLoS One. 2011 Feb 11;6(2):e17016.}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A human pathology-related mutation prevents import of an aminoacyl-tRNA synthetase into mitochondria}, author = {M Messmer and C Florentz and H Schwenzer and G C Scheper and M S van der Knaap and L Marechal-Drouard and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21121901}, doi = {10.1042/BJ20101902}, issn = {1470-8728 (Electronic) 0264-6021 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {Biochem J}, volume = {433}, number = {3}, pages = {441-446}, abstract = {Mutations in the nuclear gene coding for the mitochondrial aspartyl-tRNA synthetase, a key enzyme for mitochondrial translation, are correlated with leukoencephalopathy. A Ser to Gly mutation is located in the predicted targeting signal of the protein. We demonstrate in the present study, by in vivo and in vitro approaches, that this pathology-related mutation impairs the import process across mitochondrial membranes.}, note = {Messmer, Marie Florentz, Catherine Schwenzer, Hagen Scheper, Gert C van der Knaap, Marjo S Marechal-Drouard, Laurence Sissler, Marie Research Support, Non-U.S. Gov't England The Biochemical journal Biochem J. 2011 Jan 14;433(3):441-6.}, keywords = {Aspartate-tRNA Ligase/*genetics/*metabolism Cell Line Humans Leukoencephalopathies/etiology/genetics Mitochondria/*metabolism *Mutation, FLORENTZ, Missense Protein Transport, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNase P: At last, the key finds its lock.}, author = {B Masquida and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/21803972}, doi = {10.1261/rna.2841511}, isbn = {21803972}, year = {2011}, date = {2011-01-01}, journal = {RNA}, volume = {17}, number = {9}, pages = {1615-1618}, abstract = {Apart from the ribosome, the crystal structure of the bacterial RNase P in complex with a tRNA, reported by Reiter and colleagues recently, constitutes the first example of a multiple turnover RNA enzyme. Except in rare exceptions, RNase P is ubiquitous and, like the ribosome, is older than the initial branch point of the phylogenetic tree. Importantly, the structure shows how the RNA and the protein moieties cooperate to process the pre-tRNA substrates. The catalytic site comprises some critical RNA residues spread over the secondary structure but gathered in a compact volume next to the protein, which helps recognize and orient the substrate. The discussion here outlines some important aspects of that crystal structure, some of which could apply to RNA molecules in general.}, keywords = {RNase P Multiple-turnover ribozyme RNA–RNA interactions Molecular evolution Crystallography, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Importance of the proline-rich multimerization domain on the oligomerization and nucleic acid binding properties of HIV-1 Vif}, author = {S Bernacchi and G Mercenne and C Tournaire and R Marquet and J C Paillart}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21076154}, doi = {10.1093/nar/gkq979}, issn = {1362-4962 (Electronic) 0305-1048 (Linking)}, year = {2011}, date = {2011-01-01}, journal = {Nucleic Acids Res}, volume = {39}, number = {6}, pages = {2404-2415}, abstract = {The HIV-1 viral infectivity factor (Vif) is required for productive infection of non-permissive cells, including most natural HIV-1 targets, where it counteracts the antiviral activities of the cellular cytosine deaminases APOBEC-3G (A3G) and A3F. Vif is a multimeric protein and the conserved proline-rich domain (161)PPLP(164) regulating Vif oligomerization is crucial for its function and viral infectivity. Here, we expressed and purified wild-type Vif and a mutant protein in which alanines were substituted for the proline residues of the (161)PPLP(164) domain. Using dynamic light scattering, circular dichroism and fluorescence spectroscopy, we established the impact of these mutations on Vif oligomerization, secondary structure content and nucleic acids binding properties. In vitro, wild-type Vif formed oligomers of five to nine proteins, while Vif AALA formed dimers and/or trimers. Up to 40% of the unbound wild-type Vif protein appeared to be unfolded, but binding to the HIV-1 TAR apical loop promoted formation of beta-sheets. Interestingly, alanine substitutions did not significantly affect the secondary structure of Vif, but they diminished its binding affinity and specificity for nucleic acids. Dynamic light scattering showed that Vif oligomerization, and interaction with folding-promoting nucleic acids, favor formation of high molecular mass complexes. These properties could be important for Vif functions involving RNAs.}, note = {Bernacchi, Serena Mercenne, Gaelle Tournaire, Clemence Marquet, Roland Paillart, Jean-Christophe England Nucleic acids research Nucleic Acids Res. 2011 Mar 1;39(6):2404-15. Epub 2010 Nov 13.}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{cellot_neurons_2010, title = {Neurons are able to internalize soluble carbon nanotubes: new opportunities or old risks?}, author = {Giada Cellot and Laura Ballerini and Maurizio Prato and Alberto Bianco}, doi = {10.1002/smll.201000906}, issn = {1613-6829}, year = {2010}, date = {2010-12-01}, journal = {Small (Weinheim an Der Bergstrasse, Germany)}, volume = {6}, number = {23}, pages = {2630--2633}, keywords = {carbon, Cell Line, Cells, Cultured, Humans, I2CT, Nanotubes, Neurons, Team-Bianco, tumor}, pubstate = {published}, tppubtype = {article} } @article{noordegraaf_functional_2010, title = {Functional redundancy of Langerhans cells and Langerin+ dermal dendritic cells in contact hypersensitivity}, author = {Madelon Noordegraaf and Vincent Flacher and Patrizia Stoitzner and Björn E Clausen}, doi = {10.1038/jid.2010.223}, issn = {1523-1747}, year = {2010}, date = {2010-12-01}, journal = {The Journal of Investigative Dermatology}, volume = {130}, number = {12}, pages = {2752--2759}, abstract = {The relative roles of Langerhans cells (LC), dermal dendritic cells (DC), and, in particular, the recently discovered Langerin(+) dermal DC subset in the induction and control of contact hypersensitivity (CHS) responses remain controversial. Using an inducible mouse model, in which LC and other Langerin(+) DC can be depleted by injection of diphtheria toxin, we previously reported impaired transport of topically applied antigen to draining lymph nodes and reduced CHS in the absence of all Langerin(+) skin DC. In this study, we demonstrate that mice with a selective depletion of LC exhibit attenuated CHS only upon sensitization with a low hapten dose but not with a high hapten dose. In contrast, when painting a higher concentration of hapten onto the skin, which leads to increased antigen dissemination into the dermis, CHS is still diminished in mice lacking all Langerin(+) skin DC. Taken together, these data suggest that the magnitude of a CHS reaction depends on the number of skin DC, which have access to the hapten, rather than on the presence or absence of a particular skin DC population. LC and (Langerin(+)) dermal DC thus seem to have a redundant function in regulating CHS.}, keywords = {Animal, Animals, Antigen, Antigens, C-Type, CHS, contact, CONTACT HYPERSENSITIVITY, Dendritic Cells, DEPLETION, DERMAL DENDRITIC CELLS, Dermatitis, DERMIS, Diphtheria Toxin, Disease Models, Epidermis, function, Gene Knock-In Techniques, Genetics, Growth, HAPTEN, Haptens, Heparin-binding EGF-like Growth Factor, Hypersensitivity, Immunology, Inbred C57BL, INDUCTION, Intercellular Signaling Peptides and Proteins, LACKING, Langerhans Cells, LECTIN, Lectins, LYMPH, LYMPH NODE, Lymph Nodes, Mannose-Binding Lectins, metabolism, Mice, mouse, Mutant Strains, Organ Culture Techniques, pathology, Peptides, Poisons, Protein, Proteins, RESPONSES, signaling, Skin, Surface, Team-Mueller, Toxicity}, pubstate = {published}, tppubtype = {article} } @article{al-jamal_enhanced_2010, title = {Enhanced cellular internalization and gene silencing with a series of cationic dendron-multiwalled carbon nanotube:siRNA complexes}, author = {Khuloud T Al-Jamal and Francesca M Toma and Açelya Yilmazer and Hanene Ali-Boucetta and Antonio Nunes and Maria-Antonia Herrero and Bowen Tian and Ayad Eddaoudi and Ayad Eddaoui and Wafa' T Al-Jamal and Alberto Bianco and Maurizio Prato and Kostas Kostarelo}, doi = {10.1096/fj.09-141036}, issn = {1530-6860}, year = {2010}, date = {2010-11-01}, journal = {FASEB journal: official publication of the Federation of American Societies for Experimental Biology}, volume = {24}, number = {11}, pages = {4354--4365}, abstract = {One of the major obstacles to the clinical development of gene silencing by small interfering RNA (siRNA) is its effective cytoplasmic delivery. Carbon nanotubes have been proposed as novel nanomaterials that can offer significant advantages for the intracellular delivery of nucleic acids, such as siRNA. We recently demonstrated in a proof-of-principle study that amino-functionalized multiwalled carbon nanotubes (f-MWNT) can effectively deliver in vivo an siRNA sequence, triggering cell apoptosis that results in human lung xenograft eradication and prolonged survival. In the present study, we demonstrate how a newly synthesized series of polycationic dendron-MWNT constructs with a precisely tailored number of amino functions (dendron generations) can complex and effectively deliver double-stranded siRNA to achieve gene silencing in vitro. A systematic comparison between the f-MWNT series in terms of cellular uptake, cytotoxicity, and siRNA complexation is offered. Significant improvement in siRNA delivery with the dendron-MWNT conjugates is shown, and gene silencing was obtained in 2 human cell lines using 2 different siRNA sequences. The study reveals that through f-MWNT structure-biological function analysis novel nanotube-based siRNA transfer vectors can be designed with minimal cytotoxicity and effective delivery and gene-silencing capabilities.}, keywords = {Biological Transport, carbon, Cations, Cell Line, Cell Survival, Gene Silencing, HeLa Cells, Humans, I2CT, Models, Molecular, Nanotubes, RNA, Small Interfering, Team-Bianco, Transfection, tumor}, pubstate = {published}, tppubtype = {article} } @article{van_den_bossche_efficient_2010, title = {Efficient receptor-independent intracellular translocation of aptamers mediated by conjugation to carbon nanotubes}, author = {Jeroen Van den Bossche and Wafa' T Al-Jamal and Bowen Tian and Antonio Nunes and Chiara Fabbro and Alberto Bianco and Maurizio Prato and Kostas Kostarelos}, doi = {10.1039/c0cc02092c}, issn = {1364-548X}, year = {2010}, date = {2010-10-01}, journal = {Chemical Communications (Cambridge, England)}, volume = {46}, number = {39}, pages = {7379--7381}, abstract = {We have covalently grafted aptamers onto carboxylated carbon nanotubes to design a novel vector system that can easily translocate into the cytosol of different cell types independent of receptor-mediated uptake. We propose the use of carbon nanotubes for the efficient intracellular delivery of biologically active aptamers for potential therapeutic applications.}, keywords = {Aptamers, Base Sequence, Biological Transport, carbon, Cell Line, Cell Surface, DNA Primers, Electron, Electrophoresis, Humans, I2CT, Microscopy, Nanotubes, Nucleotide, Polyacrylamide Gel, Receptors, Team-Bianco, Transmission, tumor}, pubstate = {published}, tppubtype = {article} } @article{RH2010, title = {A heterodimeric complex of the LRR proteins LRIM1 and APL1C regulates complement-like immunity in Anopheles gambiae}, author = {Richard H Baxter and Stefanie Steinert and Y Chelliah and Gloria Volohonsky and Elena A Levashina and J Deisenhofer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/20826443}, year = {2010}, date = {2010-09-08}, volume = {107}, number = {39}, pages = {16817-22}, abstract = {The leucine-rich repeat (LRR) proteins LRIM1 and APL1C control the function of the complement-like protein TEP1 in Anopheles mosquitoes. The molecular structure of LRIM1 and APL1C and the basis of their interaction with TEP1 represent a new type of innate immune complex. The LRIM1/APL1C complex specifically binds and solubilizes a cleaved form of TEP1 without an intact thioester bond. The LRIM1 and APL1C LRR domains have a large radius of curvature, glycosylated concave face, and a novel C-terminal capping motif. The LRIM1/APL1C complex is a heterodimer with a single intermolecular disulfide bond. The structure of the LRIM1/APL1C heterodimer reveals an interface between the two LRR domains and an extensive C-terminal coiled-coil domain. We propose that a cleaved form of TEP1 may act as a convertase for activation of other TEP1 molecules and that the LRIM1/APL1C heterodimer regulates formation of this TEP1 convertase.}, keywords = {APL1C, LRIM1}, pubstate = {published}, tppubtype = {article} } @article{nicolas_identification_2010, title = {Identification of rep-associated factors in herpes simplex virus type 1-induced adeno-associated virus type 2 replication compartments.}, author = {Armel Nicolas and Nathalie Alazard-Dany and Coline Biollay and Loredana Arata and Nelly Jolinon and Lauriane Kuhn and Myriam Ferro and Sandra K Weller and Alberto L Epstein and Anna Salvetti and Anna Greco}, doi = {10.1128/JVI.00725-10}, issn = {1098-5514 0022-538X 0022-538X}, year = {2010}, date = {2010-09-01}, journal = {Journal of virology}, volume = {84}, number = {17}, pages = {8871--8887}, abstract = {Adeno-associated virus (AAV) is a human parvovirus that replicates only in cells coinfected with a helper virus, such as adenovirus or herpes simplex virus type 1 (HSV-1). We previously showed that nine HSV-1 factors are able to support AAV rep gene expression and genome replication. To elucidate the strategy of AAV replication in the presence of HSV-1, we undertook a proteomic analysis of cellular and HSV-1 factors associated with Rep proteins and thus potentially recruited within AAV replication compartments (AAV RCs). This study resulted in the identification of approximately 60 cellular proteins, among which factors involved in DNA and RNA metabolism represented the largest functional categories. Validation analyses indicated that the cellular DNA replication enzymes RPA, RFC, and PCNA were recruited within HSV-1-induced AAV RCs. Polymerase delta was not identified but subsequently was shown to colocalize with Rep within AAV RCs even in the presence of the HSV-1 polymerase complex. In addition, we found that AAV replication is associated with the recruitment of components of the Mre11/Rad50/Nbs1 complex, Ku70 and -86, and the mismatch repair proteins MSH2, -3, and -6. Finally, several HSV-1 factors were also found to be associated with Rep, including UL12. We demonstrated for the first time that this protein plays a role during AAV replication by enhancing the resolution of AAV replicative forms and AAV particle production. Altogether, these analyses provide the basis to understand how AAV adapts its replication strategy to the nuclear environment induced by the helper virus.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{samori_potentiometric_2010, title = {Potentiometric titration as a straightforward method to assess the number of functional groups on shortened carbon nanotubes}, author = {Cristian Samorì and Raquel Sainz and Cécilia Ménard-Moyon and Francesca M Toma and Enrica Venturelli and Prabhpreet Singh and Marco Ballestri and Maurizio Prato and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S0008622310001806}, doi = {10.1016/j.carbon.2010.03.015}, issn = {0008-6223}, year = {2010}, date = {2010-08-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {48}, number = {9}, pages = {2447--2454}, abstract = {The conditions for oxidizing multi-walled carbon nanotubes to shorten them to a narrow length distribution have been optimized. One of the most difficult achievements is to fully characterize this material from a chemical point of view, and to find a good quantitative correlation among different techniques. Herein, we report on the combination of different methods to determine the number of functional groups generated during strong acid treatment and a further amidation reaction. A good correlation was found using the colorimetric Kaiser test, thermogravimetric analysis and potentiometric argentometric titration. The final technique, used for the first time in this field, is highly versatile and, being non-destructive, allows a complete recovery of the starting material. Short carbon nanotubes are particularly useful for applications in biomedicine, and the control and precise assessment of their functionalization is critical when used as carriers for therapeutic molecules.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{silverman_serpins_2010, title = {Serpins flex their muscle: I. Putting the clamps on proteolysis in diverse biological systems}, author = {Gary A Silverman and James C Whisstock and Stephen P Bottomley and James A Huntington and Dion Kaiserman and Cliff J Luke and Stephen C Pak and Jean-Marc Reichhart and Phillip I Bird}, doi = {10.1074/jbc.R110.112771}, issn = {1083-351X}, year = {2010}, date = {2010-08-01}, journal = {J. Biol. Chem.}, volume = {285}, number = {32}, pages = {24299--24305}, abstract = {Serpins compose the largest superfamily of peptidase inhibitors and are well known as regulators of hemostasis and thrombolysis. Studies using model organisms, from plants to vertebrates, now show that serpins and their unique inhibitory mechanism and conformational flexibility are exploited to control proteolysis in molecular pathways associated with cell survival, development, and host defense. In addition, an increasing number of non-inhibitory serpins are emerging as important elements within a diversity of biological systems by serving as chaperones, hormone transporters, or anti-angiogenic factors.}, keywords = {Animals, Biological, Caenorhabditis elegans, Cell Death, Cell Differentiation, Cell Survival, Homeostasis, Humans, Immunity, Innate, M3i, Mice, Models, Phenotype, reichhart, Serpins, Transgenes, transgenic}, pubstate = {published}, tppubtype = {article} } @article{whisstock_serpins_2010, title = {Serpins flex their muscle: II. Structural insights into target peptidase recognition, polymerization, and transport functions}, author = {James C Whisstock and Gary A Silverman and Phillip I Bird and Stephen P Bottomley and Dion Kaiserman and Cliff J Luke and Stephen C Pak and Jean-Marc Reichhart and James A Huntington}, doi = {10.1074/jbc.R110.141408}, issn = {1083-351X}, year = {2010}, date = {2010-08-01}, journal = {J. Biol. Chem.}, volume = {285}, number = {32}, pages = {24307--24312}, abstract = {Inhibitory serpins are metastable proteins that undergo a substantial conformational rearrangement to covalently trap target peptidases. The serpin reactive center loop contributes a majority of the interactions that serpins make during the initial binding to target peptidases. However, structural studies on serpin-peptidase complexes reveal a broader set of contacts on the scaffold of inhibitory serpins that have substantial influence on guiding peptidase recognition. Structural and biophysical studies also reveal how aberrant serpin folding can lead to the formation of domain-swapped serpin multimers rather than the monomeric metastable state. Serpin domain swapping may therefore underlie the polymerization events characteristic of the serpinopathies. Finally, recent structural studies reveal how the serpin fold has been adapted for non-inhibitory functions such as hormone binding.}, keywords = {Animals, Biological, Biological Transport, Biophysics, Catalytic Domain, Hormones, Humans, Kinetics, M3i, Models, Peptide Hydrolases, Protein Binding, Protein Conformation, Protein Structure, reichhart, Serpins, Substrate Specificity, Tertiary, Thrombin}, pubstate = {published}, tppubtype = {article} } @article{MK2010, title = {The Major Yolk Protein Vitellogenin Interferes with the Anti-Plasmodium Response in the Malaria Mosquito Anopheles gambiae}, author = {Martin K Rono and Miranda M Whitten and M Oulad-Abdelghani and Elena A Levashina and Eric Marois}, url = {http://www.ncbi.nlm.nih.gov/pubmed/20652016}, year = {2010}, date = {2010-07-20}, journal = {PLoS Biol.}, volume = {10}, number = {7}, pages = {e1000434}, abstract = {When taking a blood meal on a person infected with malaria, female Anopheles gambiae mosquitoes, the major vector of human malaria, acquire nutrients that will activate egg development (oogenesis) in their ovaries. Simultaneously, they infect themselves with the malaria parasite. On traversing the mosquito midgut epithelium, invading Plasmodium ookinetes are met with a potent innate immune response predominantly controlled by mosquito blood cells. Whether the concomitant processes of mosquito reproduction and immunity affect each other remains controversial. Here, we show that proteins that deliver nutrients to maturing mosquito oocytes interfere with the antiparasitic response. Lipophorin (Lp) and vitellogenin (Vg), two nutrient transport proteins, reduce the parasite-killing efficiency of the antiparasitic factor TEP1. In the absence of either nutrient transport protein, TEP1 binding to the ookinete surface becomes more efficient. We also show that Lp is required for the normal expression of Vg, and for later Plasmodium development at the oocyst stage. Furthermore, our results uncover an inhibitory role of the Cactus/REL1/REL2 signaling cassette in the expression of Vg, but not of Lp. We reveal molecular links that connect reproduction and immunity at several levels and provide a molecular basis for a long-suspected trade-off between these two processes.}, keywords = {anti-Plasmodium response, M3i, marois, vittelogenin}, pubstate = {published}, tppubtype = {article} } @article{fabre_noncovalent_2010, title = {Noncovalent assembly of ferrocene on modified gold surfaces mediated by uracil–adenine base pairs}, author = {Bruno Fabre and Soraya Ababou-Girard and Prabhpreet Singh and Jitendra Kumar and Sandeep Verma and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S1388248110001451}, doi = {10.1016/j.elecom.2010.03.045}, issn = {1388-2481}, year = {2010}, date = {2010-06-01}, urldate = {2020-04-01}, journal = {Electrochemistry Communications}, volume = {12}, number = {6}, pages = {831--834}, abstract = {Redox-active ferrocene was assembled on gold surfaces through the hydrogen bonding interactions between adenine-substituted ferrocene and a uracil-terminated organothiol monolayer. The surface coverage of ferrocene Γ could be varied from ca. 4×10−11 to 2.0×10−10molcm−2 by diluting the thiol-modified uracil derivative with inert 1-octanethiol. A decrease in the apparent electron transfer rate constant for ferrocene, kapp, from ca. 50 to 10s−1 was observed upon increasing Γ.}, keywords = {Electrochemistry, Ferrocene, Gold surface, I2CT, Monolayer, Nucleobases, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{kostarelos_complement_2010, title = {Complement monitoring of carbon nanotubes}, author = {K Kostarelos and A Bianco and M Prato}, url = {https://www.nature.com/articles/nnano.2010.110}, doi = {10.1038/nnano.2010.110}, issn = {1748-3395}, year = {2010}, date = {2010-06-01}, urldate = {2020-04-01}, journal = {Nature Nanotechnology}, volume = {5}, number = {6}, pages = {382--383}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{matskevich_drosophila_2010b, title = {The Drosophila PRR GNBP3 assembles effector complexes involved in antifungal defenses independently of its Toll-pathway activation function}, author = {Alexey A Matskevich and Jessica Quintin and Dominique Ferrandon}, doi = {10.1002/eji.200940164}, issn = {1521-4141}, year = {2010}, date = {2010-05-01}, journal = {Eur. J. Immunol.}, volume = {40}, number = {5}, pages = {1244--1254}, abstract = {The Drosophila Toll-signaling pathway controls the systemic antifungal host response. Gram-negative binding protein 3 (GNBP3), a member of the beta-glucan recognition protein family senses fungal infections and activates this pathway. A second detection system perceives the activity of proteolytic fungal virulence factors and redundantly activates Toll. GNBP3(hades) mutant flies succumb more rapidly to Candida albicans and to entomopathogenic fungal infections than WT flies, despite normal triggering of the Toll pathway via the virulence detection system. These observations suggest that GNBP3 triggers antifungal defenses that are not dependent on activation of the Toll pathway. Here, we show that GNBP3 agglutinates fungal cells. Furthermore, it can activate melanization in a Toll-independent manner. Melanization is likely to be an essential defense against some fungal infections given that the entomopathogenic fungus Beauveria bassiana inhibits the activity of the main melanization enzymes, the phenol oxidases. Finally, we show that GNBP3 assembles "attack complexes", which comprise phenoloxidase and the necrotic serpin. We propose that Drosophila GNBP3 targets fungi immediately at the inception of the infection by bringing effector molecules in direct contact with the invading microorganisms.}, keywords = {Agglutination, Animals, Beauveria, Beauveria/immunology, Candida albicans, Candida albicans/immunology, Carrier Proteins, Carrier Proteins/*immunology/pharmacology, Drosophila melanogaster/*immunology/microbiology, Drosophila Proteins/*immunology/pharmacology/physiology, Enzyme Activation, ferrandon, Fungal, Fungi, Fungi/*immunology, Hemolymph, Hemolymph/immunology, M3i, Melanins, Melanins/*physiology, Monophenol Monooxygenase, Monophenol Monooxygenase/physiology, Multiprotein Complexes, Multiprotein Complexes/physiology, Recombinant Fusion Proteins, Recombinant Fusion Proteins/pharmacology, Serpins, Serpins/physiology, Spores, Toll-Like Receptors, Toll-Like Receptors/immunology}, pubstate = {published}, tppubtype = {article} } @article{pmid19801499, title = {HIV-1 regulation of latency in the monocyte-macrophage lineage and in CD4+ T lymphocytes}, author = {Laetitia Redel and Valentin Le Douce and Thomas Cherrier and Céline Marban and Andrea Janossy and Dominique Aunis and Carine Van Lint and Olivier Rohr and Christian Schwartz}, doi = {10.1189/jlb.0409264}, issn = {1938-3673}, year = {2010}, date = {2010-04-01}, urldate = {2010-04-01}, journal = {J Leukoc Biol}, volume = {87}, number = {4}, pages = {575--588}, abstract = {The introduction in 1996 of the HAART raised hopes for the eradication of HIV-1. Unfortunately, the discovery of latent HIV-1 reservoirs in CD4+ T cells and in the monocyte-macrophage lineage proved the optimism to be premature. The long-lived HIV-1 reservoirs constitute a major obstacle to the eradication of HIV-1. In this review, we focus on the establishment and maintenance of HIV-1 latency in the two major targets for HIV-1: the CD4+ T cells and the monocyte-macrophage lineage. Understanding the cell-type molecular mechanisms of establishment, maintenance, and reactivation of HIV-1 latency in these reservoirs is crucial for efficient therapeutic intervention. A complete viral eradication, the holy graal for clinicians, might be achieved by strategic interventions targeting latently and productively infected cells. We suggest that new approaches, such as the combination of different kinds of proviral activators, may help to reduce dramatically the size of latent HIV-1 reservoirs in patients on HAART.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{marega_two-dimensional_2010, title = {Two-dimensional diffusion-ordered NMR spectroscopy as a tool for monitoring functionalized carbon nanotube purification and composition}, author = {Riccardo Marega and Vincent Aroulmoji and Massimo Bergamin and Luigi Feruglio and Francesca Dinon and Alberto Bianco and Erminio Murano and Maurizio Prato}, doi = {10.1021/nn100257h}, issn = {1936-086X}, year = {2010}, date = {2010-04-01}, journal = {ACS nano}, volume = {4}, number = {4}, pages = {2051--2058}, abstract = {Functionalized carbon nanotube (CNT) derivatives are currently under thorough investigation in different biomedical investigations. In this field of research, the composition of sample either in terms of covalently attached or physisorbed moieties can greatly affect the observed results and hamper the comparison between different studies. Therefore, the availability of a fast and reliable analytical technique to assess both the type of interaction (covalent vs noncovalent) and the composition of CNT conjugates is of great importance. Here we describe that the two-dimensional diffusion-ordered (DOSY) NMR spectroscopy is extremely useful to discriminate between conjugated and unconjugated polyethylene glycol groups in samples obtained by condensation with oxidized single-walled carbon nanotubes (SWNTs). This fast and nondestructive technique allows us to follow the removal of unconjugated polyethylene glycol chains during the purification. In particular, DOSY analysis reveal that about 1/3 (wt %) of the polyethylene glycol used for the condensation remained physisorbed to functionalized SWNTs after dialysis. Complete elimination of physisorbed polyethylene glycol was achieved using diafiltration.}, keywords = {carbon, Diffusion, I2CT, Magnetic Resonance Spectroscopy, Nanotubes, Polyethylene Glycols, Solubility, Team-Bianco, Temperature, water}, pubstate = {published}, tppubtype = {article} } @article{lacotte_identification_2010, title = {Identification of new pathogenic players in lupus: autoantibody-secreting cells are present in nephritic kidneys of (NZBxNZW)F1 mice}, author = {Stéphanie Lacotte and Hélène Dumortier and Marion Décossas and Jean-Paul Briand and Sylviane Muller}, doi = {10.4049/jimmunol.0902595}, issn = {1550-6606}, year = {2010}, date = {2010-04-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {184}, number = {7}, pages = {3937--3945}, abstract = {An important hallmark of systemic lupus erythematosus is the production of autoantibodies specific for nuclear Ags, among which nucleosomes and their constituents, DNA and histones. It is widely admitted that some of these autoantibodies contribute largely in lupus pathogenesis because of their nephritogenic potential. However, the underlying mechanisms are still debated. In this study, we analyzed the autoimmune response against histone H2B during the course of the disease in lupus-prone (NZBxNZW)F1 mice, both in lymphoid organs and kidneys, and we assessed its potential involvement in lupus pathogenicity. We found that the N-terminal region of histone H2B represents a preferential target for circulating autoantibodies, which kinetics of appearance positively correlates with disease development. Furthermore, immunization of preautoimmune (NZBxNZW)F1 mice with H2B peptide 1-25 accelerates the disease. Kidney eluates from diseased (NZBxNZW)F1 mice do contain IgG Abs reacting with this peptide, and this H2B sequence was found to be accessible to specific Ab probes in Ag-containing deposits detected in nephritic kidneys. Finally, compared with control normal mice and to young preautoimmune (NZBxNZW)F1 animals, the frequency of cells secreting autoantibodies reacting with peptide 1-25 was significantly raised in the spleen and bone marrow and most importantly on a pathophysiological point of view, locally, in nephritic kidneys of diseased (NZBxNZW)F1 mice. Altogether our results demonstrate the existence in (NZBxNZW)F1 mice of both a systemic and local B cell response targeting the N-terminal region of histone H2B, and highlight the potential implication of this nuclear domain in lupus pathology.}, keywords = {Animals, Autoantibodies, Autoantigens, B-Lymphocytes, Dumortier, Enzyme-Linked Immunosorbent Assay, Female, Histones, I2CT, Immunoblotting, Immunohistochemistry, Inbred BALB C, Inbred NZB, Lupus Nephritis, Mice, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{samori_enhanced_2010, title = {Enhanced anticancer activity of multi-walled carbon nanotube-methotrexate conjugates using cleavable linkers}, author = {Cristian Samorì and Hanene Ali-Boucetta and Raquel Sainz and Chang Guo and Francesca Maria Toma and Chiara Fabbro and Tatiana da Ros and Maurizio Prato and Kostas Kostarelos and Alberto Bianco}, doi = {10.1039/b923560d}, issn = {1364-548X}, year = {2010}, date = {2010-03-01}, journal = {Chemical Communications (Cambridge, England)}, volume = {46}, number = {9}, pages = {1494--1496}, abstract = {Methotrexate was tethered to multi-walled carbon nanotubes through different cleavable linkers exploiting the ammonium functionalities introduced by 1,3-dipolar cycloaddition reaction of azomethine ylides to the nanotubes. The new nanobio-hybrid conjugates were internalized into human breast cancer cells and it was shown that the cytotoxic activity was strongly dependent on the presence and type of linker.}, keywords = {Antineoplastic Agents, Azo Compounds, carbon, Cell Line, Cross-Linking Reagents, Humans, I2CT, Methotrexate, Nanotubes, Team-Bianco, Thiosemicarbazones, tumor}, pubstate = {published}, tppubtype = {article} } @article{geotti-bianchini_replacement_2010, title = {Replacement of Ala by Aib improves structuration and biological stability in thymine-based α-nucleopeptides}, author = {Piero Geotti-Bianchini and Alessandro Moretto and Cristina Peggion and Julien Beyrath and Alberto Bianco and Fernando Formaggio}, url = {https://pubs.rsc.org/en/content/articlelanding/2010/ob/b920211k}, doi = {10.1039/B920211K}, issn = {1477-0539}, year = {2010}, date = {2010-03-01}, urldate = {2020-03-31}, journal = {Organic & Biomolecular Chemistry}, volume = {8}, number = {6}, pages = {1315--1321}, abstract = {Three thymine-based nucleo-heptapeptides, each containing two nucleo-amino acids and zero, one or four Aib residues, respectively, have been synthesized. A single Aib residue is enough to promote the adoption of a helical structure in our nucleopeptides and to increase significantly their resistance towards enzymatic degradation. The insertion of four Aib residues, out of seven residues in the sequence, affords a rigid, 310-helical nucleopeptide that is substantially unaffected by serum enzymes and is not cytotoxic.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{al-jamal_systemic_2010, title = {Systemic antiangiogenic activity of cationic poly-L-lysine dendrimer delays tumor growth}, author = {Khuloud T Al-Jamal and Wafa’ T Al-Jamal and Simon Akerman and Jennifer E Podesta and Açelya Yilmazer and John A Turton and Alberto Bianco and Neil Vargesson and Chryso Kanthou and Alexander T Florence and Gillian M Tozer and Kostas Kostarelos}, url = {https://www.pnas.org/content/107/9/3966}, doi = {10.1073/pnas.0908401107}, issn = {0027-8424, 1091-6490}, year = {2010}, date = {2010-03-01}, urldate = {2020-04-01}, journal = {Proceedings of the National Academy of Sciences}, volume = {107}, number = {9}, pages = {3966--3971}, abstract = {This study describes the previously unreported intrinsic capacity of poly-L-lysine (PLL) sixth generation (G6) dendrimer molecules to exhibit systemic antiangiogenic activity that could lead to solid tumor growth arrest. The PLL-dendrimer-inhibited tubule formation of SVEC4-10 murine endothelial cells and neovascularization in the chick embryo chick chorioallantoic membrane (CAM) assay. Intravenous administration of the PLL-dendrimer molecules into C57BL/6 mice inhibited vascularisation in Matrigel plugs implanted subcutaneously. Antiangiogenic activity was further evidenced using intravital microscopy of tumors grown within dorsal skinfold window chambers. Reduced vascularization of P22 rat sarcoma implanted in the dorsal window chamber of SCID mice was observed following tail vein administration (i.v.) of the PLL dendrimers. Also, the in vivo toxicological profile of the PLL-dendrimer molecules was shown to be safe at the dose regime studied. The antiangiogenic activity of the PLL dendrimer was further shown to be associated with significant suppression of B16F10 solid tumor volume and delayed tumor growth. Enhanced apoptosis/necrosis within tumors of PLL-dendrimer-treated animals only and reduction in the number of CD31 positive cells were observed in comparison to protamine treatment. This study suggests that PLL-dendrimer molecules can exhibit a systemic antiangiogenic activity that may be used for therapy of solid tumors, and in combination with their capacity to carry other therapeutic or diagnostic agents may potentially offer capabilities for the design of theranostic systems.}, keywords = {angiogenesis, cancer, I2CT, nanoparticle, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ben-valid_polyaniline-coated_2010, title = {Polyaniline-coated single-walled carbon nanotubes: synthesis, characterization and impact on primary immune cells}, author = {Shoshana Ben-Valid and Hélène Dumortier and Marion Décossas and Ruthy Sfez and Moreno Meneghetti and Alberto Bianco and Shlomo Yitzchaik}, url = {https://pubs.rsc.org/en/content/articlelanding/2010/jm/b921828a}, doi = {10.1039/B921828A}, issn = {1364-5501}, year = {2010}, date = {2010-03-01}, urldate = {2020-04-01}, journal = {Journal of Materials Chemistry}, volume = {20}, number = {12}, pages = {2408--2417}, abstract = {Functionalized carbon nanotubes are increasingly exploited as innovative components for the development of advanced biomedical devices. In this study we report a novel synthetic route for the formation of single-walled carbon nanotube (SWCNT)–polyaniline (PANI) hybrids by in situ chemical polymerization. The surfactant sodium dodecylsulfate (SDS) is used as a template for monomer assembly and polymerization. The resulting composite preserves the surfactant and is characterized by a tight binding between SWCNTs and PANI. Having the idea of integrating these new types of SWCNT conjugates into advanced biomedical tools (i.e. implantable multi-electrode arrays), we explored their potential impact on the viability and function of cells from the immune system. We have compared the cytotoxic effects of SWCNT-COOH, SWCNT/SDS and SWCNT/SDS/PANI on mouse spleen cells and macrophages. The results indicate that biocompatibility of the different SWCNT conjugates is dependent both on the doses used and the type of cells.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{flacher_epidermal_2010, title = {Epidermal Langerhans cells rapidly capture and present antigens from C-type lectin-targeting antibodies deposited in the dermis}, author = {Vincent Flacher and Christoph H Tripp and Patrizia Stoitzner and Bernhard Haid and Susanne Ebner and Barbara Del Frari and Franz Koch and Chae Gyu Park and Ralph M Steinman and Juliana Idoyaga and Nikolaus Romani}, doi = {10.1038/jid.2009.343}, issn = {1523-1747}, year = {2010}, date = {2010-03-01}, journal = {The Journal of Investigative Dermatology}, volume = {130}, number = {3}, pages = {755--762}, abstract = {Antigen-presenting cells can capture antigens that are deposited in the skin, including vaccines given subcutaneously. These include different dendritic cells (DCs) such as epidermal Langerhans cells (LCs), dermal DCs, and dermal langerin+ DCs. To evaluate access of dermal antigens to skin DCs, we used mAb to two C-type lectin endocytic receptors, DEC-205/CD205 and langerin/CD207. When applied to murine and human skin explant cultures, these mAbs were efficiently taken up by epidermal LCs. In addition, anti-DEC-205 targeted langerin+ CD103+ and langerin- CD103- mouse dermal DCs. Unexpectedly, intradermal injection of either mAb, but not isotype control, resulted in strong and rapid labeling of LCs in situ, implying that large molecules can diffuse through the basement membrane into the epidermis. Epidermal LCs targeted in vivo by ovalbumin-coupled anti-DEC-205 potently presented antigen to CD4+ and CD8+ T cells in vitro. However, to our surprise, LCs targeted through langerin were unable to trigger T-cell proliferation. Thus, epidermal LCs have a major function in uptake of lectin-binding antibodies under standard vaccination conditions.}, keywords = {Animals, Antibodies, antibody, Antigen, Antigen Presentation, ANTIGEN PRESENTING CELLS, Antigen-Presenting Cells, Antigens, BASEMENT MEMBRANE, C-Type, C-type lectin, CD103, CD8+ T cells, Cell Division, Cell Movement, Cells, Culture, Cultured, cytology, Dendritic Cells, DERMATOLOGY, DERMIS, Epidermal Cells, Epidermis, function, Human, Humans, Immunology, in situ, IN VITRO, In vivo, Inbred BALB C, Inbred C57BL, Injections, Intradermal, Langerhans Cells, LECTIN, Lectins, mAb, Mannose-Binding Lectins, Membrane, Mice, Monoclonal, mouse, murine, Pharmacology, Proliferation, Protein, Receptor, Skin, Surface, T CELLS, T-CELLS, T-Lymphocytes, Team-Mueller, Vaccination, vaccine, Vaccines}, pubstate = {published}, tppubtype = {article} } @article{menard-moyon_functionalized_2010, title = {Functionalized carbon nanotubes for probing and modulating molecular functions}, author = {Cécilia Ménard-Moyon and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, doi = {10.1016/j.chembiol.2010.01.009}, issn = {1879-1301}, year = {2010}, date = {2010-02-01}, journal = {Chemistry & Biology}, volume = {17}, number = {2}, pages = {107--115}, abstract = {Carbon nanotubes (CNTs) entered the domain of biological research a few years ago, creating a significant amount of interest due to their extraordinary physicochemical properties. The integration of CNT-based strategies with biology necessitates a multidisciplinary approach that requires competences in the diverse fields of chemistry, physics, and life sciences. In the biomedical domain CNTs are extensively explored as novel drug delivery systems for therapy and diagnosis. Additionally, CNTs can also be designed as new tools for modulation of molecular functions, by directly affecting various biological processes or by interaction with bioactive molecules. The aim of this review is to discuss how CNTs can be exploited as new probes for molecular functions. The different sections illustrate various applications of CNTs, including gene silencing, surface cell interactions via glycoproteins, biosensing, intracellular drug delivery using an atomic force microscopy tip-based nanoinjector, modulation of antibody/antigen interaction and enzyme activity, and blocking of ion channels.}, keywords = {Antibodies, Antigens, Atomic Force, Biosensing Techniques, carbon, Drug Delivery Systems, enzymes, Glycoproteins, I2CT, Ion Channels, Microscopy, Nanotubes, RNA, Small Interfering, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Synthesis and antimicrobial evaluation of amphiphilic neamine derivatives}, author = {I Baussanne and A Bussiere and S Halder and C Ganem-Elbaz and M Ouberai and M Riou and J M Paris and E Ennifar and M P Mingeot-Leclercq and J L Decout}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20000576}, isbn = {20000576}, year = {2010}, date = {2010-01-01}, journal = {J Med Chem}, volume = {53}, number = {1}, pages = {119-27}, abstract = {The aminoglycoside antibiotics bind to the 16S bacterial rRNA and disturb the protein synthesis. One to four hydroxyl functions of the small aminoglycoside neamine were capped with phenyl, naphthyl, pyridyl, or quinolyl rings. The 3',4'- (6), 3',6- (7a), and the 3',4',6- (10a) 2-naphthylmethylene derivatives appeared to be active against sensitive and resistant Staphylococcus aureus strains. 10a also showed marked antibacterial activities against Gram (-) bacteria, including strains expressing enzymes modifying aminoglycosides, efflux pumps, or rRNA methylases. 7a and 10a revealed a weak and aspecific binding to a model bacterial 16S rRNA. Moreover, as compared to neomycin B, 10a showed a lower ability to decrease (3)H leucine incorporation into proteins in Pseudomonas aeruginosa. All together, our results suggest that the 3',4',6-tri-2-naphthylmethylene neamine derivative 10a should act against Gram (-) bacteria through a mechanism different from inhibition of protein synthesis, probably by membrane destabilization.}, note = {1520-4804 (Electronic) 0022-2623 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cation-dependent cleavage of the duplex form of the subtype-B HIV-1 RNA dimerization initiation site}, author = {E Ennifar and P Walter and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20460458}, isbn = {20460458}, year = {2010}, date = {2010-01-01}, journal = {Nucleic Acids Res}, volume = {38}, number = {17}, pages = {5807-5811}, abstract = {The crystal structure of subtype-B HIV-1 genomic RNA Dimerization Initiation Site duplex revealed chain cleavage at a specific position resulting in 3'-phosphate and 5'-hydroxyl termini. A crystallographic analysis showed that Ba(2+), Mn(2+), Co(2+) and Zn(2+) bind specifically on a guanine base close to the cleaved position. The crystal structures also point to a necessary conformational change to induce an 'in-line' geometry at the cleavage site. In solution, divalent cations increased the rate of cleavage with pH/pKa compensation, indicating that a cation-bound hydroxide anion is responsible for the cleavage. We propose a 'Trojan horse' mechanism, possibly of general interest, wherein a doubly charged cation hosted near the cleavage site as a 'harmless' species is further transformed in situ into an 'aggressive' species carrying a hydroxide anion.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal article}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The canonical helix of urea oligomers at atomic resolution: insights into folding-induced axial organization}, author = {L Fischer and P Claudon and N Pendem and E Miclet and C Didierjean and E Ennifar and G Guichard}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20039243}, isbn = {20039243}, year = {2010}, date = {2010-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {49}, number = {6}, pages = {1067-70}, note = {1521-3773 (Electronic) 1433-7851 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of HIV-1 reverse transcriptase bound to a non-nucleoside inhibitor with a novel mechanism of action}, author = {S Freisz and G Bec and M Radi and P Wolff and E Crespan and L Angeli and P Dumas and G Maga and M Botta and E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20135654}, isbn = {20135654}, year = {2010}, date = {2010-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {49}, number = {10}, pages = {1805-8}, note = {1521-3773 (Electronic) 1433-7851 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {3-Ring/*chemistry Humans Indoles/*chemistry *Models, Anti-HIV Agents/*chemistry/therapeutic use Crystallography, Biological *Models, DUMAS, ENNIFAR, Molecular Molecular Structure Nitriles/*chemistry Pyridones/*chemistry Pyrimidines/*chemistry Reverse Transcriptase Inhibitors/*chemistry/therapeutic use Vinyl Compounds/*chemistry, Unité ARN, X-Ray HIV Protease Inhibitors/*chemistry/metabolism/pharmacology HIV Reverse Transcriptase/*chemistry/metabolism Heterocyclic Compounds}, pubstate = {published}, tppubtype = {article} } @article{, title = {The "Phantom Effect" of the Rexinoid LG100754: structural and functional insights}, author = {Y Sato and N Ramalanjaona and T Huet and N Potier and J Osz and P Antony and C Peluso-Iltis and P Poussin-Courmontagne and E Ennifar and Y Mely and A Dejaegere and D Moras and N Rochel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21152046}, doi = {10.1371/journal.pone.0015119}, isbn = {ISBN/1932-6203 (Electronic) 1932-6203 (Linking)}, year = {2010}, date = {2010-01-01}, journal = {PLoS One}, volume = {5}, number = {11}, pages = {e15119}, abstract = {Retinoic acid receptors (RARs) and Retinoid X nuclear receptors (RXRs) are ligand-dependent transcriptional modulators that execute their biological action through the generation of functional heterodimers. RXR acts as an obligate dimer partner in many signalling pathways, gene regulation by rexinoids depending on the liganded state of the specific heterodimeric partner. To address the question of the effect of rexinoid antagonists on RAR/RXR function, we solved the crystal structure of the heterodimer formed by the ligand binding domain (LBD) of the RARalpha bound to its natural agonist ligand (all-trans retinoic acid, atRA) and RXRalpha bound to a rexinoid antagonist (LG100754). We observed that RARalpha exhibits the canonical agonist conformation and RXRalpha an antagonist one with the C-terminal H12 flipping out to the solvent. Examination of the protein-LG100754 interactions reveals that its propoxy group sterically prevents the H12 associating with the LBD, without affecting the dimerization or the active conformation of RAR. Although LG100754 has been reported to act as a 'phantom ligand' activating RAR in a cellular context, our structural data and biochemical assays demonstrate that LG100754 mediates its effect as a full RXR antagonist. Finally we show that the 'phantom ligand effect' of the LG100754 is due to a direct binding of the ligand to RAR that stabilizes coactivator interactions thus accounting for the observed transcriptional activation of RAR/RXR.}, keywords = {Animals Binding Sites Binding, Competitive Fluorescence Polarization Humans Ligands Mice Models, DUMAS, ENNIFAR, Molecular Protein Binding Protein Multimerization Protein Structure, Retinoic Acid/*chemistry/genetics/metabolism Recombinant Proteins/chemistry/metabolism Retinoid X Receptors/*chemistry/genetics/metabolism Retinoids/*chemistry/metabolism Scattering, Small Angle Tetrahydronaphthalenes/*chemistry/metabolism Tretinoin/*chemistry/metabolism X-Ray Diffraction, Tertiary Receptor Cross-Talk Receptors, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{parietti_rituximab_2010, title = {Rituximab treatment overcomes reduction of regulatory iNKT cells in patients with rheumatoid arthritis}, author = {Véronique Parietti and Hélène Chifflot and Jean Sibilia and Sylviane Muller and Fanny Monneaux}, doi = {10.1016/j.clim.2009.11.007}, issn = {1521-7035}, year = {2010}, date = {2010-01-01}, journal = {Clinical Immunology (Orlando, Fla.)}, volume = {134}, number = {3}, pages = {331--339}, abstract = {Invariant natural killer T (iNKT) cells are a subset of T cells that recognize glycolipid antigens presented by the CD1d molecule. Accumulating evidences showed that iNKT cells are implicated in the regulatory mechanisms that control autoimmunity. We evaluated the number of circulating iNKT cells in patients with rheumatoid arthritis (RA) by flow cytometry and performed a longitudinal analysis of iNKT cell frequency in RA patients who were given an anti-CD20 therapy. Significantly lower iNKT cell numbers were measured in the blood from RA patients compared to healthy individuals (ptextless0.0001) and low iNKT cell frequencies were rather associated with an active disease. In RA patients who received rituximab treatment, iNKT cell number was increased in relation to the clinical outcome. We demonstrated that the number of iNKT cells is altered in RA patients and that following rituximab therapy, clinical remission of RA is associated with an increase of iNKT cell frequency.}, keywords = {Adult, Age Factors, Aged, Antibodies, Antirheumatic Agents, arthritis, Female, Flow Cytometry, Humans, I2CT, Longitudinal Studies, Male, Middle Aged, Monneaux, Monoclonal, Murine-Derived, Natural Killer T-Cells, Nonparametric, rheumatoid, Rituximab, Sex Factors, Statistics, Team-Dumortier, Young Adult}, pubstate = {published}, tppubtype = {article} } @article{baeza_garcia_involvement_2010, title = {Involvement of the cytokine MIF in the snail host immune response to the parasite Schistosoma mansoni}, author = {Alvaro Baeza Garcia and Raymond J Pierce and Benjamin Gourbal and Elisabeth Werkmeister and Dominique Colinet and Jean-Marc Reichhart and Colette Dissous and Christine Coustau}, doi = {10.1371/journal.ppat.1001115}, issn = {1553-7374}, year = {2010}, date = {2010-01-01}, journal = {PLoS Pathog.}, volume = {6}, number = {9}, pages = {e1001115}, abstract = {We have identified and characterized a Macrophage Migration Inhibitory Factor (MIF) family member in the Lophotrochozoan invertebrate, Biomphalaria glabrata, the snail intermediate host of the human blood fluke Schistosoma mansoni. In mammals, MIF is a widely expressed pleiotropic cytokine with potent pro-inflammatory properties that controls cell functions such as gene expression, proliferation or apoptosis. Here we show that the MIF protein from B. glabrata (BgMIF) is expressed in circulating immune defense cells (hemocytes) of the snail as well as in the B. glabrata embryonic (Bge) cell line that has hemocyte-like features. Recombinant BgMIF (rBgMIF) induced cell proliferation and inhibited NO-dependent p53-mediated apoptosis in Bge cells. Moreover, knock-down of BgMIF expression in Bge cells interfered with the in vitro encapsulation of S. mansoni sporocysts. Furthermore, the in vivo knock-down of BgMIF prevented the changes in circulating hemocyte populations that occur in response to an infection by S. mansoni miracidia and led to a significant increase in the parasite burden of the snails. These results provide the first functional evidence that a MIF ortholog is involved in an invertebrate immune response towards a parasitic infection and highlight the importance of cytokines in invertebrate-parasite interactions.}, keywords = {Amino Acid, Animals, Apoptosis, Biomphalaria, Blotting, Cell Proliferation, Cells, Cricetinae, Cultured, Hemocytes, Host-Parasite Interactions, Humans, Liver, M3i, Macrophage Migration-Inhibitory Factors, messenger, Oocysts, Recombinant Proteins, reichhart, Reverse Transcriptase Polymerase Chain Reaction, RNA, Schistosoma mansoni, Schistosomiasis mansoni, Sequence Homology, Small Interfering, Western}, pubstate = {published}, tppubtype = {article} } @article{paquette_caspase-mediated_2010, title = {Caspase-mediated cleavage, IAP binding, and ubiquitination: linking three mechanisms crucial for Drosophila NF-kappaB signaling}, author = {Nicholas Paquette and Meike Broemer and Kamna Aggarwal and Li Chen and Marie Husson and Deniz Ertürk-Hasdemir and Jean-Marc Reichhart and Pascal Meier and Neal Silverman}, doi = {10.1016/j.molcel.2009.12.036}, issn = {1097-4164}, year = {2010}, date = {2010-01-01}, journal = {Mol. Cell}, volume = {37}, number = {2}, pages = {172--182}, abstract = {Innate immune responses are critical for the immediate protection against microbial infection. In Drosophila, infection leads to the rapid and robust production of antimicrobial peptides through two NF-kappaB signaling pathways-IMD and Toll. The IMD pathway is triggered by DAP-type peptidoglycan, common to most Gram-negative bacteria. Signaling downstream from the peptidoglycan receptors is thought to involve K63 ubiquitination and caspase-mediated cleavage, but the molecular mechanisms remain obscure. We now show that PGN stimulation causes caspase-mediated cleavage of the imd protein, exposing a highly conserved IAP-binding motif (IBM) at its neo-N terminus. A functional IBM is required for the association of cleaved IMD with the ubiquitin E3-ligase DIAP2. Through its association with DIAP2, IMD is rapidly conjugated with K63-linked polyubiquitin chains. These results mechanistically connect caspase-mediated cleavage and K63 ubiquitination in immune-induced NF-kappaB signaling.}, keywords = {Alleles, Amino Acid Motifs, Animals, Biological, Caspases, Inhibitor of Apoptosis Proteins, M3i, MAP Kinase Kinase Kinases, Models, NF-kappa B, reichhart, Sequence Alignment, Signal Transduction, Ubiquitin-Protein Ligases, Ubiquitination}, pubstate = {published}, tppubtype = {article} } @article{pospisilik_drosophila_2010b, title = {Drosophila genome-wide obesity screen reveals hedgehog as a determinant of brown versus white adipose cell fate}, author = {Andrew J Pospisilik and Daniel Schramek and Harald Schnidar and Shane J F Cronin and Nadine T Nehme and Xiaoyun Zhang and Claude Knauf and Patrice D Cani and Karin Aumayr and Jelena Todoric and Martina Bayer and Arvand Haschemi and Vijitha Puviindran and Krisztina Tar and Michael Orthofer and Gregory G Neely and Georg Dietzl and Armen Manoukian and Martin Funovics and Gerhard Prager and Oswald Wagner and Dominique Ferrandon and Fritz Aberger and Chi-chung Hui and Harald Esterbauer and Josef M Penninger}, doi = {10.1016/j.cell.2009.12.027}, issn = {1097-4172}, year = {2010}, date = {2010-01-01}, journal = {Cell}, volume = {140}, number = {1}, pages = {148--160}, abstract = {Over 1 billion people are estimated to be overweight, placing them at risk for diabetes, cardiovascular disease, and cancer. We performed a systems-level genetic dissection of adiposity regulation using genome-wide RNAi screening in adult Drosophila. As a follow-up, the resulting approximately 500 candidate obesity genes were functionally classified using muscle-, oenocyte-, fat-body-, and neuronal-specific knockdown in vivo and revealed hedgehog signaling as the top-scoring fat-body-specific pathway. To extrapolate these findings into mammals, we generated fat-specific hedgehog-activation mutant mice. Intriguingly, these mice displayed near total loss of white, but not brown, fat compartments. Mechanistically, activation of hedgehog signaling irreversibly blocked differentiation of white adipocytes through direct, coordinate modulation of early adipogenic factors. These findings identify a role for hedgehog signaling in white/brown adipocyte determination and link in vivo RNAi-based scanning of the Drosophila genome to regulation of adipocyte cell fate in mammals.}, keywords = {Adipocytes, Adipogenesis, Animals, Brown, Brown/metabolism, Cyclic AMP, Cyclic AMP/metabolism, Drosophila Proteins/*metabolism, ferrandon, Glucocorticoids, Glucocorticoids/metabolism, Hedgehog Proteins, Hedgehog Proteins/*metabolism, Humans, Knockout, M3i, Mice, Muscle Cells, Muscle Cells/metabolism, Obesity, Obesity/*genetics, Repressor Proteins, Repressor Proteins/genetics, White, White/metabolism}, pubstate = {published}, tppubtype = {article} } @article{romani_targeting_2010, title = {Targeting of antigens to skin dendritic cells: possibilities to enhance vaccine efficacy}, author = {Nikolaus Romani and Martin Thurnher and Juliana Idoyaga and Ralph M Steinman and Vincent Flacher}, doi = {10.1038/icb.2010.39}, issn = {1440-1711}, year = {2010}, date = {2010-01-01}, journal = {Immunology and Cell Biology}, volume = {88}, number = {4}, pages = {424--430}, abstract = {Vaccinations in medicine are commonly administered through the skin. Therefore, the vaccine is immunologically processed by antigen-presenting cells of the skin. There is recent evidence that the clinically less often used intradermal route is effective; in cases even superior to the conventional subcutaneous or intramuscular route. Professional antigen-presenting cells of the skin comprise epidermal Langerhans cells (CD207/langerin(+)), dermal langerin(-) and dermal langerin(+) dendritic cells (DCs). In human skin, langerin(-) dermal DCs can be further subdivided on the basis of their reciprocal CD1a and CD14 expression. The relative contributions of these subsets to the generation of immunity or tolerance are still unclear. Langerhans cells in human skin seem to be specialized for induction of cytotoxic T lymphocytes. Likewise, mouse Langerhans cells are capable of cross-presentation and of protecting against experimental tumours. It is desirable to harness these properties for immunotherapy. A promising strategy to dramatically improve the outcome of vaccinations is 'antigen targeting'. Thereby, the vaccine is delivered directly and selectively to defined types of skin DCs. Targeting is achieved by means of coupling antigen to antibodies that recognize cell surface receptors on DCs. This approach is being widely explored. Little is known, however, about the events that take place in the skin and the DCs subsets involved therein. This topic will be discussed in this article.}, keywords = {Animals, Antibodies, antibody, Antigen, ANTIGEN PRESENTING CELLS, Antigen-Presenting Cells, Antigens, C-Type, CD, CD14, CD1a, CROSS-PRESENTATION, Dendritic Cells, DERMATOLOGY, Expression, Human, Humans, Immunity, Immunotherapy, INDUCTION, Intradermal, Langerhans Cells, Lectins, Lymphocytes, Mannose-Binding Lectins, mouse, Receptor, Skin, SUBSETS, T-Lymphocytes, Team-Mueller, tolerance, Vaccination, vaccine, Vaccines}, pubstate = {published}, tppubtype = {article} } @article{zhang_asparaginyl_2010, title = {The asparaginyl hydroxylase factor inhibiting HIF-1alpha is an essential regulator of metabolism.}, author = {Na Zhang and Zhenxing Fu and Sarah Linke and Johana Chicher and Jeffrey J Gorman and DeeAnn Visk and Gabriel G Haddad and Lorenz Poellinger and Daniel J Peet and Frank Powell and Randall S Johnson}, doi = {10.1016/j.cmet.2010.03.001}, issn = {1932-7420 1550-4131 1550-4131}, year = {2010}, date = {2010-01-01}, journal = {Cell metabolism}, volume = {11}, number = {5}, pages = {364--378}, abstract = {Factor inhibiting HIF-1alpha (FIH) is an asparaginyl hydroxylase. Hydroxylation of HIF-alpha proteins by FIH blocks association of HIFs with the transcriptional coactivators CBP/p300, thus inhibiting transcriptional activation. We have created mice with a null mutation in the FIH gene and found that it has little or no discernable role in mice in altering classical aspects of HIF function, e.g., angiogenesis, erythropoiesis, or development. Rather, it is an essential regulator of metabolism: mice lacking FIH exhibit reduced body weight, elevated metabolic rate, hyperventilation, and improved glucose and lipid homeostasis and are resistant to high-fat-diet-induced weight gain and hepatic steatosis. Neuron-specific loss of FIH phenocopied some of the major metabolic phenotypes of the global null animals: those mice have reduced body weight, increased metabolic rate, and enhanced insulin sensitivity and are also protected against high-fat-diet-induced weight gain. These results demonstrate that FIH acts to a significant degree through the nervous system to regulate metabolism.}, keywords = {alpha Subunit/*antagonists & inhibitors/genetics, Animals, Asparagine/genetics/metabolism, Dietary Fats/pharmacology, Fatty Liver/etiology, Glucose/metabolism, Hyperventilation/etiology, Hypoxia-Inducible Factor 1, Insulin/metabolism, Knockout, Lipid Metabolism, Mice, Mixed Function Oxygenases/deficiency/genetics/*metabolism, PPSE, Transcriptional Activation, Weight Gain}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 Vif binds to APOBEC3G mRNA and inhibits its translation}, author = {G Mercenne and S Bernacchi and D Richer and G Bec and S Henriet and J C Paillart and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19910370}, isbn = {19910370}, year = {2010}, date = {2010-01-01}, journal = {Nucleic Acids Res}, volume = {38}, number = {2}, pages = {633-646}, abstract = {The HIV-1 viral infectivity factor (Vif) allows productive infection of non-permissive cells (including most natural HIV-1 targets) by counteracting the cellular cytosine deaminases APOBEC-3G (hA3G) and hA3F. The Vif-induced degradation of these restriction factors by the proteasome has been extensively studied, but little is known about the translational repression of hA3G and hA3F by Vif, which has also been proposed to participate in Vif function. Here, we studied Vif binding to hA3G mRNA and its role in translational repression. Filter binding assays and fluorescence titration curves revealed that Vif tightly binds to hA3G mRNA. Vif overall binding affinity was higher for the 3'UTR than for the 5'UTR, even though this region contained at least one high affinity Vif binding site (apparent K(d) = 27 +/- 6 nM). Several Vif binding sites were identified in 5' and 3'UTRs using RNase footprinting. In vitro translation evidenced that Vif inhibited hA3G translation by two mechanisms: a main time-independent process requiring the 5'UTR and an additional time-dependent, UTR-independent process. Results using a Vif protein mutated in the multimerization domain suggested that the molecular mechanism of translational control is more complicated than a simple physical blockage of scanning ribosomes.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {3' Untranslated Regions 5' Untranslated Regions Binding Sites Cytidine Deaminase/*genetics/metabolism Humans Mutation *Protein Biosynthesis Protein Footprinting RNA, Fluorescence vif Gene Products, Human Immunodeficiency Virus/genetics/*metabolism, MARQUET, Messenger/*metabolism Spectrometry, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {5-Modified-2'-dU and 2'-dC as mutagenic anti HIV-1 proliferation agents: synthesis and activity}, author = {Y El Safadi and J C Paillart and G Laumond and A M Aubertin and A Burger and R Marquet and V Vivet-Boudou}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20112915}, isbn = {20112915}, year = {2010}, date = {2010-01-01}, journal = {J Med Chem}, volume = {53}, number = {4}, pages = {1534-1545}, abstract = {With the goal of limiting HIV-1 proliferation by increasing the mutation rate of the viral genome, we synthesized a series of pyrimidine nucleoside analogues modified in position 5 of the aglycone moiety but unmodified on the sugar part. The synthetic strategies allow us to prepare the targeted compounds directly from commercially available nucleosides. All compounds were tested for their ability to reduce HIV-1 proliferation in cell culture. Two of them (5-hydroxymethyl-2'-dU (1c) and 5-hydroxymethyl-2'-dC (2c)) displayed a moderate antiviral activity in single passage experiments. The same two compounds plus two additional ones (5-carbamoyl-2'-dU (1a) and 5-carbamoylmethyl-2'-dU (1b)) were potent inhibitors of HIV-1 RT activity in serial passage assays, in which they induced a progressive loss of HIV-1 replication. In addition, viruses collected after seven passages in the presence of 1c and 2c replicated very poorly after withdrawal of these compounds, consistent with the accumulation of deleterious mutations in the HIV-1 genome.}, note = {1520-4804 (Electronic) 0022-2623 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Anti-HIV Agents/*chemical synthesis/chemistry/pharmacology Cell Line, MARQUET, PAILLART, Tumor Cell Survival/drug effects Genome}, pubstate = {published}, tppubtype = {article} } @article{menard-moyon_alluring_2010, title = {The alluring potential of functionalized carbon nanotubes in drug discovery}, author = {Cécilia Ménard-Moyon and Enrica Venturelli and Chiara Fabbro and Cristian Samorì and Tatiana Da Ros and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, doi = {10.1517/17460441.2010.490552}, issn = {1746-0441}, year = {2010}, date = {2010-01-01}, journal = {Expert Opinion on Drug Discovery}, volume = {5}, number = {7}, pages = {691--707}, abstract = {IMPORTANCE OF THE FIELD: The possibility of carbon nanotube integration into living systems for therapeutic and diagnostic purposes has opened the way to explore their applications in drug delivery and discovery. A wide variety of chemical approaches has been developed to functionalize carbon nanotubes with therapeutic molecules towards different biomedical uses. AREAS COVERED IN THIS REVIEW: This review covers the recent advances in the development of functionalized carbon nanotubes to offer improvements for different diseases, in particular for cancer therapy. WHAT THE READER WILL GAIN: Functionalized carbon nanotubes are able to transport therapeutic agents. Targeted methodologies using carbon nanotube-based conjugates have been investigated to improve the efficacy of some drugs. The capacity of such nanomaterials to seamlessly translocate into cells with alternative various mechanisms and their pharmacokinetic properties is also discussed. TAKE HOME MESSAGE: Although at its infancy, functionalized carbon nanotubes are very promising as a new nanomedicine platform in the field of drug discovery and delivery. They have the capacity to cross biological barriers and can be eliminated via renal and/or fecal excretion. They can transport small drug molecules while maintaining - and in some cases improving - their therapeutic efficacy.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {The cspA mRNA is a thermosensor that modulates translation of the cold-shock protein CspA}, author = {A M Giuliodori and F Di Pietro and S Marzi and B Masquida and R Wagner and P Romby and C O Gualerzi and C L Pon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20129052}, isbn = {20129052}, year = {2010}, date = {2010-01-01}, journal = {Mol Cell}, volume = {37}, number = {1}, pages = {21-33}, abstract = {Cold induction of cspA, the paradigm Escherichia coli cold-shock gene, is mainly subject to posttranscriptional control, partly promoted by cis-acting elements of its transcript, whose secondary structure at 37 degrees C and at cold-shock temperature has been elucidated here by enzymatic and chemical probing. The structures, which were also validated by mutagenesis, demonstrate that cspA mRNA undergoes a temperature-dependent structural rearrangement, likely resulting from stabilization in the cold of an otherwise thermodynamically unstable folding intermediate. At low temperature, the "cold-shock" structure is more efficiently translated and somewhat less susceptible to degradation than the 37 degrees C structure. Overall, our data shed light on a molecular mechanism at the basis of the cold-shock response, indicating that cspA mRNA is able to sense temperature downshifts, adopting functionally distinct structures at different temperatures, even without the aid of trans-acting factors. Unlike with other previously studied RNA thermometers, these structural rearrangements do not result from melting of hairpin structures.}, note = {1097-4164 (Electronic) 1097-2765 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {5' Untranslated Regions Acclimatization *Cold Temperature Escherichia coli/*genetics/metabolism Escherichia coli Proteins/genetics/*physiology Gene Expression Regulation, Bacterial Heat-Shock Proteins/genetics/*physiology Models, Genetic Nucleic Acid Conformation *Protein Biosynthesis RNA, Messenger/chemistry/*physiology, ROMBY, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{partidos_immunomodulatory_2009, title = {Immunomodulatory consequences of ODN CpG-polycation complexes}, author = {Charalambos D Partidos and Johan Hoebeke and Sébastien Wieckowski and Olivier Chaloin and Alberto Bianco and Emmanuel Moreau and Jean-Paul Briand and Claude Desgranges and Sylviane Muller}, doi = {10.1016/j.ymeth.2009.03.005}, issn = {1095-9130}, year = {2009}, date = {2009-12-01}, journal = {Methods (San Diego, Calif.)}, volume = {49}, number = {4}, pages = {328--333}, abstract = {Immunostimulatory ODN CpGs have extensively been tested as adjuvants and immunotherapeutics and hold a lot of promise for human use. In our studies we took advantage of their negative charge to study their biological activities after being complexed with carbon nanotubes, a novel vector for vaccine delivery and Tat protein of HIV, a target protein for therapeutic or prophylactic intervention. In the case of carbon nanotubes, ODN CpGs were able to form stable complexes based on charge interaction and exert increased immunostimulatory activity in vitro. With regard to the Tat protein, ODN CpGs were shown to bind effectively through the basic domain of the protein representing residues 44-61. Moreover, using surface Plasmon Resonance Technology and an in vitro cellular system, ODN CpGs were shown to inhibit the interaction of Tat protein with the transactivation responsive element, a bulged RNA hairpin structure. However, when ODN CpGs were complexed with Tat they readily increased the apoptotic properties of this protein as studied in CD3-stimulated Jurkat cells. Overall, our findings together with published data support the view that for harnessing the beneficial effects of ODN CpGs a careful consideration has to be given depending on the target intervention.}, keywords = {Animals, carbon, CpG Islands, Humans, I2CT, Immunologic Factors, Nanotubes, Oligodeoxyribonucleotides, Polyamines, Team-Bianco, Transcriptional Activation}, pubstate = {published}, tppubtype = {article} } @article{shia_toll-dependent_2009, title = {Toll-dependent antimicrobial responses in Drosophila larval fat body require Spätzle secreted by haemocytes}, author = {Alice K H Shia and Marcus Glittenberg and Gavin Thompson and Alexander N R Weber and Jean-Marc Reichhart and Petros Ligoxygakis}, doi = {10.1242/jcs.049155}, issn = {1477-9137}, year = {2009}, date = {2009-12-01}, journal = {J. Cell. Sci.}, volume = {122}, number = {Pt 24}, pages = {4505--4515}, abstract = {In Drosophila, the humoral response characterised by the synthesis of antimicrobial peptides (AMPs) in the fat body (the equivalent of the mammalian liver) and the cellular response mediated by haemocytes (blood cells) engaged in phagocytosis represent two major reactions that counter pathogens. Although considerable analysis has permitted the elucidation of mechanisms pertaining to the two responses individually, the mechanism of their coordination has been unclear. To characterise the signals with which infection might be communicated between blood cells and fat body, we ablated circulating haemocytes and defined the parameters of AMP gene activation in larvae. We found that targeted ablation of blood cells influenced the levels of AMP gene expression in the fat body following both septic injury and oral infection. Expression of the AMP gene drosomycin (a Toll target) was blocked when expression of the Toll ligand Spätzle was knocked down in haemocytes. These results show that in larvae, integration of the two responses in a systemic reaction depend on the production of a cytokine (spz), a process that strongly parallels the mammalian immune response.}, keywords = {Animals, Enterococcus faecalis, Escherichia coli, Fat Body, Hemocytes, Larva, M3i, reichhart, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{hetru_nf-kappab_2009, title = {NF-kappaB in the immune response of Drosophila}, author = {Charles Hetru and Jules A Hoffmann}, doi = {10.1101/cshperspect.a000232}, issn = {1943-0264}, year = {2009}, date = {2009-12-01}, journal = {Cold Spring Harb Perspect Biol}, volume = {1}, number = {6}, pages = {a000232}, abstract = {The nuclear factor kappaB (NF-kappaB) pathways play a major role in Drosophila host defense. Two recognition and signaling cascades control this immune response. The Toll pathway is activated by Gram-positive bacteria and by fungi, whereas the immune deficiency (Imd) pathway responds to Gram-negative bacterial infection. The basic mechanisms of recognition of these various types of microbial infections by the adult fly are now globally understood. Even though some elements are missing in the intracellular pathways, numerous proteins and interactions have been identified. In this article, we present a general picture of the immune functions of NF-kappaB in Drosophila with all the partners involved in recognition and in the signaling cascades.}, keywords = {Animals, bacteria, Fungi, Gene Expression Regulation, hoffmann, M3i, NF-kappa B}, pubstate = {published}, tppubtype = {article} } @article{nicolau_proteomic_2009, title = {Proteomic investigation of enzymes involved in 2-ethylhexyl nitrate biodegradation in Mycobacterium austroafricanum IFP 2173.}, author = {Elodie Nicolau and Lauriane Kuhn and Rémy Marchal and Yves Jouanneau}, doi = {10.1016/j.resmic.2009.09.017}, issn = {1769-7123 0923-2508}, year = {2009}, date = {2009-12-01}, journal = {Research in microbiology}, volume = {160}, number = {10}, pages = {838--847}, abstract = {2-Ethyhexyl nitrate (2-EHN) is a synthetic chemical used as a diesel fuel additive, which is recalcitrant to biodegradation. In this study, the enzymes involved in 2-EHN degradation were investigated in Mycobacterium austroafricanum IFP 2173. Using two-dimensional gel electrophoresis and a shotgun proteomic approach, a total of 398 proteins appeared to be more abundant in cells exposed to 2-EHN than in acetate-grown cells. This set of proteins includes multiple isoenzymes of the beta-oxidation pathway, two alcohol and one aldehyde dehydrogenase, as well as four cytochromes P450, including one CYP153 which functions as an alkane hydroxylase. Strain IFP 2173 was also found to contain two alkB-like genes encoding putative membrane-bound alkane hydroxylases. RT-PCR experiments showed that the gene encoding the CYP153 protein, as well as alkB genes, were expressed on 2-EHN. These findings are discussed in the light of a recently proposed 2-EHN degradation pathway involving an initial attack by an alkane hydroxylase and one turn of beta-oxidation, leading to the accumulation of a gamma-lactone as a dead-end product.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{G2009, title = {Focusing on complement in the antiparasitic defense of mosquitoes}, author = {Gloria Volohonsky and Stefanie Steinert and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/19853513}, year = {2009}, date = {2009-10-21}, journal = {Trends Parasitol.}, volume = {26}, number = {1}, pages = {1-3}, abstract = {Malaria is an infectious disease caused by Plasmodium and transmitted to humans by the Anopheles mosquitoes. The mosquito immune system predominantly targets Plasmodium at the ookinete stage, and efficiently eliminates the majority of invading parasites. Identification of the components of the mosquito complement system now provides new focus for studies on the activation and control of this pathway, whose manipulation is expected to block malaria transmission at the vector level.}, keywords = {ookinete}, pubstate = {published}, tppubtype = {article} } @article{SA2009, title = {Dissecting the genetic basis of resistance to malaria parasites in Anopheles gambiae}, author = {Stéphanie A Blandin and R Wang-Sattler and Marina Lamacchia and Julien Gagneur and G Lycett and Y Ning and Elena A Levashina and Lars M Steinmetz}, url = {http://www.ncbi.nlm.nih.gov/pubmed/19797663}, year = {2009}, date = {2009-10-02}, journal = {Science}, volume = {326}, number = {5949}, pages = {147-50}, abstract = {The ability of Anopheles gambiae mosquitoes to transmit Plasmodium parasites is highly variable between individuals. However, the genetic basis of this variability has remained unknown. We combined genome-wide mapping and reciprocal allele-specific RNA interference (rasRNAi) to identify the genomic locus that confers resistance to malaria parasites and demonstrated that polymorphisms in a single gene encoding the antiparasitic thioester-containing protein 1 (TEP1) explain a substantial part of the variability in parasite killing. The link between TEP1 alleles and resistance to malaria may offer new tools for controlling malaria transmission. The successful application of rasRNAi in Anopheles suggests that it could also be applied to other organisms where RNAi is feasible to dissect complex phenotypes to the level of individual quantitative trait alleles.}, keywords = {blandin, M3i, TEP1}, pubstate = {published}, tppubtype = {article} } @article{zacchigna_vitro_2009, title = {In vitro behavior of multifunctionalized fullerene-warfarin conjugates}, author = {Marina Zacchigna and Cedric Klumpp and Maurizio Prato and Alberto Bianco}, doi = {10.1166/jnn.2009.1551}, issn = {1533-4880}, year = {2009}, date = {2009-10-01}, journal = {Journal of Nanoscience and Nanotechnology}, volume = {9}, number = {10}, pages = {6210--6221}, abstract = {In this study we have covalently linked the anticoagulant warfarin to polyfunctionalized fullerenes. The objective was to explore the possibility of modifying the biological profile of a drug by covalent binding to functionalized fullerene. We have chosen warfarin as a model compound because it a widely used drug. We have analyzed the stability in vitro of the conjugates and found that the drug is released from the carbon support only after incubation in mouse plasma.}, keywords = {Animals, Anticoagulants, Fullerenes, I2CT, In Vitro Techniques, Mice, Spectrophotometry, Team-Bianco, Ultraviolet, Warfarin}, pubstate = {published}, tppubtype = {article} } @article{kostarelos_promises_2009, title = {Promises, facts and challenges for carbon nanotubes in imaging and therapeutics}, author = {K Kostarelos and A Bianco and M Prato}, doi = {10.1038/nnano.2009.241}, issn = {1748-3395}, year = {2009}, date = {2009-10-01}, journal = {Nature Nanotechnology}, volume = {4}, number = {10}, pages = {627--633}, abstract = {The use of carbon nanotubes in medicine is now at the crossroads between a proof-of-principle concept and an established preclinical candidate for a variety of therapeutic and diagnostic applications. Progress towards clinical trials will depend on the outcomes of efficacy and toxicology studies, which will provide the necessary risk-to-benefit assessments for carbon-nanotube-based materials. Here we focus on carbon nanotubes that have been studied in preclinical animal models, and draw attention to the promises, facts and challenges of these materials as they transition from research to the clinical phase. We address common questions regarding the use of carbon nanotubes in disease imaging and therapy, and highlight the opportunities and challenges ahead.}, keywords = {Animals, carbon, Diagnostic Imaging, Drug Evaluation, Humans, I2CT, Nanomedicine, Nanotubes, Preclinical, Team-Bianco, therapeutics}, pubstate = {published}, tppubtype = {article} } @article{chamouard_diminution_2009, title = {Diminution of Circulating CD4+CD25 high Ŧ cells in naïve Crohn's disease}, author = {Patrick Chamouard and Fanny Monneaux and Zoe Richert and Anne-Claire Voegeli and Thomas Lavaux and Marie Pierre Gaub and René Baumann and Pierre Oudet and Sylviane Muller}, doi = {10.1007/s10620-008-0590-6}, issn = {1573-2568}, year = {2009}, date = {2009-10-01}, journal = {Digestive Diseases and Sciences}, volume = {54}, number = {10}, pages = {2084--2093}, abstract = {Crohn's disease is considered to be caused either by an excess of T-cell effector functions and/or by a defective regulatory T-cell compartment. The aim of this study was to assess in Crohn's disease the frequency of circulating CD4(+)CD25(high) T cells that possess regulatory T-cell functions and CD4(+)CD25(low) T cells that contain activated T cells. Flow cytometry of peripheral blood was used to assess CD4(+)CD25(high) and CD4(+)CD25(low) T-cell frequencies in a cohort of 66 patients with Crohn's disease in comparison to 19 patients with ulcerative colitis and 31 healthy individuals enrolled as controls. The CD4(+)CD25(high) T-cell frequency was significantly lowered in naïve Crohn's disease (P = 0.013) and in ulcerative colitis (P = 0.001). CD4(+)CD25(low) T-cell frequency was increased in Crohn's disease (P = 0.0001) and in ulcerative colitis (P = 0.0002). Both CD4(+)CD25(high) and CD4(+)CD25(low) T-cell frequencies are altered in naïve Crohn's disease resulting in an imbalance between both populations and a relative contraction of the CD4(+)CD25(high) T-cell population.}, keywords = {Adult, Aged, Blood Cell Count, CD4 Antigens, Colitis, Crohn Disease, Female, Flow Cytometry, Humans, I2CT, Interleukin-2 Receptor alpha Subunit, Lymphocyte Subsets, Male, Middle Aged, Monneaux, Regulatory, T-Lymphocytes, Team-Dumortier, Ulcerative}, pubstate = {published}, tppubtype = {article} } @article{mishima_n-terminal_2009, title = {The N-terminal domain of Drosophila Gram-negative binding protein 3 (GNBP3) defines a novel family of fungal pattern recognition receptors}, author = {Yumiko Mishima and Jessica Quintin and Vishukumar Aimanianda and Christine Kellenberger and Franck Coste and Cecile Clavaud and Charles Hetru and Jules A Hoffmann and Jean-Paul Latgé and Dominique Ferrandon and Alain Roussel}, doi = {10.1074/jbc.M109.034587}, issn = {1083-351X}, year = {2009}, date = {2009-10-01}, journal = {J. Biol. Chem.}, volume = {284}, number = {42}, pages = {28687--28697}, abstract = {Gram-negative binding protein 3 (GNBP3), a pattern recognition receptor that circulates in the hemolymph of Drosophila, is responsible for sensing fungal infection and triggering Toll pathway activation. Here, we report that GNBP3 N-terminal domain binds to fungi upon identifying long chains of beta-1,3-glucans in the fungal cell wall as a major ligand. Interestingly, this domain fails to interact strongly with short oligosaccharides. The crystal structure of GNBP3-Nter reveals an immunoglobulin-like fold in which the glucan binding site is masked by a loop that is highly conserved among glucan-binding proteins identified in several insect orders. Structure-based mutagenesis experiments reveal an essential role for this occluding loop in discriminating between short and long polysaccharides. The displacement of the occluding loop is necessary for binding and could explain the specificity of the interaction with long chain structured polysaccharides. This represents a novel mechanism for beta-glucan recognition.}, keywords = {Animals, beta-Glucans, Bombyx, Carrier Proteins, Crystallography, ferrandon, Fungal Proteins, Hemolymph, hoffmann, ligands, M3i, Molecular Conformation, Mutagenesis, Polysaccharides, Protein Structure, Secondary, Tertiary, X-Ray}, pubstate = {published}, tppubtype = {article} } @article{singh_synthesis_2009, title = {Synthesis and characterization of nucleobase-carbon nanotube hybrids}, author = {Prabhpreet Singh and Jitendra Kumar and Francesca Maria Toma and Jesus Raya and Maurizio Prato and Bruno Fabre and Sandeep Verma and Alberto Bianco}, doi = {10.1021/ja905041b}, issn = {1520-5126}, year = {2009}, date = {2009-09-01}, journal = {Journal of the American Chemical Society}, volume = {131}, number = {37}, pages = {13555--13562}, abstract = {We report the synthesis and characterization of adenine-single-walled carbon nanotube (SWCNT) hybrid materials, where for the first time nucleobases are covalently attached to the exosurface of SWCNTs. The structural properties of all hybrids have been characterized using usual spectroscopic and microscopic techniques. The degree of functional groups for functionalized SWCNTs (f-SWCNTs) 2a and 2b is one adenine group for each 26 and 37 carbon atoms, respectively. Solid-state magic angle spinning (13)C NMR spectroscopy (MAS NMR) and electrochemistry have been also applied for the characterization of these f-SWCNTs. AFM images of f-SWCNT 2b showed an interesting feature of horizontally aligned nanotubes along the surface when deposited on highly oriented pyrolytic graphite surface. Furthermore, we evaluated the coordinating ability of these hybrid materials toward silver ions, and interestingly, we found a pattern of silver nanoparticles localized over the surface of the carbon nanotube network. The presence of aligned and randomly oriented CNTs and their ability to coordinate with metal ions make this class of materials very interesting for applications in the development of novel electronic devices and as new supports for different catalytic transformations.}, keywords = {Adenine, Amides, Amines, Biosensing Techniques, carbon, Catalysis, Electrochemistry, Graphite, I2CT, Magnetic Resonance Spectroscopy, Nanotubes, Nanowires, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_host_2009b, title = {Host tolerance versus resistance and microbial virulence in the host-pathogen equation}, author = {Dominique Ferrandon}, doi = {10.1016/j.chom.2009.08.010}, issn = {1934-6069}, year = {2009}, date = {2009-09-01}, journal = {Cell Host Microbe}, volume = {6}, number = {3}, pages = {203--205}, abstract = {To deal with an infection, the organism resorts to nonmutually exclusive strategies: resistance, that is, neutralization or destruction of the pathogen; or tolerance, the ability to withstand damages inflicted by the pathogen or by host defense. In this issue of Cell Host & Microbe, Shinzawa et al. (2009) identify p38-mediated phagocytic encapsulation as a potential tolerance mechanism.}, keywords = {Animals, ferrandon, Host-Pathogen Interactions, Immune Tolerance, M3i, Salmonella typhimurium, Virulence}, pubstate = {published}, tppubtype = {article} } @article{singh_organic_2009, title = {Organic functionalisation and characterisation of single-walled carbon nanotubes}, author = {Prabhpreet Singh and Stéphane Campidelli and Silvia Giordani and Davide Bonifazi and Alberto Bianco and Maurizio Prato}, doi = {10.1039/b518111a}, issn = {1460-4744}, year = {2009}, date = {2009-08-01}, journal = {Chemical Society Reviews}, volume = {38}, number = {8}, pages = {2214--2230}, abstract = {Since carbon nanotubes (CNTs) display unique structures and remarkable physical properties, a variety of applications have emerged in both materials and life sciences. In terms of applications, the functionalisation of nanotubes is extremely important, as it increases their solubility and processability, and combines the unique properties of single-walled carbon nanotubes (SWCNTs) with those of other classes of materials. A number of methods have been developed, which can be divided into two major approaches: (1) non-covalent supramolecular modifications, and (2) covalent functionalisation. In this tutorial review, we survey the covalent modification of SWCNTs with organic moieties, and illustrate the major analytical techniques routinely used to characterise the functionalised materials.}, keywords = {Alkylation, carbon, Esterification, Free Radicals, Halogenation, I2CT, Microscopy, Nanotubes, Oxidation-Reduction, Spectrum Analysis, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{herrero_synthesis_2009, title = {Synthesis and characterization of a carbon nanotube-dendron series for efficient siRNA delivery}, author = {Antonia M Herrero and Francesca M Toma and Khuloud T Al-Jamal and Kostas Kostarelos and Alberto Bianco and Tatiana Da Ros and Fouzia Bano and Loredana Casalis and Giacinto Scoles and Maurizio Prato}, doi = {10.1021/ja903316z}, issn = {1520-5126}, year = {2009}, date = {2009-07-01}, journal = {Journal of the American Chemical Society}, volume = {131}, number = {28}, pages = {9843--9848}, abstract = {A new series of dendron-functionalized multiwalled carbon nanotube (MWNT) derivatives, characterized by the presence of numerous positively charged tetraalkyl ammonium salts at the periphery of the dendron, has been synthesized. The positive charges on the MWNT surface, coupled with the unique ability of carbon nanotubes (CNTs) to penetrate cell membranes, make the new derivatives potentially ideal vectors for siRNA delivery. Using a fluorescently labeled, noncoding siRNA sequence, we demonstrate that cytoplasmic delivery of the nucleic acid is remarkably increased throughout the different dendron generations. The work reported here highlights the fact that dendron-functionalized CNTs can be rationally designed as efficient carriers of siRNA that can eventually lead to gene silencing.}, keywords = {Acrylates, Animals, Azo Compounds, Biological Transport, carbon, Cytoplasm, Dendrimers, Drug Carriers, Ethylenediamines, Gene Silencing, HeLa Cells, Humans, I2CT, Nanotubes, Polyamines, RNA, Small Interfering, Solubility, Team-Bianco, Thiosemicarbazones, Transfection, water}, pubstate = {published}, tppubtype = {article} } @article{marega_diffusion-ordered_2009, title = {Diffusion-ordered NMR spectroscopy in the structural characterization of functionalized carbon nanotubes}, author = {Riccardo Marega and Vincent Aroulmoji and Francesca Dinon and Lisa Vaccari and Silvia Giordani and Alberto Bianco and Erminio Murano and Maurizio Prato}, doi = {10.1021/ja902728w}, issn = {1520-5126}, year = {2009}, date = {2009-07-01}, journal = {Journal of the American Chemical Society}, volume = {131}, number = {25}, pages = {9086--9093}, abstract = {The emerging applications of functionalized carbon nanotubes (CNTs) in various research domains necessitate the use of many different analytical techniques to confirm their structural modifications in a fast and reliable manner. Thus far, NMR spectroscopy has not been among the main tools for characterization of organically modified carbon nanostructures. (1)H analysis is limited because the signals in these derivatives are typically weak and broad, resulting in uncertainties of a few parts per million, and because of the strong interference of residual solvent signals. To overcome these limitations, we investigated the applicability of proton NMR spectroscopy based on gradient-edited diffusion pulse sequences (1D diffusion-ordered spectroscopy, DOSY) in the characterization of CNT derivatives. In general, diffusion NMR experiments allow the separation of NMR signals of different species present in a mixture, according to their own diffusion coefficients, merging spectroscopy information with size analysis. In the present study, a selected set of CNT derivatives was synthesized and analyzed using 1D DOSY experiments by applying strong magnetic field gradients (up to 42.6 G cm(-1)). Colorimetric tests (i.e., Kaiser test) and TGA analysis support the NMR findings, which are related to isolated and/or bundled short SWNTs, on the basis of TEM and AFM characterization. The overall results show that the diffusion-based NMR spectroscopy is a fast and promising approach for the characterization of covalently modified CNT derivatives.}, keywords = {carbon, Diffusion, I2CT, Magnetic Resonance Spectroscopy, Nanotubes, Oxidation-Reduction, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bosisio_ultrasound_2009, title = {Ultrasound biomicroscopy: a powerful tool probing murine lymph node size in vivo}, author = {M R Bosisio and C Maisonneuve and S Gregoire and A Kettaneh and C G Mueller and S L Bridal}, year = {2009}, date = {2009-07-01}, journal = {Ultrasound Med.Biol.}, volume = {35}, number = {1879-291X (Electronic)}, pages = {1209--1216}, abstract = {Invasive cell-counting in lymph node (LN) is the current reference to assess LN changes due to inflammation, immunodeficiency and cancer in murine models. This work evaluates whether ultrasound biomicroscopy (UBM) can measure LN size alterations noninvasively for a large range of sizes (0.1 mm3 to 22 mm3). Correlation was assessed (rho = 0.91, p textless 0.0001) between invasive cell count and LN volume estimated with UBM (24, 2 to 28-week-old, C57BL/6 mice; 13 same-strain, transgenic mice presenting LN hyperplasia). UBM LN modification screening was applied in a skin-graft rejection model and compared with cell-counting (15 mice). UBM LN-size follow-up with fine temporal sampling was demonstrated from 9 d of age (minimum area 0.13 mm2). Reliability (intraclass correlation coefficient [ICC] textgreater 0.84) and variability of UBM evaluations compared favourably with invasive cell count. UBM provides a noninvasive alternative to cell-counting in mice for early detection and longitudinal screening of LN modifications. This can enable significant reduction in the number of mice and exploration of LNs that would be too small to dissect for cell count}, keywords = {Acoustic, Animals, Axilla, cancer, Cell Count, Female, Graft Rejection, Hyperplasia, immunodeficiency, In vivo, Inbred C57BL, inflammation, LYMPH, LYMPH NODE, Lymph Nodes, Male, methods, Mice, Microscopy, murine, Observer Variation, pathology, SKIN GRAFT, Skin Transplantation, Team-Mueller, transgenic, TRANSGENIC MICE, ultrasonography}, pubstate = {published}, tppubtype = {article} } @article{atteia_proteomic_2009, title = {A proteomic survey of Chlamydomonas reinhardtii mitochondria sheds new light on the metabolic plasticity of the organelle and on the nature of the alpha-proteobacterial mitochondrial ancestor.}, author = {Ariane Atteia and Annie Adrait and Sabine Brugière and Marianne Tardif and Robert van Lis and Oliver Deusch and Tal Dagan and Lauriane Kuhn and Brigitte Gontero and William Martin and Jérôme Garin and Jacques Joyard and Norbert Rolland}, doi = {10.1093/molbev/msp068}, issn = {1537-1719 0737-4038}, year = {2009}, date = {2009-07-01}, journal = {Molecular biology and evolution}, volume = {26}, number = {7}, pages = {1533--1548}, abstract = {Mitochondria play a key role in the life and death of eukaryotic cells, yet the full spectrum of mitochondrial functions is far from being fully understood, especially in photosynthetic organisms. To advance our understanding of mitochondrial functions in a photosynthetic cell, an extensive proteomic survey of Percoll-purified mitochondria from the metabolically versatile, hydrogen-producing green alga Chlamydomonas reinhardtii was performed. Different fractions of purified mitochondria from Chlamydomonas cells grown under aerobic conditions were analyzed by nano-liquid chromatography-electrospray ionization-mass spectrometry after protein separation on sodium dodecyl sulfate polyacrylamide gel electrophoresis or on blue-native polyacrylamide gel electrophoresis. Of the 496 nonredundant proteins identified, 149 are known or predicted to reside in other cellular compartments and were thus excluded from the molecular and evolutionary analyses of the Chlamydomonas proteome. The mitochondrial proteome of the photosynthetic alga reveals important lineage-specific differences with other mitochondrial proteomes, reflecting the high metabolic diversity of the organelle. Some mitochondrial metabolic pathways in Chlamydomonas appear to combine typical mitochondrial enzymes and bacterial-type ones, whereas others are unknown among mitochondriate eukaryotes. The comparison of the Chlamydomonas proteins to their identifiable homologs predicted from 354 sequenced genomes indicated that Arabidopsis is the most closely related nonalgal eukaryote. Furthermore, this phylogenomic analysis shows that free-living alpha-proteobacteria from the metabolically versatile orders Rhizobiales and Rhodobacterales better reflect the gene content of the ancestor of the chlorophyte mitochondria than parasitic alpha-proteobacteria with reduced and specialized genomes.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{kraut_peptide_2009, title = {Peptide storage: are you getting the best return on your investment? Defining optimal storage conditions for proteomics samples.}, author = {Alexandra Kraut and Marlène Marcellin and Annie Adrait and Lauriane Kuhn and Mathilde Louwagie and Sylvie Kieffer-Jaquinod and Dorothée Lebert and Christophe D Masselon and Alain Dupuis and Christophe Bruley and Michel Jaquinod and Jérôme Garin and Maighread Gallagher-Gambarelli}, doi = {10.1021/pr900095u}, issn = {1535-3893 1535-3893}, year = {2009}, date = {2009-07-01}, journal = {Journal of proteome research}, volume = {8}, number = {7}, pages = {3778--3785}, abstract = {To comply with current proteomics guidelines, it is often necessary to analyze the same peptide samples several times. Between analyses, the sample must be stored in such a way as to conserve its intrinsic properties, without losing either peptides or signal intensity. This article describes two studies designed to define the optimal storage conditions for peptide samples between analyses. With the use of a label-free strategy, peptide conservation was compared over a 28-day period in three different recipients: standard plastic tubes, glass tubes, and low-adsorption plastic tubes. The results of this study showed that standard plastic tubes are unsuitable for peptide storage over the period studied. Glass tubes were found to perform better than standard plastic, but optimal peptide recovery was achieved using low-adsorption plastic tubes. The peptides showing poor recovery following storage were mainly hydrophobic in nature. The differences in peptide recovery between glass and low-adsorption plastic tubes were further studied using isotopically labeled proteins. This study allowed accurate comparison of peptide recovery between the two tube types within the same LC-MS run. The results of the label-free study were confirmed. Further, it was possible to demonstrate that peptide recovery in low-adsorption plastic tubes was optimal whatever the peptide concentration stored.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{flacher_targeting_2009, title = {Targeting of epidermal Langerhans cells with antigenic proteins: attempts to harness their properties for immunotherapy}, author = {Vincent Flacher and Florian Sparber and Christoph H Tripp and Nikolaus Romani and Patrizia Stoitzner}, doi = {10.1007/s00262-008-0563-9}, issn = {1432-0851}, year = {2009}, date = {2009-07-01}, journal = {Cancer immunology, immunotherapy: CII}, volume = {58}, number = {7}, pages = {1137--1147}, abstract = {Langerhans cells, a subset of skin dendritic cells in the epidermis, survey peripheral tissue for invading pathogens. In recent functional studies it was proven that Langerhans cells can present exogenous antigen not merely on major histocompatibility complexes (MHC)-class II molecules to CD4+ T cells, but also on MHC-class I molecules to CD8+ T cells. Immune responses against topically applied antigen could be measured in skin-draining lymph nodes. Skin barrier disruption or co-application of adjuvants was required for maximal induction of T cell responses. Cytotoxic T cells induced by topically applied antigen inhibited tumor growth in vivo, thus underlining the potential of Langerhans cells for immunotherapy. Here we review recent work and report novel observations relating to the potential use of Langerhans cells for immunotherapy. We investigated the potential of epicutaneous immunization strategies in which resident skin dendritic cells are loaded with tumor antigen in situ. This contrasts with current clinical approaches, where dendritic cells generated from progenitors in blood are loaded with tumor antigen ex vivo before injection into cancer patients. In the current study, we applied either fluorescently labeled protein antigen or targeting antibodies against DEC-205/CD205 and langerin/CD207 topically onto barrier-disrupted skin and examined antigen capture and transport by Langerhans cells. Protein antigen could be detected in Langerhans cells in situ, and they were the main skin dendritic cell subset transporting antigen during emigration from skin explants. Potent in vivo proliferative responses of CD4+ and CD8+ T cells were measured after epicutaneous immunization with low amounts of protein antigen. Targeting antibodies were mainly transported by langerin+ migratory dendritic cells of which the majority represented migratory Langerhans cells and a smaller subset the new langerin+ dermal dendritic cell population located in the upper dermis. The preferential capture of topically applied antigen by Langerhans cells and their ability to induce potent CD4+ and CD8+ T cell responses emphasizes their potential for epicutaneous immunization strategies.}, keywords = {Active, Animals, Antibodies, antibody, Antigen, Antigens, BLOOD, C-Type, cancer, CD, CD4-Positive T-Lymphocytes, CD4+ T cells, CD8-Positive T-Lymphocytes, CD8+ T cells, Dendritic Cells, DERMATOLOGY, DERMIS, Epidermis, Growth, Human, Humans, immune response, IMMUNE-RESPONSES, Immunization, Immunology, Immunotherapy, in situ, In vivo, Inbred BALB C, Inbred C57BL, INDUCTION, Langerhans Cells, LECTIN, Lectins, LYMPH, LYMPH NODE, Lymph Nodes, Major Histocompatibility Complex, Mannose-Binding Lectins, metabolism, methods, MHC class I, MHC class I molecules, Mice, Neoplasm, Neoplasms, OVALBUMIN, Patients, PROGENITORS, Protein, Proteins, RESPONSES, review, Skin, T CELLS, T-CELLS, Team-Mueller, therapy, tumor}, pubstate = {published}, tppubtype = {article} } @article{hamrita_proteomics-based_2009, title = {Proteomics-based identification of alpha1-antitrypsin and haptoglobin precursors as novel serum markers in infiltrating ductal breast carcinomas.}, author = {Bechr Hamrita and Karim Chahed and Mounir Trimeche and Christelle Lemaitre Guillier and Philippe Hammann and Anouar Chaïeb and Sadok Korbi and Lotfi Chouchane}, doi = {10.1016/j.cca.2009.03.033}, issn = {1873-3492 0009-8981}, year = {2009}, date = {2009-06-01}, journal = {Clinica chimica acta; international journal of clinical chemistry}, volume = {404}, number = {2}, pages = {111--118}, abstract = {BACKGROUND: The identification of pathological markers of breast cancer for either diagnosis, treatment response or for survival is of critical importance. METHODS: Serum protein profiling using 2-DE separations coupled to matrix-assisted laser desorption ionization mass spectrometry has been used to explore protein alterations in patients with infiltrating ductal breast carcinomas (IDCA). Sera from 39 breast cancer patients and 40 healthy controls were selected for screening study using 2-DE combined with MS. The protein expression patterns obtained after the depletion of high abundance proteins was determined by coomassie blue G-250 stain after 2-DE electrophoresis. RESULTS: Six proteins that expressed differentially in the IDCA group were found. The expression levels of four isoforms corresponding to haptoglobin precursor and two isoforms of alpha1-antitrypsin precursor (alpha1-AT) were upregulated in sera from breast cancer patients. There was an increased expression of both proteins in the sera of patients with various tumor stages (I, II, III) in comparison to healthy women. Applying immunohistochemistry, we further validated alpha1-AT immunoreactivity in 51 formalin-fixed paraffin-embedded sections of breast tumors. Enhanced expression of alpha1-AT like activity has been found in IDCA breast tumors, as well as, in different histological types of breast cancer. No significant association has been found with lymph node occurrence, while in high tumor categories a tendency to an increased expression of alpha1-AT has been found, thereby suggesting a possible role of this protein in tumor growth. CONCLUSIONS: These proteins may constitute new and useful markers of breast cancer that offer a clue to a better understanding of inflammatory pathways and carcinogenesis events linked to breast cancer progression.}, note = {Place: Netherlands}, keywords = {80 and over, Adult, Aged, alpha 1-Antitrypsin/*blood, Amino Acid Sequence, Biomarkers, Breast Neoplasms/blood/*pathology, Carcinoma, Ductal/blood/*pathology, Electrophoresis, Female, Gel, Haptoglobins/*analysis, Humans, Mass, Matrix-Assisted Laser Desorption-Ionization, Middle Aged, Molecular Sequence Data, PPSE, Protein Isoforms/blood, proteomics, Spectrometry, Tumor/*blood, Two-Dimensional}, pubstate = {published}, tppubtype = {article} } @article{podesta_antitumor_2009, title = {Antitumor activity and prolonged survival by carbon-nanotube-mediated therapeutic siRNA silencing in a human lung xenograft model}, author = {Jennifer E Podesta and Khuloud T Al-Jamal and Antonia M Herrero and Bowen Tian and Hanene Ali-Boucetta and Vikas Hegde and Alberto Bianco and Maurizio Prato and Kostas Kostarelos}, doi = {10.1002/smll.200801572}, issn = {1613-6829}, year = {2009}, date = {2009-05-01}, journal = {Small (Weinheim an Der Bergstrasse, Germany)}, volume = {5}, number = {10}, pages = {1176--1185}, abstract = {Carbon nanotubes are novel nanomaterials that are thought to offer potential benefits to a variety of biomedical and clinical applications. In this study, the treatment of a human lung carcinoma model in vivo using siRNA sequences leading to cytotoxicity and cell death is carried out using either cationic liposomes (DOTAP:cholesterol) or amino-functionalized multi-walled carbon nanotubes (MWNT - NH(+)(3)). Validation for the most cytotoxic siRNA sequence using a panel of human carcinoma and murine cells reveals that the proprietary siTOX sequence is human specific and can lead to significant cytotoxic activities delivered both by liposome or MWNT - NH(+)(3) in vitro. A comparative study using both types of vector indicates that only MWNT - NH(+)(3):siRNA complexes administered intratumorally can elicit delayed tumor growth and increased survival of xenograft-bearing animals. siTOX delivery via the cationic MWNT - NH(+)(3) is biologically active in vivo by triggering an apoptotic cascade, leading to extensive necrosis of the human tumor mass. This suggests that carbon-nanotube-mediated delivery of siRNA by intratumoral administration leads to successful and statistically significant suppression of tumor volume, followed by a concomitant prolongation of survival of human lung tumor-bearing animals. The direct comparison between carbon nanotubes and liposomes demonstrates the potential advantages offered by carbon nanotubes for the intracellular delivery of therapeutic agents in vivo. The present work may act as the impetus for further studies to explore the therapeutic capacity of chemically functionalized carbon nanotubes to deliver siRNA directly into the cytoplasm of target cells and achieve effective therapeutic silencing in various disease indications where local delivery is feasible or desirable.}, keywords = {Animals, Antineoplastic Agents, Apoptosis, carbon, Cell Line, Cell Proliferation, Electrophoresis, Gene Silencing, Humans, I2CT, Liposomes, Lung Neoplasms, Mice, Nanomedicine, Nanotubes, RNA, Small Interfering, Survival Analysis, Team-Bianco, tumor, Xenograft Model Antitumor Assays}, pubstate = {published}, tppubtype = {article} } @article{schett_b_2009, title = {B cell epitopes of the heterogeneous nuclear ribonucleoprotein A2: identification of a new specific antibody marker for active lupus disease}, author = {G Schett and H Dumortier and E Hoefler and S Muller and G Steiner}, doi = {10.1136/ard.2007.087502}, issn = {1468-2060}, year = {2009}, date = {2009-05-01}, journal = {Annals of the Rheumatic Diseases}, volume = {68}, number = {5}, pages = {729--735}, abstract = {OBJECTIVES: Autoantibody formation and T cell reactivity against the heterogeneous nuclear ribonucleoprotein A2 (hnRNP-A2) has been observed in patients with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). Since no differences in epitope recognition were reported and the usefulness of anti-hnRNP-A2 antibodies as diagnostic markers of SLE is unknown, it was our objective to characterise linear B cell epitopes of hnRNP-A2 and to relate the anti-hnRNP-A2 antibody responses to disease activity and clinical features of SLE. METHODS: Sequential serum samples from 15 patients with SLE and sera from patients with other rheumatic diseases and healthy subjects were investigated by ELISA for autoantibody reactivities against a set of 13 overlapping peptides spanning the RNA-binding region of hnRNP-A2. Antibody reactivity against the complete protein was determined by western immunoblotting and ELISA. SLE disease activity was assessed by European Consensus Lupus Activity Measure scores, by SLE Index scores and the British Isles Lupus Assessment index. RESULTS: Anti-peptide antibody reactivities were found in 60% of SLE sera but in only 5% of control samples, and were mainly directed to four peptides, one of which (p155-175) appeared to be immunodominant. Antibodies to p155-175 were exclusively seen in patients with SLE and correlated with clinical disease activity as well as kidney and skin involvement. No correlations were found for the other anti-peptide antibody responses. CONCLUSION: Peptide p155-175 encompasses a disease-specific immunodominant epitope of hnRNP-A2. Since antibodies to p155-175 correlate with disease activity and nephritis, they may be useful as markers for active SLE.}, keywords = {Autoantibodies, B-Lymphocyte, Biomarkers, Dumortier, Enzyme-Linked Immunosorbent Assay, Epitopes, Female, Follow-Up Studies, Heterogeneous-Nuclear Ribonucleoprotein Group A-B, Humans, I2CT, Lupus Erythematosus, Male, Rheumatic Diseases, Severity of Illness Index, Systemic, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{bernay_discovering_2009, title = {Discovering new bioactive neuropeptides in the striatum secretome using in vivo microdialysis and versatile proteomics.}, author = {Benoît Bernay and Marie-Claude Gaillard and Vilém Guryca and Anouk Emadali and Lauriane Kuhn and Anne Bertrand and Isabelle Detraz and Carole Carcenac and Marc Savasta and Emmanuel Brouillet and Jérôme Garin and Jean-Marc Elalouf}, doi = {10.1074/mcp.M800501-MCP200}, issn = {1535-9484 1535-9476 1535-9476}, year = {2009}, date = {2009-05-01}, journal = {Molecular & cellular proteomics : MCP}, volume = {8}, number = {5}, pages = {946--958}, abstract = {The striatum, a major component of the brain basal nuclei, is central for planning and executing voluntary movements and undergoes lesions in neurodegenerative disorders such as Huntington disease. To perform highly integrated tasks, the striatum relies on a complex network of communication within and between brain regions with a key role devoted to secreted molecules. To characterize the rat striatum secretome, we combined in vivo microdialysis together with proteomics analysis of trypsin digests and peptidomics studies of native fragments. This versatile approach, carried out using different microdialysis probes and mass spectrometer devices, allowed evidencing with high confidence the expression of 88 proteins and 100 processed peptides. Their secretory pathways were predicted by in silico analysis. Whereas high molecular weight proteins were mainly secreted by the classical mode (94%), low molecular weight proteins equally used classical and non-classical modes (53 and 47%, respectively). In addition, our results suggested alternative secretion mechanisms not predicted by bioinformatics tools. Based on spectrum counting, we performed a relative quantification of secreted proteins and peptides in both basal and neuronal depolarization conditions. This allowed detecting a series of neuropeptide precursors and a 6-fold increase for neurosecretory protein VGF and proenkephalin (PENK) levels. A focused investigation and a long peptide experiment led to the identification of new secreted non-opioid PENK peptides, referred to as PENK 114-133, PENK 239-260, and PENK 143-185. Moreover we showed that injecting synthetic PENK 114-133 and PENK 239-260 into the striatum robustly increased glutamate release in this region. Thus, the combination of microdialysis and versatile proteomics methods shed new light on the secreted protein repertoire and evidenced novel neuropeptide transmitters.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{fraiture_two_2009, title = {Two mosquito LRR proteins function as complement control factors in the TEP1-mediated killing of Plasmodium}, author = {Malou Fraiture and Richard H G Baxter and Stefanie Steinert and Yogarany Chelliah and Cécile Frolet and Wilber Quispe-Tintaya and Jules A Hoffmann and Stéphanie A Blandin and Elena A Levashina}, doi = {10.1016/j.chom.2009.01.005}, issn = {1934-6069}, year = {2009}, date = {2009-03-01}, journal = {Cell Host Microbe}, volume = {5}, number = {3}, pages = {273--284}, abstract = {Plasmodium development within Anopheles mosquitoes is a vulnerable step in the parasite transmission cycle, and targeting this step represents a promising strategy for malaria control. The thioester-containing complement-like protein TEP1 and two leucine-rich repeat (LRR) proteins, LRIM1 and APL1, have been identified as major mosquito factors that regulate parasite loads. Here, we show that LRIM1 and APL1 are required for binding of TEP1 to parasites. RNAi silencing of the LRR-encoding genes results in deposition of TEP1 on Anopheles tissues, thereby depleting TEP1 from circulation in the hemolymph and impeding its binding to Plasmodium. LRIM1 and APL1 not only stabilize circulating TEP1, they also stabilize each other prior to their interaction with TEP1. Our results indicate that three major antiparasitic factors in mosquitoes jointly function as a complement-like system in parasite killing, and they reveal a role for LRR proteins as complement control factors.}, keywords = {Animals, Anopheles, APL1, Biological, blandin, Complement System Proteins, Hemolymph, hoffmann, Immunologic Factors, LRIM1, M3i, Models, Plasmodium, Protein Binding, Proteins, TEP1}, pubstate = {published}, tppubtype = {article} } @article{bianco_potential_2009, title = {Potential usefulness of carbon nanotubes for cancer therapy}, author = {Alberto Bianco}, doi = {10.1051/medsci/2009252125}, issn = {0767-0974}, year = {2009}, date = {2009-02-01}, journal = {Medecine Sciences: M/S}, volume = {25}, number = {2}, pages = {125--127}, keywords = {carbon, Graphite, Humans, I2CT, Nanotubes, Neoplasms, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{kemp_antiviral_2009, title = {Antiviral immunity in drosophila}, author = {Cordula Kemp and Jean-Luc Imler}, doi = {10.1016/j.coi.2009.01.007}, issn = {1879-0372}, year = {2009}, date = {2009-02-01}, journal = {Current Opinion in Immunology}, volume = {21}, number = {1}, pages = {3--9}, abstract = {Genetic analysis of the drosophila antiviral response indicates that RNA interference plays a major role. This contrasts with the situation in mammals, where interferon-induced responses mediate innate antiviral host-defense. An inducible response also contributes to antiviral immunity in drosophila, and similarities in the sensing and signaling of viral infection are becoming apparent between drosophila and mammals. In particular, DExD/H box helicases appear to play a crucial role in the cytosolic detection of viral RNAs in flies and mammals.}, keywords = {Animals, Argonaute Proteins, Caspases, DEAD-box RNA Helicases, Evolution, Gene Expression Regulation, Host-Pathogen Interactions, imler, M3i, Membrane Proteins, Molecular, Nuclear Proteins, Ribonuclease III, RNA, RNA Helicases, RNA Interference, RNA Virus Infections, RNA Viruses, RNA-Induced Silencing Complex, Viral, Virulence}, pubstate = {published}, tppubtype = {article} } @article{fauny_entire_2009, title = {The entire zebrafish blastula-gastrula margin acts as an organizer dependent on the ratio of Nodal to BMP activity}, author = {Jean-Daniel Fauny and Bernard Thisse and Christine Thisse}, url = {http://www.ncbi.nlm.nih.gov/pubmed/19855023}, doi = {10.1242/dev.039693}, issn = {1477-9129}, year = {2009}, date = {2009-01-01}, urldate = {2011-10-24}, journal = {Development (Cambridge, England)}, volume = {136}, number = {22}, pages = {3811--3819}, abstract = {Formation of the vertebrate embryo is known to depend on the activity of organizing centers. The dorsal Spemann organizer is the source of growth factor antagonists that participate in the creation of signaling gradients. In various species, the existence of head, trunk and trunk-tail inducers has been proposed to explain the formation of different parts of the embryo along the anteroposterior (A/P) axis. In zebrafish, two organizing centers have been described, the dorsal and tail organizers, located at the dorsal and ventral gastrula margins, respectively. Here, we report that organizer functions are executed not only by the dorsal and ventral margins, but also by all parts of the blastula-gastrula margin. The position of different marginal territories along the dorsoventral axis defines the A/P nature of the structures they are able to organize. At the molecular level, we show that this organizing activity results from the simultaneous activation of BMP and Nodal signaling pathways. Furthermore, the A/P character of the organized structures is not defined by absolute levels but instead by the ratio of BMP and Nodal activities. Rather than resulting from the activity of discrete centers, organization of the zebrafish embryo depends on the activity of the entire margin acting as a continuous and global organizer that is established by a gradual ventral-to-dorsal modulation of the ratio of marginal BMP to Nodal activity.}, keywords = {Animals, Blastula, Bone Morphogenetic Proteins, Embryonic, Gastrula, I2CT, Imagerie, Nodal Protein, Organizers, Zebrafish, Zebrafish Proteins}, pubstate = {published}, tppubtype = {article} } @article{monneaux_molecular_2009, title = {Molecular therapies for systemic lupus erythematosus: clinical trials and future prospects}, author = {Fanny Monneaux and Sylviane Muller}, doi = {10.1186/ar2711}, issn = {1478-6362}, year = {2009}, date = {2009-01-01}, journal = {Arthritis Research & Therapy}, volume = {11}, number = {3}, pages = {234}, abstract = {The prognosis of patients with systemic lupus erythematosus has greatly improved since treatment regimens combining corticosteroids and immunosuppressive medications have been widely adopted in therapeutic strategies given to these patients. Immune suppression is evidently efficient but also leads to higher susceptibility to infectious and malignant diseases. Toxic effects and sometimes unexpectedly dramatic complications of current therapies have been progressively reported. Identifying novel molecular targets therefore remains an important issue in the treatment of lupus. The aim of this review article is to highlight emerging pharmacological options and new therapeutic avenues for lupus with a particular focus on non-antibody molecular strategies.}, keywords = {Animals, Clinical Trials as Topic, Forecasting, Genetic Therapy, Humans, I2CT, Lupus Erythematosus, Monneaux, Systemic, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{lacotte_cxcr3_2009, title = {CXCR3, inflammation, and autoimmune diseases}, author = {Stéphanie Lacotte and Susana Brun and Sylviane Muller and Hélène Dumortier}, doi = {10.1111/j.1749-6632.2009.04813.x}, issn = {1749-6632}, year = {2009}, date = {2009-01-01}, journal = {Annals of the New York Academy of Sciences}, volume = {1173}, pages = {310--317}, abstract = {CXCR3 is a G protein-coupled, seven-transmembrane receptor that binds and is activated by the three IFN-gamma-inducible chemokines of the CXC family named CXCL9, CXCL10, and CXCL11. These chemokines are not constitutively expressed but are up-regulated in a proinflammatory cytokine milieu. Consequently, their major function is to selectively recruit immune cells at inflammation sites, but they also play a role in angiogenesis mechanisms. In the last few years, strong experimental and clinical evidence has been obtained supporting the idea that the CXCR3 pathway is involved in the development of autoimmune diseases, especially by creating local amplification loops of inflammation in target organs, thereby inducing worsening of clinical manifestations. This article briefly reviews what we know today about the nature and functions of CXCR3, with special emphasis on its involvement in two main rheumatic systemic autoimmune diseases, namely rheumatoid arthritis and systemic lupus erythematosus.}, keywords = {Animals, arthritis, Biological, Chemokine CXCL10, Chemokine CXCL11, Chemokine CXCL9, CXCR3, Dumortier, Humans, I2CT, inflammation, Lupus Erythematosus, Models, Receptors, rheumatoid, Systemic, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{berry_viral_2009, title = {Viral suppressors of RNA silencing hinder exogenous and endogenous small RNA pathways in Drosophila}, author = {Bassam Berry and Safia Deddouche and Doris Kirschner and Jean-Luc Imler and Christophe Antoniewski}, doi = {10.1371/journal.pone.0005866}, issn = {1932-6203}, year = {2009}, date = {2009-01-01}, journal = {PloS One}, volume = {4}, number = {6}, pages = {e5866}, abstract = {BACKGROUND: In plants and insects, RNA interference (RNAi) is the main responder against viruses and shapes the basis of antiviral immunity. Viruses counter this defense by expressing viral suppressors of RNAi (VSRs). While VSRs in Drosophila melanogaster were shown to inhibit RNAi through different modes of action, whether they act on other silencing pathways remained unexplored. METHODOLOGY/PRINCIPAL FINDINGS: Here we show that expression of various plant and insect VSRs in transgenic flies does not perturb the Drosophila microRNA (miRNA) pathway; but in contrast, inhibits antiviral RNAi and the RNA silencing response triggered by inverted repeat transcripts, and injection of dsRNA or siRNA. Strikingly, these VSRs also suppressed transposon silencing by endogenous siRNAs (endo-siRNAs). CONCLUSIONS/SIGNIFICANCE: Our findings identify VSRs as tools to unravel small RNA pathways in insects and suggest a cosuppression of antiviral RNAi and endo-siRNA silencing by viruses during fly infections.}, keywords = {Animals, Antiviral Agents, Crosses, Double-Stranded, Gene Silencing, Genetic, Genetically Modified, Heterozygote, imler, Invertebrate, M3i, Photoreceptor Cells, Reverse Transcriptase Polymerase Chain Reaction, RNA, RNA Interference, Transgenes}, pubstate = {published}, tppubtype = {article} } @article{gaillard_carbon_2009, title = {Carbon Nanotubes Carrying Cell-Adhesion Peptides do not Interfere with Neuronal Functionality}, author = {Claire Gaillard and Giada Cellot and Shouping Li and Francesca Maria Toma and Hélène Dumortier and Giampiero Spalluto and Barbara Cacciari and Maurizio Prato and Laura Ballerini and Alberto Bianco}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adma.200900050}, doi = {10.1002/adma.200900050}, issn = {1521-4095}, year = {2009}, date = {2009-01-01}, urldate = {2020-03-31}, journal = {Advanced Materials}, volume = {21}, number = {28}, pages = {2903--2908}, abstract = {Water-soluble carbon nanotubes functionalized with cell-adhesion peptides do not affect the viability of different cell types, including Jurkat cells, splenocytes, and neurons. They also do not modify the neuronal morphology and basic functions, thus representing a promising candidate for the exploitation of novel drug-delivery systems or for designing a new generation of self-assembling nerve bridges.}, keywords = {Carbon nanotubes, Cytotoxicity, I2CT, mammalian cells, Neurons, Peptides, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{garrett_identification_2009, title = {Identification and analysis of serpin-family genes by homology and synteny across the 12 sequenced Drosophilid genomes}, author = {Matthew Garrett and Ane Fullaondo and Laurent Troxler and Gos Micklem and David Gubb}, doi = {10.1186/1471-2164-10-489}, issn = {1471-2164}, year = {2009}, date = {2009-01-01}, journal = {BMC Genomics}, volume = {10}, pages = {489}, abstract = {BACKGROUND: The Drosophila melanogaster genome contains 29 serpin genes, 12 as single transcripts and 17 within 6 gene clusters. Many of these serpins have a conserved "hinge" motif characteristic of active proteinase inhibitors. However, a substantial proportion (42%) lacks this motif and represents non-inhibitory serpin-fold proteins of unknown function. Currently, it is not known whether orthologous, inhibitory serpin genes retain the same target proteinase specificity within the Drosophilid lineage, nor whether they give rise to non-inhibitory serpin-fold proteins or other, more diverged, proteins. RESULTS: We collated 188 orthologues to the D. melanogaster serpins from the other 11 Drosophilid genomes and used synteny to find further family members, raising the total to 226, or 71% of the number of orthologues expected assuming complete conservation across all 12 Drosophilid species. In general the sequence constraints on the serpin-fold itself are loose. The critical Reactive Centre Loop (RCL) sequence, including the target proteinase cleavage site, is strongly conserved in inhibitory serpins, although there are 3 exceptional sets of orthologues in which the evolutionary constraints are looser. Conversely, the RCL of non-inhibitory serpin orthologues is less conserved, with 3 exceptions that presumably bind to conserved partner molecules. We derive a consensus hinge motif, for Drosophilid inhibitory serpins, which differs somewhat from that of the vertebrate consensus. Three gene clusters appear to have originated in the melanogaster subgroup, Spn28D, Spn77B and Spn88E, each containing one inhibitory serpin orthologue that is present in all Drosophilids. In addition, the Spn100A transcript appears to represent a novel serpin-derived fold. CONCLUSION: In general, inhibitory serpins rarely change their range of proteinase targets, except by a duplication/divergence mechanism. Non-inhibitory serpins appear to derive from inhibitory serpins, but not the reverse. The conservation of different family members varied widely across the 12 sequenced Drosophilid genomes. An approach considering synteny as well as homology was important to find the largest set of orthologues.}, keywords = {Animals, bioinformatic, Comparative Genomic Hybridization, Conserved Sequence, DNA, Drosophilidae, Evolution, Genome, Insect, Molecular, Multigene Family, Sequence Alignment, Sequence Analysis, Serpins, Synteny}, pubstate = {published}, tppubtype = {article} } @incollection{leclerc_phagocytosis_2009, title = {Phagocytosis in Drosophila melanogaster Immune Response.}, author = {V Leclerc and I Caldelari and N Veresceaghina and J-M Reichhart}, year = {2009}, date = {2009-01-01}, booktitle = {Phagocyte-pathogen Interactions}, volume = {33}, pages = {513--521}, publisher = {DG Russel and S. Gordon}, address = {Whasington, DC}, edition = {ASM Press}, keywords = {M3i, reichhart, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {incollection} } @article{cronin_genome-wide_2009b, title = {Genome-wide RNAi screen identifies genes involved in intestinal pathogenic bacterial infection}, author = {Shane J F Cronin and Nadine T Nehme and Stefanie Limmer and Samuel Liegeois and Andrew J Pospisilik and Daniel Schramek and Andreas Leibbrandt and Ricardo Matos de Simoes and Susanne Gruber and Urszula Puc and Ingo Ebersberger and Tamara Zoranovic and Gregory G Neely and Arndt von Haeseler and Dominique Ferrandon and Josef M Penninger}, doi = {10.1126/science.1173164}, issn = {1095-9203}, year = {2009}, date = {2009-01-01}, journal = {Science}, volume = {325}, number = {5938}, pages = {340--343}, abstract = {Innate immunity represents the first line of defense in animals. We report a genome-wide in vivo Drosophila RNA interference screen to uncover genes involved in susceptibility or resistance to intestinal infection with the bacterium Serratia marcescens. We first employed whole-organism gene suppression, followed by tissue-specific silencing in gut epithelium or hemocytes to identify several hundred genes involved in intestinal antibacterial immunity. Among the pathways identified, we showed that the JAK-STAT signaling pathway controls host defense in the gut by regulating stem cell proliferation and thus epithelial cell homeostasis. Therefore, we revealed multiple genes involved in antibacterial defense and the regulation of innate immunity.}, keywords = {*Genome, *RNA Interference, Animal, Animals, Cell Proliferation, Drosophila melanogaster/*genetics/immunology/*microbiology, Drosophila Proteins/genetics/metabolism, Epithelial Cells, Epithelial Cells/cytology/physiology, ferrandon, Genetically Modified, Genome, Hemocytes, Hemocytes/immunology/metabolism/microbiology, Homeostasis, Immunity, Innate, Innate/*genetics, Insect, Intestinal Mucosa, Intestinal Mucosa/cytology/immunology/metabolism/microbiology, Janus Kinases, Janus Kinases/genetics/metabolism, M3i, Models, RNA Interference, Serratia Infections, Serratia Infections/genetics/*immunology/microbiology, Serratia marcescens, Serratia marcescens/*immunology/physiology, Signal Transduction, STAT Transcription Factors, STAT Transcription Factors/genetics/metabolism, Stem Cells, Stem Cells/cytology/physiology}, pubstate = {published}, tppubtype = {article} } @article{F2009, title = {RNAi in the malaria vector, Anopheles gambiae}, author = {Flaminia Catteruccia and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/19495688}, year = {2009}, date = {2009-01-01}, journal = {Methods Mol Biol.}, volume = {555}, pages = {63-75}, abstract = {Malaria is a disease that kills more than a million people each year in tropical and subtropical countries. The disease is caused by Plasmodium parasites and is transmitted to humans exclusively by mosquitoes of the genus Anopheles. The lack of functional approaches has hampered study of the biological networks that determine parasite transmission by the insect vector. The recent discovery of RNA interference and its adaptation to mosquitoes is now providing crucial tools for the dissection of vector-parasite interactions and for the analysis of aspects of mosquito biology influencing the vectorial capacity. Two RNAi approaches have been established in mosquitoes: transient gene silencing by direct injection of double-stranded RNA, and stable expression of hairpin RNAs from transgenes integrated in the genome. Here we describe these methods in detail, providing information about their use and limitations.}, keywords = {RNAi}, pubstate = {published}, tppubtype = {article} } @article{, title = {Distinct beta-clamp interactions govern the activities of the Y family PolIV DNA polymerase}, author = { J. Wagner and H. Etienne and R. P. Fuchs and A. Cordonnier and D. Burnouf}, year = {2009}, date = {2009-01-01}, journal = {Mol Microbiol}, volume = {74}, number = {5}, pages = {1143-51}, abstract = {The prototypic Y family DNA polymerase IV (PolIV) of Escherichia coli is involved in multiple replication-associated processes including spontaneous mutagenesis, translesion synthesis (TLS), cell fitness, survival under stressful conditions and checkpoint like functions. It interacts physically and functionally with the replisome's beta processivity clamp through the canonical PolIV C-terminal peptide (CTP). A second interaction that involves a portion of the little finger (LF) domain of PolIV has been structurally described. Here we show that the LF-beta interaction stabilizes the clamp-polymerase complex in vitro and is necessary for the access of PolIV to ongoing replication forks in vivo. However, in contrast to the CTP-beta, the LF-beta interaction is dispensable for the role of the polymerase in TLS. This discloses two independent modes of action for PolIV and, in turn, uncovers a novel way by which the cell may regulate the potentially deleterious effect of such low fidelity polymerases during replication.}, note = {1365-2958 (Electronic) 0950-382X (Linking) Journal Article}, keywords = {DUMAS}, pubstate = {published}, tppubtype = {article} } @article{bianco_les_2009, title = {Les nanotubes de carbone : un nouvel outil contre le cancer}, author = {Alberto Bianco}, url = {http://www.ipubli.inserm.fr/handle/10608/6569}, doi = {10.1051/medsci/2009252125}, issn = {1958-5381}, year = {2009}, date = {2009-01-01}, urldate = {2020-04-01}, journal = {M/S. Médecine sciences [ISSN papier : 0767-0974 ; ISSN numérique : 1958-5381], 2009, Vol. 25, N° 2; p. 125-127}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNA-independent Pretransfer Editing by Class I Leucyl-tRNA Synthetase}, author = {B Zhu and P Yao and M Tan and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19068478}, isbn = {19068478}, year = {2009}, date = {2009-01-01}, journal = {J Biol Chem}, volume = {284}, number = {6}, pages = {3418-3424}, abstract = {Aminoacyl-tRNA synthetases catalyze the formation of aminoacyl-tRNA in a two-step reaction starting with amino acid activation followed by aminoacyl group transfer to tRNA. To clear mistakes that occasionally occur, some of these enzymes carry out editing activities, acting on the misactivated amino acid (pretransfer editing) or after the transfer on the tRNA (post-transfer editing). The post-transfer editing pathway of leucyl-tRNA synthetase has been extensively studied by structural and biochemical approaches. Here, we report the finding of a tRNA-independent pretransfer editing pathway in leucyl-tRNA synthetases from Aquifex aeolicus. Using a CP1-mutant defective in its post-transfer editing function, we showed that this new editing pathway is distinct from the post-transfer editing site and may occur at the synthetic catalytic site, as recently proposed for other aminoacyl-tRNA synthetases.}, note = {0021-9258 (Print) Journal Article}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleic acids: how high resolution structural biology help us to understand Darwinian evolution}, author = {E Westhof and D J Patel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19482468}, isbn = {19482468}, year = {2009}, date = {2009-01-01}, journal = {Curr Opin Struct Biol}, volume = {19}, number = {3}, pages = {235-238}, note = {1879-033X (Electronic) Editorial Introductory Journal Article}, keywords = {*Evolution, Molecular Humans Molecular Biology/*methods Nucleic Acids/*chemistry/metabolism Periodicals as Topic, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Distinct beta-clamp interactions govern the activities of the Y family PolIV DNA polymerase}, author = {J Wagner and H Etienne and R P Fuchs and A Cordonnier and D Burnouf}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19843218}, isbn = {19843218}, year = {2009}, date = {2009-01-01}, journal = {Mol Microbiol}, volume = {74}, number = {5}, pages = {1143-1151}, abstract = {The prototypic Y family DNA polymerase IV (PolIV) of Escherichia coli is involved in multiple replication-associated processes including spontaneous mutagenesis, translesion synthesis (TLS), cell fitness, survival under stressful conditions and checkpoint like functions. It interacts physically and functionally with the replisome's beta processivity clamp through the canonical PolIV C-terminal peptide (CTP). A second interaction that involves a portion of the little finger (LF) domain of PolIV has been structurally described. Here we show that the LF-beta interaction stabilizes the clamp-polymerase complex in vitro and is necessary for the access of PolIV to ongoing replication forks in vivo. However, in contrast to the CTP-beta, the LF-beta interaction is dispensable for the role of the polymerase in TLS. This discloses two independent modes of action for PolIV and, in turn, uncovers a novel way by which the cell may regulate the potentially deleterious effect of such low fidelity polymerases during replication.}, note = {1365-2958 (Electronic) 0950-382X (Linking) Journal Article}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Analogues nucleosidiques inhibiteurs de la retrotranscriptase}, author = {V Vivet-Boudou and R Marquet and A Burger}, isbn = {09/01787 (fr)}, year = {2009}, date = {2009-01-01}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Crystallographic model quality at a glance}, author = {L Urzhumtseva and P V Afonine and P D Adams and A Urzhumtsev}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19237753}, isbn = {19237753}, year = {2009}, date = {2009-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {65}, number = {Pt 3}, pages = {297-300}, abstract = {A crystallographic macromolecular model is typically characterized by a list of quality criteria, such as R factors, deviations from ideal stereochemistry and average B factors, which are usually provided as tables in publications or in structural databases. In order to facilitate a quick model-quality evaluation, a graphical representation is proposed. Each key parameter such as R factor or bond-length deviation from;ideal values' is shown graphically as a point on a;ruler'. These rulers are plotted as a set of lines with the same origin, forming a hub and spokes. Different parts of the rulers are coloured differently to reflect the frequency (red for a low frequency, blue for a high frequency) with which the corresponding values are observed in a reference set of structures determined previously. The points for a given model marked on these lines are connected to form a polygon. A polygon that is strongly compressed or dilated along some axes reveals unusually low or high values of the corresponding characteristics. Polygon vertices in;red zones' indicate parameters which lie outside typical values.}, note = {1399-0047 (Electronic) Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A short motif in Drosophila SECIS Binding Protein 2 provides differential binding affinity to SECIS RNA hairpins}, author = {A Takeuchi and D Schmitt and C Chapple and E Babaylova and G Karpova and R Guigo and A Krol and C Allmang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19223320}, isbn = {19223320}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {7}, pages = {2126-2141}, abstract = {Selenoproteins contain the amino acid selenocysteine which is encoded by a UGA Sec codon. Recoding UGA Sec requires a complex mechanism, comprising the cis-acting SECIS RNA hairpin in the 3'UTR of selenoprotein mRNAs, and trans-acting factors. Among these, the SECIS Binding Protein 2 (SBP2) is central to the mechanism. SBP2 has been so far functionally characterized only in rats and humans. In this work, we report the characterization of the Drosophila melanogaster SBP2 (dSBP2). Despite its shorter length, it retained the same selenoprotein synthesis-promoting capabilities as the mammalian counterpart. However, a major difference resides in the SECIS recognition pattern: while human SBP2 (hSBP2) binds the distinct form 1 and 2 SECIS RNAs with similar affinities, dSBP2 exhibits high affinity toward form 2 only. In addition, we report the identification of a K (lysine)-rich domain in all SBP2s, essential for SECIS and 60S ribosomal subunit binding, differing from the well-characterized L7Ae RNA-binding domain. Swapping only five amino acids between dSBP2 and hSBP2 in the K-rich domain conferred reversed SECIS-binding properties to the proteins, thus unveiling an important sequence for form 1 binding.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {ERIANI, KROL, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Frequency and isostericity of RNA base pairs}, author = {J Stombaugh and C L Zirbel and E Westhof and N B Leontis}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19240142}, isbn = {19240142}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {7}, pages = {2294-2312}, abstract = {Most of the hairpin, internal and junction loops that appear single-stranded in standard RNA secondary structures form recurrent 3D motifs, where non-Watson-Crick base pairs play a central role. Non-Watson-Crick base pairs also play crucial roles in tertiary contacts in structured RNA molecules. We previously classified RNA base pairs geometrically so as to group together those base pairs that are structurally similar (isosteric) and therefore able to substitute for each other by mutation without disrupting the 3D structure. Here, we introduce a quantitative measure of base pair isostericity, the IsoDiscrepancy Index (IDI), to more accurately determine which base pair substitutions can potentially occur in conserved motifs. We extract and classify base pairs from a reduced-redundancy set of RNA 3D structures from the Protein Data Bank (PDB) and calculate centroids (exemplars) for each base combination and geometric base pair type (family). We use the exemplars and IDI values to update our online Basepair Catalog and the Isostericity Matrices (IM) for each base pair family. From the database of base pairs observed in 3D structures we derive base pair occurrence frequencies for each of the 12 geometric base pair families. In order to improve the statistics from the 3D structures, we also derive base pair occurrence frequencies from rRNA sequence alignments.}, note = {1362-4962 (Electronic) Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.}, keywords = {ase Pairing Base Sequence Models, Bacterial/chemistry RNA, Molecular Nucleic Acid Conformation RNA/*chemistry RNA, Ribosomal/chemistry Sequence Alignment Sequence Analysis, RNA, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comparative genomics of protoploid Saccharomycetaceae}, author = {J L Souciet and B Dujon and C Gaillardin and M Johnston and P V Baret and P Cliften and D J Sherman and J Weissenbach and E Westhof and P Wincker and C Jubin and J Poulain and V Barbe and B Segurens and F Artiguenave and V Anthouard and B Vacherie and M E Val and R S Fulton and P Minx and R Wilson and P Durrens and G Jean and C Marck and T Martin and M Nikolski and T Rolland and M L Seret and S Casaregola and L Despons and C Fairhead and G Fischer and I Lafontaine and V Leh and M Lemaire and J de Montigny and C Neuveglise and A Thierry and I Blanc-Lenfle and C Bleykasten and J Diffels and E Fritsch and L Frangeul and A Goeffon and N Jauniaux and R Kachouri-Lafond and C Payen and S Potier and L Pribylova and C Ozanne and G F Richard and C Sacerdot and M L Straub and E Talla}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19525356}, isbn = {19525356}, year = {2009}, date = {2009-01-01}, journal = {Genome Res}, volume = {19}, number = {10}, pages = {1696-1709}, abstract = {Our knowledge of yeast genomes remains largely dominated by the extensive studies on Saccharomyces cerevisiae and the consequences of its ancestral duplication, leaving the evolution of the entire class of hemiascomycetes only partly explored. We concentrate here on five species of Saccharomycetaceae, a large subdivision of hemiascomycetes, that we call "protoploid" because they diverged from the S. cerevisiae lineage prior to its genome duplication. We determined the complete genome sequences of three of these species: Kluyveromyces (Lachancea) thermotolerans and Saccharomyces (Lachancea) kluyveri (two members of the newly described Lachancea clade), and Zygosaccharomyces rouxii. We included in our comparisons the previously available sequences of Kluyveromyces lactis and Ashbya (Eremothecium) gossypii. Despite their broad evolutionary range and significant individual variations in each lineage, the five protoploid Saccharomycetaceae share a core repertoire of approximately 3300 protein families and a high degree of conserved synteny. Synteny blocks were used to define gene orthology and to infer ancestors. Far from representing minimal genomes without redundancy, the five protoploid yeasts contain numerous copies of paralogous genes, either dispersed or in tandem arrays, that, altogether, constitute a third of each genome. Ancient, conserved paralogs as well as novel, lineage-specific paralogs were identified.}, note = {1549-5469 (Electronic) 1088-9051 (Linking) Comparative Study Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't}, keywords = {DNA Transposable Elements/genetics/physiology Eremothecium/genetics Gene Duplication Genes, Fungal Genomics/*methods Inteins/genetics Kluyveromyces/genetics Molecular Sequence Data Open Reading Frames/genetics Phylogeny RNA, Fungal/genetics *Genome, Unité ARN, Untranslated/genetics Saccharomyces/genetics Saccharomycetales/*genetics Spliceosomes/metabolism Zygosaccharomyces/genetics, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular mechanisms of recombination restriction in the envelope gene of the human immunodeficiency virus}, author = {E Simon-Loriere and R Galetto and M Hamoudi and J Archer and P Lefeuvre and D P Martin and D L Robertson and M Negroni}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19424420}, isbn = {19424420}, year = {2009}, date = {2009-01-01}, journal = {PLoS Pathog}, volume = {5}, number = {5}, pages = {e1000418}, abstract = {The ability of pathogens to escape the host's immune response is crucial for the establishment of persistent infections and can influence virulence. Recombination has been observed to contribute to this process by generating novel genetic variants. Although distinctive recombination patterns have been described in many viral pathogens, little is known about the influence of biases in the recombination process itself relative to selective forces acting on newly formed recombinants. Understanding these influences is important for determining how recombination contributes to pathogen genome and proteome evolution. Most previous research on recombination-driven protein evolution has focused on relatively simple proteins, usually in the context of directed evolution experiments. Here, we study recombination in the envelope gene of HIV-1 between primary isolates belonging to subtypes that recombine naturally in the HIV/AIDS pandemic. By characterizing the early steps in the generation of recombinants, we provide novel insights into the evolutionary forces that shape recombination patterns within viral populations. Specifically, we show that the combined effects of mechanistic processes that determine the locations of recombination breakpoints across the HIV-1 envelope gene, and purifying selection acting against dysfunctional recombinants, can explain almost the entire distribution of breakpoints found within this gene in nature. These constraints account for the surprising paucity of recombination breakpoints found in infected individuals within this highly variable gene. Thus, the apparent randomness of HIV evolution via recombination may in fact be relatively more predictable than anticipated. In addition, the dominance of purifying selection in localized areas of the HIV genome defines regions where functional constraints on recombinants appear particularly strong, pointing to vulnerable aspects of HIV biology.}, note = {1553-7374 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A structural view of translation initiation in bacteria}, author = {A Simonetti and S Marzi and L Jenner and A Myasnikov and P Romby and G Yusupova and B P Klaholz and M Yusupov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19011758}, isbn = {19011758}, year = {2009}, date = {2009-01-01}, journal = {Cell Mol Life Sci}, volume = {66}, number = {3}, pages = {423-436}, abstract = {The assembly of the protein synthesis machinery occurs during translation initiation. In bacteria, this process involves the binding of messenger RNA(mRNA) start site and fMet-tRNA(fMet) to the ribosome, which results in the formation of the first codon-anticodon interaction and sets the reading frame for the decoding of the mRNA. This interaction takes place in the peptidyl site of the 30S ribosomal subunit and is controlled by the initiation factors IF1, IF2 and IF3 to form the 30S initiation complex. The binding of the 50S subunit and the ejection of the IFs mark the irreversible transition to the elongation phase. Visualization of these ligands on the ribosome has been achieved by cryo-electron microscopy and X-ray crystallography studies, which has helped to understand the mechanism of translation initiation at the molecular level. Conformational changes associated with different functional states provide a dynamic view of the initiation process and of its regulation.}, note = {1420-9071 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Agarose gel facilitates enzyme crystal soaking with a ligand analog.}, author = {C Sauter and C Balg and A Moreno and K Dhouib and A Théobald-Dietrich and R Chenevert and R Giege and B Lorber}, url = {http://scripts.iucr.org/cgi-bin/paper?he5431}, isbn = {10.1107/S0021889809003446}, year = {2009}, date = {2009-01-01}, journal = {J Appl Cryst}, volume = {42}, pages = {279-283}, abstract = {Orthorhombic crystals of the enzyme aspartyl-tRNA synthetase (AspRS) were prepared in agarose gel, a chemical alternative to microgravity or nano-volume drops. Besides providing a convection-free medium, the network of the polysaccharide improved the stability of the crystalline lattice during soaking with L-aspartol adenylate, a synthetic and non-hydrolysable analog of the catalytic intermediate aspartyl adenylate. When crystals were embedded in the polysaccharide matrix the ligand reached their surfaces more uniformly. Gel-grown crystals exhibited well defined reflections even at high resolution and low mosaicity values, despite their fairly high solvent content and the relatively harsh flash cooling procedure. By contrast, soaked AspRS crystals prepared in solution broke apart within 10-30 s after the ligand was introduced into the mother liquor, and subsequently these fragments became an amorphous precipitate. A general objection to the use of gels in protein crystallization is that chemical interactions may occur between the polysaccharide matrix and proteins or ligands. The example of AspRS shows that this is not a major concern. This method may be generally applicable to crystal soaking with other small molecules or heavy atoms.}, keywords = {Crystallization Agarose gel Ligands Inhibitors Soaking, FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[The atomic structure of the ribosome into the spotlight]}, author = {P Romby and S Marzi and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19951677}, isbn = {19951677}, year = {2009}, date = {2009-01-01}, journal = {Med Sci (Paris)}, volume = {25}, number = {11}, pages = {977-981}, note = {0767-0974 (Print) 0767-0974 (Linking) Journal Article}, keywords = {ROMBY, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {An overview of RNAs with regulatory functions in gram-positive bacteria}, author = {P Romby and E Charpentier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19859665}, isbn = {19859665}, year = {2009}, date = {2009-01-01}, journal = {Cell Mol Life Sci}, volume = {67}, number = {2}, pages = {217-237}, abstract = {During the last decade, RNA molecules with regulatory functions on gene expression have benefited from a renewed interest. In bacteria, recent high throughput computational and experimental approaches have led to the discovery that 10-20% of all genes code for RNAs with critical regulatory roles in metabolic, physiological and pathogenic processes. The trans-acting RNAs comprise the noncoding RNAs, RNAs with a short open reading frame and antisense RNAs. Many of these RNAs act through binding to their target mRNAs while others modulate protein activity or target DNA. The cis-acting RNAs include regulatory regions of mRNAs that can respond to various signals. These RNAs often provide the missing link between sensing changing conditions in the environment and fine-tuning the subsequent biological responses. Information on their various functions and modes of action has been well documented for gram-negative bacteria. Here, we summarize the current knowledge of regulatory RNAs in gram-positive bacteria.}, note = {1420-9071 (Electronic) 1420-682X (Linking) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Bacterial Gram-Positive Bacteria/*genetics/pathogenicity RNA, Bacterial/genetics/*metabolism RNA, Gene Expression Regulation, ROMBY, Unité ARN, Untranslated/genetics/*metabolism Virulence/genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Discovery of chiral cyclopropyl dihydro-alkylthio-benzyl-oxopyrimidine (S-DABO) derivatives as potent HIV-1 reverse transcriptase inhibitors with high activity against clinically relevant mutants}, author = {M Radi and G Maga and M Alongi and L Angeli and A Samuele and S Zanoli and L Bellucci and A Tafi and G Casaluce and G Giorgi and M Armand-Ugon and E Gonzalez and J A Este and M Baltzinger and G Bec and P Dumas and E Ennifar and M Botta}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19140683}, isbn = {19140683}, year = {2009}, date = {2009-01-01}, journal = {J Med Chem}, volume = {52}, number = {3}, pages = {840-851}, abstract = {The role played by stereochemistry in the C2-substituent (left part) on the S-DABO scaffold for anti-HIV-1 activity has been investigated for the first time. A series of S-DABO analogues, where the double bond in the C2-substituent is replaced by an enantiopure isosteric cyclopropyl moiety, has been synthesized, leading to the identification of a potent lead compound endowed with picomolar activity against RT (wt) and nanomolar activity against selected drug-resistant mutants. Molecular modeling calculation, enzymatic studies, and surface plasmon resonance experiments allowed us to rationalize the biological behavior of the synthesized compounds, which act as mixed-type inhibitors of HIV-1 RT K103N, with a preferential association to the enzyme-substrate complex. Taken together, our data show that the right combination of stereochemistry on the left and right parts (C6-substituent) of the S-DABO scaffold plays a key role in the inhibition of both wild-type and drug-resistant enzymes, especially the K103N mutant.}, note = {1520-4804 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Unravelling the importance of microRNAs during hepatitis C virus infection in the human liver}, author = {S Pfeffer and T F Baumert}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19467729}, isbn = {19467729}, year = {2009}, date = {2009-01-01}, journal = {J Hepatol}, volume = {51}, number = {3}, pages = {606-609}, note = {0168-8278 (Print) Comment Journal Article}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {New metrics for comparing and assessing discrepancies between RNA 3D structures and models}, author = {M Parisien and J A Cruz and E Westhof and F Major}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19710185}, isbn = {19710185}, year = {2009}, date = {2009-01-01}, journal = {RNA}, volume = {15}, number = {10}, pages = {1875-1885}, abstract = {To benchmark progress made in RNA three-dimensional modeling and assess newly developed techniques, reliable and meaningful comparison metrics and associated tools are necessary. Generally, the average root-mean-square deviations (RMSDs) are quoted. However, RMSD can be misleading since errors are spread over the whole molecule and do not account for the specificity of RNA base interactions. Here, we introduce two new metrics that are particularly suitable to RNAs: the deformation index and deformation profile. The deformation index is calibrated by the interaction network fidelity, which considers base-base-stacking and base-base-pairing interactions within the target structure. The deformation profile highlights dissimilarities between structures at the nucleotide scale for both intradomain and interdomain interactions. Our results show that there is little correlation between RMSD and interaction network fidelity. The deformation profile is a tool that allows for rapid assessment of the origins of discrepancies.}, note = {1469-9001 (Electronic) 1355-8382 (Linking) Comparative Study Journal Article Research Support, Non-U.S. Gov't}, keywords = {28S/*chemistry Rats, Animals Base Pairing Calibration *Models, Molecular *Nucleic Acid Conformation RNA, Ribosomal, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {SECIS-binding protein 2, a key player in selenoprotein synthesis, is an intrinsically disordered protein}, author = {V Olieric and P Wolff and A Takeuchi and G Bec and C Birck and M Vitorino and B Kieffer and A Beniaminov and G Cavigiolio and E Theil and C Allmang and A Krol and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19467292}, isbn = {19467292}, year = {2009}, date = {2009-01-01}, journal = {Biochimie}, volume = {91}, number = {8}, pages = {1003-1009}, abstract = {Selenocysteine (Sec) is co-translationally incorporated into selenoproteins at a reprogrammed UGA codon. In mammals, this requires a dedicated machinery comprising a stem-loop structure in the 3' UTR RNA (the SECIS element) and the specific SECIS Binding Protein 2. In this report, disorder-prediction methods and several biophysical techniques showed that ca. 70% of the SBP2 sequence is disordered, whereas the RNA binding domain appears to be folded and functional. These results are consistent with a recent report on the role of the Hsp90 chaperone for the folding of SBP2 and other functionally unrelated proteins bearing an RNA binding domain homologous to SBP2.}, note = {1638-6183 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {ENNIFAR, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A fast selenium derivatization strategy for crystallization and phasing of RNA structures}, author = {V Olieric and U Rieder and K Lang and A Serganov and C Schulze-Briese and R Micura and P Dumas and E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19228585}, isbn = {19228585}, year = {2009}, date = {2009-01-01}, journal = {RNA}, volume = {15}, number = {4}, pages = {707-715}, abstract = {Site-specific 2'-methylseleno RNA labeling is a promising tool for tackling the phase problem in RNA crystallography. We have developed an efficient strategy for crystallization and structure determination of RNA and RNA/protein complexes based on preliminary crystallization screening of 2'-OCH(3)-modified RNA sequences, prior to the replacement of 2'-OCH(3) groups with their 2'-SeCH(3) counterparts. The method exploits the similar crystallization properties of 2'-OCH(3)- and 2'-SeCH(3)-modified RNAs and has been successfully validated for two test cases. In addition, our data show that 2'-SeCH(3)-modified RNA have an increased resistance to X-ray radiolysis in comparison with commonly used 5-halogen-modified RNA, which permits collection of experimental electron density maps of remarkable quality.}, note = {1469-9001 (Electronic) Journal Article Research Support, Non-U.S. Gov't Validation Studies}, keywords = {Base Sequence Crystallography, ENNIFAR, Unité ARN, X-Ray/*methods HIV-1/chemistry/genetics Nucleic Acid Conformation RNA/*chemistry Selenium/*metabolism Staining and Labeling/*methods}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Lentiviral-based vector and its use in directed evolution of genomic regions, genes and polynucleotides.}, author = {M Negroni and S Gallois-Montbrun and P Rossolillo and V Di Bartolo and G Uze and E Simon-Loriere and R Marquet and V Vivet-Boudou}, isbn = {EP 09290856.5}, year = {2009}, date = {2009-01-01}, abstract = {The present invention concerns a method of directing evolution of a target polynucleotide of interest for obtaining variants of this target polynucleotide, as well as a method to generate genetic variability by preparing a cell library. This invention also relates to a method to isolate or to screen variants of a polynucleotide or variants of a protein able to impact the phenotype of a cell or to confer a desired phenotype to target cells, and to identify these polynucleotide variants or protein variants responsible for this phenotype.}, keywords = {MARQUET, NEGRONI, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {A pH-responsive riboregulator}, author = {G Nechooshtan and M Elgrably-Weiss and A Sheaffer and E Westhof and S Altuvia}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19933154}, isbn = {19933154}, year = {2009}, date = {2009-01-01}, journal = {Genes Dev}, volume = {23}, number = {22}, pages = {2650-2662}, abstract = {The locus alx, which encodes a putative transporter, was discovered previously in a screen for genes induced under extreme alkaline conditions. Here we show that the RNA region preceding the alx ORF acts as a pH-responsive element, which, in response to high pH, leads to an increase in alx expression. Under normal growth conditions this RNA region forms a translationally inactive structure, but when exposed to high pH, a translationally active structure is formed to produce Alx. Formation of the active structure occurs while transcription is in progress under alkaline conditions and involves pausing of RNA polymerase at two distinct sites. Alkali increases the longevity of pausing at these sites and thereby interferes with formation of the inactive structure and promotes folding of the active one. The alx locus represents the first example of a pH-responsive riboregulator of gene expression, introducing a novel regulatory mechanism that involves RNA folding dynamics driven by pH.}, note = {1549-5477 (Electronic) 0890-9369 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Bacterial Hydrogen-Ion Concentration Mutation/genetics Nucleic Acid Conformation RNA Precursors/chemistry RNA, Bacterial/chemistry/genetics, Base Pairing Conserved Sequence Escherichia coli/*genetics/*metabolism Escherichia coli Proteins/genetics *Gene Expression Regulation, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure-function insights into prokaryotic and eukaryotic translation initiation}, author = {A G Myasnikov and A Simonetti and S Marzi and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19493673}, isbn = {19493673}, year = {2009}, date = {2009-01-01}, journal = {Curr Opin Struct Biol}, volume = {19}, number = {3}, pages = {300-309}, abstract = {Translation initiation is the rate-limiting and most complexly regulated step of protein synthesis in prokaryotes and eukaryotes. In the last few years, cryo-electron microscopy has provided several novel insights into the universal process of translation initiation. Structures of prokaryotic 30S and 70S ribosomal initiation complexes with initiator transfer RNA (tRNA), messenger RNA (mRNA), and initiation factors have recently revealed the mechanism of initiator tRNA recruitment to the assembling ribosomal machinery, involving molecular rearrangements of the ribosome and associated factors. First three-dimensional pictures of the particularly complex eukaryotic translation initiation machinery have been obtained, revealing how initiation factors tune the ribosome for recruiting the mRNA. A comparison of the available prokaryotic and eukaryotic structures shows that--besides significant differences--some key ribosomal features are universally conserved.}, note = {1879-033X (Electronic) 0959-440X (Linking) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Amino Acid Sequence Eukaryotic Cells/*chemistry/*metabolism Humans Molecular Sequence Data *Peptide Chain Initiation, Transfer/chemistry/metabolism Ribosomes/chemistry/genetics/metabolism, Translational Peptide Initiation Factors/chemistry/metabolism Prokaryotic Cells/*chemistry/*metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Crc global regulator binds to an unpaired A-rich motif at the Pseudomonas putida alkS mRNA coding sequence and inhibits translation initiation}, author = {R Moreno and S Marzi and P Romby and F Rojo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19825982}, isbn = {19825982}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {22}, pages = {7678-7690}, abstract = {Crc is a key global translational regulator in Pseudomonads that orchestrates the hierarchy of induction of several catabolic pathways for amino acids, sugars, hydrocarbons or aromatic compounds. In the presence of amino acids, which are preferred carbon sources, Crc inhibits translation of the Pseudomonas putida alkS and benR mRNAs, which code for transcriptional regulators of genes required to assimilate alkanes (hydrocarbons) and benzoate (an aromatic compound), respectively. Crc binds to the 5'-end of these mRNAs, but the sequence and/or structure recognized, and the way in which it inhibits translation, were unknown. We have determined the secondary structure of the alkS mRNA 5'-end through its sensitivity to several ribonucleases and chemical reagents. Footprinting and band-shift assays using variant alkS mRNAs have shown that Crc specifically binds to a short unpaired A-rich sequence located adjacent to the alkS AUG start codon. This interaction is stable enough to prevent formation of the translational initiation complex. A similar Crc-binding site was localized at benR mRNA, upstream of the Shine-Dalgarno sequence. This allowed predicting binding sites at other Crc-regulated genes, deriving a consensus sequence that will help to validate new Crc targets and to discriminate between direct and indirect effects of this regulator.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {5' Untranslated Regions Bacterial Proteins/genetics/*metabolism Binding Sites Nucleic Acid Conformation *Peptide Chain Initiation, Messenger/chemistry/metabolism Repressor Proteins/*metabolism Trans-Activators/*genetics, ROMBY, Translational Pseudomonas putida/*genetics RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The ribozyme core of group II introns: a structure in want of partners}, author = {F Michel and M Costa and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19299141}, isbn = {19299141}, year = {2009}, date = {2009-01-01}, journal = {Trends Biochem Sci}, volume = {34}, number = {4}, pages = {189-199}, abstract = {Group II introns contain a large ribozyme, which catalyzes self-splicing, and the coding sequence of a reverse transcriptase, the function of which is to cooperate with the ribozyme to achieve genomic mobility. Despite its lack of substrates for both steps of the splicing process, the crystal structure of a group II ribozyme reveals the location of two metal ions most likely to be involved in catalysis; the RNA structure that binds to these ions results from the bending of a local motif by the folding of the rest of the ribozyme. The stage is now set to determine where the intron-encoded protein binds to its partner and whether the spliceosome uses a counterpart of the group II catalytic center to excise nuclear pre-messenger introns.}, note = {0968-0004 (Print) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Tertiary network in mammalian mitochondrial tRNAAsp revealed by solution probing and phylogeny}, author = {M Messmer and J Putz and T Suzuki and C Sauter and M Sissler and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19767615}, isbn = {19767615}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {20}, pages = {6881-6895}, abstract = {Primary and secondary structures of mammalian mitochondrial (mt) tRNAs are divergent from canonical tRNA structures due to highly skewed nucleotide content and large size variability of D- and T-loops. The nonconservation of nucleotides involved in the expected network of tertiary interactions calls into question the rules governing a functional L-shaped three-dimensional (3D) structure. Here, we report the solution structure of human mt-tRNA(Asp) in its native post-transcriptionally modified form and as an in vitro transcript. Probing performed with nuclease S1, ribonuclease V1, dimethylsulfate, diethylpyrocarbonate and lead, revealed several secondary structures for the in vitro transcribed mt-tRNA(Asp) including predominantly the cloverleaf. On the contrary, the native tRNA(Asp) folds into a single cloverleaf structure, highlighting the contribution of the four newly identified post-transcriptional modifications to correct folding. Reactivities of nucleotides and phosphodiester bonds in the native tRNA favor existence of a full set of six classical tertiary interactions between the D-domain and the variable region, forming the core of the 3D structure. Reactivities of D- and T-loop nucleotides support an absence of interactions between these domains. According to multiple sequence alignments and search for conservation of Leontis-Westhof interactions, the tertiary network core building rules apply to all tRNA(Asp) from mammalian mitochondria.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Asp/*chemistry/*metabolism Transcription, Base Sequence Databases, FLORENTZ, FRUGIER, Genetic, Nucleic Acid Humans Molecular Sequence Data Nucleic Acid Conformation Phylogeny RNA/*chemistry/*metabolism RNA, SAUTER, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pathology-related mutation A7526G (A9G) helps in the understanding of the 3D structural core of human mitochondrial tRNA(Asp)}, author = {M Messmer and A Gaudry and M Sissler and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19535463}, isbn = {19535463}, year = {2009}, date = {2009-01-01}, journal = {RNA}, volume = {15}, number = {8}, pages = {1462-1468}, abstract = {More than 130 mutations in human mitochondrial tRNA (mt-tRNA) genes have been correlated with a variety of neurodegenerative and neuromuscular disorders. Their molecular impacts are of mosaic type, affecting various stages of tRNA biogenesis, structure, and/or functions in mt-translation. Knowledge of mammalian mt-tRNA structures per se remains scarce however. Primary and secondary structures deviate from classical tRNAs, while rules for three-dimensional (3D) folding are almost unknown. Here, we take advantage of a myopathy-related mutation A7526G (A9G) in mt-tRNA(Asp) to investigate both the primary molecular impact underlying the pathology and the role of nucleotide 9 in the network of 3D tertiary interactions. Experimental evidence is presented for existence of a 9-12-23 triple in human mt-tRNA(Asp) with a strongly conserved interaction scheme in mammalian mt-tRNAs. Mutation A7526G disrupts the triple interaction and in turn reduces aspartylation efficiency.}, note = {1469-9001 (Electronic) Letter Research Support, Non-U.S. Gov't}, keywords = {Asp/*chemistry/*genetics/metabolism Transfer RNA Aminoacylation/genetics, Binding Sites/genetics Humans Kinetics Mitochondrial Myopathies/genetics/metabolism/pathology Models, FLORENTZ, Missense Nucleic Acid Conformation RNA/*chemistry/*genetics/metabolism RNA, Molecular Mutation, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Peculiar inhibition of human mitochondrial aspartyl-tRNA synthetase by adenylate analogs}, author = {M Messmer and S P Blais and C Balg and R Chenevert and L Grenier and P Lague and C Sauter and M Sissler and R Giege and J Lapointe and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19254750}, isbn = {19254750}, year = {2009}, date = {2009-01-01}, journal = {Biochimie}, volume = {91}, number = {5}, pages = {596-603}, abstract = {Human mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs), the enzymes which esterify tRNAs with the cognate specific amino acid, form mainly a different set of proteins than those involved in the cytosolic translation machinery. Many of the mt-aaRSs are of bacterial-type in regard of sequence and modular structural organization. However, the few enzymes investigated so far do have peculiar biochemical and enzymological properties such as decreased solubility, decreased specific activity and enlarged spectra of substrate tRNAs (of same specificity but from various organisms and kingdoms), as compared to bacterial aaRSs. Here the sensitivity of human mitochondrial aspartyl-tRNA synthetase (AspRS) to small substrate analogs (non-hydrolysable adenylates) known as inhibitors of Escherichia coli and Pseudomonas aeruginosa AspRSs is evaluated and compared to the sensitivity of eukaryal cytosolic human and bovine AspRSs. L-aspartol-adenylate (aspartol-AMP) is a competitive inhibitor of aspartylation by mitochondrial as well as cytosolic mammalian AspRSs, with K(i) values in the micromolar range (4-27 microM for human mt- and mammalian cyt-AspRSs). 5'-O-[N-(L-aspartyl)sulfamoyl]adenosine (Asp-AMS) is a 500-fold stronger competitive inhibitor of the mitochondrial enzyme than aspartol-AMP (10nM) and a 35-fold lower competitor of human and bovine cyt-AspRSs (300 nM). The higher sensitivity of human mt-AspRS for both inhibitors as compared to either bacterial or mammalian cytosolic enzymes, is not correlated with clear-cut structural features in the catalytic site as deduced from docking experiments, but may result from dynamic events. In the scope of new antibacterial strategies directed against aaRSs, possible side effects of such drugs on the mitochondrial human aaRSs should thus be considered.}, note = {1638-6183 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Adenosine Monophosphate/*analogs & derivatives/*pharmacology Animals Aspartate-tRNA Ligase/*antagonists & inhibitors/*chemistry/metabolism Catalytic Domain Cattle Humans Mitochondria/*drug effects/*enzymology Molecular Structure Structure-Activity Relationship, FLORENTZ, SAUTER, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal growth of proteins, nucleic acids, and viruses in gels}, author = {B Lorber and C Sauter and A Théobald-Dietrich and A Moreno and P Schellenberger and M C Robert and B Capelle and S Sanglier and N Potier and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20005247}, isbn = {20005247}, year = {2009}, date = {2009-01-01}, journal = {Prog Biophys Mol Biol}, volume = {101}, number = {1-3}, pages = {13-25}, abstract = {Medium-sized single crystals with perfect habits and no defect producing intense and well-resolved diffraction patterns are the dream of every protein crystallographer. Crystals of biological macromolecules possessing these characteristics can be prepared within a medium in which mass transport is restricted to diffusion. Chemical gels (like polysiloxane) and physical gels (such as agarose) provide such an environment and are therefore suitable for the crystallisation of biological macromolecules. Instructions for the preparation of each type of gel are given to urge crystal growers to apply diffusive media for enhancing crystallographic quality of their crystals. Examples of quality enhancement achieved with silica and agarose gels are given. Results obtained with other substances forming gel-like media (such as lipidic phases and cellulose derivatives) are presented. Finally, the use of gels in combination with capillary tubes for counter-diffusion experiments is discussed. Methods and techniques implemented with proteins can also be applied to nucleic acids and nucleoprotein assemblies such as viruses.}, note = {1873-1732 (Electronic) 0079-6107 (Linking) Journal article}, keywords = {FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selenoprotein function and muscle disease}, author = {A Lescure and M Rederstorff and A Krol and P Guicheney and V Allamand}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19285112}, isbn = {19285112}, year = {2009}, date = {2009-01-01}, journal = {Biochim Biophys Acta-Gen Subj}, volume = {1790}, number = {11}, pages = {1569-1574}, abstract = {The crucial role of the trace element selenium in livestock and human health, in particular in striated muscle function, has been well established but the underlying molecular mechanisms remain poorly understood. Over the last decade, identification of the full repertoire of selenium-containing proteins has opened the way towards a better characterization of these processes. Two selenoproteins have mainly been investigated in muscle, namely SelW and SelN. Here we address their involvement in muscle development and maintenance, through the characterization of various cellular or animal models. In particular, mutations in the SEPN1 gene encoding selenoprotein N (SelN) cause a group of neuromuscular disorders now referred to as SEPN1-related myopathy. Recent findings on the functional consequences of these mutations suggest an important contribution of SelN to the regulation of oxidative stress and calcium homeostasis. Importantly, the conclusions of these experiments have opened new avenues of investigations that provide grounds for the development of therapeutic approaches.}, note = {0006-3002 (Print) 0006-3002 (Linking) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Biological Muscle Proteins/genetics/physiology Muscles/physiology Muscular Diseases/*etiology/genetics Mutation/physiology Selenoproteins/genetics/*physiology, KROL Animals Calcium/metabolism Humans Intracellular Fluid/metabolism Models, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{wilkins_differences_2009, title = {Differences in hydroxylation and binding of Notch and HIF-1alpha demonstrate substrate selectivity for factor inhibiting HIF-1 (FIH-1).}, author = {Sarah E Wilkins and Jaana Hyvärinen and Johana Chicher and Jeffrey J Gorman and Daniel J Peet and Rebecca L Bilton and Peppi Koivunen}, doi = {10.1016/j.biocel.2009.01.005}, issn = {1878-5875 1357-2725}, year = {2009}, date = {2009-01-01}, journal = {The international journal of biochemistry & cell biology}, volume = {41}, number = {7}, pages = {1563--1571}, abstract = {FIH-1, factor inhibiting hypoxia-inducible factor-1 (HIF-1), regulates oxygen sensing by hydroxylating an asparagine within HIF-alpha. It also hydroxylates asparagines in many proteins containing ankyrin repeats, including Notch1-3, p105 and I?B?. Relative binding affinity and hydroxylation rate are crucial determinants of substrate selection and modification. We determined the contributions of substrate sequence composition and length and of oxygen concentration to the FIH-1-binding and/or hydroxylation of Notch1-4 and compared them with those for HIF-1alpha. We also demonstrated hydroxylation of two asparagines in Notch2 and 3, corresponding to Sites 1 and 2 of Notch1, by mass spectrometry for the first time. Our data demonstrate that substrate length has a much greater influence on FIH-1-dependent hydroxylation of Notch than of HIF-1alpha, predominantly through binding affinity rather than maximal reaction velocity. The K(m) value of FIH-1 for Notch1, textless 0.2 microM, is at least 250-fold lower than that of 50 microM for HIF-1alpha. Site 1 of Notch1-3 appeared the preferred site of FIH-1 hydroxylation in these substrates. Interestingly, binding of Notch4 to FIH-1 was observed with an affinity almost 10-fold lower than for Notch1-3, but no hydroxylation was detected. Importantly, we demonstrate that the K(m) of FIH-1 for oxygen at the preferred Site 1 of Notch1-3, 10-19 microM, is an order of magnitude lower than that for Site 2 or HIF-1alpha. Hence, at least during in vitro hydroxylation, Notch is likely to become efficiently hydroxylated by FIH-1 even under relatively severe hypoxic conditions, where HIF-1alpha hydroxylation would be reduced.}, note = {Place: Netherlands}, keywords = {alpha Subunit/*metabolism, Amino Acid Sequence, Animals, Asparagine/metabolism, Humans, Hydroxylation, Hypoxia-Inducible Factor 1, Kinetics, Mice, Mixed Function Oxygenases, Molecular Sequence Data, Notch/chemistry/*metabolism, Oxygen/metabolism, Peptides/chemistry/metabolism, PPSE, Protein Binding, Receptors, Recombinant Proteins/metabolism, Repressor Proteins/*metabolism, Substrate Specificity}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Exploring the }, author = {J Kondo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19764470}, isbn = {19764470}, year = {2009}, date = {2009-01-01}, journal = {Tanpakushitsu Kakusan Koso}, volume = {54}, number = {11}, pages = {1356-1362}, note = {0039-9450 (Print) 0039-9450 (Linking) Journal Article Review}, keywords = {Anti-Bacterial Agents/adverse effects/pharmacology Bacteria/drug effects *Binding Sites Crystallography, Transfer RNA, Unité ARN, Untranslated Ribosomes/chemistry/*genetics/*physiology, WESTHOF, X-Ray Hearing Disorders/genetics Humans Mutation Protein Biosynthesis/genetics *RNA/genetics RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {The A-rich RNA sequences of HIV-1 pol are important for the synthesis of viral cDNA}, author = {C P Keating and M K Hill and D J Hawkes and R P Smyth and C Isel and S Y Le and A C Palmenberg and J A Marshall and R Marquet and G J Nabel and J Mak}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19106143}, isbn = {19106143}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {3}, pages = {945-956}, abstract = {The bias of A-rich codons in HIV-1 pol is thought to be a record of hypermutations in viral genomes that lack biological functions. Bioinformatic analysis predicted that A-rich sequences are generally associated with minimal local RNA structures. Using codon modifications to reduce the amount of A-rich sequences within HIV-1 genomes, we have reduced the flexibility of RNA sequences in pol to analyze the functional significance of these A-rich 'structurally poor' RNA elements in HIV-1 pol. Our data showed that codon modification of HIV-1 sequences led to a suppression of virus infectivity by 5-100-fold, and this defect does not correlate with, viral entry, viral protein expression levels, viral protein profiles or virion packaging of genomic RNA. Codon modification of HIV-1 pol correlated with an enhanced dimer stability of the viral RNA genome, which was associated with a reduction of viral cDNA synthesis both during HIV-1 infection and in a cell free reverse transcription assay. Our data provided direct evidence that the HIV-1 A-rich pol sequence is not merely an evolutionary artifact of enzyme-induced hypermutations, and that HIV-1 has adapted to rely on A-rich RNA sequences to support the synthesis of viral cDNA during reverse transcription, highlighting the utility of using 'structurally poor' RNA domains in regulating biological process.}, note = {1362-4962 (Electronic) Journal Article Research Support, N.I.H., Intramural Research Support, Non-U.S. Gov't}, keywords = {Adenine/analysis Base Sequence Cell Line Codon DNA, Complementary/*biosynthesis DNA, MARQUET, PAILLART, pol HIV-1/*genetics/physiology Humans Nucleic Acid Conformation RNA, Ribonucleic Acid *Reverse Transcription Viral Proteins/metabolism Virion/metabolism Virus Internalization Virus Replication, Unité ARN, Viral/*biosynthesis Dimerization *Genes, Viral/*chemistry *Regulatory Sequences}, pubstate = {published}, tppubtype = {article} } @article{, title = {Large telomerase RNA, telomere length heterogeneity and escape from senescence in Candida glabrata}, author = {R Kachouri-Lafond and B Dujon and E Gilson and E Westhof and C Fairhead and M T Teixeira}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19840797}, isbn = {19840797}, year = {2009}, date = {2009-01-01}, journal = {FEBS Lett}, volume = {583}, number = {22}, pages = {3605-3610}, abstract = {Telomerase, the key enzyme essential for the maintenance of eukaryotic chromosome ends, contains a reverse transcriptase and an RNA that provides the template for the synthesis of telomeric repeats. Here, we characterize the telomerase subunits in the hemiascomycete yeast Candida glabrata. We propose a secondary structure model for the telomerase RNA that is the largest described to date. Telomerase deletion mutants show a progressive shortening of telomeres and a modest loss of viability. Frequent post-senescence survivors emerge that possess long telomeric repeat tracts. We suggest that the high telomere length heterogeneity accounts for this distinct senescence phenotype.}, note = {1873-3468 (Electronic) 0014-5793 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Base Sequence Blotting, Fungal/chemistry/*genetics Sequence Homology, Fungal/genetics Flow Cytometry Gene Deletion Molecular Sequence Data Mutation Nucleic Acid Conformation RNA/chemistry/*genetics RNA, Nucleic Acid Telomerase/chemistry/*genetics Telomere/*genetics, Southern Candida glabrata/enzymology/*genetics/growth & development Cell Division DNA, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNAdb 2009: compilation of tRNA sequences and tRNA genes}, author = {F Juhling and M Morl and R K Hartmann and M Sprinzl and P F Stadler and J Putz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18957446}, isbn = {18957446}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {Database issue}, pages = {D159-162}, abstract = {One of the first specialized collections of nucleic acid sequences in life sciences was the 'compilation of tRNA sequences and sequences of tRNA genes' (http://www.trna.uni-bayreuth.de). Here, an updated and completely restructured version of this compilation is presented (http://trnadb.bioinf.uni-leipzig.de). The new database, tRNAdb, is hosted and maintained in cooperation between the universities of Leipzig, Marburg, and Strasbourg. Reimplemented as a relational database, tRNAdb will be updated periodically and is searchable in a highly flexible and user-friendly way. Currently, it contains more than 12 000 tRNA genes, classified into families according to amino acid specificity. Furthermore, the implementation of the NCBI taxonomy tree facilitates phylogeny-related queries. The database provides various services including graphical representations of tRNA secondary structures, a customizable output of aligned or un-aligned sequences with a variety of individual and combinable search criteria, as well as the construction of consensus sequences for any selected set of tRNAs.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A novel stem loop control element-dependent UGA read-through system without translational selenocysteine incorporation in Drosophila}, author = {M Hirosawa-Takamori and D Ossipov and S V Novoselov and A A Turanov and Y Zhang and V N Gladyshev and A Krol and G Vorbruggen and H Jackle}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18772345}, isbn = {18772345}, year = {2009}, date = {2009-01-01}, journal = {Faseb J}, volume = {23}, number = {1}, pages = {107-113}, abstract = {Translational read-through of the UGA stop codon is an evolutionarily conserved feature that most prominently represents the basis of selenoprotein biosynthesis. It requires a specific cis-acting stem loop control element, termed SECIS, which is located in the 3'-untranslated region of eukaryotic selenoprotein mRNAs. In a search for novel factors underlying the SECIS-directed UGA read-through process, we identified an evolutionary conserved GTPase-activating protein, termed GAPsec. We show that the activity of the Drosophila GAPsec (dGAPsec) is necessary to support SECIS-dependent UGA read-through activity in flies and the mouse homolog mGAPsec in mice tissue culture cells. However, selenoprotein biosynthesis is not impaired in flies that lack dGAPsec activity. The results indicate that GAPsec is part of a novel SECIS-dependent translational read-through system that does not involve selenocysteine incorporation.}, note = {1530-6860 (Electronic) Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't}, keywords = {KROL, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Tumultuous relationship between the human immunodeficiency virus type 1 viral infectivity factor (Vif) and the human APOBEC-3G and APOBEC-3F restriction factors}, author = {S Henriet and G Mercenne and S Bernacchi and J C Paillart and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19487726}, isbn = {19487726}, year = {2009}, date = {2009-01-01}, journal = {Microbiol Mol Biol Rev}, volume = {73}, number = {2}, pages = {211-232}, abstract = {The viral infectivity factor (Vif) is dispensable for human immunodeficiency virus type 1 (HIV-1) replication in so-called permissive cells but is required for replication in nonpermissive cell lines and for pathogenesis. Virions produced in the absence of Vif have an aberrant morphology and an unstable core and are unable to complete reverse transcription. Recent studies demonstrated that human APOBEC-3G (hA3G) and APOBEC-3F (hA3F), which are selectively expressed in nonpermissive cells, possess strong anti-HIV-1 activity and are sufficient to confer a nonpermissive phenotype. Vif induces the degradation of hA3G and hA3F, suggesting that its main function is to counteract these cellular factors. Most studies focused on the hypermutation induced by the cytidine deaminase activity of hA3G and hA3F and on their Vif-induced degradation by the proteasome. However, recent studies suggested that several mechanisms are involved both in the antiviral activity of hA3G and hA3F and in the way Vif counteracts these antiviral factors. Attempts to reconcile the studies involving Vif in virus assembly and stability with these recent findings suggest that hA3G and hA3F partially exert their antiviral activity independently of their catalytic activity by destabilizing the viral core and the reverse transcription complex, possibly by interfering with the assembly and/or maturation of the viral particles. Vif could then counteract hA3G and hA3F by excluding them from the viral assembly intermediates through competition for the viral genomic RNA, by regulating the proteolytic processing of Pr55(Gag), by enhancing the efficiency of the reverse transcription process, and by inhibiting the enzymatic activities of hA3G and hA3F.}, note = {1098-5557 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Exploring the "motion" = "function" of the ribosomal A-site molecular switch]}, author = { J. Kondo}, year = {2009}, date = {2009-01-01}, journal = {Tanpakushitsu Kakusan Koso}, volume = {54}, number = {11}, pages = {1356-62}, note = {0039-9450 (Print) 0039-9450 (Linking) Journal Article Review}, keywords = {*Binding, *RNA/genetics, Agents/adverse, Anti-Bacterial, Bacteria/drug, Biosynthesis/genetics, Crystallography, Disorders/genetics, effects, effects/pharmacology, Hearing, Humans, Mutation, Protein, Ribosomes/chemistry/*genetics/*physiology, RNA, Sites, Transfer, Untranslated, WESTHOF, X-Ray}, pubstate = {published}, tppubtype = {article} } @article{, title = {A short guide for molecular dynamics simulations of RNA systems}, author = {Y Hashem and P Auffinger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18930152}, isbn = {18930152}, year = {2009}, date = {2009-01-01}, journal = {Methods}, volume = {47}, number = {3}, pages = {187-197}, abstract = {As a result of important methodological advances and of the rapid growth of experimental data, the number of molecular dynamics (MD) simulations related to RNA systems has significantly increased. However, such MD simulations are not straightforward and great care has to be exerted during the setup stage in order to choose the appropriate MD package, force fields and ionic conditions. Furthermore, the choice and a correct evaluation of the main characteristics of the starting structure are primordial for the generation of informative and reliable MD trajectories since experimental structures are not void of inaccuracies and errors. The aim of this review is to provide, through numerous examples, practical guidelines for the setup of MD simulations, the choice of ionic conditions and the detection and correction of experimental inaccuracies in order to start MD simulations of nucleic acid systems under the best auspices.}, note = {1095-9130 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The unforeseeable hammerhead ribozyme}, author = {C Hammann and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20948624}, doi = {10.3410/B1-6}, issn = {1757-594X (Electronic) 1757-594X (Linking)}, year = {2009}, date = {2009-01-01}, journal = {F1000 Biol Rep}, volume = {1}, pages = {6}, abstract = {Despite its small size, the complex behavior of the hammerhead ribozyme keeps surprising us, even more than 20 years after its discovery. Here, we summarize recent developments in the field, in particular the discovery of the first split hammerhead ribozyme.}, note = {Hammann, Christian Westhof, Eric England F1000 biology reports F1000 Biol Rep. 2009 Jan 21;1. pii: 6.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Kinetics of deoxy-CTP incorporation opposite a dG-C8-N-2-aminofluorene adduct by a high-fidelity DNA polymerase}, author = { D. Y. Burnouf and J. E. Wagner}, year = {2009}, date = {2009-01-01}, journal = {J Mol Biol}, volume = {386}, number = {4}, pages = {951-61}, abstract = {The model carcinogen N-2-acetylaminofluorene covalently binds to the C8 position of guanine to form two adducts, the N-(2'-deoxyguanosine-8-yl)-aminofluorene (G-AF) and the N-2-(2'-deoxyguanosine-8-yl)-acetylaminofluorene (G-AAF). Although they are chemically closely related, their biological effects are strongly different and they are processed by different damage tolerance pathways. G-AF is bypassed by replicative and high-fidelity polymerases, while specialized polymerases ensure synthesis past of G-AAF. We used the DNA polymerase I fragment of a Bacillus stearothermophilus strain as a model for a high-fidelity polymerase to study the kinetics of incorporation of deoxy-CTP (dCTP) opposite a single G-AF. Pre-steady-state kinetic experiments revealed a drastic reduction in dCTP incorporation performed by the G-AF-modified ternary complex. Two populations of these ternary complexes were identified: (i) a minor productive fraction (20%) that readily incorporates dCTP opposite the G-AF adduct with a rate similar to that measured for the adduct-free ternary complexes and (ii) a major fraction of unproductive complexes (80%) that slowly evolve into productive ones. In the light of structural data, we suggest that this slow rate reflects the translocation of the modified base within the active site, from the pre-insertion site into the insertion site. By making this translocation rate limiting, the G-AF lesion reveals a novel kinetic step occurring after dNTP binding and before chemistry.}, note = {1089-8638 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Adducts, Bacillus, Catalytic, Cytidine, Deoxyguanosine/*metabolism, DNA, DNA-Directed, Domain, DUMAS, Elements, Fluorenes/*metabolism, Guanine, Kinetics, Oligonucleotides/metabolism, Phosphorothioate, Polymerase/*metabolism, Specificity, stearothermophilus/enzymology, Substrate, Titrimetry, Triphosphate/*metabolism}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Transfer RNA Aminoacylation and Modified Nucleosides}, author = {R Giege and J Lapointe}, editor = {H Grosjean}, url = {http://www.landesbioscience.com/curie/chapter/4164}, year = {2009}, date = {2009-01-01}, booktitle = {DNA and RNA Modification Enzymes: Structure, Mechanism, Function and Evolution}, publisher = {Landes Bioscience}, address = {Georgetown, Texas, U.S.A}, abstract = {Among RNAs, the transfer RNAs are those showing the highest level of posttranscriptional modifications. After an overview on early data, the chapter discusses the present knowledge on the role modified nucleosides have on tRNA structure and function with emphasis on tRNA aminoacylation. The concept of tRNA aminoacylation identity will be outlined and the cases discussed where individual modified nucleosides act either as positive determinants (for recognition by the cognate synthetases) or negative antideterminants (preventing recognition by a noncognate synthetase). Furthermore, the collective participation of the ensemble of modified nucleosides in a given tRNA will also be analyzed. Evolutionary aspects will be illustrated by the unprecedented property of a paralog of bacterial glutamyl‑tRNA synthetase restricted to the catalytic module of the synthetase, that aminoacylates the Q‑base of bacterial tRNAAsp. This has evolutionary implications suggesting that modern tRNA originated by duplication of an ancestral minihelix and finds support with the existence of sequence similarities between the anticodon stem‑loop of tRNAAsp and the accepting end of tRNAGlu. Altogether and contrarily to a common belief, posttranscriptional modifications in tRNA play an active role in a majority of aminoacylation systems, although in many cases by indirect structure‑dependent effects.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Transfer RNA recognition by synthetases}, author = {R Giege and G Eriani}, url = {http://www.els.net/WileyCDA/ElsArticle/refId-a0000531.html}, doi = {10.1002/9780470015902.a0000531.pub2}, year = {2009}, date = {2009-01-01}, booktitle = {Encyclopedia of Life Sciences}, publisher = {John Wiley & Sons}, abstract = {Fidelity of transfer ribonucleic acid (tRNA) charging by amino acids ensures correct translation of the genetic code into proteins. Charging is catalysed by a set of enzymes known as aminoacyl-tRNA synthetases. Owing to the degeneracy of the genetic code, some of the different tRNAs have the same amino acid attached to them. Specificity of the charging reaction is ensured by positive elements, the identity determinants unique to each tRNA and responsible for its recognition by the cognate synthetase, and negative elements, the antideterminants that prevent false recognitions. To fulfil the aminoacylation specificity and prevent noncognate aminoacyl-tRNA delivery to the ribosome, some synthetases also mediate proofreading reactions that increase fidelity of the tRNA charging. In such reactions, misactivated amino acids or mischarged tRNAs are checked in specific sites and noncognate products are hydrolysed.}, keywords = {Aminoacyl-tRNA synthetase Genetic code Protein synthesis RNA recognition tRNA, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {On the role of an unusual tRNAGly isoacceptor in Staphylococcus aureus}, author = {S Giannouli and A Kyritsis and N Malissovas and H D Becker and C Stathopoulos}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19014993}, isbn = {19014993}, year = {2009}, date = {2009-01-01}, journal = {Biochimie}, volume = {91}, number = {3}, pages = {344-351}, abstract = {In the available Staphylococcus aureus genomes, four different genes have been annotated to encode tRNA(Gly) isoacceptors. Besides their prominent role in protein synthesis, some of them also participate in the formation of pentaglycine bridges during cell wall synthesis. However, until today, it is not known how many and which of them are actually involved in this essential procedure. In the present study we identified, apart from the four annotated tRNA(Gly) genes, a putative pseudogene which encodes and expresses an unusual fifth tRNA(Gly) isoacceptor in S. aureus (as detected via RT-PCR and subsequent direct sequencing analysis). All the in vitro transcribed tRNA(Gly) molecules (including the "pseudogene-encoded" tRNA(Gly)) can be efficiently aminoacylated by the recombinant S. aureus glycyl-tRNA synthetase. Furthermore, bioinformatic analysis suggests that the "pseudo"-tRNA(Gly(UCC)) identified in the present study and two of the annotated isoacceptors bearing the same anticodon carry specific sequence elements that do not favour the strong interaction with EF-Tu that proteinogenic tRNAs would promote. This observation was verified by the differential capacity of Gly-tRNA(Gly) molecules to form ternary complexes with activated S. aureus EF-Tu.GTP. These tRNA(Gly) molecules display high sequence similarities with their S. epidermidis orthologs which also actively participate in cell wall synthesis. Both bioinformatic and biochemical data suggest that in S. aureus these three glycylated tRNA(Gly) isoacceptors that are weak EF-Tu binders, possibly escape protein synthesis and serve as glycine donors for the formation of pentaglycine bridges that are essential for stabilization of the staphylococcal cell wall.}, note = {1638-6183 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Bacterial Glycine-tRNA Ligase/genetics/*metabolism Peptide Elongation Factor Tu/metabolism RNA, Gly/*genetics/*metabolism Recombinant Proteins/metabolism Sequence Analysis, KERN Anticodon/metabolism Computational Biology/methods Genes, RNA Staphylococcus aureus/*genetics/*metabolism Transfer RNA Aminoacylation/genetics, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The role of mRNA structure in translational control in bacteria}, author = {T Geissmann and S Marzi and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19885993}, isbn = {19885993}, year = {2009}, date = {2009-01-01}, journal = {RNA Biol}, volume = {6}, number = {2}, pages = {153-160}, abstract = {During the past few years, our knowledge on RNA-based regulation in many organisms has tremendously increased. In bacteria, although transcriptional regulatory proteins remain key players in gene regulation, a wide variety of post-transcriptional regulatory mechanisms discovered highlights the importance of the mRNA structure in the regulation of gene expression. RNA-dependent regulation largely contributes to rapidly adapt the bacterial metabolism in response to environmental changes, stress and in establishment of virulence. Bacteria exploit the extraordinary ability of mRNA to fold into different structures in response to various signals (environmental cues, ligand binding). Induced mRNA conformational rearrangements can potentially regulate transcription, translation and mRNA stability. The present review focuses on the structures of regulatory regions of mRNA that have evolved to permit productive interactions with trans-acting regulators, such as protein or non-coding RNAs. Finally, we describe how particular properties of these regulatory complexes regulate translation initiation.}, note = {1555-8584 (Electronic) 1547-6286 (Linking) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Amino Acid, Amino Acid Sequence Bacteria/*genetics Bacterial Proteins/chemistry/genetics Molecular Sequence Data *Nucleic Acid Conformation *Protein Biosynthesis RNA, Messenger/*chemistry Sequence Homology, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A search for small noncoding RNAs in Staphylococcus aureus reveals a conserved sequence motif for regulation}, author = {T Geissmann and C Chevalier and M J Cros and S Boisset and P Fechter and C Noirot and J Schrenzel and P Francois and F Vandenesch and C Gaspin and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19786493}, isbn = {19786493}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {21}, pages = {7239-7257}, abstract = {Bioinformatic analysis of the intergenic regions of Staphylococcus aureus predicted multiple regulatory regions. From this analysis, we characterized 11 novel noncoding RNAs (RsaA-K) that are expressed in several S. aureus strains under different experimental conditions. Many of them accumulate in the late-exponential phase of growth. All ncRNAs are stable and their expression is Hfq-independent. The transcription of several of them is regulated by the alternative sigma B factor (RsaA, D and F) while the expression of RsaE is agrA-dependent. Six of these ncRNAs are specific to S. aureus, four are conserved in other Staphylococci, and RsaE is also present in Bacillaceae. Transcriptomic and proteomic analysis indicated that RsaE regulates the synthesis of proteins involved in various metabolic pathways. Phylogenetic analysis combined with RNA structure probing, searches for RsaE-mRNA base pairing, and toeprinting assays indicate that a conserved and unpaired UCCC sequence motif of RsaE binds to target mRNAs and prevents the formation of the ribosomal initiation complex. This study unexpectedly shows that most of the novel ncRNAs carry the conserved C-rich motif, suggesting that they are members of a class of ncRNAs that target mRNAs by a shared mechanism.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Bacillus subtilis/genetics/metabolism Base Sequence Computational Biology Conserved Sequence Gene Expression Profiling *Gene Expression Regulation, Bacterial Molecular Sequence Data Proteomics RNA Stability RNA, Genetic, Messenger/metabolism RNA, ROMBY, Unité ARN, Untranslated/*chemistry/genetics/metabolism Staphylococcus aureus/*genetics Transcription}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Retroviruses}, author = {R Galetto and M Negroni}, editor = {C Cameron and M Gotte and K Raney}, url = {http://www.springerlink.com/content/w873085p48771805}, year = {2009}, date = {2009-01-01}, booktitle = {Viral Genome Replication}, pages = {109-128}, publisher = {Springer-Verlag}, abstract = {Retroviruses are a large group of enveloped RNA viruses infecting vertebrates. The viral particles are spherical and acquire their envelope during budding from the infected cell. The lipid bilayer therefore contains cellular proteins as well as the viral envelope glycoproteins. These glycoproteins are constituted by a transmembrane subunit (TM) associated to the surface protein (SU), present on the virion. Underneath the membrane is a spherical shell constituted by the matrix (MA) protein. Internally is the viral capsid, whose shape varies in different viruses, constituted by the CA protein. This core contains the retroviral enzymes (the reverse transcriptase, RT, the integrase, IN, and the protease, PR), together with the genomic RNA, coated by the nucleocapsid protein (NC).}, note = {DOI: 10.1007/b135974_6}, keywords = {NEGRONI, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Structure des nucléotides et des acides nucléiques}, author = {V Fritsch and E Westhof}, editor = {J H Weil}, url = {http://www.unitheque.com/Livre/dunod/Sciences_sup/Biochimie_generale-31587.html}, year = {2009}, date = {2009-01-01}, booktitle = {Biochemie Générale 11e Edition}, pages = {619-624}, publisher = {Dunod}, address = {Paris}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Molecular adaptation in RNA complexes}, author = {V Fritsch and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19523895}, isbn = {19523895}, year = {2009}, date = {2009-01-01}, journal = {Structure}, volume = {17}, number = {6}, pages = {784-786}, abstract = {In this issue, Batey and colleagues show that the purine riboswitch is able to bind purine analogs by exploiting various strategies, including lateral shifts of the bases and, most importantly, changes in ligand tautomeric form.}, note = {1878-4186 (Electronic) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast mitochondrial Gln-tRNA(Gln) is generated by a GatFAB-mediated transamidation pathway involving Arc1p-controlled subcellular sorting of cytosolic GluRS}, author = {M Frechin and B Senger and M Brayé and D Kern and R P Martin and H D Becker}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19417106}, isbn = {19417106}, year = {2009}, date = {2009-01-01}, journal = {Genes Dev}, volume = {23}, number = {9}, pages = {1119-1130}, abstract = {It is impossible to predict which pathway, direct glutaminylation of tRNA(Gln) or tRNA-dependent transamidation of glutamyl-tRNA(Gln), generates mitochondrial glutaminyl-tRNA(Gln) for protein synthesis in a given species. The report that yeast mitochondria import both cytosolic glutaminyl-tRNA synthetase and tRNA(Gln) has challenged the widespread use of the transamidation pathway in organelles. Here we demonstrate that yeast mitochondrial glutaminyl-tRNA(Gln) is in fact generated by a transamidation pathway involving a novel type of trimeric tRNA-dependent amidotransferase (AdT). More surprising is the fact that cytosolic glutamyl-tRNA synthetase ((c)ERS) is imported into mitochondria, where it constitutes the mitochondrial nondiscriminating ERS that generates the mitochondrial mischarged glutamyl-tRNA(Gln) substrate for the AdT. We show that dual localization of (c)ERS is controlled by binding to Arc1p, a tRNA nuclear export cofactor that behaves as a cytosolic anchoring platform for (c)ERS. Expression of Arc1p is down-regulated when yeast cells are switched from fermentation to respiratory metabolism, thus allowing increased import of (c)ERS to satisfy a higher demand of mitochondrial glutaminyl-tRNA(Gln) for mitochondrial protein synthesis. This novel strategy that enables a single protein to be localized in both the cytosol and mitochondria provides a new paradigm for regulation of the dynamic subcellular distribution of proteins between membrane-separated compartments.}, note = {1549-5477 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Amino Acyl/*metabolism RNA-Binding Proteins/*metabolism Saccharomyces cerevisiae/*enzymology/*metabolism Saccharomyces cerevisiae Proteins/*metabolism Transferases/*metabolism, Fungal Glutamate-tRNA Ligase/*metabolism Glutamic Acid/metabolism Mitochondria/*enzymology Protein Binding Protein Transport RNA, KERN Cytoplasm/enzymology Gene Expression Regulation, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Translating organellar glutamine codons: a case by case scenario?}, author = {M Frechin and A M Duchene and H D Becker}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19106621}, isbn = {19106621}, year = {2009}, date = {2009-01-01}, journal = {RNA Biol}, volume = {6}, number = {1}, pages = {31-34}, abstract = {Aminoacyl-tRNAs are generally formed by direct attachment of an amino acid to tRNAs by aminoacyl-tRNA synthetases, but glutaminyl-tRNA (Q-tRNA) is an exception to this rule. Glutaminyl-tRNA(Gln) (Q-tRNA(Q)) is formed by this direct pathway in the eukaryotic cytosol and in a small subset of bacteria, but is formed by an indirect transamidation pathway in most bacteria and archaea. To date it is almost impossible to predict what pathway generates organellar Q-tRNA(Q) in a given eukaryote. All eukaryotic genomes sequenced so far, display a single glutaminyl-tRNA synthetase (QRS) gene which is at least responsible for the cytosolic QRS activity, as well as a gene coding for a mitochondrial ortholog of the essential GatB subunit of the tRNA-dependent amidotransferase (AdT). Indeed, QRS activity was found in protozoan mitochondria while AdT activity was characterized in plant organelles. The pathway for Q-tRNA(Q) synthesis in yeast and mammals mitochondria is still questionable.}, note = {1555-8584 (Electronic) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Biological Nitrogenous Group Transferases/metabolism Plants/metabolism RNA, KERN Amino Acyl-tRNA Synthetases/metabolism Animals Chloroplasts/metabolism Codon Cytosol/metabolism Glutamate-tRNA Ligase/metabolism Glutamine/*chemistry Mitochondria/metabolism Models, Messenger/metabolism RNA, Transfer/metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Organisation et expression des genomes des organites}, author = {C Florentz}, editor = {J H Weil}, url = {http://www.unitheque.com/Livre/dunod/Sciences_sup/Biochimie_generale-31587.html}, year = {2009}, date = {2009-01-01}, booktitle = {Biochemie Generale 11e edition}, pages = {619-624}, publisher = {Dunod}, address = {Paris}, keywords = {FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Ribosomal initiation complexes probed by toeprinting and effect of trans-acting translational regulators in bacteria}, author = {P Fechter and C Chevalier and G Yusupova and M Yusupov and P Romby and S Marzi}, editor = {A Serganov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19381565}, doi = {10.1007/978-1-59745-558-9_18}, isbn = {19381565}, year = {2009}, date = {2009-01-01}, booktitle = {Riboswitches: Methods and Protocols}, volume = {540}, pages = {247-263}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Toeprinting was developed to study the formation of ribosomal initiation complexes in bacteria. This approach, based on the inhibition of reverse transcriptase elongation, was used to monitor the effect of ribosomal components and translational factors on the formation of the active ribosomal initiation complex. Moreover, this method offers an easy way to study in vitro how mRNA conformational changes alter ribosome binding at the initiation site. These changes can be induced either by environmental cues (temperature, ion concentration), or by the binding of metabolites, regulatory proteins, and trans-acting RNAs. An experimental guide is given to follow the different steps of the formation of ribosomal initiation complexes in Escherichia coli and Staphylococcus aureus, and to monitor the mechanism of action of several regulators on translation initiation in vitro. Protocols to prepare the ribosome and the subunits are also given for Thermus thermophilus, Staphylococcus aureus, and Escherichia coli.}, keywords = {Bacteria/*genetics/*metabolism Bacterial Proteins/*metabolism Base Sequence Escherichia coli/metabolism Molecular Biology/*methods Molecular Sequence Data *Peptide Chain Initiation, Bacterial/chemistry/genetics Ribosomes/*metabolism Staphylococcus aureus/metabolism Thermus thermophilus/metabolism, ROMBY, Translational RNA, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Cytomegalovirus microRNAs}, author = {L Dolken and S Pfeffer and U H Koszinowski}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19291384}, isbn = {19291384}, year = {2009}, date = {2009-01-01}, journal = {Virus Genes}, volume = {38}, number = {3}, pages = {355-364}, abstract = {MicroRNAs (miRNAs) are small, non-coding RNA molecules that regulate gene expression at a post-transcriptional level in virtually all eukaryotic organisms and some viruses, particularly herpesviruses. miRNAs are non-immunogenic, stealthy tools for viruses to regulate their as well as host gene expression. The human cytomegalovirus (HCMV) is the major cause of morbidity in immunocompromised patients and allogenic bone-marrow or organ-transplant recipients and the leading cause of congenital birth defects. HCMV miRNAs may provide valuable targets for new urgently needed antiviral drugs. This review focuses on recent findings for viral miRNAs expressed by cytomegaloviruses (CMV) including data from human, chimpanzee, and murine CMV. These are discussed in the context of findings for other viruses to highlight potentially conserved roles exerted by viral miRNAs.}, note = {0920-8569 (Print) Journal Article}, keywords = {PFEFFER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Microfluidic chips for the crystallization of biomacromolecules by counter-diffusion and on-chip crystal X-ray analysis}, author = {K Dhouib and C Khan Malek and W Pfleging and B Gauthier-Manuel and R Duffait and G Thuillier and R Ferrigno and L Jacquamet and J Ohana and J L Ferrer and A Théobald-Dietrich and R Giege and B Lorber and C Sauter}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19417908}, isbn = {19417908}, year = {2009}, date = {2009-01-01}, journal = {Lab Chip}, volume = {9}, number = {10}, pages = {1412-1421}, abstract = {Microfluidic devices were designed to perform on micromoles of biological macromolecules and viruses the search and the optimization of crystallization conditions by counter-diffusion, as well as the on-chip analysis of crystals by X-ray diffraction. Chips composed of microchannels were fabricated in poly-dimethylsiloxane (PDMS), poly-methyl-methacrylate (PMMA) and cyclo-olefin-copolymer (COC) by three distinct methods, namely replica casting, laser ablation and hot embossing. The geometry of the channels was chosen to ensure that crystallization occurs in a convection-free environment. The transparency of the materials is compatible with crystal growth monitoring by optical microscopy. The quality of the protein 3D structures derived from on-chip crystal analysis by X-ray diffraction using a synchrotron radiation was used to identify the most appropriate polymers. Altogether the results demonstrate that for a novel biomolecule, all steps from the initial search of crystallization conditions to X-ray diffraction data collection for 3D structure determination can be performed in a single chip.}, note = {1473-0197 (Print) 1473-0189 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {FRUGIER, SAUTER, Unité ARN, X-Ray/*instrumentation Dimethylpolysiloxanes/chemistry Macromolecular Substances/*chemistry Microfluidic Analytical Techniques/*instrumentation Polymethyl Methacrylate/chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {The dynamic landscapes of RNA architecture}, author = {J A Cruz and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19239882}, doi = {S0092-8674(09)00139-1 [pii] 10.1016/j.cell.2009.02.003}, issn = {1097-4172 (Electronic) 0092-8674 (Linking)}, year = {2009}, date = {2009-01-01}, journal = {Cell}, volume = {136}, number = {4}, pages = {604-609}, abstract = {A wealth of information on RNA folding and ribonucleoprotein assembly has emerged from analyses of structures and from the use of innovative biophysical tools. Although integrating data obtained from static structures with dynamic measurements presents major challenges, such efforts are opening new vistas on the RNA folding landscape.}, note = {Cruz, Jose Almeida Westhof, Eric Research Support, Non-U.S. Gov't United States Cell Cell. 2009 Feb 20;136(4):604-9.}, keywords = {Animals Base Sequence Humans Models, Molecular Molecular Sequence Data Nucleic Acid Conformation RNA/*chemistry Thermodynamics, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Probing mRNA structure and sRNA-mRNA interactions in bacteria using enzymes and lead(II)}, author = {C Chevalier and T Geissmann and A C Helfer and P Romby}, editor = {A Serganov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19381563}, doi = {978-1-59745-558-9_16}, isbn = {19381563}, year = {2009}, date = {2009-01-01}, booktitle = {Riboswitches: Methods and Protocols}, volume = {540}, pages = {215-232}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Enzymatic probing and lead(II)-induced cleavages have been developed to study the secondary structure of RNA molecules either free or engaged in complex with different ligands. Using a combination of probes with different specificities (unpaired vs. paired regions), it is possible to get information on the accessibility of each nucleotide, on the binding site of a ligand (noncoding RNAs, protein, metabolites), and on RNA conformational changes that accompanied ligand binding or environmental conditions (temperature, pH, ions, etc.). The detection of the cleavages can be conducted by two different ways, which are chosen according to the length of the studied RNA. The first method uses end-labeled RNA molecules and the second one involves primer extension by reverse transcriptase. We provide here an experimental procedure that was designed to map the structure of mRNA and mRNA-sRNA interaction in vitro.}, keywords = {Bacterial/chemical synthesis RNA, Base Sequence Chemical Fractionation Enzymes/*metabolism Hydrolysis/drug effects Lead/*pharmacology Molecular Biology/*methods Molecular Sequence Data Nucleic Acid Conformation/drug effects RNA/metabolism RNA, Messenger/*chemistry/genetics/*metabolism RNA, ROMBY, Unité ARN, Untranslated/chemistry/genetics/*metabolism Staphylococcus aureus/drug effects/*metabolism}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {SECISaln, a web-based tool for the creation of structure-based alignments of eukaryotic SECIS elements}, author = {C E Chapple and R Guigo and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19179357}, isbn = {19179357}, year = {2009}, date = {2009-01-01}, journal = {Bioinformatics}, volume = {25}, number = {5}, pages = {674-675}, abstract = {SUMMARY: Selenoproteins contain the 21st amino acid selenocysteine which is encoded by an inframe UGA codon, usually read as a stop. In eukaryotes, its co-translational recoding requires the presence of an RNA stem-loop structure, the SECIS element in the 3 untranslated region of (UTR) selenoprotein mRNAs. Despite little sequence conservation, SECIS elements share the same overall secondary structure. Until recently, the lack of a significantly high number of selenoprotein mRNA sequences hampered the identification of other potential sequence conservation. In this work, the web-based tool SECISaln provides for the first time an extensive structure-based sequence alignment of SECIS elements resulting from the well-defined secondary structure of the SECIS RNA and the increased size of the eukaryotic selenoproteome. We have used SECISaln to improve our knowledge of SECIS secondary structure and to discover novel, conserved nucleotide positions and we believe it will be a useful tool for the selenoprotein and RNA scientific communities. AVAILABILITY: SECISaln is freely available as a web-based tool at http://genome.crg.es/software/secisaln/.}, note = {1460-2059 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {KROL, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selenoprotein N is dynamically expressed during mouse development and detected early in muscle precursors}, author = {P Castets and S Maugenre and C Gartioux and M Rederstorff and A Krol and A Lescure and S Tajbakhsh and V Allamand and P Guicheney}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19698141}, isbn = {19698141}, year = {2009}, date = {2009-01-01}, journal = {BMC Dev Biol}, volume = {9}, pages = {46}, abstract = {BACKGROUND: In humans, mutations in the SEPN1 gene, encoding selenoprotein N (SelN), are involved in early onset recessive neuromuscular disorders, referred to as SEPN1-related-myopathies. The mechanisms behind these pathologies are poorly understood since the function of SelN remains elusive. However, previous results obtained in humans and more recently in zebrafish pointed to a potential role for SelN during embryogenesis. Using qRT-PCR, Western blot and whole mount in situ hybridization, we characterized in detail the spatio-temporal expression pattern of the murine Sepn1 gene during development, focusing particularly on skeletal muscles. RESULTS: In whole embryos, Sepn1 transcripts were detected as early as E5.5, with expression levels peaking at E12.5, and then strongly decreasing until birth. In isolated tissues, only mild transcriptional variations were observed during development, whereas a striking reduction of the protein expression was detected during the perinatal period. Furthermore, we demonstrated that Sepn1 is expressed early in somites and restricted to the myotome, the sub-ectodermal mesenchyme and the dorsal root ganglia at mid-gestation stages. Interestingly, Sepn1 deficiency did not alter somitogenesis in embryos, suggesting that SelN is dispensable for these processes in mouse. CONCLUSION: We characterized for the first time the expression pattern of Sepn1 during mammalian embryogenesis and we demonstrated that its differential expression is most likely dependent on major post-transcriptional regulations. Overall, our data strongly suggest a potential role for selenoprotein N from mid-gestation stages to the perinatal period. Interestingly, its specific expression pattern could be related to the current hypothesis that selenoprotein N may regulate the activity of the ryanodine receptors.}, note = {1471-213X (Electronic) 1471-213X (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Developmental Humans Mice Muscle Proteins/genetics/*metabolism Muscle, KROL Animals Embryo, LESCURE, Mammalian/metabolism *Gene Expression Regulation, Skeletal/*embryology Myoblasts/metabolism Ryanodine Receptor Calcium Release Channel/metabolism Selenoproteins/genetics/*metabolism Zebrafish/embryology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Kinetics of deoxy-CTP incorporation opposite a dG-C8-N-2-aminofluorene adduct by a high-fidelity DNA polymerase}, author = {D Y Burnouf and J E Wagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19150355}, isbn = {19150355}, year = {2009}, date = {2009-01-01}, journal = {J Mol Biol}, volume = {386}, number = {4}, pages = {951-961}, abstract = {The model carcinogen N-2-acetylaminofluorene covalently binds to the C8 position of guanine to form two adducts, the N-(2'-deoxyguanosine-8-yl)-aminofluorene (G-AF) and the N-2-(2'-deoxyguanosine-8-yl)-acetylaminofluorene (G-AAF). Although they are chemically closely related, their biological effects are strongly different and they are processed by different damage tolerance pathways. G-AF is bypassed by replicative and high-fidelity polymerases, while specialized polymerases ensure synthesis past of G-AAF. We used the DNA polymerase I fragment of a Bacillus stearothermophilus strain as a model for a high-fidelity polymerase to study the kinetics of incorporation of deoxy-CTP (dCTP) opposite a single G-AF. Pre-steady-state kinetic experiments revealed a drastic reduction in dCTP incorporation performed by the G-AF-modified ternary complex. Two populations of these ternary complexes were identified: (i) a minor productive fraction (20%) that readily incorporates dCTP opposite the G-AF adduct with a rate similar to that measured for the adduct-free ternary complexes and (ii) a major fraction of unproductive complexes (80%) that slowly evolve into productive ones. In the light of structural data, we suggest that this slow rate reflects the translocation of the modified base within the active site, from the pre-insertion site into the insertion site. By making this translocation rate limiting, the G-AF lesion reveals a novel kinetic step occurring after dNTP binding and before chemistry.}, note = {1089-8638 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cluster analysis for phasing with molecular replacement: a feasibility study}, author = {A Buehler and L Urzhumtseva and V Y Lunin and A Urzhumtsev}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19564684}, isbn = {19564684}, year = {2009}, date = {2009-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {65}, number = {Pt 7}, pages = {644-650}, abstract = {Molecular replacement can fail to find a solution, namely a unique orientation and position of a search model, even when many search models are tested under various conditions. Simultaneous use of the results of these searches may help in the solution of such difficult structures. A closeness between the peaks of several calculated rotation functions may identify the model orientation. The largest and most compact cluster of such peaks usually corresponds to models which are oriented similarly to the molecule under study. A search for the optimal translation may be more problematic and both individual translation functions and straightforward cluster analysis in the space of geometric parameters such as rotation angles and translation vectors may give no result. An improvement may be obtained by performing cluster analysis of the peaks of several translation functions in phase-set space. In this case, the Fourier maps computed using the observed structure-factor magnitudes and the phases calculated from differently positioned models are compared. Again, as a rule, the largest and the most compact cluster corresponds to the correct solution. The result of the updated procedure is no longer a single search model but an averaged Fourier map.}, note = {1399-0047 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The RNA structure alignment ontology}, author = {J W Brown and A Birmingham and P E Griffiths and F Jossinet and R Kachouri-Lafond and R Knight and B F Lang and N Leontis and G Steger and J Stombaugh and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19622678}, doi = {rna.1601409 [pii] 10.1261/rna.1601409}, issn = {1469-9001 (Electronic) 1355-8382 (Linking)}, year = {2009}, date = {2009-01-01}, journal = {RNA}, volume = {15}, number = {9}, pages = {1623-1631}, abstract = {Multiple sequence alignments are powerful tools for understanding the structures, functions, and evolutionary histories of linear biological macromolecules (DNA, RNA, and proteins), and for finding homologs in sequence databases. We address several ontological issues related to RNA sequence alignments that are informed by structure. Multiple sequence alignments are usually shown as two-dimensional (2D) matrices, with rows representing individual sequences, and columns identifying nucleotides from different sequences that correspond structurally, functionally, and/or evolutionarily. However, the requirement that sequences and structures correspond nucleotide-by-nucleotide is unrealistic and hinders representation of important biological relationships. High-throughput sequencing efforts are also rapidly making 2D alignments unmanageable because of vertical and horizontal expansion as more sequences are added. Solving the shortcomings of traditional RNA sequence alignments requires explicit annotation of the meaning of each relationship within the alignment. We introduce the notion of "correspondence," which is an equivalence relation between RNA elements in sets of sequences as the basis of an RNA alignment ontology. The purpose of this ontology is twofold: first, to enable the development of new representations of RNA data and of software tools that resolve the expansion problems with current RNA sequence alignments, and second, to facilitate the integration of sequence data with secondary and three-dimensional structural information, as well as other experimental information, to create simultaneously more accurate and more exploitable RNA alignments.}, note = {Brown, James W Birmingham, Amanda Griffiths, Paul E Jossinet, Fabrice Kachouri-Lafond, Rym Knight, Rob Lang, B Franz Leontis, Neocles Steger, Gerhard Stombaugh, Jesse Westhof, Eric Letter Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. United States RNA (New York, N.Y.) RNA. 2009 Sep;15(9):1623-31. Epub 2009 Jul 21.}, keywords = {Animals Base Sequence Humans Models, Biological Molecular Sequence Data Nucleic Acid Conformation Phylogeny RNA/*analysis/chemistry Sequence Alignment/*methods/trends Sequence Analysis, Nucleic Acid *Software, RNA/methods Sequence Homology, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Plasmodial aspartyl-tRNA synthetases and peculiarities in Plasmodium falciparum}, author = {T Bour and A Akaddar and B Lorber and S Blais and C Balg and E Candolfi and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19443655}, isbn = {19443655}, year = {2009}, date = {2009-01-01}, journal = {J Biol Chem}, volume = {284}, number = {28}, pages = {18893-18903}, abstract = {Distinctive features of aspartyl-transfer RNA (tRNA) synthetases (AspRS) from the protozoan Plasmodium genus are described. These apicomplexan AspRSs contain 29-31 amino acid insertions in their anticodon binding domains, a remarkably long N-terminal appendix that varies in size from 110 to 165 amino acids and two potential initiation codons. This article focuses on the atypical functional and structural properties of Plasmodium falciparum cytosolic AspRS, the causative parasite of human malaria. This species encodes a 626 or 577 amino acids AspRS depending on whether initiation starts on the first or second in-frame initiation codon. The longer protein has poor solubility and a propensity to aggregate. Production of the short version was favored as shown by the comparison of the recombinant protein with endogenous AspRS. Comparison of the tRNA aminoacylation activity of wild-type and mutant parasite AspRSs with those of yeast and human AspRSs revealed unique properties. The N-terminal extension contains a motif that provides unexpectedly strong RNA binding to plasmodial AspRS. Furthermore, experiments demonstrated the requirement of the plasmodial insertion for AspRS dimerization and, therefore, tRNA aminoacylation and other putative functions. Implications for the parasite biology are proposed. These data provide a robust background for unraveling the precise functional properties of the parasite AspRS and for developing novel lead compounds against malaria, targeting its idiosyncratic domains.}, note = {0021-9258 (Print) 0021-9258 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Amino Acid, Amino Acid Sequence Amino Acids/chemistry Animals Aspartate-tRNA Ligase/*metabolism Aspartic Acid/chemistry Base Sequence Cloning, FRUGIER, Molecular Cytoplasm/metabolism Dimerization Fungal Proteins/chemistry Humans Kinetics Molecular Sequence Data Plasmodium falciparum Protein Structure, Tertiary Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Function and ribosomal localization of aIF6, a translational regulator shared by archaea and eukarya}, author = {D Benelli and S Marzi and C Mancone and T Alonzi and A la Teana and P Londei}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19036786}, isbn = {19036786}, year = {2009}, date = {2009-01-01}, journal = {Nucleic Acids Res}, volume = {37}, number = {1}, pages = {256-267}, abstract = {The translation factor IF6 is shared by the Archaea and the Eukarya, but is not found in Bacteria. The properties of eukaryal IF6 (eIF6) have been extensively studied, but remain somewhat elusive. eIF6 behaves as a ribosome-anti-association factor and is involved in miRNA-mediated gene silencing; however, it also seems to participate in ribosome synthesis and export. Here we have determined the function and ribosomal localization of the archaeal (Sulfolobus solfataricus) IF6 homologue (aIF6). We find that aIF6 binds specifically to the 50S ribosomal subunits, hindering the formation of 70S ribosomes and strongly inhibiting translation. aIF6 is uniformly expressed along the cell cycle, but it is upregulated following both cold- and heat shock. The aIF6 ribosomal binding site lies in the middle of the 30-S interacting surface of the 50S subunit, including a number of critical RNA and protein determinants involved in subunit association. The data suggest that the IF6 protein evolved in the archaeal-eukaryal lineage to modulate translational efficiency under unfavourable environmental conditions, perhaps acquiring additional functions during eukaryotic evolution.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {23S/chemistry/metabolism Ribosomal Proteins/metabolism Ribosome Subunits, Archaeal Proteins/analysis/chemistry/*metabolism Base Sequence Binding Sites Cell Cycle Cloning, Archaeal/*metabolism Ribosomes/metabolism Sulfolobus solfataricus/*genetics/metabolism, Large, Molecular Eukaryotic Initiation Factors/chemistry Models, Molecular Molecular Sequence Data Prokaryotic Initiation Factors/analysis/chemistry/*metabolism *Protein Biosynthesis RNA, Ribosomal, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of monomeric yeast and mammalian Sec61 complexes interacting with the translating ribosome}, author = {T Becker and S Bhushan and A Jarasch and J P Armache and S Funes and F Jossinet and J Gumbart and T Mielke and O Berninghausen and K Schulten and E Westhof and R Gilmore and E C Mandon and R Beckmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19933108}, isbn = {19933108}, year = {2009}, date = {2009-01-01}, journal = {Science}, volume = {326}, number = {5958}, pages = {1369-1373}, abstract = {The trimeric Sec61/SecY complex is a protein-conducting channel (PCC) for secretory and membrane proteins. Although Sec complexes can form oligomers, it has been suggested that a single copy may serve as an active PCC. We determined subnanometer-resolution cryo-electron microscopy structures of eukaryotic ribosome-Sec61 complexes. In combination with biochemical data, we found that in both idle and active states, the Sec complex is not oligomeric and interacts mainly via two cytoplasmic loops with the universal ribosomal adaptor site. In the active state, the ribosomal tunnel and a central pore of the monomeric PCC were occupied by the nascent chain, contacting loop 6 of the Sec complex. This provides a structural basis for the activity of a solitary Sec complex in cotranslational protein translocation.}, note = {1095-9203 (Electronic) 0036-8075 (Linking) Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.}, keywords = {Animals Binding Sites Cryoelectron Microscopy Dogs Image Processing, Computer-Assisted Membrane Proteins/*chemistry/*metabolism/ultrastructure Models, Molecular *Protein Biosynthesis Protein Conformation Protein Multimerization Protein Structure, Secondary *Protein Transport Proteins/chemistry/*metabolism/ultrastructure Ribosomes/*metabolism/ultrastructure Saccharomyces cerevisiae Proteins/*chemistry/*metabolism/ultrastructure, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Isolation, crystallization and preliminary X-ray analysis of the transamidosome, a ribonucleoprotein involved in asparagine formation}, author = {M Bailly and M Blaise and B Lorber and S Thirup and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19478435}, isbn = {19478435}, year = {2009}, date = {2009-01-01}, journal = {Acta Crystallogr F Struct Biol Commun}, volume = {65}, number = {Pt 6}, pages = {577-581}, abstract = {Thermus thermophilus deprived of asparagine synthetase synthesizes Asn on tRNA(Asn) via a tRNA-dependent pathway involving a nondiscriminating aspartyl-tRNA synthetase that charges Asp onto tRNA(Asn) prior to conversion of the Asp to Asn by GatCAB, a tRNA-dependent amidotransferase. This pathway also constitutes the route of Asn-tRNA(Asn) formation by bacteria and archaea deprived of asparaginyl-tRNA synthetase. The partners involved in tRNA-dependent Asn formation in T. thermophilus assemble into a ternary complex called the transamidosome. This particule produces Asn-tRNA(Asn) in the presence of free Asp, ATP and an amido-group donor. Crystals of the transamidosome from T. thermophilus were obtained in the presence of PEG 4000 in MES-NaOH buffer pH 6.5. They belonged to the primitive monoclinic space group P2(1), with unit-cell parameters a = 115.9}, note = {1744-3091 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Amino Acyl/genetics/metabolism RNA, Asn/*biosynthesis Ribonucleoproteins/*isolation & purification/*metabolism Scattering, KERN FLORENTZ Asparagine/*biosynthesis Aspartate-tRNA Ligase/*chemistry/genetics/*metabolism Crystallization Data Collection Escherichia coli/genetics Light RNA, Radiation Statistics as Topic Thermus thermophilus/genetics/metabolism Transfer RNA Aminoacylation X-Ray Diffraction, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Bruton's tyrosine kinase is involved in miR-346-related regulation of IL-18 release by lipopolysaccharide-activated rheumatoid fibroblast-like synoviocytes}, author = {G Alsaleh and G Suffert and N Semaan and T Juncker and L Frenzel and J E Gottenberg and J Sibilia and S Pfeffer and D Wachsmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19342689}, isbn = {19342689}, year = {2009}, date = {2009-01-01}, journal = {J Immunol}, volume = {182}, number = {8}, pages = {5088-5097}, abstract = {MicroRNAs (miRNAs) have emerged as key players in the regulation of expression of target mRNAs expression. They have been associated with diverse biological processes, and recent studies have demonstrated that miRNAs play a role in inflammatory responses. We reported previously that LPS-activated fibroblast-like synoviocytes (FLS) isolated from rheumatoid arthritis (RA) patients express IL-18 mRNA but they do not release IL-18. Based on the observation that this inhibition was due to a rapid degradation of IL-18 mRNA, our group has conducted a study to identify miRNAs that could play a role in the "antiinflammatory" response of LPS-activated RA FLS. LPS challenge modulated the expression of 63 miRNAs as assessed by microarray analysis. Fifteen miRNAs were up-regulated, including miR-346, for which overexpression upon LPS treatment was validated by quantitative RT-PCR. We then transfected FLS with an antisense oligonucleotide targeting miR-346 and found that, in these conditions, IL-18 release could be measured upon LPS activation of FLS. Moreover, we also demonstrated that miR-346 indirectly regulated IL-18 release by indirectly inhibiting LPS-induced Bruton's tyrosine kinase expression in LPS-activated RA FLS. These findings suggest that miRNAs function as regulators that help to fine-tune the inflammatory response in RA.}, note = {1550-6606 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {arthritis, Cultured Gene Expression Regulation/*genetics Humans Interleukin-18/biosynthesis/genetics/*secretion Lipopolysaccharides/*pharmacology MicroRNAs/*genetics Oligonucleotide Array Sequence Analysis Protein-Tyrosine Kinases/genetics/*metabolism RNA Interference RNA, Messenger/genetics Synovial Membrane/drug effects/*metabolism/secretion, PFEFFER, Rheumatoid/chemically induced/genetics/*metabolism Base Sequence Cell Line Cells, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The selenium to selenoprotein pathway in eukaryotes: more molecular partners than anticipated}, author = {C Allmang and L Wurth and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19285539}, isbn = {19285539}, year = {2009}, date = {2009-01-01}, journal = {Biochim Biophys Acta-Gen Subj}, volume = {1790}, number = {11}, pages = {1415-1423}, abstract = {The amino acid selenocysteine (Sec) is the major biological form of the trace element selenium. Sec is co-translationally incorporated in selenoproteins. There are 25 selenoprotein genes in humans, and Sec was found in the active site of those that have been attributed a function. This review will discuss how selenocysteine is synthesized and incorporated into selenoproteins in eukaryotes. Sec biosynthesis from serine on the tRNA(Sec) requires four enzymes. Incorporation of Sec in response to an in-frame UGA codon, otherwise signaling termination of translation, is achieved by a complex recoding machinery to inform the ribosomes not to stop at this position on the mRNA. A number of the molecular partners acting in this machinery have been identified but their detailed mechanism of action has not been deciphered yet. Here we provide an overview of the literature in the field. Particularly striking is the higher than originally envisaged number of factors necessary to synthesize Sec and selenoproteins. Clearly, selenoprotein synthesis is an exciting and very active field of research.}, note = {0006-3002 (Print) 0006-3002 (Linking) Journal Article Review}, keywords = {Animals Base Sequence Eukaryota/genetics/*metabolism Humans Metabolic Networks and Pathways/genetics/physiology Models, Biological Selenium/*metabolism Selenocysteine/biosynthesis Selenoproteins/biosynthesis/genetics/*metabolism, ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{guichard_rationally-designed_2009, title = {Rationally-designed multivalent architectures for mimicking homotrimers of CD40L, a member of the TNF superfamily}, author = {Gilles Guichard and Nathalie Trouche and Sébastien Wieckowski and Weimin Sun and Olivier Chaloin and Alberto Bianco and Johan Hoebeke and Pascal Schneider and Sylvie Fournel}, doi = {10.1007/978-0-387-73657-0_157}, issn = {0065-2598}, year = {2009}, date = {2009-01-01}, journal = {Advances in Experimental Medicine and Biology}, volume = {611}, pages = {355--357}, keywords = {Biopolymers, CD40 Ligand, I2CT, Models, Molecular, Molecular Mimicry, Team-Bianco, X-Ray Diffraction}, pubstate = {published}, tppubtype = {article} } @article{geotti-bianchini_conformationally_2009, title = {Conformationally controlled, thymine-based alpha-nucleopeptides}, author = {Piero Geotti-Bianchini and Marco Crisma and Cristina Peggion and Alberto Bianco and Fernando Formaggio}, doi = {10.1039/b822789f}, issn = {1359-7345}, year = {2009}, date = {2009-01-01}, journal = {Chemical Communications (Cambridge, England)}, number = {22}, pages = {3178--3180}, abstract = {Rigid peptide backbones and backbone-to-side chain H-bonds permit the design of alpha-nucleopeptides with known 3D-structure; thymine-thymine base pairing is also observed.}, keywords = {I2CT, Magnetic Resonance Spectroscopy, Oligopeptides, Protein Conformation, Team-Bianco, thymine, X-Ray Diffraction}, pubstate = {published}, tppubtype = {article} } @article{geotti-bianchini_synthesis_2009, title = {Synthesis and 3D-structure of conformationally controlled nucleo-peptides}, author = {Piero Geotti-Bianchini and Marco Crisma and Cristina Peggion and Alberto Bianco and Fernando Formaggio}, doi = {10.1007/978-0-387-73657-0_16}, issn = {0065-2598}, year = {2009}, date = {2009-01-01}, journal = {Advances in Experimental Medicine and Biology}, volume = {611}, pages = {37--38}, keywords = {I2CT, Models, Molecular, Nucleoproteins, Peptides, Protein Conformation, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure-function insights into prokaryotic and eukaryotic translation initiation}, author = {A G Myasnikov and A Simonetti and S Marzi and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19493673}, isbn = {19493673}, year = {2009}, date = {2009-01-01}, journal = {Curr Opin Struct Biol}, volume = {19}, number = {3}, pages = {300-9}, abstract = {Translation initiation is the rate-limiting and most complexly regulated step of protein synthesis in prokaryotes and eukaryotes. In the last few years, cryo-electron microscopy has provided several novel insights into the universal process of translation initiation. Structures of prokaryotic 30S and 70S ribosomal initiation complexes with initiator transfer RNA (tRNA), messenger RNA (mRNA), and initiation factors have recently revealed the mechanism of initiator tRNA recruitment to the assembling ribosomal machinery, involving molecular rearrangements of the ribosome and associated factors. First three-dimensional pictures of the particularly complex eukaryotic translation initiation machinery have been obtained, revealing how initiation factors tune the ribosome for recruiting the mRNA. A comparison of the available prokaryotic and eukaryotic structures shows that--besides significant differences--some key ribosomal features are universally conserved.}, note = {1879-033X (Electronic) 0959-440X (Linking) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Amino Acid Sequence Eukaryotic Cells/*chemistry/*metabolism Humans Molecular Sequence Data *Peptide Chain Initiation, mino Acid Sequence Eukaryotic Cells/*chemistry/*metabolism Humans Molecular Sequence Data *Peptide Chain Initiation, ROMBY, Transfer/chemistry/metabolism Ribosomes/chemistry/genetics/metabolism, Translational Peptide Initiation Factors/chemistry/metabolism Prokaryotic Cells/*chemistry/*metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{DW2008, title = {Molecular and cellular components of the mating machinery in Anopheles gambiae females}, author = {D W Rogers and Miranda M Whitten and Janis Thailayil and Julien Soichot and Elena A Levashina and Flaminia Catteruccia}, url = {http://www.ncbi.nlm.nih.gov/pubmed/19036921}, year = {2008}, date = {2008-12-09}, journal = {Proc Natl Acad Sci U S A.}, volume = {105}, number = {49}, pages = {19390-5}, abstract = {Anopheles gambiae mosquitoes are the principal vectors of malaria. A major determinant of the capacity of these mosquitoes as disease vectors is their high reproductive rate. Reproduction depends on a single insemination, which profoundly changes the behavior and physiology of females. To identify factors and mechanisms relevant to the fertility of A. gambiae, we performed a comprehensive analysis of the molecular and cellular machinery associated with copulation in females. Initial whole-body microarray experiments comparing virgins with females at 2 h, 6 h, and 24 h after mating detected large transcriptional changes. Analysis of tissue localization identified a subset of genes whose expression was strikingly regulated by mating in the lower reproductive tract and, surprisingly, the gut. In the atrium of virgin females, where the male seminal fluid is received, our studies revealed a "mating machinery" consisting of molecular and structural components that are turned off or collapse after copulation, suggesting that this tissue loses its competence for further insemination. In the sperm storage organ, we detected a number of mating-responsive genes likely to have a role in the maintenance and function of stored sperm. These results identify genes and mechanisms regulating the reproductive biology of A. gambiae females, highlighting considerable differences with Drosophila melanogaster. Our data inform vector control strategies and reveal promising targets for the manipulation of fertility in field populations of these important disease vectors.}, keywords = {reproduction}, pubstate = {published}, tppubtype = {article} } @article{deddouche_dexd/h-box_2008, title = {The DExD/Ħ-box helicase Dicer-2 mediates the induction of antiviral activity in drosophila}, author = {Safia Deddouche and Nicolas Matt and Aidan Budd and Stefanie Mueller and Cordula Kemp and Delphine Galiana-Arnoux and Catherine Dostert and Christophe Antoniewski and Jules A Hoffmann and Jean-Luc Imler}, doi = {10.1038/ni.1664}, issn = {1529-2916}, year = {2008}, date = {2008-12-01}, journal = {Nature Immunology}, volume = {9}, number = {12}, pages = {1425--1432}, abstract = {Drosophila, like other invertebrates and plants, relies mainly on RNA interference for its defense against viruses. In flies, viral infection also triggers the expression of many genes. One of the genes induced, Vago, encodes a 18-kilodalton cysteine-rich polypeptide. Here we provide genetic evidence that the Vago gene product controlled viral load in the fat body after infection with drosophila C virus. Induction of Vago was dependent on the helicase Dicer-2. Dicer-2 belongs to the same DExD/H-box helicase family as do the RIG-I-like receptors, which sense viral infection and mediate interferon induction in mammals. We propose that this family represents an evolutionary conserved set of sensors that detect viral nucleic acids and direct antiviral responses.}, keywords = {Amino Acid, Animals, Electrophoresis, Fat Body, Gene Expression Regulation, Genetic, Genetically Modified, hoffmann, Humans, imler, M3i, matt, Phylogeny, Polyacrylamide Gel, Reverse Transcriptase Polymerase Chain Reaction, Ribonuclease III, RNA Helicases, Sequence Homology, Transcription, Virus Diseases}, pubstate = {published}, tppubtype = {article} } @article{yandar_immunological_2008, title = {Immunological profile of a Plasmodium vivax AMA-1 N-terminus peptide-carbon nanotube conjugate in an infected Plasmodium berghei mouse model}, author = {Nubia Yandar and Giorgia Pastorin and Maurizio Prato and Alberto Bianco and Manuel Elkin Patarroyo and José Manuel Lozano}, url = {http://www.sciencedirect.com/science/article/pii/S0264410X08010736}, doi = {10.1016/j.vaccine.2008.08.014}, issn = {0264-410X}, year = {2008}, date = {2008-10-01}, urldate = {2020-03-31}, journal = {Vaccine}, volume = {26}, number = {46}, pages = {5864--5873}, abstract = {We have covalently conjugated an N-terminus Plasmodium vivax apical membrane antigen-1 (AMA-1) peptide to functionalized carbon nanotubes (f-CNT). Immunological characterization of this molecular conjugate revealed that the immunogen-AMA-1 peptide was appropriately presented after being conjugated to CNTs as well as being recognized by BALB/c polyclonal antibodies. Subsequent experiments lead us to assess the AMA-1 peptide alone, as well as the CNT-peptide conjugate regarding rodent malarial infection. Remarkably, the peptide effectively controlled and delayed Plasmodium berghei-challenged animals’ parasitaemia. The peptide-CNT conjugate displayed similar immunological properties to the peptide alone by protecting or delaying malarial infection. The peptide presentation by f-CNT to the immune system thus constitutes a promising approach for synthetic malarial vaccine formulation since the immunogen peptide conformation is well preserved.}, keywords = {Apical membrane antigen 1 (AMA-1), Functionalized carbon nanotubes (-CNT), I2CT, Synthetic vaccine delivery system, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{el_chamy_sensing_2008, title = {Sensing of 'danger signals' and pathogen-associated molecular patterns defines binary signaling pathways 'upstream' of Toll}, author = {L El Chamy and V Leclerc and I Caldelari and J-M Reichhart}, doi = {10.1038/ni.1643}, issn = {1529-2916}, year = {2008}, date = {2008-10-01}, journal = {Nat. Immunol.}, volume = {9}, number = {10}, pages = {1165--1170}, abstract = {In drosophila, molecular determinants from fungi and Gram-positive bacteria are detected by circulating pattern-recognition receptors. Published findings suggest that such pattern-recognition receptors activate as-yet-unidentified serine-protease cascades that culminate in the cleavage of Spätzle, the endogenous Toll receptor ligand, and trigger the immune response. We demonstrate here that the protease Grass defines a common activation cascade for the detection of fungi and Gram-positive bacteria mediated by pattern-recognition receptors. The serine protease Persephone, shown before to be specific for fungal detection in a cascade activated by secreted fungal proteases, was also required for the sensing of proteases elicited by bacteria in the hemolymph. Hence, Persephone defines a parallel proteolytic cascade activated by 'danger signals' such as abnormal proteolytic activities.}, keywords = {Animals, Fungi, Genetically Modified, Gram-Positive Bacteria, Gram-Positive Bacterial Infections, In Situ Hybridization, M3i, Mycoses, Pattern Recognition, Peptide Hydrolases, Receptors, reichhart, ROMBY, Serine Endopeptidases, Signal Transduction, Toll-Like Receptors, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{kwan_dermal-type_2008b, title = {Dermal-type macrophages expressing CD209/DC-SIGN show inherent resistance to dengue virus growth}, author = {Wing-Hong Kwan and Erika Navarro-Sanchez and Hélène Dumortier and Marion Decossas and Hortense Vachon and Flavia Barreto dos Santos and Hervé W Fridman and Félix A Rey and Eva Harris and Philippe Despres and Christopher G Mueller}, doi = {10.1371/journal.pntd.0000311}, issn = {1935-2735}, year = {2008}, date = {2008-10-01}, journal = {PLoS neglected tropical diseases}, volume = {2}, number = {10}, pages = {e311}, abstract = {BACKGROUND: An important question in dengue pathogenesis is the identity of immune cells involved in the control of dengue virus infection at the site of the mosquito bite. There is evidence that infection of immature myeloid dendritic cells plays a crucial role in dengue pathogenesis and that the interaction of the viral envelope E glycoprotein with CD209/DC-SIGN is a key element for their productive infection. Dermal macrophages express CD209, yet little is known about their role in dengue virus infection. METHODS AND FINDINGS: Here, we showed that dermal macrophages bound recombinant envelope E glycoprotein fused to green fluorescent protein. Because dermal macrophages stain for IL-10 in situ, we generated dermal-type macrophages from monocytes in the presence of IL-10 to study their infection by dengue virus. The macrophages were able to internalize the virus, but progeny virus production was undetectable in the infected cells. In addition, no IFN-alpha was produced in response to the virus. The inability of dengue virus to grow in the macrophages was attributable to accumulation of internalized virus particles into poorly-acidified phagosomes. CONCLUSIONS: Aborting infection by viral sequestration in early phagosomes would present a novel means to curb infection of enveloped virus and may constitute a prime defense system to prevent dengue virus spread shortly after the bite of the infected mosquito.}, keywords = {Adhesion, adhesion molecules, C-Type, Cell Adhesion, Cell Adhesion Molecules, Cell Line, Cell Surface, Cells, Chemistry, Cultured, Dendritic Cells, Dengue, Dengue virus, Gene Expression, Genetics, GLYCOPROTEIN, Growth, growth & development, Humans, ICAM-3, IFN ALPHA, IL-10, IL10, IMMATURE, Immunology, in situ, infection, LECTIN, Lectins, Macrophage, Macrophages, metabolism, METHOD, methods, monocyte, Monocytes, myeloid dendritic cells, pathogenesis, Phagosomes, PRODUCTION, Protein, Protein Binding, Proteins, Receptor, Receptors, Resistance, Skin, Team-Mueller, Viral Envelope Proteins, virology, virus}, pubstate = {published}, tppubtype = {article} } @article{geotti-bianchini_design_2008, title = {Design and synthesis of intrinsically cell-penetrating nucleopeptides}, author = {Piero Geotti-Bianchini and Julien Beyrath and Olivier Chaloin and Fernando Formaggio and Alberto Bianco}, doi = {10.1039/b811639c}, issn = {1477-0539}, year = {2008}, date = {2008-10-01}, journal = {Organic & Biomolecular Chemistry}, volume = {6}, number = {20}, pages = {3661--3663}, abstract = {Nucleopeptides, which are constituted of alpha-amino acids bearing nucleobases at their side chains, are able to penetrate into cells and to reach the nucleus without cytotoxic effects.}, keywords = {Amino Acid Sequence, Cell Line, Cells, Drug Design, Humans, I2CT, Molecular Sequence Data, Peptides, Purines, Pyrimidines, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bringel_lactobacillus_2008, title = {Lactobacillus plantarum response to inorganic carbon concentrations: PyrR2-dependent and -independent transcription regulation of genes involved in arginine and nucleotide metabolism.}, author = {Françoise Bringel and Philippe Hammann and Valérie Kugler and Florence Arsène-Ploetze}, doi = {10.1099/mic.0.2008/018184-0}, issn = {1350-0872 1350-0872}, year = {2008}, date = {2008-09-01}, journal = {Microbiology (Reading, England)}, volume = {154}, number = {Pt 9}, pages = {2629--2640}, abstract = {Lactobacillus plantarum susbp. plantarum is a capnophilic Gram-positive heterotroph with optimal growth in 4 % CO(2)-enriched air. At low inorganic carbon (C(i)) concentrations, the pyr genes encoding the enzymes of the pyrimidine biosynthetic pathway were overexpressed, in agreement with a previous study showing that these genes are regulated at the transcription level in response to C(i) via a PyrR(2)-mediated mechanism. A previous study of high-CO(2)-requiring (HCR) mutants revealed an unknown genetic link between arginine regulation and C(i)-dependent nutritional needs. To better understand L. plantarum's adaptation to C(i) availability, additional C(i)-responsive genes were sought in the arginine biosynthetic pathway (arg and car genes) using slot-blot hybridization and a proteomic differential 2D gel electrophoresis (DIGE) global approach. Besides the nine pyr-encoded proteins, 16 new Icr (inorganic-carbon-regulated) proteins accumulated differentially in response to C(i) availability, suggesting that the C(i) response involves several metabolic pathways and adaptation processes. Among these Icr proteins only argininosuccinate lyase, encoded by argH, was involved in arginine biosynthesis. Three proteins involved in the purine biosynthetic pathway and nucleotide conversion, adenylate kinase (Adk), GMP synthase (GuaA), and IMP dehydrogenase (GuaB), accumulated differentially in response to changes in C(i) levels. Expression of the Icr protein-encoding genes argH and guaB was regulated at the transcription level or by RNA stability in response to C(i) availability, as previously demonstrated for the pyr genes. However, PyrR(2) was not essential for the C(i)-regulated transcription of argH and guaB, demonstrating that PyrR(2) modulates only a subset of C(i)-regulated genes. These results suggest that the C(i) response may involve at least two regulatory mechanisms in L. plantarum.}, note = {Place: England}, keywords = {Arginine/*biosynthesis, Argininosuccinate Lyase/genetics, Bacterial, Bacterial Proteins/*genetics, Bacterial/genetics, Carbon Compounds, Carbon Dioxide/metabolism, Electrophoresis, Gel, Gene Expression Regulation, Genetic, IMP Dehydrogenase/genetics, Inorganic/*metabolism, Lactobacillus plantarum/enzymology/genetics/*metabolism, Mass, Matrix-Assisted Laser Desorption-Ionization, Nucleotides/*biosynthesis, Pentosyltransferases/*genetics, PPSE, proteomics, Repressor Proteins/*genetics, Reverse Transcriptase Polymerase Chain Reaction, RNA, Spectrometry, Transcription, Two-Dimensional}, pubstate = {published}, tppubtype = {article} } @article{lacerda_carbon-nanotube_2008, title = {Carbon-nanotube shape and individualization critical for renal excretion}, author = {Lara Lacerda and Maria A Herrero and Kerrie Venner and Alberto Bianco and Maurizio Prato and Kostas Kostarelos}, doi = {10.1002/smll.200800323}, issn = {1613-6829}, year = {2008}, date = {2008-08-01}, journal = {Small (Weinheim an Der Bergstrasse, Germany)}, volume = {4}, number = {8}, pages = {1130--1132}, keywords = {Animals, Biological Transport, carbon, Electron, Female, Glomerular Filtration Rate, I2CT, Inbred BALB C, Kidney Glomerulus, Mice, Microscopy, Nanoparticles, nanotechnology, Nanotubes, Team-Bianco, Transmission}, pubstate = {published}, tppubtype = {article} } @article{EA2008, title = {Host-parasite interactions: the balance of trade}, author = {Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/18620075}, year = {2008}, date = {2008-07-24}, journal = {Curr Opin Microbiol.}, volume = {11}, number = {4}, pages = {338-9}, keywords = {Host-parasite}, pubstate = {published}, tppubtype = {article} } @article{SA2008, title = {Antimalarial responses in Anopheles gambiae: from a complement-like protein to a complement-like pathway}, author = {Stéphanie A Blandin and Eric Marois and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/18541213}, year = {2008}, date = {2008-06-12}, journal = {Cell Host Microbe.}, volume = {3}, number = {6}, pages = {364-74}, abstract = {Malaria transmission between humans depends on the ability of Anopheles mosquitoes to support Plasmodium development. New perspectives in vector control are emerging from understanding the mosquito immune system, which plays critical roles in parasite recognition and killing. A number of factors controlling this process have been recently identified, and key among them is TEP1, a homolog of human complement factor C3 whose binding to the parasite surface targets it for subsequent killing. Here, we review our current knowledge of mosquito factors that respond to Plasmodium infection and elaborate on the activity and mode of action of the TEP1 complement-like pathway.}, keywords = {blandin, M3i, marois, TEP1}, pubstate = {published}, tppubtype = {article} } @article{fabre_covalent_2008, title = {Covalent assembly and micropatterning of functionalized multiwalled carbon nanotubes to monolayer-modified Si(111) surfaces}, author = {Bruno Fabre and Fanny Hauquier and Cyril Herrier and Giorgia Pastorin and Wei Wu and Alberto Bianco and Maurizio Prato and Philippe Hapiot and Dodzi Zigah and Mauro Prasciolu and Lisa Vaccari}, doi = {10.1021/la800358w}, issn = {0743-7463}, year = {2008}, date = {2008-06-01}, journal = {Langmuir: the ACS journal of surfaces and colloids}, volume = {24}, number = {13}, pages = {6595--6602}, abstract = {Multiwalled carbon nanotubes (MWNTs) covalently bound to monocrystalline p-type Si(111) surfaces have been prepared by attaching soluble amine-functionalized MWNTs onto a preassembled undecanoic acid monolayer using carbodiimide coupling. SEM analysis of these functionalized surfaces shows that the bound MWNTs are parallel to the surface rather than perpendicular. The voltammetric and electrochemical impedance spectroscopy measurements reveal that the electron transfer at the MWNT-modified surface is faster than that observed at a MWNT-free alkyl monolayer. We have also demonstrated that it is possible to prepare MWNT micropatterns using this surface amidation reaction and a "reagentless" UV photolithography technique. Following this approach, MWNT patterns surrounded by n-dodecyl areas have been produced and the local electrochemical properties of these micropatterned surfaces have been examined by scanning electrochemical microscopy. In particular, it is demonstrated that the MWNT patterns allow a faster charge transfer which is consistent with the results obtained for the uniformly modified surfaces.}, keywords = {Atomic Force, carbon, Electrochemistry, Electron, I2CT, Microscopy, Nanotubes, scanning, Silicon, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{li_adsorption_2008, title = {Adsorption of carbon nanotubes on active carbon microparticles}, author = {Shouping Li and Wei Wu and Stéphane Campidelli and Veronika Sarnatskaïa and Maurizio Prato and Arlette Tridon and Andrey Nikolaev and Vladimir Nikolaev and Alberto Bianco and Elisaveta Snezhkova}, url = {http://www.sciencedirect.com/science/article/pii/S0008622308001462}, doi = {10.1016/j.carbon.2008.03.010}, issn = {0008-6223}, year = {2008}, date = {2008-06-01}, urldate = {2020-03-31}, journal = {Carbon}, volume = {46}, number = {7}, pages = {1091--1095}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{parietti_function_2008, title = {Function of CD4+,CD25+ Treg cells in MRL/lpr mice is compromised by intrinsic defects in antigen-presenting cells and effector Ŧ cells}, author = {Véronique Parietti and Fanny Monneaux and Marion Décossas and Sylviane Muller}, doi = {10.1002/art.23464}, issn = {0004-3591}, year = {2008}, date = {2008-06-01}, journal = {Arthritis and Rheumatism}, volume = {58}, number = {6}, pages = {1751--1761}, abstract = {OBJECTIVE: Naturally occurring CD4+,CD25+ Treg cells are central in the maintenance of peripheral tolerance. Impaired activity and/or a lower frequency of these cells is involved in the emergence of autoimmunity. We undertook this study to analyze relative proportions and functional alterations of Treg cells in MRL/lpr mice. METHODS: The frequency of CD4+,CD25+ T cells in the peripheral blood of healthy and autoimmune mice was compared by flow cytometry. The capacity of CD4+,CD25+ T cells to inhibit the proliferation and cytokine secretion of CD4+,CD25- T cells was assessed after polyclonal activation. RESULTS: MRL/lpr mice exhibited a normal percentage of CD4+,CD25 high T cells, and forkhead box P3 messenger RNA and protein expression in Treg cells was not altered. However, MRL/lpr Treg cells displayed a reduced capacity to suppress proliferation and to inhibit interferon-gamma secretion by syngeneic effector CD4+,CD25- T cells, as compared with syngeneic cocultures of CBA/J T cells. Moreover, effector MRL/lpr CD4+,CD25- T cells were substantially less susceptible to suppression even when cultured with CBA/J or MRL/lpr Treg cells. Crossover experiments led us to conclude that in MRL/lpr mice, each partner engaged in T cell regulation displays altered functions. Molecules involved in suppressive mechanisms (CTLA-4 and CD80/CD86) are underexpressed, and antigen-presenting cells (APCs) produce raised levels of interleukin-6, which is known to abrogate suppression. CONCLUSION: Our results suggest that although the frequency and phenotype of Treg cells in MRL/lpr mice are similar to those in normal mice, Treg cells in MRL/lpr mice are not properly stimulated by APCs and are unable to suppress proinflammatory cytokine secretion from effector T cells.}, keywords = {Animal, Animals, Antigen-Presenting Cells, Antigens, B7-1 Antigen, B7-2 Antigen, CD, Cell Communication, Cells, Coculture Techniques, CTLA-4 Antigen, Cultured, Disease Models, Female, I2CT, Interleukin-1, Interleukin-2 Receptor alpha Subunit, Lupus Erythematosus, Mice, Monneaux, Regulatory, Systemic, T-Lymphocyte Subsets, T-Lymphocytes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{kostarelos_hype_2008, title = {Hype around nanotubes creates unrealistic hopes}, author = {Kostas Kostarelos and Alberto Bianco and Maurizio Prato}, doi = {10.1038/453280c}, issn = {1476-4687}, year = {2008}, date = {2008-05-01}, journal = {Nature}, volume = {453}, number = {7193}, pages = {280}, keywords = {Adult Stem Cells, carbon, Humans, I2CT, Nanomedicine, Nanotubes, Reproducibility of Results, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{lacerda_tissue_2008, title = {Tissue histology and physiology following intravenous administration of different types of functionalized multiwalled carbon nanotubes}, author = {Lara Lacerda and Hanene Ali-Boucetta and Maria A Herrero and Giorgia Pastorin and Alberto Bianco and Maurizio Prato and Kostas Kostarelos}, doi = {10.2217/17435889.3.2.149}, issn = {1748-6963}, year = {2008}, date = {2008-04-01}, journal = {Nanomedicine (London, England)}, volume = {3}, number = {2}, pages = {149--161}, abstract = {BACKGROUND: Carbon nanotubes (CNTs) constitute one of the most important types of nanomaterials, increasingly gaining interest as tools for nanomedicine applications, such as sensors, implants or delivery systems. Our groups have reported previously that chemical functionalization of CNTs can lead to their almost complete elimination from the body of animals through the urinary excretion route. The administration of CNTs may, however, impact the physiological function of organs through which CNTs traverse or accumulate. AIM: The present study addresses the short-term impact (first 24 h) of intravenous administration of various types of multiwalled nanotubes (MWNTs) on the physiology of healthy mice. MATERIALS & METHODS: Nonfunctionalized, purified MWNTs (pMWNTs) and different types of water-dispersible, functionalized MWNTs (f-MWNTs) were tail-vein injected. Histological examination of tissues (kidney, liver, spleen and lung) harvested 24 h post-administration indicated that organ accumulation depended on the degree of ammonium (NH(3)(+)) functionalization at the f-MWNT surface. RESULTS: The higher the degree of functionalization of MWNT-NH(3)(+), the less their accumulation in tissues. pMWNTs coated with autologous serum proteins prior to injection accumulated almost entirely in the lung and liver in large dark clusters. Moreover, various indicators of serum and urine analyses also confirmed that MWNT-NH(3)(+) injections did not induce any physiological abnormality in all major organs within the first 24 h post-injection. Interestingly, no abnormalities were observed either for f-MWNTs highly functionalized with carboxylate groups (diethylentriaminepentaacetic-functionalized MWNTs) or by upscaling to the highest doses ever injected so far in vivo (20 mg/kg). CONCLUSION: The high degree of f-MWNT functionalization responsible for adequate individualization of nanotubes and not the nature of the functional groups was the critical factor leading to less tissue accumulation and normal tissue physiology at least within the first 24 h post-administration, even at the highest carbon nanotube doses ever administered in any study today.}, keywords = {Animals, carbon, Female, I2CT, Inbred BALB C, Injections, Intravenous, Mice, Nanotubes, Organ Specificity, Team-Bianco, Tissue Distribution}, pubstate = {published}, tppubtype = {article} } @article{shen_orf1spcs_2008, title = {Orf1/SpcS chaperones ExoS for type three secretion by Pseudomonas aeruginosa.}, author = {Da-Kang Shen and Lauriane Quenee and Mariette Bonnet and Lauriane Kuhn and Madiha Derouazi and Daniele Lamotte and Bertrand Toussaint and Benoit Polack}, doi = {10.1016/S0895-3988(08)60014-8}, issn = {0895-3988 0895-3988}, year = {2008}, date = {2008-04-01}, journal = {Biomedical and environmental sciences : BES}, volume = {21}, number = {2}, pages = {103--109}, abstract = {OBJECTIVE: Pseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type III secretion system (TTSS) to inject effector proteins directly into the cytosol of target cells to subvert the host cell's functions. Specialized bacterial chaperones are required for effective secretion of some effectors. To identify the chaperone of ExoS, the representative effector secreted by the TTSS of P. aeruginosa, we analyzed the role of a postulated chaperone termed Orf1. METHODS: By allelic exchange, we constructed the mutant with the deletion of gene Orf1. Analysis of secreted and cell-associated fractions was performed by SDS-PAGE and Western blotting. Using strain expressing in trans Orf1, tagged by V5 polypeptide and histidine, protein-protein interaction was determined by affinity resin pull-down assay in combination with MALDI-TOF. The role of Orf1 in the expression of exoS was evaluated by gene reporter analysis. RESULTS: Pull-down assay showed that Orf1 binds to ExoS and ExoT. Secretion profile analysis showed that Orf1 was necessary for the optimal secretion of ExoS and ExoT. However, Orf1 had no effect on the expression of exoS. CONCLUSION: Orf1 is important for the secretion of ExoS probably by maintaining ExoS in a secretion-competent conformation. We propose to name Orf1 as SpcS for "specific Pseudomonas chaperone for ExoS".}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{flacher_expression_2008, title = {Expression of langerin/CD207 reveals dendritic cell heterogeneity between inbred mouse strains}, author = {Vincent Flacher and Patrice Douillard and Smina Aït-Yahia and Patrizia Stoitzner and Valérie Clair-Moninot and Nikolaus Romani and Sem Saeland}, doi = {10.1111/j.1365-2567.2007.02785.x}, issn = {1365-2567}, year = {2008}, date = {2008-03-01}, journal = {Immunology}, volume = {123}, number = {3}, pages = {339--347}, abstract = {Langerin/CD207 is expressed by a subset of dendritic cells (DC), the epithelial Langerhans cells. However, langerin is also detected among lymphoid tissue DC. Here, we describe striking differences in langerin-expressing cells between inbred mouse strains. While langerin+ cells are observed in comparable numbers and with comparable phenotypes in the epidermis, two distinct DC subsets bear langerin in peripheral, skin-draining lymph nodes of BALB/c mice (CD11c(high) CD8alpha(high) and CD11c(low) CD8alpha(low)), whereas only the latter subset is present in C57BL/6 mice. The CD11c(high) subset is detected in mesenteric lymph nodes and spleen of BALB/c mice, but is virtually absent from C57BL/6 mice. Similar differences are observed in other mouse strains. CD11c(low) langerin+ cells represent skin-derived Langerhans cells, as demonstrated by their high expression of DEC-205/CD205, maturation markers, and recruitment to skin-draining lymph nodes upon imiquimod-induced inflammation. It will be of interest to determine the role of lymphoid tissue-resident compared to skin-derived langerin+ DC.}, keywords = {Animals, Antigen, Antigens, C-Type, CD, Cell Surface, Dendritic Cells, DERMATOLOGY, Epidermis, Expression, Immunology, Immunophenotyping, Inbred Strains, inflammation, Langerhans Cells, LECTIN, Lectins, LYMPH, LYMPH NODE, Lymph Nodes, Lymphoid Tissue, Mannose-Binding Lectins, Maturation, metabolism, Mice, Minor Histocompatibility Antigens, mouse, Phenotype, Protein, Receptor, Receptors, Species Specificity, SPLEEN, SUBSETS, Surface, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dicistronic tRNA-5S rRNA genes in Yarrowia lipolytica: an alternative TFIIIA-independent way for expression of 5S rRNA genes}, author = {J Acker and C Ozanne and R Kachouri-Lafond and C Gaillardin and C Neuveglise and C Marck}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18790808}, isbn = {18790808}, year = {2008}, date = {2008-01-01}, journal = {Nucleic Acids Res}, volume = {36}, number = {18}, pages = {5832-5844}, abstract = {In eukaryotes, genes transcribed by RNA polymerase III (Pol III) carry their own internal promoters and as such, are transcribed as individual units. Indeed, a very few cases of dicistronic Pol III genes are yet known. In contrast to other hemiascomycetes, 5S rRNA genes of Yarrowia lipolytica are not embedded into the tandemly repeated rDNA units, but appear scattered throughout the genome. We report here an unprecedented genomic organization: 48 over the 108 copies of the 5S rRNA genes are located 3' of tRNA genes. We show that these peculiar tRNA-5S rRNA dicistronic genes are expressed in vitro and in vivo as Pol III transcriptional fusions without the need of the 5S rRNA gene-specific factor TFIIIA, the deletion of which displays a viable phenotype. We also report the existence of a novel putative non-coding Pol III RNA of unknown function about 70 nucleotide-long (RUF70), the 13 genes of which are devoid of internal Pol III promoters and located 3' of the 13 copies of the tDNA-Trp (CCA). All genes embedded in the various dicistronic genes, fused 5S rRNA genes, RUF70 genes and their leader tRNA genes appear to be efficiently transcribed and their products correctly processed in vivo.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Purification of T. thermophilus translation initiation factors and characterization of the corresponding ribosome complexes by filter-binding assays}, author = {A Simonetti and S Marzi and A Myasnikov and A Fabbretti and M Yusupov and C Gualerzi and B Klaholz}, url = {https://protocolexchange.researchsquare.com/article/nprot-477/v1}, doi = {10.1038/nprot.2008.130}, year = {2008}, date = {2008-01-01}, keywords = {filter-binding assay, IF1, IF2, initiation factor purification, Ribosome, ROMBY, Thermus thermophilus, translation initiation, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Dicistronic tRNA-5S rRNA genes in Yarrowia lipolytica: an alternative TFIIIA-independent way for expression of 5S rRNA genes}, author = { J. Acker and C. Ozanne and R. Kachouri-Lafond and C. Gaillardin and C. Neuveglise and C. Marck}, year = {2008}, date = {2008-01-01}, journal = {Nucleic Acids Res}, volume = {36}, number = {18}, pages = {5832-44}, abstract = {In eukaryotes, genes transcribed by RNA polymerase III (Pol III) carry their own internal promoters and as such, are transcribed as individual units. Indeed, a very few cases of dicistronic Pol III genes are yet known. In contrast to other hemiascomycetes, 5S rRNA genes of Yarrowia lipolytica are not embedded into the tandemly repeated rDNA units, but appear scattered throughout the genome. We report here an unprecedented genomic organization: 48 over the 108 copies of the 5S rRNA genes are located 3' of tRNA genes. We show that these peculiar tRNA-5S rRNA dicistronic genes are expressed in vitro and in vivo as Pol III transcriptional fusions without the need of the 5S rRNA gene-specific factor TFIIIA, the deletion of which displays a viable phenotype. We also report the existence of a novel putative non-coding Pol III RNA of unknown function about 70 nucleotide-long (RUF70), the 13 genes of which are devoid of internal Pol III promoters and located 3' of the 13 copies of the tDNA-Trp (CCA). All genes embedded in the various dicistronic genes, fused 5S rRNA genes, RUF70 genes and their leader tRNA genes appear to be efficiently transcribed and their products correctly processed in vivo.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{ali-boucetta_multiwalled_2008, title = {Multiwalled carbon nanotube-doxorubicin supramolecular complexes for cancer therapeutics}, author = {Hanene Ali-Boucetta and Khuloud T Al-Jamal and David McCarthy and Maurizio Prato and Alberto Bianco and Kostas Kostarelos}, doi = {10.1039/b712350g}, issn = {1359-7345}, year = {2008}, date = {2008-01-01}, journal = {Chemical Communications (Cambridge, England)}, number = {4}, pages = {459--461}, abstract = {Multiwalled carbon nanotube aqueous dispersions using block copolymers are able to form supramolecular complexes with the aromatic chromophore and anticancer agent doxorubicin via pi-pi stacking and enhance its cytotoxic activity.}, keywords = {Antineoplastic Agents, Breast Neoplasms, carbon, Cultured, Doxorubicin, Electron, Humans, I2CT, Microscopy, Nanotubes, Team-Bianco, Transmission, Tumor Cells}, pubstate = {published}, tppubtype = {article} } @article{prato_functionalized_2008, title = {Functionalized carbon nanotubes in drug design and discovery}, author = {Maurizio Prato and Kostas Kostarelos and Alberto Bianco}, doi = {10.1021/ar700089b}, issn = {1520-4898}, year = {2008}, date = {2008-01-01}, journal = {Accounts of Chemical Research}, volume = {41}, number = {1}, pages = {60--68}, abstract = {Carbon nanotubes (CNTs) have been proposed and actively explored as multipurpose innovative carriers for drug delivery and diagnostic applications. Their versatile physicochemical features enable the covalent and noncovalent introduction of several pharmaceutically relevant entities and allow for rational design of novel candidate nanoscale constructs for drug development. CNTs can be functionalized with different functional groups to carry simultaneously several moieties for targeting, imaging, and therapy. Among the most interesting examples of such multimodal CNT constructs described in this Account is one carrying a fluorescein probe together with the antifungal drug amphotericin B or fluorescein and the antitumor agent methotrexate. The biological action of the drug in these cases is retained or, as in the case of amphotericin B constructs, enhanced, while CNTs are able to reduce the unwanted toxicity of the drug administered alone. Ammonium-functionalized CNTs can also be considered very promising vectors for gene-encoding nucleic acids. Indeed, we have formed stable complexes between cationic CNTs and plasmid DNA and demonstrated the enhancement of the gene therapeutic capacity in comparison to DNA alone. On the other hand, CNTs conjugated with antigenic peptides can be developed as a new and effective system for synthetic vaccine applications. What makes CNTs quite unique is their ability, first shown by our groups in 2004, to passively cross membranes of many different types of cells following a translocation mechanism that has been termed the nanoneedle mechanism. In that way, CNTs open innumerable possibilities for future drug discovery based on intracellular targets that have been hard to reach until today. Moreover, adequately functionalized CNTs as those shown in this Account can be rapidly eliminated from the body following systemic administration offering further encouragment for their development. CNT excretion rates and accumulation in organs and any reactivity with the immune system will determine the CNT safety profile and, consequently, any further pharmaceutical development. Caution is advised about the need for systematic data on the long-term fate of these very interesting and versatile nano-objects in correlation with the type of CNT material used. CNTs are gradually plyaing a bigger and more important role in the emerging field of nanomedicine; however, we need to guarantee that the great opportunities they offer will be translated into feasible and safe constructs to be included in drug discovery and development pipelines.}, keywords = {Animals, carbon, Communicable Diseases, Drug Carriers, Drug Design, Genetic Therapy, Humans, I2CT, Immunization, Nanotubes, Neoplasms, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{goulev_scalloped_2008, title = {SCALLOPED interacts with YORKIE, the nuclear effector of the hippo tumor-suppressor pathway in Drosophila}, author = {Youlian Goulev and Jean Daniel Fauny and Beatriz Gonzalez-Marti and Domenico Flagiello and Joël Silber and Alain Zider}, url = {http://www.ncbi.nlm.nih.gov/pubmed/18313299}, doi = {10.1016/j.cub.2008.02.034}, issn = {0960-9822}, year = {2008}, date = {2008-01-01}, urldate = {2011-10-24}, journal = {Current Biology: CB}, volume = {18}, number = {6}, pages = {435--441}, abstract = {In Drosophila, SCALLOPED (SD) belongs to a family of evolutionarily conserved proteins characterized by the presence of a TEA/ATTS DNA-binding domain [1, 2]. SD physically interacts with the product of the vestigial (vg) gene, where the dimer functions as a master gene controlling wing formation [3, 4]. The VG-SD dimer activates the transcription of several specific wing genes, including sd and vg themselves [5, 6]. The dimer drives cell-cycle progression by inducing expression of the dE2F1 transcription factor [7], which regulates genes involved in DNA replication and cell-cycle progression. Recently, YORKIE (YKI) was identified as a transcriptional coactivator that is the downstream effector of the Hippo signaling pathway, which controls cell proliferation and apoptosis in Drosophila[8]. We identified SD as a partner for YKI. We show that interaction between YKI and SD increases SD transcriptional activity both ex vivo in Drosophila S2 cells and in vivo in Drosophila wing discs and promotes YKI nuclear localization. We also show that YKI overexpression induces vg and dE2F1 expression and that proliferation induced by YKI or by a dominant-negative form of FAT in wing disc is significantly reduced in a sd hypomorphic mutant context. Contrary to YKI, SD is not required in all imaginal tissues. This indicates that YKI-SD interaction acts in a tissue-specific fashion and that other YKI partners must exist.}, keywords = {Animals, Cell Proliferation, Drosophila, Drosophila Proteins, HeLa Cells, Humans, I2CT, Imagerie, Intracellular Signaling Peptides and Proteins, Morphogenesis, Nuclear Proteins, Protein Kinases, Protein-Serine-Threonine Kinases, Signal Transduction, Trans-Activators, Transcription Factors, Tumor Suppressor Proteins, Wing}, pubstate = {published}, tppubtype = {article} } @article{muller_spliceosomal_2008, title = {Spliceosomal peptide P140 for immunotherapy of systemic lupus erythematosus: results of an early phase II clinical trial}, author = {Sylviane Muller and Fanny Monneaux and Nicolas Schall and Rasho K Rashkov and Boycho A Oparanov and Philippe Wiesel and Jean-Marie Geiger and Robert Zimmer}, doi = {10.1002/art.24027}, issn = {0004-3591}, year = {2008}, date = {2008-01-01}, journal = {Arthritis and Rheumatism}, volume = {58}, number = {12}, pages = {3873--3883}, abstract = {OBJECTIVE: To assess the safety, tolerability, and efficacy of spliceosomal peptide P140 (IPP-201101; sequence 131-151 of the U1-70K protein phosphorylated at Ser140), which is recognized by lupus CD4+ T cells, in the treatment of patients with systemic lupus erythematosus (SLE). METHODS: An open-label, dose-escalation phase II study was conducted in two centers in Bulgaria. Twenty patients (2 male and 18 female) with moderately active SLE received 3 subcutaneous (SC) administrations of a clinical batch of P140 peptide at 2-week intervals. Clinical evaluation was performed using approved scales. A panel of autoantibodies, including antinuclear antibodies, antibodies to extractable nuclear antigens (U1 RNP, SmD1, Ro/SSA, La/SSB), and antibodies to double-stranded DNA (anti-dsDNA), chromatin, cardiolipin, and peptides of the U1-70K protein, was tested by enzyme-linked immunosorbent assay (ELISA). The plasma levels of C-reactive protein, total Ig, IgG, IgG subclasses, IgM, IgA, and IgE, and of the cytokines interleukin-2 and tumor necrosis factor alpha were measured by ELISA and nephelometry. RESULTS: IgG anti-dsDNA antibody levels decreased by at least 20% in 7 of 10 patients who received 3 x 200 microg IPP-201101 (group 1), but only in 1 patient in the group receiving 3 x 1,000 microg IPP-201101 (group 2). Physician's global assessment of disease activity scores and scores on the SLE Disease Activity Index were significantly decreased in group 1. The changes occurred progressively in the population of responders, increased in magnitude during the treatment period, and were sustained. No clinical or biologic adverse effects were observed in the individuals, except for some local irritation at the highest concentration. CONCLUSION: IPP-201101 was found to be safe and well tolerated by subjects. Three SC doses of IPP-201101 at 200 microg significantly improved the clinical and biologic status of lupus patients.}, keywords = {Adolescent, Adult, Aged, Antibodies, Antinuclear, C-Reactive Protein, DNA, Female, Humans, I2CT, Immunotherapy, Lupus Erythematosus, Male, Middle Aged, Monneaux, Peptide Fragments, Peptides, Severity of Illness Index, Spliceosomes, Systemic, Team-Dumortier, Treatment Outcome, Young Adult}, pubstate = {published}, tppubtype = {article} } @article{dieker_apoptosis-linked_2008, title = {Apoptosis-linked changes in the phosphorylation status and subcellular localization of the spliceosomal autoantigen U1-70K}, author = {J Dieker and B Cisterna and F Monneaux and M Decossas and J van der Vlag and M Biggiogera and S Muller}, doi = {10.1038/sj.cdd.4402312}, issn = {1350-9047}, year = {2008}, date = {2008-01-01}, journal = {Cell Death and Differentiation}, volume = {15}, number = {4}, pages = {793--804}, abstract = {Apoptosis consists of highly regulated pathways involving post-translational modifications and cleavage of proteins leading to sequential inactivation of the main cellular processes. Here, we focused on the apoptotic processing of one of the essential components of the mRNA splicing machinery, the U1-70K snRNP protein. We found that at an early stage of apoptosis, before the cleavage of the C-terminal part of the protein by caspase-3, the basal phosphorylation of the Ser140 residue located within the RNA recognition motif, increases very significantly. A caspase-dependent, PP1-mediated dephosphorylation of other serine residues takes place in a subset of U1-70K proteins. The U1-70K protein phosphorylated at Ser140 is clustered in heterogeneous ectopic RNP-derived structures, which are finally extruded in apoptotic bodies. The elaborate processing of the spliceosomal U1-70K protein we identified might play an important role in the regulated breakdown of the mRNA splicing machinery during early apoptosis. In addition, these specific changes in the phosphorylation/dephosphorylation balance and the subcellular localization of the U1-70K protein might explain why the region encompassing the Ser140 residue becomes a central autoantigen during the autoimmune disease systemic lupus erythematosus.}, keywords = {Apoptosis, Autoantigens, Autoimmunity, Caspase 3, Chromatin, HeLa Cells, Humans, I2CT, Jurkat Cells, Lupus Erythematosus, Monneaux, Phosphorylation, Post-Translational, Protein Phosphatase 1, Protein Processing, Protein Transport, Recombinant Proteins, Ribonucleoprotein, RNA Splicing, Serine, Spliceosomes, Systemic, Team-Dumortier, Time Factors, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{kwan_dermal-type_2008, title = {Dermal-type macrophages expressing CD209/DC-SIGN show inherent resistance to dengue virus growth}, author = {Wing-Hong Kwan and Erika Navarro-Sanchez and Hélène Dumortier and Marion Decossas and Hortense Vachon and Flavia Barreto dos Santos and Hervé W Fridman and Félix A Rey and Eva Harris and Philippe Despres and Christopher G Mueller}, doi = {10.1371/journal.pntd.0000311}, issn = {1935-2735}, year = {2008}, date = {2008-01-01}, journal = {PLoS neglected tropical diseases}, volume = {2}, number = {10}, pages = {e311}, abstract = {BACKGROUND: An important question in dengue pathogenesis is the identity of immune cells involved in the control of dengue virus infection at the site of the mosquito bite. There is evidence that infection of immature myeloid dendritic cells plays a crucial role in dengue pathogenesis and that the interaction of the viral envelope E glycoprotein with CD209/DC-SIGN is a key element for their productive infection. Dermal macrophages express CD209, yet little is known about their role in dengue virus infection. METHODS AND FINDINGS: Here, we showed that dermal macrophages bound recombinant envelope E glycoprotein fused to green fluorescent protein. Because dermal macrophages stain for IL-10 in situ, we generated dermal-type macrophages from monocytes in the presence of IL-10 to study their infection by dengue virus. The macrophages were able to internalize the virus, but progeny virus production was undetectable in the infected cells. In addition, no IFN-alpha was produced in response to the virus. The inability of dengue virus to grow in the macrophages was attributable to accumulation of internalized virus particles into poorly-acidified phagosomes. CONCLUSIONS: Aborting infection by viral sequestration in early phagosomes would present a novel means to curb infection of enveloped virus and may constitute a prime defense system to prevent dengue virus spread shortly after the bite of the infected mosquito.}, keywords = {C-Type, Cell Adhesion Molecules, Cell Line, Cell Surface, Cells, Cultured, Dengue, Dengue virus, Dumortier, Gene Expression, Humans, I2CT, Lectins, Macrophages, Protein Binding, Receptors, Skin, Team-Dumortier, Team-Mueller, Viral Envelope Proteins}, pubstate = {published}, tppubtype = {article} } @article{lacerda_dynamic_2008, title = {Dynamic Imaging of Functionalized Multi-Walled Carbon Nanotube Systemic Circulation and Urinary Excretion}, author = {L Lacerda and A Soundararajan and R Singh and G Pastorin and K T Al‐Jamal and J Turton and P Frederik and M A Herrero and S Li and A Bao and D Emfietzoglou and S Mather and W T Phillips and M Prato and A Bianco and B Goins and K Kostarelos}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adma.200702334}, doi = {10.1002/adma.200702334}, issn = {1521-4095}, year = {2008}, date = {2008-01-01}, urldate = {2020-03-31}, journal = {Advanced Materials}, volume = {20}, number = {2}, pages = {225--230}, abstract = {Intravenously administered multi-walled carbon nanotubes, functionalized with DTPA and radiolabeled with Indium-111, were dynamically tracked in vivo using a microSingle Photon Emission Tomography scanner. Imaging showed that nanotubes enter the systemic blood circulation and within 5 min begin to permeate through the renal glomerular filtration system into the bladder.}, keywords = {Biomedical applications, Carbon nanotubes, I2CT, multiwalled, Nanomaterials, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{lacotte_interfacing_2008, title = {Interfacing Functionalized Carbon Nanohorns with Primary Phagocytic Cells}, author = {Stéphanie Lacotte and Ainara García and Marion Décossas and Wafa' T Al‐Jamal and Shouping Li and Kostas Kostarelos and Sylviane Muller and Maurizio Prato and Hélène Dumortier and Alberto Bianco}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adma.200702753}, doi = {10.1002/adma.200702753}, issn = {1521-4095}, year = {2008}, date = {2008-01-01}, urldate = {2020-03-31}, journal = {Advanced Materials}, volume = {20}, number = {12}, pages = {2421--2426}, abstract = {Functionalized carbon nanohorns (f-CNH) are uptaken by macrophages, without affecting cell viability. f-CNH induce the production of reactive oxygen species and of pro-inflammatory cytokines. The level of inflammation, although moderate, should be taken into consideration when using f-CNH for drug delivery. However, it could be exploited as an intrinsic f-CNH adjuvant function for biomedical applications requiring some activation of the immune system.}, keywords = {carbon nanostructures, Cells, Cytotoxicity, Drug delivery, I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{goto_akirins_2008, title = {Akirins are highly conserved nuclear proteins required for NF-kappaB-dependent gene expression in drosophila and mice}, author = {Akira Goto and Kazufumi Matsushita and Viola Gesellchen and Laure El Chamy and David Kuttenkeuler and Osamu Takeuchi and Jules A Hoffmann and Shizuo Akira and Michael Boutros and Jean-Marc Reichhart}, doi = {10.1038/ni1543}, issn = {1529-2916}, year = {2008}, date = {2008-01-01}, journal = {Nat. Immunol.}, volume = {9}, number = {1}, pages = {97--104}, abstract = {During a genome-wide screen with RNA-mediated interference, we isolated CG8580 as a gene involved in the innate immune response of Drosophila melanogaster. CG8580, which we called Akirin, encoded a protein that acted in parallel with the NF-kappaB transcription factor downstream of the Imd pathway and was required for defense against Gram-negative bacteria. Akirin is highly conserved, and the human genome contains two homologs, one of which was able to rescue the loss-of-function phenotype in drosophila cells. Akirins were strictly localized to the nucleus. Knockout of both Akirin homologs in mice showed that one had an essential function downstream of the Toll-like receptor, tumor necrosis factor and interleukin (IL)-1beta signaling pathways leading to the production of IL-6. Thus, Akirin is a conserved nuclear factor required for innate immune responses.}, keywords = {Animals, Cell Line, Embryo, Fibroblasts, hoffmann, Humans, Immunity, Innate, Interleukin-1beta, M3i, Mammalian, Mice, NF-kappa B, Nuclear Proteins, Proteins, reichhart, Signal Transduction, Toll-Like Receptors, transgenic, Tumor Necrosis Factor-alpha}, pubstate = {published}, tppubtype = {article} } @article{roetzer_candida_2008b, title = {Candida glabrata environmental stress response involves Saccharomyces cerevisiae Msn2/4 orthologous transcription factors}, author = {Andreas Roetzer and Christa Gregori and Ann Marie Jennings and Jessica Quintin and Dominique Ferrandon and Geraldine Butler and Karl Kuchler and Gustav Ammerer and Christoph Schüller}, doi = {10.1111/j.1365-2958.2008.06301.x}, issn = {1365-2958}, year = {2008}, date = {2008-01-01}, journal = {Mol. Microbiol.}, volume = {69}, number = {3}, pages = {603--620}, abstract = {We determined the genome-wide environmental stress response (ESR) expression profile of Candida glabrata, a human pathogen related to Saccharomyces cerevisiae. Despite different habitats, C. glabrata, S. cerevisiae, Schizosaccharomyces pombe and Candida albicans have a qualitatively similar ESR. We investigate the function of the C. glabrata syntenic orthologues to the ESR transcription factor Msn2. The C. glabrata orthologues CgMsn2 and CgMsn4 contain a motif previously referred to as HD1 (homology domain 1) also present in Msn2 orthologues from fungi closely related to S. cerevisiae. We show that regions including this motif confer stress-regulated intracellular localization when expressed in S. cerevisiae. Site-directed mutagenesis confirms that nuclear export of CgMsn2 in C. glabrata requires an intact HD1. Transcript profiles of CgMsn2/4 mutants and CgMsn2 overexpression strains show that they regulate a part of the CgESR. CgMsn2 complements a S. cerevisiae msn2 null mutant and in stressed C. glabrata cells, rapidly translocates from the cytosol to the nucleus. CgMsn2 is required for full resistance against severe osmotic stress and rapid and full induction of trehalose synthesis genes (TPS1, TPS2). Constitutive activation of CgMsn2 is detrimental for C. glabrata. These results establish an Msn2-regulated general stress response in C. glabrata.}, keywords = {Animals, Candida glabrata, Candidiasis, DNA-Binding Proteins, ferrandon, Fungal, Fungal Proteins, Gene Expression Profiling, Gene Expression Regulation, Genetic, Humans, M3i, Oligonucleotide Array Sequence Analysis, Osmotic Pressure, Regulon, Saccharomyces cerevisiae Proteins, Transcription, Transcription Factors, Virulence, Yeasts}, pubstate = {published}, tppubtype = {article} } @article{huszar_drosophila_2008, title = {Drosophila viruses and the study of antiviral host-defense}, author = {Tünde Huszar and Jean-Luc Imler}, doi = {10.1016/S0065-3527(08)00406-5}, issn = {0065-3527}, year = {2008}, date = {2008-01-01}, journal = {Advances in Virus Research}, volume = {72}, pages = {227--265}, abstract = {The fruit fly Drosophila melanogaster is a powerful model to study host-pathogen interactions. Most studies so far have focused on extracellular pathogens such as bacteria and fungi. More recently, viruses have come to the front, and RNA interference was shown to play a critical role in the control of viral infections in drosophila. We review here our current knowledge on drosophila viruses. A diverse set of RNA viruses belonging to several families (Rhabdoviridae, Dicistroviridae, Birnaviridae, Reoviridae, Errantiviridae) has been reported in D. melanogaster. By contrast, no DNA virus has been recovered up to now. The drosophila viruses represent powerful tools to study virus-cell interactions in vivo. Analysis of the literature however reveals that for many of them, important gaps exist in our understanding of their replication cycle, genome organization, morphology or pathogenesis. The data obtained in the past few years on antiviral defense mechanisms in drosophila, which point to evolutionary conserved pathways, highlight the potential of the D. melanogaster model to study antiviral innate immunity and to better understand the complex interaction between arthropod-borne viruses and their insect vectors.}, keywords = {Animals, Host-Pathogen Interactions, imler, Immunity, Innate, Insect Viruses, M3i, RNA Interference, RNA Viruses}, pubstate = {published}, tppubtype = {article} } @article{SA2008b, title = {Reverse Genetics Analysis of Antiparasitic Responses in the Malaria Vector, Anopheles gambiae}, author = {Stéphanie A Blandin and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/18370165}, year = {2008}, date = {2008-01-01}, journal = {Methods Mol Biol.}, volume = {415}, pages = {365-77}, abstract = {Anopheles mosquitoes are the major vectors of human malaria parasites. Mosquito-parasite interactions are critical for disease transmission and therefore represent a potential target for malaria control strategies. Mosquitoes mount potent antiparasitic responses, and identification of mosquito factors that limit parasite development is one of the major objectives in the field. To address this question, we have developed a convenient reverse genetics approach by injection of double-stranded RNA (dsRNA) in adult mosquitoes, to evaluate the function of candidate genes in mosquito antiparasitic responses.}, keywords = {blandin, M3i, RNAi}, pubstate = {published}, tppubtype = {article} } @article{barbaroux_epidermal_2008, title = {Epidermal receptor activator of NF-kappaB ligand controls Langerhans cells numbers and proliferation}, author = {J B Barbaroux and M Beleut and C Brisken and C G Mueller and R W Groves}, year = {2008}, date = {2008-01-01}, journal = {Journal of Immunology}, volume = {181}, number = {1550-6606 (Electronic)}, pages = {1103--1108}, abstract = {Langerhans cells (LC) are the dendritic APC population of the epidermis, where they reside for long periods and are self-replicating. The molecular signals underlying these characteristics are unknown. The TNF superfamily member receptor activator of NF-kappaB ligand (RANKL, TNFSF11) has been shown to sustain viability of blood dendritic cells in addition to its role in promoting proliferation and differentiation of several cell types, notably osteoclasts. In this study, we have studied expression of the RANKL system in skin and have defined a key role for this molecule in LC homeostasis. In vitro and in vivo, human KC expressed RANKL and epidermal LC expressed cell surface RANK. In vitro, RANKL sustained CD34(+) progenitor-derived LC viability following 72-h cultures in cytokine-free medium (79.5 +/- 1% vs 55.2 +/- 5.7% live cells, respectively; n = 4; p textless 0.05). In vivo, RANKL-deficient mice displayed a marked reduction in epidermal LC density (507.1 +/- 77.2 vs 873.6 +/- 41.6 LC per mm(2); n = 9; p textless 0.05) and their proliferation was impaired without a detectable effect on apoptosis. These data indicate a key role for the RANKL system in the regulation of LC survival within the skin and suggest a regulatory role for KC in the maintenance of epidermal LC homeostasis}, keywords = {APC, Apoptosis, BLOOD, Cell Count, Cell Proliferation, Cell Survival, Culture, cytology, Dendritic Cells, DERMATOLOGY, Differentiation, Epidermis, Expression, Homeostasis, Human, Humans, Immunology, IN VITRO, In vivo, KERATINOCYTES, Langerhans Cells, ligand, metabolism, Mice, NF-kappa B, NF-kappaB, OSTEOCLAST, Osteoclasts, Proliferation, Protein, rank, RANK ligand, Receptor, Receptor Activator of Nuclear Factor-kappa B, Regulation, Signal Transduction, Skin, survival, Team-Mueller, viability}, pubstate = {published}, tppubtype = {article} } @incollection{romani_langerhans_2008, title = {Langerhans cells - dendritic cells of the epidermis and other epithelia}, author = {N Romani and S Ebner and V Flacher and C H Tripp and C Heufler and B E Clausen and P Stoitzner}, editor = {S Saeland}, year = {2008}, date = {2008-01-01}, booktitle = {Recent Advances in Skin Immunology}, publisher = {Research Signpost}, address = {Trivandrum, Kerala, India}, abstract = {Langerhans cells are dendritic cells that reside in epithelia, formeost in the epidermis. Like dendritic cells from non-epithelial tissues or from the blood, they form a functional bridge between the innate and the adaptive immune system. Although Langerhans cells have first been described 140 years ago, only recently has a lively scientific debate arisen as to their functional role in vivo, i.e., in the living organism. This is mainly due to the advent of modern, sophisticated experimental models that allow to tackle hitherto unaddressed problems. It is not yet entirely clear whether an immunogenic or a tolerogenic function of Langerhans cells prevails in vivo. Here, we attempt to summarize and discuss the current knowledge on the immunobiology of Langerhans cells with emphasis on their role in vivo.}, keywords = {BLOOD, Dendritic Cells, Epidermis, Epithelium, function, Immune System, Immunology, In vivo, Langerhans Cells, Skin, Team-Mueller}, pubstate = {published}, tppubtype = {incollection} } @article{tripp_lymph_2008, title = {The lymph vessel network in mouse skin visualised with antibodies against the hyaluronan receptor LYVE-1}, author = {Christoph H Tripp and Bernhard Haid and Vincent Flacher and Michael Sixt and Hannes Peter and Julia Farkas and Robert Gschwentner and Lydia Sorokin and Nikolaus Romani and Patrizia Stoitzner}, doi = {10.1016/j.imbio.2008.07.025}, issn = {0171-2985}, year = {2008}, date = {2008-01-01}, journal = {Immunobiology}, volume = {213}, number = {9-10}, pages = {715--728}, abstract = {Langerhans cells and dermal dendritic cells migrate to the draining lymph nodes through dermal lymphatic vessels. They do so in the steady-state and under inflammatory conditions. Peripheral T cell tolerance or T cell priming, respectively, are the consequences of migration. The nature of dendritic cell-containing vessels was mostly defined by electron microscopy or by their lack of blood endothelial markers. Selective markers for murine lymph endothelium were hitherto rare or not available. Here, we utilised recently developed antibodies against the murine hyaluronan receptor, LYVE-1, to study the lymph vessel network in mouse skin in more detail. In hairless skin from the ears, lymph vessels were spread out in a horizontal plane. They formed anastomoses, and they possessed frequent blind endings that were occasionally open. Lymph vessels were wider than blood vessels, which were identified by their strong CD31 expression. In body wall skin LYVE-1 reactive vessels did not extend laterally but they dived straight down into the deeper dermis. There, they are connected to each other and formed a network similar to ear skin. The number and width of lymph vessels did not grossly change upon inflammatory stimuli such as skin explant culture or tape stripping. There were also no marked changes in caliber in response to the TLR 7/8 ligand Imiquimod. Double-labelling experiments of cultured skin showed that most of the strongly cell surface MHC II-expressing (i.e. activated) dendritic cells were confined to the lymph vessels. Langerin/CD207(+) cells within this population appeared later than dermal dendritic cells, i.e. langerin-negative cells. Comparable results were obtained after stimulating the skin in vivo with the TLR 7/8 ligand Imiquimod or by tape stripping. In untreated skin (i.e. steady state) a few MHC II(+) and Langerin/CD207(+) cells, presumably migrating skin dendritic cells including epidermal Langerhans cells, were consistently observed within the lymph vessels. The novel antibody reagents may serve as important tools to further study the dendritic cell traffic in the skin under physiological conditions as well as in conditions of adoptive dendritic cell transfer in immunotherapy.}, keywords = {anatomy & histology, Animals, Antibodies, antibody, BLOOD, Blood Vessels, CD31, Cell Movement, Culture, cytology, Dendritic Cells, DERMAL DENDRITIC CELLS, DERMATOLOGY, DERMIS, EAR, electron microscopy, ENDOTHELIUM, Expression, GLYCOPROTEIN, Glycoproteins, hyaluronan, imiquimod, Immunology, Immunotherapy, In vivo, Inbred BALB C, Inbred C57BL, Langerhans Cells, ligand, LYMPH, LYMPH NODE, Lymph Nodes, LYMPHATIC VESSEL, Lymphatic Vessels, LYVE-1, Membrane Transport Proteins, metabolism, MHC, Mice, migration, mouse, murine, physiology, priming, Protein, Receptor, Skin, tape stripping, Team-Mueller, tolerance}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comprehensive features of natural and in vitro selected GNRA tetraloop-binding receptors}, author = { C. Geary and S. Baudrey and L. Jaeger}, year = {2008}, date = {2008-01-01}, journal = {Nucleic Acids Res}, volume = {36}, number = {4}, pages = {1138-52}, abstract = {Specific recognitions of GNRA tetraloops by small helical receptors are among the most widespread long-range packing interactions in large ribozymes. However, in contrast to GYRA and GAAA tetraloops, very few GNRA/receptor interactions have yet been identified to involve GGAA tetraloops in nature. A novel in vitro selection scheme based on a rigid self-assembling tectoRNA scaffold designed for isolation of intermolecular interactions with A-minor motifs has yielded new GGAA tetraloop-binding receptors with affinity in the nanomolar range. One of the selected receptors is a novel 12 nt RNA motif, (CCUGUG. AUCUGG), that recognizes GGAA tetraloop hairpin with a remarkable specificity and affinity. Its physical and chemical characteristics are comparable to those of the well-studied '11nt' GAAA tetraloop receptor motif. A second less specific motif (CCCAGCCC. GAUAGGG) binds GGRA tetraloops and appears to be related to group IC3 tetraloop receptors. Mutational, thermodynamic and comparative structural analysis suggests that natural and in vitro selected GNRA receptors can essentially be grouped in two major classes of GNRA binders. New insights about the evolution, recognition and structural modularity of GNRA and A-minor RNA-RNA interactions are proposed.}, note = {1362-4962 (Electronic) Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S.}, keywords = {Acid, Adenine/chemistry, Analysis, Base, Conformation, Data, dimerization, directed, Evolution, KROL, Models, Molecular, Nucleic, RNA, RNA/*chemistry/classification, Sequence, Thermodynamics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Recognition of tRNALeu by Aquifex aeolicus leucyl-tRNA synthetase during the aminoacylation and editing steps}, author = {P Yao and B Zhu and S Jaeger and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18367476}, isbn = {18367476}, year = {2008}, date = {2008-01-01}, journal = {Nucleic Acids Res}, volume = {36}, number = {8}, pages = {2728-2738}, abstract = {Recognition of tRNA by the cognate aminoacyl-tRNA synthetase during translation is crucial to ensure the correct expression of the genetic code. To understand tRNA(Leu) recognition sets and their evolution, the recognition of tRNA(Leu) by the leucyl-tRNA synthetase (LeuRS) from the primitive hyperthermophilic bacterium Aquifex aeolicus was studied by RNA probing and mutagenesis. The results show that the base A73; the core structure of tRNA formed by the tertiary interactions U8-A14, G18-U55 and G19-C56; and the orientation of the variable arm are critical elements for tRNA(Leu) aminoacylation. Although dispensable for aminoacylation, the anticodon arm carries discrete editing determinants that are required for stabilizing the conformation of the post-transfer editing state and for promoting translocation of the tRNA acceptor arm from the synthetic to the editing site.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Amino Acids/chemistry Anticodon/chemistry Bacteria/enzymology/genetics Base Sequence Iodine Leucine-tRNA Ligase/chemistry/*metabolism Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Protein Footprinting RNA, ERIANI, Leu/*chemistry/genetics/metabolism Ribonucleases Substrate Specificity *Transfer RNA Aminoacylation, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Toward predicting self-splicing and protein-facilitated splicing of group I introns}, author = {Q Vicens and P J Paukstelis and E Westhof and A M Lambowitz and T R Cech}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18768647}, isbn = {18768647}, year = {2008}, date = {2008-01-01}, journal = {RNA}, volume = {14}, number = {10}, pages = {2013-2029}, abstract = {In the current era of massive discoveries of noncoding RNAs within genomes, being able to infer a function from a nucleotide sequence is of paramount interest. Although studies of individual group I introns have identified self-splicing and nonself-splicing examples, there is no overall understanding of the prevalence of self-splicing or the factors that determine it among the >2300 group I introns sequenced to date. Here, the self-splicing activities of 12 group I introns from various organisms were assayed under six reaction conditions that had been shown previously to promote RNA catalysis for different RNAs. Besides revealing that assessing self-splicing under only one condition can be misleading, this survey emphasizes that in vitro self-splicing efficiency is correlated with the GC content of the intron (>35% GC was generally conductive to self-splicing), and with the ability of the introns to form particular tertiary interactions. Addition of the Neurospora crassa CYT-18 protein activated splicing of two nonself-splicing introns, but inhibited the second step of self-splicing for two others. Together, correlations between sequence, predicted structure and splicing begin to establish rules that should facilitate our ability to predict the self-splicing activity of any group I intron from its sequence.}, note = {1469-9001 (Electronic) Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't}, keywords = {Base Composition Base Sequence Catalysis *Introns Neurospora crassa/enzymology *Nucleic Acid Conformation *RNA Splicing RNA, Catalytic/*chemistry RNA, Messenger/*chemistry Tyrosine-tRNA Ligase/*chemistry, Unité ARN, WESTHOF, WESTHOF Base Composition Base Sequence Catalysis *Introns Neurospora crassa/enzymology *Nucleic Acid Conformation *RNA Splicing RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {Program CRYC3D for geometric computing in crystallography}, author = {L Urzhumtseva and A Urzhumtsev}, url = {http://scripts.iucr.org/cgi-bin/paper?kk5021}, isbn = {10.1107/S0021889808003415}, year = {2008}, date = {2008-01-01}, journal = {J Appl Cryst}, volume = {41}, pages = {479-480}, abstract = {The computer program CRYC3D works with three-dimensional crystallographic geometric objects or groups of them and calculates their basic geometric characteristics by a simple click in the menu. In particular, this includes vector operations in both direct and reciprocal spaces and cell transformations. Collecting basic crystallographic operations in a single and simple program helps crystallographers to avoid looking for `fast-and-dirty' scripts or using large and unwieldy packages and may be useful in everyday work. When running the program in its principal mode, macro-operations are accompanied by a list of elementary geometric operations. This feature, together with the presence of a single-command mode and online help, may be useful also as a teaching tool.}, keywords = {Computer programs Crystallographic computing Teaching Tcl/Tk, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Handling mammalian mitochondrial tRNAs and aminoacyl-tRNA synthetases for functional and structural characterization}, author = {M Sissler and B Lorber and M Messmer and A Schaller and J Putz and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18241799}, isbn = {18241799}, year = {2008}, date = {2008-01-01}, journal = {Methods}, volume = {44}, number = {2}, pages = {176-189}, abstract = {The mammalian mitochondrial (mt) genome codes for only 13 proteins, which are essential components in the process of oxidative phosphorylation of ADP into ATP. Synthesis of these proteins relies on a proper mt translation machinery. While 22 tRNAs and 2 rRNAs are also coded by the mt genome, all other factors including the set of aminoacyl-tRNA synthetases (aaRSs) are encoded in the nucleus and imported. Investigation of mammalian mt aminoacylation systems (and mt translation in general) gains more and more interest not only in regard of evolutionary considerations but also with respect to the growing number of diseases linked to mutations in the genes of either mt-tRNAs, synthetases or other factors. Here we report on methodological approaches for biochemical, functional, and structural characterization of human/mammalian mt-tRNAs and aaRSs. Procedures for preparation of native and in vitro transcribed tRNAs are accompanied by recommendations for specific handling of tRNAs incline to structural instability and chemical fragility. Large-scale preparation of mg amounts of highly soluble recombinant synthetases is a prerequisite for structural investigations that requires particular optimizations. Successful examples leading to crystallization of four mt-aaRSs and high-resolution structures are recalled and limitations discussed. Finally, the need for and the state-of-the-art in setting up an in vitro mt translation system are emphasized. Biochemical characterization of a subset of mammalian aminoacylation systems has already revealed a number of unprecedented peculiarities of interest for the study of evolution and forensic research. Further efforts in this field will certainly be rewarded by many exciting discoveries.}, note = {1046-2023 (Print) Journal article}, keywords = {FLORENTZ, FLORENTZ GIEGE, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of the 30S translation initiation complex}, author = {A Simonetti and S Marzi and A G Myasnikov and A Fabbretti and M Yusupov and C O Gualerzi and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18758445}, isbn = {18758445}, year = {2008}, date = {2008-01-01}, journal = {Nature}, volume = {455}, number = {7211}, pages = {416-420}, abstract = {Translation initiation, the rate-limiting step of the universal process of protein synthesis, proceeds through sequential, tightly regulated steps. In bacteria, the correct messenger RNA start site and the reading frame are selected when, with the help of initiation factors IF1, IF2 and IF3, the initiation codon is decoded in the peptidyl site of the 30S ribosomal subunit by the fMet-tRNA(fMet) anticodon. This yields a 30S initiation complex (30SIC) that is an intermediate in the formation of the 70S initiation complex (70SIC) that occurs on joining of the 50S ribosomal subunit to the 30SIC and release of the initiation factors. The localization of IF2 in the 30SIC has proved to be difficult so far using biochemical approaches, but could now be addressed using cryo-electron microscopy and advanced particle separation techniques on the basis of three-dimensional statistical analysis. Here we report the direct visualization of a 30SIC containing mRNA, fMet-tRNA(fMet) and initiation factors IF1 and GTP-bound IF2. We demonstrate that the fMet-tRNA(fMet) is held in a characteristic and precise position and conformation by two interactions that contribute to the formation of a stable complex: one involves the transfer RNA decoding stem which is buried in the 30S peptidyl site, and the other occurs between the carboxy-terminal domain of IF2 and the tRNA acceptor end. The structure provides insights into the mechanism of 70SIC assembly and rationalizes the rapid activation of GTP hydrolysis triggered on 30SIC-50S joining by showing that the GTP-binding domain of IF2 would directly face the GTPase-activated centre of the 50S subunit.}, note = {1476-4687 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Cryoelectron Microscopy Crystallography, Messenger/chemistry/genetics/metabolism RNA, Met/chemistry/genetics/metabolism/ultrastructure Ribosome Subunits/chemistry/metabolism/ultrastructure Ribosomes/chemistry/*metabolism/*ultrastructure Thermus thermophilus/*enzymology/genetics/*ultrastructure, Molecular Multiprotein Complexes/*chemistry/genetics/metabolism/*ultrastructure *Peptide Chain Initiation, ROMBY, Transfer, Translational Prokaryotic Initiation Factor-1/chemistry/genetics/metabolism/ultrastructure Prokaryotic Initiation Factor-2/chemistry/genetics/metabolism/ultrastructure Protein Conformation RNA, Unité ARN, X-Ray Guanosine Triphosphate/chemistry/metabolism Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Post-Translational Modifications Guard Yeast from Misaspartylation}, author = {M Ryckelynck and C A Paulus and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/pubmed/18956885}, isbn = {18956891}, year = {2008}, date = {2008-01-01}, journal = {Biochemistry}, volume = {47}, number = {47}, pages = {12476-12482}, abstract = {Yeast aspartyl-tRNA synthetase (AspRS) is downregulated at the post-transcriptional level. This complex retro-inhibition mechanism causes the cell to equilibrate cellular concentrations of tRNA (Asp), AspRS, and its encoding mRNA. This strategy hinders AspRS accumulation to keep misacylation of heterologous tRNAs under control. Here, the AspRS concentration was increased artificially in vivo but did not generate tRNA (Asn) and/or tRNA (Glu) misaspartylation or the logical consecutive post-translational stress. This work allowed the detection of an additional subtle cellular lock capable of blocking AspRS toxicity. This study revealed the presence of post-translational modifications in the N-terminal extension of AspRS. We hypothesize that by neutralizing the lysine-rich motif contained in this domain, the cell mobilizes an additional strategy that considerably reduces the probability of the enzyme binding and aspartylating noncognate tRNAs and thus harming its own translation.}, note = {1520-4995 (Electronic) 0006-2960 (Linking) Journal article}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Exploring the complex world of RNA regulation}, author = {P Romby and E G Wagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18072939}, isbn = {18072939}, year = {2008}, date = {2008-01-01}, journal = {Biol Cell}, volume = {100}, number = {1}, pages = {e1-3}, note = {1768-322X (Electronic) Editorial}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ex vivo correction of selenoprotein N deficiency in rigid spine muscular dystrophy caused by a mutation in the selenocysteine codon}, author = {M Rederstorff and V Allamand and P Guicheney and C Gartioux and P Richard and D Chaigne and A Krol and A Lescure}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18025044}, isbn = {18025044}, year = {2008}, date = {2008-01-01}, journal = {Nucleic Acids Res}, volume = {36}, number = {1}, pages = {237-244}, abstract = {Premature termination of translation due to nonsense mutations is a frequent cause of inherited diseases. Therefore, many efforts were invested in the development of strategies or compounds to selectively suppress this default. Selenoproteins are interesting candidates considering the idiosyncrasy of the amino acid selenocysteine (Sec) insertion mechanism. Here, we focused our studies on SEPN1, a selenoprotein gene whose mutations entail genetic disorders resulting in different forms of muscular diseases. Selective correction of a nonsense mutation at the Sec codon (UGA to UAA) was undertaken with a corrector tRNA(Sec) that was engineered to harbor a compensatory mutation in the anticodon. We demonstrated that its expression restored synthesis of a full-length selenoprotein N both in HeLa cells and in skin fibroblasts from a patient carrying the mutated Sec codon. Readthrough of the UAA codon was effectively dependent on the Sec insertion machinery, therefore being highly selective for this gene and unlikely to generate off-target effects. In addition, we observed that expression of the corrector tRNA(Sec) stabilized the mutated SEPN1 transcript that was otherwise more subject to degradation. In conclusion, our data provide interesting evidence that premature termination of translation due to nonsense mutations is amenable to correction, in the context of the specialized selenoprotein synthesis mechanism.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Amino Acid-Specific/*genetics Selenocysteine/metabolism Selenoproteins/biosynthesis/*deficiency/*genetics Transgenes, KROL Codon/chemistry *Codon, LESCURE, Nonsense Fibroblasts/metabolism Hela Cells Humans Muscle Proteins/biosynthesis/*deficiency/*genetics Muscular Atrophy, Spinal/*genetics/metabolism RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Implications of recombination for HIV diversity}, author = {B C Ramirez and E Simon-Loriere and R Galetto and M Negroni}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18308413}, isbn = {18308413}, year = {2008}, date = {2008-01-01}, journal = {Virus Res}, volume = {134}, number = {1-2}, pages = {64-73}, abstract = {The human immunodeficiency virus (HIV) population is characterised by extensive genetic variability that results from high error and recombination rates of the reverse transcription process, and from the fast turnover of virions in HIV-infected individuals. Among the viral variants encountered at the global scale, recombinant forms are extremely abundant. Some of these recombinants (known as circulating recombinant forms) become fixed and undergo rapid expansion in the population. The reasons underlying their epidemiological success remain at present poorly understood and constitute a fascinating area for future research to improve our understanding of immune escape, pathogenicity and transmission. Recombinant viruses are generated during reverse transcription as a consequence of template switching between the two genetically different genomic RNAs present in a heterozygous virus. Recombination can thereby generate shortcuts in evolution by producing mosaic reverse transcription products of parental genomes. Therefore, in a single infectious cycle multiple mutations that are positively selected can be combined or, conversely, negatively selected mutations can be removed. Recombination is therefore involved in different aspects of HIV evolution, adaptation to its host, and escape from antiviral treatments.}, note = {0168-1702 (Print) 0168-1702 (Linking) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {*Genetic Variation Genome, Genetic, NEGRONI, Unité ARN, Viral HIV/classification/*genetics/immunology/isolation & purification HIV Infections/drug therapy/immunology/transmission/virology Humans *Recombination}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dual-targeted tRNA-dependent amidotransferase ensures both mitochondrial and chloroplastic Gln-tRNAGln synthesis in plants}, author = {C Pujol and M Bailly and D Kern and L Marechal-Drouard and H Becker and A M Duchene}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18441100}, isbn = {18441100}, year = {2008}, date = {2008-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {105}, number = {17}, pages = {6481-6485}, abstract = {Aminoacyl-tRNAs are generally formed by direct attachment of an amino acid to tRNAs by aminoacyl-tRNA synthetases, but Gln-tRNA is an exception to this rule. Gln-tRNA(Gln) is formed by this direct pathway in the eukaryotic cytosol and in protists or fungi mitochondria but is formed by an indirect transamidation pathway in most of bacteria, archaea, and chloroplasts. We show here that the formation of Gln-tRNA(Gln) is also achieved by the indirect pathway in plant mitochondria. The mitochondrial-encoded tRNA(Gln), which is the only tRNA(Gln) present in mitochondria, is first charged with glutamate by a nondiscriminating GluRS, then is converted into Gln-tRNA(Gln) by a tRNA-dependent amidotransferase (AdT). The three subunits GatA, GatB, and GatC are imported into mitochondria and assemble into a functional GatCAB AdT. Moreover, the mitochondrial pathway of Gln-tRNA(Gln) formation is shared with chloroplasts as both the GluRS, and the three AdT subunits are dual-imported into mitochondria and chloroplasts.}, note = {1091-6490 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Amino Acyl/*biosynthesis Solanum tuberosum/*enzymology, KERN Arabidopsis/*enzymology Cell Extracts Chloroplasts/*enzymology Cytosol/enzymology Glutamate-tRNA Ligase/metabolism Glutamine/*biosynthesis Mitochondria/*enzymology Nitrogenous Group Transferases/*metabolism Protein Subunits/metabolism Protein Transport RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {The GIR1 branching ribozymes}, author = {H Nielsen and B Beckert and B Masquida and S D Johansen}, editor = {D M J Lilley and F Eckstein}, url = {http://pubs.rsc.org/en/content/chapter/bk9780854042531-00229/978-0-85404-253-1}, isbn = {0.1039/9781847557988-00229}, year = {2008}, date = {2008-01-01}, booktitle = {Ribozymes and RNA catalysis}, pages = {229-252}, publisher = {Royal Society of Chemistry}, address = {Cambridge}, abstract = {The GIR1 branching ribozyme is a ca. 179 nt ribozyme with structural resemblance to group I ribozymes.1 It is found within a complex type of group I introns, termed the twin-ribozyme introns.2 Rather than splicing, it catalyses a branching reaction in which the 2′OH of an internal residue is involve.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {RNA switches regulate initiation of translation in bacteria}, author = {S Marzi and P Fechter and C Chevalier and P Romby and T Geissmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18953726}, isbn = {18953726}, year = {2008}, date = {2008-01-01}, journal = {Biol Chem}, volume = {389}, number = {5}, pages = {585-598}, abstract = {A large variety of RNA-based mechanisms have been uncovered in all living organisms to regulate gene expression in response to internal and external changes, and to rapidly adapt cell growth in response to these signals. In bacteria, structural elements in the 5' leader regions of mRNAs have direct effects on translation initiation of the downstream coding sequences. The docking and unfolding of these mRNAs on the 30S subunit are critical steps in the initiation process directly modulating and timing translation. Structural elements can also undergo conformational changes in response to environmental cues (i.e., temperature sensors) or upon binding of a variety of trans-acting factors, such as metabolites, non-coding RNAs or regulatory proteins. These RNA switches can temporally regulate translation, leading either to repression or to activation of protein synthesis.}, note = {1431-6730 (Print) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Animals Bacteria/*genetics Genes, Bacterial/genetics/*immunology RNA, Messenger/biosynthesis/genetics Ribosomes/immunology, ROMBY, Switch/genetics/*immunology Humans Peptide Chain Initiation, Translational/genetics/*immunology Protein Biosynthesis/genetics/*immunology RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {An Investigation of the Crystallogenesis of an Icosahedral RNA Plant Virus with Solubility Phase Diagrams}, author = {B Lorber and J Witz}, url = {http://pubs.acs.org/doi/abs/10.1021/cg700722b?prevSearch=[Contrib%3A+Lorber+B]&searchHistoryKey=}, isbn = {10.1021/cg700722b}, year = {2008}, date = {2008-01-01}, journal = {Crystal Growth & Design}, volume = {8}, number = {5}, pages = {1522-1529}, abstract = {The formation of crystals of the quasi-spherical tomato bushy stunt virus (TBSV, family Tombusviridae) was investigated to understand how identical cubic crystals with a dodecahedral habit grow in the presence of two chemically different precipitants, ammonium sulfate and polyethylene glycol (PEG). Two-dimensional solubility phase diagrams were established to identify the zones in which the virus crystallizes or precipitates. In the presence of the salt the solubility of the virus decreases when temperature increases. The reverse occurs in the presence of PEG. Virions stored during several years as crystals or as amorphous precipitates readily solubilize upon dilution and recrystallize under appropriate conditions like freshly purified ones. Light scattering measurements confirm that both crystallizing agents promote attractive interactions between viral particles. On a molar basis PEG-8000 is about 70-fold more effective than ammonium sulfate in insolubilizing TBSV. In the presence of each precipitant, heterogeneous nucleation takes places in a precipitate and crystal growth proceeds via an Ostwald ripening mechanism. The growth rate is controlled by the dissolution of the precipitate. Crystals with the largest dimensions always grow at apparently low supersaturation.}, keywords = {GIEGE FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Formation of two-dimensional crystals of icosahedral RNA viruses}, author = {B Lorber and M Adrian and J Witz and M Erhardt and J R Harris}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17466523}, isbn = {17466523}, year = {2008}, date = {2008-01-01}, journal = {Micron}, volume = {39}, number = {4}, pages = {431-446}, abstract = {The formation of 2D arrays of three small icosahedral RNA viruses with known 3D structures (tomato bushy stunt virus, turnip yellow mosaic virus and bromegrass mosaic virus) has been investigated to determine the role of each component of a negative staining solution containing ammonium molybdate and polyethylene glycol. Virion association was monitored by dynamic light scattering (DLS) and virus array formation was visualised by conventional transmission electron microscopy and cryo-electron microscopy after negative staining. The structural properties of viral arrays prepared in vitro were compared to those of microcrystals found in the leaves of infected plants. A novel form of macroscopic 3D crystals of turnip yellow mosaic virus has been grown in the negative staining solution. On the basis of the experimental results, the hypothesis is advanced that microscopic arrays might be planar crystallisation nuclei. The formation of 2D crystals and the enhancing effect of polyethylene glycol on the self-organisation of virions at the air/water interface are discussed. SYNOPSIS: The formation of 2D arrays of icosahedral viruses was investigated by spectroscopic and transmission electron microscopic methods.}, note = {0968-4328 (Print) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Electron Molybdenum/pharmacology Organometallic Compounds/pharmacology Polyethylene Glycols/pharmacology Scattering, GIEGE FLORENTZ Bromovirus/*ultrastructure Crystallization Light Lycopersicon esculentum/*virology Microscopy, Radiation Tombusvirus/*ultrastructure Tymovirus/*ultrastructure, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{violette_exploring_2008, title = {Exploring helical folding of oligoureas during chain elongation by high-resolution magic-angle-spinning (HRMAS) NMR spectroscopy}, author = {Aude Violette and Nathalie Lancelot and Alexander Poschalko and Martial Piotto and Jean-Paul Briand and Jesus Raya and Karim Elbayed and Alberto Bianco and Gilles Guichard}, doi = {10.1002/chem.200701923}, issn = {0947-6539}, year = {2008}, date = {2008-01-01}, journal = {Chemistry (Weinheim an Der Bergstrasse, Germany)}, volume = {14}, number = {13}, pages = {3874--3882}, abstract = {The development of novel folding oligomers (foldamers) for biological and biomedical applications requires both precise structural information and appropriate methods to detect folding propensity. However, the synthesis and the systematic conformational investigation of large arrays of oligomers to determine the influence of factors, such as chain length, side chains, and surrounding environment, on secondary structure can be quite tedious. Herein, we show for 2.5-helical N,N'-linked oligoureas (gamma-peptide lineage) that the whole process of foldamer characterization can be accelerated by using high-resolution magic-angle-spinning (HRMAS) NMR spectroscopy. This was achieved by monitoring a simple descriptor of conformational homogeneity (e.g., chemical shift difference between diastereotopic main chain CH2 protons) at different stages of oligourea chain growth on a solid support. HRMAS NMR experiments were conducted on two sets of oligoureas, ranging from dimer to hexamer, immobilized on DEUSS, a perdeuterated poly(oxyethylene)-based solid support swollen in solvents of low to high polarity. One evident advantage of the method is that only minute amount of material is required. In addition, the resonance of the deuterated resin is almost negligeable. On-bead NOESY spectra of high quality and with resolution comparable to that of liquid samples were obtained for longer oligomers, thus allowing detailed structural characterization.}, keywords = {I2CT, Magnetic Resonance Spectroscopy, Molecular Structure, Solvents, Team-Bianco, Urea}, pubstate = {published}, tppubtype = {article} } @article{, title = {Virus and Protein Crystallization under Hypergravity}, author = {B Lorber}, url = {http://pubs.acs.org/doi/abs/10.1021/cg800073t?prevSearch=%255BContrib%253A%2BLorber%2BB%255D&searchHistoryKey=}, year = {2008}, date = {2008-01-01}, journal = {Crystal Growth & Design}, volume = {8}, number = {8}, pages = {2964-2969}, abstract = {One small RNA plant virus and three monomeric and small molecular mass proteins (Mr = 14500−22200) were crystallized in a centrifuge at gravity levels between 1000 and 22000 g under conditions where controls at unit gravity are soluble. Solubility measurements indicate that all crystals have grown in solutions that are metastable, i.e., insufficiently supersaturated to nucleate under normal conditions. Upon centrifugation, particle sedimentation generates a concentration gradient. At highest local concentration, supersaturation is shifted beyond supersolubility and the critical driving force required for nucleation is overcome. A simple procedure has been implemented for sample microvolumes. The habit of protein crystals grown under hypergravity diverges from that of reference crystals but their unit cells are unchanged. Centrifugation is applicable to the crystallization of dilute samples of biological particles with a wide range of sizes, from large viruses to small proteins. It provides a means to control the onset of nucleation and in some cases to accelerate crystal growth.}, note = {10.1021/cg800073t}, keywords = {GIEGE FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{bianco_opportunities_2008, title = {Opportunities and challenges of carbon-based nanomaterials for cancer therapy}, author = {Alberto Bianco and Kostas Kostarelos and Maurizio Prato}, doi = {10.1517/17425247.5.3.331}, issn = {1742-5247}, year = {2008}, date = {2008-01-01}, journal = {Expert Opinion on Drug Delivery}, volume = {5}, number = {3}, pages = {331--342}, abstract = {The possibility of incorporating carbon-based nanomaterials into living systems has opened the way for the investigation of their potential applications in the emerging field of nanomedicine. A wide variety of different nanomaterials based on allotropic forms of carbon, such as nanotubes, nanohorns and nanodiamonds, are currently being explored towards different biomedical applications. In this review, we discuss the recent advances in the development of these novel nanomaterials for cancer therapy. A comparison between the characteristics, the advantages, the drawbacks, the benefits and the risks associated with these novel biocompatible forms of carbon is presented here.}, keywords = {Animals, carbon, Electron, Humans, I2CT, Microscopy, Nanomedicine, Nanostructures, Nanotubes, Neoplasms, Pharmaceutical, Team-Bianco, Technology, Transmission}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular basis for the role of selenium in muscle development and function}, author = {A Lescure and M Deniziak and M Rederstorff and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18357550}, isbn = {18357550}, year = {2008}, date = {2008-01-01}, journal = {Chem Biodivers}, volume = {5}, number = {3}, pages = {408-413}, note = {1612-1880 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {KROL, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {AKAP149 binds to HIV-1 reverse transcriptase and is involved in the reverse transcription}, author = {J Lemay and P Maidou-Peindara and R Cancio and E Ennifar and G Coadou and G Maga and J C Rain and R Benarous and L X Liu}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18786546}, isbn = {18786546}, year = {2008}, date = {2008-01-01}, journal = {J Mol Biol}, volume = {383}, number = {4}, pages = {783-796}, abstract = {Like all retroviruses, human immunodeficiency virus type 1 (HIV-1) undergoes reverse transcription during its replication cycle. The cellular cofactors potentially involved in this process still remain to be identified. We show here that A-kinase anchoring protein 149 (AKAP149) interacts with HIV-1 reverse transcriptase (RT) in both the yeast two-hybrid system and human cells. The AKAP149 binding site has been mapped to the RNase H domain of HIV-1 RT. AKAP149 silencing by RNA interference in HIV-1-infected cells inhibited viral replication at the reverse transcription step. We selected single-point mutants of RT defective for AKAP149 binding and demonstrated that mutant G462R, despite retaining significant intrinsic RT activity in vitro, failed to carry out HIV-1 reverse transcription correctly in infected cells. This suggests that the interaction between RT and AKAP149 in infected cells may play an important role in HIV-1 reverse transcription.}, note = {1089-8638 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {DUMAS A Kinase Anchor Proteins/chemistry/genetics/*metabolism Amino Acid Sequence Animals Binding Sites Cell Line HIV Reverse Transcriptase/chemistry/genetics/*metabolism HIV-1/enzymology/genetics Humans Models, ENNIFAR, Molecular Molecular Sequence Data Mutagenesis Protein Binding Protein Structure, Tertiary RNA Interference Recombinant Fusion Proteins/genetics/metabolism Reverse Transcription/*physiology Two-Hybrid System Techniques Virion/metabolism Virus Replication/physiology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {HuR interacts with human immunodeficiency virus type 1 reverse transcriptase, and modulates reverse transcription in infected cells}, author = {J Lemay and P Maidou-Peindara and T Bader and E Ennifar and J C Rain and R Benarous and L X Liu}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18544151 http://www.retrovirology.com/content/5/1/47}, isbn = {18544151}, year = {2008}, date = {2008-01-01}, journal = {Retrovirology}, volume = {5}, pages = {47}, abstract = {Reverse transcription of the genetic material of human immunodeficiency virus type 1 (HIV-1) is a critical step in the replication cycle of this virus. This process, catalyzed by reverse transcriptase (RT), is well characterized at the biochemical level. However, in infected cells, reverse transcription occurs in a multiprotein complex - the reverse transcription complex (RTC) - consisting of viral genomic RNA associated with viral proteins (including RT) and, presumably, as yet uncharacterized cellular proteins. Very little is known about the cellular proteins interacting with the RTC, and with reverse transcriptase in particular. We report here that HIV-1 reverse transcription is affected by the levels of a nucleocytoplasmic shuttling protein - the RNA-binding protein HuR. A direct protein-protein interaction between RT and HuR was observed in a yeast two-hybrid screen and confirmed in vitro by homogenous time-resolved fluorescence (HTRF). We mapped the domain interacting with HuR to the RNAse H domain of RT, and the binding domain for RT to the C-terminus of HuR, partially overlapping the third RRM RNA-binding domain of HuR. HuR silencing with specific siRNAs greatly impaired early and late steps of reverse transcription, significantly inhibiting HIV-1 infection. Moreover, by mutagenesis and immunoprecipitation studies, we could not detect the binding of HuR to the viral RNA. These results suggest that HuR may be involved in and may modulate the reverse transcription reaction of HIV-1, by an as yet unknown mechanism involving a protein-protein interaction with HIV-1 RT.}, note = {1742-4690 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {DUMAS Antigens, ENNIFAR, Small Interfering/genetics/metabolism RNA, Surface/*metabolism Binding Sites Cell Line Chromatin Immunoprecipitation Fluoroimmunoassay Gene Silencing HIV Reverse Transcriptase/*metabolism HIV-1/*physiology Humans Protein Interaction Domains and Motifs *Protein Interaction Mapping RNA, Unité ARN, Viral/metabolism RNA-Binding Proteins/*metabolism Two-Hybrid System Techniques *Virus Replication}, pubstate = {published}, tppubtype = {article} } @article{, title = {The bacterial and mitochondrial ribosomal A-site molecular switches possess different conformational substates}, author = {J Kondo and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18346970}, isbn = {18346970}, year = {2008}, date = {2008-01-01}, journal = {Nucleic Acids Res}, volume = {36}, number = {8}, pages = {2654-2666}, abstract = {The A site of the small ribosomal subunit participates in the fidelity of decoding by switching between two states, a resting 'off' state and an active decoding 'on' state. Eight crystal structures of RNA duplexes containing two minimal decoding A sites of the Homo sapiens mitochondrial wild-type, the A1555G mutant or bacteria have been solved. The resting 'off' state of the mitochondrial wild-type A site is surprisingly different from that of the bacterial A site. The mitochondrial A1555G mutant has two types of the 'off' states; one is similar to the mitochondrial wild-type 'off' state and the other is similar to the bacterial 'off' state. Our present results indicate that the dynamics of the A site in bacteria and mitochondria are different, a property probably related to the small number of tRNAs used for decoding in mitochondria. Based on these structures, we propose a hypothesis for the molecular mechanism of non-syndromic hearing loss due to the mitochondrial A1555G mutation.}, note = {1362-4962 (Electronic) Journal Article}, keywords = {Bacterial/*chemistry RNA, Crystallography, Messenger/chemistry RNA, Molecular Nucleic Acid Conformation Point Mutation RNA/*chemistry/genetics RNA, Ribosomal/*chemistry RNA, Transfer/chemistry, Unité ARN, WESTHOF, WESTHOF Crystallography, X-Ray Hearing Loss/genetics Humans Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Patterson-guided ab initio analysis of structures with helical symmetry}, author = {J Kondo and L Urzhumtseva and A Urzhumtsev}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18931415}, isbn = {18931415}, year = {2008}, date = {2008-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {64}, number = {Pt 10}, pages = {1078-1091}, abstract = {Patterson maps, which have a peak for intermolecular vectors between two molecules linked by a pseudo-translation, are widely used for structure solution. However, these maps may contain other peaks that indicate additional important information. In particular, if a molecule has internal symmetry, the Patterson maps may have a peak even when the relation between two molecules is other than a pure translation. A special and frequent case is a crystal that consists of molecules with pseudo-helical symmetry, like RNA or DNA, packed more or less parallel to each other. For such pairs of molecules, the Patterson peak does not simply link the molecular centres but is shifted along the helical axis. The shift is proportional to the rotation between the two molecules. The known step of the helix may be sufficient to obtain a system of linear equations whose solution gives an approximate position for the molecule. This technique may provide crucial information when molecular replacement fails to find a solution or suggests a number of candidates. Instead of repeating numerous molecular-replacement trials varying the model, a model may be positioned directly in place and be modified and refined directly. This Patterson-based search for molecular position has been tested with several solved crystals and has assisted in structure solution of RNA duplexes containing Homo sapiens cytoplasmic or mitochondrial ribosomal decoding sites (A sites).}, note = {1399-0047 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {DNA/chemistry Humans Models, Molecular *Nucleic Acid Conformation RNA/chemistry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The different role of high-affinity and low-affinity metal ions in cleavage by a tertiary stabilized cis hammerhead ribozyme from tobacco ringspot virus}, author = {N Kisseleva and A Khvorova and E Westhof and O Schiemann and A D Wolfson}, url = {http://online.liebertpub.com/doi/abs/10.1089/oli.2008.0129}, isbn = {10.1089/oli.2008.0129.}, year = {2008}, date = {2008-01-01}, journal = {Oligonucleotides}, volume = {18}, number = {2}, pages = {101-110}, abstract = {The aim of this study was to investigate the dependence of the observed cleavage rates (kobs) of a tertiary stabilized hammerhead ribozyme (tsHHRz) and of a minimal hammerhead ribozyme (mHHRz), both derived from tobacco ringspot virus, on the type and concentration of divalent metal ions in order to interpret the functional role of high-affinity ions detected by electron paramagnetic resonance (EPR). To measure the fast cleavage of the cis tsHHRz, a new method using chemically synthesized fluorescent-labeled RNAs has been developed. The tsHHRz cleavage rate is up to 20-fold faster than that of the mHHRz under similar conditions. The presence of Mn2+ ions leads to a 60-fold faster cleavage than in the presence of Mg2+ ions. The functional role of the high-affinity ion was evaluated using neomycin B inhibition studies. Neomycin B reduces the cleavage activity of both ribozymes but the inhibitory effect on tsHHRz is much weaker than that on the mHHRz. EPR data had shown that neomycin B displaces both low-affinity and high-affinity Mn2+ ions from the mHHRz, but only low-affinity ions from tsHHRz. Inhibition of the tsHHRz activity may be due to the displacement of weakly bound Me2+ ions required for the local folding leading to cleavage, whereas both the high-affinity ion required for folding and the weakly bound ions are replaced in the mHHRz. The high-affinity metal ion is required for the stabilization of the global HHRz structure, but is not involved in catalysis or stabilization of the transient state.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Milestones in RNA molecular dynamics simulations}, author = {Y Hashem and E Westhof and P Auffinger}, editor = {T Schwede and M C Peitsch}, url = {http://ebooks.worldscinet.com/ISBN/9789812778789/9789812778789_0013.html}, year = {2008}, date = {2008-01-01}, booktitle = {Computational structural biology}, volume = {13}, pages = {363-399}, publisher = {World Scientific Publishing Co}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Crystallogenesis trends of free and liganded aminoacyl-tRNA synthetases}, author = {R Giege and E Touze and B Lorber and A Théobald-Dietrich and C Sauter}, url = {http://pubs.acs.org/doi/abs/10.1021/cg8007766?prevSearch=%255BContrib%253A%2BSauter%2Bc%255D&searchHistoryKey=}, isbn = {10.1021/cg8007766}, year = {2008}, date = {2008-01-01}, journal = {Crystal Growth & Design}, volume = {8}, number = {12}, pages = {4297-4306}, abstract = {Aminoacyl-tRNA synthetases (aaRSs) catalyze the attachment of amino acids on transfer RNAs (tRNAs) and thus enable the correct expression of the genetic code during protein synthesis. After a brief historical review of early aaRS crystallizations we present results of a survey of the crystallization conditions of 220 aaRSs (either free or bound to small ligands) and of 59 aaRS:tRNA complexes whose structures are hosted by the RCSB Protein Data Bank. These structures at resolutions between 4.5 and 1.2 Å are those of synthetases from the three kingdoms of life. Their phylogenic distribution is heterogeneous: most stem from bacteria and archaea and some from eukarya and organelles. Interesting correlations are found between biochemical, physical−chemical, and crystallographic parameters. They highlight common and more specific features of the crystallogenesis of free or liganded aaRSs. The effects of the chemical nature of the crystallant(s), of ligands and other additives, as well as of the presence of impurities and of the presence of flexible polypeptide domains on protein or protein/RNA crystallization are examined. Features that favor the growth of crystals diffracting X-rays at high resolution are discussed.}, keywords = {FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Toward a more complete view of tRNA biology}, author = {R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18836497}, isbn = {18836497}, year = {2008}, date = {2008-01-01}, journal = {Nat Struct Mol Biol}, volume = {15}, number = {10}, pages = {1007-1014}, abstract = {Transfer RNAs are ancient molecules present in all domains of life. In addition to translating the genetic code into protein and defining the second genetic code together with aminoacyl-tRNA synthetases, tRNAs act in many other cellular functions. Robust phenomenological observations on the role of tRNAs in translation, together with massive sequence and crystallographic data, have led to a deeper physicochemical understanding of tRNA architecture, dynamics and identity. In vitro studies complemented by cell biology data already indicate how tRNA behaves in cellular environments, in particular in higher Eukarya. From an opposite approach, reverse evolution considerations suggest how tRNAs emerged as simplified structures from the RNA world. This perspective discusses what basic questions remain unanswered, how these answers can be obtained and how a more rational understanding of the function and dysfunction of tRNA can have applications in medicine and biotechnology.}, note = {1545-9985 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {GIEGE Animals Biomedical Technology Biotechnology Evolution Humans Nucleic Acid Conformation Nucleotides/metabolism RNA, Transfer/*chemistry/genetics/*metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comprehensive features of natural and in vitro selected GNRA tetraloop-binding receptors}, author = {C Geary and S Baudrey and L Jaeger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18158305}, isbn = {18158305}, year = {2008}, date = {2008-01-01}, journal = {Nucleic Acids Res}, volume = {36}, number = {4}, pages = {1138-1152}, abstract = {Specific recognitions of GNRA tetraloops by small helical receptors are among the most widespread long-range packing interactions in large ribozymes. However, in contrast to GYRA and GAAA tetraloops, very few GNRA/receptor interactions have yet been identified to involve GGAA tetraloops in nature. A novel in vitro selection scheme based on a rigid self-assembling tectoRNA scaffold designed for isolation of intermolecular interactions with A-minor motifs has yielded new GGAA tetraloop-binding receptors with affinity in the nanomolar range. One of the selected receptors is a novel 12 nt RNA motif, (CCUGUG. AUCUGG), that recognizes GGAA tetraloop hairpin with a remarkable specificity and affinity. Its physical and chemical characteristics are comparable to those of the well-studied '11nt' GAAA tetraloop receptor motif. A second less specific motif (CCCAGCCC. GAUAGGG) binds GGRA tetraloops and appears to be related to group IC3 tetraloop receptors. Mutational, thermodynamic and comparative structural analysis suggests that natural and in vitro selected GNRA receptors can essentially be grouped in two major classes of GNRA binders. New insights about the evolution, recognition and structural modularity of GNRA and A-minor RNA-RNA interactions are proposed.}, note = {1362-4962 (Electronic) Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S.}, keywords = {KROL Adenine/chemistry Base Sequence Dimerization Directed Molecular Evolution Models, Molecular Molecular Sequence Data Nucleic Acid Conformation RNA/*chemistry/classification Sequence Analysis, RNA Thermodynamics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding of aminoglycoside antibiotics to the duplex form of the HIV-1 genomic RNA dimerization initiation site}, author = {S Freisz and K Lang and R Micura and P Dumas and E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18435520}, isbn = {18435520}, year = {2008}, date = {2008-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {47}, number = {22}, pages = {4110-4113}, note = {1521-3773 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {DUMAS Aminoglycosides/*chemistry/pharmacology Anti-Bacterial Agents/*chemistry/pharmacology Base Sequence Dimerization Genome, ENNIFAR, Unité ARN, Viral HIV-1/*drug effects Humans *Nucleic Acid Conformation RNA, Viral/*chemistry Virus Replication/*drug effects}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Footprinting}, author = {P Fechter and P Romby}, editor = {S Muller}, url = {http://books.google.fr/books?id=4xOLLuqKrFgC&pg=PA13&hl=fr&source=gbs_toc_r&cad=4#v=onepage&q&f=false}, doi = {10.1002/9783527622528.ch2}, year = {2008}, date = {2008-01-01}, booktitle = {Nucleic Acids from A to Z: a concise encyclopedia}, pages = {109}, publisher = {WILEY-VCH Verlag GmbH & Co}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Chemical and enzymatic structure mapping of RNAs}, author = {P Fechter and P Romby}, editor = {S Muller}, url = {http://books.google.fr/books?id=4xOLLuqKrFgC&printsec=frontcover&dq=inauthor:%22Sabine+M%C3%BCller%22&hl=fr&sa=X&ei=K2KNT-CQFtDf8QO9qaDJCw&ved=0CDcQ6AEwAQ#v=onepage&q&f=false}, year = {2008}, date = {2008-01-01}, booktitle = {Nucleic Acids from A to Z: a concise encyclopedia}, pages = {41-43}, publisher = {WILEY-VCH Verlag GmbH & Co}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Staphylococcus aureus endoribonuclease III purification and properties}, author = {C Chevalier and E Huntzinger and P Fechter and S Boisset and F Vandenesch and P Romby and T Geissmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=19161850}, isbn = {19161850}, year = {2008}, date = {2008-01-01}, journal = {Methods Enzymol}, volume = {447}, pages = {309-327}, abstract = {Staphylococcus aureus ribonuclease III (Sa-RNase III) belongs to the enzyme family known to process double-stranded RNAs consisting of two turns of the RNA helix. Although the enzyme is thought to play a role in ribosomal RNA processing and gene regulation, the deletion of the rnc gene in S. aureus does not affect cell growth in rich medium. S. aureus RNase III acts in concert with regulatory RNAIII to repress the expression of several mRNAs encoding virulence factors. The action of the RNase is most likely to initiate the degradation of repressed mRNAs leading to an irreversible repression. In this chapter, we describe the overexpression and purification of recombinant RNase III from S. aureus, and we show that its biochemical properties are similar to the orthologous enzyme from Escherichia coli. Both enzymes similarly recognize and cleave different RNA substrates and RNA-mRNA duplexes.}, note = {1557-7988 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Hsp90 chaperone controls the biogenesis of L7Ae RNPs through conserved machinery}, author = {S Boulon and N Marmier-Gourrier and B Pradet-Balade and L Wurth and C Verheggen and B E Jady and B Rothé and C Pescia and M C Robert and T Kiss and B Bardoni and A Krol and C Branlant and C Allmang and E Bertrand and B Charpentier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18268104}, issn = {0021-9525}, year = {2008}, date = {2008-01-01}, journal = {J Cell Biol}, volume = {180}, number = {3}, pages = {579-595}, abstract = {RNA-binding proteins of the L7Ae family are at the heart of many essential ribonucleoproteins (RNPs), including box C/D and H/ACA small nucleolar RNPs, U4 small nuclear RNP, telomerase, and messenger RNPs coding for selenoproteins. In this study, we show that Nufip and its yeast homologue Rsa1 are key components of the machinery that assembles these RNPs. We observed that Rsa1 and Nufip bind several L7Ae proteins and tether them to other core proteins in the immature particles. Surprisingly, Rsa1 and Nufip also link assembling RNPs with the AAA + adenosine triphosphatases hRvb1 and hRvb2 and with the Hsp90 chaperone through two conserved adaptors, Tah1/hSpagh and Pih1. Inhibition of Hsp90 in human cells prevents the accumulation of U3, U4, and telomerase RNAs and decreases the levels of newly synthesized hNop58, hNHP2, 15.5K, and SBP2. Thus, Hsp90 may control the folding of these proteins during the formation of new RNPs. This suggests that Hsp90 functions as a master regulator of cell proliferation by allowing simultaneous control of cell signaling and cell growth.}, note = {1540-8140 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {ERIANI, KROL Adenosine Triphosphatases/genetics/metabolism Cell Cycle Proteins/genetics/metabolism Cell Proliferation Conserved Sequence/genetics DNA Helicases/genetics/metabolism DNA-Binding Proteins/genetics/metabolism Evolution, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Decreased aminoacylation in pathology-related mutants of mitochondrial tRNATyr is associated with structural perturbations in tRNA architecture}, author = {L Bonnefond and C Florentz and R Giege and J Rudinger-Thirion}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18268021}, isbn = {18268021}, year = {2008}, date = {2008-01-01}, journal = {RNA}, volume = {14}, number = {4}, pages = {641-648}, abstract = {A growing number of human pathologies are ascribed to mutations in mitochondrial tRNA genes. Here, we report biochemical investigations on three mt-tRNA(Tyr) molecules with point substitutions associated with diseases. The mutations occur in the atypical T- and D-loops at positions homologous to those involved in the tertiary interaction network of canonical tRNAs. They do not correspond to tyrosine identity positions and likely do not contact the mitochondrial tyrosyl-tRNA synthetase during the aminoacylation process. The impact of these substitutions on mt-tRNA(Tyr) tyrosylation and structure was investigated using the corresponding tRNA transcripts. In vitro tyrosylation efficiency is decreased 600-fold for mutant A22G (mitochondrial gene mutation T5874C), 40-fold for G15A (C5877T), and is without significant effect on U54C (A5843G). Comparative solution probings with lead and nucleases on mutant and wild-type tRNA(Tyr) molecules reveal a greater sensitivity to single-strand specific probes for mutants G15A and A22G. For both transcripts, the mutation triggers a structural destabilization in the D-loop that propagates toward the anticodon arm and thus hinders efficient tyrosylation. Further probing analysis combined with phylogenetic data support the participation of G15 and A22 in the tertiary network of human mt-tRNA(Tyr) via nonclassical Watson-Crick G15-C48 and G13-A22 pairings. In contrast, the pathogenic effect of the tyrosylable mutant U54C, where structure is only marginally affected, has to be sought at another level of the tRNA(Tyr) life cycle.}, note = {1469-9001 (Electronic) In Vitro Journal Article Research Support, Non-U.S. Gov't}, keywords = {Base Sequence Humans Mitochondrial Diseases/genetics/metabolism Models, FLORENTZ, FRUGIER, Molecular Molecular Sequence Data Nucleic Acid Conformation *Point Mutation RNA/*chemistry/*genetics/metabolism RNA Stability RNA, Transfer, Tyr/*chemistry/*genetics/metabolism *Transfer RNA Aminoacylation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of glutamyl-queuosine tRNAAsp synthetase complexed with L-glutamate: structural elements mediating tRNA-independent activation of glutamate and glutamylation of tRNAAsp anticodon}, author = {M Blaise and V Olieric and C Sauter and B Lorber and B Roy and S Karmakar and R Banerjee and H D Becker and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18602926}, isbn = {18602926}, year = {2008}, date = {2008-01-01}, journal = {J Mol Biol}, volume = {381}, number = {5}, pages = {1224-1237}, abstract = {Glutamyl-queuosine tRNA(Asp) synthetase (Glu-Q-RS) from Escherichia coli is a paralog of the catalytic core of glutamyl-tRNA synthetase (GluRS) that catalyzes glutamylation of queuosine in the wobble position of tRNA(Asp). Despite important structural similarities, Glu-Q-RS and GluRS diverge strongly by their functional properties. The only feature common to both enzymes consists in the activation of Glu to form Glu-AMP, the intermediate of transfer RNA (tRNA) aminoacylation. However, both enzymes differ by the mechanism of selection of the cognate amino acid and by the mechanism of its activation. Whereas GluRS selects l-Glu and activates it only in the presence of the cognate tRNA(Glu), Glu-Q-RS forms Glu-AMP in the absence of tRNA. Moreover, while GluRS transfers the activated Glu to the 3' accepting end of the cognate tRNA(Glu), Glu-Q-RS transfers the activated Glu to Q34 located in the anticodon loop of the noncognate tRNA(Asp). In order to gain insight into the structural elements leading to distinct mechanisms of amino acid activation, we solved the three-dimensional structure of Glu-Q-RS complexed to Glu and compared it to the structure of the GluRS.Glu complex. Comparison of the catalytic site of Glu-Q-RS with that of GluRS, combined with binding experiments of amino acids, shows that a restricted number of residues determine distinct catalytic properties of amino acid recognition and activation by the two enzymes. Furthermore, to explore the structural basis of the distinct aminoacylation properties of the two enzymes and to understand why Glu-Q-RS glutamylates only tRNA(Asp) among the tRNAs possessing queuosine in position 34, we performed a tRNA mutational analysis to search for the elements of tRNA(Asp) that determine recognition by Glu-Q-RS. The analyses made on tRNA(Asp) and tRNA(Asn) show that the presence of a C in position 38 is crucial for glutamylation of Q34. The results are discussed in the context of the evolution and adaptation of the tRNA glutamylation system.}, note = {1089-8638 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Adenosine Triphosphate/metabolism Amino Acid Sequence Amino Acyl-tRNA Synthetases/*chemistry Anticodon/*metabolism Base Sequence Binding Sites Catalysis Conserved Sequence Crystallography, Asp/*chemistry/genetics Regulatory Sequences, FFRUGIER, Molecular Molecular Sequence Data Nucleic Acid Conformation Nucleoside Q/*chemistry Protein Structure, Ribonucleic Acid/*genetics Thermus thermophilus/enzymology, SAUTER, Secondary RNA, Transfer, Unité ARN, X-Ray Escherichia coli/*enzymology Escherichia coli Proteins/*chemistry Glutamic Acid/*chemistry Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Distinctive structures between chimpanzee and human in a brain noncoding RNA}, author = {A Beniaminov and E Westhof and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18511501}, isbn = {18511501}, year = {2008}, date = {2008-01-01}, journal = {RNA}, volume = {14}, number = {7}, pages = {1270-1275}, abstract = {Human accelerated region 1 (HAR1) is a short DNA region identified recently to have evolved the most rapidly among highly constrained regions since the divergence from our common ancestor with chimpanzee. It is transcribed as part of a noncoding RNA specifically expressed in the developing human neocortex. Employing a panoply of enzymatic and chemical probes, our analysis of HAR1 RNA proposed a secondary structure model differing from that published. Most surprisingly, we discovered that the substitutions between the chimpanzee and human sequences led the human HAR1 RNA to adopt a cloverleaf-like structure instead of an extended and unstable hairpin in the chimpanzee sequence. Thus, the rapid evolutionary changes resulted in a profound rearrangement of HAR1 RNA structure. Altogether, our results provide a structural context for elucidating HAR1 RNA function.}, note = {1469-9001 (Electronic) Letter Research Support, Non-U.S. Gov't}, keywords = {Animals Base Sequence Brain/*metabolism Evolution, Human Humans Molecular Sequence Data Nucleic Acid Conformation Pan troglodytes/*genetics RNA, Molecular Genome, Unité ARN, Untranslated/*chemistry, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular modelling of the GIR1 branching ribozyme gives new insight into evolution of structurally related ribozymes}, author = {B Beckert and H Nielsen and C Einvik and S D Johansen and E Westhof and B Masquida}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18219270}, isbn = {18219270}, year = {2008}, date = {2008-01-01}, journal = {EMBO J}, volume = {27}, number = {4}, pages = {667-678}, abstract = {Twin-ribozyme introns contain a branching ribozyme (GIR1) followed by a homing endonuclease (HE) encoding sequence embedded in a peripheral domain of a group I splicing ribozyme (GIR2). GIR1 catalyses the formation of a lariat with 3 nt in the loop, which caps the HE mRNA. GIR1 is structurally related to group I ribozymes raising the question about how two closely related ribozymes can carry out very different reactions. Modelling of GIR1 based on new biochemical and mutational data shows an extended substrate domain containing a GoU pair distinct from the nucleophilic residue that dock onto a catalytic core showing a different topology from that of group I ribozymes. The differences include a core J8/7 region that has been reduced and is complemented by residues from the pre-lariat fold. These findings provide the basis for an evolutionary mechanism that accounts for the change from group I splicing ribozyme to the branching GIR1 architecture. Such an evolutionary mechanism can be applied to other large RNAs such as the ribonuclease P.}, note = {1460-2075 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Catalytic/*chemistry, Evolution, Molecular *Models, Molecular RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Advances in the structural understanding of Vif proteins}, author = {P Barraud and J C Paillart and R Marquet and C Tisne}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18336256}, isbn = {18336256}, year = {2008}, date = {2008-01-01}, journal = {Curr HIV Res}, volume = {6}, number = {2}, pages = {91-99}, abstract = {The multidomain HIV-1 Vif protein recruits several cellular partners to achieve neutralization of the antiviral activity of APOBEC3 proteins. Vif neutralizes APOBEC3G and APOBEC3F predominantly by forming an E3 ubiquitin ligase with Cullin5, ElonginB and ElonginC that targets these proteins for degradation by the ubiquitin-proteasome pathway. Vif associates with the Cullin5-ElonginB-ElonginC complex by binding directly to ElonginC via its SOCS-box motif and to Cullin5 via hydrophobic residues within a zinc-binding region formed by a conserved HCCH motif. The HIV-1 Vif-Cullin5-ElonginBC complex is then able to ubiquitinate the APOBEC3G factor bound to Vif by its N-terminal domain. In this review, we summarize the current knowledge about the structural determinants of Vif that allow it to interact with cellular and viral partners.}, note = {1873-4251 (Electronic) Journal Article Research Support, Non-U.S. Gov't Review}, keywords = {Amino Acid Motifs Gene Products, MARQUET, PAILLART, Tertiary Simian immunodeficiency virus/*genetics, Unité ARN, vif/*genetics/*metabolism HIV-1/*genetics HIV-2/*genetics Humans Protein Binding Protein Structure}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNA-dependent asparagine formation in prokaryotes: Characterization, isolation and structural and functional analysis of a ribonucleoprotein particle generating Asn-tRNA(Asn)}, author = {M Bailly and M Blaise and H Roy and M Deniziak and B Lorber and C Birck and H D Becker and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18241796}, isbn = {18241796}, year = {2008}, date = {2008-01-01}, journal = {Methods}, volume = {44}, number = {2}, pages = {146-163}, abstract = {In some living organisms the 20 aa-tRNA species participating in protein synthesis are not charged by a complete set of 20 aminoacyl-tRNA synthetases. In prokaryotes, the deficiency of asparaginyl- and/or glutaminyl-tRNA synthetases is compensated by another aminoacyl-tRNA synthetase of relaxed specificity that mischarges the orphan tRNA and by an enzyme that converts the amino acid into that homologous to the tRNA. In Thermus thermophilus Asn-tRNA(Asn) is formed indirectly via a two-step pathway whereby tRNA(Asn) is mischarged with Asp that will subsequently be amidated into Asn by an amidotransferase. The non-discriminating aspartyl-tRNA synthetase, the trimeric GatCAB tRNA-dependent amidotransferase and the tRNA(Asn) promoting this pathway assemble into a ribonucleoprotein particle termed transamidosome. This article deals with the methods and techniques employed to clone the genes encoding the enzymes and the tRNA involved in this pathway, to express them in Escherichia coli, to isolate them on a large scale, and to transcribe and produce mg quantities of pure tRNA(Asn)in vitro. The approaches designed especially for this system include (i) clustering of the ORFs encoding the subunits of the heterotrimeric GatCAB that are sprinkled in the genome into an artificial operon, and (ii) the self-cleavage of the tRNA(Asn) transcript starting with U in 5' position through fusion with a hammerhead ribozyme. Further, the crystallization of the free enzymes is described and the characterization of their assembly with tRNA(Asn) into a ribonucleoprotein particle, as well as the investigation of the catalytic mechanism of Asn-tRNA(Asn) formation by the complex are reported.}, note = {1046-2023 (Print) Journal article}, keywords = {KERN GIEGE FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identifying the important HIV-1 recombination breakpoints}, author = {J Archer and J W Pinney and J Fan and E Simon-Loriere and E J Arts and M Negroni and D L Robertson}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18787691}, isbn = {18787691}, year = {2008}, date = {2008-01-01}, journal = {PLoS Comput Biol}, volume = {4}, number = {9}, pages = {e1000178}, abstract = {Recombinant HIV-1 genomes contribute significantly to the diversity of variants within the HIV/AIDS pandemic. It is assumed that some of these mosaic genomes may have novel properties that have led to their prevalence, particularly in the case of the circulating recombinant forms (CRFs). In regions of the HIV-1 genome where recombination has a tendency to convey a selective advantage to the virus, we predict that the distribution of breakpoints--the identifiable boundaries that delimit the mosaic structure--will deviate from the underlying null distribution. To test this hypothesis, we generate a probabilistic model of HIV-1 copy-choice recombination and compare the predicted breakpoint distribution to the distribution from the HIV/AIDS pandemic. Across much of the HIV-1 genome, we find that the observed frequencies of inter-subtype recombination are predicted accurately by our model. This observation strongly indicates that in these regions a probabilistic model, dependent on local sequence identity, is sufficient to explain breakpoint locations. In regions where there is a significant over- (either side of the env gene) or under- (short regions within gag, pol, and most of env) representation of breakpoints, we infer natural selection to be influencing the recombination pattern. The paucity of recombination breakpoints within most of the envelope gene indicates that recombinants generated in this region are less likely to be successful. The breakpoints at a higher frequency than predicted by our model are approximately at either side of env, indicating increased selection for these recombinants as a consequence of this region, or at least part of it, having a tendency to be recombined as an entire unit. Our findings thus provide the first clear indication of the existence of a specific portion of the genome that deviates from a probabilistic null model for recombination. This suggests that, despite the wide diversity of recombinant forms seen in the viral population, only a minority of recombination events appear to be of significance to the evolution of HIV-1.}, note = {1553-7358 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Computational Biology Evolution, env Genetic Variation Genome, Genetic, Genetic Models, Molecular Genes, NEGRONI, Statistical RNA, Unité ARN, Viral HIV Infections/virology HIV-1/*genetics Humans *Models, Viral/genetics *Recombination}, pubstate = {published}, tppubtype = {article} } @article{lacerda_cell-penetrating_2007, title = {Cell-penetrating CNTs for delivery of therapeutics}, author = {Lara Lacerda and Simona Raffa and Maurizio Prato and Alberto Bianco and Kostas Kostarelos}, url = {http://www.sciencedirect.com/science/article/pii/S174801320770172X}, doi = {10.1016/S1748-0132(07)70172-X}, issn = {1748-0132}, year = {2007}, date = {2007-12-01}, urldate = {2020-03-31}, journal = {Nano Today}, volume = {2}, number = {6}, pages = {38--43}, abstract = {The use of carbon-based nanostructures, such as carbon nanotubes, in biomedicine is increasingly attracting attention. One key advantage of carbon nanotubes is their ability to translocate through plasma membranes, allowing their use for the delivery of therapeutically active molecules in a manner that resembles cell-penetrating peptides. Moreover, exploitation of their unique electrical, optical, thermal, and spectroscopic properties in a biological context is hoped to yield great advances in the detection, monitoring, and therapy of disease. Here we offer a speculative overview of the general principles behind the mechanism of carbon nanotube penetration of the plasma membrane and a snapshot of the different therapeutic modalities based on these fascinating nanostructures that are currently being investigated.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{lacerda_carbon_2007, title = {Carbon nanotube cell translocation and delivery of nucleic acidsin vitro and in vivo}, author = {Lara Lacerda and Alberto Bianco and Maurizio Prato and Kostas Kostarelos}, url = {https://pubs.rsc.org/en/content/articlelanding/2008/jm/b711554g}, doi = {10.1039/B711554G}, issn = {1364-5501}, year = {2007}, date = {2007-12-01}, urldate = {2020-03-31}, journal = {Journal of Materials Chemistry}, volume = {18}, number = {1}, pages = {17--22}, abstract = {In the last few years, the carbon nanotube (CNT) field has seen a new direction of investigation growing rapidly, along with the interest of more researchers from diverse fields of expertise interested in this new material in an attempt to exploit their properties in biomedical applications. Here we describe the most recently reported work on the application of CNT for gene encoding nucleic acid (DNA and RNA) delivery purposes by using in vitro and in vivo models. Several groups have now successfully observed the cellular internalisation of nucleic acids with the aid of CNT following very different protocols. The main processes for the internalisation pathways and intracellular release of the nucleic acids are here reviewed. Furthermore, we have just started to see some initial studies of in vivo work using siRNA-CNT conjugates to achieve silencing in tumour tissue. Admittedly, it is still very early days for the technology, but future studies are necessary, and will surely appear, in order to determine the feasibility of bringing the CNT closer to the clinic.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_drosophila_2007b, title = {The Drosophila systemic immune response: sensing and signalling during bacterial and fungal infections}, author = {Dominique Ferrandon and Jean-Luc Imler and Charles Hetru and Jules A Hoffmann}, year = {2007}, date = {2007-11-01}, journal = {Nat Rev Immunol}, volume = {7}, pages = {862--74}, abstract = {A hallmark of the potent, multifaceted antimicrobial defence of Drosophila melanogaster is the challenge-induced synthesis of several families of antimicrobial peptides by cells in the fat body. The basic mechanisms of recognition of various types of microbial infections by the adult fly are now understood, often in great detail. We have further gained valuable insight into the infection-induced gene reprogramming by nuclear factor-kappaB (NF-kappaB) family members under the dependence of complex intracellular signalling cascades. The striking parallels between the adult fly response and mammalian innate immune defences described below point to a common ancestry and validate the relevance of the fly defence as a paradigm for innate immunity.}, keywords = {Animals, Bacterial Infections/*immunology/microbiology, Drosophila melanogaster/genetics/*immunology/microbiology, ferrandon, hoffmann, imler, Immunity, M3i, Mycoses/*immunology/microbiology, Natural/genetics, Signal Transduction/genetics/*immunology}, pubstate = {published}, tppubtype = {article} } @article{marmagne_high_2007, title = {A high content in lipid-modified peripheral proteins and integral receptor kinases features in the arabidopsis plasma membrane proteome.}, author = {Anne Marmagne and Myriam Ferro and Thierry Meinnel and Christophe Bruley and Lauriane Kuhn and Jérome Garin and Hélène Barbier-Brygoo and Geneviève Ephritikhine}, doi = {10.1074/mcp.M700099-MCP200}, issn = {1535-9476 1535-9476}, year = {2007}, date = {2007-11-01}, journal = {Molecular & cellular proteomics : MCP}, volume = {6}, number = {11}, pages = {1980--1996}, abstract = {The proteomics of plasma membrane has brought to date only scarce and partial information on the actual protein repertoire. In this work, the plant plasma membrane proteome of Arabidopsis thaliana was investigated. A highly purified plasma membrane fraction was washed by NaCl and Na2CO3 salts, and the insoluble fractions were further analyzed by nano-LC-MS/MS. With 446 proteins identified, we hereby describe the largest plasma membrane proteome diversity reported so far. Half of the proteins were predicted to display transmembrane domains and/or to be anchored to the membrane, validating a posteriori the pertinence of the approach. A fine analysis highlighted two main specific and novel features. First, the main functional category is represented by a majority of as yet unreported signaling proteins, including 11% receptor-like kinases. Second, 16% of the identified proteins are predicted to be lipid-modified, specifically involving double lipid linkage through N-terminal myristoylation, S-palmitoylation, C-terminal prenylation, or glycosylphosphatidylinositol anchors. Thus, our approach led for the first time to the identification of a large number of peripheral proteins as part of the plasma membrane and allowed the functionality of the plasma membrane in the cell context to be reconsidered.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{beutler_genetic_2007, title = {Genetic analysis of resistance to viral infection}, author = {Bruce Beutler and Celine Eidenschenk and Karine Crozat and Jean-Luc Imler and Osamu Takeuchi and Jules A Hoffmann and Shizuo Akira}, doi = {10.1038/nri2174}, issn = {1474-1741}, year = {2007}, date = {2007-10-01}, journal = {Nature Reviews. Immunology}, volume = {7}, number = {10}, pages = {753--766}, abstract = {As machines that reprogramme eukaryotic cells to suit their own purposes, viruses present a difficult problem for multicellular hosts, and indeed, have become one of the central pre-occupations of the immune system. Unable to permanently outpace individual viruses in an evolutionary footrace, higher eukaryotes have evolved broadly active mechanisms with which to sense viruses and suppress their proliferation. These mechanisms have recently been elucidated by a combination of forward and reverse genetic methods. Some of these mechanisms are clearly ancient, whereas others are relatively new. All are remarkably adept at discriminating self from non-self, and allow the host to cope with what might seem an impossible predicament.}, keywords = {Animals, Antiviral Agents, Disease Susceptibility, Drug Resistance, Eukaryotic Cells, hoffmann, Humans, imler, Immunity, M3i, Mutation, Viral, Virus Diseases, viruses}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_ubiquitin-proteasome:_2007b, title = {Ubiquitin-proteasome: pallbearer carries the deceased to the grave}, author = {Dominique Ferrandon}, doi = {10.1016/j.immuni.2007.10.003}, issn = {1074-7613}, year = {2007}, date = {2007-10-01}, journal = {Immunity}, volume = {27}, number = {4}, pages = {541--544}, abstract = {Phagocytosis is a complex process that involves multiple cellular functions. In this issue of Immunity, Silva et al. (2007) report that a protein ubiquitylation complex and the proteasome are required for the clearance of apoptotic cells in Drosophila.}, keywords = {*Models, Animals, Apoptosis, Apoptosis/*physiology, ferrandon, Immunological, M3i, Macrophages, Macrophages/immunology/metabolism, Models, Phagocytosis, Phagocytosis/*physiology, Proteasome Endopeptidase Complex, Proteasome Endopeptidase Complex/*metabolism, ubiquitin, Ubiquitin/*metabolism}, pubstate = {published}, tppubtype = {article} } @article{SA2007, title = {Phagocytosis in mosquito immune responses}, author = {Stéphanie A Blandin and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17850478}, year = {2007}, date = {2007-10-01}, journal = {Immunol Rev.}, volume = {219}, pages = {8-16}, abstract = {Anopheles mosquitoes are the only vectors of human malaria parasites. Mosquito-parasite interactions are critical for disease transmission and therefore are a potential target for malaria control strategies. Mosquitoes mount potent immune responses that efficiently limit proliferation of a variety of infectious agents, including microbial pathogens and malaria parasites. The recent completion of the Anopheles gambiae genome sequencing project combined with the development of the powerful RNA interference-based gene silencing helped to identify major players of the immune defenses and uncovered evolutionarily conserved mechanisms in the anti-bacterial and anti-Plasmodium responses. The anti-bacterial responses are based on phagocytosis at early steps of infections, followed, several hours later, by the synthesis of anti-microbial peptides. The principal regulators of anti-parasitic responses are predominantly synthesized by the mosquito blood cells; however, the exact molecular mechanisms of parasite killing remain unclear. Several regulators of phagocytosis are also required for efficient parasite killing. Here, we summarize our current knowledge of the anti-bacterial and anti-parasitic responses, with the particular emphasis on the role of phagocytosis in mosquito immunity.}, keywords = {blandin, M3i, Phagocytosis, RNAi}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_[new_2007b, title = {[New mechanism for detection of infections using the innate immune system of animals]}, author = {Dominique Ferrandon and Marie Gottar and Vanessa Gobert}, doi = {10.1051/medsci/20072389707}, issn = {0767-0974}, year = {2007}, date = {2007-09-01}, journal = {Med Sci (Paris)}, volume = {23}, number = {8-9}, pages = {707--709}, keywords = {Animal, Animals, Drosophila/immunology, ferrandon, Gram-Positive Bacteria, Gram-Positive Bacteria/pathogenicity, Gram-Positive Bacterial Infections, Gram-Positive Bacterial Infections/immunology, Humans, Immune System, infection, Infection/*diagnosis/*immunology, M3i, Models}, pubstate = {published}, tppubtype = {article} } @article{RH2007, title = {Structural basis for conserved complement factor-like function in the antimalarial protein TEP1}, author = {Richard H Baxter and C I Chang and Y Chelliah and Stéphanie A Blandin and Elena A Levashina and J Deisenhofer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17606907}, year = {2007}, date = {2007-07-10}, journal = {Proc Natl Acad Sci U S A.}, volume = {104}, number = {28}, pages = {11615-20}, abstract = {Thioester-containing proteins (TEPs) are a major component of the innate immune response of insects to invasion by bacteria and protozoa. TEPs form a distinct clade of a superfamily that includes the pan-protease inhibitors alpha(2)-macroglobulins and vertebrate complement factors. The essential feature of these proteins is a sequestered thioester bond that, after cleavage in a protease-sensitive region of the protein, is activated and covalently binds to its target. Recently, TEP1 from the malarial vector Anopheles gambiae was shown to mediate recognition and killing of ookinetes from the malarial parasite Plasmodium berghei, a model for the human malarial parasite Plasmodium falciparum. Here, we present the crystal structure of the TEP1 isoform TEP1r. Although the overall protein fold of TEP1r resembles that of complement factor C3, the TEP1r domains are repositioned to stabilize the inactive conformation of the molecule (containing an intact thioester) in the absence of the anaphylotoxin domain, a central component of complement factors. The structure of TEP1r provides a molecular basis for the differences between TEP1 alleles TEP1r and TEP1s, which correlate with resistance of A. gambiae to infection by P. berghei.}, keywords = {blandin, M3i, TEP1}, pubstate = {published}, tppubtype = {article} } @article{muller_dicing_2007, title = {Dicing with viruses: microRNAs as antiviral factors}, author = {Stefanie Müller and Jean-Luc Imler}, doi = {10.1016/j.immuni.2007.07.003}, issn = {1074-7613}, year = {2007}, date = {2007-07-01}, journal = {Immunity}, volume = {27}, number = {1}, pages = {1--3}, abstract = {In plants and invertebrates, Dicer genes play a critical role against infections by RNA viruses. In this issue, Otsuka et al. (2007) report that Dicer mutant mice are hypersusceptible to infection by the RNA virus VSV.}, keywords = {Animals, DEAD-box RNA Helicases, Endoribonucleases, imler, M3i, MicroRNAs, Ribonuclease III, RNA Interference, RNA Virus Infections}, pubstate = {published}, tppubtype = {article} } @article{kwan_lps_2007, title = {LPS induces rapid IL-10 release by M-CSF-conditioned tolerogenic dendritic cell precursors}, author = {W H Kwan and C Boix and N Gougelet and W H Fridman and C G Mueller}, year = {2007}, date = {2007-07-01}, journal = {Journal of Leukocyte Biology}, volume = {82}, number = {0741-5400 (Print)}, pages = {133--141}, abstract = {Dendritic cells (DC) obtained by culturing myeloid precursors in GM-CSF undergo maturation and induce an efficient T cell response when stimulated with microbial products. DC precursors themselves also recognize microbial products, and it remains unclear how these stimulated DC precursors modulate the immune response. We show here that M-CSF-conditioned human DC precursors responded to LPS, Mycobacteria bovis, and inflammatory cytokines by a rapid and robust production of IL-10, largely superior to that observed with immature DC or monocytes. The endogenous IL-10 restrained the DC precursors from converting into professional APC, as blocking the IL-10 receptor in the presence of LPS resulted in the formation of efficient T cell stimulators. LPS stimulation concomitant with DC differentiation gave rise to immature DC, which were tolerant to a secondary LPS exposure. Furthermore, the LPS-activated DC precursors reduced bystander DC maturation and anti-CD3/CD28-triggered T cell activation. These data suggest that when exposed to inflammatory or microbial signals, M-CSF-conditioned DC precursors can participate in the modulation of inflammation and immune response by rapid release of IL-10}, keywords = {Activation, APC, Cell Differentiation, COLONY-STIMULATING FACTOR, cytokine, Cytokines, cytology, Dendritic Cells, Differentiation, GM-CSF, Human, Humans, IL-10, IL10, IMMATURE, immune response, Immune Tolerance, Immunity, Immunology, inflammation, interleukin 10, Interleukin-10, lipopolysaccharide, Lipopolysaccharides, LPS, Macrophage, Macrophage Colony-Stimulating Factor, Maturation, metabolism, MODULATION, monocyte, Monocytes, MYCOBACTERIA, Mycobacterium, Myeloid Cells, Pharmacology, precursor, PRODUCTION, Protein, Receptor, Secondary, T CELL ACTIVATION, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{RM2007, title = {Evolutionary dynamics of immune-related genes and pathways in disease-vector mosquitoes}, author = {R M Waterhouse and E V Kriventseva and Stephan Meister and Z Xi and K S Alvarez and L C Bartholomay and Carolina Barillas-Mury and G Bian and Stéphanie A Blandin and B M Christensen and Y Dong and H Jiang and M R Kanost and A C Koutsos and Elena A Levashina and J Li and Petros Ligoxygakis and R M Maccallum and G F Mayhew and A Mendes and K Michel and M A Osta and S Paskewitz and S W Shin and D Vlachou and L Wang and W Wei and L Zheng and Z Zou and D W Severson and A S Raikhel and Fotis C Kafatos and G Dimopoulos and E M Zdobnov and G K Christophides}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17588928}, year = {2007}, date = {2007-06-22}, journal = {Science}, volume = {316}, pages = {5832}, abstract = {Mosquitoes are vectors of parasitic and viral diseases of immense importance for public health. The acquisition of the genome sequence of the yellow fever and Dengue vector, Aedes aegypti (Aa), has enabled a comparative phylogenomic analysis of the insect immune repertoire: in Aa, the malaria vector Anopheles gambiae (Ag), and the fruit fly Drosophila melanogaster (Dm). Analysis of immune signaling pathways and response modules reveals both conservative and rapidly evolving features associated with different functional gene categories and particular aspects of immune reactions. These dynamics reflect in part continuous readjustment between accommodation and rejection of pathogens and suggest how innate immunity may have evolved.}, keywords = {blandin, Evolution, M3i, Phylogeny}, pubstate = {published}, tppubtype = {article} } @article{habib_cutting_2007b, title = {Cutting Edge: Small Molecule CD40 Ligand Mimetics Promote Control of Parasitemia and Enhance Ŧ Cells Producing IFN-γ during Experimental Trypanosoma cruzi Infection}, author = {Mohammed Habib and Magali Noval Rivas and Mustapha Chamekh and Sébastien Wieckowski and Weimin Sun and Alberto Bianco and Nathalie Trouche and Olivier Chaloin and Hélène Dumortier and Michel Goldman and Gilles Guichard and Sylvie Fournel and Bernard Vray}, url = {https://www.jimmunol.org/content/178/11/6700}, doi = {10.4049/jimmunol.178.11.6700}, issn = {0022-1767, 1550-6606}, year = {2007}, date = {2007-06-01}, urldate = {2020-03-31}, journal = {The Journal of Immunology}, volume = {178}, number = {11}, pages = {6700--6704}, abstract = {Host resistance to Trypanosoma cruzi infection depends on a type 1 response characterized by a strong production of IL-12 and IFN-γ. Amplifying this response through CD40 triggering results in control of parasitemia. Two newly synthesized molecules (textbackslashtextless3 kDa) mimicking trimeric CD40L (mini CD40Ls-1 and -2) bind to CD40, activate murine dendritic cells, and elicit IL-12 production. Wild-type but not CD40 knockout mice exhibited a sharp decrease of parasitemia and mortality when inoculated with T. cruzi mixed with miniCD40Ls. Moreover, the immunosuppression induced by T. cruzi infection was impaired in mice treated with miniCD40Ls, as shown by proliferation of splenic lymphocytes, percentage of CD8+ T cells, and IFN-γ production. Mice surviving T. cruzi infection in the presence of miniCD40L-1 were immunized against a challenge infection. Our results indicate that CD40L mimetics are effective in vivo and promote the control of T. cruzi infection by overcoming the immunosuppression usually induced by the parasites.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{habib_cutting_2007, title = {Cutting edge: small molecule CD40 ligand mimetics promote control of parasitemia and enhance Ŧ cells producing IFN-gamma during experimental Trypanosoma cruzi infection}, author = {Mohammed Habib and Magali Noval Rivas and Mustapha Chamekh and Sébastien Wieckowski and Weimin Sun and Alberto Bianco and Nathalie Trouche and Olivier Chaloin and Hélène Dumortier and Michel Goldman and Gilles Guichard and Sylvie Fournel and Bernard Vray}, doi = {10.4049/jimmunol.178.11.6700}, issn = {0022-1767}, year = {2007}, date = {2007-06-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {178}, number = {11}, pages = {6700--6704}, abstract = {Host resistance to Trypanosoma cruzi infection depends on a type 1 response characterized by a strong production of IL-12 and IFN-gamma. Amplifying this response through CD40 triggering results in control of parasitemia. Two newly synthesized molecules (textless3 kDa) mimicking trimeric CD40L (mini CD40Ls(-1) and (-2)) bind to CD40, activate murine dendritic cells, and elicit IL-12 production. Wild-type but not CD40 knockout mice exhibited a sharp decrease of parasitemia and mortality when inoculated with T. cruzi mixed with miniCD40Ls. Moreover, the immunosuppression induced by T. cruzi infection was impaired in mice treated with miniCD40Ls, as shown by proliferation of splenic lymphocytes, percentage of CD8(+) T cells, and IFN-gamma production. Mice surviving T. cruzi infection in the presence of miniCD40L(-1) were immunized against a challenge infection. Our results indicate that CD40L mimetics are effective in vivo and promote the control of T. cruzi infection by overcoming the immunosuppression usually induced by the parasites.}, keywords = {Animals, CD40 Antigens, CD40 Ligand, Cell Line, Cells, Chagas Disease, Cultured, Dumortier, I2CT, Inbred BALB C, Inbred C57BL, Interferon-gamma, Knockout, Mice, Molecular Mimicry, Parasitemia, T-Lymphocyte Subsets, Team-Bianco, Team-Dumortier, Trypanosoma cruzi}, pubstate = {published}, tppubtype = {article} } @article{T2007, title = {Malaria Plasmodium agent induces alteration in the head proteome of their Anopheles mosquito host}, author = {T Lefevre and F Thomas and A Schwartz and Elena A Levashina and Stéphanie A Blandin and J-P Brizard and Le L Bourligu and E Demettre and F Renaud and D G Biron}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17464940}, year = {2007}, date = {2007-06-01}, journal = {Proteomics}, volume = {7}, number = {11}, pages = {1908-15}, abstract = {Despite increasing evidence of behavioural manipulation of their vectors by pathogens, the underlying mechanisms causing infected vectors to act in ways that benefit pathogen transmission remain enigmatic in most cases. Here, 2-D DIGE coupled with MS were employed to analyse and compare the head proteome of mosquitoes (Anopheles gambiae sensu stricto (Giles)) infected with the malarial parasite (Plasmodium berghei) with that of uninfected mosquitoes. This approach detected altered levels of 12 protein spots in the head of mosquitoes infected with sporozoites. These proteins were subsequently identified using MS and functionally classified as belonging to metabolic, synaptic, molecular chaperone, signalling, and cytoskeletal groups. Our results indicate an altered energy metabolism in the head of sporozoite-infected mosquitoes. Some of the up-/down-regulated proteins identified, such as synapse-associated protein, 14-3-3 protein and calmodulin, have previously been shown to play critical roles in the CNS of both invertebrates and vertebrates. Furthermore, a heat shock response (HSP 20) and a variation of cytoarchitecture (tropomyosins) have been shown. Discovery of these proteins sheds light on potential molecular mechanisms that underlie behavioural modifications and offers new insights into the study of intimate interactions between Plasmodium and its Anopheles vector.}, keywords = {blandin, M3i, Proteome}, pubstate = {published}, tppubtype = {article} } @article{weber_role_2007, title = {Role of the Spatzle Pro-domain in the generation of an active toll receptor ligand}, author = {Alexander N R Weber and Monique Gangloff and Martin C Moncrieffe and Yann Hyvert and Jean-Luc Imler and Nicholas J Gay}, doi = {10.1074/jbc.M700068200}, issn = {0021-9258}, year = {2007}, date = {2007-05-01}, journal = {The Journal of Biological Chemistry}, volume = {282}, number = {18}, pages = {13522--13531}, abstract = {The cytokine Spätzle is the ligand for Drosophila Toll, the prototype of an important family of membrane receptors that function in embryonic patterning and innate immunity. A dimeric precursor of Spätzle is processed by an endoprotease to produce a form (C-106) that cross-links Toll receptor ectodomains and establishes signaling. Here we show that before processing the pro-domain of Spätzle is required for correct biosynthesis and secretion. We mapped two loss-of-function mutations of Spätzle to a discrete site in the pro-domain and showed that the phenotype arises because of a defect in biosynthesis rather than signaling. We also report that the pro-domain and C-106 remain associated after cleavage and that this processed complex signals with the same characteristics as the C-terminal fragment. These results suggest that before activation the determinants on C-106 that bind specifically to Toll are sequestered by the pro-domain and that proteolytic processing causes conformational rearrangements that expose these determinants and enables binding to Toll. Furthermore, we show that the pro-domain is released when the Toll extracellular domain binds to the complex, a finding that has implications for the generation of a signaling-competent Toll dimer.}, keywords = {Animals, Cytokines, dimerization, imler, ligands, M3i, Post-Translational, Protein Binding, Protein Processing, Protein Structure, Signal Transduction, Tertiary, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{lelong_crl-rpos_2007, title = {The Crl-RpoS regulon of Escherichia coli.}, author = {Cécile Lelong and Kryssia Aguiluz and Sylvie Luche and Lauriane Kuhn and Jérôme Garin and Thierry Rabilloud and Johannes Geiselmann}, doi = {10.1074/mcp.M600191-MCP200}, issn = {1535-9476 1535-9476}, year = {2007}, date = {2007-04-01}, journal = {Molecular & cellular proteomics : MCP}, volume = {6}, number = {4}, pages = {648--659}, abstract = {The RpoS subunit of RNA polymerase controls the expression of numerous genes involved in stationary phase and in response to different stress conditions. The regulatory protein Crl increases the activity of RpoS by direct interaction with the RpoS holoenzyme. To define the extent of the Crl regulon, we used two-dimensional SDS-PAGE to measure the role of Crl in regulating the expression of the Escherichia coliproteome in stationary phase at 30 degrees C. By comparing the proteome of four strains (wild type, crl(-), rpoS(-), and crl(-)rpoS(-)), we observed that the intensity of 74 spots was modified in at least one mutant context. 62 spots were identified by mass spectrometry and correspond to 40 distinct proteins. They were classified in four main categories: DNA metabolism, central metabolism, response to environmental modifications, and miscellaneous. Three proteins were specifically involved in quorum sensing: TnaA (the tryptophanase that converts tryptophan to indole), WrbA (Trp repressor-binding protein), and YgaG (homologous to LuxS, autoinducer-2 synthase). Because little is known about the regulation of Crl expression, we investigated the influence of diffusible molecules on the expression of Crl. Using Western blotting experiments, we showed that, at 30 degrees C, a diffusible molecule(s) produced during the transition phase between the exponential and stationary phases induces a premature expression of Crl. Indole was tested as one of the potential candidates: at 37 degrees C, it is present in the extracellular medium at a constant concentration, but at 30 degrees C, its concentration peaks during the transition phase. When indole was added to the culture medium, it also induced prematurely the expression of Crl at both the transcriptional and translational levels in a Crl-dependent manner. Crl may thus be considered a new environmental sensor via the indole concentration.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{paclet_regulation_2007, title = {Regulation of phagocyte NADPH oxidase activity: identification of two cytochrome b558 activation states.}, author = {Marie-Hélène Paclet and Sylvie Berthier and Lauriane Kuhn and Jérôme Garin and Françoise Morel}, doi = {10.1096/fj.06-6852com}, issn = {1530-6860 0892-6638}, year = {2007}, date = {2007-04-01}, journal = {FASEB journal : official publication of the Federation of American Societies for Experimental Biology}, volume = {21}, number = {4}, pages = {1244--1255}, abstract = {Activation of the phagocyte NADPH oxidase (phox) requires the association of cytosolic proteins (p67-phox, p47-phox, p40-phox, and Rac1/2) with the membrane cytochrome b558, leading to a hemoprotein conformation change. To clarify this mechanism, the phagocyte NADPH oxidase complex was isolated through cytochrome b558 purification after three chromatographic steps. The purified neutrophil complex was constitutively active in the absence of an amphiphile agent with a maximum turnover (125 mol O2(-) x s(-1) x mol heme b(-1)), indicating that cytochrome b558 has been activated by cytosolic proteins and is in an "open conformation," able to transfer a maximum rate of electrons. In contrast, the phox complex prepared with B lymphocyte cytosol shows a lower constitutive turnover (approximately 50 mol O2(-) x s(-1) x mol heme b(-1)). Analysis of phox complex components by Western blot and mass spectrometry showed the presence of cytosolic factors (especially p67-phox) and structural proteins (moesin, ezrin). To investigate the difference in activity of phox complexes, we evaluated the effect of MRP8 and MRP14, specifically expressed in neutrophils, on the activity of the B lymphocyte complex. MRPs induce the switch between the partially and the fully "open" cytochrome b558 conformation. Moreover, their effect was independent of p67-phox. Data point out two potential cytochrome b558 activation states.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{lanquar_15n-metabolic_2007, title = {15N-metabolic labeling for comparative plasma membrane proteomics in Arabidopsis cells.}, author = {Viviane Lanquar and Lauriane Kuhn and Françoise Lelièvre and Mehdi Khafif and Christelle Espagne and Christophe Bruley and Hélène Barbier-Brygoo and Jérôme Garin and Sébastien Thomine}, doi = {10.1002/pmic.200600791}, issn = {1615-9853 1615-9853}, year = {2007}, date = {2007-03-01}, journal = {Proteomics}, volume = {7}, number = {5}, pages = {750--754}, abstract = {An important goal for proteomic studies is the global comparison of proteomes from different genotypes, tissues, or physiological conditions. This has so far been mostly achieved by densitometric comparison of spot intensities after protein separation by 2-DE. However, the physicochemical properties of membrane proteins preclude the use of 2-DE. Here, we describe the use of in vivo labeling by the stable isotope 15N as an alternative approach for comparative membrane proteomic studies in plant cells. We confirm that 15N-metabolic labeling of proteins is possible and efficient in Arabidopsis suspension cells. Quantification of 14N versus 15N MS signals reflects the relative abundance of 14N and 15N proteins in the sample analyzed. We describe the use of 15N-metabolic labeling to perform a partial comparative analysis of Arabidopsis cells following cadmium exposure. By focusing our attention on plasma membrane proteins, we were able to confidently identify proteins showing up to 5-fold regulation compared to unexposed cells. This study provides a proof of principle that 15N-metabolic labeling is a useful technique for comparative membrane proteome studies.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{kostarelos_cellular_2007, title = {Cellular uptake of functionalized carbon nanotubes is independent of functional group and cell type}, author = {Kostas Kostarelos and Lara Lacerda and Giorgia Pastorin and Wei Wu and Sébastien Wieckowski and Jacqueline Luangsivilay and Sylvie Godefroy and Davide Pantarotto and Jean-Paul Briand and Sylviane Muller and Maurizio Prato and Alberto Bianco}, doi = {10.1038/nnano.2006.209}, issn = {1748-3395}, year = {2007}, date = {2007-02-01}, journal = {Nature Nanotechnology}, volume = {2}, number = {2}, pages = {108--113}, keywords = {Animals, carbon, Cell Membrane, Cells, Cultured, Diffusion, Humans, I2CT, Nanotubes, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {A present-day aminoacyl-tRNA synthetase with ancestral editing properties}, author = {B Zhu and M W Zhao and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17095543}, isbn = {17095543}, year = {2007}, date = {2007-01-01}, journal = {RNA}, volume = {13}, number = {1}, pages = {15-21}, abstract = {Leucyl-, isoleucyl-, and valyl-tRNA synthetases form a subgroup of related aminoacyl-tRNA synthetases that attach similar amino acids to their cognate tRNAs. To prevent amino acid misincorporation during translation, these enzymes also hydrolyze mischarged tRNAs through a post-transfer editing mechanism. Here we show that LeuRS from the deep-branching bacterium Aquifex aeolicus edits the complete set of aminoacylated tRNAs generated by the three enzymes: Ile-tRNA(Ile), Val-tRNA(Ile), Val-tRNA(Val), Thr-tRNA(Val), and Ile-tRNA(Leu). This unusual enlarged editing property was studied in a model of a primitive editing system containing a composite minihelix carrying the triple leucine, isoleucine, and valine identity mimicking the primitive tRNA precursor. We found that the freestanding LeuRS editing domain can edit this precursor in contrast to IleRS and ValRS editing domains. These results suggest that A. aeolicus LeuRS carries editing properties that seem more primitive than those of IleRS and ValRS. They suggest that the A. aeolicus editing domain has preserved the ambiguous editing property from the ancestral common editing domain or, alternatively, that this plasticity results from a specific metabolic adaptation.}, note = {1355-8382 (Print) Journal Article Research Support, Non-U.S. Gov't}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Anopheles gambiae miRNAs as actors of defence reaction against Plasmodium invasion}, author = {F Winter and S Edaye and A Huttenhofer and C Brunel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17933784}, isbn = {17933784}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {20}, pages = {6953-6962}, abstract = {The path Plasmodium takes across the Anopheles midgut constitutes the major bottleneck during the malaria transmission cycle. In the present study, using a combination of shot-gun cloning and bioinformatic analysis, we have identified 18 miRNAs from Anopheles gambiae including three miRNAs unique to mosquito. Twelve of them are expressed ubiquitously across the body, independently of gender, while the other six exhibited an expression pattern restricted to the digestive system. Strikingly, the expression patterns of four miRNAs, including the three unique to mosquito, are affected by the presence of Plasmodium. We also show that knocking down Dicer1 and Ago1 mRNAs led to an increased sensitivity to Plasmodium infection. Altogether, these data support an involvement of miRNAs as new layers in the regulation of Anopheles defence reaction.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {BRUNEL Animals Anopheles gambiae/*genetics/*immunology/parasitology Digestive System/immunology/metabolism/parasitology Female *Gene Expression Profiling Gene Library Gene Silencing Male MicroRNAs/*immunology Plasmodium/*immunology Ribonuclease III/genetics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A tale in molecular recognition: the hammerhead ribozyme}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17089350}, isbn = {17089350}, year = {2007}, date = {2007-01-01}, journal = {J Mol Recognit}, volume = {20}, number = {1}, pages = {1-3}, abstract = {A new crystal structure of the hammerhead ribozyme demonstrates the influence of peripheral tertiary contacts on the local conformations around the active site. This structure resolves many conflicting results obtained on reduced systems.}, note = {0952-3499 (Print) Journal Article}, keywords = {Animals Base Sequence Catalysis Molecular Sequence Data Nucleic Acid Conformation RNA, Catalytic/chemistry/genetics/*metabolism Schistosoma mansoni/chemistry, Unité ARN, WESTHOF, WESTHOF Animals Base Sequence Catalysis Molecular Sequence Data Nucleic Acid Conformation RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {In search of new inhibitors of HIV-1 replication: synthesis and study of 1-(2'-Deoxy-beta-D-Ribofuranosyl)-1,2,4-triazole-3-carboxamide as a selective viral mutagenic agent}, author = {V Vivet-Boudou and J C Paillart and A Burger and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18066893}, isbn = {18066893}, year = {2007}, date = {2007-01-01}, journal = {Nucleosides Nucleotides Nucleic Acids}, volume = {26}, number = {6-7}, pages = {743-746}, abstract = {With the emergence of HIV strains resistant or cross-resistant to nearly all antiretroviral regimen, novel therapy approaches have to be considered. As a part of our current work on viral mutagenic compounds, we prepared 1-(2' -deoxy-beta-D-ribofuranosyl)-1,2,4-triazole-3-carboxamide (2' -deoxy-ribavirin) and its 5' -triphosphate derivative. The nucleoside mutagenic activity was evaluated on HIV-1 NL4-3 in CEMx174 cell culture. After 2.5 months, no reduction on HIV-1 viability was observed. On the other hand, in vitro experiments with purified HIV-1 RT demonstrated that the triphosphate analog can be incorporated opposite to several natural nucleosides.}, note = {1525-7770 (Print) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Antiviral Agents/*chemical synthesis/chemistry/*pharmacology HIV-1/*drug effects/*physiology Imidazoles/*chemical synthesis/chemistry/*pharmacology Mutagenesis/*drug effects Virus Replication/*drug effects, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Disorder Can Exist inside Well-Diffracting Crystals}, author = {E Touze and B Lorber and M Deniziak and H D Becker and D Kern and R Giege and C Sauter}, url = {http://pubs.acs.org/doi/abs/10.1021/cg7007057?prevSearch=Touz%25C3%25A9&searchHistoryKey=}, year = {2007}, date = {2007-01-01}, journal = {Crystal Growth & Design}, volume = {7}, number = {11}, pages = {2195-2197}, abstract = {Unlike other glutaminyl-tRNA synthetases, the one from radioresistant bacterium Deinococcus radiodurans (Dr-GlnRS) possesses an additional C-terminal extension of 220 residues that shares some homology with the subunit of another enzyme of the translation machinery. Dr-GlnRS has been crystallized in an orthorhombic space group. The crystals diffract X-rays to a resolution of 2 Å. The determination of the structure of this atypical GlnRS showed that its N- and C-terminal appendices, which encompass in total one third of the proteinメs 852 amino acids, are actually disordered in the crystal lattice. This example demonstrates that macromolecule crystallization can tolerate large flexible regions in the solvent channels as long as they do not interfere with the packing contacts. This intriguing case is analyzed and discussed in light of current crystallogenesis strategies.}, note = {DOI: 10.1021/cg7007057}, keywords = {FRUGIER, GIEGE KERN FLORENTZ, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vitro dimerization of human immunodeficiency virus type 1 (HIV-1) spliced RNAs}, author = {L Sinck and D Richer and J Howard and M Alexander and D F Purcell and R Marquet and J C Paillart}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17925344}, doi = {rna.678307 [pii] 10.1261/rna.678307}, issn = {1469-9001 (Electronic) 1355-8382 (Linking)}, year = {2007}, date = {2007-01-01}, journal = {RNA}, volume = {13}, number = {12}, pages = {2141-2150}, abstract = {The human immunodeficiency virus type 1 (HIV-1) packages its genomic RNA as a dimer of homologous RNA molecules that has to be selected among a multitude of cellular and viral RNAs. Interestingly, spliced viral mRNAs are packaged into viral particles with a relatively low efficiency despite the fact that they contain most of the extended packaging signal found in the 5' untranslated region of the genomic RNA, including the dimerization initiation site (DIS). As a consequence, HIV-1 spliced viral RNAs can theoretically homodimerize and heterodimerize with the genomic RNA, and thus they should directly compete with genomic RNA for packaging. To shed light on this issue, we investigated for the first time the in vitro dimerization properties of spliced HIV-1 RNAs. We found that singly spliced (env, vpr) and multispliced (tat, rev, and nef) RNA fragments are able to dimerize in vitro, and to efficiently form heterodimers with genomic RNA. Chemical probing experiments and inhibition of RNA dimerization by an antisense oligonucleotide directed against the DIS indicated that the DIS is structurally functional in spliced HIV-1 RNA, and that RNA dimerization occurs through a loop-loop interaction. In addition, by combining in vitro transcription and dimerization assays, we show that heterodimers can be efficiently formed only when the two RNA fragments are synthesized simultaneously, in the same environment. Together, our results support a model in which RNA dimerization would occur during transcription in the nucleus and could thus play a major role in splicing, transport, and localization of HIV-1 RNA.}, note = {Sinck, Lucile Richer, Delphine Howard, Jane Alexander, Marina Purcell, Damian F J Marquet, Roland Paillart, Jean-Christophe Research Support, Non-U.S. Gov't United States RNA (New York, N.Y.) RNA. 2007 Dec;13(12):2141-50. Epub 2007 Oct 9.}, keywords = {Dimerization Drug Stability Genes, env Genes, MARQUET, Messenger/genetics RNA, nef Genes, PAILLART, tat HIV-1/*genetics Humans *RNA Splicing RNA, Unité ARN, Viral Genes, Viral/*genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mitochondrial aspartyl-tRNA synthetase deficiency causes leukoencephalopathy with brain stem and spinal cord involvement and lactate elevation}, author = {G C Scheper and T van der Klok and R J van Andel and C G van Berkel and M Sissler and J Smet and T I Muravina and S V Serkov and G Uziel and M Bugiani and R Schiffmann and I Krageloh-Mann and J A Smeitink and C Florentz and R Van Coster and J C Pronk and M S van der Knaap}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17384640}, isbn = {17384640}, year = {2007}, date = {2007-01-01}, journal = {Nat Genet}, volume = {39}, number = {4}, pages = {534-539}, abstract = {Leukoencephalopathy with brain stem and spinal cord involvement and lactate elevation (LBSL) has recently been defined based on a highly characteristic constellation of abnormalities observed by magnetic resonance imaging and spectroscopy. LBSL is an autosomal recessive disease, most often manifesting in early childhood. Affected individuals develop slowly progressive cerebellar ataxia, spasticity and dorsal column dysfunction, sometimes with a mild cognitive deficit or decline. We performed linkage mapping with microsatellite markers in LBSL families and found a candidate region on chromosome 1, which we narrowed by means of shared haplotypes. Sequencing of genes in this candidate region uncovered mutations in DARS2, which encodes mitochondrial aspartyl-tRNA synthetase, in affected individuals from all 30 families. Enzyme activities of mutant proteins were decreased. We were surprised to find that activities of mitochondrial complexes from fibroblasts and lymphoblasts derived from affected individuals were normal, as determined by different assays.}, note = {1061-4036 (Print) Journal Article}, keywords = {FLORENTZ, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {From macrofluidies to microfluidies for the crystallization of biological macromolecules}, author = {C Sauter and K Dhouib and B Lorber}, url = {http://pubs.acs.org/doi/abs/10.1021/cg700955f?prevSearch=sauter&searchHistoryKey=}, year = {2007}, date = {2007-01-01}, journal = {Crystal Growth & Design}, volume = {7}, number = {11}, pages = {2247-2250}, abstract = {This review presents the goals and principles of microfluidic technologies applied to the crystallization of biological macromolecules. A comparison of the devices that are available commercially or described in the literature summarizes the current state-of-the-art in microfluidics. A novel chip based on the counter-diffusion of solute molecules playing the role of crystallization agents is described. Inside the microfluidic channels composing the chip mass transport essentially occurs by diffusion. The chip is made of a rigid polymer that is impermeable to gases and compatible with crystal examination and monitoring in polarized light. The selected material is also transparent to X-rays, and three-dimensional protein structures can be determined from crystals contained inside this device using X-ray diffraction data collected on a synchrotron source. The outstanding quality of the electron-density maps demonstrates that on-chip crystal analysis is feasible. The replacement of conventional crystallization setups by inexpensive microfluidic chips for screening best crystallization agents and automated crystal diffraction analysis is discussed.}, note = {DOI: 10.1021/cg700955f}, keywords = {FRUGIER, GIEGE FLORENTZ, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dinucleotide TpT and Its 2'-O-Me Analogue Possess Different Backbone Conformations and Flexibilities but Similar Stacked Geometries}, author = {G P Santini and C Pakleza and P Auffinger and C Moriou and A Favre and P Clivio and J A Cognet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17625827}, isbn = {17625827}, year = {2007}, date = {2007-01-01}, journal = {J Phys Chem B}, volume = {111}, number = {31}, pages = {9400-9409}, abstract = {UV irradiation at 254 nm of 2'-O,5-dimethyluridylyl(3'-5')-2'-O,5-dimethyluridine (1a) and of natural thymidylyl(3'-5')thymidine (1b) generates the same photoproducts (CPD and (6-4)PP; responsible for cell death and skin cancer). The ratios of quantum yields of photoproducts obtained from 1a (determined herein) to that from 1b are in a proportion close to the approximately threefold increase of stacked dinucleotides for 1a compared with those of 1b (from previous circular dichroism results). 1a and 1b however are endowed with different predominant sugar conformations, C3'-endo (1a) and C2'-endo (1b). The present investigation of the stacked conformation of these molecules, by unrestrained state-of-the-art molecular simulation in explicit solvent and salt, resolves this apparent paradox and suggests the following main conclusions. Stacked dinucleotides 1a and 1b adopt the main characteristic features of a single-stranded A and B form, respectively, where the relative positions of the backbone and the bases are very different. Unexpectedly, the geometry of the stacking of two thymine bases, within each dinucleotide, is very similar and is in excellent agreement with photochemical and circular dichroism results. Analyses of molecular dynamics trajectories with conformational adiabatic mapping show that 1a and 1b explore two different regions of conformational space and possess very different flexibilities. Therefore, even though their base stacking is very similar, these molecules possess different geometrical, mechanical, and dynamical properties that may account for the discrepancy observed between increased stacking and increased photoproduct formations. The computed average stacked conformations of 1a and 1b are well-defined and could serve as starting models to investigate photochemical reactions with quantum dynamics simulations.}, note = {1520-6106 (Print) Journal article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural elements defining elongation factor Tu mediated suppression of codon ambiguity}, author = {H Roy and H D Becker and M H Mazauric and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17478519}, isbn = {17478519}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {10}, pages = {3420-3430}, abstract = {In most prokaryotes Asn-tRNA(Asn) and Gln-tRNA(Gln) are formed by amidation of aspartate and glutamate mischarged onto tRNA(Asn) and tRNA(Gln), respectively. Coexistence in the organism of mischarged Asp-tRNA(Asn) and Glu-tRNA(Gln) and the homologous Asn-tRNA(Asn) and Gln-tRNA(Gln) does not, however, lead to erroneous incorporation of Asp and Glu into proteins, since EF-Tu discriminates the misacylated tRNAs from the correctly charged ones. This property contrasts with the canonical function of EF-Tu, which is to non-specifically bind the homologous aa-tRNAs, as well as heterologous species formed in vitro by aminoacylation of non-cognate tRNAs. In Thermus thermophilus that forms the Asp-tRNA(Asn) intermediate by the indirect pathway of tRNA asparaginylation, EF-Tu must discriminate the mischarged aminoacyl-tRNAs (aa-tRNA). We show that two base pairs in the tRNA T-arm and a single residue in the amino acid binding pocket of EF-Tu promote discrimination of Asp-tRNA(Asn) from Asn-tRNA(Asn) and Asp-tRNA(Asp) by the protein. Our analysis suggests that these structural elements might also contribute to rejection of other mischarged aa-tRNAs formed in vivo that are not involved in peptide elongation. Additionally, these structural features might be involved in maintaining a delicate balance of weak and strong binding affinities between EF-Tu and the amino acid and tRNA moieties of other elongator aa-tRNAs.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Amino Acyl/*chemistry/metabolism RNA, Asn/*chemistry/metabolism RNA, Asp/chemistry/metabolism Thermus thermophilus/genetics *Transfer RNA Aminoacylation, KERN Base Pairing *Codon Escherichia coli Proteins/metabolism Models, Molecular Peptide Elongation Factor Tu/*chemistry/metabolism Protein Binding RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{pmid17245431, title = {Recruitment of chromatin-modifying enzymes by CTIP2 promotes HIV-1 transcriptional silencing}, author = {Céline Marban and Stella Suzanne and Franck Dequiedt and Stéphane de Walque and Laetitia Redel and Carine Van Lint and Dominique Aunis and Olivier Rohr}, doi = {10.1038/sj.emboj.7601516}, issn = {0261-4189}, year = {2007}, date = {2007-01-01}, urldate = {2007-01-01}, journal = {EMBO J}, volume = {26}, number = {2}, pages = {412--423}, abstract = {Following entry and reverse transcription, the HIV-1 genome is integrated into the host genome. In contrast to productively infected cells, latently infected cells frequently harbor HIV-1 genomes integrated in heterochromatic structures, allowing persistence of transcriptionally silent proviruses. Microglial cells are the main HIV-1 target cells in the central nervous system and constitute an important reservoir for viral pathogenesis. In the present work, we show that, in microglial cells, the co-repressor COUP-TF interacting protein 2 (CTIP2) recruits a multienzymatic chromatin-modifying complex and establishes a heterochromatic environment at the HIV-1 promoter. We report that CTIP2 recruits histone deacetylase (HDAC)1 and HDAC2 to promote local histone H3 deacetylation at the HIV-1 promoter region. In addition, DNA-bound CTIP2 also associates with the histone methyltransferase SUV39H1, which increases local histone H3 lysine 9 methylation. This allows concomitant recruitment of HP1 proteins to the viral promoter and formation of local heterochromatin, leading to HIV-1 silencing. Altogether, our findings uncover new therapeutic opportunities for purging latent HIV-1 viruses from their cellular reservoirs.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Anopheles gambiae miRNAs as actors of defence reaction against Plasmodium invasion}, author = {F Winter and S Edaye and A Huttenhofer and C Brunel}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17933784}, isbn = {17933784}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {20}, pages = {6953-62}, abstract = {The path Plasmodium takes across the Anopheles midgut constitutes the major bottleneck during the malaria transmission cycle. In the present study, using a combination of shot-gun cloning and bioinformatic analysis, we have identified 18 miRNAs from Anopheles gambiae including three miRNAs unique to mosquito. Twelve of them are expressed ubiquitously across the body, independently of gender, while the other six exhibited an expression pattern restricted to the digestive system. Strikingly, the expression patterns of four miRNAs, including the three unique to mosquito, are affected by the presence of Plasmodium. We also show that knocking down Dicer1 and Ago1 mRNAs led to an increased sensitivity to Plasmodium infection. Altogether, these data support an involvement of miRNAs as new layers in the regulation of Anopheles defence reaction.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {BRUNEL Animals Anopheles gambiae/*genetics/*immunology/parasitology Digestive System/immunology/metabolism/parasitology Female *Gene Expression Profiling Gene Library Gene Silencing Male MicroRNAs/*immunology Plasmodium/*immunology Ribonuclease III/genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural requirements for nucleocapsid protein-mediated dimerization of avian leukosis virus RNA}, author = {M B Ali and F Chaminade and I Kanevsky and E Ennifar and L Josset and D Ficheux and J L Darlix and P Fosse}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17706668}, isbn = {17706668}, year = {2007}, date = {2007-01-01}, journal = {J Mol Biol}, volume = {372}, number = {4}, pages = {1082-96}, abstract = {The avian leukosis virus (ALV) belongs to the alpha group of retroviruses that are widespread in nature. The 5'-untranslated region of ALV genome contains the L3 element that is important for virus infectivity and the formation of an unstable RNA dimer in vitro. The L3 sequence is predicted to fold into a long stem-loop structure with two internal loops and an apical one. Phylogenetic analysis predicts that the L3 stem-loop is conserved in alpharetroviruses. Furthermore, a significant selection mechanism maintains a palindrome in the apical loop. The nucleocapsid protein of the alpharetroviruses (NCp12) is required for RNA dimer formation and replication in vivo. It is not known whether L3 can be an NCp12-mediated RNA dimerization site able to bind NCp12 with high affinity. Here, we report that NCp12 chaperones formation of a stable ALV RNA dimer through L3. To investigate the NCp12-mediated L3 dimerization reaction, we performed site-directed mutagenesis, gel retardation and heterodimerization assays and analysis of thermostability of dimeric RNAs. We show that the affinity of NCp12 for L3 is lower than its affinity for the microPsi RNA packaging signal. Results show that conservation of a long stem-loop structure and a loop-loop interaction are not required for NCp12-mediated L3 dimerization. We show that the L3 apical stem-loop is sufficient to form an extended duplex and the whole stem-loop L3 cannot be converted by NCp12 into a duplex extending throughout L3. Three-dimensional modelling of the stable L3 dimer supports the notion that the extended duplex may represent the minimal dimer linkage structure found in the genomic RNA.}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoglycoside binding to the HIV-1 RNA dimerization initiation site: thermodynamics and effect on the kissing-loop to duplex conversion}, author = {S Bernacchi and S Freisz and C Maechling and B Spiess and R Marquet and P Dumas and E Ennifar}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17942426}, isbn = {17942426}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {21}, pages = {7128-39}, abstract = {Owing to a striking, and most likely fortuitous, structural and sequence similarity with the bacterial 16 S ribosomal A site, the RNA kissing-loop complex formed by the HIV-1 genomic RNA dimerization initiation site (DIS) specifically binds 4,5-disubstituted 2-deoxystreptamine (2-DOS) aminoglycoside antibiotics. We used chemical probing, molecular modeling, isothermal titration calorimetry (ITC) and UV melting to investigate aminoglycoside binding to the DIS loop-loop complex. We showed that apramycin, an aminoglycoside containing a bicyclic moiety, also binds the DIS, but in a different way than 4,5-disubstituted 2-DOS aminoglycosides. The determination of thermodynamic parameters for various aminoglycosides revealed the role of the different rings in the drug-RNA interaction. Surprisingly, we found that the affinity of lividomycin and neomycin for the DIS (K(d) approximately 30 nM) is significantly higher than that obtained in the same experimental conditions for their natural target, the bacterial A site (K(d) approximately 1.6 microM). In good agreement with their respective affinity, aminoglycoside increase the melting temperature of the loop-loop interaction and also block the conversion from kissing-loop complex to extended duplex. Taken together, our data might be useful for selecting new molecules with improved specificity and affinity toward the HIV-1 DIS RNA.}, note = {1362-4962 (Electronic)}, keywords = {DUMAS, ENNIFAR, MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Influence of C-5 halogenation of uridines on hairpin versus duplex RNA folding.}, author = {E Ennifar and S Bernacchi and P Wolff and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17630326}, isbn = {17630326}, year = {2007}, date = {2007-01-01}, journal = {RNA}, volume = {13}, number = {9}, pages = {1445-52}, abstract = {Halogenation of bases is a widespread method used for solving crystal structures of nucleic acids. However, this modification may have important consequences on RNA folding and thus on the success of crystallization. We have used a combination of UV thermal melting, steady-state fluorescence, X-ray crystallography, and gel electrophoresis techniques to study the influence of uridine halogenation (bromination or iodination) on the RNA folding. The HIV-1 Dimerization Initiation Site is an RNA hairpin that can adopt an alternative duplex conformation and was used as a model. We have shown that, unexpectedly, the RNA hairpin/duplex ratio is strongly dependent not only on the presence but also on the position of halogenation.}, keywords = {ENNIFAR, MARQUET, RNA HIV fluorescence halogen crystal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{monneaux_peptide-based_2007, title = {Peptide-based therapy in lupus: promising data}, author = {Fanny Monneaux and Sylviane Muller}, doi = {10.1007/978-0-387-72005-0_11}, issn = {0065-2598}, year = {2007}, date = {2007-01-01}, journal = {Advances in Experimental Medicine and Biology}, volume = {601}, pages = {105--112}, abstract = {Systemic lupus erythematosus (SLE) is a multisystem chronic inflammatory disease of multifactorial aetiology, characterized by inflammation and damage of various tissues and organs. Current treatments of the disease are mainly based on immunosuppressive drugs such as corticosteroids and cyclophosphamide. Although these treatments have reduced mortality and morbidity, they cause a non-specific immune suppression. To avoid these side effects, our efforts should focus on the development of alternative therapeutic strategies, which consist, for example in specific T cell targeting using autoantigen-derived peptides identified as sequences encompassing major epitopes.}, keywords = {Adrenal Cortex Hormones, Animals, Autoantigens, Autoimmune Diseases, Cyclophosphamide, Epitopes, Humans, I2CT, Immune System, Immunosuppressive Agents, inflammation, Lupus Erythematosus, Monneaux, Peptides, Systemic, T-Lymphocytes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{, title = {Using X-ray absorption spectra to monitor specific radiation damage to anomalously scattering atoms in macromolecular crystallography}, author = {V Olieric and E Ennifar and A Meents and M Fleurant and C Besnard and P Pattison and M Schiltz and C Schulze-Briese and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17582167}, isbn = {17582167}, year = {2007}, date = {2007-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {63}, number = {Pt 7}, pages = {759-68}, abstract = {Radiation damage in macromolecular crystals is not suppressed even at 90 K. This is particularly true for covalent bonds involving an anomalous scatterer (such as bromine) at the 'peak wavelength'. It is shown that a series of absorption spectra recorded on a brominated RNA faithfully monitor the extent of cleavage. The continuous spectral changes during irradiation preserve an 'isosbestic point', each spectrum being a linear combination of 'zero' and 'infinite' dose spectra. This easily yields a good estimate of the partial occupancy of bromine at any intermediate dose. The considerable effect on the near-edge features in the spectra of the crystal orientation versus the beam polarization has also been examined and found to be in good agreement with a previous study. Any significant influence of the (C-Br bond/beam polarization) angle on the cleavage kinetics of bromine was also searched for, but was not detected. These results will be useful for standard SAD/MAD experiments and for the emerging 'radiation-damage-induced phasing' method exploiting both the anomalous signal of an anomalous scatterer and the 'isomorphous' signal resulting from its cleavage.}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{parietti_regulatory_2007, title = {Regulatory Ŧ cells and systemic lupus erythematosus}, author = {Véronique Parietti and Hélène Chifflot and Sylviane Muller and Fanny Monneaux}, doi = {10.1196/annals.1422.007}, issn = {0077-8923}, year = {2007}, date = {2007-01-01}, journal = {Annals of the New York Academy of Sciences}, volume = {1108}, pages = {64--75}, abstract = {Regulatory T cells, especially CD4+CD25+ T cells, "natural killer" T cells and gammadelta T cells, are central in the maintenance of peripheral tolerance and the protection from the development of autoimmune diseases. Numerical or functional modifications of these cell populations were demonstrated to lead to the breakdown of tolerance and the emergence of autoimmunity. Involvement of regulatory T cells in the pathogenesis of systemic autoimmune diseases, such as systemic lupus erythematosus, might be of first importance. In murine models and patients with lupus, these regulatory T cells seem to be reduced in number. Functional deficiencies have also been described in a few studies. A better knowledge of regulatory T cell functional properties in systemic autoimmune diseases is essential to manipulate these cells and hopefully to restore immune tolerance.}, keywords = {Animals, Autoimmunity, Humans, I2CT, Immune Tolerance, Lupus Erythematosus, Monneaux, Regulatory, Systemic, T-Lymphocyte Subsets, T-Lymphocytes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{, title = {Conformations of Flanking Bases in HIV-1 RNA DIS Kissing Complexes Studied by Molecular Dynamics}, author = {K Reblova and E Fadrna and J Sarzynska and T Kulinski and P Kulhanek and E Ennifar and J Koca and J Sponer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17704156}, isbn = {17704156}, year = {2007}, date = {2007-01-01}, journal = {Biophys J}, volume = {93}, number = {11}, pages = {3932-3949}, abstract = {Explicit solvent molecular dynamics simulations (in total almost 800 ns including locally enhanced sampling runs) were applied with different ion conditions and with two force fields (AMBER and CHARMM) to characterize typical geometries adopted by the flanking bases in the RNA kissing-loop complexes. We focus on flanking base positions in multiple x-ray and NMR structures of HIV-1 DIS kissing complexes and kissing complex from the large ribosomal subunit of Haloarcula marismortui. An initial x-ray open conformation of bulged-out bases in HIV-1 DIS complexes, affected by crystal packing, tends to convert to a closed conformation formed by consecutive stretch of four stacked purine bases. This is in agreement with those recent crystals where the packing is essentially avoided. We also observed variants of the closed conformation with three stacked bases, while nonnegligible populations of stacked geometries with bulged-in bases were detected, too. The simulation results reconcile differences in positions of the flanking bases observed in x-ray and NMR studies. Our results suggest that bulged-out geometries are somewhat more preferred, which is in accord with recent experiments showing that they may mediate tertiary contacts in biomolecular assemblies or allow binding of aminoglycoside antibiotics.}, note = {0006-3495 (Print) Journal article}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{monneaux_spliceosome_2007, title = {[The spliceosome and its interest for lupus therapy]}, author = {F Monneaux and S Muller}, doi = {10.1016/j.revmed.2007.05.003}, issn = {0248-8663}, year = {2007}, date = {2007-01-01}, journal = {La Revue De Medecine Interne}, volume = {28}, number = {10}, pages = {725--728}, abstract = {INTRODUCTION: The spliceosome, which is a particle containing a molecule of U-RNA and proteins that are specific to each U ribonuclear particle (U-snRNP) or common to every U-snRNPs, is one of the numerous nuclear targets recognized by the antibodies (Abs) and CD4+ T cells from patients with systemic lupus erythematosus and lupus mice. EXEGESIS: We recently characterized a peptide from the spliceosomal protein U1-70K (sequence 131-151), which is recognized by the Abs and CD4+ T cells from lupus mice and patients. This peptide contains a conserved RNP1 motif, which is also present in other spliceosomal proteins targeted by the Abs from individuals with lupus. We further showed that peptide 131-151 containing a phosphoserine at position 140 (peptide P140) possessed tolerogenic properties in lupus mice and was recognized by the Abs and CD4+ T cells from lupus patients. CONCLUSION: Thanks to its RNP1 motif, the peptide P140 might play an important role in the initiation and perpetuation steps of the humoral and cellular immune response diversification in lupus individuals. Therapeutic and particularly immunomodulating properties of P140 peptide are being evaluated in humans (a phase III clinical trial will be undertaken in the next weeks).}, keywords = {Amino Acid Motifs, Animals, Antibodies, CD4-Positive T-Lymphocytes, Conserved Sequence, DNA, Epitopes, Haplotypes, Humans, I2CT, Immune Tolerance, Inbred MRL lpr, Inbred NZB, Lupus Erythematosus, Mice, Monneaux, Phosphoserine, Protein, Recombinant, Ribonucleoprotein, Sequence Analysis, Serine, Spliceosomes, Systemic, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{woods_influenza_2007, title = {Influenza virus-induced type I interferon leads to polyclonal B-cell activation but does not break down B-cell tolerance}, author = {Anne Woods and Fanny Monneaux and Pauline Soulas-Sprauel and Sylviane Muller and Thierry Martin and Anne-Sophie Korganow and Jean-Louis Pasquali}, doi = {10.1128/JVI.00839-07}, issn = {0022-538X}, year = {2007}, date = {2007-01-01}, journal = {Journal of Virology}, volume = {81}, number = {22}, pages = {12525--12534}, abstract = {The link between infection and autoimmunity is not yet well understood. This study was designed to evaluate if an acute viral infection known to induce type I interferon production, like influenza, can by itself be responsible for the breakdown of immune tolerance and for autoimmunity. We first tested the effects of influenza virus on B cells in vitro. We then infected different transgenic mice expressing human rheumatoid factors (RF) in the absence or in the constitutive presence of the autoantigen (human immunoglobulin G [IgG]) and young lupus-prone mice [(NZB x NZW)F(1)] with influenza virus and looked for B-cell activation. In vitro, the virus induces B-cell activation through type I interferon production by non-B cells but does not directly stimulate purified B cells. In vivo, both RF and non-RF B cells were activated in an autoantigen-independent manner. This activation was abortive since IgM and IgM-RF production levels were not increased in infected mice compared to uninfected controls, whether or not anti-influenza virus human IgG was detected and even after viral rechallenge. As in RF transgenic mice, acute viral infection of (NZB x NZW)F(1) mice induced only an abortive activation of B cells and no increase in autoantibody production compared to uninfected animals. Taken together, these experiments show that virus-induced acute type I interferon production is not able by itself to break down B-cell tolerance in both normal and autoimmune genetic backgrounds.}, keywords = {Animals, Antibody Formation, Autoantibodies, Autoantigens, Autoimmunity, B-Lymphocytes, Humans, I2CT, Immune Tolerance, Immunoglobulin M, Inbred Strains, Influenza A virus, Interferon Type I, Lymphocyte Activation, Mice, Monneaux, Rheumatoid Factor, Team-Dumortier, transgenic}, pubstate = {published}, tppubtype = {article} } @article{monneaux_importance_2007, title = {Importance of spliceosomal RNP1 motif for intermolecular Ŧ-B cell spreading and tolerance restoration in lupus}, author = {Fanny Monneaux and Véronique Parietti and Jean-Paul Briand and Sylviane Muller}, doi = {10.1186/ar2317}, issn = {1478-6362}, year = {2007}, date = {2007-01-01}, journal = {Arthritis Research & Therapy}, volume = {9}, number = {5}, pages = {R111}, abstract = {We previously demonstrated the importance of the RNP1 motif-bearing region 131-151 of the U1-70K spliceosomal protein in the intramolecular T-B spreading that occurs in MRL/lpr lupus mice. Here, we analyze the involvement of RNP1 motif in the development and prevention of naturally-occurring intermolecular T-B cell diversification. We found that MRL/lpr peripheral blood lymphocytes proliferated in response to peptides containing or corresponding exactly to the RNP1 motif of spliceosomal U1-70K, U1-A and hnRNP-A2 proteins. We also demonstrated that rabbit antibodies to peptide 131-151 cross-reacted with U1-70K, U1-A and hnRNP-A2 RNP1-peptides. These antibodies recognized the U1-70K and U1-A proteins, and also U1-C and SmD1 proteins, which are devoid of RNP1 motif. Repeated administration of phosphorylated peptide P140 into MRL/lpr mice abolished T-cell response to several peptides from the U1-70K, U1-A and SmD1 proteins without affecting antibody and T-cell responses to foreign (viral) antigen in treated mice challenged with infectious virus. These results emphasized the importance of the dominant RNP1 region, which seems to be central in the activation cascade of B and T cells reacting with spliceosomal RNP1+ and RNP1- spliceosomal proteins. The tolerogenic peptide P140, which is recognized by lupus patients' CD4+ T cells and known to protect MRL/lpr mice, is able to thwart emergence of intermolecular T-cell spreading in treated animals.}, keywords = {Amino Acid Motifs, Amino Acid Sequence, Animals, B-Lymphocytes, I2CT, Immune Tolerance, Inbred MRL lpr, Lupus Erythematosus, Mice, Molecular Sequence Data, Monneaux, Ribonucleoproteins, RNA-Binding Proteins, Saccharomyces cerevisiae Proteins, Spliceosomes, Systemic, T-Lymphocytes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of new noncoding RNAs in Listeria monocytogenes and prediction of mRNA targets}, author = { P. Mandin and F. Repoila and M. Vergassola and T. Geissmann and P. Cossart}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {3}, pages = {962-74}, abstract = {To identify noncoding RNAs (ncRNAs) in the pathogenic bacterium Listeria monocytogenes, we analyzed the intergenic regions (IGRs) of strain EGD-e by in silico-based approaches. Among the twelve ncRNAs found, nine are novel and specific to the Listeria genus, and two of these ncRNAs are expressed in a growth-dependent manner. Three of the ncRNAs are transcribed in opposite direction to overlapping open reading frames (ORFs), suggesting that they act as antisense on the corresponding mRNAs. The other ncRNA genes appear as single transcription units. One of them displays five repeats of 29 nucleotides. Five of these new ncRNAs are absent from the non-pathogenic species L. innocua, raising the possibility that they might be involved in virulence. To predict mRNA targets of the ncRNAs, we developed a computational method based on thermodynamic pairing energies and known ncRNA-mRNA hybrids. Three ncRNAs, including one of the putative antisense ncRNAs, were predicted to have more than one mRNA targets. Several of them were shown to bind efficiently to the ncRNAs suggesting that our in silico approach could be used as a general tool to search for mRNA targets of ncRNAs.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {5', Assay, Bacterial, Base, Biology, Computational, Data, DNA, Electrophoretic, Flanking, Genes, Genomics, Intergenic/chemistry, Listeria, Messenger/chemistry/*metabolism, Mobility, Molecular, monocytogenes/*genetics/metabolism, Region, RNA, ROMBY, Sequence, Shift, Untranslated/analysis/*genetics/metabolism}, pubstate = {published}, tppubtype = {article} } @article{lacerda_intracellular_2007, title = {Intracellular Trafficking of Carbon Nanotubes by Confocal Laser Scanning Microscopy}, author = {L Lacerda and G Pastorin and D Gathercole and J Buddle and M Prato and A Bianco and K Kostarelos}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adma.200790051}, doi = {10.1002/adma.200790051}, issn = {1521-4095}, year = {2007}, date = {2007-01-01}, urldate = {2020-03-31}, journal = {Advanced Materials}, volume = {19}, number = {14}, pages = {1789--1789}, keywords = {Biomedical materials, Cells, Drug delivery, fluorescence, I2CT, Nanotubes, single-walled, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @incollection{gubb_drosophila_2007, title = {Drosophila Serpins: Regulatory Cascades in Innate Immunity and Morphogenesis}, author = {David Gubb and Andrew S Robertson and Laurent Troxler and Jean-Marc Reichhart}, year = {2007}, date = {2007-01-01}, booktitle = {Molecular and Cellular Aspects of the Serpinopathies and Disorders in Serpin Activity}, pages = {207--227}, publisher = {Silverman GA and Lomas DA}, address = {London UK}, edition = {World Scientific Pub.}, keywords = {bioinformatic, innate immunity, M3i, Morphogenesis, regulatory Cascades, reichhart, Serpins}, pubstate = {published}, tppubtype = {incollection} } @article{nehme_model_2007b, title = {A model of bacterial intestinal infections in Drosophila melanogaster}, author = {Nadine T Nehme and Samuel Liégeois and Beatrix Kele and Philippe Giammarinaro and Elizabeth Pradel and Jules A Hoffmann and Jonathan J Ewbank and Dominique Ferrandon}, doi = {10.1371/journal.ppat.0030173}, issn = {1553-7374}, year = {2007}, date = {2007-01-01}, journal = {PLoS Pathog.}, volume = {3}, number = {11}, pages = {e173}, abstract = {Serratia marcescens is an entomopathogenic bacterium that opportunistically infects a wide range of hosts, including humans. In a model of septic injury, if directly introduced into the body cavity of Drosophila, this pathogen is insensitive to the host's systemic immune response and kills flies in a day. We find that S. marcescens resistance to the Drosophila immune deficiency (imd)-mediated humoral response requires the bacterial lipopolysaccharide O-antigen. If ingested by Drosophila, bacteria cross the gut and penetrate the body cavity. During this passage, the bacteria can be observed within the cells of the intestinal epithelium. In such an oral infection model, the flies succumb to infection only after 6 days. We demonstrate that two complementary host defense mechanisms act together against such food-borne infection: an antimicrobial response in the intestine that is regulated by the imd pathway and phagocytosis by hemocytes of bacteria that have escaped into the hemolymph. Interestingly, bacteria present in the hemolymph elicit a systemic immune response only when phagocytosis is blocked. Our observations support a model wherein peptidoglycan fragments released during bacterial growth activate the imd pathway and do not back a proposed role for phagocytosis in the immune activation of the fat body. Thanks to the genetic tools available in both host and pathogen, the molecular dissection of the interactions between S. marcescens and Drosophila will provide a useful paradigm for deciphering intestinal pathogenesis.}, keywords = {Animal, Animals, Disease Models, Electron, ferrandon, fluorescence, Hemolymph, hoffmann, Host-Pathogen Interactions, Immunohistochemistry, Intestines, M3i, Microscopy, Reverse Transcriptase Polymerase Chain Reaction, Serratia Infections, Serratia marcescens, Transmission}, pubstate = {published}, tppubtype = {article} } @article{croker_atp-sensitive_2007, title = {ATP-sensitive potassium channels mediate survival during infection in mammals and insects}, author = {Ben Croker and Karine Crozat and Michael Berger and Yu Xia and Sosathya Sovath and Lana Schaffer and Ioannis Eleftherianos and Jean-Luc Imler and Bruce Beutler}, doi = {10.1038/ng.2007.25}, issn = {1546-1718}, year = {2007}, date = {2007-01-01}, journal = {Nature Genetics}, volume = {39}, number = {12}, pages = {1453--1460}, abstract = {Specific homeostatic mechanisms confer stability in innate immune responses, preventing injury or death from infection. Here we identify, from a screen of N-ethyl-N-nitrosourea-mutagenized mice, a mutation causing both profound susceptibility to infection by mouse cytomegalovirus and approximately 20,000-fold sensitization to lipopolysaccharide (LPS), poly(I.C) and immunostimulatory (CpG) DNA. The LPS hypersensitivity phenotype is not suppressed by mutations in Myd88, Trif, Tnf, Tnfrsf1a, Ifnb, Ifng or Stat1, genes contributing to LPS responses, and results from an abnormality extrinsic to hematopoietic cells. The phenotype is due to a null allele of Kcnj8, encoding Kir6.1, a protein that combines with SUR2 to form an ATP-sensitive potassium channel (K(ATP)) expressed in coronary artery smooth muscle and endothelial cells. In Drosophila melanogaster, suppression of dSUR by RNA interference similarly causes hypersensitivity to infection by flock house virus. Thus, K(ATP) evolved to serve a homeostatic function during infection, and in mammals it prevents coronary artery vasoconstriction induced by cytokines dependent on TLR and/or MDA5 immunoreceptors.}, keywords = {Animals, ATP-Binding Cassette Transporters, Cloning, Coronary Vessels, Crosses, Ethylnitrosourea, Genetic, Homozygote, imler, infection, Inwardly Rectifying, KATP Channels, Lipopolysaccharides, M3i, Mice, Molecular, Mutagenesis, Potassium Channels, Sulfonylurea Receptors}, pubstate = {published}, tppubtype = {article} } @article{galiana-arnoux_immunite_2007, title = {Immunité antivirale chez la drosophile}, author = {Delphine Galiana-Arnoux and Safia Deddouche and Jean-Luc Imler}, url = {http://www.biologie-journal.org/10.1051/jbio:2007906}, doi = {10.1051/jbio:2007906}, issn = {1295-0661}, year = {2007}, date = {2007-01-01}, urldate = {2015-11-26}, journal = {Journal de la Société de Biologie}, volume = {201}, number = {4}, pages = {359--365}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{galiana-arnoux_[antiviral_2007, title = {[Antiviral immunity in drosophila]}, author = {Delphine Galiana-Arnoux and Safia Deddouche and Jean-Luc Imler}, doi = {10.1051/jbio:2007906}, issn = {1295-0661}, year = {2007}, date = {2007-01-01}, journal = {Journal De La Société De Biologie}, volume = {201}, number = {4}, pages = {359--365}, abstract = {Viral diseases represent a constant threat and an important cause of mortality worldwide. We have developed a model to study the response to RNA virus infection in the fruit-fly drosophila. This insect is a good model to study the genetic bases of innate immunity, which constitutes the first level of host-defense in animals. We have shown that viral infection in drosophila triggers a response different from that to bacterial or fungal infections. Our data at this stage point to the existence of at least two types of antiviral defense mechanisms. On one hand, viral infection triggers a JAK-STAT dependent transcriptional response that leads to the expression of antiviral molecules that remain to be characterized. On the other hand, viral RNAs are recognized by Dicer-2 and degraded in siRNAs, thus inducing RNA interference and degradation of viral RNAs. Strikingly, the drosophila antiviral response evokes by some aspects the interferon response in mammals (JAK-STAT pathway) and antiviral defenses in plants (RNA interference).}, keywords = {imler, M3i}, pubstate = {published}, tppubtype = {article} } @article{wang-sattler_mosaic_2007, title = {Mosaic genome architecture of the Anopheles gambiae species complex}, author = {Rui Wang-Sattler and Stephanie A Blandin and Ye Ning and Claudia Blass and Guimogo Dolo and Yeya T Touré and Alessandra delle Torre and Gregory C Lanzaro and Lars M Steinmetz and Fotis C Kafatos and Liangbiao Zheng}, doi = {10.1371/journal.pone.0001249}, issn = {1932-6203}, year = {2007}, date = {2007-01-01}, journal = {PLoS ONE}, volume = {2}, number = {11}, pages = {e1249}, abstract = {BACKGROUND: Attempts over the last three decades to reconstruct the phylogenetic history of the Anopheles gambiae species complex have been important for developing better strategies to control malaria transmission. METHODOLOGY: We used fingerprint genotyping data from 414 field-collected female mosquitoes at 42 microsatellite loci to infer the evolutionary relationships of four species in the A. gambiae complex, the two major malaria vectors A. gambiae sensu stricto (A. gambiae s.s.) and A. arabiensis, as well as two minor vectors, A. merus and A. melas. PRINCIPAL FINDINGS: We identify six taxonomic units, including a clear separation of West and East Africa A. gambiae s.s. S molecular forms. We show that the phylogenetic relationships vary widely between different genomic regions, thus demonstrating the mosaic nature of the genome of these species. The two major malaria vectors are closely related and closer to A. merus than to A. melas at the genome-wide level, which is also true if only autosomes are considered. However, within the Xag inversion region of the X chromosome, the M and two S molecular forms are most similar to A. merus. Near the X centromere, outside the Xag region, the two S forms are highly dissimilar to the other taxa. Furthermore, our data suggest that the centromeric region of chromosome 3 is a strong discriminator between the major and minor malaria vectors. CONCLUSIONS: Although further studies are needed to elucidate the basis of the phylogenetic variation among the different regions of the genome, the preponderance of sympatric admixtures among taxa strongly favor introgression of different genomic regions between species, rather than lineage sorting of ancestral polymorphism, as a possible mechanism.}, keywords = {Animals, Anopheles gambiae, Artificial, Bacterial, Biological Evolution, blandin, Chromosomes, Female, Genetic Markers, Genetic Variation, Genome, M3i, Microsatellite Repeats, Mosaicism}, pubstate = {published}, tppubtype = {article} } @article{govin_pericentric_2007, title = {Pericentric heterochromatin reprogramming by new histone variants during mouse spermiogenesis.}, author = {Jérôme Govin and Emmanuelle Escoffier and Sophie Rousseaux and Lauriane Kuhn and Myriam Ferro and Julien Thévenon and Raffaella Catena and Irwin Davidson and Jérôme Garin and Saadi Khochbin and Cécile Caron}, doi = {10.1083/jcb.200604141}, issn = {0021-9525 1540-8140 0021-9525}, year = {2007}, date = {2007-01-01}, journal = {The Journal of cell biology}, volume = {176}, number = {3}, pages = {283--294}, abstract = {During male germ cell postmeiotic maturation, dramatic chromatin reorganization occurs, which is driven by completely unknown mechanisms. For the first time, we describe a specific reprogramming of mouse pericentric heterochromatin. Initiated when histones undergo global acetylation in early elongating spermatids, this process leads to the establishment of new DNA packaging structures organizing the pericentric regions in condensing spermatids. Five new histone variants were discovered, which are expressed in late spermiogenic cells. Two of them, which we named H2AL1 and H2AL2, specifically mark the pericentric regions in condensing spermatids and participate in the formation of new nucleoprotein structures. Moreover, our investigations also suggest that TH2B, an already identified testis-specific H2B variant of unknown function, could provide a platform for the structural transitions accompanying the incorporation of these new histone variants.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{mueller_critical_2007, title = {Critical role of monocytes to support normal B cell and diffuse large B cell lymphoma survival and proliferation}, author = {C G Mueller and C Boix and W H Kwan and C Daussy and E Fournier and W H Fridman and T J Molina}, year = {2007}, date = {2007-01-01}, journal = {Journal of Leukocyte Biology}, volume = {82}, number = {0741-5400 (Print)}, pages = {567--575}, abstract = {Large B cell lymphomas can comprise numerous CD14+ cells in the tumor stroma, which raises the question of whether monocytes can support B cell survival and proliferation. We show that the coculture of monocytes with B cells from peripheral blood or from diffuse large B cell lymphoma enabled prolonged B cell survival. Under these conditions, diffuse large lymphoma B cells proliferated, and addition of B cell-activating factor of the TNF family (BAFF) and IL-2 enhanced cell division. Monocytes and dendritic cells (DC) had similar antiapoptotic activity on healthy B cells but displayed differences with respect to B cell proliferation. Monocytes and cord blood-derived CD14+ cells promoted B cell proliferation in the presence of an anti-CD40 stimulus, whereas DC supported B cell proliferation when activated through the BCR. DC and CD14+ cells were able to induce plasmocyte differentiation. When B cells were activated via the BCR or CD40, they released the leukocyte attractant CCL5, and this chemokine is one of the main chemokines expressed in diffuse large B cell lymphoma. The data support the notion that large B cell lymphoma recruit monocytes via CCL5 to support B cell survival and proliferation}, keywords = {Activation, Antigen, Antigens, B CELL ACTIVATION, B CELLS, B-Cell, B-Cell Activation Factor Receptor, B-Lymphocytes, Biological, BLOOD, CC, CD14, CD40, Cell Division, Cell Proliferation, Cell Survival, Chemokine CCL5, chemokines, Coculture, cytology, Dendritic Cells, Differentiation, Diffuse, Enzyme-Linked Immunosorbent Assay, Flow Cytometry, Human, Humans, IL-2, Immunoenzyme Techniques, Interleukin-2, Large B-Cell, Lymph Nodes, LYMPHOMA, metabolism, monocyte, Monocytes, Myeloid Cells, pathology, Proliferation, Protein, Receptor, Reverse Transcriptase Polymerase Chain Reaction, survival, Team-Mueller, tumor, Tumor Markers}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of new noncoding RNAs in Listeria monocytogenes and prediction of mRNA targets}, author = {P Mandin and F Repoila and M Vergassola and T Geissmann and P Cossart}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17259222}, isbn = {17259222}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {3}, pages = {962-74}, abstract = {To identify noncoding RNAs (ncRNAs) in the pathogenic bacterium Listeria monocytogenes, we analyzed the intergenic regions (IGRs) of strain EGD-e by in silico-based approaches. Among the twelve ncRNAs found, nine are novel and specific to the Listeria genus, and two of these ncRNAs are expressed in a growth-dependent manner. Three of the ncRNAs are transcribed in opposite direction to overlapping open reading frames (ORFs), suggesting that they act as antisense on the corresponding mRNAs. The other ncRNA genes appear as single transcription units. One of them displays five repeats of 29 nucleotides. Five of these new ncRNAs are absent from the non-pathogenic species L. innocua, raising the possibility that they might be involved in virulence. To predict mRNA targets of the ncRNAs, we developed a computational method based on thermodynamic pairing energies and known ncRNA-mRNA hybrids. Three ncRNAs, including one of the putative antisense ncRNAs, were predicted to have more than one mRNA targets. Several of them were shown to bind efficiently to the ncRNAs suggesting that our in silico approach could be used as a general tool to search for mRNA targets of ncRNAs.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Bacterial Genomics Listeria monocytogenes/*genetics/metabolism Molecular Sequence Data RNA, Intergenic/chemistry Electrophoretic Mobility Shift Assay Genes, Messenger/chemistry/*metabolism RNA, ROMBY 5' Flanking Region Base Sequence Computational Biology DNA, Untranslated/analysis/*genetics/metabolism}, pubstate = {published}, tppubtype = {article} } @article{, title = {Anopheles gambiae miRNAs as actors of defence reaction against Plasmodium invasion}, author = { F. Winter and S. Edaye and A. Huttenhofer and C. Brunel}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {20}, pages = {6953-62}, abstract = {The path Plasmodium takes across the Anopheles midgut constitutes the major bottleneck during the malaria transmission cycle. In the present study, using a combination of shot-gun cloning and bioinformatic analysis, we have identified 18 miRNAs from Anopheles gambiae including three miRNAs unique to mosquito. Twelve of them are expressed ubiquitously across the body, independently of gender, while the other six exhibited an expression pattern restricted to the digestive system. Strikingly, the expression patterns of four miRNAs, including the three unique to mosquito, are affected by the presence of Plasmodium. We also show that knocking down Dicer1 and Ago1 mRNAs led to an increased sensitivity to Plasmodium infection. Altogether, these data support an involvement of miRNAs as new layers in the regulation of Anopheles defence reaction.}, note = {1362-4962 (Electronic) 0305-1048 (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {*Gene, Animals, Anopheles, BRUNEL, Digestive, Expression, Female, gambiae/*genetics/*immunology/parasitology, Gene, III/genetics, Library, Male, MicroRNAs/*immunology, Plasmodium/*immunology, Profiling, Ribonuclease, silencing, System/immunology/metabolism/parasitology}, pubstate = {published}, tppubtype = {article} } @article{, title = {A structure-based approach for targeting the HIV-1 genomic RNA dimerization initiation site}, author = {E Ennifar and J C Paillart and S Bernacchi and P Walter and P Pale and J L Decout and R Marquet and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17434658}, isbn = {17434658}, year = {2007}, date = {2007-01-01}, journal = {Biochimie}, volume = {89}, number = {10}, pages = {1195-1203}, abstract = {Dimerization of the genomic RNA is an important step of the HIV-1 replication cycle. The Dimerization Initiation Site (DIS) promotes dimerization of the viral genome by forming a loop-loop complex between two DIS hairpins. Crystal structures of the DIS loop-loop complex revealed an unexpected and strong similitude with the bacterial 16S ribosomal aminoacyl-tRNA site (A site), which is the target of aminoglycoside antibiotics. As a consequence of these structural and sequence similarities, the HIV-1 DIS also binds some aminoglycosides, not only in vitro, but also ex vivo, in lymphoid cells and in viral particles. Crystal structures of the DIS loop-loop in complex with several aminoglycoside antibiotics provide a detailed-view of the DIS/drug interaction and reveal some hints about possible modifications to increase the drug affinity and/or specificity.}, keywords = {ENNIFAR, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Vif is a RNA chaperone that could temporally regulate RNA dimerization and the early steps of HIV-1 reverse transcription}, author = {S Henriet and L Sinck and G Bec and R J Gorelick and R Marquet and J C Paillart}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17660191}, isbn = {17660191}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {15}, pages = {5141-5153}, abstract = {HIV-1 Vif (viral infectivity factor) is associated with the assembly complexes and packaged at low level into the viral particles, and is essential for viral replication in non-permissive cells. Viral particles produced in the absence of Vif exhibit structural defects and are defective in the early steps of reverse transcription. Here, we show that Vif is able to anneal primer tRNA(Lys3) to the viral RNA, to decrease pausing of reverse transcriptase during (-) strand strong-stop DNA synthesis, and to promote the first strand transfer. Vif also stimulates formation of loose HIV-1 genomic RNA dimers. These results indicate that Vif is a bona fide RNA chaperone. We next studied the effects of Vif in the presence of HIV-1 NCp, which is a well-established RNA chaperone. Vif inhibits NCp-mediated formation of tight RNA dimers and hybridization of tRNA(Lys3), while it has little effects on NCp-mediated strand transfer and it collaborates with nucleocapsid (NC) to increase RT processivity. Thus, Vif might negatively regulate NC-assisted maturation of the RNA dimer and early steps of reverse transcription in the assembly complexes, but these inhibitory effects would be relieved after viral budding, thanks to the limited packaging of Vif in the virions.}, note = {1362-4962 (Electronic) Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't}, keywords = {Amino Acyl/metabolism RNA, Capsid Proteins/metabolism DNA, gag/metabolism Gene Products, Human Immunodeficiency Virus, Human Immunodeficiency Virus vif Gene Products, MARQUET, PAILLART, Single-Stranded/biosynthesis Dimerization Gene Products, Transfer, Unité ARN, vif/*metabolism HIV-1/*genetics Molecular Chaperones/*metabolism RNA, Viral/*metabolism *Reverse Transcription Viral Proteins/metabolism gag Gene Products}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV controls the selective packaging of genomic, spliced viral and cellular RNAs into virions through different mechanisms}, author = {L Houzet and J C Paillart and F Smagulova and S Maurel and Z Morichaud and R Marquet and M Mougel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17426127}, isbn = {17426127}, year = {2007}, date = {2007-01-01}, journal = {Nucleic Acids Res}, volume = {35}, number = {8}, pages = {2695-2704}, abstract = {In addition to genomic RNA, HIV-1 particles package cellular and spliced viral RNAs. In order to determine the encapsidation mechanisms of these RNAs, we determined the packaging efficiencies and specificities of genomic RNA, singly and fully spliced HIV mRNAs and different host RNAs species: 7SL RNA, U6 snRNA and GAPDH mRNA using RT-QPCR. Except GAPDH mRNA, all RNAs are selectively encapsidated. Singly spliced RNAs, harboring the Rev-responsible element, and fully spliced viral RNAs, which do not contain this motif, are enriched in virions to similar levels, even though they are exported from the nucleus by different routes. Deletions of key motifs (SL1 and/or SL3) of the packaging signal of genomic RNA indicate that HIV and host RNAs are encapsidated through independent mechanisms, while genomic and spliced viral RNA compete for the same trans-acting factor due to the presence of the 5' common exon containing the TAR, poly(A) and U5-PBS hairpins. Surprisingly, the RNA dimerization initiation site (DIS/SL1) appears to be the main packaging determinant of genomic RNA, but is not involved in packaging of spliced viral RNAs, suggesting a functional interaction with intronic sequences. Active and selective packaging of host and spliced viral RNAs provide new potential functions to these RNAs in the early stages of the virus life cycle.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA and DNA binding properties of HIV-1 Vif protein: a fluorescence study}, author = {S Bernacchi and S Henriet and P Dumas and J C Paillart and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17609216}, isbn = {17609216}, year = {2007}, date = {2007-01-01}, journal = {J Biol Chem}, volume = {282}, number = {36}, pages = {26361-26368}, abstract = {The HIV-1 viral infectivity factor (Vif) is a small basic protein essential for viral fitness and pathogenicity. Some "non-permissive" cell lines cannot sustain replication of Vif(-) HIV-1 virions. In these cells, Vif counteracts the natural antiretroviral activity of the DNA-editing enzymes APOBEC3G/3F. Moreover, Vif is packaged into viral particles through a strong interaction with genomic RNA in viral nucleoprotein complexes. To gain insights into determinants of this binding process, we performed the first characterization of Vif/nucleic acid interactions using Vif intrinsic fluorescence. We determined the affinity of Vif for RNA fragments corresponding to various regions of the HIV-1 genome. Our results demonstrated preferential and moderately cooperative binding for RNAs corresponding to the 5'-untranslated region of HIV-1 (5'-untranslated region) and gag (cooperativity parameter omega approximately 65-80, and K(d) = 45-55 nM). In addition, fluorescence spectroscopy allowed us to point out the TAR apical loop and a short region in gag as primary strong affinity binding sites (K(d) = 9.5-14 nM). Interestingly, beside its RNA binding properties, the Vif protein can also bind the corresponding DNA oligonucleotides and their complementary counterparts with an affinity similar to the one observed for the RNA sequences, while other DNA sequences displayed reduced affinity. Taken together, our results suggest that Vif binding to RNA and DNA offers several non-exclusive ways to counteract APOBEC3G/3F factors, in addition to the well documented Vif-induced degradation by the proteasome and to the Vif-mediated repression of translation of these antiviral factors.}, keywords = {5' Untranslated Regions/genetics/immunology/metabolism Binding Sites/physiology Cytidine Deaminase Cytosine Deaminase/immunology/metabolism DNA, gag/genetics/immunology/metabolism Gene Products, Human Immunodeficiency Virus, MARQUET, Natural/physiology Nucleoside Deaminases/immunology/metabolism Oligonucleotides/genetics/immunology/metabolism Protein Binding/physiology Protein Biosynthesis/physiology RNA, PAILLART, Unité ARN, vif/genetics/immunology/*metabolism Genome, Viral/genetics/immunology/*metabolism DNA-Binding Proteins/genetics/immunology/*metabolism Gene Products, Viral/genetics/immunology/*metabolism RNA-Binding Proteins/genetics/immunology/*metabolism Repressor Proteins/genetics/immunology/*metabolism vif Gene Products, Viral/physiology HIV Long Terminal Repeat/physiology HIV-1/genetics/immunology/*metabolism/pathogenicity Humans Immunity}, pubstate = {published}, tppubtype = {article} } @article{, title = {Synthesis of a neamine dimer targeting the dimerization initiation site of HIV-1 RNA}, author = {A Bodlenner and A Alix and J M Weibel and P Pale and E Ennifar and J C Paillart and P Walter and R Marquet and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17915882}, isbn = {17915882}, year = {2007}, date = {2007-01-01}, journal = {Org Lett}, volume = {9}, number = {22}, pages = {4415-4418}, abstract = {A neamine dimer designed to bind to a specific sequence of HIV-1 RNA has been synthesized. Starting from neomycin B (1), a five-step synthesis efficiently provided a key protected neamine monomer 6 (28%). From the latter, coupling reactions with activated diacids gave dimers. After deprotection, a neamine dimer was obtained as the hexachlorohydrate salt 15 with 13% overall yield over nine steps.}, keywords = {ENNIFAR, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{klumpp_multifunctionalised_2007, title = {Multifunctionalised cationic fullerene adducts for gene transfer: design, synthesis and DNA complexation}, author = {Cédric Klumpp and Lara Lacerda and Olivier Chaloin and Tatiana Da Ros and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, doi = {10.1039/b708435h}, issn = {1359-7345}, year = {2007}, date = {2007-01-01}, journal = {Chemical Communications (Cambridge, England)}, number = {36}, pages = {3762--3764}, abstract = {Cationic poly-N,N-dimethylfulleropyrrolidinium derivatives have been designed and synthesised to complex plasmid DNA for gene delivery.}, keywords = {DNA, Electrophoresis, Fullerenes, Gene Transfer Techniques, I2CT, Molecular Structure, Plasmids, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structured mRNAs regulate translation initiation by binding to the platform of the ribosome}, author = {S Marzi and A G Myasnikov and A Serganov and C Ehresmann and P Romby and M Yusupov and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17889647}, isbn = {17889647}, year = {2007}, date = {2007-01-01}, journal = {Cell}, volume = {130}, number = {6}, pages = {1019-31}, abstract = {Gene expression can be regulated at the level of initiation of protein biosynthesis via structural elements present at the 5' untranslated region of mRNAs. These folded mRNA segments may bind to the ribosome, thus blocking translation until the mRNA unfolds. Here, we report a series of cryo-electron microscopy snapshots of ribosomal complexes directly visualizing either the mRNA structure blocked by repressor protein S15 or the unfolded, active mRNA. In the stalled state, the folded mRNA prevents the start codon from reaching the peptidyl-tRNA (P) site inside the ribosome. Upon repressor release, the mRNA unfolds and moves into the mRNA channel allowing translation initiation. A comparative structure and sequence analysis suggests the existence of a universal stand-by site on the ribosome (the 30S platform) dedicated for binding regulatory 5' mRNA elements. Different types of mRNA structures may be accommodated during translation preinitiation and regulate gene expression by transiently stalling the ribosome.}, note = {0092-8674 (Print) Comparative Study Journal Article Research Support, Non-U.S. Gov't}, keywords = {5' Untranslated Regions Amino Acid Sequence Base Sequence Binding Sites Cryoelectron Microscopy Escherichia coli/*genetics/metabolism Escherichia coli Proteins/chemistry/genetics/*metabolism *Gene Expression Regulation, Amino Acid Sequence Homology, Bacterial Models, Bacterial/chemistry/*metabolism RNA, Messenger/*metabolism RNA, Molecular Molecular Sequence Data Mutation Nucleic Acid Conformation *Peptide Chain Initiation, Nucleic Acid Structural Homology, Protein Time Factors, Ribonucleic Acid Ribosomal Proteins/chemistry/genetics/*metabolism Ribosomes/chemistry/*metabolism/ultrastructure Sequence Homology, ROMBY, Transfer/metabolism Regulatory Sequences, Translational Protein Binding Protein Conformation RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Twisting of mRNA reversibly blocks its translation by the ribosome]}, author = {S Marzi and P Romby and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17937901}, isbn = {17937901}, year = {2007}, date = {2007-01-01}, journal = {Med Sci (Paris)}, volume = {23}, number = {10}, pages = {881-3}, note = {0767-0974 (Print) News}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Functional analysis of the translation factor aIF2/5B in the thermophilic archaeon Sulfolobus solfataricus}, author = {E Maone and M Di Stefano and A Berardi and D Benelli and S Marzi and A La Teana and P Londei}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17608795}, doi = {10.1111/j.1365-2958.2007.05820.x}, isbn = {17608795}, year = {2007}, date = {2007-01-01}, journal = {Mol Microbiol}, volume = {65}, number = {3}, pages = {700-13}, abstract = {The protein IF2/eIF5B is one of the few translation initiation factors shared by all three primary domains of life (bacteria, archaea, eukarya). Despite its phylogenetic conservation, the factor is known to present marked functional divergences in the bacteria and the eukarya. In this work, the function in translation of the archaeal homologue (aIF2/5B) has been analysed in detail for the first time using a variety of in vitro assays. The results revealed that the protein is a ribosome-dependent GTPase which strongly stimulates the binding of initiator tRNA to the ribosomes even in the absence of other factors. In agreement with this finding, aIF2/5B enhances the translation of both leadered and leaderless mRNAs when expressed in a cell-free protein-synthesizing system. Moreover, the degree of functional conservation of the IF2-like factors in the archaeal and bacterial lineages was investigated by analysing the behaviour of 'chimeric' proteins produced by swapping domains between the Sulfolobus solfataricus aIF2/5B factor and the IF2 protein of the thermophilic bacterium Bacillus stearothermophilus. Beside evidencing similarities and differences between the archaeal and bacterial factors, these experiments have provided insight into the common role played by the IF2/5B proteins in all extant cells.}, note = {0950-382X (Print) 0950-382X (Linking) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Archaeal Hydrolysis Peptide Chain Initiation, Met/metabolism Recombinant Fusion Proteins/metabolism Ribosomes/metabolism Sulfolobus solfataricus/genetics/*metabolism, Molecular Conserved Sequence GTP Phosphohydrolases/metabolism Gene Expression Genes, ROMBY, Secondary RNA, Transfer, Translational Peptide Initiation Factors/chemistry/isolation & purification/*metabolism Protein Binding *Protein Biosynthesis Protein Structure, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{lacerda_carbon_2006, title = {Carbon nanotubes as nanomedicines: from toxicology to pharmacology}, author = {Lara Lacerda and Alberto Bianco and Maurizio Prato and Kostas Kostarelos}, doi = {10.1016/j.addr.2006.09.015}, issn = {0169-409X}, year = {2006}, date = {2006-12-01}, journal = {Advanced Drug Delivery Reviews}, volume = {58}, number = {14}, pages = {1460--1470}, abstract = {Various biomedical applications of carbon nanotubes have been proposed in the last few years leading to the emergence of a new field in diagnostics and therapeutics. Most of these applications will involve the administration or implantation of carbon nanotubes and their matrices into patients. The toxicological and pharmacological profile of such carbon nanotube systems developed as nanomedicines will have to be determined prior to any clinical studies undertaken. This review brings together all the toxicological and pharmacological in vivo studies that have been carried out using carbon nanotubes, to offer the first summary of the state-of-the-art in the pharmaceutical development of carbon nanotubes on the road to becoming viable and effective nanomedicines.}, keywords = {Animals, carbon, Humans, I2CT, Nanomedicine, Nanotubes, Pharmacokinetics, Pharmacology, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{gottar_dual_2006, title = {Dual detection of fungal infections in Drosophila via recognition of glucans and sensing of virulence factors}, author = {Marie Gottar and Vanessa Gobert and Alexey A Matskevich and Jean-Marc Reichhart and Chengshu Wang and Tariq M Butt and Marcia Belvin and Jules A Hoffmann and Dominique Ferrandon}, doi = {10.1016/j.cell.2006.10.046}, issn = {0092-8674}, year = {2006}, date = {2006-12-01}, journal = {Cell}, volume = {127}, number = {7}, pages = {1425--1437}, abstract = {The Drosophila immune system discriminates between various types of infections and activates appropriate signal transduction pathways to combat the invading microorganisms. The Toll pathway is required for the host response against fungal and most Gram-positive bacterial infections. The sensing of Gram-positive bacteria is mediated by the pattern recognition receptors PGRP-SA and GNBP1 that cooperate to detect the presence of infections in the host. Here, we report that GNBP3 is a pattern recognition receptor that is required for the detection of fungal cell wall components. Strikingly, we find that there is a second, parallel pathway acting jointly with GNBP3. The Drosophila Persephone protease activates the Toll pathway when proteolytically matured by the secreted fungal virulence factor PR1. Thus, the detection of fungal infections in Drosophila relies both on the recognition of invariant microbial patterns and on monitoring the effects of virulence factors on the host.}, keywords = {Animals, Antibody Formation, Beauveria, Candida albicans, Carrier Proteins, Cellular, ferrandon, Glucans, hoffmann, Immunity, Immunological, M3i, Metarhizium, Models, Polysaccharides, reichhart, Serine Endopeptidases, Signal Transduction, Virulence Factors}, pubstate = {published}, tppubtype = {article} } @article{shiao_fz2_2006, title = {Fz2 and cdc42 mediate melanization and actin polymerization but are dispensable for Plasmodium killing in the mosquito midgut}, author = {Shin-Hong Shiao and Miranda M A Whitten and Daniel Zachary and Jules A Hoffmann and Elena A Levashina}, doi = {10.1371/journal.ppat.0020133}, issn = {1553-7374}, year = {2006}, date = {2006-12-01}, journal = {PLoS Pathog.}, volume = {2}, number = {12}, pages = {e133}, abstract = {The midgut epithelium of the mosquito malaria vector Anopheles is a hostile environment for Plasmodium, with most parasites succumbing to host defenses. This study addresses morphological and ultrastructural features associated with Plasmodium berghei ookinete invasion in Anopheles gambiae midguts to define the sites and possible mechanisms of parasite killing. We show by transmission electron microscopy and immunofluorescence that the majority of ookinetes are killed in the extracellular space. Dead or dying ookinetes are surrounded by a polymerized actin zone formed within the basal cytoplasm of adjacent host epithelial cells. In refractory strain mosquitoes, we found that formation of this zone is strongly linked to prophenoloxidase activation leading to melanization. Furthermore, we identify two factors controlling both phenomena: the transmembrane receptor frizzled-2 and the guanosine triphosphate-binding protein cell division cycle 42. However, the disruption of actin polymerization and melanization by double-stranded RNA inhibition did not affect ookinete survival. Our results separate the mechanisms of parasite killing from subsequent reactions manifested by actin polymerization and prophenoloxidase activation in the A. gambiae-P. berghei model. These latter processes are reminiscent of wound healing in other organisms, and we propose that they represent a form of wound-healing response directed towards a moribund ookinete, which is perceived as damaged tissue.}, keywords = {Actins, Animals, Anopheles, Carrier Proteins, cdc42 GTP-Binding Protein, Double-Stranded, Electron, Frizzled Receptors, Gastrointestinal Tract, hoffmann, Host-Parasite Interactions, Immunity, Innate, Insect Vectors, Intestinal Mucosa, M3i, Melanins, Microarray Analysis, Microscopy, Plasmodium berghei, Polymers, Protozoan, RNA, scanning, telomerase}, pubstate = {published}, tppubtype = {article} } @article{govin_post-meiotic_2006, title = {Post-meiotic shifts in HSPA2/HSP70.2 chaperone activity during mouse spermatogenesis.}, author = {Jérôme Govin and Cécile Caron and Emmanuelle Escoffier and Myriam Ferro and Lauriane Kuhn and Sophie Rousseaux and Edward M Eddy and Jérôme Garin and Saadi Khochbin}, doi = {10.1074/jbc.M608147200}, issn = {0021-9258 1083-351X 0021-9258}, year = {2006}, date = {2006-12-01}, journal = {The Journal of biological chemistry}, volume = {281}, number = {49}, pages = {37888--37892}, abstract = {HSPA2 (formerly HSP70.2) is a testis-specific member of the HSP70 family known to play a critical role in the completion of meiosis during male germ cell differentiation. Although abundantly present in post-meiotic cells, its function during spermiogenesis remained obscure. Here, using a global proteomic approach to identify genome-organizing proteins in condensing spermatids, we discovered an unexpected role for HSPA2, which acquires new functions and becomes tightly associated with major spermatid DNA-packaging proteins, transition proteins 1 and 2. Hence, HSPA2 is identified here as the first transition protein chaperone, and these data shed a new light on the yet totally unknown process of genome-condensing structure assembly in spermatids.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{flacher_human_2006, title = {Human Langerhans cells express a specific TLR profile and differentially respond to viruses and Gram-positive bacteria}, author = {Vincent Flacher and Marielle Bouschbacher and Estelle Verronèse and Catherine Massacrier and Vanja Sisirak and Odile Berthier-Vergnes and Blandine de Saint-Vis and Christophe Caux and Colette Dezutter-Dambuyant and Serge Lebecque and Jenny Valladeau}, doi = {10.4049/jimmunol.177.11.7959}, issn = {0022-1767}, year = {2006}, date = {2006-12-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {177}, number = {11}, pages = {7959--7967}, abstract = {Dendritic cells (DC) are APCs essential for the development of primary immune responses. In pluristratified epithelia, Langerhans cells (LC) are a critical subset of DC which take up Ags and migrate toward lymph nodes upon inflammatory stimuli. TLR allow detection of pathogen-associated molecular patterns (PAMP) by different DC subsets. The repertoire of TLR expressed by human LC is uncharacterized and their ability to directly respond to PAMP has not been systematically investigated. In this study, we show for the first time that freshly purified LC from human skin express mRNA encoding TLR1, TLR2, TLR3, TLR5, TLR6 and TLR10. In addition, keratinocytes ex vivo display TLR1-5, TLR7, and TLR10. Accordingly, highly enriched immature LC efficiently respond to TLR2 agonists peptidoglycan and lipoteichoic acid from Gram-positive bacteria, and to dsRNA which engages TLR3. In contrast, LC do not directly sense TLR7/8 ligands and LPS from Gram-negative bacteria, which signals through TLR4. TLR engagement also results in cytokine production, with marked differences depending on the PAMP detected. TLR2 and TLR3 ligands increase IL-6 and IL-8 production, while dsRNA alone stimulates TNF-alpha release. Strikingly, only peptidoglycan triggers IL-10 secretion, thereby suggesting a specific function in tolerance to commensal Gram-positive bacteria. However, LC do not produce IL-12p70 or type I IFNs. In conclusion, human LC are equipped with TLR that enable direct detection of PAMP from viruses and Gram-positive bacteria, subsequent phenotypic maturation, and differential cytokine production. This implies a significant role for LC in the control of skin immune responses.}, keywords = {bacteria, Double-Stranded, Gram-Positive Bacteria, Human, Humans, Interleukin-6, Interleukin-8, Langerhans Cells, Reverse Transcriptase Polymerase Chain Reaction, RNA, Skin, Team-Mueller, TLR4, TLR7, Toll-Like Receptors, Tumor Necrosis Factor-alpha, viruses}, pubstate = {published}, tppubtype = {article} } @article{hauquier_carbon_2006, title = {Carbon nanotube-functionalized silicon surfaces with efficient redox communication}, author = {Fanny Hauquier and Giorgia Pastorin and Philippe Hapiot and Maurizio Prato and Alberto Bianco and Bruno Fabre}, doi = {10.1039/b610559a}, issn = {1359-7345}, year = {2006}, date = {2006-11-01}, journal = {Chemical Communications (Cambridge, England)}, number = {43}, pages = {4536--4538}, abstract = {Sidewall functionalized multi-walled carbon nanotubes can be covalently bound parallel to a silicon surface via a self-assembled acid-terminated monolayer used as an organic molecular glue.}, keywords = {carbon, I2CT, Nanotubes, Oxidation-Reduction, Silicon, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{frolet_boosting_2006, title = {Boosting NF-kappaB-dependent basal immunity of Anopheles gambiae aborts development of Plasmodium berghei}, author = {Cécile Frolet and Martine Thoma and Stéphanie A Blandin and Jules A Hoffmann and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17045818}, doi = {10.1016/j.immuni.2006.08.019}, issn = {1074-7613}, year = {2006}, date = {2006-10-01}, journal = {Immunity}, volume = {25}, number = {4}, pages = {677--685}, abstract = {Anopheles gambiae, the major vector for the protozoan malaria parasite Plasmodium falciparum, mounts powerful antiparasitic responses that cause marked parasite loss during midgut invasion. Here, we showed that these antiparasitic defenses were composed of pre- and postinvasion phases and that the preinvasion phase was predominantly regulated by Rel1 and Rel2 members of the NF-kappaB transcription factors. Concurrent silencing of Rel1 and Rel2 decreased the basal expression of the major antiparasitic genes TEP1 and LRIM1 and abolished resistance of Anopheles to the rodent malaria parasite P. berghei. Conversely, depletion of a negative regulator of Rel1, Cactus, prior to infection, enhanced the basal expression of TEP1 and of other immune factors and completely prevented parasite development. Our findings uncover the crucial role of the preinvasion defense in the elimination of parasites, which is at least in part based on circulating blood molecules.}, keywords = {Animals, Anopheles gambiae, blandin, Gene Expression, Gene Expression Regulation, Genes, hoffmann, Immunity, Insect, M3i, NF-kappa B, Plasmodium berghei, telomerase}, pubstate = {published}, tppubtype = {article} } @article{evans_immune_2006, title = {Immune pathways and defence mechanisms in honey bees Apis mellifera}, author = {J D Evans and K Aronstein and Y P Chen and Charles Hetru and Jean-Luc Imler and H Jiang and M Kanost and G J Thompson and Z Zou and D Hultmark}, doi = {10.1111/j.1365-2583.2006.00682.x}, issn = {0962-1075}, year = {2006}, date = {2006-10-01}, journal = {Insect Molecular Biology}, volume = {15}, number = {5}, pages = {645--656}, abstract = {Social insects are able to mount both group-level and individual defences against pathogens. Here we focus on individual defences, by presenting a genome-wide analysis of immunity in a social insect, the honey bee Apis mellifera. We present honey bee models for each of four signalling pathways associated with immunity, identifying plausible orthologues for nearly all predicted pathway members. When compared to the sequenced Drosophila and Anopheles genomes, honey bees possess roughly one-third as many genes in 17 gene families implicated in insect immunity. We suggest that an implied reduction in immune flexibility in bees reflects either the strength of social barriers to disease, or a tendency for bees to be attacked by a limited set of highly coevolved pathogens.}, keywords = {Animals, Bees, Carrier Proteins, Genome, imler, Immunity, Insect, Janus Kinases, M3i, Multigene Family, Serine Endopeptidases, Signal Transduction, STAT Transcription Factors, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{romani_epidermal_2006, title = {Epidermal Langerhans cells--changing views on their function in vivo}, author = {Nikolaus Romani and Susanne Ebner and Christoph H Tripp and Vincent Flacher and Franz Koch and Patrizia Stoitzner}, doi = {10.1016/j.imlet.2006.05.010}, issn = {0165-2478}, year = {2006}, date = {2006-08-01}, journal = {Immunology Letters}, volume = {106}, number = {2}, pages = {119--125}, abstract = {New experimental models and methods have rendered the field of Langerhans cells very lively. An interesting and productive scientific debate as to the functions of Langerhans cells in vivo is currently going on. We have not yet reached the point where the "pros" would weigh out the "cons", or vice versa. There is good evidence for a lack of Langerhans cell function and for down-regulatory Langerhans cell function in some models. On the other hand, there is also evidence for an active immunogenic and tolerogenic role of Langerhans cells. These recent developments will be discussed.}, keywords = {Animals, Epidermal Cells, Epidermis, function, Humans, Immune Tolerance, Immunological, In vivo, Langerhans Cells, Models, REVIEW/EDITORIAL, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{dumortier_functionalized_2006b, title = {Functionalized carbon nanotubes are non-cytotoxic and preserve the functionality of primary immune cells}, author = {Hélène Dumortier and Stéphanie Lacotte and Giorgia Pastorin and Riccardo Marega and Wei Wu and Davide Bonifazi and Jean-Paul Briand and Maurizio Prato and Sylviane Muller and Alberto Bianco}, doi = {10.1021/nl061160x}, issn = {1530-6984}, year = {2006}, date = {2006-07-01}, journal = {Nano Letters}, volume = {6}, number = {7}, pages = {1522--1528}, abstract = {Carbon nanotubes are emerging as innovative tools in nanobiotechnology. However, their toxic effects on environment and health have become an issue of strong concern. In the present study, we address the impact of functionalized carbon nanotubes (f-CNTs) on cells of the immune system. We have prepared two types of f-CNTs, following the 1,3-dipolar cycloaddition reaction (f-CNTs 1 and 2) and the oxidation/amidation treatment (f-CNTs 3 and 4), respectively. We have found that both types of f-CNTs are uptaken by B and T lymphocytes as well as macrophages in vitro, without affecting cell viability. Subsequently, the functionality of the different cells was analyzed carefully. We discovered that f-CNT 1, which is highly water soluble, did not influence the functional activity of immunoregulatory cells. f-CNT 3, which instead possesses reduced solubility and forms mainly stable water suspensions, preserved lymphocytes' functionality while provoking secretion of proinflammatory cytokines by macrophages.}, keywords = {Amides, B-Lymphocytes, Biotechnology, carbon, Cell Survival, Cytokines, I2CT, Macrophages, Molecular Structure, Nanotubes, Oxidation-Reduction, T-Lymphocytes, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pastorin_design_2006, title = {Design and activity of cationic fullerene derivatives as inhibitors of acetylcholinesterase}, author = {Giorgia Pastorin and Silvia Marchesan and Johan Hoebeke and Tatiana Da Ros and Laurence Ehret-Sabatier and Jean-Paul Briand and Maurizio Prato and Alberto Bianco}, doi = {10.1039/b604361e}, issn = {1477-0520}, year = {2006}, date = {2006-07-01}, journal = {Organic & Biomolecular Chemistry}, volume = {4}, number = {13}, pages = {2556--2562}, abstract = {Four different regioisomers of cationic bis-N,N-dimethylfulleropyrrolidinium salts have been prepared and evaluated as inhibitors of the enzymatic activity of acetylcholinesterase. These fullerene-based derivatives were found to be noncompetitive inhibitors of acetylthiocholine hydrolysis. Molecular modelling was used to describe the possible interactions between the fullerene cage and the amino acids surrounding the cavity of the enzyme. The cationic C(60) derivatives used in this study represent a new class of molecules potentially able to modulate the enzymatic activity of acetylcholinesterase.}, keywords = {Acetylcholinesterase, Binding Sites, Cations, Cholinesterase Inhibitors, Drug Design, Fullerenes, I2CT, Models, Molecular, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bianco_hydrolysis_2006, title = {Hydrolysis Rate of Functionalized Fullerenes Bearing Alkoxysilanes: A Comparative Study}, author = {Alberto Bianco and Michele Maggini and Marco Nogarole and Gianfranco Scorrano}, url = {https://chemistry-europe.onlinelibrary.wiley.com/doi/abs/10.1002/ejoc.200600084}, doi = {10.1002/ejoc.200600084}, issn = {1434-193X}, year = {2006}, date = {2006-07-01}, urldate = {2020-03-31}, journal = {European Journal of Organic Chemistry}, volume = {2006}, number = {13}, pages = {2934--2941}, abstract = {Abstract Soluble fulleropyrrolidines bearing a trialkoxysilyl functionality (methoxy, ethoxy, butoxy, and isopropoxy) have been prepared and characterized. The hydrolysis rate constant for each fulleropyrrolidine was measured with 1H NMR spectroscopy by following the disappearance of selected resonances of the fullerene substrate under the conditions (HCl/H2O/THF) used for the preparation of fullerene-doped sol?gel glasses. It has been found that fulleropyrrolidine 1, bearing the trimethoxysilyl group, hydrolyzes faster than substrates 2?7 and should be the reagent of choice to minimize aggregation of the fullerene spheroid in sol?gel glassy matrices. The triethoxysilyl derivative 2, our benchmark fulleropyrrolidine for incorporation in sol?gel glasses, has the secondfastest hydrolysis rate. (? Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)}, keywords = {Alkoxysilanes, Fullerenes, Fulleropyrrolidines, I2CT, Sol–gel, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{dumortier_functionalized_2006, title = {Functionalized carbon nanotubes are non-cytotoxic and preserve the functionality of primary immune cells}, author = {Hélène Dumortier and Stéphanie Lacotte and Giorgia Pastorin and Riccardo Marega and Wei Wu and Davide Bonifazi and Jean-Paul Briand and Maurizio Prato and Sylviane Muller and Alberto Bianco}, doi = {10.1021/nl061160x}, issn = {1530-6984}, year = {2006}, date = {2006-07-01}, journal = {Nano Letters}, volume = {6}, number = {7}, pages = {1522--1528}, abstract = {Carbon nanotubes are emerging as innovative tools in nanobiotechnology. However, their toxic effects on environment and health have become an issue of strong concern. In the present study, we address the impact of functionalized carbon nanotubes (f-CNTs) on cells of the immune system. We have prepared two types of f-CNTs, following the 1,3-dipolar cycloaddition reaction (f-CNTs 1 and 2) and the oxidation/amidation treatment (f-CNTs 3 and 4), respectively. We have found that both types of f-CNTs are uptaken by B and T lymphocytes as well as macrophages in vitro, without affecting cell viability. Subsequently, the functionality of the different cells was analyzed carefully. We discovered that f-CNT 1, which is highly water soluble, did not influence the functional activity of immunoregulatory cells. f-CNT 3, which instead possesses reduced solubility and forms mainly stable water suspensions, preserved lymphocytes' functionality while provoking secretion of proinflammatory cytokines by macrophages.}, keywords = {Amides, B-Lymphocytes, Biotechnology, carbon, Cell Survival, Cytokines, Dumortier, I2CT, Macrophages, Molecular Structure, Nanotubes, Oxidation-Reduction, T-Lymphocytes, Team-Bianco, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{galiana-arnoux_essential_2006, title = {Essential function in vivo for Dicer-2 in host defense against RNA viruses in drosophila}, author = {Delphine Galiana-Arnoux and Catherine Dostert and Anette Schneemann and Jules A Hoffmann and Jean-Luc Imler}, doi = {10.1038/ni1335}, issn = {1529-2908}, year = {2006}, date = {2006-06-01}, journal = {Nature Immunology}, volume = {7}, number = {6}, pages = {590--597}, abstract = {The fruit fly Drosophila melanogaster is a model system for studying innate immunity, including antiviral host defense. Infection with drosophila C virus triggers a transcriptional response that is dependent in part on the Jak kinase Hopscotch. Here we show that successful infection and killing of drosophila with the insect nodavirus flock house virus was strictly dependent on expression of the viral protein B2, a potent inhibitor of processing of double-stranded RNA mediated by the essential RNA interference factor Dicer. Conversely, flies with a loss-of-function mutation in the gene encoding Dicer-2 (Dcr-2) showed enhanced susceptibility to infection by flock house virus, drosophila C virus and Sindbis virus, members of three different families of RNA viruses. These data demonstrate the importance of RNA interference for controlling virus replication in vivo and establish Dcr-2 as a host susceptibility locus for virus infections.}, keywords = {Animals, Genetically Modified, hoffmann, imler, M3i, Mutation, Nodaviridae, Ribonuclease III, RNA, RNA Helicases, RNA Interference, RNA Viruses, Viral, Viral Proteins, Virus Replication}, pubstate = {published}, tppubtype = {article} } @article{chen_weckle_2006, title = {Weckle is a zinc finger adaptor of the toll pathway in dorsoventral patterning of the Drosophila embryo}, author = {Li-Ying Chen and Juinn-Chin Wang and Yann Hyvert and Hui-Ping Lin and Norbert Perrimon and Jean-Luc Imler and Jui-Chou Hsu}, doi = {10.1016/j.cub.2006.05.050}, issn = {0960-9822}, year = {2006}, date = {2006-06-01}, journal = {Current biology: CB}, volume = {16}, number = {12}, pages = {1183--1193}, abstract = {BACKGROUND: The Drosophila Toll pathway takes part in both establishment of the embryonic dorsoventral axis and induction of the innate immune response in adults. Upon activation by the cytokine Spätzle, Toll interacts with the adaptor proteins DmMyD88 and Tube and the kinase Pelle and triggers degradation of the inhibitor Cactus, thus allowing the nuclear translocation of the transcription factor Dorsal/Dif. weckle (wek) was previously identified as a new dorsal group gene that encodes a putative zinc finger transcription factor. However, its role in the Toll pathway was unknown. RESULTS: Here, we isolated new wek alleles and demonstrated that cactus is epistatic to wek, which in turn is epistatic to Toll. Consistent with this, Wek localizes to the plasma membrane of embryos, independently of Toll signaling. Wek homodimerizes and associates with Toll. Moreover, Wek binds to and localizes DmMyD88 to the plasma membrane. Thus, Wek acts as an adaptor to assemble/stabilize a Toll/Wek/DmMyD88/Tube complex. Remarkably, unlike the DmMyD88/tube/pelle/cactus gene cassette of the Toll pathway, wek plays a minimal role, if any, in the immune defense against Gram-positive bacteria and fungi. CONCLUSIONS: We conclude that Wek is an adaptor to link Toll and DmMyD88 and is required for efficient recruitment of DmMyD88 to Toll. Unexpectedly, wek is dispensable for innate immune response, thus revealing differences in the Toll-mediated activation of Dorsal in the embryo and Dif in the fat body of adult flies.}, keywords = {Adaptor Proteins, Animals, Antigens, Biological, Body Patterning, Cell Membrane, Differentiation, dimerization, DNA-Binding Proteins, Embryo, Epistasis, Genetic, imler, Immunity, Immunologic, Innate, M3i, Models, Mutation, Nonmammalian, Phenotype, Phosphoproteins, Receptors, Signal Transducing, Toll-Like Receptors, Transcription Factors, Zinc Fingers}, pubstate = {published}, tppubtype = {article} } @article{brosson_proteomic_2006, title = {Proteomic analysis of the eukaryotic parasite Encephalitozoon cuniculi (microsporidia): a reference map for proteins expressed in late sporogonial stages.}, author = {Damien Brosson and Lauriane Kuhn and Frédéric Delbac and Jérôme Garin and Christian P Vivarès and Catherine Texier}, doi = {10.1002/pmic.200500796}, issn = {1615-9853 1615-9853}, year = {2006}, date = {2006-06-01}, journal = {Proteomics}, volume = {6}, number = {12}, pages = {3625--3635}, abstract = {The microsporidian Encephalitozoon cuniculi is a unicellular obligate intracellular parasite considered as an emerging opportunistic human pathogen. The differentiation phase of its life cycle leads to the formation of stress-resistant spores. The E. cuniculi genome (2.9 Mbp) having been sequenced, we undertook a descriptive proteomic study of a spore-rich cell population isolated from culture supernatants. A combination of 2-DE and 2-DE-free techniques was applied to whole-cell protein extracts. Protein identification was performed using an automated MALDI-TOF-MS platform and a nanoLC-MS/MS instrument. A reference 2-DE map of about 350 major spots with multiple isoforms was obtained, and for the first time in microsporidia, a large set of unique proteins (177) including proteins with unknown function in a proportion of 25.6% was identified. The data are mainly discussed with reference to secretion and spore structural features, energy and carbohydrate metabolism, cell cycle control and parasite survival in the environment.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{MM2006, title = {Cell biology, compartmentalization and immunology of mosquito midgut – malaria parasite interactions}, author = {Miranda M Whitten and S H Shiao and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/16542314}, year = {2006}, date = {2006-04-01}, journal = {Parasite Immunol.}, volume = {28}, number = {4}, pages = {121-30}, abstract = {The malaria parasite Plasmodium has an absolute requirement for both a vertebrate and a mosquito host in order to complete its life cycle, and its interactions with the latter provide the focus for this review. The mosquito midgut represents one of the most challenging environments for the survival and development of Plasmodium, and is thus also one of the most attractive sites for novel targeted malaria control strategies. During their attempts to cross the midgut epithelium en route to the salivary glands, motile ookinetes are swiftly detected and labelled by mosquito recognition factors and targeted for destruction by a variety of immune responses that recruit killing factors both from the midgut and from other tissues in the surrounding body cavity. The exact interplay between these factors and the parasite is highly species- and strain-specific, as are the timing and the route of parasite invasion. These features are paramount to determining the success of the infection and the vector competence of the mosquito. Here we discuss recent advances in genomic analyses, coupled with detailed microscopical investigations, which are helping to unravel the identity and roles of the major players of these complex systems.}, keywords = {mosquito midgut}, pubstate = {published}, tppubtype = {article} } @article{sarry_early_2006, title = {The early responses of Arabidopsis thaliana cells to cadmium exposure explored by protein and metabolite profiling analyses.}, author = {Jean-Emmanuel Sarry and Lauriane Kuhn and Céline Ducruix and Alexandra Lafaye and Christophe Junot and Véronique Hugouvieux and Agnès Jourdain and Olivier Bastien and Julie B Fievet and Dominique Vailhen and Badia Amekraz and Christophe Moulin and Eric Ezan and Jérôme Garin and Jacques Bourguignon}, doi = {10.1002/pmic.200500543}, issn = {1615-9853 1615-9853}, year = {2006}, date = {2006-04-01}, journal = {Proteomics}, volume = {6}, number = {7}, pages = {2180--2198}, abstract = {To get more insight into plant cell response to cadmium (Cd) stress, both proteomic and metabolomic "differential display" analyses were performed on Arabidopsis thaliana cells exposed to different concentrations of the toxic chemical. After a 24 h treatment, soluble proteins extracted from untreated and treated cells were separated by 2-D-PAGE and image analyses were performed to quantify and compare protein levels. Proteins up- and down-regulated in response to Cd were identified by MS and mapped into specific metabolic pathways and cellular processes, highlighting probable activation of the carbon, nitrogen, and sulfur metabolic pathways. For some of these proteins, Northern blot and RT-PCR analyses were performed to test transcript accumulation in response to Cd. In parallel, metabolite profiling analyses by LC coupled to ESI MS were initiated to better characterize the metabolic adaptation to the chemical stress. This study revealed that the main variation at the metabolite level came from the presence of six different families of phytochelatins, in A. thaliana cells treated with Cd, whose accumulation increases with Cd concentrations. Taken together these data provide an overview of the molecular and cellular changes elicited by Cd exposure.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{levine_analysis_2006, title = {Analysis of the dynamic Bacillus subtilis Ser/Thr/Tyr phosphoproteome implicated in a wide variety of cellular processes.}, author = {Alain Lévine and Françoise Vannier and Cédric Absalon and Lauriane Kuhn and Peter Jackson and Elaine Scrivener and Valérie Labas and Joëlle Vinh and Patrick Courtney and Jérôme Garin and Simone J Séror}, doi = {10.1002/pmic.200500352}, issn = {1615-9853 1615-9853}, year = {2006}, date = {2006-04-01}, journal = {Proteomics}, volume = {6}, number = {7}, pages = {2157--2173}, abstract = {The physiological role of proteins phosphorylated on serine/threonine/tyrosine (Ser/Thr/Tyr) residues or the identity of the corresponding kinases and phosphatases is generally poorly understood in bacteria. As a first step in analysing the importance of such phosphorylation, we sought to establish the nature of the Ser/Thr/Tyr phosphoproteome in Bacillus subtilis, using in vivo labelling with [(32)P]-orthophosphate, one-unit pH 2-DE, combined with MS. Highly reproducible 2-D profiles of phosphoproteins were obtained with early stationary-phase cells. The 2-D profiles contained at least 80 clearly labelled spots in the pH range 4-7. Forty-six spots were analysed by MS (confirmed in most cases by LC-MS/MS), identifying a total of 29 different proteins, with 19 identified for the first time as bacterial phosphoproteins. These phosphoproteins are implicated in a wide variety of cellular processes, including carbon and energy metabolism, transport, stress and development. Significant changes to the profiles were obtained as a result of cold, heat or osmotic shock, demonstrating that, in stationary-phase cells, the phosphoproteome is dynamic. An initial comparative study indicated that at least 25 [(32)P]-labelled spots were also stained by Pro-Q Diamond, with apparently six additional phosphoproteins uniquely detected by Pro-Q.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{bianco_solid-phase_2006, title = {Solid-phase synthesis of CD40L mimetics}, author = {Alberto Bianco and Sylvie Fournel and Sébastien Wieckowski and Johan Hoebeke and Gilles Guichard}, doi = {10.1039/b601528j}, issn = {1477-0520}, year = {2006}, date = {2006-04-01}, journal = {Organic & Biomolecular Chemistry}, volume = {4}, number = {8}, pages = {1461--1463}, abstract = {The C3-symmetric molecule has been previously shown to mimic CD40 ligand (CD40L) homotrimers and to display effector functions. This molecule consists of a cyclic hexapeptide core containing the repetition of the D-Ala-L-Lys motif. The side chains of the lysine residues have been modified by appending the CD40L-derived sequence 143Lys-Gly-Tyr-Tyr146 via a 6-aminohexanoic acid residue as a spacer. The present report describes a general solid-phase synthesis approach to and related trimeric architectures. In addition, their CD40 binding properties as well as their effector functions have been evaluated.}, keywords = {Apoptosis, CD40 Ligand, Cell Line, Chromatography, Combinatorial Chemistry Techniques, High Pressure Liquid, Humans, I2CT, Molecular Mimicry, Molecular Structure, Protein Binding, Team-Bianco, tumor}, pubstate = {published}, tppubtype = {article} } @article{tasis_chemistry_2006, title = {Chemistry of carbon nanotubes}, author = {Dimitrios Tasis and Nikos Tagmatarchis and Alberto Bianco and Maurizio Prato}, doi = {10.1021/cr050569o}, issn = {0009-2665}, year = {2006}, date = {2006-03-01}, journal = {Chemical Reviews}, volume = {106}, number = {3}, pages = {1105--1136}, keywords = {Animals, Biopolymers, carbon, Chemical Phenomena, Chemistry, I2CT, Molecular Structure, Nanotubes, Solubility, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pastorin_double_2006, title = {Double functionalization of carbon nanotubes for multimodal drug delivery}, author = {Giorgia Pastorin and Wei Wu and Sébastien Wieckowski and Jean-Paul Briand and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, doi = {10.1039/b516309a}, issn = {1359-7345}, year = {2006}, date = {2006-03-01}, journal = {Chemical Communications (Cambridge, England)}, number = {11}, pages = {1182--1184}, abstract = {Multi-walled carbon nanotubes have been covalently functionalized via 1,3-dipolar cycloaddition of azomethine ylides with orthogonally protected amino functions that can be selectively deprotected and subsequently modified with drugs and fluorescent probes.}, keywords = {Azo Compounds, carbon, Cyclization, Drug Carriers, I2CT, Microscopy, Nanotubes, Pharmaceutical Preparations, Scanning Tunneling, Solubility, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{brough_60fullerene-pyrrolidine-n-oxides_2006, title = {[60]fullerene-pyrrolidine-N-oxides}, author = {Peter Brough and Cedric Klumpp and Alberto Bianco and Stephane Campidelli and Maurizio Prato}, doi = {10.1021/jo052388s}, issn = {0022-3263}, year = {2006}, date = {2006-03-01}, journal = {The Journal of Organic Chemistry}, volume = {71}, number = {5}, pages = {2014--2020}, abstract = {Eight members of a new family of fullerene derivatives, [60]fulleropyrrolidine-N-oxides, have been synthesized and characterized. Facile oxidation, by a peracid, of the parent [60]fulleropyrrolidine gave clean conversions into the product molecules, in which the tertiary amine is transformed into a quaternary amine bearing an oxygen atom. The reaction is very selective, favoring the nitrogen atom of the pyrrolidine ring in preference to epoxidation of the fullerene cage. The 1H NMR shows an AB quartet splitting pattern, characteristic of nonequivalent hydrogens in the pyrrolidine ring and at a chemical shift displacement of 0.8 ppm downfield. Other methods of characterization are described, including MS, differential scanning calorimetry, thermogravimetric analysis, HPLC, UV/vis, and IR. Conclusive evidence for the formation of an N-oxide moiety is provided by the synthesis, oxidation, and NMR characterization of a novel [60]fulleropyrrolidine containing a 15N isotope, showing an 85 ppm downfield heteroatom chemical shift. Preliminary details of the effects of substitution on the reactivity of the pyrrolidine ring are also reported.}, keywords = {Chromatography, Fullerenes, High Pressure Liquid, I2CT, Nitrogen, Oxidation-Reduction, Oxides, Pyrrolidines, Spectrum Analysis, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{klumpp_functionalized_2006, title = {Functionalized carbon nanotubes as emerging nanovectors for the delivery of therapeutics}, author = {Cédric Klumpp and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, doi = {10.1016/j.bbamem.2005.10.008}, issn = {0006-3002}, year = {2006}, date = {2006-03-01}, journal = {Biochimica Et Biophysica Acta}, volume = {1758}, number = {3}, pages = {404--412}, abstract = {Functionalized carbon nanotubes (f-CNT) are emerging as a new family of nanovectors for the delivery of different types of therapeutic molecules. The application of CNT in the field of carrier-mediated delivery has become possible after the recent discovery of their capacity to penetrate into the cells. CNT can be loaded with active molecules by forming stable covalent bonds or supramolecular assemblies based on noncovalent interactions. Once the cargos are carried into various cells, tissues and organs they are able to express their biological function. In this review, we will describe the potential of f-CNT to deliver different types of therapeutic molecules.}, keywords = {carbon, DNA, Drug Carriers, Drug Delivery Systems, Humans, I2CT, Nanotubes, RNA, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{peuvel-fanget_enp1_2006, title = {EnP1 and EnP2, two proteins associated with the Encephalitozoon cuniculi endospore, the chitin-rich inner layer of the microsporidian spore wall.}, author = {Isabelle Peuvel-Fanget and Valérie Polonais and Damien Brosson and Catherine Texier and Lauriane Kuhn and Pierre Peyret and Christian Vivarès and Frédéric Delbac}, doi = {10.1016/j.ijpara.2005.10.005}, issn = {0020-7519 0020-7519}, year = {2006}, date = {2006-03-01}, journal = {International journal for parasitology}, volume = {36}, number = {3}, pages = {309--318}, abstract = {Microsporidia are obligate intracellular parasites forming environmentally resistant spores that harbour a rigid cell wall. This wall comprises an outer layer or exospore and a chitin-rich inner layer or endospore. So far, only a chitin deacetylase-like protein has been shown to localize to the Encephalitozoon cuniculi endospore and either one or two proteins have been clearly assigned to the exospore in two Encephalitozoon species: SWP1 in E. cuniculi, SWP1 and SWP2 in Encephalitozoon intestinalis. Here, we report the identification of two new spore wall proteins in E. cuniculi, EnP1 and EnP2, the genes of which are both located on chromosome I (ECU01_0820 and ECU01_1270, respectively) and have no known homologue. Detected by immunoscreening of an E. cuniculi cDNA library, enp1 is characterized by small-sized 5' and 3' untranslated regions and is highly expressed throughout the whole intracellular cycle. The encoded basic 40 kDa antigen displays a high proportion of cysteine residues, arguing for a significant role of disulfide bridges in spore wall assembly. EnP2 is a 22 kDa serine-rich protein that is predicted to be O-glycosylated and glycosylated phosphatidyl inositol-anchored. Although having been identified by mass spectrometry of a dithiothreitol-soluble fraction, this protein contains only two cysteine residues. Mouse polyclonal antibodies were raised against EnP1 and EnP2 recombinant proteins produced in Escherichia coli Our immunolocalisation data indicate that EnP1 and EnP2 are targeted to the cell surface as early as the onset of sporogony and are finally associated with the chitin-rich layer of the wall in mature spores.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{durand_lipoteichoic_2006, title = {Lipoteichoic acid increases TLR and functional chemokine expression while reducing dentin formation in in vitro differentiated human odontoblasts}, author = {Stéphanie H Durand and Vincent Flacher and Annick Roméas and Florence Carrouel and Evelyne Colomb and Claude Vincent and Henry Magloire and Marie-Lise Couble and Françoise Bleicher and Marie-Jeanne Staquet and Serge Lebecque and Jean-Christophe Farges}, doi = {10.4049/jimmunol.176.5.2880}, issn = {0022-1767}, year = {2006}, date = {2006-03-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {176}, number = {5}, pages = {2880--2887}, abstract = {Gram-positive bacteria entering the dentinal tissue during the carious process are suspected to influence the immune response in human dental pulp. Odontoblasts situated at the pulp/dentin interface are the first cells encountered by these bacteria and therefore could play a crucial role in this response. In the present study, we found that in vitro-differentiated odontoblasts constitutively expressed the pattern recognition receptor TLR1-6 and 9 genes but not TLR7, 8, and 10. Furthermore, lipoteichoic acid (LTA), a wall component of Gram-positive bacteria, triggered the activation of the odontoblasts. LTA up-regulated the expression of its own receptor TLR2, as well as the production of several chemokines. In particular, an increased amount of CCL2 and CXCL10 was detected in supernatants from LTA-stimulated odontoblasts, and those supernatants augmented the migration of immature dendritic cells in vitro compared with controls. Clinical relevance of these observations came from immunohistochemical analysis showing that CCL2 was expressed in vivo by odontoblasts and blood vessels present under active carious lesions but not in healthy dental pulps. In contrast with this inflammatory response, gene expression of major dentin matrix components (type I collagen, dentin sialophosphoprotein) and TGF-beta1 was sharply down-regulated in odontoblasts by LTA. Taken together, these data suggest that odontoblasts activated through TLR2 by Gram-positive bacteria LTA are able to initiate an innate immune response by secreting chemokines that recruit immature dendritic cells while down-regulating their specialized functions of dentin matrix synthesis and mineralization.}, keywords = {Activation, Analysis, bacteria, Biosynthesis, BLOOD, Blood Vessels, Cell Differentiation, Cells, Chemistry, chemokines, COLLAGEN, Cultured, CXCL10, cytology, Dendritic Cells, DENTAL PULP, Dentin, development, Down-Regulation, Expression, extracellular, EXTRACELLULAR MATRIX, Extracellular Matrix Proteins, function, Gene, Gene Expression, Genes, Genetics, Gram-Positive Bacteria, Human, Humans, IMMATURE, Immunology, IN VITRO, In vivo, Innate immune response, lipopolysaccharide, Lipopolysaccharides, metabolism, migration, Odontoblasts, Organ Culture Techniques, Pharmacology, physiology, PRODUCTION, Protein, Proteins, Receptor, recognition, synthesis, Team-Mueller, Teichoic Acids, TLR7, Toll-Like Receptor 2, Up-Regulation}, pubstate = {published}, tppubtype = {article} } @article{singh_tissue_2006, title = {Tissue biodistribution and blood clearance rates of intravenously administered carbon nanotube radiotracers}, author = {Ravi Singh and Davide Pantarotto and Lara Lacerda and Giorgia Pastorin and Cédric Klumpp and Maurizio Prato and Alberto Bianco and Kostas Kostarelos}, doi = {10.1073/pnas.0509009103}, issn = {0027-8424}, year = {2006}, date = {2006-02-01}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {103}, number = {9}, pages = {3357--3362}, abstract = {Carbon nanotubes (CNT) are intensively being developed for biomedical applications including drug and gene delivery. Although all possible clinical applications will require compatibility of CNT with the biological milieu, their in vivo capabilities and limitations have not yet been explored. In this work, water-soluble, single-walled CNT (SWNT) have been functionalized with the chelating molecule diethylentriaminepentaacetic (DTPA) and labeled with indium ((111)In) for imaging purposes. Intravenous (i.v.) administration of these functionalized SWNT (f-SWNT) followed by radioactivity tracing using gamma scintigraphy indicated that f-SWNT are not retained in any of the reticuloendothelial system organs (liver or spleen) and are rapidly cleared from systemic blood circulation through the renal excretion route. The observed rapid blood clearance and half-life (3 h) of f-SWNT has major implications for all potential clinical uses of CNT. Moreover, urine excretion studies using both f-SWNT and functionalized multiwalled CNT followed by electron microscopy analysis of urine samples revealed that both types of nanotubes were excreted as intact nanotubes. This work describes the pharmacokinetic parameters of i.v. administered functionalized CNT relevant for various therapeutic and diagnostic applications.}, keywords = {Animals, carbon, Electron, Female, Half-Life, I2CT, Inbred BALB C, Indium Radioisotopes, Injections, Intravenous, Mice, Microscopy, Molecular Structure, Nanotubes, Pentetic Acid, Team-Bianco, Tissue Distribution, Transmission}, pubstate = {published}, tppubtype = {article} } @article{bischoff_downregulation_2006, title = {Downregulation of the Drosophila immune response by peptidoglycan-recognition proteins SC1 and SC2}, author = {Vincent Bischoff and Cécile Vignal and Bernard Duvic and Ivo G Boneca and Jules A Hoffmann and Julien Royet}, doi = {10.1371/journal.ppat.0020014}, issn = {1553-7374}, year = {2006}, date = {2006-02-01}, journal = {PLoS Pathog.}, volume = {2}, number = {2}, pages = {e14}, abstract = {Peptidoglycan-recognition proteins (PGRPs) are evolutionarily conserved molecules that are structurally related to bacterial amidases. Several Drosophila PGRPs have lost this enzymatic activity and serve as microbe sensors through peptidoglycan recognition. Other PGRP family members, such as Drosophila PGRP-SC1 or mammalian PGRP-L, have conserved the amidase function and are able to cleave peptidoglycan in vitro. However, the contribution of these amidase PGRPs to host defense in vivo has remained elusive so far. Using an RNA-interference approach, we addressed the function of two PGRPs with amidase activity in the Drosophila immune response. We observed that PGRP-SC1/2-depleted flies present a specific over-activation of the IMD (immune deficiency) signaling pathway after bacterial challenge. Our data suggest that these proteins act in the larval gut to prevent activation of this pathway following bacterial ingestion. We further show that a strict control of IMD-pathway activation is essential to prevent bacteria-induced developmental defects and larval death.}, keywords = {Animals, Antimicrobial Cationic Peptides, bacteria, Carrier Proteins, Down-Regulation, hoffmann, Larva, M3i, RNA Interference, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{leclerc_prophenoloxidase_2006, title = {Prophenoloxidase activation is not required for survival to microbial infections in Drosophila}, author = {Vincent Leclerc and Nadège Pelte and Laure El Chamy and Cosimo Martinelli and Petros Ligoxygakis and Jules A Hoffmann and Jean-Marc Reichhart}, doi = {10.1038/sj.embor.7400592}, issn = {1469-221X}, year = {2006}, date = {2006-02-01}, journal = {EMBO Rep.}, volume = {7}, number = {2}, pages = {231--235}, abstract = {The antimicrobial defence of Drosophila relies on cellular and humoral processes, of which the inducible synthesis of antimicrobial peptides has attracted interest in recent years. Another potential line of defence is the activation, by a proteolytic cascade, of phenoloxidase, which leads to the production of quinones and melanin. However, in spite of several publications on this subject, the contribution of phenoloxidase activation to resistance to infections has not been established under appropriate in vivo conditions. Here, we have isolated the first Drosophila mutant for a prophenoloxidase-activating enzyme (PAE1). In contrast to wild-type flies, PAE1 mutants fail to activate phenoloxidase in the haemolymph following microbial challenge. Surprisingly, we find that these mutants are as resistant to infections as wild-type flies, in the total absence of circulating phenoloxidase activity. This raises the question with regard to the precise function of phenoloxidase activation in defence, if any.}, keywords = {Animals, Bacterial Infections, Catechol Oxidase, Enzyme Activation, Enzyme Precursors, Gram-Negative Bacteria, Gram-Positive Bacteria, Hemolymph, hoffmann, Immunity, Innate, M3i, Mutation, reichhart, Survival Rate}, pubstate = {published}, tppubtype = {article} } @article{sarry_dynamics_2006, title = {Dynamics of Arabidopsis thaliana soluble proteome in response to different nutrient culture conditions.}, author = {Jean-Emmanuel Sarry and Lauriane Kuhn and Pascaline Le Lay and Jérôme Garin and Jacques Bourguignon}, doi = {10.1002/elps.200500561}, issn = {0173-0835 0173-0835}, year = {2006}, date = {2006-02-01}, journal = {Electrophoresis}, volume = {27}, number = {2}, pages = {495--507}, abstract = {In an effort to determine the best extraction procedure compatible with the high-reproducible 2-DE, different methods of soluble protein extraction from Arabidopsis cell culture suspensions grown in Gamborg B5 medium were tested. A reference 2-DE map was established for this soluble extract revealing 1184 spots. The most abundant protein spots were excised, trypsin-digested, and mass spectra obtained via MALDI-TOF and/or LC coupled to ESI-MS. Three hundred and thirty one proteins were identified and their functions were defined based on sequence comparisons and classified in different protein families. In order to analyze the impact of culture medium on the Arabidopsis proteome, we performed the 2-DE map from Arabidopsis cell suspensions cultured in another growth medium Murashige and Skoog (M-S) and 327 major spots were identified. Using PDQuest imaging analysis, significant increases in the amount of several housekeeping enzymes, stress/defense proteins, and heat shock proteins were found in M-S medium. Modified expression of certain proteins and detection of new isoforms involved in nitrate assimilation, nitrogen, and sulfur metabolism were also observed in the M-S medium. This study provides the first 2-DE maps of the soluble proteome of Arabidopsis cell suspensions. The comparative analysis of the Arabidopsis proteome in respect to different nutrient supplies shows that the culture medium may significantly influence the expression pattern of major soluble proteins in Arabidopsis cells. This work also constitutes an important step for further proteomic analysis concerning cell responses to abiotic or biotic stresses.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular contacts between antibiotics and the 30S ribosomal particle}, author = {J Wirmer and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17116475}, isbn = {17116475}, year = {2006}, date = {2006-01-01}, journal = {Methods Enzymol}, volume = {415}, pages = {180-202}, abstract = {Crystal structures of complexes between ribosomal particles and antibiotics have pinned down very precisely the discrete binding sites of several classes of antibiotics inhibiting protein synthesis. The crystal structures of complexes between various antibiotics and ribosomal particles show definitively that ribosomal RNAs (rRNAs), rather than ribosomal proteins, are overwhelmingly targeted. The antibiotics are found at messenger RNA or transfer RNA binding sites and, most importantly, at pivot locations that are key for the structural rearrangements during the molecular mechanical steps in initiation, elongation, or termination of protein synthesis. We focus here on the 30S particle. Structurally, the antibiotics interact in many ways with RNA: (i) only with the phosphate groups (streptomycin); (ii) mainly with bases (hygromycin, spectinomycin); (iii) with a mixture of both (paromomycin, Geneticin); (iv) via magnesium ions (tetracycline) or a protein side chain (streptomycin). The antibiotics can mimic base stacking (pactamycin) or form pseudo-base pairing interactions with ribosomal bases (paromomycin and related aminoglycosides). Resistance strategies (mutations or methylations in rRNA or enzymatic modifications of the antibiotics) can generally be understood on the basis of the intermolecular contacts made between the antibiotics and rRNA residues in the crystal structures. In humans, toxicity of ribosomal antibiotics is most likely due, at least in part, to the sensitivity of mitochondrial ribosomes, since mitochondria evolved from a bacterial ancestor. Antibiotic families (e.g., aminoglycosides) form a set of invariant H-bonds to defined rRNA residues. When such residues are conserved in bacteria, but not in eukaryotes, resistance of eukaryotic ribosomes is observed. The structural knowledge, together with comparative genomic analysis, should allow for the development of new broad-spectrum antibiotics with higher selectivity toward bacterial ribosomes and less toxicity on eukaryotic cytoplasmic and mitochondrial ribosomes.}, note = {0076-6879 (Print) Journal Article Research Support, Non-U.S. Gov't}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cooperation between Reverse Transcriptase and Integrase during Reverse Transcription and Formation of the Preintegrative Complex of Ty1}, author = {M Wilhelm and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17031000}, isbn = {17031000}, year = {2006}, date = {2006-01-01}, journal = {Eukaryot Cell}, volume = {5}, number = {10}, pages = {1760-1769}, abstract = {Reverse transcriptase (RT) and integrase (IN) play a central role in the replication and transposition of retroelements. Increasing evidence suggests that the interaction between these two enzymes is functional and plays an important role in replication. In the yeast Saccharomyces cerevisiae retrotransposon Ty1, the interaction of IN with RT is critical for the formation of an active conformation of RT. We show here that the RT associated with VLPs is active only if it is in close interaction with IN. To probe the IN-RT cis-trans relationship, we have used a complementation assay based on coexpressing two transposons. We show that IN acts in cis to activate RT and that a functional integrase provided in trans is not able to complement replication and transposition defects of IN deletion or IN active-site mutant elements. Our data support a model in which IN not only interacts closely with RT during reverse transcription but also remains associated with RT during the formation of the preintegrative complex.}, note = {1535-9778 (Print) Journal Article}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The ribosomal decoding site and antibiotics}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16876930}, isbn = {16876930}, year = {2006}, date = {2006-01-01}, journal = {Biochimie}, volume = {88}, number = {8}, pages = {931-933}, note = {0300-9084 (Print) Editorial}, keywords = {Anti-Bacterial Agents/metabolism/*pharmacology Bacteria/*drug effects/genetics/growth & development Binding Sites/drug effects Protein Biosynthesis/drug effects/genetics RNA Amino Acyl/*metabolism, Bacterial/*metabolism RNA, Transfer, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleoside and nucleotide inhibitors of HIV-1 replication}, author = {V Vivet-Boudou and J Didierjean and C Isel and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16389458}, isbn = {16389458}, year = {2006}, date = {2006-01-01}, journal = {Cell Mol Life Sci}, volume = {63}, number = {2}, pages = {163-186}, abstract = {HIV-1 reverse transcriptase (RT) is one of the main targets for antiviral therapy. Two classes of RT inhibitors can be distinguished: those that are nucleoside or nucleotide analogues (sharing the common NRTIs abbreviation) and those that are not. This review focuses on the NRTIs, which are highly efficient in slowing down viral replication and are used in combination regimens. Unfortunately, the current inhibitors do not completely suppress viral replication and due to the high capacity of adaptation of HIV, allow the selection of drug-resistant viruses. Resistance mechanisms to NRTIs have been extensively investigated and can be divided into two types: improved discrimination of a nucleotide analogue relative to the natural substrate or increased phosphorolytic cleavage of an analogue-blocked primer. This knowledge is important both for the development of new drugs designed to target resistant strains and for the development of new antiviral strategies. The NRTIs currently in clinical trials and new developments in this area are also reviewed.}, note = {1420-682X (Print) Journal Article Review}, keywords = {Biological Mutation Nucleosides/chemistry/pharmacology Nucleotides/chemistry/pharmacology Protein Structure, Drug Resistance, MARQUET, PAILLART, Tertiary Reverse Transcriptase Inhibitors/*chemistry/*pharmacology Virus Replication, Unité ARN, Viral/*genetics HIV Infections/drug therapy HIV-1/drug effects/*enzymology/genetics Humans Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {A molecular dynamics simulation study of an aminoglycoside/A-site RNA complex: conformational and hydration patterns}, author = {A C Vaiana and E Westhof and P Auffinger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16824662}, isbn = {16824662}, year = {2006}, date = {2006-01-01}, journal = {Biochimie}, volume = {88}, number = {8}, pages = {1061-1073}, abstract = {Aminoglycoside antibiotics interfere with the translation mechanism by binding to the tRNA decoding site of the 16S ribosomal RNA. Crystallographic structures of aminoglycosides bound to A-site systems clarified many static aspects of RNA-ligand interactions. To gain some insight on the dynamic aspects of recognition phenomena, we conducted molecular dynamics simulations of the aminoglycoside paromomycin bound to a eubacterial ribosomal decoding A-site oligonucleotide. Results from 25 ns of simulation time revealed that: (i) the neamine part of the antibiotic represents the main anchor for binding, (ii) additional sugar rings provide limited and fragile contacts, (iii) long-resident water molecules present at the drug/RNA interface are involved in the recognition phenomena. The combination of MD simulations together with systematic structural information offers striking insights into the molecular recognition processes underlying RNA/aminoglycoside binding. Important methodological considerations related to the use of medium resolution starting structures and associated sampling problems are thoroughly discussed.}, note = {0300-9084 (Print) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of the ribosome-bound cricket paralysis virus IRES RNA}, author = {M Schuler and S R Connell and A Lescoute and J Giesebrecht and M Dabrowski and B Schroeer and T Mielke and P A Penczek and E Westhof and C M Spahn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17115051}, isbn = {17115051}, year = {2006}, date = {2006-01-01}, journal = {Nat Struct Mol Biol}, volume = {13}, number = {12}, pages = {1092-1096}, abstract = {Internal ribosome entry sites (IRESs) facilitate an alternative, end-independent pathway of translation initiation. A particular family of dicistroviral IRESs can assemble elongation-competent 80S ribosomal complexes in the absence of canonical initiation factors and initiator transfer RNA. We present here a cryo-EM reconstruction of a dicistroviral IRES bound to the 80S ribosome. The resolution of the cryo-EM reconstruction, in the subnanometer range, allowed the molecular structure of the complete IRES in its active, ribosome-bound state to be solved. The structure, harboring three pseudoknot-containing domains, each with a specific functional role, shows how defined elements of the IRES emerge from a compactly folded core and interact with the key ribosomal components that form the A, P and E sites, where tRNAs normally bind. Our results exemplify the molecular strategy for recruitment of an IRES and reveal the dynamic features necessary for internal initiation.}, note = {1545-9993 (Print) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Dispositif microfluidique pour la cristallisation et l'analyse cristallographique de molécules.}, author = {C Sauter and B Lorber and A Théobald-Dietrich and R Giege and C Khan-Malek and B Gauthier-Manuel and G Thuillier and R Ferrigno}, isbn = {EP 2040809 A2}, year = {2006}, date = {2006-01-01}, keywords = {FRUGIER, GIEGE, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Idiosyncratic behaviour of tRNA-like structures in translation of plant Viral RNA genomes}, author = {J Rudinger-Thirion and R C Olsthoorn and R Giege and S Barends}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16337653}, isbn = {16337653}, year = {2006}, date = {2006-01-01}, journal = {J Mol Biol}, volume = {355}, number = {5}, pages = {873-878}, abstract = {Tobacco mosaic virus (TMV) and Nemesia ring necrosis virus (NeRNV) belong to the Tobamoviridae and Tymoviridae families, respectively. Although their RNAs present different 5'-untranslated regions and different family-specific genomic organizations, they share common 3'-ends organized into three consecutive pseudoknot structures followed by a histidylatable tRNA-like structure (TLS). We investigate here whether the histidine residue becomes incorporated into viral proteins and if the TLSs of TMV and NeRNV play a role in viral translation. Our results indicate that, regardless of the genomic context, the histidine moiety does not become incorporated in proteins via ribosomal translation, and that disruption of the TLS in either viral RNA does not perturb the viral translation patterns. In the light of the present data and of previous results on tymoviral TLS(Val) and bromoviral TLS(Tyr) showing differential effects on translation, we suggest that the key role for the TLS in promoting translation initiation appears to be dictated by the TLS architecture and identity.}, note = {0022-2836 (Print) Journal Article}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The role of RNAs in the regulation of virulence-gene expression}, author = {P Romby and F Vandenesch and E G Wagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16529986}, isbn = {16529986}, year = {2006}, date = {2006-01-01}, journal = {Curr Opin Microbiol}, volume = {9}, number = {2}, pages = {229-236}, abstract = {Bacterial pathogens sense their environment, and in response, virulence genes are induced or repressed through spatial and temporal regulation. They are also subjected to stress conditions, which require appropriate responses. Recent research has revealed that RNAs are key regulators in pathogens. Small RNAs regulate the translation and/or stability of mRNAs that encode virulence proteins, or proteins with roles in adaptive responses, which are triggered by environmental cues and stresses. In most cases, these small RNAs act directly on target RNAs by an antisense mechanism. Other small RNAs act indirectly, by sequestration of regulatory proteins. Direct sensing of environmental signals can occur through induced structural changes in mRNAs.}, note = {1369-5274 (Print) Journal Article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Proximity of conserved U6 and U2 snRNA elements to the 5' splice site region in activated spliceosomes}, author = {B M Rhode and K Hartmuth and E Westhof and R Luhrmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16688215}, isbn = {16688215}, year = {2006}, date = {2006-01-01}, journal = {EMBO J}, volume = {25}, number = {11}, pages = {2475-2486}, abstract = {Major structural changes occur in the spliceosome during its catalytic activation, which immediately precedes the splicing of pre-mRNA. Whereas changes in snRNA conformation are well documented at the level of secondary RNA-RNA interactions, little is known about the tertiary structure of this RNA-RNA network, which comprises the spliceosome's catalytic core. Here, we have used the hydroxyl-radical probe Fe-BABE, tethered to the tenth nucleotide (U(+10)) of the 5' end of a pre-mRNA intron, to map RNA-RNA proximities in spliceosomes. These studies revealed that several conserved snRNA regions are close to U(+10) in activated spliceosomes, namely (i) the U6 snRNA ACAGAG-box region, (ii) portions of the U6 intramolecular stem-loop (U6-ISL) including a nucleotide implicated in the first catalytic step (U74), and (iii) the region of U2 that interacts with the branch point. These data constrain the relative orientation of these structural elements with respect to U(+10) in the activated spliceosome. Upon conversion of the activated spliceosome to complex C, the accessibility of U6-ISL to hydroxyl-radical cleavage is altered, suggesting rearrangements after the first catalytic step.}, note = {0261-4189 (Print) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Biocrystallographica, a package for doing crystallography with Mathematica}, author = { N. Ambert and J. Vanwinsberghe and P. Dumas}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Understanding the importance of selenium and selenoproteins in muscle function}, author = {M Rederstorff and A Krol and A Lescure}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16314926}, isbn = {16314926}, year = {2006}, date = {2006-01-01}, journal = {Cell Mol Life Sci}, volume = {63}, number = {1}, pages = {52-59}, abstract = {Selenium is an essential trace element. In cattle, selenium deficiency causes dysfunction of various organs, including skeletal and cardiac muscles. In humans as well, lack of selenium is associated with many disorders, but despite accumulation of clinical reports, muscle diseases are not generally considered on the list. The goal of this review is to establish the connection between clinical observations and the most recent advances obtained in selenium biology. Recent results about a possible role of selenium-containing proteins in muscle formation and repair have been collected. Selenoprotein N is the first selenoprotein linked to genetic disorders consisting of different forms of congenital muscular dystrophies. Understanding the muscle disorders associated with selenium deficiency or selenoprotein N dysfunction is an essential step in defining the causes of the disease and obtaining a better comprehension of the mechanisms involved in muscle formation and maintenance.}, note = {DOI: 10.1007/s00018-005-5313-y}, keywords = {KROL Selenium Selenoprotein Muscle disorders Congenital muscular dystrophy SEPN1, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Expression, purification, and characterization of a new heterotetramer structure of leucyl-tRNA synthetase from Aquifex aeolicus in Escherichia coli}, author = {N Olieric and G Bey and H Nierengarten and E D Wang and D Moras and G Eriani and J Cavarelli}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16256368}, isbn = {16256368}, year = {2006}, date = {2006-01-01}, journal = {Protein Expr Purif}, volume = {47}, number = {1}, pages = {1-9}, abstract = {Aminoacyl-tRNA synthetases are key players in the interpretation of the genetic code. They constitute a textbook example of multi-domain proteins including insertion and terminal functional modules appended to one of the two class-specific active site domains. The non-catalytic domains usually have distinct roles in the aminoacylation reaction. Aquifex aeolicus leucyl-tRNA synthetase (LeuRS) is composed of a separated catalytic site and tRNA anticodon-binding site, which would represent one of the closest relics of the primordial aminoacyl-tRNA synthetase. Moreover, the essential catalytic site residues are split into the two different subunits. In all other class-I aminoacyl-tRNA synthetases, those two functional polypeptides are nowadays fused into a single protein chain. In this work, we report the isolation and the characterization, in Escherichia coli, of a novel oligomeric form (alphabeta)2 for A. aeolicus LeuRS, which is present in addition to the alphabeta heterodimer. A. aeolicus (alphabeta)2 LeuRS has been characterized by biochemical and biophysical methods. Native gel electrophoresis, mass spectrometry, analytical ultracentrifugation, and kinetic analysis confirmed that the (alphabeta)2 enzyme was a stable and active entity. By mass spectrometry we confirmed that the heterodimer alphabeta can bind one tRNALeu molecule whereas the heterotetramer (alphabeta)2 can bind two tRNALeu molecules. Active site titration and aminoacylation assays showed that two functional active sites are found per heterotetramer, suggesting that this molecular species might exist and be active in vivo. All those data suggest that the existence of the heterotetramer is certainly not an artifact of overexpression in E. coli.}, note = {1046-5928 (Print) Journal Article}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A genome scale location analysis of human Staf/ZNF143-binding sites suggests a widespread role for human Staf/ZNF143 in mammalian promoters}, author = {E Myslinski and M A Gerard and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17092945}, isbn = {17092945}, year = {2006}, date = {2006-01-01}, journal = {J Biol Chem}, volume = {281}, number = {52}, pages = {39953-39962}, abstract = {Staf was originally identified as the transcriptional activator of Xenopus tRNA(Sec) and small nuclear (sn) RNA-type genes. Recently, transcription of seven human (h) protein coding genes was reported to be activated by the human ortholog hStaf/ZNF143. Here we have used a combined in silico and biochemical approach to identify 1175 conserved hStaf/ZNF143-binding sites (SBS) distributed in 938 promoters of four mammalian genomes. The SBS shows a significant positional preference and occurs mostly within 200 bp upstream of the transcription start site. Chromatin immunoprecipitation assays with 295 of the promoters established that 90% contain bona fide SBS. By extrapolating the values of this mapping to the full sizes of the mammalian genomes, we can infer the existence of at least 2500 SBS distributed in 2000 promoters. This unexpected large number strongly suggests that SBS constitutes one of the most widespread transcription factor-binding sites in mammalian promoters. Furthermore, we demonstrated that the presence of the SBS alone is sufficient to direct expression of a luciferase reporter gene, suggesting that hStaf/ZNF143 can recruit per se the transcription machinery.}, note = {0021-9258 (Print) Journal Article}, keywords = {KROL, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Docking of aminoglycosides to hydrated and flexible RNA}, author = {N Moitessier and E Westhof and S Hanessian}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16451068}, isbn = {16451068}, year = {2006}, date = {2006-01-01}, journal = {J Med Chem}, volume = {49}, number = {3}, pages = {1023-1033}, abstract = {Although much effort has been devoted to the development of programs suited for the docking of ligands to proteins, much less progress has been achieved in the nucleic acid field. We have developed a unique approach for docking aminoglycosides to RNA considering the flexibility of these macromolecules using conformational ensembles and accounting for the role of the first hydration shell. This concept, successfully implemented in AutoDock, relies on the computation of the intermolecular interaction energy that accounts for the presence of dynamically bound water molecules to the RNA. As an application, a set of 11 aminoglycosides was docked with an average root-mean-square deviation (RMSD) of 1.41 A to be compared with an average RMSD of 3.25 A when the original AutoDock protocol was used.}, note = {0022-2623 (Print) Journal Article}, keywords = {Molecular Nucleic Acid Conformation RNA/*chemistry Research Support, Non-U.S. Gov't Thermodynamics Water/*chemistry, Unité ARN, WESTHOF Aminoglycosides/*chemistry Anti-Bacterial Agents/*chemistry Crystallography, X-Ray *Models}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {A Modular and Hierarchical Approach for All-Atom RNA Modeling}, author = {B Masquida and E Westhof}, editor = {Cech Atkins T J Gesteland R. F.}, url = {http://cshmonographs.org/index.php/monographs/article/viewArticle/3749}, doi = {10.1101/087969739.43.659}, year = {2006}, date = {2006-01-01}, booktitle = {The RNA World, Third Edition}, pages = {659-681}, publisher = {Cold Spring Harbor Laboratory Press}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {The RNA polymerase III-dependent family of genes in hemiascomycetes: comparative RNomics, decoding strategies, transcription and evolutionary implications}, author = {C Marck and R Kachouri-Lafond and I Lafontaine and E Westhof and B Dujon and H Grosjean}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16600899}, isbn = {16600899}, year = {2006}, date = {2006-01-01}, journal = {Nucleic Acids Res}, volume = {34}, number = {6}, pages = {1816-1835}, abstract = {We present the first comprehensive analysis of RNA polymerase III (Pol III) transcribed genes in ten yeast genomes. This set includes all tRNA genes (tDNA) and genes coding for SNR6 (U6), SNR52, SCR1 and RPR1 RNA in the nine hemiascomycetes Saccharomyces cerevisiae, Saccharomyces castellii, Candida glabrata, Kluyveromyces waltii, Kluyveromyces lactis, Eremothecium gossypii, Debaryomyces hansenii, Candida albicans, Yarrowia lipolytica and the archiascomycete Schizosaccharomyces pombe. We systematically analysed sequence specificities of tRNA genes, polymorphism, variability of introns, gene redundancy and gene clustering. Analysis of decoding strategies showed that yeasts close to S.cerevisiae use bacterial decoding rules to read the Leu CUN and Arg CGN codons, in contrast to all other known Eukaryotes. In D.hansenii and C.albicans, we identified a novel tDNA-Leu (AAG), reading the Leu CUU/CUC/CUA codons with an unusual G at position 32. A systematic 'p-distance tree' using the 60 variable positions of the tRNA molecule revealed that most tDNAs cluster into amino acid-specific sub-trees, suggesting that, within hemiascomycetes, orthologous tDNAs are more closely related than paralogs. We finally determined the bipartite A- and B-box sequences recognized by TFIIIC. These minimal sequences are nearly conserved throughout hemiascomycetes and were satisfactorily retrieved at appropriate locations in other Pol III genes.}, note = {1362-4962 (Electronic) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The interaction networks of structured RNAs}, author = {A Lescoute and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17135184}, isbn = {17135184}, year = {2006}, date = {2006-01-01}, journal = {Nucleic Acids Res}, volume = {34}, number = {22}, pages = {6587-6604}, abstract = {All pairwise interactions occurring between bases which could be detected in three-dimensional structures of crystallized RNA molecules are annotated on new planar diagrams. The diagrams attempt to map the underlying complex networks of base-base interactions and, especially, they aim at conveying key relationships between helical domains: co-axial stacking, bending and all Watson-Crick as well as non-Watson-Crick base pairs. Although such wiring diagrams cannot replace full stereographic images for correct spatial understanding and representation, they reveal structural similarities as well as the conserved patterns and distances between motifs which are present within the interaction networks of folded RNAs of similar or unrelated functions. Finally, the diagrams could help devising methods for meaningfully transforming RNA structures into graphs amenable to network analysis.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {16S/chemistry RNA, 23S/chemistry Ribonuclease P/chemistry, Base Pairing Base Sequence Introns *Models, Catalytic/chemistry RNA, Molecular Molecular Sequence Data Nucleic Acid Conformation RNA/*chemistry RNA, Ribosomal, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The A-minor motifs in the decoding recognition process}, author = {A Lescoute and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16889885}, isbn = {16889885}, year = {2006}, date = {2006-01-01}, journal = {Biochimie}, volume = {88}, number = {8}, pages = {993-999}, abstract = {The formation of A-minor motifs, mediated by adenines binding into the shallow/minor groove of stacked and helical Watson-Crick base pairs, is described. The conformations of the bacterial ribosomal decoding A site in various crystal structures are reviewed. The adenines A1492 and A1493 of the A site are seen either tucked in within the internal loop or bulging out and poised for interaction. This dynamic equilibrium contributes to the decoding process of the codon:anticodon base pairings. Aminoglycoside antibiotics lock the conformation of the A site in a single state with bulged-out adenines and thereby disrupt regulation of the decoding process.}, note = {0300-9084 (Print) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Topology of three-way junctions in folded RNAs}, author = {A Lescoute and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16373494}, isbn = {16373494}, year = {2006}, date = {2006-01-01}, journal = {RNA}, volume = {12}, number = {1}, pages = {83-93}, abstract = {The three-way junctions contained in X-ray structures of folded RNAs have been compiled and analyzed. Three-way junctions with two helices approximately coaxially stacked can be divided into three main families depending on the relative lengths of the segments linking the three Watson-Crick helices. Each family has topological characteristics with some conservation in the non-Watson-Crick pairs within the linking segments as well as in the types of contacts between the segments and the helices. The most populated family presents tertiary interactions between two helices as well as extensive shallow/minor groove contacts between a linking segment and the third helix. On the basis of the lengths of the linking segments, some guidelines could be deduced for choosing a topology for a three-way junction on the basis of a secondary structure. Examples and prediction bas'ed on those rules are discussed.}, note = {1355-8382 (Print) Journal Article}, keywords = {Base Pairing Base Sequence *Crystallography Models, Molecular *Nucleic Acid Conformation RNA/*chemistry/ultrastructure, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The building blocks and motifs of RNA architecture}, author = {N B Leontis and A Lescoute and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16713707}, isbn = {16713707}, year = {2006}, date = {2006-01-01}, journal = {Curr Opin Struct Biol}, volume = {16}, number = {3}, pages = {279-287}, abstract = {RNA motifs can be defined broadly as recurrent structural elements containing multiple intramolecular RNA-RNA interactions, as observed in atomic-resolution RNA structures. They constitute the modular building blocks of RNA architecture, which is organized hierarchically. Recent work has focused on analyzing RNA backbone conformations to identify, define and search for new instances of recurrent motifs in X-ray structures. One current view asserts that recurrent RNA strand segments with characteristic backbone configurations qualify as independent motifs. Other considerations indicate that, to characterize modular motifs, one must take into account the larger structural context of such strand segments. This follows the biologically relevant motivation, which is to identify RNA structural characteristics that are subject to sequence constraints and that thus relate RNA architectures to sequences.}, note = {0959-440X (Print) Journal Article Review}, keywords = {ase Pairing Base Sequence Computational Biology/methods Conserved Sequence *Models, Extramural Research Support, Molecular Nucleic Acid Conformation RNA/*chemistry Research Support, N.I.H., Non-U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The RNA Ontology Consortium: an open invitation to the RNA community}, author = {N B Leontis and R B Altman and H M Berman and S E Brenner and J W Brown and D R Engelke and S C Harvey and S R Holbrook and F Jossinet and S E Lewis and F Major and D H Mathews and J S Richardson and J R Williamson and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16484377}, isbn = {16484377}, year = {2006}, date = {2006-01-01}, journal = {RNA}, volume = {12}, number = {4}, pages = {533-541}, abstract = {The aim of the RNA Ontology Consortium (ROC) is to create an integrated conceptual framework-an RNA Ontology (RO)-with a common, dynamic, controlled, and structured vocabulary to describe and characterize RNA sequences, secondary structures, three-dimensional structures, and dynamics pertaining to RNA function. The RO should produce tools for clear communication about RNA structure and function for multiple uses, including the integration of RNA electronic resources into the Semantic Web. These tools should allow the accurate description in computer-interpretable form of the coupling between RNA architecture, function, and evolution. The purposes for creating the RO are, therefore, (1) to integrate sequence and structural databases; (2) to allow different computational tools to interoperate; (3) to create powerful software tools that bring advanced computational methods to the bench scientist; and (4) to facilitate precise searches for all relevant information pertaining to RNA. For example, one initial objective of the ROC is to define, identify, and classify RNA structural motifs described in the literature or appearing in databases and to agree on a computer-interpretable definition for each of these motifs. To achieve these aims, the ROC will foster communication and promote collaboration among RNA scientists by coordinating frequent face-to-face workshops to discuss, debate, and resolve difficult conceptual issues. These meeting opportunities will create new directions at various levels of RNA research. The ROC will work closely with the PDB/NDB structural databases and the Gene, Sequence, and Open Biomedical Ontology Consortia to integrate the RO with existing biological ontologies to extend existing content while maintaining interoperability.}, note = {1355-8382 (Print) Letter}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Facets of environmental nuclear toxicology}, author = {J J Leguay and R Giege and M T Menager and E Ansoborlo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17079066}, isbn = {17079066}, year = {2006}, date = {2006-01-01}, journal = {Biochimie}, volume = {88}, number = {11}, pages = {1513-1514}, note = {0300-9084 (Print) Editorial}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Two conformational states in the crystal structure of the Homo sapiens cytoplasmic ribosomal decoding A site}, author = {J Kondo and A Urzhumtsev and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16452297}, isbn = {16452297}, year = {2006}, date = {2006-01-01}, journal = {Nucleic Acids Res}, volume = {34}, number = {2}, pages = {676-685}, abstract = {The decoding A site of the small ribosomal subunit is an RNA molecular switch, which monitors codon-anticodon interactions to guarantee translation fidelity. We have solved the crystal structure of an RNA fragment containing two Homo sapiens cytoplasmic A sites. Each of the two A sites presents a different conformational state. In one state, adenines A1492 and A1493 are fully bulged-out with C1409 forming a wobble-like pair to A1491. In the second state, adenines A1492 and A1493 form non-Watson-Crick pairs with C1409 and G1408, respectively while A1491 bulges out. The first state of the eukaryotic A site is, thus, basically the same as in the bacterial A site with bulging A1492 and A1493. It is the state used for recognition of the codon/anticodon complex. On the contrary, the second state of the H.sapiens cytoplasmic A site is drastically different from any of those observed for the bacterial A site without bulging A1492 and A1493.}, note = {1362-4962 (Electronic) Journal Article}, keywords = {16S/chemistry RNA, 18S/*chemistry Research Support, Animals Comparative Study Crystallography, Bacterial/chemistry RNA, Molecular Nebramycin/analogs & derivatives/chemistry Nucleic Acid Conformation RNA, Non-U.S. Gov't Ribosomes/chemistry Tetrahymena thermophila/genetics, Protozoan/chemistry RNA, Ribosomal, Unité ARN, WESTHOF, X-Ray Genetic Code Humans *Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of the homo sapiens cytoplasmic ribosomal decoding site complexed with apramycin}, author = {J Kondo and B Francois and A Urzhumtsev and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16596680}, isbn = {16596680}, year = {2006}, date = {2006-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {45}, number = {20}, pages = {3310-3314}, note = {0570-0833 (Print) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of the bacterial ribosomal decoding site complexed with amikacin containing the gamma-amino-alpha-hydroxybutyryl (haba) group}, author = {J Kondo and B Francois and R J Russell and J B Murray and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16806634}, isbn = {16806634}, year = {2006}, date = {2006-01-01}, journal = {Biochimie}, volume = {88}, number = {8}, pages = {1027-1031}, abstract = {Amikacin is the 4,6-linked aminoglycoside modified at position N1 of the 2-deoxystreptamine ring (ring II) by the L-haba group. In the present study, the crystal structure of a complex between oligonucleotide containing the bacterial ribosomal A site and amikacin has been solved at 2.7 A resolution. Amikacin specifically binds to the A site in practically the same way as its parent compound kanamycin. In addition, the L-haba group interacts with the upper side of the A site through two direct contacts, O2*.H-N4(C1496) and N4*-H.O6(G1497). The present crystal structure shows how the introduction of the L-haba group on ring II of aminoglycoside is an effective mutation for obtaining a higher affinity to the bacterial A site.}, note = {0300-9084 (Print) Journal article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding of human SLBP on the 3'-UTR of histone precursor H4-12 mRNA induces structural rearrangements that enable U7 snRNA anchoring}, author = {S Jaeger and F Martin and J Rudinger-Thirion and R Giege and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16982637}, isbn = {16982637}, year = {2006}, date = {2006-01-01}, journal = {Nucleic Acids Res}, volume = {34}, number = {17}, pages = {4987-4995}, abstract = {In metazoans, cell-cycle-dependent histones are produced from poly(A)-lacking mRNAs. The 3' end of histone mRNAs is formed by an endonucleolytic cleavage of longer precursors between a conserved stem-loop structure and a purine-rich histone downstream element (HDE). The cleavage requires at least two trans-acting factors: the stem-loop binding protein (SLBP), which binds to the stem-loop and the U7 snRNP, which anchors to histone pre-mRNAs by annealing to the HDE. Using RNA structure-probing techniques, we determined the secondary structure of the 3'-untranslated region (3'-UTR) of mouse histone pre-mRNAs H4-12, H1t and H2a-614. Surprisingly, the HDE is embedded in hairpin structures and is therefore not easily accessible for U7 snRNP anchoring. Probing of the 3'-UTR in complex with SLBP revealed structural rearrangements leading to an overall opening of the structure especially at the level of the HDE. Electrophoretic mobility shift assays demonstrated that the SLBP-induced opening of HDE actually facilitates U7 snRNA anchoring on the histone H4-12 pre-mRNAs 3' end. These results suggest that initial binding of the SLBP functions in making the HDE more accessible for U7 snRNA anchoring.}, note = {1362-4962 (Electronic) Journal Article}, keywords = {ERIANI, FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding of neomycin-class aminoglycoside antibiotics to mutant ribosomes with alterations in the A site of 16S rRNA}, author = {S N Hobbie and P Pfister and C Bruell and P Sander and B Francois and E Westhof and E C Bottger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16569869}, isbn = {16569869}, year = {2006}, date = {2006-01-01}, journal = {Antimicrob Agents Chemother}, volume = {50}, number = {4}, pages = {1489-1496}, abstract = {Aminoglycoside antibiotics that bind to the aminoacyl-tRNA site (A site) of the ribosome are composed of a common neamine core in which a glycopyranosyl ring is attached to position 4 of a 2-deoxystreptamine moiety. The core is further substituted by one (ribostamycin), two (neomycin and paromomycin), or three (lividomycin A) additional sugars attached to position 5 of the 2-deoxystreptamine. To study the role of rings III, IV, and V in aminoglycoside binding, we used isogenic Mycobacterium smegmatis DeltarrnB mutants carrying homogeneous populations of mutant ribosomes with alterations in the 16S rRNA A site. MICs were determined to investigate drug-ribosome interactions, and the results were compared with that of the previously published crystal structure of paromomycin bound to the ribosomal A site. Our analysis demonstrates that the stacking interaction between ring I and G1491 is largely sequence independent, that rings III and IV each increase the strength of drug binding to the ribosome, that ring IV of the 6'-NH3+ aminoglycosides compensates for loss of interactions between ring II and U1495 and between ring III and G1491, that the aminoglycosides rely on pseudo-base pairing between ring I and A1408 for binding independently of the number of sugar rings attached to the neamine core, that addition of ring V to the 6'-OH 4,5-aminoglycoside paromomycin does not alter the mode of binding, and that alteration of the U1406.U1495 wobble base pair to the Watson-Crick interaction pair 1406C-1495G yields ribosomal drug susceptibilities to 4,5-aminoglycosides comparable to those seen with the wild-type A site.}, note = {0066-4804 (Print) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mg2+ dependency of HIV-1 reverse transcription, inhibition by nucleoside analogues and resistance}, author = {V Goldschmidt and J Didierjean and B Ehresmann and C Ehresmann and C Isel and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16394022}, isbn = {16394022}, year = {2006}, date = {2006-01-01}, journal = {Nucleic Acids Res}, volume = {34}, number = {1}, pages = {42-52}, abstract = {Metal ions are essential for DNA polymerase and RNase H activities of HIV-1 reverse transcriptase (RT). RT studies are routinely performed at 6-8 mM Mg2+, despite the fact that the in vivo concentration might be as low as 0.2 mM. We studied the influence of MgCl2 and ATP, which likely binds a significant fraction of the magnesium pool in vivo, on the DNA polymerase and RNase H activities of HIV-1 RT, its inhibition by nucleoside RT inhibitors (NRTIs) and primer unblocking by AZT-resistant RT. At low Mg2+ concentration, reverse transcription of a natural template strongly increased despite a dramatically reduced intrinsic polymerase activity under such conditions. Low Mg2+ concentrations affected the RNA stability and indirectly decreased its degradation by the RNase H activity. The reduced RNA degradation prevented premature dissociation of the template and primer strands that otherwise generated dead-end DNA products. In addition, low Mg2+ dramatically decreased the incorporation of NRTIs into DNA and increased nucleotide excision by AZT-resistant RT. The latter effect is also most likely owing to the diminished cleavage of the RNA template. Thus, differences in the free Mg2+ concentration between different cell types or during the cell cycle might strongly affect HIV-1 replication and its inhibition.}, note = {1362-4962 (Electronic) Journal Article}, keywords = {Adenosine Triphosphate/pharmacology Anti-HIV Agents/*pharmacology DNA/biosynthesis DNA Primers DNA, Calf Thymus/metabolism Zidovudine/pharmacology, MARQUET, Non-U.S. Gov't Reverse Transcriptase Inhibitors/*pharmacology *Reverse Transcription/drug effects Ribonuclease H, PAILLART, Single-Stranded/biosynthesis Drug Resistance, Unité ARN, Viral HIV-1 Reverse Transcriptase/*metabolism Magnesium/*pharmacology Nucleosides/pharmacology Research Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transfer RNAs and the RNA world}, author = {R Giege}, url = {none}, year = {2006}, date = {2006-01-01}, journal = {Bol Inf Assoc Nac Bioquimicos (portuguais)}, volume = {8}, pages = {5-8}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The early history of tRNA recognition by aminoacyl-tRNA synthetases}, author = {R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17206068}, isbn = {17206068}, year = {2006}, date = {2006-01-01}, journal = {J Biosci}, volume = {31}, number = {4}, pages = {477-488}, note = {0250-5991 (Print) Journal Article}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Regulatory RNAs as mediators of virulence gene expression in bacteria}, author = {T Geissmann and M Possedko and E Huntzinger and P Fechter and C Ehresmann and P Romby}, editor = {F B Hofmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16594609}, isbn = {16594609}, year = {2006}, date = {2006-01-01}, booktitle = {Handbook of Experimental Pharmacology}, volume = {173}, pages = {9-43}, publisher = {Springer}, abstract = {Bacteria exploit functional diversity of RNAs in a wide range of regulatory mechanisms to control gene expression. In last few years, small RNA molecules have been discovered at a staggering rate in bacteria, mainly in Escherichia coli. While functions of many of these RNA molecules are still not known, several of them behave as key effectors of adaptive responses, such as environmental cue recognition, stress response, and virulence control. Most fascinating, perhaps, is the discovery that mRNAs behave as direct sensors of small molecules or of environmental cues. The astonishing diversity of RNA-dependent regulatory mechanisms is linked to the dynamic properties and versatility of the RNA structure. In this review, we relate several recent studies in different bacterial pathogens that illustrate the diverse roles of RNA to control virulence gene expression.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Loss of a primordial identity element for a mammalian mitochondrial aminoacylation system}, author = {A Fender and C Sauter and M Messmer and J Putz and R Giege and C Florentz and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16597625}, isbn = {16597625}, year = {2006}, date = {2006-01-01}, journal = {J Biol Chem}, volume = {281}, number = {23}, pages = {15980-15986}, abstract = {In mammalian mitochondria the translational machinery is of dual origin with tRNAs encoded by a simplified and rapidly evolving mitochondrial (mt) genome and aminoacyl-tRNA synthetases (aaRS) coded by the nuclear genome, and imported. Mt-tRNAs are atypical with biased sequences, size variations in loops and stems, and absence of residues forming classical tertiary interactions, whereas synthetases appear typical. This raises questions about identity elements in mt-tRNAs and adaptation of their cognate mt-aaRSs. We have explored here the human mt-aspartate system in which a prokaryotic-type AspRS, highly similar to the Escherichia coli enzyme, recognizes a bizarre tRNA(Asp). Analysis of human mt-tRNA(Asp) transcripts confirms the identity role of the GUC anticodon as in other aspartylation systems but reveals the non-involvement of position 73. This position is otherwise known as the site of a universally conserved major aspartate identity element, G73, also known as a primordial identity signal. In mt-tRNA(Asp), position 73 can be occupied by any of the four nucleotides without affecting aspartylation. Sequence alignments of various AspRSs allowed placing Gly-269 at a position occupied by Asp-220, the residue contacting G73 in the crystallographic structure of E. coli AspRS-tRNA(Asp) complex. Replacing this glycine by an aspartate renders human mt-AspRS more discriminative to G73. Restriction in the aspartylation identity set, driven by a rapid mutagenic rate of the mt-genome, suggests a reverse evolution of the mt-tRNA(Asp) identity elements in regard to its bacterial ancestor.}, note = {0021-9258 (Print) Journal Article}, keywords = {Acylation Base Sequence Humans Kinetics Mutagenesis Nucleic Acid Conformation Plasmids RNA | Non-U.S. Gov't, Asp/chemistry/genetics/*metabolism Research Support, FLORENTZ, FRUGIER, Non-U.S. Gov't, SAUTER, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Polymorphism of bulged-out residues in HIV-1 RNA DIS kissing complex and structure comparison with solution studies}, author = {E Ennifar and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16403527}, isbn = {16403527}, year = {2006}, date = {2006-01-01}, journal = {J Mol Biol}, volume = {356}, number = {3}, pages = {771-782}, abstract = {All retroviruses encapsidate their genome as a dimer of homologous single-stranded RNAs. The dimerization initiation site (DIS) of human immunodeficiency virus type 1 (HIV-1) is located in the 5'-untranslated region of the viral genome and consists of a hairpin with a 6 nt self-complementary loop sequence. Genomic RNA dimerization, a crucial step for virion infectivity, is promoted by the formation of a loop-loop complex (or kissing complex) between two DIS hairpins. Crystal structures for the subtypes A, B and F of the HIV-1 DIS kissing complex have now been solved at 2.3 A, 1.9 A and 1.6 A, respectively. They revealed a polymorphism of bulged-out residues showing clearly that their conformation is not a mere consequence of crystal packing. They also provide more insights into ion binding, hydration, and RNA conformation and flexibility. In particular, we observed the binding of spermine to the loop-loop helix, which displaced a magnesium cation important for subtype A DIS dimerization. The excellent agreement between X-ray structures and the results of chemical probing and interference data on larger viral RNA fragments shows that the crystal structures are relevant for the DIS kissing complex present in solution and in viral particles. Accordingly, these structures will be helpful for designing new drugs derived from aminoglycoside antibiotics and targeted against the RNA dimerization step of the viral life-cycle.}, note = {0022-2836 (Print) Journal Article}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Einstein's tongue for teaching crystallography to biologists}, author = {P Dumas and J Vanwinsberghe and V Cura}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {A single tRNA base pair mediates bacterial tRNA-dependent biosynthesis of asparagine}, author = {M Bailly and S Giannouli and M Blaise and C Stathopoulos and D Kern and H D Becker}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17074748}, isbn = {17074748}, year = {2006}, date = {2006-01-01}, journal = {Nucleic Acids Res}, volume = {34}, number = {21}, pages = {6083-6094}, abstract = {In many prokaryotes and in organelles asparagine and glutamine are formed by a tRNA-dependent amidotransferase (AdT) that catalyzes amidation of aspartate and glutamate, respectively, mischarged on tRNAAsn and tRNAGln. These pathways supply the deficiency of the organism in asparaginyl- and glutaminyl-tRNA synthtetases and provide the translational machinery with Asn-tRNAAsn and Gln-tRNAGln. So far, nothing is known about the structural elements that confer to tRNA the role of a specific cofactor in the formation of the cognate amino acid. We show herein, using aspartylated tRNAAsn and tRNAAsp variants, that amidation of Asp acylating tRNAAsn is promoted by the base pair U1-A72 whereas the G1-C72 pair and presence of the supernumerary nucleotide U20A in the D-loop of tRNAAsp prevent amidation. We predict, based on comparison of tRNAGln and tRNAGlu sequence alignments from bacteria using the AdT-dependent pathway to form Gln-tRNAGln, that the same combination of nucleotides also rules specific tRNA-dependent formation of Gln. In contrast, we show that the tRNA-dependent conversion of Asp into Asn by archaeal AdT is mainly mediated by nucleotides G46 and U47 of the variable region. In the light of these results we propose that bacterial and archaeal AdTs use kingdom-specific signals to catalyze the tRNA-dependent formations of Asn and Gln.}, note = {1362-4962 (Electronic) Journal Article Research Support, Non-U.S. Gov't}, keywords = {KERN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Molecular dynamics simulations of RNA systems: importance of the initial conditions}, author = {P Auffinger}, editor = {J Sponer and F Lankas}, url = {http://www-ibmc.u-strasbg.fr/arn/Westhof/publ_West/bib2006/r2006_PAuffinger_jsponer.pdf}, year = {2006}, date = {2006-01-01}, booktitle = {Computational Studies of DNA and RNA}, pages = {283-300}, publisher = {Springer Verlag}, series = {Challenges and Advances in Computational Chemistry and Physics, vol. 2}, abstract = {Several factors can lead to the generation of accurate and informative molecular dynamics simulations. Among them, the choice of an appropriate starting structure is primordial. Unfortunately, experimental structures are not void of inaccuracies and errors. The aim of this review is to provide practical guidelines for detecting and correcting such inaccuracies in order to start molecular dynamics simulations of nucleic acid systems under the best possible conditions.}, keywords = {Unité ARN, WESTHOF Molecular dynamics simulation RNA Nucleic acids Interatomic interactions}, pubstate = {published}, tppubtype = {inbook} } @misc{, title = {Biocrystallographica, a package for doing crystallography with Mathematica}, author = {N Ambert and J Vanwinsberghe and P Dumas}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS, Unité ARN}, pubstate = {published}, tppubtype = {misc} } @article{, title = {Selenoprotein synthesis: UGA does not end the story}, author = {C Allmang and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16737768}, isbn = {16737768}, year = {2006}, date = {2006-01-01}, journal = {Biochimie}, volume = {88}, number = {11}, pages = {1561-1571}, abstract = {It is well established that the beneficial effects of the trace element selenium are mediated by its major biological product, the amino acid selenocysteine, present in the active site of selenoproteins. These fulfill different functions, as varied as oxidation-reduction of metabolites in bacteria, reduction of reactive oxygen species, control of the redox status of the cell or thyroid hormone maturation. This review will focus on the singularities of the selenocysteine biosynthesis pathway and its unique incorporation mechanism into eukaryal selenoproteins. Selenocysteine biosynthesis from serine is achieved on tRNA(Sec) and requires four proteins. As this amino acid is encoded by an in-frame UGA codon, otherwise signaling termination of translation, ribosomes must be told not to stop at this position in the mRNA. Several molecular partners acting in cis or in trans have been identified, but their knowledge has not enabled yet to firmly establish the molecular events underlying this mechanism. Data suggest that other, so far uncharacterized factors might exist. In this survey, we attempted to compile all the data available in the literature and to describe the latest developments in the field.}, note = {0300-9084 (Print) Journal Article}, keywords = {KROL, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {SECIS RNAs and K-turn binding proteins. A survey of evolutionary conserved RNA and protein motifs.}, author = {C Allmang and A Krol}, editor = {D L Hatfield and M J Berry and V N Gladyshev}, url = {http://www.springerlink.com/content/m854145547561671}, isbn = {10.1007/0-387-33827-6_5}, year = {2006}, date = {2006-01-01}, booktitle = {Selenium, Its Molecular Biology and Role in Human Health}, pages = {51-61}, publisher = {Springer US}, abstract = {The SelenoCysteine Insertion Sequence (SECIS) is a stem-loop structure residing in the 3? untranslated region of all selenoprotein mRNAs. Its presence is mandatory to allow the ribosome to readthrough the UGA selenocysteine codon. The SECIS RNA possesses a well-defined secondary structure. Four consecutive non-Watson-Crick base pairs, with a central tandem of sheared G.A/A.G base pairs, constitute the functional motif of the SECIS RNA which is recognized by the SECIS binding protein SBP2. The tandem of sheared base pairs is part of a recurrent motif, the kink-turn (K-turn), occurring in a variety of different RNAs. The K-turn is a helix-internal loop-helix composed of a non-Watson-Crick stem containing the G.A base pairs and a canonical stem. The internal loop between the stems is always asymmetrical and usually contains three unpaired nucleotides on one strand and none on the other. We propose here that the SECIS RNA must represent a K-turn variant with regard to the limited structural differences that distinguish it from consensus K-turns. Work by others showed that ribosomal protein L30 also binds the SECIS RNA in a specific manner. L30 and SBP2 are members of a family of proteins sharing the same RNA-binding domain called L7A/L30. All proteins possessing this fold recognize K-turn RNAs. Three structures of RNA-protein complexes containing the L7A/L30 protein fold and cognate K-turn RNAs have been solved. In light of the interaction principles governing these RNA-protein complexes, we discuss how L30 can recognize the SECIS RNA. Collectively, all the findings suggest that the L7A/L30 protein fold and the K-turn are ancient structural motifs that have evolved various functions, from pre-mRNA splicing to protein synthesis.}, keywords = {KROL, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {A single homozygous point mutation in a 3'untranslated region motif of selenoprotein N mRNA causes SEPN1-related myopathy}, author = {V Allamand and P Richard and A Lescure and C Ledeuil and D Desjardin and N Petit and C Gartioux and A Ferreiro and A Krol and N Pellegrini and J A Urtizberea and P Guicheney}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16498447}, isbn = {16498447}, year = {2006}, date = {2006-01-01}, journal = {EMBO Rep}, volume = {7}, number = {4}, pages = {450-454}, abstract = {Mutations in the SEPN1 gene encoding the selenoprotein N (SelN) have been described in different congenital myopathies. Here, we report the first mutation in the selenocysteine insertion sequence (SECIS) of SelN messenger RNA, a hairpin structure located in the 3' untranslated region, in a patient presenting a classical although mild form of rigid spine muscular dystrophy. We detected a significant reduction in both mRNA and protein levels in the patient's skin fibroblasts. The SECIS element is crucial for the insertion of selenocysteine at the reprogrammed UGA codon by recruiting the SECIS-binding protein 2 (SBP2), and we demonstrated that this mutation abolishes SBP2 binding to SECIS in vitro, thereby preventing co-translational incorporation of selenocysteine and SelN synthesis. The identification of this mutation affecting a conserved base in the SECIS functional motif thereby reveals the structural basis for a novel pathological mechanism leading to SEPN1-related myopathy.}, note = {1469-221X (Print) Journal article}, keywords = {KROL, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Einstein's tongue for teaching crystallography to biologists}, author = { P. Dumas and J. Vanwinsberghe and V. Cura}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS}, pubstate = {published}, tppubtype = {misc} } @article{campidelli_functionalization_2006, title = {Functionalization of CNT: synthesis and applications in photovoltaics and biology}, author = {Stéphane Campidelli and Cédric Klumpp and Alberto Bianco and Dirk M Guldi and Maurizio Prato}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/poc.1052}, doi = {10.1002/poc.1052}, issn = {1099-1395}, year = {2006}, date = {2006-01-01}, urldate = {2020-03-31}, journal = {Journal of Physical Organic Chemistry}, volume = {19}, number = {8-9}, pages = {531--539}, abstract = {Here, we review part of the work carried out in our laboratories on carbon nanotube functionalization. Both covalent (sidewall derivatization) and non-covalent (using π-π interactions) functionalization have been used to solubilize carbon nanotubes (NTs). The combination of NTs with various electron donors, mainly using the supramolecular approach, led to a new generation of donor-acceptor nanohybrids which can be used for the development of carbon-based photovoltaic cells. Covalent functionalization has been successfully applied for preparation of water soluble nanotubes and further derivatization of the nanotubes with bioactive molecules hold great promise for application in drug, vaccine and gene delivery. Copyright © 2006 John Wiley & Sons, Ltd.}, keywords = {Carbon nanotubes, Cells, Drug delivery, electron transfer, Functionalization, I2CT, Peptides, photovoltaic, Team-Bianco, Toxicity, Vectors}, pubstate = {published}, tppubtype = {article} } @article{lacerda_luminescence_2006, title = {Luminescence of Functionalized Carbon Nanotubes as a Tool to Monitor Bundle Formation and Dissociation in Water: The Effect of Plasmid-DNA Complexation}, author = {L Lacerda and G Pastorin and W Wu and M Prato and A Bianco and K Kostarelos}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adfm.200500569}, doi = {10.1002/adfm.200500569}, issn = {1616-3028}, year = {2006}, date = {2006-01-01}, urldate = {2020-03-31}, journal = {Advanced Functional Materials}, volume = {16}, number = {14}, pages = {1839--1846}, abstract = {Functionalized carbon nanotubes (f-CNTs) are explored as novel nanomaterials for biomedical applications. UV-vis luminescence of aqueous dispersions of CNT–NH3+ and CNT–NH–Ac (NH–Ac: acetamido) is observed using standard laboratory spectrophotometric instrumentation, and the measured fluorescence intensity is correlated with the aggregation state of the f-CNTs: a high intensity indicates improved f-CNT individualization and dispersion, while a decrease in fluorescence intensity indicates a higher degree of nanotube aggregation and bundling as a result of varying the sodium dodecyl sulfate (SDS) concentrations and pH in the aqueous phase. Moreover, utilization of this relationship between fluorescence intensity and the state of f-CNT aggregation is carried out to elucidate the interactions between f-CNTs and gene-encoding plasmid DNA (pDNA). pDNA is shown to interact with CNT–NH3+ primarily through electrostatic interactions that lead concomitantly to a higher degree of f-CNT bundling. The CNT–NH3+/pDNA interactions are successfully competed by SDS/f-CNT surface interactions, resulting in the displacement of pDNA. These studies provide exemplification of the use of fluorescence spectrophotometry to accurately describe the aggregation state of water-soluble f-CNTs. Characterization of the complexes between pDNA and f-CNTs elucidates the opportunities and limitations of such supramolecular systems as potential vectors for gene transfer.}, keywords = {Carbon nanotubes, DNA, I2CT, Luminescence, multiwalled, single-walled, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Biocrystallographica, a package for doing crystallography with Mathematica}, author = {N Ambert and J Vanwinsberghe and P Dumas}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS}, pubstate = {published}, tppubtype = {misc} } @misc{, title = {Biocrystallographica, a package for doing crystallography with Mathematica}, author = {N Ambert and J Vanwinsberghe and P Dumas}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS}, pubstate = {published}, tppubtype = {misc} } @incollection{, title = {Molecular dynamics simulations of RNA systems: importance of the initial conditions}, author = {P Auffinger}, editor = {J Sponer and F Lankas}, year = {2006}, date = {2006-01-01}, booktitle = {Computational Studies of DNA and RNA}, pages = {283-300}, publisher = {Springer Verlag}, series = {Challenges and Advances in Computational Chemistry and Physics, vol. 2}, abstract = {Several factors can lead to the generation of accurate and informative molecular dynamics simulations. Among them, the choice of an appropriate starting structure is primordial. Unfortunately, experimental structures are not void of inaccuracies and errors. The aim of this review is to provide practical guidelines for detecting and correcting such inaccuracies in order to start molecular dynamics simulations of nucleic acid systems under the best possible conditions.}, keywords = {WESTHOF Molecular dynamics simulation RNA Nucleic acids Interatomic interactions}, pubstate = {published}, tppubtype = {incollection} } @article{pelte_immune_2006, title = {Immune challenge induces N-terminal cleavage of the Drosophila serpin Necrotic}, author = {Nadège Pelte and Andrew S Robertson and Zhen Zou and Didier Belorgey and Timothy R Dafforn and Haobo Jiang and David Lomas and Jean-Marc Reichhart and David Gubb}, doi = {10.1016/j.ibmb.2005.10.004}, issn = {0965-1748}, year = {2006}, date = {2006-01-01}, journal = {Insect Biochem. Mol. Biol.}, volume = {36}, number = {1}, pages = {37--46}, abstract = {The Drosophila Necrotic protein is a serine proteinase inhibitor, which regulates the Toll-mediated innate immune response. Necrotic specifically inhibits an extracellular serine proteinase cascade leading to activation of the Toll ligand, Spätzle. Necrotic carries a polyglutamine extension amino-terminal to the core serpin structure. We show here that cleavage of this N-terminal extension occurs following immune challenge. This modification is blocked in PGRP-SA(semmelweiss) mutants after Gram-positive bacterial challenge and in persephone mutants after fungal or Gram-positive bacterial challenge, indicating that activation of either of the Toll pathway upstream branches induces N-terminal cleavage of the serpin. The absolute requirement of persephone gene product for this cleavage indicates that Gram-positive bacteria activate a redundant set of proteinases upstream of Toll. Both full-length Necrotic and the core serpin are active inhibitors of a range of serine proteinases: the highest affinity being for cathepsin G and elastases. We found a 13-fold increase in the specificity of the core serpin over that of full-length Necrotic for one of the tested proteinases (porcine pancreatic elastase). This finding indicates that cleavage of the Necrotic amino-terminal extension might modulate Toll activation following the initial immune response.}, keywords = {Animals, Gene Expression Regulation, M3i, Protein Conformation, reichhart, Serpins, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{galiana-arnoux_toll-like_2006, title = {Toll-like receptors and innate antiviral immunity}, author = {Delphine Galiana-Arnoux and Jean-Luc Imler}, doi = {10.1111/j.1399-0039.2006.00583.x}, issn = {0001-2815}, year = {2006}, date = {2006-01-01}, journal = {Tissue Antigens}, volume = {67}, number = {4}, pages = {267--276}, abstract = {Viral infections are first detected by a set of innate immunity receptors that detect primary infections by pathogens, and trigger a transcriptional response. Among the induced target genes, type I interferons (IFNs) are central to the antiviral response of the host. The receptors and signaling pathways that mediate the strong induction of the synthesis of these cytokines have long remained elusive. In the past few years, Toll-like receptors (TLRs) emerged as important sensors of infections. Several TLRs participate in the recognition of virus infection, interacting in particular with viral nucleic acids. Upon activation, TLRs interact with different cytosolic adapter molecules and activate transcription factors of the nuclear factor-kappaB and IFN regulatory factor families that concur to mediate induction of IFN-alpha/beta and other inflammatory cytokines. In addition to the transmembrane TLRs, cytosolic helicases also detect viral nucleic acids, and trigger type I IFN synthesis.}, keywords = {Animals, Humans, imler, Immunity, Innate, M3i, Signal Transduction, Toll-Like Receptors, Virus Diseases}, pubstate = {published}, tppubtype = {article} } @misc{, title = {Einstein's tongue for teaching crystallography to biologists}, author = {P Dumas and J Vanwinsberghe and V Cura}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS}, pubstate = {published}, tppubtype = {misc} } @misc{, title = {Einstein's tongue for teaching crystallography to biologists}, author = {P Dumas and J Vanwinsberghe and V Cura}, year = {2006}, date = {2006-01-01}, volume = {2006}, publisher = {IUCR}, keywords = {DUMAS}, pubstate = {published}, tppubtype = {misc} } @article{marmey_cd14_2006, title = {CD14 and CD169 expression in human lymph nodes and spleen: specific expansion of CD14+C}, author = {B Marmey and C Boix and J B Barbaroux and M C Dieu-Nosjean and J Diebold and J Audouin and W H Fridman and C G Mueller and T J Molina}, year = {2006}, date = {2006-01-01}, journal = {Hum.Pathol.}, volume = {37}, number = {0046-8177 (Print)}, pages = {68--77}, abstract = {The mononuclear phagocyte system of human lymphoid tissue comprises macrophages and dendritic cells (DCs). The heterogeneity of the non-DC mononuclear phagocyte population in human lymphoid tissue has been little addressed. Here, we studied the expression of 2 monocyte-derived markers, CD14 and CD169 (sialoadhesin), in reactive human lymphoid tissue as well as in a series of 51 B-cell lymphomas by immunohistochemistry on paraffin-embedded tissue. We confirmed that lymph node sinusoidal monocyte-derived cells were the only population staining for CD169. Although most sinusoidal histiocytes also expressed CD14, monocyte-derived cells with phagocytosis such as erythrophagocytosis, anthracosis, or tingible bodies macrophage lacked CD14 and CD169. Among B-cell lymphomas, splenic marginal zone lymphoma was the only one associated with an expansion of the CD14(+)CD169(+) cells in the cords. With respect to nodal B-cell lymphomas, CD14(+) cells were rare among B-chronic lymphocytic leukemia, follicular lymphoma (FL), mantle cell lymphoma (MCL). However, strikingly, we found a strong expansion of CD14(+)CD169(-) cells in numerous diffuse large B-cell lymphomas (DLBCLs), except in cases associated with numerous mitoses, apoptotic bodies, and tingible bodies macrophages. When cultivated in granulocyte/macrophage colony stimulating factor/interleukin 4, DLBCL purified CD14(+) cells differentiate into plasmacytoid cells, expressing DC-specific intercellular adhesion molecule 3-grabbing nonintegrin, suggesting dendritic cell differentiation potential. Our observation fits well with the lymph node and host response cluster signatures described in the gene profiling signatures of DLBCL. However, the role of this CD14(+) population that may constitute a microenvironment-related marker of this subgroup of DLBCL remains to be determined}, keywords = {Adhesion, Antigen, Antigens, B-Cell, Biological, CD14, Cell Differentiation, CELL SEPARATION, Dendritic Cells, Differentiation, Diffuse, Direct, Expression, Flow Cytometry, Fluorescent Antibody Technique, Gene, GLYCOPROTEIN, Glycoproteins, granulocyte/macrophage-colony, Human, Humans, Immunoenzyme Techniques, Immunohistochemistry, Immunologic, Large B-Cell, leukemia, LYMPH, LYMPH NODE, Lymph Nodes, Lymphadenitis, Lymphoid Tissue, LYMPHOMA, Macrophage, Macrophages, Membrane, Membrane Glycoproteins, metabolism, Monocytes, pathology, Phagocytosis, Receptor, Receptors, SIALOADHESIN, SPLEEN, Team-Mueller, tumor, Tumor Markers}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cooperation between Reverse Transcriptase and Integrase during Reverse Transcription and Formation of the Preintegrative Complex of Ty1}, author = { M. Wilhelm and F. X. Wilhelm}, year = {2006}, date = {2006-01-01}, journal = {Eukaryot Cell}, volume = {5}, number = {10}, pages = {1760-9}, abstract = {Reverse transcriptase (RT) and integrase (IN) play a central role in the replication and transposition of retroelements. Increasing evidence suggests that the interaction between these two enzymes is functional and plays an important role in replication. In the yeast Saccharomyces cerevisiae retrotransposon Ty1, the interaction of IN with RT is critical for the formation of an active conformation of RT. We show here that the RT associated with VLPs is active only if it is in close interaction with IN. To probe the IN-RT cis-trans relationship, we have used a complementation assay based on coexpressing two transposons. We show that IN acts in cis to activate RT and that a functional integrase provided in trans is not able to complement replication and transposition defects of IN deletion or IN active-site mutant elements. Our data support a model in which IN not only interacts closely with RT during reverse transcription but also remains associated with RT during the formation of the preintegrative complex.}, note = {1535-9778 (Print) Journal Article}, keywords = {MARQUET}, pubstate = {published}, tppubtype = {article} } @article{barbaroux_tumor_2006, title = {Tumor necrosis factor-alpha- and IL-4-independent development of Langerhans cell-like dendritic cells from M-CSF-conditioned precursors}, author = {Jean-Baptiste Barbaroux and Wing-Hong Kwan and Jean-Pierre Allam and Natalija Novak and Thomas Bieber and Wolf H Fridman and Richard Groves and Chris G Mueller}, doi = {10.1038/sj.jid.5700023}, issn = {0022-202X}, year = {2006}, date = {2006-01-01}, journal = {The Journal of Investigative Dermatology}, volume = {126}, number = {1}, pages = {114--120}, abstract = {GM-CSF and transforming growth factor beta (TGFbeta ) are required for the generation of Langerhans cells (LC), members of the dendritic cell (DC) family. Tumor necrosis factor alpha (TNFalpha) and IL-4 can enhance LC differentiation from human monocytes or CD34(+) progenitors. Here, we show that M-CSF-cultured DC precursors derived from CD34(+) progenitors resemble dermal CD14(+) cells and readily convert to LC-like DC in GM-CSF/TGFbeta. The cells express Langerin, CD1a, and CCR6, migrate in response to CCR6 ligand CCL20, and contain Birbeck granules. TNFalpha and IL-4, added separately or together, have an inhibitory effect on LC differentiation. Cells differentiated in the presence of IL-4 and TNFalpha express low levels of CCR7. This suggests that M-CSF-conditioned DC precursors retain the capacity to efficiently undergo a differentiation program, giving rise to LC-like DC solely through the effect of GM-CSF and TGFbeta.}, keywords = {Antigens, C-Type, Carrier Proteins, CC, CCR6, CD, CD1, CD34, Cell Differentiation, Chemokine, Chemokine CCL20, chemokines, Cytokines, DERMIS, FRANZ, Granulocyte-Macrophage Colony-Stimulating Factor, Hematopoietic Stem Cells, Humans, IL-4, Interleukin-4, Langerhans Cells, Lectins, Lipopolysaccharide Receptors, M-CSF, Macrophage Colony-Stimulating Factor, Macrophage Inflammatory Proteins, Mannose-Binding Lectins, Membrane Glycoproteins, murine, RANK ligand, Receptor Activator of Nuclear Factor-kappa B, Receptors, Surface, Team-Mueller, TNF ALPHA, Tumor Necrosis Factor-alpha}, pubstate = {published}, tppubtype = {article} } @article{Ennifar2006, title = {Targeting the dimerization initiation site of HIV-1 RNA with aminoglycosides: from crystal to cell}, author = {E Ennifar and J C Paillart and A Bodlenner and P Walter and J M Weibel and A M Aubertin and P Pale and P Dumas and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16679451}, doi = {10.1093/nar/gkl317 }, year = {2006}, date = {2006-01-01}, journal = {Nucleic Acids Research}, volume = {34}, number = {8}, pages = {2328-39}, abstract = {The kissing-loop complex that initiates dimerization of genomic RNA is crucial for Human Immunodeficiency Virus Type 1 (HIV-1) replication. We showed that owing to its strong similitude with the bacterial ribosomal A site it can be targeted by aminoglycosides. Here, we present its crystal structure in complex with neamine, ribostamycin, neomycin and lividomycin. These structures explain the specificity for 4,5-disubstituted 2-deoxystreptamine (DOS) derivatives and for subtype A and subtype F kissing-loop complexes, and provide a strong basis for rational drug design. As a consequence of the different topologies of the kissing-loop complex and the A site, these aminoglycosides establish more contacts with HIV-1 RNA than with 16S RNA. Together with biochemical experiments, they showed that while rings I, II and III confer binding specificity, rings IV and V are important for affinity. Binding of neomycin, paromomycin and lividomycin strongly stabilized the kissing-loop complex by bridging the two HIV-1 RNA molecules. Furthermore, in situ footprinting showed that the dimerization initiation site (DIS) of HIV-1 genomic RNA could be targeted by these aminoglycosides in infected cells and virions, demonstrating its accessibility.}, keywords = {Aminoglycosides/*chemistry Anti-HIV Agents/*chemistry Binding Sites Cell Line Crystallography, ENNIFAR, MARQUET, Molecular RNA, Non-U.S. Gov't Virion/chemistry, PAILLART, Unité ARN, Viral/*chemistry Research Support, X-Ray Dimerization Drug Delivery Systems HIV-1/*genetics Humans Models}, pubstate = {published}, tppubtype = {article} } @article{fournel_c3-symmetric_2005, title = {C3-symmetric peptide scaffolds are functional mimetics of trimeric CD40L}, author = {Sylvie Fournel and Sébastien Wieckowski and Weimin Sun and Nathalie Trouche and Hélène Dumortier and Alberto Bianco and Olivier Chaloin and Mohammed Habib and Jean-Christophe Peter and Pascal Schneider and Bernard Vray and René E Toes and Rienk Offringa and Cornelis J M Melief and Johan Hoebeke and Gilles Guichard}, doi = {10.1038/nchembio746}, issn = {1552-4450}, year = {2005}, date = {2005-12-01}, journal = {Nature Chemical Biology}, volume = {1}, number = {7}, pages = {377--382}, abstract = {Interaction between CD40, a member of the tumor necrosis factor receptor (TNFR) superfamily, and its ligand CD40L, a 39-kDa glycoprotein, is essential for the development of humoral and cellular immune responses. Selective blockade or activation of this pathway provides the ground for the development of new treatments against immunologically based diseases and malignancies. Like other members of the TNF superfamily, CD40L monomers self-assemble around a threefold symmetry axis to form noncovalent homotrimers that can each bind three receptor molecules. Here, we report on the structure-based design of small synthetic molecules with C3 symmetry that can mimic CD40L homotrimers. These molecules interact with CD40, compete with the binding of CD40L to CD40, and reproduce, to a certain extent, the functional properties of the much larger homotrimeric soluble CD40L. Architectures based on rigid C3-symmetric cores may thus represent a general approach to mimicking homotrimers of the TNF superfamily.}, keywords = {Animals, Apoptosis, Biological, CD40 Antigens, CD40 Ligand, Cell Line, Dumortier, Humans, I2CT, Inbred BALB C, Mice, Models, Molecular Mimicry, Molecular Structure, Peptides, Protein Conformation, Protein Structure, Quaternary, Structure-Activity Relationship, Team-Bianco, Team-Dumortier, Time Factors, tumor}, pubstate = {published}, tppubtype = {article} } @article{hayer_aberrant_2005, title = {Aberrant expression of the autoantigen heterogeneous nuclear ribonucleoprotein-A2 (RA33) and spontaneous formation of rheumatoid arthritis-associated anti-RA33 autoantibodies in TNF-alpha transgenic mice}, author = {Silvia Hayer and Makiyeh Tohidast-Akrad and Silva Haralambous and Beatrice Jahn-Schmid and Karl Skriner and Sylvie Trembleau and Hélène Dumortier and Serafin Pinol-Roma and Kurt Redlich and Georg Schett and Sylviane Muller and George Kollias and Josef Smolen and Günter Steiner}, doi = {10.4049/jimmunol.175.12.8327}, issn = {0022-1767}, year = {2005}, date = {2005-12-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {175}, number = {12}, pages = {8327--8336}, abstract = {Human TNF-alpha transgenic (hTNFtg) mice develop erosive arthritis closely resembling rheumatoid arthritis (RA). To investigate mechanisms leading to pathological autoimmune reactions in RA, we examined hTNFtg animals for the presence of RA-associated autoantibodies including Abs to citrullinated epitopes (anti-cyclic citrullinated peptide), heterogeneous nuclear ribonucleoprotein (hnRNP)-A2 (anti-RA33), and heat shock proteins (hsp) (anti-hsp). Although IgM anti-hsp Abs were detected in 40% of hTNFtg and control mice, IgG anti-hsp Abs were rarely seen, and anti-cyclic citrullinated peptide Abs were not seen at all. In contrast, textgreater50% of hTNFtg mice showed IgG anti-RA33 autoantibodies, which became detectable shortly after the onset of arthritis. These Abs were predominantly directed to a short epitope, which was identical with an epitope previously described in MRL/lpr mice. Incidence of anti-RA33 was significantly decreased in mice treated with the osteoclast inhibitor osteoprotegerin and also in c-fos-deficient mice lacking osteoclasts. Pronounced expression of hnRNP-A2 and a smaller splice variant was seen in joints of hTNFtg mice, whereas expression was low in control animals. Although the closely related hnRNP-A1 was also overexpressed, autoantibodies to this protein were infrequently detected. Because expression of hnRNP-A2 in thymus, spleen, brain, and lung was similar in hTNFtg and control mice, aberrant expression appeared to be restricted to the inflamed joint. Finally, immunization of hTNFtg mice with recombinant hnRNP-A2 or a peptide harboring the major B cell epitope aggravated arthritis. These findings suggest that overproduction of TNF-alpha leads to aberrant expression of hnRNP-A2 in the rheumatoid joint and subsequently to autoimmune reactions, which may enhance the inflammatory and destructive process.}, keywords = {Animals, Antibody Formation, arthritis, Autoantibodies, Autoantigens, Dumortier, Gene Expression Regulation, Heterogeneous-Nuclear Ribonucleoprotein Group A-B, Humans, I2CT, Joints, Mice, rheumatoid, Team-Dumortier, Tissue Distribution, transgenic, Tumor Necrosis Factor-alpha}, pubstate = {published}, tppubtype = {article} } @article{reichhart_tip_2005, title = {Tip of another iceberg: Drosophila serpins}, author = {Jean-Marc Reichhart}, doi = {10.1016/j.tcb.2005.10.001}, issn = {0962-8924}, year = {2005}, date = {2005-12-01}, journal = {Trends Cell Biol.}, volume = {15}, number = {12}, pages = {659--665}, abstract = {Serpins are serine protease inhibitors with a conserved structure that have been identified in nearly all species and act as suicide substrates by binding covalently to their target proteases. Serpins regulate various physiological processes and defence mechanisms. In humans, several serpin mutations are linked to diseases. The genome of Drosophila melanogaster encodes 29 serpins and even more serine proteases. To date, three serpins have been investigated in detail. Spn27A controls the Toll pathway during early development and is involved in defence reactions in adult flies. SPN42DaA is an inhibitor of furin, a subtilisin-like convertase that is required for pro-protein maturation. Spn43Ac controls the Toll pathway during the immune response. In each case, Drosophila genetics has shed new light on the function of these serine protease inhibitors.}, keywords = {Animals, Immunity, M3i, Protein Conformation, reichhart, Serine Proteinase Inhibitors, Serpins}, pubstate = {published}, tppubtype = {article} } @article{bianco_applications_2005, title = {Applications of carbon nanotubes in drug delivery}, author = {Alberto Bianco and Kostas Kostarelos and Maurizio Prato}, doi = {10.1016/j.cbpa.2005.10.005}, issn = {1367-5931}, year = {2005}, date = {2005-12-01}, journal = {Current Opinion in Chemical Biology}, volume = {9}, number = {6}, pages = {674--679}, abstract = {The development of new and efficient drug delivery systems is of fundamental importance to improve the pharmacological profiles of many classes of therapeutic molecules. Many different types of drug delivery systems are currently available. Within the family of nanomaterials, carbon nanotubes (CNT) have emerged as a new alternative and efficient tool for transporting and translocating therapeutic molecules. CNT can be functionalised with bioactive peptides, proteins, nucleic acids and drugs, and used to deliver their cargos to cells and organs. Because functionalised CNT display low toxicity and are not immunogenic, such systems hold great potential in the field of nanobiotechnology and nanomedicine.}, keywords = {Biological, carbon, Drug Delivery Systems, Electron, HeLa Cells, Humans, I2CT, Microscopy, Models, Nanotubes, Nucleic Acids, Peptides, scanning, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{monneaux_selective_2005, title = {Selective modulation of CD4+ Ŧ cells from lupus patients by a promiscuous, protective peptide analog}, author = {Fanny Monneaux and Johan Hoebeke and Christelle Sordet and Céline Nonn and Jean-Paul Briand and Bernard Maillère and Jean Sibilia and Sylviane Muller}, doi = {10.4049/jimmunol.175.9.5839}, issn = {0022-1767}, year = {2005}, date = {2005-11-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {175}, number = {9}, pages = {5839--5847}, abstract = {A peptide encompassing residues 131-151 of the spliceosomal U1-70K protein and its analog phosphorylated at Ser140 were synthesized as potential candidates for the treatment of patients with lupus. Studies in the MRL/lpr and (NZB x NZW)F1 lupus models have demonstrated that these sequences contain a CD4+ T cell epitope but administration of the phosphorylated peptide only ameliorates the clinical manifestations of treated MRL/lpr mice. Binding assays with soluble HLA class II molecules and molecular modeling experiments indicate that both peptides behave as promiscuous epitopes and bind to a large panel of human DR molecules. In contrast to normal T cells and T cells from non-lupus autoimmune patients, we found that PBMCs from 40% of lupus patients selected randomly and CFSE-labeled CD4+ T cells proliferate in response to peptide 131-151. Remarkably, however, we observed that phosphorylation of Ser140 prevents CD4+ T cells proliferation but not secretion of regulatory cytokines, suggesting a striking immunomodulatory effect of phosphorylated analog on lupus CD4+ T cells that was unique to patients. The analog might act as an activator of regulatory T cells or as a partial agonist of TCR.}, keywords = {Amino Acid Sequence, CD4-Positive T-Lymphocytes, HLA-DR Antigens, Humans, I2CT, Interleukin-10, Lupus Erythematosus, Lymphocyte Activation, Molecular Sequence Data, Monneaux, Peptide Fragments, Phosphorylation, Ribonucleoprotein, Systemic, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{kocks_eater_2005, title = {Eater, a transmembrane protein mediating phagocytosis of bacterial pathogens in Drosophila}, author = {Christine Kocks and Ju Hyun Cho and Nadine Nehme and Johanna Ulvila and Alan M Pearson and Marie Meister and Charles Strom and Stephanie L Conto and Charles Hetru and Lynda M Stuart and Thilo Stehle and Jules A Hoffmann and Jean-Marc Reichhart and Dominique Ferrandon and Mika Rämet and Alan R B Ezekowitz}, doi = {10.1016/j.cell.2005.08.034}, issn = {0092-8674}, year = {2005}, date = {2005-10-01}, journal = {Cell}, volume = {123}, number = {2}, pages = {335--346}, abstract = {Phagocytosis is a complex, evolutionarily conserved process that plays a central role in host defense against infection. We have identified a predicted transmembrane protein, Eater, which is involved in phagocytosis in Drosophila. Transcriptional silencing of the eater gene in a macrophage cell line led to a significant reduction in the binding and internalization of bacteria. Moreover, the N terminus of the Eater protein mediated direct microbial binding which could be inhibited with scavenger receptor ligands, acetylated, and oxidized low-density lipoprotein. In vivo, eater expression was restricted to blood cells. Flies lacking the eater gene displayed normal responses in NF-kappaB-like Toll and IMD signaling pathways but showed impaired phagocytosis and decreased survival after bacterial infection. Our results suggest that Eater is a major phagocytic receptor for a broad range of bacterial pathogens in Drosophila and provide a powerful model to address the role of phagocytosis in vivo.}, keywords = {Amino Acid, Amino Acid Motifs, Animals, Bacterial Infections, Cell Surface, Embryo, Escherichia coli, ferrandon, Flow Cytometry, Frameshift Mutation, Genes, Histidine, hoffmann, In Situ Hybridization, Insect, Insect Proteins, M3i, Macrophages, Membrane Proteins, messenger, Nonmammalian, Open Reading Frames, Phagocytosis, Receptors, reichhart, RNA, RNA Interference, Sequence Homology, Serratia marcescens}, pubstate = {published}, tppubtype = {article} } @article{wu_targeted_2005, title = {Targeted delivery of amphotericin B to cells by using functionalized carbon nanotubes}, author = {Wei Wu and Sébastien Wieckowski and Giorgia Pastorin and Monica Benincasa and Cédric Klumpp and Jean-Paul Briand and Renato Gennaro and Maurizio Prato and Alberto Bianco}, doi = {10.1002/anie.200501613}, issn = {1433-7851}, year = {2005}, date = {2005-10-01}, journal = {Angewandte Chemie (International Ed. in English)}, volume = {44}, number = {39}, pages = {6358--6362}, keywords = {Amphotericin B, Antifungal Agents, carbon, Drug Carriers, Drug Delivery Systems, Fungi, Humans, I2CT, Jurkat Cells, Molecular Structure, Nanotubes, Particle Size, Solubility, Surface Properties, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bianco_efficient_2005, title = {Efficient solid-phase synthesis of fullero-peptides using Merrifield strategy}, author = {Alberto Bianco}, doi = {10.1039/b504659a}, issn = {1359-7345}, year = {2005}, date = {2005-07-01}, journal = {Chemical Communications (Cambridge, England)}, number = {25}, pages = {3174--3176}, abstract = {Boc-protected l-fulleropyrrolidino-glutamic acid was readily prepared and employed for the synthesis of fullerene-containing peptides using the solid-phase Boc chemistry developed by Merrifield.}, keywords = {Chromatography, Fullerenes, High Pressure Liquid, I2CT, Mass Spectrometry, Peptides, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{LF2005, title = {In Vivo Identification of Novel Regulators and Conserved Pathways of Phagocytosis in A. gambiae}, author = {L F Moita and R Wang and K Michel and Stéphanie A Blandin and T Zimmermann and Elena A Levashina and Fotis C Kafatos}, url = {http://www.ncbi.nlm.nih.gov/pubmed/16039580}, year = {2005}, date = {2005-07-01}, journal = {Immunity.}, volume = {23}, number = {1}, pages = {65-73}, abstract = {Anopheles gambiae uses effective immune responses, including phagocytosis, to fight microbial infection. We have developed a semiquantitative phagocytosis test and used it in conjunction with dsRNA gene silencing to test the in vivo roles of 71 candidate genes in phagocytosis of Escherichia coli and Staphylococcus aureus. Here, we show that inactivation of 26 genes changes the phagocytic activity by more than 45% and that two pathways similar to those that mediate apoptotic cell removal in Caenorhabditis elegans are used in A. gambiae for phagocytosis of microorganisms. Simultaneous inactivation of the identified regulators of phagocytosis and conserved components defining each signaling pathway permitted provisional assignment of the novel regulators to one or the other pathway. Pathway inactivation enhances at least three times the ability of E. coli and S. aureus to proliferate in the mosquito. Interestingly, mosquito survival is not compromised even if both pathways are perturbed simultaneously.}, keywords = {blandin, M3i, RNAi}, pubstate = {published}, tppubtype = {article} } @article{berthier-vergnes_tnf-alpha_2005, title = {TNF-alpha enhances phenotypic and functional maturation of human epidermal Langerhans cells and induces IL-12 p40 and IP-10/CXCL-10 production}, author = {Odile Berthier-Vergnes and Fabienne Bermond and Vincent Flacher and Catherine Massacrier and Daniel Schmitt and Josette Péguet-Navarro}, doi = {10.1016/j.febslet.2005.04.087}, issn = {0014-5793}, year = {2005}, date = {2005-07-01}, journal = {FEBS letters}, volume = {579}, number = {17}, pages = {3660--3668}, abstract = {Dendritic cells (DC) play a central role in immunity/tolerance decision, depending on their activation/maturation state. TNF-alpha is largely produced in the skin under inflammatory conditions. However, it still remains to be defined how TNF-alpha modulates the activation status of human LC, the most specialized DC controlling skin immunity. Here, we reported that fresh immature LC, highly purified from healthy human skin and exposed for two days to TNF-alpha under serum-free conditions, expressed up-regulated level of co-stimulatory molecules (CD40, CD54, CD86), maturation markers (CD83, DC-LAMP), CCR7 lymph node homing receptor, and down-regulated Langerin level, in a dose-dependent manner. This mature phenotype is closely associated with enhanced LC allostimulatory capacity. Furthermore, TNF-alpha significantly increased the number of viable LC and decreased their spontaneous apoptosis. More importantly, TNF-alpha induced LC to produce both IFN-gamma-inducible-protein IP-10/CXCL10, a Th1-attracting chemokine and IL-12 p40. Bioactive IL-12 p70 was never detected, even after additional CD40 stimulus. The results implicate LC as an effective target through which TNF-alpha may up- or down-regulate the inflammatory skin reactions.}, keywords = {Antigens, Apoptosis, C-Type, CD, Cell Differentiation, Cells, Chemokine CXCL10, chemokines, Cultured, CXC, Epidermal Cells, HLA-DR Antigens, Humans, Hypersensitivity, Interleukin-12, Interleukin-12 Subunit p40, Langerhans Cells, Lectins, Mannose-Binding Lectins, Phenotype, Protein Subunits, Surface, T-Lymphocytes, Team-Mueller, Tumor Necrosis Factor-alpha}, pubstate = {published}, tppubtype = {article} } @article{bianco_carbon_2005, title = {Carbon nanotubes: on the road to deliver}, author = {Alberto Bianco and Johan Hoebeke and Kostas Kostarelos and Maurizio Prato and Charalambos D Partidos}, doi = {10.2174/1567201054367959}, issn = {1567-2018}, year = {2005}, date = {2005-07-01}, journal = {Current Drug Delivery}, volume = {2}, number = {3}, pages = {253--259}, abstract = {Over the last few years, considerable advances have been made in the field of nanotechnology. The advent of carbon nanotube functionalization has paved the way for their potential application as a delivery system of diverse molecules such as peptides, proteins, plasmid DNA, and synthetic oligodeoxynucleotides. This opens new therapeutic and preventive opportunities to combat diseases. The scope of this review is to summarize our recent work in this rapidly growing field.}, keywords = {carbon, CpG Islands, DNA, Drug Carriers, I2CT, nanotechnology, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pastorin_functionalized_2005, title = {Functionalized Carbon Nanotubes: Towards the Delivery of Therapeutic Molecules}, author = {Giorgia Pastorin and Kostas Kostarelos and Maurizio Prato and Alberto Bianco}, url = {http://www.ingentaselect.com/rpsv/cgi-bin/cgi?ini=xref&body=linker&reqdoi=10.1166/jbn.2005.017}, doi = {10.1166/jbn.2005.017}, issn = {15507033, 00000000}, year = {2005}, date = {2005-06-01}, urldate = {2020-11-26}, journal = {Journal of Biomedical Nanotechnology}, volume = {1}, number = {2}, pages = {133--142}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{brosson_putative_2005, title = {The putative chitin deacetylase of Encephalitozoon cuniculi: a surface protein implicated in microsporidian spore-wall formation.}, author = {Damien Brosson and Lauriane Kuhn and Gérard Prensier and Christian P Vivarès and Catherine Texier}, doi = {10.1016/j.femsle.2005.04.031}, issn = {0378-1097 0378-1097}, year = {2005}, date = {2005-06-01}, journal = {FEMS microbiology letters}, volume = {247}, number = {1}, pages = {81--90}, abstract = {Microsporidia are fungal-like unicellular eukaryotes which develop as obligate intracellular parasites. They differentiate into resistant spores that are protected by a thick cell wall composed of glycoproteins and chitin. Despite an extensive description of the fibrillar structure of this wall, very little is known about its protein components and deposit mechanisms. In this study on the human pathogen Encephalitozoon cuniculi, we identify by mass spectrometry the target of polyclonal antibodies previously raised against a 33-kDa protein located at the outer face of the parasite plasma membrane. This 254-amino acid protein is encoded by the ECU11_0510 open reading frame and presents two isoforms of 33 and 55 kDa. Sequence analysis supports an assignment to the polysaccharide deacetylase family with a suspected chitin deacetylase activity (EcCDA). As demonstrated by TEM studies, EcCDA is present at the plasma membrane of the early stages of E. cuniculi life-cycle. At the sporoblast stage, the enzyme accumulates especially in paramural bodies which are convolutions of the plasma membrane opened to the wall. The identification of an EcCDA homologue in the insect parasite Antonospora locustae (ex Nosema locustae) suggests a widespread distribution of this enzyme among Microsporidia. This characterization of a new microsporidian surface protein creates new perspectives to understand spore wall formation and spore resistance.}, keywords = {PPSE}, pubstate = {published}, tppubtype = {article} } @article{kwan_dendritic_2005, title = {Dendritic cell precursors are permissive to dengue virus and human immunodeficiency virus infection}, author = {Wing-Hong Kwan and Anna-Marija Helt and Concepción Marañón and Jean-Baptiste Barbaroux and Anne Hosmalin and Eva Harris and Wolf H Fridman and Chris G F Mueller}, doi = {10.1128/JVI.79.12.7291-7299.2005}, issn = {0022-538X}, year = {2005}, date = {2005-06-01}, journal = {Journal of Virology}, volume = {79}, number = {12}, pages = {7291--7299}, abstract = {CD14(+) interstitial cells reside beneath the epidermis of skin and mucosal tissue and may therefore play an important role in viral infections and the shaping of an antiviral immune response. However, in contrast to dendritic cells (DC) or blood monocytes, these antigen-presenting cells (APC) have not been well studied. We have previously described long-lived CD14(+) cells generated from CD34(+) hematopoietic progenitors, which may represent model cells for interstitial CD14(+) APC. Here, we show that these cells carry DC-SIGN and differentiate into immature DC in the presence of granulocyte-macrophage colony-stimulating factor. We have compared the CD14(+) cells and the DC derived from these cells with respect to dengue virus and human immunodeficiency virus type 1 (HIV-1) infection. Both cell types are permissive to dengue virus infection, but the CD14(+) cells secrete the anti-inflammatory cytokine interleukin 10 and no tumor necrosis factor alpha. Regarding HIV, the CD14(+) cells are permissive to HIV-1, release higher p24 levels than the derived DC, and more efficiently activate HIV Pol-specific CD8(+) memory T cells. The CD14(+) DC precursors infected with either virus retain their DC differentiation potential. The results suggest that interstitial CD14(+) APC may contribute to HIV-1 and dengue virus infection and the shaping of an antiviral immune response.}, keywords = {ANTIGEN PRESENTING CELLS, Antigen-Presenting Cells, APC, BLOOD, CD8-Positive T-Lymphocytes, Cell Differentiation, Cells, COLONY-STIMULATING FACTOR, Cultured, Dendritic Cells, Dengue virus, Differentiation, Epidermis, Hematopoietic Stem Cells, HIV, HIV-1, Human, Humans, IMMATURE, immunodeficiency, infection, interleukin 10, Interleukin-10, Lipopolysaccharide Receptors, MEMORY T CELLS, monocyte, Monocytes, Necrosis, precursor, PROGENITORS, Skin, T CELLS, Team-Mueller, tumor, Tumor Necrosis Factor, viral Infection, virus}, pubstate = {published}, tppubtype = {article} } @article{fauny_drosophila_2005, title = {Drosophila Lipid Storage Droplet 2 gene (Lsd-2) is expressed and controls lipid storage in wing imaginal discs}, author = {Jean Daniel Fauny and Joël Silber and Alain Zider}, url = {http://www.ncbi.nlm.nih.gov/pubmed/15704138}, doi = {10.1002/dvdy.20277}, issn = {1058-8388}, year = {2005}, date = {2005-03-01}, urldate = {2011-10-24}, journal = {Developmental Dynamics: An Official Publication of the American Association of Anatomists}, volume = {232}, number = {3}, pages = {725--732}, abstract = {Lipid droplets are the major neutral lipid storage organelles in higher eukaryotes. The PAT domain proteins (Perilipin, ADRP [adipose differentiation related protein], and TIP47 [tail-interacting 47-kDa protein]) are associated with these structures. Perilipin and ADRP are involved in the regulation of lipid storage and metabolism in mammals. Two genes encoding PAT proteins, Drosophila Lipid Storage Droplet 2 Gene (Lsd-2) and Lsd-2, have been identified in Drosophila. Lsd-2 is expressed in fat bodies and in the female germ line and is involved in lipid storage in these tissues. We showed that Lsd-2 is expressed in third-instar wing imaginal discs in Drosophila, with higher levels in the wing pouch, which corresponds to the presumptive wing region of the wing disc. This specific expression pattern is correlated with a high level of neutral lipid accumulation. We also showed that neutral lipid deposition in the wing disc is severely reduced in an Lsd-2 mutant and is increased with Lsd-2 overexpression. Finally, we showed that overexpression of the vestigial (vg) pro-wing gene induces Lsd-2 expression, suggesting that Lsd-2 mediates a vg role during wing formation. Our results suggest that Lsd-2 function is not restricted to tissues directly involved in lipid storage and could play additional roles during development.}, keywords = {Animals, Biological, Drosophila, Drosophila Proteins, Embryo, Fat Body, Genes, I2CT, Imagerie, Insect, Larva, Lipid Metabolism, Metamorphosis, Mutation, Nonmammalian, Nuclear Proteins, Phosphoproteins, Wing}, pubstate = {published}, tppubtype = {article} } @article{irving_new_2005, title = {New insights into Drosophila larval haemocyte functions through genome-wide analysis}, author = {Phil Irving and Jean-Michel Ubeda and Daniel Doucet and Laurent Troxler and Marie Lagueux and Daniel Zachary and Jules A Hoffmann and Charles Hetru and Marie Meister}, doi = {10.1111/j.1462-5822.2004.00462.x}, issn = {1462-5814}, year = {2005}, date = {2005-03-01}, journal = {Cell. Microbiol.}, volume = {7}, number = {3}, pages = {335--350}, abstract = {Drosophila blood cells or haemocytes comprise three cell lineages, plasmatocytes, crystal cells and lamellocytes, involved in immune functions such as phagocytosis, melanisation and encapsulation. Transcriptional profiling of activities of distinct haemocyte populations and from naive or infected larvae, was performed to find genes contributing to haemocyte functions. Of the 13 000 genes represented on the microarray, over 2500 exhibited significantly enriched transcription in haemocytes. Among these were genes encoding integrins, peptidoglycan recognition proteins (PGRPs), scavenger receptors, lectins, cell adhesion molecules and serine proteases. One relevant outcome of this analysis was the gain of new insights into the lamellocyte encapsulation process. We showed that lamellocytes require betaPS integrin for encapsulation and that they transcribe one prophenoloxidase gene enabling them to produce the enzyme necessary for melanisation of the capsule. A second compelling observation was that following infection, the gene encoding the cytokine Spatzle was uniquely upregulated in haemocytes and not the fat body. This shows that Drosophila haemocytes produce a signal molecule ready to be activated through cleavage after pathogen recognition, informing distant tissues of infection.}, keywords = {Animals, bioinformatic, Catechol Oxidase, Cell Lineage, Enzyme Precursors, Escherichia coli, Fat Body, Gene Expression Profiling, Genome, Hemocytes, hoffmann, Integrin alpha Chains, Integrins, Larva, M3i, Micrococcus luteus}, pubstate = {published}, tppubtype = {article} } @article{singh_binding_2005, title = {Binding and condensation of plasmid DNA onto functionalized carbon nanotubes: toward the construction of nanotube-based gene delivery vectors}, author = {Ravi Singh and Davide Pantarotto and David McCarthy and Olivier Chaloin and Johan Hoebeke and Charalambos D Partidos and Jean-Paul Briand and Maurizio Prato and Alberto Bianco and Kostas Kostarelos}, doi = {10.1021/ja0441561}, issn = {0002-7863}, year = {2005}, date = {2005-03-01}, journal = {Journal of the American Chemical Society}, volume = {127}, number = {12}, pages = {4388--4396}, abstract = {Carbon nanotubes (CNTs) constitute a class of nanomaterials that possess characteristics suitable for a variety of possible applications. Their compatibility with aqueous environments has been made possible by the chemical functionalization of their surface, allowing for exploration of their interactions with biological components including mammalian cells. Functionalized CNTs (f-CNTs) are being intensively explored in advanced biotechnological applications ranging from molecular biosensors to cellular growth substrates. We have been exploring the potential of f-CNTs as delivery vehicles of biologically active molecules in view of possible biomedical applications, including vaccination and gene delivery. Recently we reported the capability of ammonium-functionalized single-walled CNTs to penetrate human and murine cells and facilitate the delivery of plasmid DNA leading to expression of marker genes. To optimize f-CNTs as gene delivery vehicles, it is essential to characterize their interactions with DNA. In the present report, we study the interactions of three types of f-CNTs, ammonium-functionalized single-walled and multiwalled carbon nanotubes (SWNT-NH3+; MWNT-NH3+), and lysine-functionalized single-walled carbon nanotubes (SWNT-Lys-NH3+), with plasmid DNA. Nanotube-DNA complexes were analyzed by scanning electron microscopy, surface plasmon resonance, PicoGreen dye exclusion, and agarose gel shift assay. The results indicate that all three types of cationic carbon nanotubes are able to condense DNA to varying degrees, indicating that both nanotube surface area and charge density are critical parameters that determine the interaction and electrostatic complex formation between f-CNTs with DNA. All three different f-CNT types in this study exhibited upregulation of marker gene expression over naked DNA using a mammalian (human) cell line. Differences in the levels of gene expression were correlated with the structural and biophysical data obtained for the f-CNT:DNA complexes to suggest that large surface area leading to very efficient DNA condensation is not necessary for effective gene transfer. However, it will require further investigation to determine whether the degree of binding and tight association between DNA and nanotubes is a desirable trait to increase gene expression efficiency in vitro or in vivo. This study constitutes the first thorough investigation into the physicochemical interactions between cationic functionalized carbon nanotubes and DNA toward construction of carbon nanotube-based gene transfer vector systems.}, keywords = {carbon, Cations, DNA, Electron, Gene Transfer Techniques, Genetic Vectors, I2CT, Lysine, Microscopy, Nanotubes, Plasmids, Quaternary Ammonium Compounds, scanning, Surface Plasmon Resonance, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{royet_sensing_2005, title = {Sensing and signaling during infection in Drosophila}, author = {Julien Royet and Jean-Marc Reichhart and Jules A Hoffmann}, doi = {10.1016/j.coi.2004.12.002}, issn = {0952-7915}, year = {2005}, date = {2005-02-01}, journal = {Curr. Opin. Immunol.}, volume = {17}, number = {1}, pages = {11--17}, abstract = {Most of the progress in dissecting the Drosophila antimicrobial response over the past decade has centered around intracellular signaling pathways in immune response tissues and expression of genes encoding antimicrobial peptide genes. The past few years, however, have witnessed significant advances in our understanding of the recognition of microbial invaders and subsequent activation of signaling cascades. In particular, the roles of peptidoglycan recognition proteins, which have known homologues in mammals, have been recognized and examined at the structural and functional levels.}, keywords = {Animals, Antimicrobial Cationic Peptides, Bacterial Infections, Gene Expression Regulation, Genome, hoffmann, Immunity, Innate, M3i, reichhart, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reverse transcriptase and integrase of the Saccharomyces cerevisiae Ty1 element}, author = {F X Wilhelm and M Wilhelm and A Gabriel}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16093680}, isbn = {16093680}, year = {2005}, date = {2005-01-01}, journal = {Cytogenet Genome Res}, volume = {110}, number = {1-4}, pages = {269-87}, abstract = {Integrase (IN) and reverse transcriptase (RT) play a central role in transposition of retroelements. The mechanism of integration by IN and the steps of the replication process mediated by RT are briefly described here. Recently, active recombinant forms of Ty1 IN and RT have been obtained. This has allowed a more detailed understanding of their biochemical and structural properties and has made possible combined in vitro and in vivo analyses of their functions. A focus of this review is to discuss some of the results obtained thus far with these two recombinant proteins and to propose future directions.}, note = {1424-859x Journal Article}, keywords = {MARQUET}, pubstate = {published}, tppubtype = {article} } @article{, title = {Leucyl-tRNA synthetase from the ancestral bacterium Aquifex aeolicus contains relics of synthetase evolution}, author = {M W Zhao and B Zhu and R Hao and M G Xu and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15775966}, isbn = {15775966}, year = {2005}, date = {2005-01-01}, journal = {EMBO J}, volume = {24}, number = {7}, pages = {1430-1439}, abstract = {The editing reactions catalyzed by aminoacyl-tRNA synthetases are critical for the faithful protein synthesis by correcting misactivated amino acids and misaminoacylated tRNAs. We report that the isolated editing domain of leucyl-tRNA synthetase from the deep-rooted bacterium Aquifex aeolicus (alphabeta-LeuRS) catalyzes the hydrolytic editing of both mischarged tRNA(Leu) and minihelix(Leu). Within the domain, we have identified a crucial 20-amino-acid peptide that confers editing capacity when transplanted into the inactive Escherichia coli LeuRS editing domain. Likewise, fusion of the beta-subunit of alphabeta-LeuRS to the E. coli editing domain activates its editing function. These results suggest that alphabeta-LeuRS still carries the basic features from a primitive synthetase molecule. It has a remarkable capacity to transfer autonomous active modules, which is consistent with the idea that modern synthetases arose after exchange of small idiosyncratic domains. It also has a unique alphabeta-heterodimeric structure with separated catalytic and tRNA-binding sites. Such an organization supports the tRNA/synthetase coevolution theory that predicts sequential addition of tRNA and synthetase domains.}, note = {0261-4189 (Print) Journal Article}, keywords = {Amino Acid Sequence Bacteria/*genetics Comparative Study Escherichia coli/genetics *Evolution, ERIANI, Molecular Leucine-tRNA Ligase/*genetics/*metabolism Models, Molecular Molecular Sequence Data Protein Biosynthesis/*genetics Protein Structure, Non-U.S. Gov't Sequence Alignment, Tertiary/genetics Research Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of integrase in reverse transcription of the Saccharomyces cerevisiae retrotransposon Ty1}, author = {M Wilhelm and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15947198}, isbn = {15947198}, year = {2005}, date = {2005-01-01}, journal = {Eukaryot Cell}, volume = {4}, number = {6}, pages = {1057-1065}, abstract = {Reverse transcriptase (RT) with its associated RNase H (RH) domain and integrase (IN) are key enzymes encoded by retroviruses and retrotransposons. Several studies have implied a functional role of the interaction between IN and RT during the replication of retroviral and retrotransposon genomes. In this study, IN deletion mutants were used to investigate the role of IN on the RT activity of the yeast Saccharomyces cerevisiae retrotransposon Ty1. We have identified two domains of Ty1 integrase which have effects on RT activity in vivo. The deletion of a domain spanning amino acid residues 233 to 520 of IN increases the exogenous specific activity of RT up to 20-fold, whereas the removal of a region rich in acidic amino acid residues between residues 521 and 607 decreases its activity. The last result complements our observation that an active recombinant RT protein can be obtained if a small acidic tail mimicking the acidic domain of IN is fused to the RT-RH domain. We suggest that interaction between these acidic amino acid residues of IN and a basic region of RT could be critical for the correct folding of RT and for the formation of an active conformation of the enzyme.}, note = {1535-9778 Journal Article}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reverse transcriptase and integrase of the Saccharomyces cerevisiae Ty1 element}, author = {F X Wilhelm and M Wilhelm and A Gabriel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16093680}, isbn = {16093680}, year = {2005}, date = {2005-01-01}, journal = {Cytogenet Genome Res}, volume = {110}, number = {1-4}, pages = {269-287}, abstract = {Integrase (IN) and reverse transcriptase (RT) play a central role in transposition of retroelements. The mechanism of integration by IN and the steps of the replication process mediated by RT are briefly described here. Recently, active recombinant forms of Ty1 IN and RT have been obtained. This has allowed a more detailed understanding of their biochemical and structural properties and has made possible combined in vitro and in vivo analyses of their functions. A focus of this review is to discuss some of the results obtained thus far with these two recombinant proteins and to propose future directions.}, note = {1424-859x Journal Article}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Crystallographic Studies of Complexes between the Ribosomal Decoding Site and Aminoglycosides Antibiotics}, author = {E Westhof and Q Vicens}, editor = {N Niimura and H Mizuno and J Helliwell and E Westhof}, url = {http://pure.au.dk/portal/en/publications/crystallographic-studies-of-complexes-between-the-ribosomal-decoding-site-and-aminoglycoside-antibiotics%284a6ee78e-3d0a-43ba-9f46-4bdebbe3eb51%29.html}, year = {2005}, date = {2005-01-01}, booktitle = {Hydrogen- and Hydration-Sensitive Structural Biology}, pages = {125-133}, publisher = {KubaPro Co. Ltd.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Nucleic acids RNA: from architecture to recognition and catalysis}, author = {E Westhof and D J Patel}, url = {http://www-ibmc.u-strasbg.fr/upr9002/westhof/PDF/r2005_EWesthof_COSB.pdf}, isbn = {15922589}, year = {2005}, date = {2005-01-01}, journal = {Curr Opin Struct Biol}, volume = {15}, number = {3}, pages = {309-312}, note = {0959-440x Editorial}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {From RNAi to epigenomes: how RNA rules the world}, author = {E Westhof and W Filipowicz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15651038}, isbn = {15651038}, year = {2005}, date = {2005-01-01}, journal = {Chembiochem}, volume = {6}, number = {2}, pages = {441-443}, note = {1439-4227 (Print) Journal Article}, keywords = {Animals Epigenesis, Genetic *Genome MicroRNAs/genetics/metabolism *Rna *RNA Interference RNA, Plant, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Muttaiya Sundaralingam (1931-2004): A Pioneering Stereochemist and Crystallographer of Nucleic Acids}, author = {E Westhof}, url = {http://www.sciencedirect.com/science/article/pii/S0969212605000870}, doi = {10.1016/j.str.2005.02.009}, year = {2005}, date = {2005-01-01}, journal = {Structure}, volume = {13}, number = {3}, pages = {343–345}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular recognition between the ribosomal decoding site and natural or non-natural aminoglycosides}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17150632}, isbn = {17150632}, year = {2005}, date = {2005-01-01}, journal = {Nucleic Acids Symp Ser}, number = {49}, pages = {59-60}, abstract = {Crystal structures of complexes between ribosomal particles and antibiotics have pinned down very precisely the discrete binding sites of several classes of antibiotics inhibiting protein synthesis. The crystal structures of complexes between various antibiotics and ribosomal particles show definitively that ribosomal RNAs, rather than ribosomal proteins, are overwhelmingly targeted. The comparative analysis of various aminoglycosides bound to the same site has been used to decipher the contribution of each functional group to the RNA/aminoglycoside complex and to attempt to rationalize various biochemical and microbiological data as well as some resistance and toxicity mechanisms at the molecular level. Tight packing of atoms in direct van der Waals contact is central and a prerequisite to specific recognition. Water molecules participate in the assembly by linking hydrophilic groups belonging to both components. The hydration shells around nucleic acid base pairs tend to be conserved and maintained whatever the environment. Two regions are crucial for specificity and resistance: A1408 and the non-Watson-Crick pair U1406-U1495.}, note = {1746-8272 (Electronic) Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Lessons from crystals grown in the Advanced Protein Crystallisation Facility for conventional crystallisation applied to structural biology}, author = {A Vergara and B Lorber and C Sauter and R Giege and A Zagari}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16150532}, isbn = {16150532}, year = {2005}, date = {2005-01-01}, journal = {Biophys Chem}, volume = {118}, number = {2-3}, pages = {102-112}, abstract = {The crystallographic quality of protein crystals that were grown in microgravity has been compared to that of crystals that were grown in parallel on earth gravity under otherwise identical conditions. A goal of this comparison was to assess if a more accurate 3D-structure can be derived from crystallographic analysis of the former crystals. Therefore, the properties of crystals prepared with the Advanced Protein Crystallisation Facility (APCF) on earth and in orbit during the last decade were evaluated. A statistical analysis reveals that about half of the crystals produced under microgravity had a superior X-ray diffraction limit with respect of terrestrial controls. Eleven protein structures could be determined at previously unachieved resolutions using crystals obtained in the APCF. Microgravity induced features of the most relevant structures are reported. A second goal of this study was to identify the cause of the crystal quality enhancement useful for structure determination. No correlations between the effect of microgravity and other system-dependent parameters, such as isoelectric point or crystal solvent content, were found except the reduced convection during the crystallisation process. Thus, crystal growth under diffusive regime appears to be the key parameter explaining the beneficial effect of microgravity on crystal quality. The mimicry of these effects on earth in gels or in capillary tubes is discussed and the practical consequences for structural biology highlighted.}, note = {0301-4622 (Print) Journal Article Review}, keywords = {FRUGIER, GIEGE Animals *Biological Sciences Crystallization Equipment Design Humans Protein Conformation Proteins/*chemistry Research Support, Non-U.S. Gov't X-Ray Diffraction, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {MAO: a Multiple Alignment Ontology for nucleic acid and protein sequences}, author = {J D Thompson and S R Holbrook and K Katoh and P Koehl and D Moras and E Westhof and O Poch}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16043635}, isbn = {16043635}, year = {2005}, date = {2005-01-01}, journal = {Nucleic Acids Res}, volume = {33}, number = {13}, pages = {4164-4171}, abstract = {The application of high-throughput techniques such as genomics, proteomics or transcriptomics means that vast amounts of heterogeneous data are now available in the public databases. Bioinformatics is responding to the challenge with new integrated management systems for data collection, validation and analysis. Multiple alignments of genomic and protein sequences provide an ideal environment for the integration of this mass of information. In the context of the sequence family, structural and functional data can be evaluated and propagated from known to unknown sequences. However, effective integration is being hindered by syntactic and semantic differences between the different data resources and the alignment techniques employed. One solution to this problem is the development of an ontology that systematically defines the terms used in a specific domain. Ontologies are used to share data from different resources, to automatically analyse information and to represent domain knowledge for non-experts. Here, we present MAO, a new ontology for multiple alignments of nucleic and protein sequences. MAO is designed to improve interoperation and data sharing between different alignment protocols for the construction of a high quality, reliable multiple alignment in order to facilitate knowledge extraction and the presentation of the most pertinent information to the biologist.}, note = {1362-4962 Journal Article}, keywords = {Controlled, Databases, DNA/*methods Sequence Analysis, Genetic Humans Interleukin-1/genetics Internet Research Support, Non-U.S. Gov't Sequence Alignment/*methods Sequence Analysis, Protein/*methods Sequence Analysis, RNA/*methods *Software Systems Integration Vocabulary, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Evidence for the existence in mRNAs of a hairpin element responsible for ribosome dependent pyrrolysine insertion into proteins}, author = {A Théobald-Dietrich and R Giege and J Rudinger-Thirion}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16164991}, isbn = {16164991}, year = {2005}, date = {2005-01-01}, journal = {Biochimie}, volume = {87}, number = {9-10}, pages = {813-817}, abstract = {In the methanogenic archae Methanosarcina barkeri, insertion of pyrrolysine, the 22nd amino acid, results from the decoding of an amber UAG codon in the mRNA of monomethylamine methyltransferases (MtmB). Sequence comparisons combined with structural enzymatic and chemical probing on M. barkeri MtmB1 mRNA demonstrate the presence of a hairpin motif located immediately after the redefined UAG codon. This structure of 86 nucleotides differs slightly from a proposal given in the literature and comprises four successive stems separated by three internal loops and closed by a large apical loop. Sequence alignments of MtmB mRNAs of different Methanosarcinacae reveal a conservation of the motif in both sequence and folding levels. The functional role of this motif as a signal leading to pyrrolysine insertion is discussed.}, note = {0300-9084 Journal article}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Mitochondrial aminoacyl-tRNA synthetases}, author = {M Sissler and J Putz and F Fasiolo and C Florentz}, editor = {M Ibba and C Francklyn and S Cusak}, url = {http://www.ncbi.nlm.nih.gov/books/NBK6033}, year = {2005}, date = {2005-01-01}, booktitle = {The Aminoacyl-tRNA Synthetases}, publisher = {Landes Bioscience}, abstract = {Mitochondria and chloroplasts have their own genomes that encode a small number of proteins whose synthesis depends on translation machineries of multiple origin. Whereas tRNAs, rRNAs and some ribosomal proteins are often encoded by the organellar genome, all other factors and in particular aminoacyl-tRNA synthetases (aaRSs) are nuclear encoded, synthesized in the cytosol and imported. Thus, two to three sets of aaRSs coexist in eukaryotic cells, namely cytosolic, mitochondrial and chloroplastic versions. Here, the diversity in the structural and functional properties of organellar aaRSs is illustrated by mammalian mitochondrial aaRSs (size, oligomeric structure, efficiency of aminoacylation, cross reactions, identity sets). Additionally, means by which nuclear genes encode cytosolic, mitochondrial and chloroplastic aaRSs are reviewed on the basis of database exploration on fully sequenced (although not completely annotated) genomes of Homo sapiens, Saccharomyces cerevisiae, Caenorabditis elegans, Drosophila melanogaster and Arabidopsis thaliana.}, keywords = {FLORENTZ, FLORENTZ FASIOLO, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {An intricate RNA structure with two tRNA-derived motifs directs complex formation between yeast aspartyl-tRNA synthetase and its mRNA}, author = {M Ryckelynck and B Masquida and R Giege and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16257416}, isbn = {16257416}, year = {2005}, date = {2005-01-01}, journal = {J Mol Biol}, volume = {354}, number = {3}, pages = {614-629}, abstract = {Accurate translation of genetic information necessitates the tuned expression of a large group of genes. Amongst them, controlled expression of the enzymes catalyzing the aminoacylation of tRNAs, the aminoacyl-tRNA synthetases, is essential to insure translational fidelity. In the yeast Saccharomyces cerevisiae, expression of aspartyl-tRNA synthetase (AspRS) is regulated in a process necessitating recognition of the 5' extremity of AspRS messenger RNA (mRNA(AspRS)) by its translation product and adaptation to the cellular tRNA(Asp) concentration. Here, we have established the folding of the approximately 300 nucleotides long 5' end of mRNA(AspRS) and identified the structural signals involved in the regulation process. We show that the regulatory region in mRNA(AspRS) folds in two independent and symmetrically structured domains spaced by two single-stranded connectors. Domain I displays a tRNA(Asp) anticodon-like stem-loop structure with mimics of the aspartate identity determinants, that is restricted in domain II to a short double-stranded helix. The overall mRNA structure, based on enzymatic and chemical probing, supports a three-dimensional model where each monomer of yeast AspRS binds one individual domain and recognizes the mRNA structure as it recognizes its cognate tRNA(Asp). Sequence comparison of yeast genomes shows that the features within the mRNA recognized by AspRS are conserved in different Saccharomyces species. In the recognition process, the N-terminal extension of each AspRS subunit plays a crucial role in anchoring the tRNA-like motifs of the mRNA on the synthetase.}, note = {0022-2836 (Print) Journal Article}, keywords = {Aspartate-tRNA Ligase/genetics/*metabolism Base Sequence DNA Footprinting Gene Deletion Models, FRUGIER, Messenger/*chemistry/*metabolism RNA, Molecular Molecular Sequence Data Nucleic Acid Conformation Protein Structure, Non-U.S. Gov't Saccharomyces cerevisiae/*enzymology/*genetics Sequence Alignment Sequence Homology, Nucleic Acid Solubility, Tertiary RNA, Transfer/*chemistry/*metabolism Research Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNAs and tRNA mimics as cornerstones of aminoacyl-tRNA synthetase regulations}, author = {M Ryckelynck and R Giege and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15925436}, isbn = {15925436}, year = {2005}, date = {2005-01-01}, journal = {Biochimie}, volume = {87}, number = {9-10}, pages = {835-845}, abstract = {Structural plasticity of transfer RNA (tRNA) molecules is essential for interactions with their biological partners in aminoacylation reactions and during ribosome-dependent protein synthesis. This holds true when tRNAs are recruited for other functions than translation. Here we review regulation pathways where tRNAs and tRNA mimics play a pivotal role. We further discuss the importance of the identity signals used in aminoacylation that are also required to specify regulatory mechanisms. Such mechanisms are diverse and intervene in transcription, splicing and translation. Altogether, the review highlights the many manners architectural features of tRNA were selected by evolution to control biological key processes.}, note = {0300-9084 Journal article}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Functional Hammerhead Ribozymes Naturally Encoded in the Genome of Arabidopsis thaliana}, author = {R Przybilski and S Graf and A Lescoute and W Nellen and E Westhof and G Steger and C Hammann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15937227}, isbn = {15937227}, year = {2005}, date = {2005-01-01}, journal = {Plant Cell}, volume = {17}, number = {7}, pages = {1877-1885}, abstract = {The hammerhead ribozyme (HHRz) is an autocatalytic RNA motif found in subviral plant pathogens and transcripts of repetitive DNA sequences in animals. Here, we report the discovery and characterization of unique HHRzs encoded in a plant genome. Two novel sequences were identified on chromosome IV of Arabidopsis thaliana in a database search, which took into account recently defined structural requirements. The HHRzs are expressed in several tissues and coexist in vivo as both cleaved and noncleaved species. In vitro, both sequences cleave efficiently at physiological Mg(2+) concentrations, indicative of functional loop-loop interactions. Kinetic analysis of loop nucleotide variants was used to determine a three-dimensional model of these tertiary interactions. Based on these results, on the lack of infectivity of hammerhead-carrying viroids in Arabidopsis, and on extensive sequence comparisons, we propose that the ribozyme sequences did not invade this plant by horizontal transfer but have evolved independently to perform a specific, yet unidentified, biological function.}, note = {1040-4651 Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutagenesis of 16S rRNA C1409-G1491 base-pair differentiates between 6'OH and 6'NH3+ aminoglycosides}, author = {P Pfister and S Hobbie and C Brull and N Corti and A Vasella and E Westhof and E C Bottger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15670597}, isbn = {15670597}, year = {2005}, date = {2005-01-01}, journal = {J Mol Biol}, volume = {346}, number = {2}, pages = {467-475}, abstract = {Using a single rRNA allelic Gram-positive model system, we systematically mutagenized 16S rRNA positions 1409 and 1491 to probe the functional relevance of structural interactions between aminoglycoside antibiotics and the A-site rRNA that were suggested by X-ray crystallography. At the structural level, the interaction of the 2-deoxystreptamine aminoglycosides with the rRNA base-pair C1409-G1491 has been suggested to involve the following features: (i) ring I of the disubstituted 2-deoxystreptamines stacks upon G1491 and H-bonds to the Watson-Crick edge of A1408; (ii) ring III of the 4,5-disubstituted aminoglycosides shows hydrogen bonding to G1491. However, we found that mutants with altered 16S rRNA bases 1409 and 1491 discriminated poorly between 4,5-disubstituted and 4,6-disubstituted 2-deoxystreptamines, but differentially affected aminoglycosides with a hydroxyl group versus an ammonium group at position 6' of ring I, e.g. G1491U conferred high-level drug resistance to paromomycin and geneticin, but not to neomycin, tobramycin or gentamicin.}, note = {0022-2836 Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @book{, title = {Hydrogen- and Hydration-Sensitive Structural Biology}, editor = {N Niimura and H Mizuno and J Helliwell and E Westhof}, year = {2005}, date = {2005-01-01}, publisher = {Kubapro Co, Ltd, Tokyo}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {book} } @article{, title = {An mRNA is capped by a 2', 5' lariat catalyzed by a group I-like ribozyme}, author = {H Nielsen and E Westhof and S Johansen}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16141078}, isbn = {16141078}, year = {2005}, date = {2005-01-01}, journal = {Science}, volume = {309}, number = {5740}, pages = {1584-1587}, abstract = {Twin-ribozyme introns are formed by two ribozymes belonging to the group I family and occur in some ribosomal RNA transcripts. The group I-like ribozyme, GIR1, liberates the 5' end of a homing endonuclease messenger RNA in the slime mold Didymium iridis. We demonstrate that this cleavage occurs by a transesterification reaction with the joining of the first and the third nucleotide of the messenger by a 2',5'-phosphodiester linkage. Thus, a group I-like ribozyme catalyzes an RNA branching reaction similar to the first step of splicing in group II introns and spliceosomal introns. The resulting short lariat, by forming a protective 5' cap, might have been useful in a primitive RNA world.}, note = {1095-9203 (Electronic) Journal Article}, keywords = {ase Sequence Catalysis Endonucleases/biosynthesis/*genetics Esterification *Introns Molecular Sequence Data RNA Caps/*chemistry *RNA Splicing RNA, Catalytic/chemistry/*metabolism Research Support, Non-U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Conformational transition of initiation factor 2 from the GTP- to GDP-bound state visualized on the ribosome}, author = {A G Myasnikov and S Marzi and A Simonetti and A M Giuliodori and C O Gualerzi and G Yusupova and M Yusupov and B P Klaholz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16284619}, isbn = {16284619}, year = {2005}, date = {2005-01-01}, journal = {Nat Struct Mol Biol}, volume = {12}, number = {12}, pages = {1145-1149}, abstract = {Initiation of protein synthesis is a universally conserved event that requires initiation factors IF1, IF2 and IF3 in prokaryotes. IF2 is a GTPase essential for binding initiator transfer RNA to the 30S ribosomal subunit and recruiting the 50S subunit into the 70S initiation complex. We present two cryo-EM structures of the assembled 70S initiation complex comprising mRNA, fMet-tRNA(fMet) and IF2 with either a non-hydrolyzable GTP analog or GDP. Transition from the GTP-bound to the GDP-bound state involves substantial conformational changes of IF2 and of the entire ribosome. In the GTP analog-bound state, IF2 interacts mostly with the 30S subunit and extends to the initiator tRNA in the peptidyl (P) site, whereas in the GDP-bound state IF2 steps back and adopts a 'ready-to-leave' conformation. Our data also provide insights into the molecular mechanism guiding release of IF1 and IF3.}, note = {1545-9993 Journal article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effects of macromolecular impurities and of crystallization method on the quality of eubacterial aspartyl-tRNA synthetase crystals}, author = {A Moreno and A Théobald-Dietrich and B Lorber and C Sauter and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15930641}, isbn = {15930641}, year = {2005}, date = {2005-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {61}, number = {Pt 6}, pages = {789-792}, abstract = {Although macromolecular purity is thought to be essential for the growth of flawless protein crystals, only a few studies have investigated how contaminants alter the crystallization process and crystal quality. Likewise, the outcome of a crystallization process may vary with the crystallization method. Here, it is reported how these two variables affect the crystallogenesis of aspartyl-tRNA synthetase from the eubacterium Thermus thermophilus. This homodimeric enzyme (Mr=130,000) possesses a multi-domain architecture and crystallizes either in a monoclinic or an orthorhombic habit. Minute amounts of protein impurities alter to a different extent the growth of each crystal form. The best synthetase crystals are only obtained when the crystallizing solution is either enclosed in capillaries or immobilized in agarose gel. In these two environments convection is reduced with regard to that existing in an unconstrained solution.}, note = {0907-4449 Journal Article}, keywords = {FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Modeling the architecture of structural RNAs within a modular and hierarchical framework}, author = {B Masquida and E Westhof}, editor = {R K Hartmann and A Bindereif and A Schon and E Westhof}, url = {http://onlinelibrary.wiley.com/book/10.1002/9783527619504}, year = {2005}, date = {2005-01-01}, booktitle = {Handbook of RNA Biochemistry}, pages = {536-545}, publisher = {WIiley-Vch Verlag}, keywords = {Modeling Architecture Framework Physicochemical forces Amino acids, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Transfer RNA modifications and DNA editing in HIV-1 reverse transcription}, author = {R Marquet and F Dardel}, editor = {H Grosjean}, url = {http://www.springerlink.com/content/ft91mpx8xv7dq265}, doi = {10.1007/b106366}, year = {2005}, date = {2005-01-01}, booktitle = {Fine-tuning of RNA functions by Modification and Editing}, volume = {12}, pages = {401-429}, publisher = {Springer-Verlag}, abstract = {Reverse transcription is a central step in HIV-1 replication that represents a typical case of interplay between viral and cellular factors. HIV-1 diverts a cellular tRNA, tRNALys3, to prime reverse transcription. The post-transcriptional modifications of tRNALys3 are crucial for completion of reverse transcription. In some HIV-1 isolates, they are required for efficient initiation of (–) strand DNA synthesis, and in all strains, methylation of A58 is required to allow productive strand transfer during (+) strand DNA synthesis. On the other hand, some human cell types have evolved an innate antiretroviral mechanism by promoting extensive deamination of the (–) strand DNA during reverse transcription. In the absence of viral defence, this hyper-editing induces DNA degradation and lethal mutagenesis of the viral DNA. However, Vif, one of the HIV-1 "accessory" proteins, is able to inhibit DNA deamination by preventing incorporation of the editing enzymes APOBEC3G and APOBEC3F into the viral particles.}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Perspectives for pure and applied protein crystallogenesis studies}, author = {B Lorber}, url = {http://pubs.acs.org/doi/abs/10.1021/cg030065o}, year = {2005}, date = {2005-01-01}, journal = {Crystal Growth & Design}, volume = {5}, number = {1}, pages = {17-19}, abstract = {The unpredictability of protein crystallization is essentially due to our ignorance of the correlations existing between architectural features of these macromolecules and physical chemical properties of the crystallization medium in which their crystals nucleate and grow. Some ideas are suggested to guide crystal growth and bioinformatics specialists in their quest of relationships that might help to rationalize the general approach of protein crystal growth.}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Riboswitch structures: purine ligands replace tertiary contacts}, author = {A Lescoute and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15664510}, isbn = {15664510}, year = {2005}, date = {2005-01-01}, journal = {Chem Biol}, volume = {12}, number = {1}, pages = {10-13}, note = {1074-5521 Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Recurrent structural RNA motifs, Isostericity Matrices and sequence alignments}, author = {A Lescoute and N B Leontis and C Massire and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15860776}, isbn = {15860776}, year = {2005}, date = {2005-01-01}, journal = {Nucleic Acids Res}, volume = {33}, number = {8}, pages = {2395-2409}, abstract = {The occurrences of two recurrent motifs in ribosomal RNA sequences, the Kink-turn and the C-loop, are examined in crystal structures and systematically compared with sequence alignments of rRNAs from the three kingdoms of life in order to identify the range of the structural and sequence variations. Isostericity Matrices are used to analyze structurally the sequence variations of the characteristic non-Watson-Crick base pairs for each motif. We show that Isostericity Matrices for non-Watson-Crick base pairs provide important tools for deriving the sequence signatures of recurrent motifs, for scoring and refining sequence alignments, and for determining whether motifs are conserved throughout evolution. The systematic use of Isostericity Matrices identifies the positions of the insertion or deletion of one or more nucleotides relative to the structurally characterized examples of motifs and, most importantly, specifies whether these changes result in new motifs. Thus, comparative analysis coupled with Isostericity Matrices allows one to produce and refine structural sequence alignments. The analysis, based on both sequence and structure, permits therefore the evaluation of the conservation of motifs across phylogeny and the derivation of rules of equivalence between structural motifs. The conservations observed in Isostericity Matrices form a predictive basis for identifying motifs in sequences.}, note = {1362-4962 Journal Article}, keywords = {Base Pairing Models, Molecular Nucleic Acid Conformation RNA, Non-U.S. Gov't Research Support, Nucleic Acid, P.H.S. *Sequence Alignment Sequence Analysis, Ribosomal/*chemistry Research Support, RNA/*methods Sequence Homology, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{kostarelos_carbon_2005, title = {Carbon nanotube-mediated delivery of peptides and genes to cells: translating nanobiotechnology to therapeutics}, author = {K Kostarelos and L Lacerda and C D Partidos and M Prato and A Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S1773224705500054}, doi = {10.1016/S1773-2247(05)50005-4}, issn = {1773-2247}, year = {2005}, date = {2005-01-01}, urldate = {2020-03-31}, journal = {Journal of Drug Delivery Science and Technology}, volume = {15}, number = {1}, pages = {41--47}, abstract = {During the last few years, there has been a tremendous amount of optimism and expectation about nanotechnology and its impact on various fields including medicine and pharmaceutical development. One of the most promising materials being developed during the nanotechnological renaissance we are currently experiencing is the carbon nanotube. Before any biology-related application can even be envisaged, the aqueous solubility of carbon nanotubes has to be resolved. Recently, a variety of methodologies have been proposed which lead to biologically compatible carbon nanotubes. Covalent functionalization of their surface is one methodology, allowing the first attempts towards applications in the field of nanomedicine. The possibility of incorporating functionalized carbon nanotubes into cells and the biological milieu offers numerous advantages for potential applications in biology and pharmacology. One of the most promising is their utilization as a new carrier system for the delivery of therapeutic molecules. In the present article, the first attempts to transform carbon nanotubes from biologically incompatible nanomaterials to biologically relevant components of advanced therapeutics and the ensuing novel structures obtained in our laboratories are presented.}, keywords = {Carbon nanotubes, gene delivery, gene therapy, I2CT, Nanomedicine, Peptide delivery, Team-Bianco, Vaccination}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallographic studies of Homo sapiens ribosomal decoding A site complexed with aminoglycosides}, author = {J Kondo and B Francois and A Urzhumtsev and E Westhof}, url = {http://nass.oxfordjournals.org/content/49/1/253.abstract}, year = {2005}, date = {2005-01-01}, journal = {Nucleic Acids Symp Ser}, volume = {49}, number = {1}, pages = {253-254}, abstract = {Aminoglycosides are highly effective antibacterial drugs that decrease translation accuracy by binding to the aminoacyl-tRNA decoding site (A site) of 16S ribosomal RNA. On the other hand, they are highly toxic to mammals through kidney and ear-associated illnesses by binding to ribosomal A sites. To understand the mechanism of toxicity of aminoglycosides to mammals at atomic level, crystallographic studies have been carried out with a number of Homo sapiens mitochondrial and cytoplasmic A sites complexed with aminoglycosides. Several X-ray diffraction data sets were successfully collected. Initial phases of mitochondrial A site with tobramycin and cytoplasmic A site with paromomycin were derived by the molecular replacement method. Refinements of atomic parameters are now under progress.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding of manganese(II) to a tertiary stabilized hammerhead ribozyme as studied by electron paramagnetic resonance spectroscopy}, author = {N Kisseleva and A Khvorova and E Westhof and O Schiemann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15611296}, isbn = {15611296}, year = {2005}, date = {2005-01-01}, journal = {RNA}, volume = {11}, number = {1}, pages = {1-6}, abstract = {Electron paramagnetic resonance (EPR) spectroscopy is used to study the binding of MnII ions to a tertiary stabilized hammer-head ribozyme (tsHHRz) and to compare it with the binding to the minimal hammerhead ribozyme (mHHRz). Continuous wave EPR measurements show that the tsHHRz possesses a single high-affinity MnII binding site with a KD of < or =10 nM at an NaCl concentration of 0.1 M. This dissociation constant is at least two orders of magnitude smaller than the KD determined previously for the single high-affinity MnII site in the mHHRz. In addition, whereas the high-affinity MnII is displaced from the mHHRz upon binding of the aminoglycoside antibiotic neomycin B, it is not from the tsHHRz. Despite these pronounced differences in binding, a comparison between the electron spin echo envelope modulation and hyperfine sublevel correlation spectra of the minimal and tertiary stabilized HHRz demonstrates that the structure of both binding sites is very similar. This suggests that the MnII is located in both ribozymes between the bases A9 and G10.1 of the sheared G. A tandem base pair, as shown previously and in detail for the mHHRz. Thus, the much stronger MnII binding in the tsHHRz is attributed to the interaction between the two external loops, which locks in the RNA fold, trapping the MnII in the tightly bound conformation, whereas the absence of long-range loop-loop interactions in the mHHRz leads to more dynamical and open conformations, decreasing MnII binding.}, note = {1355-8382 Letter}, keywords = {Base Sequence Binding Sites Electron Spin Resonance Spectroscopy Framycetin/chemistry/metabolism Kinetics Manganese/*metabolism Nucleic Acid Conformation RNA, Catalytic/*chemistry/*metabolism Research Support, Non-U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Asparaginyl-tRNA Synthetase: Pathways and Evolutionary History of tRNA Asparaginylation}, author = {D Kern and H Roy and H D Becker}, editor = {M Ibba and C Francklyn and S Cusack}, url = {http://www.landesbioscience.com/curie/chapter/1823}, year = {2005}, date = {2005-01-01}, booktitle = {The Aminoacyl-tRNA Synthetases}, pages = {193-209}, publisher = {Landes Bioscience}, abstract = {Asparaginylation of tRNA presents the unusual character that it can be performed by two distinct pathways. A direct, and probably modern one, consisting in direct attachment of Asn on tRNAAsn by asparaginyl-tRNA synthetase (AsnRS), and an indirect, likely ancestral one, in which a mischarged Asp-tRNAAsn, synthesized by a non-discriminating aspartyl-tRNA synthetase (AspRS), is converted into Asn-tRNAAsn by a tRNA-dependent amidotransferase (AdT). This chapter describes the structural and functional properties of enzymes and tRNAs involved in both pathways. AsnRS, the key player in the direct route, is grouped with AspRS and lysyl-tRNA synthetase (LysRS) in subclass IIb aminoacyl-tRNA synthetases displaying a similar modular organization. Despite a remarkable extend of structural and functional similarities, AsnRS and AspRS adapted their mode of recognition to ensure selective binding of the cognate amino acid. Conversely, the enzymes of the indirect pathway (AspRS and AdT) exhibit relaxed specificity. The non-discriminating AspRS is involved in tRNA aspartylation and asparaginylation, whereas AdT can ensure glutaminylation of tRNA in addition to asparaginylation. These features suggest the close functional interrelation between primitive pathways of tRNA aminoacylation and amino acid synthesis. The origin of AsnRS and the interrelation between the direct and indirect pathways of tRNA asparaginylation are investigated on the basis of the distribution of both pathways in the living kingdom and an extended phylogenic analysis.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Crystal quality and differential crystal-growth behaviour of three proteins crystallized in gel at high hydrostatic pressure}, author = {A Kadri and B Lorber and C Charron and M C Robert and B Capelle and M Damak and G Jenner and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15930640}, isbn = {15930640}, year = {2005}, date = {2005-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {61}, number = {Pt 6}, pages = {784-788}, abstract = {Pressure is a non-invasive physical parameter that can be used to control and influence protein crystallization. It is also found that protein crystals of superior quality can be produced in gel. Here, a novel crystallization strategy combining hydrostatic pressure and agarose gel is described. Comparative experiments were conducted on hen and turkey egg-white lysozymes and the plant protein thaumatin. Crystals could be produced under up to 75-100 MPa (lysozymes) and 250 MPa (thaumatin). Several pressure-dependent parameters were determined, which included solubility and supersaturation of the proteins, number, size and morphology of the crystals, and the crystallization volume. Exploration of three-dimensional phase diagrams in which pH and pressure varied identified growth conditions where crystals had largest size and best morphology. As a general trend, nucleation and crystal-growth kinetics are altered and nucleation is always enhanced under pressure. Further, solubility of the lysozymes increases with pressure while that of thaumatin decreases. Likewise, changes in crystallization volumes at high and atmospheric pressure are opposite, being positive for the lysozymes and negative for thaumatin. Crystal quality was estimated by analysis of Bragg reflection profiles and X-ray topographs. While the quality of lysozyme crystals deteriorates as pressure increases, that of thaumatin crystals improves, with more homogeneous crystal morphology suggesting that pressure selectively dissociates ill-formed nuclei. Analysis of the thaumatin structure reveals a less hydrated solvent shell around the protein when pressure increases, with approximately 20% less ordered water molecules in crystals grown at 150 MPa when compared with those grown at atmospheric pressure (0.1 MPa). Noticeably, the altered water distribution is seen in depressurized crystals, indicating that pressure triggers a stable structural alteration on the protein surface while its polypeptide backbone remains essentially unaltered.}, note = {0907-4449 Journal Article}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A surprisingly large RNase P RNA in Candida glabrata}, author = {R Kachouri and V Stribinskis and Y Zhu and K S Ramos and E Westhof and Y Li}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15987816}, isbn = {15987816}, year = {2005}, date = {2005-01-01}, journal = {RNA}, volume = {11}, number = {7}, pages = {1064-1072}, abstract = {We have found an extremely large ribonuclease P (RNase P) RNA (RPR1) in the human pathogen Candida glabrata and verified that this molecule is expressed and present in the active enzyme complex of this hemiascomycete yeast. A structural alignment of the C. glabrata sequence with 36 other hemiascomycete RNase P RNAs (abbreviated as P RNAs) allows us to characterize the types of insertions. In addition, 15 P RNA sequences were newly characterized by searching in the recently sequenced genomes Candida albicans, C. glabrata, Debaryomyces hansenii, Eremothecium gossypii, Kluyveromyces lactis, Kluyveromyces waltii, Naumovia castellii, Saccharomyces kudriavzevii, Saccharomyces mikatae, and Yarrowia lipolytica; and by PCR amplification for other Candida species (Candida guilliermondii, Candida krusei, Candida parapsilosis, Candida stellatoidea, and Candida tropicalis). The phylogenetic comparative analysis identifies a hemiascomycete secondary structure consensus that presents a conserved core in all species with variable insertions or deletions. The most significant variability is found in C. glabrata P RNA in which three insertions exceeding in total 700 nt are present in the Specificity domain. This P RNA is more than twice the length of any other homologous P RNAs known in the three domains of life and is eight times the size of the smallest. RNase P RNA, therefore, represents one of the most diversified noncoding RNAs in terms of size variation and structural diversity.}, note = {1355-8382 Journal Article}, keywords = {Ascomycota/classification/genetics Base Sequence Candida glabrata/chemistry/*enzymology/genetics/*metabolism Comparative Study Conserved Sequence DNA, Chemical Molecular Sequence Data Mutation Nucleic Acid Conformation Phylogeny RNA, Fungal Databases, Fungal Genome, Fungal Models, Fungal/*chemistry/genetics/isolation & purification/metabolism Research Support, Genetic Genes, Non-U.S. Gov't Ribonuclease P/*chemistry/genetics/metabolism Sequence Homology, Nucleic Acid Variation (Genetics), Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence to Structure (S2S): display, manipulate and interconnect RNA data from sequence to structure}, author = {F Jossinet and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15905274}, isbn = {15905274}, year = {2005}, date = {2005-01-01}, journal = {Bioinformatics}, volume = {21}, number = {15}, pages = {3320-3321}, abstract = {SUMMARY: Efficient RNA sequence manipulations (like multiple alignments) need to be constrained by rules of RNA structure folding. The structural knowledge has increased dramatically in the last years with the accumulation of several large RNA structures like those of the bacterial ribosome subunits. However, no tool in the RNA community provides an easy way to link and integrate progress made at the sequence level using the available three-dimensional information. S2S proposes a framework in which an user can easily display, manipulate, and interconnect heterogeneous RNA data like multiple sequence alignments, secondary and tertiary structures. S2S has been implemented with the Java language and has been developed and tested under UNIX systems like Linux and MacOSX. AVAILABILITY: S2S is available at http://bioinformatics.org/S2S/.}, note = {1367-4803 Journal article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Translational operator of mRNA on the ribosome: how repressor proteins exclude ribosome binding}, author = {L Jenner and P Romby and B Rees and C Schulze-Briese and M Springer and C Ehresmann and B Ehresmann and D Moras and G Yusupova and M Yusupov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15802605}, isbn = {15802605}, year = {2005}, date = {2005-01-01}, journal = {Science}, volume = {308}, number = {5718}, pages = {120-123}, abstract = {The ribosome of Thermus thermophilus was cocrystallized with initiator transfer RNA (tRNA) and a structured messenger RNA (mRNA) carrying a translational operator. The path of the mRNA was defined at 5.5 angstroms resolution by comparing it with either the crystal structure of the same ribosomal complex lacking mRNA or with an unstructured mRNA. A precise ribosomal environment positions the operator stem-loop structure perpendicular to the surface of the ribosome on the platform of the 30S subunit. The binding of the operator and of the initiator tRNA occurs on the ribosome with an unoccupied tRNA exit site, which is expected for an initiation complex. The positioning of the regulatory domain of the operator relative to the ribosome elucidates the molecular mechanism by which the bound repressor switches off translation. Our data suggest a general way in which mRNA control elements must be placed on the ribosome to perform their regulatory task.}, note = {1095-9203 Journal Article}, keywords = {16S/chemistry/metabolism RNA, Bacterial/*chemistry/metabolism RNA, Messenger/*chemistry/metabolism RNA, Met/chemistry/metabolism *Regulatory Sequences, Molecular Nucleic Acid Conformation *Protein Biosynthesis RNA, Non-U.S. Gov't Ribosomal Proteins/metabolism Ribosomes/*metabolism Thermus thermophilus/genetics/*metabolism Threonine-tRNA Ligase/genetics/metabolism, Ribonucleic Acid Repressor Proteins/*metabolism Research Support, Ribosomal, ROMBY, ROMBY Bacterial Proteins/metabolism Base Pairing Binding Sites Crystallization Crystallography, Transfer, Unité ARN, X-Ray Fourier Analysis Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Expression of metazoan replication-dependent histone genes}, author = {S Jaeger and S Barends and R Giege and G Eriani and F Martin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16164992}, isbn = {16164992}, year = {2005}, date = {2005-01-01}, journal = {Biochimie}, volume = {87}, number = {9-10}, pages = {827-834}, abstract = {Histone proteins are essential components of eukaryotic chromosomes. In metazoans, they are produced from the so-called replication-dependent histone genes. The biogenesis of histones is tightly coupled to DNA replication in a stoichiometric manner because an excess of histones is highly toxic for the cell. Therefore, a strict cell cycle-regulation of critical factors required for histone expression ensures exclusive S-phase expression. This review focuses on the molecular mechanisms responsible for such a fine expression regulation. Among these, a large part will be dedicated to post-transcriptional events occurring on histone mRNA, like histone mRNA 3' end processing, nucleo-cytoplasmic mRNA export, translation and mRNA degradation. Many factors are involved, including an RNA-binding protein called HBP, also called SLBP (for hairpin- or stem-loop-binding protein) that binds to a conserved hairpin located in the 3' UTR part of histone mRNA. HBP plays a pivotal role in the expression of histone genes since it is necessary for most of the steps of histone mRNA metabolism in the cell. Moreover, the strict S-phase expression pattern of histones is achieved through a fine cell cycle-regulation of HBP. A large part of the discussion will be centered on the critical role of HBP in histone biogenesis.}, note = {0300-9084 Journal Article}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Probing RNA structures with enzymes and chemicals in vitro and in vivo}, author = {E Huntzinger and F Winter and H Moine and C Ehresmann and P Romby}, editor = {R K Hartmann and A Bindereif and A Schon and E Westhof}, url = {http://onlinelibrary.wiley.com/doi/10.1002/9783527619504.ch10/summary}, year = {2005}, date = {2005-01-01}, booktitle = {Handbook of RNA biochemistry}, volume = {1}, pages = {151-171}, publisher = {Wiley-Vch Verlag}, keywords = {ROMBY, ROMBY thiouridine 6-thioguanosine structures relationships variants, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Staphylococcus aureus RNAIII and the endoribonuclease III coordinately regulate spa gene expression}, author = {E Huntzinger and S Boisset and C Saveanu and Y Benito and T Geissmann and A Namane and G Lina and J Etienne and B Ehresmann and C Ehresmann and A Jacquier and F Vandenesch and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15678100}, isbn = {15678100}, year = {2005}, date = {2005-01-01}, journal = {EMBO J}, volume = {24}, number = {4}, pages = {824-835}, abstract = {Staphylococcus aureus RNAIII is one of the largest regulatory RNAs, which controls several virulence genes encoding exoproteins and cell-wall-associated proteins. One of the RNAIII effects is the repression of spa gene (coding for the surface protein A) expression. Here, we show that spa repression occurs not only at the transcriptional level but also by RNAIII-mediated inhibition of translation and degradation of the stable spa mRNA by the double-strand-specific endoribonuclease III (RNase III). The 3' end domain of RNAIII, partially complementary to the 5' part of spa mRNA, efficiently anneals to spa mRNA through an initial loop-loop interaction. Although this annealing is sufficient to inhibit in vitro the formation of the translation initiation complex, the coordinated action of RNase III is essential in vivo to degrade the mRNA and irreversibly arrest translation. Our results further suggest that RNase III is recruited for targeting the paired RNAs. These findings add further complexity to the expression of the S. aureus virulon.}, note = {0261-4189 Journal Article}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Analysis of the contribution of individual substituents in 4,6-aminoglycoside-ribosome interaction}, author = {S N Hobbie and P Pfister and C Brull and E Westhof and E C Bottger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16304180}, isbn = {16304180}, year = {2005}, date = {2005-01-01}, journal = {Antimicrob Agents Chemother}, volume = {49}, number = {12}, pages = {5112-5118}, abstract = {The 4,6-disubstituted 2-deoxystreptamines interfere with protein biosynthesis by specifically targeting the ribosomal A site. These drugs show subtle variations in the chemical groups of rings I, II, and III. In the present study we used site-directed mutagenesis to generate mutant strains of Mycobacterium smegmatis mc(2)155 SMR5 DeltarrnB with mutations in its single rRNA allele, rrnA. This genetic procedure gives rise to strains carrying homogeneous populations of mutant ribosomes and was used to study the contribution of individual chemical substituents to the binding of 4,6-disubstituted aminoglycosides. X-ray crystal structures of geneticin and tobramycin complexed to oligonucleotides containing the minimal bacterial ribosomal A site were used for interpretation of MICs determined for a panel of 4,6-aminoglycosides, including tobramycin, kanamycin A, kanamycin B, amikacin, gentamicin, and geneticin. Surprisingly, the considerable differences present within ring III did not seem to alter the interaction of the drug with the ribosome, as determined by site-directed mutagenesis of the A site. In contrast, subtle variations in ring I significantly influenced binding: (i) a 4'-hydroxyl moiety participates in the proper drug target interaction; and (ii) a 2'-amino group contributes an additional positive charge to ring I, making the drug less susceptible to any kind of sequence alteration within the decoding site, most notably, to conformational changes induced by transversion of U1495 to 1495A. The 4-amino-2-hydroxyl-1-oxobutyl extension at position 1 of ring II of amikacin provides an additional anchor and renders amikacin less dependent on the structural conformation of nucleotide U1406 compared to the dependencies of other kanamycins. Overall, the set of interactions forming the complex between drug substituents and nucleotides of the A site constitutes a network in which the interactions can partly compensate for each other when they are disrupted.}, note = {0066-4804 (Print) Journal Article}, keywords = {Aminoglycosides/chemistry/*pharmacology Anti-Bacterial Agents/chemistry/*pharmacology Binding Sites DNA, Bacterial/genetics Drug Design Mutation Mycobacterium smegmatis/*drug effects/genetics/metabolism RNA, Bacterial/genetics Research Support, Non-U.S. Gov't Ribosomes/chemistry/drug effects/*metabolism Structure-Activity Relationship, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{dumortier_antigen_2005, title = {Antigen presentation by an immature myeloid dendritic cell line does not cause CTL deletion in vivo, but generates CD8+ central memory-like Ŧ cells that can be rescued for full effector function}, author = {Hélène Dumortier and Geertje J D van Mierlo and Deirdre Egan and Willem van Ewijk and René E M Toes and Rienk Offringa and Cornelis J M Melief}, doi = {10.4049/jimmunol.175.2.855}, issn = {0022-1767}, year = {2005}, date = {2005-01-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {175}, number = {2}, pages = {855--863}, abstract = {Immature dendritic cells (DC), in contrast to their mature counterparts, are incapable of mobilizing a CD8+ CTL response, and, instead, have been reported to induce CTL tolerance. We directly addressed the impact of immature vs mature DC on CTL responses by infusing adenovirus peptide-loaded DC (of the D1 cell line) into mice that had received adenovirus-specific naive TCR-transgenic CD8+ T cells. Whereas i.v. injection of mature DC triggered vigorous CTL expansion, immature DC elicited little proliferation involving only a minority of the TCR-transgenic CTL. Even though the latter CTL developed effector functions, including cytolytic activity and proinflammatory cytokine secretion, these cells differed significantly from CTL primed by mature DC in that they did not exhibit down-regulation of CD62L and CCR7, receptors involved in trapping of T cells in the lymphoid organs. Interestingly, adoptive transfer of CTL effector cells harvested after priming by either mature or immature DC into naive recipient mice, followed by exposure to adenovirus, yielded quantitatively and qualitatively indistinguishable CTL memory responses. Therefore, in vivo priming of naive CD8+ T cells by immature DC, although failing to induce a full-blown, systemic CTL response, resulted in the formation of central memory-like T cells that were able to expand and produce IFN-gamma upon secondary antigenic stimulation.}, keywords = {Adenovirus E1A Proteins, Animals, Antigen, Antigen Presentation, CD8-Positive T-Lymphocytes, Cell Differentiation, Cell Line, Cell Movement, Clonal Deletion, Cytotoxic, Cytotoxicity, Dendritic Cells, Down-Regulation, Dumortier, Epitopes, Female, I2CT, Immunologic, Immunologic Memory, Inbred C57BL, Lipopolysaccharides, Lymphocyte Activation, Mice, Myeloid Cells, Receptors, Regulatory, T-Cell, T-Lymphocyte, T-Lymphocytes, Team-Dumortier, transgenic}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cooperative and specific binding of Vif to the 5' region of HIV-1 genomic RNA}, author = {S Henriet and D Richer and S Bernacchi and E Decroly and R Vigne and B Ehresmann and C Ehresmann and J C Paillart and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16236319}, isbn = {16236319}, year = {2005}, date = {2005-01-01}, journal = {J Mol Biol}, volume = {354}, number = {1}, pages = {55-72}, abstract = {The viral infectivity factor (Vif) protein of human immunodeficiency virus type 1 (HIV-1) is essential for viral replication in vivo. Packaging of Vif into viral particles is mediated by an interaction with viral genomic RNA and association with viral nucleoprotein complexes. Despite recent findings on the RNA-binding properties of Vif suggesting that Vif could be involved in retroviral assembly, no RNA sequence or structure specificity has been determined so far. To gain further insight into the mechanisms by which Vif might regulate viral replication, we studied the interactions of Vif with HIV-1 genomic RNA in vitro. Using extensive biochemical analysis, we have measured the affinity of recombinant Vif proteins for synthetic RNAs corresponding to various regions of the HIV-1 genome. We found that recombinant Vif proteins bind specifically to HIV-1 viral RNA fragments corresponding to the 5'-untranslated region (5'-UTR), gag and the 5' part of pol (K(d) between 45 nM and 65 nM). RNA encompassing nucleotides 1-497 or 499-996 of the HIV-1 genomic RNA bind 9+/-2 and 21+/-3 Vif molecules, respectively, and at least some of these proteins bind in a cooperative manner (Hill constant alpha(H) = 2.3). In contrast, RNAs corresponding to other parts of the HIV-1 genome or heterologous RNAs showed poor binding capacity and weak cooperativity (K(d) > 200 nM). Moreover, RNase T1 footprinting revealed a hierarchical binding of Vif, pointing to TAR and the poly(A) stem-loop structures as primary strong affinity targets, and downstream structures as secondary sites with moderate affinity. Taken together, our findings suggest that Vif may assist other proteins to maintain a correct folding of the genomic RNA in order to facilitate its packaging and further steps such as reverse transcription. Interestingly, our results suggest also that Vif could bind the viral RNA in order to protect it from the action of the antiviral factor APOBEC-3G/3F.}, note = {0022-2836 (Print) Journal Article}, keywords = {5' Untranslated Regions/chemistry/*metabolism Base Sequence Electrophoretic Mobility Shift Assay Gene Products, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, vif/*metabolism HIV Long Terminal Repeat HIV-1/*genetics/*metabolism Hela Cells Humans Molecular Sequence Data Nucleic Acid Conformation Protein Binding RNA, Viral/chemistry/*metabolism RNA-Binding Proteins/metabolism Recombinant Proteins/metabolism Research Support}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {RNA structure: pseudoknots}, author = {A P Gultyaev and C W A Pleij and E Westhof}, url = {http://www.els.net/WileyCDA/ElsArticle/refId-a0003134.html}, doi = {10.1038/npg.els.0003134}, year = {2005}, date = {2005-01-01}, booktitle = {Encyclopedia of Life Sciences}, publisher = {John Wiley & Sons}, abstract = {An RNA pseudoknot results from Watson–Crick base pairing interactions of a single-stranded segment, occurring between two paired sequence fragments, with a complementary sequence that is not located between the paired fragments. RNA pseudoknots adopt different folding topologies and are an essential part of various functional RNA molecules.}, keywords = {RNA folding RNA secondary structure RNA tertiary structure ribozyme ribosome reprogramming frameshifting, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Role of Arc1p in the modulation of yeast glutamyl-tRNA synthetase activity}, author = {J S Graindorge and B Senger and D Tritch and G Simos and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15667228}, isbn = {15667228}, year = {2005}, date = {2005-01-01}, journal = {Biochemistry}, volume = {44}, number = {4}, pages = {1344-1352}, abstract = {Yeast methionyl-tRNA synthetase (MetRS) and glutamyl-tRNA synthetase (GluRS) possess N-terminal extensions that bind the cofactor Arc1p in trans. The strength of GluRS-Arc1p interaction is high enough to allow copurification of the two macromolecules in a 1:1 ratio, in contrast to MetRS. Deletion analysis from the C-terminal end of the GluRS appendix combined with previous N-terminal deletions of GluRS allows restriction of the Arc1p binding site to the 110-170 amino acid region of GluRS. This region has been shown to correspond to a novel protein-protein interaction domain present in both GluRS and Arc1p but not in MetRS [Galani, K., Grosshans, H., Deinert, K., Hurt, E. C., and Simos, G. (2001) EMBO J. 20, 6889-6898]. The GluRS apoenzyme fails to show significant kinetics of tRNA aminoacylation and charges unfractionated yeast tRNA at a level 10-fold reduced compared to Arc1p-bound GluRS. The K(m) values for tRNA(Glu) measured in the ATP-PP(i) exchange were similar for the two forms of GluRS, whereas k(cat) is increased 2-fold in the presence of Arc1p. Band-shift analysis revealed a 100-fold increase in tRNA binding affinity when Arc1p is bound to GluRS. This increase requires the RNA binding properties of the full-length Arc1p since Arc1p N domain leaves the K(d) of GluRS for tRNA unchanged. Transcripts of yeast tRNA(Glu) were poor substrates for measuring tRNA aminoacylation and could not be used to clarify whether Arc1p has a specific effect on the tRNA charging reaction.}, note = {0006-2960 Journal Article}, keywords = {FASIOLO Adenosine Triphosphate/chemistry/metabolism Amino Acid Sequence Aminoacylation Base Sequence Diphosphates/chemistry/metabolism Enzyme Activation Gene Expression Regulation, Fungal Glutamate-tRNA Ligase/isolation & purification/*metabolism Kinetics Molecular Sequence Data Peptide Fragments/chemistry/metabolism Protein Binding Protein Structure, Fungal/genetics/metabolism RNA, Genetic, Glu/genetics/metabolism RNA-Binding Proteins/*chemistry/isolation & purification/metabolism Research Support, Non-U.S. Gov't Saccharomyces cerevisiae/enzymology/genetics/metabolism Saccharomyces cerevisiae Proteins/*chemistry/isolation & purification/metabolism Transcription, Tertiary RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Deletion of EFL1 Results in Heterogeneity of the 60S GTPase-associated rRNA Conformation}, author = {J S Graindorge and J C Rousselle and B Senger and P Lenormand and A Namane and F Lacroute and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16095611}, isbn = {16095611}, year = {2005}, date = {2005-01-01}, journal = {J Mol Biol}, volume = {352}, number = {2}, pages = {355-369}, abstract = {Previous work suggested that the release of the nucleolar Tif6 from nascent 60S subunits occurs in the cytoplasm and requires the cytoplasmic EF-2-like GTPase, Efl1. To check whether this release involves an rRNA structural rearrangement mediated by Efl1, we analyzed the rRNA conformation of the GTPase center of 80S ribosomes in three contexts: wild-type, Deltaefl1 and a dominant suppressor R1 of Deltaefl1. This analysis was restricted to domain II and VI of 25S rRNA. The rRNA analysis of R1 ribosomes allows us to distinguish the effects due to depletion of Efl1 from the resulting nucleolar deficit of Tif6. Efl1 inhibits the EF-2 GTPase activity, suggesting that the two proteins share a similar ribosome-binding site. The 80S ribosomes from either type failed to show any difference of conformation in the two rRNA domains analyzed. However, the same analysis performed on the pool of free 60S subunits reveals several rRNA conformational differences between wild-type and Deltaefl1 subunits, whereas that from the suppressor strain is similar to wild-type. This suggests that the nucleolar deficit of Tif6 during assembly of the 60S preribosomes is responsible for the changes in rRNA conformation observed in Deltaefl1 60S subunits. We also purified 60S preribosomes from the three genetic contexts by TAP-tagging Tif6. The protein content of 60S preribosomes associated with Tif6p in a Deltaefl1 strain are obtained at a lower yield but have, surprisingly, a protein composition that is a priori similar to that of wild-type and the suppressor strain.}, note = {0022-2836 Journal Article}, keywords = {FASIOLO, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Aspartyl-tRNA synthetases}, author = {R Giege and B Rees}, editor = {M Ibba and C Francklyn and S Cusack}, url = {http://makeover.landesbioscience.com/curie/chapter/1877}, year = {2005}, date = {2005-01-01}, booktitle = {The Aminoacyl-tRNA Synthetases}, pages = {1-26}, publisher = {Landes Biosciences}, abstract = {Aspartyl-tRNA synthetases (AspRSs) belong to subclass IIb of synthetases. The subunits of these dimeric proteins have a conserved modular architecture in the three kingdoms of life, comprising a C-terminal active site domain linked by a short hinge domain to an N-terminal anticodon-binding domain. An additional flexible domain is appended at the N-terminus of eukaryotic AspRSs that helps to better anchor the tRNA on the synthetase body. Eubacterial AspRSs are characterized by an insertion module in the active site domain, while archaeal AspRSs display the smallest structures. Sequence and especially three-dimensional structure comparisons indicate mimicries between AspRS modules and various other proteins. X-ray structures of AspRSs complexed with their ligands, separately or in combination, and results of site directed mutagenesis on tRNA and synthetase provide a detailed mechanistic understanding of the tRNA aspartylation reaction. In this process, accompanied by conformational changes in the synthetase and the interacting tRNAAsp, AspRS conserved residues and class and subclass defining motifs, together with identity determinants in tRNAAsp, play crucial roles.}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {tRNA-balanced expression of a eukaryal aminoacyl-tRNA synthetase by an mRNA-mediated pathway}, author = {M Frugier and M Ryckelynck and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16113655}, isbn = {16113655}, year = {2005}, date = {2005-01-01}, journal = {EMBO Rep}, volume = {6}, number = {9}, pages = {860-865}, abstract = {Aminoacylation of transfer RNAs is a key step during translation. It is catalysed by the aminoacyl-tRNA synthetases (aaRSs) and requires the specific recognition of their cognate substrates, one or several tRNAs, ATP and the amino acid. Whereas the control of certain aaRS genes is well known in prokaryotes, little is known about the regulation of eukaryotic aaRS genes. Here, it is shown that expression of AspRS is regulated in yeast by a feedback mechanism that necessitates the binding of AspRS to its messenger RNA. This regulation leads to a synchronized expression of AspRS and tRNA(Asp). The correlation between AspRS expression and mRNA(AspRS) and tRNA(Asp) concentrations, as well as the presence of AspRS in the nucleus, suggests an original regulation mechanism. It is proposed that the surplus of AspRS, not sequestered by tRNA(Asp), is imported into the nucleus where it binds to mRNA(AspRS) and thus inhibits its accumulation.}, note = {1469-221x Journal article}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Introduction to non-watson-crick base pairs and RNA folding}, author = {V Fritsch and E Westhof}, editor = {D Chatenay and S Cocco and R Monasson and D Thieffry and J Dalibard}, url = {http://www.sciencedirect.com/science/article/pii/S0924809905800290}, doi = {10.1016/S0924-8099(05)80029-0}, year = {2005}, date = {2005-01-01}, booktitle = {Multiple aspects of DNA: from Biophysics to Bioinformatics. Lecture Notes of the Les Houches Summer School 2004}, volume = {82}, pages = {41-72}, publisher = {Elsevier}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Crystal structures of complexes between aminoglycosides and decoding A site oligonucleotides: role of the number of rings and positive charges in the specific binding leading to miscoding}, author = {B Francois and R J Russell and J B Murray and F Aboul-ela and B Masquida and Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16214802}, isbn = {16214802}, year = {2005}, date = {2005-01-01}, journal = {Nucleic Acids Res}, volume = {33}, number = {17}, pages = {5677-5690}, abstract = {The crystal structures of six complexes between aminoglycoside antibiotics (neamine, gentamicin C1A, kanamycin A, ribostamycin, lividomycin A and neomycin B) and oligonucleotides containing the decoding A site of bacterial ribosomes are reported at resolutions between 2.2 and 3.0 A. Although the number of contacts between the RNA and the aminoglycosides varies between 20 and 31, up to eight direct hydrogen bonds between rings I and II of the neamine moiety are conserved in the observed complexes. The puckered sugar ring I is inserted into the A site helix by stacking against G1491 and forms a pseudo base pair with two H-bonds to the Watson-Crick sites of the universally conserved A1408. This central interaction helps to maintain A1492 and A1493 in a bulged-out conformation. All these structures of the minimal A site RNA complexed to various aminoglycosides display crystal packings with intermolecular contacts between the bulging A1492 and A1493 and the shallow/minor groove of Watson-Crick pairs in a neighbouring helix. In one crystal, one empty A site is observed. In two crystals, two aminoglycosides are bound to the same A site with one bound specifically and the other bound in various ways in the deep/major groove at the edge of the A sites.}, note = {1362-4962 (Electronic) Journal Article}, keywords = {16S/*chemistry Research Support, Adenine/chemistry Aminoglycosides/*chemistry Anti-Bacterial Agents/*chemistry Anticodon/chemistry Base Sequence Binding Sites Codon/chemistry Crystallography, Molecular Oligoribonucleotides/*chemistry Paromomycin/analogs & derivatives/chemistry RNA, Non-U.S. Gov't Ribosomes/chemistry Ribostamycin/chemistry, Ribosomal, Unité ARN, WESTHOF, X-Ray Framycetin/chemistry Gentamicins/chemistry Kanamycin/chemistry *Models}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Preparation and handing of RNA crystals}, author = {B Francois and A Lescoute and A Werner and B Masquida}, editor = {R K Hartmann and A Bindereif and A Schon and E Westhof}, url = {http://onlinelibrary.wiley.com/book/10.1002/9783527619504}, year = {2005}, date = {2005-01-01}, booktitle = {Handbook of RNA Biochemistry}, pages = {438-452}, publisher = {Wiley-Vch Verlag, Weinheim}, keywords = {Unité ARN, WESTHOF Crystals Preparation Programs Characterization Contaminants}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Participation a la mise à jour de l'ouvrage}, author = {C Florentz}, editor = {J H Weil}, url = {none}, year = {2005}, date = {2005-01-01}, booktitle = {Biochimie Generale, 10e edition}, publisher = {Dunod}, keywords = {FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Arginyl-tRNA synthetase}, author = {G Eriani and J Cavarelli}, editor = {M Ibba and C Francklyn and S Cusack}, url = {http://www.landesbioscience.com/curie/chapter/1284}, year = {2005}, date = {2005-01-01}, booktitle = {The Aminoacyl-tRNA Synthetases}, publisher = {Landes Bioscience}, abstract = {Determination of the crystal structures of arginyl-tRNA synthetase, either in the free state or engaged in complexes with the other partners of the arginylation reaction, led to fundamental progress in understanding the sequence-structure-function relationship of this catalytic reaction. The structures reveal unexpected results simplifying and organizing the collected biological information but also illustrating the inherent complexity of this macromolecular recognition process. ArgRS specifically recognizes the D-loop and the anticodon of tRNAArg using dedicated modules. Conformational changes which occur upon substrates binding have been visualized at the atomic level. While ArgRS requires its cognate tRNA for the first step of the aminoacylation reaction, the crystal structures reveal that (i) L-arginine binding controls the correct positioning of the CCA end of tRNAArg and that (ii) tRNAArg binding produces conformational changes of the ATP-binding cleft. In this review, results from extensive investigations preformed by several groups are summarized.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Purification, crystallization and preliminary X-ray diffraction studies of the archaeal virus resolvase SIRV2}, author = {E Ennifar and J Basquin and R Birkenbihl and D Suck}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16511081}, isbn = {16511081}, year = {2005}, date = {2005-01-01}, journal = {Acta Crystallogr F Struct Biol Commun}, volume = {61}, number = {Pt 5}, pages = {507-509}, abstract = {The Holliday junction (or four-way junction) is the universal DNA intermediate whose interaction with resolving proteins is one of the major events in the recombinational process. These proteins, called DNA junction-resolving enzymes or resolvases, bind to the junction and catalyse DNA cleavage, promoting the release of two DNA duplexes. SIRV2 Hjc, a viral resolvase infecting a thermophylic archaeon, has been cloned, expressed and purified. Crystals have been obtained in space group C2, with unit-cell parameters a = 147.8}, note = {1744-3091 (Electronic) Journal Article}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Threonyl-tRNA synthetase: a multifunctional enzyme in E.coli.}, author = {A C Dock-Bregeon and P Romby and M Springer}, editor = {M Ibba and C Francklyn and S Cusack}, url = {http://www.landesbioscience.com/curie/chapter/1876}, year = {2005}, date = {2005-01-01}, booktitle = {The aminoacyl-tRNA synthetases}, pages = {163-176}, publisher = {Landes Bioscience}, abstract = {Aminoacyl-tRNA synthetases were discovered in the mid-nineteenfifties1 as モactivating enzymesヤ that yielded モ… an enzyme bound, carboxyl-activated, aminoacid- AMP compoundヤ. These activating enzymes were fractionated such that Hoagland was able to say モseparate enzymes are involved in the activation of several aminoacids.ヤ2 At about the same time, F. Crick proposed his adaptor hypothesis.3 This brilliant hypothesis was confirmed when Hoagland et al4 discovered that their モactivating enzymesヤ were able to attach an amino acid to a small RNA, transfer RNA, that had all the properties of Crickメs adaptor.}, keywords = {ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Inhibition of human immunodeficiency virus type 1 reverse transcriptase, RNase H, and integrase activities by hydroxytropolones}, author = {J Didierjean and C Isel and F Querre and J F Mouscadet and A M Aubertin and J Y Valnot and S R Piettre and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16304149}, isbn = {16304149}, year = {2005}, date = {2005-01-01}, journal = {Antimicrob Agents Chemother}, volume = {49}, number = {12}, pages = {4884-4894}, abstract = {Human immunodeficiency virus type I reverse transcriptase (RT) possesses distinct DNA polymerase and RNase H sites, whereas integrase (IN) uses the same active site to perform 3'-end processing and strand transfer of the proviral DNA. These four enzymatic activities are essential for viral replication and require metal ions. Two Mg2+ ions are present in the RT polymerase site, and one or two Mg2+ ions are required for the catalytic activities of RNase H and IN. We tested the possibility of inhibition of the RT polymerase and RNase H as well as the IN 3'-end processing and transfer activities of purified enzymes by a series of 3,7-dihydroxytropolones designed to target two Mg2+ ions separated by approximately 3.7 angstroms. The RT polymerase and IN 3' processing and strand transfer activities were inhibited at submicromolar concentrations, while the RNase H activity was inhibited in the low micromolar range. In all cases, the lack of inhibition by tropolones and O-methylated 3,7-dihydroxytropolones was consistent with the active molecules binding the metal ions in the active site. In addition, inhibition of the DNA polymerase activity was shown to depend on the Mg2+ concentration. Furthermore, selective inhibitors were identified for several of the activities tested, leaving some potential for design of improved inhibitors. However, all tested compounds exhibited cellular toxicity that presently limits their applications.}, note = {0066-4804 (Print) Journal Article}, keywords = {Calf Thymus/*antagonists & inhibitors Tropolone/analogs & derivatives/*pharmacology, Enzyme Inhibitors/*pharmacology HIV Integrase/*metabolism HIV Integrase Inhibitors/chemistry/pharmacology HIV-1/*drug effects/enzymology HIV-1 Reverse Transcriptase/*antagonists & inhibitors Humans Research Support, MARQUET, Non-U.S. Gov't Ribonuclease H, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization and preliminary X-ray crystallographic study of the wild type and two mutants of the CP1 hydrolytic domain from Aquifex aeolicus leucyl-tRNA synthetase}, author = {V Cura and N Olieric and A Guichard and E D Wang and D Moras and G Eriani and J Cavarelli}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16511190}, isbn = {16511190}, year = {2005}, date = {2005-01-01}, journal = {Acta Crystallogr F Struct Biol Commun}, volume = {61}, number = {Pt 10}, pages = {899-901}, abstract = {The editing or hydrolytic CP1 domain of leucyl-tRNA synthetase (LeuRS) hydrolyses several misactivated amino acids. The CP1 domain of Aquifex aeolicus LeuRS was expressed, purified and crystallized by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. Crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 38.8}, note = {1744-3091 (Electronic) Journal Article}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Activation of the hetero-octameric ATP phosphoribosyl transferase through subunit interface rearrangement by a tRNA synthetase paralog}, author = {K S Champagne and M Sissler and Y Larrabee and S Doublie and C S Francklyn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16051603}, isbn = {16051603}, year = {2005}, date = {2005-01-01}, journal = {J Biol Chem}, volume = {280}, number = {40}, pages = {34096-34104}, abstract = {ATP phosphoribosyl transferase (ATP-PRT) joins ATP and 5-phosphoribosyl-1-pyrophosphate (PRPP) in a highly regulated reaction that initiates histidine biosynthesis. The unusual hetero-octameric version of ATP-PRT includes four HisG(S) catalytic subunits based on the periplasmic binding protein fold and four HisZ regulatory subunits that resemble histidyl-tRNA synthetases. Here, we present the first structure of a PRPP-bound ATP-PRT at 2.9 A and provide a structural model for allosteric activation based on comparisons with other inhibited and activated ATP-PRTs from both the hetero-octameric and hexameric families. The activated state of the octameric enzyme is characterized by an interstitial phosphate ion in the HisZ-HisG interface and new contacts between the HisZ motif 2 loop and the HisG(S) dimer interface. These contacts restructure the interface to recruit conserved residues to the active site, where they activate pyrophosphate to promote catalysis. Additionally, mutational analysis identifies the histidine binding sites within a region highly conserved between HisZ and the functional HisRS. Through the oligomerization and functional re-assignment of protein domains associated with aminoacylation and phosphate binding, the HisZ-HisG octameric ATP-PRT acquired the ability to initiate the synthesis of a key metabolic intermediate in an allosterically regulated fashion.}, note = {0021-9258 (Print) Journal Article}, keywords = {Extramural Research Support, FLORENTZ ATP Phosphoribosyltransferase/*metabolism Allosteric Regulation Amino Acyl-tRNA Synthetases DNA Mutational Analysis Enzyme Activation *Models, N.I.H., Non-P.H.S. Research Support, P.H.S., SISSLER, Structural Phosphates/metabolism Research Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The N-terminus of HIV-1 Tat protein is essential for Tat-TAR RNA interaction}, author = {O Chaloin and J C Peter and J P Briand and B Masquida and C Desgranges and S Muller and J Hoebeke}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15723170}, isbn = {15723170}, year = {2005}, date = {2005-01-01}, journal = {Cell Mol Life Sci}, volume = {62}, number = {3}, pages = {355-361}, abstract = {The human HIV transactivator protein Tat is essential for efficient viral transcription that occurs by a complex mechanism involving interaction of Tat with the TAR RNA element. This interaction appears to require the mediation of a cellular protein, cyclin T1. However, the possibility that Tat and TAR associate in a binary Tat-TAR complex has been little investigated. Using a chemically synthesized active Tat protein, the kinetic and equilibrium parameters of its interaction with TAR were determined by surface plasmon resonance technology. Independently of partner and method of immobilization onto the sensor chip, the association (k(a) = 5-9 x 105 M(-1) s(-1)) and dissociation rate constants (k(d) = 1.7-4.3 x 10(-3) s(-1)) yielded similar equilibrium dissociation constants (K(d) = 2-8 nM). A truncated peptide encompassing residues 30-86 of Tat did not bind to TAR at all. We conclude that Tat can form a high-affinity complex with TAR in the absence of cyclin T1 and that the N-terminal domain of Tat is essential for this interaction, suggesting a conformational link between this domain and the basic domain of Tat. These results are important in our quest for developing therapeutic compounds that impair viral replication.}, note = {1420-682x Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Diversity and functional plasticity of eukaryotic selenoproteins: identification and characterization of the SelJ family}, author = {S Castellano and A V Lobanov and C Chapple and S V Novoselov and M Albrecht and D Hua and A Lescure and T Lengauer and A Krol and V N Gladyshev and R Guigo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16260744}, isbn = {16260744}, year = {2005}, date = {2005-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {102}, number = {45}, pages = {16188-16193}, abstract = {Selenoproteins are a diverse group of proteins that contain selenocysteine (Sec), the 21st amino acid. In the genetic code, UGA serves as a termination signal and a Sec codon. This dual role has precluded the automatic annotation of selenoproteins. Recent advances in the computational identification of selenoprotein genes have provided a first glimpse of the size, functions, and phylogenetic diversity of eukaryotic selenoproteomes. Here, we describe the identification of a selenoprotein family named SelJ. In contrast to known selenoproteins, SelJ appears to be restricted to actinopterygian fishes and sea urchin, with Cys homologues only found in cnidarians. SelJ shows significant similarity to the jellyfish J1-crystallins and with them constitutes a distinct subfamily within the large family of ADP-ribosylation enzymes. Consistent with its potential role as a structural crystallin, SelJ has preferential and homogeneous expression in the eye lens in early stages of zebrafish development. A structural role for SelJ would be in contrast to the majority of known selenoenzymes. The unusually highly restricted phylogenetic distribution of SelJ, its specialization, and the comparative analysis of eukaryotic selenoproteomes reveal the diversity and functional plasticity of selenoproteins and point to a mosaic evolution of the use of Sec in proteins.}, note = {0027-8424 (Print) Journal Article}, keywords = {Extramural Research Support, KROL Adenosine Diphosphate Ribose/metabolism Animals Fish Proteins/chemistry/genetics/*physiology Genome Mice NIH 3T3 Cells Phylogeny Promoter Regions (Genetics) Proteome Research Support, LESCURE, N.I.H., Non-P.H.S. Selenoproteins/chemistry/genetics/*physiology Tetraodontiformes/*genetics, Non-U.S. Gov't Research Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Evolution of the tRNA(Tyr)/TyrRS aminoacylation systems}, author = {L Bonnefond and R Giege and J Rudinger-Thirion}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16164994}, isbn = {16164994}, year = {2005}, date = {2005-01-01}, journal = {Biochimie}, volume = {87}, number = {9-10}, pages = {873-883}, abstract = {The tRNA identity rules ensuring fidelity of translation are globally conserved throughout evolution except for tyrosyl-tRNA synthetases (TyrRSs) that display species-specific tRNA recognition. This discrimination originates from the presence of a conserved identity pair, G1-C72, located at the top of the acceptor stem of tRNA(Tyr) from eubacteria that is invariably replaced by an unusual C1-G72 pair in archaeal and eubacterial tRNA(Tyr). In addition to the key role of pair 1-72 in tyrosylation, discriminator base A73, the anticodon triplet and the large variable region (present in eubacterial tRNA(Tyr) but not found in eukaryal tRNA(Tyr)) contribute to tyrosylation with variable strengths. Crystallographic structures of two tRNA(Tyr)/TyrRS complexes revealed different interaction modes in accordance with the phylum-specificity. Recent functional studies on the human mitochondrial tRNA(Tyr)/TyrRS system indicates strong deviations from the canonical tyrosylation rules. These differences are discussed in the light of the present knowledge on TyrRSs.}, note = {0300-9084 Journal Article}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Human mitochondrial TyrRS disobeys the tyrosine identity rules}, author = {L Bonnefond and M Frugier and R Giege and J Rudinger-Thirion}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15840810}, isbn = {15840810}, year = {2005}, date = {2005-01-01}, journal = {RNA}, volume = {11}, number = {5}, pages = {558-562}, abstract = {Human tyrosyl-tRNA synthetase from mitochondria (mt-TyrRS) presents dual sequence features characteristic of eubacterial and archaeal TyrRSs, especially in the region containing amino acids recognizing the N1-N72 tyrosine identity pair. This would imply that human mt-TyrRS has lost the capacity to discriminate between the G1-C72 pair typical of eubacterial and mitochondrial tRNATyr and the reverse pair C1-G72 present in archaeal and eukaryal tRNATyr. This expectation was verified by a functional analysis of wild-type or mutated tRNATyr molecules, showing that mt-TyrRS aminoacylates with similar catalytic efficiency its cognate tRNATyr with G1-C72 and its mutated version with C1-G72. This provides the first example of a TyrRS lacking specificity toward N1-N72 and thus of a TyrRS disobeying the identity rules. Sequence comparisons of mt-TyrRSs across phylogeny suggest that the functional behavior of the human mt-TyrRS is conserved among all vertebrate mt-TyrRSs.}, note = {1355-8382 Journal Article}, keywords = {Amino Acid Sequence Base Sequence Catalytic Domain Humans Mitochondria/*enzymology Molecular Sequence Data RNA, FRUGIER, Non-U.S. Gov't Substrate Specificity Tyrosine/genetics/*metabolism Tyrosine-tRNA Ligase/chemistry/*metabolism, Transfer, Tyr/genetics/*metabolism Research Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Toward the Full Set of Human Mitochondrial Aminoacyl-tRNA Synthetases: Characterization of AspRS and TyrRS}, author = {L Bonnefond and A Fender and J Rudinger-Thirion and R Giege and C Florentz and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15779907}, isbn = {15779907}, year = {2005}, date = {2005-01-01}, journal = {Biochemistry}, volume = {44}, number = {12}, pages = {4805-4816}, abstract = {The human mitochondrion possesses a translational machinery devoted to the synthesis of 13 proteins. While the required tRNAs and rRNAs are produced by transcription of the mitochondrial genome, all other factors needed for protein synthesis are synthesized in the cytosol and imported. This is the case for aminoacyl-tRNA synthetases, the enzymes which esterify their cognate tRNA with the specific amino acid. The genes for the full set of cytosolic aaRSs are well defined, but only nine genes for mitochondrial synthetases are known. Here we describe the genes for human mitochondrial aspartyl- and tyrosyl-tRNA synthetases and the initial characterization of the enzymes. Both belong to the expected class of synthetases, have a dimeric organization, and aminoacylate Escherichia coli tRNAs as well as in vitro transcribed human mitochondrial tRNAs. Genes for the remaining missing synthetases were also found with the exception of glutaminyl-tRNA synthetase. Their sequence analysis confirms and further extends the view that, except for lysyl- and glycyl-tRNA synthetases, human mitochondrial and cytosolic enzymes are coded by two different sets of genes.}, note = {0006-2960 Journal Article}, keywords = {FLORENTZ, FRUGIER, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Glu-Q-tRNA(Asp) synthetase coded by the yadB gene, a new paralog of aminoacyl-tRNA synthetase that glutamylates tRNA(Asp) anticodon}, author = {M Blaise and H D Becker and J Lapointe and C Cambillau and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16164993}, isbn = {16164993}, year = {2005}, date = {2005-01-01}, journal = {Biochimie}, volume = {87}, number = {9-10}, pages = {847-861}, abstract = {Analysis of the completed genome sequences revealed presence in various bacteria of an open reading frame (ORF) encoding a polypeptide chain presenting important similarities with the catalytic domain of glutamyl-tRNA synthetases but deprived of the C-terminal anticodon-binding domain. This paralog of glutamyl-tRNA synthetases, the YadB protein, activates glutamate in the absence of tRNA and transfers the activated glutamate not on tRNA(Glu) but instead on tRNA(Asp). It has been shown that tRNA(Asp) is able to accept two amino acids: aspartate charged by aspartyl-tRNA synthetase and glutamate charged by YadB. The functional properties of YadB contrast with those of the canonical glutamyl-tRNA synthetases, which activate Glu only in presence of the cognate tRNA before aminoacylation of the 3'-end of tRNA. Biochemical approaches and mass spectrometry investigations revealed that YadB transfers the activated glutamate on the cyclopenthene-diol ring of the modified nucleoside queuosine posttranscriptionally inserted at the wobble position of the anticodon-loop to form glutamyl-queuosine. Unstability of the ester bond between the glutamate residue and the cyclopenthene-diol (half-life 7.5 min) explains why until now this modification escaped detection. Among Escherichia coli tRNAs containing queuosine in the wobble position, only tRNA(Asp) is substrate of YadB. Sequence comparison reveals a structural mimicry between the anticodon-stem and loop of tRNA(Asp) and the amino acid acceptor-stem of tRNA(Glu). YadB, renamed glutamyl-Q-tRNA(Asp) synthetase, constitutes the first enzyme structurally related to aminoacyl-tRNA synthetases which catalyzes a hypermodification in tRNA, and whose function seems to be conserved among prokaryotes. The discovery of glutamyl-Q-tRNA(Asp) synthetase breaks down the current paradigm according to which the catalytic domain of aminoacyl-tRNA synthetases recognizes the amino acid acceptor-stem of tRNA and aminoacylates the 3'-terminal ribose. The evolutionary significance of the existence of an aminoacyl-tRNA synthetase paralog dedicated to the hypermodification of a tRNA anticodon will be discussed.}, note = {0300-9084 Journal Article}, keywords = {GIEGE KERN, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of crystals of the Hjc resolvase from Archaeoglobus fulgidus grown in gel by counter-diffusion.}, author = {C Biertümpfel and J Basquin and R Birkenbihl and D Suck and C Sauter}, url = {http://scripts.iucr.org/cgi-bin/paper?za5109}, doi = {10.1107/S1744309105018269}, year = {2005}, date = {2005-01-01}, journal = {Acta Crystallogr F Struct Biol Commun}, volume = {61}, number = {7}, pages = {684-687}, abstract = {Holliday junction-resolving enzymes are ubiquitous proteins that play a key role in DNA repair and reorganization by homologous recombination. The Holliday junction-cutting enzyme (Hjc) from the archaeon Archaeoglobus fulgidus is a member of this group. The first Hjc crystals were obtained by conventional sparse-matrix screening. They exhibited an unusually elongated unit cell and their X-ray characterization required special care to avoid spot overlaps along the c* axis. The use of an arc appended to the goniometric head allowed proper orientatation of plate-like crystals grown in agarose gel by counter-diffusion. Thus, complete diffraction data were collected at 2.7 Å resolution using synchrotron radiation. They belong to space group P3121 or P3221, with unit-cell parameters a = b = 37.4}, keywords = {FRUGIER, GIEGE Holliday junction-cutting enzyme Hjc Resolvase Counter-duffusion Agarose gel, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mechanism of hairpin-duplex conversion for the HIV-1 dimerization initiation site}, author = {S Bernacchi and E Ennifar and K Toth and P Walter and J Langowski and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16169845}, isbn = {16169845}, year = {2005}, date = {2005-01-01}, journal = {J Biol Chem}, volume = {280}, number = {48}, pages = {40112-40121}, abstract = {We have used the dimerization initiation site of HIV-1 genomic RNA as a model to investigate hairpin-duplex interconversion with a combination of fluorescence, UV melting, gel electrophoresis, and x-ray crystallographic techniques. Fluorescence studies with molecular beacons and crystallization experiments with 23-nucleotide dimerization initiation site fragments showed that the ratio of hairpin to duplex formed after annealing in water essentially depends on RNA concentration and not on cooling kinetics. With natural sequences allowing to form the most stable duplex, and thus also the loop-loop complex (or "kissing complex"), concentrations as low as 3 mum in strands are necessary to obtain a majority of the hairpin form. With a mutated sequence preventing kissing complex formation, a majority of hairpins was even obtained at 80 mum in strands. However, this did not prevent an efficient conversion from hairpin to duplex in the presence of salts. Kinetic considerations are in favor of duplex formation from intermediates involving hairpins engaged in cruciform dimers rather than from free strands. The very first step of formation of such a cruciform intermediate could be trapped in a crystal structure. This mechanism might be significant for the dynamics of small RNAs beyond the strict field of HIV-1.}, note = {0021-9258 (Print) Journal Article}, keywords = {Base Sequence Binding Sites Crystallography, Drug Electrophoresis Genome, ENNIFAR, Fluorescence Spectrophotometry Temperature Thermodynamics Time Factors Ultraviolet Rays Virus Replication, MARQUET, Non-U.S. Gov't Spectrometry, PAILLART, Unité ARN, Viral HIV-1/*chemistry Kinetics Molecular Sequence Data *Nucleic Acid Conformation Protein Binding RNA/chemistry RNA, Viral/*chemistry Research Support, X-Ray Dimerization Dose-Response Relationship}, pubstate = {published}, tppubtype = {article} } @article{, title = {8-vinyl-deoxyadenosine, an alternative fluorescent nucleoside analog to 2'-deoxyribosyl-2-aminopurine with improved properties}, author = {N Ben Gaied and N Glasser and N Ramalanjaona and H Beltz and P Wolff and R Marquet and A Burger and Y Mely}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15718302}, isbn = {15718302}, year = {2005}, date = {2005-01-01}, journal = {Nucleic Acids Res}, volume = {33}, number = {3}, pages = {1031-1039}, abstract = {We report here the synthesis and the spectroscopic characterization of 8-vinyl-deoxyadenosine (8vdA), a new fluorescent analog of deoxyadenosine. 8vdA was found to absorb and emit in the same wavelength range as 2'-deoxyribosyl-2-aminopurine (2AP), the most frequently used fluorescent nucleoside analog. Though the quantum yield of 8vdA is similar to that of 2AP, its molar absorption coefficient is about twice, enabling a more sensitive detection. Moreover, the fluorescence of 8vdA was found to be sensitive to temperature and solvent but not to pH (around neutrality) or coupling to phosphate groups. Though 8vdA is base sensitive and susceptible to depurination, the corresponding phosphoramidite was successfully prepared and incorporated in oligonucleotides of the type d(CGT TTT XNX TTT TGC) where N = 8vdA and X = A, T or C. The 8vdA-labeled oligonucleotides gave more stable duplexes than the corresponding 2AP-labeled sequences when X = A or T, indicating that 8vdA is less perturbing than 2AP and probably adopts an anti conformation to preserve the Watson-Crick H-bonding. In addition, the quantum yield of 8vdA is significantly higher than 2AP in all tested oligonucleotides in both their single strand and duplex states. The steady-state and time-resolved fluorescence parameters of 8vdA and 2AP were found to depend similarly on the nature of their flanking residues and on base pairing, suggesting that their photophysics are governed by similar mechanisms. Taken together, our data suggest that 8vdA is a non perturbing nucleoside analog that may be used with improved sensitivity for the same applications as 2AP.}, note = {1362-4962 Journal Article}, keywords = {ENNIFAR, MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of a Folding Intermediate Reveals the Interplay Between Core and Peripheral Elements in RNA Folding}, author = {N J Baird and E Westhof and H Qin and T Pan and T R Sosnick}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=16115647}, isbn = {16115647}, year = {2005}, date = {2005-01-01}, journal = {J Mol Biol}, volume = {352}, number = {3}, pages = {712-722}, abstract = {Though the molecular architecture of many native RNA structures has been characterized, the structures of folding intermediates are poorly defined. Here, we present a nucleotide-level model of a highly structured equilibrium folding intermediate of the specificity domain of the Bacillus subtilis RNase P RNA, obtained using chemical and nuclease mapping, circular dichroism spectroscopy, small-angle X-ray scattering and molecular modeling. The crystal structure indicates that the 154 nucleotide specificity domain is composed of several secondary and tertiary structural modules. The structure of the intermediate contains modules composed of secondary structures and short-range tertiary interactions, implying a sequential order of tertiary structure formation during folding. The intermediate lacks the native core and several long-range interactions among peripheral regions, such as a GAAA tetraloop and its receptor. Folding to the native structure requires the local rearrangement of a T-loop in the core in concert with the formation of the GAAA tetraloop-receptor interaction. The interplay of core and peripheral structure formation rationalizes the high degree of cooperativity observed in the folding transition leading to the native structure.}, note = {0022-2836 Journal article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Molecular Dynamic Simulations of RNA Systems}, author = {P Auffinger and A Vaiana}, editor = {R K Hartmann and A Bindereif and A Schon and E Westhof}, url = {http://onlinelibrary.wiley.com/book/10.1002/9783527619504}, doi = {10.1002/9783527619504.ch34}, year = {2005}, date = {2005-01-01}, booktitle = {Handbook of RNA Biochemistry}, pages = {560-576}, publisher = {Wiley-Vch Verlag, Weinheim}, keywords = {Simulations RNA systems Advances Solvation Backbone, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {Hydrogens in RNA as visualized by molecular dynamics simulations}, author = {P Auffinger and A Vaiana}, editor = {N Niimura and H Mizuno and J Helliwel and E Westhof}, url = {http://www-ibmc.u-strasbg.fr/upr9002/westhof/PDF/r2005_Pauffinger_Niimura.pdf}, year = {2005}, date = {2005-01-01}, booktitle = {Hydrogen- and Hydration- Sensitive Structural Biology}, pages = {227-234}, publisher = {KubaPro Co, Ltd}, abstract = {Following several methodological developments, molecular dynamics simulations are now able to reproduce essential features of the solvation shell of biological molecules deduced from X-ray crystallography. Here we how molecular dynamics simulations can complement experimental data by providing clues about the position and orientation of the mobile hydrogen atoms found in RNA systems, namely those belonging to the 2メ-hydroxyl groups and to the water molecules.}, keywords = {Unité ARN, WESTHOF Nucleic acids X-ray crystallography Neutron crystallography Solvation Hydration}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Transfer RNA recognition by class I lysyl-tRNA synthetase from the Lyme disease pathogen Borrelia burgdorferi}, author = {A Ambrogelly and M Frugier and M Ibba and D Soll and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15862301}, isbn = {15862301}, year = {2005}, date = {2005-01-01}, journal = {FEBS Lett}, volume = {579}, number = {12}, pages = {2629-2634}, abstract = {Borrelia burgdorferi and other spirochetes contain a class I lysyl-tRNA synthetase (LysRS), in contrast to most eubacteria that have a canonical class II LysRS. We analyzed tRNA(Lys) recognition by B. burgdorferi LysRS, using two complementary approaches. First, the nucleotides of B. burgdorferi tRNA(Lys) in contact with B. burgdorferi LysRS were determined by enzymatic footprinting experiments. Second, the kinetic parameters for a series of variants of the B. burgdorferi tRNA(Lys) were then determined during aminoacylation by B. burgdorferi LysRS. The identity elements were found to be mostly located in the anticodon and in the acceptor stem. Transplantation of the identified identity elements into the Escherichia coli tRNA(Asp) scaffold endowed lysylation activity on the resulting chimera, indicating that a functional B. burgdorferi lysine tRNA identity set had been determined.}, note = {0014-5793 Journal Article}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{dostert_jak-stat_2005, title = {The Jak-STAT signaling pathway is required but not sufficient for the antiviral response of drosophila}, author = {Catherine Dostert and Emmanuelle Jouanguy and Phil Irving and Laurent Troxler and Delphine Galiana-Arnoux and Charles Hetru and Jules A Hoffmann and Jean-Luc Imler}, doi = {10.1038/ni1237}, issn = {1529-2908}, year = {2005}, date = {2005-01-01}, journal = {Nature Immunology}, volume = {6}, number = {9}, pages = {946--953}, abstract = {The response of drosophila to bacterial and fungal infections involves two signaling pathways, Toll and Imd, which both activate members of the transcription factor NF-kappaB family. Here we have studied the global transcriptional response of flies to infection with drosophila C virus. Viral infection induced a set of genes distinct from those regulated by the Toll or Imd pathways and triggered a signal transducer and activator of transcription (STAT) DNA-binding activity. Genetic experiments showed that the Jak kinase Hopscotch was involved in the control of the viral load in infected flies and was required but not sufficient for the induction of some virus-regulated genes. Our results indicate that in addition to Toll and Imd, a third, evolutionary conserved innate immunity pathway functions in drosophila and counters viral infection.}, keywords = {Animals, bioinformatic, DNA-Binding Proteins, Genetic, Genetically Modified, hoffmann, imler, Insect Viruses, Janus Kinase 1, M3i, Male, Oligonucleotide Array Sequence Analysis, Promoter Regions, Protein-Tyrosine Kinases, Signal Transduction, STAT1 Transcription Factor, Trans-Activators}, pubstate = {published}, tppubtype = {article} } @article{martinelli_evolution_2005, title = {Evolution and integration of innate immune systems from fruit flies to man: lessons and questions}, author = {Cosimo Martinelli and Jean-Marc Reichhart}, doi = {10.1179/096805105X37411}, issn = {0968-0519}, year = {2005}, date = {2005-01-01}, journal = {J. Endotoxin Res.}, volume = {11}, number = {4}, pages = {243--248}, abstract = {Despite broad differences in morphology, ecology and behavior, the fruit fly Drosophila melanogaster and humans show a remarkably high degree of conservation for many molecular, cellular, and developmental aspects of their biology. During the last decade, similarities have also been discovered in some of the mechanisms regulating their innate immune system. These parallels regard mainly the Toll-like receptor family and the intracellular signaling pathways involved in the control of the immune response. However, if the overall similarities are important, the detailed pathogen recognition mechanisms differ significantly between fly and humans, highlighting a complicated evolutionary history of the metazoan innate defenses. In this review, we will discuss the main similarities and differences between the two types of organisms. We hope that this current knowledge will be used as a starting point for a more comprehensive view of innate immunity within the broad variety of metazoan phyla.}, keywords = {Animals, Biological Evolution, Cell Surface, Forecasting, Humans, Immunity, Immunological, Innate, M3i, Membrane Glycoproteins, Models, Receptors, reichhart, Signal Transduction, Toll-Like Receptor 5, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{weber_ligand-receptor_2005, title = {Ligand-receptor and receptor-receptor interactions act in concert to activate signaling in the Drosophila toll pathway}, author = {Alexander N R Weber and Martin C Moncrieffe and Monique Gangloff and Jean-Luc Imler and Nicholas J Gay}, doi = {10.1074/jbc.M502074200}, issn = {0021-9258}, year = {2005}, date = {2005-01-01}, journal = {The Journal of Biological Chemistry}, volume = {280}, number = {24}, pages = {22793--22799}, abstract = {In Drosophila, the signaling pathway mediated by the Toll receptor is critical for the establishment of embryonic dorso-ventral pattern and for innate immune responses to bacterial and fungal pathogens. Toll is activated by high affinity binding of the cytokine Spätzle, a dimeric ligand of the cystine knot family. In vertebrates, a related family of Toll-like receptors play a critical role in innate immune responses. Despite the importance of this family of receptors, little is known about the biochemical events that lead to receptor activation and signaling. Here, we show that Spätzle binds to the N-terminal region of Toll and, using biophysical methods, that the binding is complex. The two binding events that cause formation of the cross-linked complex are non-equivalent: the first Toll ectodomain binds Spätzle with an affinity 3-fold higher than the second molecule suggesting that pathway activation involves negative cooperativity. We further show that the Toll ectodomains are able to form low affinity dimers in solution and that juxtamembrane sequences of Toll are critical for the activation or derepression of the pathway. These results, taken together, suggest a mechanism of signal transduction that requires both ligand-receptor and receptor-receptor interactions.}, keywords = {Amino Acid, Animals, Biophysical Phenomena, Biophysics, Body Patterning, Calorimetry, Cell Line, Cell Surface, Cross-Linking Reagents, Cytokines, dimerization, Electrophoresis, Humans, imler, ligands, Luciferases, M3i, Membrane Glycoproteins, Polyacrylamide Gel, Protein Binding, Protein Structure, Receptors, Recombinant Proteins, Sequence Homology, Signal Transduction, Tertiary, Time Factors, Toll-Like Receptors, Ultracentrifugation}, pubstate = {published}, tppubtype = {article} } @article{imler_antimicrobial_2005, title = {Antimicrobial peptides in Drosophila: structures, activities and gene regulation}, author = {Jean-Luc Imler and Philippe Bulet}, doi = {10.1159/000086648}, issn = {1660-2242}, year = {2005}, date = {2005-01-01}, journal = {Chemical Immunology and Allergy}, volume = {86}, pages = {1--21}, abstract = {The production of antimicrobial peptides (AMPs) is an important aspect of host-defence in multicellular organisms. Biochemical analysis of the hemolymph of the fruit-fly Drosophila melanogaster and other Diptera has led to the discovery of eight classes of AMPs. These peptides can be grouped into three families based on their main biological targets, gram-positive bacteria (defensin), gram-negative bacteria (cecropins, drosocin, attacins, diptericin, MPAC), or fungi (drosomycin, metchnikowin). Drosophila AMPs are synthesized by the fat body in response to infection, and secreted into the blood. Most of them can also be induced in surface epithelia in a tissue-specific manner. Finally, some of them are constitutively expressed in defined tissues, such as the salivary glands or the reproductive tract. We review here the structures and activities of these AMPs, as well as the signalling cascades, which lead to their induction upon detection of infectious non-self.}, keywords = {Animals, Antimicrobial Cationic Peptides, Defensins, Gene Expression Regulation, Genes, Glycopeptides, imler, Immunity, Innate, Insect, Insect Proteins, M3i, Molecular Structure, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reverse transcriptase and integrase of the Saccharomyces cerevisiae Ty1 element}, author = { F. X. Wilhelm and M. Wilhelm and A. Gabriel}, year = {2005}, date = {2005-01-01}, journal = {Cytogenet Genome Res}, volume = {110}, number = {1-4}, pages = {269-87}, abstract = {Integrase (IN) and reverse transcriptase (RT) play a central role in transposition of retroelements. The mechanism of integration by IN and the steps of the replication process mediated by RT are briefly described here. Recently, active recombinant forms of Ty1 IN and RT have been obtained. This has allowed a more detailed understanding of their biochemical and structural properties and has made possible combined in vitro and in vivo analyses of their functions. A focus of this review is to discuss some of the results obtained thus far with these two recombinant proteins and to propose future directions.}, note = {1424-859x Journal Article}, keywords = {MARQUET}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of integrase in reverse transcription of the Saccharomyces cerevisiae retrotransposon Ty1}, author = { M. Wilhelm and F. X. Wilhelm}, year = {2005}, date = {2005-01-01}, journal = {Eukaryot Cell}, volume = {4}, number = {6}, pages = {1057-65}, abstract = {Reverse transcriptase (RT) with its associated RNase H (RH) domain and integrase (IN) are key enzymes encoded by retroviruses and retrotransposons. Several studies have implied a functional role of the interaction between IN and RT during the replication of retroviral and retrotransposon genomes. In this study, IN deletion mutants were used to investigate the role of IN on the RT activity of the yeast Saccharomyces cerevisiae retrotransposon Ty1. We have identified two domains of Ty1 integrase which have effects on RT activity in vivo. The deletion of a domain spanning amino acid residues 233 to 520 of IN increases the exogenous specific activity of RT up to 20-fold, whereas the removal of a region rich in acidic amino acid residues between residues 521 and 607 decreases its activity. The last result complements our observation that an active recombinant RT protein can be obtained if a small acidic tail mimicking the acidic domain of IN is fused to the RT-RH domain. We suggest that interaction between these acidic amino acid residues of IN and a basic region of RT could be critical for the correct folding of RT and for the formation of an active conformation of the enzyme.}, note = {1535-9778 Journal Article}, keywords = {MARQUET}, pubstate = {published}, tppubtype = {article} } @article{fournel_c3-symmetric_2005b, title = {C3-symmetric peptide scaffolds are functional mimetics of trimeric CD40L}, author = {Sylvie Fournel and Sébastien Wieckowski and Weimin Sun and Nathalie Trouche and Hélène Dumortier and Alberto Bianco and Olivier Chaloin and Mohammed Habib and Jean-Christophe Peter and Pascal Schneider and Bernard Vray and René E Toes and Rienk Offringa and Cornelis J M Melief and Johan Hoebeke and Gilles Guichard}, doi = {10.1038/nchembio746}, issn = {1552-4450}, year = {2005}, date = {2005-01-01}, journal = {Nature Chemical Biology}, volume = {1}, number = {7}, pages = {377--382}, abstract = {Interaction between CD40, a member of the tumor necrosis factor receptor (TNFR) superfamily, and its ligand CD40L, a 39-kDa glycoprotein, is essential for the development of humoral and cellular immune responses. Selective blockade or activation of this pathway provides the ground for the development of new treatments against immunologically based diseases and malignancies. Like other members of the TNF superfamily, CD40L monomers self-assemble around a threefold symmetry axis to form noncovalent homotrimers that can each bind three receptor molecules. Here, we report on the structure-based design of small synthetic molecules with C3 symmetry that can mimic CD40L homotrimers. These molecules interact with CD40, compete with the binding of CD40L to CD40, and reproduce, to a certain extent, the functional properties of the much larger homotrimeric soluble CD40L. Architectures based on rigid C3-symmetric cores may thus represent a general approach to mimicking homotrimers of the TNF superfamily.}, keywords = {Animals, Apoptosis, Biological, CD40 Antigens, CD40 Ligand, Cell Line, Humans, I2CT, Inbred BALB C, Mice, Models, Molecular Mimicry, Molecular Structure, Peptides, Protein Conformation, Protein Structure, Quaternary, Structure-Activity Relationship, Team-Bianco, Time Factors, tumor}, pubstate = {published}, tppubtype = {article} } @article{bianco_biomedical_2005, title = {Biomedical applications of functionalised carbon nanotubes}, author = {Alberto Bianco and Kostas Kostarelos and Charalambos D Partidos and Maurizio Prato}, doi = {10.1039/b410943k}, issn = {1359-7345}, year = {2005}, date = {2005-01-01}, journal = {Chemical Communications (Cambridge, England)}, number = {5}, pages = {571--577}, abstract = {The organic functionalisation of carbon nanotubes can improve substantially their solubility and biocompatibility profile; as a consequence, their manipulation and integration into biological systems has become possible so that functionalised carbon nanotubes hold currently strong promise as novel systems for the delivery of drugs, antigens and genes.}, keywords = {Antigens, carbon, Chemical, Drug Delivery Systems, Gene Transfer Techniques, Humans, I2CT, Models, Molecular Structure, nanotechnology, Nanotubes, Team-Bianco, Vaccines}, pubstate = {published}, tppubtype = {article} } @article{bianco_cationic_2005, title = {Cationic carbon nanotubes bind to CpG oligodeoxynucleotides and enhance their immunostimulatory properties}, author = {Alberto Bianco and Johan Hoebeke and Sylvie Godefroy and Olivier Chaloin and Davide Pantarotto and Jean-Paul Briand and Sylviane Muller and Maurizio Prato and Charalambos D Partidos}, doi = {10.1021/ja044293y}, issn = {0002-7863}, year = {2005}, date = {2005-01-01}, journal = {Journal of the American Chemical Society}, volume = {127}, number = {1}, pages = {58--59}, abstract = {Functionalized cationic carbon nanotubes are able to form a stable complex with CpG ODN based on charge interaction and to increase the immunostimulatory activity of CpG motifs.}, keywords = {Adjuvants, Animals, carbon, Cations, CpG Islands, I2CT, Immunologic, Interferon-gamma, Interleukin-6, Kinetics, Lymphocytes, Mice, Nanotubes, oligonucleotides, Surface Plasmon Resonance, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{van_mierlo_activation_2004, title = {Activation of dendritic cells that cross-present tumor-derived antigen licenses CD8+ CTL to cause tumor eradication}, author = {Geertje J D van Mierlo and Zita F H M Boonman and Hélène M H Dumortier and Annemieke Th den Boer and Marieke F Fransen and Jan Nouta and Ellen I H van der Voort and Rienk Offringa and René E M Toes and Cornelis J M Melief}, doi = {10.4049/jimmunol.173.11.6753}, issn = {0022-1767}, year = {2004}, date = {2004-12-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {173}, number = {11}, pages = {6753--6759}, abstract = {The fate of naive CD8(+) T cells is determined by the environment in which they encounter MHC class I presented peptide Ags. The manner in which tumor Ags are presented is a longstanding matter of debate. Ag presentation might be mediated by tumor cells in tumor draining lymph nodes or via cross-presentation by professional APC. Either pathway is insufficient to elicit protective antitumor immunity. We now demonstrate using a syngeneic mouse tumor model, expressing an Ag derived from the early region 1A of human adenovirus type 5, that the inadequate nature of the antitumor CTL response is not due to direct Ag presentation by the tumor cells, but results from presentation of tumor-derived Ag by nonactivated CD11c(+) APC. Although this event results in division of naive CTL in tumor draining lymph nodes, it does not establish a productive immune response. Treatment of tumor-bearing mice with dendritic cell-stimulating agonistic anti-CD40 mAb resulted in systemic efflux of CTL with robust effector function capable to eradicate established tumors. For efficacy of anti-CD40 treatment, CD40 ligation of host APC is required because adoptive transfer of CD40-proficient tumor-specific TCR transgenic CTL into CD40-deficient tumor-bearing mice did not lead to productive antitumor immunity after CD40 triggering in vivo. CpG and detoxified LPS (MPL) acted similarly as agonistic anti-CD40 mAb with respect to CD8(+) CTL efflux and tumor eradication. Together these results indicate that dendritic cells, depending on their activation state, orchestrate the outcome of CTL-mediated immunity against tumors, leading either to an ineffective immune response or potent antitumor immunity.}, keywords = {Adenovirus E1A Proteins, Animals, Antibodies, Antigen-Presenting Cells, Antigens, CD11c Antigen, CD40 Antigens, Cross-Priming, Cultured, Cytotoxic, Cytotoxicity, Dendritic Cells, Dumortier, Epitopes, Experimental, I2CT, Immunologic, Inbred C57BL, Injections, Intralesional, Intravenous, Knockout, Male, Mice, Monoclonal, Neoplasms, T-Lymphocyte, T-Lymphocytes, Team-Dumortier, transgenic, tumor, Tumor Cells, Viral}, pubstate = {published}, tppubtype = {article} } @article{bischoff_function_2004, title = {Function of the drosophila pattern-recognition receptor PGRP-SD in the detection of Gram-positive bacteria}, author = {Vincent Bischoff and Cécile Vignal and Ivo G Boneca and Tatiana Michel and Jules A Hoffmann and Julien Royet}, doi = {10.1038/ni1123}, issn = {1529-2908}, year = {2004}, date = {2004-11-01}, journal = {Nat. Immunol.}, volume = {5}, number = {11}, pages = {1175--1180}, abstract = {The activation of an immune response requires recognition of microorganisms by host receptors. In drosophila, detection of Gram-positive bacteria is mediated by cooperation between the peptidoglycan-recognition protein-SA (PGRP-SA) and Gram-negative binding protein 1 (GNBP1) proteins. Here we show that some Gram-positive bacterial species activate an immune response in a PGRP-SA- and GNBP1-independent manner, indicating that alternative receptors exist. Consistent with this, we noted that PGRP-SD mutants were susceptible to some Gram-positive bacteria and that a loss-of-function mutation in PGRP-SD severely exacerbated the PGRP-SA and GNBP1 mutant phenotypes. These data indicate that PGRP-SD can function as a receptor for Gram-positive bacteria and shows partial redundancy with the PGRP-SA-GNBP1 complex.}, keywords = {Animals, Carrier Proteins, Cell Surface, Gram-Positive Bacteria, Gram-Positive Bacterial Infections, hoffmann, M3i, Mutation, Mycoses, Receptors, Staphylococcus aureus, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{bianco_carbon_2004, title = {Carbon nanotubes for the delivery of therapeutic molecules}, author = {Alberto Bianco}, doi = {10.1517/17425247.1.1.57}, issn = {1742-5247}, year = {2004}, date = {2004-11-01}, journal = {Expert Opinion on Drug Delivery}, volume = {1}, number = {1}, pages = {57--65}, abstract = {Functionalised carbon nanotubes (f-CNTs) are emerging as new tools in the field of nanobiotechnology and nanomedicine. This is because they can be easily manipulated and modified by encapsulation with biopolymers or by covalent linking of solubilising groups to the external walls and tips. The possibility of incorporating f-CNTs into biological systems has opened the way to the exploration of their potential applications in biology and medicinal chemistry. Within the different fields of applications (i.e., biosensors, composite materials, molecular electronics), one use of CNTs is as new carrier systems for the delivery of therapeutic molecules. Research discussed in this review is focused on recent advances in the development of CNT technology for the delivery of drugs, antigens and genes.}, keywords = {carbon, Drug Carriers, Gene Transfer Techniques, I2CT, Nanotubes, Pharmaceutical Preparations, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{monneaux_intramolecular_2004, title = {Intramolecular Ŧ cell spreading in unprimed MRL/lpr mice: importance of the U1-70k protein sequence 131-151}, author = {Fanny Monneaux and Véronique Parietti and Jean-Paul Briand and Sylviane Muller}, doi = {10.1002/art.20510}, issn = {0004-3591}, year = {2004}, date = {2004-10-01}, journal = {Arthritis and Rheumatism}, volume = {50}, number = {10}, pages = {3232--3238}, abstract = {OBJECTIVE: To analyze spontaneous T cell spreading against determinants of the U1-70K protein in young autoimmune MRL/lpr lupus mice, in comparison with the T cell spreading occurring in normal BALB/c mice immunized with peptide 131-151 of this protein. METHODS: Peripheral blood lymphocytes (PBLs) from both unprimed MRL/lpr mice and immunized BALB/c mice were tested for their ability to proliferate ex vivo in response to 18 overlapping peptides of the U1-70K spliceosomal protein, using assays for lymphocyte proliferation and secretion of interleukin-2. RESULTS: The proliferative response to peptides of the U1-70K protein evolved rapidly in MRL/lpr mice tested at different ages. At least 5 peptides were recognized by PBLs from 8-week-old autoimmune mice, whereas a different peptide was recognized by PBLs from MRL/lpr mice at 12 weeks of age. At 15 weeks, the proliferative response was weak or negative when assessed with any of the test peptides. At least 2 major peptides recognized by MRL/lpr PBLs were also recognized by PBLs generated in the BALB/c mice primed with peptide 131-151. We further demonstrated that, in preautoimmune MRL/lpr mice, repeated administration of phosphorylated peptide 131-151 (called P140), which was shown previously to be protective, transiently abolished T cell intramolecular spreading to other regions of the 70K protein. CONCLUSION: This is the first study to demonstrate that intramolecular T cell spreading effectively occurs in MRL/lpr mice with lupus, and that region 131-151 is important in the cascade of events observed in the murine lupus response. This sequence might originate a mechanism of tolerance spreading that leads to the beneficial effect observed in MRL/lpr mice after treatment with the phosphorylated peptide 131-151.}, keywords = {Animals, Cell Division, Female, I2CT, Immunization, Inbred BALB C, Inbred MRL lpr, Lupus Erythematosus, Lymphocytes, Mice, Monneaux, Peptides, Phosphorylation, Ribonucleoprotein, Systemic, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{levy_peptidomic_2004, title = {Peptidomic and proteomic analyses of the systemic immune response of Drosophila}, author = {Francine Levy and David Rabel and Maurice Charlet and Philippe Bulet and Jules A Hoffmann and Laurence Ehret-Sabatier}, doi = {10.1016/j.biochi.2004.07.007}, issn = {0300-9084}, year = {2004}, date = {2004-10-01}, journal = {Biochimie}, volume = {86}, number = {9-10}, pages = {607--616}, abstract = {Insects have developed an efficient host defense against microorganisms, which involves humoral and cellular mechanisms. Numerous data highlight similarities between defense responses of insects and innate immunity of mammals. The fruit fly, Drosophila melanogaster, is a favorable model system for the analysis of the first line defense against microorganisms. Taking advantages of improvements in mass spectrometry (MS), two-dimensional (2D) gel electrophoresis and bioinformatics, differential analyses of blood content (hemolymph) from immune-challenged versus control Drosophila were performed. Two strategies were developed: (i) peptidomic analyses through matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS and high performance liquid chromatography for molecules below 15 kDa, and (ii) proteomic studies based on 2D gel electrophoresis, MALDI-TOF fingerprinting and database searches, for compounds of greater molecular masses. The peptidomic strategy led to the detection of a large number of peptides induced in the hemolymph of challenged flies as compared to controls. Of these, 28 were characterized, amongst which were antimicrobial peptides. The 2D gel electrophoresis strategy led to the detection of 70 spots differentially regulated by at least fivefold after microbial infection. This approach yielded the identity of a series of proteins that were related to the Drosophila immune response, such as proteases, protease inhibitors, prophenoloxydase-activating enzymes, serpins and a Gram-negative binding protein-like protein. This strategy also brought to light new candidates with a potential function in the immune response (odorant-binding protein, peptidylglycine alpha-hydroxylating monooxygenase and transferrin). Interestingly, several molecules resulting from the cleavage of proteins were detected after a fungal infection. Together, peptidomic and proteomic analyses represent new tools to characterize molecules involved in the innate immune reactions of Drosophila.}, keywords = {Animals, hoffmann, Immunity, M3i, proteomics}, pubstate = {published}, tppubtype = {article} } @article{poschalko_deuss_2004, title = {DEUSS: A Perdeuterated Poly(oxyethylene)-Based Resin for Improving HRMAS NMR Studies of Solid-Supported Molecules}, author = {Alexander Poschalko and Nathalie Lancelot and Julien Marin and Virginie Larras and David Limal and Karim Elbayed and Jesus Raya and Martial Piotto and Jean-Paul Briand and Gilles Guichard and Alberto Bianco}, url = {https://chemistry-europe.onlinelibrary.wiley.com/doi/abs/10.1002/chem.200400373}, doi = {10.1002/chem.200400373}, issn = {0947-6539}, year = {2004}, date = {2004-09-01}, urldate = {2020-03-31}, journal = {Chemistry – A European Journal}, volume = {10}, number = {18}, pages = {4532--4537}, abstract = {Abstract A novel resin called DEUSS (perdeuterated poly(oxyethylene)-based solid support) has been prepared by anionic polymerization of deuterated [D4]ethylene oxide, followed by cross-linking with deuterated epichlorohydrin. DEUSS can be suspended in a wide range of solvents including organic and aqueous solutions, in which it displays a high swelling capacity. As measured by proton HRMAS of the swollen polymer, the signal intensity of the oxyethylene protons is reduced by a factor of 110 relative to the corresponding nondeuterated poly(oxyethylene)poly(oxypropylene) (POEPOP) resin, thus facilitating detailed HRMAS NMR studies of covalently linked molecules. This 1H NMR invisible matrix was used for the solid-phase synthesis of peptides, oligoureas, and a series of amides as well as their characterization by HRMAS NMR spectroscopy. On-bead NMR spectra of high quality and with resolution comparable to that of liquid samples were obtained and readily interpreted. The complete absence of the parasite resin signals will be of great advantage, for example, for the optimization of multistep solid-phase stereoselective reactions, and for the conformational study of resin-bound molecules in a large variety of solvents.}, keywords = {combinatorial chemistry, I2CT, NMR spectroscopy, oligoureas, Peptides, poly(oxyethylene), Resins, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{MJ2004, title = {Immune responses and parasite transmission in blood-feeding insects}, author = {M J Lehane and S Aksoy and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/15324734}, year = {2004}, date = {2004-09-01}, journal = {Trends Parasitol.}, volume = {20}, number = {4}, pages = {433-9}, abstract = {The detailed model of insect immunity being built for Drosophila, allied to mass sequencing programs for blood-feeding insects, has led to advances in our understanding of the interaction between pathogens and insect vectors. An outline of insect immunity is given here based on the Drosophila studies, which is used as a framework to discuss recent work on Plasmodium-mosquito and Trypanosoma-tsetse interactions.}, keywords = {parasite transmission}, pubstate = {published}, tppubtype = {article} } @article{pantarotto_synthesis_2004, title = {Synthesis and biological properties of fullerene-containing amino acids and peptides}, author = {Davide Pantarotto and Nikos Tagmatarchis and Alberto Bianco and Maurizio Prato}, issn = {1389-5575}, year = {2004}, date = {2004-09-01}, journal = {Mini Reviews in Medicinal Chemistry}, volume = {4}, number = {7}, pages = {805--814}, abstract = {Organofullerene derivatives have shown a great potential in a wide variety of biological activities such as DNA photocleavage, HIV-protease inhibition, neuroprotection and apoptosis. Among the plethora of functionalized organofullerenes that have been synthesized, fullerene-based amino acids are particularly appealing for structural studies and biological applications. When the fullerene-framework is incorporated into peptides, its original properties can be substantially modified. In addition, the water-solubility of the fullerene derivatives is enhanced, which makes such molecules amenable to biological studies. In this review, recent advances in the growing field of medicinal chemistry of fullerene derivatives will be discussed. Emphasis will be given to the synthesis of the biggest unnatural amino acid 3,4-fulleroproline (Fpr) and its derivatives. For example, Fpr derivatives have been found to interact with different hydrolytic enzymes and selectively discriminate between rationally designed peptides. Fullerene-based peptides have been found to substantially activate enzymes involved in the oxidative deamination of biogenic amines. In addition, their membranotropic properties and effects on the structure and permeability of the lipid bilayer of phosphatidylcholine liposomes as well as the transmembrane transport of bivalent metal ions have been studied. Finally, applications in medicinal chemistry of such types of amino acids and peptides will be highlighted.}, keywords = {Amino Acids, Animals, Fullerenes, Humans, I2CT, Molecular Structure, Peptides, Solubility, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{S2004, title = {Curing mosquitoes to control malaria ?}, author = {Stéphanie A Blandin and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/15361335}, year = {2004}, date = {2004-08-02}, journal = {Med Sci (Paris).}, volume = {20}, number = {8-9}, pages = {740-2}, keywords = {blandin, M3i, population control}, pubstate = {published}, tppubtype = {article} } @article{EA2004, title = {Immune responses in Anopheles gambiae}, author = {Elena A Levashina}, year = {2004}, date = {2004-07-01}, journal = {Insect Biochem Mol Biol.}, volume = {34}, number = {7}, pages = {673-8}, abstract = {Transmission of human malaria requires a successful development of Plasmodium parasites in anopheline mosquitoes. Insects have developed efficient immune responses to oppose microbial and eukaryotic invaders. The completion of the sequencing of the Anopheles genome provides a wealth of information on putative immune genes that are homologous to components of the Drosophila and mammalian immune systems. In this review, we will summarize our present knowledge of immune responses in the mosquito Anopheles gambiae and attempt a comparative analysis of insect immune systems.}, keywords = {insect immune system}, pubstate = {published}, tppubtype = {article} } @article{lancelot_measurement_2004, title = {Measurement of scaled residual dipolar couplings in proteins using variable-angle sample spinning}, author = {Nathalie Lancelot and Karim Elbayed and Alberto Bianco and Martial Piotto}, doi = {10.1023/B:JNMR.0000032548.60663.1f}, issn = {0925-2738}, year = {2004}, date = {2004-07-01}, journal = {Journal of biomolecular NMR}, volume = {29}, number = {3}, pages = {259--269}, abstract = {NMR spectra of ubiquitin in the presence of bicelles at a concentration of 25% w/v have been recorded under sample spinning conditions for different angles of rotation. For an axis of rotation equal to the magic angle, the (1)H/(15)N HSQC recorded without any (1)H decoupling in the indirect dimension corresponds to the classical spectrum obtained on a protein in an isotropic solution and allows the measurement of scalar J-couplings (1) J (NH). For an angle of rotation smaller than the magic angle, the bicelles orient with their normal perpendicular to the spinning axis, whereas for an angle of rotation greater than the magic angle the bicelles orient with their normal along the spinning axis. This bicelle alignment creates anisotropic conditions that give rise to the observation of residual dipolar couplings in ubiquitin. The magnitude of these dipolar couplings depends directly on the angle that the rotor makes with the main magnetic field. By changing this angle in a controlled manner, residual dipolar couplings can be either scaled up or down thus offering the possibility to study simultaneously a wide range of dipolar couplings in the same sample.}, keywords = {anisotropy, I2CT, Magnetic Resonance Spectroscopy, Magnetics, Models, Phospholipid Ethers, Proteins, Statistical, Team-Bianco, Temperature, ubiquitin}, pubstate = {published}, tppubtype = {article} } @article{irving_is_2004, title = {Is innate enough? The innate immune response in Drosophila}, author = {Phil Irving and Laurent Troxler and Charles Hetru}, issn = {1631-0691}, year = {2004}, date = {2004-06-01}, journal = {C. R. Biol.}, volume = {327}, number = {6}, pages = {557--570}, abstract = {In recent years, the innate immune system has emerged from the shadow of adaptive immune responses as a major area of research in its own right. One of the most significant model systems that has been used to investigate this phenomenon has been the fruit fly, Drosophila melanogaster. Exploration of the differential immune response presented by Drosophila led to the discovery of important signalling events and transduction pathways, which were thereafter shown to be specific for the type of infecting pathogen. These factors and pathways were subsequently found to have homologues in many other organisms, including those with adaptive immune responses. In light of the present status of studies in innate immunity, this review describes the current state of understanding of the Drosophila immune response.}, keywords = {Animals, bioinformatic, Immunity, Innate, M3i, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{EA2004b, title = {Bacterial alpha2-macroglobulins: colonization factors acquired by horizontal gene transfer from the metazoan genome ?}, author = {A Budd and Stéphanie A Blandin and Elena A Levashina and T J Gibson}, url = {http://www.ncbi.nlm.nih.gov/pubmed/15186489}, year = {2004}, date = {2004-05-26}, journal = {Genome Biol.}, volume = {5}, number = {6}, pages = {R38}, abstract = {BACKGROUND: Invasive bacteria are known to have captured and adapted eukaryotic host genes. They also readily acquire colonizing genes from other bacteria by horizontal gene transfer. Closely related species such as Helicobacter pylori and Helicobacter hepaticus, which exploit different host tissues, share almost none of their colonization genes. The protease inhibitor alpha2-macroglobulin provides a major metazoan defense against invasive bacteria, trapping attacking proteases required by parasites for successful invasion. RESULTS: Database searches with metazoan alpha2-macroglobulin sequences revealed homologous sequences in bacterial proteomes. The bacterial alpha2-macroglobulin phylogenetic distribution is patchy and violates the vertical descent model. Bacterial alpha2-macroglobulin genes are found in diverse clades, including purple bacteria (proteobacteria), fusobacteria, spirochetes, bacteroidetes, deinococcids, cyanobacteria, planctomycetes and thermotogae. Most bacterial species with bacterial alpha2-macroglobulin genes exploit higher eukaryotes (multicellular plants and animals) as hosts. Both pathogenically invasive and saprophytically colonizing species possess bacterial alpha2-macroglobulins, indicating that bacterial alpha2-macroglobulin is a colonization rather than a virulence factor. CONCLUSIONS: Metazoan alpha2-macroglobulins inhibit proteases of pathogens. The bacterial homologs may function in reverse to block host antimicrobial defenses. Alpha2-macroglobulin was probably acquired one or more times from metazoan hosts and has then spread widely through other colonizing bacterial species by more than 10 independent horizontal gene transfers. yfhM-like bacterial alpha2-macroglobulin genes are often found tightly linked with pbpC, encoding an atypical peptidoglycan transglycosylase, PBP1C, that does not function in vegetative peptidoglycan synthesis. We suggest that YfhM and PBP1C are coupled together as a periplasmic defense and repair system. Bacterial alpha2-macroglobulins might provide useful targets for enhancing vaccine efficacy in combating infections.}, keywords = {alpha2-macroglobulins, blandin, M3i}, pubstate = {published}, tppubtype = {article} } @article{leclerc_immune_2004, title = {The immune response of Drosophila melanogaster}, author = {Vincent Leclerc and Jean-Marc Reichhart}, issn = {0105-2896}, year = {2004}, date = {2004-04-01}, journal = {Immunol. Rev.}, volume = {198}, pages = {59--71}, abstract = {The response of the fruit fly Drosophila melanogaster to various microorganism infections relies on a multilayered defense. The epithelia constitute a first and efficient barrier. Innate immunity is activated when microorganisms succeed in entering the body cavity of the fly. Invading microorganisms are killed by the combined action of cellular and humoral processes. They are phagocytosed by specialized blood cells, surrounded by toxic melanin, or lysed by antibacterial peptides secreted into the hemolymph by fat body cells. During the last few years, research has focused on the mechanisms of microbial recognition by various pattern recognition receptors and of the subsequent induction of antimicrobial peptide expression. The cellular arm of the Drosophila innate immune system, which was somehow neglected, now constitutes the new frontier.}, keywords = {Animals, Cell Surface, Immunity, Immunological, Innate, M3i, Models, Receptors, reichhart, Signal Transduction, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{S2004b, title = {Thioester-containing proteins in insect immunity}, author = {Stéphanie A Blandin and Elena A Levashina}, year = {2004}, date = {2004-02-01}, journal = {Mol Immunol.}, volume = {40}, number = {12}, pages = {903-8}, abstract = {Here, we discuss the role of thioester-containing proteins in innate immune responses of insects. TEPs are represented by multi-member families both in the fruitfly, Drosophila melanogaster, and in the mosquito, Anopheles gambiae. Phylogenetic analysis of the family suggests that in these two dipteran species evolution of TEPs followed independent scenarios as a result of specific adaptation to distinct ecological environments. Research on these two relatively simple model systems, which lack adaptive immunity, may provide new insights into the evolutionary origins and functions of this important protein family.}, keywords = {blandin, M3i, TEP1}, pubstate = {published}, tppubtype = {article} } @article{, title = {On the use of low-frequency normal modes to enforce collective movements in refining macromolecular structural models}, author = { M. Delarue and P. Dumas}, year = {2004}, date = {2004-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {101}, number = {18}, pages = {6957-62}, abstract = {As more and more structures of macromolecular complexes get solved in different conditions, it has become apparent that flexibility is an inherent part of their biological function. Normal mode analysis using simplified models of proteins such as the elastic network model has proved very effective in showing that many of the structural transitions derived from a survey of the Protein Data Bank can be explained by just a few of the lowest-frequency normal modes. In this work, normal modes are used to carry out medium- or low-resolution structural refinement, enforcing collective and large-amplitude movements that are beyond the reach of existing methods. Refinement is carried out in reciprocal space with respect to the normal mode amplitudes, by using standard conjugate-gradient minimization. Several tests on synthetic diffraction data whose mode concentration follows the one of real movements observed in the Protein Data Bank have shown that the radius of convergence is larger than the one of rigid-body refinement. Tests with experimental diffraction data for the same protein in different environments also led to refined structural models showing drastic reduction of the rms deviation with the target model. Because the structural transition is described by very few parameters, over-fitting of real experimental data is easily detected by using a cross-validation test. The method has also been applied to the refinement of atomic models into molecular envelopes and could readily be used to fit large macromolecular complex rearrangements into cryo-electron microscopy-reconstructed images as well as small-angle x-ray scattering-derived envelopes.}, note = {0027-8424 Journal Article}, keywords = {*Models, Carrier, coli, Computer, Diffraction, DUMAS, Escherichia, Gov't, Molecular, Non-U.S., Proteins/*chemistry, Proteins/chemistry, Simulation, Support, X-Ray}, pubstate = {published}, tppubtype = {article} } @incollection{, title = {Detection and quantification of modified nucleotides in RNA using thin-layer chromatography}, author = { H. Grosjean and G. Keith and L. Droogmans}, editor = { J.M. Gott}, year = {2004}, date = {2004-01-01}, booktitle = {RNA Interference, Editing, and Modification: Methods and Protocols}, volume = {265}, pages = {357-91}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Identification of a modified nucleotide and its localization within an RNA molecule is a difficult task. Only direct sequencing of purified RNA molecules and high-performance liquid chromatography mass spectrometry analysis of purified RNA fragments allow determination of both the type and location of a given modified nucleotide within an RNA of 50-150 nt in length. The objective of this chapter is to describe in detail a few simple procedures that we have found particularly suited for the detection, localization, and quantification of modified nucleotides within an RNA of known sequence. The methods can also be used to reveal the enzymatic activity of a particular RNA-modifying enzyme in vitro or in vivo. The procedures are based on the use of radiolabeled RNA (with [32P], [14C], or [3H]) or [32P]-postlabeled oligonucleotides and two-dimensional thin-layer chromatography of labeled nucleotides on cellulose plates. This chapter provides useful maps of the migration characteristics of 70 modified nucleotides on thin-layer cellulose plates.}, keywords = {&, 5', acids, and, Base, Chromatography, Composition, Deoxyribonucleotides/chemistry/isolation, DNA/chemistry/genetics/isolation, Endoribonucleases, Gov't, Indicators, Isotope, KEITH, Labeling/methods, Layer/methods, Non-U.S., Nucleic, Oligoribonucleotides/chemistry/isolation, Peptide, purification, Radioisotopes, Reagents, Regions/chemistry, Ribonucleotides/*analysis, RNA/*genetics/isolation, Support, Thin, Untranslated}, pubstate = {published}, tppubtype = {incollection} } @article{, title = {The meaning of the methylation of genomic DNA in the regulation of gene expression levels}, author = { A. Popiela and G. Keith and A. Borzecki and G. Popiela and M. Manowiec and M. Gabrys}, year = {2004}, date = {2004-01-01}, journal = {Eur J Gynaecol Oncol}, volume = {25}, number = {2}, pages = {145-9}, abstract = {INTRODUCTION: Methylation of genomic DNA is one of the major mechanisms that deactivates genes and regulates their tissue-specific transcription levels. Its patterns are based on clonal inheritance that occurs in the early stages of embryogenesis. All changes in the DNA methylation levels occurring especially in the promoter region of the genes, which involve hypo- as well as hyper-methylation, lead to cell differentiation and growth disorders. Therefore it can become an impulse that initiates different pathological processes including carcinogenesis. MATERIAL AND METHODS: The purpose of this review was to present the recent knowledge concerning methylation of genomic DNA based on recent references and authors' experience. RESULTS AND CONCLUSION: Genome stability disorders could be caused either by mutations, which damage the structure of the genes and have not been formerly removed, or as the consequence of an epigenetic mechanism. Methylation plays a decisive role in the activity of many genes and could be a natural weapon of an organism against the expression of foreign genetic material that degrades the original genome structure.}, note = {0392-2936 Journal Article Review Review, Tutorial}, keywords = {*DNA, *Gene, Expression, Human, KEITH, Methylation, Neoplasms/*genetics, Neoplastic, Regulation}, pubstate = {published}, tppubtype = {article} } @article{monneaux_peptide-based_2004, title = {Peptide-based immunotherapy of systemic lupus erythematosus}, author = {Fanny Monneaux and Sylviane Muller}, doi = {10.1016/S1568-9972(03)00061-2}, issn = {1568-9972}, year = {2004}, date = {2004-01-01}, journal = {Autoimmunity Reviews}, volume = {3}, number = {1}, pages = {16--24}, abstract = {Current drug-based therapy for systemic lupus erythematosus (SLE) are non-specific and often counterbalanced by adverse effects. Current research aims at developing specific treatments that target deleterious cells only and not the whole immune system. This strategy requires the identification of sequences derived from major lupus autoantigens, responsible for the activation of autoreactive B and T cells. This review summarizes the identification and characterization of peptides, which are able to modulate T cells ex vivo, and describes the promising results obtained after administration of some of these peptides in lupus mice. Although these therapeutic trials are encouraging, the precise mode of action of peptide-based immunotherapy is still elusive. Here, we discuss the possible mechanisms leading to T-cell tolerance induction and the feasibility of extending the success of peptide-based therapy from animal models to human.}, keywords = {Animals, Antibodies, Antinuclear, Epitopes, Humans, I2CT, Immunotherapy, Lupus Erythematosus, Mice, Monneaux, Peptides, Systemic, T-Lymphocytes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{, title = {Wheat (Triticum vulgare) chloroplast nuclease ChSI exhibits 5' flap structure-specific endonuclease activity}, author = { A. Przykorska and K. Solecka and K. Olszak and G. Keith and B. Nawrot and E. Kuligowska}, year = {2004}, date = {2004-01-01}, journal = {Biochemistry}, volume = {43}, number = {35}, pages = {11283-94}, abstract = {The structure-specific ChSI nuclease from wheat (Triticum vulgare) chloroplast stroma has been previously purified and characterized in our laboratory. It is a single-strand-specific DNA and RNA endonuclease. Although the enzyme has been initially characterized and used as a structural probe, its biological function is still unknown. Localization of the ChSI enzyme inside chloroplasts, possessing their own DNA that is generally highly exposed to UV light and often affected by numerous redox reactions and electron transfer processes, might suggest, however, that this enzyme could be involved in DNA repair. The repair of some types of DNA damage has been shown to proceed through branched DNA intermediates which are substrates for the structure-specific DNA endonucleases. Thus we tested the substrate specificity of ChSI endonuclease toward various branched DNAs containing 5' flap, 5' pseudoflap, 3' pseudoflap, or single-stranded bulged structural motifs. It appears that ChSI has a high 5' flap structure-specific endonucleolytic activity. The catalytic efficiency (k(cat)/K(M)) of the enzyme is significantly higher for the 5' flap substrate than for single-stranded DNA. The ChSI 5' flap activity was inhibited by high concentrations of Mg(2+), Mn(2+), Zn(2+), or Ca(2+). However, low concentrations of divalent cations could restore the loss of ChSI activity as a consequence of EDTA pretreatment. In contrast to other known 5' flap nucleases, the chloroplast enzyme ChSI does not possess any 5'-->3' exonuclease activity on double-stranded DNA. Therefore, we conclude that ChSI is a 5' flap structure-specific endonuclease with nucleolytic activity toward single-stranded substrates.}, note = {0006-2960 Journal Article}, keywords = {&, Acid, Catalysis, Chloroplasts/*enzymology, Conformation, Desorption-Ionization, DNA, Endonucleases/*chemistry/isolation, Exonucleases/chemistry/metabolism, Flap, Gov't, Hydrolysis, KEITH, Kinetics, Laser, Mass, Matrix-Assisted, Non-U.S., Nucleic, Oligonucleotides/chemical, Plant/chemistry/metabolism, purification/*metabolism, Relationship, Single-Stranded/chemistry/metabolism, Specificity, Spectrometry, Structure-Activity, Substrate, Support, synthesis/metabolism, Thermodynamics, Triticum/*enzymology}, pubstate = {published}, tppubtype = {article} } @article{pantarotto_functionalized_2004, title = {Functionalized Carbon Nanotubes for Plasmid DNA Gene Delivery}, author = {Davide Pantarotto and Ravi Singh and David McCarthy and Mathieu Erhardt and Jean-Paul Briand and Maurizio Prato and Kostas Kostarelos and Alberto Bianco}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/anie.200460437}, doi = {10.1002/anie.200460437}, issn = {1521-3773}, year = {2004}, date = {2004-01-01}, urldate = {2020-03-31}, journal = {Angewandte Chemie International Edition}, volume = {43}, number = {39}, pages = {5242--5246}, abstract = {Genetic vaccination and gene therapy research could benefit from the application of carbon nanotubes. Functionalized, positively charged, water-soluble carbon nanotubes are able to penetrate into cells (see figure) and can transport plasmid DNA by formation of noncovalent DNA–nanotube complexes. Such nanotubes can be used as novel nonviral delivery systems for gene transfer.}, keywords = {Carbon nanotubes, gene delivery, I2CT, plasmid DNA, supramolecular chemistry, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {Internal ribosome entry site structural motifs conserved among mammalian fibroblast growth factor 1 alternatively spliced mRNAs}, author = { Y. Martineau and C. Le Bec and L. Monbrun and V. Allo and I. M. Chiu and O. Danos and H. Moine and H. Prats and A. C. Prats}, year = {2004}, date = {2004-01-01}, journal = {Mol Cell Biol}, volume = {24}, number = {17}, pages = {7622-35}, abstract = {Fibroblast growth factor 1 (FGF-1) is a powerful angiogenic factor whose gene structure contains four promoters, giving rise to a process of alternative splicing resulting in four mRNAs with alternative 5' untranslated regions (5' UTRs). Here we have identified, by using double luciferase bicistronic vectors, the presence of internal ribosome entry sites (IRESs) in the human FGF-1 5' UTRs, particularly in leaders A and C, with distinct activities in mammalian cells. DNA electrotransfer in mouse muscle revealed that the IRES present in the FGF-1 leader A has a high activity in vivo. We have developed a new regulatable TET OFF bicistronic system, which allowed us to rule out the possibility of any cryptic promoter in the FGF-1 leaders. FGF-1 IRESs A and C, which were mapped in fragments of 118 and 103 nucleotides, respectively, are flexible in regard to the position of the initiation codon, making them interesting from a biotechnological point of view. Furthermore, we show that FGF-1 IRESs A of murine and human origins show similar IRES activity profiles. Enzymatic and chemical probing of the FGF-1 IRES A RNA revealed a structural domain conserved among mammals at both the nucleotide sequence and RNA structure levels. The functional role of this structural motif has been demonstrated by point mutagenesis, including compensatory mutations. These data favor an important role of IRESs in the control of FGF-1 expression and provide a new IRES structural motif that could help IRES prediction in 5' UTR databases.}, note = {0270-7306 Journal Article}, keywords = {(Genetics), *5', *Alternative, *Nucleic, *Promoter, 1/*genetics, Acid, Alignment, Animals, Base, Cell, Conformation, Data, EHRESMANN, Factor, Fibroblast, Gene, Genes, Genetic, Gov't, Growth, Human, Line, Messenger/chemistry/*genetics/metabolism, Mice, Molecular, Muscle, Mutagenesis, Non-U.S., Regions, Ribosomes/*metabolism, RNA, Sequence, Site-Directed, Skeletal/cytology/physiology, Splicing, Structural/genetics, Support, Techniques, Transfer, Untranslated, Vectors}, pubstate = {published}, tppubtype = {article} } @article{, title = {A simple epigenetic method for the diagnosis and classification of brain tumors}, author = {R Zukiel and S Nowak and A M Barciszewska and I Gawronska and G Keith and M Z Barciszewska}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15037658}, isbn = {15037658}, year = {2004}, date = {2004-01-01}, journal = {Mol Cancer Res}, volume = {2}, number = {3}, pages = {196-202}, abstract = {The new, simple, and reliable method for the diagnosis of brain tumors is described. It is based on a TLC quantitative determination of 5-methylcytosine (m(5)C) in relation to its damage products of DNA from tumor tissue. Currently, there is evidence that oxidative stress through reactive oxygen species (ROS) plays an important role in the etiology and progression of several human diseases. Oxidative damage of DNA, lipids, and proteins is deleterious for the cell. m(5)C, along with other basic components of DNA, is the target for ROS, which results in the appearance of new modified nucleic acid bases. If so, m(5)C residue constitutes a mutational hotspot position, whether it occurs within a nucleotide sequence of a structural gene or a regulatory region. Here, we show the results of the analysis of 82 DNA samples taken from brain tumor tissues. DNA was isolated and hydrolyzed into nucleotides, which, after labeling with [gamma-(32)P]ATP, were separated on TLC. Chromatograms were evaluated using PhosphorImager and the amounts of 5-methyldeoxycytosine (m(5)dC) were calculated as a ratio (R) of m(5)dC to m(5)dC + deoxycytosine + deoxythymidine spot intensities. The R value could not only be a good diagnostic marker for brain tumors but also a factor differentiating low-grade and high-grade gliomas. Therefore, DNA methylation pattern might be a useful tool to give a primary diagnosis of a brain tumor or as a marker for the early detection of the relapse of the disease. This method has several advantages over those existing nowadays.}, note = {1541-7786 Journal Article}, keywords = {Genetic Female Human Male Middle Aged Oxidative Stress Reactive Oxygen Species/metabolism Sensitivity and Specificity Support, KEITH 5-Methylcytosine/*analysis Adult Aged Brain Neoplasms/*classification/*diagnosis/genetics/pathology Chromatography, Neoplasm/*chemistry/*metabolism *Epigenesis, Non-U.S. Gov't, Thin Layer *DNA Methylation DNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Two distinct domains of the beta subunit of Aquifex aeolicus leucyl-tRNA synthetase are involved in tRNA binding as revealed by a three-hybrid selection}, author = {Y G Zheng and H Wei and C Ling and F Martin and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15208367}, isbn = {15208367}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {11}, pages = {3294-3303}, abstract = {The Aquifex aeolicus alphabeta-LeuRS is the only known heterodimeric class Ia aminoacyl-tRNA synthetase. In this study, we investigated the function of the beta subunit which is believed to bind tRNA(Leu). A yeast three-hybrid system was constructed on the basis of the interaction of the beta subunit with its cognate tRNA(Leu). Then, seven mutated beta subunits exhibiting impaired tRNA binding capacities were selected out from a randomly mutated library. Two mutations were identified in the class Ia-helix-bundle-domain, which might interact with the D-hairpin of the tRNA analogous to other class Ia tRNA:synthetases complexes. The five other mutations were found in the LeuRS-specific C-terminal domain of which the folding is still unknown. tRNA affinity measurements and kinetic analyses performed on the isolated beta subunits and on the co-expressed alphabeta-heterodimers showed for all the mutants an effect in tRNA affinity in the ground state. In addition, an effect on the transition state of the aminoacylation reaction was observed for a 21-residues deletion mutant of the C-terminal end. These results show that the genetic approach of the three hybrid system is widely applicable and is a powerful tool for the investigation of tRNA:synthetase interactions.}, note = {1362-4962 Evaluation Studies Journal Article}, keywords = {Amino Acid Sequence Binding Sites Gram-Negative Bacteria/*enzymology Kinetics Leucine-tRNA Ligase/*chemistry/genetics/*metabolism Molecular Sequence Data Mutation Protein Structure, ERIANI, Leu/*metabolism Saccharomyces cerevisiae/genetics *Two-Hybrid System Techniques, Tertiary Protein Subunits/chemistry RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Single processing center models for human Dicer and bacterial RNase III}, author = {H Zhang and F A Kolb and L Jaskiewicz and E Westhof and W Filipowicz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15242644}, isbn = {15242644}, year = {2004}, date = {2004-01-01}, journal = {Cell}, volume = {118}, number = {1}, pages = {57-68}, abstract = {Dicer is a multidomain ribonuclease that processes double-stranded RNAs (dsRNAs) to 21 nt small interfering RNAs (siRNAs) during RNA interference, and excises microRNAs from precursor hairpins. Dicer contains two domains related to the bacterial dsRNA-specific endonuclease, RNase III, which is known to function as a homodimer. Based on an X-ray structure of the Aquifex aeolicus RNase III, models of the enzyme interaction with dsRNA, and its cleavage at two composite catalytic centers, have been proposed. We have generated mutations in human Dicer and Escherichia coli RNase III residues implicated in the catalysis, and studied their effect on RNA processing. Our results indicate that both enzymes have only one processing center, containing two RNA cleavage sites and generating products with 2 nt 3' overhangs. Based on these and other data, we propose that Dicer functions through intramolecular dimerization of its two RNase III domains, assisted by the flanking RNA binding domains, PAZ and dsRBD.}, note = {0092-8674 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Base Sequence Comparative Study Conserved Sequence Dimerization Endoribonucleases/*chemistry/genetics/isolation & purification/*metabolism Escherichia coli/enzymology Human Manganese/metabolism MicroRNAs/metabolism Models, Double-Stranded/chemistry/*metabolism RNA, Molecular Molecular Sequence Data Molecular Weight Mutagenesis, Non-U.S. Gov't, Post-Transcriptional RNA, Secondary Protein Structure, Site-Directed Mutation Protein Structure, Small Interfering/metabolism Recombinant Proteins/metabolism Ribonuclease III/*chemistry/genetics/isolation & purification/*metabolism Sequence Homology, Tertiary RNA Helicases/*chemistry/genetics/isolation & purification/*metabolism *RNA Processing, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural biology. Evolution of RNA architecture}, author = {E Westhof and C Massire}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15459373}, isbn = {15459373}, year = {2004}, date = {2004-01-01}, journal = {Science}, volume = {306}, number = {5693}, pages = {62-63}, note = {1095-9203 Comment Journal Article}, keywords = {Adenine/chemistry Base Pairing Computational Biology Crystallography, Bacterial/*chemistry/metabolism RNA, Molecular Nucleic Acid Conformation RNA Precursors/metabolism RNA, Transfer/metabolism Ribonuclease P/*chemistry/metabolism Thermus thermophilus/*chemistry/enzymology, Unité ARN, WESTHOF, X-Ray Evolution}, pubstate = {published}, tppubtype = {article} } @incollection{, title = {Detection and quantification of modified nucleotides in RNA using thin-layer chromatography}, author = {H Grosjean and G Keith and L Droogmans}, editor = {J M Gott}, year = {2004}, date = {2004-01-01}, booktitle = {RNA Interference, Editing, and Modification: Methods and Protocols}, volume = {265}, pages = {357-91}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Identification of a modified nucleotide and its localization within an RNA molecule is a difficult task. Only direct sequencing of purified RNA molecules and high-performance liquid chromatography mass spectrometry analysis of purified RNA fragments allow determination of both the type and location of a given modified nucleotide within an RNA of 50-150 nt in length. The objective of this chapter is to describe in detail a few simple procedures that we have found particularly suited for the detection, localization, and quantification of modified nucleotides within an RNA of known sequence. The methods can also be used to reveal the enzymatic activity of a particular RNA-modifying enzyme in vitro or in vivo. The procedures are based on the use of radiolabeled RNA (with [32P], [14C], or [3H]) or [32P]-postlabeled oligonucleotides and two-dimensional thin-layer chromatography of labeled nucleotides on cellulose plates. This chapter provides useful maps of the migration characteristics of 70 modified nucleotides on thin-layer cellulose plates.}, keywords = {KEITH 5' Untranslated Regions/chemistry Base Composition Chromatography, Non-U.S. Gov't}, pubstate = {published}, tppubtype = {incollection} } @article{, title = {NINETY YEARS and a lot more to come}, author = {E Westhof}, url = {http://www-ibmc.u-strasbg.fr/upr9002/westhof/PDF/r2004_EWesthof_Biochimie.pdf}, isbn = {15667935}, year = {2004}, date = {2004-01-01}, journal = {Biochimie}, volume = {86}, number = {12}, pages = {865}, note = {0300-9084 Editorial}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {How to silence silencing}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15123277}, isbn = {15123277}, year = {2004}, date = {2004-01-01}, journal = {Chem Biol}, volume = {11}, number = {2}, pages = {158-160}, abstract = {Two recent reports describe the stunning crystal structures of complexes between a viral protein that suppresses RNA silencing and a 21 nucleotide small interfering (si)RNA.}, note = {1074-5521 Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Determination of thermodynamic parameters for HIV DIS type loop-loop kissing complexes}, author = {A Weixlbaumer and A Werner and C Flamm and E Westhof and R Schroeder}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15459283}, isbn = {15459283}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {17}, pages = {5126-5133}, abstract = {The HIV-1 type dimerization initiation signal (DIS) loop was used as a starting point for the analysis of the stability of Watson-Crick (WC) base pairs in a tertiary structure context. We used ultraviolet melting to determine thermodynamic parameters for loop-loop tertiary interactions and compared them with regular secondary structure RNA helices of the same sequences. In 1 M Na+ the loop-loop interaction of a HIV-1 DIS type pairing is 4 kcal/mol more stable than its sequence in an equivalent regular and isolated RNA helix. This difference is constant and sequence independent, suggesting that the rules governing the stability of WC base pairs in the secondary structure context are also valid for WC base pairs in the tertiary structure context. Moreover, the effect of ion concentration on the stability of loop-loop tertiary interactions differs considerably from that of regular RNA helices. The stabilization by Na+ and Mg2+ is significantly greater if the base pairing occurs within the context of a loop-loop interaction. The dependence of the structural stability on salt concentration was defined via the slope of a T(m)/log [ion] plot. The short base-paired helices are stabilized by 8 degrees C/log [Mg2+] or 11 degrees C/log [Na+], whereas base-paired helices forming tertiary loop-loop interactions are stabilized by 16 degrees C/log [Mg2+] and 26 degrees C/log [Na+]. The different dependence on ionic strength that is observed might reflect the contribution of specific divalent ion binding to the preformation of the hairpin loops poised for the tertiary kissing loop-loop contacts.}, note = {1362-4962 Journal Article}, keywords = {Double-Stranded/chemistry RNA, Non-U.S. Gov't Thermodynamics Ultraviolet Rays, Unité ARN, Viral/*chemistry Sodium/chemistry Support, WESTHOF Base Pairing Base Sequence Dimerization HIV-1/*genetics Macromolecular Systems Magnesium/chemistry Molecular Sequence Data Nucleic Acid Conformation RNA Stability RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mechanism of dimerization of bicoid mRNA: initiation and stabilization}, author = {C Wagner and C Ehresmann and B Ehresmann and C Brunel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14607826}, isbn = {14607826}, year = {2004}, date = {2004-01-01}, journal = {J Biol Chem}, volume = {279}, number = {6}, pages = {4560-4569}, abstract = {Dimerization of bcd mRNA was shown to be important for the formation of ribonucleoprotein particles and their localization in Drosophila embryo. The cis-element responsible for dimerization is localized in a stem-loop domain (domain III) containing two essential complementary 6-nucleotide sequences in a hairpin loop (LIIIb) and an interior loop (LIIIa). Such an RNA element can potentially generate single or double "hand-by-arm" interactions leading to open and closed complexes, respectively. The former retains the possibility of forming multimers, whereas the latter does not. We showed previously that dimerization proceeds through a two-step mechanism, which includes a transition from the reversible initiation complex into a very stable one. Here we have addressed the nature of the initial interactions and the mechanism of transition. We engineered a series of different RNA fragments with the capacity to form defined open dimers, multimers, or closed dimers. We compared their thermodynamic and kinetic behavior and mapped nucleotides involved in intermolecular interactions by enzymatic and chemical footprinting experiments and chemical modification interference. Our results indicate that the initiation step leads to a reversible open dimer, involving a more limited number of intermolecular base pairs than expected. The two loops play distinct roles in this process, and the structure of loop IIIb is more constrained than that of loop IIIa. Thus, loop IIIa appears to be the driving element of the recognition process. The initial open dimer is then converted into a stable closed dimer, possibly through a kinetically controlled mechanism.}, note = {0021-9258 Journal Article}, keywords = {BRUNEL Animals Base Sequence Dimerization Drosophila Proteins/*genetics Drosophila melanogaster/embryology/*genetics/*metabolism Homeodomain Proteins/*genetics Kinetics Molecular Sequence Data Nucleic Acid Conformation RNA Interference RNA Stability RNA, Messenger/chemistry/*genetics/*metabolism Support, Non-U.S. Gov't Support, P.H.S. Thermodynamics Trans-Activators/*genetics, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Atypical archaeal tRNA pyrrolysine transcript behaves towards EF-Tu as a typical elongator tRNA}, author = {A Théobald-Dietrich and M Frugier and R Giege and J Rudinger-Thirion}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14872064}, isbn = {14872064}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {3}, pages = {1091-1096}, abstract = {The newly discovered tRNA(Pyl) is involved in specific incorporation of pyrrolysine in the active site of methylamine methyltransferases in the archaeon Methanosarcina barkeri. In solution probing experiments, a transcript derived from tRNA(Pyl) displays a secondary fold slightly different from the canonical cloverleaf and interestingly similar to that of bovine mitochondrial tRNA(Ser)(uga). Aminoacylation of tRNA(Pyl) transcript by a typical class II synthetase, LysRS from yeast, was possible when its amber anticodon CUA was mutated into a lysine UUU anticodon. Hydrolysis protection assays show that lysylated tRNA(Pyl) can be recognized by bacterial elongation factor. This indicates that no antideterminant sequence is present in the body of the tRNA(Pyl) transcript to prevent it from interacting with EF-Tu, in contrast with the otherwise functionally similar tRNA(Sec) that mediates selenocysteine incorporation.}, note = {1362-4962 Journal Article}, keywords = {Anticodon/metabolism Base Sequence Lysine/*analogs & derivatives/*metabolism Lysine-tRNA Ligase/metabolism Methanosarcina barkeri/genetics Mitochondria/genetics Molecular Sequence Data Nucleic Acid Conformation Peptide Elongation Factor Tu/*metabolism RNA, Archaeal/chemistry/*metabolism RNA, FRUGIER, Non-U.S. Gov't Yeasts/enzymology, Ser/chemistry Selenocysteine/metabolism Support, Transfer, Transfer/chemistry/*metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Recognition of human mitochondrial tRNALeu(UUR) by its cognate leucyl-tRNA synthetase}, author = {B Sohm and M Sissler and H Park and M P King and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15123417}, isbn = {15123417}, year = {2004}, date = {2004-01-01}, journal = {J Mol Biol}, volume = {339}, number = {1}, pages = {17-29}, abstract = {Accuracy of protein synthesis depends on specific recognition and aminoacylation of tRNAs by their cognate aminoacyl-tRNA synthetases. Rules governing these processes have been established for numerous prokaryotic and eukaryotic cytoplasmic systems, but only limited information is available for human mitochondrial systems. It has been shown that the in vitro transcribed human mitochondrial tRNA(Leu(UUR)) does not fold into the expected cloverleaf, but is however aminoacylated by the human mitochondrial leucyl-tRNA synthetase. Here, the role of the structure of the amino acid acceptor branch and the anticodon branch of tRNA(Leu(UUR)) in recognition by leucyl-tRNA synthetase was investigated. The kinetic parameters for aminoacylation of wild-type and mutant tRNA(Leu(UUR)) transcripts and of native tRNA(Leu(UUR)) were determined. Solution structure probing was performed in the presence or in the absence of leucyl-tRNA synthetase and correlated with the aminoacylation kinetics for each tRNA. Replacement of mismatches in either the anticodon-stem or D-stem that are present in the wild-type tRNA(Leu(UUR)) by G-C base-pairs is sufficient to induce (i) cloverleaf folding, (ii) improved aminoacylation efficiency, and (iii) interactions with the synthetase that are similar to those with the native tRNA(Leu(UUR)). Leucyl-tRNA synthetase contacts tRNA(Leu(UUR)) in the amino acid acceptor stem, the anticodon stem, and the D-loop, which is unprecedented for a leucine aminoacylation system.}, note = {0022-2836 Journal Article}, keywords = {Cultured, FLORENTZ, FLORENTZ *Acylation Base Sequence Comparative Study Human Kinetics Leucine/metabolism Leucine-tRNA Ligase/genetics/*metabolism Mitochondria/*metabolism Molecular Sequence Data Mutation Nucleic Acid Conformation Osteosarcoma/metabolism RNA/*genetics/metabolism RNA, Genetic/*genetics Tumor Cells, Leu/genetics/*metabolism Solutions Substrate Specificity Support, Non-U.S. Gov't Support, P.H.S. Transcription, SISSLER, Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoacylation properties of pathology-related human mitochondrial tRNA(Lys) variants}, author = {M Sissler and M Helm and M Frugier and R Giege and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15100439}, isbn = {15100439}, year = {2004}, date = {2004-01-01}, journal = {RNA}, volume = {10}, number = {5}, pages = {841-853}, abstract = {In vitro transcription has proven to be a successful tool for preparation of functional RNAs, especially in the tRNA field, in which, despite the absence of post-transcriptional modifications, transcripts are correctly folded and functionally active. Human mitochondrial (mt) tRNA(Lys) deviates from this principle and folds into various inactive conformations, due to the absence of the post-transcriptional modification m(1)A9 which hinders base-pairing with U64 in the native tRNA. Unavailability of a functional transcript is a serious drawback for structure/function investigations as well as in deciphering the molecular mechanisms by which point mutations in the mt tRNA(Lys) gene cause severe human disorders. Here, we show that an engineered in vitro transcribed "pseudo-WT" tRNA(Lys) variant is efficiently recognized by lysyl-tRNA synthetase and can substitute for the WT tRNA as a valuable reference molecule. This has been exploited in a systematic analysis of the effects on aminoacylation of nine pathology-related mutations described so far. The sole mutation located in a loop of the tRNA secondary structure, A8344G, does not affect aminoacylation efficiency. Out of eight mutations located in helical domains converting canonical Watson-Crick pairs into G-U pairs or C.A mismatches, six have no effect on aminoacylation (A8296G, U8316C, G8342A, U8356C, U8362G, G8363A), and two lead to drastic decreases (5000- to 7000-fold) in lysylation efficiencies (G8313A and G8328A). This screening, allowing for analysis of the primary impact level of all mutations affecting one tRNA under comparable conditions, indicates distinct molecular origins for different disorders.}, note = {1355-8382 Journal Article}, keywords = {ERIANI, FLORENTZ, FLORENTZ GIEGE Acylation Aminoacyltransferases/*genetics Human MERRF Syndrome/genetics Mitochondria/*genetics Mitochondrial Diseases/*genetics Mutation Nucleic Acid Conformation RNA, FRUGIER, Lys/*genetics Sequence Analysis, Non-U.S. Gov't Variation (Genetics), RNA Support, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The meaning of the methylation of genomic DNA in the regulation of gene expression levels}, author = {A Popiela and G Keith and A Borzecki and G Popiela and M Manowiec and M Gabrys}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15032270}, isbn = {15032270}, year = {2004}, date = {2004-01-01}, journal = {Eur J Gynaecol Oncol}, volume = {25}, number = {2}, pages = {145-9}, abstract = {INTRODUCTION: Methylation of genomic DNA is one of the major mechanisms that deactivates genes and regulates their tissue-specific transcription levels. Its patterns are based on clonal inheritance that occurs in the early stages of embryogenesis. All changes in the DNA methylation levels occurring especially in the promoter region of the genes, which involve hypo- as well as hyper-methylation, lead to cell differentiation and growth disorders. Therefore it can become an impulse that initiates different pathological processes including carcinogenesis. MATERIAL AND METHODS: The purpose of this review was to present the recent knowledge concerning methylation of genomic DNA based on recent references and authors' experience. RESULTS AND CONCLUSION: Genome stability disorders could be caused either by mutations, which damage the structure of the genes and have not been formerly removed, or as the consequence of an epigenetic mechanism. Methylation plays a decisive role in the activity of many genes and could be a natural weapon of an organism against the expression of foreign genetic material that degrades the original genome structure.}, note = {0392-2936 Journal Article Review Review, Tutorial}, keywords = {KEITH *DNA Methylation *Gene Expression Regulation, Neoplastic Human Neoplasms/*genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pronounced instability of tandem IU base pairs in RNA}, author = {M J Serra and P E Smolter and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15037659}, isbn = {15037659}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {5}, pages = {1824-1828}, abstract = {Optical melting was used to determine the stabilities of three series of RNA oligomers containing tandem XU base pairs, GGCXUGCC (5'XU3'), GGCUXGCC (5'UX3') and GGCXXGGC/CCGUUCCG (5'XX3'), where X is either A, G or I (inosine). The helices containing tandem AU base pairs were the most stable in the first two series (5'XU3' and 5'UX3'), with an average melting temperature approximately 11 degrees C higher than the helices with tandem 5'GU3' base pairs and 25 degrees C higher than the helices with tandem 5'IU3' base pairs. For the third series (5'XX3'), the helix containing tandem GG is the most stable, with an average melting temperature approximately 2 degrees C higher than the helix with tandem AA base pairs and approximately 24 degrees C higher than the helix with tandem II base pairs. The thermodynamic stability of the oligomers with tandem IU base pairs was also investigated as a function of magnesium ion concentration. As with normal A-U or G-U tandem duplexes, the data could best be interpreted as non-specific binding of magnesium ions to the inosine-containing RNA oligonucleotides.}, note = {1362-4962 Journal Article}, keywords = {Base Pairing Inosine/*chemistry Magnesium/chemistry Nucleic Acid Conformation RNA/*chemistry RNA Stability Support, Non-P.H.S. Thermodynamics Uracil/*chemistry, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Phasing in the presence of severe site-specific radiation damage through dose-dependent modelling of heavy atoms}, author = {M Schiltz and P Dumas and E Ennifar and C Flensburg and W Paciorek and C Vonrhein and G Bricogne}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15159561}, isbn = {15159561}, year = {2004}, date = {2004-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {60}, number = {Pt 6}, pages = {1024-1031}, abstract = {The case of a brominated RNA crystal structure determination in which standard three-wavelength MAD phasing was unsuccessful because of fast X-ray-induced debromination was reinvestigated [Ennifar et al. (2002), Acta Cryst. D58, 1262-1268]. It was found that if the data are kept unmerged and if a dose-stamp is associated with each reflection measurement, dose-dependent occupancies can be refined for the Br atoms. Such a parametrization has been implemented in the macromolecular phasing program SHARP. Refining such dose-dependent occupancies on an unmerged data set gave a dramatic improvement, even for SAD phases from only the first wavelength (peak), and resulted in a good electron-density map after solvent flattening. The adverse effect of radiation damage has been turned into a beneficial one. The crucial difference is made by the use of unmerged data: phasing power is generated through the intensity differences of symmetry-related reflections recorded at different doses, i.e. corresponding to different states of the X-ray-induced debromination. This approach should prove useful in all situations of experimental phasing where site-specific radiation damage occurs unavoidably and undesirably and not only in cases in which radiation damage is purposely being created in order to demonstrate its potential usefulness.}, note = {0907-4449 Journal Article}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Architecture and folding mechanism of the Azoarcus Group I Pre-tRNA}, author = {P Rangan and B Masquida and E Westhof and S A Woodson}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15123419}, isbn = {15123419}, year = {2004}, date = {2004-01-01}, journal = {J Mol Biol}, volume = {339}, number = {1}, pages = {41-51}, abstract = {Self-splicing RNAs must evolve to function in their specific exon context. The conformation of a group I pre-tRNA(ile) from the bacterium Azoarcus was probed by ribonuclease T(1) and hydroxyl radical cleavage, and by native gel electrophoresis. Biochemical data and three-dimensional models of the pre-tRNA showed that the tRNA is folded, and that the tRNA and intron sequences form separate tertiary domains. Models of the active site before steps 1 and 2 of the splicing reaction predict that exchange of the external G-cofactor and the 3'-terminal G is accomplished by a slight conformational change in P9.0 of the Azoarcus group I intron. Kinetic assays showed that the pre-tRNA folds in minutes, much more slowly than the intron alone. The dependence of the folding kinetics on Mg(2+) and the concentration of urea, and RNase T(1) experiments showed that formation of native pre-tRNA is delayed by misfolding of P3-P9, including mispairing between residues in P9 and the tRNA. Thus, although the intron and tRNA sequences form separate domains in the native pre-tRNA, their folding is coupled via metastable non-native base-pairs. This could help prevent premature processing of the 5' and 3' ends of unspliced pre-tRNA.}, note = {0022-2836 Journal Article}, keywords = {Azoarcus/enzymology/*genetics Base Sequence Binding Sites Exoribonucleases/metabolism Hydroxyl Radical/metabolism Introns/*genetics Magnesium/chemistry Models, Bacterial/*chemistry/genetics/*metabolism RNA, Ile/chemistry/*genetics Substrate Specificity Support, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA Precursors/*genetics RNA Splice Sites/genetics RNA Splicing RNA, Non-U.S. Gov't Support, P.H.S., Transfer, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Wheat (Triticum vulgare) chloroplast nuclease ChSI exhibits 5' flap structure-specific endonuclease activity}, author = {A Przykorska and K Solecka and K Olszak and G Keith and B Nawrot and E Kuligowska}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15366938}, isbn = {15366938}, year = {2004}, date = {2004-01-01}, journal = {Biochemistry}, volume = {43}, number = {35}, pages = {11283-11294}, abstract = {The structure-specific ChSI nuclease from wheat (Triticum vulgare) chloroplast stroma has been previously purified and characterized in our laboratory. It is a single-strand-specific DNA and RNA endonuclease. Although the enzyme has been initially characterized and used as a structural probe, its biological function is still unknown. Localization of the ChSI enzyme inside chloroplasts, possessing their own DNA that is generally highly exposed to UV light and often affected by numerous redox reactions and electron transfer processes, might suggest, however, that this enzyme could be involved in DNA repair. The repair of some types of DNA damage has been shown to proceed through branched DNA intermediates which are substrates for the structure-specific DNA endonucleases. Thus we tested the substrate specificity of ChSI endonuclease toward various branched DNAs containing 5' flap, 5' pseudoflap, 3' pseudoflap, or single-stranded bulged structural motifs. It appears that ChSI has a high 5' flap structure-specific endonucleolytic activity. The catalytic efficiency (k(cat)/K(M)) of the enzyme is significantly higher for the 5' flap substrate than for single-stranded DNA. The ChSI 5' flap activity was inhibited by high concentrations of Mg(2+), Mn(2+), Zn(2+), or Ca(2+). However, low concentrations of divalent cations could restore the loss of ChSI activity as a consequence of EDTA pretreatment. In contrast to other known 5' flap nucleases, the chloroplast enzyme ChSI does not possess any 5'-->3' exonuclease activity on double-stranded DNA. Therefore, we conclude that ChSI is a 5' flap structure-specific endonuclease with nucleolytic activity toward single-stranded substrates.}, note = {0006-2960 Journal Article}, keywords = {KEITH Catalysis Chloroplasts/*enzymology DNA, Mass, Matrix-Assisted Laser Desorption-Ionization Structure-Activity Relationship Substrate Specificity Support, Non-U.S. Gov't Thermodynamics Triticum/*enzymology, Plant/chemistry/metabolism DNA, Single-Stranded/chemistry/metabolism Exonucleases/chemistry/metabolism Flap Endonucleases/*chemistry/isolation & purification/*metabolism Hydrolysis Kinetics Nucleic Acid Conformation Oligonucleotides/chemical synthesis/metabolism Spectrometry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The meaning of the methylation of genomic DNA in the regulation of gene expression levels}, author = {A Popiela and G Keith and A Borzecki and G Popiela and M Manowiec and M Gabrys}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15032270}, isbn = {15032270}, year = {2004}, date = {2004-01-01}, journal = {Eur J Gynaecol Oncol}, volume = {25}, number = {2}, pages = {145-149}, abstract = {INTRODUCTION: Methylation of genomic DNA is one of the major mechanisms that deactivates genes and regulates their tissue-specific transcription levels. Its patterns are based on clonal inheritance that occurs in the early stages of embryogenesis. All changes in the DNA methylation levels occurring especially in the promoter region of the genes, which involve hypo- as well as hyper-methylation, lead to cell differentiation and growth disorders. Therefore it can become an impulse that initiates different pathological processes including carcinogenesis. MATERIAL AND METHODS: The purpose of this review was to present the recent knowledge concerning methylation of genomic DNA based on recent references and authors' experience. RESULTS AND CONCLUSION: Genome stability disorders could be caused either by mutations, which damage the structure of the genes and have not been formerly removed, or as the consequence of an epigenetic mechanism. Methylation plays a decisive role in the activity of many genes and could be a natural weapon of an organism against the expression of foreign genetic material that degrades the original genome structure.}, note = {0392-2936 Journal Article Review Review, Tutorial}, keywords = {KEITH *DNA Methylation *Gene Expression Regulation, Neoplastic Human Neoplasms/*genetics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dimerization of retroviral RNA genomes: an inseparable pair}, author = {J C Paillart and M Shehu-Xhilaga and R Marquet and J Mak}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15152202}, isbn = {15152202}, year = {2004}, date = {2004-01-01}, journal = {Nat Rev Microbiol}, volume = {2}, number = {6}, pages = {461-472}, note = {1740-1526 Journal Article Review Review, Tutorial}, keywords = {Base Sequence Dimerization *Genome, MARQUET, Molecular Molecular Sequence Data Nucleic Acid Conformation Virus Replication/*genetics, PAILLART, Unité ARN, Viral HIV-1/*genetics/growth & development Human Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {First snapshots of the HIV-1 RNA structure in infected cells and in virions}, author = {J C Paillart and M Dettenhofer and X F Yu and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15355993}, isbn = {15355993}, year = {2004}, date = {2004-01-01}, journal = {J Biol Chem}, volume = {279}, number = {46}, pages = {48397-48403}, abstract = {With the increasing interest of RNAs in regulating a range of cell biological processes, very little is known about the structure of RNAs in tissue culture cells. We focused on the 5'-untranslated region of the human immunodeficiency virus type 1 RNA genome, a highly conserved RNA region, which contains structural domains that regulate key steps in the viral replication cycle. Up until now, structural information only came from in vitro studies. Here, we developed chemical modification assays to test nucleotide accessibility directly in infected cells and viral particles, thus circumventing possible biases and artifacts linked to in vitro assays. The secondary structure of the 5'-untranslated region in infected cells points to the existence of the various stem-loop motifs associated to distinct functions, proposed from in vitro probing, mutagenesis, and phylogeny. However, compared with in vitro data, subtle differences were observed in the dimerization initiation site hairpin, and none of the proposed long range interactions were observed between the functional domains. Moreover, no global RNA rearrangement was observed; structural differences between infected cells and viral particles were limited to the primer binding site, which became protected against chemical modification upon tRNA(3) (Lys) annealing in virions and to the main packaging signal. In addition, our data suggested that the genomic RNA could already dimerize in the cytoplasm of infected cells. Taken together, our results provided the first analysis of the dynamic of RNA structure of the human immunodeficiency virus type 1 RNA genome during virus assembly ex vivo.}, note = {0021-9258 Journal Article}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of snRNA and snRNA-type genes in the pufferfish Fugu rubripes}, author = {E Myslinski and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15087134}, isbn = {15087134}, year = {2004}, date = {2004-01-01}, journal = {Gene}, volume = {330}, pages = {149-158}, abstract = {Vertebrate snRNA and snRNA-type genes occur in independent transcription units with external promoters. The transcription level from the basal promoter is enhanced by the distal sequence element DSE. This element contains almost invariably two activator submotifs, the Staf binding site and the octamer motif, recruiting the Staf and Oct-1 transcriptional activators. In the present work, database search identified 35 snRNA and snRNA-type genes in the genome sequence of the pufferfish Fugu rubripes. Sequence comparisons of promoter elements, determination of template activities by microinjection into Xenopus oocytes and DNA binding assays of the transcriptional activators led to the surprising finding that only two Fugu genes conform to the general scheme with the expected two submotifs in the DSE. Distinctively, all the other DSEs harbor a unique Staf binding site. Also striking was the observation that the tRNA(Sec), and the snRNA genes that are tandemly repeated, are transcribed from promoter-less DSEs. Evolutionary implications of these results are discussed.}, note = {0378-1119 Journal Article}, keywords = {Amino Acid Sequence Homology, Complementary/chemistry/genetics DNA-Binding Proteins/metabolism Female Genome Molecular Sequence Data Oocytes/metabolism Promoter Regions (Genetics)/genetics Protein Binding RNA, DNA Sequence Homology, KROL Amino Acid Sequence Animals Base Sequence Binding Sites/genetics Comparative Study DNA, Nucleic Acid Takifugu/*genetics Transcription Factors/metabolism Xenopus laevis, Nucleic Acid/genetics Response Elements/genetics Sequence Alignment Sequence Analysis, Small Nuclear/*genetics Regulatory Sequences, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cell protection, resistance and invasiveness of two malignant mesotheliomas as assessed by 10K-microarray}, author = {S Mohr and G Keith and F Galateau-Salle and P Icard and B H Rihn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14732480}, isbn = {14732480}, year = {2004}, date = {2004-01-01}, journal = {Biochim Biophys Acta-Mol Basis Dis}, volume = {1688}, number = {1}, pages = {43-60}, abstract = {Malignant pleural mesothelioma (MPM) is an aggressive serosal tumor, strongly associated with former exposure to asbestos fibers and for which there is currently no effective treatment available. In human, MPM is characterized by a high local invasiveness, poor prognosis and therapeutic outcomes. In order to assess molecular changes that specify this phenotype, we performed a global gene expression profiling of human MPM. Using a 10,000-element microarray, we analyzed mRNA relative gene expression levels by comparing a mesothelioma cell line to either a pleural cell line or tumor specimens. To analyze these gene expression data, we used various bioinformatics softwares. Hierarchical clustering methods were used to group genes and samples with similar expression in an unsupervised mode. Genes of known function were further sorted by enzyme, function and pathway clusters using a supervised software (IncyteGenomics). Taken together, these data defined a molecular fingerprint of human MPM with more than 700 up- or down-regulated genes related to several traits of the malignant phenotype, specially associated with MPM invasiveness, protection and resistance to anticancer defenses. This portrait is meaningful in disease classification and management, and relevant in finding new specific markers of MPM. These molecular markers should improve the accuracy of mesothelioma diagnosis, prognosis and therapy.}, note = {0006-3002 Journal Article}, keywords = {Biological/genetics, KEITH Cell Line, Messenger/analysis Tumor Markers, Neoplastic Human Mesothelioma/etiology/genetics/*pathology Multigene Family Pleural Neoplasms/etiology/genetics/*pathology Protein Array Analysis RNA, Tumor Gene Expression Profiling Gene Expression Regulation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Microdissection, mRNA amplification and microarray: a study of pleural mesothelial and malignant mesothelioma cells}, author = {S Mohr and M C Bottin and B Lannes and A Neuville and J P Bellocq and G Keith and B H Rihn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14987796}, isbn = {14987796}, year = {2004}, date = {2004-01-01}, journal = {Biochimie}, volume = {86}, number = {1}, pages = {13-19}, abstract = {The studies of molecular alterations in tumor cells with microarrays are often hampered by inherent tissue heterogeneity. The emergence of Laser Capture Microdissection (LCM) allowed us to overcome this challenge since it gives selective access to cancer cells that are isolated from their native tissue environment. In this report, we microdissected mesothelial cells and malignant mesothelioma cells of ex vivo resected specimens using LCM. Amplified RNA from mesothelial and mesothelioma microdissected cells allowed us to measure global gene expression with 10 K-microarrays in four independent experiments. We screened 9850 annotated human genes, 1275 of which have satisfied our data analysis requirements. They included 302 overexpressed genes and 160 downregulated genes in mesothelioma microdissected cells as compared to mesothelial microdissected cells. Among them, the expression levels of eight genes, namely BF, FTL, IGFBP7, RARRES1, RARRES2, RBP1, SAT, and TXN according to HUGO nomenclature, were increased, whereas six: ALOX5AP, CLNS1A, EIF4A2, ELK3, REQ and SYPL, were found to be underexpressed in mesothelioma microdissected cells. The ferritin light polypeptide (FTL) gene overexpression was confirmed by real time quantitative PCR. Our approach allowed a comprehensive in situ examination of mesothelioma and provided an accurate way to find new marker genes that may be useful for diagnosis and treatment of malignant pleural mesothelioma.}, note = {0300-9084 Journal Article}, keywords = {KEITH Epithelium/*metabolism Female Gene Expression Profiling Gene Expression Regulation, Messenger Reverse Transcriptase Polymerase Chain Reaction, Neoplastic/*genetics Genetic Markers Human Lasers Male Mesothelioma/*genetics/metabolism Microdissection Oligonucleotide Array Sequence Analysis Pleura/*cytology/*metabolism Pleural Neoplasms/*genetics/metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Specific recognition of rpsO mRNA and 16S rRNA by Escherichia coli ribosomal protein S15 relies on both mimicry and site differentiation}, author = {N Mathy and O Pellegrini and A Serganov and D J Patel and C Ehresmann and C Portier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15101974}, isbn = {15101974}, year = {2004}, date = {2004-01-01}, journal = {Mol Microbiol}, volume = {52}, number = {3}, pages = {661-675}, abstract = {The ribosomal protein S15 binds to 16S rRNA, during ribosome assembly, and to its own mRNA (rpsO mRNA), affecting autocontrol of its expression. In both cases, the RNA binding site is bipartite with a common subsite consisting of a G*U/G-C motif. The second subsite is located in a three-way junction in 16S rRNA and in the distal part of a stem forming a pseudoknot in Escherichia coli rpsO mRNA. To determine the extent of mimicry between these two RNA targets, we determined which amino acids interact with rpsO mRNA. A plasmid carrying rpsO (the S15 gene) was mutagenized and introduced into a strain lacking S15 and harbouring an rpsO-lacZ translational fusion. Analysis of deregulated mutants shows that each subsite of rpsO mRNA is recognized by a set of amino acids known to interact with 16S rRNA. In addition to the G*U/G-C motif, which is recognized by the same amino acids in both targets, the other subsite interacts with amino acids also involved in contacts with helix H22 of 16S rRNA, in the region adjacent to the three-way junction. However, specific S15-rpsO mRNA interactions can also be found, probably with A(-46) in loop L1 of the pseudoknot, demonstrating that mimicry between the two targets is limited.}, note = {0950-382x Journal Article}, keywords = {16S/chemistry/genetics/*metabolism Recombinant Fusion Proteins/metabolism Ribosomal Proteins/chemistry/*genetics/*metabolism Sequence Alignment Support, Bacterial Models, EHRESMANN Amino Acid Sequence Base Sequence Escherichia coli Proteins/chemistry/genetics/*metabolism Gene Expression Regulation, Messenger/metabolism RNA, Molecular *Molecular Mimicry Molecular Sequence Data Mutagenesis, Non-U.S. Gov't Support, P.H.S., Ribosomal, Secondary RNA, Site-Directed Nucleic Acid Conformation Protein Structure, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Internal ribosome entry site structural motifs conserved among mammalian fibroblast growth factor 1 alternatively spliced mRNAs}, author = {Y Martineau and C Le Bec and L Monbrun and V Allo and I M Chiu and O Danos and H Moine and H Prats and A C Prats}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15314170}, isbn = {15314170}, year = {2004}, date = {2004-01-01}, journal = {Mol Cell Biol}, volume = {24}, number = {17}, pages = {7622-7635}, abstract = {Fibroblast growth factor 1 (FGF-1) is a powerful angiogenic factor whose gene structure contains four promoters, giving rise to a process of alternative splicing resulting in four mRNAs with alternative 5' untranslated regions (5' UTRs). Here we have identified, by using double luciferase bicistronic vectors, the presence of internal ribosome entry sites (IRESs) in the human FGF-1 5' UTRs, particularly in leaders A and C, with distinct activities in mammalian cells. DNA electrotransfer in mouse muscle revealed that the IRES present in the FGF-1 leader A has a high activity in vivo. We have developed a new regulatable TET OFF bicistronic system, which allowed us to rule out the possibility of any cryptic promoter in the FGF-1 leaders. FGF-1 IRESs A and C, which were mapped in fragments of 118 and 103 nucleotides, respectively, are flexible in regard to the position of the initiation codon, making them interesting from a biotechnological point of view. Furthermore, we show that FGF-1 IRESs A of murine and human origins show similar IRES activity profiles. Enzymatic and chemical probing of the FGF-1 IRES A RNA revealed a structural domain conserved among mammals at both the nucleotide sequence and RNA structure levels. The functional role of this structural motif has been demonstrated by point mutagenesis, including compensatory mutations. These data favor an important role of IRESs in the control of FGF-1 expression and provide a new IRES structural motif that could help IRES prediction in 5' UTR databases.}, note = {0270-7306 Journal Article}, keywords = {EHRESMANN *5' Untranslated Regions *Alternative Splicing Animals Base Sequence Cell Line Fibroblast Growth Factor 1/*genetics Gene Transfer Techniques Genes, Messenger/chemistry/*genetics/metabolism Ribosomes/*metabolism Sequence Alignment Support, Non-U.S. Gov't, Site-Directed *Nucleic Acid Conformation *Promoter Regions (Genetics) RNA, Skeletal/cytology/physiology Mutagenesis, Structural/genetics Genetic Vectors Human Mice Molecular Sequence Data Muscle, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Single amino acid changes in AspRS reveal alternative routes for expanding its tRNA repertoire in vivo}, author = {F Martin and S Barends and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15289581}, isbn = {15289581}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {13}, pages = {4081-4089}, abstract = {Aminoacyl-tRNA synthetases (aaRSs) are enzymes that are highly specific for their tRNA substrates. Here, we describe the expansion of a class IIb aaRS-tRNA specificity by a genetic selection that involves the use of a modified tRNA displaying an amber anticodon and the argE(amber) and lacZ(amber) reporters. The study was performed on Escherichia coli aspartyl-tRNA synthetase (AspRS) and amber tRNA(Asp). Nine AspRS mutants able to charge the amber tRNA(Asp) and to suppress the reporter genes were selected from a randomly mutated library. All the mutants exhibited a new amber tRNA(Asp) specificity in addition to the initial native tRNA(Asp). Six mutations were found in the anticodon-binding site located in the N-terminal OB-fold. The strongest suppressor was a mutation of residue Glu-93 that contacts specifically the anticodon nucleotide 34 in the crystal structure. The other mutations in the OB-fold were found at close distance from the anticodon in the so-called loop L45 and strand S1. They concern residues that do not contact tRNA(Asp) in the native complex. In addition, this study shows that suppressors can carry mutations located far from the anticodon-binding site. One such mutation was found in the synthetase hinge-module where it increases the tRNA(Asp)-charging rate, and two other mutations were found in the prokaryotic-specific insertion domain and the catalytic core. These mutants seem to act by indirect effects on the tRNA acceptor stem binding and on the conformation of the active site of the enzyme. Altogether, these data suggest the existence of various ways for modifying the mechanism of tRNA discrimination.}, note = {1362-4962 Journal Article}, keywords = {Amino Acid Substitution Anticodon/metabolism Aspartate-tRNA Ligase/*chemistry/genetics/*metabolism Aspartic Acid/metabolism Binding Sites Models, Amino Acyl/chemistry/*metabolism Support, ERIANI, Molecular Mutation Phenotype Protein Engineering Protein Structure, Non-U.S. Gov't, Tertiary RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effects of vaccine strain mutations in domain V of the internal ribosome entry segment compared in the wild type poliovirus type 1 context}, author = {C E Malnou and A Werner and A M Borman and E Westhof and K M Kean}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14672927}, isbn = {14672927}, year = {2004}, date = {2004-01-01}, journal = {J Biol Chem}, volume = {279}, number = {11}, pages = {10261-10269}, abstract = {Initiation of poliovirus (PV) protein synthesis is governed by an internal ribosome entry segment structured into several domains including domain V, which is accepted to be important in PV neurovirulence because it harbors an attenuating mutation in each of the vaccine strains developed by A. Sabin. To better understand how these single point mutations exert their effects, we placed each of them into the same genomic context, that of PV type 1. Only the mutation equivalent to the Sabin type 3 strain mutation resulted in significantly reduced viral growth both in HeLa and neuroblastoma cells. This correlated with poor translation efficiency in vitro and could be explained by a structural perturbation of the domain V of the internal ribosome entry segment, as evidenced by RNA melting experiments. We demonstrated that reduced cell death observed during infection by this mutant is due to the absence of inhibition of host cell translation. We confirmed that this shut-off is correlated principally with cleavage of eIF4GII and not eIF4GI and that this cleavage is significantly impaired in the case of the defective mutant. These data support the previously reported conclusion that the 2A protease has markedly different affinities for the two eIF4G isoforms.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence Blotting, Genetic, Messenger/metabolism Ribosomes/*genetics Support, Non-U.S. Gov't Translation, Tertiary RNA/chemistry RNA, Unité ARN, Viral Electrophoresis, Western DNA, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {A pathogenesis-associated mutation in human mitochondrial tRNALeu(UUR) leads to reduced 3'-end processing and CCA addition}, author = {L Levinger and I Oestreich and C Florentz and M Morl}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15019775}, isbn = {15019775}, year = {2004}, date = {2004-01-01}, journal = {J Mol Biol}, volume = {337}, number = {3}, pages = {535-544}, abstract = {Point mutations in mitochondrial tRNAs can cause severe multisystemic disorders such as mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes (MELAS) and myoclonus epilepsy with ragged-red fibers (MERRF). Some of these mutations impair one or more steps of tRNA maturation and protein biosynthesis including 5'-end-processing, post-transcriptional base modification, structural stability, aminoacylation, and formation of tRNA-ribosomal complexes. tRNALeu(UUR), an etiologic hot spot for such diseases, harbors 20 of more than 90 disease-associated mutations described to date. Here, the pathogenesis-associated base substitutions A3243G, T3250C, T3271C, A3302G and C3303T within this tRNA were tested for their effects on endonucleolytic 3'-end processing and CCA addition at the tRNA 3'-terminus. Whereas mutations A3243G, A3302G and C3303T reduced the efficiency of 3'-end cleavage, only the C3303T substitution was a less efficient substrate for CCA addition. These results support the view that pathogenesis may be elicited through cumulative effects of tRNA mutations: a mutation can impede several pre-tRNA processing steps, with each such reduction contributing to the overall impairment of tRNA function.}, note = {0022-2836 Journal Article}, keywords = {FLORENTZ, FLORENTZ Human Kinetics Mitochondrial Diseases/*genetics Nucleic Acid Conformation *Point Mutation RNA/*genetics/physiology *RNA 3' End Processing RNA Nucleotidyltransferases/metabolism RNA, Leu/*genetics/physiology Support, Non-U.S. Gov't Support, P.H.S., Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mitochondrial tRNA 3' end metabolism and human disease}, author = {L Levinger and M Mörl and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15477393}, isbn = {15477393}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {18}, pages = {5430-5441}, abstract = {Over 150 mutations in the mitochondrial genome have been shown to be associated with human disease. Remarkably, two-thirds of them are found in tRNA genes, which constitute only one-tenth of the mitochondrial genome. A total of 22 tRNAs punctuate the genome and are produced together with 11 mRNAs and 2 rRNAs from long polycistronic primary transcripts with almost no spacers. Pre-tRNAs thus require precise endonucleolytic excision. Furthermore, the CCA triplet which forms the 3' end of all tRNAs is not encoded, but must be synthesized by the CCA-adding enzyme after 3' end cleavage. Amino acid attachment to the CCA of mature tRNA is performed by aminoacyl-tRNA synthetases, which, like the preceding processing enzymes, are nuclear-encoded and imported into mitochondria. Here, we critically review the effectiveness and reliability of evidence obtained from reactions with in vitro transcripts that pathogenesis-associated mutant mitochondrial tRNAs can lead to deficiencies in tRNA 3' end metabolism (3' end cleavage, CCA addition and aminoacylation) toward an understanding of molecular mechanisms underlying human tRNA disorders. These defects probably contribute, individually and cumulatively, to the progression of human mitochondrial diseases.}, note = {1362-4962 Journal Article Review Review, Tutorial}, keywords = {FLORENTZ, Non-U.S. Gov't Support, P.H.S., Transfer/chemistry/*genetics/*metabolism Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Architecture of a Diels-Alderase ribozyme with a preformed catalytic pocket}, author = {S Keiper and D Bebenroth and B Seelig and E Westhof and A Jaschke}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15380182}, isbn = {15380182}, year = {2004}, date = {2004-01-01}, journal = {Chem Biol}, volume = {11}, number = {9}, pages = {1217-1227}, abstract = {Artificial ribozymes catalyze a variety of chemical reactions. Their structures and reaction mechanisms are largely unknown. We have analyzed a ribozyme catalyzing Diels-Alder cycloaddition reactions by comprehensive mutation analysis and a variety of probing techniques. New tertiary interactions involving base pairs between nucleotides of the 5' terminus and a large internal loop forming a pseudoknot fold were identified. The probing data indicate a preformed tertiary structure that shows no major changes on substrate or product binding. Based on these observations, a molecular architecture featuring a Y-shaped arrangement is proposed. The tertiary structure is formed in a rather unusual way; that is, the opposite sides of the asymmetric internal loop are clamped by the four 5'-terminal nucleotides, forming two adjacent two base-pair helices. It is proposed that the catalytic pocket is formed by a wedge within one of these helices.}, note = {1074-5521 Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {111th ENMC International Workshop on Multi-minicore Disease. 2nd International MmD Workshop, 9-11 November 2002, Naarden, The Netherlands.}, author = {H Jungbluth and A Beggs and C Bonnemann and K Bushby and C Ceuterick-de Groote and B Estournet-Mathiaud and N Goemans and P Guicheney and A Lescure and J Lunardi and F Muntoni and R Quinlivan and C Sewry and V Straub and S Treves and A Ferreiro}, url = {http://www.ncbi.nlm.nih.gov/pubmed/15482962}, isbn = {15482962}, year = {2004}, date = {2004-01-01}, journal = {Neuromuscul Disord}, volume = {14}, number = {11}, pages = {754-766}, note = {0960-8966 (Print) Congresses Research Support, Non-U.S. Gov't}, keywords = {Differential Humans Muscle Fibers/metabolism/*pathology Muscle Proteins/genetics/metabolism Muscle, KROL Diagnosis, LESCURE, Skeletal/metabolism/*pathology/physiopathology Muscular Diseases/classification/genetics/metabolism/*pathology Mutation Ryanodine Receptor Calcium Release Channel/genetics/metabolism Selenoproteins, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{montagnani_cg-rel_2004, title = {Cg-Rel, the first Rel/NF-kappaB homolog characterized in a mollusk, the Pacific oyster Crassostrea gigas.}, author = {C Montagnani and Christine Kappler and Jean-Marc Reichhart and J M Escoubas}, year = {2004}, date = {2004-01-01}, journal = {FEBS Lett.}, volume = {561}, pages = {75--82}, abstract = {We report here the identification and functional characterization of Cg-Rel, a gene encoding the Crassostrea gigas homolog of Rel/NF-kappaB transcription factors found in insects and mammals. Sequence and phylogenetic analysis showed that Cg-Rel shares the structural organization of Rel/NF-kappaB transcription factors of class II. It includes a Rel homology domain as well as a C-terminal transactivation domain (TD). Overexpression of Cg-Rel in the Drosophila S2 cell line activated the expression of a NF-kappaB-dependent reporter gene, whereas transfection with a Cg-Rel construct containing a C-terminal deletion of the TD or using a reporter gene with mutated kappaB binding sites failed to activate expression. These results suggest that Cg-Rel is a functional member of the Rel family of transcription factors, making this the sixth structurally homologous component of the Rel/NF-kappaB pathway characterized in C. gigas. Based on homology to other invertebrates' Rel/NF-kappaB cascade, the function of the oyster pathway may serve to regulate genes involved in innate defense and/or development. These findings serve to highlight a potentially important regulatory pathway to the study of oyster immunology, hence allowing comparison of the immune system in vertebrates and invertebrates, an important key issue to understand its evolution.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{, title = {Results and prospects of the yeast three-hybrid system}, author = {S Jaeger and G Eriani and F Martin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14706817}, isbn = {14706817}, year = {2004}, date = {2004-01-01}, journal = {FEBS Lett}, volume = {556}, number = {1-3}, pages = {7-12}, abstract = {In 1996, a new method, termed the yeast three-hybrid system, dedicated to selection of RNA binding proteins using a hybrid RNA molecule as bait was described. In this minireview, we summarize the results that have been obtained using this method. Indeed, approximately 20 unknown proteins have been characterized so far. The three-hybrid strategy has also been used as a tool to dissect RNA-protein interactions. The example of such a study on human histone HBP interaction with its target mRNA is described. Problems that can be encountered are addressed in a troubleshooting section. Especially, our results with tRNA binding proteins are discussed.}, note = {0014-5793 Journal Article Review Review Literature}, keywords = {ERIANI, Histones/genetics Human RNA/chemistry/genetics/metabolism RNA-Binding Proteins/chemistry/genetics/metabolism Saccharomyces cerevisiae/*genetics Support, Non-U.S. Gov't *Two-Hybrid System Techniques, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{reichhart_guest_2004, title = {Guest Editor of the Special Issue on Innate Immunity.}, author = {Jean-Marc Reichhart}, year = {2004}, date = {2004-01-01}, journal = {Mol. Immunol.}, volume = {40}, pages = {843}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{, title = {Critical residues for RNA discrimination of the histone hairpin binding protein (HBP) investigated by the yeast three-hybrid system}, author = {S Jaeger and G Eriani and F Martin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14706861}, isbn = {14706861}, year = {2004}, date = {2004-01-01}, journal = {FEBS Lett}, volume = {556}, number = {1-3}, pages = {265-270}, abstract = {The histone hairpin binding protein (HBP, also called SLBP, which stands for stem-loop binding protein) binds specifically to a highly conserved hairpin structure located in the 3' UTR of the cell-cycle-dependent histone mRNAs. HBP consists of a minimal central RNA binding domain (RBD) flanked by an N- and C-terminal domain. The yeast three-hybrid system has been used to investigate the critical residues of the human HBP involved in the binding of its target hairpin structure. By means of negative selections followed by positive selections, we isolated mutant HBP species. Our results indicate tight relationships between the RBD and the N- and C-terminal domains.}, note = {0014-5793 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Amino Acid Substitution Base Sequence Carrier Proteins/chemistry/*genetics/*metabolism Histones/genetics Human Lac Operon/genetics Molecular Sequence Data Nucleic Acid Conformation Nucleotides/chemistry/genetics/metabolism RNA/chemistry/genetics/*metabolism RNA, ERIANI, Messenger/chemistry/genetics/metabolism RNA-Binding Proteins/chemistry/genetics/metabolism Recombinant Fusion Proteins/chemistry/genetics/metabolism Saccharomyces cerevisiae/*genetics Sequence Alignment Sequence Homology, Non-U.S. Gov't Two-Hybrid System Techniques beta-Galactosidase/genetics/metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_reponse_2004b, title = {La réponse immunitaire chez la Drosophile}, author = {Dominique Ferrandon and Julien Royet}, year = {2004}, date = {2004-01-01}, journal = {Regards sur la biochimie}, volume = {1}, pages = {27--33}, keywords = {ferrandon, M3i}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Detection and quantification of modified nucleotides in RNA using thin-layer chromatography}, author = {H Grosjean and G Keith and L Droogmans}, editor = {J M Gott}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15103084}, doi = {10.1385/1-59259-775-0:357}, isbn = {15103084}, year = {2004}, date = {2004-01-01}, booktitle = {RNA Interference, Editing, and Modification: Methods and Protocols}, volume = {265}, pages = {357-391}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, abstract = {Identification of a modified nucleotide and its localization within an RNA molecule is a difficult task. Only direct sequencing of purified RNA molecules and high-performance liquid chromatography mass spectrometry analysis of purified RNA fragments allow determination of both the type and location of a given modified nucleotide within an RNA of 50-150 nt in length. The objective of this chapter is to describe in detail a few simple procedures that we have found particularly suited for the detection, localization, and quantification of modified nucleotides within an RNA of known sequence. The methods can also be used to reveal the enzymatic activity of a particular RNA-modifying enzyme in vitro or in vivo. The procedures are based on the use of radiolabeled RNA (with [32P], [14C], or [3H]) or [32P]-postlabeled oligonucleotides and two-dimensional thin-layer chromatography of labeled nucleotides on cellulose plates. This chapter provides useful maps of the migration characteristics of 70 modified nucleotides on thin-layer cellulose plates.}, keywords = {KEITH 5' Untranslated Regions/chemistry Base Composition Chromatography, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{ferrandon_sensing_2004b, title = {Sensing infection in Drosophila: Toll and beyond}, author = {Dominique Ferrandon and Jean-Luc Imler and Jules A Hoffmann}, issn = {1044-5323}, year = {2004}, date = {2004-01-01}, journal = {Semin Immunol}, volume = {16}, pages = {43--53}, abstract = {Drosophila has evolved a potent immune system that is somewhat adapted to the nature of infections through the selective activation of either one of two NF-kappa B-like signalling pathways, the Toll and IMD (Immune deficiency) pathways. In contrast to the mammalian system, the Toll receptor does not act as a pattern recognition receptor (PRR) but as a cytokine receptor. The sensing of microbial infections is achieved by at least four PRRs that belong to two distinct families: the peptidoglycan recognition proteins (PGRPs) and the Gram-negative binding proteins (GNBPs)/beta-glucan recognition proteins (beta GRPs).}, keywords = {Animals, Carrier Proteins/chemistry/immunology/physiology, Cell Surface/immunology/*physiology, Drosophila Proteins/chemistry/immunology/*physiology, Drosophila/genetics/*immunology/microbiology, ferrandon, Fungi/immunology, Gene Expression Regulation, Gram-Negative Bacterial Infections/immunology, Gram-Positive Bacterial Infections/immunology, hoffmann, imler, Immunological, Insect Proteins/chemistry/immunology/physiology, M3i, Models, Non-U.S. Gov't, Receptors, Signal Transduction/immunology/physiology, Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural variability of the initiation complex of HIV-1 reverse transcription}, author = {V Goldschmidt and J C Paillart and M Rigourd and B Ehresmann and A M Aubertin and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15194685}, isbn = {15194685}, year = {2004}, date = {2004-01-01}, journal = {J Biol Chem}, volume = {279}, number = {34}, pages = {35923-35931}, abstract = {HIV-1 reverse transcription is initiated from a tRNA(3)(Lys) molecule annealed to the viral RNA at the primer binding site (PBS), but the structure of the initiation complex of reverse transcription remains controversial. Here, we performed in situ structural probing, as well as in vitro structural and functional studies, of the initiation complexes formed by highly divergent isolates (MAL and NL4.3/HXB2). Our results show that the structure of the initiation complex is not conserved. In MAL, and according to sequence analysis in 14% of HIV-1 isolates, formation of the initiation complex is accompanied by complex rearrangements of the viral RNA, and extensive interactions with tRNA(3)(Lys) are required for efficient initiation of reverse transcription. In NL4.3, HXB2, and most isolates, tRNA(3)(Lys) annealing minimally affects the viral RNA structure and no interaction outside the PBS is required for optimal initiation of reverse transcription. We suggest that in MAL, extensive interactions with tRNA(3)(Lys) are required to drive the structural rearrangements generating the structural elements ultimately recognized by reverse transcriptase. In NL4.3 and HXB2, these elements are already present in the viral RNA prior to tRNA(3)(Lys) annealing, thus explaining that extensive interactions with the primer are not required. Interestingly, such interactions are required in HXB2 mutants designed to use a non-cognate tRNA as primer (tRNA(His)). In the latter case, the extended interactions are required to counteract a negative contribution associate with the alternate primer.}, note = {0021-9258 Journal Article}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{imler_biology_2004, title = {Biology of Toll receptors: lessons from insects and mammals}, author = {Jean-Luc Imler and Liangbiao Zheng}, doi = {10.1189/jlb.0403160}, issn = {0741-5400}, year = {2004}, date = {2004-01-01}, journal = {Journal of Leukocyte Biology}, volume = {75}, number = {1}, pages = {18--26}, abstract = {Toll receptors are type I transmembrane proteins that play important roles in development and immunity in animals. Comparison of the genomes of mouse and human on one side and of the fruitfly Drosophila and the mosquito Anopheles (two dipteran insects) on the other, revealed that the four species possess a similar number of Toll receptors (approximately 10). However, phylogenetic analyses indicate that the families of Toll receptors expanded independently in insects and mammals. We review recent results on these receptors, which point to differences in the activation and signaling between Tolls in insects and Toll-like receptors (TLRs) in mammals. Whereas mammalian TLRs appear to be solely dedicated to host-defense, insect Tolls may be predominantly linked to other functions, probably developmental.}, keywords = {Animals, Anopheles, Cell Surface, Humans, imler, M3i, Membrane Glycoproteins, Mice, Phylogeny, Plant Physiological Phenomena, Receptors, Signal Transduction, Toll-Like Receptor 5, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{, title = {Primer unblocking by HIV-1 reverse transcriptase and resistance to nucleoside RT inhibitors (NRTIs)}, author = {V Goldschmidt and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15183338}, isbn = {15183338}, year = {2004}, date = {2004-01-01}, journal = {Int J Biochem Cell Biol}, volume = {36}, number = {9}, pages = {1687-1705}, abstract = {During zidovudine and stavudine treatment, HIV-1 selects several mutations (thymidine-associated mutations, TAMs) in the reverse transcriptase gene that confer high- and moderate-levels of resistance, respectively, to these nucleoside reverse transcriptase inhibitors (NRTIs). The mechanism of the resistance provided by these mutations has long remained elusive. However, recent data showed that ATP-phosphorolysis, a reaction analogous to pyrophosphorolysis (the reverse of the nucleotide incorporation reaction) in which ATP is the pyrophosphate donor, is central to this mechanism by allowing repair of the chain-terminated primer. A detailed structural and mechanistic model accounting for the specificity of the ATP-phosphorolysis and its inhibition by the next complementary nucleotide is now available. In the context of multiresistant viruses, the TAMs are also associated with resistance to abacavir, and to a lesser extent to didanisone, zalcitabine and tenofovir. When associated with the TAMs, a dipeptide insertion in the fingers of reverse transcriptase increases the ATP-phosphorolysis of most chain terminators, stressing the increasing importance of this mechanism. However, some non-nucleoside reverse transcriptase inhibitors (NNRTIs) inhibit this process. In addition, point mutations conferring resistance to NNRTIs (Y181C and L100I) or NRTIs (K65R, L74V, and M184V) partially resensitize the resistant viruses to AZT by inhibiting ATP-phosphorolysis. These findings allow rationalizing the beneficial effects of some drug combinations and should contribute to improve drug cocktails. The development of NRTIs that would not allow the ATP-mediated excision to take place should prove beneficial for future treatments, even though high-level resistance to multiple NRTIs can ultimately develop in the absence of any significant primer unblocking.}, note = {1357-2725 Journal Article}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{de_marco_solubility_2004, title = {The solubility and stability of recombinant proteins are increased by their fusion to NusA}, author = {Valeria De Marco and Gunter Stier and Stephanie A Blandin and Ario de Marco}, doi = {10.1016/j.bbrc.2004.07.189}, issn = {0006-291X}, year = {2004}, date = {2004-01-01}, journal = {Biochem. Biophys. Res. Commun.}, volume = {322}, number = {3}, pages = {766--771}, abstract = {The new bacterial vector pETM60 enables the expression of His-tagged recombinant proteins fused to the C-terminus of NusA through a TEV protease recognition sequence. Three sequences coding for two protein domains (Xklp3A and Tep3Ag) and one membrane-bound viral protein (E8R) could not be expressed in a soluble form in bacteria. Their GST-fusions were mostly soluble but quickly degraded during purification. The same sequences cloned in pETM60 were efficiently purified by metal affinity and recovered soluble after the removal of the fusion partner. The NusA-fused constructs enabled to yield 13-20mg of fusion protein per litre of culture and 2.5-5mg of pure protein per litre of culture. Structural analysis indicated that the purified proteins were monodispersed and correctly folded. NusA has been used to raise antibodies that have been successfully used for Western blot and immunoprecipitation of NusA fusion proteins.}, keywords = {blandin, Drug Stability, Escherichia coli Proteins, Genetic Vectors, Glutathione Transferase, Kinetics, M3i, Oxidation-Reduction, Peptide Elongation Factors, Recombinant Fusion Proteins, Recombinant Proteins, Solubility, Transcription Factors, Transcriptional Elongation Factors}, pubstate = {published}, tppubtype = {article} } @article{, title = {IgG reactivity with a 100-kDa tissue and endothelial cell antigen identified as topoisomerase 1 distinguishes between limited and diffuse systemic sclerosis patients}, author = {P Garcia de la Pena-Lefebvre and Y Chanseaud and M C Tamby and J Reinbolt and F Batteux and Y Allanore and A Kahan and O Meyer and O Benveniste and O Boyer and L Guillevin and M C Boissier and L Mouthon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15183145}, isbn = {15183145}, year = {2004}, date = {2004-01-01}, journal = {Clin Immunol}, volume = {111}, number = {3}, pages = {241-251}, abstract = {We have analyzed antibody (Ab) reactivities of patients with limited systemic sclerosis (SSc) and anti-centromere Ab, patients with diffuse SSc and anti-topoisomerase 1 (anti-topo 1) Ab, patients with diffuse SSc without anti-topo 1 or anti-centromere Ab and age- and gender-matched healthy controls with normal human tissue and endothelial cell (EC) antigens. IgG reactivities with tissue antigens differed significantly between patients with anti-topo 1 Ab and patients with anti-centromere Ab. One 100-kDa band identified as topoisomerase 1 in macrovascular and microvascular EC extracts was recognized by IgG from patients with anti-topo 1 Ab and 50% of patients without specific Ab. IgG from patients with limited SSc and anti-centromere Ab, but not those of other patients or controls specifically recognized a 80-kDa band only in microvascular EC. Our results indicate that Ab from patients with limited or diffuse SSc with or without anti-topo 1 Ab exhibit specific and mutually exclusive reactivity patterns.}, note = {1521-6616 Journal Article}, keywords = {EHRESMANN Autoantibodies/*analysis Blotting, Non-U.S. Gov't, Polyacrylamide Gel Endothelial Cells/*immunology Enzyme-Linked Immunosorbent Assay Female Human Immunoglobulin G/analysis Immunoglobulin M/analysis Male Middle Aged Scleroderma, Systemic/*immunology Support, Type I/*immunology Electrophoresis, Unité ARN, Western Centromere/immunology DNA Topoisomerases}, pubstate = {published}, tppubtype = {article} } @article{blandin_complement-like_2004, title = {Complement-like protein TEP1 is a determinant of vectorial capacity in the malaria vector Anopheles gambiae}, author = {Stephanie A Blandin and Shin-Hong Shiao and Luis F Moita and Chris J Janse and Andrew P Waters and Fotis C Kafatos and Elena A Levashina}, issn = {0092-8674}, year = {2004}, date = {2004-01-01}, journal = {Cell}, volume = {116}, number = {5}, pages = {661--670}, abstract = {Anopheles mosquitoes are major vectors of human malaria in Africa. Large variation exists in the ability of mosquitoes to serve as vectors and to transmit malaria parasites, but the molecular mechanisms that determine vectorial capacity remain poorly understood. We report that the hemocyte-specific complement-like protein TEP1 from the mosquito Anopheles gambiae binds to and mediates killing of midgut stages of the rodent malaria parasite Plasmodium berghei. The dsRNA knockdown of TEP1 in adults completely abolishes melanotic refractoriness in a genetically selected refractory strain. Moreover, in susceptible mosquitoes this knockdown increases the number of developing parasites. Our results suggest that the TEP1-dependent parasite killing is followed by a TEP1-independent clearance of dead parasites by lysis and/or melanization. Further elucidation of the molecular mechanisms of TEP1-mediated parasite killing will be of great importance for our understanding of the principles of vectorial capacity in insects.}, keywords = {Animals, Anopheles, blandin, Female, Genetic, Humans, Insect Proteins, Insect Vectors, M3i, Malaria, Models, Molecular, Plasmodium berghei, Polymorphism, Protein Structure, RNA, Sequence Alignment, Tertiary}, pubstate = {published}, tppubtype = {article} } @article{, title = {Antibacterial aminoglycosides with a modified mode of binding to the ribosomal-RNA decoding site}, author = {B Francois and J Szychowski and S S Adhikari and K Pachamuthu and E E Swayze and R H Griffey and M T Migawa and E Westhof and S Hanessian}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15593140}, isbn = {15593140}, year = {2004}, date = {2004-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {43}, number = {48}, pages = {6735-6738}, note = {0570-0833 Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{blandin_mosquito_2004, title = {Mosquito immune responses against malaria parasites}, author = {Stéphanie A Blandin and Elena A Levashina}, issn = {0952-7915}, year = {2004}, date = {2004-01-01}, journal = {Curr. Opin. Immunol.}, volume = {16}, number = {1}, pages = {16--20}, abstract = {Anopheline mosquitoes are the major vectors of human malaria. Mosquito-parasite interactions are a critical aspect of disease transmission and a potential target for malaria control. Mosquitoes vary in their innate ability to support development of the malaria parasite, but the molecular mechanisms that determine vector competence are poorly understood. This area of research has been revolutionized by recent advances in the mosquito genome characterization and by the development of new tools for functional gene analysis.}, keywords = {Animals, Anopheles, blandin, Gene Library, Genes, Hemocytes, Host-Parasite Interactions, Immunity, Innate, Insect, Insect Vectors, M3i, Malaria, Plasmodium}, pubstate = {published}, tppubtype = {article} } @article{, title = {Functional idiosyncrasies of tRNA isoacceptors in cognate and noncognate aminoacylation systems}, author = {A Fender and M Sissler and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14987797}, isbn = {14987797}, year = {2004}, date = {2004-01-01}, journal = {Biochimie}, volume = {86}, number = {1}, pages = {21-29}, abstract = {The specificity of transfer RNA aminoacylation by cognate aminoacyl-tRNA synthetase is a crucial step for synthesis of functional proteins. It is established that the aminoacylation identity of a single tRNA or of a family of tRNA isoacceptors is linked to the presence of positive signals (determinants) allowing recognition by cognate synthetases and negative signals (antideterminants) leading to rejection by the noncognate ones. The completion of identity sets was generally tested by transplantation of the corresponding nucleotides into one or several host tRNAs which acquire as a consequence the new aminoacylation specificities. Such transplantation experiments were also useful to detect peculiar structural refinements required for optimal expression of a given aminoacylation identity set within a host tRNA. This study explores expression of the defined yeast aspartate identity set into different tRNA scaffolds of a same specificity, namely the four yeast tRNA(Arg) isoacceptors. The goal was to investigate whether expression of the new identity is similar due to the unique specificity of the host tRNAs or whether it is differently expressed due to their peculiar sequences and structural features. In vitro transcribed native tRNA(Arg) isoacceptors and variants bearing the aspartate identity elements were prepared and their aminoacylation properties established. The four wild-type isoacceptors are active in arginylation with catalytic efficiencies in a 20-fold range and are inactive in aspartylation. While transplanted tRNA(1)(Arg) and tRNA(4)(Arg) are converted into highly efficient substrates for yeast aspartyl-tRNA synthetase, transplanted tRNA(2)(Arg) and tRNA(3)(Arg) remain poorly aspartylated. Search for antideterminants in these two tRNAs reveals idiosyncratic features. Conversion of the single base-pair C6-G67 into G6-C67, the pair present in tRNA(Asp), allows full expression of the aspartate identity in the transplanted tRNA(2)(Arg), but not in tRNA(3)(Arg). It is concluded that the different isoacceptor tRNAs protect themselves from misaminoacylation by idiosyncratic pathways of antidetermination.}, note = {0300-9084 Journal Article}, keywords = {Amino Acyl/genetics/*metabolism Saccharomyces cerevisiae Substrate Specificity/genetics/physiology Support, Chemical Molecular Sequence Data Mutation Nucleic Acid Conformation Protein Binding/physiology RNA, FLORENTZ, GIEGE FLORENTZ Amino Acid Activation/physiology Amino Acyl-tRNA Ligases/*metabolism Anticodon Bacterial Proteins/metabolism Base Sequence Cloning, Molecular Computer Simulation Escherichia coli Models, Non-U.S. Gov't Thermus thermophilus, SISSLER, Transfer, Transfer/genetics/*metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{imler_toll-dependent_2004, title = {Toll-dependent and Toll-independent immune responses in Drosophila}, author = {Jean-Luc Imler and Dominique Ferrandon and Julien Royet and Jean-Marc Reichhart and Charles Hetru and Jules A Hoffmann}, doi = {10.1179/096805104225005887}, issn = {0968-0519}, year = {2004}, date = {2004-01-01}, journal = {Journal of Endotoxin Research}, volume = {10}, number = {4}, pages = {241--246}, abstract = {The multifaceted response of the fruitfly Drosophila melanogaster to infection by a wide range of microbes is complex and remarkably efficient. Its most prominent aspect is the immune-inducible expression of a set of potent antimicrobial peptides. Genetic analysis of the regulation of the genes encoding these peptides has led to the identification of the receptor Toll as an essential component of the fly's host defense system. In addition, these studies have revealed that the response to Gram-negative bacterial infections involves Toll-independent mechanisms, and that the sensing of infection involves two structurally distinct sets of molecules--the PGRPs and the GNBPs/betaGRPs.}, keywords = {Acute-Phase Proteins, Animals, Blood Proteins, Cell Surface, ferrandon, hoffmann, imler, Insect Proteins, M3i, Membrane Glycoproteins, Receptors, reichhart, Toll-Like Receptor 5, Toll-Like Receptors, Up-Regulation}, pubstate = {published}, tppubtype = {article} } @article{, title = {A yeast arginine specific tRNA is a remnant aspartate acceptor}, author = {A Fender and R Geslain and G Eriani and R Giege and M Sissler and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15452274}, isbn = {15452274}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {17}, pages = {5076-5086}, abstract = {High specificity in aminoacylation of transfer RNAs (tRNAs) with the help of their cognate aminoacyl-tRNA synthetases (aaRSs) is a guarantee for accurate genetic translation. Structural and mechanistic peculiarities between the different tRNA/aaRS couples, suggest that aminoacylation systems are unrelated. However, occurrence of tRNA mischarging by non-cognate aaRSs reflects the relationship between such systems. In Saccharomyces cerevisiae, functional links between arginylation and aspartylation systems have been reported. In particular, it was found that an in vitro transcribed tRNAAsp is a very efficient substrate for ArgRS. In this study, the relationship of arginine and aspartate systems is further explored, based on the discovery of a fourth isoacceptor in the yeast genome, tRNA4Arg. This tRNA has a sequence strikingly similar to that of tRNAAsp but distinct from those of the other three arginine isoacceptors. After transplantation of the full set of aspartate identity elements into the four arginine isoacceptors, tRNA4Arg gains the highest aspartylation efficiency. Moreover, it is possible to convert tRNA4Arg into an aspartate acceptor, as efficient as tRNAAsp, by only two point mutations, C38 and G73, despite the absence of the major anticodon aspartate identity elements. Thus, cryptic aspartate identity elements are embedded within tRNA4Arg. The latent aspartate acceptor capacity in a contemporary tRNAArg leads to the proposal of an evolutionary link between tRNA4Arg and tRNAAsp genes.}, note = {1362-4962 Journal Article}, keywords = {Arg/*chemistry/genetics/metabolism RNA, Asp/*chemistry/genetics/metabolism Saccharomyces cerevisiae/*genetics Sequence Alignment Support, ERIANI, ERIANI FLORENTZ GIEGE Aspartic Acid/metabolism Base Sequence *Evolution, FLORENTZ, Fungal/*chemistry/genetics/metabolism RNA, Molecular Molecular Sequence Data Point Mutation RNA, Non-U.S. Gov't, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular mimicry in translational regulation: the case of ribosomal protein S15.}, author = {C Ehresmann and B Ehresmann and E Ennifar and P Dumas and M Garber and N Mathy and A Nikulin and C Portier and D Patel and A Serganov}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17194931}, isbn = {17194931}, year = {2004}, date = {2004-01-01}, journal = {RNA Biol}, volume = {1}, number = {1}, pages = {66-73}, abstract = {Ribosomal protein S15 is highly conserved among prokaryotes. It plays a pivotal role in the assembly of the central domain of the small ribosomal subunit and regulates its own expression by a feedback mechanism at the translational level. The protein recognizes two RNA targets (rRNA and mRNA) that share only partial similarity. Its interaction with 16S rRNA has been fully characterized, while mRNA interactions and regulatory mechanisms have been extensively studied in E. coli and in T. thermophilus. Recently, we have characterized which aminoacids are involved in E. coli mRNA recognition, using an in vivo assay allowing to identify S15 mutations affecting the S15-mRNA interactions without altering 30S subunit assembly. Here, we address the following questions: Are common determinants used by S15 to recognize its rRNA and mRNA targets? What is the extent of molecular mimicry? Is the regulatory mechanism conserved? Our results indicate that specific recognition of mRNA and rRNA relies on both mimicry and site differentiation. They also highlight the high plasticity of RNA to adapt to evolutionary constraints.}, note = {Epub 2004 May 5.}, keywords = {DUMAS, ENNIFAR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genome evolution in yeasts}, author = {B Dujon and D Sherman and G Fischer and P Durrens and S Casaregola and I Lafontaine and J De Montigny and C Marck and C Neuveglise and E Talla and N Goffard and L Frangeul and M Aigle and V Anthouard and A Babour and V Barbe and S Barnay and S Blanchin and J M Beckerich and E Beyne and C Bleykasten and A Boisrame and J Boyer and L Cattolico and F Confanioleri and A De Daruvar and L Despons and E Fabre and C Fairhead and H Ferry-Dumazet and A Groppi and F Hantraye and C Hennequin and N Jauniaux and P Joyet and R Kachouri and A Kerrest and R Koszul and M Lemaire and I Lesur and L Ma and H Muller and J M Nicaud and M Nikolski and S Oztas and O Ozier-Kalogeropoulos and S Pellenz and S Potier and G F Richard and M L Straub and A Suleau and D Swennen and F Tekaia and M Wesolowski-Louvel and E Westhof and B Wirth and M Zeniou-Meyer and I Zivanovic and M Bolotin-Fukuhara and A Thierry and C Bouchier and B Caudron and C Scarpelli and C Gaillardin and J Weissenbach and P Wincker and J L Souciet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15229592}, isbn = {15229592}, year = {2004}, date = {2004-01-01}, journal = {Nature}, volume = {430}, number = {6995}, pages = {35-44}, abstract = {Identifying the mechanisms of eukaryotic genome evolution by comparative genomics is often complicated by the multiplicity of events that have taken place throughout the history of individual lineages, leaving only distorted and superimposed traces in the genome of each living organism. The hemiascomycete yeasts, with their compact genomes, similar lifestyle and distinct sexual and physiological properties, provide a unique opportunity to explore such mechanisms. We present here the complete, assembled genome sequences of four yeast species, selected to represent a broad evolutionary range within a single eukaryotic phylum, that after analysis proved to be molecularly as diverse as the entire phylum of chordates. A total of approximately 24,200 novel genes were identified, the translation products of which were classified together with Saccharomyces cerevisiae proteins into about 4,700 families, forming the basis for interspecific comparisons. Analysis of chromosome maps and genome redundancies reveal that the different yeast lineages have evolved through a marked interplay between several distinct molecular mechanisms, including tandem gene repeat formation, segmental duplication, a massive genome duplication and extensive gene loss.}, note = {1476-4687 Journal Article}, keywords = {Chromosomes, Fungal Molecular Sequence Data RNA, Fungal/*genetics *Genome, Fungal/genetics Conserved Sequence/genetics *Evolution, Molecular Gene Duplication Genes, Non-U.S. Gov't Synteny/genetics Tandem Repeat Sequences/genetics Yeasts/*classification/*genetics, Ribosomal/genetics RNA, Transfer/genetics Saccharomyces cerevisiae Proteins/genetics Support, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {An aminoacyl-tRNA synthetase-like protein encoded by the Escherichia coli yadB gene glutamylates specifically tRNAAsp}, author = {D Y Dubois and M Blaise and H D Becker and V Campanacci and G Keith and R Giege and C Cambillau and J Lapointe and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15096594}, isbn = {15096594}, year = {2004}, date = {2004-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {101}, number = {20}, pages = {7530-7535}, abstract = {The product of the Escherichia coli yadB gene is homologous to the N-terminal part of bacterial glutamyl-tRNA synthetases (GluRSs), including the Rossmann fold with the acceptor-binding domain and the stem-contact fold. This GluRS-like protein, which lacks the anticodon-binding domain, does not use tRNA(Glu) as substrate in vitro nor in vivo, but aminoacylates tRNA(Asp) with glutamate. The yadB gene is expressed in wild-type E. coli as an operon with the dksA gene, which encodes a protein involved in the general stress response by means of its action at the translational level. The fate of the glutamylated tRNA(Asp) is not known, but its incapacity to bind elongation factor Tu suggests that it is not involved in ribosomal protein synthesis. Genes homologous to yadB are present only in bacteria, mostly in Proteobacteria. Sequence alignments and phylogenetic analyses show that the YadB proteins form a distinct monophyletic group related to the bacterial and organellar GluRSs (alpha-type GlxRSs superfamily) with ubiquitous function as suggested by the similar functional properties of the YadB homologue from Neisseria meningitidis.}, note = {0027-8424 Journal Article}, keywords = {Asp/*metabolism Support, KERN GIEGE Amino Acyl-tRNA Ligases/chemistry/genetics/*metabolism Crystallography, Messenger/metabolism RNA, Non-U.S. Gov't, Tertiary RNA, Transfer, Unité ARN, X-Ray Escherichia coli/enzymology/genetics/*metabolism Escherichia coli Proteins/chemistry/genetics/*metabolism Glutamic Acid/*metabolism Peptide Elongation Factor 2/metabolism Phylogeny Protein Structure}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization and preliminary X-ray characterization of the atypical glutaminyl-tRNA synthetase from Deinococcus radiodurans}, author = {M A Deniziak and C Sauter and H D Becker and R Giege and D Kern}, url = {http://journals.iucr.org/d/issues/2004/12/02/za0130/za0130bdy.html}, isbn = {15572799}, year = {2004}, date = {2004-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {60}, number = {Pt 12 Pt 2}, pages = {2361-2363}, abstract = {The glutaminyl-tRNA synthetase (GlnRS) from the radiation-resistant bacterium Deinococcus radiodurans differs from known GlnRSs and other tRNA synthetases by the presence of an additional C-terminal domain resembling the C-terminal region of the GatB subunit of tRNA-dependent amidotransferase (AdT). This atypical synthetase was overexpressed in Escherichia coli, purified and crystallized in the presence of PEG 3350. Orthorhombic crystals were obtained that belong to space group P2(1)2(1)2(1) and diffract to 2.3 A resolution. The crystal structure was solved by molecular replacement using the structure of E. coli GlnRS as a search model.}, note = {0907-4449 Journal article}, keywords = {FRUGIER, KERN GIEGE, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {On the use of low-frequency normal modes to enforce collective movements in refining macromolecular structural models}, author = {M Delarue and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15096585}, doi = {10.1073/pnas.0400301101}, isbn = {15096585}, year = {2004}, date = {2004-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {101}, number = {18}, pages = {6957-6962}, abstract = {As more and more structures of macromolecular complexes get solved in different conditions, it has become apparent that flexibility is an inherent part of their biological function. Normal mode analysis using simplified models of proteins such as the elastic network model has proved very effective in showing that many of the structural transitions derived from a survey of the Protein Data Bank can be explained by just a few of the lowest-frequency normal modes. In this work, normal modes are used to carry out medium- or low-resolution structural refinement, enforcing collective and large-amplitude movements that are beyond the reach of existing methods. Refinement is carried out in reciprocal space with respect to the normal mode amplitudes, by using standard conjugate-gradient minimization. Several tests on synthetic diffraction data whose mode concentration follows the one of real movements observed in the Protein Data Bank have shown that the radius of convergence is larger than the one of rigid-body refinement. Tests with experimental diffraction data for the same protein in different environments also led to refined structural models showing drastic reduction of the rms deviation with the target model. Because the structural transition is described by very few parameters, over-fitting of real experimental data is easily detected by using a cross-validation test. The method has also been applied to the refinement of atomic models into molecular envelopes and could readily be used to fit large macromolecular complex rearrangements into cryo-electron microscopy-reconstructed images as well as small-angle x-ray scattering-derived envelopes.}, note = {0027-8424 Journal Article}, keywords = {DUMAS Carrier Proteins/chemistry Computer Simulation Escherichia coli Proteins/chemistry *Models, Molecular Proteins/*chemistry Support, Non-U.S. Gov't X-Ray Diffraction, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Determination of queuosine derivatives by reverse-phase liquid chromatography for the hypomodification study of Q-bearing tRNAs from various mammal liver cells}, author = {A Costa and J P Pais de Barros and G Keith and W Baranowski and J Desgres}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14751792}, isbn = {14751792}, year = {2004}, date = {2004-01-01}, journal = {J Chromatogr B Analyt Technol Biomed Life Sci}, volume = {801}, number = {2}, pages = {237-247}, abstract = {Three queuosine derivatives (Q-derivatives) have been found at position 34 of four mammalian so-called Q-tRNAs: queuosine (Q) in tRNA(Asn) and tRNA(His), mannosyl-queuosine (manQ) in tRNA(Asp), and galactosyl-queuosine (galQ) in tRNA(Tyr). An analytical procedure based on the combined means of purified tRNA isolation from liver cells and ribonucleoside analysis by reverse-phase high performance liquid chromatography coupled with real-time UV-spectrometry (RPLC-UV) was developed for the quantitative analysis of the three Q-derivatives present in total tRNA from liver tissues and liver cell cultures. Using this analytical procedure, the rates of Q-tRNA modification were studied in total tRNAs from various mammalian hepatic cells. Our results show that the four Q-tRNAs are fully modified in liver tissues from adult mammals, regardless of the mammal species. However, a lack in the Q-modification level was observed in Q-tRNAs from newborn rat liver, as well in Q-tRNAs from normal rat liver cell cultures growing in a low queuine content medium, and from a rat hepatoma cell line. It is noteworthy that in all cases of Q-tRNA hypomodification, our analytical procedure showed that tRNA(Asp) is always the least affected by the hypomodification. The biological significance of this phenomenon is discussed.}, note = {1570-0232 Journal Article}, keywords = {Amino Acyl/chemistry RNA, Asn/chemistry Rats Support, Cultured, Cultured Chickens *Chromatography, Experimental Nucleoside Q/*analogs & derivatives/*analysis RNA, High Pressure Liquid Hepatocytes/chemistry Liver/*chemistry Liver Neoplasms, KEITH Animals Cells, Non-U.S. Gov't Tumor Cells, Transfer, Transfer/*chemistry/isolation & purification RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization and preliminary X-ray diffraction data of an archaeal asparagine synthetase related to asparaginyl-tRNA synthetase}, author = {C Charron and H Roy and M Blaise and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15039580}, isbn = {15039580}, year = {2004}, date = {2004-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {60}, number = {Pt 4}, pages = {767-769}, abstract = {The archaebacterial-type asparagine synthetase A from Pyrococcus abyssi (AS-AR), which is related to asparaginyl-tRNA synthetase, was crystallized in two different conditions using the hanging-drop vapour-diffusion method. Crystals belonging to space group C2 with unit-cell parameters a = 103.6}, note = {0907-4449 Journal Article}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of thaumatin in a hexagonal space group: comparison of packing contacts in four crystal lattices}, author = {C Charron and R Giege and B Lorber}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14684896}, isbn = {14684896}, year = {2004}, date = {2004-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {60}, number = {Pt 1}, pages = {83-89}, abstract = {The intensely sweet protein thaumatin has been crystallized in a hexagonal lattice after a temperature shift from 293 to 277 K. The structure of the protein in the new crystal was solved at 1.6 A resolution. The protein fold is identical to that found in three other crystal forms grown in the presence of crystallizing agents of differing chemical natures. The proportions of lattice interactions involving hydrogen bonds, hydrophobic or ionic groups differ greatly from one form to another. Moreover, the distribution of acidic and basic residues taking part in contacts also varies. The hexagonal packing is characterized by the presence of channels parallel to the c axis that are so wide that protein molecules can diffuse through them.}, note = {0907-4449 Journal Article}, keywords = {GIEGE Comparative Study Crystallization Crystallography, Molecular Plant Proteins/*chemistry Support, Non-U.S. Gov't, Unité ARN, X-Ray Hydrogen Bonding Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reconsidering the evolution of eukaryotic selenoproteins: a novel nonmammalian family with scattered phylogenetic distribution}, author = {S Castellano and S V Novoselov and G V Kryukov and A Lescure and E Blanco and A Krol and V N Gladyshev and R Guigo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14710190}, isbn = {14710190}, year = {2004}, date = {2004-01-01}, journal = {EMBO Rep}, volume = {5}, number = {1}, pages = {71-77}, abstract = {While the genome sequence and gene content are available for an increasing number of organisms, eukaryotic selenoproteins remain poorly characterized. The dual role of the UGA codon confounds the identification of novel selenoprotein genes. Here, we describe a comparative genomics approach that relies on the genome-wide prediction of genes with in-frame TGA codons, and the subsequent comparison of predictions from different genomes, wherein conservation in regions flanking the TGA codon suggests selenocysteine coding function. Application of this method to human and fugu genomes identified a novel selenoprotein family, named SelU, in the puffer fish. The selenocysteine-containing form also occurred in other fish, chicken, sea urchin, green algae and diatoms. In contrast, mammals, worms and land plants contained cysteine homologues. We demonstrated selenium incorporation into chicken SelU and characterized the SelU expression pattern in zebrafish embryos. Our data indicate a scattered evolutionary distribution of selenoproteins in eukaryotes, and suggest that, contrary to the picture emerging from data available so far, other taxa-specific selenoproteins probably exist.}, note = {1469-221x Journal Article}, keywords = {KROL, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Microdissection, mRNA amplification and microarray: a study of pleural mesothelial and malignant mesothelioma cells}, author = { S. Mohr and M.C. Bottin and B. Lannes and A. Neuville and J.P. Bellocq and G. Keith and B.H. Rihn}, year = {2004}, date = {2004-01-01}, journal = {Biochimie}, volume = {86}, number = {1}, pages = {13-9}, abstract = {The studies of molecular alterations in tumor cells with microarrays are often hampered by inherent tissue heterogeneity. The emergence of Laser Capture Microdissection (LCM) allowed us to overcome this challenge since it gives selective access to cancer cells that are isolated from their native tissue environment. In this report, we microdissected mesothelial cells and malignant mesothelioma cells of ex vivo resected specimens using LCM. Amplified RNA from mesothelial and mesothelioma microdissected cells allowed us to measure global gene expression with 10 K-microarrays in four independent experiments. We screened 9850 annotated human genes, 1275 of which have satisfied our data analysis requirements. They included 302 overexpressed genes and 160 downregulated genes in mesothelioma microdissected cells as compared to mesothelial microdissected cells. Among them, the expression levels of eight genes, namely BF, FTL, IGFBP7, RARRES1, RARRES2, RBP1, SAT, and TXN according to HUGO nomenclature, were increased, whereas six: ALOX5AP, CLNS1A, EIF4A2, ELK3, REQ and SYPL, were found to be underexpressed in mesothelioma microdissected cells. The ferritin light polypeptide (FTL) gene overexpression was confirmed by real time quantitative PCR. Our approach allowed a comprehensive in situ examination of mesothelioma and provided an accurate way to find new marker genes that may be useful for diagnosis and treatment of malignant pleural mesothelioma.}, note = {0300-9084 Journal Article}, keywords = {Analysis, Array, Chain, Epithelium/*metabolism, Expression, Female, Gene, Genetic, Human, KEITH, Lasers, Male, Markers, Mesothelioma/*genetics/metabolism, messenger, Microdissection, Neoplasms/*genetics/metabolism, Neoplastic/*genetics, Oligonucleotide, Pleura/*cytology/*metabolism, Pleural, Polymerase, Profiling, Reaction, Regulation, Reverse, RNA, Sequence, Transcriptase}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Escherichia coli YadB gene product reveals a novel aminoacyl-tRNA synthetase like activity}, author = {V Campanacci and D Y Dubois and H D Becker and D Kern and S Spinelli and C Valencia and F Pagot and A Salomoni and S Grisel and R Vincentelli and C Bignon and J Lapointe and R Giege and C Cambillau}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15003446}, isbn = {15003446}, year = {2004}, date = {2004-01-01}, journal = {J Mol Biol}, volume = {337}, number = {2}, pages = {273-283}, abstract = {In the course of a structural genomics program aiming at solving the structures of Escherichia coli open reading frame products of unknown function, we have determined the structure of YadB at 1.5A using molecular replacement. The YadB protein is 298 amino acid residues long and displays 34% sequence identity with E.coli glutamyl-tRNA synthetase (GluRS). It is much shorter than GluRS, which contains 468 residues, and lacks the complete domain interacting with the tRNA anticodon loop. As E.coli GluRS, YadB possesses a Zn2+ located in the putative tRNA acceptor stem-binding domain. The YadB cluster uses cysteine residues as the first three zinc ligands, but has a weaker tyrosine ligand at the fourth position. It shares with canonical amino acid RNA synthetases a major functional feature, namely activation of the amino acid (here glutamate). It differs, however, from GluRSs by the fact that the activation step is tRNA-independent and that it does not catalyze attachment of the activated glutamate to E.coli tRNAGlu, but to another, as yet unknown tRNA. These results suggest thus a novel function, distinct from that of GluRSs, for the yadB gene family.}, note = {0022-2836 Journal Article}, keywords = {Amino Acid Support, Bacterial Glutamate-tRNA Ligase/chemistry/genetics/metabolism Glutamic Acid/metabolism Kinetics Ligands Models, Glu/metabolism Sequence Homology, KERN GIEGE Adenosine Monophosphate/metabolism Adenosine Triphosphate/metabolism Amino Acid Sequence Amino Acyl-tRNA Ligases/chemistry/*genetics/*metabolism Carrier Proteins/metabolism Crystallography, Molecular Molecular Sequence Data Neoplasm Proteins/metabolism Nuclear Proteins/metabolism Protein Conformation RNA, Non-U.S. Gov't Thermus thermophilus/enzymology/genetics Zinc/metabolism, Transfer, Unité ARN, X-Ray Escherichia coli/*enzymology/*genetics Escherichia coli Proteins/chemistry/*genetics/*metabolism Genes}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cell protection, resistance and invasiveness of two malignant mesotheliomas as assessed by 10K-microarray}, author = { S. Mohr and G. Keith and F. Galateau-Salle and P. Icard and B. H. Rihn}, year = {2004}, date = {2004-01-01}, journal = {Biochim Biophys Acta-Mol Basis Dis}, volume = {1688}, number = {1}, pages = {43-60}, abstract = {Malignant pleural mesothelioma (MPM) is an aggressive serosal tumor, strongly associated with former exposure to asbestos fibers and for which there is currently no effective treatment available. In human, MPM is characterized by a high local invasiveness, poor prognosis and therapeutic outcomes. In order to assess molecular changes that specify this phenotype, we performed a global gene expression profiling of human MPM. Using a 10,000-element microarray, we analyzed mRNA relative gene expression levels by comparing a mesothelioma cell line to either a pleural cell line or tumor specimens. To analyze these gene expression data, we used various bioinformatics softwares. Hierarchical clustering methods were used to group genes and samples with similar expression in an unsupervised mode. Genes of known function were further sorted by enzyme, function and pathway clusters using a supervised software (IncyteGenomics). Taken together, these data defined a molecular fingerprint of human MPM with more than 700 up- or down-regulated genes related to several traits of the malignant phenotype, specially associated with MPM invasiveness, protection and resistance to anticancer defenses. This portrait is meaningful in disease classification and management, and relevant in finding new specific markers of MPM. These molecular markers should improve the accuracy of mesothelioma diagnosis, prognosis and therapy.}, note = {0006-3002 Journal Article}, keywords = {Analysis, Array, Biological/genetics, Cell, Expression, Family, Gene, Human, KEITH, Line, Markers, Mesothelioma/etiology/genetics/*pathology, Messenger/analysis, Multigene, Neoplasms/etiology/genetics/*pathology, Neoplastic, Pleural, Profiling, Protein, Regulation, RNA, tumor}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural and biochemical analysis of sliding clamp/ligand interactions suggest a competition between replicative and translesion DNA polymerases}, author = {D Y Burnouf and V Olieric and J Wagner and S Fujii and J Reinbolt and R P Fuchs and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14729336}, isbn = {14729336}, year = {2004}, date = {2004-01-01}, journal = {J Mol Biol}, volume = {335}, number = {5}, pages = {1187-1197}, abstract = {Most DNA polymerases interact with their cognate processive replication factor through a small peptide, this interaction being absolutely required for their function in vivo. We have solved the crystal structure of a complex between the beta sliding clamp of Escherichia coli and the 16 residue C-terminal peptide of Pol IV (P16). The seven C-terminal residues bind to a pocket located at the surface of one beta monomer. This region was previously identified as the binding site of another beta clamp binding protein, the delta subunit of the gamma complex. We show that peptide P16 competitively prevents beta-clamp-mediated stimulation of both Pol IV and alpha subunit DNA polymerase activities, suggesting that the site of interaction of the alpha subunit with beta is identical with, or overlaps that of Pol IV. This common binding site for delta, Pol IV and alpha subunit is shown to be formed by residues that are highly conserved among many bacterial beta homologs, thus defining an evolutionarily conserved hydrophobic crevice for sliding clamp ligands and a new target for antibiotic drug design.}, note = {0022-2836 Journal Article}, keywords = {Bacterial/genetics Escherichia coli/*enzymology Kinetics Ligands Models, Competitive Crystallization DNA Polymerase I/metabolism DNA Polymerase III/metabolism DNA Polymerase beta/*chemistry/genetics/*metabolism DNA Replication/*genetics DNA, DUMAS *Binding, Molecular Peptide Fragments/*metabolism Proliferating Cell Nuclear Antigen/metabolism Protein Binding Protein Subunits Recombinant Proteins/chemistry/metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Secondary structure of the 3' UTR of bicoid mRNA}, author = {C Brunel and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15016447}, isbn = {15016447}, year = {2004}, date = {2004-01-01}, journal = {Biochimie}, volume = {86}, number = {2}, pages = {91-104}, abstract = {Formation of the Bicoid morphogen gradient in early Drosophila embryos requires the pre-localization of bicoid mRNA to the anterior pole of the egg. The program of bcd mRNA localization involves multiples steps and proceeds from oogenesis until early embryogenesis. This process requires cis-elements in the 3' UTR of bcd mRNA and successive and/or concomitant critical protein interactions. Furthermore, numerous RNA elements and binding proteins contribute to regulate bcd expression. In the present paper, we investigated the secondary structure of the full length 3' UTR of the bcd mRNA, using a variety of chemical and enzymatic structural probes. This RNA probing analysis allowed us to give a detailed description of the 3' UTR of the bcd mRNA and its organization into five well-defined and independent domains (I-V). One prominent result that emerges from our data is the unexpected high degree of flexibility of the different domains relative to each others. This plasticity relies upon the open conformation of the central hinge region interconnecting domains II, III, and IV + V. Otherwise, dimerization of the 3' UTR, which participates to anchoring bcd mRNA at the anterior pole of the embryo, only results in discrete and local change in domain III. Domain I that contains sites for trans-acting factors exhibiting single stranded RNA binding specificity is mainly unstructured. By contrast, each core domains (II-V) is highly organized and folds into helices interrupted by bulges and interior loops and closed by very exposed apical loops. These elements mostly built specific determinants for trans-acting factors. Besides, these findings provide a valuable database for structure/function studies.}, note = {0300-9084 Journal Article}, keywords = {Biological *Nucleic Acid Conformation RNA, BRUNEL 3' Untranslated Regions/*chemistry/genetics Animals Conserved Sequence Dimerization Drosophila Homeodomain Proteins/*genetics Models, Messenger/biosynthesis/*chemistry/genetics RNA-Binding Proteins/chemistry/genetics Regulatory Sequences, Non-U.S. Gov't Trans-Activators/*genetics, Nucleic Acid/genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A minimalist glutamyl-tRNA synthetase dedicated to aminoacylation of the tRNAAsp QUC anticodon}, author = {M Blaise and H D Becker and G Keith and C Cambillau and J Lapointe and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15150343}, isbn = {15150343}, year = {2004}, date = {2004-01-01}, journal = {Nucleic Acids Res}, volume = {32}, number = {9}, pages = {2768-2775}, abstract = {Escherichia coli encodes YadB, a protein displaying 34% identity with the catalytic core of glutamyl-tRNA synthetase but lacking the anticodon-binding domain. We show that YadB is a tRNA modifying enzyme that evidently glutamylates the queuosine residue, a modified nucleoside at the wobble position of the tRNA(Asp) QUC anticodon. This conclusion is supported by a variety of biochemical data and by the inability of the enzyme to glutamylate tRNA(Asp) isolated from an E.coli tRNA-guanosine transglycosylase minus strain deprived of the capacity to exchange guanosine 34 with queuosine. Structural mimicry between the tRNA(Asp) anticodon stem and the tRNA(Glu) amino acid acceptor stem in prokaryotes encoding YadB proteins indicates that the function of these tRNA modifying enzymes, which we rename glutamyl-Q tRNA(Asp) synthetases, is conserved among prokaryotes.}, note = {1362-4962 Journal Article}, keywords = {Asp/chemistry/genetics/*metabolism RNA, Bacterial/chemistry/genetics/metabolism RNA, Glu/chemistry/genetics/metabolism Support, KERN Acylation Anticodon/chemistry/genetics/*metabolism Base Sequence Conserved Sequence Escherichia coli/*enzymology/*genetics Evolution Glutamate-tRNA Ligase/*chemistry/genetics/*metabolism Molecular Mimicry Nucleoside Q/genetics/*metabolism Periodic Acid/pharmacology RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The three-dimensional architecture of the class I ligase ribozyme}, author = {N H Bergman and N C Lau and V Lehnert and E Westhof and D P Bartel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14730016}, isbn = {14730016}, year = {2004}, date = {2004-01-01}, journal = {RNA}, volume = {10}, number = {2}, pages = {176-184}, abstract = {The class I ligase ribozyme catalyzes a Mg(++)-dependent RNA-ligation reaction that is chemically analogous to a single step of RNA polymerization. Indeed, this ribozyme constitutes the catalytic domain of an accurate and general RNA polymerase ribozyme. The ligation reaction is also very rapid in both single- and multiple-turnover contexts and thus is informative for the study of RNA catalysis as well as RNA self-replication. Here we report the initial characterization of the three-dimensional architecture of the ligase. When the ligase folds, several segments become protected from hydroxyl-radical cleavage, indicating that the RNA adopts a compact tertiary structure. Ribozyme folding was largely, though not completely, Mg(++) dependent, with a K(1/2[Mg]) < 1 mM, and was observed over a broad temperature range (20 degrees C -50 degrees C). The hydroxyl-radical mapping, together with comparative sequence analyses and analogy to a region within 23S ribosomal RNA, were used to generate a three-dimensional model of the ribozyme. The predictive value of the model was tested and supported by a photo-cross-linking experiment.}, note = {1355-8382 Journal Article}, keywords = {Animals Cross-Linking Reagents Human Hydroxyl Radical/metabolism Light Magnesium/metabolism *Nucleic Acid Conformation RNA? WESTHOF, Catalytic/*metabolism Support, Non-U.S. Gov't Temperature, Unité ARN, WESTHOF Animals Cross-Linking Reagents Human Hydroxyl Radical/metabolism Light Magnesium/metabolism *Nucleic Acid Conformation RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNA-like structure regulates translation of Brome mosaic virus RNA}, author = {S Barends and J Rudinger-Thirion and C Florentz and R Giege and C W Pleij and B Kraal}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15047816}, isbn = {15047816}, year = {2004}, date = {2004-01-01}, journal = {J Virol}, volume = {78}, number = {8}, pages = {4003-4010}, abstract = {For various groups of plant viruses, the genomic RNAs end with a tRNA-like structure (TLS) instead of the 3' poly(A) tail of common mRNAs. The actual function of these TLSs has long been enigmatic. Recently, however, it became clear that for turnip yellow mosaic virus, a tymovirus, the valylated TLS(TYMV) of the single genomic RNA functions as a bait for host ribosomes and directs them to the internal initiation site of translation (with N-terminal valine) of the second open reading frame for the polyprotein. This discovery prompted us to investigate whether the much larger TLSs of a different genus of viruses have a comparable function in translation. Brome mosaic virus (BMV), a bromovirus, has a tripartite RNA genome with a subgenomic RNA4 for coat protein expression. All four RNAs carry a highly conserved and bulky 3' TLS(BMV) (about 200 nucleotides) with determinants for tyrosylation. We discovered TLS(BMV)-catalyzed self-tyrosylation of the tyrosyl-tRNA synthetase but could not clearly detect tyrosine incorporation into any virus-encoded protein. We established that BMV proteins do not need TLS(BMV) tyrosylation for their initiation. However, disruption of the TLSs strongly reduced the translation of genomic RNA1, RNA2, and less strongly, RNA3, whereas coat protein expression from RNA4 remained unaffected. This aberrant translation could be partially restored by providing the TLS(BMV) in trans. Intriguingly, a subdomain of the TLS(BMV) could even almost fully restore translation to the original pattern. We discuss here a model with a central and dominant role for the TLS(BMV) during the BMV infection cycle.}, note = {0022-538x Journal Article}, keywords = {ase Sequence Bromovirus/*genetics/metabolism Genetic Complementation Test Genome, FLORENTZ, FRUGIER, Genetic Triticum/virology Tyrosine/chemistry Tyrosine-tRNA Ligase/chemistry/genetics/metabolism Viral Proteins/chemistry/genetics, Non-U.S. Gov't Translation, Transfer/chemistry/genetics RNA, Unité ARN, Viral Molecular Sequence Data Nucleic Acid Conformation RNA, Viral/*chemistry/*genetics Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Halogen bonds in biological molecules}, author = {P Auffinger and F A Hays and E Westhof and P S Ho}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15557000}, isbn = {15557000}, year = {2004}, date = {2004-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {101}, number = {48}, pages = {16789-16794}, abstract = {Short oxygen-halogen interactions have been known in organic chemistry since the 1950s and recently have been exploited in the design of supramolecular assemblies. The present survey of protein and nucleic acid structures reveals similar halogen bonds as potentially stabilizing inter- and intramolecular interactions that can affect ligand binding and molecular folding. A halogen bond in biomolecules can be defined as a short CX.OY interaction (CX is a carbon-bonded chlorine, bromine, or iodine, and OY is a carbonyl, hydroxyl, charged carboxylate, or phosphate group), where the X.O distance is less than or equal to the sums of the respective van der Waals radii (3.27 A for Cl.O, 3.37A for Br.O, and 3.50 A for I.O) and can conform to the geometry seen in small molecules, with the CX.O angle approximately 165 degrees (consistent with a strong directional polarization of the halogen) and the X.OY angle approximately 120 degrees. Alternative geometries can be imposed by the more complex environment found in biomolecules, depending on which of the two types of donor systems are involved in the interaction: (i) the lone pair electrons of oxygen (and, to a lesser extent, nitrogen and sulfur) atoms or (ii) the delocalized pi -electrons of peptide bonds or carboxylate or amide groups. Thus, the specific geometry and diversity of the interacting partners of halogen bonds offer new and versatile tools for the design of ligands as drugs and materials in nanotechnology.}, note = {0027-8424 Journal Article}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Symmetric K+ and Mg2+ ion-binding sites in the 5S rRNA loop E inferred from molecular dynamics simulations}, author = {P Auffinger and L Bielecki and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14672663}, isbn = {14672663}, year = {2004}, date = {2004-01-01}, journal = {J Mol Biol}, volume = {335}, number = {2}, pages = {555-571}, abstract = {Potassium binding to the 5S rRNA loop E motif has been studied by molecular dynamics at high (1.0 M) and low (0.2 M) concentration of added KCl in the presence and absence of Mg2+. A clear pattern of seven deep groove K+ binding sites or regions, in all cases connected with guanine N7/O6 atoms belonging to GpG, GpA, and GpU steps, was identified, indicating that the LE deep groove is significantly more ionophilic than the equivalent groove of regular RNA duplexes. Among all, two symmetry-related sites (with respect to the central G.A pair) were found to accommodate K+ ions with particularly long residence times. In a preceding molecular dynamics study by Auffinger et al. in the year 2003, these two sites were described as constituting important Mg2+ binding locations. Altogether, the data suggest that these symmetric sites correspond to the loop E main ion binding regions. Indeed, they are located in the deep groove of an important ribosomal protein binding motif associated with a fragile pattern of non-Watson-Crick pairs that has certainly to be stabilized by specific Mg2+ ions in order to be efficiently recognized by the protein. Besides, the other sites accommodate monovalent ions in a more diffuse way pointing out their lesser significance for the structure and function of this motif. Ion binding to the shallow groove and backbone atoms was generally found to be of minor importance since, at the low concentration, no well defined binding site could be characterized while high K+ concentration promoted mostly unspecific potassium binding to the RNA backbone. In addition, several K+ binding sites were located in positions equivalent to water molecules from the first hydration shell of divalent ions in simulations performed with magnesium, indicating that ion binding regions are able to accommodate both mono- and divalent ionic species. Overall, the simulations provide a more precise but, at the same time, a more intricate view of the relations of this motif with its ionic surrounding.}, note = {0022-2836 Journal Article}, keywords = {5S/*chemistry/*metabolism Ribosomal Proteins/*chemistry/metabolism Support, Bacterial/chemistry RNA, Binding Sites Computer Simulation Guanine/chemistry/metabolism Hydrogen Bonding Magnesium/chemistry/*metabolism Models, Molecular Molecular Conformation Nucleic Acid Conformation Potassium/chemistry/*metabolism Protein Binding/genetics RNA, Non-U.S. Gov't Water/chemistry/metabolism, Ribosomal, Unité ARN, WESTHOF, WESTHOF Binding Sites Computer Simulation Guanine/chemistry/metabolism Hydrogen Bonding Magnesium/chemistry/*metabolism Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Anion binding to nucleic acids}, author = {P Auffinger and L Bielecki and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pubmed/15016354}, isbn = {15016354}, year = {2004}, date = {2004-01-01}, journal = {Structure}, volume = {12}, number = {3}, pages = {379-388}, abstract = {Nucleic acids are generally considered as efficient cation binders. Therefore, the likelihood that negatively charged ions might intrude their first hydration shell is rarely considered. Here, we show on the basis of (i) a survey of the Nucleic Acid Database, (ii) several structures extracted from the Cambridge Structural Database, and (iii) molecular dynamics simulations, that the nucleotide electropositive edges involving mainly amino, imino, and hydroxyl groups can cast specific anion binding sites. These binding sites constitute also good locations for the binding of the negatively charged groups of the Asp and Glu residues or the nucleic acid phosphate groups. Furthermore, it is observed in several instances that anions, like water molecules and cations, do mediate protein/nucleic acid interactions. Thus, anions as well as negatively charged groups are directly involved in specific recognition and folding phenomena involving polyanionic nucleic acids.}, note = {0969-2126 Journal Article}, keywords = {Anions/chemistry/*metabolism Computer Simulation Databases, Molecular Nucleic Acids/chemistry/*metabolism Support, Non-U.S. Gov't X-Ray Diffraction, Nucleic Acid Models, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutation and evolution of the magnesium-binding site of a class II aminoacyl-tRNA synthetase}, author = {L Ador and S Jaeger and R Geslain and F Martin and J Cavarelli and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15170340}, isbn = {15170340}, year = {2004}, date = {2004-01-01}, journal = {Biochemistry}, volume = {43}, number = {22}, pages = {7028-7037}, abstract = {Aminoacyl-tRNA synthetases contain one or three Mg(2+) ions in their catalytic sites. In addition to their role in ATP binding, these ions are presumed to play a role in catalysis by increasing the electropositivity of the alpha-phosphate and stabilizing the pentavalent transition state. In the class II aaRS, two highly conserved carboxylate residues have been shown to participate with Mg(2+) ions in binding and coordination. It is shown here that these carboxylate residues are absolutely required for the activity of Saccharomyces cerevisiae aspartyl-tRNA synthetase. Mutants of these residues exhibit pleiotropic effects on the kinetic parameters suggesting an effect at an early stage of the aminoacylation reaction, such as the binding of ATP, Mg(2+), aspartic acid, or the amino acid activation. Despite genetic selections in an APS-knockout yeast strain, we were unable to select a single active mutant of these carboxylate residues. Nevertheless, we isolated an intragenic suppressor from a combinatorial library. The active mutant showed a second substitution close to the first one, and exhibited a significant increase of the tRNA aminoacylation rate. Structural analysis suggests that the acceptor stem of the tRNA might be repositioned to give a more productive enzyme:tRNA complex. Thus, the initial defect of the activation reaction was compensated by a significant increase of the aminoacylation rate that led to cellular complementation.}, note = {0006-2960 Journal Article}, keywords = {Acylation Adenosine Triphosphate/metabolism Amino Acid Substitution Aspartate-tRNA Ligase/chemistry/genetics/*metabolism Aspartic Acid/metabolism Binding Sites Catalytic Domain Cell Death Combinatorial Chemistry Techniques Comparative Study *Evolution, Asp/metabolism Saccharomyces cerevisiae/*enzymology Support, ERIANI, Molecular Kinetics Magnesium/*metabolism Mutagenesis, Non-U.S. Gov't Transfection, Site-Directed Mutation/*genetics Peptide Library Protein Binding Protein Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural and biochemical analysis of sliding clamp/ligand interactions suggest a competition between replicative and translesion DNA polymerases}, author = { D. Y. Burnouf and V. Olieric and J. Wagner and S. Fujii and J. Reinbolt and R. P. Fuchs and P. Dumas}, year = {2004}, date = {2004-01-01}, journal = {J Mol Biol}, volume = {335}, number = {5}, pages = {1187-97}, abstract = {Most DNA polymerases interact with their cognate processive replication factor through a small peptide, this interaction being absolutely required for their function in vivo. We have solved the crystal structure of a complex between the beta sliding clamp of Escherichia coli and the 16 residue C-terminal peptide of Pol IV (P16). The seven C-terminal residues bind to a pocket located at the surface of one beta monomer. This region was previously identified as the binding site of another beta clamp binding protein, the delta subunit of the gamma complex. We show that peptide P16 competitively prevents beta-clamp-mediated stimulation of both Pol IV and alpha subunit DNA polymerase activities, suggesting that the site of interaction of the alpha subunit with beta is identical with, or overlaps that of Pol IV. This common binding site for delta, Pol IV and alpha subunit is shown to be formed by residues that are highly conserved among many bacterial beta homologs, thus defining an evolutionarily conserved hydrophobic crevice for sliding clamp ligands and a new target for antibiotic drug design.}, note = {0022-2836 Journal Article}, keywords = {*Binding, Antigen/metabolism, Bacterial/genetics, beta/*chemistry/genetics/*metabolism, Binding, Cell, coli/*enzymology, Competitive, Crystallization, DNA, DUMAS, Escherichia, Fragments/*metabolism, I/metabolism, III/metabolism, Kinetics, ligands, Models, Molecular, Nuclear, Peptide, Polymerase, Proliferating, Protein, Proteins/chemistry/metabolism, Recombinant, Replication/*genetics, Subunits}, pubstate = {published}, tppubtype = {article} } @article{, title = {IgG reactivity with a 100-kDa tissue and endothelial cell antigen identified as topoisomerase 1 distinguishes between limited and diffuse systemic sclerosis patients}, author = { P. Garcia de la Pena-Lefebvre and Y. Chanseaud and M. C. Tamby and J. Reinbolt and F. Batteux and Y. Allanore and A. Kahan and O. Meyer and O. Benveniste and O. Boyer and L. Guillevin and M. C. Boissier and L. Mouthon}, year = {2004}, date = {2004-01-01}, journal = {Clin Immunol}, volume = {111}, number = {3}, pages = {241-51}, abstract = {We have analyzed antibody (Ab) reactivities of patients with limited systemic sclerosis (SSc) and anti-centromere Ab, patients with diffuse SSc and anti-topoisomerase 1 (anti-topo 1) Ab, patients with diffuse SSc without anti-topo 1 or anti-centromere Ab and age- and gender-matched healthy controls with normal human tissue and endothelial cell (EC) antigens. IgG reactivities with tissue antigens differed significantly between patients with anti-topo 1 Ab and patients with anti-centromere Ab. One 100-kDa band identified as topoisomerase 1 in macrovascular and microvascular EC extracts was recognized by IgG from patients with anti-topo 1 Ab and 50% of patients without specific Ab. IgG from patients with limited SSc and anti-centromere Ab, but not those of other patients or controls specifically recognized a 80-kDa band only in microvascular EC. Our results indicate that Ab from patients with limited or diffuse SSc with or without anti-topo 1 Ab exhibit specific and mutually exclusive reactivity patterns.}, note = {1521-6616 Journal Article}, keywords = {Aged, Assay, Autoantibodies/*analysis, Blotting, Cells/*immunology, Centromere/immunology, DNA, EHRESMANN, Electrophoresis, Endothelial, Enzyme-Linked, Female, G/analysis, Gel, Gov't, Human, I/*immunology, Immunoglobulin, Immunosorbent, M/analysis, Male, Middle, Non-U.S., Polyacrylamide, Scleroderma, Support, Systemic/*immunology, Topoisomerases, Type, Western}, pubstate = {published}, tppubtype = {article} } @article{, title = {A simple epigenetic method for the diagnosis and classification of brain tumors}, author = { R. Zukiel and S. Nowak and A. M. Barciszewska and I. Gawronska and G. Keith and M. Z. Barciszewska}, year = {2004}, date = {2004-01-01}, journal = {Mol Cancer Res}, volume = {2}, number = {3}, pages = {196-202}, abstract = {The new, simple, and reliable method for the diagnosis of brain tumors is described. It is based on a TLC quantitative determination of 5-methylcytosine (m(5)C) in relation to its damage products of DNA from tumor tissue. Currently, there is evidence that oxidative stress through reactive oxygen species (ROS) plays an important role in the etiology and progression of several human diseases. Oxidative damage of DNA, lipids, and proteins is deleterious for the cell. m(5)C, along with other basic components of DNA, is the target for ROS, which results in the appearance of new modified nucleic acid bases. If so, m(5)C residue constitutes a mutational hotspot position, whether it occurs within a nucleotide sequence of a structural gene or a regulatory region. Here, we show the results of the analysis of 82 DNA samples taken from brain tumor tissues. DNA was isolated and hydrolyzed into nucleotides, which, after labeling with [gamma-(32)P]ATP, were separated on TLC. Chromatograms were evaluated using PhosphorImager and the amounts of 5-methyldeoxycytosine (m(5)dC) were calculated as a ratio (R) of m(5)dC to m(5)dC + deoxycytosine + deoxythymidine spot intensities. The R value could not only be a good diagnostic marker for brain tumors but also a factor differentiating low-grade and high-grade gliomas. Therefore, DNA methylation pattern might be a useful tool to give a primary diagnosis of a brain tumor or as a marker for the early detection of the relapse of the disease. This method has several advantages over those existing nowadays.}, note = {1541-7786 Journal Article}, keywords = {*DNA, *Epigenesis, 5-Methylcytosine/*analysis, Adult, Aged, and, Brain, Chromatography, DNA, Female, Genetic, Gov't, Human, KEITH, Layer, Male, Methylation, Middle, Neoplasm/*chemistry/*metabolism, Neoplasms/*classification/*diagnosis/genetics/pathology, Non-U.S., Oxidative, oxygen, Reactive, Sensitivity, Species/metabolism, Specificity, Stress, Support, Thin}, pubstate = {published}, tppubtype = {article} } @article{, title = {Determination of queuosine derivatives by reverse-phase liquid chromatography for the hypomodification study of Q-bearing tRNAs from various mammal liver cells}, author = { A. Costa and J. P. Pais de Barros and G. Keith and W. Baranowski and J. Desgres}, year = {2004}, date = {2004-01-01}, journal = {J Chromatogr B Analyt Technol Biomed Life Sci}, volume = {801}, number = {2}, pages = {237-47}, abstract = {Three queuosine derivatives (Q-derivatives) have been found at position 34 of four mammalian so-called Q-tRNAs: queuosine (Q) in tRNA(Asn) and tRNA(His), mannosyl-queuosine (manQ) in tRNA(Asp), and galactosyl-queuosine (galQ) in tRNA(Tyr). An analytical procedure based on the combined means of purified tRNA isolation from liver cells and ribonucleoside analysis by reverse-phase high performance liquid chromatography coupled with real-time UV-spectrometry (RPLC-UV) was developed for the quantitative analysis of the three Q-derivatives present in total tRNA from liver tissues and liver cell cultures. Using this analytical procedure, the rates of Q-tRNA modification were studied in total tRNAs from various mammalian hepatic cells. Our results show that the four Q-tRNAs are fully modified in liver tissues from adult mammals, regardless of the mammal species. However, a lack in the Q-modification level was observed in Q-tRNAs from newborn rat liver, as well in Q-tRNAs from normal rat liver cell cultures growing in a low queuine content medium, and from a rat hepatoma cell line. It is noteworthy that in all cases of Q-tRNA hypomodification, our analytical procedure showed that tRNA(Asp) is always the least affected by the hypomodification. The biological significance of this phenomenon is discussed.}, note = {1570-0232 Journal Article}, keywords = {*Chromatography, &, Acyl/chemistry, Amino, Animals, Asn/chemistry, Cells, Chickens, Cultured, derivatives/*analysis, Experimental, Gov't, Hepatocytes/chemistry, high, KEITH, liquid, Liver, Liver/*chemistry, Neoplasms, Non-U.S., Nucleoside, Pressure, purification, Q/*analogs, Rats, RNA, Support, Transfer, Transfer/*chemistry/isolation, tumor}, pubstate = {published}, tppubtype = {article} } @article{pantarotto_translocation_2004, title = {Translocation of bioactive peptides across cell membranes by carbon nanotubes}, author = {Davide Pantarotto and Jean-Paul Briand and Maurizio Prato and Alberto Bianco}, doi = {10.1039/b311254c}, issn = {1359-7345}, year = {2004}, date = {2004-01-01}, journal = {Chemical Communications (Cambridge, England)}, number = {1}, pages = {16--17}, abstract = {Functionalised carbon nanotubes are able to cross the cell membrane and to accumulate in the cytoplasm or reach the nucleus without being toxic for the cell up to 10 [micro sign]M.}, keywords = {3T3 Cells, Animals, carbon, Cell Membrane, Confocal, Flow Cytometry, fluorescence, I2CT, Mice, Microscopy, Nanotubes, Particle Size, Peptides, Protein Transport, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ligoxygakis_serpin_2003, title = {A serpin regulates dorsal-ventral axis formation in the Drosophila embryo}, author = {Petros Ligoxygakis and Siegfried Roth and Jean-Marc Reichhart}, issn = {0960-9822}, year = {2003}, date = {2003-12-01}, journal = {Curr. Biol.}, volume = {13}, number = {23}, pages = {2097--2102}, abstract = {Extracellular serine protease cascades have evolved in vertebrates and invertebrates to mediate rapid, local reactions to physiological or pathological cues. The serine protease cascade that triggers the Toll signaling pathway in Drosophila embryogenesis shares several organizational characteristics with those involved in mammalian complement and blood clotting. One of the hallmarks of such cascades is their regulation by serine protease inhibitors (serpins). Serpins act as suicide substrates and are cleaved by their target protease, forming an essentially irreversible 1:1 complex. The biological importance of serpins is highlighted by serpin dysfunction diseases, such as thrombosis caused by a deficiency in antithrombin. Here, we describe how a serpin controls the serine protease cascade, leading to Toll pathway activation. Female flies deficient in Serpin-27A produce embryos that lack dorsal-ventral polarity and show uniform high levels of Toll signaling. Since this serpin has been recently shown to restrain an immune reaction in the blood of Drosophila, it demonstrates that proteolysis can be regulated by the same serpin in different biological contexts.}, keywords = {Animals, Body Patterning, Cell Surface, Crosses, Female, Genetic, Immunohistochemistry, M3i, Microinjections, Receptors, reichhart, Serine Proteinase Inhibitors, Serpins, Signal Transduction, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{reichhart_tlr5_2003, title = {TLR5 takes aim at bacterial propeller}, author = {Jean-Marc Reichhart}, doi = {10.1038/ni1203-1159}, issn = {1529-2908}, year = {2003}, date = {2003-12-01}, journal = {Nat. Immunol.}, volume = {4}, number = {12}, pages = {1159--1160}, keywords = {Animals, Bacterial Physiological Phenomena, Cell Surface, Flagella, Flagellin, Humans, M3i, Membrane Glycoproteins, Receptors, reichhart, Toll-Like Receptor 5, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{royet_detection_2003, title = {Detection of peptidoglycans by NOD proteins}, author = {Julien Royet and Jean-Marc Reichhart}, issn = {0962-8924}, year = {2003}, date = {2003-12-01}, journal = {Trends Cell Biol.}, volume = {13}, number = {12}, pages = {610--614}, abstract = {Mechanisms of innate immune defense are based on the recognition of invariant microbial molecular patterns by specific receptors, followed by the activation of signaling pathways and the expression of effector molecules that will defeat the invading microorganism. Two recent reports add to the growing list of these pattern-recognition receptors by showing that the intracellular nucleotide-binding oligomerization domain 1 (NOD1) protein recognizes a diaminopimelate-containing muropeptide, a cell-wall component of Gram-negative bacteria.}, keywords = {Adaptor Proteins, Apoptosis, Carrier Proteins, Gram-Positive Bacteria, Humans, Immunity, Immunologic, Innate, M3i, Nod1 Signaling Adaptor Protein, Oligopeptides, peptidoglycan, Receptors, reichhart, Signal Transducing, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{gobert_dual_2003, title = {Dual activation of the Drosophila toll pathway by two pattern recognition receptors}, author = {Vanessa Gobert and Marie Gottar and Alexey A Matskevich and Sophie Rutschmann and Julien Royet and Marcia Belvin and Jules A Hoffmann and Dominique Ferrandon}, doi = {10.1126/science.1085432}, issn = {1095-9203}, year = {2003}, date = {2003-12-01}, journal = {Science}, volume = {302}, number = {5653}, pages = {2126--2130}, abstract = {The Toll-dependent defense against Gram-positive bacterial infections in Drosophila is mediated through the peptidoglycan recognition protein SA (PGRP-SA). A mutation termed osiris disrupts the Gram-negative binding protein 1 (GNBP1) gene and leads to compromised survival of mutant flies after Gram-positive infections, but not after fungal or Gram-negative bacterial challenge. Our results demonstrate that GNBP1 and PGRP-SA can jointly activate the Toll pathway. The potential for a combination of distinct proteins to mediate detection of infectious nonself in the fly will refine the concept of pattern recognition in insects.}, keywords = {Animals, Carrier Proteins, Cell Surface, DNA Transposable Elements, ferrandon, Gene Expression, Genes, Gram-Negative Bacteria, Gram-Positive Bacteria, Hemolymph, hoffmann, Hypocreales, Insect, Insect Proteins, M3i, Mutation, Phenotype, Receptors, Serine Endopeptidases, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{thouzeau_spheniscins_2003, title = {Spheniscins, avian beta-defensins in preserved stomach contents of the king penguin, Aptenodytes patagonicus}, author = {Cécile Thouzeau and Yvon Le Maho and Guillaume Froget and Laurence Sabatier and Céline Le Bohec and Jules A Hoffmann and Philippe Bulet}, doi = {10.1074/jbc.M306839200}, issn = {0021-9258}, year = {2003}, date = {2003-12-01}, journal = {J. Biol. Chem.}, volume = {278}, number = {51}, pages = {51053--51058}, abstract = {During the last part of egg incubation in king penguins, the male can preserve undigested food in the stomach for several weeks. This ensures survival of the newly hatched chick, in cases where the return of the foraging female from the sea is delayed. In accordance with the characterization of stress-induced bacteria, we demonstrate the occurrence of strong antimicrobial activities in preserved stomach contents. We isolated and fully characterized two isoforms of a novel 38-residue antimicrobial peptide (AMP), spheniscin, belonging to the beta-defensin subfamily. Spheniscin concentration was found to strongly increase during the period of food storage. Using a synthetic version of one of two spheniscin isoforms, we established that this peptide has a broad activity spectrum, affecting the growth of both pathogenic bacteria and fungi. Altogether, our data suggest that spheniscins and other, not yet identified, antimicrobial substances may play a role in the long term preservation of stored food in the stomach of king penguins.}, keywords = {Animals, Antimicrobial Cationic Peptides, bacteria, beta-Defensins, Birds, Feeding Behavior, Fungi, Gastrointestinal Contents, hoffmann, M3i, Male, Mass, Matrix-Assisted Laser Desorption-Ionization, Protein Isoforms, Sequence Alignment, Spectrometry}, pubstate = {published}, tppubtype = {article} } @article{bianco_solid-phase_2003, title = {Solid-phase synthesis and characterization of a novel fullerene-peptide derived from histone H3}, author = {Alberto Bianco and Davide Pantarotto and Johan Hoebeke and Jean-Paul Briand and Maurizio Prato}, doi = {10.1039/b311505d}, issn = {1477-0520}, year = {2003}, date = {2003-12-01}, journal = {Organic & Biomolecular Chemistry}, volume = {1}, number = {23}, pages = {4141--4143}, abstract = {A peptide analogue from a histone H3 protein containing the L-fulleropyrrolidino-glutamic acid has been prepared by a solid-phase approach and has been fully characterized. By molecular modelling it was verified that this peptide derivative is able to retain a binding capacity to the MHC (major histocompatibility complex) molecule similar to that of the cognate epitope.}, keywords = {Chromatography, Epitopes, Fullerenes, Glutamic Acid, High Pressure Liquid, Histones, I2CT, Models, Molecular, Molecular Structure, Peptides, Protein Structure, Team-Bianco, Tertiary}, pubstate = {published}, tppubtype = {article} } @article{poschalko_subpol_2003, title = {SUBPOL: A Novel Sucrose-Based Polymer Support for Solid-Phase Peptide Synthesis and Affinity Chromatography Applications}, author = {Alexander Poschalko and Thomas Rohr and Heinrich Gruber and Alberto Bianco and Gilles Guichard and Jean-Paul Briand and Viktoria Weber and Dieter Falkenhagen}, url = {https://pubs.acs.org/doi/10.1021/ja035874a}, doi = {10.1021/ja035874a}, issn = {0002-7863, 1520-5126}, year = {2003}, date = {2003-11-01}, urldate = {2020-11-26}, journal = {Journal of the American Chemical Society}, volume = {125}, number = {44}, pages = {13415--13426}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{goto_silencing_2003, title = {Silencing of Toll pathway components by direct injection of double-stranded RNA into Drosophila adult flies}, author = {Akira Goto and Stéphanie A Blandin and Julien Royet and Jean-Marc Reichhart and Elena A Levashina}, issn = {1362-4962}, year = {2003}, date = {2003-11-01}, journal = {Nucleic Acids Res.}, volume = {31}, number = {22}, pages = {6619--6623}, abstract = {Double-stranded RNA (dsRNA) gene interference is an efficient method to silence gene expression in a sequence-specific manner. Here we show that the direct injection of dsRNA can be used in adult Drosophila flies to disrupt function of endogenous genes in vivo. As a proof of principle, we have used this method to silence components of a major signaling cascade, the Toll pathway, which controls fruit fly resistance to fungal and Gram-positive bacterial infections. We demonstrate that the knockout is efficient only if dsRNA is injected in 4- or more day-old flies and that it lasts for at least 1 week. Furthermore, we report dsRNA-based epistatic gene analysis via injection of a mixture of two dsRNAs and propose that injection of dsRNA represents a powerful method for rapid functional analysis of genes in Drosophila melanogaster adults, particularly of those whose mutations are lethal during development.}, keywords = {Animals, blandin, Cell Surface, Double-Stranded, Epistasis, Female, Genetic, Green Fluorescent Proteins, Homeodomain Proteins, Luminescent Proteins, M3i, Phenotype, Receptors, reichhart, RNA, RNA Interference, Serpins, Signal Transduction, Time Factors, Toll-Like Receptors, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_immune_2003, title = {The immune response of Drosophila}, author = {Jules A Hoffmann}, doi = {10.1038/nature02021}, issn = {1476-4687}, year = {2003}, date = {2003-11-01}, journal = {Nature}, volume = {426}, number = {6962}, pages = {33--38}, abstract = {Drosophila mounts a potent host defence when challenged by various microorganisms. Analysis of this defence by molecular genetics has now provided a global picture of the mechanisms by which this insect senses infection, discriminates between various classes of microorganisms and induces the production of effector molecules, among which antimicrobial peptides are prominent. An unexpected result of these studies was the discovery that most of the genes involved in the Drosophila host defence are homologous or very similar to genes implicated in mammalian innate immune defences. Recent progress in research on Drosophila immune defence provides evidence for similarities and differences between Drosophila immune responses and mammalian innate immunity.}, keywords = {Animals, Cell Surface, hoffmann, Immunity, Innate, M3i, Membrane Glycoproteins, Receptors, Signal Transduction, Toll-Like Receptor 5, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{pantarotto_immunization_2003-1, title = {Immunization with Peptide-Functionalized Carbon Nanotubes Enhances Virus-Specific Neutralizing Antibody Responses}, author = {Davide Pantarotto and Charalambos D Partidos and Johan Hoebeke and Fred Brown and Ed Kramer and Jean-Paul Briand and Sylviane Muller and Maurizio Prato and Alberto Bianco}, url = {http://www.sciencedirect.com/science/article/pii/S107455210300214X}, doi = {10.1016/j.chembiol.2003.09.011}, issn = {1074-5521}, year = {2003}, date = {2003-10-01}, urldate = {2020-03-31}, journal = {Chemistry & Biology}, volume = {10}, number = {10}, pages = {961--966}, abstract = {Functionalized carbon nanotubes (CNTs) hold a lot of promise for application in medicinal chemistry. Based on a method for preparation of water-soluble CNTs, we covalently linked a neutralizing B cell epitope from the foot-and-mouth disease virus (FMDV) to mono- and bis-derivatized CNTs. Immunological characterization of these conjugates revealed that the epitope was appropriately presented after conjugation to CNTs for recognition by antibodies as measured by BIAcore technology. Moreover, peptide-carbon nanotubes elicited strong anti-peptide antibody responses in mice with no detectable cross-reactivity to the carbon nanotubes. However, only the mono-derivitized CNT conjugate induced high levels of virus-neutralizing antibodies. These findings highlight for the first time the potential of CNTs to present biologically important epitopes in an appropriate conformation both in vitro and in vivo and open up the possibility for their use in vaccine delivery.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pantarotto_immunization_2003, title = {Immunization with peptide-functionalized carbon nanotubes enhances virus-specific neutralizing antibody responses}, author = {Davide Pantarotto and Charalambos D Partidos and Johan Hoebeke and Fred Brown and Ed Kramer and Jean-Paul Briand and Sylviane Muller and Maurizio Prato and Alberto Bianco}, doi = {10.1016/j.chembiol.2003.09.011}, issn = {1074-5521}, year = {2003}, date = {2003-10-01}, journal = {Chemistry & Biology}, volume = {10}, number = {10}, pages = {961--966}, keywords = {Animals, Antibodies, Antigen-Antibody Reactions, carbon, Drug Delivery Systems, Epitopes, Foot-and-Mouth Disease Virus, I2CT, Immunization, Mice, Monoclonal, Nanotubes, Neutralization Tests, Peptides, Team-Bianco, Vaccines, Viral}, pubstate = {published}, tppubtype = {article} } @article{weber_binding_2003, title = {Binding of the Drosophila cytokine Spätzle to Toll is direct and establishes signaling}, author = {Alexander N R Weber and Servane Tauszig-Delamasure and Jules A Hoffmann and Eric Lelièvre and Hugues Gascan and Keith P Ray and Mary A Morse and Jean-Luc Imler and Nicholas J Gay}, doi = {10.1038/ni955}, issn = {1529-2908}, year = {2003}, date = {2003-08-01}, journal = {Nature Immunology}, volume = {4}, number = {8}, pages = {794--800}, abstract = {The extracellular protein Spätzle is required for activation of the Toll signaling pathway in the embryonic development and innate immune defense of Drosophila. Spätzle is synthesized as a pro-protein and is processed to a functional form by a serine protease. We show here that the mature form of Spätzle triggers a Toll-dependent immune response after injection into the hemolymph of flies. Spätzle specifically bound to Drosophila cells and to Cos-7 cells expressing Toll. Furthermore, in vitro experiments showed that the mature form of Spätzle bound to the Toll ectodomain with high affinity and with a stoichiometry of one Spätzle dimer to two receptors. The Spätzle pro-protein was inactive in all these assays, indicating that the pro-domain sequence, which is natively unstructured, acts to prevent interaction of the cytokine and its receptor Toll. These results show that, in contrast to the human Toll-like receptors, Drosophila Toll requires only an endogenous protein ligand for activation and signaling.}, keywords = {Animals, Cell Surface, hoffmann, imler, Insect Proteins, M3i, Protein Binding, Protein Structure, Receptors, Signal Transduction, Tertiary, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{hetru_drosophila_2003, title = {Drosophila melanogaster antimicrobial defense}, author = {Charles Hetru and Laurent Troxler and Jules A Hoffmann}, doi = {10.1086/374758}, issn = {0022-1899}, year = {2003}, date = {2003-06-01}, journal = {J. Infect. Dis.}, volume = {187 Suppl 2}, pages = {S327--334}, abstract = {The Drosophila melanogaster host defense is complex but remarkably efficient. It is a multifaceted response to a variety of fungal, bacterial, and parasitic invaders. Current knowledge is discussed on recognition of infectious microorganisms and on the activation of intracellular signaling cascades that concur with the expression of numerous immune-responsive genes, among which, to date, the most prominent appear to encode potent antimicrobial peptides.}, keywords = {Animal, Animals, Bacterial Infections, bioinformatic, hoffmann, Immunity, Innate, M3i, Mycoses, Parasitic Diseases, Peptides, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{bilak_toll_2003, title = {Toll and Toll-like receptors in Drosophila}, author = {Hana Bilak and S Tauszig-Delamasure and Jean-Luc Imler}, doi = {10.1042/}, issn = {0300-5127}, year = {2003}, date = {2003-06-01}, journal = {Biochemical Society Transactions}, volume = {31}, number = {Pt 3}, pages = {648--651}, abstract = {The Drosophila Toll receptor controls the immune response to Gram-positive bacteria and fungi by activating a signalling pathway partially conserved throughout evolution. The Drosophila genome encodes eight additional Toll-related receptors, most of which appear to carry out developmental rather than immune functions. One exception may be Toll-9, which shares structural and functional similarities with mammalian TLRs.}, keywords = {Animals, Biological Evolution, Cell Surface, Fungi, Genome, Gram-Negative Bacteria, Gram-Positive Bacteria, imler, M3i, Membrane Glycoproteins, Receptors, Toll-Like Receptor 5, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{rainaldi_conformational_2003, title = {Conformational analysis by HRMAS NMR spectroscopy of resin-bound homo-peptides from C(alpha)-methyl-leucine}, author = {Mario Rainaldi and Nathalie Lancelot and Karim Elbayed and Jesus Raya and Martial Piotto and Jean-Paul Briand and Bernard Kaptein and Quirinus B Broxterman and Albrecht Berkessel and Fernando Formaggio and Claudio Toniolo and Alberto Bianco}, doi = {10.1039/b303193d}, issn = {1477-0520}, year = {2003}, date = {2003-06-01}, journal = {Organic & Biomolecular Chemistry}, volume = {1}, number = {11}, pages = {1835--1837}, abstract = {A series of [L-(alphaMe)Leu]n (n = 1-5) homo-peptides have been covalently linked to Tentagel and POEPOP resins and submitted to a conformational study using HRMAS NMR spectroscopy. Whereas the mono- and dipeptide are mainly fully-extended, stable 3(10)-helical structures are formed beginning from the trimer.}, keywords = {biomolecular, I2CT, Leucine, Nuclear Magnetic Resonance, Oligopeptides, Protein Structure, Resins, Secondary, Synthetic, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pantarotto_synthesis_2003, title = {Synthesis, structural characterization, and immunological properties of carbon nanotubes functionalized with peptides}, author = {Davide Pantarotto and Charalambos D Partidos and Roland Graff and Johan Hoebeke and Jean-Paul Briand and Maurizio Prato and Alberto Bianco}, doi = {10.1021/ja034342r}, issn = {0002-7863}, year = {2003}, date = {2003-05-01}, journal = {Journal of the American Chemical Society}, volume = {125}, number = {20}, pages = {6160--6164}, abstract = {Carbon nanotubes (NTs) are becoming highly attractive molecules for applications in medicinal chemistry. The main problem of insolubility in aqueous media has been solved by developing a synthetic protocol that allows highly water-soluble carbon NTs to be obtained. As a result, biologically active peptides can be easily linked through a stable covalent bond to carbon NTs. We have demonstrated that a bound peptide from the foot-and-mouth disease virus, corresponding to the 141-159 region of the viral envelope protein VP1, retained the structural integrity and was recognized by monoclonal and polyclonal antibodies. In addition, this peptide-NT conjugate is immunogenic, eliciting antibody responses of the right specificity. Such a system could be greatly advantageous for diagnostic purposes and could find future applications in vaccine delivery.}, keywords = {B-Lymphocyte, biomolecular, Capsid Proteins, carbon, Chromatography, Epitopes, Foot-and-Mouth Disease Virus, High Pressure Liquid, I2CT, nanotechnology, Nanotubes, Nuclear Magnetic Resonance, Peptide Fragments, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{pmid12692243, title = {Recruitment of Tat to heterochromatin protein HP1 via interaction with CTIP2 inhibits human immunodeficiency virus type 1 replication in microglial cells}, author = {Olivier Rohr and Dominique Lecestre and Sylvette Chasserot-Golaz and Céline Marban and Dorina Avram and Dominique Aunis and Mark Leid and Evelyne Schaeffer}, doi = {10.1128/jvi.77.9.5415-5427.2003}, issn = {0022-538X}, year = {2003}, date = {2003-05-01}, urldate = {2003-05-01}, journal = {J Virol}, volume = {77}, number = {9}, pages = {5415--5427}, abstract = {The Tat protein of human immunodeficiency virus type 1 (HIV-1) plays a key role as inducer of viral gene expression. We report that Tat function can be potently inhibited in human microglial cells by the recently described nuclear receptor cofactor chicken ovalbumin upstream promoter transcription factor-interacting protein 2 (CTIP2). Overexpression of CTIP2 leads to repression of HIV-1 replication, as a result of inhibition of Tat-mediated transactivation. In contrast, the related CTIP1 was unable to affect Tat function and viral replication. Using confocal microscopy to visualize Tat subcellular distribution in the presence of the CTIPs, we found that overexpression of CTIP2, and not of CTIP1, leads to disruption of Tat nuclear localization and recruitment of Tat within CTIP2-induced nuclear ball-like structures. In addition, our studies demonstrate that CTIP2 colocalizes and associates with the heterochromatin-associated protein HP1alpha. The CTIP2 protein harbors two Tat and HP1 interaction interfaces, the 145-434 and the 717-813 domains. CTIP2 and HP1alpha associate with Tat to form a three-protein complex in which the 145-434 CTIP2 domain interacts with the N-terminal region of Tat, while the 717-813 domain binds to HP1. The importance of this Tat binding interface and of Tat subnuclear relocation was confirmed by analysis of CTIP2 deletion mutants. Our findings suggest that inhibition of HIV-1 expression by CTIP2 correlates with recruitment of Tat within CTIP2-induced structures and relocalization within inactive regions of the chromatin via formation of the Tat-CTIP2-HP1alpha complex. These data highlight a new mechanism of Tat inactivation through subnuclear relocalization that may ultimately lead to inhibition of viral pathogenesis.}, keywords = {ROHR, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{green_drosophila_2003, title = {Drosophila necrotic mutations mirror disease-associated variants of human serpins}, author = {Clare Green and Gemma Brown and Timothy R Dafforn and Jean-Marc Reichhart and Terri Morley and David A Lomas and David Gubb}, issn = {0950-1991}, year = {2003}, date = {2003-04-01}, journal = {Development}, volume = {130}, number = {7}, pages = {1473--1478}, abstract = {Polymerization of members of the serpin superfamily underlies diseases as diverse as cirrhosis, angioedema, thrombosis and dementia. The Drosophila serpin Necrotic controls the innate immune response and is homologous to human alpha(1)-antitrypsin. We show that necrotic mutations that are identical to the Z-deficiency variant of alpha(1)-antitrypsin form urea-stable polymers in vivo. These necrotic mutations are temperature sensitive, which is in keeping with the temperature-dependent polymerization of serpins in vitro and the role of childhood fevers in exacerbating liver disease in Z alpha-antitrypsin deficiency. In addition, we identify two nec mutations homologous to an antithrombin point mutation that is responsible for neonatal thrombosis. Transgenic flies carrying an StextgreaterF amino-acid substitution equivalent to that found in Siiyama-variant antitrypsin (nec(StextgreaterF.UAS)) fail to complement nec-null mutations and demonstrate a dominant temperature-dependent inactivation of the wild-type nec allele. Taken together, these data establish Drosophila as a powerful system to study serpin polymerization in vivo.}, keywords = {Animals, Humans, M3i, Necrosis, reichhart, Serpins, Temperature, Urea}, pubstate = {published}, tppubtype = {article} } @article{kurz_virulence_2003b, title = {Virulence factors of the human opportunistic pathogen Serratia marcescens identified by in vivo screening}, author = {Léopold C Kurz and Sophie Chauvet and Emmanuel Andrès and Marianne Aurouze and Isabelle Vallet and Gérard P F Michel and Mitch Uh and Jean Celli and Alain Filloux and Sophie De Bentzmann and Ivo Steinmetz and Jules A Hoffmann and Brett B Finlay and Jean-Pierre Gorvel and Dominique Ferrandon and Jonathan J Ewbank}, doi = {10.1093/emboj/cdg159}, isbn = {0261-4189}, year = {2003}, date = {2003-04-01}, journal = {Embo J}, volume = {22}, pages = {1451--60}, abstract = {The human opportunistic pathogen Serratia marcescens is a bacterium with a broad host range, and represents a growing problem for public health. Serratia marcescens kills Caenorhabditis elegans after colonizing the nematode's intestine. We used C.elegans to screen a bank of transposon-induced S.marcescens mutants and isolated 23 clones with an attenuated virulence. Nine of the selected bacterial clones also showed a reduced virulence in an insect model of infection. Of these, three exhibited a reduced cytotoxicity in vitro, and among them one was also markedly attenuated in its virulence in a murine lung infection model. For 21 of the 23 mutants, the transposon insertion site was identified. This revealed that among the genes necessary for full in vivo virulence are those that function in lipopolysaccharide (LPS) biosynthesis, iron uptake and hemolysin production. Using this system we also identified novel conserved virulence factors required for Pseudomonas aeruginosa pathogenicity. This study extends the utility of C.elegans as an in vivo model for the study of bacterial virulence and advances the molecular understanding of S.marcescens pathogenicity.}, keywords = {*Virulence, Animal, Caenorhabditis elegans/*microbiology, ferrandon, hoffmann, M3i, Mutation, Non-U.S. Gov't, Serratia marcescens/genetics/*pathogenicity, Support}, pubstate = {published}, tppubtype = {article} } @article{lancelot_characterization_2003, title = {Characterization of the 310-helix in model peptides by HRMAS NMR spectroscopy}, author = {Nathalie Lancelot and Karim Elbayed and Jésus Raya and Martial Piotto and Jean-Paul Briand and Fernando Formaggio and Claudio Toniolo and Alberto Bianco}, doi = {10.1002/chem.200390151}, issn = {0947-6539}, year = {2003}, date = {2003-03-01}, journal = {Chemistry (Weinheim an Der Bergstrasse, Germany)}, volume = {9}, number = {6}, pages = {1317--1323}, abstract = {A tetra- and a hepta-homopeptide from the C(alpha)-tetrasubstituted Aib (alpha-aminoisobutyric acid) residue were covalently linked to the POEPOP resin by the fragment-condensation approach. The conformational preferences of the two model peptides were determined for the first time on a solid support by means of high-resolution magic angle spinning NMR spectroscopy. The results obtained indicate that the Aib homopeptides adopt a regular 3(10)-helical structure even when they are covalently bound to a polymeric matrix, and thus confirm the remarkable conformational stability of the peptides rich in this amino acid. An ATR-FTIR spectroscopic investigation, performed in parallel, also confirmed that these polymer-bound peptides do indeed adopt a helical conformation. The results of this study open the possibility to exploit the peptide-resin conjugates based on C(alpha)-tetrasubstituted alpha-amino acids as helpful, structurally organized templates in molecular recognition studies or as catalysts in asymmetric synthesis.}, keywords = {biomolecular, Fourier Transform Infrared, I2CT, Nuclear Magnetic Resonance, Peptides, Protein Conformation, spectroscopy, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{luna_characterization_2003, title = {Characterization of three Toll-like genes from mosquito Aedes aegypti}, author = {C Luna and N T Hoa and J Zhang and S M Kanzok and S E Brown and Jean-Luc Imler and D L Knudson and L Zheng}, issn = {0962-1075}, year = {2003}, date = {2003-02-01}, journal = {Insect Molecular Biology}, volume = {12}, number = {1}, pages = {67--74}, abstract = {Three Toll-related genes (AeToll1A, AeToll1B and AeToll5) were cloned and characterized from the yellow fever vector mosquito, Aedes aegypti. All three genes exhibited high levels of amino acid sequence similarity with Drosophila melanogaster (Dm)Toll1 and DmTehao (Toll5). AeToll1A and AeToll1B are 1124 and 1076 amino acid residues long, respectively. Both contain a carboxyl extension downstream of the Toll/interleukin-1 receptor (TIR) domain. AeToll5 is 1007 residues long and, like DmTehao, lacks the carboxyl terminal extension. Expression of these three genes was examined throughout development and after immune challenge. Both AeToll1A and AeToll5, like their Drosophila counterparts, activate transcription of drosomycin promoter in both Aedes and Drosophila cell lines. Deletion of the carboxyl extension of AeToll1A did not result in a further elevated level of the antifungal response. The intracellular signalling process appears to be species specific based on two observations. (1) DmToll is completely inactive in an Aedes cell line, suggesting a higher specificity requirement for DmToll in the intracellular signalling process. (2) Only one of three amino acid residues essential for DmToll function is required for AeToll1A function.}, keywords = {Aedes, Animals, Base Sequence, Cell Surface, Chimera, Cloning, Developmental, Female, Gene Expression Regulation, Genetic, imler, Insect Proteins, M3i, Male, messenger, Models, Molecular, Mutagenesis, Promoter Regions, Receptors, Reverse Transcriptase Polymerase Chain Reaction, RNA, Sequence Alignment, Signal Transduction, Site-Directed, Transfection}, pubstate = {published}, tppubtype = {article} } @article{imler_toll_2003, title = {Toll signaling: the TIReless quest for specificity}, author = {Jean-Luc Imler and Jules A Hoffmann}, doi = {10.1038/ni0203-105}, issn = {1529-2908}, year = {2003}, date = {2003-02-01}, journal = {Nature Immunology}, volume = {4}, number = {2}, pages = {105--106}, keywords = {Animals, Cell Surface, Dendritic Cells, hoffmann, Humans, imler, Immunological, Interferon-beta, M3i, Membrane Glycoproteins, Mice, Models, Protein Structure, Receptors, Signal Transduction, Tertiary, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{monneaux_t_2003, title = {T cell recognition and therapeutic effect of a phosphorylated synthetic peptide of the 70K snRNP protein administered in MR/lpr mice}, author = {Fanny Monneaux and José Manuel Lozano and Manuel E Patarroyo and Jean-Paul Briand and Sylviane Muller}, doi = {10.1002/immu.200310002}, issn = {0014-2980}, year = {2003}, date = {2003-01-01}, journal = {European Journal of Immunology}, volume = {33}, number = {2}, pages = {287--296}, abstract = {Modifications of self antigens that occur during apoptosis might be involved in the generation of neo-antigens, which can break tolerance and induce autoimmunity. We have previously identified an epitope at residues 131-151 of the U1-70K snRNP protein, recognized by IgG antibodies and CD4+ T cells from at least two strains of lupus mice. With the aim of investigating the possible role of phosphorylation on the antigenicity of peptide 131-151 and to gain a better understanding of how this peptide can drive autoimmune response, we synthesized two peptides phosphorylated on Ser137 and 140, respectively. We show here that peptide P140 phosphorylated on Ser140 is recognized by both CD4+ T cells and antibodies from MRL/lpr mice. Furthermore, intravenous administration to lupus-prone MRL/lpr mice of P140 in saline (but not of the non-phosphorylated peptide) decreased proteinuria and anti-DNA antibody production, and significantly prolonged survival of treated mice. We further demonstrated that P140 is recognized by antibodies from lupus patients and binds to various HLA DR molecules, offering new hope for manipulating T cell response in humans.}, keywords = {Amino Acid Sequence, Animal, Animals, Autoantibodies, Autoantigens, Autoimmune Diseases, B-Lymphocytes, Cross Reactions, Disease Models, Female, HLA-DR Antigens, HLA-DR Serological Subtypes, HLA-DR1 Antigen, HLA-DR4 Antigen, Humans, I2CT, Immunization, Immunotherapy, Inbred BALB C, Inbred MRL lpr, Lupus Erythematosus, Lupus Nephritis, Mice, Molecular Sequence Data, Monneaux, Peptide Fragments, Phosphorylation, Protein Binding, Ribonucleoprotein, Systemic, T-Lymphocytes, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{bianco_can_2003, title = {Can Carbon Nanotubes be Considered Useful Tools for Biological Applications?}, author = {A Bianco and M Prato}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/adma.200301646}, doi = {10.1002/adma.200301646}, issn = {1521-4095}, year = {2003}, date = {2003-01-01}, urldate = {2020-03-31}, journal = {Advanced Materials}, volume = {15}, number = {20}, pages = {1765--1768}, abstract = {Carbon nanotubes can be made soluble in both organic solvents and in aqueous solutions by organic functionalization. In particular, soluble carbon nanotubes can be further derivatized by coupling with amino acids and bioactive peptides. Immobilization of peptides to the external walls of carbon nanotubes may find interesting applications in diagnostics, vaccine and drug delivery, or multipresentation of bioactive molecules.}, keywords = {Carbon nanotubes, Functionalization, I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{, title = {The central PPT of the yeast retrotransposon Ty1 is not essential for transposition}, author = { T. Heyman and M. Wilhelm and F. X. Wilhelm}, year = {2003}, date = {2003-01-01}, journal = {J Mol Biol}, volume = {331}, number = {2}, pages = {315-20}, abstract = {The yeast retrotransposon Ty1 has structural and functional similarities to retroviruses. We report here that, as in retroviruses, the plus-strand DNA of Ty1 is synthesized as two segments. A central DNA flap is formed during reverse transcription consecutive to elongation (with strand displacement) of the upstream segment beyond the central polypurine tract (cPPT) until the replication machinery is stopped at the central termination sequence. Comparison of wild-type and cPPT-mutant Ty1 elements shows that the mutant element lacking the central DNA flap is only twofold defective in transposition.}, note = {0022-2836 Journal Article}, keywords = {Base, cerevisiae/genetics, Data, DNA/*biosynthesis, Genetic, Gov't, Models, Molecular, Mutation, Non-U.S., Purines/*chemistry, Retroelements/*genetics, Saccharomyces, Sequence, Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Purification of turkey pancreatic phospholipase A2}, author = { R. Ben Salah and N. Zouari and J. Reinbolt and H. Mejdoub}, year = {2003}, date = {2003-01-01}, journal = {Biosci Biotechnol Biochem}, volume = {67}, number = {10}, pages = {2139-44}, abstract = {Turkey pancreatic phospholipase (TPP) has been purified from delipidated pancreases. The purification included ammonium sulfate fractionation, acidic (pH 5) treatment, followed by sequencial column chromatographies on DEAE-cellulose, Sephadex G-75, and reverse phase high pressure liquid chromatography. The purified enzyme was found to be a monomeric protein with molecular mass of 14 kDa. The optimal activity was measured at pH 8 and 37 degrees C using egg yolk emulsion as substrate. Our results show that the enzyme (TPP) was not stable for 1 h at 60 degrees C, and that bile salt and Ca2+ were required for the expression of the purified enzyme. The sequence of the N-terminal amino acids of the purified enzyme shows a very close similarity between TPP and all other known pancreatic phospholipases.}, note = {0916-8451 Journal Article}, keywords = {*Turkeys, &, A/*isolation, Acid, acids, Amino, Ammonium, and, Animals, Bile, Calcium, Chromatography, Concentration, Data, Hydrogen-Ion, Molecular, Pancreas/*enzymology, Phospholipases, purification, Salts, Sequence, Sulfate, Temperature, Weight}, pubstate = {published}, tppubtype = {article} } @article{, title = {A single internal ribosome entry site containing a G quartet RNA structure drives fibroblast growth factor 2 gene expression at four alternative translation initiation codons}, author = { S. Bonnal and C. Schaeffer and L. Creancier and S. Clamens and H. Moine and A. C. Prats and S. Vagner}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {41}, pages = {39330-6}, abstract = {The 484-nucleotide (nt) alternatively translated region (ATR) of the human fibroblast growth factor 2 (FGF-2) mRNA contains four CUG and one AUG translation initiation codons. Although the 5'-end proximal CUG codon is initiated by a cap-dependent translation process, the other four initiation codons are initiated by a mechanism of internal entry of ribosomes. We undertook here a detailed analysis of the cis-acting elements defining the FGF-2 internal ribosome entry site (IRES). A thorough deletion analysis study within the 5'-ATR led us to define a 176-nt region as being necessary and sufficient for IRES function at four codons present in a downstream 308-nt RNA segment. Unexpectedly, a single IRES module is therefore responsible for translation initiation at four distantly localized codons. The determination of the FGF-2 5'-ATR RNA secondary structure by enzymatic and chemical probing experiments showed that the FGF-2 IRES contained two stem-loop regions and a G quartet motif that constitute novel structural determinants of IRES function.}, note = {0021-9258 Journal Article}, keywords = {2/*genetics, Acid, Alternative, Base, Cell, Chain, Codon, Complementary/genetics, Conformation, Data, Deletion, DNA, Expression, Factor, Fibroblast, Gene, Gov't, Growth, Human, initiation, Initiator/genetics, Line, Messenger/*chemistry/*genetics, Molecular, Non-U.S., Nucleic, Peptide, Ribosomes/*metabolism, RNA, Sequence, Splicing, Support, Transfection}, pubstate = {published}, tppubtype = {article} } @article{, title = {Global DNA methylation in relation to hMLH1 and hMSH2 protein immunoreactivity in sporadic human endometrial carcinomas}, author = { R. Miturski and K. Postawski and A. Semczuk and M. Bogusiewicz and W. Baranowski and J. A. Jakowicki and G. Keith}, year = {2003}, date = {2003-01-01}, journal = {Int J Mol Med}, volume = {11}, number = {5}, pages = {569-74}, abstract = {Overall DNA methylation status was studied in a group of 28 sporadic human endometrial carcinomas (ECs) using the [32P]-postlabeling technique. Moreover, expression of the DNA mismatch repair proteins (hMLH1 and hMSH2) was investigated in ECs using immunohistochemistry. Mean 5-methyldeoxycytosine (m5dC) content in the studied group was 3.48+/-0.37% (range, 2.89-4.12%). The mean m5dC scores were significantly different between early (3.35+/-0.33%) and advanced (3.66+/-0.36%) endometrial neoplasms (chi2-test; p=0.03). There was a markedly increased overall DNA methylation with the degree of histological differentiation and with the infiltration of the myometrium (p<0.05). Loss of hMLH1 and hMSH2 expression was reported in 7 (25%) and 5 (18%) tumors, respectively, but the immunoreactivity did not correlate with the known clinicopathological variables of cancer. In addition, no obvious correlation was found between global m5dC content and the lack of hMLH1 and hMSH2 protein expression in human uterine tumors (p=0.97 and p=0.19 for hMLH1 and hMSH2, respectively; Spearman's rank correlation test). Our results clearly show that alterations in global DNA methylation may influence tumor progression, but they are not directly associated with the inactivation of the mismatch-repair machinery in sporadic human ECs.}, note = {1107-3756 Journal Article}, keywords = {*DNA, Base, Carcinoma/genetics/*metabolism/pathology, DNA, Endometrial, Female, Gov't, Human, Immunohistochemistry, Methylation, Mismatch, Neoplasm, Neoplasms/genetics/*metabolism/pathology, Non-U.S., Pair, Proteins/*metabolism, Proto-Oncogene, Repair, Support}, pubstate = {published}, tppubtype = {article} } @article{kambris_dmmyd88_2003, title = {DmMyD88 controls dorsoventral patterning of the Drosophila embryo}, author = {Zakaria Kambris and Hana Bilak and Rosalba D'Alessandro and Marcia Belvin and Jean-Luc Imler and Maria Capovilla}, doi = {10.1038/sj.embor.embor714}, issn = {1469-221X}, year = {2003}, date = {2003-01-01}, journal = {EMBO reports}, volume = {4}, number = {1}, pages = {64--69}, abstract = {MyD88 is an adapter protein in the signal transduction pathway mediated by interleukin-1 (IL-1) and Toll-like receptors. A Drosophila homologue of MyD88 (DmMyD88) was recently shown to be required for the Toll-mediated immune response. In Drosophila, the Toll pathway was originally characterized for its role in the dorsoventral patterning of the embryo. We found that, like Toll, DmMyD88 messenger RNA is maternally supplied to the embryo. Here we report the identification of a new mutant allele of DmMyD88, which generates a protein lacking the carboxy-terminal extension, normally located downstream of the Toll/IL-1 receptor domain. Homozygous mutant female flies lay dorsalized embryos that are rescued by expression of a transgenic DmMyD88 complementary DNA. The DmMyD88 mutation blocks the ventralizing activity of a gain-of-function Toll mutation. These results show that DmMyD88 encodes an essential component of the Toll pathway in dorsoventral pattern formation.}, keywords = {Adaptor Proteins, Alleles, Animals, Antigens, Base Sequence, Cell Surface, Complementary, Developmental, Differentiation, DNA, DNA Transposable Elements, Egg Proteins, Embryo, Exons, Female, Gene Expression Regulation, Genetically Modified, Genotype, imler, Immunity, Immunologic, Innate, Insertional, M3i, Male, messenger, Morphogenesis, Mutagenesis, Myeloid Differentiation Factor 88, Nonmammalian, Oocytes, Protein Biosynthesis, Protein Structure, Receptors, Reverse Transcriptase Polymerase Chain Reaction, RNA, Signal Transducing, Tertiary, Toll-Like Receptors, Zygote}, pubstate = {published}, tppubtype = {article} } @article{sabatier_pherokine-2_2003, title = {Pherokine-2 and -3}, author = {Laurence Sabatier and Emmanuelle Jouanguy and Catherine Dostert and Daniel Zachary and Jean-Luc Dimarcq and Philippe Bulet and Jean-Luc Imler}, issn = {0014-2956}, year = {2003}, date = {2003-01-01}, journal = {European journal of biochemistry / FEBS}, volume = {270}, number = {16}, pages = {3398--3407}, abstract = {Drosophila is a powerful model system to study the regulatory and effector mechanisms of innate immunity. To identify molecules induced in the course of viral infection in this insect, we have developed a model based on intrathoracic injection of the picorna-like Drosophila C virus (DCV). We have used MALDI-TOF mass spectrometry to compare the hemolymph of DCV infected flies and control flies. By contrast with the strong humoral response triggered by injection of bacteria or fungal spores, we have identified only one molecule induced in the hemolymph of virus infected flies. This molecule, pherokine-2 (Phk-2), is related to OS-D/A10 (Phk-1), which was previously characterized as a putative odor/pheromone binding protein specifically expressed in antennae. The virus-induced molecule is also similar to the product of the gene CG9358 (Phk-3), which is induced by septic injury. Both Phk-2 and Phk-3 are strongly expressed during metamorphosis, suggesting that they may participate in tissue-remodeling.}, keywords = {Animals, Antibody Formation, Base Sequence, Hemolymph, imler, M3i, Mass, Matrix-Assisted Laser Desorption-Ionization, Spectrometry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Extension and cleavage of the polypurine tract plus-strand primer by Ty1 reverse transcriptase}, author = { F. X. Wilhelm and M. Wilhelm and A. Gabriel}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {48}, pages = {47678-84}, abstract = {Using hybrid RNA/DNA substrates containing the polypurine tract (PPT) plus-strand primer, we have examined the interaction between the Ty1 reverse transcriptase (RT) and the plus-strand initiation complex. We show here that, although the PPT sequence is relatively resistant to RNase H cleavage, it can be cleaved internally by the polymerase-independent RNase H activity of Ty1 RT. Alternatively, this PPT can be used to initiate plus-strand DNA synthesis. We demonstrate that cleavage at the PPT/DNA junction occurs only after at least 9 nucleotides are extended. Cleavage leaves a nick between the RNA primer and the nascent plus-strand DNA. We show that Ty1 RT has a strand displacement activity beyond a gap but that the PPT is not efficiently re-utilized in vitro for another round of DNA synthesis after a first plus-strand DNA has been synthesized and cleaved at the PPT/U3 junction.}, note = {0021-9258 Journal Article}, keywords = {Base, Calf, Data, DNA, DNA/chemistry, Genetic, Gov't, H, Messenger/metabolism, Models, Molecular, Non-U.S., P.H.S., Polymerase/*chemistry, Primers, Proteins/chemistry, Purines/*chemistry, Recombinant, Replication, Retroelements/*genetics, Ribonuclease, RNA, RNA-Directed, RNA/chemistry, Sequence, Support, Templates, Thymus/chemistry, U.S., Viral}, pubstate = {published}, tppubtype = {article} } @article{, title = {Enzymes assembled from Aquifex aeolicus and Escherichia coli leucyl-tRNA synthetases}, author = {M W Zhao and R Hao and J F Chen and F Martin and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12820878}, isbn = {12820878}, year = {2003}, date = {2003-01-01}, journal = {Biochemistry}, volume = {42}, number = {25}, pages = {7694-7700}, abstract = {Aquifex aeolicus alphabeta-LeuRS is the only known heterodimeric LeuRS, while Escherichia coli LeuRS is a canonical monomeric enzyme. By using the genes encoding A. aeolicus and E. coli LeuRS as PCR templates, the genes encoding the alpha and beta subunits from A. aeolicus alphabeta-LeuRS and the equivalent amino- and carboxy-terminal parts of E. coli LeuRS (identified as alpha' and beta') were amplified and recombined using suitable plasmids. These recombinant plasmids were transformed or cotransformed into E. coli to produce five monomeric and five heterodimeric LeuRS mutants. Seven of these were successfully overexpressed in vivo and purified, while three dimeric mutants with the beta' part of E. coli LeuRS were not successfully expressed. The seven purified mutants catalyzed amino acid activation, although several exhibited reduced aminoacylation properties. Removal of the last 36 residues of the alpha subunit of the A. aeolicus enzyme was determined to be deleterious for tRNA charging. Indeed, subunit exchange showed that the cross-species-specific recognition of A. aeolicus tRNA(Leu) occurs at the alpha subunit. None of the mixed E. coli-A. aeolicus enzymes were as thermostable as the native alphabeta-LeuRS. However, the fusion of the two alpha and beta peptides from A. aeolicus as a single chain analogous to canonical LeuRS resulted in a product more resistant to heat denaturation than the original enzyme.}, note = {0006-2960 Journal Article}, keywords = {Amino Acyl-tRNA Ligases/genetics/*metabolism Escherichia coli/*enzymology Evolution, ERIANI, Horizontal Heat Kinetics Mutation RNA, Leu/*metabolism Structure-Activity Relationship Support, Molecular Gene Transfer, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Tools for the automatic identification and classification of RNA base pairs}, author = {H Yang and F Jossinet and N Leontis and L Chen and J Westbrook and H Berman and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12824344}, isbn = {12824344}, year = {2003}, date = {2003-01-01}, journal = {Nucleic Acids Res}, volume = {31}, number = {13}, pages = {3450-3460}, abstract = {Three programs have been developed to aid in the classification and visualization of RNA structure. BPViewer provides a web interface for displaying three-dimensional (3D) coordinates of individual base pairs or base pair collections. A web server, RNAview, automatically identifies and classifies the types of base pairs that are formed in nucleic acid structures by various combinations of the three edges, Watson-Crick, Hoogsteen and the Sugar edge. RNAView produces two-dimensional (2D) diagrams of secondary and tertiary structure in either Postscript, VRML or RNAML formats. The application RNAMLview can be used to rearrange various parts of the RNAView 2D diagram to generate a standard representation (like the cloverleaf structure of tRNAs) or any layout desired by the user. A 2D diagram can be rapidly reformatted using RNAMLview since all the parts of RNA (like helices and single strands) are dynamically linked while moving the selected parts. With the base pair annotation and the 2D graphic display, RNA motifs are rapidly identified and classified. A survey has been carried out for 41 unique structures selected from the NDB database. The statistics for the occurrence of each edge and of each of the 12 bp families are given for the combinations of the four bases: A, G, U and C. The program also allows for visualization of the base pair interactions by using a symbolic convention previously proposed for base pairs. The web servers for BPViewer and RNAview are available at http://ndbserver.rutgers.edu/services/. The application RNAMLview can also be downloaded from this site. The 2D diagrams produced by RNAview are available for RNA structures in the Nucleic Acid Database (NDB) at http://ndbserver.rutgers.edu/atlas/.}, note = {1362-4962 Journal Article}, keywords = {Algorithms Base Pairing Base Sequence Computer Graphics Data Interpretation, Molecular Nucleic Acid Conformation RNA/*chemistry/classification *Software Support, Non-P.H.S. Support, Non-U.S. Gov't Support, Nucleic Acid Internet Models, P.H.S., Statistical Databases, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Extension and cleavage of the polypurine tract plus-strand primer by Ty1 reverse transcriptase}, author = {F X Wilhelm and M Wilhelm and A Gabriel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14500728}, isbn = {14500728}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {48}, pages = {47678-47684}, abstract = {Using hybrid RNA/DNA substrates containing the polypurine tract (PPT) plus-strand primer, we have examined the interaction between the Ty1 reverse transcriptase (RT) and the plus-strand initiation complex. We show here that, although the PPT sequence is relatively resistant to RNase H cleavage, it can be cleaved internally by the polymerase-independent RNase H activity of Ty1 RT. Alternatively, this PPT can be used to initiate plus-strand DNA synthesis. We demonstrate that cleavage at the PPT/DNA junction occurs only after at least 9 nucleotides are extended. Cleavage leaves a nick between the RNA primer and the nascent plus-strand DNA. We show that Ty1 RT has a strand displacement activity beyond a gap but that the PPT is not efficiently re-utilized in vitro for another round of DNA synthesis after a first plus-strand DNA has been synthesized and cleaved at the PPT/U3 junction.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence DNA/chemistry DNA Primers DNA Replication Models, Calf Thymus/chemistry Support, Genetic, Genetic Molecular Sequence Data Purines/*chemistry RNA/chemistry RNA, Messenger/metabolism RNA, Non-U.S. Gov't Support, P.H.S. Templates, U.S. Gov't, Unité ARN, Viral RNA-Directed DNA Polymerase/*chemistry Recombinant Proteins/chemistry Retroelements/*genetics Ribonuclease H}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of geneticin bound to a bacterial 16S ribosomal RNA A site oligonucleotide}, author = {Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12589761}, isbn = {12589761}, year = {2003}, date = {2003-01-01}, journal = {J Mol Biol}, volume = {326}, number = {4}, pages = {1175-1188}, abstract = {Aminoglycosides are antibacterial molecules that decrease translation accuracy by binding to the decoding aminoacyl-tRNA site (A site) on 16S ribosomal RNA. We have solved the crystal structure of an RNA fragment containing the A site bound to geneticin at 2.40A resolution. Geneticin, also known as G418, is a gentamicin-related aminoglycoside: it contains three rings that are functionalized by hydroxyl, ammonium and methyl groups. The detailed comparison of the distinctive behaviour of geneticin (binding to pro- and eukaryotic A sites) with the crystallographic, biochemical and microbiological results obtained so far for aminoglycoside-A site complexes offers new insights on the system. The two sugar rings constituting the neamine part common to most of the aminoglycosides bind to the A site, as already observed in the crystal structures solved previously with paromomycin and tobramycin. The essential hydrogen bonds involving ring I (to A1408) and ring II (to the phosphate oxygen atoms of the bulged adenine bases 1492 and 1493 and to G1494) are conserved and additional contacts are observed from ring III (to phosphate oxygen atoms of G1405 and U1406). The present work illustrates a molecular basis of the range in sensitiveness exhibited by geneticin towards common point A site mutations associated to resistance phenotypes. In addition, analysis and comparisons of the structures cast light on the role played by the conserved U1406.U1495 pair in the recognition of the A site by aminoglycosides.}, note = {0022-2836 Journal Article}, keywords = {16S/*chemistry/genetics/metabolism Support, Anti-Bacterial Agents/*chemistry/metabolism Binding Sites Crystallography, Bacterial Gentamicins/*chemistry/metabolism Human Models, Molecular Molecular Structure *Nucleic Acid Conformation Oligonucleotides/*chemistry/genetics/metabolism Protein Binding *Protein Conformation RNA, Non-U.S. Gov't, Ribosomal, Unité ARN, WESTHOF, X-Ray Drug Design Genes}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular recognition of aminoglycoside antibiotics by ribosomal RNA and resistance enzymes: an analysis of x-ray crystal structures}, author = {Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12925992}, isbn = {12925992}, year = {2003}, date = {2003-01-01}, journal = {Biopolymers}, volume = {70}, number = {1}, pages = {42-57}, abstract = {The potential of RNA molecules to be used as therapeutic targets by small inhibitors is now well established. In this fascinating wide-open field, aminoglycoside antibiotics constitute the most studied family of RNA binding drugs. Within the last three years, several x-ray crystal structures were solved for aminoglycosides complexed to one of their main natural targets in the bacterial cell, the decoding aminoacyl-tRNA site (A site). Other crystallographic structures have revealed the binding modes of aminoglycosides to the three existing types of resistance-associated enzymes. The present review summarizes the various aspects of the molecular recognition of aminoglycosides by these natural RNA or protein receptors. The analysis and the comparisons of the detailed interactions offer insights that are helpful in designing new generations of antibiotics.}, note = {0006-3525 Journal Article Review Review, Tutorial}, keywords = {Aminoglycosides/*chemistry Anti-Bacterial Agents/*chemistry Binding Sites Crystallography, Chemical Models, Microbial Enzymes/*chemistry Models, Molecular Protein Binding RNA/chemistry RNA, Ribosomal/*chemistry Ribosomes/chemistry, Unité ARN, WESTHOF, X-Ray/*methods *Drug Resistance}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA as a drug target: the case of aminoglycosides}, author = {Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14523919}, isbn = {14523919}, year = {2003}, date = {2003-01-01}, journal = {Chembiochem}, volume = {4}, number = {10}, pages = {1018-1023}, note = {1439-4227 Journal Article Review Review, Tutorial}, keywords = {16S/*chemistry/drug effects/metabolism Substrate Specificity Technology, Aminoglycosides/*pharmacology Anti-Bacterial Agents/*pharmacology Binding Sites Drug Delivery Systems Models, Catalytic/chemistry/metabolism RNA, Molecular RNA/chemistry/metabolism RNA, Pharmaceutical Water/chemistry, Ribosomal, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Physical aspects of protein crystal growth investigated with the Advanced Protein Crystallization Facility in reduced-gravity environments}, author = {A Vergara and B Lorber and A Zagari and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12499533}, isbn = {12499533}, year = {2003}, date = {2003-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {59}, number = {Pt 1}, pages = {2-15}, abstract = {The physicochemical aspects of protein crystallization in reduced-gravity environments (micro g) have been investigated with the Advanced Protein Crystallization Facility during six space missions. This review summarizes the results, dealing with the mechanisms of nucleation and crystal growth and with the quality of the crystals that were obtained under reduced gravity as well as under normal gravity on earth. Statistical analyses of the experimental data strongly support the fact that micro g has a positive effect on crystallization and on crystal quality. A comparison of experiments and theories of protein crystallization in reduced-gravity environments is presented. Recommendations for improving the performance of protein crystallization experiments in micro g and on earth are discussed.}, note = {0907-4449 Journal Article Review Review Literature}, keywords = {Animals Comparative Study Computer Simulation *Crystallization Crystallography/instrumentation/methods Data Interpretation, Computer-Assisted Proteins/*chemistry Space Flight Spacecraft Support, Non-U.S. Gov't *Weightlessness, Statistical Human Kinetics Numerical Analysis, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular modeling of the three-dimensional structure of the bacterial RNase P holoenzyme}, author = {H Y Tsai and B Masquida and R Biswas and E Westhof and V Gopalan}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12507471}, isbn = {12507471}, year = {2003}, date = {2003-01-01}, journal = {J Mol Biol}, volume = {325}, number = {4}, pages = {661-675}, abstract = {Bacterial ribonuclease P (RNase P), an enzyme involved in tRNA maturation, consists of a catalytic RNA subunit and a protein cofactor. Comparative phylogenetic analysis and molecular modeling have been employed to derive secondary and tertiary structure models of the RNA subunits from Escherichia coli (type A) and Bacillus subtilis (type B) RNase P. The tertiary structure of the protein subunit of B.subtilis and Staphylococcus aureus RNase P has recently been determined. However, an understanding of the structure of the RNase P holoenzyme (i.e. the ribonucleoprotein complex) is lacking. We have now used an EDTA-Fe-based footprinting approach to generate information about RNA-protein contact sites in E.coli RNase P. The footprinting data, together with results from other biochemical and biophysical studies, have furnished distance constraints, which in turn have enabled us to build three-dimensional models of both type A and B versions of the bacterial RNase P holoenzyme in the absence and presence of its precursor tRNA substrate. These models are consistent with results from previous studies and provide both structural and mechanistic insights into the functioning of this unique catalytic RNP complex.}, note = {0022-2836 Journal Article}, keywords = {Amino Acid Sequence Base Sequence Catalytic Domain Computer Simulation Cysteine/chemistry DNA Footprinting DNA, Bacterial/chemistry/genetics/metabolism RNA, Bacterial/genetics Edetic Acid Endoribonucleases/*chemistry/genetics/metabolism Escherichia coli/*enzymology/genetics Evolution, Catalytic/*chemistry/genetics/metabolism Ribonuclease P Support, Molecular Ferrous Compounds Holoenzymes/chemistry/genetics/metabolism Hydroxyl Radical/chemistry Models, Molecular Molecular Sequence Data Mutagenesis, Non-P.H.S. Support, P.H.S., Site-Directed Nucleic Acid Conformation Protein Subunits RNA, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Proteomic consequences of a human mitochondrial tRNA mutation beyond the frame of mitochondrial translation}, author = {P Tryoen-Toth and S Richert and B Sohm and M Mine and C Marsac and A Van Dorsselaer and E Leize and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12714596}, isbn = {12714596}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {27}, pages = {24314-24323}, abstract = {Numerous severe neurodegenerative and neuromuscular disorders, characterized biochemically by strong perturbations in energy metabolism, are correlated with single point mutations in mitochondrial genes coding for transfer RNAs. Initial comparative proteomics performed on wild-type and Myoclonic Epilepsy and Ragged Red Fibers (MERRF) mitochondria from sibling human cybrid cell lines revealed the potential of this approach. Here a quantitative analysis of several hundred silver-stained spots separated by two-dimensional gel electrophoresis was performed in the specific case of a couple of mitochondria, containing or not mutation A8344G in the gene for mitochondrial tRNALys, correlated with MERRF syndrome. Computer-assisted analysis allowed us to detect 38 spots with significant quantitative variations, of which 20 could be assigned by mass spectrometry. These include nuclear encoded proteins located in mitochondria such as respiratory chain subunits, metabolic enzymes, a protein of the mitochondrial translation machinery, and cytosolic contaminants. Furthermore, Western blotting combined with mass spectrometry revealed the occurrence of numerous isoforms of pyruvate dehydrogenase subunits, with subtle changes in post-translational modifications. This comparative proteomic approach gives the first insight for nuclear encoded proteins that undergo the largest quantitative changes, and pinpoints new potential molecular partners involved in the cascade of events that connect genotype to phenotype.}, note = {0021-9258 Journal Article}, keywords = {Cultured, FLORENTZ, Genetic Tumor Cells, Human Mitochondria/genetics *Mutation Nuclear Proteins/*genetics Proteomics RNA/*genetics RNA, Non-U.S. Gov't Translation, Transfer/*genetics Structure-Activity Relationship Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structures of the Pyrococcus abyssi Sm core and its complex with RNA. Common features of RNA binding in Archaea and Eucarya.}, author = {S Thore and C Mayer and C Sauter and S Weeks and D Suck}, url = {http://www.ncbi.nlm.nih.gov/pubmed/12409299}, isbn = {12409299}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {2}, pages = {1239-1247}, abstract = {The Sm proteins are conserved in all three domains of life and are always associated with U-rich RNA sequences. Their proposed function is to mediate RNA-RNA interactions. We present here the crystal structures of Pyrococcus abyssi Sm protein (PA-Sm1) and its complex with a uridine heptamer. The overall structure of the protein complex, a heptameric ring with a central cavity, is similar to that proposed for the eukaryotic Sm core complex and found for other archaeal Sm proteins. RNA molecules bind to the protein at two different sites. They interact specifically inside the ring with three highly conserved residues, defining the uridine-binding pocket. In addition, nucleotides also interact on the surface formed by the N-terminal alpha-helix as well as a conserved aromatic residue in beta-strand 2 of the PA-Sm1 protein. The mutation of this conserved aromatic residue shows the importance of this second site for the discrimination between RNA sequences. Given the high structural homology between archaeal and eukaryotic Sm proteins, the PA-Sm1.RNA complex provides a model for how the small nuclear RNA contacts the Sm proteins in the Sm core. In addition, it suggests how Sm proteins might exert their function as modulators of RNA-RNA interactions.}, keywords = {FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Spatial and temporal expression patterns of selenoprotein genes during embryogenesis in zebrafish}, author = {C Thisse and A Degrave and G V Kryukov and V N Gladyshev and S Obrecht-Pflumio and A Krol and B Thisse and A Lescure}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12915322}, isbn = {12915322}, year = {2003}, date = {2003-01-01}, journal = {Gene Expr Patterns}, volume = {3}, number = {4}, pages = {525-532}, abstract = {Selenium is important for embryogenesis in vertebrates but little is known about the expression patterns and biological functions of most selenoprotein genes. Taking advantage of the zebrafish model, systematic analysis of selenoprotein gene expression was performed by in situ hybridization on whole-mount embryos at different developmental stages. Twenty-one selenoprotein mRNAs were analyzed and all of them exhibited expression patterns restricted to specific tissues. Moreover, we demonstrated that highly similar selenoprotein paralogs were expressed within distinct territories. Therefore, tissue- and development-specific expression patterns provided new information for selenoproteins of unknown function.}, note = {1567-133x Journal Article}, keywords = {Animals Gene Expression Regulation, Developmental In Situ Hybridization Molecular Sequence Data Proteins/*genetics/metabolism RNA Probes RNA, LESCURE, Messenger/metabolism Support, Non-U.S. Gov't Support, P.H.S. Tissue Distribution Zebrafish/*embryology, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Towards understanding human mitochondrial leucine aminoacylation identity}, author = {B Sohm and M Frugier and H Brule and K Olszak and A Przykorska and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12729737}, isbn = {12729737}, year = {2003}, date = {2003-01-01}, journal = {J Mol Biol}, volume = {328}, number = {5}, pages = {995-1010}, abstract = {Specific recognition of tRNAs by aminoacyl-tRNA synthetases is governed by sets of aminoacylation identity elements, well defined for numerous prokaryotic systems and eukaryotic cytosolic systems. Only restricted information is available for aminoacylation of human mitochondrial tRNAs, despite their particularities linked to the non-classical structures of the tRNAs and their involvement in a growing number of human neurodegenerative disorders linked to mutations in the corresponding tRNA genes. A major difficulty to be overcome is the preparation of active in vitro transcripts enabling a rational mutagenic analysis, as is currently performed for classical tRNAs. Here, structural and aminoacylation properties of in vitro transcribed tRNA(Leu(UUR)) are presented. Solution probing using a combination of enzymatic and chemical tools revealed only partial folding into an L-shaped structure, with an acceptor branch but with a floppy anticodon branch. Optimization of aminoacylation conditions allowed charging of up to 75% of molecules, showing that, despite its partially relaxed structure, in vitro transcribed tRNA(Leu(UUR)) is able to adapt to the synthetase. In addition, mutational analysis demonstrates that the discriminator base as well as residue A14 are important leucine identity elements. Thus, human mitochondrial leucylation is dependent on rules similar to those that apply in Escherichia coli. The impact of a subset of pathology-related mutations on aminoacylation and on tRNA structure, has been explored. These variants do not show significant structural rearrangements and either do not affect aminoacylation (mutations T3250C, T3271C, C3303T) or lead to marked effects. Interestingly, two variants with a mutation at the same position (A3243G and A3243T) lead to markedly different losses in aminoacylation efficiencies (tenfold and 300-fold, respectively).}, note = {0022-2836 Journal Article}, keywords = {Base Sequence Human In Vitro Leucine/*metabolism Leucine-tRNA Ligase/*metabolism Mitochondria/*metabolism Mitochondrial Diseases/genetics/metabolism Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, ERIANI, FLORENTZ, FRUGIER, Leu/chemistry/genetics/*metabolism Recombinant Proteins/genetics/metabolism Solutions Substrate Specificity Support, Non-U.S. Gov't Variation (Genetics), Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ribosomal protein S15 represses its own translation via adaptation of an rRNA-like fold within its mRNA}, author = {A Serganov and A Polonskaia and B Ehresmann and C Ehresmann and D J Patel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12682022}, isbn = {12682022}, year = {2003}, date = {2003-01-01}, journal = {EMBO J}, volume = {22}, number = {8}, pages = {1898-1908}, abstract = {The 16S rRNA-binding ribosomal protein S15 is a key component in the assembly of the small ribosomal subunit in bacteria. We have shown that S15 from the extreme thermophile Thermus thermophilus represses the translation of its own mRNA in vitro, by interacting with the leader segment of its mRNA. The S15 mRNA-binding site was characterized by footprinting experiments, deletion analysis and site-directed mutagenesis. S15 binding triggers a conformational rearrangement of its mRNA into a fold that mimics the conserved three-way junction of the S15 rRNA-binding site. This conformational change masks the ribosome entry site, as demonstrated by direct competition between the ribosomal subunit and S15 for mRNA binding. A comparison of the T.thermophilus and Escherichia coli regulation systems reveals that the two regulatory mRNA targets do not share any similarity and that the mechanisms of translational inhibition are different. Our results highlight an astonishing plasticity of mRNA in its ability to adapt to evolutionary constraints, that contrasts with the extreme conservation of the rRNA-binding site.}, note = {0261-4189 Journal Article}, keywords = {Bacterial Proteins/genetics/metabolism Binding Sites *Nucleic Acid Conformation Protein Binding Protein Footprinting RNA, Genetic, Messenger/chemistry/*metabolism Repressor Proteins/genetics/*metabolism Ribosomal Proteins/genetics/*metabolism Ribosomes/metabolism Support, P.H.S. Thermus thermophilus/genetics/metabolism *Translation, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The RNA binding protein FMRP: new connections and missing links}, author = {C Schaeffer and M Beaulande and C Ehresmann and B Ehresmann and H Moine}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12867085}, isbn = {12867085}, year = {2003}, date = {2003-01-01}, journal = {Biol Cell}, volume = {95}, number = {3-4}, pages = {221-228}, abstract = {The loss of the fragile X mental retardation protein (FMRP) is responsible for the most common cause of inherited mental retardation called the fragile X syndrome. FMRP is suspected to participate in the synaptic plasticity of neurons by acting on posttranscriptional control of gene expression. FMRP is an RNA binding protein that associates with mRNAs together with other proteins to form large ribonucleoprotein complexes. These complexes are proposed to participate in the transport, localization and translation of target mRNAs. Progress has been made recently in the identification of the mRNAs and the proteins present in these complexes and a possible connection with the micro-RNA dependent regulatory pathway has been established.}, note = {0248-4900 Journal Article Review Review, Tutorial}, keywords = {Female Fragile X Syndrome/*genetics/metabolism Human Macromolecular Systems Male MicroRNAs/genetics Nerve Tissue Proteins/*genetics/metabolism RNA, Genetic/genetics, Messenger/genetics/*metabolism RNA-Binding Proteins/*genetics/metabolism Ribonucleoproteins/genetics/metabolism Synaptic Transmission/genetics Translation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sm-like proteins in Eubacteria: the crystal structure of the Hfq protein from Escherichia coli}, author = {C Sauter and J Basquin and D Suck}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12853626}, isbn = {12853626}, year = {2003}, date = {2003-01-01}, journal = {Nucleic Acids Res}, volume = {31}, number = {14}, pages = {4091-4098}, abstract = {The Hfq protein was discovered in Escherichia coli in the early seventies as a host factor for the Qbeta phage RNA replication. During the last decade, it was shown to be involved in many RNA processing events and remote sequence homology indicated a link to spliceosomal Sm proteins. We report the crystal structure of the E.coli Hfq protein showing that its monomer displays a characteristic Sm-fold and forms a homo-hexamer, in agreement with former biochemical data. Overall, the structure of the E.coli Hfq ring is similar to the one recently described for Staphylococcus aureus. This confirms that bacteria contain a hexameric Sm-like protein which is likely to be an ancient and less specialized form characterized by a relaxed RNA binding specificity. In addition, we identified an Hfq ortholog in the archaeon Methanococcus jannaschii which lacks a classical Sm/Lsm gene. Finally, a detailed structural comparison shows that the Sm-fold is remarkably well conserved in bacteria, Archaea and Eukarya, and represents a universal and modular building unit for oligomeric RNA binding proteins.}, note = {1362-4962 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Bacteria/*genetics Crystallography, FRUGIER, Molecular Host Factor 1 Protein/chemistry/*genetics Molecular Sequence Data Protein Conformation Protein Structure, Non-U.S. Gov't, SAUTER, Secondary Ribonucleoproteins, Small Nuclear/chemistry/*genetics Sequence Alignment Sequence Homology, Unité ARN, X-Ray Dimerization Escherichia coli Proteins/chemistry/*genetics Evolution}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast tRNA(Asp) charging accuracy is threatened by the N-terminal extension of aspartyl-tRNA synthetase}, author = {M Ryckelynck and R Giege and M Frugier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12486031}, isbn = {12486031}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {11}, pages = {9683-9690}, abstract = {This study evaluates the role of the N-terminal extension from yeast aspartyl-tRNA synthetase in tRNA aspartylation. The presence of an RNA-binding motif in this extension, conserved in eukaryotic class IIb aminoacyl-tRNA synthetases, provides nonspecific tRNA binding properties to this enzyme. Here, it is assumed that the additional contacts the 70 amino acid-long appendix of aspartyl-tRNA synthetase makes with tRNA could be important in expression of aspartate identity in yeast. Using in vitro transcripts mutated at identity positions, it is demonstrated that the extension grants better aminoacylation efficiency but reduced specificity to the synthetase, increasing considerably the risk of noncognate tRNA mischarging. Yeast tRNA(Glu(UUC)) and tRNA(Asn(GUU)) were identified as the most easily mischarged tRNA species. Both have a G at the discriminator position, and their anticodon differs only by one change from the GUC aspartate anticodon.}, note = {0021-9258 Journal Article}, keywords = {Amino Acid Motifs Aspartate-tRNA Ligase/*metabolism Aspartic Acid/chemistry Base Sequence Codon Escherichia coli/metabolism Kinetics Molecular Sequence Data Nucleic Acid Conformation Nucleic Acids/chemistry Protein Structure, Asp/*chemistry Support, FRUGIER, Messenger/metabolism RNA, Non-U.S. Gov't Yeasts/metabolism, Secondary Protein Structure, Tertiary RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {When contemporary aminoacyl-tRNA synthetases invent their cognate amino acid metabolism}, author = {H Roy and H D Becker and J Reinbolt and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12874385}, isbn = {12874385}, year = {2003}, date = {2003-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {100}, number = {17}, pages = {9837-9842}, abstract = {Faithful protein synthesis relies on a family of essential enzymes called aminoacyl-tRNA synthetases, assembled in a piecewise fashion. Analysis of the completed archaeal genomes reveals that all archaea that possess asparaginyl-tRNA synthetase (AsnRS) also display a second ORF encoding an AsnRS truncated from its anticodon binding-domain (AsnRS2). We show herein that Pyrococcus abyssi AsnRS2, in contrast to AsnRS, does not sustain asparaginyl-tRNAAsn synthesis but is instead capable of converting aspartic acid into asparagine. Functional analysis and complementation of an Escherichia coli asparagine auxotrophic strain show that AsnRS2 constitutes the archaeal homologue of the bacterial ammonia-dependent asparagine synthetase A (AS-A), therefore named archaeal asparagine synthetase A (AS-AR). Primary sequence- and 3D-based phylogeny shows that an archaeal AspRS ancestor originated AS-AR, which was subsequently transferred into bacteria by lateral gene transfer in which it underwent structural changes producing AS-A. This study provides evidence that a contemporary aminoacyl-tRNA synthetase can be recruited to sustain amino acid metabolism.}, note = {0027-8424 Journal Article}, keywords = {Amino Acid Substrate Specificity Support, Amino Acid Sequence Amino Acids/*metabolism Amino Acyl-tRNA Ligases/chemistry/genetics/*metabolism Catalytic Domain/genetics Cloning, Archaeal Genes, Bacterial Models, Molecular Escherichia coli/genetics/metabolism Genes, Molecular Molecular Sequence Data Phylogeny Protein Conformation Pyrococcus/enzymology/genetics Sequence Homology, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Bacterial translational control at atomic resolution}, author = {P Romby and M Springer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12615010}, isbn = {12615010}, year = {2003}, date = {2003-01-01}, journal = {Trends Genet}, volume = {19}, number = {3}, pages = {155-161}, abstract = {Translational regulation allows rapid adaptation of protein synthesis to environmental conditions. In prokaryotes, the synthesis of many RNA-binding proteins is regulated by a translational feedback mechanism involving a competition between their natural substrate and their binding site on mRNA, which are often thought to resemble each other. This article describes the case of threonyl-tRNA synthetase, which represses the translation of its own mRNA. Recent data provide the first opportunity to describe at the atomic level both the extent and the limit of mimicry between the way this enzyme recognizes tRNA(Thr) and its regulatory site in mRNA. The data also give some clues about how the binding of the synthetase to its mRNA inhibits translation.}, note = {0168-9525 Journal Article Review Review, Tutorial}, keywords = {Bacterial *Gene Expression Regulation, Bacterial Models, Base Sequence Conserved Sequence Escherichia coli/*enzymology/*genetics *Gene Expression Regulation, Biological Models, Enzymologic Genes, Genetic, Messenger/genetics/*metabolism RNA, Molecular Molecular Mimicry Nucleic Acid Conformation Operator Regions (Genetics) RNA, Non-U.S. Gov't Threonine-tRNA Ligase/chemistry/*genetics/metabolism *Translation, ROMBY, Thr/*metabolism RNA-Binding Proteins/metabolism Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Growth sectors and crystal quality.}, author = {M C Robert and B Capelle and B Lorber}, url = {http://www.sciencedirect.com/science/article/pii/S0076687903680092}, doi = {10.1016/S0076-6879(03)68009-2}, year = {2003}, date = {2003-01-01}, journal = {Methods Enzymol}, volume = {368}, pages = {154-169}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure-function relationships of the initiation complex of HIV-1 reverse transcription: the case of mutant viruses using tRNA(His) as primer}, author = {M Rigourd and V Goldschmidt and F Brule and C D Morrow and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14500840}, isbn = {14500840}, year = {2003}, date = {2003-01-01}, journal = {Nucleic Acids Res}, volume = {31}, number = {19}, pages = {5764-5775}, abstract = {Reverse transcription of HIV-1 RNA is initiated from the 3' end of a tRNA3Lys molecule annealed to the primer binding site (PBS). An additional interaction between the anticodon loop of tRNA3Lys and a viral A-rich loop is required for efficient initiation of reverse transcription of the HIV-1 MAL isolate. In the HIV-1 HXB2 isolate, simultaneous mutations of the PBS and the A-rich loop (mutant His-AC), but not of the PBS alone (mutant His) allows the virus to stably utilize tRNA(His) as primer. However, mutant His-AC selects additional mutations during cell culture, generating successively His-AC-GAC and His-AC-AT-GAC. Here, we wanted to establish direct relationships between the evolution of these mutants in cell culture, their efficiency in initiating reverse transcription and the structure of the primer/template complexes in vitro. The initiation of reverse transcription of His and His-AC RNAs was dramatically reduced. However, His-AC-GAC RNA, which incorporated three adaptative point mutations, was reverse transcribed more efficiently than the wild type RNA. Incorporation of two additional mutations decreased the efficiency of the initiation of reverse transcription, which remained at the wild type level. Structural probing showed that even though both His-AC and His-AC-GAC RNAs can potentially interact with the anticodon loop of tRNA(His), only the latter template formed a stable interaction. Thus, our results showed that the selection of adaptative mutations by HIV-1 mutants utilizing tRNA(His) as primer was initially dictated by the efficiency of the initiation of reverse transcription, which relied on the existence of a stable interaction between the mutated A-rich loop and the anticodon loop of tRNA(His).}, note = {1362-4962 Journal Article}, keywords = {Base Sequence Comparative Study DNA, Genetic, Genetic *Transcription Initiation Site *Transcription, His/*metabolism RNA, MARQUET, Non-U.S. Gov't Templates, Post-Transcriptional RNA, Transfer, Unité ARN, Viral HIV-1/*genetics/metabolism HIV-1 Reverse Transcriptase/metabolism Kinetics Macromolecular Systems Molecular Sequence Data Mutation RNA Probes RNA Processing, Viral/*biosynthesis/genetics Sequence Alignment Structure-Activity Relationship Support, Viral/biosynthesis *Gene Expression Regulation}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effects of tRNA 3 Lys aminoacylation on the initiation of HIV-1 reverse transcription}, author = {M Rigourd and G Bec and P Benas and S F Le Grice and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12763311}, isbn = {12763311}, year = {2003}, date = {2003-01-01}, journal = {Biochimie}, volume = {85}, number = {5}, pages = {521-525}, abstract = {HIV-1 utilizes cellular tRNA(3)(Lys) to prime the initiation of reverse transcription. The selective incorporation of cytoplasmic tRNA(3)(Lys) into HIV-1 particles was recently shown to involve the lysyl-tRNA synthetase, and hence, the encapsidated tRNA(3)(Lys) is likely to be aminoacylated. Here, we tested the effect of aminoacylation on the initiation of reverse transcription. We show that HIV-1 reverse transcriptase is unable to extend lysyl-tRNA(3)(Lys). In addition, the viral polymerase does not significantly enhance the rate of tRNA deacylation, in contrast with previous studies on avian retroviruses. Thus, aminoacylation of the primer tRNA might prevent the initiation of HIV-1 reverse transcription from taking place before viral budding and maturation.}, note = {0300-9084 Journal Article}, keywords = {Acetyltransferases/metabolism *Acylation Animals Cattle HIV-1/*physiology HIV-1 Reverse Transcriptase/pharmacology RNA/genetics RNA, Genetic/drug effects/*physiology Virus Assembly, Lys/*chemistry/drug effects Support, MARQUET, Non-U.S. Gov't Transcription, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Assembly of core helices and rapid tertiary folding of a small bacterial group I ribozyme}, author = {P Rangan and B Masquida and E Westhof and S A Woodson}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12574513}, isbn = {12574513}, year = {2003}, date = {2003-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {100}, number = {4}, pages = {1574-1579}, abstract = {Compact but non-native intermediates have been implicated in the hierarchical folding of several large RNAs, but there is little information on their structure. In this article, ribonuclease and hydroxyl radical cleavage protection assays showed that base pairing of core helices stabilize a compact state of a small group I ribozyme from Azoarcus pre-tRNA(ile). Base pairing of the ribozyme core requires 10-fold less Mg(2+) than stable tertiary interactions, indicating that assembly of helices in the catalytic core represents a distinct phase that precedes the formation of native tertiary structure. Tertiary folding occurs in <100 ms at 37 degrees C. Such rapid folding is unprecedented among group I ribozymes and illustrates the association between structural complexity and folding time. A 3D model of the Azoarcus ribozyme was constructed by identifying homologous sequence motifs in rRNA. The model reveals distinct structural features, such as a large interface between the P4-P6 and P3-P9 domains, that may explain the unusual stability of the Azoarcus ribozyme and the cooperativity of folding.}, note = {0027-8424 Journal Article}, keywords = {Azoarcus/*enzymology Base Sequence Introns Magnesium/chemistry Models, Catalytic/*chemistry/genetics Support, Molecular Molecular Sequence Data Nucleic Acid Conformation *Protein Folding Protein Structure, Non-U.S. Gov't Support, P.H.S., Tertiary RNA, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The molecular basis for A-site mutations conferring aminoglycoside resistance: relationship between ribosomal susceptibility and X-ray crystal structures}, author = {P Pfister and S Hobbie and Q Vicens and E C Bottger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14523926}, isbn = {14523926}, year = {2003}, date = {2003-01-01}, journal = {Chembiochem}, volume = {4}, number = {10}, pages = {1078-1088}, abstract = {Aminoglycoside antibiotics target the 16S ribosomal RNA (rRNA) bacterial A site and induce misreading of the genetic code. Point mutations of the ribosomal A site may confer resistance to aminoglycoside antibiotics. The influence of bacterial mutations (introduced by site-directed mutagenesis) on ribosomal drug susceptibility was investigated in vivo by determination of minimal inhibitory concentrations. To determine the origin of the various resistance phenotypes at a molecular level, the in vivo results were compared with the previously published crystal structures of paromomycin, tobramycin, and geneticin bound to oligonucleotides containing the minimal A site. Two regions appear crucial for binding in the A site: the single adenine residue at position 1408 and the non-Watson-Crick U1406.U1495 pair. The effects of mutations at those positions are modulated by the nature of the substituent at position 6' (either hydroxy or ammonium group) on ring I, by the number of positive charges on the antibiotic, and by the linkage between rings I and III (either 4,5 or 4,6). In particular, the analysis demonstrates: 1) that the C1409-G1491 to A1409-U1491 polymorphism (observed in 15 % of bacteria) is not associated with resistance, which indicates that it does not affect the stacking of ring I on residue 1491, 2) that the high-level resistance to 6'-NH3+ aminoglycosides exhibited by the A1408G mutation most probably results from the inability of ring I forming a pseudo base pair with G1408, which prevents its insertion inside the A site helix, and 3) that mutations of the uracil residues forming the U1406.U1495 pair either to cytosine or to adenine residues mostly confer low to moderate levels of drug resistance, whereas the U1406C/U1495A double mutation confers high-level resistance (except for neomycin), which suggests that aminoglycoside binding to the wild-type A site and its functional consequences strongly depend on a particular geometry of the U1406.U1495 pair. The relationships between the resistance phenotypes observed in vivo and the interactions described at the molecular level define the biological importance of the different structural interactions observed by X-ray crystallography studies.}, note = {1439-4227 Journal Article}, keywords = {Aminoglycosides/*pharmacology Anti-Bacterial Agents/*pharmacology Binding Sites Comparative Study Crystallography, Bacterial/genetics Ribosomes/*chemistry/drug effects/metabolism Species Specificity Structure-Activity Relationship Support, Molecular Mutagenesis, Non-U.S. Gov't, Site-Directed Nucleic Acid Conformation Oligonucleotides/chemistry/metabolism Plasmids Point Mutation/drug effects RNA, Unité ARN, WESTHOF, X-Ray Drug Design Drug Resistance Escherichia coli/genetics/metabolism Hemagglutinins Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selenoprotein N: an endoplasmic reticulum glycoprotein with an early developmental expression pattern}, author = {N Petit and A Lescure and M Rederstorff and A Krol and B Moghadaszadeh and U M Wewer and P Guicheney}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12700173}, isbn = {12700173}, year = {2003}, date = {2003-01-01}, journal = {Hum Mol Genet}, volume = {12}, number = {9}, pages = {1045-1053}, abstract = {Rigid spine muscular dystrophy and the classical form of multiminicore disease are caused by mutations in SEPN1 gene, leading to a new clinical entity referred to as SEPN1-related myopathy. SEPN1 codes for selenoprotein N, a new member of the selenoprotein family, the function of which is still unknown. In a previous study, two isoforms were deduced from SEPN1 transcript analyses. Using polyclonal antibodies directed against SEPN1 and cDNA constructs encoding for the two isoforms, we show that the main SEPN1 gene product corresponds to a 70 kDa protein, containing a single selenocysteine residue. Subcellular fractionation experiments and endoglycosidase H sensitivity indicate that SEPN1 is a glycoprotein-localized within the endoplasmic reticulum. Immunofluorescence analyses confirm this subcellular localization and green fluorescent protein fusion experiments demonstrate the presence of an endoplasmic reticulum-addressing and -retention signal within the N-terminus. SEPN1 is present at a high level in several human fetal tissues and at a lower level in adult ones, including skeletal muscle. Its high expression in cultured myoblasts is also down-regulated in differentiating myotubes, suggesting a role for SEPN1 in early development and in cell proliferation or regeneration.}, note = {0964-6906 Journal Article}, keywords = {Cell Division/physiology Endoplasmic Reticulum/*metabolism Fetus/metabolism Fibroblasts/metabolism Human Muscle Proteins/*metabolism Protein Sorting Signals Support, LESCURE, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Global DNA methylation in relation to hMLH1 and hMSH2 protein immunoreactivity in sporadic human endometrial carcinomas}, author = {R Miturski and K Postawski and A Semczuk and M Bogusiewicz and W Baranowski and J A Jakowicki and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12684691}, isbn = {12684691}, year = {2003}, date = {2003-01-01}, journal = {Int J Mol Med}, volume = {11}, number = {5}, pages = {569-574}, abstract = {Overall DNA methylation status was studied in a group of 28 sporadic human endometrial carcinomas (ECs) using the [32P]-postlabeling technique. Moreover, expression of the DNA mismatch repair proteins (hMLH1 and hMSH2) was investigated in ECs using immunohistochemistry. Mean 5-methyldeoxycytosine (m5dC) content in the studied group was 3.48+/-0.37% (range, 2.89-4.12%). The mean m5dC scores were significantly different between early (3.35+/-0.33%) and advanced (3.66+/-0.36%) endometrial neoplasms (chi2-test; p=0.03). There was a markedly increased overall DNA methylation with the degree of histological differentiation and with the infiltration of the myometrium (p<0.05). Loss of hMLH1 and hMSH2 expression was reported in 7 (25%) and 5 (18%) tumors, respectively, but the immunoreactivity did not correlate with the known clinicopathological variables of cancer. In addition, no obvious correlation was found between global m5dC content and the lack of hMLH1 and hMSH2 protein expression in human uterine tumors (p=0.97 and p=0.19 for hMLH1 and hMSH2, respectively; Spearman's rank correlation test). Our results clearly show that alterations in global DNA methylation may influence tumor progression, but they are not directly associated with the inactivation of the mismatch-repair machinery in sporadic human ECs.}, note = {1107-3756 Journal Article}, keywords = {Base Pair Mismatch Carcinoma/genetics/*metabolism/pathology *DNA Methylation DNA Repair Endometrial Neoplasms/genetics/*metabolism/pathology Female Human Immunohistochemistry Neoplasm Proteins/*metabolism Proto-Oncogene Proteins/*metabolism Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pathology-related substitutions in human mitochondrial tRNA(Ile) reduce precursor 3' end processing efficiency in vitro}, author = {L Levinger and R Giege and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12655007}, isbn = {12655007}, year = {2003}, date = {2003-01-01}, journal = {Nucleic Acids Res}, volume = {31}, number = {7}, pages = {1904-1912}, abstract = {The human mitochondrial genome encodes 22 tRNAs interspersed among the two rRNAs and 11 mRNAs, often without spacers, suggesting that tRNAs must be efficiently excised. Numerous maternally transmitted diseases and syndromes arise from mutations in mitochondrial tRNAs, likely due to defect(s) in tRNA metabolism. We have systematically explored the effect of pathogenic mutations on tRNA(Ile) precursor 3' end maturation in vitro by 3'-tRNase. Strikingly, four pathogenic tRNA(Ile) mutations reduce 3'-tRNase processing efficiency (V(max) / K(M)) to approximately 10-fold below that of wild-type, principally due to lower V(max). The structural impact of mutations was sought by secondary structure probing and wild-type tRNA(Ile) precursor was found to fold into a canonical cloverleaf. Among the mutant tRNA(Ile) precursors with the greatest 3' end processing deficiencies, only G4309A displays a secondary structure substantially different from wild-type, with changes in the T domain proximal to the substitution. Reduced efficiency of tRNA(Ile) precursor 3' end processing, in one case associated with structural perturbations, could thus contribute to human mitochondrial diseases caused by mutant tRNAs.}, note = {1362-4962 Journal Article}, keywords = {Base Sequence DNA, FLORENTZ, Ile/*genetics/metabolism Support, Mitochondrial/*genetics Endoribonucleases/metabolism Hela Cells Human Kinetics Molecular Sequence Data Mutation RNA Precursors/genetics/metabolism *RNA Processing, Non-U.S. Gov't Support, P.H.S., Post-Transcriptional RNA, Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Analysis of RNA motifs}, author = {N B Leontis and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12831880}, isbn = {12831880}, year = {2003}, date = {2003-01-01}, journal = {Curr Opin Struct Biol}, volume = {13}, number = {3}, pages = {300-308}, abstract = {RNA motifs are directed and ordered stacked arrays of non-Watson-Crick base pairs forming distinctive foldings of the phosphodiester backbones of the interacting RNA strands. They correspond to the 'loops' - hairpin, internal and junction - that intersperse the Watson-Crick two-dimensional helices as seen in two-dimensional representations of RNA structure. RNA motifs mediate the specific interactions that induce the compact folding of complex RNAs. RNA motifs also constitute specific protein or ligand binding sites. A given motif is characterized by all the sequences that fold into essentially identical three-dimensional structures with the same ordered array of isosteric non-Watson-Crick base pairs. It is therefore crucial, when analyzing a three-dimensional RNA structure in order to identify and compare motifs, to first classify its non-Watson-Crick base pairs geometrically.}, note = {0959-440x Journal Article Review Review, Tutorial}, keywords = {*Base Pairing Binding Sites Magnetic Resonance Imaging *Nucleic Acid Conformation RNA/*chemistry RNA, Chloroplast/chemistry Support, P.H.S., U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence elements outside the hammerhead ribozyme catalytic core enable intracellular activity}, author = {A Khvorova and A Lescoute and E Westhof and S D Jayasena}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12881719}, isbn = {12881719}, year = {2003}, date = {2003-01-01}, journal = {Nat Struct Biol}, volume = {10}, number = {9}, pages = {708-712}, abstract = {The hammerhead ribozyme (HHRz) is a small, naturally occurring ribozyme that site-specifically cleaves RNA and has long been considered a potentially useful tool for gene silencing. The minimal conserved HHRz motif derived from natural sequences consists of three helices that intersect at a highly conserved catalytic core of 11 nucleotides. The presence of this motif is sufficient to support cleavage at high Mg2+ concentrations, but not at the low Mg2+ concentrations characteristic of intracellular environments. Here we demonstrate that natural HHRzs require the presence of additional nonconserved sequence elements outside of the conserved catalytic core to enable intracellular activity. These elements may stabilize the HHRz in a catalytically active conformation via tertiary interactions. HHRzs stabilized by these interactions cleave efficiently at physiological Mg2+ concentrations and are functional in vivo. The proposed role of these tertiary interacting motifs is supported by mutational, functional, structural and molecular modeling analysis of natural HHRzs.}, note = {1072-8368 Journal Article}, keywords = {Amino Acid Motifs Base Sequence Catalysis Dose-Response Relationship, Catalytic/*chemistry Sequence Homology, Drug Genes, Nucleic Acid Time Factors, Reporter Genetic Vectors Kinetics Magnesium/chemistry Molecular Sequence Data Mutation Nucleic Acid Conformation Plasmids/metabolism Protein Binding Protein Conformation RNA/metabolism RNA, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallogenesis studies of proteins in agarose gel-combined effect of high hydrostatic pressure and pH}, author = {A Kadri and G Jenner and M Damak and B Lorber and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S0022024803014714}, doi = {10.1016/S0022-0248(03)01471-4}, year = {2003}, date = {2003-01-01}, journal = {J Crystal Growth}, volume = {257}, number = {3-4}, pages = {390-402}, abstract = {The combinedeffect of both pressure and pH on the crystallization in agarosegel of thaumatin (0.1–230 MPa and pH 5.7–8.0) and the lysozymes from turkey (0.1–100 MPa and pH 4.0–6.0) and hen egg-white (0.1–75 MPa and pH 4.0–6.0) was investigated. For each condition, crystal morphology was examined and four crystal growth parameters determined (crystal number as a measure of nucleation, solubility, supersaturation and pressure-induced volume changes of crystallization). Comparison of data reveals a different crystallization behavior of thaumatin and the two lysozymes. First, thaumatin is more pressure tolerant in agarosegel than the lysozymes, with crystals growing up to 230 MPa for the former and only up to 75/100 MPa for the latter. Second, while an increase in pressure and pH increases nucleation of thaumatin crystals and decreases their solubility, one observes an inverse effect with the two lysozymes. Here, an increase in pressure but a decrease in pH increases nucleation and solubility. Third, pH influences the pressure-induced volume changes of crystallization, the strongest pH-dependence being found for turkey lysozyme. However, the volume changes are negative for thaumatin and positive for both lysozymes. These volume changes are correlated with solubility changes and are explained by pH- and pressure-induced alterations of the conformation of the three proteins, including their solvation shells.}, keywords = {GIEGE Biocrystallization pH Pressure Agarose gel Lysozyme Thaumatin, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pressure versus pH phase diagrams of two lysozymes crystallized in agarose gel}, author = {A Kadri and M Damak and B Lorber and R Giege and G Jenner}, url = {http://www.tandfonline.com/doi/abs/10.1080/08957950310001621030}, doi = {10.1080/08957950310001621030}, year = {2003}, date = {2003-01-01}, journal = {High Pressure Research}, volume = {23}, number = {4}, pages = {485-491}, abstract = {The effect of hydrostatic pressure and pH on the crystallization of tetragonal hen lysozyme crystals (HeL, M r 14,300) and hexagonal turkey lysozyme crystals (TeL, M r 14,200) in agarose gel was studied. Samples (adjusted to a pH in the range 4–6) were pressurized at 0.1–100 MPa for nine days at 293 K. The morphology and number of crystals as well as protein solubility were analyzed after depressurization. For both proteins and whatever the pH, a higher pressure resulted in greater numbers of crystals and greater solubility values. At any pressure, the solubility and number of crystals were lower at the highest pH. Thus the physical chemical parameters and the outcome of the crystallization process are more affected by pressure at pH 4.0 than at pH 6.0. In the case of HeL, either high pressure or high pH induces a transition from the initial tetragonal form to an orthorhombic one. The observed effects are related to minor differences in the macromolecular structures.}, keywords = {GIEGE Crystallization Lysozyme Protein Pressure Ph Phase diagram, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Investigating the nucleation of protein crystals with hydrostatic pressure}, author = {A Kadri and M Damak and G Jenner and B Lorber and R Giege}, url = {http://adsabs.harvard.edu/abs/2003JPCM.15.8253K}, doi = {10.1088/0953-8984/15/49/004}, year = {2003}, date = {2003-01-01}, journal = {J Phys Condens Matter}, volume = {15}, number = {49}, pages = {8253-8262}, abstract = {Hydrostatic pressure in the 0.1-75 MPa range has been used as a non-invasive tool to study the crystallization process of the tetragonal crystal form of the protein thaumatin (Mr 22 200). Crystals were prepared within agarose gel and at temperatures in the range from 283 to 303 K. The solubility, i.e. the concentration of soluble macromolecules remaining in equilibrium with the crystals, decreases when the pressure increases and when the temperature decreases. High pressure was used to probe the nucleation behaviour of thaumatin. The pressure dependence of the nucleation rate leads to an activation volume of -46.5 cm3 mol-1. It is shown that an increase in pressure decreases the enthalpy, the entropy and the free energy of crystallization of thaumatin. The data are discussed in the light of the results of crystallographic analyses and of the structure of the protein.}, keywords = {GIEGE Hydrostatic pressure Crystals Nucleation Protein, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Effect of pressure on the crystallization of proteins.}, author = {A Kadri and C Charron and M C Robert and B Capelle and G Jenner and R Giege and B Lorber}, editor = {R Winter}, url = {http://www.springer.com/chemistry/biotechnology/book/978-3-540-00977-1}, year = {2003}, date = {2003-01-01}, booktitle = {Advances in High pressure Bioscience and Biotechnology II. Proceedings of the 2nd International Conference on High Pressure Bioscience and Biotechnology, Dortmund, September 16 - 19, 2002}, pages = {175-178}, publisher = {Springer}, address = {Heidelberg}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {The central PPT of the yeast retrotransposon Ty1 is not essential for transposition}, author = {T Heyman and M Wilhelm and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12888340}, isbn = {12888340}, year = {2003}, date = {2003-01-01}, journal = {J Mol Biol}, volume = {331}, number = {2}, pages = {315-320}, abstract = {The yeast retrotransposon Ty1 has structural and functional similarities to retroviruses. We report here that, as in retroviruses, the plus-strand DNA of Ty1 is synthesized as two segments. A central DNA flap is formed during reverse transcription consecutive to elongation (with strand displacement) of the upstream segment beyond the central polypurine tract (cPPT) until the replication machinery is stopped at the central termination sequence. Comparison of wild-type and cPPT-mutant Ty1 elements shows that the mutant element lacking the central DNA flap is only twofold defective in transposition.}, note = {0022-2836 Journal Article}, keywords = {Base Sequence DNA/*biosynthesis Models, Genetic Molecular Sequence Data Mutation Purines/*chemistry Retroelements/*genetics Saccharomyces cerevisiae/genetics Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Does the HIV-1 primer activation signal interact with tRNA3(Lys) during the initiation of reverse transcription?}, author = {V Goldschmidt and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12560480}, isbn = {12560480}, year = {2003}, date = {2003-01-01}, journal = {Nucleic Acids Res}, volume = {31}, number = {3}, pages = {850-859}, abstract = {Reverse transcription of HIV-1 RNA is primed by a tRNA3(Lys) molecule bound at the primer binding site (PBS). Complex intermolecular interactions were proposed between tRNA3(Lys) and the RNA of the HIV-1 Mal isolate. Recently, an alternative interaction was proposed between the TPsiC stem of tRNA3(Lys) and a primer activation signal (PAS) of the Lai and Hxb2 RNAs, suggesting major structural variations in the reverse transcription complex of different HIV-1 strains. Here, we analyzed mutants of the Hxb2 RNA that prevent the interaction between the PAS and tRNA3(Lys) or/and a complementary sequence in the viral RNA. We compared the kinetics of reverse transcription of the wild type and mutant Hxb2 RNAs, using either tRNA3(Lys) or an 18mer oligoribonucleotide complementary to the PBS, which cannot interact with the PAS, as primers. We also used chemical probing to test the structure of the mutant and wild type RNAs, as well as the complex formed between the later RNA and tRNA3(Lys). These experiments, together with the analysis of long term replication data of mutant viruses obtained by C. Morrow and coworkers (Birmingham, USA) that use alternate tRNAs as primers, strongly suggest that the interaction between the Hxb2 PAS and tRNA3(Lys) does not exist. Instead, the effects of the vRNA mutations on reverse transcription seem to be linked to incorrect folding of the mutant RNAs.}, note = {1362-4962 Journal Article}, keywords = {Amino Acyl/chemistry/*metabolism RNA, Base Sequence Binding Sites DNA Primers DNA, Genetic, MARQUET, Non-U.S. Gov't *Transcription, Transfer, Unité ARN, Viral HIV-1/*genetics HIV-1 Reverse Transcriptase/*metabolism Kinetics Molecular Sequence Data Mutation Nucleic Acid Conformation Oligoribonucleotides RNA, Viral/biosynthesis *Gene Expression Regulation, Viral/chemistry/genetics/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {In memorium Jean Witz: the contrbution of Jean Witz to the early study of transfer ribonucleic acids}, author = {R Giege and B Lorber}, url = {http://www.sciencedirect.com/science/article/pii/S0300908403002074}, year = {2003}, date = {2003-01-01}, journal = {Biochimie}, volume = {85}, number = {12}, pages = {1235}, keywords = {GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Transfer RNA Structure and Identity}, author = {R Giege and M Frugier}, editor = {J Lapointe and L Brakier-Gringas}, url = {http://www.ncbi.nlm.nih.gov/books/NBK6236}, year = {2003}, date = {2003-01-01}, booktitle = {Translation mechanisms}, pages = {1-24}, publisher = {Landes Bioscience}, abstract = {The structure of tRNA and its relationship with the biological necessity of specific tRNA aminoacylation reactions, in other words with identity, is reviewed. New structural data show the typical L-shaped tRNA architecture in great detail and highlight how adequate rigidity and plasticity of the molecule is essential for interaction with its biological partners, in particular with aminoacyl-tRNA synthetases. Identity is ensured by a small number of nucleosides predominantly located at the two distal extremities of the tRNA molecule. In several crystallographic complexes, these residues have been shown in contact with amino acids from the synthetases. In most cases, the interaction is accompanied by a conformational change of the tRNA. Assuming that the structural framework of tRNA displays identity elements to synthetases implies that altered and/or simplified RNA architectures can fulfill this role provided they contain correctly located identity elements. This paradigm holds true in nature where atypical and tRNA-like domains have been selected by evolution. Rationale-based engineering or selection by artificial evolution of novel RNA molecules recognized and aminoacylated by synthetases also verified it.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Genetic code expansion}, author = {R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12768199}, isbn = {12768199}, year = {2003}, date = {2003-01-01}, journal = {Nat Struct Biol}, volume = {10}, number = {6}, pages = {414-416}, note = {1072-8368 Comment News}, keywords = {Amino Acyl-tRNA Ligases/*chemistry/genetics/*metabolism Codon *Genetic Code Genetic Engineering/methods Human RNA, Transfer, Transfer/*chemistry/*metabolism RNA, Tyr/chemistry/metabolism Structure-Activity Relationship Tyrosine/chemistry/*genetics/metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A yeast knockout strain to discriminate between active and inactive tRNA molecules}, author = {R Geslain and F Martin and A Camasses and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12907713}, isbn = {12907713}, year = {2003}, date = {2003-01-01}, journal = {Nucleic Acids Res}, volume = {31}, number = {16}, pages = {4729-4737}, abstract = {Here we report the construction of a yeast genetic screen designed to identify essential residues in tRNA(Arg). The system consists of a tRNA(Arg) knockout strain and a set of vectors designed to rescue and select for variants of tRNA(Arg). By plasmid shuffling we selected inactive tRNA mutants that were further analyzed by northern blotting. The mutational analysis focused on the tRNA D and anticodon loops that contact the aminoacyl-tRNA synthetase. The anticodon triplet was excluded from the analysis because of its role in decoding the Arg codons. Most of the inactivating mutations are residues involved in tertiary interactions. These mutations had dramatic effects on tRNA(Arg) abundance. Other inactivating mutations were located in the anticodon loop, where they did not affect transcription and aminoacylation but probably altered interaction with the translation machinery. No lethal effects were observed when residues 16, 20 and 38 were individually mutated, despite the fact that they are involved in sequence-specific interactions with the aminoacyl-tRNA synthetase. However, the steady-state levels of the aminoacylated forms of U20A and U20G were decreased by a factor of 3.5-fold in vivo. This suggests that, unlike in the Escherichia coli tRNA(Arg):ArgRS system where residue 20 (A) is a major identity element, in yeast this position is of limited consequence.}, note = {1362-4962 Journal Article}, keywords = {Amino Acyl-tRNA Ligases/metabolism Arginine/genetics/metabolism Base Sequence Blotting, Arg/chemistry/*genetics/metabolism Saccharomyces cerevisiae/*genetics Support, ERIANI, Molecular Hydrogen-Ion Concentration Molecular Sequence Data Mutagenesis, Non-U.S. Gov't, Northern Cloning, Site-Directed Mutation Nucleic Acid Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Limited set of amino acid residues in a class Ia aminoacyl-tRNA synthetase is crucial for tRNA binding}, author = {R Geslain and G Bey and J Cavarelli and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14690419}, isbn = {14690419}, year = {2003}, date = {2003-01-01}, journal = {Biochemistry}, volume = {42}, number = {51}, pages = {15092-15101}, abstract = {The aim of this work was to characterize crucial amino acids for the aminoacylation of tRNA(Arg) by yeast arginyl-tRNA synthetase. Alanine mutagenesis was used to probe all the side chain mediated interactions that occur between tRNA(Arg2)(ICG) and ArgRS. The effects of the substitutions were analyzed in vivo in an ArgRS-knockout strain and in vitro by measuring the aminoacylation efficiencies for two distinct tRNA(Arg) isoacceptors. Nine mutants that generate lethal phenotypes were identified, suggesting that only a limited set of side chain mediated interactions is essential for tRNA recognition. The majority of the lethal mutants was mapped to the anticodon binding domain of ArgRS, a helix bundle that is characteristic for class Ia synthetases. The alanine mutations induce drastic decreases in the tRNA charging rates, which is correlated with a loss in affinity in the catalytic site for ATP. One of those lethal mutations corresponds to an Arg residue that is strictly conserved in all class Ia synthetases. In the known crystallographic structures of complexes of tRNAs and class Ia synthetases, this invariant Arg residue stabilizes the idiosyncratic conformation of the anticodon loop. This paper also highlights the crucial role of the tRNA and enzyme plasticity upon binding. Divalent ions are also shown to contribute to the induced fit process as they may stabilize the local tRNA-enzyme interface. Furthermore, one lethal phenotype can be reverted in the presence of high Mg(2+) concentrations. In contrast with the bacterial system, in yeast arginyl-tRNA synthetase, no lethal mutation has been found in the ArgRS specific domain recognizing the Dhu-loop of the tRNA(Arg). Mutations in this domain have no effects on tRNA(Arg) aminoacylation, thus confirming that Saccharomyces cerevisiae and other fungi belong to a distinct class of ArgRS.}, note = {0006-2960 Journal Article}, keywords = {Acylation Alanine/genetics Amino Acids/*chemistry/genetics Anticodon/chemistry/genetics Arginine-tRNA Ligase/*chemistry/classification/genetics Binding Sites/genetics Genes, Arg/*chemistry Saccharomyces cerevisiae/enzymology/genetics/growth & development Saccharomyces cerevisiae Proteins/*chemistry/*genetics Support, ERIANI, Lethal Mutagenesis, Non-U.S. Gov't, Site-Directed Protein Binding/genetics Protein Structure, Tertiary/genetics RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA recognition by designed peptide fusion creates }, author = {M Frugier and R Giege and P Schimmel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12796515}, isbn = {12796515}, year = {2003}, date = {2003-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {100}, number = {13}, pages = {7471-7475}, abstract = {The genetic code was established through aminoacylations of RNA substrates that emerged as tRNAs. The 20 aminoacyl-tRNA synthetases (one for each amino acid) are ancient proteins, the active-site domain of which catalyzes formation of an aminoacyl adenylate that subsequently reacts with the 3' end of bound tRNA. Binding of tRNA depends on idiosyncratic (to the particular synthetase) domains and motifs that are fused to or inserted into the conserved active-site domain. Here we take the domain for synthesis of alanyl adenylate and fuse it to "artificial" peptide sequences (28 aa) that were shown previously to bind to the acceptor arm of tRNAAla. Certain fusions confer aminoacylation activity on tRNAAla and on hairpin microhelices modeled after its acceptor stem. Aminoacylation was sensitive to the presence of a specific G:U base pair known to be a major determinant of tRNAAla identity. Aminoacylation efficiency and specificity also depended on the specific peptide sequence. The results demonstrate that barriers to RNA-specific aminoacylations are low and can be achieved by relatively simple peptide fusions. They also suggest a paradigm for rationally designed specific aminoacylations based on peptide fusions.}, note = {0027-8424 Journal Article}, keywords = {Amino Acid Motifs Amino Acid Sequence Amino Acyl-tRNA Ligases/*chemistry Base Sequence Binding Sites Catalytic Domain Escherichia coli/genetics/metabolism *Genetic Techniques Kinetics Models, FRUGIER, Genetic Molecular Sequence Data Mutation Nucleic Acid Conformation Peptides/*chemistry Protein Binding Protein Structure, Non-U.S. Gov't Support, P.H.S. Time Factors, Tertiary RNA/chemistry Recombinant Fusion Proteins/metabolism Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast aspartyl-tRNA synthetase binds specifically its own mRNA}, author = {M Frugier and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12888345}, isbn = {12888345}, year = {2003}, date = {2003-01-01}, journal = {J Mol Biol}, volume = {331}, number = {2}, pages = {375-383}, abstract = {Dimeric class II aspartyl-tRNA synthetase (AspRS) from yeast has a modular architecture and includes an N-terminal appendix of 70 amino acid residues that protrudes from the anticodon-binding module. This extension, of predicted helical structure, is not essential for aminoacylation but contains an RNA-binding motif that promotes non-specific interactions with tRNAs. As shown here, this protein extension can also interact with the 5' end of the AspRS mRNA. In vitro, optimal binding occurs on an mRNA domain comprising part of the 87 nucleotide long 5'UTR and the sequence encoding the N-terminal appendix. At the protein side, only the appendix and the anticodon-binding module participate in the interaction between AspRS and the mRNA domain. Binding is specific, since only tRNA(Asp) can dissociate the complex. In vivo, AspRS also binds specifically this mRNA domain and in doing so triggers a reduced translation of a fused GFP mRNA. From that, a mechanism for the regulation of this eukaryotic aminoacyl-tRNA synthetase is proposed. Implications for aspartylation accuracy in yeast are given.}, note = {0022-2836 Journal Article}, keywords = {5' Untranslated Regions Amino Acid Motifs Aspartate-tRNA Ligase/*chemistry/metabolism Binding, Competitive Blotting, Drug Gene Expression Regulation, Enzymologic Genes, FRUGIER, Fungal Kinetics Luminescent Proteins/metabolism Plasmids Protein Binding Protein Structure, Messenger/metabolism RNA, Non-U.S. Gov't, Tertiary RNA/metabolism RNA, Transfer/metabolism Saccharomyces cerevisiae/metabolism Support, Unité ARN, Western Dose-Response Relationship}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ribosome assembly in eukaryotes}, author = {M Fromont-Racine and B Senger and C Saveanu and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12957375}, isbn = {12957375}, year = {2003}, date = {2003-01-01}, journal = {Gene}, volume = {313}, pages = {17-42}, abstract = {Ribosome synthesis is a highly complex and coordinated process that occurs not only in the nucleolus but also in the nucleoplasm and the cytoplasm of eukaryotic cells. Based on the protein composition of several ribosomal subunit precursors recently characterized in yeast, a total of more than 170 factors are predicted to participate in ribosome biogenesis and the list is still growing. So far the majority of ribosomal factors have been implicated in RNA maturation (nucleotide modification and processing). Recent advances gave insight into the process of ribosome export and assembly. Proteomic approaches have provided the first indications for a ribosome assembly pathway in eukaryotes and confirmed the dynamic character of the whole process.}, note = {0378-1119 Journal Article Review Review, Tutorial}, keywords = {Animals Cell Nucleolus/genetics/metabolism Eukaryotic Cells/*metabolism RNA, Ribosomal/genetics/metabolism Ribosomal Proteins/genetics/metabolism Ribosomes/genetics/*metabolism Saccharomyces cerevisiae/genetics/metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Human mitochondrial tRNAs in health and disease}, author = {C Florentz and B Sohm and P Tryoen-Toth and J Putz and M Sissler}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12943225}, isbn = {12943225}, year = {2003}, date = {2003-01-01}, journal = {Cell Mol Life Sci}, volume = {60}, number = {7}, pages = {1356-1375}, abstract = {The human mitochondrial genome encodes 13 proteins, all subunits of the respiratory chain complexes and thus involved in energy metabolism. These genes are translated by 22 transfer RNAs (tRNAs), also encoded by the mitochondrial genome, which form the minimal set required for reading all codons. Human mitochondrial tRNAs gained interest with the rapid discovery of correlations between point mutations in their genes and various neuromuscular and neurodegenerative disorders. In this review, emerging fundamental knowledge on the structure/function relationships of these particular tRNAs and an overview of the large variety of mechanisms within translation, affected by mutations, are summarized. Also, initial results on wide-ranging molecular consequences of mutations outside the frame of mitochondrial translation are highlighted. While knowledge of mitochondrial tRNAs in both health and disease increases, deciphering the intricate network of events leading different genotypes to the variety of phenotypes requires further investigation using adapted model systems.}, note = {1420-682x Journal Article Review Review, Academic}, keywords = {Base Sequence Genetic Diseases, FLORENTZ, Genetic, Inborn/*genetics Genome Human Mitochondria/*genetics Molecular Sequence Data Nucleic Acid Conformation RNA/chemistry/*genetics RNA, Messenger/genetics/metabolism RNA, Non-U.S. Gov't Translation, SISSLER, Transfer/chemistry/*genetics Reference Values Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Mitochondrial tRNA aminoacylation and human diseases.}, author = {C Florentz and M Sissler}, editor = {J Lapointe and L Brakier-Gringas}, url = {http://www.landesbioscience.com/curie/chapter/911}, year = {2003}, date = {2003-01-01}, booktitle = {Translation Mechanisms}, pages = {129-143}, publisher = {Landes Bioscience}, abstract = {The human mitochondrial (mt) genome encodes for only 13 proteins which are all subunits of transmembranar respiratory chain complexes. These complexes contribute to a major mt functions namely the synthesis of energy in the way of ATP. Translation of the mRNAs is performed by a set of 22 tRNAs, also encoded by the mt genome, and aminoacylated by nuclear encoded aminoacyl-tRNA synthetases imported into the mitochondria. More and more point mutations affecting mt tRNA genes are reported as correlated to severe neurodegenerative disorders. Since these mutations generally lead to decreased mt protein synthesis, understanding the genotype/phenotype relationships is primarily based on investigation of the possible impacts of individual mutations on various aspects of tRNA structural and functional properties. Here, the present knowledge on human mt aminoacylation systems, as well as the strategies developed to investigate mt aminoacylation, and the effects of point mutations in tRNAs on this process are reviewed. The diversity in the effects observed so far, for a same mutation as well as for various mutations, highlights the ongoing technical limitations in studying human mt aminoacylation. They also suggest that aminoacylation may be a focus for therapeutic strategies for some mutations, while the impact of other mutations needs to be searched as well at other levels of the tRNA structure/function relationship as at unforeseen levels in mitochondria.}, keywords = {FLORENTZ, SISSLER, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {A crystallographic study of the binding of 13 metal ions to two related RNA duplexes}, author = {E Ennifar and P Walter and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12736317}, isbn = {12736317}, year = {2003}, date = {2003-01-01}, journal = {Nucleic Acids Res}, volume = {31}, number = {10}, pages = {2671-2682}, abstract = {Metal ions, and magnesium in particular, are known to be involved in RNA folding by stabilizing secondary and tertiary structures, and, as cofactors, in RNA enzymatic activity. We have conducted a systematic crystallographic analysis of cation binding to the duplex form of the HIV-1 RNA dimerization initiation site for the subtype-A and -B natural sequences. Eleven ions (K+, Pb2+, Mn2+, Ba2+, Ca2+, Cd2+, Sr2+, Zn2+, Co2+, Au3+ and Pt4+) and two hexammines [Co (NH3)6]3+ and [Ru (NH3)6]3+ were found to bind to the DIS duplex structure. Although the two sequences are very similar, strong differences were found in their cation binding properties. Divalent cations bind almost exclusively, as Mg2+, at 'Hoogsteen' sites of guanine residues, with a cation-dependent affinity for each site. Notably, a given cation can have very different affinities for a priori equivalent sites within the same molecule. Surprisingly, none of the two hexammines used were able to efficiently replace hexahydrated magnesium. Instead, [Co (NH3)4]3+ was seen bound by inner-sphere coordination to the RNA. This raises some questions about the practical use of [Co (NH3)6]3+ as a [Mg (H2O)6]2+ mimetic. Also very unexpected was the binding of the small Au3+ cation exactly between the Watson-Crick sites of a G-C base pair after an obligatory deprotonation of N1 of the guanine base. This extensive study of metal ion binding using X-ray crystallography significantly enriches our knowledge on the binding of middleweight or heavy metal ions to RNA, particularly compared with magnesium.}, note = {1362-4962 Journal Article}, keywords = {Base Sequence Binding Sites/genetics Binding, Competitive Cations, Divalent/chemistry/metabolism Cobalt/chemistry/metabolism Comparative Study Crystallization Crystallography, ENNIFAR, Molecular Nucleic Acid Heteroduplexes/*chemistry/genetics/metabolism Oligoribonucleotides/chemistry/genetics/metabolism Platinum Compounds/chemistry/metabolism RNA/*chemistry/genetics/metabolism Ruthenium Compounds/chemistry/metabolism Support, Non-U.S. Gov't, Unité ARN, X-Ray Gold Compounds/chemistry/metabolism Magnesium/chemistry/metabolism Metals/*chemistry/metabolism Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {HIV-1 RNA dimerization initiation site is structurally similar to the ribosomal A site and binds aminoglycoside antibiotics}, author = {E Ennifar and J C Paillart and R Marquet and B Ehresmann and C Ehresmann and P Dumas and P Walter}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12435744}, isbn = {12435744}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {4}, pages = {2723-2730}, abstract = {Human immunodeficiency virus (HIV) genomic RNA is packaged into virions as a dimer. The first step of dimerization is the formation of a kissing-loop complex at the so-called dimerization initiation site (DIS). We found an unexpected and fortuitous resemblance between the HIV-1 DIS kissing-loop complex and the eubacterial 16 S ribosomal aminoacyl-tRNA site (A site), which is the target of aminoglycoside antibiotics. Similarities exist not only at the primary and secondary structure level but also at the tertiary structure level, as revealed by comparison of the respective DIS and A site crystal structures. Gel shift, inhibition of lead-induced cleavage, and footprinting experiments showed that paromomycin and neomycin specifically bind to the kissing-loop complex formed by the DIS, with an affinity and a geometry similar to that observed for the A site. Modeling of the aminoglycoside-DIS complex allowed us to identify antibiotic modifications likely to increase the affinity and/or the specificity for the DIS. This could be a starting point for designing antiviral drugs against HIV-1 RNA dimerization.}, note = {0021-9258 Journal Article}, keywords = {Anti-Bacterial Agents/*pharmacology Binding Sites Dimerization HIV-1/*metabolism Models, ENNIFAR, MARQUET, Molecular Neomycin/pharmacology Nucleic Acid Conformation Paromomycin/pharmacology Protein Binding RNA/metabolism *RNA, Non-U.S. Gov't Temperature Ultraviolet Rays, PAILLART, Unité ARN, Viral Ribosomes/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Functionalized self-assembled monolayer on gold for detection of human mitochondrial tRNA gene mutations}, author = {W Du and C Marsac and M Kruschina and F Ortigao and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14705775}, isbn = {14705775}, year = {2003}, date = {2003-01-01}, journal = {Anal Biochem}, volume = {322}, number = {1}, pages = {14-25}, abstract = {We developed a rapid and simple method to identify single-nucleotide polymorphisms (SNPs) in the human mitochondrial tRNA genes. This method is based on a universal, functionalized, self-assembled monolayer, XNA on Gold chip platform. A set of probes sharing a given allele-specific sequence with a single base substitution near the middle of the sequence was immobilized on chips and the chips were then hybridized with fluorescence-labeled reference targets produced by asymmetric polymerase chain reaction from patient DNA. The ratio of the hybridization signals from the reference and test targets with each probe was then calculated. A ratio of above 3 indicates the presence of a wild-type sequence and a ratio of below 0.3 indicates a mutant sequence. We tested the sensitivity of the chip for known mutations in tRNA(Leu(UUR)) and tRNA(Lys) genes and found that it can also be used to discriminate multiple mutations and heteroplasmy, two typical features of human mitochondrial DNA. The XNA on Gold biochip method is a simple and rapid microarray method that can be used to test rapidly and reliably any SNP in the mitochondrial genome or elsewhere. It will be particularly useful for detecting SNPs associated with human diseases.}, note = {0003-2697 Journal Article}, keywords = {Alleles *Gold Human MELAS Syndrome/genetics MERRF Syndrome/genetics Nucleic Acid Hybridization/genetics Oligonucleotide Array Sequence Analysis Point Mutation/*genetics Polymerase Chain Reaction Polymorphism, FLORENTZ, Non-U.S. Gov't, Single Nucleotide/*genetics RNA/*genetics RNA, Transfer/*genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Non-discriminating and discriminating aspartyl-tRNA synthetases differ in the anticodon-binding domain}, author = {C Charron and H Roy and M Blaise and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12660169}, isbn = {12660169}, year = {2003}, date = {2003-01-01}, journal = {EMBO J}, volume = {22}, number = {7}, pages = {1632-1643}, abstract = {In most organisms, tRNA aminoacylation is ensured by 20 aminoacyl-tRNA synthetases (aaRSs). In eubacteria, however, synthetases can be duplicated as in Thermus thermophilus, which contains two distinct AspRSs. While AspRS-1 is specific, AspRS-2 is non-discriminating and aspartylates tRNA(Asp) and tRNA(Asn). The structure at 2.3 A resolution of AspRS-2, the first of a non-discriminating synthetase, was solved. It differs from that of AspRS-1 but has resemblance to that of discriminating and archaeal AspRS from Pyrococcus kodakaraensis. The protein presents non-conventional features in its OB-fold anticodon-binding domain, namely the absence of a helix inserted between two beta-strands of this fold and a peculiar L1 loop differing from the large loops known to interact with tRNA(Asp) identity determinant C36 in conventional AspRSs. In AspRS-2, this loop is small and structurally homologous to that in AsnRSs, including conservation of a proline. In discriminating Pyrococcus AspRS, the L1 loop, although small, lacks this proline and is not superimposable with that of AspRS-2 or AsnRS. Its particular status is demonstrated by a loop-exchange experiment that renders the Pyrococcus AspRS non-discriminating.}, note = {0261-4189 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence *Anticodon Aspartate-tRNA Ligase/chemistry/*metabolism Binding Sites Models, Molecular Molecular Sequence Data Protein Conformation Pyrococcus/enzymology Sequence Homology, Non-U.S. Gov't Thermus thermophilus/enzymology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The modular structure of Escherichia coli threonyl-tRNA synthetase as both an enzyme and a regulator of gene expression}, author = {J Caillet and T Nogueira and B Masquida and F Winter and M Graffe and A C Dock-Bregeon and A Torres-Larios and R Sankaranarayanan and E Westhof and B Ehresmann and C Ehresmann and P Romby and M Springer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12581352}, isbn = {12581352}, year = {2003}, date = {2003-01-01}, journal = {Mol Microbiol}, volume = {47}, number = {4}, pages = {961-974}, abstract = {In addition to its role in tRNA aminoacylation, Escherichia coli threonyl-tRNA synthetase is a regulatory protein which binds a site, called the operator, located in the leader of its own mRNA and inhibits translational initiation by competing with ribosome binding. This work shows that the two essential steps of regulation, operator recognition and inhibition of ribosome binding, are performed by different domains of the protein. The catalytic and the C-terminal domain of the protein are involved in binding the two anticodon arm-like structures in the operator whereas the N-terminal domain of the enzyme is responsible for the competition with the ribosome. This is the first demonstration of a modular structure for a translational repressor and is reminiscent of that of transcriptional regulators. The mimicry between the operator and tRNA, suspected on the basis of previous experiments, is further supported by the fact that identical regions of the synthetase recognize both the operator and the tRNA anticodon arm. Based on these results, and recent structural data, we have constructed a computer-derived molecular model for the operator-threonyl-tRNA synthetase complex, which sheds light on several essential aspects of the regulatory mechanism.}, note = {0950-382x Journal Article}, keywords = {Amino Acyl/chemistry/metabolism Ribosomes/metabolism Support, Bacterial Genes, Bacterial Macromolecular Systems Models, Bacterial/chemistry/metabolism RNA, Binding Sites Binding, Competitive Escherichia coli/*enzymology/*genetics Evolution, Messenger/metabolism RNA, Molecular Gene Expression Regulation, Molecular Molecular Mimicry Molecular Structure Mutation Operator Regions (Genetics) Protein Structure, Non-U.S. Gov't Threonine-tRNA Ligase/*chemistry/genetics/*metabolism, ROMBY, Tertiary Protein Subunits RNA, Transfer, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {A single internal ribosome entry site containing a G quartet RNA structure drives fibroblast growth factor 2 gene expression at four alternative translation initiation codons}, author = {S Bonnal and C Schaeffer and L Creancier and S Clamens and H Moine and A C Prats and S Vagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12857733}, isbn = {12857733}, year = {2003}, date = {2003-01-01}, journal = {J Biol Chem}, volume = {278}, number = {41}, pages = {39330-39336}, abstract = {The 484-nucleotide (nt) alternatively translated region (ATR) of the human fibroblast growth factor 2 (FGF-2) mRNA contains four CUG and one AUG translation initiation codons. Although the 5'-end proximal CUG codon is initiated by a cap-dependent translation process, the other four initiation codons are initiated by a mechanism of internal entry of ribosomes. We undertook here a detailed analysis of the cis-acting elements defining the FGF-2 internal ribosome entry site (IRES). A thorough deletion analysis study within the 5'-ATR led us to define a 176-nt region as being necessary and sufficient for IRES function at four codons present in a downstream 308-nt RNA segment. Unexpectedly, a single IRES module is therefore responsible for translation initiation at four distantly localized codons. The determination of the FGF-2 5'-ATR RNA secondary structure by enzymatic and chemical probing experiments showed that the FGF-2 IRES contained two stem-loop regions and a G quartet motif that constitute novel structural determinants of IRES function.}, note = {0021-9258 Journal Article}, keywords = {Alternative Splicing Base Sequence Cell Line Codon, Complementary/genetics Fibroblast Growth Factor 2/*genetics Gene Expression Human Molecular Sequence Data Nucleic Acid Conformation Peptide Chain Initiation RNA, Initiator/genetics DNA, Messenger/*chemistry/*genetics Ribosomes/*metabolism Sequence Deletion Support, Non-U.S. Gov't Transfection, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Functional analysis in yeast of the Brix protein superfamily involved in the biogenesis of ribosomes}, author = {E Bogengruber and P Briza and E Doppler and H Wimmer and L Koller and F Fasiolo and B Senger and J H Hegemann and M Breitenbach}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12702244}, isbn = {12702244}, year = {2003}, date = {2003-01-01}, journal = {FEMS Yeast Res}, volume = {3}, number = {1}, pages = {35-43}, abstract = {An extensive homology search based on the sequence of the yeast protein Brx1p (biogenesis of ribosomes in Xenopus, YOL077c) revealed that it is a member of a superfamily of proteins sharing remarkable sequence similarities. Previous work on Brx1p showed that this protein is involved in the process of ribosome biogenesis [Kaser et al., Biol. Chem. 382 (2001) 1637-1647]. Brx1p is the founding member of one of the five existing eukaryotic subfamilies which are all present in yeast. Four of them are represented by one essential gene each and one family is represented by two closely related genes which can functionally replace each other but are essential together for survival. We created conditional alleles of four of the five genes which allowed us to study the effect of depletion of the respective proteins on the ribosome profiles of the strains. In this study we show that not only Brx1p but also three additional superfamily members, namely YHR088w (Rpf1p), YKR081c (Rpf2p) and the homologous proteins Ssf1p (YHR066w)/Ssf2p (YDR312w) are all involved in the multistep process of the assembly of the large ribosomal subunit. This agrees well with the fact that these three proteins, like Brx1p, are located in the nucleolus. Moreover, all four proteins closely interact functionally, because all four mutants are suppressed by the same multicopy suppressor gene.}, note = {1567-1356 Journal Article}, keywords = {Biogenesis Cell Nucleolus/*metabolism Genes, Non-U.S. Gov't, Suppressor Peptide Initiation Factors/metabolism RNA-Binding Proteins/metabolism/*physiology Ribosomal Proteins/*biosynthesis/genetics Ribosomes/*physiology Saccharomyces cerevisiae/metabolism/*physiology Saccharomyces cerevisiae Proteins/metabolism/*physiology Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Purification of turkey pancreatic phospholipase A2}, author = {R Ben Salah and N Zouari and J Reinbolt and H Mejdoub}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14586101}, isbn = {14586101}, year = {2003}, date = {2003-01-01}, journal = {Biosci Biotechnol Biochem}, volume = {67}, number = {10}, pages = {2139-2144}, abstract = {Turkey pancreatic phospholipase (TPP) has been purified from delipidated pancreases. The purification included ammonium sulfate fractionation, acidic (pH 5) treatment, followed by sequencial column chromatographies on DEAE-cellulose, Sephadex G-75, and reverse phase high pressure liquid chromatography. The purified enzyme was found to be a monomeric protein with molecular mass of 14 kDa. The optimal activity was measured at pH 8 and 37 degrees C using egg yolk emulsion as substrate. Our results show that the enzyme (TPP) was not stable for 1 h at 60 degrees C, and that bile salt and Ca2+ were required for the expression of the purified enzyme. The sequence of the N-terminal amino acids of the purified enzyme shows a very close similarity between TPP and all other known pancreatic phospholipases.}, note = {0916-8451 Journal Article}, keywords = {Amino Acid Sequence Ammonium Sulfate Animals Bile Acids and Salts Calcium Chromatography Hydrogen-Ion Concentration Molecular Sequence Data Molecular Weight Pancreas/*enzymology Phospholipases A/*isolation & purification Temperature *Turkeys, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Mg2+ binding sites of the 5S rRNA loop E motif as investigated by molecular dynamics simulations}, author = {P Auffinger and L Bielecki and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12837388}, isbn = {12837388}, year = {2003}, date = {2003-01-01}, journal = {Chem Biol}, volume = {10}, number = {6}, pages = {551-561}, abstract = {Molecular dynamics simulations have been used to investigate the binding of Mg(2+) ions to the deep groove of the eubacterial 5S rRNA loop E. The simulations suggest that long-lived and specific water-mediated interactions established between the hydrated ions and the RNA atoms lining up the binding sites contribute to the stabilization of this motif. The Mg(2+) binding specificity is modulated by two factors: (i) a required electrostatic complementarity and (ii) a structural correspondence between the hydrated ion and its binding pocket that can be estimated by its degree of dehydration and the resulting number and lifetime of the intervening water-mediated contacts. Two distinct binding modes for pentahydrated Mg(2+) ions that result in a significant freezing of the tumbling motions of the ions are described, and mechanistic details related to the stabilization of nucleic acids by divalent ions are provided.}, note = {1074-5521 Journal Article}, keywords = {5S/*chemistry/metabolism Support, Bacterial/*chemistry RNA, Binding Sites Cations, Divalent Computer Simulation Crystallization Electrostatics Hydrogen Bonding Magnesium/*chemistry/metabolism Models, Molecular Molecular Conformation Nucleic Acid Conformation RNA Stability RNA, Non-U.S. Gov't Thermodynamics Water/chemistry, Ribosomal, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{georgakilas_amino_2002, title = {Amino acid functionalisation of water soluble carbon nanotubes}, author = {Vasilios Georgakilas and Nikos Tagmatarchis and Davide Pantarotto and Alberto Bianco and Jean-Paul Briand and Maurizio Prato}, doi = {10.1039/b209843a}, issn = {1359-7345}, year = {2002}, date = {2002-12-01}, journal = {Chemical Communications (Cambridge, England)}, number = {24}, pages = {3050--3051}, abstract = {High solubility of SWNTs and MWNTs in water is obtained by organic functionalisation; derivatisation with N-protected glycine is also easily achieved.}, keywords = {Amino Acids, carbon, I2CT, Nanotubes, Solubility, Team-Bianco, water}, pubstate = {published}, tppubtype = {article} } @article{ligoxygakis_serpin_2002, title = {A serpin mutant links Toll activation to melanization in the host defence of Drosophila}, author = {Petros Ligoxygakis and Nadège Pelte and Chuanyi Ji and Vincent Leclerc and Bernard Duvic and Marcia Belvin and Haobo Jiang and Jules A Hoffmann and Jean-Marc Reichhart}, issn = {0261-4189}, year = {2002}, date = {2002-12-01}, journal = {EMBO J.}, volume = {21}, number = {23}, pages = {6330--6337}, abstract = {A prominent response during the Drosophila host defence is the induction of proteolytic cascades, some of which lead to localized melanization of pathogen surfaces, while others activate one of the major players in the systemic antimicrobial response, the Toll pathway. Despite the fact that gain-of-function mutations in the Toll receptor gene result in melanization, a clear link between Toll activation and the melanization reaction has not been firmly established. Here, we present evidence for the coordination of hemolymph-borne melanization with activation of the Toll pathway in the Drosophila host defence. The melanization reaction requires Toll pathway activation and depends on the removal of the Drosophila serine protease inhibitor Serpin27A. Flies deficient for this serpin exhibit spontaneous melanization in larvae and adults. Microbial challenge induces its removal from the hemolymph through Toll-dependent transcription of an acute phase immune reaction component.}, keywords = {Animals, Cell Surface, Hemolymph, hoffmann, infection, M3i, Melanins, Receptors, reichhart, Serpins, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{munier_pvf2_2002, title = {PVF2, a PDGF/VEGF-like growth factor, induces hemocyte proliferation in Drosophila larvae}, author = {Anne-Isabelle Munier and Daniel Doucet and Emmanuel Perrodou and Daniel Zachary and Marie Meister and Jules A Hoffmann and Charles A Janeway and Marie Lagueux}, doi = {10.1093/embo-reports/kvf242}, issn = {1469-221X}, year = {2002}, date = {2002-12-01}, journal = {EMBO Rep.}, volume = {3}, number = {12}, pages = {1195--1200}, abstract = {Blood cells play a crucial role in both morphogenetic and immunological processes in Drosophila, yet the factors regulating their proliferation remain largely unknown. In order to address this question, we raised antibodies against a tumorous blood cell line and identified an antigenic determinant that marks the surface of prohemocytes and also circulating plasmatocytes in larvae. This antigen was identified as a Drosophila homolog of the mammalian receptor for platelet-derived growth factor (PDGF)/vascular endothelial growth factor (VEGF). The Drosophila receptor controls cell proliferation in vitro. By overexpressing in vivo one of its putative ligands, PVF2, we induced a dramatic increase in circulating hemocytes. These results identify the PDGF/VEGF receptor homolog and one of its ligands as important players in Drosophila hematopoiesis.}, keywords = {Animals, Antibodies, Blotting, Cell Differentiation, Hemocytes, hoffmann, Immunohistochemistry, Larva, ligands, M3i, Platelet-Derived Growth Factor, Receptors, Vascular Endothelial Growth Factor, Western}, pubstate = {published}, tppubtype = {article} } @article{kambris_tissue_2002, title = {Tissue and stage-specific expression of the Tolls in Drosophila embryos}, author = {Zakaria Kambris and Jules A Hoffmann and Jean-Luc Imler and Maria Capovilla}, issn = {1567-133X}, year = {2002}, date = {2002-12-01}, journal = {Gene expression patterns: GEP}, volume = {2}, number = {3-4}, pages = {311--317}, abstract = {The Drosophila transmembrane receptor Toll plays a key role in specifying the dorsoventral axis of the embryo. At later stages of development, it controls the immune response of the fly to fungal and Gram-positive bacterial infections. The Drosophila genome has a total of nine Toll-like genes, including the previously characterized Toll (Toll-1) and 18-wheeler (Toll-2). Here we describe the embryonic expression patterns of the seven Toll-like genes Toll-3 through Toll-9. We find that these genes have distinct expression domains and that their expression is dynamically changing throughout embryonic development. This complex and tissue-specific regulation of Toll-like gene expression strongly suggests a role in embryonic development for most Drosophila Tolls. The evolving picture on the Toll family members in Drosophila contrasts with that of mammalian Toll-like receptors, which are predominantly expressed in immune responsive cells where their activation occurs via microbial structural determinants.}, keywords = {Animals, Blotting, Cell Surface, Gene Expression Profiling, hoffmann, imler, Larva, M3i, Multigene Family, Northern, Receptors, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{duvic_notch_2002, title = {Notch signaling controls lineage specification during Drosophila larval hematopoiesis}, author = {Bernard Duvic and Jules A Hoffmann and Marie Meister and Julien Royet}, issn = {0960-9822}, year = {2002}, date = {2002-11-01}, journal = {Curr. Biol.}, volume = {12}, number = {22}, pages = {1923--1927}, abstract = {Drosophila larval hemocytes originate from a hematopoietic organ called lymph glands, which are composed of paired lobes located along the dorsal vessel. Two mature blood cell populations are found in the circulating hemolymph: the macrophage-like plasmatocytes, and the crystal cells that contain enzymes of the immune-related melanization process. A third class of cells, called lamellocytes, are normally absent in larvae but differentiate after infection by parasites too large to be phagocytosed. Here we present evidence that the Notch signaling pathway plays an instructive role in the differentiation of crystal cells. Loss-of-function mutations in Notch result in severely decreased crystal cell numbers, whereas overexpression of Notch provokes the differentiation of high numbers of these cells. We demonstrate that, in this process, Serrate, not Delta, is the Notch ligand. In addition, Notch function is necessary for lamellocyte proliferation upon parasitization, although Notch overexpression does not result in lamellocyte production. Finally, Notch does not appear to play a role in the differentiation of the plasmatocyte lineage. This study underlines the existence of parallels in the genetic control of hematopoiesis in Drosophila and in mammals.}, keywords = {Animals, Cell Differentiation, Hematopoiesis, hoffmann, Larva, Lymphoid Tissue, M3i, Membrane Proteins, Notch, Receptors, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{naitza_drosophila_2002, title = {The Drosophila immune defense against gram-negative infection requires the death protein dFADD}, author = {Silvia Naitza and Carine Rossé and Christine Kappler and Philippe Georgel and Marcia Belvin and David Gubb and Jacques Camonis and Jules A Hoffmann and Jean-Marc Reichhart}, issn = {1074-7613}, year = {2002}, date = {2002-11-01}, journal = {Immunity}, volume = {17}, number = {5}, pages = {575--581}, abstract = {Drosophila responds to Gram-negative infections by mounting an immune response that depends on components of the IMD pathway. We recently showed that imd encodes a protein with a death domain with high similarity to that of mammalian RIP. Using a two-hybrid screen in yeast, we have isolated the death protein dFADD as a molecule that associates with IMD. Our data show that loss of dFADD function renders flies highly susceptible to Gram-negative infections without affecting resistance to Gram-positive bacteria. By genetic analysis we show that dFADD acts downstream of IMD in the pathway that controls inducibility of the antibacterial peptide genes.}, keywords = {Adaptor Proteins, Animals, Carrier Proteins, Fas-Associated Death Domain Protein, Gene Expression Regulation, Gram-Negative Bacterial Infections, hoffmann, Immunity, M3i, reichhart, Signal Transducing, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{pantarotto_solid-phase_2002, title = {Solid-phase synthesis of fullerene-peptides}, author = {Davide Pantarotto and Alberto Bianco and Federica Pellarini and Alessandro Tossi and Anna Giangaspero and Igor Zelezetsky and Jean-Paul Briand and Maurizio Prato}, doi = {10.1021/ja027603q}, issn = {0002-7863}, year = {2002}, date = {2002-10-01}, journal = {Journal of the American Chemical Society}, volume = {124}, number = {42}, pages = {12543--12549}, abstract = {The solid-phase synthesis of peptides (SPPS) containing [60]fullerene-functionalized amino acids is reported. A new amino acid, fulleropyrrolidino-glutamic acid (Fgu), is used for the SPPS of a series of analogues of different length based on the natural Leu(5)-Enkephalin and on cationic antimicrobial peptides. These fullero-peptides were prepared on different solid supports to analyze the influence of the resin on the synthesis. Optimized protocols for the coupling and deprotection procedures were determined allowing the synthesis of highly pure peptides in sufficient quantities for evaluation of biological activities. In particular, to avoid side reactions of the fullerene moiety with bases and nucleophiles, the removal of the protecting groups was performed under inert conditions (nitrogen or argon in the dark). We have encountered serious problems with the recovery of the crude compounds, especially when Fgu was inserted in the proximity of the resin core as fullero-peptides tend to remain embedded inside the resin. Eventually, all of the fullero-peptides were easily purified, and the cationic peptides were tested for their antimicrobial activities. They displayed a specific activity against the Gram-positive bacterium S. aureus and also lysed erythrocytes. The availability of a fullero-amino acid easily useable in the SPPS of fullero-peptides may thus open the way to the synthesis of new types of biologically active oligomers.}, keywords = {Amino Acids, Anti-Bacterial Agents, Anti-Infective Agents, Candida albicans, Electrospray Ionization, Enkephalin, Escherichia coli, Fluorenes, Fullerenes, I2CT, Leucine, Mass, Microbial Sensitivity Tests, Oligopeptides, Spectrometry, Staphylococcus aureus, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{cohen_study_2002, title = {Study of Antigen-Processing Steps Reveals Preferences Explaining Differential Biological Outcomes of Two HLA-A2-Restricted Immunodominant Epitopes from Human Immunodeficiency Virus Type 1}, author = {W M Cohen and A Bianco and F Connan and L Camoin and M Dalod and G Lauvau and E Ferriès and B Culmann-Penciolelli and P M van Endert and J P Briand and J Choppin and J G Guillet}, url = {https://jvi.asm.org/content/76/20/10219}, doi = {10.1128/JVI.76.20.10219-10225.2002}, issn = {0022-538X, 1098-5514}, year = {2002}, date = {2002-10-01}, urldate = {2020-03-31}, journal = {Journal of Virology}, volume = {76}, number = {20}, pages = {10219--10225}, abstract = {Cytotoxic T-lymphocyte (CTL) responses directed to different human immunodeficiency virus (HIV) epitopes vary in their protective efficacy. In particular, HIV-infected cells are much more sensitive to lysis by anti-Gag/p17(77-85)/HLA-A2 than to that by anti-polymerase/RT(476-484)/HLA-A2 CTL, because of a higher density of p17(77-85) complexes. This report describes multiple processing steps favoring the generation of p17(77-85) complexes: (i) the exact COOH-terminal cleavage of epitopes by cellular proteases occurred faster and more frequently for p17(77-85) than for RT(476-484), and (ii) the binding efficiency of the transporter associated with antigen processing was greater for p17(77-85) precursors than for the RT(476-484) epitope. Surprisingly, these peptides, which differed markedly in their antigenicity, displayed qualitatively and quantitatively similar immunogenicity, suggesting differences in the mechanisms governing these phenomena. Here, we discuss the mechanisms responsible for such differences.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{christophides_immunity-related_2002, title = {Immunity-related genes and gene families in Anopheles gambiae}, author = {George K Christophides and Evgeny Zdobnov and Carolina Barillas-Mury and Ewan Birney and Stephanie A Blandin and Claudia Blass and Paul T Brey and Frank H Collins and Alberto Danielli and George Dimopoulos and Charles Hetru and Ngo T Hoa and Jules A Hoffmann and Stefan M Kanzok and Ivica Letunic and Elena A Levashina and Thanasis G Loukeris and Gareth Lycett and Stephan Meister and Kristin Michel and Luis F Moita and Hans-Michael Müller and Mike A Osta and Susan M Paskewitz and Jean-Marc Reichhart and Andrey Rzhetsky and Laurent Troxler and Kenneth D Vernick and Dina Vlachou and Jennifer Volz and Christian von Mering and Jiannong Xu and Liangbiao Zheng and Peer Bork and Fotis C Kafatos}, url = {http://www.ncbi.nlm.nih.gov/pubmed/12364793}, doi = {10.1126/science.1077136}, issn = {1095-9203}, year = {2002}, date = {2002-10-01}, journal = {Science}, volume = {298}, number = {5591}, pages = {159--165}, abstract = {We have identified 242 Anopheles gambiae genes from 18 gene families implicated in innate immunity and have detected marked diversification relative to Drosophila melanogaster. Immune-related gene families involved in recognition, signal modulation, and effector systems show a marked deficit of orthologs and excessive gene expansions, possibly reflecting selection pressures from different pathogens encountered in these insects' very different life-styles. In contrast, the multifunctional Toll signal transduction pathway is substantially conserved, presumably because of counterselection for developmental stability. Representative expression profiles confirm that sequence diversification is accompanied by specific responses to different immune challenges. Alternative RNA splicing may also contribute to expansion of the immune repertoire.}, keywords = {Alternative Splicing, Animals, Anopheles, Apoptosis, bacteria, bioinformatic, blandin, Catechol Oxidase, Computational Biology, Enzyme Precursors, Gene Expression Regulation, Genes, Genetic, Genome, hoffmann, Immunity, Innate, Insect, Insect Proteins, M3i, Multigene Family, Peptides, Phylogeny, Plasmodium, Protein Structure, reichhart, Selection, Serine Endopeptidases, Serpins, Signal Transduction, Tertiary}, pubstate = {published}, tppubtype = {article} } @article{S2002, title = {Reverse genetics in the mosquito, Anopheles gambiae : targeted disruption of the Defensin gene}, author = {Stéphanie A Blandin and L F Moita and T Kocher and M Wilm and Fotis C Kafatos and Elena A Levashina}, url = {http://www.ncbi.nlm.nih.gov/pubmed/12189180}, year = {2002}, date = {2002-09-02}, journal = {EMBO Rep.}, volume = {3}, number = {9}, pages = {852-6}, abstract = {Anopheles gambiae, the major vector of human malaria parasite, is an important insect model to study vector-parasite interactions. Here, we developed a simple in vivo double-stranded RNA (dsRNA) knockout approach to determine the function of the mosquito antimicrobial peptide gene Defensin. We injected dsRNA into adults and observed efficient and reproducible silencing of Defensin. Analysis of the knockdown phenotype revealed that this peptide is required for the mosquito antimicrobial defense against Gram-positive bacteria. In contrast, in mosquitoes infected by Plasmodium berghei, no loss of mosquito viability and no significant effect on the development and morphology of the parasite midgut stages were observed in the absence of Defensin. We conclude that this peptide is not a major antiparasitic factor in A. gambiae in vivo. Our results open new perspectives for the study of mosquito gene function in vivo and provide a basis for genome-scale systematic functional screens by targeted gene silencing.}, keywords = {blandin, defensin, M3i}, pubstate = {published}, tppubtype = {article} } @article{nisole_anti-hiv_2002, title = {The Anti-HIV Pentameric Pseudopeptide HB-19 Binds the C-terminal End of Nucleolin and Prevents Anchorage of Virus Particles in the Plasma Membrane of Target Cells}, author = {Sébastien Nisole and Elias A Said and Claudia Mische and Marie-Christine Prevost and Bernard Krust and Philippe Bouvet and Alberto Bianco and Jean-Paul Briand and Ara G Hovanessian}, url = {http://www.jbc.org/content/277/23/20877}, doi = {10.1074/jbc.M110024200}, issn = {0021-9258, 1083-351X}, year = {2002}, date = {2002-07-01}, urldate = {2020-03-31}, journal = {Journal of Biological Chemistry}, volume = {277}, number = {23}, pages = {20877--20886}, abstract = {The multivalent pseudopeptide HB-19 that binds the cell-surface-expressed nucleolin is a potent inhibitor of human immunodeficiency virus (HIV) infection by blocking virus particle attachment and thus anchorage in the plasma membrane. We show that cross-linking of surface-bound HB-19A (like HB-19 but with a modified template) results in aggregation of HB-19A with surface nucleolin. Consistent with its specific action, HB-19A binding to different types of cells reaches saturation at concentrations that have been reported to result in inhibition of HIV infection. By using Chinese hamster ovary mutant cell lines, we confirm that the binding of HB-19A to surface nucleolin is independent of heparan and chondroitin sulfate proteoglycans. In vitro generated full-length nucleolin was found to bind HB-19A, whereas the N-terminal part containing the acidic amino acid stretches of nucleolin did not. The use of various deletion constructs of the C-terminal part of nucleolin then permitted the identification of the extreme C-terminal end of nucleolin, containing repeats of the amino acid motif, RGG, as the domain that binds HB-19A. Finally, a synthetic peptide corresponding to the last C-terminal 63 amino acids was able to inhibit HIV infection at the stage of HIV attachment to cells, thus suggesting that this domain could be functional in the HIV anchorage process.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{ligoxygakis_activation_2002, title = {Activation of Drosophila Toll during fungal infection by a blood serine protease}, author = {Petros Ligoxygakis and Nadège Pelte and Jules A Hoffmann and Jean-Marc Reichhart}, doi = {10.1126/science.1072391}, issn = {1095-9203}, year = {2002}, date = {2002-07-01}, journal = {Science}, volume = {297}, number = {5578}, pages = {114--116}, abstract = {Drosophila host defense to fungal and Gram-positive bacterial infection is mediated by the Spaetzle/Toll/cactus gene cassette. It has been proposed that Toll does not function as a pattern recognition receptor per se but is activated through a cleaved form of the cytokine Spaetzle. The upstream events linking infection to the cleavage of Spaetzle have long remained elusive. Here we report the identification of a central component of the fungal activation of Toll. We show that ethylmethane sulfonate-induced mutations in the persephone gene, which encodes a previously unknown serine protease, block induction of the Toll pathway by fungi and resistance to this type of infection.}, keywords = {Animals, Cell Surface, Chromosome Mapping, Escherichia coli, Female, Gene Expression Regulation, Genes, Gram-Positive Cocci, Hemolymph, hoffmann, Hypocreales, Insect, Insect Proteins, M3i, Male, Mutation, Protein Sorting Signals, Protein Structure, Receptors, reichhart, Serine Endopeptidases, Tertiary, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{raya_proton_2002, title = {Proton dipolar recoupling in resin-bound peptides under high-resolution magic angle spinning}, author = {Jésus Raya and Alberto Bianco and Julien Furrer and Jean-Paul Briand and Martial Piotto and Karim Elbayed}, doi = {10.1006/jmre.2002.2573}, issn = {1090-7807}, year = {2002}, date = {2002-07-01}, journal = {Journal of Magnetic Resonance (San Diego, Calif.: 1997)}, volume = {157}, number = {1}, pages = {43--51}, abstract = {Rotational resonance and radiofrequency-driven dipolar recoupling (RFDR) experiments have been used to recover the weak proton dipolar interaction present in peptides bound to swollen resins spun at the magic angle. The intensity of the correlation peaks obtained using these sequences is shown to be significantly stronger than the one obtained using the classical NOESY experiment. In addition, it is found that during the relatively long mixing times required to transfer magnetization in such soft materials, the RFDR sequence also achieves magnetization transfer via the scalar J-coupling.}, keywords = {Amino Acid Sequence, biomolecular, Foot-and-Mouth Disease Virus, I2CT, Nuclear Magnetic Resonance, Peptides, Plant, Resins, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @inbook{EA2002, title = {Thioester-Containing Proteins of Protostomes}, author = {Elena A Levashina and Stéphanie A Blandin and L F Moita and Marie Lagueux and Fotis C Kafatos}, editor = {Alan R B Ezekowitz and Jules A Hoffmann}, year = {2002}, date = {2002-06-08}, pages = {155-173}, publisher = {Humana Press Inc, Totowa, NJ}, chapter = {Infectious Disease: Innate Immunity.}, keywords = {blandin, M3i, Thioester-Containing Protein}, pubstate = {published}, tppubtype = {inbook} } @article{monneaux_epitope_2002, title = {Epitope spreading in systemic lupus erythematosus: identification of triggering peptide sequences}, author = {Fanny Monneaux and Sylviane Muller}, doi = {10.1002/art.10263}, issn = {0004-3591}, year = {2002}, date = {2002-06-01}, journal = {Arthritis and Rheumatism}, volume = {46}, number = {6}, pages = {1430--1438}, keywords = {Amino Acid Sequence, Animals, B-Lymphocyte, Epitopes, Humans, I2CT, Lupus Erythematosus, Molecular Sequence Data, Monneaux, Systemic, T-Lymphocyte, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{bates_adamdec1_2002, title = {The ADAMDEC1 (decysin) gene structure: evolution by duplication in a metalloprotease gene cluster on chromosome 8p12}, author = {Elizabeth E M Bates and Wolf H Fridman and Chris G F Mueller}, doi = {10.1007/s00251-002-0430-3}, issn = {0093-7711}, year = {2002}, date = {2002-05-01}, journal = {Immunogenetics}, volume = {54}, number = {2}, pages = {96--105}, abstract = {Members of the ADAM superfamily of metalloprotease genes are involved in a number of biological processes, including fertilization, neurogenesis, muscle development, and the immune response. These proteins have been classified into several groups. The prototypic ADAM family is comprised of a pro-domain, a metalloprotease domain, a disintegrin domain, a cysteine-rich region, a transmembrane domain, and a variable cytoplasmic tail. We recently identified a novel member of this superfamily, ADAMDEC1 (decysin). Due to the partial lack of a disintegrin domain and the total lack of a cysteine-rich domain, this protein has been placed in a novel subclass of the ADAM gene family. We have investigated the gene structure of the human and mouse ADAMDEC1 and have revealed a metalloprotease gene cluster on human Chromosome 8p12 comprising ADAMDEC1, ADAM7, and ADAM28. Our results suggest that ADAMDEC1 has arisen by partial gene duplication from an ancestral gene at this locus and has acquired a novel function. ADAMDEC1 is expressed in the immune system, by dendritic cells and macrophages. The relatedness of ADAMDEC1, ADAM7, and ADAM28 suggests that these proteases share a similar function.}, keywords = {ADAM Proteins, Amino Acid Sequence, Animals, Base Sequence, Chromosomes, Evolution, Gene Dosage, Gene Duplication, Genetic, Human, Humans, Inbred BALB C, Macaca mulatta, Membrane Glycoproteins, Metalloendopeptidases, Mice, Molecular, Molecular Sequence Data, Multigene Family, Pair 8, Promoter Regions, Sequence Alignment, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{gottar_drosophila_2002b, title = {The Drosophila immune response against Gram-negative bacteria is mediated by a peptidoglycan recognition protein}, author = {Marie Gottar and Vanessa Gobert and Tatiana Michel and Marcia Belvin and Geoffrey Duyk and Jules A Hoffmann and Dominique Ferrandon and Julien Royet}, doi = {10.1038/nature734}, isbn = {0028-0836}, year = {2002}, date = {2002-03-01}, journal = {Nature}, volume = {416}, pages = {640--644}, abstract = {The antimicrobial defence of Drosophila relies largely on the challenge-induced synthesis of an array of potent antimicrobial peptides by the fat body. The defence against Gram-positive bacteria and natural fungal infections is mediated by the Toll signalling pathway, whereas defence against Gram-negative bacteria is dependent on the Immune deficiency (IMD) pathway. Loss-of-function mutations in either pathway reduce the resistance to corresponding infections. The link between microbial infections and activation of these two pathways has remained elusive. The Toll pathway is activated by Gram-positive bacteria through a circulating Peptidoglycan recognition protein (PGRP-SA). PGRPs appear to be highly conserved from insects to mammals, and the Drosophila genome contains 13 members. Here we report a mutation in a gene coding for a putative transmembrane protein, PGRP-LC, which reduces survival to Gram-negative sepsis but has no effect on the response to Gram-positive bacteria or natural fungal infections. By genetic epistasis, we demonstrate that PGRP-LC acts upstream of the imd gene. The data on PGRP-SA with respect to the response to Gram-positive infections, together with the present report, indicate that the PGRP family has a principal role in sensing microbial infections in Drosophila.}, keywords = {Animal, Anti-Infective Agents/metabolism, Carrier Proteins/biosynthesis/genetics/*immunology, Drosophila melanogaster/genetics/*immunology/*microbiology, Drosophila Proteins/genetics/metabolism, Epistasis, Female, ferrandon, Genes, Genetic, Genetic Predisposition to Disease, Gram-Negative Bacteria/*immunology/physiology, hoffmann, Human, Insect/genetics, M3i, Messenger/genetics/metabolism, Mutation, Non-U.S. Gov't, P.H.S., Phenotype, RNA, Signal Transduction, Support, Survival Rate, Transgenes/genetics, U.S. Gov't}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_drosophila_2002, title = {Drosophila innate immunity: an evolutionary perspective}, author = {Jules A Hoffmann and Jean-Marc Reichhart}, doi = {10.1038/ni0202-121}, issn = {1529-2908}, year = {2002}, date = {2002-02-01}, journal = {Nat. Immunol.}, volume = {3}, number = {2}, pages = {121--126}, abstract = {In response to microbial infections, Drosophila mounts a multifaceted immune response involving humoral reactions that culminate in the destruction of invading organisms by lytic peptides. These defense mechanisms are activated via two distinct signaling pathways. One of these, the Toll pathway, controls resistance to fungal and Gram-positive bacterial infections, whereas the Imd pathway is responsible for defense against Gram-negative bacterial infections. Current evidence indicates that recognition of infectious nonself agents results from interactions between microbial wall components and extracellular pattern recognition proteins. We discuss here evolutionary perspectives on our present understanding of the antimicrobial defenses of Drosophila.}, keywords = {Animals, Biological Evolution, Cell Surface, hoffmann, Immunity, Immunological, Innate, M3i, Membrane Glycoproteins, Models, Receptors, reichhart, Signal Transduction, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{, title = {The crystallization of biological macromolecules under microgravity: a way to more accurate three-dimensional structures?}, author = {B Lorber}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12479400}, isbn = {12479400}, year = {2002}, date = {2002-01-01}, journal = {Biochim Biophys Acta-Proteins Proteomics}, volume = {1599}, number = {1-2}, pages = {1-8}, abstract = {The crystallization of proteins and other biological particles (including nucleic acids, nucleo-protein complexes and large assemblies such as nucleosomes, ribosomal subunits or viruses) in a microgravity environment can produce crystals having lesser defects than crystals prepared under normal gravity on earth. Such microgravity-grown crystals can diffract X-rays to a higher resolution and have a lower mosaic spread. The inferred electron density maps can be richer in details owing to which more accurate three-dimensional structure models can be built. Major results reported in this field of research are reviewed. Novel ones obtained with the Advanced Protein Crystallization Facility are presented. For structural biology, practical applications and implications associated with crystallization and crystallography onboard the International Space Station are discussed.}, note = {0006-3002 Journal Article Review Review, Tutorial}, keywords = {Animals Crystallization Human Protein Conformation Proteins/*chemistry Support, Non-U.S. Gov't *Weightlessness, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Lead(II) as a probe for investigating RNA structure in vivo}, author = {M Lindell and P Romby and E G Wagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11991646}, isbn = {11991646}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {4}, pages = {534-541}, abstract = {In this communication, we describe a simple and reliable method for RNA structure determination in vivo, using the divalent ion, lead(II), as a structural probe. Lead(II) is known to cleave RNA within single-stranded regions, loops, and bulges, whereas cleavages in double-stranded regions are weaker or absent. Because the ion easily entered bacterial cells, Escherichia coli cultures were treated by addition of 50-100 mM lead(II) acetate for 3-7 min, resulting in partial cleavage of RNA in vivo. Cleavage positions were mapped by reverse transcription analysis of total extracted RNA. Three RNAs were analyzed: tmRNA, CopT (the target of the antisense RNA CopA), and the leader region of the ompF mRNA. All three RNAs had previously been analyzed in vitro, and secondary structure models were available. The results presented here show that lead(II) cleavages in vivo yield detailed structural information for these RNAs, which was in good agreement with the models proposed based on in vitro work. These data illustrate the potential of lead(II) as a sequence-independent RNA structure probe for use in living cells.}, note = {1355-8382 Journal Article}, keywords = {Base Sequence Biochemistry/*methods Comparative Study Lead/*chemistry/*metabolism Molecular Sequence Data Nucleic Acid Conformation Porins/genetics RNA/*chemistry/*metabolism RNA, Messenger/chemistry/metabolism Support, Non-U.S. Gov't, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Novel selenoproteins identified from genomic sequence data}, author = {A Lescure and D Gautheret and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11898439}, isbn = {11898439}, year = {2002}, date = {2002-01-01}, journal = {Methods Enzymol}, volume = {347}, pages = {57-70}, note = {0076-6879 Journal Article}, keywords = {3' Untranslated Regions Animals COS Cells Codon/genetics DNA Transposable Elements DNA, Complementary/genetics Databases, Human Human Nucleic Acid Conformation Proteins/*genetics RNA, LESCURE, Messenger/chemistry/genetics Selenium Radioisotopes Support, Non-U.S. Gov't, Nucleic Acid Genome, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Protein factors mediating selenoprotein synthesis}, author = {A Lescure and D Fagegaltier and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12370018}, isbn = {12370018}, year = {2002}, date = {2002-01-01}, journal = {Curr Protein Pept Sci}, volume = {3}, number = {1}, pages = {143-151}, abstract = {The amino acid selenocysteine represents the major biological form of selenium. Both the synthesis of selenocysteine and its co-translational incorporation into selenoproteins in response to an in-frame UGA codon, require a complex molecular machinery. To decode the UGA Sec codon in eubacteria, this machinery comprises the tRNASec, the specialized elongation factor SelB and the SECIS hairpin in the selenoprotein mRNAs. SelB conveys the Sec-tRNASec to the A site of the ribosome through binding to the SECIS mRNA hairpin adjacent to the UGA Sec codon. SelB is thus a bifunctional factor, carrying functional homology to elongation factor EF-Tu in its N-terminal domain and SECIS RNA binding activity via its C-terminal extension. In archaea and eukaryotes, selenocysteine incorporation exhibits a higher degree of complexity because the SECIS hairpin is localized in the 3' untranslated region of the mRNA. In the last couple of years, remarkable progress has been made toward understanding the underlying mechanism in mammals. Indeed, the discovery of the SECIS RNA binding protein SBP2, which is not a translation factor, paved the way for the subsequent isolation of mSelB/EFSec, the mammalian homolog of SelB. In contrast to the eubacterial SelB, the specialized elongation factor mSelB/EFSec the SECIS RNA binding function. The role is carried out by SBP2 that also forms a protein-protein complex with mSelB/EFSec. As a consequence, an important difference between the eubacterial and eukaryal selenoprotein synthesis machineries is that the functions of SelB are divided into two proteins in eukaryotes. Obviously, selenoprotein synthesis represents a higher degree of complexity than anticipated, and more needs to be discovered in eukaryotes. In this review, we will focus on the structural and functional aspects of the SelB and SBP2 factors in selenoprotein synthesis.}, note = {1389-2037 Journal Article Review Review, Tutorial}, keywords = {Bacterial Proteins/genetics/*metabolism Guanosine Triphosphate/metabolism Models, Biological Peptide Elongation Factors/*metabolism Protein Binding Proteins/*biosynthesis RNA, LESCURE, Messenger/metabolism RNA, Transfer/metabolism RNA-Binding Proteins/*metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {cDNA cloning, expression pattern and RNA binding analysis of human selenocysteine insertion sequence (SECIS) binding protein 2}, author = {A Lescure and C Allmang and K Yamada and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12095701}, isbn = {12095701}, year = {2002}, date = {2002-01-01}, journal = {Gene}, volume = {291}, number = {1-2}, pages = {279-285}, abstract = {Selenocysteine and selenoprotein synthesis require a complex molecular machinery in mammals. Among the key players is the RNA-protein complex formed by the selenocysteine insertion sequence (SECIS) binding protein (SBP2) and the SECIS element, an RNA hairpin in the 3' untranslated regions of selenoprotein messenger RNAs (mRNAs). We have isolated the DNA complementary to mRNA of the human SBP2, enabling us to establish that it differs from a previously reported human SBP2-like protein. Examination of the expression pattern revealed that the human SBP2 protein is encoded by a 4 kb long mRNA that is over-expressed in testis. Compared to the rat SBP2 sequence, the human SBP2 protein displays two highly conserved domains with 92 and 95% amino acid identity, the latter one containing the RNA binding domain. The inter-domain section carries 55% sequence identity, the remainder of the SBP2 sequences showing about 65% identity, values lower than expected for two mammalian proteins. Interestingly, we could show that the binding of human SBP2 to the SECIS RNA is stimulated by the selenoprotein-specialized elongation translation factor mSelB/eEFsec.}, note = {0378-1119 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Blotting, Complementary/chemistry/genetics Female Gene Expression Human Male Molecular Sequence Data Peptide Elongation Factors/metabolism Protein Binding RNA/*metabolism RNA, DNA Sequence Homology, ERIANI, LESCURE, Messenger/genetics/metabolism RNA-Binding Proteins/genetics/*metabolism Sequence Alignment Sequence Analysis, Molecular DNA, Non-U.S. Gov't, Northern Cloning, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The non-Watson-Crick base pairs and their associated isostericity matrices}, author = {N B Leontis and J Stombaugh and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12177293}, isbn = {12177293}, year = {2002}, date = {2002-01-01}, journal = {Nucleic Acids Res}, volume = {30}, number = {16}, pages = {3497-3531}, abstract = {RNA molecules exhibit complex structures in which a large fraction of the bases engage in non-Watson-Crick base pairing, forming motifs that mediate long-range RNA-RNA interactions and create binding sites for proteins and small molecule ligands. The rapidly growing number of three-dimensional RNA structures at atomic resolution requires that databases contain the annotation of such base pairs. An unambiguous and descriptive nomenclature was proposed recently in which RNA base pairs were classified by the base edges participating in the interaction (Watson-Crick, Hoogsteen/CH or sugar edge) and the orientation of the glycosidic bonds relative to the hydrogen bonds (cis or trans). Twelve basic geometric families were identified and all 12 have been observed in crystal structures. For each base pairing family, we present here the 4 x 4 'isostericity matrices' summarizing the geometric relationships between the 16 pairwise combinations of the four standard bases, A, C, G and U. Whenever available, a representative example of each observed base pair from X-ray crystal structures (3.0 A resolution or better) is provided or, otherwise, theoretically plausible models. This format makes apparent the recurrent geometric patterns that are observed and helps identify isosteric pairs that co-vary or interchange in sequences of homologous molecules while maintaining conserved three-dimensional motifs.}, note = {1362-4962 Journal Article}, keywords = {Algorithms *Base Pairing Base Sequence Hydrogen Bonding Models, Molecular RNA/*chemistry/classification/genetics Sequence Homology Support, Non-P.H.S. Support, P.H.S. Terminology, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Motif prediction in ribosomal RNAs Lessons and prospects for automated motif prediction in homologous RNA molecules}, author = {N B Leontis and J Stombaugh and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12458088}, isbn = {12458088}, year = {2002}, date = {2002-01-01}, journal = {Biochimie}, volume = {84}, number = {9}, pages = {961-973}, abstract = {The traditional way to infer RNA secondary structure involves an iterative process of alignment and evaluation of covariation statistics between all positions possibly involved in basepairing. Watson-Crick basepairs typically show covariations that score well when examples of two or more possible basepairs occur. This is not necessarily the case for non-Watson-Crick basepairing geometries. For example, for sheared (trans Hoogsteen/Sugar edge) pairs, one base is highly conserved (always A or mostly A with some C or U), while the other can vary (G or A and sometimes C and U as well). RNA motifs consist of ordered, stacked arrays of non-Watson-Crick basepairs that in the secondary structure representation form hairpin or internal loops, multi-stem junctions, and even pseudoknots. Although RNA motifs occur recurrently and contribute in a modular fashion to RNA architecture, it is usually not apparent which bases interact and whether it is by edge-to-edge H-bonding or solely by stacking interactions. Using a modular sequence-analysis approach, recurrent motifs related to the sarcin-ricin loop of 23S RNA and to loop E from 5S RNA were predicted in universally conserved regions of the large ribosomal RNAs (16S- and 23S-like) before the publication of high-resolution, atomic-level structures of representative examples of 16S and 23S rRNA molecules in their native contexts. This provides the opportunity to evaluate the predictive power of motif-level sequence analysis, with the goal of automating the process for predicting RNA motifs in genomic sequences. The process of inferring structure from sequence by constructing accurate alignments is a circular one. The crucial link that allows a productive iteration of motif modeling and realignment is the comparison of the sequence variations for each putative pair with the corresponding isostericity matrix to determine which basepairs are consistent both with the sequence and the geometrical data.}, note = {0300-9084 Journal Article}, keywords = {Bacterial/*chemistry/genetics RNA, Base Pairing Base Sequence Catalytic Domain Conserved Sequence Databases, Factual Models, Molecular *Nucleic Acid Conformation RNA, Non-P.H.S. Support, P.H.S., Ribosomal/*chemistry/genetics Sequence Alignment Support, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The annotation of RNA motifs}, author = {N Leontis and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2448414}, doi = {10.1002%2Fcfg.213}, year = {2002}, date = {2002-01-01}, journal = {Comp Funct Genom}, volume = {3}, number = {6}, pages = {518-524}, abstract = {The recent deluge of new RNA structures, including complete atomic-resolution views of both subunits of the ribosome, has on the one hand literally overwhelmed our individual abilities to comprehend the diversity of RNA structure, and on the other hand presented us with new opportunities for comprehensive use of RNA sequences for comparative genetic, evolutionary and phylogenetic studies. Two concepts are key to understanding RNA structure: hierarchical organization of global structure and isostericity of local interactions. Global structure changes extremely slowly, as it relies on conserved long-range tertiary interactions. Tertiary RNA–RNA and quaternary RNA–protein interactions are mediated by RNA motifs, defined as recurrent and ordered arrays of non-Watson–Crick base-pairs. A single RNA motif comprises a family of sequences, all of which can fold into the same three-dimensional structure and can mediate the same interaction(s). The chemistry and geometry of base pairing constrain the evolution of motifs in such a way that random mutations that occur within motifs are accepted or rejected insofar as they can mediate a similar ordered array of interactions. The steps involved in the analysis and annotation of RNA motifs in 3D structures are: (a) decomposition of each motif into non-Watson–Crick base-pairs; (b) geometric classification of each basepair; (c) identification of isosteric substitutions for each basepair by comparison to isostericity matrices; (d) alignment of homologous sequences using the isostericity matrices to identify corresponding positions in the crystal structure; (e) acceptance or rejection of the null hypothesis that the motif is conserved.}, keywords = {Unité ARN, WESTHOF, WESTHOF RNA motif Annotation Non-Watson–Crick basepair Shallow-groove Sugar-edge Watson–Crick edge Hoogsteen edge Isostericity matrix}, pubstate = {published}, tppubtype = {article} } @article{, title = {Detailed analysis of RNA-protein interactions within the bacterial ribosomal protein L5/5S rRNA complex}, author = { A. Perederina and N. Nevskaya and O. Nikonov and A. Nikulin and P. Dumas and M. Yao and I. Tanaka and M. Garber and G. Gongadze and S. Nikonov}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {12}, pages = {1548-57}, abstract = {The crystal structure of ribosomal protein L5 from Thermus thermophilus complexed with a 34-nt fragment comprising helix III and loop C of Escherichia coli 5S rRNA has been determined at 2.5 A resolution. The protein specifically interacts with the bulged nucleotides at the top of loop C of 5S rRNA. The rRNA and protein contact surfaces are strongly stabilized by intramolecular interactions. Charged and polar atoms forming the network of conserved intermolecular hydrogen bonds are located in two narrow planar parallel layers belonging to the protein and rRNA, respectively. The regions, including these atoms conserved in Bacteria and Archaea, can be considered an RNA-protein recognition module. Comparison of the T. thermophilus L5 structure in the RNA-bound form with the isolated Bacillus stearothermophilus L5 structure shows that the RNA-recognition module on the protein surface does not undergo significant changes upon RNA binding. In the crystal of the complex, the protein interacts with another RNA molecule in the asymmetric unit through the beta-sheet concave surface. This protein/RNA interface simulates the interaction of L5 with 23S rRNA observed in the Haloarcula marismortui 50S ribosomal subunit.}, note = {1355-8382 Journal Article}, keywords = {5S/*chemistry/*metabolism, Acid, Amino, Bacterial, Base, Binding, Bonding, coli/genetics, Conformation, Data, Escherichia, Fragments/chemistry/metabolism, Gov't, Hydrogen, Models, Molecular, Non-U.S., Nucleic, Peptide, Protein, Proteins/*chemistry/*metabolism, Proteins/chemistry/metabolism, Ribosomal, RNA, Sequence, Sites, Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A survey of metazoan selenocysteine insertion sequences}, author = {A Lambert and A Lescure and D Gautheret}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12458087}, isbn = {12458087}, year = {2002}, date = {2002-01-01}, journal = {Biochimie}, volume = {84}, number = {9}, pages = {953-959}, abstract = {The computational detection of novel selenoproteins in genomic sequences is usually achieved through identification of SECIS, a conserved secondary structure element found in the 3' UTR of animal selenoprotein mRNAs. Previous studies have used "descriptors" specifying the number of base pairs and the conserved nucleotides in SECIS to identify this element. A major drawback of the "descriptor" approach is that the number of detections in current genomic or transcript databases largely exceeds the number of true selenoproteins. In this study, we use instead the ERPIN program to detect SECIS elements. ERPIN is based on a lod-score profile algorithm that uses a training-set of aligned RNA sequences as input. From an initial alignment of 44 animal SECIS sequences, we performed a series of iterative searches in which the training set was progressively enriched up to 117 confirmed SECIS elements, from a large collection of metazoan species. About 200 high-scoring candidates were also detected. We show that ERPIN scores for these candidates can be converted into expect values, thus enabling their statistical evaluation. The most interesting SECIS candidates are presented.}, note = {0300-9084 Journal Article}, keywords = {Animals Base Sequence *Conserved Sequence Databases, Genetic Human Molecular Sequence Data Nucleic Acid Conformation Proteins/*genetics Selenocysteine/*genetics/metabolism Sequence Alignment Sequence Homology, LESCURE, Nucleic Acid Software, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Evolutionarily different RNA motifs and RNA-protein complexes to achieve selenoprotein synthesis}, author = {A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12457564}, isbn = {12457564}, year = {2002}, date = {2002-01-01}, journal = {Biochimie}, volume = {84}, number = {8}, pages = {765-774}, abstract = {A wealth of RNAs or RNA motifs are instrumental in controlling a variety of post-transcriptional or post-translational regulations. In this regard, selenocysteine incorporation in response to a redefined UGA stop codon certainly constitutes an intriguing and fascinating process. Translation elongation factors specialized for selenocysteine are needed to decode UGA selenocysteine codons. Discrimination between UGA selenocysteine and UGA stop codons also necessitates selenoprotein mRNA hairpins, called SECIS, that are internal to the coding frame in eubacteria or contained in the 3' untranslated regions in archaea/eukaryotes. This dichotomy leads to SECIS RNAs with distinct sequences and structures that tether the specialized translation elongation factor in a direct or indirect fashion, depending on the location of the SECIS RNA. The scope of this review is to bring a sharper focus on the SECIS RNA structures and SECIS RNA-protein complexes involved. Obviously, the examples described here highlight once again the versatility in form and function of RNA.}, note = {0300-9084 Journal Article Review Review Literature}, keywords = {Base Sequence Codon Conserved Sequence Eubacterium Eukaryotic Cells Evolution, Molecular Models, Molecular Molecular Sequence Data Nucleic Acid Conformation Peptide Elongation Factors/metabolism Proteins/*biosynthesis/genetics RNA/chemistry/*genetics RNA-Binding Proteins/genetics/*metabolism Selenocysteine/genetics Structure-Activity Relationship, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genomic hypomethylation in patients with B-cell chronic lymphocytic leukemia}, author = {M Kowal and J Roslinski and A Dmoszynska and A Borzecki and H Borzecka and G Keith}, url = {http://pthit.pl/Acta_Haematologica_Polonica,Metylacja_DNA_Przewlekla_bialaczka_limfocytowa,117.html}, year = {2002}, date = {2002-01-01}, journal = {Acta Haematol Pol}, volume = {33}, number = {3}, pages = {323-330}, abstract = {During the last few years, increasing attention has been paid to the relationship between DNA methylation and cancer. Hypo- or hypermethylation of DNA may result in up or down regulation of genes involved in the pathogenesis of leukemias. In this study we have analyzed DNA methylation in peripheral blood and bone marrow B (CD19+) and T (CD3+) lymphocytes from untreated patients with B-cell chronic lymphocytic leukemia.}, keywords = {GIEGE Methylation DNA Chronic lymphocytic leukemia, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effects of pressure on the crystallization and the solubility of proteins in agarose gel}, author = {A Kadri and B Lorber and G Jenner and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S0022024802017074}, doi = {10.1016/S0022-0248(02)01707-4}, year = {2002}, date = {2002-01-01}, journal = {J Crystal Growth}, volume = {245}, number = {1-2}, pages = {109-120}, abstract = {Crystals of thaumatin and of lysozyme from turkey and hen egg white have been prepared in batch at 20°C under hydrostatic pressures in the 0.1–220 MPa range. The latter model protein served as a reference in this comparative study. Crystallization was performed in an agarose gel in contrast to former studies under pressure that were conducted in solution. After depressurization, the habit, number, length, shape and solubility of crystals were compared to those of control crystals that were prepared at atmospheric pressure (0.1 MPa). For the three proteins, the number of the crystals increases with pressure. For thaumatin and hen lysozyme, crystal length decreases but for turkey lysozyme it increases. While the solubility of the first protein decreases, that of both lysozymes increases. The relationship between solubility and pressure is linear in the three cases. The crystallization volumes ΔV are −11 cm3 mol−1 for thaumatin, +15 cm3 mol−1 for turkey lysozyme and +3 cm3 mol−1 for hen lysozyme. For the lysozymes, they are explained by pressure-dependent conformational changes. The structure of thaumatin appears to be less deformable under pressure.}, keywords = {GIEGE Biocrystallization Lysozyme Pressure Proteins Thaumatin Agarose gel, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Direct and indirect contributions of RNA secondary structure elements to the initiation of HIV-1 reverse transcription}, author = {V Goldschmidt and M Rigourd and C Ehresmann and S F Le Grice and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12194974}, isbn = {12194974}, year = {2002}, date = {2002-01-01}, journal = {J Biol Chem}, volume = {277}, number = {45}, pages = {43233-43242}, abstract = {Initiation of human immunodeficiency virus type 1 (HIV-1) reverse transcription requires specific recognition between the viral RNA (vRNA), tRNA(3)(Lys), which acts as primer, and reverse transcriptase (RT). The specificity of this ternary complex is mediated by intricate interactions between the HIV-1 RNA and tRNA(3)(Lys). Here, we compared the relative importance of the secondary structure elements of this complex in the initiation process. To this aim, we used the previously published three-dimensional model of the initiation complex to rationally introduce a series of deletions and substitutions in the vRNA. When necessary, we used chemical probing to check the structure of the tRNA(3)(Lys)-mutant vRNA complexes. For each of them, we measured the binding affinity of RT and the kinetics of initial extension of tRNA(3)(Lys) and of synthesis of the (-) strand strong stop DNA. Our results were overall in keeping with the three-dimensional model of the initiation complex. Surprisingly, we found that disruption of the intermolecular template-primer interactions, which are not directly recognized by RT, more severely affected reverse transcription than deletions or disruption of one of the intramolecular helices to which RT directly binds. Perturbations of the highly constrained junction between the intermolecular helix formed by the primer binding site and the 3' end of tRNA(3)(Lys) and the helix immediately upstream also had dramatic effects on the initiation of reverse transcription. Taken together, our results demonstrate the overwhelming importance of the overall three-dimensional structure of the initiation complex and identify structural elements that constitute promising targets for anti-initiation-specific drugs.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence DNA Primers DNA Replication HIV-1/*genetics HIV-1 Reverse Transcriptase/*metabolism Human Kinetics Polymerase Chain Reaction RNA, Genetic, Lys/genetics RNA, MARQUET, Non-U.S. Gov't Transcription, Transfer, Unité ARN, Viral/*chemistry/*genetics/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoacyl-tRNA synthetases: versatile players in the changing theater of translation}, author = {C Francklyn and J J Perona and J Putz and Y M Hou}, url = {http://www.ncbi.nlm.nih.gov/pubmed/12458790}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {11}, pages = {1363-1372}, abstract = {Aminoacyl-tRNA synthetases attach amino acids to the 3' termini of cognate tRNAs to establish the specificity of protein synthesis. A recent Asilomar conference (California, January 13-18, 2002) discussed new research into the structure-function relationship of these crucial enzymes, as well as a multitude of novel functions, including participation in amino acid biosynthesis, cell cycle control, RNA splicing, and export of tRNAs from nucleus to cytoplasm in eukaryotic cells. Together with the discovery of their role in the cellular synthesis of proteins to incorporate selenocysteine and pyrrolysine, these diverse functions of aminoacyl-tRNA synthetases underscore the flexibility and adaptability of these ancient enzymes and stimulate the development of new concepts and methods for expanding the genetic code.}, keywords = {GIEGE aminoacyl-tRNA synthetases genetic code, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The HIV-1 Nef protein enhances the affinity of reverse transcriptase for RNA in vitro}, author = {C Fournier and J C Cortay and C Carbonnelle and C Ehresmann and R Marquet and P Boulanger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12881637}, isbn = {12881637}, year = {2002}, date = {2002-01-01}, journal = {Virus Genes}, volume = {25}, number = {3}, pages = {255-269}, abstract = {Several viral proteins, including nucleocapsid protein, integrase, Vif, Tat, and Nef have been proposed to act as cofactors of HIV-1 reverse transcription. Using two viral RNA probes, one overlapping the primer-binding site (PBS) and the other representing the ribosomal frameshifting signal (FS) of HIV-1 RNA, we found that recombinant full-length Nef protein (NefLAI) increased the affinity of reverse transcriptase (RT) for RNA in vitro, and interacted directly with RT in protein co-precipitation assays. The effect on RT-RNA binding and the capacity of Nef to interact with RT was also observed with N-terminal deletion mutant NefDelta57 and NefSF2, although to a lesser level. NefDelta57 corresponded to the processed Nef protein present in the internal core of mature virions, and lacked the N-myristoylated N-terminus and N-terminal region implicated in virus infectivity and pathogenicity in vivo. NefSF2, a Nef allele from a highly pathogenic strain of HIV-1, differed from NefLAI by the amino acid sequence and immunoreactivity of its N-terminal domain. The effect observed with NefSF2 and NefDelta57, and data from phage biopanning experiments suggested that the RT-binding region in Nef involved the C-terminal flexible loop of its C-terminal domain, but the function in RT-RNA binding was also influenced by its N-terminal domain.}, note = {0920-8569 Journal Article}, keywords = {Amino Acid Sequence Binding Sites Electrophoretic Mobility Shift Assay Gene Products, MARQUET, nef/*metabolism HIV-1/*genetics/metabolism Human In Vitro Molecular Sequence Data Protein Footprinting RNA/genetics/*metabolism RNA-Directed DNA Polymerase/*metabolism Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular investigations on tRNAs involved in human mitochondrial disorders}, author = {C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12418552}, isbn = {12418552}, year = {2002}, date = {2002-01-01}, journal = {Biosci Rep}, volume = {22}, number = {1}, pages = {81-98}, abstract = {Over the last decade, human neurodegenerative disorders which correlate with point mutations in mitochondrial tRNA genes became more and more numerous. Both the number of mutations (more than 70) and the variety of phenotypes (cardiopathies, myopathies, encephalopathies as well as diabetes, deafness or others) render the understanding of the genotype/phenotype relationships very complex. Here we first summarize the efforts undertaken to decipher the initial impact of various mutations on the structure/function relationships of tRNAs. This includes several lines of research, namely (i) investigation of human mitochrondrial tRNA structures, (ii) comparison of disease-related and polymorphic mutations at a theoretical level, and (iii) experimental investigations of affected tRNAs in the frame of mitochondrial protein synthesis. A new approach aimed at searching for long-range effects of mitochondrial tRNA mutations on a broader global mitochondrial level will also be presented. Initial results obtained by comparative mitochondrial proteomics turn out to be very promising for deciphering unexpected molecular partners involved in the pathological status of the mitochondria.}, note = {0144-8463 Journal Article Review Review Literature}, keywords = {FLORENTZ, Human Mitochondrial Diseases/*genetics/*physiopathology Mutation Nucleic Acid Conformation RNA, Non-U.S. Gov't, Transfer/*chemistry/*genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ribozymes: the first 20 years}, author = {M J Fedor and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12419214}, isbn = {12419214}, year = {2002}, date = {2002-01-01}, journal = {Mol Cell}, volume = {10}, number = {4}, pages = {703-704}, abstract = {Twenty years have passed since the first reports that certain RNAs mediate self-splicing and precursor tRNA processing reactions in the absence of proteins. An entire field emerged to learn how RNAs that lack the chemical versatility of amino acids nonetheless assemble into enzymes that accelerate chemical reactions with efficiencies that rival those of their protein counterparts.}, note = {1097-2765 Congresses}, keywords = {Catalysis Introns/genetics RNA Precursors/genetics/metabolism RNA Splicing RNA, Catalytic/chemistry/genetics/*metabolism, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {X-ray-induced debromination of nucleic acids at the Br K absorption edge and implications for MAD phasing}, author = {E Ennifar and P Carpentier and J L Ferrer and P Walter and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12136136}, isbn = {12136136}, year = {2002}, date = {2002-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {58}, number = {Pt 8}, pages = {1262-1268}, abstract = {Multi-wavelength anomalous dispersion (MAD) using brominated derivatives is considered a common and convenient technique for solving chemically synthesized nucleic acid structures. Here, it is shown that a relatively moderate X-ray dose (of the order of 5 x 10(15) photons mm(-2)) can induce sufficient debromination to prevent structure determination. The decrease in bromine occupancy with radiation dose can be accounted for by a simple exponential, with an estimated rate constant at the absorption-peak wavelength, 7.4 (0.8) MGy, that is not significantly different from its value at the absorption-edge wavelength, 9.2 (2.6) MGy (the given e.s.d.s assess the relative closeness of the two values, not their absolute accuracy, which is probably worse). Chemically, these results (and others) are consistent with bromine cleavage resulting from direct photodissociation and/or from the action of free electrons, rather than from the action of hydroxyl radicals originating from water dissociation. The free bromine species (Br(-)) diffuse too quickly, even in amorphous ice around 100 K, to allow the determination of a diffusion coefficient. From a practical point of view, it is suggested that a single data collection with a crystal consisting of iodinated instead of brominated derivatives could provide both anomalous scattering and SIR phase information by the progressive cleavage of iodine.}, note = {0907-4449 Journal Article}, keywords = {Bromine/chemistry/radiation effects Crystallography, ENNIFAR, Non-U.S. Gov't X-Rays, Unité ARN, Viral/chemistry Support, X-Ray HIV-1/chemistry Molecular Structure Nucleic Acids/*chemistry/radiation effects RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {A 5'-3' long-range interaction in Ty1 RNA controls its reverse transcription and retrotransposition}, author = {G Cristofari and C Bampi and M Wilhelm and F X Wilhelm and J L Darlix}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12169639}, isbn = {12169639}, year = {2002}, date = {2002-01-01}, journal = {EMBO J}, volume = {21}, number = {16}, pages = {4368-4379}, abstract = {LTR-retrotransposons are abundant components of all eukaryotic genomes and appear to be key players in their evolution. They share with retroviruses a reverse transcription step during their replication cycle. To better understand the replication of retrotransposons as well as their similarities to and differences from retroviruses, we set up an in vitro model system to examine minus-strand cDNA synthesis of the yeast Ty1 LTR-retrotransposon. Results show that the 5' and 3' ends of Ty1 genomic RNA interact through 14 nucleotide 5'-3' complementary sequences (CYC sequences). This 5'-3' base pairing results in an efficient initiation of reverse transcription in vitro. Transposition of a marked Ty1 element and Ty1 cDNA synthesis in yeast rely on the ability of the CYC sequences to base pair. This 5'-3' interaction is also supported by phylogenic analysis of all full-length Ty1 and Ty2 elements present in the Saccharomyces cerevisiae genome. These novel findings lead us to propose that circularization of the Ty1 genomic RNA controls initiation of reverse transcription and may limit reverse transcription of defective retroelements.}, note = {0261-4189 Journal Article}, keywords = {Complementary/biosynthesis *Gene Expression Regulation, DNA, Fungal In Vitro Nucleic Acid Conformation Phylogeny RNA, Fungal/chemistry/*metabolism RNA, Genetic, Messenger/chemistry/*metabolism Retroelements/*genetics Saccharomyces cerevisiae/*genetics Support, Non-U.S. Gov't *Transcription, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {X-ray diffraction properties of protein crystals prepared in agarose gel under hydrostatic pressure.}, author = {C Charron and M C Robert and B Capelle and A Kadri and G Jenner and R Giege and B Lorber}, url = {http://www.sciencedirect.com/science/article/pii/S0022024802017360}, doi = {10.1016/S0022-0248(02)01736-0}, year = {2002}, date = {2002-01-01}, journal = {J Crystal Growth}, volume = {245}, number = {3-4}, pages = {321-333}, abstract = {Crystals of thaumatin and of turkey and hen lysozyme, prepared in a batch at 293K under hydrostatic pressures in the 0.1–150MPa range, have been analyzed to investigate how this parameter influences crystallization and if depressurization introduces defects in the crystalline lattice. The X-ray diffraction properties of depressurized crystals have been compared with those of unpressurized control crystals prepared under otherwise identical conditions at atmospheric pressure (0.1 MPa). Independently of pressure, the crystals of each protein belongto the same space group and have cell parameters identical to those of the controls. Their diffraction limit is more dependent upon crystal volume than upon pressure. Crystal mosaicity, expressed as the full-width at half-maximum w of the Bragg reflection profile, was used to quantify the defects in the lattice. While the quality of lysozyme crystals deteriorates as pressure increases, that of tetragonal thaumatin crystals becomes more homogeneous. The first result correlates with the apparition of cleavages oriented perpendicularily to the crystalメs c-axis. The second one is interpreted by a gradual selection of well-formed nuclei. The 3D structure of thaumatin in a crystal that was grown under a pressure of 150MPa and returned to atmospheric pressure was solved. The fold of the polypeptide chain and the distribution of water molecules was compared with those in a reference crystal grown at 0.1 MPa.The results suggest that the crystal lattice is elastic. A possible application of pressure to the preparation of high quality protein crystals is discussed in the light of these results.}, keywords = {GIEGE Crystallization Pressure Agarose gel Lysozyme Proteins Thaumatin, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal contacts engineering of aspartyl-tRNA synthetase from Thermus thermophilus: effects on crystallizability}, author = {C Charron and D Kern and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12351895}, isbn = {12351895}, year = {2002}, date = {2002-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {58}, number = {Pt 10 Pt 1}, pages = {1729-1733}, abstract = {To understand how surface residues in a protein structure influence crystal growth, packing arrangement and crystal quality, crystal surfaces were modified and crystallizability of seven different mutants investigated. The model was aspartyl-tRNA synthetase-1 from Thermus thermophilus, a homodimer (M(r) 122000) with a subunit of 580 amino acids. Engineering concerned modification of amino acids involved in packing contacts in the orthorhombic lattice (P2(1)2121) of the synthetase. Comparison of the crystallization behaviour of the mutants indicates a correlation between disruption/addition of packing interactions and crystallizability of the mutants: disruption or modification of lattice contacts prevents crystallization or leads to crystals of poor quality. In contrast, addition of potential contacts leads to well-shaped crystals of same space group and cell parameters than wild-type crystals.}, note = {0907-4449 Journal Article}, keywords = {Aspartate-tRNA Ligase/*chemistry/*genetics Base Sequence Crystallization/*methods Crystallography, Bacterial/genetics Hydrogen Bonding Macromolecular Systems Models, Molecular Molecular Structure Mutagenesis, Non-U.S. Gov't Surface Properties Thermus thermophilus/*enzymology/*genetics, Site-Directed Protein Engineering Recombinant Proteins/chemistry/genetics Support, Unité ARN, X-Ray DNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization in the presence of glycerol displaces water molecules in the structure of thaumatin}, author = {C Charron and A Kadri and M C Robert and R Giege and B Lorber}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12454465}, isbn = {12454465}, year = {2002}, date = {2002-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {58}, number = {Pt 12}, pages = {2060-2065}, abstract = {The intensely sweet protein thaumatin has been crystallized at 293 K in the presence of sodium tartrate and 25%(v/v) glycerol for X-ray diffraction data collection at 100 K. A comparison of the three-dimensional structure model derived from a crystal grown in the presence of glycerol with that of a control deprived of this additive reveals only minor changes in the overall structure but a approximately 20% reduction in the number of water molecules. X-ray topography analyses show that the overall quality of the crystals prepared in the presence of this cryoprotectant is enhanced.}, note = {0907-4449 Journal Article}, keywords = {Crystallization Crystallography, Molecular Plant Proteins/*chemistry Protein Conformation Support, Non-U.S. Gov't Sweetening Agents/*chemistry Water/*chemistry, Unité ARN, X-Ray Glycerol/*chemistry Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA loop-loop interactions as dynamic functional motifs}, author = {C Brunel and R Marquet and P Romby and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12458085}, isbn = {12458085}, year = {2002}, date = {2002-01-01}, journal = {Biochimie}, volume = {84}, number = {9}, pages = {925-944}, abstract = {RNA loop-loop interactions are frequently used to trigger initial recognition between two RNA molecules. In this review, we present selected well-documented cases that illustrate the diversity of biological processes using RNA loop-loop recognition properties. The first one is related to natural antisense RNAs that play a variety of regulatory functions in bacteria and their extra-chromosomal elements. The second one concerns the dimerization of HIV-1 genomic RNA, which is responsible for the encapsidation of a diploid RNA genome. The third one concerns RNA interactions involving double-loop interactions. These are used by the bicoid mRNA to form dimers, a property that appears to be important for mRNA localization in drosophila embryo, and by bacteriophage phi29 pRNA which forms hexamers that participate in the translocation of the DNA genome through the portal vertex of the capsid. Despite the high diversity of systems and mechanisms, some common features can be highlighted. (1) Efficient recognition requires rapid bi-molecular binding rates, regardless of the RNA pairing scheme. (2) The initial recognition is favored by particular conformations of the loops enabling a proper presentation of nucleotides (generally a restricted number) that initiate the recognition process. (3) The fate of the initial reversible loop-loop complex is dictated by both functional and structural constraints. RNA structures have evolved either to "freeze" the initial complex, or to convert it into a more stable one, which involves propagation of intermolecular interactions along topologically feasible pathways. Stabilization of the initial complex may also be assisted by proteins and/or formation of additional contacts.}, note = {0300-9084 Journal Article Review Review Literature}, keywords = {Animals Base Pairing Base Sequence Dimerization HIV-1/genetics Human Kinetics Molecular Sequence Data *Nucleic Acid Conformation RNA/genetics/*metabolism Support, MARQUET, Non-U.S. Gov't Thermodynamics, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural and functional properties of the HIV-1 RNA-tRNA(Lys)3 primer complex annealed by the nucleocapsid protein: comparison with the heat-annealed complex}, author = {F Brule and R Marquet and L Rong and M A Wainberg and B P Roques and S F Le Grice and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11873759}, isbn = {11873759}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {1}, pages = {8-15}, abstract = {The conversion of the single-stranded RNA genome into double-stranded DNA by virus-coded reverse transcriptase (RT) is an essential step of the retrovirus life cycle. In human immunodeficiency virus type 1 (HIV-1), RT uses the cellular tRNA(Lys)3 to initiate the (-) strand DNA synthesis. Placement of the primer tRNA(Lys)3 involves binding of its 3'-terminal 18 nt to a complementary region of genomic RNA termed PBS. However, the PBS sequence is not the unique determinant of primer usage and additional contacts are important. This placement is believed to be achieved in vivo by the nucleocapsid domain of Gag or by the mature protein NCp. Up to now, structural information essentially arose from heat-annealed primer-template complexes (Isel et al., J Mol Biol, 1995, 247:236-250; Isel et al., EMBO J, 1999, 18:1038-1048). Here, we investigated the formation of the primer-template complex mediated by NCp and compared structural and functional properties of heat- and NCp-annealed complexes. We showed that both heat- and NCp-mediated procedures allow comparable high yields of annealing. Then, we investigated structural features of both kinds of complexes by enzymatic probing, and we compared their relative efficiency in (-) strong stop DNA synthesis. We did not find any significant differences between these complexes, suggesting that information derived from the heat-annealed complex can be transposed to the NCp-mediated complex and most likely to complexes formed in vivo.}, note = {1355-8382 Journal Article}, keywords = {Genetic, Genetic Transcription, Lys/*chemistry/genetics/metabolism RNA, MARQUET, Non-U.S. Gov't Templates, Transfer, Unité ARN, Viral/*chemistry/genetics/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization of biological macromolecules using agarose gel.}, author = {C Biertümpfel and J Basquin and D Suck and C Sauter}, url = {http://scripts.iucr.org/cgi-bin/paper?ic0008}, isbn = {10.1107/S0907444902012738}, year = {2002}, date = {2002-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {58}, number = {10}, pages = {1657-1659}, abstract = {Gellified media prevent convection and crystal sedimentation, and provide an attractive growth environment for optimising biological crystals. Agarose gels are particularly easy to use and they are compatible with most of the common crystallization methods. They also offer new possibilities like counter-diffusion techniques. This paper gives a brief overview of their general properties and presents an application of a counter-diffusion setup combining agarose gel and capillaries to the crystallization of proteins and protein / nucleic acid complexes.}, keywords = {Agarose gel Crystallization Biological macromolecule Counter-diffusion., FRUGIER, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Melting of the solvent structure around a RNA duplex: a molecular dynamics simulation study}, author = {P Auffinger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12062380}, isbn = {12062380}, year = {2002}, date = {2002-01-01}, journal = {Biophys Chem}, volume = {95}, number = {3}, pages = {203-210}, abstract = {From three 2.4-ns molecular dynamics simulations of the r(CpG)(12) duplex conducted at 5, 25 and 37 degrees C, a strong temperature dependence of the dynamics of the water molecules and ions located in the first nucleic acid coordination shell is observed. At 5 degrees C, the highest residence times of bound water molecules exceed 1 ns while, at 37 degrees C, they decrease to 0.5 ns in agreement with available NMR data. Similar temperature dependencies are observed for the potassium ions bound to the duplex. In this temperature range, the structure of the RNA helix remains essentially unchanged. Thus, the observed alterations correspond to a 'premelting' of the solvent structure around the duplex. It is proposed that, before the nucleic acid structure melts, the entropy of the solvent increases to a point where it is no longer compensated by the enthalpic contribution of solute-solute and solute-solvent interactions. At this stage, the weakest structural elements start to melt. In other terms, the experimentally observed melting processes are preceded by a melting of the more labile solvent structure.}, note = {0301-4622 Journal Article}, keywords = {Computer Simulation Entropy Models, Double-Stranded/*chemistry Solvents/chemistry/metabolism Temperature Thermodynamics Water/*chemistry/metabolism, Molecular Molecular Structure Motion Nucleic Acid Conformation RNA, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Strucutral and dynamical characterization of nucleic acids water and ion binding sites.}, author = {P Auffinger and B Masquida and E Westhof}, editor = {T Schlick and H H Gan}, url = {http://books.google.fr/books?id=DLDdglVVhVIC&pg=PA61&lpg=PA61&dq=Structural+and+dynamical+characterization+of+nucleic+acids+water+and+ion+binding+sites.&source=bl&ots=hmiZnqX_lA&sig=PbZPEKPo3NitnY9sMj0S0TAHS58&hl=fr&sa=X&ei=e4T9T-GlMsenhAfJqrCDAg&ved=0CFQQ6AEwAA#v=onepage&q=Structural%20and%20dynamical%20characterization%20of%20nucleic%20acids%20water%20and%20ion%20binding%20sites.&f=false}, year = {2002}, date = {2002-01-01}, booktitle = {Computational methods for macromolecules: challenges and applications. Proceedings of the 3rd international workshop on algorithms for macromolecular modeling, New York, October 12-14, 2000. Lecture Notes in Computational Science and Engineering.}, volume = {24}, pages = {61-70}, publisher = {Springer}, abstract = {Recent methodological developments led to longer and more accurate molecular dynamics (MD) simulations. In parallel, methods have been designed with the purpose of characterizing water and ion binding features. Here, we give an outline of some of the methods we used in order to extract structural and dynamical information concerning the first water and ion coordination shell, from MD simulations conducted on RNA and DNA structures. Coordinates for the water and ion binding sites located in the first coordination shell of r(G=C), d(G=C), r(AU), and d(A-T) base-pairs are provided, along with calculated モpseudoヤ thermal factors.}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {The SBP2 and 15.5 kD/Snu13p proteins share the same RNA binding domain: identification of SBP2 amino acids important to SECIS RNA binding}, author = {C Allmang and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12403468}, isbn = {12403468}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {10}, pages = {1308-1318}, abstract = {Selenoprotein synthesis in eukaryotes requires the selenocysteine insertion sequence (SECIS) RNA, a hairpin in the 3' untranslated region of selenoprotein mRNAs. The SECIS RNA is recognized by the SECIS-binding protein 2 (SBP2), which is a key player in this specialized translation machinery. The objective of this work was to obtain structural insight into the SBP2-SECIS RNA complex. Multiple sequence alignment revealed that SBP2 and the U4 snRNA-binding protein 15.5 kD/Snu13p share the same RNA binding domain of the L7A/L30 family, also found in the box H/ACA snoRNP protein Nhp2p and several ribosomal proteins. In corollary, we have detected a similar secondary structure motif in the SECIS and U4 RNAs. Combining the data of the crystal structure of the 15.5 kD-U4 snRNA complex, and the SBP2/15.5 kD sequence similarities, we designed a structure-guided strategy predicting 12 SBP2 amino acids that should be critical for SECIS RNA binding. Alanine substitution of these amino acids followed by gel shift assays of the SBP2 mutant proteins identified four residues whose mutation severely diminished or abolished SECIS RNA binding, the other eight provoking intermediate down effects. In addition to identifying key amino acids for SECIS recognition by SBP2, our findings led to the proposal that some of the recognition principles governing the 15.5 kD-U4 snRNA interaction must be similar in the SBP2-SECIS RNA complex.}, note = {1355-8382 Journal Article}, keywords = {Amino Acid Spliceosomes/metabolism Structural Homology, Amino Acid Sequence Amino Acid Substitution Binding Sites Human Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, ERIANI, Non-U.S. Gov't, Protein Support, Small Nuclear/chemistry/*metabolism RNA-Binding Proteins/genetics/*metabolism Ribonucleoproteins, Small Nuclear/genetics/*metabolism Selenocysteine/metabolism Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genomic hypomethylation in patients with B-cell chronic lymphocytic leukemia}, author = {M Kowal and J Roslinski and A Dmoszynska and A Borzecki and H Borzecka and G Keith}, editor = {Editor}, url = {http://pthit.pl/Acta_Haematologica_Polonica,Metylacja_DNA_Przewlekla_bialaczka_limfocytowa,117.html}, year = {2002}, date = {2002-01-01}, journal = {Acta Haematol Pol}, volume = {33}, number = {3}, pages = {323-330}, abstract = {During the last few years, increasing attention has been paid to the relationship between DNA methylation and cancer. Hypo- or hypermethylation of DNA may result in up or down regulation of genes involved in the pathogenesis of leukemias. In this study we have analyzed DNA methylation in peripheral blood and bone marrow B (CD19+) and T (CD3+) lymphocytes from untreated patients with B-cell chronic lymphocytic leukemia.}, keywords = {GIEGE Methylation DNA Chronic lymphocytic leukemia}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Estimation of DNA methylation level in endometrial cancer tissues]}, author = { A. Popiela and M. S. Gabrys and J. Rabczynski and M. Panszczyk and G. Keith and W. Baranowski}, year = {2002}, date = {2002-01-01}, journal = {Ginekol Pol}, volume = {73}, number = {11}, pages = {966-9}, abstract = {OBJECTIVES: A level of DNA methylation plays an important role in regulation of cellular gene's expression. Estimation of DNA methylation level in endometrial neoplastic tissues compared to normal endometrial samples was the aim of this study. DESIGN: It was to be shown, that changes in methylation rate in promotory regions could lead to carcinogenesis in particular cell. Authors describe an analysis of DNA methylation level in endometrial cancer tissues compared to DNA methylation level in normal or hyperplastic endometrium. MATERIAL AND METHODS: Endometrial samples from 88 women were collected. 56 of them were classified as adenocarcinoma, 20 as hyperplastic changes, 12 as normal endometrium-control group. DNA was isolated from tissues and than prepared to pm5dC and pdC. Than we performed a statistical analysis of results. RESULTS: The median DNA methylation level was significantly higher in neoplastic tissues than in normal endometrium. There was no difference between DNA methylation level between normal endometrium and hyperplastic changes. CONCLUSIONS: Authors conclude that neoplastic endometrial tissues show high DNA methylation rate compared to normal or hyperplastic endometrium.}, note = {0017-0011 Journal Article}, keywords = {*DNA, 80, Abstract, Adenocarcinoma/chemistry/*genetics, Aged, and, Case-Control, Endometrial, English, Expression, Female, Gene, Human, Hyperplasia/genetics, Methylation, Middle, Neoplasms/chemistry/*genetics, Neoplastic, over, Regulation, Studies}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Estimation of DNA methylation level in nonendometrial uterus malignancies]}, author = { A. Popiela and M. S. Gabrys and J. Rabczynski and M. Panszczyk and G. Keith and W. Baranowski}, year = {2002}, date = {2002-01-01}, journal = {Ginekol Pol}, volume = {73}, number = {11}, pages = {962-5}, abstract = {OBJECTIVES: Rebuilding of genome structure leads to many pathological states including neoplastic malignancies. Rebuilding often occurs as a process caused by disturbances in gene silencing mechanism. DNA methylation pattern is one of the most important mechanisms connected to gene's silencing. Estimation of DNA methylation level in nonendometrial uterine neoplastic tissues compared to normal endometrial samples was the aim of this study. DESIGN: It was to be shown, that changes in methylation rate in promotory regions could lead to carcinogenesis in particular cell. Authors describe an analysis of DNA methylation level in nonendometrial neoplastic uterine tissues compared to DNA methylation level in normal endometrium. MATERIALS AND METHODS: Tissue samples from 9 women with tumor mixtus mesodermalis were collected. 12 samples were normal endometrium-control group. DNA was isolated from tissues and than we performed an estimation of DNA methylation levels. Than we performed a statistical analysis of results. RESULTS: The median DNA methylation level was significantly higher in neoplastic tissues than in normal endometrium. CONCLUSIONS: Authors conclude, that DNA methylation level is higher in neoplastic tissues, but does not correlate with clinical stage of the disease.}, note = {0017-0011 Journal Article}, keywords = {*DNA, 80, Abstract, Age, Aged, and, Case-Control, English, Expression, Factors, Female, Gene, Human, Mesodermal/genetics/*metabolism, Methylation, Middle, Mixed, Neoplasms/genetics/*metabolism, Neoplastic, over, Regulation, silencing, Studies, tumor, Uterine}, pubstate = {published}, tppubtype = {article} } @article{furrer_dynamic_2002, title = {Dynamic and magnetic susceptibility effects on the MAS NMR linewidth of a tetrapeptide bound to different resins}, author = {Julien Furrer and Karim Elbayed and Maryse Bourdonneau and Jésus Raya and David Limal and Alberto Bianco and Martial Piotto}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/mrc.970}, doi = {10.1002/mrc.970}, issn = {1097-458X}, year = {2002}, date = {2002-01-01}, urldate = {2020-03-31}, journal = {Magnetic Resonance in Chemistry}, volume = {40}, number = {2}, pages = {123--132}, abstract = {Under magic angle spinning, the NMR spectrum of the tetrapeptide Ala-Ile-Gly-Met bound to a Wang resin, and swollen in DMF, exhibits proton and carbon linewidths that are sharp enough to allow the complete characterization of the peptide using classical liquid-state NMR methods. The proton linewidths of the bound peptide remain, however, about three times larger than those of the free peptide in solution. The residual NMR linewidth originates essentially from incompletely averaged magnetic susceptibility effects due to the Wang resin. Replacing the aromatic Wang resin with a PEGA or POEPOP resin removes this effect. To investigate the contribution to line broadening of the peptide dynamics, relaxation studies were performed on the peptide bound to Wang and POEPOP resins. Copyright © 2001 John Wiley & Sons, Ltd.}, keywords = {13C NMR, 1H NMR, high-resolution magic angle spinning, I2CT, magnetic susceptibility, NMR, relaxation, solid-phase peptide synthesis, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{cremer_long-lived_2002, title = {Long-lived immature dendritic cells mediated by TRANCE-RANK interaction}, author = {I Cremer and M C Dieu-Nosjean and S Mar�chal and C Dezutter-Dambuyant and S Goddard and D Adams and N Winter and C Menetrier-Caux and C Saut�s-Fridman and W H Fridman and C G F Mueller}, year = {2002}, date = {2002-01-01}, journal = {Blood}, volume = {100}, number = {10}, pages = {3646--3655}, abstract = {Immature dendritic cells (DCs) reside in Interstitial tissues (Int-DC) or in the epidermis, where they capture antigen and, thereafter, mature and migrate to draining lymph nodes (LNs), where they present processed antigen to T cells. We have Identified Int-DCs that express both TRANCE (tumor necrosis factor-related activation-induced cytokine) and RANK (receptor activator of NF-kappaB) and have generated these cells from CD34(+) human progenitor cells using macrophage colony-stimulating factor (M-CSF). These CD34(+)-derived Int-DCs, which are related to macrophages, are long-lived, but addition of soluble RANK leads to significant reduction of cell viability and BcI-2 expression. This suggests that constitutive TRANCE-RANK interaction is responsible for CD34(+)-derived Int-DC longevity. Conversely, CD1a(+) DCs express only RANK and are short-lived. However, they can be rescued from cell death either by recombinant soluble TRANCE or by CD34(+)-derived Int-DCs. CD34(+)-derived Int-DCs mature in response to lipopolysaccharide (LPS) plus CD40 ligand (L) and become capable of CCL21/CCL19-mediated chemotaxis and naive T-cell activation. Upon maturation, they lose TRANCE, making them, like CD1a(+) DCs, dependent on exogenous TRANCE for survival. These findings provide evidence that TRANCE and RANK play important roles in the homeostasis of DCs. (C) 2002 by The American Society of Hematology}, keywords = {Activation, Antigen, CD40, CD40 Ligand, CHEMOTAXIS, Cytokines, Dendritic Cells, Epidermis, Expression, Homeostasis, Human, IMMATURE, l, ligand, lipopolysaccharide, Longevity, LPS, LYMPH, LYMPH NODE, Lymph Nodes, M-CSF, Macrophage, Macrophages, Maturation, naive, Necrosis, NF-kappaB, PROGENITOR CELLS, rank, Receptor, survival, T CELL ACTIVATION, T CELLS, Team-Mueller, TRANCE, tumor, viability}, pubstate = {published}, tppubtype = {article} } @article{, title = {Lack of genotoxicity of bitumen fumes in transgenic mouse lung}, author = { J. C. Micillino and C. Coulais and S. Binet and M. C. Bottin and G. Keith and D. Moulin and B. H. Rihn}, year = {2002}, date = {2002-01-01}, journal = {Toxicology}, volume = {170}, number = {1-2}, pages = {11-20}, abstract = {During hot application of bitumen containing materials, e.g. in hot paving or roofing, fumes are emitted that contain polycyclic aromatic compounds. Previous studies with rodents exposed to bitumen and coal-tar fume condensates showed formation of DNA adducts. In order to clarify the genotoxicity of bitumen fumes, we designed a study by using mice carrying a reporter gene for mutagenesis analysis and exposed by nose-only to a constant and reproducible aerosol of bitumen fumes. We analyzed the genotoxic activity of inhaled bitumen fumes generated under those controlled conditions through the induction of mutation and DNA adducts in Big Blue mice. Mice were exposed to bitumen fumes (100 mg/m(3) total particulate matter) 6 h per day during 5 days by nose-only in an inhalation chamber designed in our laboratory. Following a 30-day fixation period, the experiment was terminated and lung DNA was extracted for mutant frequency and adduct determinations. The mutant frequency was determined using the cII and the lacI mutant analysis systems. In, addition, 61 and 54 mutants were sequenced in control and exposed groups, respectively. The study did not show any mutation or adduct induction in the exposed group compared to the control group: cII mutant frequencies were 11.0+/-4.5x10(-5) and 11.0+/-4.8x10(-5) in control and exposed lungs, respectively. Identically, using the lacI mutation detection system, the mutant frequencies were 6.4+/-3.1x10(-5) and 5.8+/-2.0x10(-5). The mutation spectra of both series were quite similar with regard to transition and transversion frequencies. The absence of genotoxicity in the group exposed to 100 mg/m(3) bitumen is discussed with regard to dosage of inhaled polycyclic aromatic compounds and species.}, note = {0300-483x Journal Article}, keywords = {Adducts/drug, Aerosols, Animals, C57BL, Chromatography, DNA, DNA/drug, effects, effects/metabolism, Gases/*toxicity, Genes, Hydrocarbons/*toxicity, Inbred, Lac, Layer, Lung/*drug, Mice, Mutagenicity, Mutagens/*toxicity, Mutation/drug, Operon/genetics, Reporter/genetics, Tests, Thin, transgenic}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoacyl-tRNA synthetases: versatile players in the changing theater of translation}, author = { C. Francklyn and J.J. Perona and J. Putz and Y. M. Hou}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {11}, pages = {1363-1372}, abstract = {Aminoacyl-tRNA synthetases attach amino acids to the 3' termini of cognate tRNAs to establish the specificity of protein synthesis. A recent Asilomar conference (California, January 13-18, 2002) discussed new research into the structure-function relationship of these crucial enzymes, as well as a multitude of novel functions, including participation in amino acid biosynthesis, cell cycle control, RNA splicing, and export of tRNAs from nucleus to cytoplasm in eukaryotic cells. Together with the discovery of their role in the cellular synthesis of proteins to incorporate selenocysteine and pyrrolysine, these diverse functions of aminoacyl-tRNA synthetases underscore the flexibility and adaptability of these ancient enzymes and stimulate the development of new concepts and methods for expanding the genetic code.}, keywords = {Aminoacyl-tRNA, Code, Genetic, GIEGE, synthetases}, pubstate = {published}, tppubtype = {article} } @article{, title = {Microarrays as cancer keys: an array of possibilities}, author = { S. Mohr and G. D. Leikauf and G. Keith and B. H. Rihn}, year = {2002}, date = {2002-01-01}, journal = {J Clin Oncol}, volume = {20}, number = {14}, pages = {3165-75}, abstract = {Malignant transformation results from accumulation of genetic and epigenetic events. Functional studies of cancer will be crucial to our understanding of its complexity and polymorphism. There is no doubt that emerging genomic and proteomic technologies will facilitate such investigations. Microarray technology is a new and efficient approach to extract data of biomedical relevance for a wide range of applications. In cancer research, it will provide high-throughput and valuable insights into differences in an individual's tumor as compared with constitutional DNA, mRNA expression, and protein expression and activity. Across individuals, comparisons could provide tissue-specific disease signatures that provide diagnosis based on hundreds of informative genes. The resulting product should be a wealth of tumor-associated and tumor-specific biomarkers, which may help in cancer etiology, diagnosis, and therapy and ultimately lead to "molecular nosology" of cancers. This review highlights the recent developments in microarray technologies in cancer research, focuses on the results obtained so far, and describes the eventual use of microarray technology for clinical applications.}, note = {0732-183x Journal Article Review Review, Tutorial}, keywords = {(Genetics), *Gene, *Oligonucleotide, *Sequence, Aberrations, Analysis, Analysis/methods, Animals, Array, Chromosome, DNA/methods, Expression, Genotype, Gov't, Human, Mutation, Neoplasms/*genetics, Neoplastic, Oncogenes/*genetics, P.H.S., Polymorphism, Profiling/methods, Proteome/genetics, Regulation, Sequence, Support, U.S.}, pubstate = {published}, tppubtype = {article} } @article{, title = {Susceptibility of recombinant porcine endogenous retrovirus reverse transcriptase to nucleoside and non-nucleoside inhibitors}, author = { M. Wilhelm and J. A. Fishman and R. Pontikis and A. M. Aubertin and F. X. Wilhelm}, year = {2002}, date = {2002-01-01}, journal = {Cell Mol Life Sci}, volume = {59}, number = {12}, pages = {2184-90}, abstract = {Transplantation of organs, tissues or cells from pigs to humans could be a potential solution to the shortage of human organs for transplantation. Porcine endogenous retroviruses (PERVs) remain a major safety concern for porcine xenotransplantation. Thus, finding drugs that could be used as virological prophylaxis (or therapy) against PERV replication would be desirable. One of the most effective ways to block retroviral multiplication is to inhibit the enzyme reverse transcriptase (RT) which catalyzes the reverse transcription of viral RNA to proviral double-stranded DNA. We report here the cloning and expression of PERV RT and its susceptibility to several inhibitors. Our data demonstrate PERV susceptibility in vitro to the triphosphorylated nucleoside analog of zidovudine (AZT) and to ddGTP and to a lesser extent to ddTTP but almost no susceptibility to the non-nucleoside RT inhibitors tested.}, note = {1420-682x Journal Article}, keywords = {Acid, Amino, Animals, Calf, Chloride/metabolism, Chlorides/metabolism, Cloning, Compounds/metabolism, Data, DNA, DNA-Directed, endogenous, Gov't, H, Human, Inhibitors/*pharmacology, Magnesium, Manganese, Molecular, Non-U.S., Nucleosides/chemistry/*metabolism, P.H.S., Polymerase/chemistry/genetics/*metabolism, Polymerase/metabolism, Proteins/metabolism, Recombinant, Retroviruses/*enzymology, Reverse, Ribonuclease, RNA-Directed, Sequence, Sodium, structure, Support, Swine, Thymus/metabolism, Transcriptase, U.S.}, pubstate = {published}, tppubtype = {article} } @article{ligoxygakis_critical_2002, title = {Critical evaluation of the role of the Toll-like receptor 18-Wheeler in the host defense of Drosophila}, author = {Petros Ligoxygakis and Philippe Bulet and Jean-Marc Reichhart}, doi = {10.1093/embo-reports/kvf130}, issn = {1469-221X}, year = {2002}, date = {2002-01-01}, journal = {EMBO Rep.}, volume = {3}, number = {7}, pages = {666--673}, abstract = {Essential aspects of innate immune responses to microbial infections appear to be conserved between insects and mammals. In particular, in both groups, transmembrane receptors of the Toll superfamily play a crucial role in activating immune defenses. The Drosophila Toll family member 18-Wheeler had been proposed to sense Gram-negative infection and direct selective expression of peptides active against Gram-negative bacteria. Here we re-examine the role of 18-Wheeler and show that in adults it is dispensable for immune responses. In larvae, 18wheeler is required for normal fat body development, and in mutant larvae induction of all antimicrobial peptide genes, and not only of those directed against Gram-negative bacteria, is compromised. 18-Wheeler does not qualify as a pattern recognition receptor of Gram-negative bacteria.}, keywords = {Animals, Anti-Bacterial Agents, Antimicrobial Cationic Peptides, Cell Adhesion Molecules, Fat Body, Gene Expression Regulation, Genes, Immunohistochemistry, Immunologic, Insect, Insect Proteins, Larva, M3i, Mass, Matrix-Assisted Laser Desorption-Ionization, Membrane Proteins, Receptors, reichhart, Reporter, Spectrometry, Transgenes}, pubstate = {published}, tppubtype = {article} } @article{reichhart_splice-activated_2002, title = {Splice-activated UAS hairpin vector gives complete RNAi knockout of single or double target transcripts in Drosophila melanogaster}, author = {Jean-Marc Reichhart and Petros Ligoxygakis and Silvia Naitza and Gertrud Woerfel and Jean-Luc Imler and David Gubb}, doi = {10.1002/gene.10122}, issn = {1526-954X}, year = {2002}, date = {2002-01-01}, journal = {Genesis (New York, N.Y.: 2000)}, volume = {34}, number = {1-2}, pages = {160--164}, keywords = {Animals, DNA Transposable Elements, DNA-Binding Proteins, Enhancer Elements, Genetic, Genetic Vectors, Genetically Modified, imler, M3i, reichhart, Saccharomyces cerevisiae Proteins, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{tzou_over-expression_2002, title = {Over-expression of a single antimicrobial peptide restores wild-type resistance to infection in immuno-deficient Drosophila mutants.}, author = {P Tzou and Jean-Marc Reichhart and Bruno Lemaitre}, year = {2002}, date = {2002-01-01}, journal = {Proc. Natl. Acad. Sci. U S A}, volume = {99}, pages = {2152--2157}, abstract = {The analysis of the in vivo relevance of the antimicrobial peptides (AMPs) for Drosophila resistance against microbial infection is complicated by the numerous AMP genes present in this organism, as well as the redundant defence mechanism within the innate immune system. Mutants deficient for both the Imd and Toll pathway failed to express any AMP genes after infection and are extremely susceptible to both fungal and bacterial infections. To study unambiguously the in vivo role of each antimicrobial peptide in the Drosophila host defence, we have generated imd; spz double mutant flies that over-express a peptide driven by a transgene under a constitutive promoter. Our results clearly show that over-expression of certain peptides are sufficient to rescue the imd; spz susceptibility to microbial infection, supporting the important role of AMPs in Drosophila host defence.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_drosophile_2002, title = {La drosophile : un modèle pour l'étude de la réponse immunitaire des invertébrés}, author = {Dominique Ferrandon and Julien Royet}, year = {2002}, date = {2002-01-01}, journal = {Regards sur la biochimie}, volume = {1}, pages = {27--33}, keywords = {ferrandon, M3i}, pubstate = {published}, tppubtype = {article} } @incollection{royet_humoral_2002, title = {Humoral and cellular responses in textitDrosophila innate immunity}, author = {Julien Royet and Marie Meister and Dominique Ferrandon}, editor = {R A B Ezekowitz and J A Hoffmann}, year = {2002}, date = {2002-01-01}, booktitle = {Innate Immunity}, publisher = {The Humana Press Inc.}, address = {Totowa}, keywords = {ferrandon, M3i}, pubstate = {published}, tppubtype = {incollection} } @article{tauszig-delamasure_drosophila_2002, title = {Drosophila MyD88 is required for the response to fungal and Gram-positive bacterial infections}, author = {Servane Tauszig-Delamasure and Hana Bilak and Maria Capovilla and Jules A Hoffmann and Jean-Luc Imler}, doi = {10.1038/ni747}, issn = {1529-2908}, year = {2002}, date = {2002-01-01}, journal = {Nature Immunology}, volume = {3}, number = {1}, pages = {91--97}, abstract = {We report here the identification and functional characterization of DmMyD88, a gene encoding the Drosophila homolog of mammalian MyD88. DmMyD88 combines a Toll-IL-1R homology (TIR) domain and a death domain. Overexpression of DmMyD88 was sufficient to induce expression of the antifungal peptide Drosomycin, and induction of Drosomycin was markedly reduced in DmMyD88-mutant flies. DmMyD88 interacted with Toll through its TIR domain and required the death domain proteins Tube and Pelle to activate expression of Drs, which encodes Drosomycin. DmMyD88-mutant flies were highly susceptible to infection by fungi and Gram-positive bacteria, but resisted Gram-negative bacterial infection much as did wild-type flies. Phenotypic comparison of DmMyD88-mutant flies and MyD88-deficient mice showed essential differences in the control of Gram-negative infection in insects and mammals.}, keywords = {Adaptor Proteins, Amino Acid, Animals, Antigens, Antimicrobial Cationic Peptides, Cell Surface, Chromosome Mapping, Differentiation, Disease Susceptibility, Enterococcus faecalis, Epistasis, Escherichia coli, Female, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Gram-Negative Bacteria, hoffmann, Hypocreales, imler, Immunologic, Insect, Insect Proteins, M3i, Membrane Glycoproteins, Micrococcus luteus, Myeloid Differentiation Factor 88, Protein Structure, Protein-Serine-Threonine Kinases, Receptors, Recombinant Fusion Proteins, Sequence Alignment, Sequence Homology, Signal Transducing, Tertiary, Toll-Like Receptors, Transfection}, pubstate = {published}, tppubtype = {article} } @article{rutschmann_toll_2002b, title = {The Toll pathway is required for resistance to Gram-positive bacterial infections in Drosophila.}, author = {Sophie Rutschmann and A Kilinc and Dominique Ferrandon}, year = {2002}, date = {2002-01-01}, journal = {J Immunol}, volume = {168}, pages = {1542--1546}, keywords = {ferrandon, M3i}, pubstate = {published}, tppubtype = {article} } @article{imler_toll_2002, title = {Toll receptors in Drosophila: a family of molecules regulating development and immunity}, author = {Jean-Luc Imler and Jules A Hoffmann}, issn = {0070-217X}, year = {2002}, date = {2002-01-01}, journal = {Current Topics in Microbiology and Immunology}, volume = {270}, pages = {63--79}, abstract = {In recent years, Toll-like receptors (TLRs) have emerged as key receptors which detect microbes and initiate an inflammatory response. The Toll receptor was originally identified and characterized 14 years ago for its role in the embryonic development of the fruit-fly Drosophila melanogaster. Subsequently, it was also shown to be an essential component of the signaling pathway mediating the anti-fungal host defense in this model organism. New factors involved in the activation of the Toll receptor or in intracytoplasmic signaling during the immune response in Drosophila have recently been identified. The existence of significant functional differences between mammalian TLRs and Drosophila Toll receptors is also becoming apparent.}, keywords = {Animals, Cell Surface, Genetic, Gram-Negative Bacteria, hoffmann, imler, M3i, Receptors, Signal Transduction, Toll-Like Receptors, Transcription}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genomic hypomethylation in patients with B-cell chronic lymphocytic leukemia}, author = { M. Kowal and J. Roslinski and A. Dmoszynska and A. Borzecki and H. Borzecka and G. Keith}, year = {2002}, date = {2002-01-01}, journal = {Acta Haematol Pol}, volume = {33}, number = {3}, pages = {323-330}, abstract = {During the last few years, increasing attention has been paid to the relationship between DNA methylation and cancer. Hypo- or hypermethylation of DNA may result in up or down regulation of genes involved in the pathogenesis of leukemias. In this study we have analyzed DNA methylation in peripheral blood and bone marrow B (CD19+) and T (CD3+) lymphocytes from untreated patients with B-cell chronic lymphocytic leukemia.}, keywords = {Chronic, DNA, GIEGE, leukemia, lymphocytic, Methylation}, pubstate = {published}, tppubtype = {article} } @article{, title = {Leucyl-tRNA synthetase consisting of two subunits from hyperthermophilic bacteria Aquifex aeolicus}, author = {M G Xu and J F Chen and F Martin and M W Zhao and G Eriani and E D Wang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12196521}, isbn = {12196521}, year = {2002}, date = {2002-01-01}, journal = {J Biol Chem}, volume = {277}, number = {44}, pages = {41590-41596}, abstract = {In a hyperthermophilic bacterium, Aquifex aeolicus, leucyl-tRNA synthetase (LeuRS) consists of two non-identical polypeptide subunits (alpha and beta), different from the canonical LeuRS, which has a single polypeptide chain. By PCR, using genome DNA of A. aeolicus as a template, genes encoding the alpha and beta subunits were amplified and cloned in Escherichia coli. The alpha subunit could not be expressed stably in vivo, whereas the beta subunit was overproduced and purified by a simple procedure. The beta subunit was inactive in catalysis but was able to bind tRNA(Leu). Interestingly, the heterodimer alphabeta-LeuRS could be overproduced in E. coli cells containing both genes and was purified to 95% homogeneity as a hybrid dimer. The kinetics of A. aeolicus LeuRS in pre-steady and steady states and cross-recognition of LeuRS and tRNA(Leu) from A. aeolicus and E. coli were studied. Magnesium concentration, pH value, and temperature aminoacylation optima were determined to be 12 mm, 7.8, and 70 degrees C, respectively. Under optimal conditions, A. aeolicus alphabeta-LeuRS is stable up to 65 degrees C.}, note = {0021-9258 Journal Article}, keywords = {Bacteria/*enzymology Binding Sites Circular Dichroism Cloning, ERIANI, Leu Support, Molecular Enzyme Stability Escherichia coli/enzymology Kinetics Leucine-tRNA Ligase/*chemistry/genetics/isolation & purification Protein Subunits RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Susceptibility of recombinant porcine endogenous retrovirus reverse transcriptase to nucleoside and non-nucleoside inhibitors}, author = {M Wilhelm and J A Fishman and R Pontikis and A M Aubertin and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12568344}, isbn = {12568344}, year = {2002}, date = {2002-01-01}, journal = {Cell Mol Life Sci}, volume = {59}, number = {12}, pages = {2184-2190}, abstract = {Transplantation of organs, tissues or cells from pigs to humans could be a potential solution to the shortage of human organs for transplantation. Porcine endogenous retroviruses (PERVs) remain a major safety concern for porcine xenotransplantation. Thus, finding drugs that could be used as virological prophylaxis (or therapy) against PERV replication would be desirable. One of the most effective ways to block retroviral multiplication is to inhibit the enzyme reverse transcriptase (RT) which catalyzes the reverse transcription of viral RNA to proviral double-stranded DNA. We report here the cloning and expression of PERV RT and its susceptibility to several inhibitors. Our data demonstrate PERV susceptibility in vitro to the triphosphorylated nucleoside analog of zidovudine (AZT) and to ddGTP and to a lesser extent to ddTTP but almost no susceptibility to the non-nucleoside RT inhibitors tested.}, note = {1420-682x Journal Article}, keywords = {Amino Acid Sequence Animals Chlorides/metabolism Cloning, Calf Thymus/metabolism Sodium Chloride/metabolism Support, Non-U.S. Gov't Support, P.H.S. Swine, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Foreword}, author = {E Westhof}, url = {http://www.sciencedirect.com/science/article/pii/S0300908402014645}, doi = {10.1016/S0300-9084(02)01464-5}, year = {2002}, date = {2002-01-01}, journal = {Biochimie}, volume = {84}, number = {8}, pages = {687-689}, keywords = {Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Group I introns and RNA folding}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12440993}, isbn = {12440993}, year = {2002}, date = {2002-01-01}, journal = {Biochem Soc Trans}, volume = {30}, number = {Pt 6}, pages = {1149-1152}, abstract = {Before the discovery of catalytic RNA, tRNA molecules were the most studied RNA molecules for understanding RNA folding. Afterwards, group I introns, because of their stability and the fact that structural folding could be monitored by following their catalytic activity, became the molecule of choice for studying RNA architecture and folding. A major advantage of group I introns for studying the catalytic activity of RNA molecules is that catalytic activity is triggered by the addition of external guanosine cofactors. The self-splicing activity can therefore be precisely controlled. Using group I introns, several RNA motifs central to RNA-RNA self-assembly and folding were discovered. The analysis of the recent X-ray structures of the rRNA subunits indicates that several motifs present in the ribosome occur also in various group I introns.}, note = {0300-5127 Journal Article Review Review, Tutorial}, keywords = {Base Sequence *Introns Molecular Sequence Data *Nucleic Acid Conformation RNA/*chemistry RNA Splicing RNA, Catalytic, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNAML: a standard syntax for exchanging RNA information}, author = {A Waugh and P Gendron and R Altman and J W Brown and D Case and D Gautheret and S C Harvey and N Leontis and J Westbrook and E Westhof and M Zuker and F Major}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12088144}, isbn = {12088144}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {6}, pages = {707-717}, abstract = {Analyzing a single data set using multiple RNA informatics programs often requires a file format conversion between each pair of programs, significantly hampering productivity. To facilitate the interoperation of these programs, we propose a syntax to exchange basic RNA molecular information. This RNAML syntax allows for the storage and the exchange of information about RNA sequence and secondary and tertiary structures. The syntax permits the description of higher level information about the data including, but not restricted to, base pairs, base triples, and pseudoknots. A class-oriented approach allows us to represent data common to a given set of RNA molecules, such as a sequence alignment and a consensus secondary structure. Documentation about experiments and computations, as well as references to journals and external databases, are included in the syntax. The chief challenge in creating such a syntax was to determine the appropriate scope of usage and to ensure extensibility as new needs will arise. The syntax complies with the eXtensible Markup Language (XML) recommendations, a widely accepted standard for syntax specifications. In addition to the various generic packages that exist to read and interpret XML formats, an XML processor was developed and put in the open-source MC-Core library for nucleic acid and protein structure computer manipulation.}, note = {1355-8382 Journal Article}, keywords = {*Databases, Non-P.H.S. Support, Non-U.S. Gov't Support, Nucleic Acid *Nucleic Acid Conformation Programming Languages RNA/*chemistry Support, P.H.S., U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding of tobramycin leads to conformational changes in yeast tRNA(Asp) and inhibition of aminoacylation}, author = {F Walter and J Putz and R Giege and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11847123}, isbn = {11847123}, year = {2002}, date = {2002-01-01}, journal = {EMBO J}, volume = {21}, number = {4}, pages = {760-768}, abstract = {Aminoglycosides inhibit translation in bacteria by binding to the A site in the ribosome. Here, it is shown that, in yeast, aminoglycosides can also interfere with other processes of translation in vitro. Steady-state aminoacylation kinetics of unmodified yeast tRNA(Asp) transcript indicate that the complex between tRNA(Asp) and tobramycin is a competitive inhibitor of the aspartylation reaction with an inhibition constant (K(I)) of 36 nM. Addition of an excess of heterologous tRNAs did not reverse the charging of tRNA(Asp), indicating a specific inhibition of the aspartylation reaction. Although magnesium ions compete with the inhibitory effect, the formation of the aspartate adenylate in the ATP-PP(i) exchange reaction by aspartyl-tRNA synthetase in the absence of the tRNA is not inhibited. Ultraviolet absorbance melting experiments indicate that tobramycin interacts with and destabilizes the native L-shaped tertiary structure of tRNA(Asp). Fluorescence anisotropy using fluorescein-labelled tobramycin reveals a stoichiometry of one molecule bound to tRNA(Asp) with a K(D) of 267 nM. The results indicate that aminoglycosides are biologically effective when their binding induces a shift in a conformational equilibrium of the RNA.}, note = {0261-4189 Journal Article}, keywords = {Acylation Base Sequence Carbohydrate Sequence Fluorescence Polarization Molecular Sequence Data *Nucleic Acid Conformation RNA, Asp/chemistry/*metabolism Saccharomyces cerevisiae/*genetics Support, Fungal/chemistry/*metabolism RNA, Non-U.S. Gov't Tobramycin/*metabolism, Transfer, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Monitoring intermediate folding states of the td group I intron in vivo}, author = {C Waldsich and B Masquida and E Westhof and R Schroeder}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12356744}, isbn = {12356744}, year = {2002}, date = {2002-01-01}, journal = {EMBO J}, volume = {21}, number = {19}, pages = {5281-5291}, abstract = {Group I introns consist of two major structural domains, the P4-P6 and P3-P9 domains, which assemble through interactions with peripheral extensions to fold into an active ribozyme. To assess group I intron folding in vivo, we probed the structure of td wild-type and mutant introns using dimethyl sulfate. The results suggest that the majority of the intron population is in the native state in accordance with the current structural model, which was refined to include two novel tertiary contacts. The importance of the loop E motif in the P7.1-P7.2 extension in assisting ribozyme folding was deduced from modeling and mutational analyses. Destabilization of stem P6 results in a deficiency in tertiary structure formation in both major domains, while weakening of stem P7 only interferes with folding of the P3-P9 domain. The different impact of mutations on the tertiary structure suggests that they interfere with folding at different stages. These results provide a first insight into the structure of folding intermediates and suggest a putative order of events in a hierarchical folding pathway in vivo.}, note = {0261-4189 Journal Article}, keywords = {Bacterial/chemistry/genetics Escherichia coli/genetics Introns/*physiology Models, Base Sequence DNA, Molecular Molecular Sequence Data *Nucleic Acid Conformation Support, Non-U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Antisense RNAs in bacteria and their genetic elements.}, author = {E G Wagner and S Altuvia and P Romby}, editor = {J C Dunlap and C.-ting Wu}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11931231}, isbn = {11931231}, year = {2002}, date = {2002-01-01}, booktitle = {Advances in Genetics: Homology Effects.}, volume = {46}, pages = {361-398}, publisher = {Academic Press}, abstract = {Antisense RNA-mediated regulation is widespread in bacteria. Most antisense RNA control systems have been found in plasmids, phages, and transposons. Fewer examples were identified in bacterial chromosomes. This chapter summarizes our current knowledge about antisense RNAs with respect to their occurrence, their biological roles, and their diverse mechanisms of action. Examples of cis- or trans-encoded antisense RNAs are discussed, and their properties compared. Most antisense RNAs are posttranscriptionally acting inhibitors of target genes, but a few examples of activator antisense RNAs are known. The implications of RNA structure on topologically and kinetically favored binding pathways are addressed, and solutions that have evolved to permit productive interactions between intricately folded RNAs are discussed. Finally, we describe how particular properties of individual antisense/target RNA systems match their respective biological roles.}, note = {0065-2660 Review Review, Academic}, keywords = {Antisense/*genetics/metabolism RNA, Bacteria/*genetics/metabolism Bacteriophages/genetics Chromosomes, Bacterial Models, Bacterial/*genetics/metabolism Support, Bacterial/genetics Conjugation, Genetic DNA Replication/genetics DNA Transposable Elements/genetics Gene Expression Regulation, Genetic Mutation Plasmids/genetics RNA, Non-U.S. Gov't, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Crystal structure of a complex between the aminoglycoside tobramycin and an oligonucleotide containing the ribosomal decoding a site}, author = {Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12079787}, isbn = {12079787}, year = {2002}, date = {2002-01-01}, journal = {Chem Biol}, volume = {9}, number = {6}, pages = {747-755}, abstract = {Aminoglycoside antibiotics target the decoding aminoacyl site (A site) on the 16S ribosomal RNA and induce miscoding during translation. Here, we present the crystal structure, at 2.54 A resolution, of an RNA oligonucleotide containing the A site sequence complexed to the 4,6-disubstituted 2-deoxystreptamine aminoglycoside tobramycin. The three aminosugar rings making up tobramycin interact with the deep-groove atoms directly or via water molecules and stabilize a fully bulged-out conformation of adenines A(1492) and A(1493). The comparison between this structure and the one previously solved in the presence of paromomycin confirms the importance of the functional groups on the common neamine part of these two antibiotics for binding to RNA. Furthermore, the analysis of the present structure provides a molecular explanation to some of the resistance mechanisms that have spread among bacteria and rendered aminoglycoside antibiotics inefficient.}, note = {1074-5521 Journal Article}, keywords = {16S/*chemistry Support, Anti-Bacterial Agents/*chemistry Binding Sites Crystallography Escherichia coli/metabolism Models, Molecular Oligonucleotides/*chemistry Paromomycin/chemistry Protein Structure, Non-U.S. Gov't Tobramycin/*chemistry, Ribosomal, Secondary Protein Structure, Tertiary RNA, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Brownian-dynamics simulations of metal-ion binding to four-way junctions}, author = {B N van Buuren and T Hermann and S S Wijmenga and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11788713}, isbn = {11788713}, year = {2002}, date = {2002-01-01}, journal = {Nucleic Acids Res}, volume = {30}, number = {2}, pages = {507-514}, abstract = {Four-way junctions (4Hs) are important intermediates in DNA rearrangements such as genetic recombination. Under the influence of multivalent cations these molecules undergo a conformational change, from an extended planar form to a quasi-continuous stacked X-structure. Recently, a number of X-ray structures and a nuclear magnetic resonance (NMR) structure of 4Hs have been reported and in three of these the position of multivalent cations is revealed. These structures belong to two main families, characterized by the angle between the two co-axial stacked helices, which is either around +40 to +55 degrees or around -70 to -80 degrees. To investigate the role of metal-ion binding on the conformation of folded 4Hs we performed Brownian-dynamics simulations on the set of available structures. The simulations confirm the proposed metal-ion binding sites in the NMR structure and in one of the X-ray structures. Furthermore, the calculations suggest positions for metal-ion binding in the other X-ray structures. The results show a striking dependence of the ion density on the helical environment (B-helix or A-helix) and the structural family.}, note = {1362-4962 Journal Article}, keywords = {Base Pair Mismatch Base Sequence Binding Sites Cations/*metabolism *Computer Simulation DNA/*chemistry/genetics/*metabolism Diffusion Electrostatics Metals/*metabolism Models, Biomolecular *Nucleic Acid Conformation Nucleic Acid Hybridization RNA/chemistry/genetics/metabolism Recombination, Genetic/*genetics Support, Molecular Nuclear Magnetic Resonance, Non-U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural basis of translational control by Escherichia coli threonyl tRNA synthetase}, author = {A Torres-Larios and A C Dock-Bregeon and P Romby and B Rees and R Sankaranarayanan and J Caillet and M Springer and C Ehresmann and B Ehresmann and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11953757}, isbn = {11953757}, year = {2002}, date = {2002-01-01}, journal = {Nat Struct Biol}, volume = {9}, number = {5}, pages = {343-347}, abstract = {Escherichia coli threonyl-tRNA synthetase (ThrRS) represses the translation of its own messenger RNA by binding to an operator located upstream of the initiation codon. The crystal structure of the complex between the core of ThrRS and the essential domain of the operator shows that the mRNA uses the recognition mode of the tRNA anticodon loop to initiate binding. The final positioning of the operator, upon which the control mechanism is based, relies on a characteristic RNA motif adapted to the enzyme surface. The finding of other thrS operators that have this conserved motif leads to a generalization of this regulatory mechanism to a subset of Gram-negative bacteria.}, note = {1072-8368 Journal Article}, keywords = {Anticodon/genetics Base Sequence Crystallography, Bacterial/chemistry/genetics/metabolism RNA, Genetic, Messenger/chemistry/genetics/*metabolism RNA, Molecular Molecular Sequence Data Nucleic Acid Conformation Protein Conformation RNA, Non-U.S. Gov't Threonine-tRNA Ligase/*chemistry/*metabolism *Translation, ROMBY, Transfer/chemistry/genetics/metabolism Sequence Alignment Structure-Activity Relationship Support, Unité ARN, X-Ray Escherichia coli/*enzymology/genetics Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {High affinity nucleic acid aptamers for streptavidin incorporated into bi-specific capture ligands}, author = {A Tahiri-Alaoui and L Frigotto and N Manville and J Ibrahim and P Romby and W James}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12000850}, isbn = {12000850}, year = {2002}, date = {2002-01-01}, journal = {Nucleic Acids Res}, volume = {30}, number = {10}, pages = {e45}, abstract = {We have isolated 2'-Fluoro-substituted RNA aptamers that bind to streptavidin (SA) with an affinity around 7 +/- 1.8 nM, comparable with that of recently described peptide aptamers. Binding to SA was not prevented by prior saturation with biotin, enabling nucleic acid aptamers to form useful ternary complexes. Mutagenesis, secondary structure analysis, ribonuclease footprinting and deletion analysis provided evidence for the essential structural features of SA-binding aptamers. In order to provide a general method for the exploitation of these aptamers, we produced derivatives in which they were fused to the naturally structured RNA elements, CopT or CopA. In parallel, we produced derivatives of CD4-binding aptamers fused to the complementary CopA or CopT elements. When mixed, these two chimeric aptamers rapidly hybridized, by virtue of CopA-CopT complementarity, to form stable, bi-functional aptamers that we called 'adaptamers'. We show that a CD4-SA-binding adaptamer can be used to capture CD4 onto a SA-derivatized surface, illustrating their general utility as indirect affinity ligands.}, note = {1362-4962 Journal Article}, keywords = {Affinity Labels/isolation & purification Base Sequence Binding Sites Binding, Competitive Electrophoretic Mobility Shift Assay Ligands Molecular Sequence Data Nucleic Acid Conformation Oligonucleotides/chemistry/genetics/metabolism RNA/chemistry/isolation & purification/*metabolism Streptavidin/chemistry/*metabolism, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effects of magnesium ions on the stabilization of RNA oligomers of defined structures}, author = {M J Serra and J D Baird and T Dale and B L Fey and K Retatagos and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12003491}, isbn = {12003491}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {3}, pages = {307-323}, abstract = {Optical melting was used to determine the stabilities of 11 small RNA oligomers of defined secondary structure as a function of magnesium ion concentration. The oligomers included helices composed of Watson-Crick base pairs, GA tandem base pairs, GU tandem base pairs, and loop E motifs (both eubacterial and eukaryotic). The effect of magnesium ion concentration on stability was interpreted in terms of two simple models. The first assumes an uptake of metal ion upon duplex formation. The second assumes nonspecific electrostatic attraction of metal ions to the RNA oligomer. For all oligomers, except the eubacterial loop E, the data could best be interpreted as nonspecific binding of metal ions to the RNAs. The effect of magnesium ions on the stability of the eubacterial loop E was distinct from that seen with the other oligomers in two ways. First, the extent of stabilization by magnesium ions (as measured by either change in melting temperature or free energy) was three times greater than that observed for the other helical oligomers. Second, the presence of magnesium ions produces a doubling of the enthalpy for the melting transition. These results indicate that magnesium ion stabilizes the eubacterial loop E sequence by chelating the RNA specifically. Further, these results on a rather small system shed light on the large enthalpy changes observed upon thermal unfolding of large RNAs like group I introns. It is suggested that parts of those large enthalpy changes observed in the folding of RNAs may be assigned to variations in the hydration states and types of coordinating atoms in some specifically bound magnesium ions and to an increase in the observed cooperativity of the folding transition due to the binding of those magnesium ions coupling the two stems together. Brownian dynamic simulations, carried out to visualize the metal ion binding sites, reveal rather delocalized ionic densities in all oligomers, except for the eubacterial loop E, in which precisely located ion densities were previously calculated.}, note = {1355-8382 Journal Article}, keywords = {Base Pairing Heat Hydrogen Bonding Magnesium/*pharmacology Models, Molecular Nucleic Acid Conformation RNA/*metabolism RNA Stability/*drug effects Support, Non-P.H.S. Support, Non-U.S. Gov't Support, P.H.S. Thermodynamics, U.S. Gov't, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Do mRNA and rRNA binding sites of E.coli ribosomal protein S15 share common structural determinants?}, author = {A Serganov and E Ennifar and C Portier and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12126618}, isbn = {12126618}, year = {2002}, date = {2002-01-01}, journal = {J Mol Biol}, volume = {320}, number = {5}, pages = {963-978}, abstract = {Escherichia coli ribosomal protein S15 recognizes two RNA targets: a three-way junction in 16S rRNA and a pseudoknot structure on its own mRNA. Binding to mRNA occurs when S15 is expressed in excess over its rRNA target, resulting in an inhibition of translation start. The sole apparent similarity between the rRNA and mRNA targets is the presence of a G-U/G-C motif that contributes only modestly to rRNA binding but is essential for mRNA. To get more information on the structural determinants used by S15 to bind its mRNA target as compared to its rRNA site, we used site-directed mutagenesis, substitution by nucleotide analogs, footprinting experiments on both RNA and protein, and graphic modeling. The size of the mRNA-binding site could be reduced to 45 nucleotides, without loss of affinity. This short RNA preferentially folds into a pseudoknot, the formation of which depends on magnesium concentration and temperature. The size of the loop L2 that bridges the two stems of the pseudoknot through the minor groove could not be reduced below nine nucleotides. Then we showed that the pseudoknot recognizes the same side of S15 as 16S rRNA, although shielding a smaller surface area. It turned out that the G-U/G-C motif is recognized from the minor groove in both cases, and that the G-C pair is recognized in a very similar manner. However, the wobble G-U pair of the mRNA is not directly contacted by S15, as in rRNA, but is most likely involved in building a precise conformation of the RNA, essential for binding. Otherwise, unique specific features are utilized, such as the three-way junction in the case of 16S rRNA and the looped out A(-46) for the mRNA pseudoknot.}, note = {0022-2836 Journal Article}, keywords = {Binding Sites Cytosine Escherichia coli Guanosine Models, ENNIFAR, Messenger/*chemistry RNA, Molecular Nucleic Acid Conformation Protein Structure, Non-U.S. Gov't Uridine, Ribosomal/*chemistry Ribosomal Proteins/*chemistry Support, Tertiary RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cyclic PNA hexamer-based compound: modelling, synthetis and inhibition of the HIV-1 RNA dimerization process}, author = {C Schwergold and G Depecker and C Di Giorgio and N Patino and F Jossinet and B Ehresmann and R Terreux and D Cabrol-Bass and R Condom}, url = {http://www.sciencedirect.com/science/article/pii/S0040402002005276}, doi = {10.1016/S0040-4020(02)00527-6}, year = {2002}, date = {2002-01-01}, journal = {Tetrahedron}, volume = {58}, number = {28}, pages = {5675-5687}, abstract = {A cyclic molecule constituted by (i) a hexameric PNA moiety complementary to six among the nine residues of the dimerization initiation site loop of HIV-1 and (ii) a spacer tethering the N- to the C-extremities of the PNA, has been elaborated to inhibit the dimerization process of HIV-1 genome. This compound has been synthesized following a liquid-phase procedure (fully protected backbone approach). Preliminary agarose gel electrophoresis analyses have shown that the cyclic PNA conjugate is able to inhibit the HIV- I dimerization.}, keywords = {EHRESMANN RNA hexamer HIV-1 dimerization process Synthesis, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Secondary structure of the 3' terminus of hepatitis C virus minus-strand RNA}, author = {C Schuster and C Isel and I Imbert and C Ehresmann and R Marquet and M P Kieny}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12134011}, isbn = {12134011}, year = {2002}, date = {2002-01-01}, journal = {J Virol}, volume = {76}, number = {16}, pages = {8058-8068}, abstract = {The 3'-terminal ends of both the positive and negative strands of the hepatitis C virus (HCV) RNA, the latter being the replicative intermediate, are most likely the initiation sites for replication by the viral RNA-dependent RNA polymerase, NS5B. The structural features of the very conserved 3' plus [(+)] strand untranslated region [3' (+) UTR] are well established (K. J. Blight and C. M. Rice, J. Virol. 71:7345-7352, 1997). However, little information is available concerning the 3' end of the minus [(-)] strand RNA. In the present work, we used chemical and enzymatic probing to investigate the conformation of that region, which is complementary to the 5' (+) UTR and the first 74 nucleotides of the HCV polyprotein coding sequence. By combining our experimental data with computer predictions, we have derived a secondary-structure model of this region. In our model, the last 220 nucleotides, where initiation of the (+) strand RNA synthesis presumably takes place, fold into five stable stem-loops, forming domain I. Domain I is linked to an overall less stable structure, named domain II, containing the sequences complementary to the pseudoknot of the internal ribosomal entry site in the 5' (+) UTR. Our results show that, even though the (-) strand 3'-terminal region has the antisense sequence of the 5' (+) UTR, it does not fold into its mirror image. Interestingly, comparison of the replication initiation sites on both strands reveals common structural features that may play key functions in the replication process.}, note = {0022-538x Journal Article}, keywords = {Base Sequence DNA, MARQUET, Molecular Molecular Probe Techniques Molecular Sequence Data Nucleic Acid Conformation Plasmids/genetics RNA, Non-U.S. Gov't Virus Replication, Unité ARN, Viral/*chemistry/genetics Support, Viral/genetics Hepacivirus/*chemistry/genetics/physiology Human Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Towards atomic resolution with crystals grown in gel: the case of thaumatin seen at room temperature}, author = {C Sauter and B Lorber and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12112683}, isbn = {12112683}, year = {2002}, date = {2002-01-01}, journal = {Proteins}, volume = {48}, number = {2}, pages = {146-150}, abstract = {One reason for introducing a gel in the crystallization medium of proteins is its ability to reduce convection in solution. This can lead to better nucleation and growth conditions, and to crystals having enhanced diffraction properties. We report here the X-ray characterization at room temperature of high-quality crystals of the intensely sweet thaumatin prepared in a sodium tartrate solution gelified with 0.15% (m/v) agarose. Using a synchrotron radiation, these crystals diffracted to a previously unachieved resolution. A diffraction dataset was collected from four crystals at a resolution of 1.2 A with a R(sym) of 3.6% and a completeness of 99%. Refinement was carried out to a final crystallographic R-factor of 12.0%. The quality of the electron density map allowed for the observation of fine structural details in the protein and its solvation shell. Crystallization in gel might be used more generally to improve the quality of macromolecular crystals. Advantages provided by the gelified medium in the frame of structural studies are emphasized.}, note = {1097-0134 Journal Article}, keywords = {Crystallization *Crystallography, FRUGIER, Molecular Plant Proteins/*chemistry Support, Non-U.S. Gov't *Sweetening Agents Temperature Weightlessness, SAUTER, Unité ARN, X-Ray Hydrogels/*chemistry *Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Primer unblocking and rescue of DNA synthesis by azidothymidine (AZT)-resistant HIV-1 reverse transcriptase: comparison between initiation and elongation of reverse transcription and between (-) and (+) strand DNA synthesis}, author = {M Rigourd and C Ehresmann and M A Parniak and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11901149}, isbn = {11901149}, year = {2002}, date = {2002-01-01}, journal = {J Biol Chem}, volume = {277}, number = {21}, pages = {18611-18618}, abstract = {Azidothymidine (AZT) is a widely used inhibitor of type 1 human immunodeficiency virus reverse transcriptase (RT) that acts as chain terminator. Upon treatment, mutations conferring AZT resistance to RT are gradually selected. It has been shown that resistant RT is able to unblock the AZT-terminated primer by an ATP-dependent mechanism. However, this resistance mechanism has only been demonstrated for DNA-dependent DNA elongation. Here, we compared the AZT resistance of mutant RT during DNA elongation on DNA and RNA templates. We showed that, during DNA elongation, primer unblocking and rescue of DNA synthesis take place with similar rate constants on DNA and RNA templates. However, the fraction of a primer eventually repaired during RNA-dependent DNA synthesis is 2x lower compared with that of DNA-dependent synthesis, leading to reduced resistance. We also compared the initiation of reverse transcription, which uses tRNA(3)(Lys) as a primer and displays characteristic kinetic features, and the subsequent RNA-dependent elongation. Unlike during elongation, resistant RT was unable to unblock the AZT-terminated primer during initiation of (-) DNA strand synthesis. Our results demonstrate that the efficiency of primer unblocking conferred by the AZT resistance mutations greatly vary during the different steps of the provirus synthesis. These results also suggest that inhibitors specifically targeting the initiation of reverse transcription might prove to be advantageous, as compared with elongation inhibitors.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence Comparative Study DNA Primers *DNA Replication Drug Resistance, MARQUET, Microbial HIV-1 Reverse Transcriptase/*drug effects Kinetics Reverse Transcriptase Inhibitors/*pharmacology Support, Non-U.S. Gov't Zidovudine/*pharmacology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comparative proteomics as a new tool for exploring human mitochondrial tRNA disorders}, author = {T Rabilloud and J M Strub and N Carte and S Luche and A Van Dorsselaer and J Lunardi and R Giege and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11772011}, isbn = {11772011}, year = {2002}, date = {2002-01-01}, journal = {Biochemistry}, volume = {41}, number = {1}, pages = {144-150}, abstract = {More than 70 different point mutations in human mitochondrial tRNA genes are correlated with severe disorders, including fatal cardiopathies, encephalopathies, myopathies, and others. So far, investigation of the molecular impact(s) of mutations has focused on the affected tRNA itself by seeking structural and/or functional perturbations capable of interfering with synthesis of the 13 mitochondrion-encoded subunits of respiratory chain complexes. Here, a proteomic approach was used to investigate whether such mutations would affect the pattern of mitochondrial proteins at a broader level. Analysis of several hundred mitochondrial proteins from sibling cybrid cell lines by two-dimensional electrophoresis, an approach that takes into account all regulatory steps of mitochondrial and nuclear gene expression, indeed reveals a number of up- and downregulated proteins when healthy and single-point-mutation-carrying mitochondria representative of either MELAS or MERRF syndrome were compared. Assignment by mass spectrometry of the two proteins which exhibit obvious large quantitative decreases in the levels of both pathologic mitochondria identified nuclear-encoded subunits of cytochrome c oxidase, a respiratory chain complex. This clearly shows a linkage between the effects of mutations in mitochondrial tRNA genes and the steady-state level of nuclear-encoded proteins in mitochondria. It opens new routes toward a large-scale exploration of potential proteic partners involved in the genotype-phenotype correlation of mitochondrial disorders.}, note = {0006-2960 Journal Article}, keywords = {Amino Acid Sequence Cell Line Cell Nucleus/physiology Comparative Study DNA, FLORENTZ, Gel, Inborn/*metabolism Human Mitochondria/*metabolism Mitochondrial Proteins/*metabolism Molecular Sequence Data *Point Mutation Proteome RNA/*genetics RNA, Mitochondrial/physiology Electrophoresis, Non-U.S. Gov't, Transfer/*genetics Support, Two-Dimensional/methods Genetic Diseases, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Estimation of DNA methylation level in nonendometrial uterus malignancies]}, author = {A Popiela and M S Gabrys and J Rabczynski and M Panszczyk and G Keith and W Baranowski}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12722382}, isbn = {12722382}, year = {2002}, date = {2002-01-01}, journal = {Ginekol Pol}, volume = {73}, number = {11}, pages = {962-965}, abstract = {OBJECTIVES: Rebuilding of genome structure leads to many pathological states including neoplastic malignancies. Rebuilding often occurs as a process caused by disturbances in gene silencing mechanism. DNA methylation pattern is one of the most important mechanisms connected to gene's silencing. Estimation of DNA methylation level in nonendometrial uterine neoplastic tissues compared to normal endometrial samples was the aim of this study. DESIGN: It was to be shown, that changes in methylation rate in promotory regions could lead to carcinogenesis in particular cell. Authors describe an analysis of DNA methylation level in nonendometrial neoplastic uterine tissues compared to DNA methylation level in normal endometrium. MATERIALS AND METHODS: Tissue samples from 9 women with tumor mixtus mesodermalis were collected. 12 samples were normal endometrium-control group. DNA was isolated from tissues and than we performed an estimation of DNA methylation levels. Than we performed a statistical analysis of results. RESULTS: The median DNA methylation level was significantly higher in neoplastic tissues than in normal endometrium. CONCLUSIONS: Authors conclude, that DNA methylation level is higher in neoplastic tissues, but does not correlate with clinical stage of the disease.}, note = {0017-0011 Journal Article}, keywords = {80 and over Case-Control Studies *DNA Methylation English Abstract Female Gene Expression Regulation, Age Factors Aged Aged, Mesodermal/genetics/*metabolism Uterine Neoplasms/genetics/*metabolism, Neoplastic Gene Silencing Human Middle Aged Mixed Tumor, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Estimation of DNA methylation level in endometrial cancer tissues]}, author = {A Popiela and M S Gabrys and J Rabczynski and M Panszczyk and G Keith and W Baranowski}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12722383}, isbn = {12722383}, year = {2002}, date = {2002-01-01}, journal = {Ginekol Pol}, volume = {73}, number = {11}, pages = {966-969}, abstract = {OBJECTIVES: A level of DNA methylation plays an important role in regulation of cellular gene's expression. Estimation of DNA methylation level in endometrial neoplastic tissues compared to normal endometrial samples was the aim of this study. DESIGN: It was to be shown, that changes in methylation rate in promotory regions could lead to carcinogenesis in particular cell. Authors describe an analysis of DNA methylation level in endometrial cancer tissues compared to DNA methylation level in normal or hyperplastic endometrium. MATERIAL AND METHODS: Endometrial samples from 88 women were collected. 56 of them were classified as adenocarcinoma, 20 as hyperplastic changes, 12 as normal endometrium-control group. DNA was isolated from tissues and than prepared to pm5dC and pdC. Than we performed a statistical analysis of results. RESULTS: The median DNA methylation level was significantly higher in neoplastic tissues than in normal endometrium. There was no difference between DNA methylation level between normal endometrium and hyperplastic changes. CONCLUSIONS: Authors conclude that neoplastic endometrial tissues show high DNA methylation rate compared to normal or hyperplastic endometrium.}, note = {0017-0011 Journal Article}, keywords = {80 and over Case-Control Studies *DNA Methylation Endometrial Neoplasms/chemistry/*genetics English Abstract Female Gene Expression Regulation, Adenocarcinoma/chemistry/*genetics Aged Aged, Neoplastic Human Hyperplasia/genetics Middle Aged, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Detailed analysis of RNA-protein interactions within the bacterial ribosomal protein L5/5S rRNA complex}, author = {A Perederina and N Nevskaya and O Nikonov and A Nikulin and P Dumas and M Yao and I Tanaka and M Garber and G Gongadze and S Nikonov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12515387}, isbn = {12515387}, year = {2002}, date = {2002-01-01}, journal = {RNA}, volume = {8}, number = {12}, pages = {1548-1557}, abstract = {The crystal structure of ribosomal protein L5 from Thermus thermophilus complexed with a 34-nt fragment comprising helix III and loop C of Escherichia coli 5S rRNA has been determined at 2.5 A resolution. The protein specifically interacts with the bulged nucleotides at the top of loop C of 5S rRNA. The rRNA and protein contact surfaces are strongly stabilized by intramolecular interactions. Charged and polar atoms forming the network of conserved intermolecular hydrogen bonds are located in two narrow planar parallel layers belonging to the protein and rRNA, respectively. The regions, including these atoms conserved in Bacteria and Archaea, can be considered an RNA-protein recognition module. Comparison of the T. thermophilus L5 structure in the RNA-bound form with the isolated Bacillus stearothermophilus L5 structure shows that the RNA-recognition module on the protein surface does not undergo significant changes upon RNA binding. In the crystal of the complex, the protein interacts with another RNA molecule in the asymmetric unit through the beta-sheet concave surface. This protein/RNA interface simulates the interaction of L5 with 23S rRNA observed in the Haloarcula marismortui 50S ribosomal subunit.}, note = {1355-8382 Journal Article}, keywords = {5S/*chemistry/*metabolism Ribosomal Proteins/*chemistry/*metabolism Support, Amino Acid Sequence Bacterial Proteins/chemistry/metabolism Base Sequence Binding Sites Escherichia coli/genetics Hydrogen Bonding Models, Molecular Molecular Sequence Data Nucleic Acid Conformation Peptide Fragments/chemistry/metabolism Protein Conformation RNA, Non-U.S. Gov't, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A 5'-3' long-range interaction in Ty1 RNA controls its reverse transcription and retrotransposition}, author = { G. Cristofari and C. Bampi and M. Wilhelm and F. X. Wilhelm and J. L. Darlix}, year = {2002}, date = {2002-01-01}, journal = {EMBO J}, volume = {21}, number = {16}, pages = {4368-79}, abstract = {LTR-retrotransposons are abundant components of all eukaryotic genomes and appear to be key players in their evolution. They share with retroviruses a reverse transcription step during their replication cycle. To better understand the replication of retrotransposons as well as their similarities to and differences from retroviruses, we set up an in vitro model system to examine minus-strand cDNA synthesis of the yeast Ty1 LTR-retrotransposon. Results show that the 5' and 3' ends of Ty1 genomic RNA interact through 14 nucleotide 5'-3' complementary sequences (CYC sequences). This 5'-3' base pairing results in an efficient initiation of reverse transcription in vitro. Transposition of a marked Ty1 element and Ty1 cDNA synthesis in yeast rely on the ability of the CYC sequences to base pair. This 5'-3' interaction is also supported by phylogenic analysis of all full-length Ty1 and Ty2 elements present in the Saccharomyces cerevisiae genome. These novel findings lead us to propose that circularization of the Ty1 genomic RNA controls initiation of reverse transcription and may limit reverse transcription of defective retroelements.}, note = {0261-4189 Journal Article}, keywords = {*Gene, *Transcription, Acid, cerevisiae/*genetics, Complementary/biosynthesis, Conformation, DNA, Expression, Fungal, Fungal/chemistry/*metabolism, Genetic, Gov't, in, Messenger/chemistry/*metabolism, Non-U.S., Nucleic, Phylogeny, Regulation, Retroelements/*genetics, RNA, Saccharomyces, Support, vitro}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vitro evidence for a long range pseudoknot in the 5'-untranslated and matrix coding regions of HIV-1 genomic RNA}, author = {J C Paillart and E Skripkin and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11744696}, isbn = {11744696}, year = {2002}, date = {2002-01-01}, journal = {J Biol Chem}, volume = {277}, number = {8}, pages = {5995-6004}, abstract = {The 5'-untranslated leader region of human immunodeficiency virus type 1 (HIV-1) RNA contains multiple signals that control distinct steps of the viral replication cycle such as transcription, reverse transcription, genomic RNA dimerization, splicing, and packaging. It is likely that fine tuned coordinated regulation of these functions is achieved through specific RNA-protein and RNA-RNA interactions. In a search for cis-acting elements important for the tertiary structure of the 5'-untranslated region of HIV-1 genomic RNA, we identified, by ladder selection experiments, a short stretch of nucleotides directly downstream of the poly(A) signal that interacts with a nucleotide sequence located in the matrix region. Confirmation of the sequence of the interacting sites was obtained by partial or complete inhibition of this interaction by antisense oligonucleotides and by nucleotide substitutions. In the wild type RNA, this long range interaction was intramolecular, since no intermolecular RNA association was detected by gel electrophoresis with an RNA mutated in the dimerization initiation site and containing both sequences involved in the tertiary interaction. Moreover, the functional importance of this interaction is supported by its conservation in all HIV-1 isolates as well as in HIV-2 and simian immunodeficiency virus. Our results raise the possibility that this long range RNA-RNA interaction might be involved in the full-length genomic RNA selection during packaging, repression of the 5' polyadenylation signal, and/or splicing regulation.}, note = {0021-9258 Journal Article}, keywords = {5' Untranslated Regions/*genetics Base Sequence Dimerization HIV-1/*genetics Human Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Oligodeoxyribonucleotides, Antisense/pharmacology Poly A/chemistry/genetics RNA, MARQUET, Non-U.S. Gov't Virus Replication/genetics, Nucleic Acid Support, PAILLART, Unité ARN, Viral/chemistry/drug effects/*genetics Sequence Alignment Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comparative analysis of space-grown and earth-grown crystals of an aminoacyl-tRNA synthetase: space-grown crystals are more useful for structural determination}, author = {J D Ng and C Sauter and B Lorber and N Kirkland and J Arnez and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11914489}, isbn = {11914489}, year = {2002}, date = {2002-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {58}, number = {Pt 4}, pages = {645-652}, abstract = {Protein crystallization under microgravity aims at benefiting from the quasi-absence of convection and sedimentation to favor well ordered crystal nucleation and growth. The dimeric multidomain enzyme aspartyl-tRNA synthetase from Thermus thermophilus has been crystallized within dialysis reactors of the Advanced Protein Crystallization Facility in the laboratory on earth and under microgravity aboard the US Space Shuttle. A strictly comparative crystallographic analysis reveals that the crystals grown in space are superior in every respect to control crystals prepared in otherwise identical conditions on earth. They diffract X-rays more intensely and have a lower mosaicity, facilitating the process of protein structure determination. Indeed, the electron-density map calculated from diffraction data of space-grown crystals contains considerably more detail. The resulting three-dimensional structure model at 2.0 A resolution is more accurate than that produced in parallel using the data originating from earth-grown crystals. The major differences between the structures, including the better defined amino-acid side chains and the higher order of bound water molecules, are emphasized.}, note = {0907-4449 Journal Article}, keywords = {Amino Acyl-tRNA Ligases/*chemistry Comparative Study Crystallization Crystallography, FRUGIER, Molecular Support, Non-U.S. Gov't Thermus thermophilus/chemistry Water/chemistry Weightlessness, SAUTER, Unité ARN, X-Ray Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Microarrays as cancer keys: an array of possibilities}, author = {S Mohr and G D Leikauf and G Keith and B H Rihn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12118031}, isbn = {12118031}, year = {2002}, date = {2002-01-01}, journal = {J Clin Oncol}, volume = {20}, number = {14}, pages = {3165-3175}, abstract = {Malignant transformation results from accumulation of genetic and epigenetic events. Functional studies of cancer will be crucial to our understanding of its complexity and polymorphism. There is no doubt that emerging genomic and proteomic technologies will facilitate such investigations. Microarray technology is a new and efficient approach to extract data of biomedical relevance for a wide range of applications. In cancer research, it will provide high-throughput and valuable insights into differences in an individual's tumor as compared with constitutional DNA, mRNA expression, and protein expression and activity. Across individuals, comparisons could provide tissue-specific disease signatures that provide diagnosis based on hundreds of informative genes. The resulting product should be a wealth of tumor-associated and tumor-specific biomarkers, which may help in cancer etiology, diagnosis, and therapy and ultimately lead to "molecular nosology" of cancers. This review highlights the recent developments in microarray technologies in cancer research, focuses on the results obtained so far, and describes the eventual use of microarray technology for clinical applications.}, note = {0732-183x Journal Article Review Review, Tutorial}, keywords = {Animals Chromosome Aberrations *Gene Expression Profiling/methods *Gene Expression Regulation, DNA/methods Support, Neoplastic Genotype Human Mutation Neoplasms/*genetics *Oligonucleotide Array Sequence Analysis/methods Oncogenes/*genetics Polymorphism (Genetics) Proteome/genetics *Sequence Analysis, P.H.S., U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Lack of genotoxicity of bitumen fumes in transgenic mouse lung}, author = {J C Micillino and C Coulais and S Binet and M C Bottin and G Keith and D Moulin and B H Rihn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11750079}, isbn = {11750079}, year = {2002}, date = {2002-01-01}, journal = {Toxicology}, volume = {170}, number = {1-2}, pages = {11-20}, abstract = {During hot application of bitumen containing materials, e.g. in hot paving or roofing, fumes are emitted that contain polycyclic aromatic compounds. Previous studies with rodents exposed to bitumen and coal-tar fume condensates showed formation of DNA adducts. In order to clarify the genotoxicity of bitumen fumes, we designed a study by using mice carrying a reporter gene for mutagenesis analysis and exposed by nose-only to a constant and reproducible aerosol of bitumen fumes. We analyzed the genotoxic activity of inhaled bitumen fumes generated under those controlled conditions through the induction of mutation and DNA adducts in Big Blue mice. Mice were exposed to bitumen fumes (100 mg/m(3) total particulate matter) 6 h per day during 5 days by nose-only in an inhalation chamber designed in our laboratory. Following a 30-day fixation period, the experiment was terminated and lung DNA was extracted for mutant frequency and adduct determinations. The mutant frequency was determined using the cII and the lacI mutant analysis systems. In, addition, 61 and 54 mutants were sequenced in control and exposed groups, respectively. The study did not show any mutation or adduct induction in the exposed group compared to the control group: cII mutant frequencies were 11.0+/-4.5x10(-5) and 11.0+/-4.8x10(-5) in control and exposed lungs, respectively. Identically, using the lacI mutation detection system, the mutant frequencies were 6.4+/-3.1x10(-5) and 5.8+/-2.0x10(-5). The mutation spectra of both series were quite similar with regard to transition and transversion frequencies. The absence of genotoxicity in the group exposed to 100 mg/m(3) bitumen is discussed with regard to dosage of inhaled polycyclic aromatic compounds and species.}, note = {0300-483x Journal Article}, keywords = {Aerosols Animals Chromatography, Inbred C57BL Mice, Reporter/genetics Hydrocarbons/*toxicity Lac Operon/genetics Lung/*drug effects/metabolism Mice Mice, Thin Layer DNA/drug effects/metabolism DNA Adducts/drug effects Gases/*toxicity Genes, Transgenic Mutagenicity Tests Mutagens/*toxicity Mutation/drug effects, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {From conventional crystallization to better crystals from space: a review on pilot crystallogenesis studies with aspartyl-tRNA synthetases}, author = {B Lorber and A Théobald-Dietrich and C Charron and C Sauter and J D Ng and D W Zhu and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12351885}, isbn = {12351885}, year = {2002}, date = {2002-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {58}, number = {Pt 10 Pt 1}, pages = {1674-1680}, abstract = {Aspartyl-tRNA synthetases were the model proteins in pilot crystallogenesis experiments. They are homodimeric enzymes of Mr approximately 125 kDa that possess as substrates a transfer RNA, ATP and aspartate. They have been isolated from different sources and were crystallized either as free proteins or in association with their ligands. This review discusses their crystallisability with emphasis to crystal quality and structure determination. Crystallization in low diffusivity gelled media or in microgravity environments is highlighted. It has contributed to prepare high-resolution diffracting crystals with better internal order as reflected by their mosaicity. With AspRS from Thermus thermophilus, the better crystalline quality of the space-grown crystals within APCF is correlated with higher quality of the derived electron density maps. Usefulness for structural biology of targeted methods aimed to improve the intrinsic physical quality of protein crystals is highlighted.}, note = {0907-4449 Journal Article Review Review, Tutorial}, keywords = {Aspartate-tRNA Ligase/*chemistry Crystallization/*methods Crystallography, Non-U.S. Gov't Thermus thermophilus/enzymology Weightlessness, SAUTER, Unité ARN, X-Ray Molecular Structure Pilot Projects Saccharomyces cerevisiae/enzymology Space Flight Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A modular plate for the optimization of protein crystallization conditions}, author = {B Lorber and R Cudney}, url = {http://scripts.iucr.org/cgi-bin/paper?S0021889802007227}, doi = {10.1107/S0021889802007227}, year = {2002}, date = {2002-01-01}, journal = {J Appl Cryst}, volume = {35}, number = {4}, pages = {509-510}, abstract = {A modular crystallization plate that simplifies the optimization of crystallization conditions after a preliminary screening procedure has been designed and manufactured.}, keywords = {GIEGE Protein Nucleic acid Crystallization Screen Temperature., Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {X-ray diffraction properties of protein crystals prepared in agarose gel under hydrostatic pressure}, author = {B Lorber and C Charron and M C Robert and B Capelle and A Kadri and G Jenner and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S0022024802017360}, doi = {10.1016/S0022-0248(02)01736-0}, year = {2002}, date = {2002-01-01}, journal = {J Crystal Growth}, volume = {245}, number = {3-4}, pages = {321–333}, abstract = {Crystals of thaumatin and of turkey and hen lysozyme, prepared in a batch at 293 K underhydrostatic pressures in the 0.1–150 MPa range, have been analyzed to investigate how this parameter influences crystallization and if depressurization introduces defects in the crystalline lattice. The X-raydiffractionproperties of depressurized crystals have been compared with those of unpressurized control crystalspreparedunder otherwise identical conditions at atmospheric pressure (0.1 MPa). Independently of pressure, the crystals of each protein belong to the same space group and have cell parameters identical to those of the controls. Their diffraction limit is more dependent upon crystal volume than upon pressure. Crystal mosaicity, expressed as the full-width at half-maximum w of the Bragg reflection profile, was used to quantify the defects in the lattice. While the quality of lysozyme crystals deteriorates as pressure increases, that of tetragonal thaumatin crystals becomes more homogeneous. The first result correlates with the apparition of cleavages oriented perpendicularily to the crystal's c-axis. The second one is interpreted by a gradual selection of well-formed nuclei. The 3D structure of thaumatin in a crystal that was grown under a pressure of 150 MPa and returned to atmospheric pressure was solved. The fold of the polypeptide chain and the distribution of water molecules was compared with those in a reference crystal grown at 0.1 MPa.The results suggest that the crystal lattice is elastic. A possible application of pressure to the preparation of high quality proteincrystals is discussed in the light of these results.}, keywords = {Crystallization Pressure Agarose gel Lysozyme Proteins Thaumatin, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{irving_genome-wide_2001, title = {A genome-wide analysis of immune responses in Drosophila}, author = {Phil Irving and Laurent Troxler and Timothy S Heuer and Marcia Belvin and Casey Kopczynski and Jean-Marc Reichhart and Jules A Hoffmann and Charles Hetru}, doi = {10.1073/pnas.261573998}, issn = {0027-8424}, year = {2001}, date = {2001-12-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {98}, number = {26}, pages = {15119--15124}, abstract = {Oligonucleotide DNA microarrays were used for a genome-wide analysis of immune-challenged Drosophila infected with Gram-positive or Gram-negative bacteria, or with fungi. Aside from the expression of an established set of immune defense genes, a significant number of previously unseen immune-induced genes were found. Genes of particular interest include corin- and Stubble-like genes, both of which have a type II transmembrane domain; easter- and snake-like genes, which may fulfil the roles of easter and snake in the Toll pathway; and a masquerade-like gene, potentially involved in enzyme regulation. The microarray data has also helped to greatly reduce the number of target genes in large gene groups, such as the proteases, helping to direct the choices for future mutant studies. Many of the up-regulated genes fit into the current conceptual framework of host defense, whereas others, including the substantial number of genes with unknown functions, offer new avenues for research.}, keywords = {Animals, bioinformatic, Gene Expression Regulation, Genome, Gram-Negative Bacteria, hoffmann, M3i, Male, Oligonucleotide Array Sequence Analysis, reichhart, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{michel_drosophila_2001, title = {Drosophila Toll is activated by Gram-positive bacteria through a circulating peptidoglycan recognition protein}, author = {T Michel and Jean-Marc Reichhart and Jules A Hoffmann and Julien Royet}, doi = {10.1038/414756a}, issn = {0028-0836}, year = {2001}, date = {2001-12-01}, journal = {Nature}, volume = {414}, number = {6865}, pages = {756--759}, abstract = {Microbial infection activates two distinct intracellular signalling cascades in the immune-responsive fat body of Drosophila. Gram-positive bacteria and fungi predominantly induce the Toll signalling pathway, whereas Gram-negative bacteria activate the Imd pathway. Loss-of-function mutants in either pathway reduce the resistance to corresponding infections. Genetic screens have identified a range of genes involved in these intracellular signalling cascades, but how they are activated by microbial infection is largely unknown. Activation of the transmembrane receptor Toll requires a proteolytically cleaved form of an extracellular cytokine-like polypeptide, Spätzle, suggesting that Toll does not itself function as a bona fide recognition receptor of microbial patterns. This is in apparent contrast with the mammalian Toll-like receptors and raises the question of which host molecules actually recognize microbial patterns to activate Toll through Spätzle. Here we present a mutation that blocks Toll activation by Gram-positive bacteria and significantly decreases resistance to this type of infection. The mutation semmelweis (seml) inactivates the gene encoding a peptidoglycan recognition protein (PGRP-SA). Interestingly, seml does not affect Toll activation by fungal infection, indicating the existence of a distinct recognition system for fungi to activate the Toll pathway.}, keywords = {Amino Acid, Animals, Anti-Bacterial Agents, Anti-Infective Agents, Bacillus thuringiensis, Carrier Proteins, Cell Surface, Chromosome Mapping, Enterococcus faecalis, Fungi, Genes, Gram-Positive Bacteria, Hemolymph, hoffmann, Humans, Insect, Insect Proteins, M3i, Membrane Glycoproteins, Micrococcus luteus, Mutation, Receptors, reichhart, Sequence Homology, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{mueller_mannose_2001, title = {Mannose receptor ligand-positive cells express the metalloprotease decysin in the B cell follicle}, author = {C G Mueller and I Cremer and P E Paulet and S Niida and N Maeda and S Lebeque and W H Fridman and C Sautès-Fridman}, doi = {10.4049/jimmunol.167.9.5052}, issn = {0022-1767}, year = {2001}, date = {2001-11-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {167}, number = {9}, pages = {5052--5060}, abstract = {Decysin, a gene encoding a disintegrin metalloprotease, is transcribed in human dendritic cells (DC) and germinal centers (GC). We have cloned its murine homologue and show that it is processed by the endoprotease furin before secretion of the catalytic domain. We have defined the cell types that express decysin in mouse spleen in the course of an immune response to T cell-dependent Ags. Like in humans, decysin is transcribed by activated CD11c(+) DC that enter the T cell zone from the marginal zone (MZ). In the GC, decysin is expressed by follicular DC and tingible body macrophages. In addition, a MZ cell population expresses decysin and appears to migrate into the B cell follicle. The majority of these follicle-homing cells express the mannose receptor ligand, a marker for the macrophage-like MZ metallophils. The follicle-homing cells are M-CSF dependent, as they are absent in op/op mice that lack functional M-CSF. This suggests that mannose receptor ligand(+) MZ metallophils differentiate into cells that migrate from the MZ into the B cell follicle. Decysin represents the first marker for this previously unrecognized cell population of the mouse spleen, which may represent a precursor for GCDC and may be specialized in the transport of unprocessed Ag from the MZ into developing GC.}, keywords = {ADAM Proteins, Amino Acid Sequence, Animals, B-Lymphocytes, C-Type, Cell Surface, Cloning, Dendritic Cells, Follicular, Germinal Center, Humans, Inbred BALB C, Lectins, ligands, Macrophage Colony-Stimulating Factor, Macrophages, Mannose-Binding Lectins, Metalloendopeptidases, Mice, Molecular, Molecular Sequence Data, Receptors, SPLEEN, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{georgel_drosophila_2001, title = {Drosophila immune deficiency (IMD) is a death domain protein that activates antibacterial defense and can promote apoptosis}, author = {Philippe Georgel and S Naitza and Christine Kappler and Dominique Ferrandon and Daniel Zachary and C Swimmer and C Kopczynski and G Duyk and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {1534-5807}, year = {2001}, date = {2001-10-01}, journal = {Dev. Cell}, volume = {1}, number = {4}, pages = {503--514}, abstract = {We report the molecular characterization of the immune deficiency (imd) gene, which controls antibacterial defense in Drosophila. imd encodes a protein with a death domain similar to that of mammalian RIP (receptor interacting protein), a protein that plays a role in both NF-kappaB activation and apoptosis. We show that imd functions upstream of the DmIKK signalosome and the caspase DREDD in the control of antibacterial peptide genes. Strikingly, overexpression of imd leads to constitutive transcription of these genes and to apoptosis, and both effects are blocked by coexpression of the caspase inhibitor P35. We also show that imd is involved in the apoptotic response to UV irradiation. These data raise the possibility that antibacterial response and apoptosis share common control elements in Drosophila.}, keywords = {Animals, Anti-Infective Agents, Apoptosis, Bacterial Infections, Caspases, Chromosome Mapping, Cysteine Proteinase Inhibitors, DNA Damage, Female, ferrandon, Gene Expression, hoffmann, I-kappa B Kinase, Immunocompromised Host, In Situ Nick-End Labeling, Insect Proteins, M3i, Male, Mutation, Phenotype, Protein Structure, Protein-Serine-Threonine Kinases, reichhart, Tertiary}, pubstate = {published}, tppubtype = {article} } @article{vizioli_gambicin:_2001, title = {Gambicin: a novel immune responsive antimicrobial peptide from the malaria vector Anopheles gambiae}, author = {J Vizioli and Philippe Bulet and Jules A Hoffmann and Fotis C Kafatos and H M Müller and G Dimopoulos}, doi = {10.1073/pnas.221466798}, issn = {0027-8424}, year = {2001}, date = {2001-10-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {98}, number = {22}, pages = {12630--12635}, abstract = {A novel mosquito antimicrobial peptide, gambicin, and the corresponding gene were isolated in parallel through differential display-PCR, an expressed sequence tag (EST) project, and characterization of an antimicrobial activity in a mosquito cell line by reverse-phase chromatography. The 616-bp gambicin ORF encodes an 81-residue protein that is processed and secreted as a 61-aa mature peptide containing eight cysteines engaged in four disulfide bridges. Gambicin lacks sequence homology with other known proteins. Like other Anopheles gambiae antimicrobial peptide genes, gambicin is induced by natural or experimental infection in the midgut, fatbody, and hemocyte-like cell lines. Within the midgut, gambicin is predominantly expressed in the anterior part. Both local and systemic gambicin expression is induced during early and late stages of natural malaria infection. In vitro experiments showed that the 6.8-kDa mature peptide can kill both Gram-positive and Gram-negative bacteria, has a morphogenic effect on a filamentous fungus, and is marginally lethal to Plasmodium berghei ookinetes. An oxidized form of gambicin isolated from the cell line medium was more active against bacteria than the nonoxidized form from the same medium.}, keywords = {Animals, Anopheles, Anti-Bacterial Agents, Anti-Infective Agents, Base Sequence, Chromosome Mapping, hoffmann, Insect Proteins, Insect Vectors, M3i, Malaria, messenger, RNA}, pubstate = {published}, tppubtype = {article} } @article{choppin_characteristics_2001, title = {Characteristics of HIV-1 Nef Regions Containing Multiple CD8+ Ŧ Cell Epitopes: Wealth of HLA-Binding Motifs and Sensitivity to Proteasome Degradation}, author = {Jeannine Choppin and William Cohen and Alberto Bianco and Jean-Paul Briand and Francine Connan and Marc Dalod and Jean-Gérard Guillet}, url = {https://www.jimmunol.org/content/166/10/6164}, doi = {10.4049/jimmunol.166.10.6164}, issn = {0022-1767, 1550-6606}, year = {2001}, date = {2001-05-01}, urldate = {2020-03-31}, journal = {The Journal of Immunology}, volume = {166}, number = {10}, pages = {6164--6169}, abstract = {First and foremost among the many factors that influence epitope presentation are the degradation of Ag, which results in peptide liberation, and the presence of HLA class I molecules able to present the peptides to T lymphocytes. To define the regions of HIV-1 Nef that can provide multiple T cell epitopes, we analyzed the Nef sequence and determined that there are 73 peptides containing 81 HLA-binding motifs. We tested the binding of these peptides to six common HLA molecules (HLA-A2, -A3, -A24, -B7, -B8, and -B35), and we showed that most of them were efficient binders (54% of motifs), especially peptides associating with HLA-A3, -B7/35, and -B8 molecules. Nef peptides most frequently recognized by T cells of HIV-1-infected individuals were 90–97, 135–143, 71–81, 77–85, 90–100, 73–82, and 128–137. The frequency of T cell recognition was not directly related to the strength of peptide-HLA binding. The generation of Nef epitopes is crucial; therefore, we investigated the digestion by the 20S proteasome of a large peptide, Nef66–100. This fragment was efficiently cleaved, and NH2-terminally extended precursors of epitope 71–81 were recognized by T cells of an HIV-1-infected individual. These results suggest that a high frequency of T cell recognition may depend on proteasome cleavage.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{bianco_fullerene-based_2001, title = {Fullerene-based amino acids and peptides}, author = {A Bianco and T Da Ros and M Prato and C Toniolo}, doi = {10.1002/psc.313}, issn = {1075-2617}, year = {2001}, date = {2001-04-01}, journal = {Journal of Peptide Science: An Official Publication of the European Peptide Society}, volume = {7}, number = {4}, pages = {208--219}, abstract = {Recent advances in the chemistry of fullerene have allowed the synthesis of many classes of novel fullerene derivatives. Among these classes, fullerene-based amino acids and peptides are particularly interesting, both for structural studies and biological applications. In this review, we will discuss our own achievements in this rapidly growing field. In particular, the application of fulleroproline (Fpr) amino acids and peptides to medicinal chemistry and material science will be highlighted.}, keywords = {Amino Acids, Animals, Antiviral Agents, carbon, Fullerenes, Humans, I2CT, Infections, Oxidative Stress, Peptides, Proline, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{piotto_destruction_2001, title = {Destruction of Magnetization during TOCSY Experiments Performed under Magic Angle Spinning: Effect of Radial B1 Inhomogeneities}, author = {Martial Piotto and Maryse Bourdonneau and Julien Furrer and Alberto Bianco and Jésus Raya and Karim Elbayed}, url = {http://www.sciencedirect.com/science/article/pii/S1090780701922876}, doi = {10.1006/jmre.2001.2287}, issn = {1090-7807}, year = {2001}, date = {2001-03-01}, urldate = {2020-03-31}, journal = {Journal of Magnetic Resonance}, volume = {149}, number = {1}, pages = {114--118}, abstract = {TOCSY experiments performed on liquid-like samples under magic angle spinning conditions can exhibit some very peculiar behavior. In the most extreme cases, an almost complete loss of magnetization is observed. The intensity of the effect depends essentially on the ratio of the radiofrequency field strength to the speed of rotation of the sample. It is shown in this study that the periodic modulation of the B1 field in the course of the sample rotation is responsible for this effect.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{jung_microfluorometer_2001, title = {A microfluorometer assay to measure the expression of beta-galactosidase and green fluorescent protein reporter genes in single Drosophila flies}, author = {Alain C Jung and M C Criqui and Sophie Rutschmann and Jules A Hoffmann and Dominique Ferrandon}, issn = {0736-6205}, year = {2001}, date = {2001-03-01}, journal = {BioTechniques}, volume = {30}, number = {3}, pages = {594--598, 600--601}, abstract = {beta-galactosidase and green fluorescent protein (GFP) are among the most commonly used reporter genes to monitor gene expression in various organisms including Drosophila melanogaster. Their expression is usually detected in a qualitative way by direct microscopic observations of cells, tissues, or whole animals. To measure in vivo the inducibility of two antimicrobial peptide genes expressed during the Drosophila innate immune response, we have adapted two reporter gene systems based on the beta-galactosidase enzymatic activity and GFP. We have designed a 96-well microplate fluorometric assay sensitive enough to quantify the expression of both reporter genes in single flies. The assay has enabled us to process efficiently and rapidly a large number of individual mutant flies generated during an ethylmethane sulfonate saturation mutagenesis of the Drosophila genome. This method may be used in any screen that requires the quantification of reporter gene activity in individual insects.}, keywords = {Animals, beta-Galactosidase, Cytophotometry, ferrandon, Genes, Green Fluorescent Proteins, hoffmann, Luminescent Proteins, M3i, Reporter}, pubstate = {published}, tppubtype = {article} } @article{lamberty_insect_2001, title = {Insect immunity. Constitutive expression of a cysteine-rich antifungal and a linear antibacterial peptide in a termite insect.}, author = {M Lamberty and Daniel Zachary and R Lanot and C Bordereau and A Robert and Jules A Hoffmann and Philippe Bulet}, doi = {10.1074/jbc.M002998200}, issn = {0021-9258}, year = {2001}, date = {2001-02-01}, journal = {J. Biol. Chem.}, volume = {276}, number = {6}, pages = {4085--4092}, abstract = {Two novel antimicrobial peptides, which we propose to name termicin and spinigerin, have been isolated from the fungus-growing termite Pseudacanthotermes spiniger (heterometabole insect, Isoptera). Termicin is a 36-amino acid residue antifungal peptide, with six cysteines arranged in a disulfide array similar to that of insect defensins. In contrast to most insect defensins, termicin is C-terminally amidated. Spinigerin consists of 25 amino acids and is devoid of cysteines. It is active against bacteria and fungi. Termicin and spinigerin show no obvious sequence similarities with other peptides. Termicin is constitutively present in hemocyte granules and in salivary glands. The presence of termicin and spinigerin in unchallenged termites contrasts with observations in evolutionary recent insects or insects undergoing complete metamorphosis, in which antimicrobial peptides are induced in the fat body and released into the hemolymph after septic injury.}, keywords = {Amino Acid, Animals, Anti-Bacterial Agents, Antifungal Agents, Base Sequence, Chromatography, Cysteine, DNA Primers, High Pressure Liquid, hoffmann, Immunohistochemistry, Isoptera, M3i, Peptides, Protein Conformation, Recombinant Proteins, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{boulanger_immune_2001, title = {Immune response of Drosophila melanogaster to infection with the flagellate parasite Crithidia spp}, author = {Nathalie Boulanger and Laurence Ehret-Sabatier and R Brun and Daniel Zachary and Philippe Bulet and Jean-Luc Imler}, issn = {0965-1748}, year = {2001}, date = {2001-02-01}, journal = {Insect Biochemistry and Molecular Biology}, volume = {31}, number = {2}, pages = {129--137}, abstract = {Insects are able to recognize invading microorganisms and to mount an immune response to bacterial and fungal infections. Recently, the fruitfly Drosophila melanogaster has emerged as a promising invertebrate model to investigate innate immunity because of its well-characterized genetics. Insects are also vectors of numerous parasites which can trigger an immune response. We have investigated the interaction of Drosophila melanogaster with the flagellate protozoan Crithidia spp. We show that a per os parasitic infection triggers the synthesis of several antimicrobial peptides. By reverse phase HPLC and mass spectrometry, peptides were shown to be present in the hemolymph and not in the gut tissue, suggesting the presence of immune messengers between the site of the infection, namely the gut, and the fat body, the main site of synthesis for antimicrobial peptides. Interestingly, we have identified one molecule which is specifically induced in the hemolymph after infection with Crithidia, but not with bacteria, suggesting that Drosophila can discriminate between pathogens. When flagellates were injected into the hemolymph, a low synthesis of antimicrobial peptides was observed together with phagocytosis of parasites by circulating hemocytes. The data presented here suggest that Drosophila-Crithidia spp. represents an interesting model to study host defense against protozoan parasites.}, keywords = {Animals, Anti-Bacterial Agents, Antimicrobial Cationic Peptides, Crithidia, Defensins, Gene Expression, Glycopeptides, Hemocytes, imler, Insect Proteins, M3i, Phagocytosis}, pubstate = {published}, tppubtype = {article} } @article{, title = {Growth kinetics, diffraction properties and effect of agarose on the stability of a novel crystal form of Thermus thermophilus aspartyl-tRNA synthetase-1}, author = {D W Zhu and B Lorber and C Sauter and J D Ng and P Benas and C Le Grimellec and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11264584}, isbn = {11264584}, year = {2001}, date = {2001-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {57}, number = {Pt 4}, pages = {552-558}, abstract = {Growth kinetics and diffraction properties of monoclinic crystals of eubacterial Thermus thermophilus aspartyl-tRNA synthetase-1 (AspRS-1) prepared in the presence of polyethylene glycol and agarose are studied. Their solubility and two-dimensional phase diagram are compared with those of orthorhombic crystals which grow in the presence of sodium formate or ammonium sulfate. The growth mechanism of the novel crystals was monitored by atomic force microscopy. The gel stabilizes the crystal lattice under the cryogenic conditions used for structure determination at high resolution.}, note = {0907-4449 Journal Article}, keywords = {Aspartate-tRNA Ligase/*chemistry/*metabolism Crystallization Crystallography, Atomic Force Osmolar Concentration Sepharose/*metabolism Solubility Support, Non-U.S. Gov't Temperature Thermodynamics Thermus thermophilus/*enzymology, SAUTER, Unité ARN, X-Ray/methods Enzyme Stability Gels Kinetics Microscopy}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA cleavage by 1,4-diazabicyclo[2.2.2]octane-imidazole conjugates}, author = {M Zenkova and N Beloglazova and V Sil'nikov and V Vlassov and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11582799}, isbn = {11582799}, year = {2001}, date = {2001-01-01}, journal = {Methods Enzymol}, volume = {341}, pages = {468-490}, note = {0076-6879 Journal Article}, keywords = {Aza Compounds/*chemistry Base Sequence Bicyclo Compounds/*chemistry Catalysis Comparative Study Imidazoles/*chemistry Kinetics Molecular Sequence Data Nucleic Acid Conformation Oligoribonucleotides/chemistry Phosphorus Radioisotopes RNA/chemistry/*metabolism RNA, Messenger/chemistry Radioisotope Dilution Technique Ribonuclease, Non-U.S. Gov't, Pancreatic/metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reverse transcription of retroviruses and LTR retrotransposons}, author = {M Wilhelm and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11577982}, isbn = {11577982}, year = {2001}, date = {2001-01-01}, journal = {Cell Mol Life Sci}, volume = {58}, number = {9}, pages = {1246-1262}, abstract = {Retroelements are mobile genetic entities that replicate via reverse transcription of a template RNA. A key component to the life cycle of these elements is the enzyme reverse transcriptase (RT), which copies the single-stranded genomic RNA of the element into a linear double-stranded DNA that is ultimately integrated into the host genome by the element-encoded integrase. RT is a multifunctionnal enzyme which possesses RNA-dependent and DNA-dependent DNA polymerase activities as well as RNase H activity that specifically degrades the RNA strand of RNA-DNA duplexes. At some stages of the replication a strand-displacement activity of RT is also necessary. All activities are essential for the conversion of single-stranded genomic RNA into the double-stranded preintegrative DNA. This review focuses on the role of RT in the different steps of the replication process of retroelements. The features of retrotransposon replication which differ from the retroviral ones will be emphasized. In a second part of the review, the biochemical and enzymatic properties of two newly characterized retrotransposon RTs will be described. The role of the integrase domain in reverse transcriptase activity of some retroviral and retrotransposon RTs will be discussed.}, note = {1420-682x Journal Article Review Review, Academic}, keywords = {Amino Acid *Terminal Repeat Sequences, Amino Acid Sequence Animals Base Sequence Human Molecular Sequence Data Nucleic Acid Conformation RNA-Directed DNA Polymerase/*chemistry/*metabolism *Retroelements Retroviridae/*enzymology/*genetics Sequence Alignment Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Polypurine tract formation by Ty1 RNase H}, author = {M Wilhelm and O Uzun and E H Mules and A Gabriel and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11595735}, isbn = {11595735}, year = {2001}, date = {2001-01-01}, journal = {J Biol Chem}, volume = {276}, number = {50}, pages = {47695-47701}, abstract = {To better understand the mechanism by which Ty1 RNase H creates the polypurine tract (PPT) primer, we have demonstrated the polymerase-dependent hydrolytic activity of Ty1 reverse transcriptase (RT) during minus-strand synthesis. Using RNase H and polymerase mutants of the recombinant Ty1 RT protein, we show that the two domains of Ty1 RT can act independently of one another. Our results indicate that RNA/DNA substrates containing a short RNA PPT, which serve as primers for plus-strand DNA synthesis, are relatively resistant to RNase H cleavage. RNA substrates with a correct 5' end but with 3' end extending beyond the plus-strand initiation site were cleaved specifically to generate the correct 3' end of the PPT. Using long RNA/DNA duplexes containing the PPT, we show that Ty1 RT is able to make specific internal cleavages that could generate the plus-strand primer with correct 5' and 3' ends. Long RNA/DNA duplexes with mutations in the PPT or in a U-rich region upstream of the PPT, which abolish plus-strand initiation in vivo, were not cleaved specifically at the 5' end of the PPT. Our work demonstrates that the in vitro enzyme can recapitulate key processes that control proper replication in vivo.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence Binding Sites DNA/metabolism DNA Primers/pharmacology Hydrolysis Molecular Sequence Data Mutation Protein Binding *Purines RNA/metabolism RNA-Directed DNA Polymerase/*chemistry/*metabolism Recombinant Proteins/metabolism *Retroelements Ribonuclease H, Calf Thymus/*chemistry/*genetics/metabolism Support, Non-U.S. Gov't Support, P.H.S. Time Factors, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {tRNA structure}, author = {E Westhof and P Auffinger}, url = {http://www-ibmc.u-strasbg.fr/arn/Westhof/publ_West/bib2001/r2001_EWesthof_ELS.pdf}, doi = {10.1002/9780470015902.a0000527.pub2}, year = {2001}, date = {2001-01-01}, booktitle = {Encyclopedia of Life Sciences}, publisher = {John Wiley & Sons}, abstract = {Transfer ribonucleic acid (tRNA) molecules that participate in the elongation step of protein synthesis on the ribosome have a conserved secondary structure, known as the cloverleaf, and fold into a common three-dimensional architecture.}, keywords = {transfer ribonucleic acid tRNA cloverleaf structure wobble hypothesis anticodon Watson–Crick pairs non-Watson–Crick pairs hydrogen bond Hoogsteen pairs dynamics solvation magnesium, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Protein-dependent transition states for ribonucleoprotein assembly}, author = {A E Webb and M A Rose and E Westhof and K M Weeks}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11399081}, isbn = {11399081}, year = {2001}, date = {2001-01-01}, journal = {J Mol Biol}, volume = {309}, number = {5}, pages = {1087-1100}, abstract = {Native folding and splicing by the Saccharomyces cerevisiae mitochondrial bI5 group I intron RNA is facilitated by both the S. cerevisiae CBP2 and Neurospora crassa CYT-18 protein cofactors. Both protein-bI5 RNA complexes splice at similar rates, suggesting that the RNA active site structure is similar in both ribonucleoproteins. In contrast, the two proteins assemble with the bI5 RNA by distinct mechanisms and bind opposing, but partially overlapping, sides of the group I intron catalytic core. Assembly with CBP2 is limited by a slow, unimolecular RNA folding step characterized by a negligible activation enthalpy. We show that assembly with CYT-18 shows four distinctive features. (1) CYT-18 binds stably to the bI5 RNA at the diffusion controlled limit, but assembly to a catalytically active RNA structure is still limited by RNA folding, as visualized directly using time-resolved footprinting. (2) This mechanism of rapid stable protein binding followed by subsequent assembly steps has a distinctive kinetic signature: the apparent ratio of k(off) to k(on), determined in a partitioning experiment, differs from the equilibrium K(d) by a large factor. (3) Assembly with CYT-18 is characterized by a large activation enthalpy, consistent with a rate limiting conformational rearrangement. (4) Because assembly from the kinetically trapped state is faster at elevated temperature, we can identify conditions where CYT-18 accelerates (catalyzes) bI5 RNA folding relative to assembly with CBP2.}, note = {0022-2836 Journal Article}, keywords = {Allosteric Site Base Sequence Catalysis Catalytic Domain Fungal Proteins/*metabolism Hydroxyl Radical/metabolism Introns/genetics Iodine/metabolism Kinetics Models, Catalytic/chemistry/genetics/metabolism RNA-Binding Proteins/*metabolism Ribonucleoproteins/chemistry/*genetics/*metabolism Saccharomyces cerevisiae/enzymology/genetics *Saccharomyces cerevisiae Proteins Support, Molecular Neurospora crassa *Nucleic Acid Conformation Protein Binding RNA/chemistry/genetics/metabolism RNA Splicing/*genetics RNA Stability RNA, Non-U.S. Gov't Support, P.H.S., U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Catalytic RNA}, author = {F Walter and E Westhof}, url = {http://www-ibmc.u-strasbg.fr/upr9002/westhof/PDF/r2000_FWalter_ELS.pdf}, doi = {10.1038/npg.els.0000870}, year = {2001}, date = {2001-01-01}, booktitle = {Encyclopedia of Life Sciences}, publisher = {John Wiley & Sons}, abstract = {Some RNA molecules can function like enzymes and exert a catalytic action on themselves or on other molecules. These ribozymes are diverse in size and sequence and differ in important aspects of their three-dimensional structure and folding. Catalytic RNAs present a new target for drugs and can be used for inactivating unwanted RNA or DNA molecules by a specific cleavage reaction.}, keywords = {ribozyme RNA chemistry nucleic acid conformation in vitro selection antibiotics gene therapy, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Dimerization of the 3'UTR of bicoid mRNA involves a two-step mechanism}, author = {C Wagner and I Palacios and L Jaeger and D St Johnston and B Ehresmann and C Ehresmann and C Brunel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11676536}, isbn = {11676536}, year = {2001}, date = {2001-01-01}, journal = {J Mol Biol}, volume = {313}, number = {3}, pages = {511-524}, abstract = {The proper localization of bicoid (bcd) mRNA requires cis-acting signals within its 3' untranslated region (UTR) and trans-acting factors such as Staufen. Dimerization of bcd mRNA through intermolecular base-pairing between two complementary loops of domain III of the 3'UTR was proposed to be important for particle formation in the embryo. The participation in the dimerization process of each domain building the 3'UTR was evaluated by thermodynamic and kinetic analysis of various mutated and truncated RNAs. Although sequence complementarity between the two loops of domain III is required for initiating mRNA dimerization, the initial reversible loop-loop complex is converted rapidly into an almost irreversible complex. This conversion involves parts of RNA outside of domain III that promote initial recognition, and dimerization can be inhibited by sense or antisense oligonucleotides only before conversion has proceeded. Injection of the different bcd RNA variants into living Drosophila embryos shows that all elements that inhibit RNA dimerization in vitro prevent formation of localized particles containing Staufen. Particle formation appeared to be dependent on both mRNA dimerization and other element(s) in domains IV and V. Domain III of bcd mRNA could be substituted by heterologous dimerization motifs of different geometry. The resulting dimers were converted into stable forms, independently of the dimerization module used. Moreover, these chimeric RNAs were competent in forming localized particles and recruiting Staufen. The finding that the dimerization domain of bcd mRNA is interchangeable suggests that dimerization by itself, and not the precise geometry of the intermolecular interactions, is essential for the localization process. This suggests that the stabilizing interactions that are formed during the second step of the dimerization process might represent crucial elements for Staufen recognition and localization.}, note = {0022-2836 Journal Article}, keywords = {3' Untranslated Regions/*chemistry/genetics/*metabolism Animals Base Pairing Base Sequence Biological Transport Dimerization Drosophila Proteins/genetics/metabolism Drosophila melanogaster/embryology/*genetics/metabolism Genes, Biological Mutation/genetics *Nucleic Acid Conformation Oligonucleotides/chemistry/genetics/metabolism Protein Transport RNA-Binding Proteins/metabolism Regulatory Sequences, Insect/genetics Homeodomain Proteins/*genetics Kinetics Models, Non-U.S. Gov't Thermodynamics Trans-Activators/*genetics, Nucleic Acid/genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Manipulation of tRNA properties by structure-based and combinatorial in vitro approaches}, author = {S Vortler and J Putz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11642365}, isbn = {11642365}, year = {2001}, date = {2001-01-01}, journal = {Prog Nucleic Acid Res Mol Biol}, volume = {70}, pages = {291-334}, abstract = {The wide knowledge accumulated over the years on the structure and function of transfer RNAs (tRNAs) has allowed molecular biologists to decipher the rules underlying the function and the architecture of these molecules. These rules will be discussed and the implications for manipulating tRNA properties by structure-based and combinatorial in vitro approaches reviewed. Since most of the signals conferring function to tRNAs are located on the two distal extremities of their three-dimensional L shape, this implies that the structure of the RNA domain connecting these two extremities can be of different architecture and/or can be modified without disturbing individual functions. This concept is first supported by the existence in nature of RNAs of peculiar structures having tRNA properties, as well as by engineering experiments on natural tRNAs. The concept is further illustrated by examples of RNAs designed by combinatorial methods. The different procedures used to select RNAs or tRNA-mimics interacting with aminoacyl-tRNA synthetases or with elongation factors and to select tRNA-mimics aminoacylated by synthetases are presented, as well as the functional and structural characteristics of the selected molecules. Production and characteristics of aptameric RNAs fulfilling aminoacyl-tRNA synthetase functions and of RNAs selected to have affinities for amino acids are also described. Finally, properties of RNAs obtained by either the structure-based or the combinatorial methods are discussed in the light of the origin and evolution of the translation machinery, but also with a view to obtain new inhibitors targeting specific steps in translation.}, note = {0079-6603 Journal Article Review Review, Academic}, keywords = {Base Sequence *Combinatorial Chemistry Techniques In Vitro Molecular Mimicry Nucleic Acid Conformation RNA, Non-U.S. Gov't, Transfer/*chemistry/*metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of paromomycin docked into the eubacterial ribosomal decoding A site}, author = {Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11587639}, isbn = {11587639}, year = {2001}, date = {2001-01-01}, journal = {Structure}, volume = {9}, number = {8}, pages = {647-658}, abstract = {BACKGROUND: Aminoglycoside antibiotics interfere with translation in both gram-positive and gram-negative bacteria by binding to the tRNA decoding A site of the 16S ribosomal RNA. RESULTS: Crystals of complexes between oligoribonucleotides incorporating the sequence of the ribosomal A site of Escherichia coli and the aminoglycoside paromomycin have been solved at 2.5 A resolution. Each RNA fragment contains two A sites inserted between Watson-Crick pairs. The paromomycin molecules interact in an enlarged deep groove created by two bulging and one unpaired adenines. In both sites, hydroxyl and ammonium side chains of the antibiotic form 13 direct hydrogen bonds to bases and backbone atoms of the A site. In the best-defined site, 8 water molecules mediate 12 other hydrogen bonds between the RNA and the antibiotics. Ring I of paromomycin stacks over base G1491 and forms pseudo-Watson-Crick contacts with A1408. Both the hydroxyl group and one ammonium group of ring II form direct and water-mediated hydrogen bonds to the U1495oU1406 pair. The bulging conformation of the two adenines A1492 and A1493 is stabilized by hydrogen bonds between phosphate oxygens and atoms of rings I and II. The hydrophilic sites of the bulging A1492 and A1493 contact the shallow groove of G=C pairs in a symmetrical complex. CONCLUSIONS: Water molecules participate in the binding specificity by exploiting the antibiotic hydration shell and the typical RNA water hydration patterns. The observed contacts rationalize the protection, mutation, and resistance data. The crystal packing mimics the intermolecular contacts induced by aminoglycoside binding in the ribosome.}, note = {0969-2126 Journal Article}, keywords = {16S/*chemistry Ribosomes/*chemistry Spectrometry, Amino Acid Motifs Anti-Bacterial Agents/chemistry Base Sequence Binding Sites Crystallography, Mass, Matrix-Assisted Laser Desorption-Ionization Support, Molecular Molecular Sequence Data Mutation Paromomycin/*chemistry Protein Structure, Non-U.S. Gov't Tobramycin/chemistry Water/chemistry, Ribosomal, Secondary RNA, Unité ARN, X-Ray Escherichia coli/metabolism Magnetic Resonance Spectroscopy Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Statistical analysis of atomic contacts at RNA-protein interfaces}, author = {M Treger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11500966}, isbn = {11500966}, year = {2001}, date = {2001-01-01}, journal = {J Mol Recognit}, volume = {14}, number = {4}, pages = {199-214}, abstract = {Forty-five crystals of complexes between proteins and RNA molecules from the Protein Data Bank have been statistically surveyed for the number of contacts between RNA components (phosphate, ribose and the four bases) and amino acid side chains. Three groups of complexes were defined: the tRNA synthetases; the ribosomal complexes; and a third group containing a variety of complexes. The types of atomic contacts were a priori classified into ionic, neutral H-bond, C-H.O H-bond, or van der Waals interaction. All the contacts were organized into a relational database which allows for statistical analysis. The main conclusions are the following: (i) in all three groups of complexes, the most preferred amino acids (Arg, Asn, Ser, Lys) and the less preferred ones (Ala, Ile, Leu, Val) are the same; Trp and Cys are rarely observed (respectively 15 and 5 amino acids in the ensemble of interfaces); (ii) of the total number of amino acids located at the interfaces 22% are hydrophobic, 40% charged (positive 32%, negative 8%), 30% polar and 8% are Gly; (iii) in ribosomal complexes, phosphate is preferred over ribose, which is preferred over the bases, but there is no significant preference in the other two groups; (iv) there is no significant prevalence of a base type at protein-RNA interfaces, but specifically Arg and Lys display a preference for phosphate over ribose and bases; Pro and Asn prefer bases over ribose and phosphate; Met, Phe and Tyr prefer ribose over phosphate and bases. Further, Ile, Pro, Ser prefer A over the others; Leu prefers C; Asp and Gly prefer G; and Asn prefers U. Considering the contact types, the following conclusions could be drawn: (i) 23% of the contacts are via potential H-bonds (including CH.O H-bonds and ionic interactions), 72% belong to van der Waals interactions and 5% are considered as short contacts; (ii) of all potential H-bonds, 54% are standard, 33% are of the C-H.O type and 13% are ionic; (iii) the Watson-Crick sites of G, O6(G) and principally N2(G) and the hydroxyl group O2' is more often involved in H-bonds than expected; the protein main chain is involved in 32% and the side chains in 68% of the H-bonds; considering the neutral and ionic H-bonds, the following couples are more frequent than expected-base A-Ser, base G-Asp/Glu, base U-Asn. The RNA CH groups interact preferentially with oxygen atoms (62% on the main chain and 19% on the side chains); (iv) the bases are involved in 38% of all H-bonds and more than 26% of the H-bonds have the H donor group on the RNA; (v) the atom O2' is involved in 21% of all H-bonds, a number greater than expected; (vi) amino acids less frequently in direct contact with RNA components interact frequently via their main chain atoms through water molecules with RNA atoms; in contrast, those frequently observed in direct contact, except Ser, use instead their side chain atoms for water bridging interactions.}, note = {0952-3499 Journal Article}, keywords = {Amino Acids/analysis Amino Acyl-tRNA Ligases/chemistry/metabolism Animals Comparative Study Crystallography, Factual Electrostatics Hydrogen Bonding Ions Nucleotides/chemistry Phosphates/chemistry Protein Binding Protein Structure, Secondary RNA/*chemistry/*metabolism RNA-Binding Proteins/*chemistry/*metabolism Ribose/chemistry Ribosomes/chemistry/metabolism Statistics Water/metabolism, Unité ARN, X-Ray Databases}, pubstate = {published}, tppubtype = {article} } @article{, title = {Isolation, crystallization, and investigation of ribosomal protein S8 complexed with specific fragments of rRNA of bacterial or archaeal origin}, author = {S V Tishchenko and J M Vassilieva and O B Platonova and A A Serganov and N P Fomenkova and E S Mudrik and W Piendl and C Ehresmann and B Ehresmann and M B Garber}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11703173}, isbn = {11703173}, year = {2001}, date = {2001-01-01}, journal = {Biochemistry (Mosc)}, volume = {66}, number = {9}, pages = {948-953}, abstract = {The core ribosomal protein S8 binds to the central domain of 16S rRNA independently of other ribosomal proteins and is required for assembling the 30S subunit. It has been shown with E. coli ribosomes that a short rRNA fragment restricted by nucleotides 588-602 and 636-651 is sufficient for strong and specific protein S8 binding. In this work, we studied the complexes formed by ribosomal protein S8 from Thermus thermophilus and Methanococcus jannaschii with short rRNA fragments isolated from the same organisms. The dissociation constants of the complexes of protein S8 with rRNA fragments were determined. Based on the results of binding experiments, rRNA fragments of different length were designed and synthesized in preparative amounts in vitro using T7 RNA-polymerase. Stable S8-RNA complexes were crystallized. Crystals were obtained both for homologous bacterial and archaeal complexes and for hybrid complexes of archaeal protein with bacterial rRNA. Crystals of the complex of protein S8 from M. jannaschii with the 37-nucleotide rRNA fragment from the same organism suitable for X-ray analysis were obtained.}, note = {0006-2979 Journal Article}, keywords = {Archaeal/chemistry/metabolism RNA, Bacterial/chemistry/metabolism RNA, Binding Sites Crystallization Magnesium/chemistry/metabolism Methanococcus/chemistry/genetics Nucleic Acid Conformation RNA, Non-U.S. Gov't Thermus thermophilus/chemistry/genetics, Ribosomal/*chemistry/*metabolism Ribosomal Proteins/*chemistry/isolation & purification/*metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Detailed analysis of RNA-protein interactions within the ribosomal protein S8-rRNA complex from the archaeon Methanococcus jannaschii}, author = {S Tishchenko and A Nikulin and N Fomenkova and N Nevskaya and O Nikonov and P Dumas and H Moine and B Ehresmann and C Ehresmann and W Piendl and V Lamzin and M Garber and S Nikonov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11478863}, isbn = {11478863}, year = {2001}, date = {2001-01-01}, journal = {J Mol Biol}, volume = {311}, number = {2}, pages = {311-324}, abstract = {The crystal structure of ribosomal protein S8 bound to its target 16 S rRNA from a hyperthermophilic archaeon Methanococcus jannaschii has been determined at 2.6 A resolution. The protein interacts with the minor groove of helix H21 at two sites located one helical turn apart, with S8 forming a bridge over the RNA major groove. The specificity of binding is essentially provided by the C-terminal domain of S8 and the highly conserved nucleotide core, characterized by two dinucleotide platforms, facing each other. The first platform (A595-A596), which is the less phylogenetically and structurally constrained, does not directly contact the protein but has an important shaping role in inducing cross-strand stacking interactions. The second platform (U641-A642) is specifically recognized by the protein. The universally conserved A642 plays a pivotal role by ensuring the cohesion of the complex organization of the core through an array of hydrogen bonds, including the G597-C643-U641 base triple. In addition, A642 provides the unique base-specific interaction with the conserved Ser105, while the Thr106 - Thr107 peptide link is stacked on its purine ring. Noteworthy, the specific recognition of this tripeptide (Thr-Ser-Thr/Ser) is parallel to the recognition of an RNA tetraloop by a dinucleotide platform in the P4-P6 ribozyme domain of group I intron. This suggests a general dual role of dinucleotide platforms in recognition of RNA or peptide motifs. One prominent feature is that conserved side-chain amino acids, as well as conserved bases, are essentially involved in maintaining tertiary folds. The specificity of binding is mainly driven by shape complementarity, which is increased by the hydrophobic part of side-chains. The remarkable similarity of this complex with its homologue in the T. thermophilus 30 S subunit indicates a conserved interaction mode between Archaea and Bacteria.}, note = {0022-2836 Journal Article}, keywords = {16S/*chemistry/genetics/*metabolism RNA-Binding Proteins/chemistry/metabolism Ribosomal Proteins/*chemistry/*metabolism Ribosomes/chemistry/genetics/metabolism Sequence Alignment Substrate Specificity Support, Amino Acid Sequence Archaeal Proteins/chemistry/metabolism Bacteria/chemistry/genetics Base Sequence Binding Sites Conserved Sequence/genetics Crystallography, Archaeal/chemistry/genetics/metabolism RNA, Molecular Human Hydrogen Bonding Methanococcus/*chemistry/*genetics Models, Molecular Molecular Sequence Data Nucleic Acid Conformation Protein Binding Protein Structure, Non-U.S. Gov't, Ribosomal, Secondary RNA, Unité ARN, X-Ray Evolution}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Crystals grown in microgravity can yield better protein structures: the examples of thaumatin and aspartyl-tRNA synthetase.}, author = {A Théobald-Dietrich and B Lorber and J D Ng and C Sauter and C Charron and M C Robert and B Capelle and R Giege}, editor = {O Minster and B Schurmann}, url = {http://adsabs.harvard.edu/abs/2001ESASP.454.457T}, year = {2001}, date = {2001-01-01}, booktitle = {Microgravity Research and Aplications in Physical Sciences and Biotechnology, Proceedings of the First International Symposium held 10-15 September, 2000 in Sorrento, Italy.}, pages = {457-463}, publisher = {European Space Agency}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Role of conserved nucleotides in building the 16 S rRNA binding site for ribosomal protein S15}, author = {A Serganov and L Benard and C Portier and E Ennifar and M Garber and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11162092}, isbn = {11162092}, year = {2001}, date = {2001-01-01}, journal = {J Mol Biol}, volume = {305}, number = {4}, pages = {785-803}, abstract = {Ribosomal protein S15 recognizes a highly conserved target on 16 S rRNA, which consists of two distinct binding regions. Here, we used extensive site-directed mutagenesis on a Escherichia coli 16 S rRNA fragment containing the S15 binding site, to investigate the role of conserved nucleotides in protein recognition and to evaluate the relative contribution of the two sites. The effect of mutations on S15 recognition was studied by measuring the relative binding affinity, RNA probing and footprinting. The crystallographic structure of the Thermus thermophilus complex allowed molecular modelling of the E. coli complex and facilitated interpretation of biochemical data. Binding is essentially driven by site 1, which includes a three-way junction constrained by a conserved base triple and cross-strand stacking. Recognition is based mainly on shape complementarity, and the role of conserved nucleotides is to maintain a unique backbone geometry. The wild-type base triple is absolutely required for protein interaction, while changes in the conserved surrounding nucleotides are partially tolerated. Site 2, which provides functional groups in a conserved G-U/G-C motif, contributes only modestly to the stability of the interaction. Binding to this motif is dependent on binding at site 1 and is allowed only if the two sites are in the correct relative orientation. Non-conserved bulged nucleotides as well as a conserved purine interior loop, although not directly involved in recognition, are used to provide an appropriate flexibility between the two sites. In addition, correct binding at the two sites triggers conformational adjustments in the purine interior loop and in a distal region, which are known to be involved for subsequent binding of proteins S6 and S18. Thus, the role of site 1 is to anchor S15 to the rRNA, while binding at site 2 is aimed to induce a cascade of events required for subunit assembly.}, note = {0022-2836 Journal Article}, keywords = {16S/chemistry/*genetics/*metabolism RNA-Binding Proteins/chemistry/metabolism Ribosomal Proteins/chemistry/*metabolism Sequence Alignment Support, Amino Acid Sequence Base Pairing Base Sequence Binding Sites Conserved Sequence/*genetics *Escherichia coli/chemistry/genetics/metabolism Models, ENNIFAR, Molecular Molecular Sequence Data Mutation/genetics Nuclease Protection Assays Phylogeny Protein Binding Protein Conformation Purines/metabolism RNA, Non-U.S. Gov't Thermodynamics Thermus thermophilus/chemistry, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The nucle(ol)ar Tif6p and Efl1p are required for a late cytoplasmic step of ribosome synthesis}, author = {B Senger and D L Lafontaine and J S Graindorge and O Gadal and A Camasses and A Sanni and J M Garnier and M Breitenbach and E Hurt and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11779510}, isbn = {11779510}, year = {2001}, date = {2001-01-01}, journal = {Mol Cell}, volume = {8}, number = {6}, pages = {1363-1373}, abstract = {Deletion of elongation factor-like 1 (Efl1p), a cytoplasmic GTPase homologous to the ribosomal translocases EF-G/EF-2, results in nucle(ol)ar pre-rRNA processing and pre-60S subunits export defects. Efl1p interacts genetically with Tif6p, a nucle(ol)ar protein stably associated with pre-60S subunits and required for their synthesis and nuclear exit. In the absence of Efl1p, 50% of Tif6p is relocated to the cytoplasm. In vitro, the GTPase activity of Efl1p is stimulated by 60S, and Efl1p promotes the dissociation of Tif6p-60S complexes. We propose that Tif6p binds to the pre-60S subunits in the nucle(ol)us and escorts them to the cytoplasm where the GTPase activity of Efl1p triggers a late structural rearrangement, which facilitates the release of Tif6p and its recycling to the nucle(ol)us.}, note = {1097-2765 Journal Article}, keywords = {Biological Transport Cell Division Cell Nucleolus/*metabolism Cell Nucleus/*metabolism Conserved Sequence Cytoplasm/enzymology/*metabolism Enzyme Activation GTP Phosphohydrolases/chemistry/genetics/*metabolism Gene Deletion Genes, Fungal/chemistry/genetics/metabolism RNA, Non-U.S. Gov't, Post-Transcriptional RNA, Reporter/genetics Molecular Weight Phenotype Protein Subunits RNA Precursors/chemistry/genetics/metabolism *RNA Processing, Ribosomal/chemistry/genetics/metabolism Ribosomes/chemistry/*metabolism Saccharomyces cerevisiae/cytology/genetics/growth & development/*metabolism Saccharomyces cerevisiae Proteins/chemistry/genetics/metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast cytoplasmic and mitochondrial methionyl-tRNA synthetases: two structural frameworks for identical functions}, author = {B Senger and L Despons and P Walter and H Jakubowski and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11469869}, isbn = {11469869}, year = {2001}, date = {2001-01-01}, journal = {J Mol Biol}, volume = {311}, number = {1}, pages = {205-216}, abstract = {The yeast Saccharomyces cerevisiae possesses two methionyl-tRNA synthetases (MetRS), one in the cytoplasm and the other in mitochondria. The cytoplasmic MetRS has a zinc-finger motif of the type Cys-X(2)-Cys-X(9)-Cys-X(2)-Cys in an insertion domain that divides the nucleotide-binding fold into two halves, whereas no such motif is present in the mitochondrial MetRS. Here, we show that tightly bound zinc atom is present in the cytoplasmic MetRS but not in the mitochondrial MetRS. To test whether the presence of a zinc-binding site is required for cytoplasmic functions of MetRS, we constructed a yeast strain in which cytoplasmic MetRS gene was inactivated and the mitochondrial MetRS gene was expressed in the cytoplasm. Provided that methionine-accepting tRNA is overexpressed, this strain was viable, indicating that mitochondrial MetRS was able to aminoacylate tRNA(Met) in the cytoplasm. Site-directed mutagenesis demonstrated that the zinc domain was required for the stability and consequently for the activity of cytoplasmic MetRS. Mitochondrial MetRS, like cytoplasmic MetRS, supported homocysteine editing in vivo in the yeast cytoplasm. Both MetRSs catalyzed homocysteine editing and aminoacylation of coenzyme A in vitro. Thus, identical synthetic and editing functions can be carried out in different structural frameworks of cytoplasmic and mitochondrial MetRSs.}, note = {0022-2836 Journal Article}, keywords = {Acylation Amino Acid Sequence Binding Sites Coenzyme A/metabolism Comparative Study Cysteine/genetics/metabolism Cytoplasm/*enzymology Genes, Fungal/genetics Genetic Complementation Test Homocysteine/genetics/metabolism Kinetics Methionine/metabolism Methionine-tRNA Ligase/*chemistry/genetics/*metabolism Mitochondria/*enzymology Molecular Sequence Data Mutation/genetics Protein Transport RNA, Met/genetics/metabolism Saccharomyces cerevisiae/cytology/*enzymology/genetics Sequence Alignment Structure-Activity Relationship Support, Non-P.H.S. Zinc/metabolism Zinc Fingers/genetics/physiology, Non-U.S. Gov't Support, Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Hydrophobic DNA adducts in relationship to estrogen and progesterone receptors content in human uterine cancer.]}, author = { K. Postawski and E. Olech-Fudali and E. Korobowicz and J. A. Jakowicki and G. Keith and W. Baranowski}, year = {2001}, date = {2001-01-01}, journal = {Ginekol Pol}, volume = {72}, number = {9}, pages = {709-16}, abstract = {OBJECTIVE: Determination of the relationship between hydrophobic DNA adducts (A) and estrogen receptors (ER) and progesterone (PR) receptor status in uterine cancers. METHODS: Using the P1 enriched version of 32P-postlabeling for hydrophobic DNA adducts detection on polyethyleneimine (PEI) cellulose thin layer chromatograms (TLC) we examined 11 uterine cancer DNAs. The quantification of the adducts was performed by Cerenkov counting of the spots. ER and PR status was recognized histochemically and H-score estimate was performed for each investigated cancer tissue. Patterns of uterine cancer DNA adducts were compared to the maps of adducts recognized in normal human endometrium. RESULTS: In three of the studied uterine cancers there was no positive staining of ER and PR; in one case there was a weak ER staining but PR staining was negative. In ER negative tumors the A level was significantly higher than in ER positive cancers (138.1 +/- 64.1 vs. 49.7 +/- 26.8 adducts per 10(9) nucleotides, respectively, p < 0.05). Highest A levels were found in two ER and PR negative G3 metastatic tumors. Finally, in all investigated cancers there was a strong, inverse correlation between ER content and A level (r = -0.67, p < 0.03). In addition, the correlation between PR level and A was of borderline significance (r = -0.6}, note = {0017-0011 Journal Article}, keywords = {Abstract, Adducts/*analysis, Autoradiography, DNA, English, Estrogen/*analysis, Female, Human, Neoplasm/*analysis, Neoplasms/genetics/*pathology, Progesterone/*analysis, Receptors, Uterine}, pubstate = {published}, tppubtype = {article} } @article{, title = {The plant tRNA 3' processing enzyme has a broad substrate spectrum}, author = {S Schiffer and M Helm and A Théobald-Dietrich and R Giege and A Marchfelder}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11444972}, isbn = {11444972}, year = {2001}, date = {2001-01-01}, journal = {Biochemistry}, volume = {40}, number = {28}, pages = {8264-8272}, abstract = {To elucidate the minimal substrate for the plant nuclear tRNA 3' processing enzyme, we synthesized a set of tRNA variants, which were subsequently incubated with the nuclear tRNA 3' processing enzyme. Our experiments show that the minimal substrate for the nuclear RNase Z consists of the acceptor stem and T arm. The broad substrate spectrum of the nuclear RNase Z raises the possibility that this enzyme might have additional functions in the nucleus besides tRNA 3' processing. Incubation of tRNA variants with the plant mitochondrial enzyme revealed that the organellar counterpart of the nuclear enzyme has a much narrower substrate spectrum. The mitochondrial RNase Z only tolerates deletion of anticodon and variable arms and only with a drastic reduction in cleavage efficiency, indicating that the mitochondrial activity can only cleave bona fide tRNA substrates efficiently. Both enzymes prefer precursors containing short 3' trailers over extended 3' additional sequences. Determination of cleavage sites showed that the cleavage site is not shifted in any of the tRNA variant precursors.}, note = {0006-2960 Journal Article}, keywords = {Non-U.S. Gov't, Plant/genetics/*metabolism RNA, Post-Transcriptional RNA, Transfer, Tyr/genetics/*metabolism Substrate Specificity/genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The fragile X mental retardation protein binds specifically to its mRNA via a purine quartet motif}, author = {C Schaeffer and B Bardoni and J L Mandel and B Ehresmann and C Ehresmann and H Moine}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11532944}, isbn = {11532944}, year = {2001}, date = {2001-01-01}, journal = {EMBO J}, volume = {20}, number = {17}, pages = {4803-4813}, abstract = {Fragile X syndrome is caused by the absence of protein FMRP, the function of which is still poorly understood. Previous studies have suggested that FMRP may be involved in various aspects of mRNA metabolism, including transport, stability and/or translatability. FMRP was shown to interact with a subset of brain mRNAs as well as with its own mRNA; however, no specific RNA-binding site could be identified precisely. Here, we report the identification and characterization of a specific and high affinity binding site for FMRP in the RGG-coding region of its own mRNA. This site contains a purine quartet motif that is essential for FMRP binding and can be substituted by a heterologous quartet-forming motif. The specific binding of FMRP to its target site was confirmed further in a reticulocyte lysate through its ability to repress translation of a reporter gene harboring the RNA target site in the 5'-untranslated region. Our data address interesting questions concerning the role of FMRP in the post-transcriptional control of its own gene and possibly other target genes.}, note = {0261-4189 Journal Article}, keywords = {Animals Base Sequence Binding Sites Chickens Chimeric Proteins/chemistry/metabolism Fragile X Syndrome/genetics Gene Expression Regulation Human Kinetics Mental Retardation/*genetics Mice Molecular Sequence Data Nerve Tissue Proteins/*genetics/*metabolism RNA, Genetic Vertebrates Xenopus laevis, Messenger/chemistry/*genetics/*metabolism Rats Recombinant Fusion Proteins/chemistry/metabolism Sequence Alignment Sequence Homology, Non-U.S. Gov't *Translation, Nucleic Acid Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of tetragonal hen egg-white lysozyme at 0.94 A from crystals grown by the counter-diffusion method}, author = {C Sauter and F Otalora and J A Gavira and O Vidal and R Giege and J M Garcia-Ruiz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11468395}, isbn = {11468395}, year = {2001}, date = {2001-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {57}, number = {Pt 8}, pages = {1119-1126}, abstract = {Very high quality crystals of tetragonal hen egg-white lysozyme were grown in the Advanced Protein Crystallization Facility (APCF) on board the Space Shuttle using a modified free-interface diffusion (FID) reactor designed ad hoc to have a longer diffusion path. This design allows the performance of true counter-diffusion experiments. Crystals were obtained under the classical chemical conditions defined 50 y ago with NaCl as a crystallizing agent and acetate pH 4.5 as a buffer. Counter-diffusion crystallization allows a "physical" instead of chemical optimization of growth conditions: indeed, this method screens for the best supersaturation conditions in a single trial and yields crystals of very high quality. A complete diffraction data set was collected at atomic resolution from one of these crystals using synchrotron radiation at the DESY-EMBL beamlines. The overall R(merge) on intensities in the resolution range 31-0.94 A was 5.2% and the data were 98.9% complete. Refinement was carried out with the programs CNS and SHELX97 to a final crystallographic R factor of 12.26% for 72 390 reflections. A mean standard uncertainty in the atomic positions of 0.024 A was estimated from inversion of blocked least-squares matrices. 22 side chains show alternate conformations and the loop 59-75 adopts in the same crystal packing two conformations that were observed for either triclinic or tetragonal lysozyme in previous high-resolution studies. In addition to 255 water molecules, the crystallizing agent (one hexacoordinated sodium ion and five chloride anions) participates in the ordered lysozyme hydration shell.}, note = {0907-4449 Journal Article}, keywords = {Amino Acids/chemistry Animals Anions/chemistry Binding Sites Cations/chemistry Chickens Crystallization Crystallography, Molecular Muramidase/*chemistry Protein Conformation Protein Structure, Non-U.S. Gov't, SAUTER, Tertiary Support, Unité ARN, X-Ray Egg White/analysis Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallogenesis in tRNA aminoacylation systems: how packing accounts for crystallization drawbacks with yeast aspartyl-tRNA synthetase}, author = {C Sauter and B Lorber and A Théobald-Dietrich and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S0022024801010727}, isbn = {10.1016/S0022-0248(01)01072-7}, year = {2001}, date = {2001-01-01}, journal = {J Crystal Growth}, volume = {232}, number = {1-4}, pages = {399-408}, abstract = {Two active forms of homodimeric aspartyl-tRNAsynthetase from Saccharomyces cerevisiae differing in length at their N-terminus crystallize in the same orthorhombic space group (P41212) with identical cell parameters. Initial studies were hampered by the poor and anisotropic diffraction of the crystals of enzyme extracted from yeast cells. Isotropic diffraction at higher resolution was obtained when crystals were grown from an engineered protein deprived of its 70 N-terminal amino acids. The present work describes the packing contacts in crystals of the shortened protein whose structure was solved at 2.3 Å resolution. Each subunit of the enzyme develops two lattice interactions covering a surface of 670 Å2, about 7-fold smaller than that of the interface between monomers. The smallest lattice interaction, covering 150 Å2, brings the anticodon binding domain adjacent to the N-terminus of one monomer in contact with a loop from the active-site domain of a neighboring monomer. Modeling of the extension in the solvent channels shows that the 150 Å2 intermolecular contact is perturbed in protein molecules possessing a floppy appendix while their second and larger 520 Å2 contact area is unaffected. Altogether the packing organization explains the poor diffraction properties of the native enzyme crystals and the enhanced diffraction of the crystals of shortened synthetase.}, keywords = {GIEGE Crystal packing Crystal structure X-ray diffraction Growth from solutions Aspartyl-tRNA synthetase Proteins, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{monneaux_key_2001, title = {Key sequences involved in the spreading of the systemic autoimmune response to spliceosomal proteins}, author = {F Monneaux and S Muller}, doi = {10.1046/j.1365-3083.2001.00942.x}, issn = {0300-9475}, year = {2001}, date = {2001-01-01}, journal = {Scandinavian Journal of Immunology}, volume = {54}, number = {1-2}, pages = {45--54}, abstract = {Immune spreading to multiple intracellular antigens is likely to be of primary importance in organ-specific and systemic autoimmune diseases. A number of mechanisms by which immune spreading may occur from only a single autoreactive epitope have been proposed. Search for an initiator or early epitope thus represents an important area of investigation. For example, many studies have focused on the identification of epitopes recognized by the antibodies from both patients with systemic lupus erythematosus (SLE) and lupus-prone mice. Recently, an autoepitope present in the 70K U1 ribonucleo protein (RNP) and recognized by CD4+ T cells from lupus mice has also been identified. Here, we analyze the results of B- and T-cell-epitope mapping studies of several RNPs present in the spliceosome and propose a model of epitope spreading. In this model, a consensus sequence (the RNP motif) conserved in many nuclear, nucleolar and cytoplasmic antigens, might play a role as 'driver' epitope. This hypothesis is based on the observation that this sequence is recognized by CD4+ T cells from lupus mice and is often targeted by autoantibodies, very early during the course of the disease. Targeting this region that is repeated in different self-antigens, might represent an interesting strategy to interfere with the continuous T-cell stimulation and exposure to specific antigens.}, keywords = {Animals, Autoantibodies, Autoimmune Diseases, Autoimmunity, B-Lymphocyte, Epitopes, Humans, I2CT, Mice, Monneaux, Ribonucleoproteins, Spliceosomes, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{monneaux_murine_2001, title = {Murine models of systemic lupus erythematosus: B and Ŧ cell responses to spliceosomal ribonucleoproteins in MRL/Fas(lpr) and (NZB x NZW)F(1) lupus mice}, author = {F Monneaux and H Dumortier and G Steiner and J P Briand and S Muller}, doi = {10.1093/intimm/13.9.1155}, issn = {0953-8178}, year = {2001}, date = {2001-01-01}, journal = {International Immunology}, volume = {13}, number = {9}, pages = {1155--1163}, abstract = {(NZB x NZW)F(1) and MRL/Fas(lpr) lupus mice present a similar phenotype with a spectrum of autoantibodies associated with very severe nephritis. It is thought, however, that in contrast to other lupus-prone mice such as MRL/Fas(lpr) mice, (NZB x NZW)F(1) mice do not generate autoantibodies to ribonucleoproteins (RNP) Sm/RNP. In this study, we demonstrate that contrary to previous reports, the autoimmune response directed against Sm/RNP antigens also occurs in NZB x NZW mice. CD4(+) T cells from unprimed 10-week-old NZB x NZW mice proliferate and secrete IL-2 in response to peptide 131-151 of the U1-70K protein, which is known to contain a T(h) epitope recognized by CD4(+) T cells from MRL/Fas(lpr) mice. Peptide 131-151, which was found to bind I-A(k) and I-E(k) class II MHC molecules, also bound both I-A(d) and I-E(d) molecules. This result led us to also re-evaluate longitudinally the anti-Sm/RNP antibody response in NZB x NZW mice. We found that 25-week-old mice do produce antibodies reacting with several small nuclear and heterogeneous nuclear (hn) RNP proteins, such as SmD1, U1-70K and hnRNP A2/B1 proteins. The fine specificity of these antibodies was studied with overlapping synthetic peptides. The same antigenically positive and negative peptides were characterized in MRL/Fas(lpr) and NZB x NZW mice in the three proteins. This new finding can help to understand the mechanisms involved in the development of the anti-Sm/RNP antibody response and, particularly, the role played by non-MHC genes in this autoimmune response.}, keywords = {Animals, Antibody Specificity, B-Lymphocytes, Crosses, Dumortier, fas Receptor, Female, Genetic, Heterogeneous-Nuclear Ribonucleoprotein Group A-B, Heterogeneous-Nuclear Ribonucleoproteins, Histocompatibility Antigens Class II, I2CT, Immunoglobulin G, Inbred MRL lpr, Inbred NZB, Lupus Erythematosus, Mice, Monneaux, Peptide Fragments, Ribonucleoprotein, Ribonucleoproteins, Small Nuclear, Species Specificity, Spliceosomes, Systemic, T-Lymphocytes, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{, title = {From ancient remedies to modern therapeutics: pine bark uses in skin disorders revisited}, author = { B. Rihn and C. Saliou and M. C. Bottin and G. Keith and L. Packer}, year = {2001}, date = {2001-01-01}, journal = {Phytother Res}, volume = {15}, number = {1}, pages = {76-8}, abstract = {The effect of French maritime pine bark extract (PBE) on the gene expression profile of HaCaT human keratinocytes was studied using high density filter arrays. The expression profile of both control and PBE-treated cells was determined. Interestingly, PBE was shown to downregulate both calgranulin A and B genes which are known to be upregulated in psoriasis and various dermatoses. Thus, PBE could be considered in human dermatoses.}, note = {0951-418x Journal Article}, keywords = {*Plants, Cell, Diseases/drug, effects, Expression, Flavonoids/*pharmacology/therapeutic, Gene, Human, Keratinocytes/*drug, Line/drug, Medicinal, Regulation, Skin, therapy/*genetics, use}, pubstate = {published}, tppubtype = {article} } @article{, title = {DNA synthesis fidelity by the reverse transcriptase of the yeast retrotransposon Ty1}, author = { M. Boutabout and M. Wilhelm and F. X. Wilhelm}, year = {2001}, date = {2001-01-01}, journal = {Nucleic Acids Res}, volume = {29}, number = {11}, pages = {2217-22}, abstract = {The fidelity of the yeast retrotransposon Ty1 reverse transcriptase (RT) was determined by an assay based on gel electrophoresis. Steady-state kinetics analyses of deoxyribonucleotide (dNTP) incorporation at a defined primer-template site indicate that Ty1 RT misincorporates dNTP at a frequency of 0.45 x 10(-5) for the A(t):A mispair in which dATP is misincorporated opposite a template A to 6.27 x 10(-5) for the C(t):A mispair. The G(t):G and T(t):T mispairs are formed with very low efficiency. The fidelity parameters of Ty1 RT do not depend on whether RNA or DNA are copied. Relative to lentiviral RTs (HIV-1, HIV-2 or EIAV) Ty1 RT is approximately 10-fold less error prone. Our data also show that the Ty1 RT is able to recapitulate two error-generating mechanisms: extension of mismatches and non-templated addition of nucleotides at the end of a blunt-end primer-template.}, note = {1362-4962 Journal Article}, keywords = {cerevisiae/*genetics/metabolism, DNA, Fungal/genetics, Fungal/genetics/*metabolism, Genetic, Gov't, Kinetics, Non-U.S., Nucleotides/genetics/metabolism, Polymerase/*metabolism, Retroelements/*genetics, RNA, RNA-Directed, Saccharomyces, Support, Templates}, pubstate = {published}, tppubtype = {article} } @article{, title = {Biomedicine. Do G quartets orchestrate fragile X pathology?}, author = { H. Moine and J. L. Mandel}, year = {2001}, date = {2001-01-01}, journal = {Science}, volume = {294}, number = {5551}, pages = {2487-8}, note = {0036-8075 Journal Article}, keywords = {Acid, Analysis, Animals, Array, Binding, Brain/metabolism, Conformation, Crystallography, Expression, Fragile, Gene, Genetic, Human, Messenger/*chemistry/genetics/*metabolism, Mice, Nerve, Nucleic, Oligonucleotide, Protein, Proteins/chemistry/genetics/*metabolism, Regions, Regulation, RNA, Sequence, Sites, structure, Synapses/physiology, Syndrome/genetics/*metabolism, Tertiary, Tissue, Translation, Untranslated, X, X-Ray}, pubstate = {published}, tppubtype = {article} } @article{, title = {Polypurine tract formation by Ty1 RNase H}, author = { M. Wilhelm and O. Uzun and E. H. Mules and A. Gabriel and F. X. Wilhelm}, year = {2001}, date = {2001-01-01}, journal = {J Biol Chem}, volume = {276}, number = {50}, pages = {47695-701}, abstract = {To better understand the mechanism by which Ty1 RNase H creates the polypurine tract (PPT) primer, we have demonstrated the polymerase-dependent hydrolytic activity of Ty1 reverse transcriptase (RT) during minus-strand synthesis. Using RNase H and polymerase mutants of the recombinant Ty1 RT protein, we show that the two domains of Ty1 RT can act independently of one another. Our results indicate that RNA/DNA substrates containing a short RNA PPT, which serve as primers for plus-strand DNA synthesis, are relatively resistant to RNase H cleavage. RNA substrates with a correct 5' end but with 3' end extending beyond the plus-strand initiation site were cleaved specifically to generate the correct 3' end of the PPT. Using long RNA/DNA duplexes containing the PPT, we show that Ty1 RT is able to make specific internal cleavages that could generate the plus-strand primer with correct 5' and 3' ends. Long RNA/DNA duplexes with mutations in the PPT or in a U-rich region upstream of the PPT, which abolish plus-strand initiation in vivo, were not cleaved specifically at the 5' end of the PPT. Our work demonstrates that the in vitro enzyme can recapitulate key processes that control proper replication in vivo.}, note = {0021-9258 Journal Article}, keywords = {*Purines, *Retroelements, Base, Binding, Calf, Data, DNA, DNA/metabolism, Factors, Gov't, H, Hydrolysis, Molecular, Mutation, Non-U.S., P.H.S., Polymerase/*chemistry/*metabolism, Primers/pharmacology, Protein, Proteins/metabolism, Recombinant, Ribonuclease, RNA-Directed, RNA/metabolism, Sequence, Sites, Support, Thymus/*chemistry/*genetics/metabolism, time, U.S.}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reverse transcription of retroviruses and LTR retrotransposons}, author = { M. Wilhelm and F. X. Wilhelm}, year = {2001}, date = {2001-01-01}, journal = {Cell Mol Life Sci}, volume = {58}, number = {9}, pages = {1246-62}, abstract = {Retroelements are mobile genetic entities that replicate via reverse transcription of a template RNA. A key component to the life cycle of these elements is the enzyme reverse transcriptase (RT), which copies the single-stranded genomic RNA of the element into a linear double-stranded DNA that is ultimately integrated into the host genome by the element-encoded integrase. RT is a multifunctionnal enzyme which possesses RNA-dependent and DNA-dependent DNA polymerase activities as well as RNase H activity that specifically degrades the RNA strand of RNA-DNA duplexes. At some stages of the replication a strand-displacement activity of RT is also necessary. All activities are essential for the conversion of single-stranded genomic RNA into the double-stranded preintegrative DNA. This review focuses on the role of RT in the different steps of the replication process of retroelements. The features of retrotransposon replication which differ from the retroviral ones will be emphasized. In a second part of the review, the biochemical and enzymatic properties of two newly characterized retrotransposon RTs will be described. The role of the integrase domain in reverse transcriptase activity of some retroviral and retrotransposon RTs will be discussed.}, note = {1420-682x Journal Article Review Review, Academic}, keywords = {*Retroelements, *Terminal, Acid, Alignment, Amino, Animals, Base, Conformation, Data, DNA, Homology, Human, Molecular, Nucleic, Polymerase/*chemistry/*metabolism, Repeat, Retroviridae/*enzymology/*genetics, RNA-Directed, Sequence, Sequences}, pubstate = {published}, tppubtype = {article} } @article{levashina_conserved_2001, title = {Conserved role of a complement-like protein in phagocytosis revealed by dsRNA knockout in cultured cells of the mosquito, Anopheles gambiae}, author = {Elena A Levashina and L F Moita and Stéphanie A Blandin and G Vriend and Marie Lagueux and F C Kafatos}, issn = {0092-8674}, year = {2001}, date = {2001-01-01}, journal = {Cell}, volume = {104}, number = {5}, pages = {709--718}, abstract = {We characterize a novel hemocyte-specific acute phase glycoprotein from the malaria vector, Anopheles gambiae. It shows substantial structural and functional similarities, including the highly conserved thioester motif, to both a central component of mammalian complement system, factor C3, and to a pan-protease inhibitor, alpha2-macroglobulin. Most importantly, this protein serves as a complement-like opsonin and promotes phagocytosis of some Gram-negative bacteria in a mosquito hemocyte-like cell line. Chemical inactivation by methylamine and depletion by double-stranded RNA knockout demonstrate that this function is dependent on the internal thioester bond. This evidence of a complement-like function in a protostome animal adds substantially to the accumulating evidence of a common ancestry of immune defenses in insects and vertebrates.}, keywords = {alpha-Macroglobulins, Animals, Anopheles, blandin, Cells, Cloning, Complement C3, Cultured, DNA Fragmentation, Double-Stranded, Female, Genetic, Gram-Negative Bacteria, Hemocytes, Insect Proteins, M3i, Molecular, Nucleic Acid Denaturation, Phagocytosis, Protein Structure, RNA, Tertiary, Transcription}, pubstate = {published}, tppubtype = {article} } @article{imler_toll_2001, title = {Toll receptors in innate immunity}, author = {Jean-Luc Imler and Jules A Hoffmann}, issn = {0962-8924}, year = {2001}, date = {2001-01-01}, journal = {Trends in Cell Biology}, volume = {11}, number = {7}, pages = {304--311}, abstract = {Innate immunity is the first-line host defense of multicellular organisms that rapidly operates to limit infection upon exposure to infectious agents. In addition, the cells and molecules operating during this early stage of the immune response in vertebrates have a decisive impact on the shaping of the subsequent adaptive response. Genetic studies initially performed in the fruitfly Drosophila and later in mice have revealed the importance of proteins of the Toll family in the innate immune response. We present here our current understanding of the role of this evolutionary ancient family of proteins that are thought to function as cytokine receptors (Toll in Drosophila) or pattern-recognition receptors (TLRs in mammals) and activate similar, albeit non-identical, signal-transduction pathways in flies and mammals.}, keywords = {Animals, Cell Surface, hoffmann, Humans, imler, Immunity, Immunologic, Innate, M3i, Membrane Glycoproteins, Membrane Proteins, Receptors, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleotides U28-A42 and A37 in unmodified yeast tRNA(Trp) as negative identity elements for bovine tryptophanyl-tRNA synthetase}, author = { D. Carnicelli and M. Brigotti and S. Rizzi and G. Keith and L. Montanaro and S. Sperti}, year = {2001}, date = {2001-01-01}, journal = {FEBS Lett}, volume = {492}, number = {3}, pages = {238-41}, abstract = {Wild-type bovine and yeast tRNA(Trp) are efficiently aminoacylated by tryptophanyl-tRNA synthetase both from beef and from yeast. Upon loss of modified bases in the synthetic transcripts, mammalian tRNA(Trp) retains the double recognition by the two synthetases, while yeast tRNA(Trp) loses its substrate properties for the bovine enzyme and is recognised only by the cognate synthetase. By testing chimeric bovine-yeast transcripts with tryptophanyl-tRNA synthetase purified from beef pancreas, the nucleotides responsible for the loss of charging of the synthetic yeast transcript have been localised in the anticodon arm. A complete loss of charging akin to that observed with the yeast transcript requires substitution in the bovine backbone of G37 in the anticodon loop with yeast A37 and of C28-G42 in the anticodon stem with yeast U28-A42. Since A37 does not prevent aminoacylation of the wild-type yeast tRNA(Trp) by the beef enzyme, a negative combination apparently emerges in the synthetic transcript after unmasking of U28 by loss of pseudourydilation.}, note = {0014-5793 Journal Article}, keywords = {Acid, Adenine/chemistry, Animals, Base, Cattle, cerevisiae/genetics, Conformation, Data, Fungal/genetics/metabolism, Gov't, Kinetics, Ligase/*metabolism, Molecular, Non-U.S., Nucleic, RNA, Saccharomyces, Sequence, Species, Specificity, Substrate, Support, Transfer, Trp/chemistry/*metabolism, Tryptophan-tRNA, Uridine/chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Horseradish peroxidase mediates DNA and deoxyguanosine 3'-monophosphate adduct formation in the presence of ochratoxin A}, author = { S. Obrecht-Pflumio and G. Dirheimer}, year = {2001}, date = {2001-01-01}, journal = {Arch Toxicol}, volume = {75}, number = {10}, pages = {583-590}, abstract = {Ochratoxin A (OTA) gives rise to DNA and deoxyguanosine-3'-monophosphate (dGMP) adducts in vitro using mice kidney microsomes in the presence of arachidonic acid. This result points to the involvement of prostaglandin H synthases, which are present at high levels in the kidney, urinary bladder and seminal vesicles, and/or of lipoxygenases in the metabolic activation of OTA to genotoxic compounds. These enzymes have peroxidase activities. Incubation of OTA with DNA in the presence of horseradish peroxidase (HRP) and cumene hydroperoxide at pH 7.4 led to the formation of one major and three minor adducts with a total adduct level of 42 per 10(9) nucleotides. Incubation with dGMP gave a total adduct level of 159 per 10(9) nucleotides. In the presence of H2O2 instead of cumene hydroperoxide, a lower level of adducts was obtained, both with DNA and dGMP. The concentrations of HRP and co-substrate used in this paper were higher than those used by other authors who obtained negative results when they sought DNA adducts of OTA in the presence of HRP and H2O2. The main adduct we obtained with DNA incubated with HRP and OTA had the same chromatographic behaviour as that obtained when DNA or dGMP were incubated with OTA, arachidonic acid and mice kidney microsomes. However, the main adduct obtained with dGMP incubated with HRP and OTA behaved differently. These results show that OTA can be metabolized by a peroxidase to metabolically activated species that bind covalently to DNA and dGMP; however, the main adduct obtained in vitro with HRP and dGMP cannot serve as a model for one of the adducts formed by OTA with DNA because it behaves differently in two chromatographic systems.}, keywords = {GIEGE}, pubstate = {published}, tppubtype = {article} } @article{furrer_evidence_2001, title = {Evidence of secondary structure by high-resolution magic angle spinning NMR spectroscopy of a bioactive peptide bound to different solid supports}, author = {J Furrer and M Piotto and M Bourdonneau and D Limal and G Guichard and K Elbayed and J Raya and J P Briand and A Bianco}, doi = {10.1021/ja003566w}, issn = {0002-7863}, year = {2001}, date = {2001-01-01}, journal = {Journal of the American Chemical Society}, volume = {123}, number = {18}, pages = {4130--4138}, abstract = {The structure of the 19-amino acid peptide epitope, corresponding to the 141-159 sequence of capsid viral protein VP1 of foot-and-mouth disease virus (FMDV), bound to three different resins, namely, polystyrene-MBHA, PEGA, and POEPOP, has been determined by high-resolution magic angle spinning (HRMAS) NMR spectroscopy. A combination of homonuclear and heteronuclear bidimensional experiments was used for the complete peptide resonance assignment and the qualitative characterization of the peptide folding. The influence of the chemicophysical nature of the different polymers on the secondary structure of the covalently attached FMDV peptide was studied in detail. In the case of polystyrene-MBHA and polyacrylamide-PEGA resins, the analysis of the 2D spectra was hampered by missing signals and extensive overlaps, and only a propensity toward a peptide secondary structure could be derived from the assigned NOE correlations. When the FMDV peptide was linked to the polyoxyethylene-based POEPOP resin, it was found to adopt in dimethylformamide a helical conformation encompassing the C-terminal domain from residues 152 to 159. This conformation is very close to that of the free peptide previously analyzed in 2,2,2-trifluoroethanol. Our study clearly demonstrates that a regular helical structure can be adopted by a resin-bound bioactive peptide. Moreover, a change in the folding was observed when the same peptide-POEPOP conjugate was swollen in aqueous solution, displaying the same conformational features as the free peptide in water. The possibility of studying solid-supported ordered secondary structures by the HRMAS NMR technique in a wide range of solvents can be extended either to other biologically relevant peptides and proteins or to new synthetic oligomers.}, keywords = {Amino Acid Sequence, biomolecular, Capsid, Capsid Proteins, Epitopes, I2CT, Molecular Sequence Data, Nuclear Magnetic Resonance, Peptide Fragments, Plant, Protein Structure, Resins, Secondary, Solvents, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{casimir_conformational_2000, title = {Conformational restriction of the Tyr53 side-chain in the decapeptide HE}, author = {J R Casimir and K Iterbeke and W Van Den Nest and M C Trescol-Biémont and H Dumortier and S Muller and D Gerlier and C Rabourdin-Combe and D Tourwé and J Paris}, doi = {10.1034/j.1399-3011.2000.00777.x}, issn = {1397-002X}, year = {2000}, date = {2000-12-01}, journal = {The Journal of Peptide Research: Official Journal of the American Peptide Society}, volume = {56}, number = {6}, pages = {398--408}, abstract = {A series of conformationally restricted analogs of the hen egg lysozyme (HEL) decapeptide 52-61 in which the conformationally flexible Tyr53 residue was replaced by several more constrained tyrosine and phenylalanine analogs was prepared. Among these tyrosine and phenylalanine analogs were 1,2,3,4-tetrahydro-7-hydroxyisoquinoline-3-carboxylic acid (Htc), 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid (Tic), 4-amino- 1,2,4,5-tetrahydro-8-hydroxy-2-benzazepine-3-one (Hba), 4-amino-1,2,4,5-tetrahydro-2-benzazepine-3-one (Aba), 2-amino-6-hydroxytetralin-2-carboxylic acid (Hat) and 2-amino-5-hydroxyindan-2-carboxylic acid (Hai) in which the rotations around Calpha-Cbeta and Cbeta-Cgamma were restricted because of cyclization of the side-chain to the backbone. Synthesis of Pht-Hba-Gly-OH using a modification of the Flynn and de Laszlo procedure is described. Analogs of beta-methyltyrosine (beta-MeTyr) in which the side-chains were biased to particular side-chain torsional angles because of substitution at the beta-hydrogens were also prepared. These analogs of HEL[52-61] peptide were tested for their ability to bind to the major histocompatibility complex class II I-Ak molecule and to be recognized in this context by two T-cell hybridomas, specific for the parent peptide HEL[52-61]. The data showed that the conformation and also the configuration of the Tyr53 residue influenced both the binding of the peptide to I-Ak and the recognition of the peptide/I-Ak complex by a T-cell receptor.}, keywords = {Amino Acid Sequence, Animals, Antigen, Antigen-Presenting Cells, B-Lymphocytes, Chemical, Chickens, Dumortier, I2CT, Major Histocompatibility Complex, Mice, Models, Molecular Sequence Data, Muramidase, Peptide Biosynthesis, Peptides, Phenylalanine, Protein Binding, Protein Conformation, Receptors, T-Cell, Team-Dumortier, Temperature, Tyrosine}, pubstate = {published}, tppubtype = {article} } @article{bianco_multistep_2000, title = {Multistep Synthesis of 2,5-Diketopiperazines on Different Solid Supports Monitored by High Resolution Magic Angle Spinning NMR Spectroscopy}, author = {Alberto Bianco and Julien Furrer and David Limal and Gilles Guichard and Karim Elbayed and Jésus Raya and Martial Piotto and Jean-Paul Briand}, url = {https://doi.org/10.1021/cc0000489}, doi = {10.1021/cc0000489}, issn = {1520-4766}, year = {2000}, date = {2000-11-01}, urldate = {2020-03-31}, journal = {Journal of Combinatorial Chemistry}, volume = {2}, number = {6}, pages = {681--690}, abstract = {The solid-phase synthesis of 2,5-diketopiperazines containing the trans-4-hydroxy-l-proline amino acid residue (Hyp) was performed on Ellman polystyrene, polyoxyethylene-polyoxypropylene (POEPOP), polystyrene-polyoxyethylene NovaSyn, and Wang resins, respectively. The reaction pathway allowed the introduction of different functional groups around the bicyclic scaffold in a combinatorial approach, and it generated mixtures of isomers. A detailed characterization of the single reaction steps by high resolution magic angle spinning (HRMAS) NMR spectroscopy was performed. The NMR spectral resolution of the resin-bound intermediates and final products was greatly influenced by the polymer matrix. The POEPOP resin permitted to obtain HRMAS NMR spectra with a resolution comparable with that of the spectra of the molecules in solution. Moreover, configurational and conformational isomers formed during the solid-phase reaction steps could be detected and easily assigned. Therefore, the combination of the HRMAS NMR technique with the use of nonaromatic resins may become an extremely powerful tool in solid-phase organic synthesis. This approach will allow the monitoring of multistep reactions and the conception of on-bead structural studies either on small molecules or on natural and/or synthetic oligomers.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{monneaux_laboratory_2000, title = {Laboratory protocols for the identification of Th cell epitopes on self-antigens in mice with systemic autoimmune diseases}, author = {F Monneaux and S Muller}, doi = {10.1016/s0022-1759(00)00256-8}, issn = {0022-1759}, year = {2000}, date = {2000-10-01}, journal = {Journal of Immunological Methods}, volume = {244}, number = {1-2}, pages = {195--204}, abstract = {T cells play a critical role in both the immunological and clinical manifestations of systemic autoimmune diseases such as systemic lupus erythematosus (SLE). Although in normal mice multiple T cell epitopes have been characterized in several self-proteins, there is little information on the fine specificity of autoreactive T cells in lupus model mice and humans. In SLE-prone mice and humans, the only Th cell epitopes identified at the molecular level in self-antigens concern histones and nucleosomes, and the 70-kD U1-snRNP protein. T cell characterization in certain autoimmune mice such as MRL lpr/lpr and NZB/NZW mice has been largely impaired by their hyporesponsiveness in response to mitogen and minimal IL-2 secretion. In addition, MRL lpr/lpr mice also develop lymphadenopathy characterized by the progressive accumulation of functionally immature CD4(-) CD8(-) T cells. It is therefore important to optimize the methods used to measure T cell proliferation and cytokine production ex vivo in order to identify minimal activation in the presence of appropriate antigen. The protocol described in this article has been used for identifying in young MRL lpr/lpr and NZB/NZW mice a CD4(+) T cell epitope in the murine 70-kD U1-RNP protein.}, keywords = {Animals, Antigen Presentation, Antigen-Presenting Cells, Autoantigens, B-Lymphocytes, Coculture Techniques, Epitopes, Female, Flow Cytometry, I2CT, Inbred MRL lpr, Inbred NZB, Lupus Erythematosus, Lymphocyte Activation, Mice, Monneaux, Ribonucleoproteins, Small Nuclear, Systemic, T-Lymphocyte, Team-Dumortier, Th1 Cells, Th2 Cells}, pubstate = {published}, tppubtype = {article} } @article{lagueux_constitutive_2000, title = {Constitutive expression of a complement-like protein in toll and JAK gain-of-function mutants of Drosophila}, author = {Marie Lagueux and E Perrodou and Elena A Levashina and Maria Capovilla and Jules A Hoffmann}, doi = {10.1073/pnas.97.21.11427}, issn = {0027-8424}, year = {2000}, date = {2000-10-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {97}, number = {21}, pages = {11427--11432}, abstract = {We show that Drosophila expresses four genes encoding proteins with significant similarities with the thiolester-containing proteins of the complement C3/alpha(2)-macroglobulin superfamily. The genes are transcribed at a low level during all stages of development, and their expression is markedly up-regulated after an immune challenge. For one of these genes, which is predominantly expressed in the larval fat body, we observe a constitutive expression in gain-of-function mutants of the Janus kinase (JAK) hop and a reduced inducibility in loss-of-function hop mutants. We also observe a constitutive expression in gain-of-function Toll mutants. We discuss the possible roles of these novel complement-like proteins in the Drosophila host defense.}, keywords = {alpha-Macroglobulins, Amino Acid, Animals, Cell Surface, Complement C3, Esters, Genetic, hoffmann, Insect Proteins, Janus Kinases, M3i, Membrane Glycoproteins, Mutation, Protein-Tyrosine Kinases, Proteins, Receptors, Sequence Homology, Sulfhydryl Compounds, Toll-Like Receptors, Transcription, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{rutschmann_role_2000, title = {Role of Drosophila IKK gamma in a toll-independent antibacterial immune response}, author = {Sophie Rutschmann and Alain C Jung and R Zhou and N Silverman and Jules A Hoffmann and Dominique Ferrandon}, doi = {10.1038/79801}, issn = {1529-2908}, year = {2000}, date = {2000-10-01}, journal = {Nat. Immunol.}, volume = {1}, number = {4}, pages = {342--347}, abstract = {We have generated, by ethylmethane sulfonate mutagenesis, loss-of-function mutants in the Drosophila homolog of the mammalian I-kappa B kinase (IKK) complex component IKK gamma (also called NEMO). Our data show that Drosophila IKK gamma is required for the Relish-dependent immune induction of the genes encoding antibacterial peptides and for resistance to infections by Escherichia coli. However, it is not required for the Toll-DIF-dependent antifungal host defense. The results indicate distinct control mechanisms of the Rel-like transactivators DIF and Relish in the Drosophila innate immune response and show that Drosophila Toll does not signal through a IKK gamma-dependent signaling complex. Thus, in contrast to the vertebrate inflammatory response, IKK gamma is required for the activation of only one immune signaling pathway in Drosophila.}, keywords = {Animals, Antigens, Bacterial, Cell Surface, ferrandon, Gene Expression Regulation, hoffmann, I-kappa B Kinase, Immunity, Innate, Insect Proteins, M3i, Membrane Glycoproteins, Protein-Serine-Threonine Kinases, Receptors, Signal Transduction, Toll-Like Receptors, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{tauszig_toll-related_2000, title = {Toll-related receptors and the control of antimicrobial peptide expression in Drosophila}, author = {Servane Tauszig and Emmanuelle Jouanguy and Jules A Hoffmann and Jean-Luc Imler}, doi = {10.1073/pnas.180130797}, issn = {0027-8424}, year = {2000}, date = {2000-09-01}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {97}, number = {19}, pages = {10520--10525}, abstract = {Insects defend themselves against infectious microorganisms by synthesizing potent antimicrobial peptides. Drosophila has appeared in recent years as a favorable model to study this innate host defense. A genetic analysis of the regulation of the antifungal peptide drosomycin has demonstrated a key role for the transmembrane receptor Toll, which prompted the search for mammalian homologs. Two of these, Toll-like receptor (TLR)2 and TLR4, recently were shown to play a critical role in innate immunity against bacteria. Here we describe six additional Toll-related genes (Toll-3 to Toll-8) in Drosophila in addition to 18-wheeler. Two of these genes, Toll-3 and Toll-4, are expressed at a low level. Toll-6, -7, and -8, on the other hand, are expressed at high levels during embryogenesis and molting, suggesting that, like Toll and 18w, they perform developmental functions. Finally, Toll-5 is expressed only in larvae and adults. By using chimeric constructs, we have tested the capacity of the signaling Toll/IL-1R homology domains of these receptors to activate antimicrobial peptide promoters and found that only Toll and Toll-5 can activate the drosomycin promoter in transfected cells, thus demonstrating specificity at the level of the Toll/IL-1R homology domain. In contrast, none of these constructs activated antibacterial peptide promoters, suggesting that Toll-related receptors are not involved in the regulation of antibacterial peptide expression. This result was independently confirmed by the demonstration that a dominant-negative version of the kinase Pelle can block induction of drosomycin by the cytokine Spaetzle, but does not affect induction of the antibacterial peptide attacin by lipopolysaccharide.}, keywords = {Amino Acid, Animals, Anti-Bacterial Agents, Blotting, Cell Line, Cell Surface, hoffmann, imler, M3i, Membrane Glycoproteins, Multigene Family, Northern, Peptides, Receptors, Reverse Transcriptase Polymerase Chain Reaction, Sequence Homology, Toll-Like Receptor 2, Toll-Like Receptor 4, Toll-Like Receptor 5, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{dumortier_b_2000, title = {B and Ŧ cell responses to the spliceosomal heterogeneous nuclear ribonucleoproteins A2 and B1 in normal and lupus mice}, author = {H Dumortier and F Monneaux and B Jahn-Schmid and J P Briand and K Skriner and P L Cohen and J S Smolen and G Steiner and S Muller}, doi = {10.4049/jimmunol.165.4.2297}, issn = {0022-1767}, year = {2000}, date = {2000-08-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {165}, number = {4}, pages = {2297--2305}, abstract = {Autoantibodies directed against spliceosomal heterogeneous nuclear ribonucleoproteins (hnRNPs) are a typical feature of rheumatoid arthritis, systemic lupus erythematosus, and mixed-connective tissue disease. With the aim of investigating a potential pathogenic role of these Abs, we have studied the Ab response to A2/B1 hnRNPs in different murine models of lupus. The specificity of anti-A2/B1 Abs was tested with a series of 14 overlapping synthetic peptides covering the region 1-206 of A2 that contains most of the epitopes recognized by patients' Abs. A major epitope recognized very early during the course of the disease by Abs from most of MRL lpr/lpr mice but not from other lupus mice and from mice of different MHC haplotypes immunized against B1 was identified in residues 50-70. This peptide contains a highly conserved sequence RGFGFVTF also present in other hnRNPs and small nuclear ribonucleoproteins. Abs reacting with a second A2 epitope identified in residues 35-55 were detectable several weeks later, suggesting an intramolecular B cell epitope spreading during the course of the disease. We identified several T cell epitopes within the region 35-175 that generated an effective Th cell response with IL-2 and IFN-gamma secretion in nonautoimmune CBA/J mice sharing the same MHC haplotype H-2k as MRL/lpr mice. None of the peptides stimulated T cells primed in vivo with B1. Because Abs to peptide 50-70 were detected significantly earlier than Abs reacting with other A2 peptides and the protein itself, it is possible that within the protein, this segment contains residues playing an initiator role in the induction of the anti-A2/B1 and antispliceosome Ab response.}, keywords = {Amino Acid Sequence, Animals, Autoantibodies, B-Lymphocytes, Dumortier, Epitope Mapping, Female, Heterogeneous Nuclear, Heterogeneous-Nuclear Ribonucleoprotein Group A-B, Heterogeneous-Nuclear Ribonucleoproteins, Humans, I2CT, Immunoglobulin G, Inbred BALB C, Inbred C57BL, Inbred CBA, Inbred MRL lpr, Injections, Lupus Nephritis, Lymphocyte Activation, Male, Mice, Molecular Sequence Data, Monneaux, Peptide Fragments, Recombinant Proteins, Ribonucleoproteins, RNA, Spliceosomes, Subcutaneous, T-Lymphocytes, Team-Dumortier, transgenic}, pubstate = {published}, tppubtype = {article} } @article{monneaux_b_2000, title = {B and Ŧ cell immune response to small nuclear ribonucleoprotein particles in lupus mice: autoreactive CD4(+) Ŧ cells recognize a Ŧ cell epitope located within the RNP80 motif of the 70K protein}, author = {F Monneaux and J P Briand and S Muller}, doi = {10.1002/1521-4141(2000)30:8<2191::AID-IMMU2191>3.0.CO;2-R}, issn = {0014-2980}, year = {2000}, date = {2000-08-01}, journal = {European Journal of Immunology}, volume = {30}, number = {8}, pages = {2191--2200}, abstract = {Systemic lupus erythematosus is characterized by the presence of high titers of autoantibodies reacting with various components of the U1 small nuclear ribonucleoprotein particle (snRNP). It has been suggested that these antibodies are produced by an antigen-driven mechanism under the dependence of antigen-specific T cells. To investigate the role of T cell help in this process, we sought, with 20 overlapping peptides, the Th epitopes on the U1-70K snRNP in unprimed H-2(k) MRL / lpr lupus mice and immunized CBA normal mice. The peptide 131 - 151 was recognized by both IgG autoantibodies and CD4(+) T cells from 7 - 9-week-old MRL / lpr mice. In this test, antigen-presenting cells (APC) from MRL / lpr mice were required; APC from naive CBA mice failed to stimulate CD4(+) cells from MRL / lpr mice. The potential role of MRL / lpr B cells as APC, the expression of MHC class II molecules at their surface and their activation state (expression of CD69, CD80 / B7-1 and CD86 / B7-2 molecules) were studied. Peptide 131 - 151 bound both I-A(k) and I-E(k) class II molecules and favored an IL-2-positive T cell response but not IFN-gamma, IL-6 and IL-10 secretion. Segment 131 - 151 is localized within the RNP80 motif and contains residues that are highly conserved in many nuclear, nucleolar and cytoplasmic RNA binding proteins.}, keywords = {Amino Acid Motifs, Animals, Antigen-Presenting Cells, Autoimmunity, B-Lymphocytes, CD4-Positive T-Lymphocytes, Epitopes, Female, I2CT, Inbred BALB C, Inbred CBA, Inbred MRL lpr, Lupus Vulgaris, Lymphocyte Activation, Mice, Monneaux, Peptide Fragments, Ribonucleoprotein, T-Lymphocyte, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{adamkewicz_purification_2000, title = {Purification and enzymic properties of Mot1 ATPase, a regulator of basal transcription in the yeast Saccharomyces cerevisiae}, author = {J I Adamkewicz and C G Mueller and K E Hansen and W A Prud'homme and J Thorner}, doi = {10.1074/jbc.M002639200}, issn = {0021-9258}, year = {2000}, date = {2000-07-01}, journal = {The Journal of Biological Chemistry}, volume = {275}, number = {28}, pages = {21158--21168}, abstract = {The 1867-residue Mot1 protein is a member of a superfamily of ATPases, some of which are helicases, that interact with protein-nucleic acid assemblies. Mot1 is an essential regulator of RNA polymerase II-dependent transcription in vivo and dissociates TATA box-binding protein (TBP)-DNA complexes in vitro. Mot1-(His)(6) was purified to apparent homogeneity from yeast extracts. The preparation efficiently dissociated TBP.TATA complexes, suggesting that no other protein or cofactor is required. Mot1 behaved as a non-globular monomer in hydrodynamic studies, and no association was detected between differentially tagged co-expressed Mot1 constructs. ATPase activity was stimulated about 10-fold by high ionic strength or alkaline pH, or by deletion of the N-terminal TBP-binding segment, suggesting that the N-terminal domain negatively regulates the C-terminal ATPase domain (Mot1C). Correspondingly, at moderate salt concentration, Mot1 ATPase (but not Mot1C) was stimulated textgreater/=10-fold by yeast TBP, suggesting that interaction with TBP relieves a conformational constraint in Mot1. Double- or single-stranded TATA-containing DNA did not affect ATPase activity of Mot1 or Mot1C, with or without TBP. Mot1 did not exhibit detectable helicase activity in strand displacement assays using substrates with flush ends or 5'- or 3'-overhangs. Mot1-catalyzed dissociation of TBP from DNA was not prevented by a psoralen cross-link positioned immediately preceding the TATA sequence. Thus, Mot1 most likely promotes release of TBP from TATA-containing DNA by causing a structural change in TBP itself, rather than by strand unwinding.}, keywords = {Adenosine Triphosphatases, Base Sequence, Chromatography, DNA Helicases, DNA-Binding Proteins, Fungal, Gel, Gene Expression Regulation, Genetic, Kinetics, Molecular Sequence Data, Molecular Weight, Osmolar Concentration, Recombinant Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, TATA Box, TATA-Binding Protein Associated Factors, TATA-Box Binding Protein, Team-Mueller, Transcription, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{rutschmann_rel_2000, title = {The Rel protein DIF mediates the antifungal but not the antibacterial host defense in Drosophila}, author = {Sophie Rutschmann and Alain C Jung and Charles Hetru and Jean-Marc Reichhart and Jules A Hoffmann and Dominique Ferrandon}, issn = {1074-7613}, year = {2000}, date = {2000-05-01}, journal = {Immunity}, volume = {12}, number = {5}, pages = {569--580}, abstract = {We have isolated two Drosophila lines that carry point mutations in the gene coding for the NF-KB-like factor DIF. Like mutants of the Toll pathway, Dif mutant flies are susceptible to fungal but not to bacterial infections. Genetic epistasis experiments demonstrate that Dif mediates the Toll-dependent control of the inducibility of the antifungal peptide gene Drosomycin. Strikingly, DIF alone is required for the antifungal response in adults, but is redundant in larvae with Dorsal, another Rel family member. In Drosophila, Dif appears to be dedicated to the antifungal defense elicited by fungi and gram-positive bacteria. We discuss in this light the possibility that NF-KB1/p50 might be required more specifically in the innate immune response against gram-positive bacteria in mammals.}, keywords = {Animals, Antigens, Bacterial, DNA-Binding Proteins, ferrandon, Fungal, hoffmann, Immunity, Innate, M3i, reichhart, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{bianco_solid-phase_2000, title = {Solid-Phase Synthesis and Structural Characterization of Highly Substituted Hydroxyproline-Based 2,5-Diketopiperazines}, author = {Alberto Bianco and Carsten P Sonksen and Peter Roepstorff and Jean-Paul Briand}, url = {https://doi.org/10.1021/jo991818+}, doi = {10.1021/jo991818+}, issn = {0022-3263}, year = {2000}, date = {2000-04-01}, urldate = {2020-03-31}, journal = {The Journal of Organic Chemistry}, volume = {65}, number = {7}, pages = {2179--2187}, abstract = {Two general solid-phase methods for the synthesis of a new class of 2,5-diketopiperazines (DKPs) containing the trans-4-hydroxy-l-proline amino acid residue (Hyp) have been developed. An N-protected hydroxyproline methyl ester was linked through the hydroxyl function to the Ellman resin. The synthesis procedures were conceived to enable a sequence of Hyp alkylation, Hyp N-acylation, cyclization, and amide bond alkylation. Up to three different centers of molecular diversity were introduced around the DKP scaffold. Highly functionalized bicyclic compounds were obtained in good yield and purity. The alkylation of hydroxyproline αCH was performed without control of the diastereoselectivity. During the final alkylation of the backbone, amide bond epimerization at the α-carbon atoms of the two amino acid residues was observed. The structures of representative DKPs were elucidated with multidimensional NMR experiments. The described reaction pathways can be applied to the identification of heterocyclic molecule inhibitors to diverse enzyme targets.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{nisole_hb-19_2000, title = {The HB-19 pseudopeptide 5[Kψ(CH2N)PR]-TASP inhibits attachment of Ŧ lymphocyte- and macrophage-tropic HIV to permissive cells}, author = {Sébastien Nisole and Bernard Krust and Elisabeth Dam and Alberto Bianco and Nabila Seddiki and Solen Loaec and Christian Callebaut and Gilles Guichard and Sylviane Muller and J Briand and Ara Hovanessian}, doi = {10.1089/088922200309331}, year = {2000}, date = {2000-03-01}, journal = {AIDS research and human retroviruses}, volume = {16}, pages = {237--49}, abstract = {The HB-19 pseudopeptide 5[Kpsi(CH2N)PR]-TASP[psi(CH2N) indicating a reduced peptide bond], which binds the cell surface-expressed nucleolin, is a potent inhibitor of HIV infection. Here, by using primary T lymphocyte cultures and an experimental cell model to monitor HIV entry, we show that HB-19 inhibits in a dose-dependent manner both T lymphocyte- and macrophage-tropic HIV isolates. Similar positively charged control pseudopeptides have no effect on HIV infection even at high concentrations. These observations, and the fact that HB-19 has no effect on SIV-mac and HIV-1 pseudotyped with VSV envelope glycoproteins, confirm the specific nature of this inhibitor against the entry process mediated by the HIV envelope glycoproteins. Finally, association of low doses of HB-19 with beta-chemokines or AZT results in an increased inhibitory effect on HIV infection. HB-19 has no inhibitory effect when added to cells a few hours after HIV entry. On the other hand, in HB-19-pretreated cells, the inhibitory effect persists for several hours, even after washing cells to remove away the unbound pseudopeptide. Under such conditions, the attachment of HIV particles to cells is inhibited as efficiently as by neutralizing monoclonal antibodies directed against the V3 loop. In view of its specific mode of action on various HIV isolates, HB-19 represents a potential anti-HIV drug.}, keywords = {I2CT, Team-Bianco}, pubstate = {published}, tppubtype = {article} } @article{basset_phytopathogenic_2000, title = {The phytopathogenic bacteria Erwinia carotovora infects Drosophila and activates an immune response}, author = {A Basset and R S Khush and A Braun and L Gardan and F Boccard and Jules A Hoffmann and Bruno Lemaitre}, doi = {10.1073/pnas.070357597}, issn = {0027-8424}, year = {2000}, date = {2000-03-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {97}, number = {7}, pages = {3376--3381}, abstract = {Although Drosophila possesses potent immune responses, little is known about the microbial pathogens that infect Drosophila. We have identified members of the bacterial genus Erwinia that induce the systemic expression of genes encoding antimicrobial peptides in Drosophila larvae after ingestion. These Erwinia strains are phytopathogens and use flies as vectors; our data suggest that these strains have also evolved mechanisms for exploiting their insect vectors as hosts. Erwinia infections induce an antimicrobial response in Drosophila larvae with a preferential expression of antibacterial versus antifungal peptide-encoding genes. Antibacterial peptide gene expression after Erwinia infection is reduced in two Drosophila mutants that have reduced numbers of hemocytes, suggesting that blood cells play a role in regulating Drosophila antimicrobial responses and also illustrating that this Drosophila-Erwinia interaction provides a powerful model for dissecting host-pathogen relationships.}, keywords = {Animals, Bacterial, Gene Expression Regulation, Genetically Modified, hoffmann, Insect Proteins, Larva, M3i, Pectobacterium carotovorum}, pubstate = {published}, tppubtype = {article} } @article{imler_signaling_2000, title = {Signaling mechanisms in the antimicrobial host defense of Drosophila}, author = {Jean-Luc Imler and Jules A Hoffmann}, issn = {1369-5274}, year = {2000}, date = {2000-02-01}, journal = {Current Opinion in Microbiology}, volume = {3}, number = {1}, pages = {16--22}, abstract = {Drosophila has appeared in recent years as a powerful model to study innate immunity. Several papers published in the past year shed light on the role of the three Rel proteins Dorsal, Dif and Relish in the regulation of antimicrobial peptide expression. In addition, the discovery that a blood serine protease inhibitor is involved in the control of the antifungal response indicates that Toll is activated upon triggering of a proteolytic cascade and does not function as a Drosophila pattern recognition receptor.}, keywords = {Animals, Anti-Infective Agents, Cell Surface, Gene Expression Regulation, Genes, hoffmann, imler, Insect, Insect Proteins, M3i, Membrane Glycoproteins, Receptors, Signal Transduction, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization and preliminary X-ray crystallographic analysis of yeast arginyl-tRNA synthetase-yeast tRNAArg complexes}, author = { B. Delagoutte and G. Keith and D. Moras and J. Cavarelli}, year = {2000}, date = {2000-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {56}, number = {Pt 4}, pages = {492-4}, abstract = {Three different crystal forms of complexes between arginyl-tRNA synthetase from the yeast Saccharomyces cerevisae (yArgRS) and the yeast second major tRNA(Arg) (tRNA(Arg)(ICG)) isoacceptor have been crystallized by the hanging-drop vapour-diffusion method in the presence of ammonium sulfate. Crystal form II, which diffracts beyond 2.2 A resolution at the European Synchrotron Radiation Facility ID14-4 beamline, belongs to the orthorhombic space group P2(1)2(1)2, with unit-cell parameters a = 129.64}, note = {0907-4449 Journal Article}, keywords = {&, Arg/*chemistry/isolation, Arginine-tRNA, cerevisiae/enzymology/genetics, Crystallization, Crystallography, Fungal/chemistry/isolation, Gov't, Ligase/*chemistry/isolation, Non-U.S., purification/*metabolism, purification/metabolism, RNA, Saccharomyces, Support, Transfer, X-Ray}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Overall genomic DNA methylation in relation to estrogen]}, author = { K. Postawski and E. Olech-Fudali and J. A. Jakowicki and E. Korobowicz and G. Keith and W. Baranowski}, year = {2000}, date = {2000-01-01}, journal = {Ginekol Pol}, volume = {71}, number = {9}, pages = {1206-11}, abstract = {Overall genomic DNA methylation was analyzed using enzymatic digestion into nucleotides, 32P postlabeling, two-dimensional thin-layer chromatography on cellulose plates and phosphobioimaging quantitation, in relation to immunohistochemically measured estrogen (ER) and progesterone receptor (PR) status of 15 uterine cancers. Mean 5-methyldeoxycytosine (m5dC) content did not differ between ER-positive and ER-negative neoplasms. Highest values of m5dC were noted both in ER-negative and ER-positive tumors. Additionally, there was no low DNA methylation in ER negative uterine cancer tissues. Decrease of the overall genomic DNA methylation could be related to the increase of ER/PR ratio, however it was not significant in our investigation. The potential role of steroid receptors status in uterine cancer tissue is discussed.}, note = {0017-0011 Journal Article}, keywords = {80, Abstract, Aged, and, Biopsy, DNA, English, Estrogen/*metabolism, Female, Gov't, Human, Methylases/metabolism, Methylation, Middle, modification, Neoplasms/*metabolism/*pathology, Non-U.S., over, Progesterone/*metabolism, Receptors, Support, Uterine, Uterus/*metabolism/pathology}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Chemical ribonucleases. 2. Design and hydrolytic properties RNase mimetics based on diazabicyclo[2.2.2]octane with various positive charges]}, author = {M A Zenkova and A V Vlasov and D A Konevets and V N Sil'nikov and R Giege and V V Vlasov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11036527}, isbn = {11036527}, year = {2000}, date = {2000-01-01}, journal = {Bioorg Khim}, volume = {26}, number = {9}, pages = {679-685}, abstract = {A procedure was proposed allowing one to synthesize RNA mimics on the basis of conjugates of diazabicyclo[2.2.2]octane with imidazole bearing a varying number of positive charges (nDm series, where n is the number of positive charges at neutral pH, m is the code of an imidazole-containing fragment of the catalytic domain: 1, histamine; 2, histidine methyl ester). The hydrolytic activity of six compounds of this series was studied under physiological conditions using in vitro transcript of human mitochondrial tRNA(Lys) as a substrate. It was shown that the rate of RNA hydrolysis with nDm conjugates rises with an increase in the number of positive charges: an approximately 30-fold acceleration of hydrolysis was observed with an increase in the total charge of the construct from +2 to +4.}, note = {0132-3423 Journal Article}, keywords = {Bicyclo Compounds, Heterocyclic/*chemical synthesis/chemistry Catalysis Cations, Lys/chemistry/genetics Ribonucleases/*chemistry Structure-Activity Relationship, Monovalent/chemistry Drug Design English Abstract Human Hydrolysis Imidazoles/*chemical synthesis/chemistry Magnesium/chemistry Mitochondria/chemistry Molecular Mimicry Point Mutation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Evaluation of uranyl photocleavage as a probe to monitor ion binding and flexibility in RNAs}, author = {D Wittberger and C Berens and C Hammann and E Westhof and R Schroeder}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10873469}, isbn = {10873469}, year = {2000}, date = {2000-01-01}, journal = {J Mol Biol}, volume = {300}, number = {2}, pages = {339-352}, abstract = {In order to evaluate uranyl photocleavage as a tool to identify and characterize structural and dynamic properties in RNA, we compared uranyl cleavage sites in five RNA molecules with known X-ray structures, namely the hammerhead and hepatitis delta virus ribozymes, the P4-P6 domain of the Tetrahymena group I intron, as well as tRNA(Phe) and tRNA(Asp) from yeast. Uranyl photocleavage was observed at specific positions in all molecules investigated. In order to characterize the sites, photocleavage was performed in the absence and in increasing amounts of MgCl(2). Uranyl photocleavage correlates well with sites of low calculated accessibility, suggesting that uranyl ions bind in tight RNA pockets formed by close approach of phosphate groups. RNA foldings require ion binding, usually magnesium ions. Thus, upon the adoption of the native structure, uranyl ions can no longer bind well except in flexible and open to the solvent regions that can undergo induced-fit without disrupting the native fold. Uranyl photocleavage was compared to N-ethyl-N-nitrosourea and lead-induced cleavages in the context of the three-dimensional X-ray structures. Overall, the regions protected from ENU attack are sites of uranyl cleavage, indicating sites of low accessibility which can form ion binding sites. On the contrary, lead cleavages occur at flexible and accessible sites and correlate with the unspecific cleavages prevalent in dynamic and open regions. Applied in a magnesium-dependent manner, and only in combination with other backbone probing agents such as N-ethyl-N-nitrosourea, lead and Fenton cleavage, uranyl probing has the potential to reveal high-affinity metal ion environments, as well as regions involved in conformational transitions.}, note = {0022-2836 Journal Article}, keywords = {Animals Base Pairing Base Sequence Ethylnitrosourea/metabolism Hepatitis Delta Virus/genetics Hydrogen Peroxide/metabolism Introns/genetics Ions/metabolism Iron/metabolism Lead/metabolism Magnesium Chloride/pharmacology Models, Asp/chemistry/genetics/metabolism RNA, Catalytic/chemistry/genetics/metabolism RNA, Fungal/chemistry/genetics/metabolism RNA, Molecular Molecular Sequence Data *Nucleic Acid Conformation *Photolysis/drug effects Pliability RNA/*chemistry/genetics/*metabolism RNA, Non-U.S. Gov't Tetrahymena/genetics Uranyl Nitrate/*metabolism Yeasts/genetics, Phe/chemistry/genetics/metabolism RNA, Protozoan/chemistry/genetics/metabolism RNA, Transfer, Unité ARN, Viral/chemistry/genetics/metabolism Solvents Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Expression of an active form of recombinant Ty1 reverse transcriptase in Escherichia coli: a fusion protein containing the C-terminal region of the Ty1 integrase linked to the reverse transcriptase-RNase H domain exhibits polymerase and RNase H activities}, author = {M Wilhelm and M Boutabout and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10816427}, isbn = {10816427}, year = {2000}, date = {2000-01-01}, journal = {Biochem J}, volume = {348}, number = {Pt 2}, pages = {337-342}, abstract = {Replication of the Saccharomyces cerevisiae Ty1 retrotransposon requires a reverse transcriptase capable of synthesizing Ty1 DNA. The first description of an active form of a recombinant Ty1 enzyme with polymerase and RNase H activities is reported here. The Ty1 enzyme was expressed as a hexahistidine-tagged fusion protein in Escherichia coli to facilitate purification of the recombinant protein by metal-chelate chromatography. Catalytic activity of the recombinant protein was detected only when amino acid residues encoded by the integrase gene were added to the N-terminus of the reverse transcriptase-RNase H domain. This suggests that the integrase domain could play a role in proper folding of reverse transcriptase. Several biochemical properties of the Ty1 enzyme were analysed, including the effect of MgCl(2), NaCl, temperature and of the chain terminator dideoxy GTP on its polymerase activity. RNase H activity was examined by monitoring the cleavage of a RNA-DNA template-primer. Our results suggest that the distance between the RNase H and polymerase active sites corresponds to the length of a 14-nucleotide RNA-DNA heteroduplex. The recombinant protein produced in E. coli should be useful for further biochemical and structural analyses and for a better understanding of the role of integrase in the activation of reverse transcriptase.}, note = {0264-6021 Journal Article}, keywords = {Affinity Cloning, Amino Acid Support, Amino Acid Sequence Chromatography, Calf Thymus/isolation & purification/*metabolism Saccharomyces cerevisiae/*enzymology/*genetics Sequence Alignment Sequence Homology, Genetic, Molecular Codon, Non-U.S. Gov't Templates, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selection of viral RNA-derived tRNA-like structures with improved valylation activities}, author = {J Wientges and J Putz and R Giege and C Florentz and A Schwienhorst}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10821696}, isbn = {10821696}, year = {2000}, date = {2000-01-01}, journal = {Biochemistry}, volume = {39}, number = {20}, pages = {6207-6218}, abstract = {The tRNA-like structure (TLS) of turnip yellow mosaic virus (TYMV) RNA was previously shown to be efficiently charged by yeast valyl-tRNA synthetase (ValRS). This RNA has a noncanonical structure at its 3'-terminus but mimics a tRNA L-shaped fold, including an anticodon loop containing the major identity nucleotides for valylation, and a pseudoknotted amino acid accepting domain. Here we describe an in vitro selection experiment aimed (i) to verify the completeness of the valine identity set, (ii) to elucidate the impact of the pseudoknot on valylation, and (iii) to investigate whether functional communication exists between the two distal anticodon and amino acid accepting domains. Valylatable variants were selected from a pool of 2 x 10(13) RNA molecules derived from the TYMV TLS randomized in the anticodon loop nucleotides and in the length (1-6 nucleotides) and sequence of the pseudoknot loop L1. After nine rounds of selection by aminoacylation, 42 have been isolated. Among them, 17 RNAs could be efficiently charged by yeast ValRS. Their sequence revealed strong conservation of the second and the third anticodon triplet positions (A(56), C(55)) and the very 3'-end loop nucleotide C(53). A large variability of the other nucleotides of the loop was observed and no wild-type sequence was recovered. The selected molecules presented pseudoknot domains with loop L1 varying in size from 3-6 nucleotides and some sequence conservation, but did neither reveal the wild-type combination. All selected variants are 5-50 times more efficiently valylated than the wild-type TLS, suggesting that the natural viral sequence has emerged from a combination of evolutionary pressures among which aminoacylation was not predominant. This is in line with the role of the TLS in viral replication.}, note = {0006-2960 Journal Article}, keywords = {3' Untranslated Regions Acylation Anticodon/chemistry Base Sequence Cloning, FLORENTZ, Molecular Gene Library Kinetics Molecular Sequence Data Nucleic Acid Conformation Oligonucleotides/chemistry RNA, Non-U.S. Gov't Tymovirus/enzymology/genetics Valine-tRNA Ligase/chemistry Variation (Genetics), RNA Support, Transfer, Unité ARN, Val/*chemistry RNA, Viral/*chemistry Sequence Analysis}, pubstate = {published}, tppubtype = {article} } @article{, title = {Atomic Glimpses on a Billion-Year-Old Molecular Machine}, author = {E Westhof and N Leontis}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10820444}, isbn = {10820444}, year = {2000}, date = {2000-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {39}, number = {9}, pages = {1587-1591}, note = {0570-0833 Journal article}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA folding: beyond Watson-Crick pairs}, author = {E Westhof and V Fritsch}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10745012}, isbn = {10745012}, year = {2000}, date = {2000-01-01}, journal = {Structure}, volume = {8}, number = {3}, pages = {R55-65}, abstract = {Several crystal structures of RNA fragments, alone or in complex with a specific protein, have been recently solved. In addition, the structures of an artificial ribozyme, the leadzyme, and the cleavage product of a human pathogen ribozyme, have extended the structural diversity of ribozyme architectures. The attained set of folding rules and motifs expand the repertoire seen previously in tRNA structures.}, note = {0969-2126 Journal Article Review Review, Tutorial}, keywords = {Base Pairing Human Hydrogen Bonding *Nucleic Acid Conformation RNA/*chemistry Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {RNA tertiary structure}, author = {E Westhof and P Auffinger}, editor = {R A Meyers}, url = {http://eu.wiley.com/WileyCDA/WileyTitle/productCd-0471976709.html}, year = {2000}, date = {2000-01-01}, booktitle = {Encyclopedia of Analytical Chemistry: Applications, Theory, and Instrumentation}, pages = {5222-5232}, publisher = {John Wiley & Sons Ltd}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {NMR and biochemical characterization of recombinant human tRNA(Lys)3 expressed in Escherichia coli: identification of posttranscriptional nucleotide modifications required for efficient initiation of HIV-1 reverse transcription}, author = {C Tisne and M Rigourd and R Marquet and C Ehresmann and F Dardel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11073216}, isbn = {11073216}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {10}, pages = {1403-1412}, abstract = {Reverse transcription of HIV-1 viral RNA uses human tRNA(Lys)3 as a primer. Some of the modified nucleotides carried by this tRNA must play a key role in the initiation of this process, because unmodified tRNA produced in vitro is only marginally active as primer. To provide a better understanding of the contribution of base modifications in the initiation complex, we have designed a recombinant system that allows tRNA(Lys)3 expression in Escherichia coli. Because of their high level of overexpression, some modifications are incorporated at substoichiometric levels. We have purified the two major recombinant tRNA(Lys)3 subspecies, and their modified nucleotide contents have been characterized by a combination of NMR and biochemical techniques. Both species carry psis, Ds, T, t6A, and m7G. Differences are observed at position 34, within the anticodon. One fraction lacks the 5-methylaminomethyl group, whereas the other lacks the 2-thio group. Although the s2U34-containing recombinant tRNA is a less efficient primer, it presents most of the characteristics of the mammalian tRNA. On the other hand, the mnm5U34-containing tRNA has a strongly reduced activity. Our results demonstrate that the modifications that are absent in E. coli (m2G10, psi27, m5C48, m5C49, and m1A58) as well as the mnm5 group at position 34 are dispensable for initiation of reverse transcription. In contrast, the 2-thio group at position 34 seems to play an important part in this process.}, note = {1355-8382 Journal Article}, keywords = {Base Sequence DNA, Biomolecular *Nucleic Acid Conformation RNA/*chemistry/genetics/metabolism RNA, Genetic Transcription, Genetic/*genetics Virus Replication, Lys/*chemistry/genetics/metabolism Structure-Activity Relationship Support, MARQUET, Molecular Molecular Sequence Data Mutation/genetics Nuclear Magnetic Resonance, Non-U.S. Gov't Templates, Transfer, Unité ARN, Viral/biosynthesis/genetics Escherichia coli/genetics *Genetic Engineering HIV-1/*genetics/physiology Human Iodine/metabolism Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {The 2.0 A crystal structure of Thermus thermophilus methionyl-tRNA synthetase reveals two RNA-binding modules}, author = {I Sugiura and O Nureki and Y Ugaji-Yoshikawa and S Kuwabara and A Shimada and M Tateno and B Lorber and R Giege and D Moras and S Yokoyama and M Konno}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10673435}, isbn = {10673435}, year = {2000}, date = {2000-01-01}, journal = {Structure}, volume = {8}, number = {2}, pages = {197-208}, abstract = {Background: The 20 aminoacyl-tRNA synthetases are divided into two classes, I and II. The 10 class I synthetases are considered to have in common the catalytic domain structure based on the Rossmann fold, which is totally different from the class II catalytic domain structure. The class I synthetases are further divided into three subclasses, a, b and c, according to sequence homology. No conserved structural features for tRNA recognition by class I synthetases have been established. Results: We determined the crystal structure of the class Ia methionyl-tRNA synthetase (MetRS) at 2.0 A resolution, using MetRS from an extreme thermophile, Thermus thermophilus HB8. The T. thermophilus MetRS structure is in full agreement with the biochemical and genetic data from Escherichia coli MetRS. The conserved 'anticodon-binding' residues are spatially clustered on an alpha-helix-bundle domain. The Rossmann-fold and anticodon-binding domains are connected by a beta-alpha-alpha-beta-alpha topology ('SC fold') domain that contains the class I specific KMSKS motif. Conclusions: The alpha-helix-bundle domain identified in the MetRS structure is the signature of the class Ia enzymes, as it was also identified in the class Ia structures of the isoleucyl- and arginyl-tRNA synthetases. The beta-alpha-alpha-beta-alpha topology domain, which can now be identified in all known structures of the class Ia and Ib synthetases, is likely to dock with the inner side of the L-shaped tRNA, thereby positioning the anticodon stem.}, note = {0969-2126 Journal Article}, keywords = {Anticodon Catalytic Domain Crystallography, Molecular Protein Conformation Protein Folding RNA-Binding Proteins/*chemistry/metabolism Thermus thermophilus/*chemistry, Unité ARN, X-Ray Methionine-tRNA Ligase/*chemistry/metabolism Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural organization of Staf-DNA complexes}, author = {M Schaub and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10773080}, isbn = {10773080}, year = {2000}, date = {2000-01-01}, journal = {Nucleic Acids Res}, volume = {28}, number = {10}, pages = {2114-2121}, abstract = {The transactivator Staf, which contains seven contiguous zinc fingers of the C(2)-H(2)type, exerts its effects on gene expression by binding to specific targets in vertebrate small nuclear RNA (snRNA) and snRNA-type gene promoters. Here, we have investigated the interaction of the Staf zinc finger domain with the optimal Xenopus selenocysteine tRNA (xtRNA(Sec)) and human U6 snRNA (hU6) Staf motifs. Generation of a series of polypeptides containing increasing numbers of Staf zinc fingers tested in binding assays, by interference techniques and by binding site selection served to elucidate the mode of interaction between the zinc fingers and the Staf motifs. Our results provide strong evidence that zinc fingers 3-6 represent the minimal zinc finger region for high affinity binding to Staf motifs. Furthermore, we show that the binding of Staf is achieved through a broad spectrum of close contacts between zinc fingers 1-6 and xtRNA(Sec)or optimal sites or between zinc fingers 3-6 and the hU6 site. Extensive DNA major groove contacts contribute to the interaction with Staf that associates more closely with the non-template than with the template strand. Based on these findings and the structural information provided by the solved structures of other zinc finger-DNA complexes, we propose a model for the interaction between Staf zinc fingers and the xtRNA(Sec), optimal and hU6 sites.}, note = {1362-4962 Journal Article}, keywords = {Amino Acid Sequence Animals Base Sequence Binding Sites DNA-Binding Proteins/*chemistry/*metabolism Human Models, Amino Acid-Specific/*genetics Support, Genetic Trans-Activators/*chemistry/*metabolism Vertebrates Xenopus laevis Zinc Fingers, Molecular Molecular Sequence Data Nucleic Acid Conformation Plasmids/*chemistry/*metabolism Protein Conformation RNA, Non-U.S. Gov't Templates, Small Nuclear/*genetics RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The free yeast aspartyl-tRNA synthetase differs from the tRNA(Asp)-complexed enzyme by structural changes in the catalytic site, hinge region, and anticodon-binding domain}, author = {C Sauter and B Lorber and J Cavarelli and D Moras and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10873455}, isbn = {10873455}, year = {2000}, date = {2000-01-01}, journal = {J Mol Biol}, volume = {299}, number = {5}, pages = {1313-1324}, abstract = {Aminoacyl-tRNA synthetases catalyze the specific charging of amino acid residues on tRNAs. Accurate recognition of a tRNA by its synthetase is achieved through sequence and structural signalling. It has been shown that tRNAs undergo large conformational changes upon binding to enzymes, but little is known about the conformational rearrangements in tRNA-bound synthetases. To address this issue the crystal structure of the dimeric class II aspartyl-tRNA synthetase (AspRS) from yeast was solved in its free form and compared to that of the protein associated to the cognate tRNA(Asp). The use of an enzyme truncated in N terminus improved the crystal quality and allowed us to solve and refine the structure of free AspRS at 2.3 A resolution. For the first time, snapshots are available for the different macromolecular states belonging to the same tRNA aminoacylation system, comprising the free forms for tRNA and enzyme, and their complex. Overall, the synthetase is less affected by the association than the tRNA, although significant local changes occur. They concern a rotation of the anticodon binding domain and a movement in the hinge region which connects the anticodon binding and active-site domains in the AspRS subunit. The most dramatic differences are observed in two evolutionary conserved loops. Both are in the neighborhood of the catalytic site and are of importance for ligand binding. The combination of this structural analysis with mutagenesis and enzymology data points to a tRNA binding process that starts by a recognition event between the tRNA anticodon loop and the synthetase anticodon binding module.}, note = {0022-2836 Journal Article}, keywords = {Anticodon/chemistry/genetics/*metabolism Aspartate-tRNA Ligase/*chemistry/genetics/*metabolism Binding Sites Catalytic Domain Conserved Sequence/genetics Crystallization Crystallography, Asp/chemistry/genetics/*metabolism Rotation Sequence Deletion/genetics Support, Fungal/chemistry/genetics/metabolism RNA, Molecular Molecular Sequence Data Movement Nucleic Acid Conformation Protein Structure, Non-U.S. Gov't Yeasts/*enzymology/genetics, SAUTER, Secondary RNA, Transfer, Unité ARN, X-Ray Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Zinc ion mediated amino acid discrimination by threonyl-tRNA synthetase}, author = {R Sankaranarayanan and A C Dock-Bregeon and B Rees and M Bovee and J Caillet and P Romby and C S Francklyn and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10881191}, isbn = {10881191}, year = {2000}, date = {2000-01-01}, journal = {Nat Struct Biol}, volume = {7}, number = {6}, pages = {461-465}, abstract = {Accurate translation of the genetic code depends on the ability of aminoacyl-tRNA synthetases to distinguish between similar amino acids. In order to investigate the basis of amino acid recognition and to understand the role played by the zinc ion present in the active site of threonyl-tRNA synthetase, we have determined the crystal structures of complexes of an active truncated form of the enzyme with a threonyl adenylate analog or threonine. The zinc ion is directly involved in threonine recognition, forming a pentacoordinate intermediate with both the amino group and the side chain hydroxyl. Amino acid activation experiments reveal that the enzyme shows no activation of isosteric valine, and activates serine at a rate 1,000-fold less than that of cognate threonine. This study demonstrates that the zinc ion is neither strictly catalytic nor structural and suggests how the zinc ion ensures that only amino acids that possess a hydroxyl group attached to the beta-position are activated.}, note = {1072-8368 Journal Article}, keywords = {Binding Sites Catalytic Domain Crystallography, Molecular Molecular Sequence Data Protein Binding Protein Conformation Sequence Deletion/genetics Serine-tRNA Ligase/chemistry/metabolism Structure-Activity Relationship Substrate Specificity Support, Non-U.S. Gov't Threonine/analogs & derivatives/chemistry/*metabolism Threonine-tRNA Ligase/*chemistry/genetics/*metabolism Valine-tRNA Ligase/chemistry/metabolism Zinc/*metabolism, ROMBY, Unité ARN, X-Ray Dimerization Escherichia coli/*enzymology Kinetics Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Differential gene expression in mesothelioma}, author = {B H Rihn and S Mohr and S A McDowell and S Binet and J Loubinoux and F Galateau and G Keith and G D Leikauf}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11034307}, isbn = {11034307}, year = {2000}, date = {2000-01-01}, journal = {FEBS Lett}, volume = {480}, number = {2-3}, pages = {95-100}, abstract = {To investigate the molecular events controlling malignant transformation of human pleural cells, we compared constitutive gene expression of mesothelioma cells to that of pleural cells. Using cDNA microarray and high-density filter array, we assessed expression levels of > 6500 genes. Most of the highly expressed transcripts were common to both cell lines and included genes associated with stress response and DNA repair, outcomes consistent with the radio- and chemo-resistance of mesothelioma. Interestingly, of the fewer than 300 genes that differed between cell lines, most functioned in (i) macromolecule stability, (ii) cell adhesion and recognition, (iii) cell migration (invasiveness), and (iv) extended cell division. Expression levels of several of these genes were confirmed by RT-PCR and could be useful as diagnostic markers of human mesothelioma.}, note = {0014-5793 Journal Article}, keywords = {Cell Adhesion Cell Cycle Cell Division Gene Expression Profiling *Gene Expression Regulation, Cultured Xenobiotics, Neoplastic Human Mesothelioma/*genetics/metabolism Neoplasm Invasiveness Neoplasm Proteins/metabolism Oligonucleotide Array Sequence Analysis/methods Oxidative Stress Reverse Transcriptase Polymerase Chain Reaction Tumor Cells, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genotoxicity of 3-methylcholanthrene in liver of transgenic big Blue mice}, author = {B H Rihn and M C Bottin and C Coulais and R Rouget and N Monhoven and W Baranowski and A Edorh and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11152559}, isbn = {11152559}, year = {2000}, date = {2000-01-01}, journal = {Environ Mol Mutagen}, volume = {36}, number = {4}, pages = {266-273}, abstract = {Transgenic mice provide a unique tool for studying the tissue specificity and mutagenic potential of chemicals. Because 3-methylcholanthrene (3MC) was found mutagenic in bacteria, clastogenic in bone marrow, and induces DNA adducts in animals, we were interested to determinine whether this xenobiotic provokes (1) cell proliferation, (2) transcriptional activity changes, (3) DNA adducts, and (4) hepatic mutations in transgenic Big Blue mice carrying the lambdaLIZ phage shuttle vector. Big Blue C57/Bl male mice were treated with a single intraperitoneal dose of 80 mg/kg 3MC for 1, 3, 6, 14, or 30 days. Cell proliferation was checked by 5-bromo-2-deoxyuridine labeling and immunohistochemical detection. The maximal increase of the mitotic index was evidenced after 3 days (2.9 times the control value; P < 0.01). The relative nucleus area, reflecting the transcriptional activity, was also the highest in the treated group after 3 days: 1.86 times the control value, on average (P < 0.01). Four major DNA adducts, determined according to the [(32)P]-postlabeling method, were evidenced in liver DNA of treated mice, 6 days after the treatment: the spot intensities increased in a time-dependent manner. The mutant frequency of liver DNA was the highest after 14 days: 20.3 +/- 2.9 x 10(-5) in the treated vs. 7.6 +/- 2.7 x 10(-5) in the control mice (P < 0.01). Sequencing of the lambda lacI mutant plaques showed mainly G:C --> T:A and C:G --> A:T transversions. In conclusion, 3MC at first induced nuclear enlargement and a slight increase of cell proliferation in liver, followed by parallel formation of DNA adducts and mutations. This study shows how transgenic models allow in vivo evaluation of mechanistically simultaneous endpoints.}, note = {0893-6692 Journal Article}, keywords = {Animals Bacterial Proteins/genetics Base Sequence Cell Division/drug effects DNA Adducts DNA Primers *Escherichia coli Proteins Liver/cytology/*drug effects Methylcholanthrene/*toxicity Mice Mice, Inbred C57BL Mice, Non-U.S. Gov't, Transgenic Mutagens/*toxicity Mutation Organ Weight Repressor Proteins/genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Inhaled crocidolite mutagenicity in lung DNA}, author = {B Rihn and C Coulais and E Kauffer and M C Bottin and P Martin and F Yvon and J C Vigneron and S Binet and N Monhoven and G Steiblen and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10753093}, isbn = {10753093}, year = {2000}, date = {2000-01-01}, journal = {Environ Health Perspect}, volume = {108}, number = {4}, pages = {341-346}, abstract = {We used transgenic mice carrying the lacI reporter gene to study the mutagenesis potential of asbestos crocidolite. The animals were exposed by nose-only inhalation to an aerosol containing 5.75 mg/m(3) crocidolite dust for 6 hr/day and 5 consecutive days. After 1, 4, and 12 weeks, we examined four end points: the cytology of bronchoalveolar lavage, the lung load of crocidolite, the hydrophobic DNA adducts, and the mutations in the lacI reporter gene. Twelve weeks after exposure, nearly 10% of the inhaled fibers remained in the lung (227 +/- 103 ng/mg lung). There was evidence of a typical inflammatory response consisting of multinucleate macrophages at weeks 4 and 12, whereas immediately after the exposure, we observed numerous polymorphonuclear neutrophils. The mutant frequency significatively increased during the fourth week after the exposure: 13.5 [time] 10(-5) in the exposed group versus 6. 9 10(-5) in the control group. The induction factor, defined by the ratio of checked mutants of exposed mice to checked mutants of control mice, was 1.96. The mutation spectrum of control lung DNA and exposed lung DNA was similar, suggesting the possible involvement of a DNA repair decrease in crocidolite-treated animals. We used the (32)P-postlabeling method and did not detect any increase of either 5 mC or bulky adduct in treated mice. This is the first study that demonstrates asbestos mutagenicity in vivo after a nose-only inhalation.}, note = {0091-6765 Journal Article}, keywords = {Air Pollutants/*adverse effects Animals Asbestos, Alveolar/physiology Male Mice Mice, Crocidolite/administration & dosage/*adverse effects DNA Adducts/*genetics DNA Damage/*genetics Inhalation Exposure Lung/*drug effects/pathology Macrophages, Non-U.S. Gov't, Transgenic Mutagenicity Tests Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Inhibition of the initiation of HIV-1 reverse transcription by 3'-azido-3'-deoxythymidine. Comparison with elongation}, author = {M Rigourd and J M Lanchy and S F Le Grice and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10864929}, isbn = {10864929}, year = {2000}, date = {2000-01-01}, journal = {J Biol Chem}, volume = {275}, number = {35}, pages = {26944-26951}, abstract = {Initiation of human immunodeficiency virus-1 reverse transcription requires formation of a complex containing the viral RNA, primer tRNA(3)(Lys), and reverse transcriptase. Initiation, corresponding to addition of the first six nucleotides to tRNA(3)(Lys), is distinguished from elongation by its high specificity and low efficiency (processivity). Here, we compared the inhibition of initiation and elongation of reverse transcription by 3'-azido-3'-deoxythymidine 5'-triphosphate (AZTTP), the active form of 3'-azido-3'-deoxythymidine. We report the first detailed study of nucleotide binding, discrimination, and pyrophosphorolysis by the authentic initiation complex. We showed that the initiation and elongation complexes bound AZTTP and dTTP with the same affinity, while the polymerization rates were reduced by 148-160-fold during initiation. The pyrophosphorolysis rate of dTTP was reduced by the same extent, indicating that the polymerization equilibrium is the same in the two phases. The efficient unblocking of the 3'-azido-3'-deoxythymidine 5'-monophosphate (AZTMP)-terminated primer by pyrophosphorolysis significantly relieved inhibition of DNA synthesis during elongation in the presence of physiological pyrophosphate concentrations. Remarkably, although pyrophosphorolysis of dTMP and AZTMP were equally efficient during elongation, reverse transcriptase was almost totally unable to unblock the AZTMP-terminated primer during initiation. As a result, inhibition of reverse transcription by AZTTP was more efficient during initiation than elongation of reverse transcription, despite a reduced selectivity of incorporation.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence Comparative Study DNA Primers Diphosphates/metabolism HIV-1/*genetics HIV-1 Reverse Transcriptase/antagonists & inhibitors/metabolism Human Hydrolysis Kinetics Lymphocytes/drug effects/metabolism/virology Peptide Chain Elongation/*drug effects Support, Genetic/*drug effects Zidovudine/analogs & derivatives/metabolism/*pharmacology, MARQUET, Non-U.S. Gov't Thymine Nucleotides/metabolism Transcription, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Overall genomic DNA methylation in relation to estrogen]}, author = {K Postawski and E Olech-Fudali and J A Jakowicki and E Korobowicz and G Keith and W Baranowski}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11083005}, isbn = {11083005}, year = {2000}, date = {2000-01-01}, journal = {Ginekol Pol}, volume = {71}, number = {9}, pages = {1206-1211}, abstract = {Overall genomic DNA methylation was analyzed using enzymatic digestion into nucleotides, 32P postlabeling, two-dimensional thin-layer chromatography on cellulose plates and phosphobioimaging quantitation, in relation to immunohistochemically measured estrogen (ER) and progesterone receptor (PR) status of 15 uterine cancers. Mean 5-methyldeoxycytosine (m5dC) content did not differ between ER-positive and ER-negative neoplasms. Highest values of m5dC were noted both in ER-negative and ER-positive tumors. Additionally, there was no low DNA methylation in ER negative uterine cancer tissues. Decrease of the overall genomic DNA methylation could be related to the increase of ER/PR ratio, however it was not significant in our investigation. The potential role of steroid receptors status in uterine cancer tissue is discussed.}, note = {0017-0011 Journal Article}, keywords = {80 and over Biopsy DNA Methylation DNA Modification Methylases/metabolism English Abstract Female Human Middle Aged Receptors, Aged Aged, Estrogen/*metabolism Receptors, Non-U.S. Gov't Uterine Neoplasms/*metabolism/*pathology Uterus/*metabolism/pathology, Progesterone/*metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Mechanism of oligonucleotide hybridization with the 3'-terminal region of yeast tRNA(Phe)]}, author = {V A Petyuk and R Giege and V V Vlasov and M A Zenkova}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11033816}, isbn = {11033816}, year = {2000}, date = {2000-01-01}, journal = {Mol Biol (Mosk)}, volume = {34}, number = {5}, pages = {879-886}, note = {0026-8984 Journal Article}, keywords = {Base Sequence DNA Primers Electrophoresis, Phe/chemistry/*genetics, Polyacrylamide Gel Nucleic Acid Conformation Nucleic Acid Hybridization RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Invasion of strongly binding oligonucleotides into tRNA structure}, author = {V Petyuk and R Serikov and V Tolstikov and V Potapov and R Giege and M Zenkova and V Vlassov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10999254}, isbn = {10999254}, year = {2000}, date = {2000-01-01}, journal = {Nucleosides Nucleotides Nucleic Acids}, volume = {19}, number = {7}, pages = {1145-1158}, abstract = {Interaction of yeast tRNA(Phe) with oligodeoxyribonucleotides containing 5-methylcytosine, 2-aminoadenine, and 5-propynyl-2'-deoxyuridine was investigated. The modified oligonucleotides show increased binding capacity although the association rates are similar for the modified and natural oligonucleotides. The most pronounced increase in association constant (70 times) due to the incorporation of the strongly binding units was achieved in the case of oligonucleotide complementary to the sequence 65-76 of the tRNA(Phe).}, note = {1525-7770 Journal Article}, keywords = {2-Aminopurine/*analogs & derivatives/chemistry 5-Methylcytosine Cytosine/analogs & derivatives/chemistry Deoxyuridine/*analogs & derivatives/chemistry Electrophoresis, Non-U.S. Gov't Time Factors Yeasts/chemistry, Phe/*chemistry/metabolism Support, Polyacrylamide Gel Kinetics Nucleic Acid Conformation Oligodeoxyribonucleotides/chemistry Oligonucleotides/*metabolism RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vitro DNA and dGMP adducts formation caused by ochratoxin A}, author = {S Obrecht-Pflumio and G Dirheimer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10903417}, isbn = {10903417}, year = {2000}, date = {2000-01-01}, journal = {Chem Biol Interact}, volume = {127}, number = {1}, pages = {29-44}, abstract = {Ochratoxin A (OTA), a nephrotoxic and nephrocarcinogenic mycotoxin, leads to the formation of DNA adducts after administration to animals. This could be due to an epigenetic effect. In vitro assays can exclude an indirect effect, where the xenobiotic can generate, in vivo, endogenous reactive compounds which give adducts on DNA. Microsomes prepared from mice or rabbit kidney and liver, used as metabolic activators, were incubated in the presence of commercial salmon testes DNA and OTA, with NADPH or arachidonic acid used as cofactors. Upto 126 DNA adducts for 10(9) nucleotides were detected using the 32P postlabeling method after incubation with the mouse kidney system. Similar results were obtained with rabbit kidney microsomes. Using liver microsomes, the number of DNA adducts detected was much lower. When NADPH was used as a cosubstrate (to explore the cytochrome P450 metabolic pathways), with mice kidney microsomes, the adduct level was only 44% of the one obtained with arachidonic acid. These results lend support to the hypothesis of the preferential activation of OTA by the peroxidase activity of prostaglandin synthases and/or lipoxygenases to direct genotoxic metabolites, and are in agreement with the previously obtained results after in vivo treatment of mice. In order to identify the nucleotides of DNA modified by the OTA metabolites, dAMP, dGMP, dTMP and dCMP were used as substrates under the same conditions as with DNA. The adducts were found only on dGMP. The total adduct level was of 344 adducts per 10(9) nucleotides with the appearance of three major adducts in the presence of arachidonic acid. With NADPH, 271 adducts were obtained per 10(9) nucleotides, with again three major adducts, but only two of them were similar to two adducts obtained in the presence of arachidonic acid. Desferal (desferrioxamine B methanesulphonate), at a 50 microM concentration, did not reduce the adduct level. Adducts were also obtained when polydG, polydC and dG-p-dG were used as alternative substrates, whereas no adducts were obtained with polydA, polydT and polydC. The major adduct obtained after incubation of DNA with OTA, comigrated with the major adduct obtained with dGMP, in two chromatographic solvents. These results show that OTA is metabolized to genotoxic metabolite(s) which interact with the guanine residues of DNA.}, note = {0009-2797 Journal Article}, keywords = {Animals Arachidonic Acid/metabolism Carcinogens/pharmacology Chelating Agents/pharmacology Chromatography DNA Adducts/*metabolism Deferoxamine/pharmacology Deoxyguanine Nucleotides/*metabolism Female Kidney/ultrastructure Male Mice Microsomes/metabolism Microsomes, Liver/metabolism Mycotoxins/pharmacology NADP/metabolism Nucleotides/metabolism Ochratoxins/*pharmacology Peroxidase/metabolism Rabbits Spectrophotometry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of the S15-rRNA complex}, author = {A Nikulin and A Serganov and E Ennifar and S Tishchenko and N Nevskaya and W Shepard and C Portier and M Garber and B Ehresmann and C Ehresmann and S Nikonov and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10742169}, isbn = {10742169}, year = {2000}, date = {2000-01-01}, journal = {Nat Struct Biol}, volume = {7}, number = {4}, pages = {273-277}, abstract = {In bacterial ribosomes, the small (30S) ribosomal subunit is composed of 16S rRNA and 21 distinct proteins. Ribosomal protein S15 is of particular interest because it binds primarily to 16S rRNA and is required for assembly of the small subunit and for intersubunit association, thus representing a key element in the assembly of a whole ribosome. Here we report the 2.8 inverted question mark resolution crystal structure of the highly conserved S15-rRNA complex. Protein S15 interacts in the minor groove with a G-U/G-C motif and a three-way junction. The latter is constrained by a conserved base triple and stacking interactions, and locked into place by magnesium ions and protein side chains, mainly through interactions with the unique three-dimensional geometry of the backbone. The present structure gives insights into the dual role of S15 in ribosome assembly and translational regulation.}, note = {1072-8368 Journal Article}, keywords = {16S/*chemistry/genetics/*metabolism Ribosomal Proteins/*chemistry/*metabolism Structure-Activity Relationship Support, Amino Acid Sequence Base Pairing/drug effects/genetics Base Sequence Binding Sites/drug effects Conserved Sequence/genetics Crystallography, Bacterial/chemistry/genetics/metabolism RNA, ENNIFAR, Molecular Molecular Sequence Data *Nucleic Acid Conformation/drug effects Protein Conformation RNA, Non-U.S. Gov't Thermus thermophilus/*chemistry/genetics, Ribosomal, Unité ARN, X-Ray Magnesium/pharmacology Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vivo selection of functional ribosomes with variations in the rRNA-binding site of Escherichia coli ribosomal protein S8: evolutionary implications}, author = {H Moine and C L Squires and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10639126}, isbn = {10639126}, year = {2000}, date = {2000-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {97}, number = {2}, pages = {605-610}, abstract = {The highly conserved nature of rRNA sequences throughout evolution allows these molecules to be used to build philogenic trees of different species. It is unknown whether the stability of specific interactions and structural features of rRNA reflects an optimal adaptation to a functional task or an evolutionary trap. In the work reported here, we have applied an in vivo selection strategy to demonstrate that unnatural sequences do work as a functional replacement of the highly conserved binding site of ribosomal protein S8. However, growth competition experiments performed between Escherichia coli isolates containing natural and unnatural S8-binding sites showed that the fate of each isolate depended on the growth condition. In exponentially growing cells, one unnatural variant was found to be equivalent to wild type in competition experiments performed in rich media. In culture conditions leading to slow growth, however, cells containing the wild-type sequence were the ultimate winner of the competition, emphasizing that the wild-type sequence is, in fact, the most fit solution for the S8-binding site.}, note = {0027-8424http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10639126 Journal Article}, keywords = {16S/genetics/metabolism Recombinant Fusion Proteins/genetics/metabolism Ribosomal Proteins/genetics/*metabolism Ribosomes/genetics/*metabolism Spectinomycin/pharmacology Support, Base Sequence Binding Sites/genetics Binding, Competitive Cell Division/genetics Cloning, Microbial Escherichia coli/drug effects/*genetics/metabolism Evolution, Molecular Drug Resistance, Molecular Protein Binding RNA, P.H.S. Variation (Genetics), Ribosomal, Ribosomal/genetics/*metabolism RNA, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Modeling RNA tertiary structure from patterns of sequence variation}, author = {F Michel and M Costa and C Massire and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10829297}, isbn = {10829297}, year = {2000}, date = {2000-01-01}, journal = {Methods Enzymol}, volume = {317}, pages = {491-510}, note = {0076-6879 Journal Article Review Review, Tutorial}, keywords = {Base Pairing Base Sequence Models, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA, Transfer/*chemistry Sequence Alignment, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {On the wobble GoU and related pairs}, author = {B Masquida and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10668794}, isbn = {10668794}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {1}, pages = {9-15}, abstract = {The wobble GoU pairs have been implicated in several biological processes where RNA molecules play a key role. We review the geometrical and conformational properties of wobble GoU pairs on the basis of available crystal structures of RNAs at high resolution. The similarities with the wobble A+oC pairs and UoU pairs are illustrated, while the differences with the recently discovered bifurcated G x U pairs are contrasted.}, note = {1355-8382 Journal Article Review Review, Tutorial}, keywords = {*Base Pairing Models, Molecular RNA/*chemistry Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Positive and negative mutant selection in the human histone hairpin- binding protein using the yeast three-hybrid system.}, author = {F Martin and F Michel and D Zenklusen and B Muller and D Schumperli}, url = {http://www.ncbi.nlm.nih.gov/pubmed/10710426Martin}, isbn = {10710426}, year = {2000}, date = {2000-01-01}, journal = {Nucleic Acids Res}, volume = {28}, number = {7}, pages = {1594-1603.}, abstract = {We have used the yeast three-hybrid system in a positive selection for mutants of the human histone hairpin-binding protein (HBP) capable of interacting with non-canonical hairpins and in a negative selection for loss-of-binding mutants. Interestingly, all mutations from the positive selection are located in the N- and C-terminal regions flanking a minimal RNA-binding domain (RBD) previously defined between amino acids 126 and 198. Further, in vitro binding studies demonstrate that the RBD, which shows no obvious similarity to other RNA-binding motifs, has a relaxed sequence specificity compared to full-length HBP, allowing it to bind to mutant hairpin RNAs not normally found in histone genes. These findings indicate that the sequences flanking the RBD are important for restricting binding to the highly conserved histone hairpin structure. Among the loss-of-binding mutations, about half are nonsense mutations distributed throughout the N-terminal part and the RBD whereas the other half are missense mutations restricted to the RBD. Whereas the nonsense mutations permit a more precise definition of the C-terminal border of the RBD, the missense mutations identify critical residues for RNA binding within the RBD.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Growth kinetics and motions of thaumatin crystals during USML-2 and LMS microgravity missions and comparison with earth controls.}, author = {B Lorber and J D Ng and P Lautenschlager and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S002202489900398X}, doi = {10.1016/S0022-0248(99)00398-X}, year = {2000}, date = {2000-01-01}, journal = {J Crystal Growth}, volume = {208}, number = {1-4}, pages = {665-677}, abstract = {As part of a study of the effects of microgravity on protein crystallization, the growth of tetragonal crystals of thaumatin was monitored by CCD-time-lapse video in environments where convection is negligible. In Space Shuttle missions entitled United States Microgravity Laboratory-2 and Life and Microgravity Sciences, free interface diffusion and dialysis techniques were utilized to grow crystals in the advanced protein crystallization facility (APCF). Ng et al. (Acta Crystallogr. D 53 (1997) 724) have shown that the crystals recovered in these experiments are of superior crystallographic quality (at the level of their diffraction intensity, resolution, and mosaicity) with regard to earthcontrols. Here, the number of crystals, their size, growth rate, and protein solubility in microgravity were compared with data of dialysis experiments performed in parallel on earth. Image analysis shows that in microgravity about one-quarter to half of the crystals have nucleated and grown in the bulk of the solution, the remaining being attached to the walls of crystallization vessels. The growth of free-floating crystals was 2.5 times faster, has resulted in 15-fold larger crystals, and consumed more protein than that of attached crystals in earthcontrols. Distances between immobile free-floating crystals in microgravity were related to the size of the latter. Experimental results are in favor of a correlation between more favorable growth parameters in microgravity and better diffraction properties. The displacements of free-floating crystals at various velocities and in various directions on unrelated trajectories are indicative of drift and stirring motion. On the basis of an overview of reactors monitored on four APCF missions some forces causing motion are proposed. Advantages of crystallization in microgravity are discussed and recommendations for future experiments}, keywords = {Protein Crystallization Microgravity Thaumatin Growthkinetics Motion, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Convergence of natural and artificial evolution on an RNA loop-loop interaction: the HIV-1 dimerization initiation site}, author = {J S Lodmell and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10999604}, isbn = {10999604}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {9}, pages = {1267-1276}, abstract = {Loop-loop interactions among nucleic acids constitute an important form of molecular recognition in a variety of biological systems. In HIV-1, genomic dimerization involves an intermolecular RNA loop-loop interaction at the dimerization initiation site (DIS), a hairpin located in the 5' noncoding region that contains an autocomplementary sequence in the loop. Only two major DIS loop sequence variants are observed among natural viral isolates. To investigate sequence and structural constraints on genomic RNA dimerization as well as loop-loop interactions in general, we randomized several or all of the nucleotides in the DIS loop and selected in vitro for dimerization-competent sequences. Surprisingly, increasing interloop complementarity above a threshold of 6 bp did not enhance dimerization, although the combinations of nucleotides forming the theoretically most stable hexanucleotide duplexes were selected. Noncanonical interactions contributed significantly to the stability and/or specificity of the dimeric complexes as demonstrated by the overwhelming bias for noncanonical base pairs closing the loop and covariations between flanking and central loop nucleotides. Degeneration of the entire loop yielded a complex population of dimerization-competent sequences whose consensus sequence resembles that of wild-type HIV-1. We conclude from these findings that the DIS has evolved to satisfy simultaneous constraints for optimal dimerization affinity and the capacity for homodimerization. Furthermore, the most constrained features of the DIS identified by our experiments could be the basis for the rational design of DIS-targeted antiviral compounds.}, note = {1355-8382 Journal Article}, keywords = {Codon, Initiator Dimerization Directed Molecular Evolution Evolution, MARQUET, Molecular HIV-1/*chemistry/genetics Nucleic Acid Conformation RNA, Non-U.S. Gov't, Unité ARN, Viral/*chemistry/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Splicing enhancement in the yeast rp51b intron}, author = {D Libri and A Lescure and M Rosbash}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10744020}, isbn = {10744020}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {3}, pages = {352-368}, abstract = {Splicing enhancement in higher eukaryotes has been linked to SR proteins, to U1 snRNP, and to communication between splice sites across introns or exons mediated by protein-protein interactions. It has been previously shown that, in yeast, communication mediated by RNA-RNA interactions between the two ends of introns is a basis for splicing enhancement. We designed experiments of randomization-selection to isolate splicing enhancers that would work independently from RNA secondary structures. Surprisingly, one of the two families of sequences selected was essentially composed of 5' splice site variants. We show that this sequence enhances splicing independently of secondary structure, is exportable to heterologous contexts, and works in multiple copies with additive effects. The data argue in favor of an early role for splicing enhancement, possibly coincident with commitment complex formation. Genetic compensation experiments with U1 snRNA mutants suggest that U1 snRNP binding to noncanonical locations is required for splicing enhancement.}, note = {1355-8382 Journal Article}, keywords = {5' Untranslated Regions/genetics DNA Mutational Analysis Introns/*genetics Multigene Family Nucleic Acid Conformation RNA Splicing/*genetics *Regulatory Sequences, LESCURE, Non-U.S. Gov't Support, Nucleic Acid Ribonucleoprotein, P.H.S. Uridine/metabolism, U.S. Gov't, U1 Small Nuclear/chemistry/genetics Saccharomyces cerevisiae/*genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {From RNA structure to the identification of new genes: the example of selenoproteins}, author = {A Lescure and D Gautheret and D Fagegaltier and P Carbon and A Krol}, url = {http://sciencelinks.jp/j-east/article/200107/000020010701A0078173.php}, isbn = {01A0078173}, year = {2000}, date = {2000-01-01}, journal = {J Health Sci}, volume = {46}, number = {6}, pages = {405-408}, keywords = {KROL ACL, LESCURE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dynamics of the HIV-1 reverse transcription complex during initiation of DNA synthesis}, author = {J M Lanchy and C Isel and G Keith and S F Le Grice and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10766870}, isbn = {10766870}, year = {2000}, date = {2000-01-01}, journal = {J Biol Chem}, volume = {275}, number = {16}, pages = {12306-12312}, abstract = {Initiation of human immunodeficiency virus-1 (HIV-1) reverse transcription requires formation of a complex containing the viral RNA (vRNA), tRNA(3)(Lys) and reverse transcriptase (RT). The vRNA and the primer tRNA(3)(Lys) form several intermolecular interactions in addition to annealing of the primer 3' end to the primer binding site (PBS). These interactions are crucial for the efficiency and the specificity of the initiation of reverse transcription. However, as they are located upstream of the PBS, they must unwind as DNA synthesis proceeds. Here, the dynamics of the complex during initiation of reverse transcription was followed by enzymatic probing. Our data revealed reciprocal effects of the tertiary structure of the vRNA.tRNA(3)(Lys) complex and reverse transcriptase (RT) at a distance from the polymerization site. The structure of the initiation complex allowed RT to interact with the template strand up to 20 nucleotides upstream from the polymerization site. Conversely, nucleotide addition by RT modified the tertiary structure of the complex at 10-14 nucleotides from the catalytic site. The viral sequences became exposed at the surface of the complex as they dissociated from the tRNA following primer extension. However, the counterpart tRNA sequences became buried inside the complex. Surprisingly, they became exposed when mutations prevented the intermolecular interactions in the initial complex, indicating that the fate of the tRNA depended on the tertiary structure of the initial complex.}, note = {0021-9258 Journal Article}, keywords = {*Anticodon Base Sequence *DNA Replication *Hiv-1 Human Molecular Sequence Data Mutagenesis Nucleic Acid Conformation RNA, Genetic, Lys/genetics/metabolism RNA, MARQUET, Non-U.S. Gov't Templates, Transfer, Unité ARN, Viral/genetics/metabolism RNA-Directed DNA Polymerase/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {An unusual structure formed by antisense-target RNA binding involves an extended kissing complex with a four-way junction and a side-by-side helical alignment}, author = {F A Kolb and C Malmgren and E Westhof and C Ehresmann and B Ehresmann and E G Wagner and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10744017}, isbn = {10744017}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {3}, pages = {311-324}, abstract = {The antisense RNA CopA binds to the leader region of the repA mRNA (target: CopT). Previous studies on CopA-CopT pairing in vitro showed that the dominant product of antisense RNA-mRNA binding is not a full RNA duplex. We have studied here the structure of CopA-CopT complex, combining chemical and enzymatic probing and computer graphic modeling. CopI, a truncated derivative of CopA unable to bind CopT stably, was also analyzed. We show here that after initial loop-loop interaction (kissing), helix propagation resulted in an extended kissing complex that involves the formation of two intermolecular helices. By introducing mutations (base-pair inversions) into the upper stem regions of CopA and CopT, the boundaries of the two newly formed intermolecular helices were delimited. The resulting extended kissing complex represents a new type of four-way junction structure that adopts an asymmetrical X-shaped conformation formed by two helical domains, each one generated by coaxial stacking of two helices. This structure motif induces a side-by-side alignment of two long intramolecular helices that, in turn, facilitates the formation of an additional intermolecular helix that greatly stabilizes the inhibitory CopA-CopT RNA complex. This stabilizer helix cannot form in CopI-CopT complexes due to absence of the sequences involved. The functional significance of the three-dimensional models of the extended kissing complex (CopI-CopT) and the stable complex (CopA-CopT) are discussed.}, note = {1355-8382 Journal Article}, keywords = {Antisense/*metabolism RNA, Bacterial Proteins/*metabolism Base Pairing Base Sequence Binding Sites Cations, Divalent Computer Simulation Metals, Double-Stranded/metabolism RNA, Heavy/metabolism Models, Messenger/metabolism RNA, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA Stability RNA, Non-U.S. Gov't, ROMBY, Spliced Leader/metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Progression of a loop-loop complex to a four-way junction is crucial for the activity of a regulatory antisense RNA}, author = {F A Kolb and H M Engdahl and J G Slagter-Jager and B Ehresmann and C Ehresmann and E Westhof and E G Wagner and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11060041}, isbn = {11060041}, year = {2000}, date = {2000-01-01}, journal = {EMBO J}, volume = {19}, number = {21}, pages = {5905-5915}, abstract = {The antisense RNA, CopA, regulates the replication frequency of plasmid R1 through inhibition of RepA translation by rapid and specific binding to its target RNA (CopT). The stable CopA-CopT complex is characterized by a four-way junction structure and a side-by-side alignment of two long intramolecular helices. The significance of this structure for binding in vitro and control in vivo was tested by mutations in both CopA and CopT. High rates of stable complex formation in vitro and efficient inhibition in vivo required initial loop-loop complexes to be rapidly converted to extended interactions. These interactions involve asymmetric helix progression and melting of the upper stems of both RNAs to promote the formation of two intermolecular helices. Data presented here delineate the boundaries of these helices and emphasize the need for unimpeded helix propagation. This process is directional, i.e. one of the two intermolecular helices (B) must form first to allow formation of the other (B'). A binding pathway, characterized by a hierarchy of intermediates leading to an irreversible and inhibitory RNA-RNA complex, is proposed.}, note = {0261-4189 Journal Article}, keywords = {Antisense/*chemistry/*genetics/metabolism RNA, Bacterial Models, Bacterial Proteins/genetics Base Sequence Binding, Bacterial/chemistry/genetics/metabolism Support, Competitive DNA Primers/genetics Escherichia coli/chemistry/genetics/metabolism Genes, Molecular Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, Non-U.S. Gov't, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A potential RNA drug target in the hepatitis C virus internal ribosomal entry site}, author = {R Klinck and E Westhof and S Walker and M Afshar and A Collier and F Aboul-Ela}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11073218}, isbn = {11073218}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {10}, pages = {1423-1431}, abstract = {Subdomain IlId from the hepatitis C virus (HCV) internal ribosome entry site (IRES) has been shown to be essential for cap-independent translation. We have conducted a structural study of a 27-nt fragment, identical in sequence to IlId, to explore the structural features of this subdomain. The proposed secondary structure of IlId is comprised of two 3 bp helical regions separated by an internal loop and closed at one end by a 6-nt terminal loop. NMR and molecular modeling were used interactively to formulate a validated model of the three-dimensional structure of IlId. We found that this fragment contains several noncanonical structural motifs and non-Watson-Crick base pairs, some of which are common to other RNAs. In particular, a motif characteristic of the rRNA alpha-sarcin/ricin loop was located in the internal loop. The terminal loop, 5'-UUGGGU, was found to fold to form a trinucleotide loop closed by a trans-wobble U.G base pair. The sixth nucleotide was bulged out to allow stacking of this U.G pair on the adjacent helical region. In vivo mutational analysis in the context of the full IRES confirmed the importance of each structural motif within IIId for IRES function. These findings may provide clues as to host cellular proteins that play a role in IRES-directed translation and, in particular, the mechanism through which host ribosomes are sequestered for viral function.}, note = {1355-8382 Journal Article}, keywords = {Base Pairing Base Sequence Computational Biology *Drug Design Endoribonucleases/metabolism Genetic Engineering Hepacivirus/*genetics Models, Biomolecular *Nucleic Acid Conformation RNA, Genetic, Messenger/chemistry/genetics/metabolism RNA, Molecular Molecular Sequence Data Mutation Nuclear Magnetic Resonance, Nucleic Acid/*genetics Reproducibility of Results Ribosomes/*metabolism Ricin/metabolism Structure-Activity Relationship Substrate Specificity Translation, Unité ARN, Viral/*chemistry/genetics/*metabolism Regulatory Sequences}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genomic DNA methylation in the human trophoblast from recurrent spontaneous abortion-premilimary report}, author = {M Jerzak and K Potawski and G Keith and J A Jakowicki and W Baranovski}, url = {none}, year = {2000}, date = {2000-01-01}, journal = {Pol J Gyn Invest}, volume = {3}, pages = {9-12}, keywords = {ACL GIEGE, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Inhaled crocidolite mutagenicity in lung DNA}, author = { B. Rihn and C. Coulais and E. Kauffer and M. C. Bottin and P. Martin and F. Yvon and J. C. Vigneron and S. Binet and N. Monhoven and G. Steiblen and G. Keith}, year = {2000}, date = {2000-01-01}, journal = {Environ Health Perspect}, volume = {108}, number = {4}, pages = {341-6}, abstract = {We used transgenic mice carrying the lacI reporter gene to study the mutagenesis potential of asbestos crocidolite. The animals were exposed by nose-only inhalation to an aerosol containing 5.75 mg/m(3) crocidolite dust for 6 hr/day and 5 consecutive days. After 1, 4, and 12 weeks, we examined four end points: the cytology of bronchoalveolar lavage, the lung load of crocidolite, the hydrophobic DNA adducts, and the mutations in the lacI reporter gene. Twelve weeks after exposure, nearly 10% of the inhaled fibers remained in the lung (227 +/- 103 ng/mg lung). There was evidence of a typical inflammatory response consisting of multinucleate macrophages at weeks 4 and 12, whereas immediately after the exposure, we observed numerous polymorphonuclear neutrophils. The mutant frequency significatively increased during the fourth week after the exposure: 13.5 [time] 10(-5) in the exposed group versus 6. 9 10(-5) in the control group. The induction factor, defined by the ratio of checked mutants of exposed mice to checked mutants of control mice, was 1.96. The mutation spectrum of control lung DNA and exposed lung DNA was similar, suggesting the possible involvement of a DNA repair decrease in crocidolite-treated animals. We used the (32)P-postlabeling method and did not detect any increase of either 5 mC or bulky adduct in treated mice. This is the first study that demonstrates asbestos mutagenicity in vivo after a nose-only inhalation.}, note = {0091-6765 Journal Article}, keywords = {&, Adducts/*genetics, Air, Alveolar/physiology, Animals, Asbestos, Crocidolite/administration, Damage/*genetics, DNA, dosage/*adverse, effects, effects/pathology, Exposure, Gov't, Inhalation, Lung/*drug, Macrophages, Male, Mice, Mutagenicity, Non-U.S., Pollutants/*adverse, Support, Tests, transgenic}, pubstate = {published}, tppubtype = {article} } @article{, title = {Tecto-RNA: One-Dimensional Self-Assembly through Tertiary Interactions}, author = {L Jaeger and N B Leontis}, url = {http://onlinelibrary.wiley.com/doi/10.1002/1521-3773%2820000717%2939:14%3C2521:AID-ANIE2521%3E3.0.CO;2-P/abstract}, doi = {10.1002/1521-3773(20000717)39:14<2521:AID-ANIE2521>3.0.CO;2-P}, isbn = {10941124}, year = {2000}, date = {2000-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {39}, number = {14}, pages = {2521-2524}, abstract = {The modularity of natural RNA is the basis for the design of tecto-RNA, modular RNA units capable of directed self-assembly. One such modular association through specific RNA loop–receptor tertiary interactions, which leads to one-dimensional oligomer arrays, is demonstrated (see picture).}, keywords = {RNA self-organisation supramolecular chemistry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genotoxicity of 3-methylcholanthrene in liver of transgenic big Blue mice}, author = { B. H. Rihn and M. C. Bottin and C. Coulais and R. Rouget and N. Monhoven and W. Baranowski and A. Edorh and G. Keith}, year = {2000}, date = {2000-01-01}, journal = {Environ Mol Mutagen}, volume = {36}, number = {4}, pages = {266-73}, abstract = {Transgenic mice provide a unique tool for studying the tissue specificity and mutagenic potential of chemicals. Because 3-methylcholanthrene (3MC) was found mutagenic in bacteria, clastogenic in bone marrow, and induces DNA adducts in animals, we were interested to determinine whether this xenobiotic provokes (1) cell proliferation, (2) transcriptional activity changes, (3) DNA adducts, and (4) hepatic mutations in transgenic Big Blue mice carrying the lambdaLIZ phage shuttle vector. Big Blue C57/Bl male mice were treated with a single intraperitoneal dose of 80 mg/kg 3MC for 1, 3, 6, 14, or 30 days. Cell proliferation was checked by 5-bromo-2-deoxyuridine labeling and immunohistochemical detection. The maximal increase of the mitotic index was evidenced after 3 days (2.9 times the control value; P < 0.01). The relative nucleus area, reflecting the transcriptional activity, was also the highest in the treated group after 3 days: 1.86 times the control value, on average (P < 0.01). Four major DNA adducts, determined according to the [(32)P]-postlabeling method, were evidenced in liver DNA of treated mice, 6 days after the treatment: the spot intensities increased in a time-dependent manner. The mutant frequency of liver DNA was the highest after 14 days: 20.3 +/- 2.9 x 10(-5) in the treated vs. 7.6 +/- 2.7 x 10(-5) in the control mice (P < 0.01). Sequencing of the lambda lacI mutant plaques showed mainly G:C --> T:A and C:G --> A:T transversions. In conclusion, 3MC at first induced nuclear enlargement and a slight increase of cell proliferation in liver, followed by parallel formation of DNA adducts and mutations. This study shows how transgenic models allow in vivo evaluation of mechanistically simultaneous endpoints.}, note = {0893-6692 Journal Article}, keywords = {*Escherichia, Adducts, Animals, Bacterial, Base, C57BL, Cell, coli, Division/drug, DNA, effects, Gov't, Inbred, Liver/cytology/*drug, Methylcholanthrene/*toxicity, Mice, Mutagens/*toxicity, Mutation, Non-U.S., Organ, Primers, Proteins, Proteins/genetics, Repressor, Sequence, Support, transgenic, Weight}, pubstate = {published}, tppubtype = {article} } @article{, title = {Rational drug design and high-throughput techniques for RNA targets}, author = {T Hermann and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10903381}, isbn = {10903381}, year = {2000}, date = {2000-01-01}, journal = {Comb Chem High Throughput Screen}, volume = {3}, number = {3}, pages = {219-234}, abstract = {RNA molecules are the only known molecules which possess the double property of being depository of genetic information, like DNA, and of displaying catalytic activities, like protein enzymes. RNA molecules intervene in all steps of gene expression and in many other biological activities. Like proteins, RNAs achieve those biological functions by adopting intricate three-dimensional folds and architectures. Further, as in protein sequences, RNA sequences contain signatures specific for three-dimensional motifs which participate in recognition and binding. In regulatory pathways, RNA molecules exist in equilibria between transient structures differentially stabilized by effectors such as proteins or cofactors. Therefore, RNA molecules display their potential as drug targets on different levels, namely in three-dimensional folds, in structural equilibria and in RNA-protein interfaces. Several examples will be described together with the already available techniques for combinatorial synthesis and high-throughput screening of potential drug and target RNA molecules.}, note = {1386-2073 Journal Article Review Review, Tutorial}, keywords = {Base Sequence *Combinatorial Chemistry Techniques *Drug Design Drug Evaluation, Molecular Molecular Sequence Data RNA/chemistry/*drug effects, Preclinical/*methods Models, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Differential gene expression in mesothelioma}, author = { B. H. Rihn and S. Mohr and S. A. McDowell and S. Binet and J. Loubinoux and F. Galateau and G. Keith and G. D. Leikauf}, year = {2000}, date = {2000-01-01}, journal = {FEBS Lett}, volume = {480}, number = {2-3}, pages = {95-100}, abstract = {To investigate the molecular events controlling malignant transformation of human pleural cells, we compared constitutive gene expression of mesothelioma cells to that of pleural cells. Using cDNA microarray and high-density filter array, we assessed expression levels of > 6500 genes. Most of the highly expressed transcripts were common to both cell lines and included genes associated with stress response and DNA repair, outcomes consistent with the radio- and chemo-resistance of mesothelioma. Interestingly, of the fewer than 300 genes that differed between cell lines, most functioned in (i) macromolecule stability, (ii) cell adhesion and recognition, (iii) cell migration (invasiveness), and (iv) extended cell division. Expression levels of several of these genes were confirmed by RT-PCR and could be useful as diagnostic markers of human mesothelioma.}, note = {0014-5793 Journal Article}, keywords = {*Gene, Adhesion, Analysis/methods, Array, Cell, Cells, Chain, Cultured, Cycle, Division, Expression, Gene, Human, Invasiveness, Mesothelioma/*genetics/metabolism, Neoplasm, Neoplastic, Oligonucleotide, Oxidative, Polymerase, Profiling, Proteins/metabolism, Reaction, Regulation, Reverse, Sequence, Stress, Transcriptase, tumor, Xenobiotics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Search for characteristic structural features of mammalian mitochondrial tRNAs}, author = {M Helm and H Brule and D Friede and R Giege and D Putz and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11073213}, isbn = {11073213}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {10}, pages = {1356-1379}, abstract = {A number of mitochondrial (mt) tRNAs have strong structural deviations from the classical tRNA cloverleaf secondary structure and from the conventional L-shaped tertiary structure. As a consequence, there is a general trend to consider all mitochondrial tRNAs as "bizarre" tRNAs. Here, a large sequence comparison of the 22 tRNA genes within 31 fully sequenced mammalian mt genomes has been performed to define the structural characteristics of this specific group of tRNAs. Vertical alignments define the degree of conservation/variability of primary sequences and secondary structures and search for potential tertiary interactions within each of the 22 families. Further horizontal alignments ascertain that, with the exception of serine-specific tRNAs, mammalian mt tRNAs do fold into cloverleaf structures with mostly classical features. However, deviations exist and concern large variations in size of the D- and T-loops. The predominant absence of the conserved nucleotides G18G19 and T54T55C56, respectively in these loops, suggests that classical tertiary interactions between both domains do not take place. Classification of the tRNA sequences according to their genomic origin (G-rich or G-poor DNA strand) highlight specific features such as richness/poorness in mismatches or G-T pairs in stems and extremely low G-content or C-content in the D- and T-loops. The resulting 22 "typical" mammalian mitochondrial sequences built up a phylogenetic basis for experimental structural and functional investigations. Moreover, they are expected to help in the evaluation of the possible impacts of those point mutations detected in human mitochondrial tRNA genes and correlated with pathologies.}, note = {1355-8382 Journal Article}, keywords = {Acylation Animals Base Pairing Base Sequence *Computational Biology Escherichia coli/genetics Genome Human Molecular Sequence Data Multigene Family *Nucleic Acid Conformation RNA/*chemistry/genetics RNA Stability RNA, Amino Acid-Specific/*chemistry/genetics Regulatory Sequences, FLORENTZ, Non-U.S. Gov't Variation (Genetics), Nucleic Acid/genetics Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {La cristallogenèse des macromolécules biologiques.}, author = {R Giege and C Sauter and D W Zhu and J D Ng and B Lorber}, editor = {J Samarut}, url = {http://cj.sauter.free.fr/xtal/abstracts.html#31}, year = {2000}, date = {2000-01-01}, booktitle = {Images de la Recherche en Biologie Structurale}, pages = {145-151}, publisher = {Editions du CNRS, Paris}, abstract = {The goal of macromolecular crystallogenesis is to understand and control the crystallisation of proteins and other macromolecular compounds. This survey presents the novel trends in the field and discusses physical methods employed to characterise crystallisation. Similarities and differences with crystal growth of small molecules are emphasised and ways to obtain crystals of higher perfection given. Crystal engineering perspectives and frontiers towards which crystallogenesis leads are outlined.}, keywords = {SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Cleavage of RNA with synthetic ribonuclease mimics}, author = {R Giege and B Felden and M A Zenkova and V N Sil'nikov and V V Vlassov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10889986}, isbn = {10889986}, year = {2000}, date = {2000-01-01}, journal = {Methods Enzymol}, volume = {318}, pages = {147-165}, note = {0076-6879 Journal Article}, keywords = {Asp/chemistry Ribonuclease, Base Sequence Electrophoresis, Chemical Molecular Sequence Data Nucleic Acid Conformation Nucleic Acid Hybridization Phosphorylation Plasmids/metabolism RNA/chemistry/*metabolism RNA, Non-U.S. Gov't, Pancreatic/chemistry/pharmacology Ribonucleases/*chemistry/pharmacology Saccharomyces cerevisiae/genetics Spectrophotometry Support, Polyacrylamide Gel Genetic Techniques Hydrolysis Imidazoles/pharmacology Models, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vivo selection of lethal mutations reveals two functional domains in arginyl-tRNA synthetase}, author = {R Geslain and F Martin and B Delagoutte and J Cavarelli and J Gangloff and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10744027}, isbn = {10744027}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {3}, pages = {434-448}, abstract = {Using random mutagenesis and a genetic screening in yeast, we isolated 26 mutations that inactivate Saccharomyces cerevisiae arginyl-tRNA synthetase (ArgRS). The mutations were identified and the kinetic parameters of the corresponding proteins were tested after purification of the expression products in Escherichia coli. The effects were interpreted in the light of the crystal structure of ArgRS. Eighteen functional residues were found around the arginine-binding pocket and eight others in the carboxy-terminal domain of the enzyme. Mutations of these residues all act by strongly impairing the rates of tRNA charging and arginine activation. Thus, ArgRS and tRNA(Arg) can be considered as a kind of ribonucleoprotein, where the tRNA, before being charged, is acting as a cofactor that activates the enzyme. Furthermore, by using different tRNA(Arg) isoacceptors and heterologous tRNA(Asp), we highlighted the crucial role of several residues of the carboxy-terminal domain in tRNA recognition and discrimination.}, note = {1355-8382 Journal Article}, keywords = {Arginine-tRNA Ligase/chemistry/*genetics Cloning, ERIANI, Fungal Genes, Fungal/genetics Kinetics Models, Lethal/*genetics Genes, Molecular Fungal Proteins/biosynthesis/genetics Gene Expression Regulation, Molecular Mutation/*genetics Peptide Fragments/chemistry/genetics Saccharomyces cerevisiae/enzymology/genetics Support, Non-U.S. Gov't, Structural, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A domain in the N-terminal extension of class IIb eukaryotic aminoacyl-tRNA synthetases is important for tRNA binding}, author = {M Frugier and L Moulinier and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10811628}, isbn = {10811628}, year = {2000}, date = {2000-01-01}, journal = {EMBO J}, volume = {19}, number = {10}, pages = {2371-2380}, abstract = {Cytoplasmic aspartyl-tRNA synthetase (AspRS) from Saccharomyces cerevisiae is a homodimer of 64 kDa subunits. Previous studies have emphasized the high sensitivity of the N-terminal region to proteolytic cleavage, leading to truncated species that have lost the first 20-70 residues but that retain enzymatic activity and dimeric structure. In this work, we demonstrate that the N-terminal extension in yeast AspRS participates in tRNA binding and we generalize this finding to eukaryotic class IIb aminoacyl-tRNA synthetases. By gel retardation studies and footprinting experiments on yeast tRNA(Asp), we show that the extension, connected to the anticodon-binding module of the synthetase, contacts tRNA on the minor groove side of its anticodon stem. Sequence comparison of eukaryotic class IIb synthetases identifies a lysine-rich 11 residue sequence ((29)LSKKALKKLQK(39) in yeast AspRS with the consensus xSKxxLKKxxK in class IIb synthetases) that is important for this binding. Direct proof of the role of this sequence comes from a mutagenesis analysis and from binding studies using the isolated peptide.}, note = {0261-4189 Journal Article}, keywords = {Amino Acid Sequence Amino Acyl-tRNA Ligases/chemistry/*metabolism Aspartate-tRNA Ligase/chemistry/metabolism Molecular Sequence Data RNA, ERIANI, FRUGIER, Fungal/metabolism RNA, Non-U.S. Gov't, Transfer/*metabolism Saccharomyces cerevisiae/*metabolism Sequence Alignment Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identity of tRNA for yeast tyrosyl-tRNA synthetase: tyrosylation is more sensitive to identity nucleotides than to structural features}, author = {P Fechter and J Rudinger-Thirion and A Théobald-Dietrich and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10677221}, isbn = {10677221}, year = {2000}, date = {2000-01-01}, journal = {Biochemistry}, volume = {39}, number = {7}, pages = {1725-1733}, abstract = {The specific aminoacylation of tRNA by yeast tyrosyl-tRNA synthetase does not rely on the presence of modified residues in tRNA(Tyr), although such residues stabilize its structure. Thus, the major tyrosine identity determinants were searched by the in vitro approach using unmodified transcripts produced by T7 RNA polymerase. On the basis of the tyrosylation efficiency of tRNA variants, the strongest determinants are base pair C1-G72 and discriminator residue A73 (the 5'-phosphoryl group on C1, however, is unimportant for tyrosylation). The three anticodon bases G34, U35, and A36 contribute also to the tyrosine identity, but to a lesser extent, with G34 having the most pronounced effect. Mutation of the GUA tyrosine anticodon into a CAU methionine anticodon, however, leads to a loss of tyrosylation efficiency similar to that obtained after mutation of the C1-G72 or A73 determinants. Transplantation of the six determinants into four different tRNA frameworks and activity assays on heterologous Escherichia coli and Methanococcus jannaschii tRNA(Tyr) confirmed the completeness of the tyrosine set and the eukaryotic character of the C1-G72 base pair. On the other hand, it was found that tyrosine identity in yeast does not rely on fine architectural features of the tRNA, in particular the size and sequence of the D-loop. Noticeable, yeast TyrRS efficiently charges a variant of E. coli tRNA(Tyr) with a large extra-region provided its G1-C72 base pair is changed to a C1-G72 base pair. Finally, tyrosylation activity is compatible with a +1 shift of the anticodon in the 3'-direction but is strongly inhibited if this shift occurs in the opposite 5'-direction.}, note = {0006-2960 Journal Article}, keywords = {Acylation Anticodon/chemistry/metabolism Base Sequence Escherichia coli/enzymology/genetics Heat Methanococcus/enzymology/genetics Molecular Mimicry Molecular Sequence Data Nucleic Acid Denaturation RNA Processing, Fungal/chemistry/*metabolism RNA, Non-U.S. Gov't Tyrosine/chemistry/*metabolism Tyrosine-tRNA Ligase/chemistry/*metabolism, Post-Transcriptional RNA, Transfer, Tyr/chemistry/*metabolism Saccharomyces cerevisiae/*enzymology/genetics Structure-Activity Relationship Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural analysis of new local features in SECIS RNA hairpins}, author = {D Fagegaltier and A Lescure and R Walczak and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10908323}, isbn = {10908323}, year = {2000}, date = {2000-01-01}, journal = {Nucleic Acids Res}, volume = {28}, number = {14}, pages = {2679-2689}, abstract = {Decoding of the UGA selenocysteine codon for selenoprotein translation requires the SECIS element, a stem-loop motif in the 3'-UTR of the mRNA carrying short or large apical loops. In previous structural studies, we derived a secondary structure model for SECIS RNAs with short apical loops. Work from others proposed that intra-apical loop base pairing can occur in those SECIS that possess large apical loops, yielding form 2 SECIS versus the form 1 with short loops. In this work, SECIS elements arising from eight different selenoprotein mRNAs were assayed by enzymatic and/or chemical probing showing that seven can adopt form 2. Further, database searches led to the discovery in drosophila and zebrafish of SECIS elements in the selenophosphate synthetase 2, type 1 deiodinase and SelW mRNAs. Alignment of SECIS sequences not only highlighted the predominance of form 2 but also made it possible to classify the SECIS elements according to the type of selenoprotein mRNA they belong to. Interestingly, the alignment revealed that an unpaired adenine, previously thought to be invariant, is replaced by a guanine in four SECIS elements. Tested in vivo, neither the A to G nor the A to U changes at this position greatly affected the activity while the most detrimental effect was provided by a C. The putative contribution of the various SECIS motifs to function and ligand binding is discussed.}, note = {1362-4962 Journal Article}, keywords = {Animals Base Sequence COS Cells DNA/chemistry/genetics DNA, DNA Support, Factual Drosophila melanogaster/genetics Glutathione Peroxidase/genetics/metabolism Human Mice Molecular Sequence Data Mutagenesis, LESCURE, Non-U.S. Gov't Xenopus laevis, Nucleic Acid/*genetics Selenocysteine/*genetics/metabolism Sequence Alignment Sequence Analysis, Recombinant/genetics/metabolism Databases, Site-Directed Nucleic Acid Conformation Phosphotransferases/genetics RNA/chemistry/*genetics Rats Regulatory Sequences, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of mSelB, a novel mammalian elongation factor for selenoprotein translation}, author = {D Fagegaltier and N Hubert and K Yamada and T Mizutani and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10970870}, isbn = {10970870}, year = {2000}, date = {2000-01-01}, journal = {EMBO J}, volume = {19}, number = {17}, pages = {4796-4805}, abstract = {Decoding of UGA selenocysteine codons in eubacteria is mediated by the specialized elongation factor SelB, which conveys the charged tRNA(Sec) to the A site of the ribosome, through binding to the SECIS mRNA hairpin. In an attempt to isolate the eukaryotic homolog of SelB, a database search in this work identified a mouse expressed sequence tag containing the complete cDNA encoding a novel protein of 583 amino acids, which we called mSelB. Several lines of evidence enabled us to establish that mSelB is the bona fide mammalian elongation factor for selenoprotein translation: it binds GTP, recognizes the Sec-tRNA(Sec) in vitro and in vivo, and is required for efficient selenoprotein translation in vivo. In contrast to the eubacterial SelB, the recombinant mSelB alone is unable to bind specifically the eukaryotic SECIS RNA hairpin. However, complementation with HeLa cell extracts led to the formation of a SECIS-dependent complex containing mSelB and at least another factor. Therefore, the role carried out by a single elongation factor in eubacterial selenoprotein translation is devoted to two or more specialized proteins in eukaryotes.}, note = {0261-4189 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Animals Bacterial Proteins/chemistry/*metabolism/physiology Caenorhabditis elegans/genetics Drosophila/genetics Hela Cells Human Mice Molecular Sequence Data Peptide Elongation Factors/chemistry/*metabolism/physiology Protein Binding Proteins/*genetics RNA, Amino Acyl/metabolism Sequence Homology, Genetic/*physiology, Non-U.S. Gov't Translation, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The selenocysteine insertion sequence binding protein SBP is different from the Y-box protein dbpB}, author = {D Fagegaltier and N Hubert and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10727766}, isbn = {10727766}, year = {2000}, date = {2000-01-01}, journal = {Biochimie}, volume = {82}, number = {2}, pages = {117-122}, abstract = {In eukaryotes, translation of internal UGA selenocysteine codons requires the SECIS stem-loop structure in the 3'UTR of selenoprotein mRNAs. In an earlier work, we identified SBP as a selenocysteine insertion sequence (SECIS)-binding protein. Here, the yeast three-hybrid screen was employed to capture the cDNA of SBP. One candidate, satisfying the genetic screens, was identified as the already known dbpB protein. Although it was also found by another group, but with a different strategy, to carry SECIS-binding activity, further experiments enabled us to show that dbpB was unable to bind the SECIS element in vitro. Altogether, our findings led us to conclude that, under our conditions, dbpB and SBP are two distinct proteins.}, note = {0300-9084 Journal Article}, keywords = {Animals Base Sequence *CCAAT-Enhancer-Binding Proteins COS Cells Cloning, Molecular DNA-Binding Proteins/genetics/*metabolism Molecular Sequence Data Nucleic Acid Conformation RNA/metabolism RNA-Binding Proteins/genetics/*metabolism Rats Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The crystal structure of UUCG tetraloop}, author = {E Ennifar and A Nikulin and S Tishchenko and A Serganov and N Nevskaya and M Garber and B Ehresmann and C Ehresmann and S Nikonov and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11071808}, isbn = {11071808}, year = {2000}, date = {2000-01-01}, journal = {J Mol Biol}, volume = {304}, number = {1}, pages = {35-42}, abstract = {All large structured RNAs contain hairpin motifs made of a stem closed by several looped nucleotides. The most frequent loop motif is the UUCG one. This motif belongs to the tetraloop family and has the peculiarity of being highly thermodynamically stable. Here, we report the first crystal structure of two UUCG tetraloops embedded in a larger RNA-protein complex solved at 2.8 A resolution. The two loops present in the asymmetric unit are in a different crystal packing environment but, nevertheless, have an identical conformation. The observed structure is globally close to that obtained in solution by nuclear magnetic resonance. However, subtle differences point to a more detailed picture of the role played by 2'-hydroxyl groups in stabilising this tetraloop.}, note = {0022-2836 Journal Article}, keywords = {16S/*chemistry/genetics/*metabolism Ribosomal Proteins/chemistry/*metabolism Solvents Support, Base Sequence Crystallography, Biomolecular *Nucleic Acid Conformation RNA Stability RNA, ENNIFAR, Molecular Molecular Sequence Data Motion Nuclear Magnetic Resonance, Non-U.S. Gov't Thermodynamics, Ribosomal, Unité ARN, X-Ray Hydrogen Bonding Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transfer RNA-mediated editing in threonyl-tRNA synthetase. The class II solution to the double discrimination problem}, author = {A Dock-Bregeon and R Sankaranarayanan and P Romby and J Caillet and M Springer and B Rees and C S Francklyn and C Ehresmann and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11136973}, isbn = {11136973}, year = {2000}, date = {2000-01-01}, journal = {Cell}, volume = {103}, number = {6}, pages = {877-884}, abstract = {Threonyl-tRNA synthetase, a class II synthetase, uses a unique zinc ion to discriminate against the isosteric valine at the activation step. The crystal structure of the enzyme with an analog of seryl adenylate shows that the noncognate serine cannot be fully discriminated at that step. We show that hydrolysis of the incorrectly formed ser-tRNA(Thr) is performed at a specific site in the N-terminal domain of the enzyme. The present study suggests that both classes of synthetases use effectively the ability of the CCA end of tRNA to switch between a hairpin and a helical conformation for aminoacylation and editing. As a consequence, the editing mechanism of both classes of synthetases can be described as mirror images, as already seen for tRNA binding and amino acid activation.}, note = {0092-8674 Journal Article}, keywords = {Acylation Amino Acid Activation Binding Sites Crystallography, Amino Acyl/chemistry/*metabolism Serine/metabolism Support, Molecular Mutation *Nucleic Acid Conformation Protein Structure, Non-U.S. Gov't Threonine/metabolism Threonine-tRNA Ligase/*chemistry/*genetics/metabolism Zinc/metabolism, ROMBY, Tertiary *RNA Editing RNA, Transfer, Unité ARN, X-Ray Kinetics Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization and preliminary X-ray crystallographic analysis of yeast arginyl-tRNA synthetase-yeast tRNAArg complexes}, author = {B Delagoutte and G Keith and D Moras and J Cavarelli}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10739930}, isbn = {10739930}, year = {2000}, date = {2000-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {56}, number = {Pt 4}, pages = {492-494}, abstract = {Three different crystal forms of complexes between arginyl-tRNA synthetase from the yeast Saccharomyces cerevisae (yArgRS) and the yeast second major tRNA(Arg) (tRNA(Arg)(ICG)) isoacceptor have been crystallized by the hanging-drop vapour-diffusion method in the presence of ammonium sulfate. Crystal form II, which diffracts beyond 2.2 A resolution at the European Synchrotron Radiation Facility ID14-4 beamline, belongs to the orthorhombic space group P2(1)2(1)2, with unit-cell parameters a = 129.64}, note = {0907-4449 Journal Article}, keywords = {Arg/*chemistry/isolation & purification/*metabolism Saccharomyces cerevisiae/enzymology/genetics Support, Arginine-tRNA Ligase/*chemistry/isolation & purification/*metabolism Crystallization Crystallography, Fungal/chemistry/isolation & purification/metabolism RNA, Non-U.S. Gov't, Transfer, Unité ARN, X-Ray RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence analysis and modular organization of threonyl-tRNA synthetase from Thermus thermophilus and its interrelation with threonyl-tRNA synthetases of other origins}, author = {V Cura and D Moras and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10632708}, isbn = {10632708}, year = {2000}, date = {2000-01-01}, journal = {Eur J Biochem}, volume = {267}, number = {2}, pages = {379-393}, abstract = {The gene encoding threonyl-tRNA synthetase (Thr-tRNA synthetase) from the extreme thermophilic eubacterium Thermus thermophilus HB8 has been cloned and sequenced. The ORF encodes a polypeptide chain of 659 amino acids (Mr 75 550) that shares strong similarities with other Thr-tRNA synthetases. Comparative analysis with the three-dimensional structure of other subclass IIa synthetases shows it to be organized into four structural modules: two N-terminal modules specific to Thr-tRNA synthetases, a catalytic core and a C-terminal anticodon-binding module. Comparison with the three-dimensional structure of Escherichia coli Thr-tRNA synthetase in complex with tRNAThr enabled identification of the residues involved in substrate binding and catalytic activity. Analysis by atomic absorption spectrometry of the enzyme overexpressed in E. coli revealed the presence in each monomer of one tightly bound zinc atom, which is essential for activity. Despite strong similarites in modular organization, Thr-tRNA synthetases diverge from other subclass IIa synthetases on the basis of their N-terminal extensions. The eubacterial and eukaryotic enzymes possess a large extension folded into two structural domains, N1 and N2, that are not significantly similar to the shorter extension of the archaebacterial enzymes. Investigation of a truncated Thr-tRNA synthetase demonstrated that domain N1 is not essential for tRNA charging. Thr-tRNA synthetase from T. thermophilus is of the eubacterial type, in contrast to other synthetases from this organism, which exhibit archaebacterial characteristics. Alignments show conservation of part of domain N2 in the C-terminal moiety of Ala-tRNA synthetases. Analysis of the nucleotide sequence upstream from the ORF showed the absence of both any anticodon-like stem-loop structure and a loop containing sequences complementary to the anticodon and the CCA end of tRNAThr. This means that the expression of Thr-tRNA synthetase in T. thermophilus is not regulated by the translational and trancriptional mechanisms described for E. coli thrS and Bacillus subtilis thrS and thrZ. Here we discuss our results in the context of evolution of the threonylation systems and of the position of T. thermophilus in the phylogenic tree.}, note = {0014-2956 Journal Article}, keywords = {Amino Acid Motifs Amino Acid Sequence Bacterial Proteins/*genetics/*metabolism Base Sequence Cloning, Bacterial Molecular Sequence Data Phylogeny Regulatory Sequences, Molecular Enzyme Stability Escherichia coli/enzymology/genetics Evolution, Molecular Gene Expression Regulation, Non-U.S. Gov't Thermus thermophilus/*enzymology Threonine-tRNA Ligase/*genetics/*metabolism Zinc/metabolism, Nucleic Acid Sequence Analysis Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A three-dimensional perspective on exon binding by a group II self-splicing intron}, author = {M Costa and F Michel and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10990464}, isbn = {10990464}, year = {2000}, date = {2000-01-01}, journal = {EMBO J}, volume = {19}, number = {18}, pages = {5007-5018}, abstract = {We have used chemical footprinting, kinetic dissection of reactions and comparative sequence analysis to show that in self-splicing introns belonging to subgroup IIB, the sites that bind the 5' and 3' exons are connected to one another by tertiary interactions. This unanticipated arrangement, which contrasts with the direct covalent linkage that prevails in the other major subdivision of group II (subgroup IIA), results in a unique three-dimensional architecture for the complex between the exons, their binding sites and intron domain V. A key feature of the modeled complex is the presence of several close contacts between domain V and one of the intron-exon pairings. These contacts, whose existence is supported by hydroxyl radical footprinting, provide a structural framework for the known role of domain V in catalysis and its recently demonstrated involvement in binding of the 5' exon.}, note = {0261-4189 Journal Article}, keywords = {Base Sequence Catalysis *Exons Hydroxyl Radical *Introns Kinetics *Models, Catalytic/metabolism Support, Genetic, Genetic Molecular Sequence Data Nucleic Acid Conformation *RNA Splicing RNA, Non-U.S. Gov't Support, P.H.S. Temperature Transcription, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Probing RNA structure and RNA-ligand complexes with chemical probes}, author = {C Brunel and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10889976}, isbn = {10889976}, year = {2000}, date = {2000-01-01}, journal = {Methods Enzymol}, volume = {318}, pages = {3-21}, note = {0076-6879 Journal Article Review Review, Tutorial}, keywords = {Crystallography, Genetic, ROMBY, Unité ARN, X-Ray *Ligands Nucleic Acid Conformation Nucleotides/chemistry/metabolism Proteins/chemistry RNA/*chemistry/drug effects/*metabolism Transcription}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vitro evidence for the interaction of tRNA(3)(Lys) with U3 during the first strand transfer of HIV-1 reverse transcription}, author = {F Brule and G Bec and G Keith and S F Le Grice and B P Roques and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10606665}, isbn = {10606665}, year = {2000}, date = {2000-01-01}, journal = {Nucleic Acids Res}, volume = {28}, number = {2}, pages = {634-640}, abstract = {Over the course of its evolution, HIV-1 has taken maximum advantage of its tRNA(3)(Lys)primer by utilizing it in several steps of reverse transcription. Here, we have identified a conserved nonanucleotide sequence in the U3 region of HIV-1 RNA that is complementary to the anticodon stem of tRNA(3)(Lys). In order to test its possible role in the first strand transfer reaction, we applied an assay using a donor RNA corresponding to the 5'-part and an acceptor RNA spanning the 3'-part of HIV-1 RNA. In addition, we constructed two acceptor RNAs in which the nonanucleotide sequence complementary to tRNA(3)(Lys)was either substituted (S) or deleted (Delta). We used either natural tRNA(3)(Lys)or an 18 nt DNA as primer and measured the efficiency of (-) strand strong stop DNA transfer in the presence of wild-type, S or Delta acceptor RNA. Mutations in U3 did not decrease the transfer efficiency when reverse transcription was primed with the 18mer DNA. However, they significantly reduced the strand transfer efficiency in the tRNA(3)(Lys)-primed reactions. This reduction was also observed in the presence of nucleocapsid protein. These results suggest that tRNA(3)(Lys)increases (-) strand strong stop transfer by interacting with the U3 region of the genomic RNA. Sequence comparisons suggest that such long range interactions also exist in other lentiviruses.}, note = {1362-4962 Journal Article}, keywords = {Base Sequence HIV-1 Reverse Transcriptase/*metabolism Nucleic Acid Conformation Polymerase Chain Reaction RNA, Genetic, Lys/*metabolism RNA, MARQUET, Non-U.S. Gov't *Transcription, Transfer, Unité ARN, Viral/chemistry/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Positive and negative mutant selection in the human histone hairpin- binding protein using the yeast three-hybrid system.}, author = { F. Martin and F. Michel and D. Zenklusen and B. Muller and D. Schumperli}, year = {2000}, date = {2000-01-01}, journal = {Nucleic Acids Res}, volume = {28}, number = {7}, pages = {1594-603.}, abstract = {We have used the yeast three-hybrid system in a positive selection for mutants of the human histone hairpin-binding protein (HBP) capable of interacting with non-canonical hairpins and in a negative selection for loss-of-binding mutants. Interestingly, all mutations from the positive selection are located in the N- and C-terminal regions flanking a minimal RNA-binding domain (RBD) previously defined between amino acids 126 and 198. Further, in vitro binding studies demonstrate that the RBD, which shows no obvious similarity to other RNA-binding motifs, has a relaxed sequence specificity compared to full-length HBP, allowing it to bind to mutant hairpin RNAs not normally found in histone genes. These findings indicate that the sequences flanking the RBD are important for restricting binding to the highly conserved histone hairpin structure. Among the loss-of-binding mutations, about half are nonsense mutations distributed throughout the N-terminal part and the RBD whereas the other half are missense mutations restricted to the RBD. Whereas the nonsense mutations permit a more precise definition of the C-terminal border of the RBD, the missense mutations identify critical residues for RNA binding within the RBD.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Probing the structure of RNAIII, the Staphylococcus aureus agr regulatory RNA, and identification of the RNA domain involved in repression of protein A expression}, author = {Y Benito and F A Kolb and P Romby and G Lina and J Etienne and F Vandenesch}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10836788}, isbn = {10836788}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {5}, pages = {668-679}, abstract = {RNAIII, a 514-nt RNA molecule, regulates the expression of many Staphylococcus aureus genes encoding exoproteins and cell-wall-associated proteins. We have studied the structure of RNAIII in solution, using a combination of chemical and enzymatic probes. A model of the secondary structure was derived from experimental data with the help of computer simulation of RNA folding. The model contains 14 hairpin structures connected by unpaired nucleotides. The data also point to three helices formed by distant nucleotides that close off structural domains. This model was generally compatible with the results of in vivo probing experiments with dimethylsulfate in late exponential-phase cultures. Toe-printing experiments revealed that the ribosome binding site of hld, which is encoded by RNAIII, was accessible to the Escherichia coli 30S ribosomal subunit, suggesting that the in vitro structure represented a translatable form of RNAIII. We also found that, within the 3' end of RNAIII, the conserved hairpin 13 and the terminator form an intrinsic structural domain that exerts specific regulatory activity on protein A gene expression.}, note = {1355-8382 Journal Article}, keywords = {Antisense/*chemistry/genetics/metabolism RNA, Bacterial Models, Bacterial/*chemistry/genetics/metabolism Ribosomes/metabolism Staphylococcal Protein A/*genetics Staphylococcus aureus/*chemistry/*genetics/metabolism Support, Base Sequence Binding Sites/genetics DNA Primers/genetics Escherichia coli/metabolism Gene Expression Genes, Molecular Molecular Sequence Data Nucleic Acid Conformation RNA, Non-U.S. Gov't, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The crystal structure of HIV reverse-transcription primer tRNA(Lys,3) shows a canonical anticodon loop}, author = {P Benas and G Bec and G Keith and R Marquet and C Ehresmann and B Ehresmann and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11073212}, isbn = {11073212}, year = {2000}, date = {2000-01-01}, journal = {RNA}, volume = {6}, number = {10}, pages = {1347-1355}, abstract = {We have solved to 3.3 A resolution the crystal structure of the HIV reverse-transcription primer tRNA(Lys,3). The overall structure is exactly comparable to the well-known L-shape structure first revealed by yeast tRNA(Phe). In particular, it unambiguously shows a canonical anticodon loop. This contradicts previous results in short RNA fragment studies and leads us to conclude that neither frameshifting specificities of tRNA(Lys) nor tRNA(Lys,3) primer selection by HIV are due to a specific three-dimensional anticodon structure. Comparison of our structure with the results of an NMR study on a hairpin representing a nonmodified anticodon stem-loop makes plausible the conclusion that chemical modifications of the wobble base U34 to 5-methoxycarbonyl-methyl-2-thiouridine and of A37 to 2-methylthio-N-6-threonylcarbamoyl-adenosine would be responsible for a canonical 7-nt anticodon-loop structure, whereas the unmodified form would result in a noncanonical UUU short triloop. The hexagonal crystal packing is remarkable and shows tight dimers of tRNAs forming a right-handed double superhelix. Within the dimers, the tRNAs are associated head-to-tail such that the CCA end of one tRNA interacts with the anticodon of the symmetry-related tRNA. This provides us with a partial view of a codon-anticodon interaction and gives insights into the positioning of residue 37, and of its posttranscriptional modifications, relative to the first base of the codon.}, note = {1355-8382 Journal Article}, keywords = {Animals Anticodon/*chemistry/genetics Base Sequence Cattle Chickens/*genetics Crystallography, Biomolecular *Nucleic Acid Conformation RNA/*chemistry/genetics RNA, Lys/*chemistry/genetics Rabbits Support, MARQUET, Molecular Molecular Sequence Data Nuclear Magnetic Resonance, Non-U.S. Gov't, Transfer, Unité ARN, X-Ray HIV-1/*genetics Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Thermus thermophilus contains an eubacterial and an archaebacterial aspartyl-tRNA synthetase}, author = {H D Becker and H Roy and L Moulinier and M H Mazauric and G Keith and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10727213}, isbn = {10727213}, year = {2000}, date = {2000-01-01}, journal = {Biochemistry}, volume = {39}, number = {12}, pages = {3216-3230}, abstract = {Thermus thermophilus possesses two aspartyl-tRNA synthetases (AspRSs), AspRS1 and AspRS2, encoded by distinct genes. Alignment of the protein sequences with AspRSs of other origins reveals that AspRS1 possesses the structural features of eubacterial AspRSs, whereas AspRS2 is structurally related to the archaebacterial AspRSs. The structural dissimilarity between the two thermophilic AspRSs is correlated with functional divergences. AspRS1 aspartylates tRNA(Asp) whereas AspRS2 aspartylates tRNA(Asp), and tRNA(Asn) with similar efficiencies. Since Asp bound on tRNA(Asn) is converted into Asn by a tRNA-dependent aspartate amidotransferase, AspRS2 is involved in Asn-tRNA(Asn) formation. These properties relate functionally AspRS2 to archaebacterial AspRSs. The structural basis of the dual specificity of T. thermophilus tRNA(Asn) was investigated by comparing its sequence with those of tRNA(Asp) and tRNA(Asn) of strict specificity. It is shown that the thermophilic tRNA(Asn) contains the elements defining asparagine identity in Escherichia coli, part of which being also the major elements of aspartate identity, whereas minor elements of this identity are missing. The structural context that permits expression of aspartate and asparagine identities by tRNA(Asn) and how AspRS2 accommodates tRNA(Asp) and tRNA(Asn) will be discussed. This work establishes a distinct structure-function relationship of eubacterial and archaebacterial AspRSs. The structural and functional properties of the two thermophilic AspRSs will be discussed in the context of the modern and primitive pathways of tRNA aspartylation and asparaginylation and related to the phylogenetic connexion of T. thermophilus to eubacteria and archaebacteria.}, note = {0006-2960 Journal Article}, keywords = {Amino Acid Support, Asn/genetics/metabolism RNA, Asp/metabolism Sequence Alignment Sequence Homology, Molecular Consensus Sequence Escherichia coli/enzymology/genetics Human Kinetics Molecular Sequence Data Peptide Fragments/chemistry RNA, Non-U.S. Gov't Support, P.H.S. Thermus thermophilus/*enzymology/genetics, Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The heterotrimeric Thermus thermophilus Asp-tRNA(Asn) amidotransferase can also generate Gln-tRNA(Gln)}, author = {H D Becker and B Min and C Jacobi and G Raczniak and J Pelaschier and H Roy and S Klein and D Kern and D Soll}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10913601}, isbn = {10913601}, year = {2000}, date = {2000-01-01}, journal = {FEBS Lett}, volume = {476}, number = {3}, pages = {140-144}, abstract = {Thermus thermophilus strain HB8 is known to have a heterodimeric aspartyl-tRNA(Asn) amidotransferase (Asp-AdT) capable of forming Asn-tRNA(Asn) [Becker, H.D. and Kern, D. (1998) Proc. Natl. Acad. Sci. USA 95, 12832-12837]. Here we show that, like other bacteria, T. thermophilus possesses the canonical set of amidotransferase (AdT) genes (gatA, gatB and gatC). We cloned and sequenced these genes, and constructed an artificial operon for overexpression in Escherichia coli of the thermophilic holoenzyme. The overproduced T. thermophilus AdT can generate Gln-tRNA(Gln) as well as Asn-tRNA(Asn). Thus, the T. thermophilus tRNA-dependent AdT is a dual-specific Asp/Glu-AdT resembling other bacterial AdTs. In addition, we observed that removal of the 44 carboxy-terminal amino acids of the GatA subunit only inhibits the Asp-AdT activity, leaving the Glu-AdT activity of the mutant AdT unaltered; this shows that Asp-AdT and Glu-AdT activities can be mechanistically separated.}, note = {0014-5793 Journal Article}, keywords = {Amino Acid Sequence Cloning, Amino Acyl/*metabolism Recombinant Proteins/chemistry/genetics/metabolism Sequence Deletion Substrate Specificity Support, Bacterial Molecular Sequence Data Nitrogenous Group Transferases/chemistry/genetics/*metabolism Protein Structure, Bacterial/metabolism RNA, Molecular Escherichia coli/genetics Genes, Non-U.S. Gov't Thermus thermophilus/*enzymology/genetics, Quaternary RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Water and ion binding around RNA and DNA (C,G) oligomers}, author = {P Auffinger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10903858}, isbn = {10903858}, year = {2000}, date = {2000-01-01}, journal = {J Mol Biol}, volume = {300}, number = {5}, pages = {1113-1131}, abstract = {The dynamics, hydration, and ion-binding features of two duplexes, the A(r(CG)(12)) and the B(d(CG)(12)), in a neutralizing aqueous environment with 0.25 M added KCl have been investigated by molecular dynamics (MD) simulations. The regular repeats of the same C=G base-pair motif have been exploited as a statistical alternative to long MD simulations in order to extend the sampling of the conformational space. The trajectories demonstrate the larger flexibility of DNA compared to RNA helices. This flexibility results in less well defined hydration patterns around the DNA than around the RNA backbone atoms. Yet, 22 hydration sites are clearly characterized around both nucleic acid structures. With additional results from MD simulations, the following hydration scale for C=G pairs can be deduced: A-DNA<RNA (+3 H(2)O) and B-DNA<RNA (+2 H(2)O). The calculated residence times of water molecules in the first hydration shell of the helices range from 0.5 to 1 ns, in good agreement with available experimental data. Such water molecules are essentially found in the vicinity of the phosphate groups and in the DNA minor groove. The calculated number of ions that break into the first hydration shell of the nucleic acids is close to 0.5 per base-pair for both RNA and DNA. These ions form contacts essentially with the oxygen atoms of the phosphate groups and with the guanine N7 and O6 atoms; they display residence times in the deep/major groove approaching 500 ps. Further, a significant sequence-dependent effect on ion binding has been noted. Despite slight structural differences, K(+) binds essentially to GpC and not to CpG steps. These results may be of importance for understanding various sequence-dependent binding affinities. Additionally, the data help to rationalize the experimentally observed differences in gel electrophoretic mobility between RNA and DNA as due to the difference in hydration (two water molecules in favor of RNA) rather than to strong ion-binding features, which are largely similar for both nucleic acid structures.}, note = {0022-2836 Journal Article}, keywords = {Base Pairing Binding Sites Computer Simulation DNA/*chemistry/metabolism Electrophoresis, Molecular Oligonucleotides/*chemistry/*metabolism Potassium/*metabolism RNA/*chemistry/metabolism Solvents Water/*metabolism, Polyacrylamide Gel Electrostatics Hydrogen Bonding Ions Kinetics Models, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA solvation: a molecular dynamics simulation perspective}, author = {P Auffinger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11754340}, isbn = {11754340}, year = {2000}, date = {2000-01-01}, journal = {Biopolymers}, volume = {56}, number = {4}, pages = {266-274}, abstract = {With the availability of accurate methods to treat the electrostatic long-range interactions, molecular dynamics simulations have resulted in refined dynamical models of the structure of the hydration shell around RNA motifs. The models reviewed here range from basic Watson-Crick to more specific noncanonical base pairs, from "simple" double helices to RNA molecules displaying more complex tertiary folds, and from DNA/RNA hybrid double helices to RNA hybrids formed with a chemically modified strand.}, note = {0006-3525 Journal Article}, keywords = {Base Pairing *Computer Simulation Electrostatics Hydrogen Bonding Ions/chemistry Models, Molecular RNA/*chemistry Solvents/*chemistry Thermodynamics Water/chemistry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Tecto-RNA: One-Dimensional Self-Assembly through Tertiary Interactions}, author = { L. Jaeger and N. B. Leontis}, year = {2000}, date = {2000-01-01}, journal = {Angew Chem Int Ed Engl}, volume = {39}, number = {14}, pages = {2521-2524}, abstract = {The modularity of natural RNA is the basis for the design of tecto-RNA, modular RNA units capable of directed self-assembly. One such modular association through specific RNA loop–receptor tertiary interactions, which leads to one-dimensional oligomer arrays, is demonstrated (see picture).}, keywords = {Chemistry, RNA, self-organisation, supramolecular}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genomic DNA methylation in the human trophoblast from recurrent spontaneous abortion-premilimary report}, author = { M. Jerzak and K. Potawski and G. Keith and J. A. Jakowicki and W. Baranovski}, year = {2000}, date = {2000-01-01}, journal = {Pol J Gyn Invest}, volume = {3}, pages = {9-12}, keywords = {ACL, GIEGE}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genotoxicity of 3-methylcholanthrene in liver of transgenic big Blue mice}, author = {B H Rihn and M C Bottin and C Coulais and R Rouget and N Monhoven and W Baranowski and A Edorh and G Keith}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11152559}, isbn = {11152559}, year = {2000}, date = {2000-01-01}, journal = {Environ Mol Mutagen}, volume = {36}, number = {4}, pages = {266-73}, abstract = {Transgenic mice provide a unique tool for studying the tissue specificity and mutagenic potential of chemicals. Because 3-methylcholanthrene (3MC) was found mutagenic in bacteria, clastogenic in bone marrow, and induces DNA adducts in animals, we were interested to determinine whether this xenobiotic provokes (1) cell proliferation, (2) transcriptional activity changes, (3) DNA adducts, and (4) hepatic mutations in transgenic Big Blue mice carrying the lambdaLIZ phage shuttle vector. Big Blue C57/Bl male mice were treated with a single intraperitoneal dose of 80 mg/kg 3MC for 1, 3, 6, 14, or 30 days. Cell proliferation was checked by 5-bromo-2-deoxyuridine labeling and immunohistochemical detection. The maximal increase of the mitotic index was evidenced after 3 days (2.9 times the control value; P < 0.01). The relative nucleus area, reflecting the transcriptional activity, was also the highest in the treated group after 3 days: 1.86 times the control value, on average (P < 0.01). Four major DNA adducts, determined according to the [(32)P]-postlabeling method, were evidenced in liver DNA of treated mice, 6 days after the treatment: the spot intensities increased in a time-dependent manner. The mutant frequency of liver DNA was the highest after 14 days: 20.3 +/- 2.9 x 10(-5) in the treated vs. 7.6 +/- 2.7 x 10(-5) in the control mice (P < 0.01). Sequencing of the lambda lacI mutant plaques showed mainly G:C --> T:A and C:G --> A:T transversions. In conclusion, 3MC at first induced nuclear enlargement and a slight increase of cell proliferation in liver, followed by parallel formation of DNA adducts and mutations. This study shows how transgenic models allow in vivo evaluation of mechanistically simultaneous endpoints.}, note = {0893-6692 Journal Article}, keywords = {Animals Bacterial Proteins/genetics Base Sequence Cell Division/drug effects DNA Adducts DNA Primers *Escherichia coli Proteins Liver/cytology/*drug effects Methylcholanthrene/*toxicity Mice Mice, Inbred C57BL Mice, Non-U.S. Gov't, Transgenic Mutagens/*toxicity Mutation Organ Weight Repressor Proteins/genetics Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Expression of an active form of recombinant Ty1 reverse transcriptase in Escherichia coli: a fusion protein containing the C-terminal region of the Ty1 integrase linked to the reverse transcriptase-RNase H domain exhibits polymerase and RNase H activities}, author = { M. Wilhelm and M. Boutabout and F. X. Wilhelm}, year = {2000}, date = {2000-01-01}, journal = {Biochem J}, volume = {348}, number = {Pt 2}, pages = {337-42}, abstract = {Replication of the Saccharomyces cerevisiae Ty1 retrotransposon requires a reverse transcriptase capable of synthesizing Ty1 DNA. The first description of an active form of a recombinant Ty1 enzyme with polymerase and RNase H activities is reported here. The Ty1 enzyme was expressed as a hexahistidine-tagged fusion protein in Escherichia coli to facilitate purification of the recombinant protein by metal-chelate chromatography. Catalytic activity of the recombinant protein was detected only when amino acid residues encoded by the integrase gene were added to the N-terminus of the reverse transcriptase-RNase H domain. This suggests that the integrase domain could play a role in proper folding of reverse transcriptase. Several biochemical properties of the Ty1 enzyme were analysed, including the effect of MgCl(2), NaCl, temperature and of the chain terminator dideoxy GTP on its polymerase activity. RNase H activity was examined by monitoring the cleavage of a RNA-DNA template-primer. Our results suggest that the distance between the RNase H and polymerase active sites corresponds to the length of a 14-nucleotide RNA-DNA heteroduplex. The recombinant protein produced in E. coli should be useful for further biochemical and structural analyses and for a better understanding of the role of integrase in the activation of reverse transcriptase.}, note = {0264-6021 Journal Article}, keywords = {&, Acid, affinity, Alignment, Amino, Calf, cerevisiae/*enzymology/*genetics, Chromatography, Cloning, Codon, coli, Comparative, Data, DNA, DNA/metabolism, Escherichia, Frames, Fusion, Genetic, Gov't, H, Heteroduplexes/metabolism, HIV-1, Homology, Integrases/chemistry/metabolism, Kinetics, Molecular, Non-U.S., Nucleic, Open, Polymerase/chemistry/isolation, Proteins/chemistry/isolation, purification/*metabolism, purification/metabolism, Reading, Recombinant, Retroelements/*genetics, Reverse, Ribonuclease, RNA-Directed, RNA/metabolism, Saccharomyces, Sequence, Study, Support, Templates, Terminator, Thymus/isolation, Transcriptase/chemistry}, pubstate = {published}, tppubtype = {article} } @article{green_necrotic_2000, title = {The necrotic gene in Drosophila corresponds to one of a cluster of three Serpin transcripts mapping at 43A1.2.}, author = {Clare Green and Elena A Levashina and C McKimmie and Timothy R Dafforn and Jean-Marc Reichhart and David Gubb}, year = {2000}, date = {2000-01-01}, journal = {Genetics}, volume = {156}, pages = {1117--1127}, abstract = {Mutants of the necrotic (nec) gene in Drosophila melanogaster die in the late pupal stage as pharate adults, or hatch as weak, but relatively normal-looking, flies. Adults develop black melanized spots on the body and leg joints, the abdomen swells with hemolymph and flies die within three or four days of eclosion. Pleiotropic nec phenotypes include melanization and cellular necrosis. These changes are consistent with activation of one, or more, proteolytic cascades. The nec gene corresponds to Spn43Ac, one of a cluster of three putative serine proteinase inhibitors at 43A1.2, on the right arm of chromosome two. Although serpins have been implicated in the activation of many diverse pathways, lack of an individual serpin rarely causes a detectable phenotype. Absence of Spn43Ac , however, gives a clear phenotype, which will allow a mutational analysis of critical features of the molecular structure of serpins.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{reichhart_toll_2000, title = {Toll Story}, author = {Jean-Marc Reichhart and Jean-Luc Imler}, year = {2000}, date = {2000-01-01}, journal = {Médecine Sciences: M/S}, volume = {16}, pages = {1439--1442}, keywords = {imler, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{hetru_androctonin_2000, title = {Androctonin, a hydrophilic disulphide-bridged non-haemolytic anti-microbial peptide: a plausible mode of action}, author = {Charles Hetru and L Letellier and Z Oren and Jules A Hoffmann and Y Shai}, issn = {0264-6021}, year = {2000}, date = {2000-01-01}, journal = {Biochem. J.}, volume = {345 Pt 3}, pages = {653--664}, abstract = {Androctonin is a 25-residue non-haemolytic anti-microbial peptide isolated from the scorpion Androctonus australis and contains two disulphide bridges. Androctonin is different from known native anti-microbial peptides, being a relatively hydrophilic and non-amphipathic molecule. This raises the possibility that the target of androctonin might not be the bacterial membrane, shown to be a target for most amphipathic lytic peptides. To shed light on its mode of action on bacteria and its non-haemolytic activity, we synthesized androctonin, its fluorescent derivatives and its all-D-amino acid enantiomer. The enantiomer preserved high activity, suggesting a lipid-peptide interaction between androctonin and bacterial membranes. In Gram-positive and (at higher concentrations) Gram-negative bacteria, androctonin induced an immediate perturbation of the permeability properties of the cytoplasmic membrane of the bacterial energetic state, concomitant with perturbation of the morphology of the cell envelope as revealed by electron microscopy. Androctonin binds only to negatively charged lipid vesicles and induces the leakage of markers at high concentrations and with a slow kinetics, in contrast with amphipathic alpha-helical anti-microbial peptides that bind and permeate negatively charged vesicles, and to a smaller extent also zwitterionic ones. This might explain the selective lytic activity of androctonin towards bacteria but not red blood cells. Polarized attenuated total reflection-Fourier transform infrared spectroscopy revealed that androctonin adopts a beta-sheet structure in membranes and did not affect the lipid acyl chain order, which supports a detergent-like effect. The small size of androctonin, its hydrophilic character and its physicochemical properties are favourable features for its potential application as a replacement for commercially available antibiotics to which bacteria have developed resistance.}, keywords = {Adenosine Triphosphate, Anti-Bacterial Agents, Cations, Cell Membrane Permeability, Cytoplasm, Disulfides, Electron, Escherichia coli, Fluoresceins, Fluorescent Dyes, Fourier Transform Infrared, Gram-Negative Bacteria, hoffmann, Insect Proteins, Liposomes, M3i, Microbial Sensitivity Tests, Micrococcus luteus, Microscopy, oxygen, Phospholipids, Potassium, Proteins, spectroscopy}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_immunite_2000, title = {L'immunité innée : de la drosophile à l'homme}, author = {Dominique Ferrandon and Charles Hetru and Jean-Marc Reichhart and Jules A Hoffmann}, year = {2000}, date = {2000-01-01}, journal = {Pour la Science}, volume = {Dossier Hors Série Octobre}, pages = {8--12}, keywords = {ferrandon, hoffmann, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{tzou_tissue-specific_2000b, title = {Tissue-specific inducible expression of antimicrobial peptide genes in Drosophila surface epithelia}, author = {P Tzou and S Ohresser and Dominique Ferrandon and Maria Capovilla and Jean-Marc Reichhart and Bruno Lemaitre and Jules A Hoffmann and Jean-Luc Imler}, issn = {1074-7613}, year = {2000}, date = {2000-01-01}, journal = {Immunity}, volume = {13}, pages = {737--48.}, abstract = {The production of antimicrobial peptides is an important aspect of host defense in multicellular organisms. In Drosophila, seven antimicrobial peptides with different spectra of activities are synthesized by the fat body during the immune response and secreted into the hemolymph. Using GFP reporter transgenes, we show here that all seven Drosophila antimicrobial peptides can be induced in surface epithelia in a tissue-specific manner. The imd gene plays a critical role in the activation of this local response to infection. In particular, drosomycin expression, which is regulated by the Toll pathway during the systemic response, is regulated by imd in the respiratory tract, thus demonstrating the existence of distinct regulatory mechanisms for local and systemic induction of antimicrobial peptide genes in Drosophila.}, keywords = {*Genes, Animal, Anti-Infective Agents/*immunology/metabolism, Drosophila/genetics/*immunology, ferrandon, Gene Expression Regulation/*immunology, Genes, Glycoside Hydrolases/immunology, hoffmann, Human, imler, Insect, Insect Proteins/genetics/immunology, M3i, Non-U.S. Gov't, Organ Specificity, P.H.S., reichhart, Reporter, Support, Transfection, U.S. Gov't}, pubstate = {published}, tppubtype = {article} } @article{imler_lps-induced_2000, title = {LPS-induced immune response in Drosophila}, author = {Jean-Luc Imler and Servane Tauszig and Emmanuelle Jouanguy and C Forestier and Jules A Hoffmann}, issn = {0968-0519}, year = {2000}, date = {2000-01-01}, journal = {Journal of Endotoxin Research}, volume = {6}, number = {6}, pages = {459--462}, abstract = {The study of the regulation of the inducible synthesis of antimicrobial peptides in Drosophila melanogaster has established this insect as a powerful model in which to study innate immunity. In particular, the molecular characterization of the regulatory pathway controlling the antifungal peptide drosomycin has revealed the importance of Toll receptors in innate immunity. We report here that injection of LPS into flies induces an immune response, suggesting that LPS receptors are used in Drosophila to detect Gram-negative bacteria infection. We have identified in the recently sequenced genome of Drosophila eight genes coding for Toll-like receptors in addition to Toll, which may function as LPS receptors. However, overexpression of a selection of these genes in tissue-culture cells does not result in up-regulation of the antibacterial peptide genes. These results are discussed in light of the recent data from genetic screens aimed at identifying the genes controlling the antibacterial response in Drosophila.}, keywords = {Animals, Biological, Cell Line, Cell Surface, Defensins, Genes, Genetic, hoffmann, imler, Insect, Insect Proteins, Lipopolysaccharides, M3i, Membrane Glycoproteins, Models, Mutation, Promoter Regions, Receptors, Signal Transduction, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{meister_antimicrobial_2000, title = {The antimicrobial host defense of Drosophila}, author = {Marie Meister and Charles Hetru and Jules A Hoffmann}, issn = {0070-217X}, year = {2000}, date = {2000-01-01}, journal = {Curr. Top. Microbiol. Immunol.}, volume = {248}, pages = {17--36}, keywords = {Animals, Anti-Infective Agents, Fat Body, Genes, hoffmann, Insect, Insect Proteins, M3i}, pubstate = {published}, tppubtype = {article} } @article{imler_toll_2000, title = {Toll and Toll-like proteins: an ancient family of receptors signaling infection}, author = {Jean-Luc Imler and Jules A Hoffmann}, issn = {1398-1714}, year = {2000}, date = {2000-01-01}, journal = {Reviews in Immunogenetics}, volume = {2}, number = {3}, pages = {294--304}, abstract = {Innate immunity is the first-line host defense of multicellular organisms that rapidly operates to limit infection upon exposure to microbes. It involves intracellular signaling pathways in the fruit-fly Drosophila and in mammals that show striking similarities. Recent genetic and biochemical data have revealed, in particular, that proteins of the Toll family play a critical role in the immediate response to infection. We review here the recent developments on the structural and functional characterization of this evolutionary ancient and important family of proteins, which can function as cytokine receptors (Toll in Drosophila) or pattern recognition receptors (TLR4 in mammals) and activate similar, albeit non identical signal transduction pathways, in flies and mammals.}, keywords = {Adaptor Proteins, Animals, Antigens, Autoantigens, CD14, Cell Adhesion Molecules, Cell Surface, Differentiation, DNA-Binding Proteins, Gene Expression Regulation, hoffmann, I-kappa B Proteins, imler, Immunity, Immunologic, infection, Innate, Insect Proteins, Interleukin-1 Receptor-Associated Kinases, Knockout, Larva, Lipopolysaccharides, M3i, Mammals, MAP Kinase Signaling System, Membrane Glycoproteins, Membrane Proteins, Mice, Multigene Family, Myeloid Differentiation Factor 88, NF-kappa B, peptidoglycan, Phosphorylation, Post-Translational, Protein Kinases, Protein Processing, Protein Structure, Receptors, Recombinant Fusion Proteins, Signal Transducing, Signal Transduction, Teichoic Acids, Tertiary, Toll-Like Receptor 4, Toll-Like Receptor 5, Toll-Like Receptor 6, Toll-Like Receptor 9, Toll-Like Receptors, Ubiquitins}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genomic DNA methylation in the human trophoblast from recurrent spontaneous abortion-premilimary report}, author = {M Jerzak and K Potawski and G Keith and J A Jakowicki and W Baranovski}, editor = {Editor}, url = {none}, year = {2000}, date = {2000-01-01}, journal = {Pol J Gyn Invest}, volume = {3}, pages = {9-12}, keywords = {ACL GIEGE}, pubstate = {published}, tppubtype = {article} } @article{, title = {In vitro DNA and dGMP adducts formation caused by ochratoxin A}, author = { S. Obrecht-Pflumio and G. Dirheimer}, year = {2000}, date = {2000-01-01}, journal = {Chem Biol Interact}, volume = {127}, number = {1}, pages = {29-44}, abstract = {Ochratoxin A (OTA), a nephrotoxic and nephrocarcinogenic mycotoxin, leads to the formation of DNA adducts after administration to animals. This could be due to an epigenetic effect. In vitro assays can exclude an indirect effect, where the xenobiotic can generate, in vivo, endogenous reactive compounds which give adducts on DNA. Microsomes prepared from mice or rabbit kidney and liver, used as metabolic activators, were incubated in the presence of commercial salmon testes DNA and OTA, with NADPH or arachidonic acid used as cofactors. Upto 126 DNA adducts for 10(9) nucleotides were detected using the 32P postlabeling method after incubation with the mouse kidney system. Similar results were obtained with rabbit kidney microsomes. Using liver microsomes, the number of DNA adducts detected was much lower. When NADPH was used as a cosubstrate (to explore the cytochrome P450 metabolic pathways), with mice kidney microsomes, the adduct level was only 44% of the one obtained with arachidonic acid. These results lend support to the hypothesis of the preferential activation of OTA by the peroxidase activity of prostaglandin synthases and/or lipoxygenases to direct genotoxic metabolites, and are in agreement with the previously obtained results after in vivo treatment of mice. In order to identify the nucleotides of DNA modified by the OTA metabolites, dAMP, dGMP, dTMP and dCMP were used as substrates under the same conditions as with DNA. The adducts were found only on dGMP. The total adduct level was of 344 adducts per 10(9) nucleotides with the appearance of three major adducts in the presence of arachidonic acid. With NADPH, 271 adducts were obtained per 10(9) nucleotides, with again three major adducts, but only two of them were similar to two adducts obtained in the presence of arachidonic acid. Desferal (desferrioxamine B methanesulphonate), at a 50 microM concentration, did not reduce the adduct level. Adducts were also obtained when polydG, polydC and dG-p-dG were used as alternative substrates, whereas no adducts were obtained with polydA, polydT and polydC. The major adduct obtained after incubation of DNA with OTA, comigrated with the major adduct obtained with dGMP, in two chromatographic solvents. These results show that OTA is metabolized to genotoxic metabolite(s) which interact with the guanine residues of DNA.}, note = {0009-2797 Journal Article}, keywords = {Acid/metabolism, Adducts/*metabolism, Agents/pharmacology, Animals, Arachidonic, Carcinogens/pharmacology, Chelating, Chromatography, Deferoxamine/pharmacology, Deoxyguanine, DNA, Female, Kidney/ultrastructure, Liver/metabolism, Male, Mice, Microsomes, Microsomes/metabolism, Mycotoxins/pharmacology, NADP/metabolism, Nucleotides/*metabolism, Nucleotides/metabolism, Ochratoxins/*pharmacology, Peroxidase/metabolism, Rabbits, Spectrophotometry}, pubstate = {published}, tppubtype = {article} } @article{otten_sequence_1999, title = {Sequence and functional analysis of the left-hand part of the Ŧ-region from the nopaline-type Ti plasmid, pTiC58.}, author = {L Otten and J Y Salomone and A Helfer and J Schmidt and P Hammann and P De Ruffray}, doi = {10.1023/a:1006370207379}, issn = {0167-4412 0167-4412}, year = {1999}, date = {1999-12-01}, journal = {Plant molecular biology}, volume = {41}, number = {6}, pages = {765--776}, abstract = {The Agrobacterium tumefaciens nopaline strain C58 transfers a large, 29 kb T-DNA into plant cells during infection. Part of this DNA (the 'common DNA') is also found on the T-DNA of octopine strains, the remaining DNA is nopaline strain-specific. Up to now, only parts of the C58 T-DNA and related T37 T-DNA have been sequenced. We have sequenced the remainder of the nopaline-specific T-DNA (containing genes a to d) and acs to iaaM. Gene c codes for a new unknown T-DNA protein. Gene a is homologous to the agrocinopine synthase gene. Genes b, c', d and e are part of a larger family: they are related to the T-DNA genes 5, rolB, lso and 3'. Genes 5, rolB and lso induce or modify plant growth and have been called T-DNA oncogenes. Our studies show that gene 3' (located on the TR-DNA of octopine strains) is also oncogenic. Although the b-e T-DNA fragment from C58 and its individual genes lack growth-inducing activity, an a-acs deletion mutant was distinctly less virulent on Kalanchoe daigremontiana and showed reduced shoot formation on Kalanchoe tubiflora. Shoot formation could be restored by genes c and c' in co-infection experiments. Contrary to an earlier report, a C58 e gene deletion mutant was fully virulent on all plants tested.}, note = {Place: Netherlands}, keywords = {Agrobacterium tumefaciens/*genetics/pathogenicity, Bacterial/chemistry/*genetics, Bacterial/genetics, Chromosome Mapping, DNA, Gene Deletion, Genes, Genetic Complementation Test, Lycopersicon esculentum/genetics/microbiology, Medicinal/genetics/microbiology, Molecular Sequence Data, Mutation, Phylogeny, Plant Tumors/genetics/microbiology, Plants, Plasmids/chemistry/*genetics, PPSE, Sequence Analysis, Species Specificity, Tobacco/genetics/microbiology, Toxic, Virulence/genetics}, pubstate = {published}, tppubtype = {article} } @article{nisole_anti-hiv_1999, title = {The anti-HIV pseudopeptide HB-19 forms a complex with the cell-surface-expressed nucleolin independent of heparan sulfate proteoglycans}, author = {S Nisole and B Krust and C Callebaut and G Guichard and S Muller and J P Briand and A G Hovanessian}, doi = {10.1074/jbc.274.39.27875}, issn = {0021-9258}, year = {1999}, date = {1999-09-01}, journal = {The Journal of Biological Chemistry}, volume = {274}, number = {39}, pages = {27875--27884}, abstract = {The HB-19 pseudopeptide 5[Kpsi(CH(2)N)PR]-TASP, psi(CH(2)N) for reduced peptide bond, is a specific inhibitor of human immunodeficiency virus (HIV) infection in different CD4(+) cell lines and in primary T-lymphocytes and macrophages. Here, by using an experimental CD4(+) cell model to monitor HIV entry and infection, we demonstrate that HB-19 binds the cell surface and inhibits attachment of HIV particles to permissive cells. At concentrations that inhibit HIV attachment, HB-19 binds cells irreversibly, becomes complexed with the cell-surface-expressed nucleolin, and eventually results in its degradation. Accordingly, by confocal immunofluorescence microscopy, we demonstrate the drastic reduction of the cell-surface-expressed nucleolin following treatment of cells with HB-19. HIV particles can prevent the binding of HB-19 to cells and inhibit complex formation with nucleolin. Such a competition between viral particles and HB-19 is consistent with the implication of nucleolin in the process of HIV attachment to target cells. We show that another inhibitor of HIV infection, the fibroblast growth factor-2 (FGF-2) that uses cell-surface-expressed heparan sulfate proteoglycans as low affinity receptors, binds cells and blocks attachment of HIV to permissive cells. FGF-2 does not prevent the binding of HB-19 to cells and to nucleolin, and similarly HB-19 has no apparent effect on the binding of FGF-2 to the cell surface. The lack of competition between these two anti-HIV agents rules out the potential involvement of heparan sulfate proteoglycans in the mechanism of anti-HIV effect of HB-19, thus pointing out that nucleolin is its main target.}, keywords = {Anti-HIV Agents, Binding Sites, CD4-Positive T-Lymphocytes, Cell Line, Cell Membrane, Confocal, Fibroblast Growth Factor 2, Flow Cytometry, Heparan Sulfate Proteoglycans, HIV-1, Humans, Microscopy, Oligopeptides, Peptides, Phospholipid Ethers, Phosphoproteins, Proteins, RNA-Binding Proteins}, pubstate = {published}, tppubtype = {article} } @article{levashina_constitutive_1999, title = {Constitutive activation of toll-mediated antifungal defense in serpin-deficient Drosophila}, author = {Elena A Levashina and E Langley and C Green and David Gubb and M Ashburner and Jules A Hoffmann and Jean-Marc Reichhart}, issn = {0036-8075}, year = {1999}, date = {1999-09-01}, journal = {Science}, volume = {285}, number = {5435}, pages = {1917--1919}, abstract = {The antifungal defense of Drosophila is controlled by the spaetzle/Toll/cactus gene cassette. Here, a loss-of-function mutation in the gene encoding a blood serine protease inhibitor, Spn43Ac, was shown to lead to constitutive expression of the antifungal peptide drosomycin, and this effect was mediated by the spaetzle and Toll gene products. Spaetzle was cleaved by proteolytic enzymes to its active ligand form shortly after immune challenge, and cleaved Spaetzle was constitutively present in Spn43Ac-deficient flies. Hence, Spn43Ac negatively regulates the Toll signaling pathway, and Toll does not function as a pattern recognition receptor in the Drosophila host defense.}, keywords = {Animals, Antifungal Agents, Antimicrobial Cationic Peptides, Body Patterning, Cell Surface, Escherichia coli, Genes, Hemolymph, hoffmann, Insect, Insect Proteins, M3i, Membrane Glycoproteins, Micrococcus luteus, Mutagenesis, Peptides, Receptors, Recombinant Fusion Proteins, reichhart, Serine Proteinase Inhibitors, Serpins, Signal Transduction, Toll-Like Receptors, Up-Regulation}, pubstate = {published}, tppubtype = {article} } @article{manfruelli_mosaic_1999, title = {A mosaic analysis in Drosophila fat body cells of the control of antimicrobial peptide genes by the Rel proteins Dorsal and DIF}, author = {P Manfruelli and Jean-Marc Reichhart and R Steward and Jules A Hoffmann and Bruno Lemaitre}, doi = {10.1093/emboj/18.12.3380}, issn = {0261-4189}, year = {1999}, date = {1999-06-01}, journal = {EMBO J.}, volume = {18}, number = {12}, pages = {3380--3391}, abstract = {Expression of the gene encoding the antifungal peptide Drosomycin in Drosophila adults is controlled by the Toll signaling pathway. The Rel proteins Dorsal and DIF (Dorsal-related immunity factor) are possible candidates for the transactivating protein in the Toll pathway that directly regulates the drosomycin gene. We have examined the requirement of Dorsal and DIF for drosomycin expression in larval fat body cells, the predominant immune-responsive tissue, using the yeast site-specific flp/FRT recombination system to generate cell clones homozygous for a deficiency uncovering both the dorsal and the dif genes. Here we show that in the absence of both genes, the immune-inducibility of drosomycin is lost but can be rescued by overexpression of either dorsal or dif under the control of a heat-shock promoter. This result suggests a functional redundancy between both Rel proteins in the control of drosomycin gene expression in the larvae of Drosophila. Interestingly, the gene encoding the antibacterial peptide Diptericin remains fully inducible in the absence of the dorsal and dif genes. Finally, we have used fat body cell clones homozygous for various mutations to show that a linear activation cascade Spaetzle--textgreater Toll--textgreaterCactus--textgreaterDorsal/DIF leads to the induction of the drosomycin gene in larval fat body cells.}, keywords = {Animals, Anti-Infective Agents, Cell Surface, Clone Cells, DNA-Binding Proteins, Fat Body, Female, Gene Expression Regulation, Genes, hoffmann, Insect, Insect Proteins, Larva, M3i, Male, Membrane Glycoproteins, Mosaicism, Mutation, Nuclear Proteins, Phosphoproteins, Receptors, reichhart, Reporter, Signal Transduction, Toll-Like Receptors, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_phylogenetic_1999, title = {Phylogenetic perspectives in innate immunity}, author = {Jules A Hoffmann and Fotis C Kafatos and Charles A Janeway and Alan R B Ezekowitz}, issn = {0036-8075}, year = {1999}, date = {1999-05-01}, journal = {Science}, volume = {284}, number = {5418}, pages = {1313--1318}, abstract = {The concept of innate immunity refers to the first-line host defense that serves to limit infection in the early hours after exposure to microorganisms. Recent data have highlighted similarities between pathogen recognition, signaling pathways, and effector mechanisms of innate immunity in Drosophila and mammals, pointing to a common ancestry of these defenses. In addition to its role in the early phase of defense, innate immunity in mammals appears to play a key role in stimulating the subsequent, clonal response of adaptive immunity.}, keywords = {Active, Animals, Culicidae, hoffmann, Humans, Immunity, Immunological, infection, Innate, Insect Vectors, M3i, Mammals, Models, Phagocytosis, Phylogeny, Proteins}, pubstate = {published}, tppubtype = {article} } @article{lamberty_insect_1999, title = {Insect immunity. Isolation from the lepidopteran Heliothis virescens of a novel insect defensin with potent antifungal activity}, author = {M Lamberty and S Ades and S Uttenweiler-Joseph and G Brookhart and D Bushey and Jules A Hoffmann and Philippe Bulet}, issn = {0021-9258}, year = {1999}, date = {1999-04-01}, journal = {J. Biol. Chem.}, volume = {274}, number = {14}, pages = {9320--9326}, abstract = {Lepidoptera have been reported to produce several antibacterial peptides in response to septic injury. However, in marked contrast to other insect groups, no inducible antifungal molecules had been described so far in this insect order. Surprisingly, also cysteine-rich antimicrobial peptides, which predominate in the antimicrobial defense of other insects, had not been discovered in Lepidoptera. Here we report the isolation from the hemolymph of immune induced larvae of the lepidopteran Heliothis virescens of a cysteine-rich molecule with exclusive antifungal activity. We have fully characterized this antifungal molecule, which has significant homology with the insect defensins, a large family of antibacterial peptides directed against Gram-positive strains. Interestingly, the novel peptide shows also similarities with the antifungal peptide drosomycin from Drosophila. Thus, Lepidoptera appear to have built their humoral immune response against bacteria on cecropins and attacins. In addition, we report that Lepidoptera have conferred antifungal properties to the well conserved structure of antibacterial insect defensins through amino acid replacements.}, keywords = {Amino Acid, Animals, Antifungal Agents, Capillary, Chromatography, Defensins, Electrophoresis, Escherichia coli, Hemolymph, High Pressure Liquid, hoffmann, Insect Proteins, Larva, Lepidoptera, M3i, Micrococcus luteus, Proteins, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{lowenberger_insect_1999, title = {Insect immunity: molecular cloning, expression, and characterization of cDNAs and genomic DNA encoding three isoforms of insect defensin in Aedes aegypti}, author = {C A Lowenberger and C T Smartt and Philippe Bulet and M T Ferdig and D W Severson and Jules A Hoffmann and B M Christensen}, issn = {0962-1075}, year = {1999}, date = {1999-02-01}, journal = {Insect Mol. Biol.}, volume = {8}, number = {1}, pages = {107--118}, abstract = {Aedes aegypti were immune activated by injection with bacteria, and the expression of insect defensins was measured over time. Northern analyses indicated that defensin transcriptional activity continued for at least 21 days after bacterial injection, and up to 10 days after saline inoculation. Mature defensin levels in the haemolymph reached approximately 45 microM at 24 h post inoculation. cDNAs encoding the preprodefensins of three previously described mature Ae. aegypti defensins were amplified by PCR, cloned and sequenced. Genomic clones were amplified using primers designed against the cDNA sequence. Sequence comparison indicates that there is significant inter- and intra-isoform variability in the signal peptide and prodefensin sequences of defensin genes. Preprodefensin sequences of isoforms A and B are very similar, consisting of a signal peptide region of twenty amino acids, a prodefensin region of thirty-eight amino acids and a forty amino acid mature peptide domain. The sequence encoding isoform C is significantly different, comprising a signal peptide region of twenty-three amino acids, a prodefensin region of thirty-six amino acids, and the mature protein domain of forty amino acids. Analysis of the genomic clones of each isoform revealed one intron spatially conserved in the prodefensin region of all sequences. The intron in isoforms A and B is 64 nt long, and except for a 4 nt substitution in one clone, these intron sequences are identical. The intron in isoform C is 76 nt long and does not share significant identity with the intron sequences of isoforms A or B. The defensin gene mapped to chromosome 3, between two known loci, blt and LF168.}, keywords = {Aedes, Amino Acid, Animals, Base Sequence, Blotting, Chromosome Mapping, Cloning, Complementary, Defensins, DNA, Gene Expression, Hemolymph, hoffmann, M3i, Molecular, Northern, Protein Isoforms, Proteins, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Candida albicans CUG-decoding ser-tRNA has an atypical anticodon stem-loop structure}, author = { V. M. Perreau and G. Keith and W. M. Holmes and A. Przykorska and M. A. Santos and M. F. Tuite}, year = {1999}, date = {1999-01-01}, journal = {J Mol Biol}, volume = {293}, number = {5}, pages = {1039-53}, abstract = {In many Candida species, the leucine CUG codon is decoded by a tRNA with two unusual properties: it is a ser-tRNA and, uniquely, has guanosine at position 33 (G33). Using a combination of enzymatic (V1 RNase, RnI nuclease) and chemical (Pb(2+), imidazole) probing of the native Candida albicans ser-tRNACAG, we demonstrate that the overall tertiary structure of this tRNA resembles that of a ser-tRNA rather than a leu-tRNA, except within the anticodon arm where there is considerable disruption of the anticodon stem. Using non-modified in vitro transcripts of the C. albicans ser-tRNACAG carrying G, C, U or A at position 33, we demonstrate that it is specifically a G residue at this position that induces the atypical anticodon stem structure. Further quantitative evidence for an unusual structure in the anticodon arm of the G33-tRNA is provided by the observed change in kinetics of methylation of the G at position 37, by purified Escherichia coli m(1)G37 methyltransferase. We conclude that the anticodon arm distortion, induced by a guanosine base at position 33 in the anticodon loop of this novel tRNA, results in reduced decoding ability which has facilitated the evolution of this tRNA without extinction of the species encoding it.}, note = {0022-2836 Journal Article}, keywords = {*Nucleic, Acid, albicans/*genetics, Anticodon/*chemistry/*genetics/metabolism, Base, Candida, cerevisiae/genetics, Code/genetics, Conformation, Evolution, Fungal/chemistry/genetics/metabolism, Genetic, Gov't, Imidazoles/metabolism, Lead/metabolism, Methylation, Methyltransferases/metabolism, Molecular, Mutation/genetics, Non-P.H.S., Non-U.S., Nucleosides/genetics/metabolism, P.H.S., Ribonucleases/metabolism, RNA, Saccharomyces, Sequence, Ser/*chemistry/*genetics/metabolism, Solutions, Support, Transfer, tRNA, U.S.}, pubstate = {published}, tppubtype = {article} } @article{, title = {Detection and treatment of twinning: an improvement and new results}, author = {P Dumas and E Ennifar and P Walter}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10329781}, isbn = {10329781}, year = {1999}, date = {1999-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {55}, number = {Pt 6}, pages = {1179-87}, abstract = {This work deals with two aspects of the twinning problem. Firstly, an improvement of a known statistical test aimed at detecting twinning is presented and, secondly, a new parametrization of twinning is described, as well as a new method to obtain an accurate estimate of the degree of twinning. During work on crystals of the dimerization-initiation site of the HIV-1 genomic RNA, perfectly twinned crystals were obtained which were not immediately recognized as such by use of a known statistical method. This method, reminiscent of Wilson tests for the detection of centrosymmetric space groups, relies on calculation of <F2>/<F>2 or, equivalently, of <I2>/<I>2. It is shown that overlooking experimental errors may lead to erroneously large values of this index and, in turn, to ambiguous or incorrect conclusions. An immediate solution to this problem is presented. Independently, an alternative parametrization which expresses both the effect of twinning on intensities and the operation of untwinning to recover the correct intensities is proposed. A new method for estimating the degree of twinning is also presented. It is based upon maximization of the cross-correlation coefficients between intensities of all available data sets, and yields a fully analytical solution. Tests made with experimental data are quite satisfactory. It is suggested that the latter results could be used efficiently within the MIR method by allowing refinement, through one additional parameter only, of the twinning ratios of all data sets considered for phasing. Finally, the new parametrization of twinning has striking consequences in this correlation-based twinning determination: very unexpectedly, it yields a novel estimate of the 'twinning ratio' of a potentially twinned crystal which is fully independent of the data set used for the comparison.}, note = {0907-4449 Journal Article}, keywords = {ENNIFAR, HIV-1/*genetics *Nucleic Acid Conformation RNA, Unité ARN, Viral/*chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dynamical scattering and electron diffraction from thin polymer lamellar crystals - poly(tert-butylethylene sulfide)}, author = { D. L. Dorset and P. Dumas and L. Cartier and B. Lotz}, year = {1999}, date = {1999-01-01}, journal = {Acta Crystallogr A}, volume = {55}, number = {Pt 5}, pages = {901-907}, abstract = {Strong violations of Friedel symmetry are observed in hk0 electron diffraction patterns from lamellar crystals of poly(tert-butylethylene sulfide) obtained at 120 kV. These deviations are largely explained by a multislice dynamical scattering calculation based on the crystal structure model. Further improvement is found when a secondary scattering component is added, in keeping with a perfect crystallite thickness less than that of the lamellar thickness. Despite the multiple-scattering perturbations, the frustrated chain packing can still be determined by direct methods followed by Fourier refinement. However, the Friedel-related intensities must be averaged before calculation of normalized structure factors.}, note = {0108-7673 Journal article}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dynamical scattering and electron diffraction from thin polymer lamellar crystals - poly(tert-butylethylene sulfide). Erratum}, author = { D. L. Dorset and P. Dumas and L. Cartier and B. Lotz}, year = {1999}, date = {1999-01-01}, journal = {Acta Crystallogr A}, volume = {55}, number = {Pt 6}, pages = {1061}, abstract = {In the paper by Dorset et al. [Acta Cryst. (1999), A55, 901-907], the last line of page 904 should read 'ellipsisthe S atoms. However, the autocorrelation functionellipsis'.}, note = {0108-7673 Journal article}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {The crystal structure of the dimerization initiation site of genomic HIV-1 RNA reveals an extended duplex with two adenine bulges}, author = {E Ennifar and M Yusupov and P Walter and R Marquet and B Ehresmann and C Ehresmann and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10574792}, isbn = {10574792}, year = {1999}, date = {1999-01-01}, journal = {Structure}, volume = {7}, number = {11}, pages = {1439-49}, abstract = {BACKGROUND: An important step in retroviral replication is dimerization of the genomic RNA prior to encapsidation. Dimerization is initiated by the formation of a transient 'kissing-loop complex' that is thought to be subsequently matured into an extended duplex by the nucleocapsid protein (NCp). Although chemical probing and nuclear magnetic resonance spectroscopy have provided insight into the structure of the kissing-loop structure, no structural information concerning the extended-duplex state is available so far. RESULTS: The structure of a minimal HIV-1 RNA dimerization initiation site has been solved at 2.3 A resolution in two different space groups. It reveals a 22 base pair extended duplex with two noncanonical Watson-Crick-like G-A mismatches, each adjacent to a bulged-out adenine. The structure shows significant asymmetry in deep groove width and G-A base-pair conformations. A network of eight magnesium cations was clearly identified, one being unusually chelated by the 3' phosphate of each bulge across an extremely narrowed deep major groove. CONCLUSIONS: These crystal structures represent the putative matured form of the initial kissing-loop complex. They show the ability of this self-complementary RNA hairpin loop to acquire a more stable extended duplex structure. Both bulged adenines form a striking 'base grip' that could be a recognition signal, either in cis for another viral RNA sequence, or in trans for a protein, possibly the NCp. Magnesium binding might be important to promote and stabilize the observed extrahelical conformation of these bulges.}, note = {0969-2126 Journal Article}, keywords = {Adenine/*chemistry Base Pair Mismatch Base Sequence Crystallography, ENNIFAR, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA, Non-U.S. Gov't, Unité ARN, Viral/*chemistry/metabolism Support, X-Ray Dimerization HIV-1/*genetics Magnesium/metabolism Magnetic Resonance Spectroscopy Manganese/metabolism Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comments on the paper by kumagai, hibino, kawano and sugiyama (1999) FEBS lett. 450, 227-230}, author = { P. Dumas and M. Bergdoll and J. Masson}, year = {1999}, date = {1999-01-01}, journal = {FEBS Lett}, volume = {459}, number = {2}, pages = {282-3;discussion 284}, note = {0014-5793 Comment Journal Article}, keywords = {*Acetyltransferases, Antibiotics, Bacterial, Bleomycin/*metabolism/pharmacology, effects/genetics/*metabolism, Glycopeptide/metabolism/pharmacology, Mutation, Proline/*genetics/pharmacology, Proteins/genetics/*metabolism, Streptomyces/drug}, pubstate = {published}, tppubtype = {article} } @article{, title = {Kinetic analysis of the effect on Fab binding of identical substitutions in a peptide and its parent protein.}, author = {L Choulier and N Rauffer-Bruyere and M B Khalifa and F Martin and T Vernet and D Altschuh}, url = {http://www.ncbi.nlm.nih.gov/pubmed/10090739}, isbn = {10090739}, year = {1999}, date = {1999-01-01}, journal = {Biochemistry}, volume = {38}, number = {12}, pages = {3530-7}, abstract = {Monoclonal antibody 57P, which was raised against tobacco mosaic virus protein, cross-reacts with a peptide corresponding to residues 134-146 of this protein. Previous studies using peptide variants suggested that the peptide in the antibody combining site adopts a helical configuration that mimics the structure in the protein. In this study, we carried out a detailed comparison of Fab-peptide and Fab-protein interactions. The same five amino acid substitutions were introduced in the peptide (residues 134-151) and the parent protein, and the effect of these substitutions on antibody binding parameters have been measured with a Biacore instrument. Fabs that recognize epitopes located away from the site of mutations were used as indirect probes for the conformational integrity of protein antigens. Their interaction kinetics with all proteins were similar, suggesting that the substitutions had no drastic effect on their conformation. The five substitutions introduced in the peptide and the protein had minor effects on association rate constants (ka) and significant effects on dissociation rate constants (kd) of the antigen-Fab 57P interactions. In four out of five cases, the effect on binding affinity of the substitutions was identical when the epitope was presented in the form of a peptide or a protein antigen, indicating that antibody binding specifity was not affected by epitope presentation. However, ka values were about 10 times larger and kd values about 5 times larger for the peptide-Fab compared to the protein-Fab interaction, suggesting a different binding mechanism. Circular dichroism measurements performed for three of the peptides showed that they were mainly lacking structure in solution. Differences in conformational properties of the peptide and protein antigens in solution and/or in the paratope could explain differences in binding kinetics. Our results demonstrate that the peptides were able to mimic correctly some but not all properties of the protein-Fab 57P interaction and highlight the importance of quantitative analysis of both equilibrium and kinetic binding parameters in the design of synthetic vaccines and drugs.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Kinetic analysis of the effect on Fab binding of identical substitutions in a peptide and its parent protein.}, author = {L Choulier and N Rauffer-Bruyere and M B Khalifa and F Martin and T Vernet and D Altschuh}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/pubmed/10090739}, isbn = {10090739}, year = {1999}, date = {1999-01-01}, journal = {Biochemistry}, volume = {38}, number = {12}, pages = {3530-7}, abstract = {Monoclonal antibody 57P, which was raised against tobacco mosaic virus protein, cross-reacts with a peptide corresponding to residues 134-146 of this protein. Previous studies using peptide variants suggested that the peptide in the antibody combining site adopts a helical configuration that mimics the structure in the protein. In this study, we carried out a detailed comparison of Fab-peptide and Fab-protein interactions. The same five amino acid substitutions were introduced in the peptide (residues 134-151) and the parent protein, and the effect of these substitutions on antibody binding parameters have been measured with a Biacore instrument. Fabs that recognize epitopes located away from the site of mutations were used as indirect probes for the conformational integrity of protein antigens. Their interaction kinetics with all proteins were similar, suggesting that the substitutions had no drastic effect on their conformation. The five substitutions introduced in the peptide and the protein had minor effects on association rate constants (ka) and significant effects on dissociation rate constants (kd) of the antigen-Fab 57P interactions. In four out of five cases, the effect on binding affinity of the substitutions was identical when the epitope was presented in the form of a peptide or a protein antigen, indicating that antibody binding specifity was not affected by epitope presentation. However, ka values were about 10 times larger and kd values about 5 times larger for the peptide-Fab compared to the protein-Fab interaction, suggesting a different binding mechanism. Circular dichroism measurements performed for three of the peptides showed that they were mainly lacking structure in solution. Differences in conformational properties of the peptide and protein antigens in solution and/or in the paratope could explain differences in binding kinetics. Our results demonstrate that the peptides were able to mimic correctly some but not all properties of the protein-Fab 57P interaction and highlight the importance of quantitative analysis of both equilibrium and kinetic binding parameters in the design of synthetic vaccines and drugs.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of post-transcriptional modifications of primer tRNALys,3 in the fidelity and efficacy of plus strand DNA transfer during HIV-1 reverse transcription}, author = { S. Auxilien and G. Keith and S. F. Le Grice and J. L. Darlix}, year = {1999}, date = {1999-01-01}, journal = {J Biol Chem}, volume = {274}, number = {7}, pages = {4412-20}, abstract = {During HIV reverse transcription, (+) strand DNA synthesis is primed by an RNase H-resistant sequence, the polypurine tract, and continues as far as a 18-nt double-stranded RNA region corresponding to the 3' end of tRNALys,3 hybridized to the viral primer binding site (PBS). Before (+) strand DNA transfer, reverse transcriptase (RT) needs to unwind the double-stranded tRNA-PBS RNA in order to reverse-transcribe the 3' end of primer tRNALys,3. Since the detailed mechanism of (+) strand DNA transfer remains incompletely understood, we developed an in vitro system to closely examine this mechanism, composed of HIV 5' RNA, natural modified tRNALys,3, synthetic unmodified tRNALys,3 or oligonucleotides (RNA or DNA) complementary to the PBS, as well as the viral proteins RT and nucleocapsid protein (NCp7). Prior to (+) strand DNA transfer, RT stalls at the double-stranded tRNA-PBS RNA complex and is able to reverse-transcribe modified nucleosides of natural tRNALys,3. Modified nucleoside m1A-58 of natural tRNALys,3 is only partially effective as a stop signal, as RT can transcribe as far as the hyper-modified adenosine (ms2t6A-37) in the anticodon loop. m1A-58 is almost always transcribed into A, whereas other modified nucleosides are transcribed correctly, except for m7G-46, which is sometimes transcribed into T. In contrast, synthetic tRNALys,3, an RNA PBS primer, and a DNA PBS primer are completely reverse-transcribed. In the presence of an acceptor template, (+) strand DNA transfer is efficient only with templates containing natural tRNALys,3 or the RNA PBS primer. Sequence analysis of transfer products revealed frequent errors at the transfer site with synthetic tRNALys,3, not observed with natural tRNALys,3. Thus, modified nucleoside m1A-58, present in all retroviral tRNA primers, appears to be important for both efficacy and fidelity of (+) strand DNA transfer. We show that other factors such as the nature of the (-) PBS of the acceptor template and the RNase H activity of RT also influence the efficacy of (+) strand DNA transfer.}, note = {0021-9258 Journal Article}, keywords = {*RNA, *Transcription, Acid, Base, Calf, Conformation, Data, DNA, Genetic, Gov't, H, HIV-1, HIV-1/*physiology, Lys/*metabolism, Molecular, Non-U.S., Nucleic, post-transcriptional, Processing, Reverse, Ribonuclease, RNA, Sequence, Support, Templates, Thymus/metabolism, Transcriptase/metabolism, Transfer, Viral/*metabolism, Viral/metabolism}, pubstate = {published}, tppubtype = {article} } @article{dumortier_mhc_1999, title = {MHC class II gene associations with autoantibodies to U1A and SmD1 proteins}, author = {H Dumortier and M Abbal and M Fort and J P Briand and A Cantagrel and S Muller}, doi = {10.1093/intimm/11.2.249}, issn = {0953-8178}, year = {1999}, date = {1999-01-01}, journal = {International Immunology}, volume = {11}, number = {2}, pages = {249--257}, abstract = {Autoantibodies against U small nuclear ribonucleoproteins (snRNP) are frequently present in the serum of patients with systemic rheumatic diseases, and have been reported to be associated with HLA-DR and -DQ genes. To better define the role of HLA genes in the production of such antibodies, we studied immunogenetic associations with autoantibodies reacting with U1 RNP, U1A and SmD1 proteins, and synthetic peptides containing immunodominant linear epitopes of these proteins. Only two out of the 15 overlapping peptides of U1A (i.e. peptides 35-58 and 257-282) and three of 11 peptides of SmD1 (i.e. peptides 1-20, 44-67 and 97-119) were significantly recognized by patients' sera selected on the basis of their antibody positivity with RNP in immunodiffusion. The distribution of DRB1, DQB1 and DPB1 alleles among the anti-RNP antibody-positive patients (n = 28) and healthy control subjects was similar. Antibodies against U1A (tested in Western immunoblotting with HeLa cell extracts) were positively associated to DRB1*06 allele; antibodies reacting with SmD1 peptide 44-67 were negatively associated to DRB1*02 and DQB1*0602 alleles. No association was found between DPB1 alleles and antibodies reacting with U1A and SmD1 antigens. This first study reporting an association between autoantibodies reacting with U1A and SmD1 proteins (and peptides of these proteins), and immunogenetic markers suggest that the production of antibody subsets directed against different components (or regions of these proteins) bound to the same snRNP particle is associated with distinct MHC class II alleles.}, keywords = {Alleles, Antibody Specificity, Autoantibodies, Autoantigens, Autoimmune Diseases, Blotting, Dumortier, Enzyme-Linked Immunosorbent Assay, Genes, HLA-DP Antigens, HLA-DP beta-Chains, HLA-DQ Antigens, HLA-DQ beta-Chains, HLA-DR Antigens, HLA-DRB1 Chains, Humans, I2CT, MHC Class II, Peptides, Rheumatic Diseases, Ribonucleoprotein, Ribonucleoproteins, RNA-Binding Proteins, Small Nuclear, snRNP Core Proteins, Team-Dumortier, U1 Small Nuclear, Western}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization of the dimerization-initiation site of genomic HIV-1 RNA: preliminary crystallographic results}, author = {M Yusupov and P Walter and R Marquet and C Ehresmann and B Ehresmann and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10089425}, isbn = {10089425}, year = {1999}, date = {1999-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {55}, number = {Pt 1}, pages = {281-284}, abstract = {The genomic RNA of all retroviruses is encapsidated in virions as a dimer of single-stranded chains held together near their 5'-end. For HIV-1, the initial site of dimerization has been shown to be a hairpin with a nine-residue loop containing a self-complementary sequence of six residues. This structure is proposed to promote dimerization by loop-loop interaction and formation of a so-called 'kissing complex'. A 23-nucleotide RNA strand containing the loop enclosed by a seven base-pair stem has been synthesized. This oligomer was crystallized by the vapour-diffusion method at 310 K, pH 6.5, with methyl-pentanediol as the precipitant agent in the presence of MgCl2, KCl and spermine. Quasi-complete diffraction data were obtained at 2.7 A resolution with a conventional X-ray source and at 2.3 A resolution on a synchrotron beamline. The space group is P3121 or its enantiomorph P3221, with cell parameters a = b = 60. 1}, note = {0907-4449 Journal Article}, keywords = {Base Sequence Binding Sites Crystallization Crystallography, MARQUET, Non-U.S. Gov't, Unité ARN, Viral HIV-1/*chemistry/genetics Human Molecular Sequence Data Nucleic Acid Conformation RNA, Viral/*chemistry/genetics/isolation & purification Solutions Support, X-Ray Dimerization Genome}, pubstate = {published}, tppubtype = {article} } @article{, title = {Gene cloning, overproduction and purification of Escherichia coli tRNA(Arg2).}, author = {J F Wu and E D Wang and Y L Wang and G Eriani and J Gangloff}, url = {http://www.ncbi.nlm.nih.gov/pubmed/12136168}, isbn = {12136168}, year = {1999}, date = {1999-01-01}, journal = {Acta Bioch Bioph Sin}, volume = {31}, number = {3}, pages = {226-232}, abstract = {A synthetic gene encoding Escherichia coli tRNA(Arg)(2) was inserted in a plasmid under the control of an isopropyl-beta, D-thiogalactopyranoside (IPTG)-inducible promotor, pTrc99B. In E.coli MT102 transformed by the above plasmid containing the target gene. TRNA(Arg)(2) was overproduced up to 30 fold of that of the host. In the transformant the quantity contained tRNA(Arg) increased 10 times and was 70% of the total tRNA. The tRNA(Arg)(2) was purified to 88% homogeneity by passing through a DEAE-Sephacel column, and then was purified by benzyl-DEAE cellulose column chromatography to a purity of 99% with an arginylation activity of 1 600 pmole/A(260) unit. Eighteen milligrams of tRNA(Arg)(2) could be obtained from 40 mg total tRNA which was obtained from four liters of overnight culture, and the yield of the purification was 62%. The accurate kinetic constants of aminoacylation of tRNA(Arg)(2) catalyzed by arginyl-tRNA synthetase were comparable with that of tRNA(Arg) from Sigma.}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mimics of yeast tRNAAsp and their recognition by aspartyl-tRNA synthetase}, author = {A D Wolfson and A M Khvorova and C Sauter and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10508395}, isbn = {10508395}, year = {1999}, date = {1999-01-01}, journal = {Biochemistry}, volume = {38}, number = {37}, pages = {11926-11932}, abstract = {Assuming that the L-shaped three-dimensional structure of tRNA is an architectural framework allowing the proper presentation of identity nucleotides to aminoacyl-tRNA synthetases implies that altered and/or simplified RNA architectures can fulfill this role and be functional substrates of these enzymes, provided they contain correctly located identity elements. In this work, this paradigm was submitted to new experimental verification. Yeast aspartyl-tRNA synthetase was the model synthetase, and the extent to which the canonical structural framework of cognate tRNAAsp can be altered without losing its ability to be aminoacylated was investigated. Three novel architectures recognized by the synthetase were found. The first resembles that of metazoan mitochondrial tRNASer lacking the D-arm. The second lacks both the D- and T-arms, and the 5'-strand of the amino acid acceptor arm. The third structure is a construct in which the acceptor and anticodon helices are joined by two connectors. Aspartylation specificity of these RNAs is verified by the loss of aminoacylation activity upon mutation of the putative identity residues. Kinetic data indicate that the first two architectures are mimics of the whole tRNAAsp molecule, while the third one behaves as an aspartate minihelix mimic. Results confirm the primordial role of the discriminator nucleotide G73 in aspartylation and demonstrate that neither a helical structure in the acceptor domain nor the presence of a D- or T-arm is mandatory for specific aspartylation, but that activity relies on the presence of the cognate aspartate GUC sequence in the anticodon loop.}, note = {0006-2960 Journal Article}, keywords = {Acylation Aspartate-tRNA Ligase/*chemistry/metabolism Base Sequence Catalysis Cloning, Asp/*chemistry/genetics/metabolism Saccharomyces cerevisiae Support, FLORENTZ, Molecular Enzyme Activation/genetics Genetic Engineering Molecular Mimicry Molecular Sequence Data Mutagenesis, Non-U.S. Gov't, SAUTER, Site-Directed Plasmids/chemical synthesis RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A sequence immediately upstream of the plus-strand primer is essential for plus-strand DNA synthesis of the Saccharomyces cerevisiae Ty1 retrotransposon}, author = {M Wilhelm and T Heyman and M Boutabout and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10556309}, isbn = {10556309}, year = {1999}, date = {1999-01-01}, journal = {Nucleic Acids Res}, volume = {27}, number = {23}, pages = {4547-4552}, abstract = {Priming of plus-strand DNA is a critical step in reverse transcription of retroviruses and retrotransposons. All retroelements use an RNase H-resistant oligoribonucleotide spanning a purine-rich sequence (the polypurine tract or PPT) to prime plus-strand DNA synthesis. Plus-strand DNA synthesis of the yeast Saccharomyces cerevisiae Ty1-H3 retrotransposon is initiated at two sites, PPT1 and PPT2, located at the upstream boundary of the 3'-long terminal repeat and near the middle of the pol gene in the integrase coding region. The two plus-strand primers have the same purine-rich sequence GGGTGGTA. This sequence is not sufficient by itself to generate a plus-strand origin since two identical sequences located upstream of PPT2 in the integrase coding region are not used efficiently as primers for plus-strand DNA synthesis. Thus, other factors must be involved in the formation of a specific plus-strand DNA primer. We show here that mutations upstream of the PPT in a highly conserved T-rich region severely alters plus-strand DNA priming of Ty1. Our results demonstrate the importance of sequences or structural elements upstream of the PPT for initiation of plus-strand DNA synthesis.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence *DNA Primers DNA, Fungal/*biosynthesis Mutation Response Elements *Retroelements Saccharomyces cerevisiae/*genetics Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reverse transcription of the yeast Ty1 retrotransposon: the mode of first strand transfer is either intermolecular or intramolecular}, author = {M Wilhelm and M Boutabout and T Heyman and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10329158}, isbn = {10329158}, year = {1999}, date = {1999-01-01}, journal = {J Mol Biol}, volume = {288}, number = {4}, pages = {505-510}, abstract = {Replication of the yeast Ty1 retrotransposon occurs by a mechanism similar to that of retroviruses. According to the current model of retroviral reverse transcription, two strand transfers (the so-called minus-strand and plus-strand strong-stop DNA transfers) are required to produce full-length preintegrative DNA. Because two genomic RNA molecules are packaged inside the viral particles, the strand transfers can be either intra- or intermolecular. To study the mode of transfer of minus-strand strong-stop DNA during reverse transcription of the yeast Ty1 retrotransposon, we have analyzed the cDNA products that accumulate in the cytoplasmic virus-like particles of yeast cells harboring two marked Ty1 elements. Our results indicate that Ty1 minus-strand transfer occurs in a random manner with approximately similar frequencies of intra- and intermolecular transfer. It has been observed recently that intra- and intermolecular minus-strand transfer occur at similar frequencies during replication of a complex retrovirus such as HIV-1. These results together with the observation that genetic recombination occurs with a high frequency during minus-strand synthesis suggest that both packaged RNA molecules are needed for the synthesis of one minus-strand DNA.}, note = {0022-2836 Journal Article}, keywords = {Base Sequence DNA, Genetic, Non-U.S. Gov't *Transcription, Nucleic Acid *Retroelements Saccharomyces cerevisiae/*genetics Support, Single-Stranded/genetics Repetitive Sequences, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleic acids on folded architectures, molecular recognition and catalysis: Editorial overview}, author = {E Westhof and D Patel}, url = {http://www.sciencedirect.com/science/article/pii/S0959440X99800396}, doi = {10.1016/S0959-440X(99)80039-6}, year = {1999}, date = {1999-01-01}, journal = {Curr Opin Struct Biol}, volume = {9}, number = {3}, pages = {293–297}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Leadzyme RNA catalysis}, author = {E Westhof and T Hermann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10074935}, isbn = {10074935}, year = {1999}, date = {1999-01-01}, journal = {Nat Struct Biol}, volume = {6}, number = {3}, pages = {208-209}, note = {1072-8368 News}, keywords = {Base Sequence Catalysis Magnetic Resonance Spectroscopy Nucleic Acid Conformation RNA, Catalytic/chemistry/*metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Chemical diversity in RNA cleavage}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10532891}, isbn = {10532891}, year = {1999}, date = {1999-01-01}, journal = {Science}, volume = {286}, number = {5437}, pages = {61-62}, note = {0036-8075 Comment Journal Article}, keywords = {Base Pairing Catalysis Cytosine/chemistry/metabolism Hepatitis Delta Virus/*enzymology Hydrogen-Ion Concentration Imidazoles/chemistry/metabolism/pharmacology Mutagenesis Point Mutation RNA, Catalytic/*chemistry/*metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoglycoside-RNA interactions}, author = {F Walter and Q Vicens and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10600721}, isbn = {10600721}, year = {1999}, date = {1999-01-01}, journal = {Curr Opin Chem Biol}, volume = {3}, number = {6}, pages = {694-704}, abstract = {The structural and physico-chemical parameters promoting the binding of aminoglycosides to RNAs are becoming clear. The strength of the interaction is dominated by electrostatics, with the positively charged aminoglycosides displacing metal ions. Although aminoglycosides inhibit most known ribozymes, aminoglycosides or polyamines are able to catalyze specific RNA cleavage in the absence of metal ions.}, note = {1367-5931 Journal Article Review Review, Tutorial}, keywords = {Aminoglycosides Anti-Bacterial Agents/metabolism/*pharmacology Carbohydrate Sequence Molecular Sequence Data RNA/chemistry/*drug effects/metabolism RNA, Catalytic/antagonists & inhibitors/drug effects Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Variability of Substrate Specificity of Serum Antibodies Obtained from Patients with Different Autoimmune and Viral Diseases in Reaction of tRNA Hydrolysis}, author = {A V Vlassov and M Helm and C Florentz and V A Naumov and A A Breusov and V N Buneva and R Giege and G A Nevinsky}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12687113}, isbn = {12687113}, year = {1999}, date = {1999-01-01}, journal = {Russ J Immunol}, volume = {4}, number = {1}, pages = {25-32}, abstract = {Recently we have shown that the substrate specificity of catalytic IgG isolated from sera of patients with Hashimoto's thyroiditis, systemic lupus erythematosus (SLE), polyarthritis and hepatitis B for classic poly(N) homopolynucleotide substrates and for specific tRNA(Phe) with compact and stable structure was correlated with the type of disease. At the same time the cleavage specificity was different in comparison with that of all known human RNases. Here we investigated for the first time the hydrolysis by the IgGs isolated from sera of 31 patients with different diseases of the in vitro transcript of human mitochondrial tRNA(Lys) which has less stable structure as compared to tRNA(Phe). The level of activity was strongly dependent on the patient, but in general increased in the order: hepatitis B </= Hashimoto's thyroiditis < SLE. The pH dependencies and various salts effects also varied for Abs from the sera of different patients. Nevertheless, the RNase activity of all IgGs was specifically stimulated by Mg(2+) ions, that essentially completely suppress the activity of all known human RNases. In contrast to the classical substrates, no correlation between patient's IgG cleavage specificity and a specific disease was revealed; each patient demonstrated an individual repertoire of polyclonal RNA-hydrolyzing IgGs independently of the disease.}, note = {1028-7221 Journal article}, keywords = {FLORENTZ, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Features of tRNA hydrolysis by autoantibodies from blood serum of patients with certain autoimmune and virus diseases]}, author = {A V Vlasov and M Helm and V A Naumov and A A Breusov and V N Buneva and C Florentz and R Giege and G A Nevinskii}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10579192}, isbn = {10579192}, year = {1999}, date = {1999-01-01}, journal = {Mol Biol (Mosk)}, volume = {33}, number = {5}, pages = {866-872}, note = {0026-8984 Journal Article}, keywords = {Antibodies, Autoimmune/blood/*immunology Virus Diseases/blood/*immunology, Catalytic/metabolism Autoantibodies/*blood Base Sequence Human Hydrolysis Lupus Erythematosus, FLORENTZ, Systemic/blood/*immunology Molecular Sequence Data Nucleic Acid Conformation RNA, Transfer/chemistry/*metabolism Thyroiditis, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Imidazole compounds simulated active center of ribonuclease A. Synthesis and RNA cleaving activity]}, author = {V N Sil'nikov and N P Luk'ianchuk and G V Shishkin and R Giege and V V Vlasov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10347826}, isbn = {10347826}, year = {1999}, date = {1999-01-01}, journal = {Dokl Akad Nauk}, volume = {364}, number = {5}, pages = {690-694}, note = {0869-5652 Journal Article}, keywords = {Base Sequence Binding Sites *Imidazoles/chemical synthesis/chemistry/metabolism Molecular Sequence Data RNA, Pancreatic/*chemistry/metabolism Substrate Specificity, Phe/*metabolism Ribonuclease, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Maximization of selenocysteine tRNA and U6 small nuclear RNA transcriptional activation achieved by flexible utilization of a Staf zinc finger}, author = {M Schaub and E Myslinski and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10455183}, isbn = {10455183}, year = {1999}, date = {1999-01-01}, journal = {J Biol Chem}, volume = {274}, number = {35}, pages = {25042-25050}, abstract = {Transcriptional activators Staf and Oct-1 play critical roles in the activation of small nuclear RNA (snRNA) and snRNA-type gene transcription. Recently, we established that Staf binding to the human U6 snRNA (hU6) and Xenopus selenocysteine tRNA (xtRNA(Sec)) genes requires different sets of the seven C2-H2 zinc fingers. In this work, using a combination of oocyte microinjection, electrophoretic mobility shift assays, and missing nucleoside experiments with wild-type and mutant promoters, we demonstrate that the hU6 gene requires zinc fingers 2-7 for Staf binding and Oct-1 for maximal transcriptional activity. In contrast, the xtRNA(Sec) gene needs the binding of the seven Staf zinc fingers, but not Oct-1, for optimal transcriptional capacity. Mutation in the binding site for Staf zinc finger 1 in the tRNA(Sec) promoter reduced both Staf binding and transcriptional activity. Conversely, introduction of a zinc finger 1 binding site in the hU6 promoter increased Staf binding but interfered with the simultaneous Staf and Oct-1 binding, thus reducing transcriptional activity. Collectively, these results show that the differential utilization of Staf zinc finger 1 represents a new, critical determinant of the transcriptional activation mechanism for the Xenopus tRNA(Sec) and human U6 snRNA genes.}, note = {0021-9258 Journal Article}, keywords = {Amino Acid Sequence Animals Binding Sites/genetics DNA/genetics DNA-Binding Proteins/*genetics Human Hydroxyl Radical/metabolism Microinjections Molecular Sequence Data Mutation Oocytes Peptide Fragments/immunology Promoter Regions (Genetics) Protein Binding RNA, Amino Acid-Specific/*genetics Sequence Homology Support, Genetic Trans-Activation (Genetics)/*genetics Trans-Activators/*genetics Transcription Factors/genetics Xenopus Zinc Fingers/*genetics, Non-U.S. Gov't Templates, Small Nuclear/*genetics RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Flexible zinc finger requirement for binding of the transcriptional activator staf to U6 small nuclear RNA and tRNA(Sec) promoters}, author = {M Schaub and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10446199}, isbn = {10446199}, year = {1999}, date = {1999-01-01}, journal = {J Biol Chem}, volume = {274}, number = {34}, pages = {24241-24249}, abstract = {The transactivator Staf, which contains seven zinc finger motifs, exerts its effect on gene expression by binding to specific targets in small nuclear RNA (snRNA) and snRNA-type gene promoters. In this work, binding site selection allowed us to identify the 21-base pair ATTACCCATAATGCATYGCGG sequence as the high affinity consensus binding site for Staf. It shows a high sequence divergence with Staf-responsive elements in the Xenopus selenocysteine tRNA (tRNA(Sec)) and human U6 snRNA promoters. By using a combination of approaches, we analyzed the interaction of wild-type and truncated Staf zinc finger domains with the consensus, Xenopus tRNA(Sec), and human U6 sites. Two main conclusions emerged from our data. First, the data clearly indicate that zinc finger 7 does not establish base-specific contacts in Staf-DNA complexes. The second conclusion concerns zinc finger 1, which is required for the binding to the Xenopus tRNA(Sec) site but is dispensable in the case of the human U6 site. Taking into account the sequence differences in the two sites, these findings demonstrate that Staf utilizes zinc finger 1 in a rather flexible manner, illustrating how a protein can interact with DNAs containing targets of different sequences.}, note = {0021-9258 Journal Article}, keywords = {Amino Acid Sequence Animals Binding Sites DNA-Binding Proteins/chemistry/*metabolism Deoxyribonuclease I/pharmacology Human Molecular Sequence Data *Promoter Regions (Genetics) RNA, Amino Acid-Specific/*genetics Support, Non-U.S. Gov't Trans-Activators/chemistry/*metabolism Xenopus *Zinc Fingers, Small Nuclear/*genetics RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Additives for the crystallization of proteins and nucleic acids.}, author = {C Sauter and J D Ng and B Lorber and G Keith and P Brion and D W Hosseini and J M Lehn and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S0022024898008525}, year = {1999}, date = {1999-01-01}, journal = {J Crystal Growth}, volume = {196}, number = {2-4}, pages = {365-376}, abstract = {Numerous molecules have been described in literature as additives that were indispensable either for nucleation or growth of macromolecular crystals. In some cases, such additives were shown to improve the quality of the X-ray di¤raction and to extend di¤raction limits. We have investigated the e¤ects of more than Þfty compounds, belonging to several chemical families, on the crystallization of four model proteins (hen and turkey egg-white lysozymes, thaumatin, and aspartyl-tRNA synthetase from ¹hermus thermophilus). In addition, we have studied the crystallization of a ribonucleic acid from yeast, the transfer RNA speciÞc for phenylalanine in the presence of synthetic polyamines. Crystals grown in the presence of the additives were optically evaluated and X-ray di¤raction analyses were performed on selective crystals to compare their space group, cell parameters, and di¤raction limit with those of controls. Whereas no changes in space group nor cell parameters were observed for the model proteins, signiÞcant improvements in di¤raction limit were found when the transfer RNA was crystallized with certain synthetic polyamines. (1999 Elsevier Science B.V. All rights reserved.}, keywords = {Crystallogenesis Additives Polyamines, SAUTER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallogenesis studies on yeast aspartyl-tRNA synthetase: use of phase diagram to improve crystal quality}, author = {C Sauter and B Lorber and D Kern and J Cavarelli and D Moras and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10089405}, doi = {10089405}, year = {1999}, date = {1999-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {55}, number = {Pt 1}, pages = {149-156}, abstract = {Aspartyl-tRNA synthetase (AspRS) extracted from yeast is heterogeneous owing to proteolysis of its positively charged N-terminus; its crystals are of poor quality. To overcome this drawback, a rational strategy was developed to grow crystals of sufficient quality for structure determination. The strategy is based on improvement of the protein homogeneity and optimization of crystallization, taking advantage of predictions from crystal-growth theories. An active mutant lacking the first 70 residues was produced and initial crystallization conditions searched. The shape and habit of initial crystals were improved by establishing a phase diagram of protein versus crystallizing-agent concentrations. Growth of large well faceted crystals takes place at low supersaturations near the isochronic supersolubility curve. Further refinement led to reproducible growth of two crystalline forms of bipyramidal (I) or prismatic (II) habit. Both diffract X-rays better than crystals previously obtained with native AspRS. Complete data sets were collected at 3 A resolution for form I (space group P41212) and form II (space group P3221) and molecular-replacement solutions were found in both space groups.}, note = {0907-4449 Journal Article}, keywords = {Aspartate-tRNA Ligase/*chemistry/genetics/*isolation & purification Crystallization Crystallography, Fungal Saccharomyces cerevisiae/*enzymology/genetics Sequence Deletion Solutions Support, Non-U.S. Gov't, SAUTER, Unité ARN, X-Ray Genes}, pubstate = {published}, tppubtype = {article} } @article{, title = {The structure of threonyl-tRNA synthetase-tRNA(Thr) complex enlightens its repressor activity and reveals an essential zinc ion in the active site}, author = {R Sankaranarayanan and A C Dock-Bregeon and P Romby and J Caillet and M Springer and B Rees and C Ehresmann and B Ehresmann and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10319817}, isbn = {10319817}, year = {1999}, date = {1999-01-01}, journal = {Cell}, volume = {97}, number = {3}, pages = {371-381}, abstract = {E. coli threonyl-tRNA synthetase (ThrRS) is a class II enzyme that represses the translation of its own mRNA. We report the crystal structure at 2.9 A resolution of the complex between tRNA(Thr) and ThrRS, whose structural features reveal novel strategies for providing specificity in tRNA selection. These include an amino-terminal domain containing a novel protein fold that makes minor groove contacts with the tRNA acceptor stem. The enzyme induces a large deformation of the anticodon loop, resulting in an interaction between two adjacent anticodon bases, which accounts for their prominent role in tRNA identity and translational regulation. A zinc ion found in the active site is implicated in amino acid recognition/discrimination.}, note = {0092-8674 Journal Article}, keywords = {Amino Acid Support, Amino Acyl/*chemistry/genetics/*metabolism Sequence Homology, Messenger/genetics RNA, Non-U.S. Gov't Zinc/*chemistry, ROMBY, Secondary Protein Structure, Tertiary RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aminoacylated tmRNA from Escherichia coli interacts with prokaryotic elongation factor Tu}, author = {J Rudinger-Thirion and R Giege and B Felden}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10445873}, isbn = {10445873}, year = {1999}, date = {1999-01-01}, journal = {RNA}, volume = {5}, number = {8}, pages = {989-992}, note = {1355-8382 Letter}, keywords = {Ala/metabolism RNA, Amino Acyl/*metabolism Support, Bacterial/*metabolism RNA, Base Sequence Escherichia coli/*genetics Guanosine Triphosphate/metabolism Models, Genetic Molecular Sequence Data Nucleic Acid Conformation Peptide Elongation Factor Tu/*metabolism RNA, Non-U.S. Gov't Thermus thermophilus/*metabolism Time Factors, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The peculiar architectural framework of tRNASec is fully recognized by yeast AspRS}, author = {J Rudinger-Thirion and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10199566}, isbn = {10199566}, year = {1999}, date = {1999-01-01}, journal = {RNA}, volume = {5}, number = {4}, pages = {495-502}, abstract = {The wild-type transcript of Escherichia coli tRNASec, characterized by a peculiar core architecture and a large variable region, was shown to be aspartylatable by yeast AspRS. Similar activities were found for tRNASec mutants with methionine, leucine, and tryptophan anticodons. The charging efficiency of these molecules was found comparable to that of a minihelix derived from tRNAAsp and is accounted for by the presence of the discriminator residue G73, which is a major aspartate identity determinant. Introducing the aspartate identity elements from the anticodon loop (G34, U35, C36, C38) into tRNASec transforms this molecule into an aspartate acceptor with kinetic properties identical to tRNAAsp. Expression of the aspartate identity set in tRNASec is independent of the size of its variable region. The functional study was completed by footprinting experiments with four different nucleases as structural probes. Protection patterns by AspRS of transplanted tRNASec and tRNAAsp were found similar. They are modified, particularly in the anticodon loop, upon changing the aspartate anticodon into that of methionine. Altogether, it appears that recognition of a tRNA by AspRS is more governed by the presence of the aspartate identity set than by the structural framework that carries this set.}, note = {1355-8382 Journal Article}, keywords = {Amino Acid-Specific/*genetics RNA, Amino Acyl-tRNA Ligases/*genetics/metabolism Anticodon/genetics Aspartic Acid/genetics/metabolism Base Sequence Escherichia coli/genetics Fungi/*enzymology/genetics Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, Asp/genetics Selenocysteine/genetics/metabolism Support, Bacterial/genetics RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Hybridization of antisense oligonucleotides with the 3'part of tRNA(Phe)}, author = {V A Petyuk and M A Zenkova and R Giege and V V Vlassov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10050762}, isbn = {10050762}, year = {1999}, date = {1999-01-01}, journal = {FEBS Lett}, volume = {444}, number = {2-3}, pages = {217-221}, abstract = {The interaction of antisense oligodeoxyribonucleotides with yeast tRNA(Phe) was investigated. 14-15-mers complementary to the 3'-terminal sequence including the ACCA end bind to the tRNA under physiological conditions. At low oligonucleotide concentrations the binding occurs at the unique complementary site. At higher oligonucleotide concentrations, the second oligonucleotide molecule binds to the complex due to non-perfect duplex formation in the T-loop stabilized by stacking between the two bound oligonucleotides. In these complexes the acceptor stem is open and the 5'-terminal sequence of the tRNA is accessible for binding of a complementary oligonucleotide. The results prove that the efficient binding of oligonucleotides to the 3'-terminal sequence of the tRNA occurs through initial binding to the single-stranded sequence ACCA followed by invasion in the acceptor stem and strand displacement.}, note = {0014-5793 Journal Article}, keywords = {Antisense/*genetics RNA, Base Sequence Electrophoresis, Calf Thymus/metabolism Support, Fungal/genetics RNA, Non-U.S. Gov't, Phe/*genetics Ribonuclease H, Polyacrylamide Gel Molecular Sequence Data Nucleic Acid Conformation Nucleic Acid Hybridization/*genetics Oligodeoxyribonucleotides/genetics Oligonucleotides, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interaction of complementary oligonucleotides with the 3'-end of yeast tRNA(Phe)}, author = {V Petyuk and M Zenkova and R Giege and V Vlassov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10474225}, isbn = {10474225}, year = {1999}, date = {1999-01-01}, journal = {Nucleosides Nucleotides}, volume = {18}, number = {6-7}, pages = {1459-1461}, abstract = {Interaction of yeast tRNA(Phe) with oligodeoxyribonucleotides (ONs), complementary to the nucleotides 62-76 was investigated. Results of gel-mobility shift assay and RNase A probing evidence that the ONs containing the sequence complementary to the tRNA ACCA end can easily invade the hairpin structure under physiological conditions. The limiting step of association process is the tRNA unfolding.}, note = {0732-8311 Journal Article}, keywords = {Non-U.S. Gov't, Nucleic Acid Conformation RNA, Phe/*chemistry Saccharomyces cerevisiae/*genetics Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Drosophila modifier of variegation modulo gene product binds specific RNA sequences at the nucleolus and interacts with DNA and chromatin in a phosphorylation-dependent manner}, author = {L Perrin and P Romby and P Laurenti and H Berenger and S Kallenbach and H M Bourbon and J Pradel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10037720}, isbn = {10037720}, year = {1999}, date = {1999-01-01}, journal = {J Biol Chem}, volume = {274}, number = {10}, pages = {6315-6323}, abstract = {modulo belongs to the modifier of Position Effect Variegation class of Drosophila genes, suggesting a role for its product in regulating chromatin structure. Genetics assigned a second function to the gene, in protein synthesis capacity. Bifunctionality is consistent with protein localization in two distinct subnuclear compartments, chromatin and nucleolus, and with its organization in modules potentially involved in DNA and RNA binding. In this study, we examine nucleic acid interactions established by Modulo at nucleolus and chromatin and the mechanism that controls the distribution and balances the function of the protein in the two compartments. Structure/function analysis and oligomer selection/amplification experiments indicate that, in vitro, two basic terminal domains independently contact DNA without sequence specificity, whereas a central RNA Recognition Motif (RRM)-containing domain allows recognition of a novel sequence-/motif-specific RNA class. Phosphorylation moreover is shown to down-regulate DNA binding. Evidence is provided that in vivo nucleolar Modulo is highly phosphorylated and belongs to a ribonucleoprotein particle, whereas chromatin-associated protein is not modified. A functional scheme is finally proposed in which modification by phosphorylation modulates Mod subnuclear distribution and balances its function at the nucleolus and chromatin.}, note = {0021-9258 Journal Article}, keywords = {Animals Base Sequence Binding Sites/genetics Chromatin/*genetics DNA/*genetics DNA-Binding Proteins/*genetics/metabolism Drosophila/*genetics *Drosophila Proteins *Genes, Insect Insect Proteins/*genetics/metabolism Molecular Sequence Data Phosphorylation RNA/*genetics RNA-Binding Proteins/*genetics/metabolism Support, Non-U.S. Gov't, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A complex ligase ribozyme evolved in vitro from a group I ribozyme domain.}, author = { L. Jaeger and M.C. Wright and G.F. Joyce}, year = {1999}, date = {1999-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {96}, number = {26}, pages = {14712-7}, abstract = {Like most proteins, complex RNA molecules often are modular objects made up of distinct structural and functional domains. The component domains of a protein can associate in alternative combinations to form molecules with different functions. These observations raise the possibility that complex RNAs also can be assembled from preexisting structural and functional domains. To test this hypothesis, an in vitro evolution procedure was used to isolate a previously undescribed class of complex ligase ribozymes, starting from a pool of 10(16) different RNA molecules that contained a constant region derived from a large structural domain that occurs within self-splicing group I ribozymes. Attached to this constant region were three hypervariable regions, totaling 85 nucleotides, that gave rise to the catalytic motif within the evolved catalysts. The ligase ribozymes catalyze formation of a 3',5'-phosphodiester linkage between adjacent template-bound oligonucleotides, one bearing a 3' hydroxyl and the other a 5' triphosphate. Ligation occurs in the context of a Watson-Crick duplex, with a catalytic rate of 0.26 min(-1) under optimal conditions. The constant region is essential for catalytic activity and appears to retain the tertiary structure of the group I ribozyme. This work demonstrates that complex RNA molecules, like their protein counterparts, can share common structural domains while exhibiting distinct catalytic functions.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {The Candida albicans CUG-decoding ser-tRNA has an atypical anticodon stem-loop structure}, author = {V M Perreau and G Keith and W M Holmes and A Przykorska and M A Santos and M F Tuite}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10547284}, isbn = {10547284}, year = {1999}, date = {1999-01-01}, journal = {J Mol Biol}, volume = {293}, number = {5}, pages = {1039-1053}, abstract = {In many Candida species, the leucine CUG codon is decoded by a tRNA with two unusual properties: it is a ser-tRNA and, uniquely, has guanosine at position 33 (G33). Using a combination of enzymatic (V1 RNase, RnI nuclease) and chemical (Pb(2+), imidazole) probing of the native Candida albicans ser-tRNACAG, we demonstrate that the overall tertiary structure of this tRNA resembles that of a ser-tRNA rather than a leu-tRNA, except within the anticodon arm where there is considerable disruption of the anticodon stem. Using non-modified in vitro transcripts of the C. albicans ser-tRNACAG carrying G, C, U or A at position 33, we demonstrate that it is specifically a G residue at this position that induces the atypical anticodon stem structure. Further quantitative evidence for an unusual structure in the anticodon arm of the G33-tRNA is provided by the observed change in kinetics of methylation of the G at position 37, by purified Escherichia coli m(1)G37 methyltransferase. We conclude that the anticodon arm distortion, induced by a guanosine base at position 33 in the anticodon loop of this novel tRNA, results in reduced decoding ability which has facilitated the evolution of this tRNA without extinction of the species encoding it.}, note = {0022-2836 Journal Article}, keywords = {Anticodon/*chemistry/*genetics/metabolism Base Sequence Candida albicans/*genetics Evolution, Fungal/chemistry/genetics/metabolism RNA, Molecular Genetic Code/genetics Imidazoles/metabolism Lead/metabolism Methylation Mutation/genetics *Nucleic Acid Conformation Nucleosides/genetics/metabolism RNA, Non-P.H.S. Support, Non-U.S. Gov't Support, P.H.S. tRNA Methyltransferases/metabolism, Ser/*chemistry/*genetics/metabolism Ribonucleases/metabolism Saccharomyces cerevisiae/genetics Solutions Support, Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Reverse transcription of the yeast Ty1 retrotransposon: the mode of first strand transfer is either intermolecular or intramolecular}, author = { M. Wilhelm and M. Boutabout and T. Heyman and F. X. Wilhelm}, year = {1999}, date = {1999-01-01}, journal = {J Mol Biol}, volume = {288}, number = {4}, pages = {505-10}, abstract = {Replication of the yeast Ty1 retrotransposon occurs by a mechanism similar to that of retroviruses. According to the current model of retroviral reverse transcription, two strand transfers (the so-called minus-strand and plus-strand strong-stop DNA transfers) are required to produce full-length preintegrative DNA. Because two genomic RNA molecules are packaged inside the viral particles, the strand transfers can be either intra- or intermolecular. To study the mode of transfer of minus-strand strong-stop DNA during reverse transcription of the yeast Ty1 retrotransposon, we have analyzed the cDNA products that accumulate in the cytoplasmic virus-like particles of yeast cells harboring two marked Ty1 elements. Our results indicate that Ty1 minus-strand transfer occurs in a random manner with approximately similar frequencies of intra- and intermolecular transfer. It has been observed recently that intra- and intermolecular minus-strand transfer occur at similar frequencies during replication of a complex retrovirus such as HIV-1. These results together with the observation that genetic recombination occurs with a high frequency during minus-strand synthesis suggest that both packaged RNA molecules are needed for the synthesis of one minus-strand DNA.}, note = {0022-2836 Journal Article}, keywords = {*Retroelements, *Transcription, Acid, Base, cerevisiae/*genetics, DNA, Genetic, Gov't, Non-U.S., Nucleic, Repetitive, Saccharomyces, Sequence, Sequences, Single-Stranded/genetics, Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A sequence immediately upstream of the plus-strand primer is essential for plus-strand DNA synthesis of the Saccharomyces cerevisiae Ty1 retrotransposon}, author = { M. Wilhelm and T. Heyman and M. Boutabout and F. X. Wilhelm}, year = {1999}, date = {1999-01-01}, journal = {Nucleic Acids Res}, volume = {27}, number = {23}, pages = {4547-52}, abstract = {Priming of plus-strand DNA is a critical step in reverse transcription of retroviruses and retrotransposons. All retroelements use an RNase H-resistant oligoribonucleotide spanning a purine-rich sequence (the polypurine tract or PPT) to prime plus-strand DNA synthesis. Plus-strand DNA synthesis of the yeast Saccharomyces cerevisiae Ty1-H3 retrotransposon is initiated at two sites, PPT1 and PPT2, located at the upstream boundary of the 3'-long terminal repeat and near the middle of the pol gene in the integrase coding region. The two plus-strand primers have the same purine-rich sequence GGGTGGTA. This sequence is not sufficient by itself to generate a plus-strand origin since two identical sequences located upstream of PPT2 in the integrase coding region are not used efficiently as primers for plus-strand DNA synthesis. Thus, other factors must be involved in the formation of a specific plus-strand DNA primer. We show here that mutations upstream of the PPT in a highly conserved T-rich region severely alters plus-strand DNA priming of Ty1. Our results demonstrate the importance of sequences or structural elements upstream of the PPT for initiation of plus-strand DNA synthesis.}, note = {0305-1048 Journal Article}, keywords = {*DNA, *Retroelements, Base, cerevisiae/*genetics, DNA, Elements, Fungal/*biosynthesis, Gov't, Mutation, Non-U.S., Primers, response, Saccharomyces, Sequence, Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A complex ligase ribozyme evolved in vitro from a group I ribozyme domain.}, author = {L Jaeger and M C Wright and G F Joyce}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/pubmed/10611278}, isbn = {10611278}, year = {1999}, date = {1999-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {96}, number = {26}, pages = {14712-7}, abstract = {Like most proteins, complex RNA molecules often are modular objects made up of distinct structural and functional domains. The component domains of a protein can associate in alternative combinations to form molecules with different functions. These observations raise the possibility that complex RNAs also can be assembled from preexisting structural and functional domains. To test this hypothesis, an in vitro evolution procedure was used to isolate a previously undescribed class of complex ligase ribozymes, starting from a pool of 10(16) different RNA molecules that contained a constant region derived from a large structural domain that occurs within self-splicing group I ribozymes. Attached to this constant region were three hypervariable regions, totaling 85 nucleotides, that gave rise to the catalytic motif within the evolved catalysts. The ligase ribozymes catalyze formation of a 3',5'-phosphodiester linkage between adjacent template-bound oligonucleotides, one bearing a 3' hydroxyl and the other a 5' triphosphate. Ligation occurs in the context of a Watson-Crick duplex, with a catalytic rate of 0.26 min(-1) under optimal conditions. The constant region is essential for catalytic activity and appears to retain the tertiary structure of the group I ribozyme. This work demonstrates that complex RNA molecules, like their protein counterparts, can share common structural domains while exhibiting distinct catalytic functions.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{gross_dorsal-b_1999, title = {Dorsal-B, a splice variant of the Drosophila factor Dorsal, is a novel Rel/NF-kB transcriptional activator.}, author = {I Gross and Philippe Georgel and P Oertel-Buchheit and M Schnarr and Jean-Marc Reichhart}, year = {1999}, date = {1999-01-01}, journal = {Gene}, volume = {228}, pages = {233--242}, abstract = {The Drosophila transcription factor Dorsal, a member of the Rel/NF-kB family of proteins, plays a key role in the establishment of dorsoventral polarity in the early embryo and is also involved in the immune response. Here we present evidence that the primary transcript of dorsal can be alternatively spliced, generating Dorsal B, a new Rel/NF-kB family member. Dorsal and Dorsal B are identical in the N-terminal region which comprises both a DNA binding domain and a dimerization domain. However, Dorsal B lacks the nuclear localization signal located at the end of the Rel domain of Dorsal and is totally divergent in the C-terminal portion. Although Dorsal B by itself is not able to induce the expression of a kB-controlled Luciferase reporter gene, we demonstrate that its C-terminal portion has transactivating properties. Analysis of thedorsal B expression pattern indicates that the splicing is tissue-specific and excludes a putative role in early embryogenesis. On the other hand, Dorsal B synthesis is enhanced upon septic injury and this challenge induces a nuclear accumulation of the protein in fat body cells suggesting that it may be involved in the immune response.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{lowenberger_mosquito-plasmodium_1999, title = {Mosquito-Plasmodium interactions in response to immune activation of the vector}, author = {C A Lowenberger and S Kamal and J Chiles and S Paskewitz and Philippe Bulet and Jules A Hoffmann and B M Christensen}, doi = {10.1006/expr.1999.4350}, issn = {0014-4894}, year = {1999}, date = {1999-01-01}, journal = {Exp. Parasitol.}, volume = {91}, number = {1}, pages = {59--69}, abstract = {During the development of Plasmodium sp. within the mosquito midgut, the parasite undergoes a series of developmental changes. The elongated ookinete migrates through the layers of the midgut where it forms the oocyst under the basal lamina. We demonstrate here that if Aedes aegypti or Anopheles gambiae, normally susceptible to Plasmodium gallinaceum and P. berghei, respectively, are immune activated by the injection of bacteria into the hemocoel, and subsequently are fed on an infectious bloodmeal, there is a significant reduction in the prevalence and mean intensity of infection of oocysts on the midgut. Only those mosquitoes immune activated prior to, or immediately after, parasite ingestion exhibit this reduction in parasite development. Mosquitoes immune activated 2-5 days after bloodfeeding show no differences in parasite burdens compared with naive controls. Northern analyses reveal that transcriptional activity for mosquito defensins is not detected in the whole bodies of Ae. aegypti from 4 h to 10 days after ingesting P. gallinaceum, suggesting that parasite ingestion, passage from the food bolus through the midgut, oocyst formation, and subsequent release of sporozoites into the hemolymph do not induce the production of defensin. However, reverse transcriptase-PCR of RNA isolated solely from the midguts of Ae. aegypti indicates that transcription of mosquito defensins occurs in the midguts of naive mosquitoes and those ingesting an infectious or noninfectious bloodmeal. Bacteria-challenged Ae. aegypti showed high levels of mature defensin in the hemolymph that correlate with a lower prevalence and mean intensity of infection with oocysts. Because few oocysts were found on the midgut of immune-activated mosquitoes, the data suggest that some factor, induced by bacterial challenge, kills the parasite at a preoocyst stage.}, keywords = {Aedes, Animals, Anopheles, Culicidae, Defensins, Digestive System, Escherichia coli, Female, Genetic, Hemolymph, hoffmann, Insect Vectors, M3i, messenger, Micrococcus luteus, Plasmodium, Plasmodium berghei, Plasmodium gallinaceum, Proteins, Reverse Transcriptase Polymerase Chain Reaction, RNA, Transcription}, pubstate = {published}, tppubtype = {article} } @article{, title = {Kinetic analysis of the effect on Fab binding of identical substitutions in a peptide and its parent protein.}, author = { L. Choulier and N. Rauffer-Bruyere and M.B. Khalifa and F. Martin and T. Vernet and D. Altschuh}, year = {1999}, date = {1999-01-01}, journal = {Biochemistry}, volume = {38}, number = {12}, pages = {3530-7}, abstract = {Monoclonal antibody 57P, which was raised against tobacco mosaic virus protein, cross-reacts with a peptide corresponding to residues 134-146 of this protein. Previous studies using peptide variants suggested that the peptide in the antibody combining site adopts a helical configuration that mimics the structure in the protein. In this study, we carried out a detailed comparison of Fab-peptide and Fab-protein interactions. The same five amino acid substitutions were introduced in the peptide (residues 134-151) and the parent protein, and the effect of these substitutions on antibody binding parameters have been measured with a Biacore instrument. Fabs that recognize epitopes located away from the site of mutations were used as indirect probes for the conformational integrity of protein antigens. Their interaction kinetics with all proteins were similar, suggesting that the substitutions had no drastic effect on their conformation. The five substitutions introduced in the peptide and the protein had minor effects on association rate constants (ka) and significant effects on dissociation rate constants (kd) of the antigen-Fab 57P interactions. In four out of five cases, the effect on binding affinity of the substitutions was identical when the epitope was presented in the form of a peptide or a protein antigen, indicating that antibody binding specifity was not affected by epitope presentation. However, ka values were about 10 times larger and kd values about 5 times larger for the peptide-Fab compared to the protein-Fab interaction, suggesting a different binding mechanism. Circular dichroism measurements performed for three of the peptides showed that they were mainly lacking structure in solution. Differences in conformational properties of the peptide and protein antigens in solution and/or in the paratope could explain differences in binding kinetics. Our results demonstrate that the peptides were able to mimic correctly some but not all properties of the protein-Fab 57P interaction and highlight the importance of quantitative analysis of both equilibrium and kinetic binding parameters in the design of synthetic vaccines and drugs.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{dumortier_at_1998, title = {At least three linear regions but not the zinc-finger domain of U1C protein are exposed at the surface of the protein in solution and on the human spliceosomal U1 snRNP particle}, author = {H Dumortier and J Klein Gunnewiek and J P Roussel and Y van Aarssen and J P Briand and W J van Venrooij and S Muller}, doi = {10.1093/nar/26.23.5486}, issn = {0305-1048}, year = {1998}, date = {1998-12-01}, journal = {Nucleic Acids Research}, volume = {26}, number = {23}, pages = {5486--5491}, abstract = {No structural information on U1C protein either in its free state or bound to the spliceosomal U1 small nuclear ribonucleoprotein (snRNP) particle is currently available. Using rabbit antibodies raised against a complete set of 15 U1C overlapping synthetic peptides (16-30 residues long) in different immunochemical tests, linear regions exposed at the surface of free and U1 snRNP-bound U1C were identified. Epitopes within at least three regions spanning residues 31-62, 85-103 and 116-159 were recognized on free and plastic-immobilized recombinant human U1C expressed in Escherichia coli, on in vitro translated U1C protein and on U1C bound to the U1 snRNP particle present in HeLa S100 extract. Using a zinc affinity labeling method, we further showed that the N-terminal U1C peptide containing a zinc-finger motif (peptide 5-34) effectively binds65Zn2+. The N-terminal region of U1C, which is functional in U1 snRNP assembly, is apparently not located at the surface of the U1 snRNP particle.}, keywords = {Amino Acid Sequence, Animals, Dumortier, HeLa Cells, Humans, I2CT, Molecular Sequence Data, Peptide Fragments, Protein Binding, Rabbits, Ribonucleoprotein, Ribonucleoproteins, Small Nuclear, Solutions, Spliceosomes, Team-Dumortier, U1 Small Nuclear, Zinc, Zinc Fingers}, pubstate = {published}, tppubtype = {article} } @article{hoet_human_1998, title = {Human monoclonal autoantibody fragments from combinatorial antibody libraries directed to the U1snRNP associated U1C protein; epitope mapping, immunolocalization and V-gene usage}, author = {R M Hoet and J M Raats and R de Wildt and H Dumortier and S Muller and F van den Hoogen and W J van Venrooij}, doi = {10.1016/s0161-5890(98)00093-5}, issn = {0161-5890}, year = {1998}, date = {1998-11-01}, journal = {Molecular Immunology}, volume = {35}, number = {16}, pages = {1045--1055}, abstract = {To study the localization and function of the U1snRNP associated U1C protein, so far only human sera from systemic lupus erythematosus (SLE) overlap syndrome patients have been used. Here we report for the first time the isolation of human monoclonal anti-UIC autoantibody fragments from IgG derived combinatorial and semi-synthetic human antibody libraries. Two classes of human monoclonal anti-UIC (auto)antibodies were found: specific anti-U1C autoantibodies, recognizing U1C only, and cross-reactive antibodies which also react with U1A and Sm-B/B'proteins. The heavy chains (V(H)genes) of all five antibodies from the semi-synthetic libraries and two of the three U1C-specific patient derived autoantibody fragments are encoded by V(H)3 genes, in which V(H) 3-30 (DP-49) was overrepresented. The heavy chain of the two cross-reactive autoantibodies are derived from the 3-07 (DP-54) gene. Three epitope regions on the U1C protein are targeted by these antibodies. (1) Four U1C specific antibodies recognize an N-terminal region of U1C in which amino acids 30-63 are essential for recognition, (2) two antibodies recognize only the complete U1C protein, and (3) two cross-reactive and one U1C specific antibody recognize the C-terminal domain in which amino acids 98-126 are critical for recognition. The two cross-reactive antibodies (K 11 and K 15) recognize the proline-rich region of the U1C protein (amino acids 98 126) and cross-react with proline-rich regions in Sm-B/B' (amino acids 163-184) and U1A (amino acids 187-204). All 10 antibody fragments are able to immunoprecipitate the native U1snRNP particle. The two cross-reactive antibodies immunoprecipitate the other Sm containing snRNPs as well. Using confocal immunofluorescence microscopy we could show that the major part of the U1C protein is localized within the coiled body structure.}, keywords = {Amino Acid Sequence, Antibodies, Autoantibodies, Cross Reactions, Dumortier, Epitope Mapping, Genes, HeLa Cells, Humans, I2CT, Immunoglobulin, Immunoglobulin Fragments, Immunoglobulin Variable Region, Immunohistochemistry, Lupus Erythematosus, Monoclonal, Ribonucleoprotein, Ribonucleoproteins, Small Nuclear, Systemic, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{braun_analysis_1998, title = {Analysis of the Drosophila host defense in domino mutant larvae, which are devoid of hemocytes}, author = {A Braun and Jules A Hoffmann and Marie Meister}, issn = {0027-8424}, year = {1998}, date = {1998-11-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {95}, number = {24}, pages = {14337--14342}, abstract = {We have analyzed the Drosophila immune response in domino mutant larvae, which are devoid of blood cells. The domino mutants have a good larval viability, but they die as prepupae. We show that, on immune challenge, induction of the genes encoding antimicrobial peptides in the fat body is not affected significantly in the mutant larvae, indicating that hemocytes are not essential in this process. The hemocoele of domino larvae contains numerous live microorganisms, the presence of which induces a weak antimicrobial response in the fat body. A full response is observed only after septic injury. We propose that the fat body cells are activated both by the presence of microorganisms and by injury and that injury potentiates the effect of microorganisms. Survival experiments after an immune challenge showed that domino mutants devoid of blood cells maintain a wild-type resistance to septic injury. This resistance was also observed in mutant larvae in which the synthesis of antibacterial peptides is impaired (immune deficiency larvae) and in mutants that are deficient for humoral melanization (Black cells larvae). However, if domino was combined with either the immune deficiency or the Black cell mutation, the resistance to septic injury was reduced severely. These results establish the relevance of the three immune reactions: phagocytosis, synthesis of antibacterial peptides, and melanization. By working in synergy, they provide Drosophila a highly effective defense against injury and/or infection.}, keywords = {Adipose Tissue, Animals, Candida, Escherichia coli, Fungal, Genotype, Hemocytes, hoffmann, Larva, M3i, Melanins, Micrococcus luteus, Spores}, pubstate = {published}, tppubtype = {article} } @article{uttenweiler-joseph_differential_1998, title = {Differential display of peptides induced during the immune response of Drosophila: a matrix-assisted laser desorption ionization time-of-flight mass spectrometry study}, author = {S Uttenweiler-Joseph and M Moniatte and Marie Lagueux and Van A Dorsselaer and Jules A Hoffmann and Philippe Bulet}, issn = {0027-8424}, year = {1998}, date = {1998-09-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {95}, number = {19}, pages = {11342--11347}, abstract = {We have developed an approach based on a differential mass spectrometric analysis to detect molecules induced during the immune response of Drosophila, regardless of their biological activities. For this, we have applied directly matrix-assisted laser desorption/ionization MS to hemolymph samples from individual flies before and after an immune challenge. This method provided precise information on the molecular masses of immune-induced molecules and allowed the detection, in the molecular range of 1.5-11 kDa, of 24 Drosophila immune-induced molecules (DIMs). These molecules are all peptides, and four correspond to already characterized antimicrobial peptides. We have further analyzed the induction of the various peptides by immune challenge in wild-type flies and in mutants with a compromised antimicrobial response. We also describe a methodology combining matrix-assisted laser desorption ionization time-of-flight MS, HPLC, and Edman degradation, which yielded the peptide sequence of three of the DIMs. Finally, molecular cloning and Northern blot analyses revealed that one of the DIMs is produced as a prepropeptide and is inducible on a bacterial challenge.}, keywords = {Animals, bacteria, Chromatography, Cloning, Hemolymph, High Pressure Liquid, hoffmann, Immunity, Insect Proteins, M3i, Mass, Matrix-Assisted Laser Desorption-Ionization, messenger, Molecular, Peptides, Protein Precursors, RNA, Sequence Analysis, Spectrometry, Time Factors}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_drosomycin-gfp_1998, title = {A drosomycin-GFP reporter transgene reveals a local immune response in Drosophila that is not dependent on the Toll pathway}, author = {Dominique Ferrandon and Alain C Jung and M Criqui and Bruno Lemaitre and S Uttenweiler-Joseph and Lydia Michaut and Jean-Marc Reichhart and Jules A Hoffmann}, doi = {10.1093/emboj/17.5.1217}, issn = {0261-4189}, year = {1998}, date = {1998-08-01}, journal = {EMBO J.}, volume = {17}, number = {5}, pages = {1217--1227}, abstract = {A hallmark of the systemic antimicrobial response of Drosophila is the synthesis by the fat body of several antimicrobial peptides which are released into the hemolymph in response to a septic injury. One of these peptides, drosomycin, is active primarily against fungi. Using a drosomycin-green fluorescent protein (GFP) reporter gene, we now show that in addition to the fat body, a variety of epithelial tissues that are in direct contact with the external environment, including those of the respiratory, digestive and reproductive tracts, can express the antifungal peptide, suggesting a local response to infections affecting these barrier tissues. As is the case for vertebrate epithelia, insect epithelia appear to be more than passive physical barriers and are likely to constitute an active component of innate immunity. We also show that, in contrast to the systemic antifungal response, this local immune response is independent of the Toll pathway.}, keywords = {Animals, bacteria, Cell Surface, Developmental, Digestive System, Epithelium, Fat Body, Female, ferrandon, Fungal, Gene Expression Regulation, Genes, Green Fluorescent Proteins, hoffmann, Insect Proteins, Larva, Luminescent Proteins, M3i, Male, Membrane Glycoproteins, Organ Specificity, Receptors, reichhart, Reporter, Respiratory System, Spores, Toll-Like Receptors, Trachea, Transgenes}, pubstate = {published}, tppubtype = {article} } @article{bulet_differential_1998, title = {Differential display of peptides induced during the immune response of Drosophila: a matrix-assisted laser desorption ionization time-of-flight mass spectrometry study}, author = {Philippe Bulet and S Uttenweiler-Joseph and M Moniatte and Van A Dorsselaer and Jules A Hoffmann}, issn = {0277-8033}, year = {1998}, date = {1998-08-01}, journal = {J. Protein Chem.}, volume = {17}, number = {6}, pages = {528--529}, keywords = {Animals, Anti-Infective Agents, hoffmann, M3i, Mass, Matrix-Assisted Laser Desorption-Ionization, Peptide Biosynthesis, Peptides, Spectrometry}, pubstate = {published}, tppubtype = {article} } @article{shahabuddin_plasmodium_1998, title = {Plasmodium gallinaceum: differential killing of some mosquito stages of the parasite by insect defensin}, author = {M Shahabuddin and I Fields and Philippe Bulet and Jules A Hoffmann and L H Miller}, doi = {10.1006/expr.1998.4212}, issn = {0014-4894}, year = {1998}, date = {1998-05-01}, journal = {Exp. Parasitol.}, volume = {89}, number = {1}, pages = {103--112}, abstract = {We examined several insect antimicrobial peptides to study their effect on Plasmodium gallinaceum zygotes, ookinetes, oocysts, and sporozoites. Only two insect defensins-Aeschna cyanea (dragon fly) and Phormia terranovae (flesh fly)-had a profound toxic effect on the oocysts in Aedes aegypti and on isolated sporozoites. The defensins affected the oocysts in a time-dependent manner. Injecting the peptide into the hemolymph 1 or 2 days after an infectious blood meal had no significant effect on prevalence of infection or relative oocyst density per mosquito. When injected 3 days after parasite ingestion, the relative oocyst density was significantly reduced. Injection on day 4 or later damaged the developing oocysts, although the oocysts density per mosquito was not significantly different when examined on day 8. The oocysts were swollen or had extensive internal vacuolization. The peptides had no detectable effect on the early stages of the parasite: the zygotes and ookinetes tested in vitro. Both the defensins were highly toxic to isolated sporozoites in vitro as indicated by disruption of the membrane permeability barrier, a change in morphology, and loss of motility. In contrast to the toxicity of cecropin and magainin for mosquitoes, defensin, at concentrations that kill parasites, is not toxic to mosquitoes, suggesting that defensin should be studied further as a potential molecule to block sporogonic development of Plasmodium.}, keywords = {Aedes, Animals, Anti-Infective Agents, Blood Proteins, Defensins, Diptera, hoffmann, Insect Vectors, insects, M3i, Plasmodium gallinaceum, Zygote}, pubstate = {published}, tppubtype = {article} } @article{taguchi_novel_1998, title = {A novel insect defensin from the ant Formica rufa}, author = {S Taguchi and Philippe Bulet and Jules A Hoffmann}, issn = {0300-9084}, year = {1998}, date = {1998-04-01}, journal = {Biochimie}, volume = {80}, number = {4}, pages = {343--346}, abstract = {By combination of size exclusion and reversed-phase chromatography, we have isolated a novel member of insect defensin-type antimicrobial peptides from the entire bodies of bacteria-challenged Formica rufa (hymenoptera, formicidae). The molecular mass of the purified peptide was estimated to be 4120.42 by matrix-assisted laser desorption/ionization-time of flight/mass spectrometry. Sequence analysis revealed that this peptide consisted of 40 amino acid residues with six cysteines engaged in the formation of three intramolecular disulfide bridges. This peptide is unique among the arthropod defensins in terms of the presence of asparatic acid and alanine at position 33 and as C-terminal residue, respectively. In addition, this novel defensin from Formica rufa has the particularity to have no C-terminal extension in contrast to those reported for other hymenoptera defensins.}, keywords = {Amino Acid, Animals, Anti-Bacterial Agents, Ants, Chromatography, High Pressure Liquid, hoffmann, Insect Proteins, insects, M3i, Mass, Matrix-Assisted Laser Desorption-Ionization, Protein Structure, Secondary, Sequence Alignment, Sequence Homology, Spectrometry}, pubstate = {published}, tppubtype = {article} } @article{nicolas_vivo_1998, title = {In vivo regulation of the IkappaB homologue cactus during the immune response of Drosophila}, author = {E Nicolas and Jean-Marc Reichhart and Jules A Hoffmann and Bruno Lemaitre}, issn = {0021-9258}, year = {1998}, date = {1998-04-01}, journal = {J. Biol. Chem.}, volume = {273}, number = {17}, pages = {10463--10469}, abstract = {The dorsoventral regulatory gene pathway (spätzle/Toll/cactus) controls the expression of several antimicrobial genes during the immune response of Drosophila. This regulatory cascade shows striking similarities with the cytokine-induced activation cascade of NF-kappaB during the inflammatory response in mammals. Here, we have studied the regulation of the IkappaB homologue Cactus in the fat body during the immune response. We observe that the cactus gene is up-regulated in response to immune challenge. Interestingly, the expression of the cactus gene is controlled by the spätzle/Toll/cactus gene pathway, indicating that the cactus gene is autoregulated. We also show that two Cactus isoforms are expressed in the cytoplasm of fat body cells and that they are rapidly degraded and resynthesized after immune challenge. This degradation is also dependent on the Toll signaling pathway. Altogether, our results underline the striking similarities between the regulation of IkappaB and cactus during the immune response.}, keywords = {Animals, Cell Surface, DNA-Binding Proteins, Gene Expression Regulation, hoffmann, Insect Proteins, Larva, M3i, Membrane Glycoproteins, Phosphoproteins, Proto-Oncogene Proteins, Receptors, reichhart, Signal Transduction, Toll-Like Receptors, Transcription Factor RelB, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{, title = {Concomitant changes in polyamine pools and DNA methylation during growth inhibition of human colonic cancer cells}, author = { B. Duranton and G. Keith and Goss and C. Bergmann and R. Schleiffer and F. Raul}, year = {1998}, date = {1998-01-01}, journal = {Exp Cell Res}, volume = {243}, number = {2}, pages = {319-25}, abstract = {The effects of CGP 48664 and DFMO, selective inhibitors of the key enzymes of polyamine biosynthesis, namely, of S-adenosylmethionine decarboxylase (AdoMetDC) and ornithine decarboxylase (ODC), were investigated on growth, polyamine metabolism, and DNA methylation in the Caco-2 cell line. Both inhibitors caused growth inhibition and affected similarly the initial expression of the differentiation marker sucrase. In the presence of the AdoMetDC inhibitor, ODC activity and the intracellular pool of putrescine were enhanced, whereas the spermidine and spermine pools were decreased. In the presence of the ODC inhibitor, the AdoMetDC activity was enhanced and the intracellular pools of putrescine and spermidine were decreased. With both compounds, the degree of global DNA methylation was increased. Spermine and spermidine (but not putrescine) selectively inhibited cytosine-DNA methyltransferase activity. Our observations suggest that spermidine (and to a lesser extent spermine) controls DNA methylation and may represent a crucial step in the regulation of Caco-2 cell growth and differentiation.}, note = {0014-4827 Journal Article}, keywords = {*Cell, *DNA, &, Adenosylmethionine, Amidines/pharmacology, Caco-2, Cells, Decarboxylase/antagonists, Differentiation/drug, Division/drug, effects, Eflornithine/pharmacology, enzyme, Gov't, Human, Indans/pharmacology, inhibitors/metabolism, Inhibitors/pharmacology, Methylation, Non-U.S., Ornithine, Polyamines/*metabolism, Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {All in the family: structural and evolutionary relationships among three modular proteins with diverse functions and variable assembly}, author = { M. Bergdoll and L. D. Eltis and A. D. Cameron and P. Dumas and J. T. Bolin}, year = {1998}, date = {1998-01-01}, journal = {Protein Sci}, volume = {7}, number = {8}, pages = {1661-70}, abstract = {The crystal structures of three proteins of diverse function and low sequence similarity were analyzed to evaluate structural and evolutionary relationships. The proteins include a bacterial bleomycin resistance protein, a bacterial extradiol dioxygenase, and human glyoxalase I. Structural comparisons, as well as phylogenetic analyses, strongly indicate that the modern family of proteins represented by these structures arose through a rich evolutionary history that includes multiple gene duplication and fusion events. These events appear to be historically shared in some cases, but parallel and historically independent in others. A significant early event is proposed to be the establishment of metal-binding in an oligomeric ancestor prior to the first gene fusion. Variations in the spatial arrangements of homologous modules are observed that are consistent with the structural principles of three-dimensional domain swapping, but in the unusual context of the formation of larger monomers from smaller dimers or tetramers. The comparisons support a general mechanism for metalloprotein evolution that exploits the symmetry of a homooligomeric protein to originate a metal binding site and relies upon the relaxation of symmetry, as enabled by gene duplication, to establish and refine specific functions.}, note = {0961-8368 Journal Article}, keywords = {*Acetyltransferases, *Evolution, Acid, Amino, Bacterial, Burkholderia/*chemistry, Crystallography, Data, Genetic, Gov't, Homology, Human, Lactoylglutathione, Lyase/*chemistry, Models, Molecular, Non-U.S., Oxygenases/chemistry, P.H.S., Phylogeny, Protein, Proteins/*chemistry, Secondary, Sequence, structure, Support, U.S., X-Ray}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of the tRNAs which up-regulate agrostin, barley RIP and PAP-S, three ribosome-inactivating proteins of plant origin}, author = { M. Brigotti and G. Keith and A. Pallanca and D. Carnicelli and P. Alvergna and G. Dirheimer and L. Montanaro and S. Sperti}, year = {1998}, date = {1998-01-01}, journal = {FEBS Lett}, volume = {431}, number = {2}, pages = {259-62}, abstract = {Ribosome-inactivating proteins (RIP) are RNA-N-glycosidases widely diffused in plants which depurinate ribosomal RNA at a specific universally conserved position, A4324 in rat ribosomes. A small group of RIPs (cofactor-dependent RIPs) require ATP and tRNA to reach maximal activity on isolated ribosomes. The tRNA which stimulates gelonin was identified as tRNA(Trp). The present paper reports the identification of three other tRNAs which stimulate agrostin (tRNA(Ala)), barley RIP (tRNA(Ala), tRNA(Val)) and PAP-S (tRNA(Gly)), while for tritin-S no particular stimulating tRNA emerged. The sequences of tRNA(Val) and tRNA(Gly) correspond to the already known ones (rabbit and man, respectively). The tRNA(Ala) (anticodon IGC) identifies a new isoacceptor. Only the stimulating activity of the tRNA(Ala) for agrostin approaches the specificity previously observed for the couple gelonin-tRNA(Trp).}, note = {0014-5793 Journal Article}, keywords = {&, Acid, Adenosine, Base, Conformation, Data, effects/*metabolism, Gov't, Hordeum/metabolism, Hydrolases/*metabolism, Molecular, N-Glycosyl, Non-U.S., Nucleic, Plant, Plant/chemistry/isolation, Proteins/drug, purification/*metabolism, RNA, Sequence, Support, Transfer/chemistry/isolation, Triphosphate/pharmacology, Up-Regulation}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interactions between Ty1 retrotransposon RNA and the T and D regions of the tRNA(iMet) primer are required for initiation of reverse transcription in vivo}, author = { S. Friant and T. Heyman and A. S. Bystrom and M. Wilhelm and F. X. Wilhelm}, year = {1998}, date = {1998-01-01}, journal = {Mol Cell Biol}, volume = {18}, number = {2}, pages = {799-806}, abstract = {Reverse transcription of the Saccharomyces cerevisiae Ty1 retrotransposon is primed by tRNA(iMet) base paired to the primer binding site (PBS) near the 5' end of Ty1 genomic RNA. The 10-nucleotide PBS is complementary to the last 10 nucleotides of the acceptor stem of tRNA(iMet). A structural probing study of the interactions between the Ty1 RNA template and the tRNA(iMet) primer showed that besides interactions between the PBS and the 3' end of tRNA(iMet), three short regions of Ty1 RNA, named boxes 0, 1, and 2.1, interact with the T and D stems and loops of tRNA(iMet). To determine if these sequences are important for the reverse transcription pathway of the Ty1 retrotransposon, mutant Ty1 elements and tRNA(iMet) were tested for the ability to support transposition. We show that the Ty1 boxes and the complementary sequences in the T and D stems and loops of tRNA(iMet) contain bases that are critical for Ty1 retrotransposition. Disruption of 1 or 2 bp between tRNA(iMet) and box 0, 1, or 2.1 dramatically decreases the level of transposition. Compensatory mutations which restore base pairing between the primer and the template restore transposition. Analysis of the reverse transcription intermediates generated inside Ty1 virus-like particles indicates that initiation of minus-strand strong-stop DNA synthesis is affected by mutations disrupting complementarity between Ty1 RNA and primer tRNA(iMet).}, note = {0270-7306 Journal Article}, keywords = {*Retroelements, *Transcription, Acid, Base, Binding, cerevisiae, Conformation, Data, DNA, Fungal/*metabolism, Fungal/biosynthesis, Genetic, Gov't, Met/*metabolism, Molecular, Mutagenesis, Non-U.S., Nucleic, Primers, Replication, RNA, Saccharomyces, Sequence, Sites, Support, Transfer}, pubstate = {published}, tppubtype = {article} } @article{, title = {The yeast tRNA:pseudouridine synthase Pus1p displays a multisite substrate specificity}, author = { Y. Motorin and G. Keith and C. Simon and D. Foiret and G. Simos and E. Hurt and H. Grosjean}, year = {1998}, date = {1998-01-01}, journal = {RNA}, volume = {4}, number = {7}, pages = {856-69}, abstract = {We have previously shown that the yeast gene PUS1 codes for a tRNA:pseudouridine synthase and that recombinant Pus1p catalyzes, in an intron-dependent way, the formation of psi34 and psi36 in the anticodon loop of the yeast minor tRNA(Ile) in vitro (Simos G et al., 1996, EMBO J 15:2270-2284). Using a set of T7 transcripts of different tRNA genes, we now demonstrate that yeast pseudouridine synthase 1 catalyzes in vitro pseudouridine formation at positions 27 and/or 28 in several yeast cytoplasmic tRNAs and at position 35 in the intron-containing tRNA(Tyr) (anticodon GUA). Thus, Pus1p not only displays a broad specificity toward the RNA substrates, but is also capable of catalyzing the pseudouridine (psi) formation at distinct noncontiguous sites within the same tRNA molecule. The cell-free extract prepared from the yeast strain bearing disrupted gene PUS1 is unable to catalyze the formation of psi27, psi28, psi34, and psi36 in vitro, however, psi35 formation in the intron-containing tRNA(Tyr)(GUA) remains unaffected. Thus, in yeast, only one gene product accounts for tRNA pseudouridylation at positions 27, 28, 34, and 36, whereas for position 35 in tRNA(Tyr), another site-specific tRNA:pseudouridine synthase with overlapping specificity exists. Mapping of pseudouridine residues present in various tRNAs extracted from the PUS1-disrupted strain confirms the in vitro data obtained with the recombinant Pus1p. In addition, they suggest that Pus1p is implicated in modification at positions U26, U65, and U67 in vivo.}, note = {1355-8382 Journal Article}, keywords = {*RNA, cerevisiae, Cloning, Fractions/metabolism, Fungal, Fungal/metabolism, Gov't, Hydro-Lyases/biosynthesis/genetics/*metabolism, Molecular, Mutation, Non-U.S., Plant/metabolism, post-transcriptional, Precursors/*metabolism, Processing, Proteins/biosynthesis, Proteins/biosynthesis/genetics/metabolism, Pseudouridine/*biosynthesis, Recombinant, RNA, Saccharomyces, Specificity, Subcellular, Substrate, Support, Transfer/*metabolism}, pubstate = {published}, tppubtype = {article} } @article{, title = {The yeast Ty3 retrotransposon contains a 5'-3' bipartite primer-binding site and encodes nucleocapsid protein NCp9 functionally homologous to HIV-1 NCp7}, author = { C. Gabus and D. Ficheux and M. Rau and G. Keith and S. Sandmeyer and J. L. Darlix}, year = {1998}, date = {1998-01-01}, journal = {EMBO J}, volume = {17}, number = {16}, pages = {4873-80}, abstract = {Retroviruses, including HIV-1 and the distantly related yeast retroelement Ty3, all encode a nucleoprotein required for virion structure and replication. During an in vitro comparison of HIV-1 and Ty3 nucleoprotein function in RNA dimerization and cDNA synthesis, we discovered a bipartite primer-binding site (PBS) for Ty3 composed of sequences located at opposite ends of the genome. Ty3 cDNA synthesis requires the 3' PBS for primer tRNAiMet annealing to the genomic RNA, and the 5' PBS, in cis or in trans, as the reverse transcription start site. Ty3 RNA alone is unable to dimerize, but formation of dimeric tRNAiMet bound to the PBS was found to direct dimerization of Ty3 RNA-tRNAiMet. Interestingly, HIV-1 nucleocapsid protein NCp7 and Ty3 NCp9 were interchangeable using HIV-1 and Ty3 RNA template-primer systems. Our findings impact on the understanding of non-canonical reverse transcription as well as on the use of Ty3 systems to screen for anti-NCp7 drugs.}, note = {0261-4189 Journal Article}, keywords = {*Capsid, *Retroelements, Acid, Base, Binding, Capsid/*genetics, cerevisiae/*genetics, dimerization, gag/*genetics, Gene, Gov't, Homology, Met/genetics/*metabolism, Non-U.S., Nucleic, P.H.S., Products, Proteins, RNA, Saccharomyces, Sequence, Sites, Support, Transfer, U.S.}, pubstate = {published}, tppubtype = {article} } @article{reichhart_green_1998, title = {Green Balancers.}, author = {Jean-Marc Reichhart and Dominique Ferrandon}, year = {1998}, date = {1998-01-01}, journal = {D. I. S.}, volume = {81}, pages = {201--202}, abstract = {We have used the S65T green fluorescent protein (GFP ; (Chalfie et al., 1994) ; (Heim et al., 1995)) as a vital reporter to introduce a dominant innocuous marker onto the balancers of the three major chromosomes of D. melanogaster.}, keywords = {ferrandon, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{levashina_two_1998, title = {Two distinct pathways can control expression of the gene encoding the Drosophila antimicrobial peptide metchnikowin}, author = {Elena A Levashina and S Ohresser and Bruno Lemaitre and Jean-Luc Imler}, doi = {10.1006/jmbi.1998.1705}, issn = {0022-2836}, year = {1998}, date = {1998-01-01}, journal = {Journal of Molecular Biology}, volume = {278}, number = {3}, pages = {515--527}, abstract = {Metchnikowin is a recently discovered proline-rich peptide from Drosophila with antibacterial and antifungal properties. Like most other antimicrobial peptides from insects, its expression is immune-inducible. Here we present evidence that induction of metchnikowin gene expression can be mediated either by the TOLL pathway or by the imd gene product. We show that the gene remains inducible in Toll-deficient mutants, in which the antifungal response is blocked, as well as in imd mutants, which fail to mount an antibacterial response. However, in Toll-deficient;imd double mutants, metchnikowin gene expression can no longer be detected after immune challenge. Our results suggest that expression of this peptide with dual activity can be triggered by signals generated by either bacterial or fungal infection. Cloning of the metchnikowin gene revealed the presence in the 5' flanking region of several putative cis-regulatory motifs characterized in the promoters of insect immune genes: namely, Rel sites, GATA motifs, interferon consensus response elements and NF-IL6 response elements. Establishment of transgenic fly lines in which the GFP reporter gene was placed under the control of 1.5 kb of metchnikowin gene upstream sequences indicates that this fragment is able to confer full immune inducibility and tissue specificity of expression on the transgene.}, keywords = {Animals, Anti-Infective Agents, Antimicrobial Cationic Peptides, Base Sequence, Cloning, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Glycopeptides, imler, Insect, Insect Proteins, Larva, M3i, Molecular, Mutation, Peptides, Promoter Regions, Recombinant Fusion Proteins, Reporter, Restriction Mapping, Transcription}, pubstate = {published}, tppubtype = {article} } @article{lemaitre_drosophila_1997, title = {Drosophila host defense: differential induction of antimicrobial peptide genes after infection by various classes of microorganisms}, author = {Bruno Lemaitre and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0027-8424}, year = {1997}, date = {1997-12-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {94}, number = {26}, pages = {14614--14619}, abstract = {Insects respond to microbial infection by the rapid and transient expression of several genes encoding potent antimicrobial peptides. Herein we demonstrate that this antimicrobial response of Drosophila is not aspecific but can discriminate between various classes of microorganisms. We first observe that the genes encoding antibacterial and antifungal peptides are differentially expressed after injection of distinct microorganisms. More strikingly, Drosophila that are naturally infected by entomopathogenic fungi exhibit an adapted response by producing only peptides with antifungal activities. This response is mediated through the selective activation of the Toll pathway.}, keywords = {Animals, Genes, hoffmann, Immunity, Insect, M3i, Peptides, reichhart}, pubstate = {published}, tppubtype = {article} } @article{meister_antimicrobial_1997, title = {Antimicrobial peptide defense in Drosophila}, author = {Marie Meister and Bruno Lemaitre and Jules A Hoffmann}, doi = {10.1002/bies.950191112}, issn = {0265-9247}, year = {1997}, date = {1997-11-01}, journal = {Bioessays}, volume = {19}, number = {11}, pages = {1019--1026}, abstract = {Drosophila responds to a septic injury by the rapid synthesis of antimicrobial peptides. These molecules are predominantly produced by the fat body, a functional equivalent of mammalian liver, and are secreted into the hemolymph where their concentrations can reach up to 100 microM. Six distinct antibacterial peptides (plus isoforms) and one antifungal peptide have been characterized in Drosophila and their genes cloned. The induction of the gene encoding the antifungal peptide relies on the spätzle/Toll/cactus gene cassette, which is involved in the control of dorsoventral patterning in the embryo, and shows interesting structural and functional similarities with cytokine-induced activation of NF-kappa B in mammalian cells. An additional pathway, dependent on the as yet unidentified imd (for immune-deficiency) gene, is required for the full induction of the antibacterial peptide genes. Mutants deficient for the Toll and imd pathways exhibit a severely reduced survival to fungal and bacterial infections, respectively. Recent data on the molecular mechanisms underlying recognition of non-self are also discussed in this review.}, keywords = {Animals, Anti-Infective Agents, Gene Expression Regulation, Genetic, hoffmann, Insect Proteins, M3i, Models, Peptides, Promoter Regions, Signal Transduction}, pubstate = {published}, tppubtype = {article} } @article{meziere_vivo_1997, title = {In vivo Ŧ helper cell response to retro-inverso peptidomimetics}, author = {C Mézière and M Viguier and H Dumortier and R Lo-Man and C Leclerc and J G Guillet and J P Briand and S Muller}, issn = {0022-1767}, year = {1997}, date = {1997-10-01}, journal = {Journal of Immunology (Baltimore, Md.: 1950)}, volume = {159}, number = {7}, pages = {3230--3237}, abstract = {Peptide analogues containing reversed peptide bonds between each residue along the peptide sequence (retro-inverso modification) have been analyzed for their antigenic and in vivo immunogenic properties in the MHC II and Th cell response context. Two antigenic peptides were selected for this study, namely peptide 103-115 of poliovirus VP1, which is involved in the production of Abs that neutralize the infectivity of the virus, and peptide 435-446 from the third constant region of mouse heavy chain IgG2a allopeptide gamma 2ab, which mimics a corneal Ag implicated in autoimmune keratitis. In a competition assay performed in vitro using reference hybridomas of known MHC class II restriction, both retro-inverso analogues bound (although more weakly in our test) to I-Ad and/or I-Ed class II molecules. However, in both cases, this lower affinity was apparently largely compensated in vivo, as a T cell response (with IL-2 secretion), equivalent to that obtained with the wild-type peptides, was observed following immunization of BALB/c mice with the retro-inverso analogues. Moreover, these T cells proliferated and produced IL-2 in response to the cognate peptides. It is concluded that the T cell receptors of T cells primed in vivo with the retro-inverso analogues readily cross-react with parent and retro-inverso analogue-MHC complexes. The approach of using pseudopeptides containing changes involving the backbone, and not the orientation of side chains, may thus be promising to design potent immunogens for class II-restricted T cells.}, keywords = {Amino Acid Sequence, Animals, Antibodies, Antigen, Capsid, Capsid Proteins, Dumortier, Female, Helper-Inducer, Histocompatibility Antigens Class II, I2CT, Immunoglobulin Allotypes, Immunoglobulin G, Inbred BALB C, Injections, Intraperitoneal, Lymphocyte Activation, Mice, Molecular Sequence Data, Peptide Fragments, Poliovirus, Protein Binding, Receptors, T-Cell, T-Lymphocytes, Team-Dumortier, Viral}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_immune_1997, title = {Immune responsiveness in vector insects}, author = {Jules A Hoffmann}, issn = {0027-8424}, year = {1997}, date = {1997-10-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {94}, number = {21}, pages = {11152--11153}, keywords = {Animals, Anopheles, bacteria, Blood Proteins, Defensins, hoffmann, Humans, Insect Vectors, Life Cycle Stages, M3i, Malaria, Mammals, Plasmodium}, pubstate = {published}, tppubtype = {article} } @article{dimarcq_treatment_1997, title = {Treatment of l(2)mbn Drosophila tumorous blood cells with the steroid hormone ecdysone amplifies the inducibility of antimicrobial peptide gene expression}, author = {Jean-Luc Dimarcq and Jean-Luc Imler and R Lanot and Alan R B Ezekowitz and Jules A Hoffmann and C A Janeway and Marie Lagueux}, issn = {0965-1748}, year = {1997}, date = {1997-10-01}, journal = {Insect Biochemistry and Molecular Biology}, volume = {27}, number = {10}, pages = {877--886}, abstract = {Insects rely on both humoral and cellular mechanisms to defend themselves against microbial infections. The humoral response involves synthesis of a battery of potent antimicrobial peptides by the fat body and, to a lesser extent, by blood cells. The cellular response on the other hand consists of phagocytosis of small microorganisms and melanization and encapsulation of larger parasites. The l(2)mbn cell line, established from tumorous larval hemocytes, represents a system of choice to dissect the molecular events controlling cellular immunity. We report here that l(2)mbn cells can be efficiently induced to differentiate in adherent, macrophage-like cells by treatment with 20-hydroxyecdysone. Ecdysone treatment increases both the phagocytic capacity of l(2)mbn cells and their competence to express antimicrobial genes in response to immune challenge. We also report that expression of several regulatory molecules thought to be involved in the immune response is up-regulated by ecdysone in l(2)mbn cells.}, keywords = {Animals, Bacterial Infections, Cellular, Ecdysone, Gene Expression, Genes, Hemocytes, Hemolymph, hoffmann, imler, Immunity, Insect, M3i, Macrophages, Peptide Biosynthesis, Phagocytosis}, pubstate = {published}, tppubtype = {article} } @article{, title = {The new world of ribozymes.}, author = { L. Jaeger}, year = {1997}, date = {1997-01-01}, journal = {Curr Opin Struct Biol}, volume = {7}, number = {3}, pages = {324-335}, abstract = {The number of RNA molecules that have novel catalytic activities has dramatically increased during the past two years. This ribozymic boom is not due to the discovery of additional examples of natural ribozymes but rather to the development of artificial ribozymes isolated by in vitro selection and evolution techniques. The structural and functional complexities of these artificial ribozymes, however, do not match those of the larger natural ribozymes. The understanding of both RNA structure and catalysis performed by natural and artificial ribozymes paves the way for the creation of RNA molecules that are able to efficiently catalyze more complex reactions.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Proline-dependent oligomerization with arm exchange}, author = { M. Bergdoll and M. H. Remy and C. Cagnon and J. M. Masson and P. Dumas}, year = {1997}, date = {1997-01-01}, journal = {Structure}, volume = {5}, number = {3}, pages = {391-401}, abstract = {BACKGROUND: Oligomerization is often necessary for protein activity or regulation and its efficiency is fundamental for the cell. The quaternary structure of a large number of oligomers consists of protomers tightly anchored to each other by exchanged arms or swapped domains. However, nothing is known about how the arms can be kept in a favourable conformation before such an oligomerization. RESULTS: Upon examination of such quaternary structures, we observe an extremely frequent occurrence of proline residues at the point where the arm leaves the protomer. Sequence alignment and site-directed mutagenesis confirm the importance of these prolines. The conservation of these residues at the hinge regions can be explained by the constraints that they impose on polypeptide conformation and dynamics: by rigidifying the mainchain, prolines favour extended conformations of arms thus favouring oligomerization, and may prevent interaction of the arms with the core of the protomer. CONCLUSIONS: Hinge prolines can be considered as 'quaternary structure helpers'. The presence of a proline should be considered when searching for a determinant of oligomerization with arm exchange and could be used to engineer synthetic oligomers or to displace a monomers to oligomers equilibrium by mutation of this proline residue.}, note = {0969-2126 Journal Article}, keywords = {*Acetyltransferases, *Dimerization, *Protein, Acid, Alignment, Amino, Aminotransferases/chemistry, Animals, Aspartate, ATPase/chemistry, Bacterial, Binding, Cattle, Chickens, Comparative, Conformation, Data, Folding, Heart/enzymology, Human, mitochondria, Models, Molecular, Mutagenesis, Na(+)-K(+)-Exchanging, Pancreatic/chemistry, Plant, Proline/*physiology, Protein, Proteins/chemistry, Pyrophosphatases/chemistry, Ribonuclease, Sequence, Site-Directed, Structural, Study, Viral, Viruses/chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Heterogeneous terminal structure of Ty1 and Ty3 reverse transcripts}, author = {M Wilhelm and T Heyman and S Friant and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9153316}, isbn = {9153316}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {11}, pages = {2161-2166}, abstract = {A specific terminal structure of preintegrative DNA is required for transposition of retroviruses and LTR-retrotransposons. We have used an anchored PCR technique to map the 3'ends of DNA intermediates synthesized inside yeast Ty1 and Ty3 retrotransposon virus-like particles. We find that, unlike retroviruses, Ty1 replicated DNA does not have two extra base pairs at its 3'ends. In contrast some Ty3 preintegrative DNA molecules have two extra nucleotides at the 3'end of upstream and downstream long terminal repeats. Moreover we find that some molecules of replicated Ty3 DNA have more than two extra nucleotides at the 3'end of the upstream LTR. This observation could be accounted for by imprecise RNAse H cutting of the PPT sequence. The site of Ty1 and Ty3 plus-strand strong-stop DNA termination was also examined. Our results confirm that the prominent Ty1 and Ty3 plus-strand strong-stop molecules harbor 12 tRNA templated bases but also show that some Ty1 and Ty3 plus-strand strong-stop DNA molecules harbor less tRNA templated bases. We propose that these less than full length plus-strand molecules could be active intermediates in Ty retrotransposon replication.}, note = {0305-1048 Journal Article}, keywords = {Calf Thymus/metabolism Support, DNA Replication DNA, Fungal/*chemistry/metabolism *Nucleic Acid Conformation Nucleic Acid Hybridization Plasmids/chemistry/genetics/metabolism Polymerase Chain Reaction RNA, Genetic, Non-U.S. Gov't *Transcription, Transfer/chemistry Retroelements/*genetics Ribonuclease H, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nucleic acids. From self-assembly to induced-fit recognition}, author = {E Westhof and D J Patel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9204270}, isbn = {9204270}, year = {1997}, date = {1997-01-01}, journal = {Curr Opin Struct Biol}, volume = {7}, number = {3}, pages = {305-309}, note = {0959-440x Editorial Review Review, Tutorial}, keywords = {Catalytic/chemistry/metabolism, Crystallography, Electron/methods Models, Molecular Nucleic Acid Conformation Nucleic Acids/*chemistry/*metabolism Proteins/chemistry/metabolism RNA/*chemistry/*metabolism RNA, Unité ARN, X-Ray/methods DNA/chemistry Magnetic Resonance Spectroscopy/methods Microscopy}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {DNA and RNA structure prediction}, author = {E Westhof and P Auffinger and C Gaspin}, editor = {M Bishop and C Rawlings}, url = {http://www.amazon.com/DNA-Protein-Sequence-Analysis-Practical/dp/0199634637}, doi = {10.1016/S0307-4412(97)00100-3}, year = {1997}, date = {1997-01-01}, booktitle = {DNA and Protein Sequence Analysis: A Practical Approach}, volume = {14}, pages = {255-278}, publisher = {Oxford University Press}, series = {Practical Approach Series)}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Solution structure of SECIS, the mRNA element required for eukaryotic selenocysteine insertion--interaction studies with the SECIS-binding protein SBP}, author = {R Walczak and N Hubert and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9315308}, isbn = {9315308}, year = {1997}, date = {1997-01-01}, journal = {Biomed Environ Sci}, volume = {10}, number = {2-3}, pages = {177-181}, abstract = {Selenocysteine, a selenium-containing analog of cysteine, is found in the prokaryotic and eukaryotic kingdoms in active sites of enzymes involved in oxidation-reduction reactions. This aminoacid is cotranslationally incorporated at UGA codons which usually act as translation stop codons. In eukaryotes, decoding of selenocysteine necessitates the participation of the selenocysteine insertion sequence (SECIS), an element lying in the 3'-untranslated region of selenoprotein mRNAs. A detailed experimental study of the secondary structures of the SECIS elements of rat and human type 1 iodothyronine deiodinases and rat glutathione peroxidase was performed. Enzymatic and chemical structure probing led us to propose a secondary structure model, supported by sequence comparison of 23 SECIS mRNAs. The secondary structure model revealed the existence of a novel type of RNA motif composed of four consecutive non-Watson-Crick base-pairs. Using gel shift experiments, we identified in several mammalian cell type extracts the protein SBP, for SECIS-binding protein, that specifically recognizes the iodothyronine deiodinases and glutathione peroxidase SECIS elements. The structural model that we derived for the SECIS RNAs discloses RNA features possibly implicated in the binding of SBP and/or SECIS function.}, note = {0895-3988 Journal Article Review Review, Tutorial}, keywords = {Animals Human Magnetic Resonance Spectroscopy Models, Chemical Nucleic Acid Conformation RNA, Messenger/*chemistry Rats Selenocysteine/*chemistry Solutions Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence-specific cleavage of yeast tRNA(Phe) with oligonucleotides conjugated to a diimidazole construct}, author = {V Vlassov and T Abramova and T Godovikova and R Giege and V Silnikov}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9055037}, isbn = {9055037}, year = {1997}, date = {1997-01-01}, journal = {Antisense Nucleic Acid Drug Dev}, volume = {7}, number = {1}, pages = {39-42}, abstract = {Oligonucleotide derivatives conjugated to a chemical construction with two histamine residues imitating the catalytic center of ribonuclease A have been synthesized. In experiments with the conjugates complementary to the 3'-end and to the variable loop and the T loop of yeast tRNA(Phe), it was shown that the compounds can accomplish sequence-specific cleavage of the target RNA in physiologic conditions.}, note = {1087-2906 Journal Article}, keywords = {Antisense/chemical synthesis/metabolism/*pharmacology RNA, Base Sequence Imidazoles/*chemistry Molecular Sequence Data Oligonucleotides, Fungal/*drug effects RNA, Non-U.S. Gov't Yeasts/genetics, Pancreatic/pharmacology Support, Phe/chemistry/*drug effects/*metabolism Ribonuclease, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA-hydrolyzing antibodies from peripheral blood of patients with lupus erythematosus}, author = {A V Vlassov and O A Andrievskaya and T G Kanyshkova and A G Baranovsky and V A Naumov and A A Breusov and R Giege and V N Buneva and G A Nevinsky}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9275287}, isbn = {9275287}, year = {1997}, date = {1997-01-01}, journal = {Biochemistry (Mosc)}, volume = {62}, number = {5}, pages = {474-479}, abstract = {Experiments and hydrolysis of substrates with known spatial structures (such as yeast tRNAPhe, as well as normal and mutant tRNALys from human mitochondria produced by transcription of the appropriate DNA species, that is, RNA genes) were performed to study the ribonuclease activity of antibodies isolated from blood sera of patients with systemic lupus erythematosus (SLE). The antibody preparations contained two types of ribonuclease activities: the first corresponded to the specificity of ribonuclease A and was found during hydrolysis at low salt concentrations, whereas the second was stimulated by Mg2+ and displayed unique specificity toward double-stranded regions of the substrate. The possible use of the antibody preparations as tools for structural studies of conformational differences between RNA molecules was examined. In experiments with unmodified and mutant tRNALys species differing in one base found in the T-loop, we found that hydrolysis with SLE antibodies can detect small local structural changes in RNA under physiological conditions.}, note = {0006-2979 Journal Article}, keywords = {Antibodies, Catalytic/*blood Human Hydrolysis Lupus Erythematosus, Lys/chemistry/*metabolism, Systemic/blood/*immunology Nucleic Acid Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization and preliminary crystallographic analysis of ribosomal protein S8 from Thermus thermophilus}, author = {S V Tishchenko and V S Vysotskaya and N P Fomenkova and S V Nikonov and B Ehresmann and M B Garber}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9061793}, isbn = {9061793}, year = {1997}, date = {1997-01-01}, journal = {Proteins}, volume = {27}, number = {2}, pages = {309-310}, abstract = {Crystals have been obtained for recombinant ribosomal protein S8 from Thermus thermophilus produced by Escherichia coli. The protein crystals have been grown in 40 mM potassium phosphate buffer (pH 6.0) in hanging drops equilibrated against saturated ammonium sulfate (unbuffered) with 2-methyl-2,4-pentandiol (v/v). The crystals belong to the space group P4(1(3)) 2(1)2 with cell parameters a = b = 67.65 A}, note = {0887-3585 Journal Article}, keywords = {Bacterial Proteins/*chemistry Crystallization Crystallography, Non-U.S. Gov't Thermus thermophilus/*chemistry, Unité ARN, X-Ray Recombinant Fusion Proteins/chemistry Ribosomal Proteins/*chemistry Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mirror image alternative interaction patterns of the same tRNA with either class I arginyl-tRNA synthetase or class II aspartyl-tRNA synthetase}, author = {M Sissler and G Eriani and F Martin and R Giege and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9396794}, isbn = {9396794}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {24}, pages = {4899-4906}, abstract = {Gene cloning, overproduction and an efficient purification protocol of yeast arginyl-tRNA synthetase (ArgRS) as well as the interaction patterns of this protein with cognate tRNAArgand non-cognate tRNAAspare described. This work was motivated by the fact that the in vitro transcript of tRNAAspis of dual aminoacylation specificity and is not only aspartylated but also efficiently arginylated. The crystal structure of the complex between class II aspartyl-tRNA synthetase (AspRS) and tRNAAsp, as well as early biochemical data, have shown that tRNAAspis recognized by its variable region side. Here we show by footprinting with enzymatic and chemical probes that transcribed tRNAAspis contacted by class I ArgRS along the opposite D arm side, as is homologous tRNAArg, but with idiosyncratic interaction patterns. Besides protection, footprints also show enhanced accessibility of the tRNAs to the structural probes, indicative of conformational changes in the complexed tRNAs. These different patterns are interpreted in relation to the alternative arginine identity sets found in the anticodon loops of tRNAArgand tRNAAsp. The mirror image alternative interaction patterns of unmodified tRNAAspwith either class I ArgRS or class II AspRS, accounting for the dual identity of this tRNA, are discussed in relation to the class defining features of the synthetases. This study indicates that complex formation between unmodified tRNAAspand either ArgRS and AspRS is solely governed by the proteins.}, note = {0305-1048 Journal Article}, keywords = {Anticodon/chemistry Arginine-tRNA Ligase/classification/*metabolism Aspartate-tRNA Ligase/classification/*metabolism Base Sequence DNA Footprinting Escherichia coli Fungal Proteins/classification/*metabolism Models, Arg/chemistry/*metabolism RNA, Asp/chemistry/*metabolism Recombinant Fusion Proteins/metabolism Saccharomyces cerevisiae/metabolism Stereoisomerism Substrate Specificity Support, ERIANI, FLORENTZ, Fungal/chemistry/*metabolism RNA, Molecular Molecular Sequence Data *Nucleic Acid Conformation Protein Binding RNA, Non-U.S. Gov't, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ribosomal protein S15 from Thermus thermophilus--cloning, sequencing, overexpression of the gene and RNA-binding properties of the protein}, author = {A Serganov and A Rak and M Garber and J Reinbolt and B Ehresmann and C Ehresmann and M Grunberg-Manago and C Portier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9208917}, isbn = {9208917}, year = {1997}, date = {1997-01-01}, journal = {Eur J Biochem}, volume = {246}, number = {2}, pages = {291-300}, abstract = {A 6-kb DNA fragment from an extreme thermophile, Thermus thermophilus, carrying the genes for cytochrome oxidase ba3 subunit I (cbaA) and the ribosomal protein S15 (rpsO) was cloned into Escherichia coli. The gene rpsO was sequenced. The deduced amino acid sequence exhibits 59% identity to the corresponding protein from E. coli. Expression of rpsO in E. coli requires the use of a fully repressed inducible promoter because S15 from T. thermophilus is toxic for E. coli cells. When purified without denaturation from either overproducing E. coli strain or from T. thermophilus ribosomes, the S15 protein is stable and binds a cloned T. thermophilus 16S rRNA fragment (nucleotides 559-753), with low identical dissociation constants (2.5 nM), thus demonstrating that the thermophilic protein folds correctly in a mesophilic bacterium. The rRNA fragment bound corresponds in position and structure to the 16S rRNA fragment of E. coli. A similar high affinity was also found for the binding of S15 from T. thermophilus or E. coli to the corresponding E. coli 16S rRNA fragment, whereas a slightly lower affinity was observed in binding experiments between E. coli S15 and T. thermophilus 16S rRNA fragment. These results suggest that S15 from T. thermophilus recognizes similar determinants in both rRNA fragments. Competition experiments support this conclusion.}, note = {0014-2956 Journal Article}, keywords = {16S/metabolism RNA-Binding Proteins/*genetics/metabolism Ribosomal Proteins/*genetics/metabolism Sequence Homology, Amino Acid Support, Amino Acid Sequence Base Sequence Cloning, Bacterial Escherichia coli/genetics Molecular Sequence Data Plasmids RNA, Molecular DNA, Non-U.S. Gov't Thermus thermophilus/*genetics, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The modified wobble base inosine in yeast tRNAIle is a positive determinant for aminoacylation by isoleucyl-tRNA synthetase}, author = {B Senger and S Auxilien and U Englisch and F Cramer and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9204872}, isbn = {9204872}, year = {1997}, date = {1997-01-01}, journal = {Biochemistry}, volume = {36}, number = {27}, pages = {8269-8275}, abstract = {Earlier work by two independent groups has established the fact that anticodons GAU and LAU of Escherichia coli tRNAIle isoacceptors play a critical role in the tRNA identity. Yeast possesses two isoleucine transfer RNAs, a major one with anticodon IAU and a minor one with anticodon PsiAPsi which are derived from the post-transcriptional modification of AAU and UAU gene sequences, respectively. We present direct evidence which reveals that inosine is a positive determinant for yeast isoleucyl-tRNA synthetase. We also show that yeast tRNAMet with guanosine at the wobble position becomes aminoacylated with isoleucine while methionine acceptance is lost. As inosine and guanosine share the 6-keto and the N-1 hydrogen groups, this suggests that these hydrogen donor and acceptor groups are determinants for isoleucine specificity. The role of the minor tRNAIle anticodon pseudouridines in tRNA isoleucylation could not be tested directly but was deduced from a 40-fold decrease in the activity of the unmodified transcript. The presence of the NHCO structure in guanosine, inosine, pseudouridine, and lysidine suggests a unifying model of wobble base recognition by the yeast and E. coli isoleucyl-tRNA synthetase. In contrast to lysidine which switches the identity of the tRNA from methionine to isoleucine [Muramatsu, T., Nishikawa, K., Nemoto, F., Kuchino, Y., Nishimura, S., Miyazawa, T., & Yokoyama, S. (1988) Nature 336, 179-181], pseudouridine-34 does not modify the specificity of the yeast minor tRNAIle since U-34 is a strong negative determinant for yeast MetRS. Therefore, the major role of Psi-34 (in combination with Psi-36 or not) is likely in isoleucine AUA codon specificity and translational fidelity.}, note = {0006-2960 Journal Article}, keywords = {Acylation Anticodon Base Sequence Escherichia coli/genetics Inosine/*chemistry/metabolism Isoleucine-tRNA Ligase/*metabolism Molecular Sequence Data Nucleic Acid Conformation Pseudouridine/chemistry/metabolism RNA, Bacterial/chemistry/metabolism RNA, Fungal/chemistry/metabolism RNA, Ile/*chemistry/metabolism Saccharomyces cerevisiae/*genetics Structure-Activity Relationship Substrate Specificity Support, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Probing the structure of the regulatory region of human transferrin receptor messenger RNA and its interaction with iron regulatory protein-1}, author = {J Schlegl and V Gegout and B Schlager and M W Hentze and E Westhof and C Ehresmann and B Ehresmann and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9326491}, isbn = {9326491}, year = {1997}, date = {1997-01-01}, journal = {RNA}, volume = {3}, number = {10}, pages = {1159-1172}, abstract = {A portion of the 3'UTR of the human transferrin receptor mRNA mediates iron-dependent regulation of mRNA stability. The minimal RNA regulatory region contains three conserved hairpins, so-called iron responsive elements (IREs), that are recognized specifically by iron regulatory proteins (IRPs). The structure of this regulatory region and its complex with IRP-1 was probed using a combination of enzymes and chemicals. The data support the existence of an intrinsic IRE loop structure that is constrained by an internal C-G base pair. This particular structure is one of the determinants required for optimal IRP binding. IRP-1 covers one helical turn of the IRE and protects conserved residues in each of the three IREs: the bulged cytosine and nucleotides in the hairpin loops. Two essential IRP-phosphate contacts were identified by ethylation interference. Three-dimensional modeling of one IRE reveals that IRP-1 contacts several bases and the ribose-phosphate backbone located on one face in the deep groove, but contacts also exist with the shallow groove. A conformational change of the IRE loop mediated by IRP-1 binding was visualized by Pb2+-catalyzed hydrolysis. This effect is dependent on the loop structure and on the nature of the closing base pair. Within the regulatory region of transferrin receptor mRNA, IRP-1 induces reactivity changes in a U-rich hairpin loop that requires the presence of the stem-loop structure located just downstream the endonucleolytic cleavage site identified by Binder et al. (Binder R et al. 1994, EMBO J 13:1969-1980). These results provide indications of the mechanism by which IRP-1 stabilizes the transferrin receptor mRNA under iron depletion conditions.}, note = {1355-8382 Journal Article}, keywords = {Base Composition Base Sequence Binding Sites Electrophoresis, Messenger/*chemistry/metabolism RNA-Binding Proteins/*metabolism Receptors, Molecular Molecular Sequence Data Mutation *Nucleic Acid Conformation RNA, Non-U.S. Gov't, Polyacrylamide Gel Ethylnitrosourea/pharmacology Human Hydrolysis Hydroxyl Radical/metabolism Iron/metabolism Iron Regulatory Protein 1 Iron-Regulatory Proteins Iron-Sulfur Proteins/*metabolism Lead/pharmacology Models, ROMBY, Transferrin/*genetics Ribonuclease T1/metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Staf, a promiscuous activator for enhanced transcription by RNA polymerases II and III}, author = {M Schaub and E Myslinski and C Schuster and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9009278}, isbn = {9009278}, year = {1997}, date = {1997-01-01}, journal = {EMBO J}, volume = {16}, number = {1}, pages = {173-181}, abstract = {Staf is a zinc finger protein that we recently identified as the transcriptional activator of the RNA polymerase III-transcribed selenocysteine tRNA gene. In this work we demonstrate that enhanced transcription of the majority of vertebrate snRNA and snRNA-type genes, transcribed by RNA polymerases II and III, also requires Staf. DNA binding assays and microinjection of mutant genes into Xenopus oocytes showed the presence of Staf-responsive elements in the genes for human U4C, U6, Y4 and 7SK, Xenopus U1b1, U2, U5 and MRP and mouse U6 RNAs. Using recombinant Staf, we established that it mediates the activating properties of Staf-responsive elements on RNA polymerase II and III snRNA promoters in vivo. Lastly a 19 bp consensus sequence for the Staf binding site, YY(A/T)CCC(A/G)N(A/C)AT(G/C)C(A/C)YY-RCR, was derived by binding site selection. It enabled us to identify 23 other snRNA and snRNA-type genes carrying potential Staf binding sites. Altogether, our results emphasize the prime importance of Staf as a novel activator for enhanced transcription of snRNA and snRNA-type genes.}, note = {0261-4189 Journal Article}, keywords = {Animals Binding Sites Consensus Sequence DNA/metabolism DNA-Binding Proteins/*metabolism Human Mice Molecular Sequence Data Oocytes Promoter Regions (Genetics) RNA Polymerase I/*metabolism RNA Polymerase II/*metabolism RNA, Genetic Xenopus *Zinc Fingers, Non-U.S. Gov't Trans-Activation (Genetics) Trans-Activators/*metabolism *Transcription, Small Nuclear/*genetics/metabolism Recombinant Proteins/metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Strategy for RNA recognition by yeast histidyl-tRNA synthetase}, author = {J Rudinger and B Felden and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9222493}, isbn = {9222493}, year = {1997}, date = {1997-01-01}, journal = {Bioorg Med Chem}, volume = {5}, number = {6}, pages = {1001-1009}, abstract = {Histidine aminoacylation systems are of interest because of the structural diversity of the RNA substrates recognized by histidyl-tRNA synthetases. Among tRNAs participating in protein synthesis, those specific for histidine all share an additional residue at their 5'-extremities. On the other hand, tRNA-like domains at the 3'--termini of some plant viruses can also be charged by histidyl-tRNA synthetases, although they are not actors in protein synthesis. This is the case for the RNAs from tobacco mosaic virus and its satellite virus but also those of turnip yellow and brome mosaic viruses. All these RNAs have intricate foldings at their 3'-termini differing from that of canonical tRNAs and share a pseudoknotted domain which is the prerequisite for their folding into structures mimicking the overall L-shape of tRNAs. This paper gives an overview on tRNA identity and rationalizes the apparently contradictory structural and aminoacylation features of histidine-specific tRNAs and tRNA-like structures. The discussion mainly relies on histidylation data obtained with the yeast synthetase, but the conclusions are of a more universal nature. In canonical tRNA(His), the major histidine identity element is the 'minus' 1 residue, since its removal impairs histidylation and conversely its addition to a non-cognate tRNA(Asp) confers histidine identity to the transplanted molecule. Optimal expression of histidine identity depends on the chemical nature of the -1 residue and is further increased and/or modulated by the discriminator base N73 and by residues in the anticodon. In the viral tRNA-like domains, the major identity determinant -1 is mimicked by a residue from the single-stranded L1 regions of the different pseudoknots. The consequences of this mimicry for the function of minimalist RNAs derived from tRNA-like domains are discussed. The characteristics of the histidine systems illustrate well the view that the core of the amino acid accepting RNAs is a scaffold that allows proper presentation of identity nucleotides to their amino acid identity counterparts in the synthetase and that different types of scaffoldings are possible.}, note = {0968-0896 Journal Article Review Review, Tutorial}, keywords = {Amino Acyl/metabolism RNA, Base Sequence Histidine-tRNA Ligase/*metabolism Molecular Sequence Data RNA, FLORENTZ, Fungal/*metabolism RNA, Non-U.S. Gov't, Transfer, Unité ARN, Viral/metabolism Saccharomyces cerevisiae/*enzymology Substrate Specificity Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallogenesis studies in microgravity with the Advanced Protein Crystallization Facility on SpaceHab-0.}, author = {M Ries-Kautt and I Broutin and A Ducruix and W Shepard and R Kahn and N Chayen and D Blow and K Paal and W Littke and B Lorber and A Théobald-Dietrich and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S0022024897002807}, isbn = {10.1016/S0022-0248(97)00280-7}, year = {1997}, date = {1997-01-01}, volume = {181}, number = {1-2}, pages = {79–96}, abstract = {The Advanced Protein Crystallization Facility (APCF), a new protein crystallization device developed by ESA for the IML-2 Mission in 1994, was tested in its maiden flight on STS-57 Mission in SpaceHab-01 with a physico-chemical experiment on lysozyme crystallization. In pre-flight ground experiments, prior to the Shuttle Mission, the protocol for lysozyme crystallization with NaCl was based on its solubility diagram at 18°C and pH 4.5. Crystallization was conducted under microgravity in 25 APCF reactors using vapor diffusion, dialysis, and free liquid interface diffusion, with control on earth in 25 identical reactors. Identical supersaturation values were tested by the three crystallization techniques. Values of supersaturation derived from ground experiments allowed for conditions that yielded crystals in microgravity. The average number and size of crystals from the flight experiment and the earth control showed no significant difference; however many crystals were not free floating and grew on the walls of some of the protein chambers. The dialysis technique proved to be suitable, since no additional nucleation was generated by the membrane. Protein concentration measurements indicated that 13 days after activation of the experiment as much as 70–90% of the protein in supersaturated state had already crystallized. Data indicated differences in the crystallization behavior depending upon the crystallization set-up. Images of the protein chamber of 6 reactors, recorded during the flight, allowed us to evaluate the early stage of crystallization, to verify that recovered crystals had actually grown under microgravity conditions, and showed motions of crystals during the Mission. Using synchrotron radiation, resolution and rocking curve measurements of ground and space lysozyme crystals grown in APCF reactors showed no significant differences, although the values are much better than previously recorded diffraction limits and mosaicity data obtained with tetragonal lysozyme crystals grown in other set-ups and under different conditions. All controls foreseen throughout the microgravity experiment proved to be essential for the interpretation of the flight data, as concerning the effect of microgravity.}, keywords = {* Microgravity * Protein * Lysozyme * Solubility * Nucleation * Crystallization * Crystallography * Mosaicity, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interaction of tRNA with tRNA (guanosine-1)methyltransferase: binding specificity determinants involve the dinucleotide G36pG37 and tertiary structure}, author = {M Redlak and C Andraos-Selim and R Giege and C Florentz and W M Holmes}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9220956}, isbn = {9220956}, year = {1997}, date = {1997-01-01}, journal = {Biochemistry}, volume = {36}, number = {29}, pages = {8699-8709}, abstract = {The sequence G37pG36 is present in all tRNA species recognized and methylated by the Escherichia coli modification enzyme tRNA (guanosine-1)methyltransferase. We have examined whether this dinucleotide sequence provides the base specific recognition signal for this enzyme and have assessed the role of the remaining tRNA in recognition. E. coli tRNAHis and yeast tRNAAsp were substituted with G at positions 36 and 37 and were found to be excellent substrates for methylation. This suggested that the general tRNA structure can be specifically bound by the enzyme. In addition, heterologous tRNA species including fully modified tRNA1Leu are excellent inhibitors of tRNA1Leu transcript methylation. Analyses of structural variants of yeast tRNAAsp and E. coli tRNA1Leu demonstrate clearly that the core tertiary structures of tRNA are required for recognition and that G37 must be in the correct position in space relative to important contacts elsewhere in the molecule. This latter conclusion was reached because the addition of one to three stacked base pairs in the anticodon stem of tRNA1Leu dramatically alters activity. In this case, the G37 base is rotated away from the correct position in space relative to other tRNA contact sites. The acceptor stem structure is required for optimal activity since deletion of three or five base pairs is detrimental to activity; however, specific base sequence may not be important because (i) the addition of three stacked base pairs of different sequence had little effect on activity and (ii) heterologous tRNAs with little or no sequence homology in the acceptor stem are excellent substrates. Both poly G and GpG are potent and specific inhibitors of enzyme activity and are minimal substrates which can be methylated, forming m1G. Taken together, these studies suggest that 1MGT can bind the general tRNA structure and that the crucial base-pair contacts are G37 and G36.}, note = {0006-2960 Journal Article}, keywords = {Anticodon/metabolism Base Sequence Binding Sites Dinucleoside Phosphates/*metabolism Escherichia coli Kinetics Molecular Sequence Data *Nucleic Acid Conformation Poly G/*metabolism RNA, FLORENTZ, Non-P.H.S. tRNA Methyltransferases/isolation & purification/*metabolism, Non-U.S. Gov't Support, Transfer/chemistry/*metabolism Substrate Specificity Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Treatment of counterions in computer simulations of DNA.}, author = {G Ravishanker and P Auffinger and D R Langley and B Jayaram and M A Young and D L Beveridge}, editor = {K B Lipkowitz and D B Boyd}, url = {http://onlinelibrary.wiley.com/doi/10.1002/9780470125885.ch6/summary?globalMessage=0}, doi = {10.1002/9780470125885.ch6}, year = {1997}, date = {1997-01-01}, booktitle = {Reviews of Computational Chemistry}, volume = {11}, pages = {317-372}, publisher = {Wiley-VCH}, keywords = {* counterion treatment * computer simulations * simulation protocols * atomistic computer simulations * nonbonded interactions, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Rapid selection of aminoacyl-tRNAs based on biotinylation of alpha-NH2 group of charged amino acids}, author = {J Putz and J Wientges and M Sissler and R Giege and C Florentz and A Schwienhorst}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9162902}, isbn = {9162902}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {9}, pages = {1862-1863}, abstract = {A rapid selection procedure to separate low amounts of aminoacylated tRNAs from large pools of inactive variants is described. The procedure involves a three-step protocol. After initial aminoacylation of a tRNA pool, N-hydroxysuccinimide ester chemistry is applied to biotinylate the alpha-NH2 group of the amino acid bound to the 3'-end of a tRNA. The biotin tag is used to capture the derivatized tRNAs on streptavidin-conjugated magnetic beads. Variants bound to the solid phase can be amplified by RT-PCR and transcription, providing tRNAs for subsequent selection rounds.}, note = {0305-1048 Journal Article}, keywords = {Amino Acids/*chemistry Biotin/*chemistry Polymerase Chain Reaction RNA, Amino Acyl/chemistry/*isolation & purification Support, FLORENTZ, Non-U.S. Gov't, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Non-canonical interactions in a kissing loop complex: the dimerization initiation site of HIV-1 genomic RNA}, author = {J C Paillart and E Westhof and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9231899}, isbn = {9231899}, year = {1997}, date = {1997-01-01}, journal = {J Mol Biol}, volume = {270}, number = {1}, pages = {36-49}, abstract = {Retroviruses encapsidate two molecules of genomic RNA that are noncovalently linked close to their 5' ends in a region called the dimer linkage structure (DLS). The dimerization initiation site (DIS) of human immunodeficiency virus type 1 (HIV-1) constitutes the essential part of the DLS in vitro and is crucial for efficient HIV-1 replication in cell culture. We previously identified the DIS as a hairpin structure, located upstream of the major splice donor site, that contains in the loop a six-nucleotide self-complementary sequence preceded and followed by two and one purines, respectively. Two RNA monomers form a kissing loop complex via intermolecular interactions of the six nucleotide self-complementary sequence. Here, we introduced compensatory mutations in the self-complementary sequence and/or a mutation in the flanking purines. We determined the kinetics of dimerization, the thermal stabilities and the apparent equilibrium dissociation constants of wild-type and mutant dimers and used chemical probing to obtain structural information. Our results demonstrate the importance of the 5'-flanking purine and of the two central bases of the self-complementary sequence in the dimerization process. The experimental data are rationalized by triple interactions between these residues in the deep groove of the kissing helix and are incorporated into a three-dimensional model of the kissing loop dimer. In addition, chemical probing and molecular modeling favor the existence of a non-canonical interaction between the conserved adenine residues at the first and last positions in the DIS loop. Furthermore, we show that destabilization of the kissing loop complex at the DIS can be compensated by interactions involving sequences located downstream of the splice donor site of the HIV-1 genomic RNA.}, note = {0022-2836 Journal Article}, keywords = {Dimerization HIV-1/*genetics Kinetics Models, MARQUET, Molecular Mutagenesis, Non-U.S. Gov't, PAILLART, Site-Directed Nucleic Acid Conformation Purines/chemistry RNA, Unité ARN, Viral/*chemistry/genetics/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comparative analysis of thaumatin crystals grown on earth and in microgravity}, author = {J D Ng and B Lorber and R Giege and S Koszelak and J Day and A Greenwood and A McPherson}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11540583}, isbn = {11540583}, year = {1997}, date = {1997-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {53}, number = {Pt 6}, pages = {724-733}, abstract = {The protein thaumatin was studied as a model macro-molecule for crystallization in microgravity-environment experiments conducted on two US Space Shuttle missions (USML-2 and LMS). In this investigation, we have evaluated and compared the quality of space- and earth-grown thaumatin crystals using X-ray diffraction analyses, and characterized them according to crystal size, diffraction resolution limit and mosaicity. Two different approaches for growing thaumatin crystals in the microgravity environment, dialysis and liquid-liquid diffusion, were employed as a joint experiment by our two investigative teams. Thaumatin crystals grown in a microgravity environment were generally larger in volume and the total number of crystals was less, relative to crystals grown on earth. They diffracted to significantly higher resolution and with improved diffraction properties, as judged by relative plots of I/sigma versus resolution. The mosaicity of space-grown crystals was significantly less than that of crystals grown on earth. Increased concentrations of protein in the crystallization chambers in microgravity led to larger crystals. The data presented here lend further support to the idea that protein crystals of improved quality can be obtained in a microgravity environment.}, note = {0907-4449 Journal Article}, keywords = {Comparative Study Crystallization Crystallography/instrumentation/methods Crystallography, Non-P.H.S. Sweetening Agents/*chemistry *Weightlessness, Non-U.S. Gov't Support, U.S. Gov't, Unité ARN, X-Ray Particle Size Plant Proteins/*chemistry Space Flight/*instrumentation Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Visualization of RNA crystal growth by atomic force microscopy}, author = {J D Ng and Y G Kuznetsov and A J Malkin and G Keith and R Giege and A McPherson}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9185567}, isbn = {9185567}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {13}, pages = {2582-2588}, abstract = {The crystallization of transfer RNA (tRNA) was investigated using atomic force microscopy (AFM) over the temperature range from 4 to 16 degrees C, and this produced the first in situ AFM images of developing nucleic acid crystals. The growth of the (110) face of hexagonal yeast tRNAPhe crystals was observed to occur at steps on vicinal hillocks generated by multiple screw dislocation sources in the temperature range of 13.5-16 degrees C. Two-dimensional nucleation begins to dominate at 13.5 degrees C, with the appearance of three-dimensional nuclei at 12 degrees C. The changes in growth mechanisms are correlated with variations in supersaturation which is higher in the low temperature range. Growth of tRNA crystals was characterized by a strong anisotropy in the tangential step movement and transformation of growth modes on single crystals were directly observed by AFM over the narrow temperature range utilized. Finally, lattice resolution images of the molecular structure of surface layers were recorded. The implications of the strong temperature dependence of tRNAPhe crystal growth are discussed in view of improving and better controlling crystallization of nucleic acids.}, note = {0305-1048 Journal Article}, keywords = {Atomic Force RNA, Crystallization *Microscopy, Fungal/*chemistry RNA, Non-U.S. Gov't Temperature, Phe/*chemistry Saccharomyces cerevisiae/genetics Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Conformational analysis of Escherichia coli 30S ribosomes containing the single-base mutations G530U, U1498G, G1401C, and C1501G and the double-base mutation G1401C/C1501G}, author = {H Moine and K Nurse and B Ehresmann and C Ehresmann and J Ofengand}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9354641}, isbn = {9354641}, year = {1997}, date = {1997-01-01}, journal = {Biochemistry}, volume = {36}, number = {44}, pages = {13700-13709}, abstract = {Biochemical and genetic studies have pointed out the importance of several sites in 16S ribosomal RNA of Escherichia coli in the decoding process. These sites consist of the core of the decoding center (1400/1500 region) and two other segments (530 and 1050/1200 regions). To detect a possible structural link between these functionally related regions, we analyzed their sensitivity to conformational changes induced by mutations which are located in each of these regions and are known to affect the decoding process. The conformations of five segments of 16S rRNA (1-106, 406-569, 780-978, 997-1247, and 1334-1519) were analyzed by chemical probing of 30S ribosomes containing the following mutations: G530U, U1498G, G1401C, C1501G, and G1401C/C1501G. Ribosomes reconstituted with natural wild-type 16S RNA showed only minor conformational differences with respect to ribosomes isolated from cells. When 16S RNA made in vitro replaced natural 16S RNA, a slightly looser conformation of the central core region was found. Mutant ribosomes made by reconstitution with mutant 16S RNA made in vitro showed conformational effects which were in all cases localized to the region of secondary structure surrounding the site of mutation. Although the core of the decoding center (1400/1500 region) and the two other sites (530 and 1050/1200 regions) participating in the decoding function have been functionally linked, our data indicate that they are structurally independent. They also provide evidence for an unusual structure of the 1400/1500 decoding center, possibly involving noncanonical interactions. Furthermore, the absence of any conformational effect induced by the G530U mutation except at the site of mutation itself points to its direct, as opposed to indirect, involvement in the decoding function of the ribosome.}, note = {0006-2960 Journal Article}, keywords = {16S/chemical synthesis/*chemistry/genetics Ribosomes/chemistry/genetics Structure-Activity Relationship Support, Bacterial/*chemistry/genetics RNA, Base Sequence Cytosine Nucleotides/genetics Deoxyuridine Escherichia coli/genetics Guanine Nucleotides/genetics Molecular Sequence Data *Mutagenesis, Non-U.S. Gov't, Ribosomal, Site-Directed *Nucleic Acid Conformation RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The RNA binding site of S8 ribosomal protein of Escherichia coli: Selex and hydroxyl radical probing studies}, author = {H Moine and C Cachia and E Westhof and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9056763}, isbn = {9056763}, year = {1997}, date = {1997-01-01}, journal = {RNA}, volume = {3}, number = {3}, pages = {255-268}, abstract = {The RNA binding site of ribosomal protein S8 of Escherichia coli is confined to a small region within the stem of a hairpin in 16S rRNA (nt 588-605/633-651), and thus represents a model system for understanding RNA/protein interaction rules. The S8 binding site on 16S rRNA was suspected to contain noncanonical features difficult to prove with classical genetical or biochemical means. We performed in vitro iterative selection of RNA aptamers that bind S8. For the different aptamers, the interactions with the protein were probed with hydroxyl radicals. Aptamers that were recognized according to the same structural rules as wild-type RNA, but with variations not found in nature, were identified. These aptamers revealed features in the S8 binding site that had been concealed during previous characterizations by the high base conservation throughout evolution. Our data demonstrate that the core structure of the S8 binding site is composed of three interdependent bases (nt 597/641/643), with an essential intervening adenine nucleotide (position 642). The other elements important for the binding site are a base pair (598/640) above the three interdependent bases and a bulged base at position 595, the identity of which is not important. Possible implications on the geometry of the S8 binding site are discussed with the help of a three-dimensional model.}, note = {1355-8382 Journal Article}, keywords = {16S/chemistry/metabolism Ribosomal Proteins/chemistry/*metabolism Support, Autoradiography Base Sequence Binding Sites Consensus Sequence Escherichia coli Genetic Techniques Hydroxyl Radical/chemistry Models, Bacterial/*metabolism RNA, Molecular Molecular Sequence Data Nucleic Acid Conformation Phylogeny Polymerase Chain Reaction RNA, Non-U.S. Gov't, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @incollection{, title = {Treatment of counterions in computer simulations of DNA.}, author = {G Ravishanker and P Auffinger and D R Langley and B Jayaram and M A Young and D L Beveridge}, editor = {K B Lipkowitz and D B Boyd}, year = {1997}, date = {1997-01-01}, booktitle = {Reviews of Computational Chemistry}, volume = {11}, pages = {317-72}, publisher = {Wiley-VCH}, keywords = {* counterion treatment * computer simulations * simulation protocols * atomistic computer simulations * nonbonded interactions}, pubstate = {published}, tppubtype = {incollection} } @article{, title = {Phylogenetic evidence for a new tertiary interaction in bacterial RNase P RNAs}, author = {C Massire and L Jaeger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9174090}, isbn = {9174090}, year = {1997}, date = {1997-01-01}, journal = {RNA}, volume = {3}, number = {6}, pages = {553-556}, note = {1355-8382 Letter}, keywords = {Bacterial/*chemistry RNA, Base Sequence Comparative Study Databases, Catalytic/*chemistry Ribonuclease P Sequence Homology, Factual Endoribonucleases/*chemistry Molecular Sequence Data *Nucleic Acid Conformation Phylogeny RNA, Non-U.S. Gov't Variation (Genetics), Nucleic Acid Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Context dependent RNA-RNA recognition in a three-dimensional model of the 16S rRNA core}, author = {B Masquida and B Felden and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9222495}, isbn = {9222495}, year = {1997}, date = {1997-01-01}, journal = {Bioorg Med Chem}, volume = {5}, number = {6}, pages = {1021-1035}, abstract = {A 3-D model of the core of the 16S rRNA of Escherichia coli containing 328 residues has been built in the protein map derived from neutron scattering data with the help of all the available phylogenetic, biochemical, and cross-linking data. The three pseudoknots of the 16S-core cluster, through the arrangement of complex three-, four- and five-way junctions, around the neck and at the subunit interface. The roles in assembly, initiation or elongation of the three pseudoknots in ribosomal dynamics are emphasized. The 530-loop, localized on the periphery of the 30S particle, could be built with and without a pseudoknot independently of the state of the particle. The pseudoknot of the central domain controls the dynamics of an helix connected to the subunit interface which could trigger some mechanism during translation. The process of the model construction is compatible with a folding scenario in which the 5'-terminal pseudoknot controls the assembly of the central junction and the subsequent folding of the 3'-major domain. The modelling, together with the phylogenetic analysis and the experimental data, point to several potential RNA-RNA contacts which depend on the structural and sequence context in which they occur.}, note = {0968-0896 Journal Article}, keywords = {16S/*chemistry/*metabolism Ribosomal Proteins/chemistry Substrate Specificity Support, Base Sequence Escherichia coli/metabolism *Models, Molecular Molecular Sequence Data *Nucleic Acid Conformation Peptide Mapping RNA/*chemistry/*metabolism RNA, Non-U.S. Gov't, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Characterization of a thermosensitive Escherichia coli aspartyl-tRNA synthetase mutant}, author = {F Martin and G J Sharples and R G Lloyd and S Eiler and D Moras and J Gangloff and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9171418}, isbn = {9171418}, year = {1997}, date = {1997-01-01}, journal = {J Bacteriol}, volume = {179}, number = {11}, pages = {3691-3696}, abstract = {The Escherichia coli tls-1 strain carrying a mutated aspS gene (coding for aspartyl-tRNA synthetase), which causes a temperature-sensitive growth phenotype, was cloned by PCR, sequenced, and shown to contain a single mutation resulting in substitution by serine of the highly conserved proline 555, which is located in motif 3. When an aspS fragment spanning the codon for proline 555 was transformed into the tls-1 strain, it was shown to restore the wild-type phenotype via homologous recombination with the chromosomal tls-1 allele. The mutated AspRS purified from an overproducing strain displayed marked temperature sensitivity, with half-life values of 22 and 68 min (at 42 degrees C), respectively, for tRNA aminoacylation and ATP/PPi exchange activities. Km values for aspartic acid, ATP, and tRNA(Asp) did not significantly differ from those of the native enzyme; thus, mutation Pro555Ser lowers the stability of the functional configuration of both the acylation and the amino acid activation sites but has no significant effect on substrate binding. This decrease in stability appears to be related to a conformational change, as shown by gel filtration analysis. Structural data strongly suggest that the Pro555Ser mutation lowers the stability of the Lys556 and Thr557 positions, since these two residues, as shown by the crystallographic structure of the enzyme, are involved in the active site and in contacts with the tRNA acceptor arm, respectively.}, note = {0021-9193 Journal Article}, keywords = {Aspartate-tRNA Ligase/*genetics Escherichia coli/*genetics Models, ERIANI, Molecular Mutation Structure-Activity Relationship Support, Non-U.S. Gov't Temperature, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Antisense RNA control of plasmid R1 replication. The dominant product of the antisense rna-mrna binding is not a full RNA duplex}, author = {C Malmgren and E G Wagner and C Ehresmann and B Ehresmann and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9139701}, isbn = {9139701}, year = {1997}, date = {1997-01-01}, journal = {J Biol Chem}, volume = {272}, number = {19}, pages = {12508-12512}, abstract = {The replication frequency of plasmid R1 is controlled by an antisense RNA (CopA) that binds to its target site (CopT) in the leader region of repA mRNA and inhibits the synthesis of the replication initiator protein RepA. Previous studies on CopA-CopT pairing in vitro revealed the existence of a primary loop-loop interaction (kissing complex) that is subsequently converted to an almost irreversible duplex. However, the structure of more stable binding intermediates that lead to the formation of a complete duplex was speculative. Here, we investigated the interaction between CopA and CopT by using Pb(II)-induced cleavages. The kissing complex was studied using a truncated antisense RNA (CopI) that is unable to form a full duplex with CopT. Furthermore, RNase III, which is known to process the CopA-CopT complex in vivo, was used to detect the existence of a full duplex. Our data indicate that the formation of a full CopA-CopT duplex appears to be a very slow process in vitro. Unexpectedly, we found that the loop-loop interaction persists in the predominant CopA-CopT complex and is stabilized by intermolecular base pairing involving the 5'-proximal 30 nucleotides of CopA and the complementary region of CopT. This almost irreversible complex suffices to inhibit ribosome binding at the tap ribosome binding site and may be the inhibitory complex in vivo.}, note = {0021-9258 Journal Article}, keywords = {Antisense/*metabolism RNA, Bacterial Proteins/genetics/metabolism Base Sequence Copper/metabolism Electrophoresis, Messenger/*metabolism Ribonuclease III Support, Non-U.S. Gov't, Polyacrylamide Gel Endoribonucleases/metabolism Escherichia coli Lead Molecular Sequence Data Nucleic Acid Conformation Plasmids/*metabolism Pseudomonas RNA, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The new world of ribozymes.}, author = {L Jaeger}, url = {http://www.ncbi.nlm.nih.gov/pubmed/9204273}, isbn = {9204273}, year = {1997}, date = {1997-01-01}, journal = {Curr Opin Struct Biol}, volume = {7}, number = {3}, pages = {324-335}, abstract = {The number of RNA molecules that have novel catalytic activities has dramatically increased during the past two years. This ribozymic boom is not due to the discovery of additional examples of natural ribozymes but rather to the development of artificial ribozymes isolated by in vitro selection and evolution techniques. The structural and functional complexities of these artificial ribozymes, however, do not match those of the larger natural ribozymes. The understanding of both RNA structure and catalysis performed by natural and artificial ribozymes paves the way for the creation of RNA molecules that are able to efficiently catalyze more complex reactions.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Two step synthesis of (-) strong-stop DNA by avian and murine reverse transcriptases in vitro}, author = {C Isel and C Ehresmann and G Keith and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9016594}, isbn = {9016594}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {3}, pages = {545-552}, abstract = {Retroviral reverses transcriptases (RTs) are RNA- and DNA-dependent DNA polymerases that use a tRNA bound at the so-called primer binding site (PBS) located near the 5'end of the genomic RNA as primer. Thus, RTs must be able to accommodate both RNA and DNA in the primer strand. To test whether the natural primer confers some advantages to the priming process, we compared initiation of reverse transcription of avian and murine retroviral RNAs, using either their natural tRNA primer, tRNATrp and tRNAPro, respectively, or synthetic 18mer oligodeoxyribonucleotides (ODNs) and oligoribonucleotides (ORNs) complementary to their PBS. In both retroviral systems, the initial extension of ODNs was fast and processive. The initial extension of ORNs, tRNATrp and tRNAPro was much slower and distributive, giving rise to the transient accumulation of short pausing products. Synthesis of (-) strong-stop DNA was delayed when using ORNs and tRNAs, compared to ODNs. Even though ORNs and tRNAs were initially extended at the same rate, the short pausing products were more rapidly extended when using the tRNA primers. As a consequence, synthesis of (-) strong-stop DNA was much more efficient with tRNA primers, compared to ORNs. Taken together, these results suggest that the tRNA-primed synthesis of (-) strong-stop DNA is a two-step process, as already observed for HIV-1. The initiation mode corresponds to the initial non-processive nucleotide addition and extension of the short pausing products. It is more efficient with the natural primers than with ORNs. Initiation is followed by a more processive and unspecific elongation mode. Elongation is observed when the primer strand is DNA, i.e. when using the ODNs as primers or when the ORN and tRNA primers have been extended by a sufficient number (depending on the retroviral system) of deoxyribonucleotides.}, note = {0305-1048 Journal Article}, keywords = {Animals Cattle DNA Primers DNA, Avian/*enzymology Rna RNA, MARQUET, Murine/*enzymology Mice Myeloblastosis Virus, Non-U.S. Gov't, Pro RNA, Transfer, Trp RNA-Directed DNA Polymerase/*metabolism Support, Unité ARN, Viral/*biosynthesis Leukemia Virus}, pubstate = {published}, tppubtype = {article} } @article{, title = {Evidence for a hydroxide ion bridging two magnesium ions at the active site of the hammerhead ribozyme}, author = {T Hermann and P Auffinger and W G Scott and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9254698}, isbn = {9254698}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {17}, pages = {3421-3427}, abstract = {In the presence of magnesium ions, cleavage by the hammerhead ribozyme RNA at a specific residue leads to 2'3'-cyclic phosphate and 5'-OH extremities. In the cleavage reaction an activated ribose 2'-hydroxyl group attacks its attached 3'-phosphate. Molecular dynamics simulations of the crystal structure of the hammerhead ribozyme, obtained after flash-freezing of crystals under conditions where the ribozyme is active, provide evidence that a mu-bridging OH-ion is located between two Mg2+ions close to the cleavable phosphate. Constrained simulations show further that a flip from the C3'- endo to the C2'- endo conformation of the ribose at the cleavable phosphate brings the 2'-hydroxyl in proximity to both the attacked phosphorous atom and the mu-bridging OH-ion. Thus, the simulations lead to a detailed new insight into the mechanism of hammerhead ribozyme cleavage where a mu-hydroxo bridged magnesium cluster, located on the deep groove side, provides an OH-ion that is able to activate the 2'-hydroxyl nucleophile after a minor and localized conformational change in the RNA.}, note = {0305-1048 Journal Article}, keywords = {Binding Sites Crystallization Electrochemistry Freezing Hydroxides/*chemistry Magnesium/*chemistry Models, Catalytic/*chemistry/metabolism Support, Molecular Molecular Structure Nucleic Acid Conformation RNA, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Intron-dependent enzymatic formation of modified nucleosides in eukaryotic tRNAs: a review}, author = {H Grosjean and Z Szweykowska-Kulinska and Y Motorin and F Fasiolo and G Simos}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9258438}, isbn = {9258438}, year = {1997}, date = {1997-01-01}, journal = {Biochimie}, volume = {79}, number = {5}, pages = {293-302}, abstract = {In eukaryotic cells, especially in yeast, several genes encoding tRNAs contain introns. These are removed from pre-tRNAs during the maturation process by a tRNA-specific splicing machinery that is located within the nucleus at the nuclear envelope. Before and after the intron removal, several nucleoside modifications are added in a stepwise manner, but most of them are introduced prior to intron removal. Some of these early nucleoside modifications are catalyzed by intron-dependent enzymes while most of the others are catalyzed in an intron-independent manner. In the present paper, we review all known cases where the nucleoside modifications were shown to depend strictly on the presence of an intron. These are pseudouridines at anticodon positions 34, 35 and 36 and 5-methylcytosine at position 34 of several eukaryotic tRNAs. One common property of the corresponding intron-dependent modifying enzymes is that their activities are essentially dependent on the local specific architecture of the pre-tRNA molecule that comprises the anticodon stem and loop prolonged by the intron domain. Thus introns clearly serve as internal (cis-type) RNAs that guide nucleoside modifications by providing transient target sites in tRNA for selected nuclear modifying enzymes. This situation may be similar to the recently discovered (trans-type) snoRNA-guided process of ribose methylations of ribosomal RNAs within the nucleolus of eukaryotic cells.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Animals Base Sequence Eukaryotic Cells *Introns Molecular Sequence Data Nucleosides/*metabolism *RNA Processing, Non-U.S. Gov't tRNA Methyltransferases/*metabolism, Post-Transcriptional RNA, Transfer/genetics/*metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{mueller_polymerase_1997, title = {Polymerase chain reaction-based identification of a novel serpin from human dendritic cells}, author = {C G F Mueller and S Ho and C Massacrier and S Lebecque and Y J Liu}, year = {1997}, date = {1997-01-01}, journal = {European Journal of Immunology}, volume = {27}, pages = {3130--3134}, keywords = {Dendritic Cells, Human, Serpin}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence comparison of the Ty1 and Ty2 elements of the yeast genome supports the structural model of the tRNAiMet-Ty1 RNA reverse transcription initiation complex}, author = {S Friant and T Heyman and O Poch and M Wilhelm and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9200813}, isbn = {9200813}, year = {1997}, date = {1997-01-01}, journal = {Yeast}, volume = {13}, number = {7}, pages = {639-645}, abstract = {In the reverse transcription initiation complex of the yeast Ty1 retrotransposon, interaction between the template RNA and primer tRNAiMet is not limited to base pairing of the primer binding site (PBS) with ten nucleotides at the 3' end of tRNAiMet, but three regions named boxes O, 1 and 2.1 interact with the T and D stems and loops of tRNAiMet. Sequence comparison of 33 Ty1 elements and 13 closely related Ty2 elements found in the yeast genome shows that the nucleotide sequence of all elements is highly conserved in the region spanning the PBS and the three boxes. Since the domain of the template RNA encodes a portion of protein TyA, we have calculated its amino acid profile and its nucleotide profile to evaluate the role played by nucleotide sequence conservation in the selection for TyA function and in the maintenance of base pairing interactions for the priming function of Ty1 RNA. Our results show that the nucleotide sequence conservation of Ty1 RNA is constrained not only by selection for Ty1 function but also by maintenance of a given nucleotide sequence able to base pair with the tRNAiMet in the primer-template initiation complex.}, note = {0749-503x Journal Article}, keywords = {Amino Acid Sequence Base Sequence DNA Transposable Elements/*genetics Molecular Sequence Data Molecular Structure RNA, Fungal/genetics RNA, Met/*chemistry/*genetics Sequence Alignment *Sequence Analysis, Non-U.S. Gov't Yeasts/*genetics, RNA Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{mueller_polymerase_1997-1, title = {Polymerase chain reaction selects a novel disintegrin proteinase from CD40-activated germinal center dendritic cells}, author = {C G F Mueller and M C Rissoan and B Salinas and S Ait-Yahia and O Ravel and J M Bridon and F Briere and S Lebecque and Y J Liu}, year = {1997}, date = {1997-01-01}, journal = {Journal of Experimental Medicine}, volume = {186}, pages = {655--663}, keywords = {Dendritic Cells, Germinal Center, METALLOPROTEINASE, murine, Team-Mueller}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA-RNA interaction is required for the formation of specific bicoid mRNA 3' UTR-STAUFEN ribonucleoprotein particles}, author = {D Ferrandon and I Koch and E Westhof and C Nusslein-Volhard}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9130719}, isbn = {9130719}, year = {1997}, date = {1997-01-01}, journal = {EMBO J}, volume = {16}, number = {7}, pages = {1751-1758}, abstract = {The formation of the anterior pattern of the Drosophila embryo is dependent on the localization of the mRNA of the morphogen Bicoid (bcd) to the anterior pole of the egg cell. Staufen protein (STAU) is required in a late step of the localization to anchor the bcd mRNA in the anterior cytoplasm. We have shown previously that endogenous STAU associates specifically with injected bcd mRNA 3'-untranslated region (UTR), resulting in the formation of characteristic RNA-protein particles that are transported along microtubules of the mitotic spindles in a directed manner. The regions recognized by STAU in this in vivo assay are predicted to form three stem-loop structures involving large double-stranded stretches. Here, we show that the STAU interaction requires a double-stranded conformation of the stems within the RNA localization signal. In addition, base pairing between two single-stranded loops plays a major role in particle formation. This loop-loop interaction is intermolecular, not intramolecular; thus dimers or multimers of the RNA localization signal must be associated with STAU in these particles. The bcd mRNA 3' UTR can also dimerize in vitro in the absence of STAU. Thus, in addition to RNA-protein interactions, RNA-RNA interaction might be involved in the formation of ribonucleoprotein particles for transport and localization.}, note = {0261-4189 Journal Article}, keywords = {Animals Base Sequence DNA Primers Dimerization Drosophila/embryology/genetics/*physiology *Drosophila Proteins Embryo, Genetic, Messenger/*biosynthesis/*chemistry/metabolism RNA-Binding Proteins/*biosynthesis/*metabolism Ribonucleoproteins/*metabolism Trans-Activators/*biosynthesis Translation, Nonmammalian/*physiology Homeodomain Proteins/*biosynthesis Insect Proteins/biosynthesis Models, Structural Molecular Sequence Data *Nucleic Acid Conformation RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Implications of RNA structure on the annealing of a potent antisense RNA directed against the human immunodeficiency virus type 1}, author = {S Eckardt and P Romby and G Sczakiel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9335527}, isbn = {9335527}, year = {1997}, date = {1997-01-01}, journal = {Biochemistry}, volume = {36}, number = {42}, pages = {12711-12721}, abstract = {Antisense RNA-mediated regulation in bacterial systems is related to the kinetics of RNA-RNA annealing in vitro. Here, we investigated the secondary structure of alphaY69, an effective HIV-directed antisense RNA in human cells. Purified RNA preparations contain a single conformer. The global structure was identified by a cleavage experiment under native conditions using a short complementary oligonucleotide and RNase H. Structural analyses indicate a three-domain structure of alphaY69 consisting of two stem-loop elements connected by a seven-nucleotide single-stranded hinge region. Kinetic data suggest that the formation of base pairs between a CGC triplet of alphaY69 and its target RNA is essential for fast annealing. The complementary sequence stretch of the target folds into a high-energy secondary structure. The relationship between modifications in structural elements of alphaY69 and the annealing kinetics suggested that rate-limiting steps of the annealing involve a single site of alphaY69 and do not involve its 5' or 3'-end. Further, the data indicate that both initial base-specific interactions and duplex formation are dependent on the CGC triplet of the central region of alphaY69. This mechanism represents a specific and efficient way of RNA-RNA annealing that is initiated by the interaction of unstructured RNA regions.}, note = {0006-2960 Journal Article}, keywords = {Anti-HIV Agents/*chemistry/pharmacology Base Sequence Cells, Antisense/*chemistry/*pharmacology RNA, Calf Thymus Ribonuclease T1 Software Support, Cultured Electrophoresis, Genetic, Insertional Mutagenesis, Non-U.S. Gov't Thermodynamics Transcription, Polyacrylamide Gel HIV-1/*drug effects/genetics Human Kinetics Molecular Sequence Data Mutagenesis, ROMBY, Site-Directed *Nucleic Acid Conformation Oligodeoxyribonucleotides Oligoribonucleotides/*chemistry/pharmacology RNA, Unité ARN, Viral/*chemistry/drug effects Ribonuclease H}, pubstate = {published}, tppubtype = {article} } @article{, title = {Inter-domain cross-linking and molecular modelling of the hairpin ribozyme}, author = {D J Earnshaw and B Masquida and S Muller and S T Sigurdsson and F Eckstein and E Westhof and M J Gait}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9398527}, isbn = {9398527}, year = {1997}, date = {1997-01-01}, journal = {J Mol Biol}, volume = {274}, number = {2}, pages = {197-212}, abstract = {The hairpin ribozyme is a small catalytic RNA composed of two helical domains containing a small and a large internal loop and, thus, constitutes a valuable paradigm for the study of RNA structure and catalysis. We have carried out molecular modelling of the hairpin ribozyme to learn how the two domains (A and B) might fold and approach each other. To help distinguish alternative inter-domain orientations, we have chemically synthesized hairpin ribozymes containing 2'-2' disulphide linkages of known spacing (12 or 16 A) between defined ribose residues in the internal loop regions of each domain. The abilities of cross-linked ribozymes to carry out RNA cleavage under single turnover conditions were compared to the corresponding disulphide-reduced, untethered ribozymes. Ribozymes were classed in three categories according to whether their cleavage rates were marginally, moderately, or strongly affected by cross-linking. This rank order of activity guided the docking of the two domains in the molecular modelling process. The proposed three-dimensional model of the hairpin ribozyme incorporates three different crystallographically determined structural motifs: in domain A, the 5'-GAR-3'-motif of the hammerhead ribozyme, in domain B, the J4/5 motif of group I ribozymes, and connecting the two domains, a "ribose zipper", another group I ribozyme feature, formed between the hydroxyl groups of residues A10, G11 of domain A and C25, A24 of domain B. This latter feature might be key to the selection and precise orientation of the inter-domain docking necessary for the specific phosphodiester cleavage. The model provides an important basis for further studies of hairpin ribozyme structure and function.}, note = {0022-2836 Journal Article}, keywords = {Catalytic/*chemistry/metabolism Structure-Activity Relationship Support, Computer Simulation Cross-Linking Reagents/chemistry/metabolism Disulfides/chemistry/metabolism Kinetics Magnetic Resonance Spectroscopy Models, Molecular *Nucleic Acid Conformation Oligoribonucleotides/chemistry/isolation & purification RNA/chemistry RNA, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Isoalloxazine derivatives promote photocleavage of natural RNAs at G.U base pairs embedded within helices.}, author = {P Burgstaller and T Hermann and C Huber and E Westhof and M Famulok}, url = {http://www.ncbi.nlm.nih.gov/pubmed/9321652}, isbn = {9321652}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {20}, pages = {4018-4027}, abstract = {We have recently shown that isoalloxazine derivatives are able to photocleave RNA specifically at G.U base pairs embedded within a helical stack. The reaction involves the selective molecular recognition of G.U base pairs by the isoalloxazine ring and the removal of one nucleoside downstream of the uracil residue. Divalent metal ions are absolutely required for cleavage. Here we extend our studies to complex natural RNA molecules with known secondary and tertiary structures, such as tRNAs and a group I intron (td). G.U pairs were cleaved in accordance with the phylogenetically and experimentally derived secondary and tertiary structures. Tandem G.U pairs or certain G.U pairs located at a helix extremity were not affected. These new cleavage data, together with the RNA crystal structure, allowed us to perform molecular dynamics simulations to provide a structural basis for the observed specificity. We present a stable structural model for the ternary complex of the G. U-containing helical stack, the isoalloxazine molecule and a metal ion. This model provides significant new insight into several aspects of the cleavage phenomenon, mechanism and specificity for G. U pairs. Our study shows that in large natural RNAs a secondary structure motif made of an unusual base pair can be recognized and cleaved with high specificity by a low molecular weight molecule. This photocleavage reaction thus opens up the possibility of probing the accessibility of G.U base pairs, which are endowed with specific structural and functional roles in numerous structured and catalytic RNAs and interactions of RNA with proteins, in folded RNAs.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Hierarchy and dynamics of RNA folding}, author = {P Brion and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9241415}, isbn = {9241415}, year = {1997}, date = {1997-01-01}, journal = {Annu Rev Biophys Biomol Struct}, volume = {26}, pages = {113-137}, abstract = {The evidence showing that the self-assembly of complex RNAs occurs in discrete transitions, each relating to the folding of sub-systems of increasing size and complexity starting from a state with most of the secondary structure, is reviewed. The reciprocal influence of the concentration of magnesium ions and nucleotide mutations on tertiary structure is analyzed. Several observations demonstrate that detrimental mutations can be rescued by high magnesium concentrations, while stabilizing mutations lead to a lesser dependence on magnesium ion concentration. Recent data point to the central controlling and monitoring roles of RNA-binding proteins that can bind to the different folding stages, either before full establishment of the secondary structure or at the molten globule state before the cooperative transition to the final three-dimensional structure.}, note = {1056-8700 Journal Article Review Review, Tutorial}, keywords = {Animals Magnesium Mutation *Nucleic Acid Conformation RNA/*chemistry Support, Non-U.S. Gov't Tetrahymena, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Proline-dependent oligomerization with arm exchange}, author = {M Bergdoll and M H Remy and C Cagnon and J M Masson and P Dumas}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9083108}, isbn = {9083108}, year = {1997}, date = {1997-01-01}, journal = {Structure}, volume = {5}, number = {3}, pages = {391-401}, abstract = {BACKGROUND: Oligomerization is often necessary for protein activity or regulation and its efficiency is fundamental for the cell. The quaternary structure of a large number of oligomers consists of protomers tightly anchored to each other by exchanged arms or swapped domains. However, nothing is known about how the arms can be kept in a favourable conformation before such an oligomerization. RESULTS: Upon examination of such quaternary structures, we observe an extremely frequent occurrence of proline residues at the point where the arm leaves the protomer. Sequence alignment and site-directed mutagenesis confirm the importance of these prolines. The conservation of these residues at the hinge regions can be explained by the constraints that they impose on polypeptide conformation and dynamics: by rigidifying the mainchain, prolines favour extended conformations of arms thus favouring oligomerization, and may prevent interaction of the arms with the core of the protomer. CONCLUSIONS: Hinge prolines can be considered as 'quaternary structure helpers'. The presence of a proline should be considered when searching for a determinant of oligomerization with arm exchange and could be used to engineer synthetic oligomers or to displace a monomers to oligomers equilibrium by mutation of this proline residue.}, note = {0969-2126 Journal Article}, keywords = {*Acetyltransferases Amino Acid Sequence Animals Aspartate Aminotransferases/chemistry Bacterial Proteins/chemistry Cattle Chickens Comparative Study *Dimerization Human Mitochondria, Heart/enzymology Models, Molecular Molecular Sequence Data Mutagenesis, Pancreatic/chemistry Sequence Alignment Viral Structural Proteins/chemistry, Site-Directed Na(+)-K(+)-Exchanging ATPase/chemistry Plant Viruses/chemistry Proline/*physiology Protein Binding *Protein Conformation *Protein Folding Pyrophosphatases/chemistry Ribonuclease, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Major identity determinants for enzymatic formation of ribothymidine and pseudouridine in the T psi-loop of yeast tRNAs}, author = {H F Becker and Y Motorin and M Sissler and C Florentz and H Grosjean}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9417931}, isbn = {9417931}, year = {1997}, date = {1997-01-01}, journal = {J Mol Biol}, volume = {274}, number = {4}, pages = {505-518}, abstract = {Almost all transfer RNA molecules sequenced so far contain two universal modified nucleosides at positions 54 and 55, respectively: ribothymidine (T54) and pseudouridine (psi 55). To identify the tRNA elements recognized by tRNA:m5uridine-54 methyltransferase and tRNA:pseudouridine-55 synthase from the yeast Saccharomyces cerevisiae, a set of 43 yeast tRNA(Asp) mutants were used. Some variants contained point mutations, while the others included progressive reductions in size down to a tRNA minisubstrate consisting of the T psi-loop with only one G.C base-pair as stem (9-mer). All substrates (full-sized tRNA(Asp) and various minihelices) were produced in vitro by T7 transcription and tested using yeast extract (S100) as a source of enzymatic activities and S-adenosyl-L-methionine as a methyl donor. The results indicate that the minimal substrate for enzymatic formation of psi 55 is a stem/loop structure with only four G.C base-pairs in the stem, while a longer stem is required for efficient T54 formation. None of the conserved nucleotides (G53, C56, A58 and C61) and U54 for psi 55 or U55 for T54 formation can be replaced by any of the other three canonical nucleotides. Yeast tRNA:m5uridine-54 methyltransferase additionally requires the presence of a pyrimidine-60 in the loop. Interestingly, in a tRNA(Asp) variant in which the T psi-loop was permuted with the anticodon-loop, the new U32 and U33 residues derived from the T psi-loop were quantitatively converted to T32 and psi 33, respectively. Structural mapping of this variant with ethylnitrosourea confirmed that the intrinsic characteristic structure of the T psi-loop was conserved upon permutation and that the displaced anticodon-loop did not acquire a T psi-loop structure. These results demonstrate that a local conformation rather than the exact location of the U-U sequence within the tRNA architecture is the important identity determinant for recognition by yeast tRNA:m5uridine-54 methyltransferase and tRNA:pseudouridine-55 synthase.}, note = {0022-2836 Journal Article}, keywords = {Asp/chemistry/metabolism Saccharomyces cerevisiae/genetics Substrate Specificity Support, Base Sequence Conserved Sequence Intramolecular Lyases/metabolism Models, FLORENTZ, Fungal/*chemistry/metabolism RNA, Molecular Molecular Sequence Data Nucleic Acid Conformation Pseudouridine/chemistry/*metabolism RNA, Non-U.S. Gov't Uridine/*analogs & derivatives/chemistry/metabolism tRNA Methyltransferases/metabolism, SISSLER, Transfer, Transfer/*chemistry/*metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Existence of two distinct aspartyl-tRNA synthetases in Thermus thermophilus. Structural and biochemical properties of the two enzymes}, author = {H D Becker and J Reinbolt and R Kreutzer and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9220965}, isbn = {9220965}, year = {1997}, date = {1997-01-01}, journal = {Biochemistry}, volume = {36}, number = {29}, pages = {8785-8797}, abstract = {Two aspartyl-tRNA synthetases (AspRSs) were isolated from Thermus thermophilus HB8. Both are alpha2 dimers but differ in the length of their polypeptide chains (AspRS1, 68 kDa; and AspRS2, 51 kDa). Both chains start with Met and are deprived of common sequences to a significant extent. This rules out the possibility that AspRS2 is derived from AspRS1 by proteolysis, in agreement with specific recognition of each AspRS by the homologous antibodies. DNA probes derived from N-terminal amino acid sequences hybridize specifically to different genomic DNA fragments, revealing that the two AspRSs are encoded by distinct genes. Both enzymes are present in various strains from T. thermophilus and along the growth cycle of the bacteria, suggesting that they are constitutive. Kinetic investigations show that the two enzymes are specific for aspartic acid activation and tRNAAsp charging. tRNA aspartylation by the thermostable AspRSs is governed by thermodynamic parameters which values are similar to those measured for mesophilic aspartylation systems. Both thermophilic AspRSs are deprived of species specificity for tRNA aspartylation and exhibit N-terminal sequence signatures found in other AspRSs, suggesting that they are evolutionarily related to AspRSs from mesophilic prokaryotes and eukaryotes. Comparison of the efficiency of tRNA aspartylation by each enzyme under conditions approaching the physiological ones suggests that in vivo tRNAAsp charging is essentially ensured by AspRS1, although AspRS2 is the major species. The physiological significance of the two different AspRSs in T. thermophilus is discussed.}, note = {0006-2960 Journal Article}, keywords = {Amino Acid Sequence Aspartate-tRNA Ligase/chemistry/genetics/*isolation & purification Blotting, Amino Acyl/metabolism Sequence Alignment Thermus thermophilus/*enzymology/genetics, Transfer, Unité ARN, Western Catalysis Isoenzymes/chemistry/*isolation & purification Kinetics Molecular Sequence Data Molecular Weight RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {DNA- and RNA-hydrolyzing antibodies from the blood of patients with various forms of viral hepatitis}, author = {A G Baranovsky and V G Matushin and A V Vlassov and V G Zabara and V A Naumov and R Giege and V N Buneva and G A Nevinsky}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9481869}, isbn = {9481869}, year = {1997}, date = {1997-01-01}, journal = {Biochemistry (Mosc)}, volume = {62}, number = {12}, pages = {1358-1366}, abstract = {Antibodies (Abs) hydrolyzing proteins, DNA, and RNA are detected in the blood of patients with various autoimmune diseases. In the present work, homogeneous preparations of IgG Abs from the blood of the healthy donors as well as patients with A, B, C, and delta types of viral hepatitis, influenza, pneumonia, tuberculosis, tonsillitis, duodenal ulcer, and some types of cancer were purified. For the first time, the fraction of IgG and its Fab fragments of patients with viral hepatitis were shown to have high DNA- and RNA-hydrolyzing activity. In case of Abs from the healthy donors and patients with other diseases, high activity of Abs was not detected. The data obtained by various methods indicate that the activity of hepatitis Abs is an intrinsic property of the immunoglobulins. The relative rates of hydrolysis of cCMP, poly(U), poly(A), poly(C), and tRNA(Phe) by hepatitis Abs were compared with those of RNase A and other RNases from human blood. Significant differences in activities of Abs and nucleases in hydrolysis of model substrates were demonstrated. Thus, catalytically active Abs can appear in the blood of patients not only with autoimmune disorders, but with viral diseases as well.}, note = {0006-2979 Journal Article}, keywords = {Antibodies, Antinuclear/*blood/immunology Antibodies, Catalytic/*blood/isolation & purification Base Sequence Chromatography, Gel DNA/*metabolism Deoxyribonucleases/metabolism Electrophoresis, Human/blood/*immunology Human Hydrolysis Molecular Sequence Data Nucleic Acid Conformation RNA/chemistry/*metabolism Ribonucleases/metabolism Support, Non-U.S. Gov't, Polyacrylamide Gel Hepatitis, Unité ARN, Viral}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA hydration: three nanoseconds of multiple molecular dynamics simulations of the solvated tRNA(Asp) anticodon hairpin}, author = {P Auffinger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9199403}, isbn = {9199403}, year = {1997}, date = {1997-01-01}, journal = {J Mol Biol}, volume = {269}, number = {3}, pages = {326-341}, abstract = {The hydration of the tRNA(Asp) anticodon hairpin was investigated through the analysis of six 500 ps multiple molecular dynamics (MMD) trajectories generated by using the particle mesh Ewald method for the treatment of the long-range electrostatic interactions. Although similar in their dynamical characteristics, these six trajectories display different local hydration patterns reflecting the landscape of the "theoretical" conformational space being explored. The statistical view gained through the MMD strategy allowed us to characterize the hydration patterns around important RNA structural motifs such as a G-U base-pair, the anticodon U-turn, and two modified bases: pseudouridine and 1-methylguanine. The binding of ammonium counterions to the hairpin has also been investigated. No long-lived hydrogen bond between water and a 2'-hydroxyl has been observed. Water molecules with long-residence times are found bridging adjacent pro-Rp phosphate atoms. The conformation of the pseudouridine is stiffened by a water-mediated base-backbone interaction and the 1-methylguanine is additionally stabilized by long-lived hydration patterns. Such long-lived hydration patterns are essential to ensure the structural integrity of this hairpin motif. Consequently, our simulations confirm the conclusion reached from an analysis of X-ray crystal structures according to which water molecules form an integral part of nucleic acid structure. The fact that the same conclusion is reached from a static and a dynamic point of view suggests that RNA and water together constitute the biologically relevant functional entity.}, note = {0022-2836 Journal Article}, keywords = {Anticodon/*chemistry Base Composition *Computer Simulation Guanine/chemistry Hydrogen Bonding Models, Asp/*chemistry/metabolism Ribose/chemistry/metabolism Support, Molecular Nucleic Acid Conformation RNA, Non-U.S. Gov't Uridine/chemistry Water/*chemistry/metabolism, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Rules governing the orientation of the 2'-hydroxyl group in RNA}, author = {P Auffinger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9398515}, isbn = {9398515}, year = {1997}, date = {1997-01-01}, journal = {J Mol Biol}, volume = {274}, number = {1}, pages = {54-63}, abstract = {Molecular dynamics simulations reveal that, in C3'-endo sugar puckers, only three orientations are accessible to the 2'-hydroxyl groups distinctive of RNA molecules: towards (i) the O3', (ii) the O4' of the same sugar, and (iii) the shallow groove base atoms. In the rarer C2'-endo sugar puckers, orientations towards the O3' atom of the same sugar are strongly favoured. Surprisingly, in helical regions, the frequently suggested intra-strand O2'-H(n).O4'(n+1) interaction is not found. This observation led to the detection of an axial C-H.O interaction between the C2'-H2'(n) group and the O4'(n+1) atom contributing to the stabilization of RNA helical regions. Subsequent analysis of crystallographic structures of both RNA and A-DNA helices fully supports this finding. Specific hydration patterns are also thought to play a significant role in the stabilization of RNA structures. In the shallow groove of RNA, known as a favourable RNA or protein-binding region, three well-defined hydration sites are located around the O2' atom. These hydration sites, occupied by water molecules exchanging with the bulk, constitute, after dehydration, anchor points for specific interactions between RNA and nucleic acids, proteins or drugs. Therefore, the fact that the 2'-hydroxyl group is not monopolised by axial stabilization, together with its water-like behaviour, facilitates complex formation involving RNA helical regions.}, note = {0022-2836 Journal Article}, keywords = {Asp/*chemistry/*metabolism Ribose/chemistry/metabolism Support, Carbohydrate Conformation Carbon/chemistry/metabolism Hydrogen Bonding Hydroxyl Radical/*chemistry/metabolism Proteins/genetics/metabolism RNA, Non-U.S. Gov't Water/metabolism, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure and aminoacylation capacities of tRNA transcripts containing deoxyribonucleotides}, author = {R Aphasizhev and A Théobald-Dietrich and D Kostyuk and S N Kochetkov and L Kisselev and R Giege and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9257648}, isbn = {9257648}, year = {1997}, date = {1997-01-01}, journal = {RNA}, volume = {3}, number = {8}, pages = {893-904}, abstract = {The contribution of the ribose 2'-hydroxyls to RNA structure and function has been analyzed, but still remains controversial. In this work, we report the use of a mutant T7 RNA polymerase as a tool in RNA studies, applied to the aspartate and methionine tRNA aminoacylation systems from yeast. Our approach consists of determining the effect of substituting natural ribonucleotides by deoxyribonucleotides in RNA and, thereby, defining the subset of important 2'-hydroxyl groups. We show that deoxyribose-containing RNA can be folded in a global conformation similar to that of natural RNA. Melting curves of tRNAs, obtained by temperature-gradient gel electrophoresis, indicate that in deoxyribo-containing molecules, the thermal stability of the tertiary network drops down, whereas the stability of the secondary structure remains unaltered. Nuclease footprinting reveals a significant increase in the accessibility of both single- and double-stranded regions. As to the functionality of the deoxyribose-containing tRNAs, their in vitro aminoacylation efficiency indicates striking differential effects depending upon the nature of the substituted ribonucleotides. Strongest decrease in charging occurs for yeast initiator tRNA(Met) transcripts containing dG or dC residues and for yeast tRNA(Asp) transcripts with dU or dG. In the aspartate system, the decreased aminoacylation capacities can be correlated with the substitution of the ribose moieties of U11 and G27, disrupting two hydrogen bond contacts with the synthetase. Altogether, this suggests that specific 2'-hydroxyl groups in tRNAs can act as determinants specifying aminoacylation identity.}, note = {1355-8382 Journal Article}, keywords = {Asp/chemistry/genetics/metabolism RNA, Base Sequence DNA-Directed RNA Polymerases/genetics/metabolism Deoxyribonucleotides/chemistry/*metabolism Models, Genetic, Met/chemistry/genetics/metabolism Structure-Activity Relationship Support, Molecular Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, Non-U.S. Gov't *Transcription, Transfer, Transfer/*chemistry/genetics/*metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Importance of structural features for tRNA(Met) identity}, author = {R Aphasizhev and B Senger and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9149230}, isbn = {9149230}, year = {1997}, date = {1997-01-01}, journal = {RNA}, volume = {3}, number = {5}, pages = {489-497}, abstract = {We showed previously that the tRNA tertiary structure makes an important contribution to the identity of yeast tRNA(Met) (Senger B, Aphasizhev R, Walter P, Fasiolo F, 1995, J Mol Biol 249:45-58). To learn more about the role played by the tRNA framework, we analyzed the effect of some phosphodiester cleavages and 2'OH groups in tRNA binding and aminoacylation. The tRNA is inactivated provided the break occurs in the central core region responsible for the tertiary fold or in the anticodon stem/loop region. We also show that, for tRNA(Met) to bind, the anticodon loop, but not the anticodon stem, requires a ribosephosphate backbone. A tertiary mutant of yeast tRNA(Met) involving interactions from the D- and T-loop unique to the initiator species fails to be aminoacylated, but still binds to yeast methionyl-tRNA synthetase. In the presence of 10 mM MgCl2, the mutant transcript has a 3D fold significantly stabilized by about 30 degrees C over a wild-type transcript as deduced from the measure of their T(m) values. The k(cat) defect of the tRNA(Met) mutant may arise from a failure to overcome an increase of the free energetic cost of distorting the more stable tRNA structure and/or a tRNA based MetRS conformational change required for formation of transition state of aminoacylation.}, note = {1355-8382 Journal Article}, keywords = {Anticodon Base Sequence Electrophoresis, Genetic Variation (Genetics), Met/*biosynthesis/*chemistry/isolation & purification Saccharomyces cerevisiae/genetics/metabolism Support, Non-U.S. Gov't Transcription, Polyacrylamide Gel Kinetics Magnesium Chloride Models, Structural Molecular Sequence Data *Nucleic Acid Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence comparison of the Ty1 and Ty2 elements of the yeast genome supports the structural model of the tRNAiMet-Ty1 RNA reverse transcription initiation complex}, author = { S. Friant and T. Heyman and O. Poch and M. Wilhelm and F. X. Wilhelm}, year = {1997}, date = {1997-01-01}, journal = {Yeast}, volume = {13}, number = {7}, pages = {639-45}, abstract = {In the reverse transcription initiation complex of the yeast Ty1 retrotransposon, interaction between the template RNA and primer tRNAiMet is not limited to base pairing of the primer binding site (PBS) with ten nucleotides at the 3' end of tRNAiMet, but three regions named boxes O, 1 and 2.1 interact with the T and D stems and loops of tRNAiMet. Sequence comparison of 33 Ty1 elements and 13 closely related Ty2 elements found in the yeast genome shows that the nucleotide sequence of all elements is highly conserved in the region spanning the PBS and the three boxes. Since the domain of the template RNA encodes a portion of protein TyA, we have calculated its amino acid profile and its nucleotide profile to evaluate the role played by nucleotide sequence conservation in the selection for TyA function and in the maintenance of base pairing interactions for the priming function of Ty1 RNA. Our results show that the nucleotide sequence conservation of Ty1 RNA is constrained not only by selection for Ty1 function but also by maintenance of a given nucleotide sequence able to base pair with the tRNAiMet in the primer-template initiation complex.}, note = {0749-503x Journal Article}, keywords = {*Sequence, Acid, Alignment, Amino, Analysis, Base, Data, DNA, Elements/*genetics, Fungal/genetics, Gov't, Met/*chemistry/*genetics, Molecular, Non-U.S., RNA, Sequence, structure, Support, Transfer, Transposable, Yeasts/*genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Heterogeneous terminal structure of Ty1 and Ty3 reverse transcripts}, author = { M. Wilhelm and T. Heyman and S. Friant and F. X. Wilhelm}, year = {1997}, date = {1997-01-01}, journal = {Nucleic Acids Res}, volume = {25}, number = {11}, pages = {2161-6}, abstract = {A specific terminal structure of preintegrative DNA is required for transposition of retroviruses and LTR-retrotransposons. We have used an anchored PCR technique to map the 3'ends of DNA intermediates synthesized inside yeast Ty1 and Ty3 retrotransposon virus-like particles. We find that, unlike retroviruses, Ty1 replicated DNA does not have two extra base pairs at its 3'ends. In contrast some Ty3 preintegrative DNA molecules have two extra nucleotides at the 3'end of upstream and downstream long terminal repeats. Moreover we find that some molecules of replicated Ty3 DNA have more than two extra nucleotides at the 3'end of the upstream LTR. This observation could be accounted for by imprecise RNAse H cutting of the PPT sequence. The site of Ty1 and Ty3 plus-strand strong-stop DNA termination was also examined. Our results confirm that the prominent Ty1 and Ty3 plus-strand strong-stop molecules harbor 12 tRNA templated bases but also show that some Ty1 and Ty3 plus-strand strong-stop DNA molecules harbor less tRNA templated bases. We propose that these less than full length plus-strand molecules could be active intermediates in Ty retrotransposon replication.}, note = {0305-1048 Journal Article}, keywords = {*Nucleic, *Transcription, Acid, Calf, Chain, Conformation, DNA, Fungal/*chemistry/metabolism, Genetic, Gov't, H, Hybridization, Non-U.S., Nucleic, Plasmids/chemistry/genetics/metabolism, Polymerase, Reaction, Replication, Retroelements/*genetics, Ribonuclease, RNA, Support, Thymus/metabolism, Transfer/chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {The new world of ribozymes.}, author = {L Jaeger}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/pubmed/9204273}, isbn = {9204273}, year = {1997}, date = {1997-01-01}, journal = {Curr Opin Struct Biol}, volume = {7}, number = {3}, pages = {324-335}, abstract = {The number of RNA molecules that have novel catalytic activities has dramatically increased during the past two years. This ribozymic boom is not due to the discovery of additional examples of natural ribozymes but rather to the development of artificial ribozymes isolated by in vitro selection and evolution techniques. The structural and functional complexities of these artificial ribozymes, however, do not match those of the larger natural ribozymes. The understanding of both RNA structure and catalysis performed by natural and artificial ribozymes paves the way for the creation of RNA molecules that are able to efficiently catalyze more complex reactions.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_drosophila_1997, title = {Drosophila immunity.}, author = {Jules A Hoffmann and Jean-Marc Reichhart}, year = {1997}, date = {1997-01-01}, journal = {Trends in Cell Biology}, volume = {7}, pages = {309--316}, abstract = {Septic injury induces in Drosophila the rapid and transient transcription of several genes encoding potent antimicrobial peptides. Significant structural and functional similarities exist between the injury-induced signalling cascades leading to antimicrobial peptide gene expression in Drosophila and cytokine-induced expression of mammalian acute-phase proteins. Here, the authors discuss their understanding of these pathways and their relationships to those found in the mammalian cells. They also analyse non-self recognition and the role of blood cells in Drosophila host defence.}, keywords = {hoffmann, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{halimi_comparison_1996, title = {Comparison of two different methods using overlapping synthetic peptides for localizing linear B cell epitopes in the U1 snRNP-C autoantigen}, author = {H Halimi and H Dumortier and J P Briand and S Muller}, doi = {10.1016/s0022-1759(96)00171-8}, issn = {0022-1759}, year = {1996}, date = {1996-11-01}, journal = {Journal of Immunological Methods}, volume = {199}, number = {1}, pages = {77--85}, abstract = {We have compared the performances of two different approaches using overlapping synthetic peptides to identify the location of linear epitopes of the U1 snRNP-C autoantigen. The first method was based on the use of 15 overlapping peptides (16-30 residue-long) synthesized using conventional Fmoc chemistry, removed from the resin by a standard cleavage procedure, and tested by ELISA after direct coating to polyvinyl microtiter plates. The second approach used a commercial kit (SPOT) to synthesize 75 overlapping decapeptides on cellulose membrane which were assayed by a direct immunoenzymatic test. Both standard and SPOTscan methods were evaluated with antibodies raised in rabbits against synthetic peptides of U1C and sera from patients with autoimmune diseases. In addition to inherent problems linked to the SPOT synthesis (in particular the impossibility of checking the quality of peptides), a number of limitations in the SPOTscan method were identified (e.g. a certain lack of sensitivity and, in one case, the complete lack of peptide reactivity due to the removal of charged end groups at both extremities). However, we found no background with sera from autoimmune patients in the SPOTscan and the antigenic maps obtained using the two approaches generally agreed. This study shows that the SPOTscan approach represents a simple, relatively non expensive and rapid method for initial screening to identify candidate sequences that may be dominant linear epitopes in a protein. Subsequent analysis and controls should include the preparation of conventionally synthesized peptides for formal immunochemical investigations.}, keywords = {Amino Acid Sequence, Autoantigens, B-Lymphocytes, Dumortier, Enzyme-Linked Immunosorbent Assay, Epitope Mapping, Humans, I2CT, Lupus Erythematosus, Molecular Sequence Data, Peptides, Ribonucleoprotein, Systemic, Team-Dumortier, U1 Small Nuclear}, pubstate = {published}, tppubtype = {article} } @article{ehret-sabatier_characterization_1996, title = {Characterization of novel cysteine-rich antimicrobial peptides from scorpion blood}, author = {L Ehret-Sabatier and D Loew and M Goyffon and P Fehlbaum and Jules A Hoffmann and A van Dorsselaer and Philippe Bulet}, issn = {0021-9258}, year = {1996}, date = {1996-11-01}, journal = {J. Biol. Chem.}, volume = {271}, number = {47}, pages = {29537--29544}, abstract = {We have isolated, from the hemolymph of unchallenged scorpions of the species Androctonus australis, three distinct antimicrobial peptides, which we have fully characterized by Edman degradation, electrospray ionization mass spectrometry, and matrix-assisted laser desorption/ionization mass spectrometry. Two are novel molecules: (i) androctonin, a 25-residue peptide with two disulfide bridges, active against both bacteria (Gram-positive and Gram-negative) and fungi and showing marked sequence homology to tachyplesins and polyphemusins from horseshoe crabs; and (ii) buthinin, a 34-residue antibacterial (Gram-positive and Gram-negative) peptide with three disulfide bridges. The third peptide contains 37 residues and three disulfide bridges and clearly belongs to the family of anti-Gram-positive insect defensins. We have synthesized androctonin and explored its activity spectrum and mode of action.}, keywords = {Animals, Anti-Bacterial Agents, Chromatography, Cysteine, Electron, Hemolymph, Hemolysis, High Pressure Liquid, hoffmann, M3i, Mass Spectrometry, Microscopy, Peptides, Scorpions}, pubstate = {published}, tppubtype = {article} } @article{barillas-mury_immune_1996, title = {Immune factor Gambif1, a new rel family member from the human malaria vector, Anopheles gambiae}, author = {Carolina Barillas-Mury and A Charlesworth and I Gross and A Richman and Jules A Hoffmann and Fotis C Kafatos}, issn = {0261-4189}, year = {1996}, date = {1996-09-01}, journal = {EMBO J.}, volume = {15}, number = {17}, pages = {4691--4701}, abstract = {A novel rel family member, Gambif1 (gambiae immune factor 1), has been cloned from the human malaria vector, Anopheles gambiae, and shown to be most similar to Drosophila Dorsal and Dif. Gambif1 protein is translocated to the nucleus in fat body cells in response to bacterial challenge, although the mRNA is present at low levels at all developmental stages and is not induced by infection. DNA binding activity to the kappaB-like sites in the A.gambiae Defensin and the Drosophila Diptericin and Cecropin promoters is also induced in larval nuclear extracts following infection. Gambif1 has the ability to bind to kappaB-like sites in vitro. Co-transfection assays in Drosophila mbn-2 cells show that Gambif1 can activate transcription by interacting with the Drosophila Diptericin regulatory elements, but is not functionally equivalent to Dorsal in this assay. Gambif1 protein translocation to the nucleus and the appearance of kappaB-like DNA binding activity can serve as molecular markers of activation of the immune system and open up the possibility of studying the role of defence reactions in determining mosquito susceptibility/refractoriness to malaria infection.}, keywords = {Amino Acid, Animals, Anopheles, Base Sequence, Biological Transport, Cell Nucleus, Cells, Complementary, Cultured, DNA, DNA-Binding Proteins, hoffmann, Insect Proteins, Insect Vectors, M3i, NF-kappa B, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-rel, Sequence Homology, Trans-Activators, Transcriptional Activation}, pubstate = {published}, tppubtype = {article} } @article{charlet_innate_1996, title = {Innate immunity. Isolation of several cysteine-rich antimicrobial peptides from the blood of a mollusc, Mytilus edulis}, author = {Maurice Charlet and S Chernysh and H Philippe and Charles Hetru and Jules A Hoffmann and Philippe Bulet}, issn = {0021-9258}, year = {1996}, date = {1996-09-01}, journal = {J. Biol. Chem.}, volume = {271}, number = {36}, pages = {21808--21813}, abstract = {We have isolated from the blood of immune-challenged and untreated mussels (Mytilus edulis) antibacterial and antifungal peptides. We have characterized two isoforms of a novel 34-residue, cysteine-rich, peptide with potent bactericidal activity and partially characterized a novel 6.2-kDa antifungal peptide containing 12 cysteines. We report the presence of two members of the insect defensin family of antibacterial peptides and provide a phylogenetic analysis that indicates that mollusc and arthropod defensins have a common ancestry. Our data argue that circulating antimicrobial peptides represent an ancient host defense mechanism that predated the separation between molluscs and arthropods at the root of the Cambrian, about 545 million years ago.}, keywords = {Amino Acid, Animals, Anti-Infective Agents, Antifungal Agents, Bivalvia, Blood Proteins, Chromatography, Cysteine, Defensins, High Pressure Liquid, hoffmann, M3i, Molecular Weight, Phylogeny, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{richman_inducible_1996, title = {Inducible immune factors of the vector mosquito Anopheles gambiae: biochemical purification of a defensin antibacterial peptide and molecular cloning of preprodefensin cDNA}, author = {A M Richman and Philippe Bulet and Charles Hetru and Carolina Barillas-Mury and Jules A Hoffmann and Fotis C Kafalos}, issn = {0962-1075}, year = {1996}, date = {1996-08-01}, journal = {Insect Mol. Biol.}, volume = {5}, number = {3}, pages = {203--210}, abstract = {Larvae of the mosquito vector of human malaria, Anopheles gambiae, were inoculated with bacteria and extracts were biochemically fractionated by reverse-phase HPLC. Multiple induced polypeptides and antibacterial activities were observed following bacterial infection, including a member of the insect defensin family of antibacterial proteins. A cDNA encoding An. gambiae preprodefensin was isolated using PCR primers based on phylogenetically conserved sequences. The mature peptide is highly conserved, but the signal and propeptide segments are not, relative to corresponding defensin sequences of other insects. Defensin expression is induced in response to bacterial infection, in both adult and larval stages. In contrast, pupae express defensin mRNA constitutively. Defensin expression may prove a valuable molecular marker to monitor the An. gambiae host response to infection by parasitic protozoa of medical importance.}, keywords = {Amino Acid, Animals, Anopheles, Base Sequence, Blood Bactericidal Activity, Blood Proteins, Cloning, Complementary, Defensins, DNA, Escherichia coli, Female, Gene Expression, Genes, hoffmann, Insect, Insect Vectors, Larva, M3i, Micrococcus luteus, Molecular, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{lowenberger_aedes_1996, title = {Aedes aegypti: induced antibacterial proteins reduce the establishment and development of Brugia malayi}, author = {C A Lowenberger and M T Ferdig and Philippe Bulet and S Khalili and Jules A Hoffmann and B M Christensen}, doi = {10.1006/expr.1996.0066}, issn = {0014-4894}, year = {1996}, date = {1996-07-01}, journal = {Exp. Parasitol.}, volume = {83}, number = {2}, pages = {191--201}, abstract = {The effect of host immune activation on the development of Brugia malayi in one susceptible and four refractory strains of Aedes aegypti and in Armigeres subalbatus was assessed. A. aegypti that were immune activated by the injection of saline or bacteria 24 hr before feeding on a B. malayi-infected gerbil had significantly reduced prevalences and mean intensities of infection from those of naive controls when exposed to bloodmeals with low (105 mf/20 microliters) and medium (160 mf/20 microliters) microfilaremias. At a higher microfilaremia (237 mf/20 microliters) there were no significant differences in mean intensities, suggesting that the number of parasites ingested may affect the host's ability to mount an effective defense response. Because the major immune proteins in A. aegypti are defensins, we did Northern analyses of fat body RNA 8 hr after immune activation or bloodfeeding. All mosquitoes demonstrated rapid transcriptional activity for defensins following immune activation by intrathoracic inoculation with either saline or bacteria. However, no strain of A. aegypti, susceptible or refractory to B. malayi, nor Ar. subalbatus produced defensin transcripts after bloodfeeding on an uninfected or a B. malayi-infected gerbil. These data suggest that inducible immune proteins of mosquitoes can reduce the prevalence and mean intensity of infections with ingested parasites, but these proteins are not expressed routinely after parasite ingestion and midgut penetration and probably do not contribute to existing refractory mechanisms. Immune proteins such as defensins, however, represent potential candidates to genetically engineer mosquitoes for resistance to filarial worms.}, keywords = {Aedes, Analysis of Variance, Animals, Anti-Bacterial Agents, Base Sequence, Blood Proteins, Blotting, Brugia malayi, Culicidae, Defensins, DNA, Escherichia coli, Fat Body, Genetic, Gerbillinae, hoffmann, M3i, Micrococcus luteus, Microfilaria, Northern, RNA, Transcription}, pubstate = {published}, tppubtype = {article} } @article{gross_drosophila_1996, title = {Drosophila immunity: a comparative analysis of the Rel proteins dorsal and Dif in the induction of the genes encoding diptericin and cecropin}, author = {I Gross and Philippe Georgel and Christine Kappler and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0305-1048}, year = {1996}, date = {1996-04-01}, journal = {Nucleic Acids Res.}, volume = {24}, number = {7}, pages = {1238--1245}, abstract = {In Drosophila, bacterial challenge induces the rapid transcription of several genes encoding potent antibacterial peptides. The upstream sequences of the diptericin and cecropin Al genes, which have been investigated in detail, contain two, respectively one sequence element homologous to the binding site of the mammalian nuclear factor kappaB. These elements have been shown to be mandatory for immune-induced transcription of both genes. Functional studies have shown that these kappaB-related elements can be the target for the Drosophila Rel proteins dorsal and Dif. Here we present a comparative analysis of the transactivating capacities of these proteins on reporter genes fused to either the diptericin or the cecropin kappaB-related motifs. We conclude from our results: (i) the kappaB motifs of the diptericin and cecropin genes are not functionally equivalent; (ii) the dorsal and Dif proteins have distinct DNA-binding characteristics; (iii) dorsal and Dif can heterodimerize in vitro; (iv) mutants containing no copies of dorsal and a single copy of Dif retain their full capacity to express the diptericin and cecropin genes in response to challenge.}, keywords = {Animals, Antimicrobial Cationic Peptides, Base Sequence, DNA Primers, DNA-Binding Proteins, Gene Expression Regulation, Genetic, hoffmann, Insect Hormones, Insect Proteins, M3i, NF-kappa B, Nuclear Proteins, Peptides, Phosphoproteins, reichhart, Transcription, Transcription Factors, Transcriptional Activation}, pubstate = {published}, tppubtype = {article} } @article{ezekowitz_innate_1996, title = {Innate immunity}, author = {Alan R B Ezekowitz and Jules A Hoffmann}, issn = {1879-0372}, year = {1996}, date = {1996-02-01}, journal = {Curr. Opin. Immunol.}, volume = {8}, number = {1}, pages = {1--2}, keywords = {hoffmann, M3i}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_innate_1996, title = {Innate immunity in higher insects}, author = {Jules A Hoffmann and Jean-Marc Reichhart and Charles Hetru}, issn = {0952-7915}, year = {1996}, date = {1996-02-01}, journal = {Curr. Opin. Immunol.}, volume = {8}, number = {1}, pages = {8--13}, abstract = {The hallmark of the innate immune response of higher insects is the rapid and transient synthesis of a battery of broad spectrum antimicrobial peptides by the fat body. The control of the genes encoding these peptides involves cis-regulatory promoter elements homologous to sequences functional in mammalian acute-phase genes. Study of immune-deficient mutants of Drosophila has indicated that distinct pathways control the antibacterial and antifungal responses in this species. Novel receptors potentially involved in the initiation of the immune response have been recently characterized.}, keywords = {Animals, Base Sequence, Cyclic, hoffmann, Immunity, Immunologic, Immunological, Innate, insects, M3i, Models, Peptide Hydrolases, Peptides, Receptors, reichhart}, pubstate = {published}, tppubtype = {article} } @article{fehlbaum_structure-activity_1996, title = {Structure-activity analysis of thanatin, a 21-residue inducible insect defense peptide with sequence homology to frog skin antimicrobial peptides}, author = {P Fehlbaum and Philippe Bulet and S Chernysh and J P Briand and J P Roussel and L Letellier and Charles Hetru and Jules A Hoffmann}, issn = {0027-8424}, year = {1996}, date = {1996-02-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {93}, number = {3}, pages = {1221--1225}, abstract = {Immune challenge to the insect Podisus maculiventris induces synthesis of a 21-residue peptide with sequence homology to frog skin antimicrobial peptides of the brevinin family. The insect and frog peptides have in common a C-terminally located disulfide bridge delineating a cationic loop. The peptide is bactericidal and fungicidal, exhibiting the largest antimicrobial spectrum observed so far for an insect defense peptide. An all-D-enantiomer is nearly inactive against Gram-negative bacteria and some Gram-positive strains but is fully active against fungi and other Gram-positive bacteria, suggesting that more than one mechanism accounts for the antimicrobial activity of this peptide. Studies with truncated synthetic isoforms underline the role of the C-terminal loop and flanking residues for the activity of this molecule for which we propose the name thanatin.}, keywords = {Amino Acid, Amphibian Proteins, Animals, Anti-Bacterial Agents, Anti-Infective Agents, Antimicrobial Cationic Peptides, Cyclic, Fungi, Gram-Negative Bacteria, Gram-Positive Bacteria, Hemiptera, hoffmann, M3i, Mass Spectrometry, Microbial Sensitivity Tests, Peptides, Ranidae, Sequence Homology, Skin, Structure-Activity Relationship}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of RNA primers in initiation of minus-strand and plus-strand DNA synthesis of the yeast retrotransposon Ty1}, author = {S Friant and T Heyman and F X Wilhelm and M Wilhelm}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955910}, isbn = {8955910}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {674-80}, abstract = {The Ty1 retrotransposon of the yeast Saccharomyces cerevisiae is a long terminal repeat mobile genetic element that transposes through an RNA intermediate. Initiation of minus-strand and plus-strand DNA synthesis are two critical steps during reverse transcription of the retrotransposon genome. Initiation of minus-strand DNA synthesis of the Ty1 element is primed by the cytoplasmic initiator methionine tRNA base paired to the primer binding site near the 5' end of the genomic RNA. A structural probing study of the primer tRNA-Ty1 RNA binary complex reveals that besides interactions between the primer binding site and the last 10 nucleotides at the 3' end of the primer tRNA, three short regions of Ty1 RNA named box 0, box 1 and box 2.1 interact with the T and D stems and loops of the primer tRNA. Some in vivo results underline the functional importance of the nucleotide sequence of the boxes and suggest that extended interactions between genomic Ty1 RNA and the primer tRNA play a role in the reverse transcription pathway. Plus-strand DNA synthesis is initiated from an RNase H resistant oligoribonucleotide spanning a purine-rich sequence, the polypurine tract (PPT). Two sites of initiation located at the 5' boundary of the 3' long terminal repeat (PPT1) and near the middle of the TyB (pol) gene in the integrase coding sequence (PPT2) have been identified in the genome of Ty1. The two PPTs have an identical sequence, TGGGTGGTA. Mutations replacing purines by pyrimidines in this sequence significantly diminish or abolish initiation of plus-strand DNA synthesis. Ty1 elements bearing a mutated PPT2 sequence are not defective for transposition whereas mutations in PPT1 abolish transposition.}, note = {0300-9084 Journal Article}, keywords = {*DNA Replication *DNA Transposable Elements DNA, Bacterial/*metabolism Support, Complementary/metabolism Molecular Sequence Data Mutagenesis Nucleic Acid Conformation RNA/*metabolism RNA, Non-U.S. Gov't}, pubstate = {published}, tppubtype = {article} } @article{, title = {Topography of the Escherichia coli initiation factor 2/fMet-tRNA(f)(Met) complex as studied by cross-linking}, author = {G Yusupova and J Reinbolt and H Wakao and S Laalami and M Grunberg-Manago and P Romby and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8608135}, isbn = {8608135}, year = {1996}, date = {1996-01-01}, journal = {Biochemistry}, volume = {35}, number = {9}, pages = {2978-2984}, abstract = {trans-Diamminedichloroplatinum(II) was used to induce reversible cross-links between Escherichia coli initiation factor 2 (IF-2) and fMet-tRNA(f)(Met). Two distinct cross-links between IF-2 and the initiator tRNA were produced. Analysis of the cross-linking regions on both RNA and protein moieties reveals that the T arm of the tRNA is in the proximity of a region of the C-terminal domain of IF-2 (residues Asn611-Arg645). This cross-link is well-correlated with the fact that the C-domain of IF-2 contains the fMet-tRNA binding site and that the cross-linked RNA fragment precisely maps in a region which is protected by IF-2 from chemical modification and enzymatic digestion. Rather unexpectedly, a second cross-link was characterized which involves the anticodon arm of fMet-tRNA(f)(Met) and the N-terminal part of IF-2 (residues Trp215-Arg237).}, note = {0006-2960 Journal Article}, keywords = {Amino Acid Sequence Base Sequence Cisplatin/*pharmacology Cross-Linking Reagents Electrophoresis, Met/chemistry/isolation & purification/*metabolism Substrate Specificity Support, Non-U.S. Gov't, Polyacrylamide Gel Escherichia coli/drug effects/*metabolism Eukaryotic Initiation Factor-2/chemistry/isolation & purification/*metabolism Kinetics Molecular Sequence Data Nucleic Acid Conformation Peptide Fragments/chemistry/isolation & purification Protein Conformation RNA, ROMBY, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Primer selection by HIV-1 reverse transcriptase on RNA-tRNA(3Lys) and DNA-tRNA(3Lys) hybrids}, author = {G Yusupova and J M Lanchy and M Yusupov and G Keith and S F Le Grice and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8780773}, isbn = {8780773}, year = {1996}, date = {1996-01-01}, journal = {J Mol Biol}, volume = {261}, number = {3}, pages = {315-321}, abstract = {During reverse transcription of the genomic RNA of human immunodeficiency virus type 1 (HIV-1) into double-stranded DNA, reverse transcriptase (RT) must accommodate RNA-RNA, DNA-RNA, RNA-DNA and DNA-DNA hybrids as primer-template. In this study, we examined extension of RNA-tRNA3Lys, and DNA-tRNA3Lys complexes by HIV-1 RT. When the 3' end of tRNA3Lys is annealed to oligoribonucleotides, tRNA3Lys, but not the complementary RNAs, is extended by HIV-1 RT, indicating that tRNA3Lys is efficiently used as primer and RNA as template. An opposite primer usage is observed when tRNA3Lys is annealed to complementary oligodeoxyribonucleotides. In this case, the oligodeoxyribonucleotides are efficiently used as primer and tRNA3Lys as template. This result indicates that the nature of nucleic acid bound to tRNA3Lys determines which strand of the RNA-tRNA3Lys and DNA-tRNA3Lys hybrids is extended by HIV-1 RT. When an oligoribonucleotide is annealed to an unmodified transcript of tRNA3Lys, both nucleic acids are extended by HIV-1 RT, indicating that specific selection of tRNA3Lys as primer requires the post-transcriptional modifications of tRNA3Lys.}, note = {0022-2836 Journal Article}, keywords = {Base Sequence DNA, Lys/genetics/*metabolism RNA, MARQUET, Non-U.S. Gov't Support, P.H.S., Post-Transcriptional RNA, Transfer, U.S. Gov't, Unité ARN, Viral/genetics HIV-1/*enzymology/genetics HIV-1 Reverse Transcriptase Human Molecular Sequence Data *RNA Processing, Viral/genetics RNA-Directed DNA Polymerase/genetics/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural basis of ligand discrimination by two related RNA aptamers resolved by NMR spectroscopy}, author = {Y Yang and M Kochoyan and P Burgstaller and E Westhof and M Famulok}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8650546}, isbn = {8650546}, year = {1996}, date = {1996-01-01}, journal = {Science}, volume = {272}, number = {5266}, pages = {1343-1347}, abstract = {In a previous study, an RNA aptamer for the specific recognition of arginine was evolved from a parent sequence that bound citrulline specifically. The two RNAs differ at only 3 positions out of 44. The solution structures of the two aptamers complexed to their cognate amino acids have now been determined by two-dimensional nuclear magnetic resonance spectroscopy. Both aptamers contain two asymmetrical internal loops that are not well ordered in the free RNA but that fold into a compact structure upon ligand binding. Those nucleotides common to both RNAs include a conserved cluster of purine residues, three of which form an uneven plane containing a G:G pair, and two other residues nearly perpendicular to that surface. Two of the three variant nucleotides are stacked on the cluster of purines and form a triple contact to the amino acid side chain, whereas the edge of the third variant nucleotide is capping the binding pocket.}, note = {0036-8075 Journal Article}, keywords = {Arginine/chemistry/*metabolism Base Composition Base Sequence Citrulline/chemistry/*metabolism Crystallography, Molecular Molecular Sequence Data Mutation *Nucleic Acid Conformation RNA/*chemistry/genetics/*metabolism, Unité ARN, X-Ray Hydrogen Bonding Ligands Magnetic Resonance Spectroscopy Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mapping in three dimensions of regions in a catalytic RNA protected from attack by an Fe(II)-EDTA reagent}, author = {E Westhof and D Wesolowski and S Altman}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8636995}, isbn = {8636995}, year = {1996}, date = {1996-01-01}, journal = {J Mol Biol}, volume = {258}, number = {4}, pages = {600-613}, abstract = {The accessibility of the ribose groups in the phosphodiester chain of M1 RNA, the catalytic subunit of ribonuclease P from Escherichia coli, has been probed with an Fe(II)-EDTA reagent when the RNA is alone in solution, when it is in a complex with a tRNA precursor substrate, and when it is in the holoenzyme complex with its cofactor, C5 protein. The regions found to be protected under these various conditions, as well as those previously identified in other chemical probing experiments, have been mapped on a three-dimensional working model of M1 RNA and are generally compatible with the previously proposed placement of the substrate on the enzyme and with previous data and inferences regarding the interactions of C5 protein with M1 RNA. On the basis of the accessibilities of the C(4') atoms, refinements have been introduced in the model to accommodate the Fe(II)-EDTA protection data. The protein cofactor makes contact with several helical regions of the catalytic RNA on the opposite side of the surface to which substrates bind.}, note = {0022-2836 Journal Article}, keywords = {Bacterial Proteins/chemistry/metabolism Base Sequence Coenzymes/*chemistry/drug effects Computer Simulation Edetic Acid/*pharmacology Endoribonucleases/*chemistry/drug effects Escherichia coli/chemistry Ferrous Compounds/*pharmacology Magnesium/pharmacology Models, Bacterial/*chemistry/drug effects RNA, Catalytic/*chemistry/drug effects RNA, Molecular Molecular Sequence Data *Nucleic Acid Conformation Protein Binding RNA, Non-U.S. Gov't Support, P.H.S., Transfer/chemistry/metabolism Ribonuclease P Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {The structural domains of group I introns.}, author = {E Westhof and F Michel}, editor = {R Green and R Schroeder}, url = {https://catalyst.library.jhu.edu/catalog/bib_1891715}, year = {1996}, date = {1996-01-01}, booktitle = {Ribosomal Rna and Group I Introns (Molecular Biology Intelligence Unit)}, pages = {1-14}, publisher = {R G Landes Co}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Role of RNA primers in initiation of minus-strand and plus-strand DNA synthesis of the yeast retrotransposon Ty1}, author = { S. Friant and T. Heyman and F. X. Wilhelm and M. Wilhelm}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {674-80}, abstract = {The Ty1 retrotransposon of the yeast Saccharomyces cerevisiae is a long terminal repeat mobile genetic element that transposes through an RNA intermediate. Initiation of minus-strand and plus-strand DNA synthesis are two critical steps during reverse transcription of the retrotransposon genome. Initiation of minus-strand DNA synthesis of the Ty1 element is primed by the cytoplasmic initiator methionine tRNA base paired to the primer binding site near the 5' end of the genomic RNA. A structural probing study of the primer tRNA-Ty1 RNA binary complex reveals that besides interactions between the primer binding site and the last 10 nucleotides at the 3' end of the primer tRNA, three short regions of Ty1 RNA named box 0, box 1 and box 2.1 interact with the T and D stems and loops of the primer tRNA. Some in vivo results underline the functional importance of the nucleotide sequence of the boxes and suggest that extended interactions between genomic Ty1 RNA and the primer tRNA play a role in the reverse transcription pathway. Plus-strand DNA synthesis is initiated from an RNase H resistant oligoribonucleotide spanning a purine-rich sequence, the polypurine tract (PPT). Two sites of initiation located at the 5' boundary of the 3' long terminal repeat (PPT1) and near the middle of the TyB (pol) gene in the integrase coding sequence (PPT2) have been identified in the genome of Ty1. The two PPTs have an identical sequence, TGGGTGGTA. Mutations replacing purines by pyrimidines in this sequence significantly diminish or abolish initiation of plus-strand DNA synthesis. Ty1 elements bearing a mutated PPT2 sequence are not defective for transposition whereas mutations in PPT1 abolish transposition.}, note = {0300-9084 Journal Article}, keywords = {*DNA, Acid, Bacterial/*metabolism, Complementary/metabolism, Conformation, Data, DNA, Elements, Gov't, Molecular, Mutagenesis, Non-U.S., Nucleic, Replication, RNA, RNA/*metabolism, Sequence, Support, Transposable}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNA tectonics: towards RNA design}, author = {E Westhof and B Masquida and L Jaeger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9079386}, isbn = {9079386}, year = {1996}, date = {1996-01-01}, journal = {Fold Des}, volume = {1}, number = {4}, pages = {R78-88}, abstract = {Our understanding of the structural, folding and catalytic properties of RNA molecules has increased enormously in recent years. The discovery of catalytic RNA molecules by Sidney Altman and Tom Cech, the development of in vitro selection procedures, and the recent crystallizations of hammerhead ribozymes and of a large domain of an autocatalytic group 1 intron are some of the milestones that have contributed to the explosion of the RNA field. The availability of a three-dimensional model for the catalytic core of group 1 introns contributed also a heuristic drive toward the development of new techniques and approaches for unravelling RNA architecture, folding and stability. Here, we emphasize the mosaic structure of RNA and review some of the recent literature pertinent to this working framework. In the long run, RNA tectonics aims at constructing combinatorial libraries, using RNA mosaic units for creating molecules with dedicated shapes and properties.}, note = {1359-0278 Journal Article Review Review, Tutorial}, keywords = {Animals Base Sequence Drug Design Evolution, Catalytic/chemistry/genetics RNA, Molecular Models, Molecular Molecular Sequence Data Molecular Structure Nucleic Acid Conformation RNA/*chemistry/genetics RNA, Protozoan/chemistry/genetics Tetrahymena thermophila/chemistry/genetics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Extended interactions between the primer tRNAi(Met) and genomic RNA of the yeast Ty1 retrotransposon}, author = { S. Friant and T. Heyman and M. L. Wilhelm and F. X. Wilhelm}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {3}, pages = {441-9}, abstract = {Reverse transcription of the yeast Ty1 retrotransposon is primed by tRNAi(Met) base paired to the primer binding site near the 5'-end of Ty1 genomic RNA. To understand the molecular basis of the tRNAi(Met)-Ty1 RNA interaction the secondary structure of the binary complex was analysed. Enzymatic probes were used to test the conformation of tRNAi(Met) and of Ty1 RNA in the free form and in the complex. A secondary structure model of the tRNAi(Met) Ty1 RNA complex consistent with the probing data was constructed with the help of a computer program. The model shows that besides interactions between the primer binding site and the last 10 nt at the 3'-end of tRNAi(Met), three short regions of Ty1 RNA named boxes 0, 1 and 2.1 interact with the T and D stems and loops of tRNAiMet. Mutations were made in the boxes or in the complementary sequences of tRNAi(Met) to study the contribution of these sequences to formation of the complex. We find that interaction with at least one of the two boxes 0 or 1 is absolutely required for efficient annealing of the two RNAs. Sequence comparison showing that the primary sequence of the boxes is strictly conserved in Ty1 and Ty2 elements and previously published in vivo results underline the functional importance of the primary sequence of the boxes and suggest that extended interactions between genomic Ty1 RNA and the primary tRNAi(Met) play a role in the reverse transcription pathway.}, note = {0305-1048 Journal Article}, keywords = {Acid, Base, cerevisiae, Conformation, Data, Gov't, Met/genetics/*metabolism, Molecular, Mutation, Non-U.S., Nucleic, Retroelements/*genetics, RNA, RNA/genetics/*metabolism, Saccharomyces, Sequence, structure, Support, Transfer}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Refinement of protein and nucleic acid structures}, author = {E Westhof and P Dumas}, editor = {C Jones and B Mulloy and M R Sanderson}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8781248}, doi = {10.1385/0-89603-259-0:227}, isbn = {8781248}, year = {1996}, date = {1996-01-01}, booktitle = {Crystallographic Methods and Protocols}, volume = {56}, pages = {227-244}, publisher = {Springer Protocols, Humana Press}, series = {Methods in Molecular Biology}, note = {Review Review, Tutorial}, keywords = {Animals Base Sequence Crystallography, Unité ARN, X-Ray/methods DNA/*chemistry Deoxyribonucleotides/chemistry Dogfish Isoenzymes L-Lactate Dehydrogenase/chemistry *Nucleic Acid Conformation *Protein Conformation Proteins/*chemistry RNA/*chemistry Software}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {A novel RNA structural motif in the selenocysteine insertion element of eukaryotic selenoprotein mRNAs}, author = {R Walczak and E Westhof and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8634917}, isbn = {8634917}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {4}, pages = {367-379}, abstract = {In eukaryotes, co-translational insertion of selenocysteine into selenoproteins necessitates the participation of the selenocysteine insertion sequence (SECIS), an element lying in the 3'-untranslated region of selenoprotein mRNAs. We report a detailed experimental study of the secondary structures of the SECIS elements of three selenoprotein mRNAs, the rat and human type I iodothyronine deiodinase (5'DI) and rat glutathione peroxidase (GPx). Based on RNase and chemical probing, a new secondary structure model is established. It is characterized by a stem-loop structure, comprising two helices (I and II) separated by an internal loop, with an apical loop surmounting helix II. Sequence comparisons of 20 SECIS elements, arising from 2 5'DI, 13 GPx, 2 selenoprotein P, and 1 selenoprotein W mRNAs, confirm the secondary structure model. The most striking finding of the experimental study concerns a set of conserved sequences in helix II that interact to form a novel RNA structural motif consisting of a quartet composed of non-Watson-Crick base pairs 5'UGAY3': 5'UGAU3'. The potential for forming the quartet is preserved in 15 SECIS elements, but three consecutive non-Watson-Crick base pairs can nevertheless form in the other five SECIS, the central G.A tandem being invariant in all cases. A 3D model, derived by computer modeling with the use of the solution data, suggests that the base pairing interactions in the G.A tandem are of the type found in GNRA loops. The 3D model displays the quartet lying in an accessible position at the foot of helix II, which is bent at the internal loop, suggesting that the non-Watson-Crick base pair arrangement provides an unusual pattern of chemical groups for putative ligand interaction.}, note = {1355-8382 Journal Article}, keywords = {Animals Base Sequence DNA Primers Glutathione Peroxidase/chemistry Human Hydroxyl Radical Models, Messenger/*chemistry Rats Selenocysteine/chemistry/*genetics Support, Molecular Molecular Probes Molecular Sequence Data Protein Conformation Protein Structure, Non-U.S. Gov't, Secondary Proteins/chemistry/*genetics RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The environment of two metal ions surrounding the splice site of a group I intron}, author = {B Streicher and E Westhof and R Schroeder}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8665863}, isbn = {8665863}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {10}, pages = {2556-2564}, abstract = {Several divalent metal ions (Ca2+, Sr2+ and Pb2+) do not promote splicing, but instead induce cleavage at a single site in the conserved group I intron core in the absence of the guanosine cofactor at elevated pH, generating products with 5'-OH and 3'-phosphate ends. The reaction is competed by Mg2+, which does not cleave at this position, but hydrolyses the splice sites producing 3'-OH and 5'-phosphate ends. Mn2+ promotes both core cleavage and splice site hydrolysis under identical conditions, suggesting that two different metal atoms are involved, each responsible for one type of cleavage, and with different chemical and geometric requirements. Based on the core cleavage position and on the previously proposed coordination sites for Mg2+, we propose a structural location for two metal ions surrounding the splice site in the Michel-Westhof three-dimensional model of the group I intron core. The proposed location was strengthened by a first mutational analysis which supported the suggested interaction between one of the metal ions and the bulged residue in P7.}, note = {0261-4189 Journal Article}, keywords = {Bacteriophage T4/genetics Base Sequence Calcium/physiology Catalysis *Cations, Catalytic/chemistry/*metabolism Strontium/physiology Support, Divalent Comparative Study Hydrogen-Ion Concentration Hydrolysis Introns/*genetics Lead/physiology Magnesium/physiology Manganese/physiology Models, Molecular Molecular Sequence Data RNA Precursors/genetics/*metabolism RNA Splicing/*physiology RNA, Non-U.S. Gov't Thymidylate Synthase/genetics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mechanisms of inhibition of in vitro dimerization of HIV type I RNA by sense and antisense oligonucleotides}, author = {E Skripkin and J C Paillart and R Marquet and M Blumenfeld and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8910525}, isbn = {8910525}, year = {1996}, date = {1996-01-01}, journal = {J Biol Chem}, volume = {271}, number = {46}, pages = {28812-28817}, abstract = {Retroviruses display a strong selective pressure to maintain the dimeric nature of their genomic RNAs, suggesting that dimerization is essential for viral replication. Recently, we identified the cis-element required for initiation of human immunodeficiency virus type I (HIV-I) RNA dimerization in vitro. The dimerization initiation site (DIS) is a hairpin structure containing a self-complementary sequence in the loop. We proposed that dimerization is initiated by a loop-loop kissing interaction involving the self-complementary sequence present in each monomer. We tested the ability of sense and antisense oligonucleotides targeted against the DIS to interfere with a preformed viral RNA dimer. Self-dimerization and inhibition properties of the tested oligonucleotides are dictated by the nature of the loop. An RNA loop is absolutely required in the case of sense oligonucleotides, whereas the nature and the sequence of the stem is not important. They form reversible loop-loop interactions and act as competitive inhibitors. Antisense oligonucleotides are less efficient in self-dimerization and are more potent inhibitors than sense oligonucleotides. They are less sensitive to the nature of the loop than the antisense oligonucleotides. Antisense hairpins with either RNA or DNA stems are able to form highly stable and irreversible complexes with viral RNA, resulting from complete extension of base pairing initiated by loop-loop interaction.}, note = {0021-9258 Journal Article}, keywords = {Antisense/*pharmacology RNA, Biopolymers HIV-1/*genetics Nucleic Acid Conformation Oligonucleotides/*pharmacology Oligonucleotides, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*antagonists & inhibitors/chemistry Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Psoralen crosslinking between human immunodeficiency virus type 1 RNA and primer tRNA3(Lys)}, author = {E Skripkin and C Isel and R Marquet and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8602365}, isbn = {8602365}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {3}, pages = {509-514}, abstract = {Initiation of reverse transcription is a crucial step of retroviral infection. In HIV-1, it involves hybridization of the 18 3'-terminal nucleotides of the primer tRNA3(Lys) to the primer binding site (PBS) of the viral RNA. Moreover, additional interactions between the two RNAs were recently evidenced [Isel et al. (1995) J. Mol. Biol. 247, 25269-25272]. To get further information on the topology of the viral RNA/tRNA3(Lys) complex, we used psoralen to induce RNA-RNA crosslinking. A defined intermolecular crosslinked complex was obtained. The crosslinked regions were characterized by RNase T1 digestion followed by bi-dimensional gel electrophoresis. The crosslinked residues (nucleotide mcm5S2U34 and U35 in the anticodon loop of tRNA3(Lys) and UCU154 in the viral RNA upstream of the PBS) were mapped using a retardation method coupled with random hydrolysis. The formation of this crosslink depends on the same elements that are required for the formation of the extended interactions between primer and template RNAs, i.e., the modified bases of the tRNA and a conserved A-rich loop located upstream of the PBS in the genomic RNA. Therefore, the present crosslinking data provide relevant information on the topology of the template/primer binary complex.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Cross-Linking Reagents/*metabolism Ficusin/*metabolism HIV-1/*genetics Human Molecular Sequence Data Molecular Structure RNA, Lys/*genetics/metabolism RNA, MARQUET, Non-U.S. Gov't, Transfer, Unité ARN, Viral/*genetics/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Arginine aminoacylation identity is context-dependent and ensured by alternate recognition sets in the anticodon loop of accepting tRNA transcripts}, author = {M Sissler and R Giege and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8890180}, isbn = {8890180}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {18}, pages = {5069-5076}, abstract = {Yeast arginyl-tRNA synthetase recognizes the non-modified wild-type transcripts derived from both yeast tRNA(Arg) and tRNA(Asp) with equal efficiency. It discriminates its cognate natural substrate, tRNA(Arg), from non-cognate tRNA(Asp) by a negative discrimination mechanism whereby a single methyl group acts as an anti-determinant. Considering these facts, recognition elements responsible for specific arginylation in yeast have been searched by studying the in vitro arginylation properties of a series of transcripts derived from yeast tRNA(Asp), considered as an arginine isoacceptor tRNA. In parallel, experiments on similar tRNA(Arg) transcripts were performed. Unexpectedly, in the tRNA(Arg) context, arginylation is basically linked to the presence of residue C35, whereas in the tRNA(Asp) context, it is deeply related to that of C36 and G37 but is insensitive to the nucleotide at position 35. Each of these nucleotides present in one host, is absent in the other host tRNA. Thus, arginine identity is dependent on two different specific recognition sets according to the tRNA framework investigated.}, note = {0261-4189 Journal Article}, keywords = {*Anticodon Arginine/*metabolism Base Sequence Kinetics Molecular Sequence Data Nucleic Acid Conformation RNA, Arg/*chemistry/metabolism RNA, Asp/chemistry/metabolism Saccharomyces cerevisiae Support, FLORENTZ, Fungal/*chemistry/metabolism RNA, Non-U.S. Gov't, SISSLER, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The yeast protein Arc1p binds to tRNA and functions as a cofactor for the methionyl- and glutamyl-tRNA synthetases}, author = {G Simos and A Segref and F Fasiolo and K Hellmuth and A Shevchenko and M Mann and E C Hurt}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8895587}, isbn = {8895587}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {19}, pages = {5437-5448}, abstract = {Arc1p was found in a screen for components that interact genetically with Los1p, a nuclear pore-associated yeast protein involved in tRNA biogenesis. Arc1p is associated with two proteins which were identified as methionyl-tRNA and glutamyl-tRNA synthetase (MetRS and GluRS) by a new mass spectrometry method. ARC1 gene disruption leads to slow growth and reduced MetRS activity, and synthetically lethal arc1- mutants are complemented by the genes for MetRS and GluRS. Recombinant Arc1p binds in vitro to purified monomeric yeast MetRS, but not to an N-terminal truncated form, and strongly increases its apparent affinity for tRNAMet. Furthermore, Arc1p, which is allelic to the quadruplex nucleic acid binding protein G4p1, exhibits specific binding to tRNA as determined by gel retardation and UV-cross-linking. Arc1p is, therefore, a yeast protein with dual specificity: it associates with tRNA and aminoacyl-tRNA synthetases. This functional interaction may be required for efficient aminoacylation in vivo.}, note = {0261-4189 Journal Article}, keywords = {Acylation Amino Acid Sequence Cytoplasm/chemistry Genes, Amino Acid Support, Fungal/genetics Glutamate-tRNA Ligase/*metabolism Kinetics Membrane Proteins/metabolism Methionine-tRNA Ligase/*metabolism Molecular Sequence Data Mutation Nuclear Envelope/metabolism RNA, Non-U.S. Gov't Yeasts/enzymology/*genetics/growth & development, Transfer/*metabolism RNA-Binding Proteins/analysis/genetics/*metabolism Recombinant Fusion Proteins/metabolism *Saccharomyces cerevisiae Proteins Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The 16S rRNA binding site of Thermus thermophilus ribosomal protein S15: comparison with Escherichia coli S15, minimum site and structure}, author = {A A Serganov and B Masquida and E Westhof and C Cachia and C Portier and M Garber and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8903343}, isbn = {8903343}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {11}, pages = {1124-1138}, abstract = {Binding of Escherichia coli and Thermus thermophilus ribosomal proteins S15 to a 16S ribosomal RNA fragment from T. thermophilus (nt 559-753) has been investigated in detail by extensive deletion analysis, filter-binding assays, gel mobility shift, structure probing, footprinting with chemical, enzymatic, and hydroxyl radical probes. Both S15 proteins recognize two distinct sites. The first one maps in the bottom of helix 638-655/717-734 (H22) and in the three-way junction between helix 560-570/737-747 (H20), helix 571-600/606-634 (H21), and H22. The second is located in a conserved purine-rich region in the center of H22. The first site provides a higher contribution to the free energy of binding than the second one, and both are required for efficient binding. A short RNA fragment of 56 nt containing these elements binds S15 with high affinity. The structure of the rRNA is constrained by the three-way junction and requires both magnesium and S15 to be stabilized. A 3D model, derived by computer modeling with the use of experimental data, suggests that the bound form adopts a Y-shaped conformation, with a quasi-coaxial stacking of H22 on H20, and H21 forming an acute angle with H22. In this model, S15 binds to the shallow groove of the RNA on the exterior side of the Y-shaped structure, making contact with the two sites, which are separated by one helix turn.}, note = {1355-8382 Journal Article}, keywords = {16S/chemistry/genetics/*metabolism Ribosomal Proteins/*metabolism Species Specificity Support, Bacterial/chemistry/genetics/*metabolism RNA, Base Sequence Binding Sites/genetics Comparative Study Computer Simulation Conserved Sequence Escherichia coli/genetics/*metabolism Magnesium/metabolism Models, Molecular Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, Non-U.S. Gov't Thermodynamics Thermus thermophilus/genetics/*metabolism, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast tRNA(Met) recognition by methionyl-tRNA synthetase requires determinants from the primary, secondary and tertiary structure: a review}, author = {B Senger and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955903}, isbn = {8955903}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {597-604}, abstract = {The primordial role of the CAU anticodon in methionine identity of the tRNA has been established by others nearly a decade ago in Escherichia coli and yeast tRNA(Met). We show here that the CAU triplet alone is unable to confer methionine acceptance to a tRNA. This requires the contribution of the discriminatory base A73 and the non-anticodon bases of the anticodon loop. To better understand the functional communication between the anticodon and the active site, we analysed the binding and aminoacylation of tRNA(Met) based anticodon and acceptor-stem minihelices and of tRNA(Met) chimeras where the central core region of yeast tRNA(Met) is replaced by that of unusual mitochondrial forms lacking either a D-stem or a T-stem. These studies suggest that the high selectivity of the anticodon bases in tRNA(Met) implies the L-conformation of the tRNA and the presence of a D-stem. The importance of a L-structure for recognition of tRNA(Met) was also deduced from mutations of tertiary interactions known to play a general role in tRNA(Met) folding.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Amino Acid Sequence Anticodon Methionine-tRNA Ligase/*metabolism Molecular Sequence Data Nucleic Acid Conformation Protein Structure, Met/*metabolism Structure-Activity Relationship Support, Non-U.S. Gov't, Secondary Protein Structure, Tertiary RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Antideterminants present in minihelix(Sec) hinder its recognition by prokaryotic elongation factor Tu}, author = {J Rudinger and R Hillenbrandt and M Sprinzl and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8599948}, isbn = {8599948}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {3}, pages = {650-657}, abstract = {During protein biosynthesis, all aminoacylated elongator tRNAs except selenocysteine-inserting tRNA Sec form ternary complexes with activated elongation factor. tRNA Sec is bound by its own translation factor, an elongation factor analogue, e.g. the SELB factor in prokaryotes. An apparent reason for this discrimination could be related to the unusual length of tRNA Sec amino acid-acceptor branch formed by 13 bp. However, it has been recently shown that an aspartylated minihelix of 13 bp derived from yeast tRNA Asp is an efficient substrate for Thermus thermophilus EF-Tu-GTP, suggesting that features other than the length of tRNA Sec prevent its recognition by EF-Tu-GTP. A stepwise mutational analysis of a minihelix derived from tRNA Sec in which sequence elements of tRNA Asp were introduced showed that the sequence of the amino acid- acceptor branch of Escherichia coli tRNA Sec contains a specific structural element that hinders its binding to T.thermophilus EF-Tu-GTP. This antideterminant is located in the 8th, 9th and 10th bp in the acceptor branch of tRNA Sec, corresponding to the last base pair in the amino acid acceptor stem and the two first pairs in the T-stem. The function of this C7.G66/G49.U65/C50.G64 box was tested by its transplantation into a minihelix derived from tRNA Asp, abolishing its recognition by EF-Tu-GTP. The specific role of this nucleotide combination is further supported by its absence in all known prokaryotic elongator tRNAs.}, note = {0261-4189 Journal Article}, keywords = {Amino Acid-Specific/*chemistry/genetics/*metabolism RNA, Antisense/chemistry/genetics/metabolism RNA, Asp/chemistry/genetics/metabolism Saccharomyces cerevisiae/genetics/metabolism Support, Base Sequence Escherichia coli/genetics/metabolism Evolution Guanosine Triphosphate/metabolism Molecular Sequence Data Nucleic Acid Conformation Peptide Elongation Factor Tu/*metabolism RNA, Non-U.S. Gov't Thermus thermophilus/metabolism, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The expression of E.coli threonyl-tRNA synthetase is regulated at the translational level by symmetrical operator-repressor interactions}, author = {P Romby and J Caillet and C Ebel and C Sacerdot and M Graffe and F Eyermann and C Brunel and H Moine and C Ehresmann and B Ehresmann and M Springer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8918475}, isbn = {8918475}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {21}, pages = {5976-5987}, abstract = {Threonyl-tRNA synthetase from Escherichia coli represses the translation of its own mRNA by binding to the operator region located upstream from the ribosome binding site. The operator contains two stemloop structures which interact specifically with the homodimeric enzyme. Here, we provide in vitro and in vivo evidence that these two stem-loop structures are recognized by the enzyme in an analogous way and mimic the anticodon arm of E.coli tRNA(Thr). Determination of the stoichiometry of the different RNA-threonyl-tRNA synthetase complexes reveals that two tRNA(Thr) molecules bind to the enzyme whereas only one thrS operator interacts with the homodimeric enzyme. A model is presented in which the two anticodon-like domains of the operator bind symmetrically to the two tRNA(Thr) anticodon recognition sites (one per subunit) of the dimeric threonyl-tRNA synthetase. Although symmetrical operator-repressor interactions in transcriptional control are widespread, this report stresses the importance of such interactions in translational regulation of gene expression.}, note = {0261-4189 Journal Article}, keywords = {Anticodon Base Sequence Binding Sites Binding, Bacterial Gene Expression Regulation, Bacterial/chemistry/genetics/metabolism RNA, Biological Molecular Sequence Data Mutagenesis, Competitive Escherichia coli/*enzymology/*genetics/metabolism Gene Expression Regulation, Enzymologic Models, Genetic, Messenger/genetics/metabolism Repressor Proteins/genetics Support, Non-U.S. Gov't Threonine-tRNA Ligase/*genetics Translation, ROMBY, Site-Directed Nucleic Acid Conformation Operator Regions (Genetics) RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {2'-O-methyl-5-formylcytidine (f5Cm), a new modified nucleotide at the 'wobble' of two cytoplasmic tRNAs Leu (NAA) from bovine liver}, author = {J P Pais de Barros and G Keith and C El Adlouni and A L Glasser and G Mack and G Dirheimer and J Desgres}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8628682}, isbn = {8628682}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {8}, pages = {1489-1496}, abstract = {The nucleotide analysis of a cytoplasmic tRNA(Leu) isolated from bovine liver revealed the presence of an unknown modified nucleotide N. The corresponding N nucleoside was isolated by different enzymatic and chromatographic protocols from a partially purified preparation of this tRNA(Leu). Its chemical characterization was determined from its chromatographic properties, UV-absorption spectroscopy and mass spectrometric measurements, as well as from those of the borohydride reduced N nucleoside and its etheno-trimethylsilyl derivative. The structure of N was established as 2'-O-methyl-5-formylcytidine (f5CM), and its reduced derivative as 2'-O-methyl-5-hydroxy-methylcytidine (om5Cm). By sequencing the bovine liver tRNA(Leu), the structure of the anticodon was determined as f5CmAA. In addition, the nucleotide sequence showed two primary structures differing only by the nucleotide 47c which is either uridine or adenosine. The two slightly differing bovine liver tRNAs-Leu(f5CmAA) are the only tRNAs so far sequenced which contain f5Cm. The role of such a modified cytidine at the first position of the anticodon is discussed in terms of decoding properties for the UUG and UUA leucine codons. Recently, precise evidence was obtained for the presence of f5Cm at the same position in tRNAs(Leu)(NAA) isolated from rabbit and lamb liver. Therefore, the 2'-O-methyl-5-formyl modification of cytidine at position 34 could be a general feature of cytoplasmic tRNAs(Leu)(NAA) in mammals.}, note = {0305-1048 Journal Article}, keywords = {Amino Acyl/*chemistry/isolation & purification Support, Animals Base Sequence Borohydrides/chemistry Cattle Cytidine/*analogs & derivatives/chemistry/isolation & purification Cytoplasm Hela Cells Human Liver/*chemistry Mass Fragmentography Molecular Sequence Data Molecular Structure Nucleic Acid Conformation RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A loop-loop "kissing" complex is the essential part of the dimer linkage of genomic HIV-1 RNA}, author = {J C Paillart and E Skripkin and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8643617}, isbn = {8643617}, year = {1996}, date = {1996-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {93}, number = {11}, pages = {5572-5577}, abstract = {RNA-RNA interactions govern a number of biological processes. Several RNAs, including natural sense and antisense RNAs, interact by means of a two-step mechanism: recognition is mediated by a loop-loop complex, which is then stabilized by formation of an extended intermolecular duplex. It was proposed that the same mechanism holds for dimerization of the genomic RNA of human immunodeficiency virus type 1 (HIV-1), an event thought to control crucial steps of HIV-1 replication. However, whereas interaction between the partially self-complementary loop of the dimerization initiation site (DIS) of each monomer is well established, formation of the extended duplex remained speculative. Here we first show that in vitro dimerization of HIV-1 RNA is a specific process, not resulting from simple annealing of denatured molecules. Next we used mutants of the DIS to test the formation of the extended duplex. Four pairs of transcomplementary mutants were designed in such a way that all pairs can form the loop-loop "kissing" complex, but only two of them can potentially form the extended duplex. All pairs of mutants form heterodimers whose thermal stability, dissociation constant, and dynamics were analyzed. Taken together, our results indicate that, in contrast with the interactions between natural sense and antisense RNAs, no extended duplex is formed during dimerization of HIV-1 RNA. We also showed that 55-mer sense RNAs containing the DIS are able to interfere with the preformed HIV-1 RNA dimer.}, note = {0027-8424 Journal Article}, keywords = {Base Composition Base Sequence HIV-1/*genetics Heat Human Kinetics Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Nucleic Acid Denaturation Plasmids RNA, Genetic, MARQUET, Non-U.S. Gov't Thermodynamics Transcription, PAILLART, Unité ARN, Viral/biosynthesis/*chemistry/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {The use of chemical modification interference and inverse PCR mutagenesis to identify the dimerization initiation site of HIV-1 genomic RNA}, author = {J C Paillart and E Skripkin and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8786995}, isbn = {8786995}, year = {1996}, date = {1996-01-01}, journal = {Pharm Acta Helv}, volume = {71}, number = {1}, pages = {21-28}, abstract = {The retroviral genome consists of two identical RNA molecules physically linked together close to their 5' end, in a region called the Dimer Linkage Structure (DLS). Recent findings suggest that dimerization is involved in encapsidation, regulation of translation and reverse transcription. Previous in vitro studies localized the DLS of HIV-1 in a region downstream of the splice donor (SD) site. More recently, we showed that dimerization of HIV-1 RNA also involves sequences upstream of the SD site. Modification interference experiments and site-directed mutagenesis were used to identify the nucleotides required in the dimerization process of HIV-1 RNA. Our results point out a self-complementary sequence located in a hairpin loop, between the Primer Binding Site (PBS) and the SD site, as the Dimerization Initiation Site.}, note = {0031-6865 Journal Article}, keywords = {Base Sequence *Genome, MARQUET, Non-U.S. Gov't, PAILLART, Site-Directed Polymerase Chain Reaction RNA, Unité ARN, Viral HIV-1/*chemistry Human Molecular Sequence Data Mutagenesis, Viral/*chemistry Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dimerization of retroviral genomic RNAs: structural and functional implications}, author = {J C Paillart and R Marquet and E Skripkin and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955907}, isbn = {8955907}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {639-653}, abstract = {Retroviruses are a family of widespread small animal viruses at the origin of a diversity of diseases. They share common structural and functional properties such as reverse transcription of their RNA genome and integration of the proviral DNA into the host genome, and have the particularity of packaging a diploid genome. The genome of all retroviruses is composed of two homologous RNA molecules that are non-covalently linked near their 5' end in a region called the dimer linkage structure (DLS). There is now considerable evidence that a specific site (or sites) in the 5' leader region of all retroviruses, located either upstream or/and downstream of the major splice donor site, is involved in the dimer linkage. For MoMuLV and especially HIV-1, it was shown that dimerization is initiated at a stem-loop structure named the dimerization initiation site (DIS). The DIS of HIV-1 and related regions in other retroviruses corresponds to a highly conserved structure with a self-complementary loop sequence, that is involved in a typical loop-loop 'kissing' complex which can be further stabilized by long distance interactions or by conformational rearrangements. RNA interactions involved in the viral RNA dimer were postulated to regulate several key steps in retroviral cycle, such as: i) translation and encapsidation: the arrest of gag translation imposed by the highly structured DLS-encapsidation signal would leave the RNA genome available for the encapsidation machinery; and ii) recombination during reverse transcription: the presence of two RNA molecules in particles would be necessary for variability and viability of virus progeny and the ordered structure imposed by the DLS would be required for efficient reverse transcription.}, note = {0300-9084 Journal Article Review Review, Academic}, keywords = {Animals Base Sequence HIV-1/genetics Human Microscopy, DNA Support, Electron Molecular Sequence Data *Nucleic Acid Conformation RNA, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*chemistry/metabolism Rats Retroviridae/*genetics Sequence Analysis}, pubstate = {published}, tppubtype = {article} } @article{, title = {A dual role of the putative RNA dimerization initiation site of human immunodeficiency virus type 1 in genomic RNA packaging and proviral DNA synthesis}, author = {J C Paillart and L Berthoux and M Ottmann and J L Darlix and R Marquet and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8970954}, isbn = {8970954}, year = {1996}, date = {1996-01-01}, journal = {J Virol}, volume = {70}, number = {12}, pages = {8348-8354}, abstract = {In retroviruses, the genomic RNA is in the form of a 60S-70S complex composed of two identical genome-length RNA molecules tightly associated through numerous interactions. A major interaction, called the dimer linkage structure, has been found near the RNA 5' end and is probably involved in the control of translation, packaging, and recombination during proviral DNA synthesis. Recently, a small sequence corresponding to a stem-loop structure located in the 5' leader of human immunodeficiency virus type 1 (HIV-1) RNA was found to be required for the initiation of HIV-1 RNA dimerization in vitro and named the dimerization initiation site (E. Skripkin, J.-C. Paillart, R. Marquet, B. Ehresmann, and C. Ehresmann, Proc. Natl. Acad. Sci. USA 91: 4945-4949, 1994). To investigate the possible role of this 5' stem-loop in HIV-1 virion formation and infectivity, four mutant viruses were generated and analyzed in vivo. Results show that deletion of the stem-loop structure reduces infectivity by a factor of 10(3) whereas loop substitutions cause a decrease of 10- to 100-fold. The level of genomic RNA packaging was found to be decreased fivefold in mutants virions containing the stem-loop deletion and only twofold in the loop-substituted virions. Surprisingly, the second DNA strand transfer during reverse transcription was found to be severely impaired upon stem-loop deletion. Taken together, these results indicate that the stem-loop structure called the dimerization initiation site is a cis element acting on both genomic RNA packaging and synthesis of proviral DNA.}, note = {0022-538x Journal Article}, keywords = {Animals COS Cells DNA, Genetic Virion *Virus Assembly, MARQUET, Non-U.S. Gov't Transcription, Nucleic Acid Support, PAILLART, Post-Translational Proviruses/genetics *RNA, Unité ARN, Viral *Regulatory Sequences, Viral HIV-1/*genetics/physiology Human Mutagenesis Protein Processing, Viral/*biosynthesis Gene Expression Genome}, pubstate = {published}, tppubtype = {article} } @article{, title = {The crystallization of biological macromolecules from precipitates: evidence for Ostwald ripening.}, author = {J D Ng and B Lorber and J Witz and A Théobald-Dietrich and D Kern and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/0022024896003624}, isbn = {10.1016/0022-0248(96)00362-4}, year = {1996}, date = {1996-01-01}, journal = {J Crystal Growth}, volume = {168}, number = {1-4}, pages = {50-62}, abstract = {Crystals were obtained by different methods under conditions where nucleation and growth occur from precipitated macromolecular material. The phenomenon was observed with compounds of different size and nature, such as thaumatin, concanavalin A, an α-amylase, a thermostable aspartyl-tRNA synthetase, the nucleo-protein complex between a tRNAAsp transcript and its cognate yeast aspartyl-tRNA synthetase, and tomato bushy stunt virus. In each system, after a rather rapid precipitation step at high supersaturation lasting one to several days, a few microcrystals appear after prolonged equilibration at constant temperature. With α-amylase, the virus and the thermostable synthetase, crystallization is accompanied by appearance of depletion zones around the growing crystals and growth of the largest crystals at the expense of the smaller ones. These features are evidences for crystal growth by Ostwald ripening. In the case of thaumatin, concanavalin A and the nucleo-protein complex, crystallization occurs by a phase transition mechanism since it is never accompanied by the disappearance of the smallest crystals. A careful analysis with thermostable aspartyl-tRNA synthetase indicates that its crystallization at 4°C under high supersaturation starts by a phase transition mechanism with the formation of small crystals within an amorphous protein precipitate. Ostwald ripening follows over a period of up to three/four months with a growth rate of about 0.8 Å/s that is 13 times slower than that of crystals growing at 20°C in the absence of precipitate without ripening. At the end of the ripening process at 4°C, only one unique synthetase crystal remains per microassay with dimensions as large as 1 mm.}, keywords = {* Protein * Virus * Crystallization * Growth kinetics * Ostwald ripening, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Visualizing the logic behind RNA self-assembly}, author = {F Michel and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8830411}, isbn = {8830411}, year = {1996}, date = {1996-01-01}, journal = {Science}, volume = {273}, number = {5282}, pages = {1676-1677}, note = {0036-8075 Comment Journal Article}, keywords = {Animals Base Composition Crystallography, Catalytic/*chemistry RNA, Protozoan/*chemistry Tetrahymena/genetics, Unité ARN, X-Ray Introns *Nucleic Acid Conformation RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {An example of non-conservation of oligomeric structure in prokaryotic aminoacyl-tRNA synthetases. Biochemical and structural properties of glycyl-tRNA synthetase from Thermus thermophilus}, author = {M H Mazauric and J Reinbolt and B Lorber and C Ebel and G Keith and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8944770}, isbn = {8944770}, year = {1996}, date = {1996-01-01}, journal = {Eur J Biochem}, volume = {241}, number = {3}, pages = {814-826}, abstract = {Glycyl-tRNA synthetase (Gly-tRNA synthetase) from Thermus thermophilus was purified to homogeneity and with high yield using a five-step purification procedure in amounts sufficient to solve its crystallographic structure [Logan, D.T., Mazauric, M.-H., Kern, D. & Moras, D. (1995) EMBO J. 14, 4156-4167]. Molecular-mass determinations of the native and denatured protein indicate an oligomeric structure of the alpha 2 type consistent with that found for eukaryotic Gly-tRNA synthetases (yeast and Bombyx mori), but different from that of Gly-tRNA synthetases from mesophilic prokaryotes (Escherichia coli and Bacillus brevis) which are alpha 2 beta 2 tetramers. N-terminal sequencing of the polypeptide chain reveals significant identity, reaching 50% with those of the eukaryotic enzymes (B. mori, Homo sapiens, yeast and Caenorhabditis elegans) but no significant identity was found with both alpha and beta chains of the prokaryotic enzymes (E. coli, Haemophilus influenzae and Coxiella burnetii) albeit the enzyme is deprived of the N-terminal extension characterizing eukaryotic synthetases. Thus, the thermophilic Gly-tRNA synthetase combines strong structural homologies of eukaryotic Gly-tRNA synthetases with a feature of prokaryotic synthetases. Heat-stability measurements show that this synthetase keeps its ATP-PPi exchange and aminoacylation activities up to 70 degrees C. Glycyladenylate strongly protects the enzyme against thermal inactivation at higher temperatures. Unexpectedly, tRNA(Gly) does not induce protection. Cross-aminoacylations reveal that the thermophilic Gly-tRNA synthetase charges heterologous E. coli tRNA(gly(GCC)) and tRNA(Gly(GCC)) and yeast tRNA(Gly(GCC)) as efficiently as T. thermophilus tRNA(Gly). All these aminoacylation reactions are characterized by similar activation energies as deduced from Arrhenius plots. Therefore, contrary to the E. coli and H. sapiens Gly-tRNA synthetases, the prokaryotic thermophilic enzyme does not possess a strict species specificity. The results are discussed in the context of the three-dimensional structure of the synthetase and in the view of the particular evolution of the glycinylation systems.}, note = {0014-2956 Journal Article}, keywords = {Amino Acid Species Specificity Substrate Specificity Support, Amino Acid Sequence Comparative Study Enzyme Stability Eukaryotic Cells Glycine-tRNA Ligase/chemistry/*isolation & purification/metabolism Heat Kinetics Molecular Sequence Data Molecular Weight Prokaryotic Cells Protein Conformation RNA, Gly/metabolism Sequence Analysis Sequence Homology, Non-U.S. Gov't Thermodynamics Thermus thermophilus/*enzymology, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selection of tRNA(Asp) amber suppressor mutants having alanine, arginine, glutamine, and lysine identity}, author = {F Martin and J Reinbolt and G Dirheimer and J Gangloff and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8809018}, isbn = {8809018}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {9}, pages = {919-927}, abstract = {Elements that confer identity to a tRNA in the cellular environment, where all aminoacyl-tRNA synthetases are competing for substrates, may be delineated by in vivo experiments using suppressor tRNAs. Here we describe the selection of active Escherichia coli tRNAAsp amber mutants and analyze their identity. Starting from a library containing randomly mutated tRNA(CUA)Asp genes, we isolated four amber suppressors presenting either lysine, alanine, or glutamine activity. Two of them, presenting mainly alanine or lysine activity, were further submitted to a second round of mutagenesis selection in order to improve their efficiency of suppression. Eleven suppressors were isolated, each containing two or three mutations. Ten presented identities of the two parental mutants, whereas one had switched from lysine to arginine identity. Analysis of the different mutants revealed (or confirmed for some nucleotides) their role as positive and/or negative determinants in AlaRS, LysRS, and ArgRS recognition. More generally, it appears that tRNAAsp presents identity characteristics closely related to those of tRNALys, as well as a structural basis for acquiring alanine or arginine identity upon moderate mutational changes; these consist of addition or suppression of the corresponding positive or negative determinants, as well as tertiary interactions. Failure to isolate aspartic acid-inserting suppressors is probably due to elimination of the important G34 identity element and its replacement by an antideterminant when changing the anticodon of the tRNAAsp to the CUA triplet.}, note = {1355-8382 Journal Article}, keywords = {Alanine/genetics Arginine/genetics Base Sequence Escherichia coli/genetics Genes, Asp/*genetics *Selection (Genetics) Support, ERIANI, Genetic, Genetic Molecular Sequence Data *Mutation RNA, Non-U.S. Gov't *Suppression, Suppressor Glutamine/genetics Lysine/genetics Models, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Localization of the dimerization initiation site of HIV-1 genomic RNA and mechanism of dimerization.}, author = {R Marquet and J C Paillart and E Skripkin and C Ehresmann and B Ehresmann}, editor = {R H Sarma and M H Sarma}, url = {http://books.google.fr/books?hl=fr&id=SOpqAAAAMAAJ&q=Marquet#search_anchor}, year = {1996}, date = {1996-01-01}, booktitle = {Biological Structure and Dynamics: Proceedings of the Ninth Conversation in the Discipline Biomolecular Stereodynamics, held at the State University of New York at Albany, June 20-24, 1995}, volume = {2}, pages = {61-72}, publisher = {Adenine Press}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Antideterminants present in minihelix(Sec) hinder its recognition by prokaryotic elongation factor Tu}, author = {J Rudinger and R Hillenbrandt and M Sprinzl and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8599948}, isbn = {8599948}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {3}, pages = {650-657}, abstract = {During protein biosynthesis, all aminoacylated elongator tRNAs except selenocysteine-inserting tRNA Sec form ternary complexes with activated elongation factor. tRNA Sec is bound by its own translation factor, an elongation factor analogue, e.g. the SELB factor in prokaryotes. An apparent reason for this discrimination could be related to the unusual length of tRNA Sec amino acid-acceptor branch formed by 13 bp. However, it has been recently shown that an aspartylated minihelix of 13 bp derived from yeast tRNA Asp is an efficient substrate for Thermus thermophilus EF-Tu-GTP, suggesting that features other than the length of tRNA Sec prevent its recognition by EF-Tu-GTP. A stepwise mutational analysis of a minihelix derived from tRNA Sec in which sequence elements of tRNA Asp were introduced showed that the sequence of the amino acid- acceptor branch of Escherichia coli tRNA Sec contains a specific structural element that hinders its binding to T.thermophilus EF-Tu-GTP. This antideterminant is located in the 8th, 9th and 10th bp in the acceptor branch of tRNA Sec, corresponding to the last base pair in the amino acid acceptor stem and the two first pairs in the T-stem. The function of this C7.G66/G49.U65/C50.G64 box was tested by its transplantation into a minihelix derived from tRNA Asp, abolishing its recognition by EF-Tu-GTP. The specific role of this nucleotide combination is further supported by its absence in all known prokaryotic elongator tRNAs.}, note = {0261-4189 Journal Article}, keywords = {Amino Acid-Specific/*chemistry/genetics/*metabolism RNA, Antisense/chemistry/genetics/metabolism RNA, Asp/chemistry/genetics/metabolism Saccharomyces cerevisiae/genetics/metabolism Support, Base Sequence Escherichia coli/genetics/metabolism Evolution Guanosine Triphosphate/metabolism Molecular Sequence Data Nucleic Acid Conformation Peptide Elongation Factor Tu/*metabolism RNA, Non-U.S. Gov't Thermus thermophilus/metabolism, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The expression of E.coli threonyl-tRNA synthetase is regulated at the translational level by symmetrical operator-repressor interactions}, author = {P Romby and J Caillet and C Ebel and C Sacerdot and M Graffe and F Eyermann and C Brunel and H Moine and C Ehresmann and B Ehresmann and M Springer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8918475}, isbn = {8918475}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {21}, pages = {5976-5987}, abstract = {Threonyl-tRNA synthetase from Escherichia coli represses the translation of its own mRNA by binding to the operator region located upstream from the ribosome binding site. The operator contains two stemloop structures which interact specifically with the homodimeric enzyme. Here, we provide in vitro and in vivo evidence that these two stem-loop structures are recognized by the enzyme in an analogous way and mimic the anticodon arm of E.coli tRNA(Thr). Determination of the stoichiometry of the different RNA-threonyl-tRNA synthetase complexes reveals that two tRNA(Thr) molecules bind to the enzyme whereas only one thrS operator interacts with the homodimeric enzyme. A model is presented in which the two anticodon-like domains of the operator bind symmetrically to the two tRNA(Thr) anticodon recognition sites (one per subunit) of the dimeric threonyl-tRNA synthetase. Although symmetrical operator-repressor interactions in transcriptional control are widespread, this report stresses the importance of such interactions in translational regulation of gene expression.}, note = {0261-4189 Journal Article}, keywords = {Anticodon Base Sequence Binding Sites Binding, Bacterial Gene Expression Regulation, Bacterial/chemistry/genetics/metabolism RNA, Biological Molecular Sequence Data Mutagenesis, Competitive Escherichia coli/*enzymology/*genetics/metabolism Gene Expression Regulation, Enzymologic Models, Genetic, Messenger/genetics/metabolism Repressor Proteins/genetics Support, Non-U.S. Gov't Threonine-tRNA Ligase/*genetics Translation, ROMBY, Site-Directed Nucleic Acid Conformation Operator Regions (Genetics) RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {2'-O-methyl-5-formylcytidine (f5Cm), a new modified nucleotide at the 'wobble' of two cytoplasmic tRNAs Leu (NAA) from bovine liver}, author = {J P Pais de Barros and G Keith and C El Adlouni and A L Glasser and G Mack and G Dirheimer and J Desgres}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8628682}, isbn = {8628682}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {8}, pages = {1489-1496}, abstract = {The nucleotide analysis of a cytoplasmic tRNA(Leu) isolated from bovine liver revealed the presence of an unknown modified nucleotide N. The corresponding N nucleoside was isolated by different enzymatic and chromatographic protocols from a partially purified preparation of this tRNA(Leu). Its chemical characterization was determined from its chromatographic properties, UV-absorption spectroscopy and mass spectrometric measurements, as well as from those of the borohydride reduced N nucleoside and its etheno-trimethylsilyl derivative. The structure of N was established as 2'-O-methyl-5-formylcytidine (f5CM), and its reduced derivative as 2'-O-methyl-5-hydroxy-methylcytidine (om5Cm). By sequencing the bovine liver tRNA(Leu), the structure of the anticodon was determined as f5CmAA. In addition, the nucleotide sequence showed two primary structures differing only by the nucleotide 47c which is either uridine or adenosine. The two slightly differing bovine liver tRNAs-Leu(f5CmAA) are the only tRNAs so far sequenced which contain f5Cm. The role of such a modified cytidine at the first position of the anticodon is discussed in terms of decoding properties for the UUG and UUA leucine codons. Recently, precise evidence was obtained for the presence of f5Cm at the same position in tRNAs(Leu)(NAA) isolated from rabbit and lamb liver. Therefore, the 2'-O-methyl-5-formyl modification of cytidine at position 34 could be a general feature of cytoplasmic tRNAs(Leu)(NAA) in mammals.}, note = {0305-1048 Journal Article}, keywords = {Amino Acyl/*chemistry/isolation & purification Support, Animals Base Sequence Borohydrides/chemistry Cattle Cytidine/*analogs & derivatives/chemistry/isolation & purification Cytoplasm Hela Cells Human Liver/*chemistry Mass Fragmentography Molecular Sequence Data Molecular Structure Nucleic Acid Conformation RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A loop-loop "kissing" complex is the essential part of the dimer linkage of genomic HIV-1 RNA}, author = {J C Paillart and E Skripkin and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8643617}, isbn = {8643617}, year = {1996}, date = {1996-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {93}, number = {11}, pages = {5572-5577}, abstract = {RNA-RNA interactions govern a number of biological processes. Several RNAs, including natural sense and antisense RNAs, interact by means of a two-step mechanism: recognition is mediated by a loop-loop complex, which is then stabilized by formation of an extended intermolecular duplex. It was proposed that the same mechanism holds for dimerization of the genomic RNA of human immunodeficiency virus type 1 (HIV-1), an event thought to control crucial steps of HIV-1 replication. However, whereas interaction between the partially self-complementary loop of the dimerization initiation site (DIS) of each monomer is well established, formation of the extended duplex remained speculative. Here we first show that in vitro dimerization of HIV-1 RNA is a specific process, not resulting from simple annealing of denatured molecules. Next we used mutants of the DIS to test the formation of the extended duplex. Four pairs of transcomplementary mutants were designed in such a way that all pairs can form the loop-loop "kissing" complex, but only two of them can potentially form the extended duplex. All pairs of mutants form heterodimers whose thermal stability, dissociation constant, and dynamics were analyzed. Taken together, our results indicate that, in contrast with the interactions between natural sense and antisense RNAs, no extended duplex is formed during dimerization of HIV-1 RNA. We also showed that 55-mer sense RNAs containing the DIS are able to interfere with the preformed HIV-1 RNA dimer.}, note = {0027-8424 Journal Article}, keywords = {Base Composition Base Sequence HIV-1/*genetics Heat Human Kinetics Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Nucleic Acid Denaturation Plasmids RNA, Genetic, MARQUET, Non-U.S. Gov't Thermodynamics Transcription, PAILLART, Unité ARN, Viral/biosynthesis/*chemistry/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {The use of chemical modification interference and inverse PCR mutagenesis to identify the dimerization initiation site of HIV-1 genomic RNA}, author = {J C Paillart and E Skripkin and B Ehresmann and C Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8786995}, isbn = {8786995}, year = {1996}, date = {1996-01-01}, journal = {Pharm Acta Helv}, volume = {71}, number = {1}, pages = {21-28}, abstract = {The retroviral genome consists of two identical RNA molecules physically linked together close to their 5' end, in a region called the Dimer Linkage Structure (DLS). Recent findings suggest that dimerization is involved in encapsidation, regulation of translation and reverse transcription. Previous in vitro studies localized the DLS of HIV-1 in a region downstream of the splice donor (SD) site. More recently, we showed that dimerization of HIV-1 RNA also involves sequences upstream of the SD site. Modification interference experiments and site-directed mutagenesis were used to identify the nucleotides required in the dimerization process of HIV-1 RNA. Our results point out a self-complementary sequence located in a hairpin loop, between the Primer Binding Site (PBS) and the SD site, as the Dimerization Initiation Site.}, note = {0031-6865 Journal Article}, keywords = {Base Sequence *Genome, MARQUET, Non-U.S. Gov't, PAILLART, Site-Directed Polymerase Chain Reaction RNA, Unité ARN, Viral HIV-1/*chemistry Human Molecular Sequence Data Mutagenesis, Viral/*chemistry Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dimerization of retroviral genomic RNAs: structural and functional implications}, author = {J C Paillart and R Marquet and E Skripkin and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955907}, isbn = {8955907}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {639-653}, abstract = {Retroviruses are a family of widespread small animal viruses at the origin of a diversity of diseases. They share common structural and functional properties such as reverse transcription of their RNA genome and integration of the proviral DNA into the host genome, and have the particularity of packaging a diploid genome. The genome of all retroviruses is composed of two homologous RNA molecules that are non-covalently linked near their 5' end in a region called the dimer linkage structure (DLS). There is now considerable evidence that a specific site (or sites) in the 5' leader region of all retroviruses, located either upstream or/and downstream of the major splice donor site, is involved in the dimer linkage. For MoMuLV and especially HIV-1, it was shown that dimerization is initiated at a stem-loop structure named the dimerization initiation site (DIS). The DIS of HIV-1 and related regions in other retroviruses corresponds to a highly conserved structure with a self-complementary loop sequence, that is involved in a typical loop-loop 'kissing' complex which can be further stabilized by long distance interactions or by conformational rearrangements. RNA interactions involved in the viral RNA dimer were postulated to regulate several key steps in retroviral cycle, such as: i) translation and encapsidation: the arrest of gag translation imposed by the highly structured DLS-encapsidation signal would leave the RNA genome available for the encapsidation machinery; and ii) recombination during reverse transcription: the presence of two RNA molecules in particles would be necessary for variability and viability of virus progeny and the ordered structure imposed by the DLS would be required for efficient reverse transcription.}, note = {0300-9084 Journal Article Review Review, Academic}, keywords = {Animals Base Sequence HIV-1/genetics Human Microscopy, DNA Support, Electron Molecular Sequence Data *Nucleic Acid Conformation RNA, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*chemistry/metabolism Rats Retroviridae/*genetics Sequence Analysis}, pubstate = {published}, tppubtype = {article} } @article{, title = {A dual role of the putative RNA dimerization initiation site of human immunodeficiency virus type 1 in genomic RNA packaging and proviral DNA synthesis}, author = {J C Paillart and L Berthoux and M Ottmann and J L Darlix and R Marquet and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8970954}, isbn = {8970954}, year = {1996}, date = {1996-01-01}, journal = {J Virol}, volume = {70}, number = {12}, pages = {8348-8354}, abstract = {In retroviruses, the genomic RNA is in the form of a 60S-70S complex composed of two identical genome-length RNA molecules tightly associated through numerous interactions. A major interaction, called the dimer linkage structure, has been found near the RNA 5' end and is probably involved in the control of translation, packaging, and recombination during proviral DNA synthesis. Recently, a small sequence corresponding to a stem-loop structure located in the 5' leader of human immunodeficiency virus type 1 (HIV-1) RNA was found to be required for the initiation of HIV-1 RNA dimerization in vitro and named the dimerization initiation site (E. Skripkin, J.-C. Paillart, R. Marquet, B. Ehresmann, and C. Ehresmann, Proc. Natl. Acad. Sci. USA 91: 4945-4949, 1994). To investigate the possible role of this 5' stem-loop in HIV-1 virion formation and infectivity, four mutant viruses were generated and analyzed in vivo. Results show that deletion of the stem-loop structure reduces infectivity by a factor of 10(3) whereas loop substitutions cause a decrease of 10- to 100-fold. The level of genomic RNA packaging was found to be decreased fivefold in mutants virions containing the stem-loop deletion and only twofold in the loop-substituted virions. Surprisingly, the second DNA strand transfer during reverse transcription was found to be severely impaired upon stem-loop deletion. Taken together, these results indicate that the stem-loop structure called the dimerization initiation site is a cis element acting on both genomic RNA packaging and synthesis of proviral DNA.}, note = {0022-538x Journal Article}, keywords = {Animals COS Cells DNA, Genetic Virion *Virus Assembly, MARQUET, Non-U.S. Gov't Transcription, Nucleic Acid Support, PAILLART, Post-Translational Proviruses/genetics *RNA, Unité ARN, Viral *Regulatory Sequences, Viral HIV-1/*genetics/physiology Human Mutagenesis Protein Processing, Viral/*biosynthesis Gene Expression Genome}, pubstate = {published}, tppubtype = {article} } @article{, title = {The crystallization of biological macromolecules from precipitates: evidence for Ostwald ripening.}, author = {J D Ng and B Lorber and J Witz and A Théobald-Dietrich and D Kern and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/0022024896003624}, isbn = {10.1016/0022-0248(96)00362-4}, year = {1996}, date = {1996-01-01}, journal = {J Crystal Growth}, volume = {168}, number = {1-4}, pages = {50-62}, abstract = {Crystals were obtained by different methods under conditions where nucleation and growth occur from precipitated macromolecular material. The phenomenon was observed with compounds of different size and nature, such as thaumatin, concanavalin A, an α-amylase, a thermostable aspartyl-tRNA synthetase, the nucleo-protein complex between a tRNAAsp transcript and its cognate yeast aspartyl-tRNA synthetase, and tomato bushy stunt virus. In each system, after a rather rapid precipitation step at high supersaturation lasting one to several days, a few microcrystals appear after prolonged equilibration at constant temperature. With α-amylase, the virus and the thermostable synthetase, crystallization is accompanied by appearance of depletion zones around the growing crystals and growth of the largest crystals at the expense of the smaller ones. These features are evidences for crystal growth by Ostwald ripening. In the case of thaumatin, concanavalin A and the nucleo-protein complex, crystallization occurs by a phase transition mechanism since it is never accompanied by the disappearance of the smallest crystals. A careful analysis with thermostable aspartyl-tRNA synthetase indicates that its crystallization at 4°C under high supersaturation starts by a phase transition mechanism with the formation of small crystals within an amorphous protein precipitate. Ostwald ripening follows over a period of up to three/four months with a growth rate of about 0.8 Å/s that is 13 times slower than that of crystals growing at 20°C in the absence of precipitate without ripening. At the end of the ripening process at 4°C, only one unique synthetase crystal remains per microassay with dimensions as large as 1 mm.}, keywords = {* Protein * Virus * Crystallization * Growth kinetics * Ostwald ripening, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Visualizing the logic behind RNA self-assembly}, author = {F Michel and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8830411}, isbn = {8830411}, year = {1996}, date = {1996-01-01}, journal = {Science}, volume = {273}, number = {5282}, pages = {1676-1677}, note = {0036-8075 Comment Journal Article}, keywords = {Animals Base Composition Crystallography, Catalytic/*chemistry RNA, Protozoan/*chemistry Tetrahymena/genetics, Unité ARN, X-Ray Introns *Nucleic Acid Conformation RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {An example of non-conservation of oligomeric structure in prokaryotic aminoacyl-tRNA synthetases. Biochemical and structural properties of glycyl-tRNA synthetase from Thermus thermophilus}, author = {M H Mazauric and J Reinbolt and B Lorber and C Ebel and G Keith and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8944770}, isbn = {8944770}, year = {1996}, date = {1996-01-01}, journal = {Eur J Biochem}, volume = {241}, number = {3}, pages = {814-826}, abstract = {Glycyl-tRNA synthetase (Gly-tRNA synthetase) from Thermus thermophilus was purified to homogeneity and with high yield using a five-step purification procedure in amounts sufficient to solve its crystallographic structure [Logan, D.T., Mazauric, M.-H., Kern, D. & Moras, D. (1995) EMBO J. 14, 4156-4167]. Molecular-mass determinations of the native and denatured protein indicate an oligomeric structure of the alpha 2 type consistent with that found for eukaryotic Gly-tRNA synthetases (yeast and Bombyx mori), but different from that of Gly-tRNA synthetases from mesophilic prokaryotes (Escherichia coli and Bacillus brevis) which are alpha 2 beta 2 tetramers. N-terminal sequencing of the polypeptide chain reveals significant identity, reaching 50% with those of the eukaryotic enzymes (B. mori, Homo sapiens, yeast and Caenorhabditis elegans) but no significant identity was found with both alpha and beta chains of the prokaryotic enzymes (E. coli, Haemophilus influenzae and Coxiella burnetii) albeit the enzyme is deprived of the N-terminal extension characterizing eukaryotic synthetases. Thus, the thermophilic Gly-tRNA synthetase combines strong structural homologies of eukaryotic Gly-tRNA synthetases with a feature of prokaryotic synthetases. Heat-stability measurements show that this synthetase keeps its ATP-PPi exchange and aminoacylation activities up to 70 degrees C. Glycyladenylate strongly protects the enzyme against thermal inactivation at higher temperatures. Unexpectedly, tRNA(Gly) does not induce protection. Cross-aminoacylations reveal that the thermophilic Gly-tRNA synthetase charges heterologous E. coli tRNA(gly(GCC)) and tRNA(Gly(GCC)) and yeast tRNA(Gly(GCC)) as efficiently as T. thermophilus tRNA(Gly). All these aminoacylation reactions are characterized by similar activation energies as deduced from Arrhenius plots. Therefore, contrary to the E. coli and H. sapiens Gly-tRNA synthetases, the prokaryotic thermophilic enzyme does not possess a strict species specificity. The results are discussed in the context of the three-dimensional structure of the synthetase and in the view of the particular evolution of the glycinylation systems.}, note = {0014-2956 Journal Article}, keywords = {Amino Acid Species Specificity Substrate Specificity Support, Amino Acid Sequence Comparative Study Enzyme Stability Eukaryotic Cells Glycine-tRNA Ligase/chemistry/*isolation & purification/metabolism Heat Kinetics Molecular Sequence Data Molecular Weight Prokaryotic Cells Protein Conformation RNA, Gly/metabolism Sequence Analysis Sequence Homology, Non-U.S. Gov't Thermodynamics Thermus thermophilus/*enzymology, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selection of tRNA(Asp) amber suppressor mutants having alanine, arginine, glutamine, and lysine identity}, author = {F Martin and J Reinbolt and G Dirheimer and J Gangloff and G Eriani}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8809018}, isbn = {8809018}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {9}, pages = {919-927}, abstract = {Elements that confer identity to a tRNA in the cellular environment, where all aminoacyl-tRNA synthetases are competing for substrates, may be delineated by in vivo experiments using suppressor tRNAs. Here we describe the selection of active Escherichia coli tRNAAsp amber mutants and analyze their identity. Starting from a library containing randomly mutated tRNA(CUA)Asp genes, we isolated four amber suppressors presenting either lysine, alanine, or glutamine activity. Two of them, presenting mainly alanine or lysine activity, were further submitted to a second round of mutagenesis selection in order to improve their efficiency of suppression. Eleven suppressors were isolated, each containing two or three mutations. Ten presented identities of the two parental mutants, whereas one had switched from lysine to arginine identity. Analysis of the different mutants revealed (or confirmed for some nucleotides) their role as positive and/or negative determinants in AlaRS, LysRS, and ArgRS recognition. More generally, it appears that tRNAAsp presents identity characteristics closely related to those of tRNALys, as well as a structural basis for acquiring alanine or arginine identity upon moderate mutational changes; these consist of addition or suppression of the corresponding positive or negative determinants, as well as tertiary interactions. Failure to isolate aspartic acid-inserting suppressors is probably due to elimination of the important G34 identity element and its replacement by an antideterminant when changing the anticodon of the tRNAAsp to the CUA triplet.}, note = {1355-8382 Journal Article}, keywords = {Alanine/genetics Arginine/genetics Base Sequence Escherichia coli/genetics Genes, Asp/*genetics *Selection (Genetics) Support, ERIANI, Genetic, Genetic Molecular Sequence Data *Mutation RNA, Non-U.S. Gov't *Suppression, Suppressor Glutamine/genetics Lysine/genetics Models, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Localization of the dimerization initiation site of HIV-1 genomic RNA and mechanism of dimerization.}, author = {R Marquet and J C Paillart and E Skripkin and C Ehresmann and B Ehresmann}, editor = {R H Sarma and M H Sarma}, url = {http://books.google.fr/books?hl=fr&id=SOpqAAAAMAAJ&q=Marquet#search_anchor}, year = {1996}, date = {1996-01-01}, booktitle = {Biological Structure and Dynamics: Proceedings of the Ninth Conversation in the Discipline Biomolecular Stereodynamics, held at the State University of New York at Albany, June 20-24, 1995}, volume = {2}, pages = {61-72}, publisher = {Adenine Press}, keywords = {MARQUET, PAILLART, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {L'initiation de la transcription inverse du HIV-1: spécificités structurales et fonctionelles.}, author = {R Marquet}, url = {http://cat.inist.fr/?aModele=afficheN&cpsidt=2524109}, issn = {0336-1640}, year = {1996}, date = {1996-01-01}, journal = {Regard sur la Biochimie}, volume = {4}, pages = {62-69}, keywords = {MARQUET, Unité ARN, Virus HIV1 Transcription inverse Initiation transcription Article synthèse Virus immunodéficience humaine Lentivirinae Retroviridae Virus}, pubstate = {published}, tppubtype = {article} } @article{, title = {An antisense/target RNA duplex or a strong intramolecular RNA structure 5' of a translation initiation signal blocks ribosome binding: the case of plasmid R1}, author = {C Malmgren and H M Engdahl and P Romby and E G Wagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8849778}, isbn = {8849778}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {10}, pages = {1022-1032}, abstract = {Antisense RNAs in prokaryotic systems often inhibit translation of mRNAs. In some cases, this involves sequestration of Shine-Dalgarno (SD) sequences and start codons. In other cases, antisense/target RNA duplexes do not overlap these signals, but form upstream. We have performed toeprinting analyses on repA mRNA of plasmid R1, both free and in duplex with the antisense RNA, CopA. An intermolecular RNA duplex 2 nt upstream of the tap SD prevents ribosome binding. An intrastrand stem-loop at this location yields the same inhibition. Thus, stable secondary structures immediately upstream of the tap SD sequence inhibit translation, as shown by toeprinting in vitro and repA-lacZ expression in vivo. Previous work showed that repA (initiator protein) expression requires tap (leader peptide) translation. Toeprinting data confirm that the tap ribosome binding site (RBS) is accessible, whereas the repA RBS, which is sequestered by a stable stem-loop, is weakly recognized by the ribosome. Truncated CopA RNA (CopI) is unable to pair completely with target RNA, but proceeds normally to a kissing intermediate. This mutant RNA species inhibits repA expression in vivo. By a kinetic toeprint inhibition protocol, we have shown that the structure of the kissing complex is sufficient to sterically prevent ribosome binding. These results are discussed in comparison with the effect of RNA structures elsewhere in the ribosome-binding region of an mRNA.}, note = {1355-8382 Journal Article}, keywords = {Antisense/chemistry/*metabolism RNA, Bacterial Proteins/genetics Base Sequence Escherichia coli/genetics Genetic Techniques Kinetics Molecular Sequence Data Mutation *Nucleic Acid Conformation Peptide Chain Initiation/*genetics Protein Sorting Signals/genetics *Proteins R Factors/*chemistry/genetics RNA, Bacterial/chemistry/metabolism RNA, Messenger/chemistry/*metabolism Ribosomes/*metabolism Support, Non-U.S. Gov't, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {RNA structure from molecular dynamics simulations}, author = {S Louise-May and P Auffinger and E Westhof}, editor = {R H Sarma and M H Sarma}, url = {http://books.google.fr/books/about/Biological_structure_and_dynamics.html?id=SepqAAAAMAAJ&redir_esc=y}, year = {1996}, date = {1996-01-01}, booktitle = {Biological Structure and Dynamics: Proceedings of the Ninth Conversation in the Discipline Biomolecular Stereodynamics, held at the State University of New York at Albany, June 20-24, 1995}, volume = {2}, pages = {73-90}, publisher = {Adenine Press}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Calculations of nucleic acid conformations}, author = {S Louise-May and P Auffinger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8804831}, isbn = {8804831}, year = {1996}, date = {1996-01-01}, journal = {Curr Opin Struct Biol}, volume = {6}, number = {3}, pages = {289-298}, abstract = {The present computational power and sophistication of theoretical approaches to nucleic acid structural investigation are sufficient for the realization of static and dynamic models that correlate accurately with current crystallographic, NMR and solution-probing structural data, and consequently are able to provide valuable insights and predictions for a variety of nucleic acid conformational families. In molecular dynamics simulations, the year 1995 was marked by the foray of fast Ewald methods, an accomplishment resulting from several years' work in the search for an adequate treatment of the electrostatic long-range forces so primordial in nucleic acid behavior. In very large systems, and particularly in the RNA-folding field, techniques originating from artificial intelligence research, like constraint satisfaction programming or genetic algorithms, have established their utility and potential.}, note = {0959-440x Journal Article Review Review, Tutorial}, keywords = {DNA/chemistry DNA, Molecular *Nucleic Acid Conformation Nucleic Acid Hybridization Nucleosides/chemistry Protein Binding RNA/chemistry Telomere/chemistry, Superhelical/chemistry *Genetic Techniques Models, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effect of high hydrostatic pressure on nucleation and growth of protein crystals}, author = {B Lorber and G Jenner and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/0022024895003991}, isbn = {10.1016/0022-0248(95)00399-1}, year = {1996}, date = {1996-01-01}, journal = {J Crystal Growth}, volume = {158}, number = {1-2}, pages = {103-117}, abstract = {The influence of hydrostatic pressure on the nucleation and growth of protein crystals was studied. A micromethod was developed to establish a solubility phase diagram of hen egg-white lysozyme as a function of pressure and protein concentration. The pressure dependence of the formation of canonical tetragonal crystals was investigated at different precipitating agent and protein concentrations (in the range 0.6–1.2M NaCl and 10–35 mg/ml lysozyme). The apparent protein solubility significantly increases when pressure is raised from 0.1 MPa (atmospheric pressure) to 250 MPa. With an increase in pressure, the size and number of lysozyme crystals decline and a transition to urchin-like particles made of crystalline needles progressively occurs. The shape of tetragonal crystals becomes more elongated in a limited region of the phase diagram as indicated by the ratio of the lengths of the (110) and (101) faces. Single tetragonal crystals grown under high pressure diffract X-rays at high resolution. They belong to the same space group and have identical cell parameters as control crystals grown at atmospheric pressure. Changes in solubility and crystallizability are explained by pressure-induced minor reversible alterations in the protein structure.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Containerless protein crystallization in floating drops: application to crystal growth monitoring under reduced nucleation conditions}, author = {B Lorber and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/0022024896003569}, isbn = {10.1016/0022-0248(96)00356-9}, year = {1996}, date = {1996-01-01}, journal = {J Crystal Growth}, volume = {168}, number = {1-4}, pages = {204-215}, abstract = {A micromethod was developed for the batch crystallization of proteins under conditions were the solution has no contact with the container walls. Drops of crystallization solutions (5 to 100 μl) are placed at the interface between two layers of inert and non-miscible silicone fluids contained in square glass or plastic cuvettes. The densities of the fluids are either lower or higher than those of the major precipitating agents of macromolecules, including aqueous solutions containing salts, polyethylene glycols or alcohols. Several proteins and a spherical plant virus were crystallized in the temperature range 4°C–20°C using this set-up. A thermostated device was built for the dynamic control of the temperature of crystallization drops and the monitoring of crystal growth by video-microscopy. In all cases, the habit of the crystals grown in floating drops are identical to those of controls grown in sealed glass tubes without silicone fluid. The comparison of the number of crystals in drops kept under one layer of fluid and in floating drops of the same volume indicates that heterogeneous nucleation is minimized when protein crystallization is performed in floating drops. The advantages and limitations of this novel containerless crystallization method are discussed.}, keywords = {* Protein crystallization * Heterogeneous nucleation * Growth kinetics * Instrumentation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structure of 4.5S RNA in the signal recognition particle of Escherichia coli as studied by enzymatic and chemical probing}, author = {G Lentzen and H Moine and C Ehresmann and B Ehresmann and W Wintermeyer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8608448}, isbn = {8608448}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {3}, pages = {244-253}, abstract = {The structure of 4.5S RNA, the Escherichia coli homologue of the signal recognition particle (SRP) RNA, alone and in the SRP complex with protein P48 (Ffh) was probed both enzymatically and chemically. The molecule is largely resistant against single strand-specific nucleases, indicating a highly base paired structure. Reactivity appears mainly in the apical tetraloop and in one of the conserved internal loops. Although some residues are found reactive toward dimethylsulphate and kethoxal in regions predicted to be unpaired by the phylogenetic secondary structure model of 4.5S RNA, generally the reactivity is low, and some residues in internal loops are not reactive at all. RNase V1 cleaves the RNA at multiple sites that coincide with predicted helices, although the cleavages show a pronounced asymmetry. The binding of protein P48 to 4.5S RNA results in a protection of residues in the apical part of the molecule homologous to eukaryotic SRP RNA (domain IV), whereas the cleavages in the conserved apical tetraloop are not protected. Hydroxyl radical treatment reveals an asymmetric pattern of backbone reactivity; in particular, the region encompassing nucleotides 60-82, i.e., the 3' part of the conserved domain IV, is protected. The data suggest that a bend in the domain IV region, most likely at the central asymmetric internal loop, is an important element of the tertiary structure of 4.5S RNA. Hyperchromicity and lead cleavage data are consistent with the model as they reveal the unfolding of a higher-order structure between 30 and 40 degrees C. Protection by protein P48 occurs in this region of the RNA and, more strongly, in the 5' part of domain IV (nt 26-50, most strongly from 35 to 49). It is likely that P48 binds to the outside of the bent form of 4.5S RNA.}, note = {1355-8382 Journal Article}, keywords = {Bacterial Proteins/chemistry/metabolism Base Sequence Escherichia coli/*chemistry *Escherichia coli Proteins Hydroxyl Radical Lead Molecular Sequence Data *Nucleic Acid Conformation Nucleic Acid Denaturation RNA, Bacterial/*chemistry/metabolism Ribonucleases Signal Recognition Particle/*chemistry/metabolism Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {New loop-loop tertiary interactions in self-splicing introns of subgroup IC and ID: a complete 3D model of the Tetrahymena thermophila ribozyme}, author = {V Lehnert and L Jaeger and F Michel and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9000010}, isbn = {9000010}, year = {1996}, date = {1996-01-01}, journal = {Chem Biol}, volume = {3}, number = {12}, pages = {993-1009}, abstract = {BACKGROUND: Group I introns self-splice via two consecutive trans-esterification reactions in the presence of guanosine cofactor and magnesium ions. Comparative sequence analysis has established that a catalytic core of about 120 nucleotides is conserved in all known group I introns. This core is generally not sufficient for activity, however, and most self-splicing group I introns require non-conserved peripheral elements to stabilize the complete three-dimensional (3D) structure. The physico-chemical properties of group I introns make them excellent systems for unraveling the structural basis of the RNA-RNA interactions responsible for promoting the self-assembly of complex RNAs. RESULTS: We present phylogenetic and experimental evidence for the existence of three additional tertiary base pairings between hairpin loops within peripheral components of subgroup IC1 and ID introns. Each of these new long range interactions, called P13, P14 and P16, involves a terminal loop located in domain 2. Although domains 2 of IC and ID introns share very strong sequence similarity, their terminal loops interact with domains 5 and 9 (subgroup IC1) and domain 6 (subgroup ID). Based on these tertiary contacts, comparative sequence analysis, and published experimental results such as Fe(II)-EDTA protection patterns, we propose 3D models for two entire group I introns, the subgroup IC1 intron in the large ribosomal precursor RNA of Tetrahymena thermophila and the SdCob.1 subgroup ID intron found in the cytochrome b gene of Saccharomyces douglasii. CONCLUSIONS: Three-dimensional models of group I introns belonging to four different subgroups are now available. They all emphasize the modular and hierarchical organization of the architecture of group I introns and the widespread use of base-pairings between terminal hairpin loops for stabilizing the folded and active structures of large and complex RNA molecules.}, note = {1074-5521 Journal Article}, keywords = {Animals Base Composition Base Sequence Electrophoresis, Catalytic/*chemistry Sequence Alignment Support, Molecular Molecular Sequence Data Mutation/genetics Nucleic Acid Conformation RNA/chemistry/metabolism RNA Splicing/*genetics RNA, Non-U.S. Gov't Tetrahymena thermophila/*metabolism, Polyacrylamide Gel Ferrous Compounds/metabolism/pharmacology Introns/*genetics Models, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural and functional evidence that initiation and elongation of HIV-1 reverse transcription are distinct processes}, author = {J M Lanchy and C Isel and C Ehresmann and R Marquet and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9150889}, isbn = {9150889}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {11-12}, pages = {1087-1096}, abstract = {Retroviral reverse transcription starts with the extension of a cellular tRNA primer bound near the 5' end of the viral genomic RNA at a site called the primer binding site (PBS). Formation of the HIV-1 initiation complex between tRNA3(Lys), viral RNA and reverse transcriptase probably occurs during encapsidation of these components. tRNA3(Lys) is thought to be selectively packaged by interaction with the reverse transcriptase domain of the Pr160Gag-Pol precursor protein, then annealed to the PBS of viral RNA with the help of the nucleocapsid protein. tRNA3(Lys) and HIV-1 viral RNA form a highly-structured complex, with extended interactions between the two molecules. Two different modes of reverse transcription have been distinguished: initiation, a tRNA3(Lys)-specific and distributive mode of polymerization corresponding to the addition of the first five nucleotides, followed by elongation, a non-specific and processive mode of DNA synthesis. These two modes are reminiscent of the initiation and elongation processes previously observed with DNA-dependent RNA polymerases.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Amino Acyl/*biosynthesis/*chemistry RNA, Base Sequence Comparative Study HIV-1/*genetics/*metabolism HIV-1 Reverse Transcriptase/*metabolism Human Molecular Sequence Data *Nucleic Acid Conformation RNA, Genetic, MARQUET, Non-U.S. Gov't *Transcription, Transfer, Unité ARN, Viral/biosynthesis/chemistry Retroviridae/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding and kinetic properties of HIV-1 reverse transcriptase markedly differ during initiation and elongation of reverse transcription}, author = {J M Lanchy and C Ehresmann and S F Le Grice and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9003793}, isbn = {9003793}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {24}, pages = {7178-7187}, abstract = {We recently showed that primer tRNA3Lys, human immunodeficiency virus type 1 (HIV-1) RNA and HIV-1 reverse transcriptase (RT) form a specific complex of initiation of reverse transcription that can be functionally distinguished from the elongation complex, which can be obtained by substituting an 18mer oligodeoxyribonucleotide (ODN) for the natural primer (Isel et al., 1996). Here, we compared the binding properties and the single and multiple turnover kinetics of HIV-1 RT in the initiation and elongation complexes. Even though the equilibrium dissociation constants of HIV-1 RT are not very different for the two complexes, RT dissociates approximately 200-fold faster from the initiation complex. Furthermore, nucleotide incorporation by the pre-formed primer-template-RT complexes is reduced by a approximately 50-fold factor during initiation of reverse transcription, compared with elongation. As a consequence, processivity of HIV-1 RT in the initiation complex is close to unity, while it increases by four orders of magnitude during elongation, as expected for a replication enzyme. This processivity change is reminiscent of the transition from initiation to elongation of transcription. Furthermore, our results indicate that the post-transcriptional modifications of tRNA3Lys play a role similar to that of the sigma factor in transcription by the Escherichia coli RNA polymerase: they favour the formation of the specific initiation complex but do not affect the polymerization rate of the bound enzyme.}, note = {0261-4189 Journal Article}, keywords = {DNA Primers HIV-1/*enzymology HIV-1 Reverse Transcriptase/*metabolism Kinetics Protein Binding Support, Genetic, Genetic *Transcription, MARQUET, Non-U.S. Gov't Support, P.H.S. Templates, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Function of a pseudoknot in the suppression of an alternative splicing event in a group I intron}, author = {L Jaeger and E Westhof and F Michel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8915536}, isbn = {8915536}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {6}, pages = {466-473}, abstract = {Like most mitochondrial group I introns with a free-standing open reading frame (ORF) located downstream of their catalytic core, the Sd.cob, 1 intron in the gene coding for the cytochrome b of Saccharomyces douglasii mitochondria possesses a putative proximal 3' splice site. However, incubation of Sd.cob, 1 preRNA transcripts under optimal in vitro splicing conditions essentially results in splicing at the authentic, distal 3' splice junction. The mechanism by which the proximal splicing event is suppressed in vitro involves formation of a tertiary interaction which is only found in the Sd.cob, 1 intron. Core nucleotides located in loop L5 block proximal splicing by forming Watson-Crick base pairs with the nucleotide sequence of the proximal 3' splice site. This tertiary base pairing, also important for the folding of the intron into an active conformation, may be regarded as equivalent to the L9/P5, GNRA-loop/helix interaction found in more than one-third of known group I introns.}, note = {0300-9084 Journal Article}, keywords = {Alternative Splicing/*genetics Base Sequence Cytochrome b Group/genetics Electrophoresis, Genetic/genetics, Genetic/genetics Transcription, Molecular Molecular Sequence Data Mutation/genetics *Nucleic Acid Conformation RNA Precursors/metabolism RNA Splicing/genetics Saccharomyces/*metabolism Suppression, Polyacrylamide Gel Introns/genetics Mitochondria/metabolism Models, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {The structure of group I ribozymes}, author = {L Jaeger and F Michel and E Westhof}, editor = {F Eckstein and M J Lilley}, url = {http://books.google.fr/books/about/Catalytic_RNA.html?id=_n4TAQAAMAAJ&redir_esc=y}, year = {1996}, date = {1996-01-01}, booktitle = {Catalytic RNA (Nucleic Acids and Molecular Biology)}, volume = {10}, pages = {33-51}, publisher = {Springer}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Specific initiation and switch to elongation of human immunodeficiency virus type 1 reverse transcription require the post-transcriptional modifications of primer tRNA3Lys}, author = {C Isel and J M Lanchy and S F Le Grice and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8631312}, isbn = {8631312}, year = {1996}, date = {1996-01-01}, journal = {EMBO J}, volume = {15}, number = {4}, pages = {917-924}, abstract = {Initiation of RNA-dependent DNA synthesis by retroviral reverse transcriptases is generally considered as unspecific. In the case of human immunodeficiency virus type 1 (HIV-1), the natural primer is tRNA3Lys. We recently found evidence of complex interactions between tRNA3Lys and HIV-1 RNA that may be involved in the priming process. In this study, we compare the ability of natural and unmodified synthetic tRNA3Lys and 18mer oligoribo- and oligodeoxyribonucleotides complementary to the viral primer binding site to initiate replication of HIV-1 RNA using either homologous or heterologous reverse transcriptases. We show that HIV-1 RNA, HIV-1 reverse transcriptase and primer tRNA3Lys form a specific initiation complex that differs from the unspecific elongation complex formed when an oligodeoxyribonucleotide is used as primer. Modified nucleosides of tRNA3Lys are required for efficient initiation and transition to elongation. Transition from initiation to elongation, but not initiation of reverse transcription itself, is facilitated by extended primer-template interactions. Elongation, but not initiation of reverse transcription, is inhibited by Mn2+, which further differentiates these two different functional states of reverse transcriptase. These results define initiation of reverse transcription as a target to block viral replication.}, note = {0261-4189 Journal Article}, keywords = {Cell-Free System Gene Expression Regulation, Genetic *Virus Replication, Lys/*metabolism RNA, MARQUET, Non-U.S. Gov't Templates, Transfer, Unité ARN, Viral HIV-1/*genetics HIV-1 Reverse Transcriptase RNA, Viral/metabolism RNA-Directed DNA Polymerase/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Determining the conformation of RNAs in solution. Application to a retroviral system: structure of the HIV-1 primer binding site region and effect of tRNA(3Lys) binding}, author = {C Isel and C Ehresmann and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8786994}, isbn = {8786994}, year = {1996}, date = {1996-01-01}, journal = {Pharm Acta Helv}, volume = {71}, number = {1}, pages = {11-19}, abstract = {RNAs play a crucial and central role in a large variety of biological functions obviously linked to the wide variety of structures that they can adopt. Understanding the function of RNAs thus requires the knowledge of their two- and three-dimensional structures. We describe in detail the way to access the secondary structure of RNAs, by combining sequence comparison, secondary structure prediction by computer and, mainly, experimental data obtained by probing with chemicals and ribonucleases. These approaches were used to investigate secondary structure of the region containing the primer binding site of HIV-1 genomic RNA either free or involved in the binary complex with the replication primer tRNA(3Lys).}, note = {0031-6865 Journal Article}, keywords = {Amino Acid Sequence Binding Sites DNA Primers HIV-1/*chemistry Human Nucleic Acid Conformation RNA, Lys/*chemistry RNA, MARQUET, Non-U.S. Gov't, Transfer, Unité ARN, Viral/*chemistry Solutions Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Solution structure of mRNA hairpins promoting selenocysteine incorporation in Escherichia coli and their base-specific interaction with special elongation factor SELB}, author = {A Huttenhofer and E Westhof and A Bock}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8634916}, isbn = {8634916}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {4}, pages = {354-366}, abstract = {On the basis of chemical probing data, the solution structures of RNA hairpins within fdhF and fdnG mRNAs in Escherichia coli, which both promote selenocysteine incorporation at UGA codons, were derived with the help of computer modeling. We find that these mRNA hairpins contain two separate structural domains that possibly also exert two different functions. The first domain is comprised of the UGA codon, which is included within a complex and distorted double-stranded region. Thereby, release factor 2 might be prevented from binding to the UGA codon to terminate protein synthesis. The second domain is located within the apical loop of the mRNA hairpin structures. This loop region exhibits a defined tertiary structure in which no base is involved in Watson-Crick interactions. The structure of the loop is such that, following a sharp turn after G22 (A22 in fdnG mRNA), bases G23 and U24 are exposed to the solvent on the deep groove side of the supporting helix. Residues C25 and U26 close the loop with a possible single H-bonding interaction between the first and last residues of the loop, 04(U26) and N6(A21). The bulge residues U17 and U18 (in fdhF mRNA), or Ul7 only in fdnG mRNA, point their Watson-Crick positions in the same direction as loop residues G23 and U24 do, and at the same time open up the deep groove at the top of the hairpin helix. Chemical probing data demonstrate that bases G23 and U24 in both mRNA hairpins, as well as residues U17 and Ul7/U18 (for fdhF mRNA) located in a bulge 5' to the loop, are involved directly in binding to special elongation factor SELB in both mRNAs. Therefore, SELB recognizes identical bases within both mRNA hairpins despite differences in their primary sequence, consistent with the derived 3D models for these mRNAs, which exhibit similar tertiary structures. Binding of SELB to the fdhF mRNA hairpin was estimated to proceed with an apparent Kd of 30 nM.}, note = {1355-8382 Journal Article}, keywords = {Messenger/*chemistry Selenocysteine/*metabolism Solutions Sulfuric Acid Esters/chemistry Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {RNAs mediating cotranslational insertion of selenocysteine in eukaryotic selenoproteins}, author = {N Hubert and R Walczak and C Sturchler and E Myslinski and C Schuster and E Westhof and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955902}, isbn = {8955902}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {590-596}, abstract = {Selenocysteine, a selenium-containing analog of cysteine, is found in the prokaryotic and eukaryotic kingdoms in active sites of enzymes involved in oxidation-reduction reactions. Its biosynthesis and cotranslational insertion into selenoproteins is performed by an outstanding mechanism, implying the participation of several gene products. The tRNA(Sec) is one of these. In eukaryotes, its transcription mode by RNA polymerase III differs from that of classical tRNA genes, both at the level of the promoter elements and transcription factors involved. In addition, enhanced transcription is afforded by a newly characterized zinc finger activator. Not only transcription of the gene, but also the tRNA(Sec) itself is atypical since its 2D and 3D structures exhibit features which set it apart from classical tRNAs. Decoding of eukaryotic selenocysteine UGA codons requires a stem-loop structure in the 3'UTR of mRNAs, the selenocysteine insertion sequence (SECIS) element. Structure probing and sequence comparisons led us to propose a 2D structure model for the SECIS element, containing a novel RNA motif composed of four consecutive non-Watson-Crick base-pairs. A 3D model, rationalizing the accessibility data, was elaborated by computer modeling. It yields indicative or suggestive evidence for the role that could play some conserved residues and/or structural features in SECIS function. These might act as signals for interaction with SBP, the SECIS binding protein that we have characterized.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Amino Acid-Specific/chemistry/metabolism Rats Schistosoma mansoni Selenocysteine/chemistry/*metabolism Support, Animals Base Sequence Cattle Escherichia coli Human Mice Models, Molecular Molecular Sequence Data Nucleic Acid Conformation Proteins/chemistry/*metabolism RNA/chemistry/*metabolism RNA, Non-U.S. Gov't Xenopus laevis, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A protein binds the selenocysteine insertion element in the 3'-UTR of mammalian selenoprotein mRNAs}, author = {N Hubert and R Walczak and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8602359}, isbn = {8602359}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {3}, pages = {464-469}, abstract = {Several gene products are involved in co-translational insertion of selenocysteine by the tRNA(Sec). In addition, a stem-loop structure in the mRNAs coding for selenoproteins is essential to mediate the selection of the proper selenocysteine UGA codon. Interestingly, in eukaryotic selenoprotein mRNAs, this stem-loop structure, the selenocysteine insertion sequence (SECIS) element, resides in the 3'-untranslated region, far downstream of the UGA codon. In view of unravelling the underlying complex mechanism, we have attempted to detect RNA-binding proteins with specificity for the SECIS element. Using mobility shift assays, we could show that a protein, present in different types of mammalian cell extracts, possesses the capacity of binding the SECIS element of the selenoprotein glutathione peroxidase (GPx) mRNA. We have termed this protein SBP, for Secis Binding Protein. Competition experiments attested that the binding is highly specific and UV cross-linking indicated that the protein has an apparent molecular weight in the range of 60-65 kDa. Finally, some data suggest that the SECIS elements in the mRNAs of GPx and another selenoprotein, type I iodothyronine 5' deiodinase, recognize the same SBP protein. This constitutes the first report of the existence of a 3' UTR binding protein possibly involved in the eukaryotic selenocysteine insertion mechanism.}, note = {0305-1048 Journal Article}, keywords = {Animals Base Sequence DNA Transposable Elements/*genetics Molecular Sequence Data Protein Binding Proteins/*genetics/metabolism RNA, Genetic, Messenger/genetics/*metabolism Rats Selenocysteine/genetics/*metabolism Support, Non-U.S. Gov't Translation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Fe.bleomycin as a probe of RNA conformation}, author = {C E Holmes and A T Abraham and S M Hecht and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8811095}, isbn = {8811095}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {17}, pages = {3399-3406}, abstract = {Two crystallographically defined tRNAs, yeast tRNAAsp and tRNAPhe, were used as substrates for oxidative cleavage by Fe.bleomycin to facilitate definition at high resolution of the structural elements in RNAs conducive to bleomycin binding and cleavage. Yeast tRNAAsp underwent cleavage at G45 and U66; yeast tRNAPhe was cleaved at four sites, namely G19, A31, U52 and A66. Only two of these six sites involved oxidative cleavage of a 5'-G.Pyr-3' sequence, but three sites were at the junction between single- and double-stranded regions of the RNA, consistent with a binding model in which the bithiazole + C-terminal substituent of bleomycin bind to minor groove structures on the RNA. Also studied were four tRNA transcripts believed on the basis of biochemical and chemical mapping experiments to share structural elements in common with the mature tRNAs. Cleavage of these tRNAs by Fe.bleomycin gave patterns of cleavage very different from each other and than those of the mature tRNAs. This observation suggests strongly that Fe.bleomycin cannot be used for chemical mapping in the same fashion as more classical reagents, such as Pb2+ or dimethyl sulfate. However, the great sensitivity of Fe.bleomycin to changes in nucleic acid structure argues that those species which do show similar patterns of cleavage must be very close in structure.}, note = {0305-1048 Journal Article}, keywords = {Asp/chemistry RNA, Binding Sites Bleomycin/*analogs & derivatives/chemistry Models, FLORENTZ, Fungal/*chemistry RNA, Messenger/chemistry RNA, Molecular *Molecular Probes *Nucleic Acid Conformation RNA Precursors/chemistry RNA, Non-U.S. Gov't Support, P.H.S., Phe/chemistry Support, Transfer, Transfer/*chemistry RNA, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Enzymatic formation of modified nucleosides in tRNA: dependence on tRNA architecture}, author = {H Grosjean and J Edqvist and K B Straby and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8568876}, isbn = {8568876}, year = {1996}, date = {1996-01-01}, journal = {J Mol Biol}, volume = {255}, number = {1}, pages = {67-85}, abstract = {Information is still quite limited concerning the structural requirements in tRNA molecules for their post-transcriptional maturation by base and ribose modification enzymes. To address this question, we have chosen as the model system yeast tRNAAsp that has a known three-dimensional structure and the in vivo modifying machinery of the Xenopus laevis oocyte able to act on microinjected tRNA precursors. We have systematically compared the modification pattern of wild-type tRNAAsp with that of a series of structural mutants (21 altogether) altered at single or multiple positions in the D-, T-and the anticodon branch, as well as in the variable region. The experimental system allowed us to analyze the effects of structural perturbations in tRNA on the enzymatic formation of modified nucleosides at 12 locations scattered over the tRNA cloverleaf. We found that the formation of m1G37 and psi 40 in the anticodon loop and stem and psi 13 in the D-stem, were extremely sensitive to 3D perturbations. In contrast, the formation of T54, psi 55 and m1A58 in the T-loop, m5C49 in the T-stem and m2G6 in the amino acid accepting stem were essentially insensitive to change in the overall tRNA architecture; these modified nucleosides were also formed in appropriate minimalist (stems and loops) tRNA domains. The formation of m2G26 at the junction between the anticodon and the D-stem, of Q34 and manQ34 in the anticodon loop were sensitive only to drastic structural perturbation of the tRNA. Altogether, these results reflect the existence of different modes of tRNA recognition by the many different modifying enzymes. A classification of this family of maturation enzymes into two major groups, according to their sensitivities to structural perturbations in tRNA, is proposed.}, note = {0022-2836 Journal Article}, keywords = {Animals Base Sequence Isomerases/metabolism Microinjections Molecular Sequence Data Mutation *Nucleic Acid Conformation Oocytes Pentosyltransferases/metabolism RNA Processing, Asp/*chemistry/metabolism Ribonucleosides/biosynthesis/*metabolism Support, Non-U.S. Gov't Xenopus laevis tRNA Methyltransferases/*metabolism, Post-Transcriptional/*physiology RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Probing the higher order structure of RNA with peroxonitrous acid}, author = {M Gotte and R Marquet and C Isel and V E Anderson and G Keith and H J Gross and C Ehresmann and B Ehresmann and H Heumann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8706865}, isbn = {8706865}, year = {1996}, date = {1996-01-01}, journal = {FEBS Lett}, volume = {390}, number = {2}, pages = {226-228}, abstract = {Potassium peroxonitrite (ONOOK) and [Fe(EDTA)]2- were used to analyze the influence of chemically entirely different hydroxyl radical sources on tRNA cleavage profiles. [Fe(EDTA)]2- gives rise to hydroxyl radicals via a Fenton-like reaction during the oxidation of chelated Fe2+, while ONOOK generates hydroxyl radicals via its conjugate acid (ONOOH) when adding a stable alkaline solution of ONOOK in samples buffered at neutral pH. [Fe(EDTA)]2- is known to induce oxidative strand scission at sugar moieties thought to be solvent accessible, while those residues located in the 'inside' of structured RNAs are protected. Although ONOOH is neutral and significantly smaller than the metal complex, both reagents generate the same protection pattern on tRNAs, suggesting that access of the commonly formed hydroxyl radical, rather than access of its source, is the determining factor when probing the higher order structure of RNA. Strong difference in reactivity is only seen at the modified 2-thiouridine S34 of tRNA(Lys3) which shows hyperreactivity towards ONOOK treatment. This particular reaction may require interaction between the peroxonitrite anion and the thiocarbonyl group of the base, since hyperreactivity is not observed when probing the dethiolated tRNA(Lys3).}, note = {0014-5793 Journal Article}, keywords = {Animals Chelating Agents Edetic Acid Hydroxyl Radical/chemistry Molecular Probes Molecular Structure *Nitrates RNA, Fungal/chemistry RNA, Lys/chemistry RNA, MARQUET, Non-U.S. Gov't, Phe/chemistry Rabbits Saccharomyces cerevisiae/chemistry Support, Transfer, Transfer/*chemistry RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Aspartate identity of transfer RNAs}, author = {R Giege and C Florentz and D Kern and J Gangloff and G Eriani and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955904}, isbn = {8955904}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {605-623}, abstract = {Structure/function relationships accounting for specific tRNA charging by class II aspartyl-tRNA synthetases from Saccharomyces cerevisiae, Escherichia coli and Thermus thermophilus are reviewed. Effects directly linked to tRNA features are emphasized and aspects about synthetase contribution in expression of tRNA(Asp) identity are also covered. Major identity nucleotides conferring aspartate specificity to yeast, E coli and T thermophilus tRNAs comprise G34, U35, C36, C38 and G73, a set of nucleotides conserved in tRNA(Asp) molecules of other biological origin. Aspartate specificity can be enhanced by negative discrimination preventing, eg mischarging of native yeast tRNA(Asp by yeast arginyl-tRNA synthetase. In the yeast system crystallography shows that identity nucleotides are in contact with identity amino acids located in the catalytic and anticodon binding domains of the synthetase. Specificity of RNA/protein interaction involves a conformational change of the tRNA that optimizes the H-bonding potential of the identity signals on both partners of the complex. Mutation of identity nucleotides leads to decreased aspartylation efficiencies accompanied by a loss of specific H-bonds and an altered adaptation of tRNA on the synthetase. Species-specific characteristics of aspartate systems are the number, location and nature of minor identity signals. These features and the structural variations in aspartate tRNAs and synthetases are correlated with mechanistic differences in the aminoacylation reactions catalyzed by the various aspartyl-tRNA synthetases. The reality of the aspartate identity set is verified by its functional expression in a variety of RNA frameworks. Inversely a number of identities can be expressed within a tRNA(Asp) framework. From this emerged the concept of the RNA structural frameworks underlying expression of identities which is illustrated with data obtained with engineered tRNAs. Efficient aspartylation of minihelices is explained by the primordial role of G73. From this and other considerations it is suggested that aspartate identity appeared early in the history of tRNA aminoacylation systems.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Asp/*chemistry Saccharomyces cerevisiae Structure-Activity Relationship Support, Aspartate-tRNA Ligase/chemistry/metabolism Aspartic Acid/analysis Base Sequence Escherichia coli Models, ERIANI, FLORENTZ, Molecular Molecular Sequence Data Nucleic Acid Conformation RNA, Non-U.S. Gov't Thermus thermophilus, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interplay of tRNA-like structures from plant viral RNAs with partners of the translation and replication machineries}, author = {R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8901535}, isbn = {8901535}, year = {1996}, date = {1996-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {93}, number = {22}, pages = {12078-12081}, note = {0027-8424 Comment Journal Article Review Review, Tutorial}, keywords = {Amino Acyl/*physiology RNA, Base Sequence Evolution, Genetic Molecular Sequence Data Peptide Elongation Factors/metabolism Plant Viruses/*genetics RNA, Genetic Virus Replication, Molecular Models, Transfer, Unité ARN, Viral/biosynthesis Translation}, pubstate = {published}, tppubtype = {article} } @article{, title = {Extended interactions between the primer tRNAi(Met) and genomic RNA of the yeast Ty1 retrotransposon}, author = {S Friant and T Heyman and M L Wilhelm and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8602356}, isbn = {8602356}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {3}, pages = {441-449}, abstract = {Reverse transcription of the yeast Ty1 retrotransposon is primed by tRNAi(Met) base paired to the primer binding site near the 5'-end of Ty1 genomic RNA. To understand the molecular basis of the tRNAi(Met)-Ty1 RNA interaction the secondary structure of the binary complex was analysed. Enzymatic probes were used to test the conformation of tRNAi(Met) and of Ty1 RNA in the free form and in the complex. A secondary structure model of the tRNAi(Met) Ty1 RNA complex consistent with the probing data was constructed with the help of a computer program. The model shows that besides interactions between the primer binding site and the last 10 nt at the 3'-end of tRNAi(Met), three short regions of Ty1 RNA named boxes 0, 1 and 2.1 interact with the T and D stems and loops of tRNAiMet. Mutations were made in the boxes or in the complementary sequences of tRNAi(Met) to study the contribution of these sequences to formation of the complex. We find that interaction with at least one of the two boxes 0 or 1 is absolutely required for efficient annealing of the two RNAs. Sequence comparison showing that the primary sequence of the boxes is strictly conserved in Ty1 and Ty2 elements and previously published in vivo results underline the functional importance of the primary sequence of the boxes and suggest that extended interactions between genomic Ty1 RNA and the primary tRNAi(Met) play a role in the reverse transcription pathway.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Molecular Sequence Data Molecular Structure Mutation Nucleic Acid Conformation RNA/genetics/*metabolism RNA, Met/genetics/*metabolism Retroelements/*genetics Saccharomyces cerevisiae Support, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of RNA primers in initiation of minus-strand and plus-strand DNA synthesis of the yeast retrotransposon Ty1}, author = {S Friant and T Heyman and F X Wilhelm and M Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955910}, isbn = {8955910}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {674-680}, abstract = {The Ty1 retrotransposon of the yeast Saccharomyces cerevisiae is a long terminal repeat mobile genetic element that transposes through an RNA intermediate. Initiation of minus-strand and plus-strand DNA synthesis are two critical steps during reverse transcription of the retrotransposon genome. Initiation of minus-strand DNA synthesis of the Ty1 element is primed by the cytoplasmic initiator methionine tRNA base paired to the primer binding site near the 5' end of the genomic RNA. A structural probing study of the primer tRNA-Ty1 RNA binary complex reveals that besides interactions between the primer binding site and the last 10 nucleotides at the 3' end of the primer tRNA, three short regions of Ty1 RNA named box 0, box 1 and box 2.1 interact with the T and D stems and loops of the primer tRNA. Some in vivo results underline the functional importance of the nucleotide sequence of the boxes and suggest that extended interactions between genomic Ty1 RNA and the primer tRNA play a role in the reverse transcription pathway. Plus-strand DNA synthesis is initiated from an RNase H resistant oligoribonucleotide spanning a purine-rich sequence, the polypurine tract (PPT). Two sites of initiation located at the 5' boundary of the 3' long terminal repeat (PPT1) and near the middle of the TyB (pol) gene in the integrase coding sequence (PPT2) have been identified in the genome of Ty1. The two PPTs have an identical sequence, TGGGTGGTA. Mutations replacing purines by pyrimidines in this sequence significantly diminish or abolish initiation of plus-strand DNA synthesis. Ty1 elements bearing a mutated PPT2 sequence are not defective for transposition whereas mutations in PPT1 abolish transposition.}, note = {0300-9084 Journal Article}, keywords = {*DNA Replication *DNA Transposable Elements DNA, Bacterial/*metabolism Support, Complementary/metabolism Molecular Sequence Data Mutagenesis Nucleic Acid Conformation RNA/*metabolism RNA, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Usefulness of functional and structural solution data for the modeling of tRNA-like structures}, author = {B Felden and C Florentz and E Westhof and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8786997}, isbn = {8786997}, year = {1996}, date = {1996-01-01}, journal = {Pharm Acta Helv}, volume = {71}, number = {1}, pages = {3-9}, abstract = {Structures of large RNAs are not easily solved by X-ray crystallography or by NMR spectroscopy. This paper reviews the alternate methodology based on enzymatic and chemical mapping data collected on RNAs combined with graphical modeling for the construction of three-dimensional models. The different steps that lead to the establishment of the models are critically discussed. It is shown how the correctness of an RNA model can be strengthened by establishing correlations between the structure and the functionality of the molecule and its variants. Finally, the predictive potential of a model is discussed The approach is illustrated by results obtained on plant viral tRNA-like structures, and particularly on that of brome mosaic virus (BMV) RNA.}, note = {0031-6865 Journal Article Review Review, Tutorial}, keywords = {Base Sequence Models, Chemical Molecular Sequence Data RNA, FLORENTZ, Non-U.S. Gov't, Transfer/*chemistry Solutions Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A central pseudoknotted three-way junction imposes tRNA-like mimicry and the orientation of three 5' upstream pseudoknots in the 3' terminus of tobacco mosaic virus RNA}, author = {B Felden and C Florentz and R Giege and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8608444}, isbn = {8608444}, year = {1996}, date = {1996-01-01}, journal = {RNA}, volume = {2}, number = {3}, pages = {201-212}, abstract = {A three-dimensional model of the histidylable 3'-terminal tRNA-like domain of tobacco mosaic virus RNA is proposed on the basis of a comparative structural analysis, chemical and enzymatic probing, combined with graphical modeling of three RNA constructs of increasing size (38, 108, and 182 nt) derived from the 3'-terminal viral RNA sequence. The comparison between the probing patterns of the three RNAs allowed the determination of the relative orientation of these structural domains in the full-length viral tRNA-like structure. Modeling data indicate that only one of the two possible isomers of the three-way junction located at a central position of the tRNA-like domain is in agreement with structural data. Interestingly, this isomer gives rise to a molecule bearing a structural mimicry with the L-shape of canonical tRNAs. A pseudoknotted acceptor branch containing a T-like loop is located perpendicularly to an anticodon-like branch. Moreover, a single-stranded RNA stretch belonging to the pseudoknotted central core mimics a D-like loop and it is proposed that it interacts via two conserved guanosines with nucleotides of the T-like loop as found in canonical tRNAs. This model is valid for the 3' noncoding regions of tobamoviral RNAs as well as for the tRNA-like domain of the satellite tobacco mosaic virus RNA. All three molecules are substrates for yeast HisRS; however, whereas the complete viral genome is required for optimal histidylation capacities, both charging levels and affinity constants are decreased for the three RNA transcripts, suggesting that additional contacts located outside the tRNA-like domain are needed for an optimal aminoacylation process.}, note = {1355-8382 Journal Article}, keywords = {Base Sequence Computer Simulation Histidine-tRNA Ligase Models, FLORENTZ, Molecular Molecular Mimicry/*physiology Molecular Sequence Data *Nucleic Acid Conformation RNA, Non-U.S. Gov't Tobacco Mosaic Virus/*chemistry, Transfer/chemistry RNA, Unité ARN, Viral/*chemistry/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A tyrosyl-tRNA synthetase recognizes a conserved tRNA-like structural motif in the group I intron catalytic core}, author = {M G Caprara and V Lehnert and A M Lambowitz and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8978617}, isbn = {8978617}, year = {1996}, date = {1996-01-01}, journal = {Cell}, volume = {87}, number = {6}, pages = {1135-1145}, abstract = {The Neurospora crassa mitochondrial (mt) tyrosyl-tRNA synthetase (CYT-18 protein) functions in splicing group I introns, in addition to aminoacylating tRNA(Tyr). Here, we compared the CYT-18 binding sites in the N. crassa mt LSU and ND1 introns with that in N. crassa mt tRNA(Tyr) by constructing three-dimensional models based on chemical modification and RNA footprinting data. Remarkably, superimposition of the CYT-18 binding sites in the model structures revealed an extended three-dimensional overlap between the tRNA and the group I intron catalytic core. Our results provide insight into how an RNA-splicing factor can evolve from a cellular RNA-binding protein. Further, the structural similarities between group I introns and tRNAs are consistent with an evolutionary relationship and suggest a general mechanism for the evolution of complex catalytic RNAs.}, note = {0092-8674 Journal Article}, keywords = {Base Sequence Binding Sites/genetics Conserved Sequence Evolution *Introns Molecular Sequence Data Neurospora crassa Nucleic Acid Conformation Protein Conformation Protein Structure, Fungal/chemistry/metabolism/physiology RNA, Non-U.S. Gov't Support, P.H.S. Tyrosine-tRNA Ligase/*chemistry/*genetics/metabolism, Tertiary RNA Splicing/physiology RNA, Transfer, Tyr/chemistry Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization of Escherichia coli aspartyl-tRNA synthetase in its free state and in a complex with yeast tRNA(Asp).}, author = {M Boeglin and A C Dock-Brégeon and G Eriani and J Gangloff and M Ruff and A Poterszman and J C Thierry and D Moras}, url = {http://www.ncbi.nlm.nih.gov/pubmed/15299749}, isbn = {15299749}, year = {1996}, date = {1996-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {52}, number = {Pt 1}, pages = {211-214}, abstract = {Overexpressed dimeric E. coli aspartyl-tRNA synthetase (AspRS) has been crystallized in its free state and complexed with yeast tRNA(Asp). Triclinic crystals of the enzyme alone (a = 104.4}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pseudoknot and translational control in the expression of the S15 ribosomal protein}, author = {L Benard and C Philippe and B Ehresmann and C Ehresmann and C Portier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955900}, isbn = {8955900}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {568-576}, abstract = {Translational autocontrol of the expression of the ribosomal protein S15 proceeds through the transitory formation of a pseudoknot. A synopsis of the known data is used to propose a molecular model of the mechanism involved and for the role of the pseudoknot. This latter structure is able to recruit 30S ribosomal subunits to initiate translation, but also to bind S15 and to stop translation by trapping the ribosome on its loading site. Information on the S15 protein recognition of the messenger RNA site was deduced from mutational analyses and chemical probing. A comparison of this messenger site with the S15 ribosomal binding site was conducted by analysing hydroxyl radical footprintings of these two sites. The existence of two subsites in 16S RNA suggests that the ribosomal protein S15 might present either two different binding sites or at least one common subsite. Clues for the presence of a common site between the messenger and 16S RNA are given which cannot rule out that recognition specificity is linked to a few other determinants. Whether these determinants are different or not remains an open question.}, note = {0300-9084 Journal Article}, keywords = {Base Sequence Molecular Sequence Data Nucleic Acid Conformation Ribosomal Proteins/*biosynthesis/chemistry/genetics/metabolism Structure-Activity Relationship Support, Genetic, Non-U.S. Gov't *Translation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identity of prokaryotic and eukaryotic tRNA(Asp) for aminoacylation by aspartyl-tRNA synthetase from Thermus thermophilus}, author = {H D Becker and R Giege and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8652522}, isbn = {8652522}, year = {1996}, date = {1996-01-01}, journal = {Biochemistry}, volume = {35}, number = {23}, pages = {7447-7458}, abstract = {The aspartate identity of tRNA for AspRS from Thermus thermophilus has been investigated by kinetic analysis of the aspartylation reaction of different tRNA molecules and their variants as well as of tRNAPhe variants with transplanted aspartate identity elements. It is shown that G10, G34, U35, C36, C38, and G73 determine recognition and aspartylation of yeast and T.thermophilus tRNA(Asp) by the thermophilic AspRS. This set of nucleotides specifies also tRNA aspartylation in the homologous yeast and Escherichia coli systems. Structural considerations indicate that the major aspartate identity elements interact with amino acids conserved in all AspRSs. It follows that the structural features of tRNA and synthetase specifying aspartylation are mainly conserved in various structural contexts and in organisms adapted to different life conditions. Mutations of tRNA identity elements provoke drastic losses of charging in the heterologous system involving yeast tRNA(Asp) and T. thermophilus AspRS. In the homologous systems, the mutational effects are less pronounced. However, effects in E. coli and T. thermophilus exceed those in yeast which are particularly moderate, indicating variations in the individual contributions of identity elements for aspartylation in prokaryotes and eukaryotes. Analysis of multiple tRNA mutants reveals cooperativity between the cluster of determinants of the anticodon loop and the additional determinants G10 and G73 for efficient aspartylation in the thermophilic system, suggesting that conformational changes trigger formation of the functional tRNA/synthetase complex.}, note = {0006-2960 Journal Article}, keywords = {Amino Acid Substrate Specificity Support, Amino Acid Sequence Anticodon Aspartate-tRNA Ligase/chemistry/*metabolism Base Sequence Comparative Study Escherichia coli Kinetics Molecular Sequence Data Nucleic Acid Conformation RNA, Asp/biosynthesis/*metabolism RNA, Genetic, Non-U.S. Gov't Thermus thermophilus/*enzymology Transcription, Phe/biosynthesis/metabolism Recombinant Proteins/chemistry/metabolism Saccharomyces cerevisiae Sequence Homology, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {H-bond stability in the tRNA(Asp) anticodon hairpin: 3 ns of multiple molecular dynamics simulations}, author = {P Auffinger and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8842234}, isbn = {8842234}, year = {1996}, date = {1996-01-01}, journal = {Biophys J}, volume = {71}, number = {2}, pages = {940-954}, abstract = {Multiple molecular dynamics trajectories of the solvated and neutralized 17-residue tRNA(Asp) anticodon hairpin were generated for a total of 3 ns. Explicit treatment of all long-ranged electrostatic interactions by the particle mesh Ewald algorithm, as implemented in the AMBER MD software package, effected a degree of structural stabilization not previously achieved by use of a long 16-A solvent interaction truncation scheme. The increased stability of this multiple molecular dynamics set was appropriate for an in-depth analysis of the six 500-ps-long trajectories and allowed the characterization of a number of key structural interactions. The dynamical behavior of the standard Watson-Crick base pairs, the noncanonical G30-U40 "wobble" base pair, and the psi 32-C38 pseudo-base pair is presented as well as that of two C--H. O hydrogen bonds found to contribute to the array of tertiary interactions that stabilize the seven-nucleotide native loop conformation. The least mobile residue in the loop is U33, which forms the U-turn motif and which participates in several hydrogen-bonding interactions, whereas the most mobile residue is the apical residue G34 at the wobble position, a factor undoubtedly important in its biological function. The set of multiple molecular dynamics trajectories obtained does not converge on a 500-ps time scale to a unique dynamical model but instead describes an ensemble of structural microstates accessible to the system under the present simulation protocol, which is the result of local structural heterogeneity rather than of global conformational changes.}, note = {0006-3495 Journal Article}, keywords = {Anticodon/*chemistry Base Composition Base Sequence *Computer Simulation Drug Stability Hydrogen Bonding Kinetics Models, Asp/*chemistry Software Support, Molecular *Nucleic Acid Conformation RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{charlet_cloning_1996, title = {Cloning of the gene encoding the antibacterial peptide drosocin involved in Drosophila immunity. Expression studies during the immune response.}, author = {Maurice Charlet and Marie Lagueux and Jean-Marc Reichhart and Danièle Hoffmann and A Braun and Marie Meister}, year = {1996}, date = {1996-01-01}, journal = {Eur. J. Biochem.}, volume = {241}, pages = {699--706}, abstract = {A potent inducible antibacterial peptide carrying an O-glycosylated substitution has recently been isolated from Drosophila [Bulet et al. (1993) J. Biol. Chem. 268 14893-14897]. Here we report cloning studies that show that Drosophila contains a single, intronless gene, located at position 51C1-6, which encodes the precursor protein from which drosocin is processed. The upstream and the downstream sequences of the drosocin gene contain putative cis-regulatory elements similar to mammalian regulatory motifs, namely three kB-related decameric sequences. The drosocin gene is silent in naive animals, and is strongly induced with acute phase kinetics after immune challenge in larvae and adults. We have established several transgenic fly lines in which reporter genes were placed under the control of various drosocin promoter sequences. Our results indcate that 2.5 kb of upstream sequences confer inducibility and tissue specificity to the transgene, but that the level of its expression in the fat body after immune challenge}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular Dynamics Simulations of the Anticodon Hairpin of tRNAAsp: Structuring Effects of C−H···O Hydrogen Bonds and of Long-Range Hydration Forces}, author = {P Auffinger and S Louise-May and E Westhof}, url = {http://pubs.acs.org/doi/abs/10.1021/ja952494j?prevSearch=Auffinger&searchHistoryKey=}, doi = {10.1021/ja952494j}, year = {1996}, date = {1996-01-01}, journal = {J Am Chem Soc}, volume = {118}, number = {5}, pages = {1181–1189}, abstract = {The inclusion of long-range solvent interactions out to 16 Å in a molecular dynamics study of the anticodon loop of tRNAAsp led to an overall structural stabilization of the RNA hairpin tertiary interactions in a set of six independent fully solvated and neutralized 100 ps MD trajectories as compared to a shorter-ranged solvent interaction electrostatic model (8 Å). The increased structural stabilization allowed for the emergence of non-classical C−H···O hydrogen bonds in the MD trajectories. The presence of the C−H···O hydrogen bonds in the crystal structure was subsequently verified and dynamically characterized and their contribution to the preservation of the tertiary native conformation was assessed. The MD trajectories generated using a truncation distance of 16 Å for the electrostatic solute−solvent and solvent−solvent interactions, with no cutoffs applied to the electrostatic solute−solute interactions, compared to an earlier set of eight independent 100 ps MD trajectories using a smaller truncation distance of 8 Å (Auffinger, P.; Louise-May, S.; Westhof, E. J. Am. Chem. Soc. 1995, 117, 6720−6726), revealed an increase in consistency of structural characteristics between individual MD trajectories of a given set and on average a decrease in root-mean-square deviation values from the starting crystal structure. Dihedral transitions in the sugar−phosphate backbone decreased and tertiary interactions specific to the loop topology were better preserved and showed reduced dynamical fluctuation. These results emphasize the important influence of long-ranged solvation forces on the stabilization of the tertiary structure of highly charged nucleic acid systems and signify that long-ranged theoretical models may be necessary for a truly accurate description of biomacromolecular solution structure and dynamics.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{michaut_determination_1996, title = {Determination of the disulfide array of the first inducible antifungal peptide from insects : drosomycin from Drosophila melanogaster.}, author = {Lydia Michaut and P Fehlbaum and M Moniatte and Van A Dorsselaer and Jean-Marc Reichhart and Philippe Bulet}, year = {1996}, date = {1996-01-01}, journal = {FEBS Lett.}, volume = {395}, pages = {6--10}, abstract = {Drosomycin is a 44-residue antifungal peptide with four intramolecular disulfide bridges which have been isolated from immune-challenged Drosophila. To produce adequate amounts of this peptide for 3D-structure analysis, studies on the mode of action and activity spectrum, we expressed a synthetic cDNA in Saccharomyces cerevisiae. For this purpose, we used the mating factor a gene and concomitantly over-expressed the KEX2 gene to increase the yield of fully processed drosomycin. Using a combination of Edman degradation and mass spectrometry, we show that drosomycin shares the same array of intramolecular disulfide bridges than plant defensins, in addition to their sequence similarities.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{, title = {Hydration of C-H groups in tRNA}, author = {P Auffinger and S Louise-May and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9136637}, isbn = {9136637}, year = {1996}, date = {1996-01-01}, journal = {Faraday Discuss}, number = {103}, pages = {151-173}, abstract = {Molecular dynamic (MD) simulations of the anticodon hairpin of tRNA(Asp) and of the full tRNA, both in a solvent bath with neutralizing NH4+ counter-ions, have been produced with the particle mesh ewald (PME) method and the multiple molecular dynamics (MMD) strategy. The latter consists of generating uncorrelated trajectories starting from the same initial configuration but with a slightly perturbed initial velocity distribution. The 3 ns (six uncorrelated 500 ps MD trajectories) MMD set of the 17 nucleotide anticodon fragment and the single 500 ps trajectory of the 75 nucleotide tRNA were analyzed with the aim of characterizing long lived C-H.Ow interactions for the two main nucleic acid base and ribose C-H bond types. Some C-H sites present very long residence lifetimes for water molecules, especially those around the ribose H(3') and the pyrimidine H(5) atoms. The C(3')-H(3').Ow contacts occur concurrently with the strong hydration of the anionic phosphate oxygen atoms and especially with the water bridges linking successive phosphate groups along the polynucleotide chain. Therefore, these contacts are of opportunistic character and result from the geometries of the covalent structure and adjacent interactions. On the other hand, the pyrimidine H(5) atoms display a hydrophilic character with interaction geometries indicating that water contacts in which they are involved should be considered as bona fide hydrogen bonds.}, note = {1359-6640 Journal Article}, keywords = {Base Sequence Carbon/chemistry Hydrogen/chemistry Models, Non-U.S. Gov't Water/chemistry, Structural Molecular Sequence Data Nucleic Acid Conformation RNA, Transfer/*chemistry Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{lemaitre_dorsoventral_1996, title = {The dorsoventral regulatory gene cassette spätzle/Toll/cactus controls the potent antifungal response in Drosophila adults}, author = {Bruno Lemaitre and E Nicolas and Lydia Michaut and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0092-8674}, year = {1996}, date = {1996-01-01}, journal = {Cell}, volume = {86}, number = {6}, pages = {973--983}, abstract = {The cytokine-induced activation cascade of NF-kappaB in mammals and the activation of the morphogen dorsal in Drosophila embryos show striking structural and functional similarities (Toll/IL-1, Cactus/I-kappaB, and dorsal/NF-kappaB). Here we demonstrate that these parallels extend to the immune response of Drosophila. In particular, the intracellular components of the dorsoventral signaling pathway (except for dorsal) and the extracellular Toll ligand, spätzle, control expression of the antifungal peptide gene drosomycin in adults. We also show that mutations in the Toll signaling pathway dramatically reduce survival after fungal infection. Antibacterial genes are induced either by a distinct pathway involving the immune deficiency gene (imd) or by combined activation of both imd and dorsoventral pathways.}, keywords = {Animals, Antifungal Agents, Cell Surface, DNA-Binding Proteins, Fungi, Gene Expression, Genes, hoffmann, Insect, Insect Hormones, Insect Proteins, M3i, Membrane Glycoproteins, MHC Class II, Mutation, Mycoses, NF-kappa B, Phosphoproteins, Proteins, Receptors, reichhart, Signal Transduction, Toll-Like Receptors}, pubstate = {published}, tppubtype = {article} } @article{, title = {Initiation of (-) strand DNA synthesis from tRNA(3Lys) on lentiviral RNAs: implications of specific HIV-1 RNA-tRNA(3Lys) interactions inhibiting primer utilization by retroviral reverse transcriptases}, author = {E J Arts and S R Stetor and X Li and J W Rausch and K J Howard and B Ehresmann and T W North and B M Wohrl and R S Goody and M A Wainberg and S F Grice}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8816751}, isbn = {8816751}, year = {1996}, date = {1996-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {93}, number = {19}, pages = {10063-10068}, abstract = {Initiation of minus (-) strand DNA synthesis was examined on templates containing R, U5, and primer-binding site regions of the human immunodeficiency virus type 1 (HIV-1), feline immunodeficiency virus (FIV), and equine infectious anemia virus (EIAV) genomic RNA. DNA synthesis was initiated from (i) an oligoribonucleotide complementary to the primer-binding sites, (ii) synthetic tRNA(3Lys), and (iii) natural tRNA(3Lys), by the reverse transcriptases of HIV-1, FIV, EIAV, simian immunodeficiency virus, HIV type 2 (HIV-2), Moloney murine leukemia virus, and avian myeloblastosis virus. All enzymes used an oligonucleotide on wild-type HIV-1 RNA, whereas only a limited number initiated (-) strand DNA synthesis from either tRNA(3Lys). In contrast, all enzymes supported efficient tRNA(3Lys)-primed (-) strand DNA synthesis on the genomes of FIV and EIAV. This may be in part attributable to the observation that the U5-inverted repeat stem-loop of the EIAV and FIV genomes lacks an A-rich loop shown with HIV-1 to interact with the U-rich tRNA anticodon loop. Deletion of this loop in HIV-1 RNA, or disrupting a critical loop-loop complex by tRNA(3Lys) extended by 9 nt, restored synthesis of HIV-1 (-) strand DNA from primer tRNA(3Lys) by all enzymes. Thus, divergent evolution of lentiviruses may have resulted in different mechanisms to use the same host tRNA for initiation of reverse transcription.}, note = {0027-8424 Journal Article}, keywords = {Amino Acyl/chemistry/*metabolism RNA, Animals Base Sequence Cats DNA, Equine/genetics/*metabolism Kinetics Molecular Sequence Data Nucleic Acid Conformation RNA, Genetic, Non-U.S. Gov't Support, P.H.S. Templates, Transfer, U.S. Gov't, Unité ARN, Viral HIV-1/genetics/*metabolism Horses Human Infectious Anemia Virus, Viral/*biosynthesis Genome, Viral/chemistry/*metabolism RNA-Directed DNA Polymerase/*metabolism SIV/genetics/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Restoration of tRNA3Lys-primed(-)-strand DNA synthesis to an HIV-1 reverse transcriptase mutant with extended tRNAs. Implications for retroviral replication}, author = {E J Arts and M Ghosh and P S Jacques and B Ehresmann and S F Le Grice}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8621554}, isbn = {8621554}, year = {1996}, date = {1996-01-01}, journal = {J Biol Chem}, volume = {271}, number = {15}, pages = {9054-9061}, abstract = {The mechanism for the initiation of reverse transcription in human immunodeficiency virus type 1 (HIV-1) was studied utilizing a unique reverse transcriptase (RT) mutant altered in its noncatalytic p51 subunit. This mutant (p66/p51Delta13) retains full DNA- and RNA-dependent DNA polymerase activity but has reduced affinity for tRNA3Lys, the cognate HIV primer. When the ability to support(-)-strand DNA synthesis on a viral RNA template was evaluated, this mutant initiated from an 18-nucleotide (nt) oligoribo- or oligodeoxyribonucleotide primer complementary to the primer binding site (pbs). However, it failed to do so from natural and synthetic versions of tRNA3Lys. tRNA-primed(-)-strand synthesis could, however, be rescued by substituting the 76-nt tRNA3Lys with 81- and 107-nt tRNA-DNA chimeras, i.e. tRNA3Lys extended by 5 and 31 deoxyribonucleotides complementary to the viral genome upstream of the pbs. These findings imply that through interactions involving its p51 subunit, RT may be required to disrupt additional tRNA-viral RNA duplexes outside the pbs to proceed into productive(-)-strand DNA synthesis. Alternatively, specific interactions between tRNA3Lys and HIV-1 RT may be necessary for efficient initiation of(-)-strand DNA synthesis.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence DNA, Calf Thymus/metabolism Sequence Deletion Structure-Activity Relationship Support, Complementary/biosynthesis DNA, Lys/*chemistry RNA-Directed DNA Polymerase/genetics/*metabolism Recombinant Proteins Ribonuclease H, Non-U.S. Gov't Support, P.H.S., Transfer, U.S. Gov't, Unité ARN, Viral/*biosynthesis HIV-1 Reverse Transcriptase Hydrogen Bonding Molecular Sequence Data Nucleic Acid Conformation RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {Conservation in evolution for a small monomeric phenylalanyl-tRNA synthetase of the tRNA(Phe) recognition nucleotides and initial aminoacylation site}, author = {R Aphasizhev and B Senger and J U Rengers and M Sprinzl and P Walter and G Nussbaum and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8555164}, isbn = {8555164}, year = {1996}, date = {1996-01-01}, journal = {Biochemistry}, volume = {35}, number = {1}, pages = {117-123}, abstract = {We previously showed that yeast mitochondrial phenylalanyl-tRNA synthetase (MSF protein) is evolutionarily distant to the cytoplasmic counterpart based on a high degree of divergence in protein sequence, molecular mass, and quaternary structure. Using yeast cytoplasmic tRNA(Phe) which is efficiently aminoacylated by MSF protein, we report here the tRNA(Phe) primary site of aminoacylation and the identity determinants for MSF protein. As for the cytoplasmic phenylalanyl-tRNA synthetase (Sampson, J. R., Di Renzo, A. B., Behlen, L. S., & Uhlenbeck, O. C. (1989) Science 243, 1363-1366), MSF protein recognizes nucleotides from the anticodon and the acceptor end including base A73 and, as shown here, adjacent G1-C72 base pair or at least C72 base. This indicates that the way of tRNA(Phe) binding for the two phenylalanine enzymes is conserved in evolution. However, tRNA(Phe) tertiary structure seems more critical for the interaction with the cytoplasmic enzyme than with MSF protein, and unlike cytoplasmic phenylalanyl-tRNA synthetase, the small size of the monomeric MSF protein probably does not allow contacts with residue 20 at the top corner of the L molecule. We also show that MSF protein preferentially aminoacylates the terminal 2'-OH group of tRNA(Phe) but with a catalytic efficiency for tRNA(Phe)-CC-3'-deoxyadenosine reduced 100-fold from that of native tRNA(Phe), suggesting a role of the terminal 3'-OH in catalysis. The loss is only 1.5-fold when tRNA(Phe)-CC-3'-deoxyadenosine is aminoacylated by yeast cytoplasmic PheRS (Sprinzl, M., & Cramer, F. (1973) Nature 245, 3-5), indicating mechanistic differences between the two PheRS's active sites for the amino acid transfer step.}, note = {0006-2960 Journal Article}, keywords = {Non-U.S. Gov't Variation (Genetics), Nucleic Acid Substrate Specificity Support, Phe/*chemistry/*metabolism Saccharomyces cerevisiae/enzymology/genetics Sequence Homology, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {2'-O-methyl-5-formylcytidine (f5Cm), a new modified nucleotide at the 'wobble' of two cytoplasmic tRNAs Leu (NAA) from bovine liver}, author = { J. P. Pais de Barros and G. Keith and C. El Adlouni and A. L. Glasser and G. Mack and G. Dirheimer and J. Desgres}, year = {1996}, date = {1996-01-01}, journal = {Nucleic Acids Res}, volume = {24}, number = {8}, pages = {1489-96}, abstract = {The nucleotide analysis of a cytoplasmic tRNA(Leu) isolated from bovine liver revealed the presence of an unknown modified nucleotide N. The corresponding N nucleoside was isolated by different enzymatic and chromatographic protocols from a partially purified preparation of this tRNA(Leu). Its chemical characterization was determined from its chromatographic properties, UV-absorption spectroscopy and mass spectrometric measurements, as well as from those of the borohydride reduced N nucleoside and its etheno-trimethylsilyl derivative. The structure of N was established as 2'-O-methyl-5-formylcytidine (f5CM), and its reduced derivative as 2'-O-methyl-5-hydroxy-methylcytidine (om5Cm). By sequencing the bovine liver tRNA(Leu), the structure of the anticodon was determined as f5CmAA. In addition, the nucleotide sequence showed two primary structures differing only by the nucleotide 47c which is either uridine or adenosine. The two slightly differing bovine liver tRNAs-Leu(f5CmAA) are the only tRNAs so far sequenced which contain f5Cm. The role of such a modified cytidine at the first position of the anticodon is discussed in terms of decoding properties for the UUG and UUA leucine codons. Recently, precise evidence was obtained for the presence of f5Cm at the same position in tRNAs(Leu)(NAA) isolated from rabbit and lamb liver. Therefore, the 2'-O-methyl-5-formyl modification of cytidine at position 34 could be a general feature of cytoplasmic tRNAs(Leu)(NAA) in mammals.}, note = {0305-1048 Journal Article}, keywords = {&, Acid, Acyl/*chemistry/isolation, Amino, Animals, Base, Borohydrides/chemistry, Cattle, Cells, Conformation, Cytidine/*analogs, Cytoplasm, Data, derivatives/chemistry/isolation, Fragmentography, Gov't, Hela, Human, Liver/*chemistry, Mass, Molecular, Non-U.S., Nucleic, purification, RNA, Sequence, structure, Support, Transfer}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of RNA primers in initiation of minus-strand and plus-strand DNA synthesis of the yeast retrotransposon Ty1}, author = {S Friant and T Heyman and F X Wilhelm and M Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8955910}, isbn = {8955910}, year = {1996}, date = {1996-01-01}, journal = {Biochimie}, volume = {78}, number = {7}, pages = {674-80}, abstract = {The Ty1 retrotransposon of the yeast Saccharomyces cerevisiae is a long terminal repeat mobile genetic element that transposes through an RNA intermediate. Initiation of minus-strand and plus-strand DNA synthesis are two critical steps during reverse transcription of the retrotransposon genome. Initiation of minus-strand DNA synthesis of the Ty1 element is primed by the cytoplasmic initiator methionine tRNA base paired to the primer binding site near the 5' end of the genomic RNA. A structural probing study of the primer tRNA-Ty1 RNA binary complex reveals that besides interactions between the primer binding site and the last 10 nucleotides at the 3' end of the primer tRNA, three short regions of Ty1 RNA named box 0, box 1 and box 2.1 interact with the T and D stems and loops of the primer tRNA. Some in vivo results underline the functional importance of the nucleotide sequence of the boxes and suggest that extended interactions between genomic Ty1 RNA and the primer tRNA play a role in the reverse transcription pathway. Plus-strand DNA synthesis is initiated from an RNase H resistant oligoribonucleotide spanning a purine-rich sequence, the polypurine tract (PPT). Two sites of initiation located at the 5' boundary of the 3' long terminal repeat (PPT1) and near the middle of the TyB (pol) gene in the integrase coding sequence (PPT2) have been identified in the genome of Ty1. The two PPTs have an identical sequence, TGGGTGGTA. Mutations replacing purines by pyrimidines in this sequence significantly diminish or abolish initiation of plus-strand DNA synthesis. Ty1 elements bearing a mutated PPT2 sequence are not defective for transposition whereas mutations in PPT1 abolish transposition.}, note = {0300-9084 Journal Article}, keywords = {*DNA Replication *DNA Transposable Elements DNA, Bacterial/*metabolism Support, Complementary/metabolism Molecular Sequence Data Mutagenesis Nucleic Acid Conformation RNA/*metabolism RNA, Non-U.S. Gov't}, pubstate = {published}, tppubtype = {article} } @article{lemaitre_recessive_1995, title = {A recessive mutation, immune deficiency (imd), defines two distinct control pathways in the Drosophila host defense}, author = {Bruno Lemaitre and E Kromer-Metzger and Lydia Michaut and E Nicolas and Marie Meister and Philippe Georgel and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0027-8424}, year = {1995}, date = {1995-10-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {92}, number = {21}, pages = {9465--9469}, abstract = {In this paper we report a recessive mutation, immune deficiency (imd), that impairs the inducibility of all genes encoding antibacterial peptides during the immune response of Drosophila. When challenged with bacteria, flies carrying this mutation show a lower survival rate than wild-type flies. We also report that, in contrast to the antibacterial peptides, the antifungal peptide drosomycin remains inducible in a homozygous imd mutant background. These results point to the existence of two different pathways leading to the expression of two types of target genes, encoding either the antibacterial peptides or the antifungal peptide drosomycin.}, keywords = {Animals, Anti-Bacterial Agents, Antimicrobial Cationic Peptides, Bacterial Infections, Base Sequence, Gene Expression Regulation, Genes, Glycopeptides, hoffmann, Insect, Insect Hormones, Insect Proteins, M3i, Male, Mutation, Mycoses, Nucleic Acid, Peptides, Protein Binding, Recessive, Regulatory Sequences, reichhart, Reporter, Survival Analysis}, pubstate = {published}, tppubtype = {article} } @article{levashina_metchnikowin_1995, title = {Metchnikowin, a novel immune-inducible proline-rich peptide from Drosophila with antibacterial and antifungal properties}, author = {Elena A Levashina and S Ohresser and Philippe Bulet and Jean-Marc Reichhart and Charles Hetru and Jules A Hoffmann}, issn = {0014-2956}, year = {1995}, date = {1995-10-01}, journal = {Eur. J. Biochem.}, volume = {233}, number = {2}, pages = {694--700}, abstract = {One of the characteristics of the host defense of higher insects is the rapid and transient synthesis of a variety of potent antimicrobial peptides. To date, several distinct inducible antimicrobial peptides or peptide families have been totally or partially characterized. We present here the isolation and characterization of a novel 26-residue proline-rich immune-inducible peptide from Drosophila, which exhibits both antibacterial (Gram-positive) and antifungal activities. Peptide sequencing and cDNA cloning indicate the presense of two isoforms in our Drosophila Oregon strain, which differ by one residue (His compared to Arg) as a consequence of a single nucleotide change. The gene, which maps in position 52A1-2 on the right arm of the second chromosome, is expressed in the fat body after immune challenge. The novel peptide, which we propose to name metchnikowin, is a member of a family of proline-rich peptides, and we discuss the possible evolutionary relationships within this family.}, keywords = {Animals, Anti-Bacterial Agents, Antifungal Agents, Antimicrobial Cationic Peptides, bacteria, Base Sequence, Cells, Chromosome Mapping, Cloning, Cultured, Genetic, hoffmann, M3i, Molecular, Peptides, Proline, reichhart, Transcription}, pubstate = {published}, tppubtype = {article} } @article{briand_retro-inverso_1995, title = {Retro-inverso peptidomimetics as new immunological probes. Validation and application to the detection of antibodies in rheumatic diseases}, author = {J P Briand and G Guichard and H Dumortier and S Muller}, doi = {10.1074/jbc.270.35.20686}, issn = {0021-9258}, year = {1995}, date = {1995-09-01}, journal = {The Journal of Biological Chemistry}, volume = {270}, number = {35}, pages = {20686--20691}, abstract = {Retro-inverso peptides which contain NH-CO bonds instead of CO-NH peptide bonds are much more resistant to proteolysis than L-peptides. Moreover, they have been shown recently to be able to mimic natural L-peptides with respect to poly- and monoclonal antibodies (Guichard, G., Benkirane, N., Zeder-Lutz, G., Van Regenmortel, M. H. V., Briand, J. P., and Muller, S. (1994b) Proc. Natl. Acad. Sci. U.S.A. 91, 9765-9769). We have further tested the capacity of retro-inverso peptidomimetics to serve as possible targets for antibodies produced by lupus mice and by patients with rheumatic autoimmune diseases. Several retro-inverso peptides corresponding to sequences known to be recognized by autoantibodies were synthesized, namely peptides 28-45 and 130-135 of H3, 277-291 of the Ro/SSA 52-kDa protein, and 304-324 of the Ro/SSA 60-kDa protein, and tested with autoimmune sera by enzyme-linked immunosorbent assay. We have found that retro-inverso peptides are recognized as well as or even better than natural peptides by antibodies from autoimmune patients and lupus mice. This new approach may lead to important progress in the future development of immunodiagnostic assays, particularly in the case of diseases characterized by inflammatory reactions in the course of which the level of degradative enzymes is increased.}, keywords = {Amino Acid Sequence, Animals, Antibodies, Autoantibodies, Autoimmune Diseases, Dumortier, Enzyme-Linked Immunosorbent Assay, Humans, I2CT, Immunoassay, Lupus Erythematosus, Mice, Molecular Sequence Data, Monoclonal, Peptide Fragments, Peptides, Rheumatic Diseases, Stereoisomerism, Structure-Activity Relationship, Systemic, Team-Dumortier}, pubstate = {published}, tppubtype = {article} } @article{lowenberger_insect_1995, title = {Insect immunity: isolation of three novel inducible antibacterial defensins from the vector mosquito, Aedes aegypti}, author = {C Lowenberger and Philippe Bulet and Maurice Charlet and Charles Hetru and B Hodgeman and B M Christensen and Jules A Hoffmann}, issn = {0965-1748}, year = {1995}, date = {1995-07-01}, journal = {Insect Biochem. Mol. Biol.}, volume = {25}, number = {7}, pages = {867--873}, abstract = {The injection of Escherichia coli and Micrococcus luteus into the hemocoel of Aedes aegypti induces a potent antibacterial activity in the hemolymph. We have purified and fully characterized three 40-residue antibacterial peptides from the hemolymph of bacteria-challenged mosquitoes that are absent in naive mosquitoes. The peptides are potently active against Gram-positive bacteria and against one of the Gram-negative bacteria that were tested. The amino acid sequences clearly show that the three peptides are novel isoforms of the insect defensin family of antibacterial peptides. They differ from each other by one or two amino acid residues. We present here the complete amino acid sequences of the three isoforms and the activity spectrum of the predominant Aedes defensin.}, keywords = {Aedes, Amino Acid, Animals, Anti-Bacterial Agents, Blood Proteins, Defensins, Escherichia coli, Gram-Negative Bacteria, Gram-Positive Bacteria, hoffmann, Immunity, Insect Vectors, M3i, Micrococcus luteus, Sequence Homology, Stereoisomerism}, pubstate = {published}, tppubtype = {article} } @article{bulet_insect_1995, title = {Insect immunity. The inducible antibacterial peptide diptericin carries two O-glycans necessary for biological activity}, author = {Philippe Bulet and G Hegy and J Lambert and Alan van Dorsselaer and Jules A Hoffmann and Charles Hetru}, issn = {0006-2960}, year = {1995}, date = {1995-06-01}, journal = {Biochemistry}, volume = {34}, number = {22}, pages = {7394--7400}, abstract = {A bacterial challenge of larvae of the dipteran insect Phormia terranovae induces the rapid synthesis of diptericin, an antibacterial polypeptide, previously characterized at the amino acid level and indirectly by cDNA cloning studies. This 82-residue polypeptide consists of an N-terminal proline-rich domain and a central and C-terminal glycine-rich domain. Using liquid chromatography coupled to electrospray ionization-mass spectrometry, we demonstrate here that this molecule is more complex than anticipated and carries two O-substitutions on threonine residues, one in the proline-rich domain (residue 10) and one in the glycine-rich domain (residue 54). These substitutions consist of identical trisaccharides: glucose--textgreatergalactose--textgreaterN-acetylgalactosamine--textgreater(threonine). Treatment of diptericin with O-glycosidase, which selectively removes the substitutions without altering the polypeptide proper, abolishes the antibacterial activity, indicating that this posttranslational modification is essential for biological activity of the polypeptide. We also show that diptericin is posttranslationally modified by a C-terminal amidation.}, keywords = {Animals, Anti-Bacterial Agents, Carbohydrate Sequence, Carbohydrates, Diptera, Escherichia coli, Glycopeptides, Hemolymph, hoffmann, Insect Hormones, Insect Proteins, Larva, M3i, Mass Spectrometry, Plants, Trisaccharides}, pubstate = {published}, tppubtype = {article} } @article{cornet_refined_1995, title = {Refined three-dimensional solution structure of insect defensin A}, author = {B Cornet and J M Bonmatin and Charles Hetru and Jules A Hoffmann and M Ptak and F Vovelle}, issn = {0969-2126}, year = {1995}, date = {1995-05-01}, journal = {Structure}, volume = {3}, number = {5}, pages = {435--448}, abstract = {BACKGROUND: Insect defensin A is a basic 4 kDa protein secreted by Phormia terranovae larvae in response to bacterial challenges or injuries. Previous biological tests suggest that the bacterial cytoplasmic membrane is the target of defensin A. The structural study of this protein is the first step towards establishing a structure-activity relationship and forms the basis for understanding its antibiotic activity at the molecular level. RESULTS: We describe a refined model of the three-dimensional structure of defensin A derived from an extensive analysis of 786 inter-proton nuclear Overhauser effects. The backbone fold involves an N-terminal loop and an alpha-helical fragment followed by an antiparallel beta-structure. The helix and the beta-structure are connected by two of the three disulphide bridges present in defensin A, forming a so-called 'cysteine-stabilized alpha beta' (CS alpha beta) motif. The N-terminal loop, which is locally well defined, can occupy different positions with respect to the other moieties of the molecule. CONCLUSIONS: The CS alpha beta motif, which forms the core of the defensin A structure, appears to be a common organization for several families of small proteins with toxic properties. The distribution of amino acid side chains in the protein structure creates several hydrophobic or hydrophilic patches. This leads us to propose that the initial step in the action of positively charged defensin A molecules with cytoplasmic membranes may involve interactions with acidic phospholipids.}, keywords = {Amino Acid, Animals, Bacteriolysis, Chemistry, Defensins, Diptera, Gram-Positive Bacteria, hoffmann, Hydrogen Bonding, Insect Hormones, M3i, Magnetic Resonance Spectroscopy, Models, Molecular, Physical, Physicochemical Phenomena, Protein Conformation, Recombinant Proteins, Sequence Homology, Solutions, Structure-Activity Relationship}, pubstate = {published}, tppubtype = {article} } @article{georgel_drosophila_1995, title = {Drosophila immunity. A sequence homologous to mammalian interferon consensus response element enhances the activity of the diptericin promoter}, author = {Philippe Georgel and Christine Kappler and E Langley and I Gross and E Nicolas and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0305-1048}, year = {1995}, date = {1995-04-01}, journal = {Nucleic Acids Res.}, volume = {23}, number = {7}, pages = {1140--1145}, abstract = {Bacterial challenge of larvae or adults of Drosophila induces the rapid transcription of several genes encoding antibacterial peptides with a large spectrum of activity. One of these peptides, the 82-residue anti-gram negative diptericin, is encoded by a single intronless gene and we are investigating the control of expression of this gene. Previous studies using both transgenic experiments and footprint analysis have highlighted the role in the induction of this gene of a 30 nucleotide region which contains three partially overlapping motifs with sequence homology to mammalian NF-kappa B and NF-IL6 response elements and to the GAAANN sequence present in the interferon consensus response elements of some mammalian interferon-induced genes. We now show that the latter sequence binds in immune responsive tissues (fat body, blood cells) of Drosophila a approximately 45 kDa polypeptide which cross-reacts with a polyserum directed against mammalian interferon Regulatory Factor-I. Using a transfection assay of Drosophila tumorous blood cells, we show that the GAAANN sequence positively regulates the activity of the diptericin promoter. We propose that this motif cooperatively interacts with the other response elements in the regulation of the diptericin gene expression.}, keywords = {Animals, Base Sequence, CCAAT-Enhancer-Binding Proteins, DNA, DNA-Binding Proteins, Genes, Genetic, hoffmann, Immunity, Insect, Insect Hormones, Insect Proteins, interferons, Lipopolysaccharides, M3i, NF-kappa B, Nuclear Proteins, Plasmids, Promoter Regions, reichhart, Up-Regulation}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_innate_1995, title = {Innate immunity of insects}, author = {Jules A Hoffmann}, issn = {0952-7915}, year = {1995}, date = {1995-02-01}, journal = {Curr. Opin. Immunol.}, volume = {7}, number = {1}, pages = {4--10}, abstract = {Insects are particularly resistant to microorganisms. Their host-defense system relies on several innate reactions: upon injury, the immediate onset of two proteolytic cascades leading to localized blood clotting and to melanization, the latter process involving production of cytotoxic molecules (namely reactive oxygen intermediates); the phagocytosis of bacteria and the encapsulation of larger parasites by blood cells; the induced synthesis by the fat body of a battery of potent antimicrobial peptides/polypeptides which are secreted into the hemolymph where they act synergistically to kill the invading microorganisms. The insect host defence system shares many of the basic characteristics of the mammalian acute phase response, especially at the level of the coordinate control of gene expression, where similar cis-regulatory and inducible transactivators appear to play key functions. The powerful techniques developed to study the genetics of Drosophila provide a unique opportunity to dissect the development and differentiation of this primordial immune system and may contribute to our understanding of the innate immune response in higher organisms.}, keywords = {Animals, Anti-Bacterial Agents, Blood Proteins, Cellular, Defensins, Gene Expression Regulation, hoffmann, Immunity, Innate, insects, M3i, Peptides}, pubstate = {published}, tppubtype = {article} } @article{, title = {Cleavage of tRNA with imidazole and spermine imidazole constructs: a new approach for probing RNA structure}, author = {V V Vlassov and G Zuber and B Felden and J P Behr and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7667092}, isbn = {7667092}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Res}, volume = {23}, number = {16}, pages = {3161-3167}, abstract = {Hydrolysis of RNA in imidazole buffer and by spermine-imidazole conjugates has been investigated. The RNA models were yeast tRNA(Asp) and a transcript derived from the 3'-terminal sequence of tobacco mosaic virus RNA representing a minihelix capable of being enzymatically aminoacylated with histidine. Imidazole buffer and spermine-imidazole conjugates in the presence of free imidazole cleave phosphodiester bonds in the folded RNAs in a specific fashion. Imidazole buffer induces cleavages preferentially in single-stranded regions because nucleotides in these regions have more conformational freedom and can assume more easily the geometry needed for formation of the hydrolysis intermediate state. Spermine-imidazole constructs supplemented with free imidazole cleave tRNA(Asp) within single-stranded regions after pyrimidine residues with a marked preference for pyrimidine-A sequences. Hydrolysis patterns suggest a cleavage mechanism involving an attack by the imidazole residue of the electrostatically bound spermine-imidazole and by free imidazole at the most accessible single-stranded regions of the RNA. Cleavages in a viral RNA fragment recapitulating a tRNA-like domain were found in agreement with the model of this molecule that accounts for its functional properties, thus illustrating the potential of the imidazole-derived reagents as structural probes for solution mapping of RNAs. The cleavage reactions are simple to perform, provide information reflecting the state of the ribose-phosphate backbone of RNA and can be used for mapping single- and double-stranded regions in RNAs.}, note = {0305-1048 Journal Article}, keywords = {Asp/*chemistry/genetics/*metabolism RNA, Base Sequence Binding Sites Buffers Hydrolysis Imidazoles Molecular Probes Molecular Sequence Data Molecular Structure Nucleic Acid Conformation RNA, Fungal/chemistry/genetics/metabolism RNA, Non-U.S. Gov't Tobacco Mosaic Virus/genetics/metabolism, Transfer, Unité ARN, Viral/chemistry/genetics/metabolism Saccharomyces cerevisiae/genetics/metabolism Spermine Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selenocysteylation in eukaryotes necessitates the uniquely long aminoacyl acceptor stem of selenocysteine tRNA(Sec)}, author = {C Sturchler-Pierrat and N Hubert and T Totsuka and T Mizutani and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7629188}, isbn = {7629188}, year = {1995}, date = {1995-01-01}, journal = {J Biol Chem}, volume = {270}, number = {31}, pages = {18570-18574}, abstract = {Selenocysteine synthesis is achieved on a specific tRNA, tRNA(Sec), which is first charged with serine to yield seryl-tRNA(Sec). Eukaryotic tRNA(Sec) exhibits an aminoacyl acceptor stem with a unique length of 9 base pairs. Within this stem, two base pairs, G5a.U67b and U6.U67, drew our attention, whose non-Watson-Crick status is maintained in the course of evolution either through U6.U67 base conservation or base covariation at G5a.U67b. Single or double point mutations were performed, which modified the identity of either or both of the base pairs. Serylation by seryl-tRNA synthetase was unaffected by substitutions at either G5a.U67b or U6.U67. Instead, and quite surprisingly, changing G5a.U67b and U6.U67 to G5a-C67b/U6.G67 or G5a-C67b/C6-G67 gave rise to a tRNA(Sec) mutant exhibiting a gain of function in serylation. This finding sheds light on the negative influence born by a few base pairs in the acceptor stem of tRNA(Sec) on its serylation abilities. The tRNA(Sec) capacities to support selenocysteylation were next examined with regard to a possible role played by the two non-Watson-Crick base pairs and the unique length of the acceptor stem. It first emerges from our study that tRNA(Sec) transcribed in vitro is able to support selenocysteylation. Second, none of the point mutations engineered at G5a.U67b and/or U6.U67 significantly modified the selenocysteylation level. In contrast, reduction of the acceptor stem length to 8 base pairs led tRNA(Sec) to lose its ability to efficiently support selenocysteylation. Thus, our study provides strong evidence that the length of the acceptor stem is of prime importance for the serine to selenocysteine conversion step.}, note = {0021-9258 Journal Article}, keywords = {Acylation Animals Base Composition Base Sequence Cattle Eukaryotic Cells Kinetics Molecular Sequence Data Mutagenesis, Amino Acid-Specific RNA, Amino Acyl/*biosynthesis/*genetics/metabolism Selenocysteine/*biosynthesis Sequence Deletion Serine-tRNA Ligase/*metabolism Structure-Activity Relationship Support, Genetic, Non-U.S. Gov't Transcription, Site-Directed Phylogeny *RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Selenium, selenoproteines: une autre lecture du code genétique.}, author = {C Sturchler-Pierrat and P Carbon and A Krol}, url = {http://vinci.bibli.be/opac/index.php?lvl=bulletin_display&id=638}, year = {1995}, date = {1995-01-01}, journal = {Med Sci (Paris)}, volume = {11}, number = {8}, pages = {1081-1088}, abstract = {Le sélénium est un élément présent à l'état de trace, trace dont l'importance physiologique n'a été appréciée à sa juste valeur que dans un passé récent. Sa forme biologique consiste essentiellement en l'acide aminé sélénocystéine, composé n'existant pas naturellement à l'état libre dans la cellule. Une machinerie complexe mais originale, mettant en oeuvre plusieurs produits géniques, insère cet acide aminé de manière cotraductionnelle. Parmi les sélénoenzymes identifiées à l'heure actuelle chez les mammifères, les glutathion peroxydases constituent une des lignes de défense contre les agressions par les radicaux libres oxygénés; la tétra-iodothyronine-5'-désiodinase, pour sa part, active l'hormone thyroïdienne. Ces deux exemples mettent clairement en exergue l'importance cruciale du sélénium, aussi bien dans la prévention des dommages causés aux macromolécules biologiques qu'au cours du développement.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effect of macromolecular impurities on lysozyme solubility and crystallizability: dynamic light scattering, phase diagram, and crystal growth studies}, author = {M Skouri and B Lorber and R Giege and J P Munch and S J Candau}, url = {http://www.sciencedirect.com/science/article/pii/0022024895000518}, isbn = {10.1016/0022-0248(95)00051-8}, year = {1995}, date = {1995-01-01}, journal = {J Crystal Growth}, volume = {152}, number = {3}, pages = {209-220}, abstract = {The effects of macromolecular impurities on protein solubility and crystallizability were investigated by dynamic light scattering and crystal growth experiments using hen egg-white lysozyme as the model protein. In the presence of traces of protein impurities, representing no more than 2% (w/w) of the total protein, the average diffusion coefficients of the macromolecular particles found in undersaturated lysozyme solutions are significantly lower than those measured with purest lysozyme preparations. This fact is explained by the simultaneous existence of individual molecules and of large size aggregates in contaminated solutions, as indicated by the bimodal light scattering autocorrelation function. Controlled contamination experiments in which ovalbumin or conalbumin were added to purest lysozyme indicate that aggregates result from heterogeneous association of lysozyme molecules with the structurally unrelated proteins. These aggregates might become starting points for heterogeneous nucleation leading to the growth of ill-shaped microcrystals. Aggregates in under- or supersaturated lysozyme solutions containing NaCl can be eliminated by filtration over microporous membranes. As a result the number of ill-shaped crystals diminishes drastically; that of well-shaped tetragonal crystals decreases also but their size increases.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The presence of a D-stem but not a T-stem is essential for triggering aminoacylation upon anticodon binding in yeast methionine tRNA}, author = {B Senger and R Aphasizhev and P Walter and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7776375}, isbn = {7776375}, year = {1995}, date = {1995-01-01}, journal = {J Mol Biol}, volume = {249}, number = {1}, pages = {45-58}, abstract = {Dissection of the yeast cytoplasmic initiator tRNA(Met) into two helical domains, the T psi C acceptor and anticodon minihelices, failed to show anminoacylation and binding of the acceptor minihelix by the yeast methionyl-tRNA synthetase (MetRS) even in the presence of the anticodon minihelix. In contrast, based on the measure of the inhibition constant Ki, the anticodon minihelix carrying the methionine anticodon CAU is specifically bound to the synthetase and with an affinity comparable to that of the full-length tRNA. The yeast tRNA(Met) acceptor and anticodon minihelices were covalently linked using the central core sequences of either bovine mitochondrial tRNA(Ser) (AGY) lacking a D-stem or initiator tRNA(Met) from Caenorhabditis elegans lacking a T-stem. Based on modeling studies of analogous constructs performed by others, we assume that the folding and distance between the anticodon and acceptor ends of these hybrid tRNAs are identical to that of canonical tRNA. The three-quarter molecule, which includes the T-stem, has aminoacylation activity significantly more than an acceptor minihelix, while the acceptor stem/anticodon-D stem biloop has near wild-type aminoacylation activity. These results suggest that the high selectivity of the anticodon bases in tRNA(Met) depends upon the tRNA L-shape conformation and the presence of a D-arm. Protein contacts with the D-arm phosphate backbone are required for connecting anticodon recognition with the active site. These interactions probably contribute to fine tune the position of the acceptor end in the active site, allowing entry into the transition state of aminoacylation upon anticodon binding. The importance of an L structure for recognition of tRNA(Met) by yeast MetRS was also deduced from mutations of tertiary interactions known to play a general role in tRNA folding.}, note = {0022-2836 Journal Article}, keywords = {Acylation Animals Anticodon/metabolism Base Sequence Caenorhabditis elegans Cattle Molecular Sequence Data Molecular Structure Mutation RNA, Met/chemistry/genetics/*metabolism Saccharomyces cerevisiae/*metabolism Support, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Staf, a novel zinc finger protein that activates the RNA polymerase III promoter of the selenocysteine tRNA gene}, author = {C Schuster and E Myslinski and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7641696}, isbn = {7641696}, year = {1995}, date = {1995-01-01}, journal = {EMBO J}, volume = {14}, number = {15}, pages = {3777-3787}, abstract = {The selenocysteine tRNA gene (tRNA(Sec)) is atypical. Though transcribed by RNA polymerase III like all other tRNA genes, its basal promoter elements are distinct and reside essentially upstream of the coding region. In addition, transcription from the basal promoter is activated by a 15 bp activator element. In this report we describe the cloning and functional characterization of Staf (selenocysteine tRNA gene transcription activating factor), a novel Xenopus laevis transcription factor which binds to the tRNA(Sec) activator element and mediates its activation properties. The 600 amino acid Staf protein contains seven zinc fingers and a separate acidic activation domain. Seven highly conserved regions were detected between Staf and human ZNF76, a protein of unknown function, thereby aiding in predicting the locations of the functional domains of Staf. With the use of a novel expression assay in X.laevis oocytes we succeeded in demonstrating that Staf can activate the RNA polymerase III promoter of the tRNA(Sec) gene. This constitutes the first demonstration of the capacity of a cloned factor to activate RNA polymerase III transcription in vivo.}, note = {0261-4189 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Animals Base Sequence Cloning, Amino Acid-Specific/*genetics Recombinant Fusion Proteins/metabolism Sequence Analysis, DNA Sequence Homology, Messenger RNA, Molecular DNA/metabolism DNA-Binding Proteins/biosynthesis/genetics/*metabolism Gene Expression Genes, Non-U.S. Gov't Trans-Activation (Genetics)/*physiology Trans-Activators/biosynthesis/genetics/*metabolism Xenopus laevis *Zinc Fingers, Reporter Human Molecular Sequence Data Oocytes Promoter Regions (Genetics)/*genetics RNA Polymerase III/*genetics RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular dissection of the pseudoknot governing the translational regulation of Escherichia coli ribosomal protein S15}, author = {C Philippe and L Benard and C Portier and E Westhof and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7532857}, isbn = {7532857}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Res}, volume = {23}, number = {1}, pages = {18-28}, abstract = {The ribosomal protein S15 controls its own translation by binding to a mRNA region overlapping the ribosome binding site. That region of the mRNA can fold in two mutually exclusive conformations that are in dynamic equilibrium: a structure with two hairpins and a pseudoknot. A mutational analysis provided evidence for the existence and requirement of the pseudoknot for translational control in vivo and S15 recognition in vitro. In this study, we used chemical probing to analyze the structural consequences of mutations and their effect on the stem-loop/pseudoknot equilibrium. Interactions between S15 and the pseudoknot structure were further investigated by footprinting experiments. These data, combined with computer modelling and the previously published data on S15 binding and in vivo control, provide important clues on pseudoknot formation and S15 recognition. An unexpected result is that the relevant control element, here the pseudoknot form, can exist in a variety of topologically equivalent structures recognizable and shapable by S15. S15 sits on the deep groove of the co-axial stack and makes contacts with both stems, shielding the bridging adenine. The only specific sequence determinants are found in the helix common to the pseudoknot and the hairpin structures.}, note = {0305-1048 Journal Article}, keywords = {Bacterial/genetics/metabolism RNA, Base Sequence Binding Sites/genetics Escherichia coli/*genetics/metabolism Models, Genetic, Messenger/genetics/metabolism Ribosomal Proteins/*chemistry/*genetics/metabolism Support, Molecular Molecular Sequence Data Mutation Protein Conformation RNA, Non-U.S. Gov't Thermodynamics *Translation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genetic probes of ribosomal RNA function}, author = {M O'Connor and C A Brunelli and M A Firpo and S T Gregory and K R Lieberman and J S Lodmell and H Moine and D I Van Ryk and A E Dahlberg}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8722001}, isbn = {8722001}, year = {1995}, date = {1995-01-01}, journal = {Biochem Cell Biol}, volume = {73}, number = {11-12}, pages = {859-868}, abstract = {We have used a genetic approach to uncover the functional roles of rRNA in protein synthesis. Mutations were constructed in a cloned rrn operon by site-directed mutagenesis or isolated by genetic selections following random mutagenesis. We have identified mutations that affect each step in the process of translation. The data are consistent with the results of biochemical and phylogenetic analyses but, in addition, have provided novel information on regions of rRNA not previously investigated.}, note = {0829-8211 Journal Article Review Review, Tutorial}, keywords = {16S/genetics RNA, Base Sequence Molecular Sequence Data Mutation Nucleic Acid Conformation RNA Probes RNA, Messenger/genetics RNA, Non-U.S. Gov't Support, P.H.S., Ribosomal, Ribosomal/*genetics RNA, Transfer/genetics Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding of the HIV-1 nucleocapsid protein to the primer tRNA(3Lys), in vitro, is essentially not specific}, author = {Y Mely and H de Rocquigny and M Sorinas-Jimeno and G Keith and B P Roques and R Marquet and D Gerard}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7829498}, isbn = {7829498}, year = {1995}, date = {1995-01-01}, journal = {J Biol Chem}, volume = {270}, number = {4}, pages = {1650-1656}, abstract = {The nucleocapsid protein NCp7 of human immunodeficiency virus, type 1, is a key component in the viral life cycle. Since, the first common step of all reported NCp7 activities corresponds to a nucleic acid-binding step, the NCp7 binding parameters to the natural primer tRNA(3Lys) were investigated. Using NCp7 intrinsic fluorescence, we found that (i) in 0.1 M NaCl, NCp7 bound noncooperatively to tRNA(3Lys) with a Kobs = 3.2 x 10(6) M-1 association constant and a n = 6 binding site size, (ii) four ionic interactions were formed in the NCp7.tRNA(3Lys) complex, and (iii) nonelectrostatic factors provided about 60% of the binding energy. These binding parameters were not significantly altered when the natural tRNA(3Lys) was replaced by either an in vitro synthetic tRNA(3Lys) transcript, the heterologous yeast tRNA(Phe) or the structurally unrelated 5 S RNA from Escherichia coli. Moreover, the environment of the intrinsic fluorescent reporters (Trp37 and Trp61) was similar in the various complexes. Finally, experiments performed at low protein concentration provide no evidence of high affinity binding sites. Taken together, our data strongly suggested an essentially nonspecific binding of NCp7 to tRNA(3Lys) and thus did not seem to support a direct role of NCp7, per se, in the selection of tRNA(3Lys) from the pool of cellular tRNAs.}, note = {0021-9258 Journal Article}, keywords = {Amino Acid Sequence Capsid/*chemistry/*metabolism *Capsid Proteins Escherichia coli Gene Products, Amino Acyl/biosynthesis/chemistry/*metabolism Substrate Specificity Support, gag/*chemistry/*metabolism HIV-1/*metabolism Hydrogen-Ion Concentration Kinetics Magnesium Chloride/pharmacology Mathematics Models, Genetic Zinc Fingers, MARQUET, Non-U.S. Gov't Transcription, Theoretical Molecular Sequence Data Nucleic Acid Conformation Osmolar Concentration Protein Binding RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genetic selection for active E.coli amber tRNA(Asn) exclusively led to glutamine inserting suppressors}, author = {F Martin and G Eriani and J Reinbolt and G Dirheimer and J Gangloff}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7708493}, isbn = {7708493}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Res}, volume = {23}, number = {5}, pages = {779-784}, abstract = {Suppressor tRNAs are useful tools for determining identity elements which define recognition of tRNAs in vivo by their cognate aminoacyl-tRNA synthetases. This study was aimed at the isolation of active amber tRNA(Asn). Nineteen mutated tRNA(Asn)CUA having amber suppressor activity were selected by an in vivo genetic screen, and all exclusively inserted glutamine. From analysis of the different mutations it is concluded that glutamine accepting activity was obtained upon reducing the interaction strength between the first base pair of the tRNA(Asn)CUA by direct or indirect effects. Failure to isolate tRNA(Asn)CUA suppressors charged with asparagine as well as other evolutionary related amino acids is discussed.}, note = {0305-1048 Journal Article}, keywords = {Asn/*genetics Support, Base Sequence Escherichia coli/*genetics *Genes, ERIANI, Insertional RNA, Non-U.S. Gov't, Suppressor Glutamine/*genetics Molecular Sequence Data Mutagenesis, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNAs as primer of reverse transcriptases}, author = {R Marquet and C Isel and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7541250}, isbn = {7541250}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {113-124}, abstract = {Genetic elements coding for proteins that present amino acid identity with the conserved motifs of retroviral reverse transcriptases constitute the retroid family. With the exception of reverse transcriptases encoded by mitochondrial plasmids of Neurospora, all reverse transcriptases have an absolute requirement for a primer to initiate DNA synthesis. In retroviruses, plant pararetroviruses, and retrotransposons (transposons containing long terminal repeats), DNA synthesis is primed by specific tRNAs. All these retroelements contain a primer binding site presenting a Watson-Crick complementarity with the primer tRNA. The tRNAs most widely used as primers are tRNA(Trp), tRNA(Pro), tRNA(1,2Lys), tRNA(3Lys), tRNA(iMet). Other tRNAs such as tRNA(Gln), tRNA(Leu), tRNA(Ser), tRNA(Asn) and tRNA(Arg) are also occasionally used as primers. In the retroviruses and plant pararetroviruses, the primer binding site is complementary to the 3' end of the primer tRNA. In the case of retrotransposons, the primer binding site is either complementary to the 3' end or to an internal region of the primer tRNA. Additional interactions taking place between the primer tRNA and the retro-RNA outside of the primer binding site have been evidenced in the case of Rous sarcoma virus, human immunodeficiency virus type I, and yeast retrotransposon Ty1. A selective encapsidation of the primer tRNA, probably promoted by interactions with reverse transcriptase, occurs during the formation of virus or virus-like particles. Annealing of the primer tRNA to the primer binding site appears to be mediated by reverse transcriptase and/or the nucleocapsid protein. Modified nucleosides of the primer tRNA have been shown to be important for replication of the primer binding site, encapsidation of the primer (in the case of Rous sarcoma virus), and interaction with the genomic RNA (in the case of human immunodeficiency virus type I).}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Binding Sites DNA Nucleotidylexotransferase/genetics/metabolism Hepadnaviridae/genetics Introns Plant Viruses/genetics Plasmids RNA/chemistry/*metabolism RNA, MARQUET, Non-U.S. Gov't, Transfer/chemistry/*metabolism RNA, Unité ARN, Viral/genetics/*metabolism RNA-Directed DNA Polymerase/*metabolism Retroelements/genetics Retroviridae/genetics Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure of glycyl-tRNA synthetase from Thermus thermophilus}, author = {D T Logan and M H Mazauric and D Kern and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7556056}, isbn = {7556056}, year = {1995}, date = {1995-01-01}, journal = {EMBO J}, volume = {14}, number = {17}, pages = {4156-4167}, abstract = {The sequence and crystal structure at 2.75 A resolution of the homodimeric glycyl-tRNA synthetase from Thermus thermophilus, the first representative of the last unknown class II synthetase subgroup, have been determined. The three class II synthetase sequence motifs are present but the structure was essential for identification of motif 1, which does not possess the proline previously believed to be an essential class II invariant. Nevertheless, crucial contacts with the active site of the other monomer involving motif 1 are conserved and a more comprehensive description of class II now becomes possible. Each monomer consists of an active site strongly resembling that of the aspartyl and seryl enzymes, a C-terminal anticodon recognition domain of 100 residues and a third domain unusually inserted between motifs 1 and 2 almost certainly interacting with the acceptor arm of tRNA(Gly). The C-terminal domain has a novel five-stranded parallel-antiparallel beta-sheet structure with three surrounding helices. The active site residues most probably responsible for substrate recognition, in particular in the Gly binding pocket, can be identified by inference from aspartyl-tRNA synthetase due to the conserved nature of the class II active site.}, note = {0261-4189 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Binding Sites Comparative Study Crystallography, Bacterial Glycine-tRNA Ligase/*chemistry/genetics/isolation & purification Macromolecular Systems Models, Molecular Molecular Sequence Data Polymerase Chain Reaction *Protein Structure, Non-U.S. Gov't Thermus thermophilus/*enzymology, Secondary Sequence Homology, Unité ARN, X-Ray/methods DNA Probes Genes}, pubstate = {published}, tppubtype = {article} } @article{, title = {Detection of DNA adducts in declining hop plants grown on fields formerly treated with heptachlor, a persistent insecticide}, author = {A Laouedj and C Schenk and A Pfohl-Leszkowicz and G Keith and D Schontz and P Guillemaut and B Rether}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15091476}, isbn = {15091476}, year = {1995}, date = {1995-01-01}, journal = {Environ Pollut}, volume = {90}, number = {3}, pages = {409-414}, abstract = {Hop decline was observed in Alsace, eastern France, in reparcelled sugar beet fields formerly abundantly treated with an insecticide, heptachlor. Leaves were collected from 'declining hops' grown in an heptachlor-contaminated area and from 'healthy hops' grown in a soil not contaminated by heptachlor. These two samples came from hop vines treated with other usual pesticides. 'Control' hop leaves came from soil neither treated with pesticide nor contaminated with heptachlor. Hypermodified nucleotides (DNA adducts) were detected using the (32)P-postlabelling method. No detectable DNA adducts were found in the 'control' specimen, whereas eight adducts were detected in the 'healthy hops' specimen, probably due to the usual pesticide treatment. However, 16 adducts, nine of which were new adducts, could be detected in the 'declining hops' specimen. It may therefore be supposed that the presence of these hypermodified nucleotides perturbs gene expression and so contributes to the hop decline. In addition, to confirm the genotoxicity of heptachlor, it is shown that it induces DNA adducts in bean-cell suspension culture as well. Finally, it is proposed, in the case of alternate cultures scheduled in fields which were formerly treated with pesticides, adapted to other cultures, that particular attention should be given to the history of the soils.}, note = {0269-7491 Journal Article}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Footprinting of tRNA(Phe) transcripts from Thermus thermophilus HB8 with the homologous phenylalanyl-tRNA synthetase reveals a novel mode of interaction}, author = {R Kreutzer and D Kern and R Giege and J Rudinger}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8524648}, isbn = {8524648}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Res}, volume = {23}, number = {22}, pages = {4598-4602}, abstract = {The phosphates of the tRNA(Phe) transcript from Thermus thermophilus interacting with the cognate synthetase were determined by footprinting. Backbone bond protection against cleavage by iodine of the phosphorothioate-containing transcripts was found in the anticodon stem-loop, the D stem-loop and the acceptor stem and weak protection was also seen in the variable loop. Most of the protected phosphates correspond to regions around known identity elements of tRNA(Phe). Enhancement of cleavage at certain positions indicates bending of tRNAPhe upon binding to the enzyme. When applied to the three-dimensional model of tRNA(Phe) from yeast the majority of the protections occur on the D loop side of the molecule, revealing that phenylalanyl-tRNA synthetase has a rather complex and novel pattern of interaction with tRNAPhe, differing from that of other known class II aminoacyl-tRNA synthetases.}, note = {0305-1048 Journal Article}, keywords = {Base Composition Base Sequence Cloning, Genetic, Molecular Comparative Study Escherichia coli Kinetics Models, Phe/biosynthesis/chemistry/*metabolism Thermus thermophilus/*enzymology/*genetics *Transcription, Structural Molecular Sequence Data Nucleic Acid Conformation Phenylalanine-tRNA Ligase/*metabolism Protein Binding RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Postlabeling: a sensitive method for studying DNA adducts and their role in carcinogenesis}, author = {G Keith and G Dirheimer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7894079}, isbn = {7894079}, year = {1995}, date = {1995-01-01}, journal = {Curr Opin Biotechnol}, volume = {6}, number = {1}, pages = {3-11}, abstract = {The covalent binding of xenobiotics to DNA is an important trigger of the multistage process that leads to carcinogenesis. 32P-postlabeling represents a highly sensitive method for biomonitoring exposure to genotoxic agents and for cancer risk assessment; it is capable of detecting less than one DNA adduct per human genome. Recent improvements to the technique have shown that the resistance of adducted DNA to enzyme digestion may lead to an overestimation of the number of different adducts present in a sample.}, note = {0958-1669 Journal Article Review Review, Academic}, keywords = {Animals Base Sequence Cell Division *Cell Transformation, Genetic Molecular Sequence Data Mutagenesis Neoplasms/*genetics/pathology Phosphorus Radioisotopes Precancerous Conditions/genetics/pathology Radioisotope Dilution Technique Sensitivity and Specificity Support, Human Human Models, Neoplastic DNA Adducts/*analysis *Genome, Non-U.S. Gov't Xenobiotics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mobilities of modified ribonucleotides on two-dimensional cellulose thin-layer chromatography}, author = {G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7599271}, isbn = {7599271}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {142-144}, note = {0300-9084 Journal Article}, keywords = {*Chromatography, Thin Layer Purines/chemistry Pyrimidines/chemistry RNA, Transfer/chemistry Ribonucleases/metabolism Ribonucleotides/*chemistry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Initiation of reverse transcription of HIV-1: secondary structure of the HIV-1 RNA/tRNA(3Lys) (template/primer)}, author = {C Isel and C Ehresmann and G Keith and B Ehresmann and R Marquet}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7707372}, isbn = {7707372}, year = {1995}, date = {1995-01-01}, journal = {J Mol Biol}, volume = {247}, number = {2}, pages = {236-250}, abstract = {Reverse transcription of human immunodeficiency virus type-1 (HIV-1) genomic RNA is primed by tRNA(3Lys), whose 3' end 18 nucleotides are complementary to the viral primer binding site (PBS). We used chemical and enzymatic probes to test the conformation of the viral RNA and tRNA(3Lys), in their free form and in the HIV-1 RNA/tRNA(3Lys) binary complex. Extensive reactivity changes were observed in both molecules upon formation of the binary complex. In the viral RNA, reactivity changes occurred up to 69 nucleotides upstream and 72 nucleotides downstream of the PBS. A secondary structure model of the HIV-1 RNA/tRNA(3Lys) complex accounting for all probing data has been constructed. It reveals an unexpectedly complex and compact pseudoknot-like structure in which most of the anticodon loop, the 3' strand of the anticodon stem and the 5' part of the variable loop of tRNA(3Lys) interact with viral sequences 12 to 39 nucleotides upstream of the PBS. The core of the binary complex is a complex junction formed by two single-stranded sequences of tRNA(3Lys), an intramolecular viral helix, an intramolecular tRNA helix, and two intermolecular helices formed by the template/primer interaction. This junction probably highly constrains the tertiary structure of the HIV-1 RNA/tRNA(3Lys) complex. Compared to the structure of the free molecules, only the D arm of tRNA(3Lys) and a small viral stem-loop downstream of the PBS are unaffected in the binary complex. Sequence comparison reveals that the main characteristics of the binary complex model are conserved among all HIV-1 isolates.}, note = {0022-2836 Journal Article}, keywords = {Base Sequence Binding Sites Conserved Sequence HIV-1/*genetics Models, Genetic, Lys/*genetics/metabolism RNA, MARQUET, Molecular Molecular Probes Molecular Sequence Data *Nucleic Acid Conformation RNA, Non-U.S. Gov't *Transcription, Transfer, Unité ARN, Viral/*genetics/metabolism Structure-Activity Relationship Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Plus-strand DNA synthesis of the yeast retrotransposon Ty1 is initiated at two sites, PPT1 next to the 3' LTR and PPT2 within the pol gene. PPT1 is sufficient for Ty1 transposition}, author = {T Heyman and B Agoutin and S Friant and F X Wilhelm and M L Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7563090}, isbn = {7563090}, year = {1995}, date = {1995-01-01}, journal = {J Mol Biol}, volume = {253}, number = {2}, pages = {291-303}, abstract = {Long terminal repeat elements and retroviruses require primers for initiation of minus and plus-strand DNA synthesis by reverse transcriptase. Here we demonstrate genetically that plus-strand DNA synthesis of the yeast Ty1 element is initiated at two sites located at the 5' boundary of the 3' long terminal repeat (PPT1) and near the middle of the pol gene in the integrase coding sequence (PPT2). A consequence of the presence of two PPTs is that Ty1 plus-strand DNA exists as segments at some time during replication. Three fragments have been identified: the plus-strand strong-stop DNA initiated at PPT1, a downstream fragment initiated at PPT2 and an upstream fragment spanning the 5'-terminal part of Ty1 and a portion of the TyB gene. Characterization of the 3' ends of the plus-strand DNA fragments reveals (1) that the upstream fragment is elongated beyond PPT2 creating a plus-strand overlap and (2) that the majority of plus-strand strong-stop DNA fragments bear a copy of the minus-strand primer binding site in agreement with the accepted model of retroviral genomic RNA reverse transcription. The two polypurine tracts, PPT1 and PPT2, have an identical sequence GGGTGGTA. Mutations replacing purines by pyrimidines in this sequence significantly diminish or abolish initiation of plus-strand synthesis. Ty1 elements bearing a mutated PPT2 sequence are not defective for transposition whereas mutations in PPT1 abolish transposition.}, note = {0022-2836 Journal Article}, keywords = {Base Sequence Cloning, Fungal *Genes, Fungal/biosynthesis DNA, Genetic, Molecular DNA Primers *DNA Replication DNA, Non-U.S. Gov't Transcription, Nucleic Acid Restriction Mapping *Retroelements Saccharomyces cerevisiae/genetics/*virology Support, pol Genome, Unité ARN, Viral Molecular Sequence Data Poly C/analysis Polymerase Chain Reaction *Repetitive Sequences, Viral/*biosynthesis Genes}, pubstate = {published}, tppubtype = {article} } @article{, title = {An interactive framework for RNA secondary structure prediction with a dynamical treatment of constraints}, author = {C Gaspin and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7490740}, isbn = {7490740}, year = {1995}, date = {1995-01-01}, journal = {J Mol Biol}, volume = {254}, number = {2}, pages = {163-174}, abstract = {A novel approach aiding in the prediction of RNA secondary structures is presented. Although phylogenetic methods are the most successful at deriving RNA secondary structures, the are not applicable when the number of sequences or the sequence variability is too low. Methods based on energy minimization are therefore of great interest. However, some of the suboptimal RNA secondary structures computed with classic methods are unsaturated structures, i.e. some structures are included into others. Thus, the incorporation of constraints during the process of folding is not possible, while the incorporation of constraints before the process of folding often introduces a bias into the energy function. This paper describes a new procedure which allows for the incorporation of constraints before and during the process of RNA folding. SAPSSARN is an interactive program which offers a framework, both to specify a secondary structure through a set of folding constraints and to compute all the supoptimal saturated RNA secondary structures which satisfy all the folding constraints. At the start, it relies on the computation of the probabilities of pairing of each base with all others according to McCaskill's algorithm. The constraint satisfaction formulation of the problem deals dynamically with a chosen set of folding constraints and, finally, a search algorithm computes all the suboptimal saturated secondary structures which satisfy those folding constraints. Within such a framework, it is possible to test new ideas about RNA folding and secondary structures, including pseudoknots, can be computed. The program is illustrated with RNA sequences on which we obtained results in agreement with known structures by using a protocol which mimics the hierarchical folding of RNA molecules.}, note = {0022-2836 Journal Article}, keywords = {Algorithms Base Sequence Endoribonucleases/*chemistry Escherichia coli/chemistry Introns Models, Bacterial/*chemistry RNA, Catalytic/*chemistry Ribonuclease P *Software Support, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA/*chemistry RNA, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural specificity of nuclease from wheat chloroplasts stroma}, author = {J Gabryszuk and G Keith and M Monko and E Kuligowska and G Dirheimer and J W Szarkowski and A Przykorska}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8643343}, isbn = {8643343}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Symp Ser}, number = {33}, pages = {115-9}, abstract = {A single-strand-specific nuclease from wheat chloroplasts (ChS nuclease) was tested as a tool for RNA secondary and tertiary structure investigations, using yeast tRNA(Phe) and yeast tRNA(Asp) as models. In tRNA(Phe) the nuclease introduced main primary cleavages at positions U33, A35 and A36 in the anticodon-loop and G18 and G19 in the D-loop. In tRNA(Asp) the main primary cleavages occurred at positions U33, G34 and U35 in the anticodon-loop and the lower one at position C20:1 in the D-loop. No primary cleavages were observed within the double-stranded stems. Because ChS nuclease has (i) a low molecular weight, (ii) a wide pH range of action (5.0 to 7.5) (iii) no divalent cation requirement in the reaction mixture and (iv) can be obtained as a pure protein in rather large quantities it appeared to be a very good tool for secondary and tertiary structural studies of RNAs.}, note = {0261-3166 Journal Article}, keywords = {Asp/chemistry/genetics/metabolism RNA, Base Sequence Binding Sites Chloroplasts/*enzymology Endonucleases/isolation & purification/*metabolism Molecular Sequence Data Nucleic Acid Conformation RNA/chemistry/metabolism RNA, Fungal/chemistry/genetics/metabolism RNA, Non-U.S. Gov't Triticum/*enzymology, Phe/chemistry/genetics/metabolism Substrate Specificity Support, Transfer}, pubstate = {published}, tppubtype = {article} } @article{, title = {Native bovine selenocysteine tRNA(Sec) secondary structure as probed by two plant single-strand-specific nucleases}, author = {J Gabryszuk and A Przykorska and M Monko and E Kuligowska and C Sturchler and A Krol and G Dirheimer and J W Szarkowski and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7665090}, isbn = {7665090}, year = {1995}, date = {1995-01-01}, journal = {Gene}, volume = {161}, number = {2}, pages = {259-263}, abstract = {Two single-strand-specific nucleases, discovered in plants, have been used to investigate the secondary and tertiary structures of the native bovine liver selenocysteine tRNA(Sec). To check the possible influence of nucleotide modifications on these structures, we compared the results obtained with the fully modified tRNA to the unmodified transcript prepared by in vitro T7 transcription of the Xenopus laevis tRNA(Sec) gene. We found that the structures in solution of the native tRNA(Sec) and the transcript are very similar despite some differences in accessibility to the enzymatic probes. Indeed, the modified anticodon-loop of native bovine tRNA(Sec), containing 5-methylcarboxymethyluridine (mcm5U34) and N6-isopentenyladenosine (i6A37), is less accessible to Rn nuclease than that of the transcript: the intensity of bands representing cuts at A36 and A38 is much lower as compared to those of the transcript, whereas no cuts were found at the level of i6A37 in the anticodon loop of the native molecule. Surprisingly, the variable arm of the native molecule has been found to be more susceptible to single-strand-specific nuclease action, suggesting a looser structure of the variable arm in native bovine tRNA(Sec) than in the transcript.}, note = {0378-1119 Journal Article}, keywords = {Amino Acid-Specific/*chemistry/*genetics Support, Animals Anticodon/chemistry/genetics Base Sequence Cattle Comparative Study Endonucleases Liver/chemistry Molecular Sequence Data Molecular Structure *Nucleic Acid Conformation Plants/enzymology RNA, Non-U.S. Gov't Xenopus laevis, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural specificity of nuclease from wheat chloroplasts stroma}, author = {J Gabryszuk and G Keith and M Monko and E Kuligowska and G Dirheimer and J W Szarkowski and A Przykorska}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8643343}, isbn = {8643343}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Symp Ser}, number = {33}, pages = {115-119}, abstract = {A single-strand-specific nuclease from wheat chloroplasts (ChS nuclease) was tested as a tool for RNA secondary and tertiary structure investigations, using yeast tRNA(Phe) and yeast tRNA(Asp) as models. In tRNA(Phe) the nuclease introduced main primary cleavages at positions U33, A35 and A36 in the anticodon-loop and G18 and G19 in the D-loop. In tRNA(Asp) the main primary cleavages occurred at positions U33, G34 and U35 in the anticodon-loop and the lower one at position C20:1 in the D-loop. No primary cleavages were observed within the double-stranded stems. Because ChS nuclease has (i) a low molecular weight, (ii) a wide pH range of action (5.0 to 7.5) (iii) no divalent cation requirement in the reaction mixture and (iv) can be obtained as a pure protein in rather large quantities it appeared to be a very good tool for secondary and tertiary structural studies of RNAs.}, note = {0261-3166 Journal Article}, keywords = {Asp/chemistry/genetics/metabolism RNA, Base Sequence Binding Sites Chloroplasts/*enzymology Endonucleases/isolation & purification/*metabolism Molecular Sequence Data Nucleic Acid Conformation RNA/chemistry/metabolism RNA, Fungal/chemistry/genetics/metabolism RNA, Non-U.S. Gov't Triticum/*enzymology, Phe/chemistry/genetics/metabolism Substrate Specificity Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The class II aminoacyl-tRNA synthetases and their active site: evolutionary conservation of an ATP binding site}, author = {G Eriani and J Cavarelli and F Martin and L Ador and B Rees and J C Thierry and J Gangloff and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7783225}, isbn = {7783225}, year = {1995}, date = {1995-01-01}, journal = {J Mol Evol}, volume = {40}, number = {5}, pages = {499-508}, abstract = {Previous sequence analyses have suggested the existence of two distinct classes of aminoacyl-tRNA synthetase. The partition was established on the basis of exclusive sets of sequence motifs (Eriani et al. [1990] Nature 347:203-306). X-ray studies have now well defined the structural basis of the two classes: the class I enzymes share with dehydrogenases and kinases the classic nucleotide binding fold called the Rossmann fold, whereas the class II enzymes possess a different fold, not found elsewhere, built around a six-stranded antiparallel beta-sheet. The two classes of synthetases catalyze the same global reaction that is the attachment of an amino acid to the tRNA, but differ as to where on the terminal adenosine of the tRNA the amino acid is placed: class I enzymes act on the 2' hydroxyl whereas the class II enzymes prefer the 3' hydroxyl group. The three-dimensional structure of aspartyl-tRNA synthetase from yeast, a typical class II enzyme, is described here, in relation to its function. The crucial role of the sequence motifs in substrate binding and enzyme structure is high-lighted. Overall these results underline the existence of an intimate evolutionary link between the aminoacyl-tRNA synthetases, despite their actual structural diversity.}, note = {0022-2844 Journal Article}, keywords = {Amino Acid Species Specificity Structure-Activity Relationship, Asp/metabolism Saccharomyces cerevisiae/enzymology Sequence Alignment Sequence Homology, ERIANI, Molecular Molecular Sequence Data Protein Binding Protein Conformation RNA, Transfer, Transfer/metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Comparative study of DNA adducts levels in white sucker fish (Catostomus commersoni) from the basin of the St. Lawrence River (Canada)}, author = {C el Adlouni and J Tremblay and P Walsh and J Lagueux and J Bureau and D Laliberte and G Keith and D Nadeau and G G Poirier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8594417}, isbn = {8594417}, year = {1995}, date = {1995-01-01}, journal = {Mol Cell Biochem}, volume = {148}, number = {2}, pages = {133-138}, abstract = {The levels of DNA adducts in the hepatic tissue of the white sucker fish species Catostomus commersoni were determined by 32P-postlabelling. The fish were caught at four sites: two sites near the city of Windsor (Quebec, Canada) on the St. Francois River, a downstream tributary of the St. Lawrence River, and two sites in the St. Lawrence River itself, near the city of Montreal (Quebec, Canada). The latter sites are known to be contaminated by many pollutants including polycyclic aromatic hydrocarbons. Total adduct levels in all fish ranged from 25.1-178.0 adducts per 10(9) nucleotides. White sucker from the selected sites of the St. Lawrence River had a significantly higher mean level of DNA adducts than those of the St. Francois River (129.4 vs 56.8, respectively). These results suggest that the effluents of many heavy industries (e.g. from a Soderberg aluminium plant) flowing in the St. Lawrence River are more likely to produce genotoxic damage to fish than those released in one of its tributary, and mainly associated to the activities of a small town and a nearby pulp and paper mill.}, note = {0300-8177 Journal Article}, keywords = {Animals Comparative Study Cypriniformes/genetics/*metabolism DNA Adducts/*analysis *DNA Damage Fresh Water Industrial Waste Liver/chemistry Polycyclic Hydrocarbons/adverse effects Quebec Support, Chemical/*adverse effects, Non-U.S. Gov't Water Pollutants, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A pseudoknot is required for efficient translational initiation and regulation of the Escherichia coli rpsO gene coding for ribosomal protein S15}, author = {C Ehresmann and C Philippe and E Westhof and L Benard and C Portier and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8722030}, isbn = {8722030}, year = {1995}, date = {1995-01-01}, journal = {Biochem Cell Biol}, volume = {73}, number = {11-12}, pages = {1131-1140}, abstract = {Escherichia coli ribosomal protein S15 down regulates its own synthesis by binding to its mRNA in a region overlapping the ribosome binding site, called the translational operator. This binding stabilizes a pseudoknot structure that exists in equilibrium with two stem-loop structures. When synthesized in excess over 16S rRNA, S15 binds to its translational operator and traps the ribosome on its loading site in a transient state, preventing the formation of the active ternary (30S-mRNA-rRNA(f)Met) complex. This inhibition can be suppressed by 16S rRNA, which displaces S15 from the mRNA. An extensive mutational analysis showed that the pseudoknot is the structural element required for S15 recognition and in vivo translational control. Specific sequence determinants are located in limited regions of the structure formed by the pseudoknot. An unexpected result is that the pseudoknot can exist in a variety of topologically equivalent structures recognizable and shapable by S15. Based on footprinting experiments and computer graphic modelling, S15 shields the two stems of the pseudoknot, sitting in the major groove of the coaxial stack.}, note = {0829-8211 Journal Article Review Review, Tutorial}, keywords = {Bacterial Genetic Code Molecular Sequence Data Nucleic Acid Conformation *Peptide Chain Initiation Ribosomal Proteins/*genetics *Translation, Bacterial/*physiology *Genes, Base Sequence Escherichia coli/*genetics Gene Expression Regulation, Genetic, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Primary, secondary and tertiary structures of tRNAs.}, author = {G Dirheimer and G Keith and P Dumas and E Westhof}, editor = {D Soll and U L RajBhandary}, url = {http://books.google.fr/books?hl=fr&lr=&id=jx6mGLkB-E0C&oi=fnd&pg=PA93&dq=Primary,+secondary+and+tertiary+structures+of+tRNAs.&ots=7GJsXp6gAB&sig=F_uhTMBSTQE7VvPN6XpmbNO_V00#v=onepage&q=Primary%2C%20secondary%20and%20tertiary%20structures%20of%20tRNAs.&f=false}, year = {1995}, date = {1995-01-01}, booktitle = {tRNA: Structure, Biosynthesis and Function}, pages = {93-126}, publisher = {American Society for Microbiology, Washington DC}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Variations in tRNA modifications, particularly of their queuine content in higher eukaryotes. Its relation to malignancy grading}, author = {G Dirheimer and W Baranowski and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7599283}, isbn = {7599283}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {99-103}, abstract = {Literature references dealing with the variations in the modification level of nucleosides in total eukaryotic tRNAs as a function of different physiological status and after drug administration as well as in sequenced cytoplasmic tRNAs between normal and tumor cells and in SV40-transformed cells are reviewed. In addition, special attention is given to guanine replacement of queuine in the first position of the anticodon of tRNAs. A correlation between the level of this undermodification in cancer tissues and the malignancy grading could be found in human ovarian tumors, confirming the results reported in several laboratories for lymphomas and lung cancer tissues. Indeed tRNAs from primary and metastatic human ovarian malignant tumors are Q deficient as compared to tRNAs from normal tissues or benign tumors: thus queuine deficiency increases with malignancy and grading of differentiation.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Animals Cell Transformation, Neoplastic/genetics Female Guanine/*analogs & derivatives/analysis Human Neoplasms/*genetics/pathology Ovarian Neoplasms/*genetics/pathology Purines/analysis Pyrimidines/analysis RNA Processing, Non-U.S. Gov't, Post-Transcriptional RNA, Transfer/*chemistry/metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization of threonyl-tRNA synthetase from Thermus thermophilus and preliminary crystallographic data}, author = {V Cura and D Kern and A Mitschler and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7589494}, isbn = {7589494}, year = {1995}, date = {1995-01-01}, journal = {FEBS Lett}, volume = {374}, number = {1}, pages = {110-112}, abstract = {Threonyl-tRNA synthetase from Thermus thermophilus (ttTRS) has been overproduced in Escherichia coli, purified and crystallized in solutions containing ammonium sulfate and glycerol. The crystals grew in the orthorhombic space group C222(1) with unit cell dimensions a = 119.5 A}, note = {0014-5793 Journal Article}, keywords = {Cloning, Molecular Crystallization Crystallography, Non-U.S. Gov't Temperature Thermus thermophilus/*enzymology Threonine-tRNA Ligase/*chemistry/genetics, Unité ARN, X-Ray Escherichia coli Solubility Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Stabilised secondary structure at a ribosomal binding site enhances translational repression in E. coli}, author = {C Brunel and P Romby and C Sacerdot and M de Smit and M Graffe and J Dondon and J van Duin and B Ehresmann and C Ehresmann and M Springer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7563089}, isbn = {7563089}, year = {1995}, date = {1995-01-01}, journal = {J Mol Biol}, volume = {253}, number = {2}, pages = {277-290}, abstract = {The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase is negatively autoregulated at the translational level. The negative feedback is due to the binding of the synthetase to an operator site on its own mRNA located upstream of the initiation codon. The present work describes the characterisation of operator mutants that have the rare property of enhancing repression. These mutations cause (1) a low basal level of expression, (2) a temperature-dependent expression, and (3) an increased capacity of the synthetase to repress its own expression at low temperature. Surprisingly, this enhancement of repression is not explained by an increase of affinity of the mutant operators for the enzyme but by the formation, at low temperature, of a few supplementary base-pairs between the ribosomal binding site and a normally single-stranded domain of the operator. Although this additional base-pairing only slightly inhibits ribosome binding in the absence of repressor, simple thermodynamic considerations indicate that this is sufficient to increase repression. This increase is explained by the competition between the ribosome and repressor for overlapping regions of the mRNA. When the ribosomal binding site is base-paired, the ribosome cannot bind while the repressor can, giving the repressor the advantage in the competition. Thus, the existence of an open versus base-paired equilibrium in a ribosomal binding site of a translational operator amplifies the magnitude of control. This molecular amplification device might be an essential component of translational control considering the low free repressor/ribosome ratio of the low affinity of translational repressors for their target operators.}, note = {0022-2836 Journal Article}, keywords = {Bacterial *Gene Expression Regulation, Base Composition Base Sequence Binding Sites Comparative Study Enzyme Repression Escherichia coli/genetics/*metabolism Gene Expression Regulation, Enzymologic Homeostasis Kinetics Mathematics Models, Genetic *Translation, Genetic beta-Galactosidase/biosynthesis, Messenger/biosynthesis/*chemistry/*metabolism Recombinant Proteins/biosynthesis Ribosomes/*metabolism Support, Non-U.S. Gov't Temperature Threonine-tRNA Ligase/*biosynthesis Transcription, ROMBY, Site-Directed *Nucleic Acid Conformation RNA, Theoretical Molecular Sequence Data Mutagenesis, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural specificity of nuclease from wheat chloroplasts stroma}, author = {J Gabryszuk and G Keith and M Monko and E Kuligowska and G Dirheimer and J W Szarkowski and A Przykorska}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8643343}, isbn = {8643343}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Symp Ser}, number = {33}, pages = {115-9}, abstract = {A single-strand-specific nuclease from wheat chloroplasts (ChS nuclease) was tested as a tool for RNA secondary and tertiary structure investigations, using yeast tRNA(Phe) and yeast tRNA(Asp) as models. In tRNA(Phe) the nuclease introduced main primary cleavages at positions U33, A35 and A36 in the anticodon-loop and G18 and G19 in the D-loop. In tRNA(Asp) the main primary cleavages occurred at positions U33, G34 and U35 in the anticodon-loop and the lower one at position C20:1 in the D-loop. No primary cleavages were observed within the double-stranded stems. Because ChS nuclease has (i) a low molecular weight, (ii) a wide pH range of action (5.0 to 7.5) (iii) no divalent cation requirement in the reaction mixture and (iv) can be obtained as a pure protein in rather large quantities it appeared to be a very good tool for secondary and tertiary structural studies of RNAs.}, note = {0261-3166 Journal Article}, keywords = {Asp/chemistry/genetics/metabolism RNA, Base Sequence Binding Sites Chloroplasts/*enzymology Endonucleases/isolation & purification/*metabolism Molecular Sequence Data Nucleic Acid Conformation RNA/chemistry/metabolism RNA, Fungal/chemistry/genetics/metabolism RNA, Non-U.S. Gov't Triticum/*enzymology, Phe/chemistry/genetics/metabolism Substrate Specificity Support, Transfer}, pubstate = {published}, tppubtype = {article} } @article{, title = {Multiple Molecular Dynamics Simulations of the Anticodon Loop of tRNAAsp in Aqueous Solution with Counterions}, author = {P Auffinger and S Louise-May and E Westhof}, url = {http://pubs.acs.org/doi/abs/10.1021/ja00130a011}, doi = {10.1021/ja00130a011}, year = {1995}, date = {1995-01-01}, journal = {J Am Chem Soc}, volume = {117}, number = {25}, pages = {6720-6726}, abstract = {In a systematic search for a stable protocol with which to extend our dynamical investigations, a nanosecond of molecular dynamics simulations of the solvated anticodon loop of tRNAASp consisting of ten unique trajectories was obtained by slight modifications to the starting conditions. These changes produced divergent trajectories which varied widely in structural and dynamical characteristics. However, the properties of these trajectories could not be directly correlated to the slight modifications introduced in the system, and thus, questions were raised regarding the probity of the standard protocol we utilized. Instead of a detailed analysis of the results, the multiple molecular dynamics (MD) approach was used as a diagnostic for estimating the reliability of the set of trajectories generated and the extent of relevant biochemical information which can be extracted from it. We address here issues concerning critical evaluation of molecular dynamics methodology and detection of protocol instabilities. We infer that an ensemble of initial uncorrelated trajectories should be generated in order to investigate the constancy of structural and dynamical properties of the system under study.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A simple test for evaluating the truncation effects in simulations of systems involving charged groups}, author = {P Auffinger and D L Beveridge}, url = {http://www.sciencedirect.com/science/article/pii/000926149500065C}, doi = {10.1016/0009-2614(95)00065-C}, year = {1995}, date = {1995-01-01}, journal = {Chem Phys Lett}, volume = {234}, number = {4-6}, pages = {413-415}, abstract = {We report the results from a molecular dynamics (MD) simulation on a 1.0 M aqueous NaCl solution using the classical SPC water model and a treatment of long-range electrostatic forces involving a 9–16 Å twin-range cut-off. This study was undertaken to explore the importance of truncation effects in simulations involving charged species which employ cut-offs at longer range than typically used in MD. Artefacts were discerned in the ion-ion radial distribution functions at ≈ 16 Å, indicating that increasing the length scale of the truncation scheme does not necessarily lead to reliable trajectories. Our results indicate that ムcharge group-charge groupメ radial distribution functions are a sensitive test for the detection of artefacts due to truncation and it is suggested that they should be used as a routine screening device in the analysis of MD systems involving charged groups.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Variations in tRNA modifications, particularly of their queuine content in higher eukaryotes. Its relation to malignancy grading}, author = { G. Dirheimer and W. Baranowski and G. Keith}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {99-103}, abstract = {Literature references dealing with the variations in the modification level of nucleosides in total eukaryotic tRNAs as a function of different physiological status and after drug administration as well as in sequenced cytoplasmic tRNAs between normal and tumor cells and in SV40-transformed cells are reviewed. In addition, special attention is given to guanine replacement of queuine in the first position of the anticodon of tRNAs. A correlation between the level of this undermodification in cancer tissues and the malignancy grading could be found in human ovarian tumors, confirming the results reported in several laboratories for lymphomas and lung cancer tissues. Indeed tRNAs from primary and metastatic human ovarian malignant tumors are Q deficient as compared to tRNAs from normal tissues or benign tumors: thus queuine deficiency increases with malignancy and grading of differentiation.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {&, Animals, Cell, derivatives/analysis, Female, Gov't, Guanine/*analogs, Human, Neoplasms/*genetics/pathology, Neoplastic/genetics, Non-U.S., Ovarian, post-transcriptional, Processing, Purines/analysis, Pyrimidines/analysis, RNA, Support, Transfer/*chemistry/metabolism, Transformation}, pubstate = {published}, tppubtype = {article} } @article{, title = {Detection of DNA adducts in declining hop plants grown on fields formerly treated with heptachlor, a persistent insecticide}, author = {A Laouedj and C Schenk and A Pfohl-Leszkowicz and G Keith and D Schontz and P Guillemaut and B Rether}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15091476}, isbn = {15091476}, year = {1995}, date = {1995-01-01}, journal = {Environ Pollut}, volume = {90}, number = {3}, pages = {409-14}, abstract = {Hop decline was observed in Alsace, eastern France, in reparcelled sugar beet fields formerly abundantly treated with an insecticide, heptachlor. Leaves were collected from 'declining hops' grown in an heptachlor-contaminated area and from 'healthy hops' grown in a soil not contaminated by heptachlor. These two samples came from hop vines treated with other usual pesticides. 'Control' hop leaves came from soil neither treated with pesticide nor contaminated with heptachlor. Hypermodified nucleotides (DNA adducts) were detected using the (32)P-postlabelling method. No detectable DNA adducts were found in the 'control' specimen, whereas eight adducts were detected in the 'healthy hops' specimen, probably due to the usual pesticide treatment. However, 16 adducts, nine of which were new adducts, could be detected in the 'declining hops' specimen. It may therefore be supposed that the presence of these hypermodified nucleotides perturbs gene expression and so contributes to the hop decline. In addition, to confirm the genotoxicity of heptachlor, it is shown that it induces DNA adducts in bean-cell suspension culture as well. Finally, it is proposed, in the case of alternate cultures scheduled in fields which were formerly treated with pesticides, adapted to other cultures, that particular attention should be given to the history of the soils.}, note = {0269-7491 Journal Article}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Plus-strand DNA synthesis of the yeast retrotransposon Ty1 is initiated at two sites, PPT1 next to the 3' LTR and PPT2 within the pol gene. PPT1 is sufficient for Ty1 transposition}, author = { T. Heyman and B. Agoutin and S. Friant and F. X. Wilhelm and M. L. Wilhelm}, year = {1995}, date = {1995-01-01}, journal = {J Mol Biol}, volume = {253}, number = {2}, pages = {291-303}, abstract = {Long terminal repeat elements and retroviruses require primers for initiation of minus and plus-strand DNA synthesis by reverse transcriptase. Here we demonstrate genetically that plus-strand DNA synthesis of the yeast Ty1 element is initiated at two sites located at the 5' boundary of the 3' long terminal repeat (PPT1) and near the middle of the pol gene in the integrase coding sequence (PPT2). A consequence of the presence of two PPTs is that Ty1 plus-strand DNA exists as segments at some time during replication. Three fragments have been identified: the plus-strand strong-stop DNA initiated at PPT1, a downstream fragment initiated at PPT2 and an upstream fragment spanning the 5'-terminal part of Ty1 and a portion of the TyB gene. Characterization of the 3' ends of the plus-strand DNA fragments reveals (1) that the upstream fragment is elongated beyond PPT2 creating a plus-strand overlap and (2) that the majority of plus-strand strong-stop DNA fragments bear a copy of the minus-strand primer binding site in agreement with the accepted model of retroviral genomic RNA reverse transcription. The two polypurine tracts, PPT1 and PPT2, have an identical sequence GGGTGGTA. Mutations replacing purines by pyrimidines in this sequence significantly diminish or abolish initiation of plus-strand synthesis. Ty1 elements bearing a mutated PPT2 sequence are not defective for transposition whereas mutations in PPT1 abolish transposition.}, note = {0022-2836 Journal Article}, keywords = {*DNA, *Genes, *Repetitive, *Retroelements, Acid, Base, C/analysis, cerevisiae/genetics/*virology, Chain, Cloning, Data, DNA, Fungal, Fungal/biosynthesis, Genes, Genetic, Genome, Gov't, Mapping, Molecular, Non-U.S., Nucleic, pol, Poly, Polymerase, Primers, Reaction, Replication, Restriction, Saccharomyces, Sequence, Sequences, Support, Transcription, Viral, Viral/*biosynthesis}, pubstate = {published}, tppubtype = {article} } @article{, title = {Variations in tRNA modifications, particularly of their queuine content in higher eukaryotes. Its relation to malignancy grading}, author = {G Dirheimer and W Baranowski and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7599283}, isbn = {7599283}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {99-103}, abstract = {Literature references dealing with the variations in the modification level of nucleosides in total eukaryotic tRNAs as a function of different physiological status and after drug administration as well as in sequenced cytoplasmic tRNAs between normal and tumor cells and in SV40-transformed cells are reviewed. In addition, special attention is given to guanine replacement of queuine in the first position of the anticodon of tRNAs. A correlation between the level of this undermodification in cancer tissues and the malignancy grading could be found in human ovarian tumors, confirming the results reported in several laboratories for lymphomas and lung cancer tissues. Indeed tRNAs from primary and metastatic human ovarian malignant tumors are Q deficient as compared to tRNAs from normal tissues or benign tumors: thus queuine deficiency increases with malignancy and grading of differentiation.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Animals Cell Transformation, Neoplastic/genetics Female Guanine/*analogs & derivatives/analysis Human Neoplasms/*genetics/pathology Ovarian Neoplasms/*genetics/pathology Purines/analysis Pyrimidines/analysis RNA Processing, Non-U.S. Gov't, Post-Transcriptional RNA, Transfer/*chemistry/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A simple test for evaluating the truncation effects in simulations of systems involving charged groups}, author = {P Auffinger and D L Beveridge}, url = {http://www.sciencedirect.com/science/article/pii/000926149500065C}, doi = {10.1016/0009-2614(95)00065-C}, year = {1995}, date = {1995-01-01}, journal = {Chem Phys Lett}, volume = {234}, number = {4-6}, pages = {413-415}, abstract = {We report the results from a molecular dynamics (MD) simulation on a 1.0 M aqueous NaCl solution using the classical SPC water model and a treatment of long-range electrostatic forces involving a 9–16 Å twin-range cut-off. This study was undertaken to explore the importance of truncation effects in simulations involving charged species which employ cut-offs at longer range than typically used in MD. Artefacts were discerned in the ion-ion radial distribution functions at ≈ 16 Å, indicating that increasing the length scale of the truncation scheme does not necessarily lead to reliable trajectories. Our results indicate that ムcharge group-charge groupメ radial distribution functions are a sensitive test for the detection of artefacts due to truncation and it is suggested that they should be used as a routine screening device in the analysis of MD systems involving charged groups.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Variations in tRNA modifications, particularly of their queuine content in higher eukaryotes. Its relation to malignancy grading}, author = {G Dirheimer and W Baranowski and G Keith}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7599283}, isbn = {7599283}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {99-103}, abstract = {Literature references dealing with the variations in the modification level of nucleosides in total eukaryotic tRNAs as a function of different physiological status and after drug administration as well as in sequenced cytoplasmic tRNAs between normal and tumor cells and in SV40-transformed cells are reviewed. In addition, special attention is given to guanine replacement of queuine in the first position of the anticodon of tRNAs. A correlation between the level of this undermodification in cancer tissues and the malignancy grading could be found in human ovarian tumors, confirming the results reported in several laboratories for lymphomas and lung cancer tissues. Indeed tRNAs from primary and metastatic human ovarian malignant tumors are Q deficient as compared to tRNAs from normal tissues or benign tumors: thus queuine deficiency increases with malignancy and grading of differentiation.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Animals Cell Transformation, Neoplastic/genetics Female Guanine/*analogs & derivatives/analysis Human Neoplasms/*genetics/pathology Ovarian Neoplasms/*genetics/pathology Purines/analysis Pyrimidines/analysis RNA Processing, Non-U.S. Gov't, Post-Transcriptional RNA, Transfer/*chemistry/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A simple test for evaluating the truncation effects in simulations of systems involving charged groups}, author = {P Auffinger and D L Beveridge}, editor = {Editor}, url = {http://www.sciencedirect.com/science/article/pii/000926149500065C}, doi = {10.1016/0009-2614(95)00065-Ci}, year = {1995}, date = {1995-01-01}, journal = {Chem Phys Lett}, volume = {234}, number = {4-6}, pages = {413-415}, abstract = {We report the results from a molecular dynamics (MD) simulation on a 1.0 M aqueous NaCl solution using the classical SPC water model and a treatment of long-range electrostatic forces involving a 9–16 Å twin-range cut-off. This study was undertaken to explore the importance of truncation effects in simulations involving charged species which employ cut-offs at longer range than typically used in MD. Artefacts were discerned in the ion-ion radial distribution functions at ≈ 16 Å, indicating that increasing the length scale of the truncation scheme does not necessarily lead to reliable trajectories. Our results indicate that ムcharge group-charge groupメ radial distribution functions are a sensitive test for the detection of artefacts due to truncation and it is suggested that they should be used as a routine screening device in the analysis of MD systems involving charged groups.}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mobilities of modified ribonucleotides on two-dimensional cellulose thin-layer chromatography}, author = { G. Keith}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {142-4}, note = {0300-9084 Journal Article}, keywords = {*Chromatography, Layer, Purines/chemistry, Pyrimidines/chemistry, Ribonucleases/metabolism, Ribonucleotides/*chemistry, RNA, Thin, Transfer/chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Postlabeling: a sensitive method for studying DNA adducts and their role in carcinogenesis}, author = { G. Keith and G. Dirheimer}, year = {1995}, date = {1995-01-01}, journal = {Curr Opin Biotechnol}, volume = {6}, number = {1}, pages = {3-11}, abstract = {The covalent binding of xenobiotics to DNA is an important trigger of the multistage process that leads to carcinogenesis. 32P-postlabeling represents a highly sensitive method for biomonitoring exposure to genotoxic agents and for cancer risk assessment; it is capable of detecting less than one DNA adduct per human genome. Recent improvements to the technique have shown that the resistance of adducted DNA to enzyme digestion may lead to an overestimation of the number of different adducts present in a sample.}, note = {0958-1669 Journal Article Review Review, Academic}, keywords = {*Cell, *Genome, Adducts/*analysis, and, Animals, Base, Cell, Conditions/genetics/pathology, Data, Dilution, Division, DNA, Genetic, Gov't, Human, Models, Molecular, Mutagenesis, Neoplasms/*genetics/pathology, Neoplastic, Non-U.S., Phosphorus, Precancerous, Radioisotope, Radioisotopes, Sensitivity, Sequence, Specificity, Support, Technique, Transformation, Xenobiotics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural specificity of nuclease from wheat chloroplasts stroma}, author = { J. Gabryszuk and G. Keith and M. Monko and E. Kuligowska and G. Dirheimer and J. W. Szarkowski and A. Przykorska}, year = {1995}, date = {1995-01-01}, journal = {Nucleic Acids Symp Ser}, number = {33}, pages = {115-9}, abstract = {A single-strand-specific nuclease from wheat chloroplasts (ChS nuclease) was tested as a tool for RNA secondary and tertiary structure investigations, using yeast tRNA(Phe) and yeast tRNA(Asp) as models. In tRNA(Phe) the nuclease introduced main primary cleavages at positions U33, A35 and A36 in the anticodon-loop and G18 and G19 in the D-loop. In tRNA(Asp) the main primary cleavages occurred at positions U33, G34 and U35 in the anticodon-loop and the lower one at position C20:1 in the D-loop. No primary cleavages were observed within the double-stranded stems. Because ChS nuclease has (i) a low molecular weight, (ii) a wide pH range of action (5.0 to 7.5) (iii) no divalent cation requirement in the reaction mixture and (iv) can be obtained as a pure protein in rather large quantities it appeared to be a very good tool for secondary and tertiary structural studies of RNAs.}, note = {0261-3166 Journal Article}, keywords = {&, Acid, Asp/chemistry/genetics/metabolism, Base, Binding, Chloroplasts/*enzymology, Conformation, Data, Endonucleases/isolation, Fungal/chemistry/genetics/metabolism, Gov't, Molecular, Non-U.S., Nucleic, Phe/chemistry/genetics/metabolism, purification/*metabolism, RNA, RNA/chemistry/metabolism, Sequence, Sites, Specificity, Substrate, Support, Transfer, Triticum/*enzymology}, pubstate = {published}, tppubtype = {article} } @article{, title = {Genetic probes of ribosomal RNA function}, author = { M. O'Connor and C. A. Brunelli and M. A. Firpo and S. T. Gregory and K. R. Lieberman and J. S. Lodmell and H. Moine and D. I. Van Ryk and A. E. Dahlberg}, year = {1995}, date = {1995-01-01}, journal = {Biochem Cell Biol}, volume = {73}, number = {11-12}, pages = {859-68}, abstract = {We have used a genetic approach to uncover the functional roles of rRNA in protein synthesis. Mutations were constructed in a cloned rrn operon by site-directed mutagenesis or isolated by genetic selections following random mutagenesis. We have identified mutations that affect each step in the process of translation. The data are consistent with the results of biochemical and phylogenetic analyses but, in addition, have provided novel information on regions of rRNA not previously investigated.}, note = {0829-8211 Journal Article Review Review, Tutorial}, keywords = {16S/genetics, Acid, Base, Conformation, Data, Gov't, Messenger/genetics, Molecular, Mutation, Non-U.S., Nucleic, P.H.S., Probes, Ribosomal, Ribosomal/*genetics, RNA, Sequence, Support, Transfer/genetics, U.S.}, pubstate = {published}, tppubtype = {article} } @article{, title = {Detection of DNA adducts in declining hop plants grown on fields formerly treated with heptachlor, a persistent insecticide}, author = { A. Laouedj and C. Schenk and A. Pfohl-Leszkowicz and G. Keith and D. Schontz and P. Guillemaut and B. Rether}, year = {1995}, date = {1995-01-01}, journal = {Environ Pollut}, volume = {90}, number = {3}, pages = {409-14}, abstract = {Hop decline was observed in Alsace, eastern France, in reparcelled sugar beet fields formerly abundantly treated with an insecticide, heptachlor. Leaves were collected from 'declining hops' grown in an heptachlor-contaminated area and from 'healthy hops' grown in a soil not contaminated by heptachlor. These two samples came from hop vines treated with other usual pesticides. 'Control' hop leaves came from soil neither treated with pesticide nor contaminated with heptachlor. Hypermodified nucleotides (DNA adducts) were detected using the (32)P-postlabelling method. No detectable DNA adducts were found in the 'control' specimen, whereas eight adducts were detected in the 'healthy hops' specimen, probably due to the usual pesticide treatment. However, 16 adducts, nine of which were new adducts, could be detected in the 'declining hops' specimen. It may therefore be supposed that the presence of these hypermodified nucleotides perturbs gene expression and so contributes to the hop decline. In addition, to confirm the genotoxicity of heptachlor, it is shown that it induces DNA adducts in bean-cell suspension culture as well. Finally, it is proposed, in the case of alternate cultures scheduled in fields which were formerly treated with pesticides, adapted to other cultures, that particular attention should be given to the history of the soils.}, note = {0269-7491 Journal Article}, keywords = {}, pubstate = {published}, tppubtype = {article} } @article{, title = {A novel approach to introduce site-directed specific cross-links within RNA-protein complexes. Application to the Escherichia coli threonyl-tRNA synthetase/translational operator complex}, author = {M Zenkova and C Ehresmann and J Caillet and M Springer and G Karpova and B Ehresmann and P Romby}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7544283}, isbn = {7544283}, year = {1995}, date = {1995-01-01}, journal = {Eur J Biochem}, volume = {231}, number = {3}, pages = {726-735}, abstract = {We describe a methodology which allows the introduction of a photoactivatable azido group at specific internal positions of any RNA in order to identify the neighboring elements of an interacting protein. The first step involves site-directed modification of the target RNA with an antisense oligodeoxyribonucleotide bearing, at its 3' or 5' phosphate, a 4-[-N-(2-chloroethyl)-N-methylamino]benzylmethylamino group. Position N7 of a guanine residue located in the close vicinity of the hybrid is the main target for alkylation. The antisense oligodeoxyribonucleotide is then removed by acidic pH treatment and a photoreactive reagent (2,4-dinitro-5-fluorophenylazide) is condensed to the modified nucleotide. This method was used to induce specific cross-links between Escherichia coli threonyl-tRNA synthetase and the leader region of threonyl-tRNA synthetase mRNA, which is involved in translational feedback regulation. Control experiments revealed that the modification affects neither the structure of the mRNA nor the interaction with the enzyme. More than 50% of the modified mRNA complexed with threonyl-tRNA synthetase can be cross-linked to the enzyme, depending on the nucleotide modified.}, note = {0014-2956 Journal Article}, keywords = {Alkylation Base Sequence Cross-Linking Reagents Escherichia coli/enzymology/*genetics Molecular Sequence Data Nucleic Acid Conformation *Operator Regions (Genetics) RNA, Bacterial/*chemistry/genetics RNA-Binding Proteins/*chemistry/genetics Support, Genetic, Non-U.S. Gov't Threonine-tRNA Ligase/*genetics *Translation, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {The Use of Molecular Dynamics Simulations for Modelling Nucleic Acids}, author = {E Westhof and C Rubin-Carrez and V Fritsch}, editor = {J M Goodfellow}, url = {http://onlinelibrary.wiley.com/doi/10.1002/9783527615339.ch5/summary}, doi = {10.1002/9783527615339.ch5}, year = {1995}, date = {1995-01-01}, booktitle = {Computer Modelling in Molecular Biology}, pages = {103-131}, publisher = {Wiley-VCH}, keywords = {* molecular dynamics simulations * nucleic acids * potential energy function * treatment of solvent * counterions, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Nucleic acids: Diversity, folding, and stability of nucleic acid structures.}, author = {E Westhof and D Patel}, url = {http://www.sciencedirect.com/science/article/pii/0959440X95800883}, doi = {10.1016/0959-440X(95)80088-3}, year = {1995}, date = {1995-01-01}, journal = {Curr Opin Struct Biol}, volume = {5}, number = {3}, pages = {279–281}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Prediction and experimental investigation of RNA secondary and tertiary foldings.}, author = {E Westhof and F Michel}, editor = {K Nagai and I W Mattaj}, url = {http://ukcatalogue.oup.com/product/9780199635047.do}, year = {1995}, date = {1995-01-01}, booktitle = {RNA-Protein Interactions: Frontiers in Molecular Biology}, pages = {25-51}, publisher = {IRL Press at Oxford University Press}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @inbook{, title = {On the role of single-stranded adenines in RNA-RNA recognition.}, author = {E Westhof}, editor = {A Pullman and J Jortner and B Pullman}, url = {http://www.springer.com/life+sciences/biochemistry+%26+biophysics/book/978-0-7923-3102-5}, year = {1995}, date = {1995-01-01}, booktitle = {Modelling of Biomolecular Structures and Mechanisms (Jerusalem Symposia)}, pages = {305-313}, publisher = {Kluwer Academic Publishers}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{lemaitre_functional_1995, title = {Functional analysis and regulation of nuclear import of dorsal during the immune response in Drosophila}, author = {Bruno Lemaitre and Marie Meister and S Govind and Philippe Georgel and R Steward and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0261-4189}, year = {1995}, date = {1995-01-01}, journal = {EMBO J.}, volume = {14}, number = {3}, pages = {536--545}, abstract = {In addition to its function in embryonic development, the NF-kappa B/rel-related gene dorsal (dl) of Drosophila is expressed in larval and adult fat body where its RNA expression is enhanced upon injury. Injury also leads to a rapid nuclear translocation of dl from the cytoplasm in fat body cells. Here we present data which strongly suggest that the nuclear localization of dl during the immune response is controlled by the Toll signaling pathway, comprising gene products that participate in the intracellular part of the embryonic dorsoventral pathway. We also report that in mutants such as Toll or cactus, which exhibit melanotic tumor phenotypes, dl is constitutively nuclear. Together, these results point to a potential link between the Toll signaling pathway and melanotic tumor induction. Although dl has been shown previously to bind to kappa B-related motifs within the promoter of the antibacterial peptide coding gene diptericin, we find that injury-induced expression of diptericin can occur in the absence of dl. Furthermore, the melanotic tumor phenotype of Toll and cactus is not dl dependent. These data underline the complexity of the Drosophila immune response. Finally, we observed that like other rel proteins, dl can control the level of its own transcription.}, keywords = {Animals, Anti-Bacterial Agents, Anti-Infective Agents, Antimicrobial Cationic Peptides, Biological Transport, Cell Nucleus, Cell Surface, DNA-Binding Proteins, Fat Body, Gene Expression Regulation, Genetic, hoffmann, Immunity, Immunohistochemistry, Insect Hormones, Insect Proteins, M3i, Melanins, Membrane Glycoproteins, Mutation, Neoplasms, Nuclear Proteins, Phosphoproteins, Receptors, reichhart, Signal Transduction, Toll-Like Receptors, Transcription, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mobilities of modified ribonucleotides on two-dimensional cellulose thin-layer chromatography}, author = {G Keith}, editor = {Editor}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7599271}, isbn = {7599271}, year = {1995}, date = {1995-01-01}, journal = {Biochimie}, volume = {77}, number = {1-2}, pages = {142-4}, note = {0300-9084 Journal Article}, keywords = {*Chromatography, Thin Layer Purines/chemistry Pyrimidines/chemistry RNA, Transfer/chemistry Ribonucleases/metabolism Ribonucleotides/*chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {The role of Lys378 and Lys381 on the enzyme activity of E. coli arginyl-tRNA synthetase.}, author = {E D Wang and Y W Huang and Y L Wang and G Eriani and J Gangloff}, url = {none}, year = {1995}, date = {1995-01-01}, journal = {Acta Bioch Bioph Sin}, volume = {27}, pages = {37-43}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Lys306 of E. coli arginyl-tRNA synthetase is necessary for the activity of this enzyme.}, author = {E D Wang and W L Gu and Y L Wang and G Eriani and J Gangloff}, url = {none}, year = {1995}, date = {1995-01-01}, journal = {Acta Bioch Bioph Sin}, volume = {27}, pages = {123-128}, keywords = {ERIANI, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{meister_insect_1994, title = {Insect immunity. A transgenic analysis in Drosophila defines several functional domains in the diptericin promoter}, author = {Marie Meister and A Braun and Christine Kappler and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0261-4189}, year = {1994}, date = {1994-12-01}, journal = {EMBO J.}, volume = {13}, number = {24}, pages = {5958--5966}, abstract = {Diptericins are antibacterial polypeptides which are strongly induced in the fat body and blood cells of dipteran insects in response to septic injury. The promoter of the single-copy, intronless diptericin gene of Drosophila contains several nucleotide sequences homologous to mammalian cis-regulatory motifs involved in the control of acute phase response genes. Extending our previous studies on the expression of the diptericin gene, we now report a quantitative analysis of the contribution of various putative regulatory elements to the bacterial inducibility of this gene, based on the generation of 60 transgenic fly lines carrying different elements fused to a reporter gene. Our data definitively identify two Kappa B-related motifs in the proximal promoter as the sites conferring inducibility and tissue-specific expression to the diptericin gene. These motifs alone, however, mediate only minimal levels of expression. Additional proximal regulatory elements are necessary to attain some 20% of the full response and we suspect a role for sequences homologous to mammalian IL6 response elements and interferon-gamma responsive sites in this up-regulation. The transgenic experiments also reveal the existence of a distal regulatory element located upstream of -0.6 kb which increases the level of expression by a factor of five.}, keywords = {Animals, Anti-Infective Agents, Base Sequence, beta-Galactosidase, DNA Mutational Analysis, Female, Gene Expression Regulation, Genetic, Genetically Modified, Germ Cells, hoffmann, Insect Hormones, Insect Proteins, M3i, Male, Models, Nucleic Acid, Promoter Regions, Recombinant Fusion Proteins, reichhart, Repetitive Sequences, Transformation}, pubstate = {published}, tppubtype = {article} } @article{fehlbaum_insect_1994, title = {Insect immunity. Septic injury of Drosophila induces the synthesis of a potent antifungal peptide with sequence homology to plant antifungal peptides}, author = {P Fehlbaum and Philippe Bulet and L Michaut and Marie Lagueux and W F Broekaert and Charles Hetru and Jules A Hoffmann}, issn = {0021-9258}, year = {1994}, date = {1994-12-01}, journal = {J. Biol. Chem.}, volume = {269}, number = {52}, pages = {33159--33163}, abstract = {In response to a septic injury (pricking with a bacteria-soaked needle) larvae and adults of Drosophila produce considerable amounts of a 44-residue peptide containing 8 cysteines engaged in intramolecular disulfide bridges. The peptide is synthesized in the fat body, a functional homologue of the mammalian liver, and secreted into the blood of the insect. It exhibits potent antifungal activity but is inactive against bacteria. This novel inducible peptide, which we propose to name drosomycin, shows a significant homology with a family of 5-kDa cysteine-rich plant antifungal peptides recently isolated from seeds of Brassicaceae. This finding underlines that plants and insects can rely on similar molecules in their innate host defense.}, keywords = {Amino Acid, Animals, Antifungal Agents, Base Sequence, Cloning, Complementary, DNA, hoffmann, Insect Proteins, M3i, Male, messenger, Microbial Sensitivity Tests, Molecular, Peptide Biosynthesis, Peptides, Plants, Protein Biosynthesis, Protein Precursors, Proteins, RNA, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_staufen_1994b, title = {Staufen protein associates with the 3'UTR of bicoid mRNA to form particles that move in a microtubule-dependent manner}, author = {Dominique Ferrandon and L Elphick and C Nüsslein-Volhard and St D Johnston}, issn = {0092-8674}, year = {1994}, date = {1994-12-01}, journal = {Cell}, volume = {79}, number = {7}, pages = {1221--1232}, abstract = {Staufen protein is required in order to anchor bicoid (bcd) mRNA at the anterior pole of the Drosophila egg. Here we show that staufen protein colocalizes with bcd mRNA at the anterior, and that this localization depends upon its association with the mRNA. Upon injection into the embryo, bcd transcripts specifically interact with staufen, and we have mapped the sequences required to three regions of the 3'UTR, each of which is predicted to form a long stem-loop. The resulting staufen-bcd 3'UTR complexes form particles that show a microtubule-dependent localization. Since staufen is also transported with oskar (osk) mRNA during oogenesis, staufen associates specifically with both osk and bcd mRNAs to mediate their localizations, but at two distinct stages of development.}, keywords = {Animals, Base Sequence, Cell Polarity, ferrandon, Homeodomain Proteins, Insect Hormones, M3i, messenger, Microtubules, Nucleic Acid Conformation, Oocytes, RNA, RNA-Binding Proteins, Trans-Activators}, pubstate = {published}, tppubtype = {article} } @article{auble_mot1_1994, title = {Mot1, a global repressor of RNA polymerase II transcription, inhibits TBP binding to DNA by an ATP-dependent mechanism}, author = {D T Auble and K E Hansen and C G Mueller and W S Lane and J Thorner and S Hahn}, doi = {10.1101/gad.8.16.1920}, issn = {0890-9369}, year = {1994}, date = {1994-08-01}, journal = {Genes & Development}, volume = {8}, number = {16}, pages = {1920--1934}, abstract = {Basal transcription of many genes in yeast is repressed by Mot1, an essential protein which is a member of the Snf2/Swi2 family of conserved nuclear factors. ADI is an ATP-dependent inhibitor of TATA-binding protein (TBP) binding to DNA that inhibits transcription in vitro. Here we demonstrate that ADI is encoded by the MOT1 gene. Mutation of MOT1 abolishes ADI activity and derepresses basal transcription in vitro and in vivo. Recombinant Mot1 removes TBP from DNA and Mot1 contains an ATPase activity which is essential for its function. Genetic interactions between Mot1 and TBP indicate that their functions are interlinked in vivo. These results provide a general model for understanding the mechanism of action of a large family of nuclear factors involved in processes such as transcription and DNA repair.}, keywords = {Adenosine Triphosphatases, Adenosine Triphosphate, Amino Acid Sequence, Base Sequence, Biological, DNA, DNA Helicases, DNA Probes, DNA-Binding Proteins, Fungal, Fungal Proteins, Genes, Genetic, Models, Molecular Sequence Data, Mutagenesis, Repressor Proteins, RNA Polymerase II, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Site-Directed, TATA Box, TATA-Binding Protein Associated Factors, TATA-Box Binding Protein, Team-Mueller, Transcription, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{cociancich_novel_1994, title = {Novel inducible antibacterial peptides from a hemipteran insect, the sap-sucking bug Pyrrhocoris apterus}, author = {S Cociancich and A Dupont and G Hegy and R Lanot and F Holder and Charles Hetru and Jules A Hoffmann and Philippe Bulet}, issn = {0264-6021}, year = {1994}, date = {1994-06-01}, journal = {Biochem. J.}, volume = {300 ( Pt 2)}, pages = {567--575}, abstract = {Insects belonging to the recent orders of the endopterygote clade (Lepidoptera, Diptera, Hymenoptera and Coleoptera) respond to bacterial challenge by the rapid and transient synthesis of a battery of potent antibacterial peptides which are secreted into their haemolymph. Here we present the first report on inducible antibacterial molecules in the sap-sucking bug Pyrrhocoris apterus, a representative species of the Hemiptera, which predated the Endoptergotes by at least 50 million years in evolution. We have isolated and characterized from immune blood of this species three novel peptides or polypeptides: (i) a 43-residue cysteine-rich anti-(Gram-positive bacteria) peptide which is a new member of the family of insect defensins; (ii) a 20-residue proline-rich peptide carrying an O-glycosylated substitution (N-acetylgalactosamine), active against Gram-negative bacteria; (iii) a 133-residue glycine-rich polypeptide also active against Gram-negative bacteria. The proline-rich peptide shows high sequence similarities with drosocin, an O-glycosylated antibacterial peptide from Drosophila, and also with the N-terminal domain of diptericin, an inducible 9 kDa antibacterial peptide from members of the order Diptera, whereas the glycine-rich peptide has similarities with the glycine-rich domain of diptericin. We discuss the evolutionary aspects of these findings.}, keywords = {Amino Acid, Animals, Anti-Bacterial Agents, Antimicrobial Cationic Peptides, Blood Proteins, Chromatography, Defensins, Gas Chromatography-Mass Spectrometry, Gel, Gram-Negative Bacteria, Gram-Positive Bacteria, Hemiptera, Hemolymph, hoffmann, Insect Proteins, M3i, Peptides, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{dimarcq_characterization_1994, title = {Characterization and transcriptional profiles of a Drosophila gene encoding an insect defensin. A study in insect immunity}, author = {Jean-Luc Dimarcq and Danièle Hoffmann and Marie Meister and Philippe Bulet and R Lanot and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0014-2956}, year = {1994}, date = {1994-04-01}, journal = {Eur. J. Biochem.}, volume = {221}, number = {1}, pages = {201--209}, abstract = {Insect defensins are a family of 4-kDa, cationic, inducible antibacterial peptides which bear six cysteine residues engaged in three intramolecular disulfide bridges. They owe their name to certain sequence similarities with defensins from mammalian neutrophiles and macrophages. We report the characterization of a novel defensin isoform from Drosophila and the cloning of the gene encoding a preprodefensin. The gene, which is intronless and present in a single copy/haploid genome, maps at position 46CD on the right arm of the second chromosome. The analysis of the upstream region of the gene reveals the presence of multiple putative cis-regulatory sequences similar to mammalian regulatory motifs of acute-phase-response genes. Transcriptional profiles indicate that the Drosophila defensin gene is induced by bacterial challenge with acute-phase kinetics. It is also expressed in the absence of immune challenge during metamorphosis. These and other data on the Drosophila defensin gene lead us to suggest that insect and mammalian defensins have evolved independently.}, keywords = {Animals, Base Sequence, Blood Proteins, Chromosome Mapping, Cloning, Complementary, Defensins, DNA, Gene Expression, Genetic, Gram-Positive Bacteria, hoffmann, Larva, M3i, Molecular, Molecular Structure, Nucleic Acid, Protein Precursors, Regulatory Sequences, reichhart, Transcription}, pubstate = {published}, tppubtype = {article} } @article{cociancich_inducible_1994, title = {The inducible antibacterial peptides of insects}, author = {S Cociancich and Philippe Bulet and Charles Hetru and Jules A Hoffmann}, issn = {0169-4758}, year = {1994}, date = {1994-04-01}, journal = {Parasitol. Today (Regul. Ed.)}, volume = {10}, number = {4}, pages = {132--139}, abstract = {Insects respond to bacterial challenge by the rapid and transient synthesis of a large number of potent antibacterial peptides that are active against many different bacteria. Two families of inducible antibacterial peptides are well characterized: the cecropins and the insect defensins. A rapidly increasing number of proline- and glycine-rich peptides are reported from various insect species together with cecropins and insect defensins. In this review, Stéphane Cociancich, Philippe Bulet, Charles Hetru and Jules A. Hoffmann give an update of our current information on the induced antibacterial peptides.}, keywords = {hoffmann, M3i}, pubstate = {published}, tppubtype = {article} } @incollection{meister_immune_1994, title = {Immune Gene Expression in Drosophila}, author = {Marie Meister and Philippe Georgel and Bruno Lemaitre and Christine Kappler and Marie Lagueux and Jean-Marc Reichhart and Jules A Hoffmann}, year = {1994}, date = {1994-01-01}, booktitle = {Phylogenetic Perspectives in Immunity : The Insect Host Defense,}, publisher = {Hoffmann JA, Janeway Jr CA, Natori S.}, address = {Austin, Georgetown}, edition = {R.G. Landes Company}, keywords = {hoffmann, M3i, reichhart}, pubstate = {published}, tppubtype = {incollection} } @article{, title = {Threonyl-tRNA synthetase from Thermus thermophilus: purification and some structural and kinetic properties}, author = {J Zheltonosova and E Melnikova and M Garber and J Reinbolt and D Kern and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8031907}, isbn = {8031907}, year = {1994}, date = {1994-01-01}, journal = {Biochimie}, volume = {76}, number = {1}, pages = {71-77}, abstract = {Threonyl-tRNA synthetase (ThrRS) has been isolated from an extreme thermophile Thermus thermophilus strain HB8. The enzyme was purified to electrophoretic homogeneity by combinations of column chromatographies on DEAE-Sepharose, S-Sepharose, ACA-44 Ultrogel and HA-Ultrogel. Seventeen mg of purified enzyme were obtained from 1 kg of biomass. In parallel, purified aspartyl- and phenylalanyl-tRNA synthetases were obtained. The purified ThrRS is composed of two identical subunits with a molecular mass of about 77,000 (virtually the same as E coli ThrRS). The N-terminal sequence has been determined. The homology between the first 45 amino acid residues of ThrRS from T thermophilus and E coli is about 29%. A comparative study of tRNA(Thr) charging by ThrRS from E coli and T thermophilus reveals a similar efficiency of the reaction in both homologous systems. This efficiency remains unchanged for aminoacylation of tRNA(Thr) from T thermophilus by the heterologous ThrRS from E coli, but decreases 700 times for aminoacylation of E coli tRNA(Thr) by ThrRS from T thermophilus.}, note = {0300-9084 Journal Article}, keywords = {Affinity Chromatography, Amino Acid Thermus thermophilus/*enzymology Threonine-tRNA Ligase/chemistry/*isolation & purification/metabolism, Amino Acid Sequence Aspartate-tRNA Ligase/chemistry/isolation & purification Chromatography, Gel Comparative Study Electrophoresis, Polyacrylamide Gel Escherichia coli/cytology/*enzymology Kinetics Molecular Sequence Data Phenylalanine-tRNA Ligase/chemistry/isolation & purification Sequence Alignment Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transfer RNA binding protein in the nucleus of Saccharomyces cerevisiae}, author = {M L Wilhelm and J Reinbolt and J Gangloff and G Dirheimer and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8050578}, isbn = {8050578}, year = {1994}, date = {1994-01-01}, journal = {FEBS Lett}, volume = {349}, number = {2}, pages = {260-264}, abstract = {A yeast nuclear protein that binds to tRNA was identified using a RNA mobility shift assay. Northwestern blotting and N-terminal sequencing experiments indicate that this tRNA-binding protein is identical to zuotin which has previously been shown to bind to Z-DNA [(1992) EMBO J. 11, 3787-3796]. Labeled tRNA and poly(dG-m5dC) stabilized in the Z-DNA form identify the same protein on a Northwestern blot. In a gel retardation assay poly(dG-m5dC) in the Z-form strongly diminishes the binding of tRNA to zuotin. These studies establish that zuotin is able to bind to both tRNA and Z-DNA. Zuotin may be transiently associated with tRNA in the nucleus of yeast cells and play a role in its processing or transport to the cytoplasm.}, note = {0014-5793 Journal Article}, keywords = {Amino Acid Sequence Cell Nucleus/*metabolism Chromatography, Fungal/*isolation & purification RNA, High Pressure Liquid DNA/metabolism DNA-Binding Proteins/genetics/*metabolism Fungal Proteins/genetics/*metabolism Molecular Sequence Data RNA, Transfer/*isolation & purification Saccharomyces cerevisiae/*metabolism *Saccharomyces cerevisiae Proteins, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast Ty1 retrotransposon: the minus-strand primer binding site and a cis-acting domain of the Ty1 RNA are both important for packaging of primer tRNA inside virus-like particles}, author = {M Wilhelm and F X Wilhelm and G Keith and B Agoutin and T Heyman}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7527135}, isbn = {7527135}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {22}, pages = {4560-4565}, abstract = {Reverse transcription of the yeast retrotransposon Ty1 is primed by the cytoplasmic initiator methionine tRNA (tRNA(iMet)). The primer tRNA(iMet) is packaged inside virus-like particles (VLPs) and binds to a 10 nucleotides minus-strand primer binding site, the (-)PBS, complementary to its 3' acceptor stem. We have found that three short sequences of the Ty1 RNA (box 1, box 2.1 and box 2.2) located 3' to the (-)PBS are complementary to other regions of the primer tRNA(iMet) (T psi C and DHU stems and loops). Reconstitution of reverse transcription in vitro with T7 transcribed Ty1 RNA species and tRNA(iMet) purified from yeast cells shows that the boxes do not affect the efficiency of reverse transcription. Thus the role of the boxes on packaging of the primer tRNA(iMet) into the VLPs was investigated by analysing the level of tRNA(iMet) packaged into mutant VLPs. Specific nucleotide changes in the (-)PBS or in the boxes that do not change the protein coding sequence but disrupt the complementarity with the primer tRNA(iMet) within the VLPs. We propose that base pairing between the primer tRNA(iMet) and the Ty1 RNA is of major importance for tRNA(iMet) packaging into the VLPs. Moreover the intactness of the boxes is essential for retrotransposition as shown by the transposition defect of a Ty1 element harboring an intact (-)PBS and mutated boxes.}, note = {0305-1048 Journal Article}, keywords = {Amino Acid Sequence Base Sequence Binding Sites Cloning, Fungal/*genetics RNA, Genetic, Met/*genetics Retroelements/*genetics/physiology Retroviridae/genetics Saccharomyces cerevisiae/*genetics Support, Molecular Molecular Sequence Data Mutation/physiology RNA/*genetics RNA, Non-U.S. Gov't Transcription, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Three-dimensional working model of M1 RNA, the catalytic RNA subunit of ribonuclease P from Escherichia coli}, author = {E Westhof and S Altman}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7515186}, isbn = {7515186}, year = {1994}, date = {1994-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {91}, number = {11}, pages = {5133-5137}, abstract = {A three-dimensional model of M1 RNA, the catalytic RNA subunit of RNase P from Escherichia coli, was constructed with the aid of a computer. The modeling process took into account data from chemical and enzymatic protection experiments, phylogenetic analysis, studies of the activities of mutants, and the kinetics of reactions catalyzed by the binding of substrate to M1 RNA. The model provides a plausible picture of the binding to M1 RNA of the tRNA domain of a precursor tRNA substrate. The scissile bond and adjacent segments of the aminoacyl acceptor stem of a precursor tRNA substrate can fit into a cleft that leads to the phylogenetically conserved, central part of the structure.}, note = {0027-8424 Journal Article}, keywords = {Bacterial/*chemistry/metabolism RNA, Base Sequence Computer Graphics Computer Simulation Endoribonucleases/*chemistry/metabolism Escherichia coli/*enzymology Molecular Sequence Data *Nucleic Acid Conformation RNA, Catalytic/*chemistry/metabolism Ribonuclease P Substrate Specificity Support, Non-U.S. Gov't Support, P.H.S., U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The ribosomal protein S8 from Thermus thermophilus VK1. Sequencing of the gene, overexpression of the protein in Escherichia coli and interaction with rRNA}, author = {V Vysotskaya and S Tischenko and M Garber and D Kern and M Mougel and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7519982}, isbn = {7519982}, year = {1994}, date = {1994-01-01}, journal = {Eur J Biochem}, volume = {223}, number = {2}, pages = {437-445}, abstract = {The gene of the ribosomal protein S8 from Thermus thermophilus VK1 has been isolated from a genomic library by hybridization of an oligonucleotide coding for the N-terminal amino acid sequence of the protein, amplified by PCR and sequenced. Nucleotide sequence reveals an open reading frame coding for a protein of 138 amino acid residues (M(r) 15,839). The codon usage shows that 94% of the codons possess G or C in the third position, and agrees with the preferential usage of codons of high G+C content in the bacteria of the genus Thermus. The amino acid sequence of the protein shows 48% identity with the protein from Escherichia coli. Ribosomal protein S8 from T. thermophilus has been expressed in E. coli under the control of the T7 promoter and purified to homogeneity by heat treatment of the extract followed by cation-exchange chromatography. Conditions were defined in which T. thermophilus protein S8 binds specifically an homologous 16S rRNA fragment containing the putative S8 binding site with an apparent association constant of 5 x 10(7) M-1. The overexpressed protein binds the rRNA with the same affinity as that extracted from T. thermophilus, indicating that the thermophilic protein is correctly folded in E. coli. The specificity of this binding is dependent on the ionic strength. The protein S8 from T. thermophilus recognizes the E. coli rRNA binding sites as efficiently as the S8 protein from E. coli. This result agrees with sequence comparisons of the S8 binding site on the small subunit rRNA from E. coli and from T. thermophilus, showing strong similarities in the regions involved in the interaction. It suggests that the structural features responsible for the recognition are conserved in the mesophilic and thermophilic eubacteria, despite structural peculiarities in the thermophilic partners conferring thermostability.}, note = {0014-2956 Journal Article}, keywords = {16S/*metabolism Recombinant Proteins/metabolism Ribosomal Proteins/chemistry/*genetics/isolation & purification/metabolism Sequence Alignment Support, Amino Acid Sequence Base Sequence Blotting, Bacterial Molecular Sequence Data Molecular Weight Nucleic Acid Hybridization Polymerase Chain Reaction Promoter Regions (Genetics) Protein Binding Protein Structure, Bacterial/chemistry/genetics/isolation & purification Escherichia coli/genetics/metabolism *Gene Expression *Genes, Bacterial/metabolism RNA, Genetic, Molecular DNA, Non-U.S. Gov't Thermus thermophilus/*genetics Transcription, Ribosomal, Secondary RNA, Southern Cloning, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Common structural features of the Ro RNP associated hY1 and hY5 RNAs}, author = {C W van Gelder and J P Thijssen and E C Klaassen and C Sturchler and A Krol and W J van Venrooij and G J Pruijn}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8041611}, isbn = {8041611}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {13}, pages = {2498-2506}, abstract = {The secondary structures of human hY1 and hY5 RNAs were determined using both chemical modification techniques and enzymatic structure probing. The results indicate that both for hY1 and for hY5 RNA the secondary structure largely corresponds to the structure predicted by sequence alignment and computerized energy-minimization. However, some important deviations were observed. In the case of hY1 RNA, two regions forming a predicted helix appeared to be single-stranded. Furthermore, the pyrimidine-rich region of hY1 RNA appeared to be very resistant to reagents under native conditions, although it was accessible to chemical reagents under semi-denaturing conditions. This may point to yet unidentified tertiary interactions for this region of hY1 RNA. In the case of hY5 RNA, two neighbouring internal loops in the predicted structure appeared to form one large internal loop.}, note = {0305-1048 Journal Article}, keywords = {Animals Autoantigens/*genetics/metabolism Base Sequence Cloning, Molecular Dna Human Molecular Sequence Data *Nucleic Acid Conformation RNA Probes RNA, Non-U.S. Gov't, Ribosomal/*chemistry/metabolism Ribonucleoproteins/*genetics/metabolism Sequence Alignment Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A three-dimensional model for the hammerhead ribozyme based on fluorescence measurements}, author = {T Tuschl and C Gohlke and T M Jovin and E Westhof and F Eckstein}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7973630}, isbn = {7973630}, year = {1994}, date = {1994-01-01}, journal = {Science}, volume = {266}, number = {5186}, pages = {785-789}, abstract = {For the understanding of the catalytic function of the RNA hammerhead ribozyme, a three-dimensional model is essential but neither a crystal nor a solution structure has been available. Fluorescence resonance energy transfer (FRET) was used to study the structure of the ribozyme in solution in order to establish the relative spatial orientation of the three constituent Watson-Crick base-paired helical segments. Synthetic constructs were labeled with the fluorescence donor (5-carboxyfluorescein) and acceptor (5-carboxytetramethylrhodamine) located at the ends of the strands constituting the ribozyme molecule. The acceptor helix in helix pairs I and III and in II and III was varied in length from 5 to 11 and 5 to 9 base pairs, respectively, and the FRET efficiencies were determined and correlated with a reference set of labeled RNA duplexes. The FRET efficiencies were predicted on the basis of vector algebra analysis, as a function of the relative helical orientations in the ribozyme constructs, and compared with experimental values. The data were consistent with a Y-shaped arrangement of the ribozyme with helices I and II in close proximity and helix III pointing away. These orientational constraints were used for molecular modeling of a three-dimensional structure of the complete ribozyme.}, note = {0036-8075 Journal Article}, keywords = {Base Composition Base Sequence Energy Transfer Fluoresceins Least-Squares Analysis *Models, Catalytic/*chemistry Rhodamines Software Support, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A three-dimensional model of hepatitis delta virus ribozyme based on biochemical and mutational analyses}, author = {N K Tanner and S Schaff and G Thill and E Petit-Koskas and A M Crain-Denoyelle and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7922369}, isbn = {7922369}, year = {1994}, date = {1994-01-01}, journal = {Curr Biol}, volume = {4}, number = {6}, pages = {488-498}, abstract = {BACKGROUND: Hepatitis delta virus (HDV), which has a single-stranded RNA genome about 1700 nucleotides long, is a satellite virus of hepatitis B, and is associated with a high incidence of fulminant hepatitis and death in infected humans. Like certain pathogenic subviral RNAs that infect plants, HDV RNA features a closed-circular conformation, a rolling-circle mechanism of replication and RNA-catalyzed self-cleaving reactions of both genomic and anti-genomic strands in vitro. The catalytic domains cannot be folded into either the hammerhead or hairpin secondary-structure motifs that have been found in other self-cleaving RNAs. RESULTS: A pseudoknot secondary-structure model has been suggested for the catalytic domain (ribozyme) of HDV RNA. We conducted extensive mutational analyses of regions of the HDV ribozyme predicted in this model to be single stranded, and found that several of them are important for catalytic activity. We used these data, sequence comparisons between different isolates and previously published structural analyses to produce a computer graphic model of the three-dimensional architecture of the HDV ribozyme. CONCLUSIONS: Our model supports the pseudoknotted structure and rationalizes several observations relating to the lengths of the various stems and the sequence requirements of the single-stranded regions. It also provides insight into the catalytic mechanism of the HDV ribozyme. We specifically propose that residues C75, U20 and C21 form the basis of the catalytic region and are close to the cleavable phosphate.}, note = {0960-9822 Journal Article}, keywords = {Base Sequence DNA, Catalytic/*chemistry/genetics/metabolism RNA, Molecular Molecular Sequence Data Mutagenesis, Non-U.S. Gov't, Nucleic Acid Support, Site-Directed Nucleic Acid Conformation RNA, Unité ARN, Viral/chemistry/genetics/metabolism Sequence Homology, Viral/genetics Hepatitis Delta Virus/*enzymology/genetics Human Kinetics Models}, pubstate = {published}, tppubtype = {article} } @article{, title = {Intron-dependent formation of pseudouridines in the anticodon of Saccharomyces cerevisiae minor tRNA(Ile)}, author = {Z Szweykowska-Kulinska and B Senger and G Keith and F Fasiolo and H Grosjean}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7925304}, isbn = {7925304}, year = {1994}, date = {1994-01-01}, journal = {EMBO J}, volume = {13}, number = {19}, pages = {4636-4644}, abstract = {We have isolated and sequenced the minor species of tRNA(Ile) from Saccharomyces cerevisiae. This tRNA contains two unusual pseudouridines (psi s) in the first and third positions of the anticodon. As shown earlier by others, this tRNA derives from two genes having an identical 60 nt intron. We used in vitro procedures to study the structural requirements for the conversion of the anticodon uridines to psi 34 and psi 36. We show here that psi 34/psi 36 modifications require the presence of the pre-tRNA(Ile) intron but are not dependent upon the particular base at any single position of the anticodon. The conversion of U34 to psi 34 occurs independently from psi 36 synthesis and vice versa. However, psi 34 is not formed when the middle and the third anticodon bases of pre-tRNA(Ile) are both substituted to yield ochre anticodon UUA. This ochre pre-tRNA(Ile) mutant has the central anticodon uridine modified to psi 35 as is the case for S.cerevisiae SUP6 tyrosine-inserting ochre suppressor tRNA. In contrast, neither the first nor the third anticodon pseudouridine is formed, when the ochre (UUA) anticodon in the pre-tRNA(Tyr) is substituted with the isoleucine UAU anticodon. A synthetic mini-substrate consisting of the anticodon stem and loop and the wild-type intron of pre-tRNA(Ile) is sufficient to fully modify the anticodon U34 and U36 into psi s. This is the first example of the tRNA intron sequence, rather than the whole tRNA or pre-tRNA domain, being the main determinant of nucleoside modification.}, note = {0261-4189 Journal Article}, keywords = {Anticodon/*metabolism Base Sequence Introns/*physiology Molecular Sequence Data Nucleic Acid Conformation Pseudouridine/*biosynthesis RNA Processing, Fungal/*metabolism RNA, Ile/*metabolism Saccharomyces cerevisiae/*genetics Support, Non-U.S. Gov't, Post-Transcriptional/physiology RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Base modification pattern at the wobble position of Xenopus selenocysteine tRNA(Sec)}, author = {C Sturchler and A Lescure and G Keith and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8031393}, isbn = {8031393}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {8}, pages = {1354-1358}, abstract = {We examined the base modification pattern of Xenopus tRNA(Sec) using microinjection into Xenopus oocytes, with particular focus on the wobble base U34 at the first position of the anticodon. We found that U34 becomes modified to mcm5U34 (5-methylcarboxymethyluridine) in the oocyte cytoplasm in a rather complex manner. When the tRNA(Sec) gene is injected into Xenopus oocyte nuclei, psi 55 and m1A58 are readily obtained, but not mcm5U34. This will appear only upon cytoplasmic injection of the gene product arising from the first nuclear injection. In contrast, tRNA(Sec) produced by in vitro transcription with T7 RNA polymerase readily acquires i6A37, psi 55, m1A58, and mcm5U34. The latter is obtained after direct nuclear or cytoplasmic injections. It has been reported by others that mcm5Um, a 2'-O-methylated derivative of mcm5U34, also exists in rat and bovine tRNA(Sec). With both the gene product and the in vitro transcript, and using the sensitive RNase T2 assay, we were unable to detect under our conditions the presence of a dinucleotide carrying mcm5Um and that would be therefore refractory to hydrolysis. We showed that the unusual mcm5U acquisition pathway does not result from impairment of nucleocytoplasmic transport. Rather, these data can be interpreted to mean that the modification is performed by a tRNA(Sec) specific enzyme, limiting in the oocyte cytoplasm.}, note = {0305-1048 Journal Article}, keywords = {Amino Acid-Specific/chemistry/*genetics/metabolism Selenocysteine/*metabolism Support, Animals *Anticodon Base Composition Base Sequence Microinjections Molecular Sequence Data Mutagenesis Nucleic Acid Conformation RNA, LESCURE, Non-U.S. Gov't Uridine Xenopus laevis, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identification of the primary site of the human immunodeficiency virus type 1 RNA dimerization in vitro}, author = {E Skripkin and J C Paillart and R Marquet and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8197162}, isbn = {8197162}, year = {1994}, date = {1994-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {91}, number = {11}, pages = {4945-4949}, abstract = {The diploid genome of all retroviruses is made of two homologous copies of RNA intimately associated near their 5' end, in a region called the dimer linkage structure. Dimerization of genomic RNA is thought to be important for crucial functions of the retroviral life cycle (reverse transcription, translation, encapsidation). Previous in vitro studies mapped the dimer linkage structure of human immunodeficiency virus type 1 (HIV-1) in a region downstream of the splice donor site, containing conserved purine tracts that were postulated to mediate dimerization, through purine quartets. However, we recently showed that dimerization of HIV-1 RNA also involves sequences upstream of the splice donor site. Here, we used chemical modification interference to identify nucleotides that are required in unmodified form for dimerization of a RNA fragment containing nucleotides 1-707 of HIV-1 RNA. These nucleotides map exclusively in a restricted area upstream of the splice donor site and downstream of the primer binding site. They are centered around a palindromic sequence (GUGCAC279) located in a hairpin loop. Our results support a model in which dimer formation is initiated by the annealing of the palindromic sequences, possibly by a loop-loop interaction between the two monomers. Further experiments show that the deletion of the stem-loop or base substitutions in the loop abolish dimerization, despite the presence of the previously postulated dimer linkage structure. On the other hand, deletions of the purine tracts downstream of the splice donor site do not prevent dimerization. Therefore, we conclude that the palindromic region represents the dimerization initiation site of genomic RNA.}, note = {0027-8424 Journal Article}, keywords = {Base Sequence Binding Sites Biopolymers HIV-1/*genetics Molecular Sequence Data Nucleic Acid Conformation Nucleotides/chemistry RNA, MARQUET, Non-U.S. Gov't, Nucleic Acid Support, PAILLART, Unité ARN, Viral/*chemistry Repetitive Sequences}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transfer RNA profiling: a new method for the identification of pathogenic Candida species}, author = {M A Santos and C el-Adlouni and A D Cox and J M Luz and G Keith and M F Tuite}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7941747}, isbn = {7941747}, year = {1994}, date = {1994-01-01}, journal = {Yeast}, volume = {10}, number = {5}, pages = {625-636}, abstract = {A new molecular taxonomic method applicable to the identification of medically important Candida species and other yeast species has been developed. It is based on the electrophoretic pattern of total tRNA samples (a 'tRNA profile') isolated from Candida species and generated using high-resolution semi-denaturing urea-polyacrylamide gel electrophoresis and methylene blue staining. Species-specific tRNA profiles for the species C. albicans, C. tropicalis, C. parapsilosis, C. guilliermondii, C. glabrata and Pichia guilliermondii were obtained. Detailed studies with the major human pathogen of the Candida genus, C. albicans, demonstrated that the tRNA profile for a given species was both reproducible and strain-independent; seven different C. albicans strains generated identical tRNA profiles. Minor strain-specific heterogeneities in the tRNA profiles of C. guilliermondii and C. parapsilosis were detected, but in neither case did they significantly alter the species-specific diagnostic tRNA profile. The potential of this method in clarifying taxonomic anomalies was demonstrated by the finding that Type I and Type II strains of C. stellatoidea generate very different tRNA profiles, with that of a Type II strain being identical to the C. albicans tRNA profile. This method offers a number of advantages over current electrophoretic karyotype methods for species identification, both within the Candida genus and with yeast species in general.}, note = {0749-503x Journal Article}, keywords = {Candida/classification/*genetics/pathogenicity Electrophoresis, Fungal RNA, Non-U.S. Gov't, Polyacrylamide Gel Genetic Markers RNA, Transfer/*analysis Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Histidylation by yeast HisRS of tRNA or tRNA-like structure relies on residues -1 and 73 but is dependent on the RNA context}, author = {J Rudinger and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7800496}, isbn = {7800496}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {23}, pages = {5031-5037}, abstract = {Residue G-1 and discriminator base C73 are the major histidine identity elements in prokaryotes. Here we evaluate the importance of these two nucleotides in yeast histidine aminoacylation identity. Deletion of G-1 in yeast tRNA(His) transcript leads to a drastic loss of histidylation specificity (about 500-fold). Mutation of discriminator base A73, common to all yeast tRNA(His) species, into G73 has a more moderate but still significant effect with a 22-fold decrease in histidylation specificity. Changes at position 36 in the anticodon loop has negligible effect on histidylation. The role of residues -1 and 73 for specific aminoacylation by yeast HisRS was further investigated by studying the histidylation capacities of seven minihelices derived from the Turnip Yellow Mosaic Virus tRNA-like structure. Changes in the nature of nucleotides -1 and 73 modulate this activity but do not suppress it. The optimal mini-substrate for HisRS presents a G.A mismatch at the position equivalent to residues G-1.A73 in yeast tRNA(His), confirms the importance of this structural feature in yeast histidine identity. The fact that the minisubstrates contain a pseudoknot in which position -1 is mimicked by an internal nucleotide from the pseudoknot highlights further the necessity of a stacking interaction of this position over the amino acid accepting branch of the tRNA during the aminoacylation process. Individual transplantation of G-1 or A73 into yeast tRNA(Asp) transcript improves the histidylation efficiency of the engineered tRNA(Asp). However, a tRNA(Asp) transcript presenting simultaneously both residues G-1 and A73 becomes a less good substrate for HisRS, suggesting the importance of the structural context and/or the presence of antideterminants for an optimal expression of these two identity elements.}, note = {0305-1048 Journal Article}, keywords = {Anticodon/genetics Base Sequence Genes, Asp/genetics RNA, FLORENTZ, His/*chemistry/*genetics RNA, Non-U.S. Gov't Tymovirus/genetics Yeasts/enzymology, Synthetic/genetics Histidine-tRNA Ligase/*metabolism Kinetics Molecular Sequence Data *Nucleic Acid Conformation Point Mutation/physiology RNA, Transfer, Unité ARN, Viral/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {A single methyl group prevents the mischarging of a tRNA}, author = {J Putz and C Florentz and F Benseler and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7634096}, isbn = {7634096}, year = {1994}, date = {1994-01-01}, journal = {Nat Struct Biol}, volume = {1}, number = {9}, pages = {580-582}, note = {1072-8368 Letter}, keywords = {Arginine/*genetics Arginine-tRNA Ligase/metabolism Base Sequence Methylation Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, Asp/chemistry/*genetics/metabolism, FLORENTZ, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural elements of rps0 mRNA involved in the modulation of translational initiation and regulation of E. coli ribosomal protein S15}, author = {C Philippe and L Benard and F Eyermann and C Cachia and S V Kirillov and C Portier and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8041615}, isbn = {8041615}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {13}, pages = {2538-2546}, abstract = {Previous experiments showed that S15 inhibits its own translation by binding to its mRNA in a region overlapping the ribosome loading site. This binding was postulated to stabilize a pseudoknot structure that exists in equilibrium with two stem-loops and to trap the ribosome on its mRNA loading site in a transitory state. In this study, we investigated the effect of mutations in the translational operator on: the binding of protein S15, the formation of the 30S/mRNA/tRNA(fMet) ternary initiation complex, the ability of S15 to inhibit the formation of this ternary complex. The results were compared to in vivo expression and repression rates. The results show that (1) the pseudoknot is required for S15 recognition and translational control; (2) mRNA and 16S rRNA efficiently compete for S15 binding and 16S rRNA suppresses the ability of S15 to inhibit the formation of the active ternary complex; (3) the ribosome binds more efficiently to the pseudoknot than to the stem-loop; (4) sequences located between nucleotides 12 to 47 of the S15 coding phase enhances the efficiency of ribosome binding in vitro; this is correlated with enhanced in vivo expression and regulation rates.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Cloning, Genetic, Messenger/*chemistry/metabolism RNA, Molecular Escherichia coli/*genetics Kinetics Lac Operon Molecular Sequence Data Mutation Nucleic Acid Conformation Operon Protein Binding RNA, Non-U.S. Gov't *Translation, Ribosomal/metabolism Ribosomal Proteins/*genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutational analysis of the bipartite dimer linkage structure of human immunodeficiency virus type 1 genomic RNA}, author = {J C Paillart and R Marquet and E Skripkin and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7961663}, isbn = {7961663}, year = {1994}, date = {1994-01-01}, journal = {J Biol Chem}, volume = {269}, number = {44}, pages = {27486-27493}, abstract = {The genome of all retroviruses consists in two homologous RNA molecules associated near their 5' end in a region called the dimer linkage structure. Dimerization of genomic RNA is thought to be important for several functions of the retroviral cycle such as encapsidation, reverse transcription, and translation. In human immunodeficiency virus type 1 (HIV-1), a region downstream of the splice donor site was initially postulated to mediate dimerization. However, we recently showed that the dimerization initiation site is located upstream of the splice donor site and suggested that dimerization may initiate through a loop-loop interaction. Here, we show that single base mutations in the palindromic loop of the dimerization initiation site completely abolish dimerization, while introduction of compensatory mutations restores the process. Furthermore, two single nucleotide mutants that are unable to form homodimers efficiently codimerize, while the wild type RNA and the compensatory mutant, which both form homodimers, are unable to codimerize. These results unambiguously prove the interaction between the palindromic loops of each monomer. By contrast, none of the deletions that we introduced downstream of the splice donor site abolishes dimerization. However, deletions of two purine tracts located in the vicinity of the initiation codon of the gag gene significantly decrease the thermal stability of the HIV-1 RNA dimer.}, note = {0021-9258 Journal Article}, keywords = {Base Sequence DNA Mutational Analysis HIV-1/*chemistry Heat Hydrogen Bonding Molecular Sequence Data Nucleic Acid Conformation Nucleic Acid Denaturation RNA, MARQUET, Non-U.S. Gov't, PAILLART, Unité ARN, Viral/*chemistry Structure-Activity Relationship Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular recognition of the identity-determinant set of isoleucine transfer RNA from Escherichia coli}, author = {O Nureki and T Niimi and T Muramatsu and H Kanno and T Kohno and C Florentz and R Giege and S Yokoyama}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8114089}, isbn = {8114089}, year = {1994}, date = {1994-01-01}, journal = {J Mol Biol}, volume = {236}, number = {3}, pages = {710-724}, abstract = {Molecular recognition of Escherichia coli tRNA(Ile) by the cognate isoleucyl-tRNA synthetase (IleRS) was studied by analyses of chemical footprinting with N-nitroso-N-ethylurea and aminoacylation kinetics of variant tRNA(Ile) transcripts prepared with bacteriophage T7 RNA polymerase. IleRS binds to the acceptor, dihydrouridine (D), and anticodon stems as well as to the anticodon loop. The "complete set" of determinants for the tRNA(Ile) identity consists of most of the nucleotides in the anticodon loop (G34, A35, U36, t6A37 and A38), the discriminator nucleotide (A73), and the base-pairs in the middle of the anticodon, D and acceptor stems (C29.G41, U12.A23 and C4.G69, respectively). As for the tertiary base-pairs, two are indispensable for the isoleucylation activity, whereas the others are dispensable. Correspondingly, some of the phosphate groups of these dispensable tertiary base-pair residues were shown to be exposed to N-nitroso-N-ethylurea when tRNA(Ile) was bound with IleRS. Furthermore, deletion of the T psi C-arm only slightly impaired the tRNA(Ile) activity. Thus, it is proposed that the recognition by IleRS of all the widely distributed identity determinants is coupled with a global conformational change that involves the loosening of a particular set of tertiary base-pairs of tRNA(Ile).}, note = {0022-2836 Journal Article}, keywords = {Anticodon/chemistry Base Composition Base Sequence Binding Sites Computer Graphics Escherichia coli/genetics/*metabolism Genes, Bacterial Genes, FLORENTZ, Ile/*chemistry/metabolism Support, Molecular Molecular Sequence Data *Nucleic Acid Conformation Nucleic Acid Denaturation RNA, Non-U.S. Gov't, Structural, Synthetic Isoleucine-tRNA Ligase/*metabolism Models, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutations in helix 34 of Escherichia coli 16 S ribosomal RNA have multiple effects on ribosome function and synthesis}, author = {H Moine and A E Dahlberg}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7966269}, isbn = {7966269}, year = {1994}, date = {1994-01-01}, journal = {J Mol Biol}, volume = {243}, number = {3}, pages = {402-412}, abstract = {Helix 34 of E. coli 16 S rRNA (1046 to 1067 and 1189 to 1211) has been proposed to participate directly in the termination of translation at UGA stop codons. We have constructed mutations in this helix in plasmid-encoded rDNA to explore the specific functional roles of the sequence UCAUCA (1199 to 1204) and a secondary structure also involving positions 1054 and 1057-1058. The rRNA mutations were analyzed for their effects on in vivo translational accuracy (stop codon readthrough and frameshifting) as well as growth rate, ribosome synthesis and incorporation into polysomes. Mutations at positions 1054, 1057, 1058, 1199 and 1200 had significant effects on translational accuracy, causing non-specific readthrough of all three stop codons as well as enhanced +1 and -1 frameshifting. Mutations at 1202 and 1203, however, had no effect. The incorporation of deleterious mutant subunits into 70 S ribosomes and polysomes was severely reduced and was associated with a slower growth rate and increased synthesis of host-encoded ribosomes. These data support the proposal that helix 34 is an essential component of the decoding center of the 30 S ribosomal subunit and is not restricted in function to UGA-codon specific termination.}, note = {0022-2836 Journal Article}, keywords = {16S/*chemistry/genetics Ribosomes/*metabolism Support, Base Sequence Codon, Genetic beta-Galactosidase/genetics, Non-U.S. Gov't Support, P.H.S. *Translation, Ribosomal, Terminator Escherichia coli/*genetics/growth & development Molecular Sequence Data *Mutation *Nucleic Acid Conformation RNA, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Slippery substrates}, author = {F Michel and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7656007}, isbn = {7656007}, year = {1994}, date = {1994-01-01}, journal = {Nat Struct Biol}, volume = {1}, number = {1}, pages = {5-7}, note = {1072-8368 News}, keywords = {Animals Binding Sites Models, Catalytic/*chemistry/genetics/metabolism Substrate Specificity Tetrahymena/enzymology/genetics, Molecular Nucleic Acid Conformation RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transcription factors required for the expression of Xenopus laevis selenocysteine tRNA in vitro}, author = {W Meissner and I Wanandi and P Carbon and A Krol and K H Seifart}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8127703}, isbn = {8127703}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {4}, pages = {553-559}, abstract = {It has previously been reported that transcription in vivo of the tRNA(Sec) gene requires three promoter elements, a PSE and a TATA-box upstream of the coding region which are functionally interchangeable with the U6 snRNA gene counterparts and an internal B-block, resembling that of classical tRNA genes (1). We have established an in vitro transcription system from HeLa cells in which three factors, which are either essential for or stimulate transcription were identified. Apart from the TATA-binding protein TBP, the PSE-binding protein PBP was found to be essentially required for expression of the gene. Depletion of PBP from cell extracts by PSE-oligonucleotides abolished tRNA(Sec) transcription, which could be reconstituted by readdition of partially purified PBP. Addition of increasing amounts of recombinant human TBP to an S100 extract stimulated transcription of the tRNA(Sec), the mouse U6 snRNA and the human Y3 genes, an effect which was not observed in the case of a TATA-less tRNA gene. Purified human TFIIA strongly stimulated tRNA(Sec) transcription in a fashion depending on the concentration of TBP. Surprisingly, partially purified TFIIIC was shown to be dispensable for transcription in vitro and unable to bind the B-block of this gene in vitro, although its sequence matches the consensus for this element. Collectively, these data suggest that the mechanism by which transcription complexes are formed on the tRNA(Sec) gene is dramatically different from that observed for classical tRNA genes and much more resembles that observed for externally controlled pol III genes.}, note = {0305-1048 Journal Article}, keywords = {Amino Acid-Specific/*genetics Support, Animals Base Sequence DNA-Binding Proteins/*physiology Human Molecular Sequence Data Oligonucleotide Probes RNA, Genetic Xenopus laevis, Non-U.S. Gov't TATA Box Transcription Factors/*genetics Transcription, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {DRAWNA: a program for drawing schematic views of nucleic acids}, author = {C Massire and C Gaspin and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7529557}, isbn = {7529557}, year = {1994}, date = {1994-01-01}, journal = {J Mol Graph}, volume = {12}, number = {3}, pages = {201-206, 196}, abstract = {A program for drawing automatically exact and schematic views of nucleic acids is described. The program is written in C ANSI and uses the Silicon Graphics GL and Xirisw libraries within the X11/Motif environment. Through menus, the user can choose, specify, and manipulate in real time the three-dimensional views to be displayed. Drawing options include partitioning of structures into differently colored or shaped fragments, representation of backbones as flat or with conic-section ribbons, display of paired or free bases as rods, and display of surfaces as filled or outlined and stereo or depth-cued views.}, note = {0263-7855 Journal Article}, keywords = {Computer Graphics DNA/*chemistry *Models, Molecular Models, Theoretical *Nucleic Acid Conformation RNA/*chemistry *Software, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Dimerization of human immunodeficiency virus type 1 RNA involves sequences located upstream of the splice donor site}, author = {R Marquet and J C Paillart and E Skripkin and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8121797}, isbn = {8121797}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {2}, pages = {145-151}, abstract = {The retroviral genome consists of two homologous RNA molecules associated close to their 5' ends. We studied the spontaneous dimerization of four HIV-1 RNA fragments (RNAs 1-707, 1-615, 311-612, and 311-415) containing the previously defined dimerization domain, and a RNA fragment (RNA 1-311) corresponding to the upstream sequences. Significant dimerization of all RNAs is observed on agarose gels when magnesium is included in the electrophoresis buffer. In contrast to dimerization of RNAs 311-612 and 311-415, dimerization of RNAs 1-707, 1-615 and 1-311 strongly depends on the size of the monovalent cation present in the incubation buffer. Also, dimerization of RNAs 1-707, 1-615, and 1-311 is 10 times faster than that of RNAs 311-612 and 311-415. The dimers formed by the latter RNAs are substantially more stable than that of RNA 1-615, while RNA 1-311 dimer is 5-7 degrees C less stable than RNA 1-615 dimer. These results indicate that dimerization of HIV-1 genomic RNA involves elements located upstream of the splice donor site (position 305), i.e. outside of the previously defined dimerization domain.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Cations HIV-1/*genetics Kinetics Macromolecular Systems Magnesium Models, Chemical Models, Genetic Molecular Sequence Data RNA Splicing RNA, MARQUET, Non-U.S. Gov't Thermodynamics, PAILLART, Unité ARN, Viral/*chemistry Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallisation of the glycyl-tRNA synthetase from Thermus thermophilus and initial crystallographic data}, author = {D T Logan and V Cura and J P Touzel and D Kern and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8071996}, isbn = {8071996}, year = {1994}, date = {1994-01-01}, journal = {J Mol Biol}, volume = {241}, number = {5}, pages = {732-735}, abstract = {The glycyl-tRNA synthetase from Thermus thermophilus is a dimer of molecular mass 115 kDa, which has been crystallised using the vapour diffusion method from 5 to 7% polyethylene glycol 6000, 0.8 to 1.4 M NaCl at protein concentrations of 2 to 8 mg/ml. Nucleation is carried out at 4 degrees C and crystals are subsequently transferred to 15 degrees C to maximise growth. Crystals are truncated rhombohedra measuring on average 0.4 mm x 0.4 mm x 0.2 mm, which appear within a few days and reach full size in one to two months. GlyRS crystallises in two closely related space groups, P2(1)2(1)2(1) and C2,2,2(1), both with the same cell a = 125 A}, note = {0022-2836 Journal Article}, keywords = {Crystallization Crystallography, Unité ARN, X-Ray Glycine-tRNA Ligase/*chemistry Molecular Structure Thermus thermophilus/*enzymology}, pubstate = {published}, tppubtype = {article} } @article{, title = {The N-terminal domain of the human TATA-binding protein plays a role in transcription from TATA-containing RNA polymerase II and III promoters}, author = {A Lescure and Y Lutz and D Eberhard and X Jacq and A Krol and I Grummt and I Davidson and P Chambon and L Tora}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7510635}, isbn = {7510635}, year = {1994}, date = {1994-01-01}, journal = {EMBO J}, volume = {13}, number = {5}, pages = {1166-1175}, abstract = {In eukaryotes, the TATA box binding protein (TBP) is an integral component of the transcription initiation complexes of all three classes of nuclear RNA polymerases. In this study we have investigated the role of the N-terminal region of human TBP in transcription initiation from RNA polymerase (Pol) I, II and III promoters by using three monoclonal antibodies (mAbs). Each antibody recognizes a distinct epitope in the N-terminal domain of human TBP. We demonstrate that these antibodies differentially affect transcription from distinct classes of promoters. One antibody, mAb1C2, and a synthetic peptide comprising its epitope selectively inhibited in vitro transcription from TATA-containing, but not from TATA-less promoters, irrespective of whether they were transcribed by Pol II or Pol III. Transcription by Pol I, on the other hand, was not affected. Two other antibodies and their respective epitope peptides did not affect transcription from any of the promoters tested. Order of addition experiments indicate that mAb1C2 did not prevent binding of TBP to the TATA box or the formation of the TBP-TFIIA-TFIIB complex but rather inhibited a subsequent step of preinitiation complex formation. These data suggest that a defined region within the N-terminal domain of human TBP may be involved in specific protein-protein interactions required for the assembly of functional preinitiation complexes on TATA-containing, but not on TATA-less promoters.}, note = {0261-4189 Journal Article}, keywords = {Antibodies, Genetic, LESCURE, Non-U.S. Gov't *TATA Box TATA-Box Binding Protein Transcription Factor TFIIA Transcription Factor TFIIB Transcription Factor TFIID Transcription Factors/biosynthesis/isolation & purification/*metabolism *Transcription, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Involvement of a GNRA tetraloop in long-range RNA tertiary interactions}, author = {L Jaeger and F Michel and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7510342}, isbn = {7510342}, year = {1994}, date = {1994-01-01}, journal = {J Mol Biol}, volume = {236}, number = {5}, pages = {1271-1276}, abstract = {Terminal loops with a GNRA consensus sequence are widespread in RNA. It has been suggested that these loops act as "anchors" during tertiary folding, by interacting in a sequence-specific way with helices at distant locations along the molecule. We now show that a GUGA loop changes state upon disruption of the tertiary architecture of a self-splicing group I intron. Successful replacement of the postulated loop-helix contact by classical base-pairing points to binding of the loop into the shallow (minor) groove of the helix, as also indicated by partial restoration of ribozyme stability upon a specific double nucleotide substitution.}, note = {0022-2836 Journal Article}, keywords = {Bacteriophage T4 Base Sequence Hydrogen Bonding Introns Molecular Sequence Data *Nucleic Acid Conformation RNA/*chemistry RNA, Non-U.S. Gov't, Unité ARN, Viral/chemistry Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure and site-directed mutagenesis of a bleomycin resistance protein and their significance for drug sequestering}, author = { P. Dumas and M. Bergdoll and C. Cagnon and J. M. Masson}, year = {1994}, date = {1994-01-01}, journal = {EMBO J}, volume = {13}, number = {11}, pages = {2483-92}, abstract = {The antibiotic bleomycin, a strong DNA cutting agent, is naturally produced by actinomycetes which have developed a resistance mechanism against such a lethal compound. The crystal structure, at 2.3 A resolution, of a bleomycin resistance protein of 14 kDa reveals a structure in two halves with the same alpha/beta fold despite no sequence similarity. The crystal packing shows compact dimers with a hydrophobic interface and involved in mutual chain exchange. Two independent solution studies (analytical centrifugation and light scattering) showed that this dimeric form is not a packing artefact but is indeed the functional one. Furthermore, light scattering also showed that one dimer binds two antibiotic molecules as expected. A crevice located at the dimer interface, as well as the results of a site-directed mutagenesis study, led to a model wherein two bleomycin molecules are completely sequestered by one dimer. This provides a novel insight into antibiotic resistance due to drug sequestering, and probably also into drug transport and excretion.}, note = {0261-4189 Journal Article}, keywords = {*Acetyltransferases, &, Acid, Amino, Bacterial, Bacterial/*genetics, Base, Binding, Bleomycin/*metabolism/pharmacology, Conformation, Crystallization, Crystallography, Data, Drug, Fusion, Genes, Gov't, Microbial/genetics, Models, Molecular, Mutagenesis, Non-U.S., Protein, Proteins/*chemistry/genetics/isolation, Proteins/isolation, purification, purification/metabolism, Recombinant, Relationship, Resistance, Secondary, Sequence, Site-Directed, Sites, Structural, structure, Structure-Activity, Support, X-Ray}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast serine isoacceptor tRNAs: variations of their content as a function of growth conditions and primary structure of the minor tRNA(Ser)GCU}, author = {T Heyman and B Agoutin and C Fix and G Dirheimer and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8033992}, isbn = {8033992}, year = {1994}, date = {1994-01-01}, journal = {FEBS Lett}, volume = {347}, number = {2-3}, pages = {143-146}, abstract = {The primary structure of Saccharomyces cerevisiae tRNA(Ser)GCU is presented (EMBL database accession No. X74268 S. cerevisiae tRNA-Ser). In addition, quantitation of the relative amounts of serine isoaccepting tRNAs in yeast grown on different media showed that the minor tRNA(Ser)GCU decreased while the major tRNA(Ser)AGA increased as the growth rate and the cellular protein content increased. The minor species, tRNA(Ser)CGA and tRNA(Ser)UGA, were not separated by our gel system, however, taken together they appeared to vary in the same way as tRNA(Ser)GCU. These data suggest a growth rate dependence of yeast tRNAs similar to that previously described for E. coli tRNAs.}, note = {0014-5793 Journal Article}, keywords = {Anticodon Base Sequence Culture Media Galactose Molecular Sequence Data Nucleic Acid Conformation Nucleic Acid Hybridization RNA Probes RNA, Fungal/*chemistry RNA, Ser/analysis/*chemistry Saccharomyces cerevisiae/*genetics/*growth & development, Transfer, Transfer/*chemistry RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallogenesis of biological macromolecules: facts and perspectives}, author = {R Giege and B Lorber and A Théobald-Dietrich}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15299382}, isbn = {15299382}, year = {1994}, date = {1994-01-01}, journal = {Acta Crystallogr D Biol Crystallogr}, volume = {50}, number = {Pt 4}, pages = {339-350}, abstract = {This paper gives an overview of the science of crystals of biological macromolecules. The historical background of the field is outlined and the main achievements and open problems are discussed from both biological and physical-chemical viewpoints. Selected results, including data from the authors, illustrate this overview. The perspectives of crystallogenesis for structural biology, but also more general trends, are presented.}, note = {0907-4449 Journal Article}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {The determination of the secondary structures of RNA as a constraint satisfaction problem.}, author = {C Gaspin and E Westhof}, editor = {S Schulze-Kremer}, url = {http://books.google.fr/books?hl=de&id=VmFSNNm7k6cC&q=Westhof#v=snippet&q=Westhof&f=false}, year = {1994}, date = {1994-01-01}, booktitle = {Advances in Molecular Bioinformatics (Frontiers in Artificial Intelligence and Applications)}, pages = {103-122}, publisher = {IOS Press}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Identity switches between tRNAs aminoacylated by class I glutaminyl- and class II aspartyl-tRNA synthetases}, author = {M Frugier and D Soll and R Giege and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8060999}, isbn = {8060999}, year = {1994}, date = {1994-01-01}, journal = {Biochemistry}, volume = {33}, number = {33}, pages = {9912-9921}, abstract = {High-resolution X-ray structures for the tRNA/aminoacyl-tRNA synthetase complexes between Escherichia coli tRNAGln/GlnRS and yeast tRNAAsp/AspRS have been determined. Positive identity nucleotides that direct aminoacylation specificity have been defined in both cases; E. coli tRNAGln identity is governed by 10 elements scattered in the tRNA structure, while specific aminoacylation of yeast tRNAAsp is dependent on 5 positions. Both identity sets are partially overlapping and share 3 nucleotides. Interestingly, the two enzymes belong to two different classes described for aminoacyl-tRNA synthetases. The class I glutaminyl-tRNA synthetase and the class II aspartyl-tRNA synthetase recognize their cognate tRNA from opposite sides. Mutants derived from glutamine and aspartate tRNAs have been created by progressively introducing identity elements from one tRNA into the other one. Glutaminylation and aspartylation assays of the transplanted tRNAs show that identity nucleotides from a tRNA originally aminoacylated by a synthetase from one class are still recognized if they are presented to the enzyme in a structural framework corresponding to a tRNA aminoacylated by a synthetase belonging to the other class. The simple transplantation of the glutamine identity set into tRNAAsp is sufficient to obtain glutaminylatable tRNA, but additional subtle features seem to be important for the complete conversion of tRNAGln in an aspartylatable substrate. This study defines C38 in yeast tRNAAsp as a new identity nucleotide for aspartylation. We show also in this paper that, during the complex formation, aminoacyl-tRNA synthetases are at least partially responsible for conformational changes which involve structural constraints in tRNA molecules.}, note = {0006-2960 Journal Article}, keywords = {Acylation Aspartate-tRNA Ligase/chemistry/*metabolism Base Sequence Crystallization Escherichia coli/*enzymology/genetics Glutamate-tRNA Ligase/chemistry/*metabolism Kinetics Molecular Sequence Data Molecular Structure Mutation Nucleic Acid Conformation RNA, Asp/chemistry/*metabolism RNA, ERIANI, FLORENTZ, FRUGIER, Gln/chemistry/*metabolism Saccharomyces cerevisiae/*enzymology/genetics Support, Non-U.S. Gov't Support, P.H.S., Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Synthetic polyamines stimulate in vitro transcription by T7 RNA polymerase}, author = {M Frugier and C Florentz and M W Hosseini and J M Lehn and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8052534}, isbn = {8052534}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {14}, pages = {2784-2790}, abstract = {The influence of nine synthetic polyamines on in vitro transcription with T7 RNA polymerase has been studied. The compounds used were linear or macrocyclic tetra- and hexaamine, varying in their size, shape and number of protonated groups. Their effect was tested on different types of templates, all presenting the T7 RNA promoter in a double-stranded form followed by sequences encoding short transcripts (25 to 35-mers) either on single- or double-stranded synthetic oligodeoxyribonucleotides. All polyamines used stimulate transcription of both types of templates at levels dependent on their size, shape, protonation degree, and concentration. For each compound, an optimal concentration could be defined; above this concentration, transcription inhibition occurred. Highest stimulation (up to 12-fold) was obtained by the largest cyclic compound called [38]N6C10.}, note = {0305-1048 Journal Article}, keywords = {Bacteriophage T7/enzymology Base Sequence Comparative Study DNA-Directed RNA Polymerases/drug effects/*metabolism Kinetics Molecular Sequence Data Molecular Structure Nucleic Acid Conformation Oligodeoxyribonucleotides Polyamines/chemistry/*pharmacology Promoter Regions (Genetics) RNA, ERIANI, FLORENTZ, FRUGIER, Genetic Transcription, Genetic/*drug effects, Non-U.S. Gov't Templates, Transfer, Unité ARN, Val/*biosynthesis/chemistry Structure-Activity Relationship Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Efficient aminoacylation of resected RNA helices by class II aspartyl-tRNA synthetase dependent on a single nucleotide}, author = {M Frugier and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8187774}, isbn = {8187774}, year = {1994}, date = {1994-01-01}, journal = {EMBO J}, volume = {13}, number = {9}, pages = {2218-2226}, abstract = {We show here that small RNA helices which recapitulate part or all of the acceptor stem of yeast aspartate tRNA are efficiently aminoacylated by cognate class II aspartyl-tRNA synthetase. Aminoacylation is strongly dependent on the presence of the single-stranded G73 'discriminator' identity nucleotide and is essentially insensitive to the sequence of the helical region. Substrates which contain as few as 3 bp fused to G73CCAOH are aspartylated. Their charging is insensitive to the sequence of the loop closing the short helical domains. Aminoacylation of the aspartate mini-helix is not stimulated by a hairpin helix mimicking the anticodon domain and containing the three major anticodon identity nucleotides. A thermodynamic analysis demonstrates that enzyme interactions with G73 in the resected RNA substrates and in the whole tRNA are the same. Thus, if the resected RNA molecules resemble in some way the earliest substrates for aminoacylation with aspartate, then the contemporary tRNA(Asp) has quantitatively retained the influence of the major signal for aminoacylation in these substrates.}, note = {0261-4189 Journal Article}, keywords = {Acylation Anticodon Aspartate-tRNA Ligase/*metabolism Aspartic Acid/metabolism Base Sequence Evolution Molecular Sequence Data Nucleic Acid Conformation RNA, ERIANI, FLORENTZ, FRUGIER, Fungal/chemistry/*metabolism Substrate Specificity Support, Non-U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A histidine accepting tRNA-like fold at the 3'-end of satellite tobacco mosaic virus RNA}, author = {B Felden and C Florentz and A McPherson and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8065897}, isbn = {8065897}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {15}, pages = {2882-2886}, abstract = {A model of secondary structure is proposed for the 3'-terminal sequence of the satellite tobacco mosaic virus (STMV) RNA on the basis of phylogenetic comparisons with tobacco mosaic virus (TMV) genomic RNA. Sequence homologies and compensatory base changes found between the two related viral RNAs imply that the 3'-end of STMV RNA folds into a tRNA-like domain similar to that found in the TMV RNA. Accordingly, functional assays showed that STMV RNA can be aminoacylated in vitro with histidine by yeast histidyl-tRNA synthetase to plateaus reaching 30%. Histidylation properties of STMV RNA were compared to those of TMV RNA and of a canonical yeast tRNA(His) transcript which both are chargeable to nearly 100% plateau levels. Kinetic data indicate an excellent catalytic efficiency of STMV RNA charging expressed as Vmax/Km ratio, quasi-equivalent to that of TMV RNA, and only 17-fold reduced as compared to that of the yeast tRNAHis transcript. Biological implications of the structural mimicry between the tRNA-like regions of TMV and STMV RNAs are discussed in the light of the relationships of a satellite virus with its helper virus. This is the first report on a chargeable tRNA-like structure at the 3'-end of a satellite virus RNA.}, note = {0305-1048 Journal Article}, keywords = {Acylation Base Sequence Comparative Study Histidine/*metabolism Histidine-tRNA Ligase/metabolism Kinetics Molecular Sequence Data *Nucleic Acid Conformation Phylogeny RNA, FLORENTZ, His/metabolism RNA, Non-U.S. Gov't Tobacco Mosaic Virus/*genetics, Transfer, Transfer/*chemistry RNA, Unité ARN, Viral/*chemistry Saccharomyces cerevisiae/enzymology Sequence Homology Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Solution structure of the 3'-end of brome mosaic virus genomic RNAs. Conformational mimicry with canonical tRNAs}, author = {B Felden and C Florentz and R Giege and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8289279}, isbn = {8289279}, year = {1994}, date = {1994-01-01}, journal = {J Mol Biol}, volume = {235}, number = {2}, pages = {508-531}, abstract = {The conformation of the last 201 nucleotides located at the 3'-end of brome mosaic virus (BMV) RNAs was investigated in solution using different chemical and enzymatic probes. Bases were probed with dimethylsulfate (which methylates N-1 positions of A, N-3 positions of C and N-7 positions of G), a carbodiimide (which modifies N-1 positions of G and N-3 positions of U) and diethylpyrocarbonate (which modifies N-7 positions of A). Ribonucleases T1, U2 and S1 were used to map unpaired nucleotides and ribonuclease V1 to monitor paired bases or stacked nucleotides. Cleavage or modification sites were detected by gel electrophoresis either indirectly by analyzing DNA sequence patterns generated by primer extension with reverse transcriptase of the modified RNAs or by direct identification within the statistical cleavage patterns of the RNA. On the basis of these biochemical results, an atomic model was built by computer modeling and its stereochemistry refined. The deduced secondary structure of the RNA confirms data previously proposed by others but contains additional base-pairs (A27-U32, A28-G31, G41-A134, G64-C68, U80-A99, G81-A98, G88-U91, G100-U126, U104-U125, G162-G166 and A172-A191), one new tertiary long-range interaction (U103-U164) and a small triple helical conformation with (G41-A134)-A18 and (C42-G133)-A17 interactions. The new secondary structure also indicates the existence of a second pseudoknot involving pairing between residues A181 to A184 and residues U197 to U194, outside the domain conferring tyrosylation ability to BMV RNA. The main outcome from the model stems from its intricate folding, which allows a new assignment for the domains mimicking the anticodon- and D-loop regions of tRNA. Interestingly, the stem and loop region found structurally to be analogous to the anticodon arm of tRNA(Tyr) does not contain the tyrosine anticodon involved in the aminoacylation process. The structural analogies with canonical tRNA(Tyr) illustrate the functional mimicry existing between the BMV RNA structure and canonical tRNA(Tyr) that allows for their efficient aminoacylation by tyrosyl-tRNA synthetase. This structural model rationalizes mutagenic and footprinting data that have established the importance of specific regions of the viral RNA for recognition by its replicase, (ATP,CTP):tRNA nucleotidyl-transferase and yeast tyrosyl-tRNA synthetase. The new fold has biological implications that can be used as a predictive tool for elaborating new experiments.}, note = {0022-2836 Journal Article}, keywords = {Base Sequence Bromovirus/*genetics Computer Simulation Models, FLORENTZ, Genetic Models, Molecular Molecular Sequence Data Nucleic Acid Conformation RNA, Non-U.S. Gov't, Transfer, Tyr/*chemistry RNA, Unité ARN, Viral/*chemistry Solutions Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystal structure and site-directed mutagenesis of a bleomycin resistance protein and their significance for drug sequestering}, author = {P Dumas and M Bergdoll and C Cagnon and J M Masson}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7516875}, isbn = {7516875}, year = {1994}, date = {1994-01-01}, journal = {EMBO J}, volume = {13}, number = {11}, pages = {2483-2492}, abstract = {The antibiotic bleomycin, a strong DNA cutting agent, is naturally produced by actinomycetes which have developed a resistance mechanism against such a lethal compound. The crystal structure, at 2.3 A resolution, of a bleomycin resistance protein of 14 kDa reveals a structure in two halves with the same alpha/beta fold despite no sequence similarity. The crystal packing shows compact dimers with a hydrophobic interface and involved in mutual chain exchange. Two independent solution studies (analytical centrifugation and light scattering) showed that this dimeric form is not a packing artefact but is indeed the functional one. Furthermore, light scattering also showed that one dimer binds two antibiotic molecules as expected. A crevice located at the dimer interface, as well as the results of a site-directed mutagenesis study, led to a model wherein two bleomycin molecules are completely sequestered by one dimer. This provides a novel insight into antibiotic resistance due to drug sequestering, and probably also into drug transport and excretion.}, note = {0261-4189 Journal Article}, keywords = {*Acetyltransferases Amino Acid Sequence Bacterial Proteins/*chemistry/genetics/isolation & purification/metabolism Base Sequence Binding Sites Bleomycin/*metabolism/pharmacology Crystallization Crystallography, Bacterial/*genetics Models, Microbial/genetics Genes, Molecular Molecular Sequence Data Mutagenesis, Non-U.S. Gov't, Secondary Recombinant Fusion Proteins/isolation & purification Structure-Activity Relationship Support, Site-Directed Protein Conformation Protein Structure, Structural, Unité ARN, X-Ray Drug Resistance}, pubstate = {published}, tppubtype = {article} } @article{, title = {The active site of yeast aspartyl-tRNA synthetase: structural and functional aspects of the aminoacylation reaction}, author = {J Cavarelli and G Eriani and B Rees and M Ruff and M Boeglin and A Mitschler and F Martin and J Gangloff and J C Thierry and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8313877}, isbn = {8313877}, year = {1994}, date = {1994-01-01}, journal = {EMBO J}, volume = {13}, number = {2}, pages = {327-337}, abstract = {The crystal structures of the various complexes formed by yeast aspartyl-tRNA synthetase (AspRS) and its substrates provide snapshots of the active site corresponding to different steps of the aminoacylation reaction. Native crystals of the binary complex tRNA-AspRS were soaked in solutions containing the two other substrates, ATP (or its analog AMPPcP) and aspartic acid. When all substrates are present in the crystal, this leads to the formation of the aspartyl-adenylate and/or the aspartyl-tRNA. A class II-specific pathway for the aminoacylation reaction is proposed which explains the known functional differences between the two classes while preserving a common framework. Extended signature sequences characteristic of class II aaRS (motifs 2 and 3) constitute the basic functional unit. The ATP molecule adopts a bent conformation, stabilized by the invariant Arg531 of motif 3 and a magnesium ion coordinated to the pyrophosphate group and to two class-invariant acidic residues. The aspartic acid substrate is positioned by a class II invariant acidic residue, Asp342, interacting with the amino group and by amino acids conserved in the aspartyl synthetase family. The amino acids in contact with the substrates have been probed by site-directed mutagenesis for their functional implication.}, note = {0261-4189 Journal Article}, keywords = {Acylation Adenosine Triphosphate/metabolism Amino Acid Sequence Animals Aspartate-tRNA Ligase/chemistry/genetics/*metabolism Binding Sites Computer Graphics Human Molecular Sequence Data Mutagenesis, Amino Acid Structure-Activity Relationship Support, ERIANI, Non-U.S. Gov't, Site-Directed Saccharomyces cerevisiae/*enzymology Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Modulation of the suppression efficiency and amino acid identity of an artificial yeast amber isoleucine transfer RNA in Escherichia coli by a G-U pair in the anticodon stem}, author = {V Buttcher and B Senger and S Schumacher and J Reinbolt and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8166708}, isbn = {8166708}, year = {1994}, date = {1994-01-01}, journal = {Biochem Biophys Res Commun}, volume = {200}, number = {1}, pages = {370-377}, abstract = {The artificial amber suppressor corresponding to the major isoleucine tRNA from yeast (pVBt5), when expressed in E. coli, is a poor suppressor of the amber mutation lacIam181-Z. By analysing mutant forms, we could show that this was due to the presence of a U30-G40 wobble pair in the anticodon stem of the yeast tRNA and not to the level of the heterologously expressed tRNA. Efficient suppressors were obtained by restoring a normal U30-A40 or G30-C40 Watson-Crick pair. In vivo the mutant forms are exclusively charged by the bacterial lysyl-tRNA synthetase (LysRS), whereas the original yeast amber tRNA is charged at a low level by E. coli glutaminyl-tRNA synthetase (GlnRS) and LysRS. The inversion of the U30-G40 pair also induces a loss of the Gln identity. We conclude from these experiments that the U30-G40 base pair constitutes a negative determinant for LysRS interaction which operates either at the level of complex formation or at the catalytic step. As no direct contacts are seen between GlnRS and positions 30-40 of the complexed homologous tRNA, the U30-G40 pair of pVBt5 is believed to influence aminoacylation by GlnRS indirectly, probably at the level of the anticodon loop conformation by favouring an optimal apposition of the anticodon nucleotides with the protein.}, note = {0006-291x Journal Article}, keywords = {Amino Acyl-tRNA Ligases/metabolism Anticodon/*genetics Base Composition Base Sequence Chromosomes, Artificial, Bacterial Guanine Inversion (Genetics) Lysine-tRNA Ligase/metabolism Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Plasmids RNA, Genetic Tetrahydrofolate Dehydrogenase/biosynthesis/genetics/isolation & purification Uracil, Gln/chemistry/genetics RNA, Ile/chemistry/*genetics RNA, Lys/chemistry/*genetics Saccharomyces cerevisiae/*genetics *Suppression, Structural, Transfer, Unité ARN, Yeast Escherichia coli/*genetics Genes}, pubstate = {published}, tppubtype = {article} } @article{, title = {Replication control of plasmid R1: disruption of an inhibitory RNA structure that sequesters the repA ribosome-binding site permits tap-independent RepA synthesis}, author = {P Blomberg and H M Engdahl and C Malmgren and P Romby and E G Wagner}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7520116}, isbn = {7520116}, year = {1994}, date = {1994-01-01}, journal = {Mol Microbiol}, volume = {12}, number = {1}, pages = {49-60}, abstract = {The replication frequency of plasmid R1 is controlled by an antisense RNA, CopA, that inhibits the synthesis of the replication initiator protein, RepA, at the post-transcriptional level. This inhibition is indirect and affects translation of a leader peptide reading frame (tap). Translation of tap is required for repA translation (Blomberg et al., 1992). Here we asked whether an RNA stem-loop sequestering the repA ribosome-binding site blocks tap translation-independent repA expression. Destabilization of this structure resulted in tap-independent RepA synthesis, concomitant with a loss of CopA-mediated inhibition; thus, CopA acts at the level of tap translation. Structure probing of RepA mRNAs confirmed that the introduced mutations induced a local destabilization in the repA ribosome-binding site stem-loop. An increased spacing between the repA Shine-Dalgarno region and the start codon permitted even higher repA expression. In Incl alpha/IncB plasmids, an RNA pseudoknot acts as an activator for rep translation. We suggest that the regulatory pathway in plasmid R1 does not involve an activator RNA pseudoknot.}, note = {0950-382x Journal Article}, keywords = {Antisense/chemistry/*physiology RNA, Bacterial Models, Bacterial Proteins/genetics/*metabolism Base Sequence Binding Sites *DNA Replication *Gene Expression Regulation, Bacterial/*genetics Reading Frames Ribosomes/*metabolism Sequence Alignment Support, Genetic, Genetic Molecular Sequence Data Mutagenesis Nucleic Acid Conformation Peptides/*genetics/physiology *Proteins R Factors/*genetics RNA, Non-U.S. Gov't Translation, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutational analysis of the pseudoknot structure of the S15 translational operator from Escherichia coli}, author = {L Benard and C Philippe and L Dondon and M Grunberg-Manago and B Ehresmann and C Ehresmann and C Portier}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7830558}, isbn = {7830558}, year = {1994}, date = {1994-01-01}, journal = {Mol Microbiol}, volume = {14}, number = {1}, pages = {31-40}, abstract = {Expression of rpsO, the gene encoding the small ribosomal protein S15, is autoregulated at the translational level by S15, which binds to its mRNA in a region overlapping the ribosome-binding site. By measuring the effect of mutations on the expression of a translational rpsO-lacZ fusion and the S15 binding affinity for the translational operator, the formation of a pseudoknot in the operator site in vivo is fully demonstrated and appears to be a prerequisite for S15 binding. The mutational analysis suggests also that specific determinants for S15 binding are located in very limited regions of the structure formed by the pseudoknot. It is deduced that a specific pseudoknot conformation is a key element for autoregulation.}, note = {0950-382x Journal Article}, keywords = {Bacterial Molecular Sequence Data *Nucleic Acid Conformation Plasmids Protein Binding RNA, Bacteriophage lambda/genetics Base Sequence Binding Sites Codon DNA Mutational Analysis Escherichia coli/*genetics/metabolism *Gene Expression Regulation, Genetic beta-Galactosidase/biosynthesis, Genetic Translation, Messenger/*chemistry/*metabolism Recombinant Fusion Proteins/biosynthesis Restriction Mapping Ribosomal Proteins/biosynthesis/*genetics/metabolism Support, Non-U.S. Gov't Transcription, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Eukaryotic selenocysteine inserting tRNA species support selenoprotein synthesis in Escherichia coli}, author = {C Baron and C Sturchler and X Q Wu and H J Gross and A Krol and A Bock}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8036149}, isbn = {8036149}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {12}, pages = {2228-2233}, abstract = {Although the tRNA species directing selenocysteine insertion in prokaryotes differ greatly in their primary structure from that of their eukaryotic homologues they share very similar three-dimensional structures. To analyse whether this conservation of the overall shape of the molecules reflects a conservation of their functional interactions it was tested whether the selenocysteine inserting tRNA species from Homo sapiens supports selenoprotein synthesis in E. coli. It was found that the expression of the human tRNA(Sec) gene in E.coli can complement a lesion in the tRNA(Sec) gene of this organism. Transcripts of the Homo sapiens and Xenopus laevis tRNA(Sec) genes synthesised in vitro were amino-acylated by the E.coli seryl-tRNA ligase although at a very low rate and the resulting seryl-tRNA(Sec) was bound to and converted into selenocysteyl-tRNA(Sec) by the selenocysteine synthase of this organism. Selenocysteyl-tRNA(Sec) from both eukaryotes was able to form a complex with translation factor SELB from E.coli. Although the mechanism of selenocysteine incorporation into seleno-proteins appears to be rather different in E.coli and in vertebrates, we observe here a surprising conservation of functions over an enormous evolutionary distance.}, note = {0305-1048 Journal Article}, keywords = {Amino Acid-Specific/chemistry/*genetics Selenocysteine/chemistry Serine-tRNA Ligase/metabolism Support, Animals Bacterial Proteins/metabolism Base Sequence Cloning, Molecular Escherichia coli/*genetics Genetic Complementation Test Human Nucleic Acid Conformation Peptide Elongation Factors/metabolism Proteins/*biosynthesis/genetics RNA, Non-U.S. Gov't Xenopus laevis, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Deficiency of queuine, a highly modified purine base, in transfer RNAs from primary and metastatic ovarian malignant tumors in women}, author = {W Baranowski and G Dirheimer and J A Jakowicki and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8044797}, isbn = {8044797}, year = {1994}, date = {1994-01-01}, journal = {Cancer Res}, volume = {54}, number = {16}, pages = {4468-4471}, abstract = {The tRNAs from rapidly growing tissues, particularly from neoplasia, often exhibit queuine deficiency. In order to check whether different kinds of ovarian tumors display queuine deficiencies we have analyzed tRNA samples from 16 ovarian malignancies. The tRNAs from histologically normal myometrium (4 samples) and myoma (6 samples) were taken as healthy tissue and benign tumor references. Queuine deficiency was determined by an exchange assay using [8-3H]guanine and tRNA:guanine transglycosylase from Escherichia coli. The mean values of queuine deficiencies in tRNAs were: 10.95 +/- 2.21 (SD) pmol/A260 in gonadal and germ cell tumors (5 cases); 23.75 +/- 7.89 pmol/A260 in primary epithelial tumors (9 cases); and 34.58 +/- 7.18 pmol/A260 in metastatic tumors (2 cases). These values displayed statistically significant differences (P = 0.0003, Kruskal-Wallis test). The queuine deficiencies in tRNAs significantly increased when moving from well-differentiated through moderately differentiated to poorly differentiated tumors, with the highest values found in poorly differentiated metastatic tumors (P = 0.0002, Kruskal-Wallis test). Queuine deficiency determination in tRNAs is proposed as a factor for clinical outcome prognosis of ovarian malignancies.}, note = {0008-5472 Journal Article}, keywords = {Adolescent Adult Female Guanine/*analogs & derivatives/analysis Human Middle Aged Ovarian Neoplasms/*chemistry/pathology RNA, Neoplasm/*chemistry RNA, Non-U.S. Gov't, Transfer/*chemistry Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of conserved nucleotides in building the 16S rRNA binding site of E. coli ribosomal protein S8}, author = {C Allmang and M Mougel and E Westhof and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7937081}, isbn = {7937081}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {18}, pages = {3708-3714}, abstract = {Ribosomal protein S8 specifically recognizes a helical and irregular region of 16S rRNA that is highly evolutionary constrained. Despite its restricted size, the precise conformation of this region remains a question of debate. Here, we used chemical probing to analyze the structural consequences of mutations in this RNA region. These data, combined with computer modelling and previously published data on protein binding were used to investigate the conformation of the RNA binding site. The experimental data confirm the model in which adenines A595, A640 and A642 bulge out in the deep groove. In addition to the already proposed non canonical U598-U641 interaction, the structure is stabilized by stacking interactions (between A595 and A640) and an array of hydrogen bonds involving bases and the sugar phosphate backbone. Mutations that alter the ability to form these interdependent interactions result in a local destabilization or reorganization. The specificity of recognition by protein S8 is provided by the irregular and distorted backbone and the two bulged adenines 640 and 642 in the deep groove. The third adenine (A595) is not a direct recognition site but must adopt a bulged position. The U598-U641 pair should not be directly in contact with the protein.}, note = {0305-1048 Journal Article}, keywords = {16S/*chemistry/*metabolism Ribosomal Proteins/*metabolism, Base Sequence Binding Sites Computer Simulation *Conserved Sequence Escherichia coli/metabolism Models, Molecular Molecular Sequence Data *Nucleic Acid Conformation Point Mutation/physiology RNA, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast serine isoacceptor tRNAs: variations of their content as a function of growth conditions and primary structure of the minor tRNA(Ser)GCU}, author = { T. Heyman and B. Agoutin and C. Fix and G. Dirheimer and G. Keith}, year = {1994}, date = {1994-01-01}, journal = {FEBS Lett}, volume = {347}, number = {2-3}, pages = {143-6}, abstract = {The primary structure of Saccharomyces cerevisiae tRNA(Ser)GCU is presented (EMBL database accession No. X74268 S. cerevisiae tRNA-Ser). In addition, quantitation of the relative amounts of serine isoaccepting tRNAs in yeast grown on different media showed that the minor tRNA(Ser)GCU decreased while the major tRNA(Ser)AGA increased as the growth rate and the cellular protein content increased. The minor species, tRNA(Ser)CGA and tRNA(Ser)UGA, were not separated by our gel system, however, taken together they appeared to vary in the same way as tRNA(Ser)GCU. These data suggest a growth rate dependence of yeast tRNAs similar to that previously described for E. coli tRNAs.}, note = {0014-5793 Journal Article}, keywords = {&, Acid, Anticodon, Base, cerevisiae/*genetics/*growth, Conformation, Culture, Data, development, Fungal/*chemistry, Galactose, Hybridization, Media, Molecular, Nucleic, Probes, RNA, Saccharomyces, Sequence, Ser/analysis/*chemistry, Transfer, Transfer/*chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Deficiency of queuine, a highly modified purine base, in transfer RNAs from primary and metastatic ovarian malignant tumors in women}, author = { W. Baranowski and G. Dirheimer and J. A. Jakowicki and G. Keith}, year = {1994}, date = {1994-01-01}, journal = {Cancer Res}, volume = {54}, number = {16}, pages = {4468-71}, abstract = {The tRNAs from rapidly growing tissues, particularly from neoplasia, often exhibit queuine deficiency. In order to check whether different kinds of ovarian tumors display queuine deficiencies we have analyzed tRNA samples from 16 ovarian malignancies. The tRNAs from histologically normal myometrium (4 samples) and myoma (6 samples) were taken as healthy tissue and benign tumor references. Queuine deficiency was determined by an exchange assay using [8-3H]guanine and tRNA:guanine transglycosylase from Escherichia coli. The mean values of queuine deficiencies in tRNAs were: 10.95 +/- 2.21 (SD) pmol/A260 in gonadal and germ cell tumors (5 cases); 23.75 +/- 7.89 pmol/A260 in primary epithelial tumors (9 cases); and 34.58 +/- 7.18 pmol/A260 in metastatic tumors (2 cases). These values displayed statistically significant differences (P = 0.0003, Kruskal-Wallis test). The queuine deficiencies in tRNAs significantly increased when moving from well-differentiated through moderately differentiated to poorly differentiated tumors, with the highest values found in poorly differentiated metastatic tumors (P = 0.0002, Kruskal-Wallis test). Queuine deficiency determination in tRNAs is proposed as a factor for clinical outcome prognosis of ovarian malignancies.}, note = {0008-5472 Journal Article}, keywords = {&, Adolescent, Adult, Aged, derivatives/analysis, Female, Gov't, Guanine/*analogs, Human, Middle, Neoplasm/*chemistry, Neoplasms/*chemistry/pathology, Non-U.S., Ovarian, RNA, Support, Transfer/*chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transfer RNA profiling: a new method for the identification of pathogenic Candida species}, author = { M. A. Santos and C. el-Adlouni and A. D. Cox and J. M. Luz and G. Keith and M. F. Tuite}, year = {1994}, date = {1994-01-01}, journal = {Yeast}, volume = {10}, number = {5}, pages = {625-36}, abstract = {A new molecular taxonomic method applicable to the identification of medically important Candida species and other yeast species has been developed. It is based on the electrophoretic pattern of total tRNA samples (a 'tRNA profile') isolated from Candida species and generated using high-resolution semi-denaturing urea-polyacrylamide gel electrophoresis and methylene blue staining. Species-specific tRNA profiles for the species C. albicans, C. tropicalis, C. parapsilosis, C. guilliermondii, C. glabrata and Pichia guilliermondii were obtained. Detailed studies with the major human pathogen of the Candida genus, C. albicans, demonstrated that the tRNA profile for a given species was both reproducible and strain-independent; seven different C. albicans strains generated identical tRNA profiles. Minor strain-specific heterogeneities in the tRNA profiles of C. guilliermondii and C. parapsilosis were detected, but in neither case did they significantly alter the species-specific diagnostic tRNA profile. The potential of this method in clarifying taxonomic anomalies was demonstrated by the finding that Type I and Type II strains of C. stellatoidea generate very different tRNA profiles, with that of a Type II strain being identical to the C. albicans tRNA profile. This method offers a number of advantages over current electrophoretic karyotype methods for species identification, both within the Candida genus and with yeast species in general.}, note = {0749-503x Journal Article}, keywords = {Candida/classification/*genetics/pathogenicity, Electrophoresis, Fungal, Gel, Genetic, Gov't, Markers, Non-U.S., Polyacrylamide, RNA, Support, Transfer/*analysis}, pubstate = {published}, tppubtype = {article} } @article{, title = {Mutations in helix 34 of Escherichia coli 16 S ribosomal RNA have multiple effects on ribosome function and synthesis}, author = { H. Moine and A. E. Dahlberg}, year = {1994}, date = {1994-01-01}, journal = {J Mol Biol}, volume = {243}, number = {3}, pages = {402-12}, abstract = {Helix 34 of E. coli 16 S rRNA (1046 to 1067 and 1189 to 1211) has been proposed to participate directly in the termination of translation at UGA stop codons. We have constructed mutations in this helix in plasmid-encoded rDNA to explore the specific functional roles of the sequence UCAUCA (1199 to 1204) and a secondary structure also involving positions 1054 and 1057-1058. The rRNA mutations were analyzed for their effects on in vivo translational accuracy (stop codon readthrough and frameshifting) as well as growth rate, ribosome synthesis and incorporation into polysomes. Mutations at positions 1054, 1057, 1058, 1199 and 1200 had significant effects on translational accuracy, causing non-specific readthrough of all three stop codons as well as enhanced +1 and -1 frameshifting. Mutations at 1202 and 1203, however, had no effect. The incorporation of deleterious mutant subunits into 70 S ribosomes and polysomes was severely reduced and was associated with a slower growth rate and increased synthesis of host-encoded ribosomes. These data support the proposal that helix 34 is an essential component of the decoding center of the 30 S ribosomal subunit and is not restricted in function to UGA-codon specific termination.}, note = {0022-2836 Journal Article}, keywords = {*Mutation, *Nucleic, *Translation, &, 16S/*chemistry/genetics, Acid, Base, beta-Galactosidase/genetics, Codon, coli/*genetics/growth, Conformation, Data, development, Escherichia, Genetic, Gov't, Molecular, Non-U.S., P.H.S., Ribosomal, Ribosomes/*metabolism, RNA, Sequence, Support, Terminator, U.S.}, pubstate = {published}, tppubtype = {article} } @article{, title = {Transfer RNA binding protein in the nucleus of Saccharomyces cerevisiae}, author = { M. L. Wilhelm and J. Reinbolt and J. Gangloff and G. Dirheimer and F. X. Wilhelm}, year = {1994}, date = {1994-01-01}, journal = {FEBS Lett}, volume = {349}, number = {2}, pages = {260-4}, abstract = {A yeast nuclear protein that binds to tRNA was identified using a RNA mobility shift assay. Northwestern blotting and N-terminal sequencing experiments indicate that this tRNA-binding protein is identical to zuotin which has previously been shown to bind to Z-DNA [(1992) EMBO J. 11, 3787-3796]. Labeled tRNA and poly(dG-m5dC) stabilized in the Z-DNA form identify the same protein on a Northwestern blot. In a gel retardation assay poly(dG-m5dC) in the Z-form strongly diminishes the binding of tRNA to zuotin. These studies establish that zuotin is able to bind to both tRNA and Z-DNA. Zuotin may be transiently associated with tRNA in the nucleus of yeast cells and play a role in its processing or transport to the cytoplasm.}, note = {0014-5793 Journal Article}, keywords = {*Saccharomyces, &, Acid, Amino, Cell, cerevisiae, cerevisiae/*metabolism, Chromatography, Data, DNA-Binding, DNA/metabolism, Fungal, Fungal/*isolation, high, liquid, Molecular, Nucleus/*metabolism, Pressure, Proteins, Proteins/genetics/*metabolism, purification, RNA, Saccharomyces, Sequence, Transfer/*isolation}, pubstate = {published}, tppubtype = {article} } @article{, title = {Yeast Ty1 retrotransposon: the minus-strand primer binding site and a cis-acting domain of the Ty1 RNA are both important for packaging of primer tRNA inside virus-like particles}, author = { M. Wilhelm and F. X. Wilhelm and G. Keith and B. Agoutin and T. Heyman}, year = {1994}, date = {1994-01-01}, journal = {Nucleic Acids Res}, volume = {22}, number = {22}, pages = {4560-5}, abstract = {Reverse transcription of the yeast retrotransposon Ty1 is primed by the cytoplasmic initiator methionine tRNA (tRNA(iMet)). The primer tRNA(iMet) is packaged inside virus-like particles (VLPs) and binds to a 10 nucleotides minus-strand primer binding site, the (-)PBS, complementary to its 3' acceptor stem. We have found that three short sequences of the Ty1 RNA (box 1, box 2.1 and box 2.2) located 3' to the (-)PBS are complementary to other regions of the primer tRNA(iMet) (T psi C and DHU stems and loops). Reconstitution of reverse transcription in vitro with T7 transcribed Ty1 RNA species and tRNA(iMet) purified from yeast cells shows that the boxes do not affect the efficiency of reverse transcription. Thus the role of the boxes on packaging of the primer tRNA(iMet) into the VLPs was investigated by analysing the level of tRNA(iMet) packaged into mutant VLPs. Specific nucleotide changes in the (-)PBS or in the boxes that do not change the protein coding sequence but disrupt the complementarity with the primer tRNA(iMet) within the VLPs. We propose that base pairing between the primer tRNA(iMet) and the Ty1 RNA is of major importance for tRNA(iMet) packaging into the VLPs. Moreover the intactness of the boxes is essential for retrotransposition as shown by the transposition defect of a Ty1 element harboring an intact (-)PBS and mutated boxes.}, note = {0305-1048 Journal Article}, keywords = {Acid, Amino, Base, Binding, cerevisiae/*genetics, Cloning, Data, Fungal/*genetics, Genetic, Gov't, Met/*genetics, Molecular, Mutation/physiology, Non-U.S., Retroelements/*genetics/physiology, Retroviridae/genetics, RNA, RNA/*genetics, Saccharomyces, Sequence, Sites, Support, Transcription, Transfer}, pubstate = {published}, tppubtype = {article} } @article{georgel_insect_1993, title = {Insect immunity: the diptericin promoter contains multiple functional regulatory sequences homologous to mammalian acute-phase response elements}, author = {Philippe Georgel and Marie Meister and Christine Kappler and Bruno Lemaitre and Jean-Marc Reichhart and Jules A Hoffmann}, doi = {10.1006/bbrc.1993.2508}, issn = {0006-291X}, year = {1993}, date = {1993-12-01}, journal = {Biochem. Biophys. Res. Commun.}, volume = {197}, number = {2}, pages = {508--517}, abstract = {We are using the diptericin gene as a model system to study the control of expression of the genes encoding antibacterial peptides during the Drosophila immune reaction. In order to investigate the putative regulatory regions in the diptericin promoter, we performed DNaseI footprinting experiments combined with gel-shift assays in two inducible systems: the larval fat body and a tumorous Drosophila blood cell line. Our results confirm the importance of kappa B-like elements previously described in the immune response of insects and reveal for the first time the involvement of other regions containing sequences homologous to mammalian acute-phase response elements.}, keywords = {Acute-Phase Proteins, Animals, Anti-Infective Agents, Base Sequence, Cell Line, Deoxyribonuclease I, DNA-Binding Proteins, Genetic, hoffmann, Insect Hormones, Insect Proteins, Larva, M3i, Mammals, NF-kappa B, Nucleic Acid, Oligonucleotide Probes, Polymerase Chain Reaction, Promoter Regions, Regulatory Sequences, reichhart}, pubstate = {published}, tppubtype = {article} } @article{bulet_novel_1993, title = {A novel inducible antibacterial peptide of Drosophila carries an O-glycosylated substitution}, author = {Philippe Bulet and Jean-Luc Dimarcq and Charles Hetru and Marie Lagueux and Maurice Charlet and G Hegy and Alan Van Dorsselaer and Jules A Hoffmann}, issn = {0021-9258}, year = {1993}, date = {1993-07-01}, journal = {J. Biol. Chem.}, volume = {268}, number = {20}, pages = {14893--14897}, abstract = {One of the facets of the host defense of higher insects is the rapid and transient synthesis, following bacterial challenge or trauma, of a battery of potent antibacterial peptides (Steiner, H., Hultmark, D., Engström, A., Bennich, H., and Boman, H. G. (1981) Nature 292, 246-248). The best characterized of these peptides are the cecropins (ibid.), 4-kDa peptides devoid of cysteines, and the insect defensins (Hoffmann, J. A., and Hetru, C. (1992) Immunol. Today 13, 411-415), 4-kDa peptides with three intramolecular disulfide bridges. Several other inducible antibacterial peptides have been characterized only at the level of their amino acid sequences (Hoffmann, J. A., Dimarcq, J. L., and Bulet, P. (1992) Médecine & Sciences 8, 432-439). We report here the isolation of a novel 19-residue proline-rich inducible antibacterial peptide from Drosophila. In contrast to all previous reports on antibacterial peptides, this molecule carries a substitution as evidenced by molecular mass determinations; our data show that this reflects the O-glycosylation of a Thr residue by an N-acetylgalactosamine plus a galactose. A synthetic nonsubstituted peptide of identical amino acid sequence has an activity several times lower (5-10) than the native compound. Our data suggest that this substitution represents a post-translational modification essential for the full biological activity of this novel peptide.}, keywords = {Animals, Anti-Bacterial Agents, Base Sequence, Carbohydrates, Cloning, DNA, Escherichia coli, Gas Chromatography-Mass Spectrometry, Glycopeptides, Glycosylation, hoffmann, M3i, Molecular}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_humoral_1993, title = {The humoral antibacterial response of Drosophila}, author = {Jules A Hoffmann and Charles Hetru and Jean-Marc Reichhart}, issn = {0014-5793}, year = {1993}, date = {1993-06-01}, journal = {FEBS Lett.}, volume = {325}, number = {1-2}, pages = {63--66}, abstract = {Drosophila, like other insects, responds to the injection of bacteria by the rapid and transient synthesis of a battery of potent antibacterial peptides. Only a few of these peptides have been fully characterized to date. We review our recent data on the control of the expression of a gene encoding one of the induced peptides, i.e. diptericin. Our data highlight the role of proximal cis-regulatory motifs similar to regulatory elements binding NF-kappa B and NF-IL6 in promoters of some immune genes of mammals. We argue that the Drosophila host defense is homologous to the mammalian acute phase response.}, keywords = {Animals, Anti-Bacterial Agents, Base Sequence, Genes, hoffmann, Insect, Insect Hormones, Insect Proteins, M3i, reichhart, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{kappler_insect_1993, title = {Insect immunity. Two 17 bp repeats nesting a kappa B-related sequence confer inducibility to the diptericin gene and bind a polypeptide in bacteria-challenged Drosophila}, author = {Christine Kappler and Marie Meister and Marie Lagueux and E Gateff and Jules A Hoffmann and Jean-Marc Reichhart}, issn = {0261-4189}, year = {1993}, date = {1993-04-01}, journal = {EMBO J.}, volume = {12}, number = {4}, pages = {1561--1568}, abstract = {The Drosophila diptericin gene codes for a 9 kDa antibacterial peptide and is rapidly and transiently expressed in larvae and adults after bacterial challenge. It is also induced in a tumorous Drosophila blood cell line by the addition of lipopolysaccharide (LPS). The promoter of this gene contains two 17 bp repeats located closely upstream of the TATA-box and harbouring a decameric kappa B-related sequence. This study reports that the replacement of the two 17 bp repeats by random sequences abolishes bacteria inducibility in transgenic fly lines. In transfected tumorous blood cells, the replacement of both or either of the 17 bp motifs reduces dramatically LPS inducibility, whereas multiple copies significantly increase the level of transcriptional activation by LPS challenge. A specific DNA-protein binding activity is evidenced in cytoplasmic and nuclear extracts of induced blood cells and fat body. It is absent in controls. It is proposed that induction of the diptericin gene mediated by the two 17 bp repeats occurs via a mechanism similar to that of mammalian NF-kappa B.}, keywords = {Animals, Anti-Bacterial Agents, Base Sequence, Cloning, Gene Expression Regulation, Genes, Genetic, Genetically Modified, hoffmann, Insect, Insect Hormones, Insect Proteins, Lipopolysaccharides, M3i, messenger, Molecular, NF-kappa B, Nucleic Acid, Oligodeoxyribonucleotides, Promoter Regions, Regulatory Sequences, reichhart, RNA, Transfection}, pubstate = {published}, tppubtype = {article} } @article{reichhart_expression_1993, title = {Expression and nuclear translocation of the rel/NF-kappa B-related morphogen dorsal during the immune response of Drosophila}, author = {Jean-Marc Reichhart and Philippe Georgel and Marie Meister and Bruno Lemaitre and Christine Kappler and Jules A Hoffmann}, issn = {0764-4469}, year = {1993}, date = {1993-01-01}, journal = {C. R. Acad. Sci. III, Sci. Vie}, volume = {316}, number = {10}, pages = {1218--1224}, abstract = {The rel/NF-kappa B-related morphogen dorsal is a maternally expressed gene which is involved in the control of the dorso-ventral axis during early embryogenesis of Drosophila. We show that this gene is also expressed in the fat body of larvae and adults of Drosophila as well as in a tumorous blood cell line: its expression is noticeably enhanced upon bacterial (or lipopolysaccharide) challenge. This challenge also induces within 15-30 min a nuclear translocation of the dorsal protein. The genes encoding inducible antibacterial peptides in Drosophila contain kappa B-related nucleotide sequences and we show that the dorsal protein can bind to such motifs and sequence-specifically transactivate a reporter gene in co-transfection experiments with a Drosophila cell line. However, in dl1 mutants, in the absence of dorsal protein, the genes encoding antibacterial peptides retain their inducibility, suggesting a multifactorial control. The results indicate that in addition to its role in embryogenesis, dorsal is involved in the immune response of Drosophila. They also strengthen the analogy between the mammalian acute phase response and the insect immune response.}, keywords = {Animals, Blotting, Cellular, Gene Expression, Genes, Genetic, hoffmann, Immunity, Insect, M3i, NF-kappa B, Northern, reichhart, translocation, Zygote}, pubstate = {published}, tppubtype = {article} } @article{, title = {Unusual structure of the double-stranded RNA associated with the '447' cytoplasmic male sterility in Vicia faba}, author = { P. Pfeiffer and J. L. Jung and J. Heitzler and G. Keith}, year = {1993}, date = {1993-01-01}, journal = {J Gen Virol}, volume = {74}, number = {Pt 6}, pages = {1167-73}, abstract = {The 16.7 kbp dsRNA specific to the '447' cytoplasmic male sterility (CMS) line of Vicia faba was labelled in vitro with [alpha-32P]ATP and poly(A) polymerase, and by T4 RNA ligase-mediated addition of [32P]pCp. Analysis of the reaction products under denaturing conditions revealed in both cases extensive labelling of a 4.5 kb ssRNA, already detected in previous experiments in which the RNA-dependent RNA polymerase associated with the dsRNA was allowed to pursue RNA synthesis on preinitiated complexes. Mobility shift analysis of total pCp-labelled dsRNA revealed not two but three different 3' termini. The most prominent sequencing pattern corresponded to the 4.5 kb ssRNA, indicating that this RNA species has a preferentially accessible, free 3' OH extremity. Northern blot analysis of the denatured dsRNA confirmed that the 4.5 kb ssRNA is a subgenomic mRNA and detected its counterpart of about 12 kb. Nearly all 16.7 kbp dsRNA molecules featured an interrupted positive-sense strand, indicating a marked prevalence of transcription over replication complexes. This unusual strategy of transcription by a strand displacement mechanism, following initiation at an internal discontinuity, is compared with that of other dsRNA viruses or defective viruses, and is discussed in relation to the expression of the CMS trait.}, note = {0022-1317 Journal Article}, keywords = {*Plants, &, Base, Bodies, Cytoplasm, Data, Extrachromosomal, Fabaceae/*genetics, Genetic, Inclusion, Infertility/genetics, Inheritance/*genetics, Medicinal, Molecular, Pollen/genetics, purification, Replicase/metabolism, RNA, RNA/*genetics/isolation, Sequence, Transcription, Viral}, pubstate = {published}, tppubtype = {article} } @article{, title = {A very poorly expressed tRNA(Ser) is highly concentrated together with replication primer initiator tRNA(Met) in the yeast Ty1 virus-like particles}, author = { P. Pochart and B. Agoutin and C. Fix and G. Keith and T. Heyman}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {7}, pages = {1517-21}, abstract = {The analysis of the tRNAs associated to the virus-like particles produced by the Ty1 element revealed the specific packaging of three major tRNA species, in about equal amounts: the replication primer initiator tRNA(Met), the tRNA(Ser)AGA and a tRNA undetected until now as an expressed species in yeast. The latter tRNA is coded by the already described tDNA(Ser)GCT. This tRNA is enriched more than 150 fold in the particles as compared to its content in total cellular tRNA where it represents less than 0.1% (initiator tRNA(Met) and tRNA(Ser)AGA being 11 and 4 fold enriched respectively). This tRNA is the only species coded by the tDNA(Ser)GCT gene which is found in three copies per genome since no other corresponding expressed tRNA could be detected. This gene is thus very poorly expressed. The high concentration of tRNA(Ser)GCU in the particles compared to its very low cellular content led us to consider its possible implication in Ty specific processes.}, note = {0305-1048 Journal Article}, keywords = {&, Acid, Base, cerevisiae/metabolism, Conformation, Data, development, DNA, Electrophoresis, Elements/*physiology, Gel, Met/metabolism, Molecular, Nucleic, Retroviridae/*growth, RNA, Saccharomyces, Sequence, Ser/*metabolism, Transfer, Transposable, Two-Dimensional, Viral/*metabolism}, pubstate = {published}, tppubtype = {article} } @article{, title = {High levels of 2',5'-oligoadenylate synthetase and 2',5'-oligoadenylate-dependent endonuclease in human trophoblast}, author = {G Y Zhang and B Beltchev and A Fournier and Y H Zhang and A Malassine and C Bisbal and B Ehresmann and C Ehresmann and J L Darlix and M N Thang}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8457385}, isbn = {8457385}, year = {1993}, date = {1993-01-01}, journal = {AIDS Res Hum Retroviruses}, volume = {9}, number = {2}, pages = {189-196}, abstract = {Human placenta contains a high level of 2',5'-oligoadenylate (2-5A) synthetase activity of the 100-kD form of the enzyme. About 20% of the placental 2-5A synthetase activity was found to be cytosolic, whereas the remaining 80% was released by 0.5 M KCl in the presence of detergent. Most of the enzyme activity was localized in trophoblast cells, which also contain a high level of 2-5A-dependent RNase L activity. The purified trophoblast 100-kD 2-5A synthetase was shown to be activated by human immunodeficiency virus type 1 (HIV-1) 5' RNA 1-311 and 1-707, which both contain the TAR and primer binding site (PBS) structured regions. These two HIV-1 RNAs activated human trophoblast 2-5A synthetase at the same level as poly(I).poly (C), a standard highly efficient activator of the enzyme, and at the same optimal concentration. On the contrary, HIV-1 RNA 311-618, a poorly structured region missing TAR and PBS, was shown to be a poor activator of the enzyme. The specific cellular location of the 2-5A synthetase and its efficient activation by HIV 5' RNA favors the idea that the trophoblast 2-5A system negatively controls HIV replication in trophoblasts.}, note = {0889-2229 Journal Article}, keywords = {2', 5'-Oligoadenylate Synthetase/*metabolism Endoribonucleases/*metabolism Enzyme Activation/drug effects Female HIV Infections/transmission HIV-1/*physiology Human In Vitro Maternal-Fetal Exchange Pregnancy RNA, Non-U.S. Gov't Trophoblasts/*enzymology/*microbiology Virus Replication, Unité ARN, Viral/pharmacology Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Identity elements of tRNA(Trp). Identification and evolutionary conservation}, author = {H Xue and W Shen and R Giege and J T Wong}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8486627}, isbn = {8486627}, year = {1993}, date = {1993-01-01}, journal = {J Biol Chem}, volume = {268}, number = {13}, pages = {9316-9322}, abstract = {In this study, the varying reactivities of Bacillus subtilis tryptophanyl-tRNA synthetase toward prokaryotic, eukaryotic, and halophile tRNAs were employed to define the potential identity elements on tRNA(Trp). On this basis mutagenesis was performed to obtain, through in vivo heterologous expression in Escherichia coli and in vitro transcription with T7 RNA polymerase, mutant B. subtilis tRNA(Trp) for comparison with the wild-type. These comparisons served to establish G73 and the anticodon as major identity elements, and A1-U72, G5-C68, and A9 as minor identity elements. While the tryptophanyl-tRNA synthetase from B. subtilis and E. coli require G73 to function, replacement of G73 by A73 favors the enzyme from yeast. This change points to the variation of the identity elements for the same amino acid among different organisms. The similarity in these elements between B. subtilis and E. coli tryptophanyl-tRNA synthetase, however, suggests that identity elements on tRNA, like the active centers on enzymes, undergo evolutionary change at slower rates than less essential portions of the macromolecule.}, note = {0021-9258 Journal Article}, keywords = {Animals Bacillus subtilis/*genetics Base Sequence Cattle Cloning, Bacterial Halobacterium/genetics Kinetics Liver/physiology Molecular Sequence Data Nucleic Acid Conformation Nucleic Acid Denaturation RNA, Molecular Comparative Study Escherichia coli/*genetics *Evolution Genes, Non-U.S. Gov't Triticum/genetics Tryptophan-tRNA Ligase/metabolism, Nucleic Acid Support, Structural, Transfer, Trp/chemistry/*genetics/metabolism Saccharomyces cerevisiae/genetics Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Nuclease footprinting of human immunodeficiency virus reverse transcriptase/tRNA(Lys-3) complexes}, author = {B M Wohrl and B Ehresmann and G Keith and S F Le Grice}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7685766}, isbn = {7685766}, year = {1993}, date = {1993-01-01}, journal = {J Biol Chem}, volume = {268}, number = {18}, pages = {13617-13624}, abstract = {Nuclease footprinting has been used to probe features of binary complexes of type 1 human immunodeficiency virus reverse transcriptase (HIV-1 RT) with both natural and synthetic preparations of its cognate replication primer, tRNA(Lys-3). In addition to heterodimeric RT (p66/p51), ribonucleoprotein complexes containing either the p66 or p51 subunit were analyzed. Footprinting experiments employed both structure- and sequence-specific nucleases. Our results indicate a similar mode of interaction for the three RT preparations tested, suggesting contact with each loop of the tRNA primer (D, anticodon, and T psi C), as well as minor perturbation of the anticodon stem. Although there is little evidence for extensive disruption of the 3'-acceptor stem. RNase A footprinting data with natural and synthetic tRNA suggests that potential base pairing between the T psi C and D loops is disrupted in the presence of RT.}, note = {0021-9258 Journal Article}, keywords = {Anticodon Base Sequence HIV-1/*enzymology HIV-1 Reverse Transcriptase Hydrolysis Molecular Sequence Data Nucleic Acid Conformation RNA, Double-Stranded/metabolism RNA, Lys/chemistry/*metabolism RNA-Directed DNA Polymerase/*metabolism Recombinant Proteins/metabolism Ribonuclease, Non-U.S. Gov't Support, P.H.S., Pancreatic/metabolism Support, Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Unique secondary and tertiary structural features of the eucaryotic selenocysteine tRNA(Sec)}, author = {C Sturchler and E Westhof and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8464694}, isbn = {8464694}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {5}, pages = {1073-1079}, abstract = {Cotranslational insertion of selenocysteine into selenoenzymes is mediated by a specialized transfer RNA, the tRNA(Sec). We have carried out the determination of the solution structure of the eucaryotic tRNA(Sec). Based on the enzymatic and chemical probing approach, we show that the secondary structure bears a few unprecedented features like a 9 bp aminoacid-, a 4 bp thymine- and a 6 bp dihydrouridine-stems. Surprisingly, the eighth nucleotide, although being a uridine, is base-paired and cannot therefore correspond to the single-stranded invariant U8 found in all tRNAs. Rather, experimental evidence led us to propose that the role of the invariant U8 is actually played by the tenth nucleotide which is an A, numbered A8 to indicate this fact. The experimental data therefore demonstrate that the cloverleaf structure we derived experimentally resembles the hand-folded model proposed by Bock et al (ref. 3). Using the solution data and computer modelling, we derived a three-dimensional structure model which shows some unique aspects. Basically, A8, A14, U21 form a novel type of tertiary interaction in which A8 interacts with the Hoogsteen sites of A14 which itself forms a Watson-Crick pair with U21. No coherent model containing the canonical 15-48 interaction could be derived. Thus, the number of tertiary interactions appear to be limited, leading to an uncoupling of the variable stem from the rest of the molecule.}, note = {0305-1048 Journal Article}, keywords = {Amino Acid-Specific/*chemistry *Selenocysteine Support, Animals Base Sequence Cloning, Molecular Computer Simulation DNA, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA, Non-U.S. Gov't Xenopus laevis, Single-Stranded Models, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Synthesis and ribosome binding properties of model mRNAs modified with undecagold cluster}, author = {E Skripkin and G Yusupova and M Yusupov and P Kessler and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8305524}, isbn = {8305524}, year = {1993}, date = {1993-01-01}, journal = {Bioconjug Chem}, volume = {4}, number = {6}, pages = {549-553}, abstract = {The synthesis and purification of short model messenger RNAs modified with undecagold cluster are described. A monoamino undecagold cluster was introduced on the oxidized 3' cis-glycol group of the mRNA followed by reduction of the formed Schiff's base. The stability of the modified mRNA under the conditions used for in vitro messenger RNA translation is studied. The possibility of the formation of a specific translational initiation complex with bacterial ribosomes and modified mRNAs is shown. The results of these experiments indicate that the attachment of an undecagold cluster to a mRNA is a useful tool for electron microscopic and crystallographic studies.}, note = {1043-1802 Journal Article}, keywords = {Base Sequence Comparative Study Escherichia coli/metabolism Gold/metabolism Models, Genetic/drug effects/genetics, Messenger/*chemical synthesis/chemistry/*metabolism RNA, Met/metabolism Ribosomes/*metabolism Support, Molecular Molecular Sequence Data Organometallic Compounds/*chemical synthesis/chemistry/*metabolism RNA, Non-U.S. Gov't Translation, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {An operational RNA code for amino acids and possible relationship to genetic code}, author = {P Schimmel and R Giege and D Moras and S Yokoyama}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7692438}, isbn = {7692438}, year = {1993}, date = {1993-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {90}, number = {19}, pages = {8763-8768}, abstract = {RNA helical oligonucleotides that recapitulate the acceptor stems of transfer RNAs, and that are devoid of the anticodon trinucleotides of the genetic code, are aminoacylated by aminoacyl tRNA synthetases. The specificity of aminoacylation is sequence dependent, and both specificity and efficiency are generally determined by only a few nucleotides proximal to the amino acid attachment site. This sequence/structure-dependent aminoacylation of RNA oligonucleotides constitutes an operational RNA code for amino acids. To a rough approximation, members of the two different classes of tRNA synthetases are, like tRNAs, organized into two major domains. The class-defining conserved domain containing the active site incorporates determinants for recognition of RNA mini-helix substrates. This domain may reflect the primordial synthetase, which was needed for expression of the operational RNA code. The second synthetase domain, which generally is less or not conserved, provides for interactions with the second domain of tRNA, which incorporates the anticodon. The emergence of the genetic from the operational RNA code could occur when the second domain of synthetases was added with the anticodon-containing domain of tRNAs.}, note = {0027-8424 Journal Article Review Review, Tutorial}, keywords = {*Amino Acid Sequence Amino Acids/*metabolism Amino Acyl-tRNA Ligases/metabolism Base Sequence Conserved Sequence Escherichia coli/enzymology/genetics *Genetic Code Nucleic Acid Conformation Oligoribonucleotides RNA/*genetics RNA, Non-U.S. Gov't Support, P.H.S., Transfer/*genetics/metabolism Support, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Non-standard translational events in Candida albicans mediated by an unusual seryl-tRNA with a 5'-CAG-3' (leucine) anticodon}, author = {M A Santos and G Keith and M F Tuite}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8440250}, isbn = {8440250}, year = {1993}, date = {1993-01-01}, journal = {EMBO J}, volume = {12}, number = {2}, pages = {607-616}, abstract = {From in vitro translation studies we have previously demonstrated the existence of an apparent efficient UAG (amber) suppressor tRNA in the dimorphic fungus Candida albicans (Santos et al., 1990). Using an in vitro assay for termination codon readthrough the tRNA responsible was purified to homogeneity from C.albicans cells. The determined sequence of the purified tRNA predicts a 5'-CAG-3' anticodon that should decode the leucine codon CUG and not the UAG termination codon as originally hypothesized. However, the tRNA(CAG) sequence shows greater nucleotide homology with seryl-tRNAs from the closely related yeast Saccharomyces cerevisiae than with leucyl-tRNAs from the same species. In vitro tRNA-charging studies demonstrated that the purified tRNA(CAG) is charged with Ser. The gene encoding the tRNA was cloned from C.albicans by a PCR-based strategy and DNA sequence analysis confirmed both the structure of the tRNA(CAG) and the absence of any introns in the tRNA gene. The copy number of the tRNA(CAG) gene (1-2 genes per haploid genome) is in agreement with the relatively low abundance (< 0.5% total tRNA) of this tRNA. In vitro translation studies revealed that the purified tRNA(CAG) could induce apparent translational bypass of all three termination codons. However, peptide mapping of in vitro translation products demonstrated that the tRNA(CAG) induces translational misreading in the amino-terminal region of two RNA templates employed, namely the rabbit alpha- and beta-globin mRNAs. These results suggest that the C.albicans tRNA(CAG) is not an 'omnipotent' suppressor tRNA but rather may mediate a novel non-standard translational event in vitro during the translation of the CUG codon. The possible nature of this non-standard translation event is discussed in the context of both the unusual structural features of the tRNA(CAG) and its in vitro behaviour.}, note = {0261-4189 Journal Article}, keywords = {*Anticodon Base Sequence Candida albicans/*genetics Cloning, Fungal Genes, Fungal Leucine/*genetics Molecular Sequence Data Nucleic Acid Conformation RNA, Fungal/chemistry/genetics/isolation & purification RNA, Genetic, Molecular DNA, Non-U.S. Gov't *Translation, Ser/chemistry/*genetics/isolation & purification Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Kinetic properties of pure overproduced Bacillus subtilis phenylalanyl-tRNA synthetase do not favour its in vivo inhibition by ochratoxin A}, author = {A Roth and G Eriani and G Dirheimer and J Gangloff}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8325392}, isbn = {8325392}, year = {1993}, date = {1993-01-01}, journal = {FEBS Lett}, volume = {326}, number = {1-3}, pages = {87-91}, abstract = {Ochratoxine A (OTA) inhibits growth of Bacillus subtilis at pHs below 7. Since OTA is a phenylalanine analogue, this effect could be due to inhibition of phenylalanine-tRNA synthetase (PheRS) by competition of this mycotoxin with the amino acid. Homogeneous PheRS was purified from Bacillus subtilis and from E. coli transformed with the PheRS gene. The latter produced about 40 times more PheRS than B. subtilis. The Km and Ki values of PheRS, respectively, for phenylalanine and OTA were measured and their concentrations within the cell determined. It appears that the concentration of OTA in the cell, in spite of a 25-fold accumulation, remained too low to significantly compete with phenylalanine. This does not suggest PheRS to be the target of OTA in cell growth and protein synthesis inhibition in Bacillus subtilis. It was also shown that the 2-3-fold increase of PheRS in OTA-treated cells is not due to phenylalanine-controlled attenuation regulation.}, note = {0014-5793 Journal Article}, keywords = {Bacillus subtilis/drug effects/*enzymology Binding, Bacterial, Competitive Escherichia coli/enzymology/genetics Kinetics Macromolecular Systems Ochratoxins/*pharmacology Phenylalanine/metabolism Phenylalanine-tRNA Ligase/antagonists & inhibitors/*metabolism Support, ERIANI, Non-U.S. Gov't Transformation, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The complete amino acid sequence of ribosomal protein S8 from Thermus thermophilus}, author = {J Reinbolt and I Eliseikina and S Sedelnilkova and M Garber and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8427638}, isbn = {8427638}, year = {1993}, date = {1993-01-01}, journal = {J Protein Chem}, volume = {12}, number = {1}, pages = {79-83}, abstract = {Protein S8 from Thermus thermophilus consists of 138 amino acids of M(r) 15,840. Its primary structure was established using peptide sequences from two different digests. Protein S8 from T. thermophilus shares a high percentage of identity with protein S8 from Thermus aquaticus. There are some consensus sequences between proteins S8 from eubacteria, archebacteria, chloroplasts, and cyanelles.}, note = {0277-8033 Journal Article}, keywords = {Amino Acid Serine Endopeptidases/chemistry Thermus/chemistry Thermus thermophilus/*chemistry, Amino Acid Sequence Hydrolysis Metalloendopeptidases/chemistry Molecular Sequence Data Ribosomal Proteins/*chemistry Sequence Homology, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Additive, cooperative and anti-cooperative effects between identity nucleotides of a tRNA}, author = {J Putz and J D Puglisi and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8335008}, isbn = {8335008}, year = {1993}, date = {1993-01-01}, journal = {EMBO J}, volume = {12}, number = {7}, pages = {2949-2957}, abstract = {We have investigated the functional relationship between nucleotides in yeast tRNAAsp that are important for aspartylation by yeast aspartyl-tRNA synthetase. Transcripts of tRNAAsp with two or more mutations at identity positions G73, G34, U35, C36 and base pair G10-U25 have been prepared and the steady-state kinetics of their aspartylation were measured. Multiple mutations affect the catalytic activities of the synthetase mainly at the level of the catalytic constant, kcat. Kinetic data were expressed as free energy variation at transition state of these multiple mutants and comparison of experimental values with those calculated from results on single mutants defined three types of relationships between the identity nucleotides of this tRNA. Nucleotides located far apart in the three-dimensional structure of the tRNA act cooperatively whereas nucleotides of the anticodon triplet act either additively or anti-cooperatively. These results are related to the specific interactions of functional groups on identity nucleotides with amino acids in the protein as revealed by the crystal structure of the tRNAAsp/aspartyl-tRNA synthetase complex. These relationships between identity nucleotides may play an important role in the biological function of tRNAs.}, note = {0261-4189 Journal Article}, keywords = {Acylation Aspartate-tRNA Ligase/metabolism Aspartic Acid/chemistry/genetics Base Sequence Catalysis Kinetics Molecular Sequence Data Mutation Nucleic Acid Conformation Nucleotides/*metabolism RNA, Asp/chemistry/genetics/*metabolism Saccharomyces cerevisiae/enzymology Support, FLORENTZ, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Rye nuclease I as a tool for structural studies of tRNAs with large variable arms}, author = { C. el Adlouni and G. Keith and G. Dirheimer and J. W. Szarkowski and A. Przykorska}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {4}, pages = {941-7}, abstract = {A single-strand-specific nuclease from rye germ (Rn nuclease I) was used for secondary and tertiary structure investigations of tRNAs with large variable arms (class II tRNAs). We have studied the structure in solution of two recently sequenced tRNA(Leu): yeast tRNA(Leu)(ncm5UmAA) and bovine tRNA(Leu)(XmAA) as well as yeast tRNA(Leu)(UAG), tRNA(Leu)(m5CAA) and tRNA(Ser)(IGA). The latter is the only tRNA with a long variable arm for which the secondary and tertiary structure has already been studied by use of chemical probes and computer modelling. The data obtained in this work showed that the general model of class II tRNAs proposed by others for tRNA(Ser) can be extended to tRNAs(Leu) as well. However interesting differences in the structure of tRNAs(Leu) versus tRNA(Ser)(IGA) were also noticed. The main difference was observed in the accessibility of the variable loops to nucleolytic attack of Rn nuclease I: variable loops of all studied tRNA(Leu) species were cut by Rn nuclease I, while that of yeast tRNA(Ser)(IGA) was not. This could be due to differences in stability of the variable arms and the lengths of their loops which are 3 and 4 nucleotides in tRNA(Ser)(IGA) and tRNAs(Leu) respectively.}, note = {0305-1048 Journal Article}, keywords = {*Nucleotidases, Acid, Animals, Anticodon, Base, Cattle, cereale/*enzymology, cerevisiae, Conformation, Data, Gov't, Leu/chemistry, Molecular, Non-U.S., Nucleic, RNA, Saccharomyces, Secale, Sequence, Ser/chemistry, Support, Transfer, Transfer/*chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Influence of tRNA tertiary structure and stability on aminoacylation by yeast aspartyl-tRNA synthetase}, author = {J D Puglisi and J Putz and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8441619}, isbn = {8441619}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {1}, pages = {41-49}, abstract = {Mutations have been designed that disrupt the tertiary structure of yeast tRNA(Asp). The effects of these mutations on both tRNA structure and specific aspartylation by yeast aspartyl-tRNA synthetase were assayed. Mutations that disrupt tertiary interactions involving the D-stem or D-loop result in destabilization of the base-pairing in the D-stem, as monitored by nuclease digestion and chemical modification studies. These mutations also decrease the specificity constant (kcat/Km) for aspartylation by aspartyl-tRNA synthetase up to 10(3)-10(4) fold. The size of the T-loop also influences tRNA(Asp) structure and function; change of its T-loop to a tetraloop (-UUCG-) sequence results in a denatured D-stem and an almost 10(4) fold decrease of kcat/Km for aspartylation. The negative effects of these mutations on aspartylation activity are significantly alleviated by additional mutations that stabilize the D-stem. These results indicate that a critical role of tertiary structure in tRNA(Asp) for aspartylation is the maintenance of a base-paired D-stem.}, note = {0305-1048 Journal Article}, keywords = {Acylation Aspartate-tRNA Ligase/*metabolism Base Sequence Kinetics Molecular Sequence Data Mutation *Nucleic Acid Conformation RNA, FLORENTZ, Non-U.S. Gov't Temperature, Transfer/*chemistry/metabolism Saccharomyces cerevisiae/enzymology Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence, overproduction and crystallization of aspartyl-tRNA synthetase from Thermus thermophilus. Implications for the structure of prokaryotic aspartyl-tRNA synthetases}, author = {A Poterszman and P Plateau and D Moras and S Blanquet and M H Mazauric and R Kreutzer and D Kern}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8319804}, isbn = {8319804}, year = {1993}, date = {1993-01-01}, journal = {FEBS Lett}, volume = {325}, number = {3}, pages = {183-186}, abstract = {The genes of aspartyl-tRNA synthetase (AspRS) from two Thermus thermophilus strain VK-1 and HB8, have been cloned and sequenced. Their nucleotidic sequences code for the same protein which displays the three characteristic motifs of class II aminoacyl-tRNA synthetases. This enzyme shows 50% identity with Escherichia coli AspRS, over the totality of the chain (580 amino acids). A comparison with the eukaryotic yeast cytoplasmic AspRS indicates the presence in the prokaryotic AspRS of an extra domain between motifs 2 and 3 much larger than in the eukaryotic ones. When its gene is under the control of the tac promoter of the expression vector pKK223-3, the protein is efficiently overexpressed as a thermostable protein in E. coli. It can be further purified to homogeneity using a heat treatment followed by a single anion exchange chromatography. Single crystals of the pure protein, diffracting at least to 2.2 A resolution (space group P2(1)2(1)2(1)}, note = {0014-5793 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Aspartate-tRNA Ligase/chemistry/*genetics/metabolism Base Sequence Cloning, Bacterial Models, Molecular Crystallization Genes, Molecular Molecular Sequence Data Oligodeoxyribonucleotides Sequence Homology, Non-U.S. Gov't Thermus thermophilus/*enzymology/genetics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Synthetic RNA-cleaving molecules mimicking ribonuclease A active center. Design and cleavage of tRNA transcripts}, author = {M A Podyminogin and V V Vlassov and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7507235}, isbn = {7507235}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {25}, pages = {5950-5956}, abstract = {RNA cleaving molecules were synthesized by conjugating imidazole residues imitating the essential imidazoles in the active center of pancreatic ribonuclease to an intercalating compound, derivative of phenazine capable of binding to the double stranded regions of polynucleotides. Action of the molecules on tRNA was investigated. It was found, that some of the compounds bearing two imidazole residues cleave tRNA under physiological conditions. The cleavage reaction shows a bell-shaped pH dependence with a maximum at pH 7.0 indicating participation of protonated and non-protonated imidazole residues in the process. Under the conditions stabilizing the tRNA structure, a tRNAAsp transcript was cleaved preferentially at the junctions of the stem and loop regions of the cloverleaf tRNA fold, at the five positions C56, C43, C20.1, U13, and U8, with a marked preference for C56. This cleavage pattern is consistent with a hydrolysis mechanism involving non-covalent binding of the compounds to the double-stranded regions of tRNA followed by an attack of the imidazole residues at the juxtaposed flexible single-stranded regions of the molecule. The compounds provide new probes for the investigation of RNA structure in solution and potential reactive groups for antisense oligonucleotide derivatives.}, note = {0305-1048 Journal Article}, keywords = {Asp/*metabolism Ribonuclease, Base Sequence Binding Sites Hydrogen-Ion Concentration Molecular Sequence Data Nucleic Acid Conformation RNA/metabolism RNA, Genetic, Non-U.S. Gov't Temperature Transcription, Pancreatic/antagonists & inhibitors/chemical synthesis/*metabolism Substrate Specificity Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A very poorly expressed tRNA(Ser) is highly concentrated together with replication primer initiator tRNA(Met) in the yeast Ty1 virus-like particles}, author = {P Pochart and B Agoutin and C Fix and G Keith and T Heyman}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8386834}, isbn = {8386834}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {7}, pages = {1517-1521}, abstract = {The analysis of the tRNAs associated to the virus-like particles produced by the Ty1 element revealed the specific packaging of three major tRNA species, in about equal amounts: the replication primer initiator tRNA(Met), the tRNA(Ser)AGA and a tRNA undetected until now as an expressed species in yeast. The latter tRNA is coded by the already described tDNA(Ser)GCT. This tRNA is enriched more than 150 fold in the particles as compared to its content in total cellular tRNA where it represents less than 0.1% (initiator tRNA(Met) and tRNA(Ser)AGA being 11 and 4 fold enriched respectively). This tRNA is the only species coded by the tDNA(Ser)GCT gene which is found in three copies per genome since no other corresponding expressed tRNA could be detected. This gene is thus very poorly expressed. The high concentration of tRNA(Ser)GCU in the particles compared to its very low cellular content led us to consider its possible implication in Ty specific processes.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence DNA Transposable Elements/*physiology Electrophoresis, Gel, Met/metabolism RNA, Ser/*metabolism RNA, Transfer, Two-Dimensional Molecular Sequence Data Nucleic Acid Conformation RNA, Unité ARN, Viral/*metabolism Retroviridae/*growth & development Saccharomyces cerevisiae/metabolism}, pubstate = {published}, tppubtype = {article} } @article{, title = {Ribosomal protein S15 from Escherichia coli modulates its own translation by trapping the ribosome on the mRNA initiation loading site}, author = {C Philippe and F Eyermann and L Benard and C Portier and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7685101}, isbn = {7685101}, year = {1993}, date = {1993-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {90}, number = {10}, pages = {4394-4398}, abstract = {From genetic and biochemical evidence, we previously proposed that S15 inhibits its own translation by binding to its mRNA in a region overlapping the ribosome loading site. This binding was postulated to stabilize a pseudoknot structure that exists in equilibrium with two stem-loops. Here, we use "toeprint" experiments with Moloney murine leukemia virus reverse transcriptase to analyze the effect of S15 on the formation of the ternary mRNA-30S-tRNA(fMet) complex. We show that the binding of the 30S subunit on the mRNA stops reverse transcriptase near position +10, corresponding to the 3' terminus of the pseudoknot, most likely by stabilizing the pseudoknot conformation. Furthermore, S15 is found to stabilize the binary 30S-mRNA complex. When the ternary 30S-mRNA-tRNA(fMet) complex is formed, a toeprint is observed at position +17. This toeprint progressively disappears when the ternary complex is formed in the presence of increasing concentrations of S15, while a shift from position +17 to position +10 is observed. Beside, RNase T1 footprinting experiments reveal the simultaneous binding of S15 and 30S subunit on the mRNA. Otherwise, we show by filter binding assays that initiator tRNA remains bound to the 30S subunit even in the presence of S15. Our results indicate that S15 prevents the formation of a functional ternary 30S-mRNA-tRNA(fMet) complex, the ribosome being trapped in a preternary 30S-mRNA-tRNA(fMet) complex.}, note = {0027-8424 Journal Article}, keywords = {Bacterial Molecular Sequence Data Nucleic Acid Conformation Operator Regions (Genetics) *Peptide Chain Initiation RNA, Bacterial/genetics RNA, Base Sequence Escherichia coli/*genetics *Gene Expression Regulation, Messenger/genetics RNA, Met/metabolism Ribosomal Proteins/*genetics Ribosomes/*metabolism Structure-Activity Relationship Support, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Unusual structure of the double-stranded RNA associated with the '447' cytoplasmic male sterility in Vicia faba}, author = {P Pfeiffer and J L Jung and J Heitzler and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7685375}, isbn = {7685375}, year = {1993}, date = {1993-01-01}, journal = {J Gen Virol}, volume = {74}, number = {Pt 6}, pages = {1167-1173}, abstract = {The 16.7 kbp dsRNA specific to the '447' cytoplasmic male sterility (CMS) line of Vicia faba was labelled in vitro with [alpha-32P]ATP and poly(A) polymerase, and by T4 RNA ligase-mediated addition of [32P]pCp. Analysis of the reaction products under denaturing conditions revealed in both cases extensive labelling of a 4.5 kb ssRNA, already detected in previous experiments in which the RNA-dependent RNA polymerase associated with the dsRNA was allowed to pursue RNA synthesis on preinitiated complexes. Mobility shift analysis of total pCp-labelled dsRNA revealed not two but three different 3' termini. The most prominent sequencing pattern corresponded to the 4.5 kb ssRNA, indicating that this RNA species has a preferentially accessible, free 3' OH extremity. Northern blot analysis of the denatured dsRNA confirmed that the 4.5 kb ssRNA is a subgenomic mRNA and detected its counterpart of about 12 kb. Nearly all 16.7 kbp dsRNA molecules featured an interrupted positive-sense strand, indicating a marked prevalence of transcription over replication complexes. This unusual strategy of transcription by a strand displacement mechanism, following initiation at an internal discontinuity, is compared with that of other dsRNA viruses or defective viruses, and is discussed in relation to the expression of the CMS trait.}, note = {0022-1317 Journal Article}, keywords = {Base Sequence Cytoplasm Extrachromosomal Inheritance/*genetics Fabaceae/*genetics Inclusion Bodies, Genetic, Medicinal Pollen/genetics RNA/*genetics/isolation & purification RNA Replicase/metabolism Transcription, Unité ARN, Viral Infertility/genetics Molecular Sequence Data *Plants}, pubstate = {published}, tppubtype = {article} } @article{, title = {Correlation between the location of antigenic sites and the prediction of turns in proteins}, author = {J L Pellequer and E Westhof and M H Van Regenmortel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7688347}, isbn = {7688347}, year = {1993}, date = {1993-01-01}, journal = {Immunol Lett}, volume = {36}, number = {1}, pages = {83-99}, abstract = {In the present study, we developed new turn scales based on the occurrence of amino acids at each of the four positions of a turn using a structural database comprised of 87 proteins. We found that the scales correctly predicted a fraction of the turn regions in proteins with approximately 80% confidence. We used the turn scales for predicting the location of antigenic sites in proteins. The method was developed with the specific aim of predicting only a few peaks for each protein (two or three). We found that it leads to a high level of accurate prediction (70% of correct prediction of known epitopes). Our method should be useful for selecting protein regions to be synthesized in order to produce anti-peptide antibodies cross-reacting with the parent protein.}, note = {0165-2478 Journal Article}, keywords = {Amino Acid Sequence Databases, Factual Epitopes/*chemistry Molecular Sequence Data Protein Conformation Protein Folding *Protein Structure, Secondary Proteins/*chemistry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {PREDITOP: a program for antigenicity prediction}, author = {J L Pellequer and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7509182}, isbn = {7509182}, year = {1993}, date = {1993-01-01}, journal = {J Mol Graph}, volume = {11}, number = {3}, pages = {204-210, 191-202}, abstract = {A program (PREDITOP) for predicting the location of antigenic regions (or epitopes) on proteins is described. This program and the associated ones are written in Turbo Pascal and run on IBM-PC compatibles. The program contains 22 normalized scales, corresponding to hydrophilicity, accessibility, flexibility, or secondary structure propensities. New scales are easily implemented. An hydrophobic moment procedure has also been implemented in order to determine amphiphilic helices. The program generates a result file where the values represent a particular physicochemical aspect of the studied protein. PREDITOP can display one or several result files by simple graphical super-imposition. Curve combinations can be done by the ADDITIO or MULTIPLI routines which create a new result file by adding or multiplying previously calculated files representing several propensities. The program is useful and efficient for identifying potential antigenic regions in a protein with the aim of raising antibodies against synthesized peptides which cross-react with the native protein.}, note = {0263-7855 Journal Article}, keywords = {Antigens/*chemistry Computer Graphics Epitopes/chemistry Proteins/*chemistry/*immunology *Software Software Design, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Spontaneous dimerization of retroviral MoMuLV RNA}, author = {J Paoletti and M Mougel and N Tounekti and P M Girard and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8286441}, isbn = {8286441}, year = {1993}, date = {1993-01-01}, journal = {Biochimie}, volume = {75}, number = {8}, pages = {681-686}, abstract = {The genome of the Moloney murine leukemia virus (MoMuLV) is composed of two identical RNA molecules joined at their 5' ends by the dimer linkage structure (DLS). Dimerization sequences are located within the PSI encapsidation domain. We present here an overview of the work we have performed on spontaneous dimerization of a MoMuLV RNA fragment encompassing the PSI domain in order to understand the mechanism by which retroviral RNA dimerization takes place. We present kinetical, thermodynamical and conformational evidence which leads to the conclusion that the PSI domain is a structurally independent domain and that conformational changes are triggered by the dimerization process. We conclude that at least one particular region (nucleotides 278-309) of the RNA is directly involved in the process while the conformation of some other regions is changed probably because of a long-range effect.}, note = {0300-9084 Journal Article}, keywords = {Base Sequence Biopolymers Molecular Sequence Data Moloney murine leukemia virus/*genetics Nucleic Acid Conformation RNA, Non-U.S. Gov't, Unité ARN, Viral/*chemistry Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Methylation of DNA in tissue of ovarian malignant tumors in women]}, author = { W. Baranowski and J. Tomaszewski and G. Keith}, year = {1993}, date = {1993-01-01}, journal = {Ginekol Pol}, volume = {64}, number = {4}, pages = {169-73}, abstract = {DNA methylation level from woman ovarian malignant tissues was investigated by 32P postlabeling of the single nucleotides, separation on TLC with followed autoradiography and Cerenkov counting. Slight differences in DNA methylation extent were found in malignant ovarian tumors as compared to normal myometrium and benign uterine tumor [myomas]. Lowest level of m5dC in relation to C and also in relation to total DNA was found in carcinoma embryonal tissue whereas maximal DNA methylation level was obtained in folliculoma tissue. This preliminary report needs further investigation of particularly genes methylation.}, note = {0017-0011 Journal Article}, keywords = {Abstract, Adult, Aged, Carcinoma/genetics, Cell, DNA, English, Female, Human, Methylation, Middle, Neoplasm/*metabolism, Neoplasms/*genetics, Ovarian, Sertoli, Tumor/genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {A gene encoding arginyl-tRNA synthetase is located in the upstream region of the lysA gene in Brevibacterium lactofermentum: regulation of argS-lysA cluster expression by arginine}, author = {J A Oguiza and M Malumbres and G Eriani and A Pisabarro and L M Mateos and F Martin and J F Martin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8226683}, isbn = {8226683}, year = {1993}, date = {1993-01-01}, journal = {J Bacteriol}, volume = {175}, number = {22}, pages = {7356-7362}, abstract = {The Brevibacterium lactofermentum argS gene, which encodes an arginyl-tRNA synthetase, was identified in the upstream region of the lysA gene. The cloned gene was sequenced; it encodes a 550-amino-acid protein with an M(r) of 59,797. The deduced amino acid sequence showed 28% identical and 49% similar residues when compared with the sequence of the Escherichia coli arginyl-tRNA synthetase. The B. lactofermentum enzyme showed the highly conserved motifs of class I aminoacyl-tRNA synthetases. Expression of the argS gene in B. lactofermentum and E. coli resulted in an increase in aminoacyl-tRNA synthetase activity, correlated with the presence in sodium dodecyl sulfate-polyacrylamide gels of a clear protein band that corresponds to this enzyme. One single transcript of about 3,000 nucleotides and corresponding to the B. lactofermentum argS-lysA operon was identified. The transcription of these genes is repressed by lysine and induced by arginine, showing an interesting pattern of biosynthetic interlock between the pathways of both amino acids in corynebacteria.}, note = {0021-9193 Journal Article}, keywords = {Amino Acid Support, Amino Acid Sequence Amino Acyl-tRNA Ligases/genetics Arginine/*pharmacology Arginine-tRNA Ligase/biosynthesis/*genetics Bacteria/enzymology Brevibacterium/*enzymology/*genetics Carboxy-Lyases/*genetics Cloning, Bacterial Molecular Sequence Data Molecular Weight *Multigene Family Plasmids Restriction Mapping Sequence Homology, Bacterial/*drug effects Gene Expression Regulation, Enzymologic/drug effects *Genes, ERIANI, Molecular Comparative Study Escherichia coli/genetics/growth & development Fungi/enzymology Gene Expression Regulation, Non-U.S. Gov't, Structural, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Unusual deficiency of the modified purine base queuine in transfer ribonucleic acid from the human placenta as tested by enzymatic assay}, author = { W. Baranowski and J. Tomaszewski and G. Keith}, year = {1993}, date = {1993-01-01}, journal = {Am J Obstet Gynecol}, volume = {169}, number = {3}, pages = {581-2}, abstract = {Transfer ribonucleic acid from rapidly growing tissues, particularly from neoplasia, is partially deficient in queuine, a highly modified transfer ribonucleic acid constituent. By means of an enzymatic assay we also found a queuine deficiency (14%) in human placenta transfer ribonucleic acid despite its high concentrations in the amniotic fluid. Proposed cause and significance of the results are discussed.}, note = {0002-9378 Journal Article}, keywords = {&, derivatives/chemistry, Female, Guanine/*analogs, Human, Placenta/*chemistry, Pregnancy, RNA, Transfer/*chemistry}, pubstate = {published}, tppubtype = {article} } @article{, title = {Promoter strength and structure dictate module composition in RNA polymerase III transcriptional activator elements}, author = {E Myslinski and C Schuster and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7693950}, isbn = {7693950}, year = {1993}, date = {1993-01-01}, journal = {J Mol Biol}, volume = {234}, number = {2}, pages = {311-318}, abstract = {RNA polymerase III transcription of genes with external promoters only (e.g. U6 snRNA) or containing in addition an internal B box (selenocysteine tRNA(Sec)) is stimulated by upstream elements; a distal sequence element (DSE) for U6 or an activator element in the tRNA(Sec) gene. In contrast to the composite structure of the DSE which requires an octamer motif, the Xenopus tRNA(Sec) activator element contains an SPH motif only. In vivo transcription is optimally stimulated by SPH in an absolute octamer-independent manner since adding octamer does not induce superstimulation. Experiments performed in the work presented here led to the following observations. Co-operation between SPH and octamer motifs can be detected in two distinct cases: first when these motifs are placed in front of B box-less tRNA(Sec) or U6 external promoters and second, if either element of the external promoter (proximal sequence element or TATA element), or the SPH motif itself, are altered. Altogether, our data provide evidence that an SPH motif can function alone in an optimized promoter only. In contrast, an octamer becomes indispensable when the basal promoter is weak or disabled. It follows that module composition of Pol III transcriptional activator elements is dependent on the structure and strength of the promoter. This reveals the existence of cross-talk between activator and promoter elements, mediated by the bound transcription factors, which are thus able to compensate for each other in order to allow successful assembly of the transcription complex.}, note = {0022-2836 Journal Article}, keywords = {Animals Base Sequence Molecular Sequence Data Mutagenesis, Genetic/*physiology Xenopus laevis, Non-U.S. Gov't Transcription, Nucleic Acid/*physiology Selenocysteine/genetics Support, Site-Directed Oocytes/metabolism Promoter Regions (Genetics)/*genetics RNA/*genetics RNA Polymerase III/*metabolism RNA, Small Nuclear/genetics RNA, Transfer/genetics Regulatory Sequences, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Point mutations 5' to the tRNA selenocysteine TATA box alter RNA polymerase III transcription by affecting the binding of TBP}, author = {E Myslinski and C Schuster and J Huet and A Sentenac and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8290344}, isbn = {8290344}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {25}, pages = {5852-5858}, abstract = {The selenocysteine tRNA(Sec) gene possesses two external promoter elements, one of which is constituted by a strong TATA box. Point mutant analysis performed in this study led to the conclusion that the functional TATA promoter actually encompasses the sequence -34 GGGTATAAAAGG-23. Individual changes at T-31 do not affect transcription much. Position T-29 is less permissive to mutation since transversion to a G, for example, is less well tolerated than at T-31. Interestingly, a double point mutation, converting GG(-33/-32) to TT, causes abrogation of transcription in vivo and severe reduction of transcription in vitro with human TBP. Therefore, data obtained underscore the fact that, in the Xenopus tRNA(Sec), these two Gs are an integral part of the TATA promoter. Gel retardation experiments indicate that the GG to TT substitution, which led human TBP to lose its ability to support efficient transcription in vitro, correlates with the appearance of an altered pattern of retarded complexes. Altogether, the data presented in this report support a model in which TBP interacts directly with the TATA element of the tRNA(Sec) gene, in contrast to the type of interaction proposed for classical TATA-less tRNA genes.}, note = {0305-1048 Journal Article}, keywords = {Amino Acid-Specific/*genetics/metabolism Recombinant Proteins/metabolism Support, Animals Base Sequence DNA/metabolism DNA-Binding Proteins/*metabolism Molecular Sequence Data *Point Mutation Protein Binding RNA Polymerase III/*metabolism RNA, Genetic Xenopus laevis, Non-U.S. Gov't *TATA Box TATA-Box Binding Protein Transcription Factors/*metabolism *Transcription, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Automatic display of RNA secondary structures}, author = {G Muller and C Gaspin and A Etienne and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7507400}, isbn = {7507400}, year = {1993}, date = {1993-01-01}, journal = {Comput Appl Biosci}, volume = {9}, number = {5}, pages = {551-561}, abstract = {A set of programs written in C language with the GL library and under UNIX has been developed for generating compact, pleasant and non-overlapping displays of secondary structures of ribonucleic acids. The first program, rnasearch, implements a new search procedure that dynamically rearranges overlapping portions of the two-dimensional drawing while preserving clear and readable displays of the two-dimensional structure. The algorithm is fast (the execution time for the command rnasearch is 38.6 s for the 16S rRNA of Escherichia coli with 1542 bases), accepts outputs from two-dimensional prediction programs and therefore allows for rapid comparison between the various two-dimensional folds generated. A second program, rnadisplay, allows the graphical display of the computed two-dimensional structures on a graphics workstation. Otherwise, it is possible to obtain a paper output of the two-dimensional structure by using the program print2D which builds a Postscript file. Moreover the two-dimensional drawing can be labelled for representing data coming from chemical modifications and/or enzymatic cleavages. Application to a few secondary structures such as RNaseP, 5S rRNA and 16S rRNA are given.}, note = {0266-7061 Journal Article}, keywords = {16S/chemistry/genetics RNA, 5S/chemistry/genetics Ribonuclease P *Software, Algorithms Bacillus subtilis/chemistry/genetics Base Sequence *Computer Graphics Endoribonucleases/genetics Escherichia coli/chemistry/genetics Molecular Sequence Data Nucleic Acid Conformation RNA/*chemistry/genetics RNA, Bacterial/genetics RNA, Catalytic/genetics RNA, Ribosomal, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Conformational analysis of the 5' leader and the gag initiation site of Mo-MuLV RNA and allosteric transitions induced by dimerization}, author = {M Mougel and N Tounekti and J L Darlix and J Paoletti and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8233816}, isbn = {8233816}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {20}, pages = {4677-4684}, abstract = {Dimerization of genomic RNA is a key step in the retroviral life cycle and has been postulated to be involved in the regulation of translation, encapsidation and reverse transcription. Here, we have derived a secondary structure model of nucleotides upstream from psi and of the gag initiation region of Mo-MuLV RNA in monomeric and dimeric forms, using chemical probing, sequence comparison and computer prediction. The 5' domain is extensively base-paired and interactions take place between U5 and 5' leader sequences. The U5-PBS subdomain can fold in two mutually exclusive conformations: a very stable and extended helical structure (E form) in which 17 of the 18 nucleotides of the PBS are paired, or an irregular three-branch structure (B form) in which 10 nucleotides of the PBS are paired. The dimeric RNA adopts the B conformation. The monomeric RNA can switch from the E to the B conformation by a thermal treatment. If the E to B transition is associated to dimerization, it may facilitate annealing of the primer tRNAPro to the PBS by lowering the free energy required for melting the PBS. Furthermore, dimerization induces allosteric rearrangements around the SD site and the gag initiation region.}, note = {0305-1048 Journal Article}, keywords = {Allosteric Regulation Base Sequence Biopolymers *Genes, gag Molecular Sequence Data Moloney murine leukemia virus/*genetics *Nucleic Acid Conformation RNA, Non-U.S. Gov't, Nucleic Acid Support, Unité ARN, Viral/*chemistry/genetics Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{, title = {Minimal 16S rRNA binding site and role of conserved nucleotides in Escherichia coli ribosomal protein S8 recognition}, author = {M Mougel and C Allmang and F Eyermann and C Cachia and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7689052}, isbn = {7689052}, year = {1993}, date = {1993-01-01}, journal = {Eur J Biochem}, volume = {215}, number = {3}, pages = {787-792}, abstract = {Escherichia coli ribosomal protein S8 was previously shown to bind a 16S rRNA fragment (nucleotides 584-756) with the same affinity as the complete 16S rRNA, and to shield an irregular helical region (region C) [Mougel, M., Eyermann, F., Westhof, E., Romby, P., Expert-Bezancon, Ebel, J. P., Ehresmann, B. & Ehresmann, C. (1987). J. Mol. Biol. 198, 91-107]. Region C was postulated to display characteristic features: three bulged adenines (A595, A640 and A642), a non-canonical U598-U641 pair surrounded by two G.C pairs. In order to delineate the minimal RNA binding site, deletions were introduced by site-directed mutagenesis and short RNA fragments were synthesized. Their ability to bind S8 was assayed by filter binding. Our results show that the RNA binding site can be restricted to a short helical stem (588-605/633-651) containing region C. The second part of the work focused on region C and on the role of conserved nucleotides as potential determinants of S8 recognition. Single and double mutations were introduced by site-directed mutagenesis in fragment 584-756, and their effect on S8 binding was measured. It was found that the three bulged positions are essential and that adenines are required at positions 640 and 642. U598 is also crucial and the highly conserved G597.C643 pair cannot be inverted. These conserved nucleotides are either directly involved in the recognition process as direct contacts or required to maintain a specific conformation. The strong evolutionary pressure and the small number of positive mutants stress the high stringency of the recognition process.}, note = {0014-2956 Journal Article}, keywords = {16S/*metabolism Ribosomal Proteins/genetics/*metabolism Support, Adenine/metabolism Bacterial Proteins/metabolism Base Composition Base Sequence Binding Sites Conserved Sequence Escherichia coli/*metabolism Molecular Sequence Data Mutation Nucleic Acid Conformation RNA, Bacterial/metabolism RNA, Non-U.S. Gov't, Ribosomal, ROMBY, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Overproduction and purification of native and queuine-lacking Escherichia coli tRNA(Asp). Role of the wobble base in tRNA(Asp) acylation}, author = {F Martin and G Eriani and S Eiler and D Moras and G Dirheimer and J Gangloff}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8263943}, isbn = {8263943}, year = {1993}, date = {1993-01-01}, journal = {J Mol Biol}, volume = {234}, number = {4}, pages = {965-974}, abstract = {Escherichia coli tRNA(Asp) was overproduced in E. coli up to 15-fold from a synthetic tRNA(Asp) gene placed in a plasmid under the dependence of an isopropyl-beta,D-thiogalactopyranoside-inducible promoter. Purification to nearly homogeneity (95%) was achieved after two HPLC DEAE-cellulose columns. E. coli tRNA(Asp)[G34] (having guanine instead of queuine at position 34) was obtained by the same procedure except that it was overproduced in a strain lacking the enzyme responsible for queuine modification. Nucleoside analysis showed that, except for the replacement of Q34 by G34 in mutant-derived tRNA(Asp), the base modification levels of both tRNAs are the same as those in wild-type E. coli tRNA(Asp). Kinetic properties of tRNA(Asp)[Q34] and [G34] with yeast AspRS compared to those in the homologous reactions in yeast and E. coli clearly indicate that the major identity elements are the same in both organisms: the conserved discriminant base and the anticodon triplet. In connection with this, we explored by site-directed mutagenesis the functional role of the interactions which, as revealed by the crystallographic structure, occur between the wobble base of yeast tRNA(Asp) and two residues of yeast AspRS. Their absence strongly affected aspartylation and the kd of tRNA(Asp). Each contact individually restores almost completely the wild-type acylation properties of the enzyme; thus, wobble base recognition in yeast appears to be more protected against mutational events than in E. coli, where only one contact is thought to occur at position 34.}, note = {0022-2836 Journal Article}, keywords = {*Amino Acid Activation Anticodon Aspartate-tRNA Ligase/metabolism Base Composition Base Sequence Cloning, Asp/chemistry/*metabolism Saccharomyces cerevisiae/metabolism Structure-Activity Relationship Support, ERIANI, Molecular Comparative Study Crystallography, Non-U.S. Gov't, Transfer, Unité ARN, X-Ray Escherichia coli/*metabolism Guanine/*analogs & derivatives/chemistry Molecular Sequence Data Nucleic Acid Conformation RNA}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Higher Order Structures of HIV-1 RNA as Sites for Drug Actions.}, author = {R Marquet and C Ehresmann and B Ehresmann}, editor = {S T Crooke and B Lebleu}, url = {http://www.crcpress.com/product/isbn/9780849347054}, year = {1993}, date = {1993-01-01}, booktitle = {Antisense Research and Applications}, pages = {401-414}, publisher = {CRC Press}, keywords = {MARQUET, Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {Influence of impurities on protein crystallization: the case of lysozyme.}, author = {B Lorber and M Skouri and J P Munch and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/S0022024807801242}, isbn = {10.1016/S0022-0248(07)80124-2}, year = {1993}, date = {1993-01-01}, journal = {J Crystal Growth}, volume = {128}, number = {1-4, part 2}, pages = {1203-1211}, abstract = {Several batches of hen egg white lysozyme were compared on the basis of their biochemical purity and homogeneity as well as of their ability to crystallize in the tetragonal space group in the presence of sodium chloride and sodium acetate at pH 4.5. Trace amounts (< 2% (w/w)) of detectable protein impurities and inactive lysozyme molecules interfere with the nucleation and crystal growth processes. The presence of impurities significantly decreases solubility of lysozyme preparations. The intentional addition of ovalbumin or bovine serum albumin to pure lysozyme is correlated with an increase of the proportion of twinned crystals. Thus, reproductibility of lysozyme crystallization is dependent upon the presence of impurities.}, note = {Proceedings of the Tenth International Conference on Crystal Growth}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Sequence of tRNA(Asp) from Thermus thermophilus HB8}, author = {G Keith and M Yusupov and C Briand and D Moras and D Kern and C Brion}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7692402}, isbn = {7692402}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {18}, pages = {4399}, note = {0305-1048 Journal Article}, keywords = {Asp/chemistry/*genetics Thermus thermophilus/*genetics, Bacterial/chemistry/genetics RNA, Base Sequence Molecular Sequence Data Nucleic Acid Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The primary structure of cytoplasmic initiator tRNA(Met) from Schizosaccharomyces pombe}, author = {G Keith and J Heitzler and C el Adlouni and A L Glasser and C Fix and J Desgres and G Dirheimer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8332511}, isbn = {8332511}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {12}, pages = {2949}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Molecular Sequence Data RNA, Fungal/*chemistry RNA, Met/*chemistry Schizosaccharomyces/*genetics, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Monitoring of the cooperative unfolding of the sunY group I intron of bacteriophage T4. The active form of the sunY ribozyme is stabilized by multiple interactions with 3' terminal intron components}, author = {L Jaeger and E Westhof and F Michel}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8230218}, isbn = {8230218}, year = {1993}, date = {1993-01-01}, journal = {J Mol Biol}, volume = {234}, number = {2}, pages = {331-346}, abstract = {We have studied the mechanism by which the 3' terminal domain of the sunY intron of bacteriophage T4 activates the group I ribozyme core of this intron, from which it is separated by some 800 nucleotides. As shown by monitoring either UV absorbance or self-splicing reaction kinetics as a function of temperature, intron transcripts undergo highly cooperative unfolding/inactivation upon heating: the two methods yield similar estimates of the thermodynamic parameters associated with this process. Such cooperativity makes it possible in turn to assess the energetic contribution of specific interactions to the overall structure, by comparing the sensitivity to heat inactivation of molecules carrying various nucleotide substitutions. By combining this approach with chemical modification, we have probed several proven or putative interactions between the core and 3' terminal domain of the intron and conclude that the role of the 3' terminal domain is to stabilize the active form of the ribozyme. Interestingly, the P9.0 interaction, which brings 3' terminal nucleotides next to the core site that binds the guanosine cofactor of the self-splicing reaction, is now shown to be composed in fact of two distinct pairings. An isolated base-pair (P9.0a), involving a residue located only six nucleotides upstream of the 3' splice site, participates in the stabilization of the ribozyme and appears to persist during the second stage of self-splicing (exon ligation). In contrast, formation of the previously demonstrated P9.0b pairing, which involves the two penultimate intron nucleotides, contributes no additional stability and results in no detectable rearrangement of the core structure. Implications for the concept of a static ribozyme are discussed in the light of a slightly revised three-dimensional model of the sunY intron.}, note = {0022-2836 Journal Article}, keywords = {Bacteriophage T4/*genetics Base Sequence Enzyme Activation Enzyme Stability Introns/*physiology Models, Catalytic/drug effects/*metabolism/radiation effects RNA, Molecular Molecular Sequence Data Nucleic Acid Denaturation/physiology RNA, Non-U.S. Gov't Thermodynamics Ultraviolet Rays, Unité ARN, Viral/drug effects/*metabolism/radiation effects Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Modified nucleotides of tRNA(3Lys) modulate primer/template loop-loop interaction in the initiation complex of HIV-1 reverse transcription}, author = {C Isel and R Marquet and G Keith and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7503978}, isbn = {7503978}, year = {1993}, date = {1993-01-01}, journal = {J Biol Chem}, volume = {268}, number = {34}, pages = {25269-25272}, abstract = {In all retroviruses, reverse transcription is primed by a tRNA whose 3' end 18 nucleotides are complementary to the so called viral primer binding site. Previous work showed that reverse transcription of HIV-1 RNA is initiated by tRNA(3Lys). Using a variety of chemical and enzymatic structural probes, we investigated the interactions between HIV-1 RNA and its natural primer tRNA(3Lys). In addition to the predictable contacts between the viral primer binding site and the 3' end of tRNA(3Lys), a specific interaction takes place between an A-rich loop located upstream of the primer binding site region and the anticodon loop of tRNA(3Lys). This AAAA/Umcm5s2UUU loop-loop interaction is not observed when the natural primer is replaced by an in vitro synthesized tRNA(3Lys) transcript. Furthermore, dethiolation of the modified nucleotide mcm5s2U at position 34 of tRNA(3Lys) strongly destabilizes this interaction. Sequence and structure comparisons indicate that the primer/template loop-loop interaction is conserved in all HIV-1 isolates, and possibly also in HIV-2 and SIV.}, note = {0021-9258 Journal Article}, keywords = {Anticodon/genetics/metabolism Base Sequence Binding Sites Comparative Study DNA Primers HIV-1/genetics/*metabolism HIV-1 Reverse Transcriptase HIV-2/genetics/metabolism Molecular Sequence Data Nucleic Acid Conformation RNA, Genetic, Genetic *Transcription, Lys/*metabolism RNA, MARQUET, Non-U.S. Gov't Templates, Transfer, Unité ARN, Viral/*biosynthesis RNA-Directed DNA Polymerase/*metabolism SIV/genetics/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {An unusual RNA tertiary interaction has a role for the specific aminoacylation of a transfer RNA}, author = {Y M Hou and E Westhof and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8341698}, isbn = {8341698}, year = {1993}, date = {1993-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {90}, number = {14}, pages = {6776-6780}, abstract = {The nucleotides in a tRNA that specifically interact with the cognate aminoacyl-tRNA synthetase have been found largely located in the helical stems, the anticodon, or the discriminator base, where they vary from one tRNA to another. The conserved and semiconserved nucleotides that are responsible for the tRNA tertiary structure have been shown to have little role in synthetase recognition. Here we report that aminoacylation of Escherichia coli tRNA(Cys) depends on the anticodon, the discriminator base, and a tertiary interaction between the semiconserved nucleotides at positions 15 and 48. While all other tRNAs contain a purine at position 15 and a complementary pyrimidine at position 48 that establish the tertiary interaction known as the Levitt pair, E. coli tRNA(Cys) has guanosine -15 and -48. Replacement of guanosine -15 or -48 with cytidine virtually eliminates aminoacylation. Structural analyses with chemical probes suggest that guanosine -15 and -48 interact through hydrogen bonds between the exocyclic N-2 and ring N-3 to stabilize the joining of the two long helical stems of the tRNA. This tertiary interaction is different from the traditional base pairing scheme in the Levitt pair, where hydrogen bonds would form between N-1 and O-6. Our results provide evidence for a role of RNA tertiary structure in synthetase recognition.}, note = {0027-8424 Journal Article}, keywords = {Amino Acyl-tRNA Ligases/*metabolism Base Sequence Escherichia coli/*chemistry/genetics/metabolism Hydrogen Bonding Models, Cys/*chemistry/genetics/metabolism Support, Molecular Molecular Sequence Data *Nucleic Acid Conformation RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The secondary structure of the 5'-noncoding region of beet necrotic yellow vein virus RNA 3: evidence for a role in viral RNA replication}, author = {D Gilmer and C Allmang and C Ehresmann and H Guilley and K Richards and G Jonard and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8464729}, isbn = {8464729}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {6}, pages = {1389-1395}, abstract = {Secondary structure-sensitive chemical and enzymatic probes have been used to produce a model for the folding of the first 312 residues of the long 5'-noncoding region of beet necrotic yellow vein virus RNA 3. The structure consists of two major domains, one of which includes long distance base-pairing interactions between two short sequence elements (Box I and Box II) situated between positions 237 and 292 and complementary elements (Box I' and II') near the 5'-terminus. Previous studies have shown that base pairing between these sequence elements (in either the plus-strand or minus-strand RNA) is important for RNA 3 accumulation during infection. RNA 3 transcripts were produced containing mutations which preferentially disrupted Box II-II' base pairing in either the plus- or minus-strand. In infection experiments, transcripts with mutations which disrupted the Box II-II' interaction in the plus-strand structure replicated less efficiently than mutants in which the Box II-II' interaction was disrupted in the minus-strand. These findings indicate that the complex 5'-proximal plus-strand structure to which the Box II-II' interaction contributes comprises at least part of the promoter for plus-strand RNA synthesis.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Gene Expression Hydrogen Bonding Molecular Sequence Data Mutagenesis, Messenger/genetics RNA, Site-Directed Nucleic Acid Conformation Plant Viruses/*genetics/ultrastructure Promoter Regions (Genetics) RNA Viruses/*genetics/ultrastructure RNA, Unité ARN, Viral/chemistry/*genetics/ultrastructure Vegetables *Virus Replication}, pubstate = {published}, tppubtype = {article} } @article{, title = {tRNA structure and aminoacylation efficiency}, author = {R Giege and J D Puglisi and C Florentz}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8341800}, isbn = {8341800}, year = {1993}, date = {1993-01-01}, journal = {Prog Nucleic Acid Res Mol Biol}, volume = {45}, pages = {129-206}, note = {0079-6603 Journal Article Review Review, Academic}, keywords = {Acylation Amino Acids/metabolism Amino Acyl-tRNA Ligases/metabolism Animals Base Sequence Dna Human Molecular Sequence Data *Nucleic Acid Conformation RNA, FLORENTZ, Non-U.S. Gov't, Transfer/*chemistry/metabolism Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The TYMV tRNA-like structure}, author = {R Giege and C Florentz and T W Dreher}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8268257}, isbn = {8268257}, year = {1993}, date = {1993-01-01}, journal = {Biochimie}, volume = {75}, number = {7}, pages = {569-582}, abstract = {The genomic RNA from turnip yellow mosaic virus presents a 3'-end functionally and structurally related to tRNAs. This report summarizes our knowledge about the peculiar structure of the tRNA-like domain and its interaction with tRNA specific proteins, like RNAse P, tRNA nucleotidyl-transferase, aminoacyl-tRNA synthetases, and elongation factors. It discusses also the biological role of this structure in the viral life cycle. A brief survey of our knowledge of other tRNA mimicries in biological systems, as well as their relevance for understanding canonical tRNA, will also be presented.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Base Sequence Molecular Sequence Data *Nucleic Acid Conformation RNA, FLORENTZ, Transfer/*chemistry/metabolism RNA, Unité ARN, Viral/*chemistry/metabolism Tymovirus/*genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Triple aminoacylation specificity of a chimerized transfer RNA}, author = {M Frugier and C Florentz and P Schimmel and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8268184}, isbn = {8268184}, year = {1993}, date = {1993-01-01}, journal = {Biochemistry}, volume = {32}, number = {50}, pages = {14053-14061}, abstract = {We report here the rational design and construction of a chimerized transfer RNA with tripartite aminoacylation specificity. A yeast aspartic acid specific tRNA was transformed into a highly efficient acceptor of alanine and phenylalanine and a moderate acceptor of valine. The transformation was guided by available knowledge of the requirements for aminoacylation by each of the three amino acids and was achieved by iterative changes in the local sequence context and the structural framework of the variable loop and the two variable regions of the dihydrouridine loop. The changes introduced to confer efficient acceptance of the three amino acids eliminate aminoacylation with aspartate. The interplay of determinants and antideterminants for different specific aminoacylations, and the constraints imposed by the structural framework, suggest that a tRNA with an appreciable capacity for more than three efficient aminoacylations may be inherently difficult to achieve.}, note = {0006-2960 Journal Article}, keywords = {Acylation Alanine/metabolism Base Sequence Chimera Escherichia coli/genetics Molecular Sequence Data Mutation Nucleic Acid Conformation Phenylalanine/metabolism RNA, Asp/chemistry/genetics/*metabolism Saccharomyces cerevisiae/genetics Support, ERIANI, FLORENTZ, FRUGIER, Non-U.S. Gov't Support, P.H.S. Valine/metabolism, Transfer, U.S. Gov't, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular dynamics simulations of poly(dA).poly(dT): comparisons between implicit and explicit solvent representations}, author = {V Fritsch and G Ravishanker and D L Beveridge and E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8218922}, isbn = {8218922}, year = {1993}, date = {1993-01-01}, journal = {Biopolymers}, volume = {33}, number = {10}, pages = {1537-1552}, abstract = {The program AMBER 3.0 has been used to generate molecular dynamics trajectories of a poly(dA).poly(dT) decamer. The simulations were performed using different methods to treat solvent effects. Results of a simulation including 18 counterions NH4+ and 4109 water molecules under (N, P, T) conditions were compared to simulation runs with implicit solvent representation in which solvent screening effects were represented by the use of a sigmoidal distance-dependent dielectric function. In the latter case, the system was simulated under microcanonical (N, V, E) and canonical (N, V, T) conditions. For the fully hydrated system simulation, a preequilibration protocol was developed since it was observed that long and progressive periods of heating and equilibration on the overall system were necessary in order to avoid energetic collisions between the solute and the solvent molecules, leading to severe irreversible deformation of the solute. A detailed analysis of DNA conformations, sugar puckers, and stability of the hydrogen bonds, Watson-Crick and three-center H bonds, is reported. The results show that DNA remains essentially in the B conformer with a tendency in the hydrated model to adopt a slightly distorted, unwound, and stretched conformation in comparison to standard B-DNA. Concerning sugar puckers, the mean pseudorotation phases of the adenine residues are systematically higher than those of the thymine residues, except in the case of the hydrated model for which a articular behavior is observed for the adenine strand. In this case, the terminal bases oscillate between C2'-endo and O4'-endo and the central ones stay in the C3'-endo domain. The mean lifetimes of the internal Watson-Crick H-bond (A) HN6.O4(T) are also dependent on the base pairs included in the calculation, excepted for the implicit solvent simulation at constant temperature. The three-center H bonds have very small mean lifetimes in all three cases of MD simulation. In the minor groove of the hydrated model, a spine of hydration is found as observed by x-ray crystallography and other theoretical simulations. On the basis of the rms deviations, it appears that the fully hydrated simulation has not reached a plateau at the end of the run, while the implicit simulation at constant energy seems to have converged. At constant temperature, very large oscillations in rms deviations are observed.}, note = {0006-3525 Journal Article}, keywords = {Base Sequence Chemistry, Physical Comparative Study Computer Simulation Molecular Sequence Data Poly dA-dT/*chemistry Solvents Thermodynamics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Non-canonical substrates of aminoacyl-tRNA synthetases: the tRNA-like structure of brome mosaic virus genomic RNA}, author = {B Felden and C Florentz and E Westhof and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8199250}, isbn = {8199250}, year = {1993}, date = {1993-01-01}, journal = {Biochimie}, volume = {75}, number = {12}, pages = {1143-1157}, abstract = {A 3-D model of the tyrosylable tRNA-like domain of the genomic brome mosaic virus RNAs was built by computer modelling based on solution probing of the molecule with different chemical and enzymatic reagents. This model encompasses four major structural domains, including two peculiar substructures oriented perpendicularly and mimicking a tRNA structure, and a fifth domain which makes the connection with the rest of the viral RNA. After recalling the different steps that led to the present structural knowledge of the BMV tRNA-like domain, we review its novel structural features revealed by the modelling and that did not appear in older versions of 3-D models of this structure. These features comprise additional base-pairs, hairpin loops, new tertiary long-range interactions, and a second pseudoknot. The main goal of this paper is to strengthen the validity of the model by establishing correlations between the putative 3-D conformation and the functional properties of the domain. For that, we show how the present structural model rationalises mutagenic and footprinting data that have established the importance of specific regions of the RNA for its recognition and aminoacylation by yeast tyrosyl-tRNA synthetase. We discuss further how the model corroborates mutational analyses performed to understand recognition of this RNA domain by the (ATP,CTP):tRNA nucleotidyl-transferase and by the viral replicase. The published mutants of the BMV tRNA-like domain fall into two classes. In one class, the mutants leave unchanged the overall architecture of the molecule, thereby affecting functions directly. In the second class, the overall architecture of the mutants is perturbed, and thus functions are affected indirectly.}, note = {0300-9084 Journal Article Review Review, Tutorial}, keywords = {Base Sequence Bromovirus/*genetics Computer Simulation Genome, FLORENTZ, Molecular Molecular Sequence Data Mutation/genetics Nucleic Acid Conformation Promoter Regions (Genetics) RNA, Non-U.S. Gov't Tyrosine-tRNA Ligase/chemistry/*metabolism, Transfer/*chemistry/genetics/metabolism RNA, Unité ARN, Viral Models, Viral/*chemistry/genetics/metabolism Structure-Activity Relationship Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Role of dimerization in yeast aspartyl-tRNA synthetase and importance of the class II invariant proline}, author = {G Eriani and J Cavarelli and F Martin and G Dirheimer and D Moras and J Gangloff}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8248175}, isbn = {8248175}, year = {1993}, date = {1993-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {90}, number = {22}, pages = {10816-10820}, abstract = {Cytoplasmic aspartyl-tRNA synthetase (AspRS; EC 6.1.1.12) from yeast is, as are most class II synthetases, an alpha 2 dimer. The only invariant amino acid in signature motif 1 of this class is Pro-273; this residue is located at the dimer interface. To understand the role of Pro-273 in the conserved dimeric configuration, we tested the effect of a Pro-273-->Gly (P273G) substitution on the catalytic properties of homo- and heterodimeric AspRS. Heterodimers of AspRS were produced in vivo by overexpression of their respective subunit variants from plasmid-encoded genes and purified to homogeneity in one HPLC step. The homodimer containing the P273G shows an 80% inactivation of the enzyme and an affinity decrease for its cognate tRNA(Asp) of one order of magnitude. The P273G-mutated subunit recovered wild-type enzymatic properties when associated with a native subunit or a monomer otherwise inactivated having an intact dimeric interface domain. These results, which can be explained by the crystal structure of the native enzyme complexed with its substrates, confirm the structural importance of Pro-273 for dimerization and clearly establish the functional interdependence of the AspRS subunits. More generally, the dimeric conformation may be a structural prerequisite for the activity of mononucleotide binding sites constructed from antiparallel beta strands.}, note = {0027-8424 Journal Article}, keywords = {Asp/metabolism Saccharomyces cerevisiae/chemistry Structure-Activity Relationship Support, Aspartate-tRNA Ligase/*chemistry Fungal Proteins/chemistry Kinetics Macromolecular Systems Models, ERIANI, Molecular Mutagenesis, Non-U.S. Gov't, Site-Directed Proline/chemistry Protein Binding Protein Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Rye nuclease I as a tool for structural studies of tRNAs with large variable arms}, author = {C el Adlouni and G Keith and G Dirheimer and J W Szarkowski and A Przykorska}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8383845}, isbn = {8383845}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {4}, pages = {941-947}, abstract = {A single-strand-specific nuclease from rye germ (Rn nuclease I) was used for secondary and tertiary structure investigations of tRNAs with large variable arms (class II tRNAs). We have studied the structure in solution of two recently sequenced tRNA(Leu): yeast tRNA(Leu)(ncm5UmAA) and bovine tRNA(Leu)(XmAA) as well as yeast tRNA(Leu)(UAG), tRNA(Leu)(m5CAA) and tRNA(Ser)(IGA). The latter is the only tRNA with a long variable arm for which the secondary and tertiary structure has already been studied by use of chemical probes and computer modelling. The data obtained in this work showed that the general model of class II tRNAs proposed by others for tRNA(Ser) can be extended to tRNAs(Leu) as well. However interesting differences in the structure of tRNAs(Leu) versus tRNA(Ser)(IGA) were also noticed. The main difference was observed in the accessibility of the variable loops to nucleolytic attack of Rn nuclease I: variable loops of all studied tRNA(Leu) species were cut by Rn nuclease I, while that of yeast tRNA(Ser)(IGA) was not. This could be due to differences in stability of the variable arms and the lengths of their loops which are 3 and 4 nucleotides in tRNA(Ser)(IGA) and tRNAs(Leu) respectively.}, note = {0305-1048 Journal Article}, keywords = {Animals Anticodon Base Sequence Cattle Molecular Sequence Data Nucleic Acid Conformation *Nucleotidases RNA, Leu/chemistry RNA, Non-U.S. Gov't, Ser/chemistry Saccharomyces cerevisiae Secale cereale/*enzymology Support, Transfer, Transfer/*chemistry RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Translational regulation of the Escherichia coli threonyl-tRNA synthetase gene: structural and functional importance of the thrS operator domains}, author = {C Brunel and P Romby and H Moine and J Caillet and M Grunberg-Manago and M Springer and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8199252}, isbn = {8199252}, year = {1993}, date = {1993-01-01}, journal = {Biochimie}, volume = {75}, number = {12}, pages = {1167-1179}, abstract = {Previous work showed that E coli threonyl-tRNA synthetase (ThrRS) binds to the leader region of its own mRNA and represses its translation by blocking ribosome binding. The operator consists of four distinct domains, one of them (domain 2) sharing structural analogies with the anticodon arm of the E coli tRNA(Thr). The regulation specificity can be switched by using tRNA identity rules, suggesting that the operator could be recognized by ThrRS as a tRNA-like structure. In the present paper, we investigated the relative contribution of the four domains to the regulation process by using deletions and point mutations. This was achieved by testing the effects of the mutations on RNA conformation (by probing experiments), on ThrRS recognition (by footprinting experiments and measure of the competition with tRNA(Thr) for aminoacylation), on ribosome binding and ribosome/ThrRS competition (by toeprinting experiments). It turns out that: i) the four domains are structurally and functionally independent; ii) domain 2 is essential for regulation and contains the major structural determinants for ThrRS binding; iii) domain 4 is involved in control and ThrRS recognition, but to a lesser degree than domain 2. However, the previously described analogies with the acceptor-like stem are not functionally significant. How it is recognized by ThrRS remains to be resolved; iv) domain 1, which contains the ribosome loading site, is not involved in ThrRS recognition. The binding of ThrRS probably masks the ribosome binding site by steric hindrance and not by direct contacts. This is only achieved when ThrRS interacts with both domains 2 and 4; and v) the unpaired domain 3, which connects domains 2 and 4, is not directly involved in ThrRS recognition. It should serve as an articulation to provide an appropriate spacing between domains 2 and 4. Furthermore, it is possibly involved in ribosome binding.}, note = {0300-9084 Journal Article}, keywords = {Bacterial/*genetics Molecular Sequence Data Mutation Nucleic Acid Conformation *Operator Regions (Genetics) Point Mutation Protein Structure, Base Sequence Escherichia coli/*enzymology/genetics Gene Deletion Gene Expression Regulation, Genetic, Messenger/chemistry/metabolism RNA, Met/chemistry/metabolism Ribosomes/metabolism Structure-Activity Relationship Support, Non-U.S. Gov't Threonine-tRNA Ligase/chemistry/*genetics/metabolism Translation, ROMBY, Secondary RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Functional sites in the 5' region of human immunodeficiency virus type 1 RNA form defined structural domains}, author = {F Baudin and R Marquet and C Isel and J L Darlix and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8429553}, isbn = {8429553}, year = {1993}, date = {1993-01-01}, journal = {J Mol Biol}, volume = {229}, number = {2}, pages = {382-397}, abstract = {The 5' region of HIV-1 RNA contains functional elements involved in key steps of the retroviral cycle, such as genomic RNA transcription, splicing, translation, dimerization or initiation of reverse transcription. In the present work, we investigated the conformation of the first 500 nucleotides covering the RNA leader and the 5' gag coding sequences of HIV-MAL, using chemical probing. We provide detailed information on almost each nucleotide at one of their Watson-Crick positions and on position N-7 of purines. Experiments were conducted on two in vitro transcribed RNA fragments (1 to 707 and 1 to 311). A secondary structure model was derived by combining the experimental data, computer predictions and sequence comparison. Under conditions favoring dimerization (high salt concentration), HIV-1 RNA folds into independent structural domains that can be related to defined functional regions. The first domain corresponds to TAR forming a stable stem-loop. Intrinsic structural features are found to stabilize the TAR hairpin loop. The second domain (nucleotides 56 to 299) contains the PBS sequence, which is located in a stable subdomain constrained by a four stem junction (nucleotides 139 to 218). Although the MAL isolate has an insertion near the PBS, probably resulting from the duplication of a 23-nucleotide sequence, the structural organization of this subdomain is conserved in all other HIV-1 isolates. The third domain (nucleotides 300 to 404) contains the splice donor site, packaging and dimerization elements and the AUG initiation codon of gag. A major result is the structural versatility of this region. Two mutually exclusive structures, both equally in agreement with probing data, could modulate the different functions involving this domain. The reduced accessibility of the gag translational initiation site possibly accounts for the low efficiency of the in vitro translation of the dimer. Finally, the 5' gag coding sequences form a metastable domain.}, note = {0022-2836 Journal Article}, keywords = {Animals Base Sequence Electrophoresis, Genetic, MARQUET, Non-U.S. Gov't Translation, Polyacrylamide Gel HIV-1/*genetics Molecular Sequence Data Nucleic Acid Conformation RNA, Unité ARN, Viral/*chemistry/metabolism Rabbits Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Solution structure of selenocysteine-inserting tRNA(Sec) from Escherichia coli. Comparison with canonical tRNA(Ser)}, author = {C Baron and E Westhof and A Bock and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8510147}, isbn = {8510147}, year = {1993}, date = {1993-01-01}, journal = {J Mol Biol}, volume = {231}, number = {2}, pages = {274-292}, abstract = {Selenocysteine-inserting tRNAs (or tRNA(Sec)) are structurally untypical tRNAs that are charged by seryl-tRNA synthetase before being recognized by the selenocysteine synthase that converts serine into selenocysteine. tRNA(Sec) from Escherichia coli contains 95 nucleotides and is the longest tRNA known to date, in contrast to canonical tRNA(Ser), 88 nucleotides-long. We have studied its solution conformation by chemical and enzymatic probing. Global structural features were obtained by cobra venom and S1 nuclease mapping, as well as by probing with Pb2+. Accessibilities of phosphate groups were measured by ethylnitrosourea probing. Information about positions in bases involved in Watson-Crick pairing, in stacking or in tertiary interactions were obtained by chemical probing with dimethylsulfate, diethylpyrocarbonate, kethoxal and carbodiimide. On the basis of these chemical data, a three-dimensional model was constructed by computer modeling and compared to that of canonical tRNA(Ser). tRNA(Sec) resembles tRNA(Ser) at the level of its T-arm and anticodon-arm conformations, as well as at the joining of the D- and T-loops by a tertiary Watson-Crick G19-C56 interaction. Its extra-long variable arm is a double-stranded structure closed by a four nucleotide loop that is linked to the body of the tRNA in a way different from that found in tRNA(Ser). As anticipated from the peculiar features of the sequence in the D-loop and at the junction of amino acid and D-arms, tRNA(Sec) possesses a novel but restricted set of tertiary interactions in the core of its three-dimensional structure: a G8-A21-U14 triple pair and a novel interaction between C16 of the D-loop and C59 of the T-loop. A third triple interaction involving C15-G20a-G48 is suggested but some experimental evidence for it is still lacking. It is furthermore concluded that the D-arm has six base-pairs instead of three, as in canonical class II tRNA(Ser), with the D-loop containing only four nucleotides. Finally, the amino acid accepting arm forms a stack of eight Watson-Crick base-pairs (instead of 7 in other tRNAs). The biological relevance of this model with regard to interaction with seryl-tRNA synthetase and enzymes from the selenocysteine metabolism is discussed.}, note = {0022-2836 Journal Article}, keywords = {Adenine/chemistry Aspergillus Nuclease S1/pharmacology Base Sequence Comparative Study Escherichia coli/*chemistry Guanine/chemistry Lead/pharmacology Models, Amino Acid-Specific/*chemistry/drug effects RNA, Molecular Molecular Sequence Data Nucleic Acid Conformation RNA, Non-U.S. Gov't, Nucleic Acid Support, Ser/*chemistry/drug effects Selenocysteine/*metabolism Sequence Homology, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {[Methylation of DNA in tissue of ovarian malignant tumors in women]}, author = {W Baranowski and J Tomaszewski and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8282237}, isbn = {8282237}, year = {1993}, date = {1993-01-01}, journal = {Ginekol Pol}, volume = {64}, number = {4}, pages = {169-173}, abstract = {DNA methylation level from woman ovarian malignant tissues was investigated by 32P postlabeling of the single nucleotides, separation on TLC with followed autoradiography and Cerenkov counting. Slight differences in DNA methylation extent were found in malignant ovarian tumors as compared to normal myometrium and benign uterine tumor [myomas]. Lowest level of m5dC in relation to C and also in relation to total DNA was found in carcinoma embryonal tissue whereas maximal DNA methylation level was obtained in folliculoma tissue. This preliminary report needs further investigation of particularly genes methylation.}, note = {0017-0011 Journal Article}, keywords = {Adult Carcinoma/genetics DNA, Neoplasm/*metabolism English Abstract Female Human Methylation Middle Aged Ovarian Neoplasms/*genetics Sertoli Cell Tumor/genetics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Unusual deficiency of the modified purine base queuine in transfer ribonucleic acid from the human placenta as tested by enzymatic assay}, author = {W Baranowski and J Tomaszewski and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=8372867}, isbn = {8372867}, year = {1993}, date = {1993-01-01}, journal = {Am J Obstet Gynecol}, volume = {169}, number = {3}, pages = {581-582}, abstract = {Transfer ribonucleic acid from rapidly growing tissues, particularly from neoplasia, is partially deficient in queuine, a highly modified transfer ribonucleic acid constituent. By means of an enzymatic assay we also found a queuine deficiency (14%) in human placenta transfer ribonucleic acid despite its high concentrations in the amniotic fluid. Proposed cause and significance of the results are discussed.}, note = {0002-9378 Journal Article}, keywords = {Female Guanine/*analogs & derivatives/chemistry Human Placenta/*chemistry Pregnancy RNA, Transfer/*chemistry, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Non-standard translational events in Candida albicans mediated by an unusual seryl-tRNA with a 5'-CAG-3' (leucine) anticodon}, author = { M. A. Santos and G. Keith and M. F. Tuite}, year = {1993}, date = {1993-01-01}, journal = {EMBO J}, volume = {12}, number = {2}, pages = {607-16}, abstract = {From in vitro translation studies we have previously demonstrated the existence of an apparent efficient UAG (amber) suppressor tRNA in the dimorphic fungus Candida albicans (Santos et al., 1990). Using an in vitro assay for termination codon readthrough the tRNA responsible was purified to homogeneity from C.albicans cells. The determined sequence of the purified tRNA predicts a 5'-CAG-3' anticodon that should decode the leucine codon CUG and not the UAG termination codon as originally hypothesized. However, the tRNA(CAG) sequence shows greater nucleotide homology with seryl-tRNAs from the closely related yeast Saccharomyces cerevisiae than with leucyl-tRNAs from the same species. In vitro tRNA-charging studies demonstrated that the purified tRNA(CAG) is charged with Ser. The gene encoding the tRNA was cloned from C.albicans by a PCR-based strategy and DNA sequence analysis confirmed both the structure of the tRNA(CAG) and the absence of any introns in the tRNA gene. The copy number of the tRNA(CAG) gene (1-2 genes per haploid genome) is in agreement with the relatively low abundance (< 0.5% total tRNA) of this tRNA. In vitro translation studies revealed that the purified tRNA(CAG) could induce apparent translational bypass of all three termination codons. However, peptide mapping of in vitro translation products demonstrated that the tRNA(CAG) induces translational misreading in the amino-terminal region of two RNA templates employed, namely the rabbit alpha- and beta-globin mRNAs. These results suggest that the C.albicans tRNA(CAG) is not an 'omnipotent' suppressor tRNA but rather may mediate a novel non-standard translational event in vitro during the translation of the CUG codon. The possible nature of this non-standard translation event is discussed in the context of both the unusual structural features of the tRNA(CAG) and its in vitro behaviour.}, note = {0261-4189 Journal Article}, keywords = {*Anticodon, *Translation, &, Acid, albicans/*genetics, Base, Candida, Cloning, Conformation, Data, DNA, Fungal, Fungal/chemistry/genetics/isolation, Genes, Genetic, Gov't, Leucine/*genetics, Molecular, Non-U.S., Nucleic, purification, RNA, Sequence, Ser/chemistry/*genetics/isolation, Support, Transfer}, pubstate = {published}, tppubtype = {article} } @article{, title = {The primary structure of cytoplasmic initiator tRNA(Met) from Schizosaccharomyces pombe}, author = { G. Keith and J. Heitzler and C. el Adlouni and A. L. Glasser and C. Fix and J. Desgres and G. Dirheimer}, year = {1993}, date = {1993-01-01}, journal = {Nucleic Acids Res}, volume = {21}, number = {12}, pages = {2949}, note = {0305-1048 Journal Article}, keywords = {Base, Data, Fungal/*chemistry, Met/*chemistry, Molecular, RNA, Schizosaccharomyces/*genetics, Sequence, Transfer}, pubstate = {published}, tppubtype = {article} } @article{bulet_novel_1992, title = {A novel insect defensin mediates the inducible antibacterial activity in larvae of the dragonfly Aeschna cyanea (Paleoptera, Odonata)}, author = {Philippe Bulet and S Cociancich and M Reuland and F Sauber and R Bischoff and G Hegy and Van A Dorsselaer and Charles Hetru and Jules A Hoffmann}, issn = {0014-2956}, year = {1992}, date = {1992-11-01}, journal = {Eur. J. Biochem.}, volume = {209}, number = {3}, pages = {977--984}, abstract = {The injection of low doses of bacteria into the aquatic larvae of dragonflies (Aeschna cyanea, Odonata, Paleoptera) induces the appearance in their hemolymph of a potent antibacterial activity. We have isolated a 38-residue peptide from this hemolymph which is strongly active against Gram-positive bacteria and also shows activity against one of the Gram-negative bacteria which was tested. The peptide is a novel member of the insect defensin family of inducible antibacterial peptides, which had so far only been reported from the higher insect orders believed to have evolved 100 million years after the Paleoptera. Aeschna defensin is more potent than defensin from the dipteran Phormia, from which its structure differs in several interesting aspects, which are discussed in the paper.}, keywords = {Animals, Anti-Bacterial Agents, Antimicrobial Cationic Peptides, Blood Bactericidal Activity, Blood Proteins, Defensins, Hemolymph, hoffmann, Insect Proteins, insects, Larva, M3i, Mass Spectrometry, Peptides}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_insect_1992, title = {Insect defensins: inducible antibacterial peptides}, author = {Jules A Hoffmann and Charles Hetru}, doi = {10.1016/0167-5699(92)90092-L}, issn = {0167-5699}, year = {1992}, date = {1992-10-01}, journal = {Immunol. Today}, volume = {13}, number = {10}, pages = {411--415}, abstract = {In response to bacterial challenge or trauma, insects produce a battery of bactericidal or bacteriostatic molecules with a broad spectrum of activity against Gram-positive and/or Gram-negative bacteria; most are small-sized cationic peptides. This review focuses on insect defensins, a large group of inducible antibacterial peptides that are present both in ancient and recent insect orders. This immune response of insects shares many of the characteristics of the mammalian acute phase response.}, keywords = {Amino Acid, Animals, Bacterial Infections, Blood Bactericidal Activity, Blood Proteins, Defensins, hoffmann, insects, M3i, Peptides, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{reichhart_expression_1992, title = {Expression and secretion of active insect defensin, an inducible antibacterial peptide from the fleshfly Phormia terranovae.}, author = {Jean-Marc Reichhart and I Petit and M Legrain and Jean-Luc Dimarcq and E Keppi and J -P Lecocq and Jules A Hoffmann and T Achstetter}, year = {1992}, date = {1992-01-01}, journal = {Intern. J. of Invertebrate Reprod. and Develop.}, volume = {21}, pages = {15--24}, abstract = {Insect defensin A is an inducible antibacterial peptide of the fleshfly Phormia terranovae and has been recently characterized as a 40 residue basic peptide with six cysteines engaged in three intramolecular disulfide bridges. We report the expression of this peptide in Saccharomyces cerevisiae as a fusion protein carrying at its N-terminus leader sequences which are derived from the precursor of the yeast pheromone mating factor a (MFa1). These sequences allow the biologically active peptide to be secreted at high levels in a correctly processed form. Thus heterologous production of defensin A circumvents laborious purification of minute amounts of the peptide from its natural source. The insect defensin pro peptide can substitute for the MFa1 pro peptide, indicating that yeast can process an insect pro sequence.}, keywords = {hoffmann, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{ferrandon_localisation_1992, title = {The localisation of the maternal signals that determine the anterior-posterior axis in the Drosophila embryo}, author = {Dominique Ferrandon and St D Johnston}, year = {1992}, date = {1992-01-01}, journal = {Seminars in Developmental Biology}, volume = {3}, pages = {3--13}, keywords = {ferrandon, M3i}, pubstate = {published}, tppubtype = {article} } @article{reichhart_insect_1992, title = {Insect immunity: developmental and inducible activity of the Drosophila diptericin promoter}, author = {Jean-Marc Reichhart and Marie Meister and Jean-Luc Dimarcq and Daniel Zachary and Danièle Hoffmann and C Ruiz and G Richards and Jules A Hoffmann}, issn = {0261-4189}, year = {1992}, date = {1992-01-01}, journal = {EMBO J.}, volume = {11}, number = {4}, pages = {1469--1477}, abstract = {Diptericins are 9 kDa inducible antibacterial peptides initially isolated from immune haemolymph of Phormia (Diptera). Following the isolation of a Drosophila cDNA encoding a diptericin homologue, we have now cloned a genomic fragment containing the Drosophila diptericin gene. To dissect the regulation of this gene, we have transformed flies with a fusion gene in which the reporter beta-galactosidase gene is under the control of 2.2 kb upstream sequences of the diptericin gene. We show that such a fusion gene is inducible by injection of live bacteria or complete Freund's adjuvant and respects the tissue specific expression pattern of the resident diptericin gene. Our analysis reveals at least four distinct phases in the regulation of this gene: young larvae, late third instar larvae, pupae and adults. This complexity may be related to the presence in the upstream sequences of multiple copies of response elements previously characterized in genes encoding acute phase response proteins in mammals (e.g. NK-kappa B, NF-kappa B related, NF-IL6 response elements).}, keywords = {Acute-Phase Proteins, Adipose Tissue, Animals, Base Sequence, beta-Galactosidase, Embryo, Gene Expression Regulation, Genetic, hoffmann, Insect Hormones, Insect Proteins, M3i, Mammals, Nonmammalian, Oligodeoxyribonucleotides, Promoter Regions, Recombinant Fusion Proteins, reichhart, Restriction Mapping}, pubstate = {published}, tppubtype = {article} } @article{, title = {Presence and coding properties of 2'-O-methyl-5-carbamoylmethyluridine (ncm5Um) in the wobble position of the anticodon of tRNA(Leu) (U*AA) from brewer's yeast}, author = { A. L. Glasser and C. el Adlouni and G. Keith and E. Sochacka and A. Malkiewicz and M. Santos and M. F. Tuite and J. Desgres}, year = {1992}, date = {1992-01-01}, journal = {FEBS Lett}, volume = {314}, number = {3}, pages = {381-5}, abstract = {The unknown modified nucleoside U* has been isolated by enzymatic and HPLC protocols from tRNA(Leu) (U*AA) recently discovered in brewer's yeast. The pure U* nucleoside has been characterized by electron impact mass spectroscopy, and comparison of its chromatographic and UV-absorption properties with those of appropriate synthetic compounds. The structure of U* was established as 2'-O-methyl-5-carbamoylmethyluridine (ncm5Um). The yeast tRNA(Leu) (U*AA) is the only tRNA so far sequenced which has been shown to contain ncm5Um. The location of such a modified uridine at the first position of the anticodon restricts the decoding property to A of the leucine UUA codon.}, note = {0014-5793 Journal Article}, keywords = {*Anticodon, &, Analysis, cerevisiae/*genetics, Chromatography, derivatives/analysis/chemistry/genetics, Fungal, Fungal/genetics, Gov't, high, Leu/*genetics, liquid, Mass, Molecular, Non-U.S., Pressure, Proteins/biosynthesis, RNA, Saccharomyces, Spectrophotometry, Spectrum, structure, Support, Transfer, Ultraviolet, Uridine/*analogs}, pubstate = {published}, tppubtype = {article} } @article{bonmatin_two-dimensional_1992, title = {Two-dimensional 1H NMR study of recombinant insect defensin A in water: resonance assignments, secondary structure and global folding}, author = {J M Bonmatin and J L Bonnat and X Gallet and F Vovelle and M Ptak and Jean-Marc Reichhart and Jules A Hoffmann and E Keppi and M Legrain and T Achstetter}, issn = {0925-2738}, year = {1992}, date = {1992-01-01}, journal = {J. Biomol. NMR}, volume = {2}, number = {3}, pages = {235--256}, abstract = {A 500 MHz 2D 1H NMR study of recombinant insect defensin A is reported. This defense protein of 40 residues contains 3 disulfide bridges, is positively charged and exhibits antibacterial properties. 2D NMR maps of recombinant defensin A were fully assigned and secondary structure elements were localized. The set of NOE connectivities, 3JNH-alpha H coupling constants as well as 1H/2H exchange rates and delta delta/delta T temperature coefficients of NH protons strongly support the existence of an alpha-helix (residues 14-24) and of an antiparallel beta-sheet (residues 27-40). Models of the backbone folding were generated by using the DISMAN program and energy refined by using the AMBER program. This was done on the basis of: (i) 133 selected NOEs, (ii) 21 dihedral restraints from 3JNH-alpha H coupling constants, (iii) 12 hydrogen bonds mostly deduced from 1H/2H exchange rates or temperature coefficients, in addition to 9 initial disulfide bridge covalent constraints. The two secondary structure elements and the two bends connecting them involve approximately 70% of the total number of residues, which impose some stability in the C-terminal part of the molecule. The remaining N-terminal fragment forms a less well defined loop. This spatial organization, in which a beta-sheet is linked to an alpha-helix by two disulfide bridges and to a large loop by a third disulfide bridge, is rather similar to that found in scorpion charybdotoxin and seems to be partly present in several invertebrate toxins.}, keywords = {Animals, Defensins, hoffmann, Hydrogen, Insect Hormones, insects, M3i, Magnetic Resonance Spectroscopy, Models, Molecular, Protein Conformation, Recombinant Proteins, reichhart, Saccharomyces cerevisiae, Thermodynamics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural specificity of Rn nuclease I as probed on yeast tRNA(Phe) and tRNA(Asp)}, author = { A. Przykorska and C. el Adlouni and G. Keith and J. W. Szarkowski and G. Dirheimer}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {4}, pages = {659-63}, abstract = {A single-strand-specific nuclease from rye germ (Rn nuclease I) was characterized as a tool for secondary and tertiary structure investigation of RNAs. To test the procedure, yeast tRNA(Phe) and tRNA(Asp) for which the tertiary structures are known, as well as the 3'-half of tRNA(Asp) were used as substrates. In tRNA(Phe) the nuclease introduced main primary cuts at positions U33 and A35 of the anticodon loop and G18 and G19 of the D loop. No primary cuts were observed within the double stranded stems. In tRNA(Asp) the main cuts occurred at positions U33, G34, U35, C36 of the anticodon loop and G18 and C20:1 positions in the D loop. No cuts were observed in the T loop in intact tRNA(Asp) but strong primary cleavages occurred at positions psi 55, C56, A57 within that loop in the absence of the tertiary interactions between T and D loops (use of 3'-half tRNA(Asp)). These results show that Rn nuclease I is specific for exposed single-stranded regions.}, note = {0305-1048 Journal Article}, keywords = {Acid, Asp/chemistry/genetics/*metabolism, Base, cereale, Composition, Conformation, Data, Gov't, Molecular, Non-U.S., Nucleic, Pancreatic/*metabolism, Phe/chemistry/genetics/*metabolism, Ribonuclease, RNA, Secale, Sequence, Specificity, Substrate, Support, Transfer, Yeasts/genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Use of a dot blot hybridization method for identification of pure tRNA species on different membranes}, author = { J. Heitzler and L. Marechal-Drouard and G. Dirheimer and G. Keith}, year = {1992}, date = {1992-01-01}, journal = {Biochim Biophys Acta-Gene Regul Mech}, volume = {1129}, number = {3}, pages = {273-7}, abstract = {The characterization of a tRNA in purification procedures usually involves aminoacylation assays but recently, the hybridization by dot blot with specific oligonucleotides as probes has been used for the tRNA identification. We present here an optimization of a dot blot hybridization method for the tRNA detection by comparing the efficiency of eight different nylon membranes. Neutral 0.22 microns porosity membranes (Nytran, Biodine A) give the best detection efficiency when small quantities of material (less than 40 ng of tRNA) are dotted on filter; by contrast, neutral 0.45 microns porosity membranes (such as Hybond N) are the most efficient when larger quantities of tRNA are dotted on the filter. The described technique allows to detect less than 20 pg of a pure tRNA species. Its use in the identification of Saccharomyces cerevisiae initiator tRNA(Met) in counter-current distribution fractions is shown.}, note = {0006-3002 Journal Article}, keywords = {*Membranes, Acid, Artificial, Autoradiography, cerevisiae/genetics, Fungal/genetics, Gov't, Hybridization, Met/genetics, Non-U.S., Nucleic, RNA, Saccharomyces, Support, Transfer, Transfer/*genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pleiotropic effect of a point mutation in the yeast SUP4-o tRNA gene: in vivo pre-tRNA processing in S. cerevisiae}, author = {M L Wilhelm and G Keith and C Fix and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1542574}, isbn = {1542574}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {4}, pages = {791-796}, abstract = {The expression of mutant tyrosine-inserting ochre suppressor SUP4-o tRNA genes in vivo in S. cerevisiae was examined as a basis for further studies of tRNA transcription and processing. In vivo yeast precursor tRNAs have been identified by filter hybridization and primer extension analysis. We have previously shown that a mutant SUP4-o tRNA gene with a C52----A52 transversion at positive 52 (C52----A52(+IVS) allele) was transcribed but that the primary transcript was not processed correctly. We show here that 5' and 3' end processing as well as splicing are defective for this mutant but that the 5' end processing is restored when the intron is removed from the gene by oligonucleotide directed mutagenesis (C52----A52(-IVS) allele). Our results imply that the C52----A52 transversion by itself cannot account for the lack of susceptibility to RNase P cleavage but that the overall tertiary structure of the mutant tRNA precursor is destabilized by the intron/anticodon stem. A second consequence of the C52----A52 transversion is to prevent complete maturation of the tRNA precursor at its 3' end since intermediates containing incompletely processed 3' trailers accumulate in the yeast cells transformed with the C52----A52(-IVS) allele. A correct structure of the T stem might therefore define a structural feature required for the recognition of the 3' processing activity.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Blotting, Fungal/*genetics Genes, Fungal/genetics/metabolism RNA, Northern Genes, Suppressor/*genetics Introns/genetics Molecular Sequence Data Mutation/genetics RNA Precursors/*metabolism RNA, Transfer, Tyr/*genetics/metabolism Saccharomyces cerevisiae/*genetics/metabolism, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Rapid transfer of small RNAs from a polyacrylamide gel onto a nylon membrane using a gel dryer}, author = {M L Wilhelm and W Baranowski and G Keith and F X Wilhelm}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1508703}, isbn = {1508703}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {15}, pages = {4106}, note = {0305-1048 Journal Article}, keywords = {5S/*isolation & purification RNA, Acrylic Resins Human Nylons RNA, Ribosomal, Small Nuclear/*isolation & purification RNA, Transfer/*isolation & purification Yeasts/genetics, Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Some tertiary motifs of RNA foldings}, author = {E Westhof and F Michel}, editor = {D M J Lilley and H Heumann and D Suck}, url = {http://www.amazon.com/Structural-Analysis-Protein-Nucleic-Complexes-Advances/dp/0817627766}, year = {1992}, date = {1992-01-01}, booktitle = {Structural Tools for the Analysis of Protein-Nucleic Acid Complexes (Advances in Life Sciences)}, pages = {255-267}, publisher = {Birkhauser Verlag, Basel}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {RNA pseudoknots}, author = {E Westhof and L Jaeger}, url = {http://www.sciencedirect.com/science/article/pii/0959440X9290221R}, doi = {10.1016/0959-440X(92)90221-R}, year = {1992}, date = {1992-01-01}, journal = {Curr Opin Struct Biol}, volume = {2}, number = {3}, pages = {327-333}, abstract = {RNA pseudoknots result from Watson-Crick base pairing involving a stretch of bases located between paired strands and a distal single-stranded region. Recently, significant advances in our understanding of their structural and functional aspects have been accomplished. At the structural level, modelling and NMR studies have shown that a defined subset of pseudoknots may be considered as tertiary motifs in RNA foldings. At the functional level, there is evidence that the realm of functions encompassed by RNA pseudoknots extends from the control of translation in prokaryotes, retroviruses and coronaviruses to the control of catalytic activity in ribozymes and the control of replication in some plant viruses.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Westhof's rule}, author = {E Westhof}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1641036}, isbn = {1641036}, year = {1992}, date = {1992-01-01}, journal = {Nature}, volume = {358}, number = {6386}, pages = {459-460}, note = {0028-0836 Letter}, keywords = {*Hydrogen Bonding *Nucleic Acid Conformation, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effect of dimerization on the conformation of the encapsidation Psi domain of Moloney murine leukemia virus RNA}, author = {N Tounekti and M Mougel and C Roy and R Marquet and J L Darlix and J Paoletti and B Ehresmann and C Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1731069}, isbn = {1731069}, year = {1992}, date = {1992-01-01}, journal = {J Mol Biol}, volume = {223}, number = {1}, pages = {205-220}, abstract = {In Moloney murine leukemia virus, the encapsidation Psi element was shown to be necessary and sufficient to promote packaging of viral RNA, and to be required for dimerization. The conformation of the Psi domain (nucleotides 215 to 565) was investigated in solution by chemical probing. The four bases were monitored at one of their Watson-Crick positions with dimethylsulfate at cytosine N3 and adenosine N1, and with a carbodiimide derivative at guanosine N1 and uridine N3. Position N7 of adenine residues was probed with diethylpyrocarbonate. The analyses were conducted on in vitro transcribed fragments corresponding either to the isolated Psi domain or to the 5'-terminal 725 nucleotides. The RNA fragments were analyzed in their monomeric and dimeric forms. A secondary structure model was derived from probing data, computer prediction and sequence analysis of related murine retroviruses. One major result is that Psi forms an independent and highly structured domain. Dimerization induces an extensive reduction of reactivity in region 278 to 309 that can be interpreted as the result of intermolecular interactions and/or intramolecular conformational rearrangements. A second region (around position 215) was shown to display discrete reactivity changes upon dimerization. These two regions represent likely elements of dimerization. More unexpectedly, reactivity changes (essentially enhancement of reactivity) were also detected in another part of Psi (around position 480) not believed to contain elements of dimerization. These reactivity changes could be interpreted as dimerization-induced allosteric transitions.}, note = {0022-2836 Journal Article}, keywords = {Base Sequence Comparative Study Hydrogen Bonding Macromolecular Systems Molecular Sequence Data Molecular Structure Moloney murine leukemia virus/*ultrastructure Nucleic Acid Conformation Phylogeny RNA, MARQUET, Non-U.S. Gov't, Unité ARN, Viral/chemistry/*ultrastructure Sequence Alignment Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {An additional long-range interaction in human U1 snRNA}, author = {C Sturchler and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1532853}, isbn = {1532853}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {6}, pages = {1215-1221}, abstract = {We present evidence for the existence of an additional long-range interaction in vertebrate U1 snRNAs. By submitting human U1 snRNP, HeLa nuclear extracts, authentic human or X. laevis in vitro transcribed U1 snRNAs to RNase V1, a nuclease specific for double-stranded regions, cleavages occurred in the sequence psi psi ACC (positions 5-9) residing in the 5' terminal region of the RNA. The RNase V1 sensitive region is insensitive to single-stranded probes, something unexpected knowing that it was considered single-stranded in order to base-pair to pre-mRNA 5' splice site. We have identified the sequence GGUAG (positions 132-136) as the only possible 3' partner. Mutants, either abolishing or restoring the interaction between the partners, coupled to an RNase V1 assay, served to substantiate this base-pairing model. The presence of this additional helix, even detected in nuclear extracts under in vitro splicing conditions, implies that a conformational change must occur to release a free U1 snRNA 5' end.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Binding Sites Hela Cells Human Molecular Sequence Data Mutagenesis, Site-Directed Nucleic Acid Conformation RNA, Small Nuclear, Small Nuclear/*chemistry/metabolism Ribonucleases/metabolism Ribonucleoproteins/metabolism Ribonucleoproteins, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Interaction between lysozyme molecules under precrystallization conditions studied by light scattering.}, author = {M Skouri and J P Munch and B Lorber and R Giege and S Candau}, url = {http://www.sciencedirect.com/science/article/pii/0022024892902214}, isbn = {10.1016/0022-0248(92)90221-4}, year = {1992}, date = {1992-01-01}, journal = {J Crystal Growth}, volume = {122}, number = {1-4}, pages = {14-20}, abstract = {Lightscattering experiments were undertaken on lysozymeunder various solvent conditions. When the protein is undersaturated, attractive interparticular interactions are detected. They are enhanced when the temperature is decreased, but are much weaker in NaCl solutions in which the protein crystallizes than in ammonium sulfate solutions in which it forms amorphous precipitates. When the protein in a NaCl solution is brought to supersaturation by a temperature decrease, lightscattering measurements indicate the simultaneous presence of two scatter populations which can be assimilated to individual lysozymemolecules and to large particles. Kinetic experiments in which the temperature is quenched rapidly indicate that the apparent hydrodynamic radius of the large particles increases regularly with time up to a plateau value of about 2600A˚.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The anticodon triplet is not sufficient to confer methionine acceptance to a transfer RNA}, author = {B Senger and L Despons and P Walter and F Fasiolo}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1438273}, isbn = {1438273}, year = {1992}, date = {1992-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {89}, number = {22}, pages = {10768-10771}, abstract = {Previous work suggested that the presence of the anticodon CAU alone was enough to confer methionine acceptance to a tRNA. Conversions of Escherichia coli nonmethionine tRNAs to a methionine-accepting species were obtained by substitutions reconstructing the whole methionine anticodon loop together with preservation (or introduction) of the acceptor stem base A73. We show here that the CAU triplet alone is unable to confer methionine acceptance when transplanted into a yeast aspartic tRNA. Both non-anticodon bases of the anticodon loop of yeast tRNA(Met) and A73 are required in addition to CAU for methionine acceptance. The importance of these non-anticodon bases in other CAU-containing tRNA frameworks was also established. These specific non-anticodon base interactions make a substantial thermodynamic contribution to the methionine acceptance of a transfer RNA.}, note = {0027-8424 Journal Article}, keywords = {Anticodon/genetics/*metabolism Base Sequence Kinetics Methionine/*metabolism Models, Genetic, Met/genetics/*metabolism Saccharomyces cerevisiae/*genetics Support, Non-U.S. Gov't Transcription, Structural Molecular Sequence Data Nucleic Acid Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Determinant nucleotides of yeast tRNA(Asp) interact directly with aspartyl-tRNA synthetase}, author = {J Rudinger and J D Puglisi and J Putz and D Schatz and F Eckstein and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1631068}, isbn = {1631068}, year = {1992}, date = {1992-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {89}, number = {13}, pages = {5882-5886}, abstract = {The interaction of wild-type and mutant yeast tRNA(Asp) transcripts with yeast aspartyl-tRNA synthetase (AspRS; EC 6.1.1.12) has been probed by using iodine cleavage of phosphorothioate-substituted transcripts. AspRS protects phosphates in the anticodon (G34, U35), D-stem (U25), and acceptor end (G73) that correspond to determinant nucleotides for aspartylation. This protection, as well as that in anticodon stem (C29, U40, G41) and D-stem (U11 to U13), is consistent with direct interaction of AspRS at these phosphates. Other protection, in the variable loop (G45), D-loop (G18, G19), and T-stem and loop (G53, U54, U55), as well as enhanced reactivity at G37, may result from conformational changes of the transcript upon binding to AspRS. Transcripts mutated at determinant positions showed a loss of phosphate protection in the region of the mutation while maintaining the global protection pattern. The ensemble of results suggests that aspartylation specificity arises from both protein-base and protein-phosphate contacts and that different regions of tRNA(Asp) interact independently with AspRS. A mutant transcript of yeast tRNA(Phe) that contains the set of identity nucleotides for specific aspartylation gave a phosphate protection pattern strikingly similar to that of wild-type tRNA(Asp). This confirms that a small number of nucleotides within a different tRNA sequence context can direct specific interaction with synthetase.}, note = {0027-8424 Journal Article}, keywords = {Asp/chemistry/*metabolism Saccharomyces cerevisiae/enzymology Structure-Activity Relationship Support, Aspartate-tRNA Ligase/*metabolism Base Sequence Binding Sites DNA Mutational Analysis Fungal Proteins/metabolism Molecular Sequence Data Protein Binding RNA, FLORENTZ, Fungal/chemistry/metabolism RNA, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Efficient mischarging of a viral tRNA-like structure and aminoacylation of a minihelix containing a pseudoknot: histidinylation of turnip yellow mosaic virus RNA}, author = {J Rudinger and C Florentz and T Dreher and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1579487}, isbn = {1579487}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {8}, pages = {1865-1870}, abstract = {Mischarging of the valine specific tRNA-like structure of turnip yellow mosaic virus (TYMV) RNA has been tested in the presence of purified arginyl-, aspartyl-, histidinyl-, and phenylalanyl-tRNA synthetases from bakers' yeast. Important mischarging of a 264 nucleotide-long transcript was found with histidinyl-tRNA synthetase which can acylate this fragment up to a level of 25% with a loss of specificity (expressed as Vmax/KM ratios) of only 100 fold as compared to a yeast tRNA(His) transcript. Experiments on transcripts of various lengths indicate that the minimal valylatable fragment (n = 88) is the most efficient substrate for histidinyl-tRNA synthetase, with kinetic characteristics similar to those found for the control tRNA(His) transcript. Mutations in the anticodon or adjacent to the 3' CCA that severely affect the valylation capacity of the 264 nucleotide long TYMV fragment are without negative effect on its mischarging, and for some cases even improve its efficiency. A short fragment (n = 42) of the viral RNA containing the pseudoknot and corresponding to the amino acid accepting branch of the molecule is an efficient histidine acceptor.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence Codon/genetics Histidine-tRNA Ligase/*metabolism Kinetics Molecular Sequence Data Mosaic Viruses/enzymology/genetics/*metabolism Mutation/genetics Nucleic Acid Conformation RNA, FLORENTZ, His/*metabolism RNA, Non-P.H.S. Yeasts/enzymology, Non-U.S. Gov't Support, Transfer, U.S. Gov't, Unité ARN, Viral/*metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Molecular mimicry in translational control of E. coli threonyl-tRNA synthetase gene. Competitive inhibition in tRNA aminoacylation and operator-repressor recognition switch using tRNA identity rules}, author = {P Romby and C Brunel and J Caillet and M Springer and M Grunberg-Manago and E Westhof and C Ehresmann and B Ehresmann}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1280807}, isbn = {1280807}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {21}, pages = {5633-5640}, abstract = {We previously showed that: (i) E.coli threonyl-tRNA synthetase (ThrRS) binds to the leader of its mRNA and represses translation by preventing ribosome binding to its loading site; (ii) the translational operator shares sequence and structure similarities with tRNA(Thr); (iii) it is possible to switch the specificity of the translational control from ThrRS to methionyl-tRNA synthetase (MetRS) by changing the CGU anticodon-like sequence to CAU, the tRNA(Met) anticodon. Here, we show that the wild type (CGU) and the mutated (CAU) operators act as competitive inhibitors of tRNA(Thr) and tRNA(fMet) for aminoacylation catalyzed by E.coli ThrRS and MetRS, respectively. The apparent Kd of the MetRS/CAU operator complex is one order magnitude higher than that of the ThrRS/CGU operator complex. Although ThrRS and MetRS shield the anticodon- and acceptor-like domains of their respective operators, the relative contribution of these two domains differs significantly. As in the threonine system, the interaction of MetRS with the CAU operator occludes ribosome binding to its loading site. The present data demonstrate that the anticodon-like sequence is one major determinant for the identity of the operator and the regulation specificity. It further shows that the tRNA-like operator obeys to tRNA identity rules.}, note = {0305-1048 Journal Article}, keywords = {Acylation Anticodon Base Sequence Binding, Bacterial Methionine-tRNA Ligase/metabolism Molecular Sequence Data Mutation Nucleic Acid Conformation *Operator Regions (Genetics) RNA, Bacterial/metabolism RNA, Competitive Escherichia coli/enzymology/*genetics *Gene Expression Regulation, Genetic, Met/metabolism RNA, Non-U.S. Gov't Threonine-tRNA Ligase/antagonists & inhibitors/*genetics/metabolism Translation, ROMBY, Thr/metabolism Repressor Proteins/*metabolism Ribosomes/metabolism Support, Transfer, Transfer/*metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Alternating d(G-A) sequences form a parallel-stranded DNA homoduplex}, author = {K Rippe and V Fritsch and E Westhof and T M Jovin}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1396571}, isbn = {1396571}, year = {1992}, date = {1992-01-01}, journal = {EMBO J}, volume = {11}, number = {10}, pages = {3777-3786}, abstract = {The oligonucleotides d[(G-A)7G] and d[(G-A)12G] self-associate under physiological conditions (10 mM MgCl2, neutral pH) into a stable double-helical structure (psRR-DNA) in which the two polypurine strands are in a parallel orientation in contrast to the antiparallel disposition of conventional B-DNA. We have characterized psRR-DNA by gel electrophoresis, UV absorption, vacuum UV circular dichroism, monomer-excimer fluorescence of oligonucleotides end-labelled with pyrene, and chemical probing with diethyl pyrocarbonate and dimethyl sulfate. The duplex is stable at pH 4-9, suggesting that the structure is compatible with, but does not require, protonation of the A residues. The data support a model derived from force-field analysis in which the parallel-stranded d(G-A)n helix is right-handed and constituted of alternating, symmetrical Gsyn.Gsyn and Aanti.Aanti base pairs with N1H.O6 and N6H.N7 hydrogen bonds, respectively. This dinucleotide structure may be the source of a negative peak observed at 190 nm in the vacuum UV CD spectrum, a feature previously reported only for left-handed Z-DNA. The related sequence d[(GAAGGA)4G] also forms a parallel-stranded duplex but one that is less stable and probably involves a slightly different secondary structure. We discuss the potential intervention of psRR-DNA in recombination, gene expression and the stabilization of genomic structure.}, note = {0261-4189 Journal Article}, keywords = {Base Sequence Circular Dichroism DNA/*chemistry Hydrogen Bonding Hydrogen-Ion Concentration Indicators and Reagents Magnesium Chloride Models, Fluorescence Spectrophotometry, MARQUET, Molecular Molecular Sequence Data *Nucleic Acid Conformation Oligodeoxyribonucleotides/*chemistry Spectrometry, PAILLART, Ultraviolet Structure-Activity Relationship Thermodynamics, Unité ARN, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {Structural specificity of Rn nuclease I as probed on yeast tRNA(Phe) and tRNA(Asp)}, author = {A Przykorska and C el Adlouni and G Keith and J W Szarkowski and G Dirheimer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1542562}, isbn = {1542562}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {4}, pages = {659-663}, abstract = {A single-strand-specific nuclease from rye germ (Rn nuclease I) was characterized as a tool for secondary and tertiary structure investigation of RNAs. To test the procedure, yeast tRNA(Phe) and tRNA(Asp) for which the tertiary structures are known, as well as the 3'-half of tRNA(Asp) were used as substrates. In tRNA(Phe) the nuclease introduced main primary cuts at positions U33 and A35 of the anticodon loop and G18 and G19 of the D loop. No primary cuts were observed within the double stranded stems. In tRNA(Asp) the main cuts occurred at positions U33, G34, U35, C36 of the anticodon loop and G18 and C20:1 positions in the D loop. No cuts were observed in the T loop in intact tRNA(Asp) but strong primary cleavages occurred at positions psi 55, C56, A57 within that loop in the absence of the tertiary interactions between T and D loops (use of 3'-half tRNA(Asp)). These results show that Rn nuclease I is specific for exposed single-stranded regions.}, note = {0305-1048 Journal Article}, keywords = {Asp/chemistry/genetics/*metabolism RNA, Base Composition Base Sequence Molecular Sequence Data Nucleic Acid Conformation RNA, Non-U.S. Gov't Yeasts/genetics, Pancreatic/*metabolism Secale cereale Substrate Specificity Support, Phe/chemistry/genetics/*metabolism Ribonuclease, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Effect of conformational features on the aminoacylation of tRNAs and consequences on the permutation of tRNA specificities}, author = {V Perret and C Florentz and J D Puglisi and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1640453}, isbn = {1640453}, year = {1992}, date = {1992-01-01}, journal = {J Mol Biol}, volume = {226}, number = {2}, pages = {323-333}, abstract = {The structure and function of in vitro transcribed tRNA(Asp) variants with inserted conformational features characteristic of yeast tRNA(Phe), such as the length of the variable region or the arrangement of the conserved residues in the D-loop, have been investigated. Although they exhibit significant conformational alterations as revealed by Pb2+ treatment, these variants are still efficiently aspartylated by yeast aspartyl-tRNA synthetase. Thus, this synthetase can accommodate a variety of tRNA conformers. In a second series of variants, the identity determinants of yeast tRNA(Phe) were transplanted into the previous structural variants of tRNA(Asp). The phenylalanine acceptance of these variants improves with increasing the number of structural characteristics of tRNA(Phe), suggesting that phenylalanyl-tRNA synthetase is sensitive to the conformational frame embedding the cognate identity nucleotides. These results contrast with the efficient transplantation of tRNA(Asp) identity elements into yeast tRNA(Phe). This indicates that synthetases respond differently to the detailed conformation of their tRNA substrates. Efficient aminoacylation is not only dependent on the presence of the set of identity nucleotides, but also on a precise conformation of the tRNA.}, note = {0022-2836 Journal Article}, keywords = {Amino Acid Activation Aspartate-tRNA Ligase/*metabolism Base Sequence Molecular Sequence Data Nucleic Acid Conformation Phenylalanine-tRNA Ligase/*metabolism RNA, Asp/metabolism/*ultrastructure RNA, FLORENTZ, Fungal/metabolism/ultrastructure RNA, Non-U.S. Gov't, Phe/metabolism/*ultrastructure Saccharomyces cerevisiae Structure-Activity Relationship Substrate Specificity Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The initiation accuracy of the SV40 early transcription is determined by the functional domains of two TATA elements}, author = {M Pauly and M Treger and E Westhof and P Chambon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1312710}, isbn = {1312710}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {5}, pages = {975-982}, abstract = {To locate the boundaries of the TATA element in the SV40 early promoter, point mutations have been constructed such as to cover the whole T + A-rich region of the replication origin. The effects of these mutations on the rate of transcription in vivo show that this region actually contains two TATA elements I and II, each independently directing the accurate initiation of transcription from a specified set of start sites, EES1 and EES2, respectively. The sequence of TATA element I fits best with the compiled 'consensus' sequence found in eukaryotic gene promoters and is the most efficient in directing transcription initiation. Mutations which improve this fit can still increase the rate of transcription, confirming the theory of a correlation between the nucleotide sequence of a TATA element and its functional efficiency. Moreover, some mutations which simultaneously modify the angle of DNA curvature in the T + A-rich promoter region and the rate of transcription reveal a correlation between DNA bending and transcription initiation.}, note = {0305-1048 Journal Article}, keywords = {Base Sequence DNA Mutational Analysis DNA, Genetic/*genetics, Non-U.S. Gov't TATA Box/*genetics Transcription, Unité ARN, Viral/*genetics Hela Cells Human Molecular Sequence Data Nucleic Acid Conformation Simian virus 40/*genetics Support, Viral/chemistry/genetics Electrophoresis Genes, WESTHOF}, pubstate = {published}, tppubtype = {article} } @article{, title = {The major RNA in prosomes of HeLa cells and duck erythroblasts is tRNA(Lys,3)}, author = {H G Nothwang and O Coux and G Keith and I Silva-Pereira and K Scherrer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1579498}, isbn = {1579498}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {8}, pages = {1959-1965}, abstract = {Two-dimensional gel electrophoresis of HeLa cell prosomal RNAs, 3'-end labeled by RNA ligase, revealed one prominent spot. Determination of a partial sequence at the 3'-end indicated full homology to the 18 nucleotides at the 3'-end of tRNA(Lys,3) from rabbit, the bovine and the human species. An oligonucleotide complementary to the 3'-end of tRNA(Lys,3) hybridized on Northern blots with prosomal RNA from both HeLa cells and duck erythroblasts. In two-dimensional PAGE, the major pRNA of HeLa cells co-migrated with bovine tRNA(Lys,3). Reconstitution of the CCA 3'-end of RNA from both human and duck prosomes, by tRNA-nucleotidyl-transferase, confirmed the tRNA character of this type of RNA. Furthermore, it revealed at least one additional tRNA band about 85 nt long among the prosomal RNA from both species. Finally, confirming an original property of prosomal RNA, we show that in vitro synthesized tRNA(Lys,3) hybridizes stably to duck globin mRNA, and to poly(A)(+)- and poly(A)(-)-RNA from HeLa cells.}, note = {0305-1048 Journal Article}, keywords = {Animals Base Sequence Blotting, Gel, Lys/*analysis/metabolism Ribonucleoproteins/*chemistry/drug effects Support, Non-U.S. Gov't Zinc/pharmacology, Northern Ducks Electrophoresis, Transfer, Two-Dimensional Erythroblasts Hela Cells Human Molecular Sequence Data RNA Nucleotidyltransferases/metabolism RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Optimal tRNA((Ser)Sec) gene activity requires an upstream SPH motif}, author = {E Myslinski and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1311068}, isbn = {1311068}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {2}, pages = {203-209}, abstract = {The X. laevis tRNA((Ser)Sec) gene is different from the other tRNA genes in that its promoter contains two external elements, a PSE and a TATA box functionally equivalent to those of the U6 snRNA gene. Of the two internal promoters governing classical tRNA gene transcription, only subsists the internal B box. In this report, we show that the tRNA((Ser)Sec) contains in addition an activator element (AE) which we have mapped by extensive mutagenesis. Activation is only dependent on a 15 bp fragment residing between -209 and -195 and containing an SPH motif. In vitro, this element forms a complex with a nuclear protein which is different from the TEF-1 transcriptional activator that binds the SV40 Sph motifs. This AE is versatile since it shows capacity of activating a variety of genes in vivo, including U1 and U6 snRNAs and HSV thymidine kinase. Unexpectedly for an snRNA-related gene, the tRNA((Ser)Sec) is deprived of octamer or octamer-like motifs. The X.laevis tRNA((Ser)Sec) gene represents the first example of a Pol III snRNA-type gene whose activation of transcription is completely octamer-independent.}, note = {0305-1048 Journal Article}, keywords = {Amino Acid-Specific/*genetics Recombinant Fusion Proteins/genetics/metabolism Simian virus 40/*genetics Support, Animals Base Sequence DNA Mutational Analysis DNA-Binding Proteins/genetics Enhancer Elements (Genetics)/*genetics/physiology Gene Expression Regulation/*genetics Molecular Sequence Data Promoter Regions (Genetics)/genetics RNA, Non-U.S. Gov't TATA Box/genetics Xenopus laevis/*genetics, Small Nuclear/genetics RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The differential transcriptional activity of two amphibian U1 small-nuclear RNA genes correlates with structural differences in the proximal sequence element}, author = {S Murgo and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1735429}, isbn = {1735429}, year = {1992}, date = {1992-01-01}, journal = {Eur J Biochem}, volume = {203}, number = {3}, pages = {443-447}, abstract = {We previously analyzed the transcription of an axolotl U1 small-nuclear RNA (snRNA) gene (AmU1) by microinjection into Xenopus laevis oocytes. In such an assay, AmU1 showed a low template activity compared to that of an X. laevis U1 snRNA gene (XlU1B2). Swapping the proximal sequence element (PSE) with that of XlU1B2 was required for AmU1 to acquire a transcription level equal to that of XlU1B2. In the present work, we examine the functional importance of the nucleotides that are common or different in both PSEs with the aim of identifying which nucleotides within the Xenopus U1 PSE are critical for this enhancement of Ambystoma mexicanum U1 snRNA transcription. The PSE mutation analysis showed that the central, phylogenetically conserved C-58/C-57 doublet is absolutely required for U1 promoter activity. In the 3' portion of this element, a CGC to ATG change (positions -54/-52) which partially restores the XlU1B2 PSE sequence, enables the AmU1 gene to gain the same transcriptional activity as XlU1B2. Remarkably, in this clustered point mutation, the sole C-54 to A-54 change is sufficient to obtain this increased level. Therefore, the activity of the AmU1 gene in injected Xenopus oocytes is strongly affected by a single sequence difference between AmU1 and XlU1B2 PSEs. This finding underscores the crucial importance of the nucleotide identity at position -54 to the function of the Xenopus U1 PSE.}, note = {0014-2956 Journal Article}, keywords = {Ambystoma Animals Base Sequence Microinjections Molecular Sequence Data Mutagenesis, Genetic Xenopus laevis, Non-U.S. Gov't *Transcription, Nucleic Acid Sequence Homology, Nucleic Acid Support, Site-Directed Phylogeny RNA, Small Nuclear/*genetics *Regulatory Sequences, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Activation of the catalytic core of a group I intron by a remote 3' splice junction}, author = {F Michel and L Jaeger and E Westhof and R Kuras and F Tihy and M Q Xu and D A Shub}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1644285}, isbn = {1644285}, year = {1992}, date = {1992-01-01}, journal = {Genes Dev}, volume = {6}, number = {8}, pages = {1373-1385}, abstract = {Over 1000 nucleotides may separate the ribozyme core of some group I introns from their 3' splice junctions. Using the sunY intron of bacteriophage T4 as a model system, we have investigated the mechanisms by which proximal splicing events are suppressed in vitro, as well as in vivo. Exon ligation as well as cleavage at the 5' splice site are shown to require long-range pairing between one of the peripheral components of the ribozyme core and some of the nucleotides preceding the authentic 3' splice junction. Consistent with our three-dimensional modeling of the entire sunY ribozyme, we propose that this novel interaction is necessary to drive 5' exon-core transcripts into an active conformation. A requirement for additional stabilizing interactions, either RNA-based or mediated by proteins, appears to be a general feature of group I self-splicing. A role for these interactions in mediating putative alternative splicing events is discussed.}, note = {0890-9369 Journal Article}, keywords = {Base Sequence DNA Mutational Analysis Escherichia coli/genetics Genes, Catalytic/genetics/*metabolism RNA, Messenger/genetics/*metabolism Support, P.H.S. T-Phages/genetics, U.S. Gov't, Unité ARN, Viral/*genetics Introns/genetics/*physiology Magnesium/metabolism Molecular Sequence Data Mutation/genetics Nucleic Acid Conformation Plasmids/genetics RNA Splicing/*genetics RNA}, pubstate = {published}, tppubtype = {article} } @article{, title = {Preparative isolation of angiotensin-converting enzyme from human lung.}, author = {Q C Meng and S J King and K E Branham and L J DeLucas and B Lorber and S Oparil}, url = {http://www.ncbi.nlm.nih.gov/pubmed/1332983}, year = {1992}, date = {1992-01-01}, journal = {J Chromatogr}, volume = {579}, number = {1}, pages = {63-71}, abstract = {Angiotensin-converting enzyme from human lung was purified to apparent homogeneity using a five-step purification procedure consisting of ammonium sulfate precipitation, ion-exchange chromatography on DEAE Sephadex A-50, gel permeation on Sephadex G-200, chromatofocusing on a polybuffer exchange (PBE 94) column and high-performance liquid chromatographic gel permeation on a Bio-Sil TSK-250 column. This procedure gave an approximately 700-fold purification with a 20% yield compared to a 550-fold purification and a 1% yield with an affinity chromatography-based procedure. The 20-fold greater yield of the five-step procedure offers a major advantage for preparative use in the structural characterization of angiotensin-converting enzyme.}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {article} } @inbook{, title = {Preparation and handling of biological macromolecules for crystallization.}, author = {B Lorber and R Giege}, editor = {A Ducruix and R Giegé}, url = {http://www.worldcat.org/title/crystallization-of-nucleic-acids-and-proteins-a-practical-approach/oclc/24009434}, year = {1992}, date = {1992-01-01}, booktitle = {Crystallization of Nucleic Acids and Proteins: A Practical Approach}, pages = {19-45}, publisher = {Oxford University Press, USA}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {inbook} } @article{, title = {A versatile reactor for temperature controlled crystallization of biological macromolecules.}, author = {B Lorber and R Giege}, url = {http://www.sciencedirect.com/science/article/pii/002202489290240J}, isbn = {10.1016/0022-0248(92)90240-J}, year = {1992}, date = {1992-01-01}, journal = {J Crystal Growth}, volume = {122}, number = {1-4}, pages = {168-175}, abstract = {Accurate control of crystallization conditions, and accordingly reproducibility of experiments, is often hampered by the lack of adequate instrumentation in the laboratory. A versatile and inexpensive crystallization reactor allowing crystal growth by vapor phase equilibration (sitting, hanging or sandwiched drops) or by the batch technique is described here. In this reactor, temperature is controlled accurately by a Peltier device and the crystallization process monitored by a time-lapse video recorder. An application for evaluating the growth kinetics of tetragonal lysozyme crystals as a function of temperature is given. It was observed that the apparent growth rate decreased when temperature was lowered, although under such conditions supersaturation was increased and crystals appeared sooner.}, keywords = {FRUGIER, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {A factor with Sp1 DNA-binding specificity stimulates Xenopus U6 snRNA in vivo transcription by RNA polymerase III}, author = {A Lescure and G Tebb and I W Mattaj and A Krol and P Carbon}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1453450}, isbn = {1453450}, year = {1992}, date = {1992-01-01}, journal = {J Mol Biol}, volume = {228}, number = {2}, pages = {387-394}, abstract = {We have previously shown that transcription of the Xenopus U6 snRNA gene by RNA polymerase III is stimulated in injected Xenopus oocytes by an activator element termed the DSE, which contains an octamer sequence. Data presented here reveal that the DSE contains, in addition, a GC-rich sequence capable of binding Sp1. Both elements are required to obtain wild-type levels of U6 transcription in vivo. The Xenopus U6 DSE exhibits optimal activation properties only when positioned at its normal location upstream from the start site. The U6 Sp1 motif binds the mammalian Sp1 transcriptional activator independently of the Oct-1 protein in vitro. Those mutations that lead to a reduced transcription level in vivo abolish the binding of Sp1 in vitro. Thus, transcriptional stimulation through the Xenopus U6 Sp1 motif is likely to be mediated by a protein with DNA-binding specificity identical to mammalian Sp1. These findings support the notion that RNA polymerase II and III transcription complexes share transactivators.}, note = {0022-2836 Journal Article}, keywords = {Animals Base Sequence Binding Sites Dna Molecular Sequence Data RNA Polymerase III/*metabolism RNA, Genetic Xenopus, LESCURE, Non-U.S. Gov't Transcription Factor, Nucleic Acid Support, Small Nuclear/*genetics *Regulatory Sequences, Sp1/metabolism *Transcription, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The proximal promoter and the start site cooperate to specify correct U1 snRNA transcription initiation by RNA polymerase II}, author = {A Lescure and S Murgo and P Carbon and A Krol}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1579449}, isbn = {1579449}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {7}, pages = {1573-1578}, abstract = {In this work, we attempted to gain insight into the detailed mechanism allowing correct transcription initiation of U1 snRNA genes by RNA polymerase II. Abolition of the CA motif residing at -1/+1 in the Xenopus U1 gene leads to a loss of the ability of the promoter to direct accurate initiation. A discrete site is selected only if a purine preceded by a pyrimidine is positioned at 58/57 bp downstream of the center of the PSE. The PSE alone is unable to designate a discrete initiation site. Rather, it serves to set the location of an initiation window without discriminating suitable from unsuitable initiation sites. The latter role is devoted to a PyPu sequence positioned at -1/+1. Therefore, it is the concomitant action of the PSE and an essential PyPu positioned at the proper distance from this promoter that specifies correct U1 snRNA transcription initiation by RNA polymerase II.}, note = {0305-1048 Journal Article}, keywords = {Animals Base Sequence DNA/metabolism DNA Mutational Analysis Molecular Sequence Data Promoter Regions (Genetics)/*genetics RNA Polymerase II/*metabolism RNA, Genetic/*genetics Xenopus laevis/genetics, KROL, LESCURE, Non-U.S. Gov't Transcription, Small Nuclear/*genetics Support, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Inhibition of murine leukemia viruses by nuclease-resistant alpha-oligonucleotides}, author = {M Lavignon and N Tounekti and B Rayner and J L Imbach and G Keith and J Paoletti and C Malvy}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1292779}, isbn = {1292779}, year = {1992}, date = {1992-01-01}, journal = {Antisense Res Dev}, volume = {2}, number = {4}, pages = {315-324}, abstract = {We studied the antiviral activity of nuclease-resistant alpha-anomeric oligonucleotides. An alpha-oligonucleotide (20-mer) targeted to the primer binding site (PBS) of murine retroviruses inhibited viral spreading. The inhibition only occurred when the cells had been electropermeabilized in the presence of the oligonucleotide. The PBS sequence is involved in reverse transcription and in translation. The data suggest that the oligonucleotide could perturb reverse transcription activity. Thus, either the oligonucleotide induced a decrease in initiation or it inhibited the extension of the minus or plus strands DNA during reverse transcription. These results show that reverse transcription may be an interesting target for antisense oligonucleotides.}, note = {1050-5261 Journal Article}, keywords = {3T3 Cells Animals Base Sequence Binding Sites Culture Media Friend murine leukemia virus/*drug effects/genetics/physiology Mice Molecular Sequence Data Moloney murine leukemia virus/*drug effects/genetics/physiology Oligonucleotides, Antisense/metabolism/*pharmacology RNA, Genetic/drug effects, Genetic/drug effects Translation, Messenger/chemistry/genetics/metabolism RNA, Non-U.S. Gov't Transcription, Unité ARN, Viral/chemistry/genetics/metabolism Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Use of a dot blot hybridization method for identification of pure tRNA species on different membranes}, author = {J Heitzler and L Marechal-Drouard and G Dirheimer and G Keith}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1536878}, isbn = {1536878}, year = {1992}, date = {1992-01-01}, journal = {Biochim Biophys Acta-Gene Regul Mech}, volume = {1129}, number = {3}, pages = {273-277}, abstract = {The characterization of a tRNA in purification procedures usually involves aminoacylation assays but recently, the hybridization by dot blot with specific oligonucleotides as probes has been used for the tRNA identification. We present here an optimization of a dot blot hybridization method for the tRNA detection by comparing the efficiency of eight different nylon membranes. Neutral 0.22 microns porosity membranes (Nytran, Biodine A) give the best detection efficiency when small quantities of material (less than 40 ng of tRNA) are dotted on filter; by contrast, neutral 0.45 microns porosity membranes (such as Hybond N) are the most efficient when larger quantities of tRNA are dotted on the filter. The described technique allows to detect less than 20 pg of a pure tRNA species. Its use in the identification of Saccharomyces cerevisiae initiator tRNA(Met) in counter-current distribution fractions is shown.}, note = {0006-3002 Journal Article}, keywords = {Artificial Nucleic Acid Hybridization RNA, Autoradiography *Membranes, Fungal/genetics RNA, Met/genetics Saccharomyces cerevisiae/genetics Support, Non-U.S. Gov't, Transfer, Transfer/*genetics RNA, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {The specificity of translational control switched with transfer RNA identity rules}, author = {M Graffe and J Dondon and J Caillet and P Romby and C Ehresmann and B Ehresmann and M Springer}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1372129}, isbn = {1372129}, year = {1992}, date = {1992-01-01}, journal = {Science}, volume = {255}, number = {5047}, pages = {994-996}, abstract = {The interaction of Escherichia coli threonyl-transfer RNA (tRNA) synthetase with the leader sequence of its own messenger RNA inhibits ribosome binding, resulting in negative translational feedback regulation. The leader sequence resembles the substrate (tRNA(Thr)) of the enzyme, and the nucleotides that mediate the correct recognition of the leader and the tRNA may be the same. A mutation suggested by tRNA identity rules that switches the resemblance of the leader sequence from tRNA(Thr) to tRNA(Met) causes the translation of the threonyl-tRNA synthetase messenger RNA to become regulated by methionyl-tRNA synthetase. This identity swap in the leader messenger RNA indicates that tRNA identity rules may be extended to interactions of synthetases with other RNAs.}, note = {0036-8075 Journal Article}, keywords = {Bacterial Genes, Bacterial Molecular Sequence Data Nucleic Acid Conformation RNA, Bacterial Proteins/metabolism Base Sequence DNA Mutational Analysis *Gene Expression Regulation, Bacterial/metabolism RNA, Genetic, Messenger/*metabolism/ultrastructure RNA, Non-U.S. Gov't Threonine-tRNA Ligase/*genetics/metabolism *Translation, ROMBY, Structural, Thr/*metabolism Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Presence and coding properties of 2'-O-methyl-5-carbamoylmethyluridine (ncm5Um) in the wobble position of the anticodon of tRNA(Leu) (U*AA) from brewer's yeast}, author = {A L Glasser and C el Adlouni and G Keith and E Sochacka and A Malkiewicz and M Santos and M F Tuite and J Desgres}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1468572}, isbn = {1468572}, year = {1992}, date = {1992-01-01}, journal = {FEBS Lett}, volume = {314}, number = {3}, pages = {381-385}, abstract = {The unknown modified nucleoside U* has been isolated by enzymatic and HPLC protocols from tRNA(Leu) (U*AA) recently discovered in brewer's yeast. The pure U* nucleoside has been characterized by electron impact mass spectroscopy, and comparison of its chromatographic and UV-absorption properties with those of appropriate synthetic compounds. The structure of U* was established as 2'-O-methyl-5-carbamoylmethyluridine (ncm5Um). The yeast tRNA(Leu) (U*AA) is the only tRNA so far sequenced which has been shown to contain ncm5Um. The location of such a modified uridine at the first position of the anticodon restricts the decoding property to A of the leucine UUA codon.}, note = {0014-5793 Journal Article}, keywords = {*Anticodon Chromatography, Fungal/genetics RNA, High Pressure Liquid Fungal Proteins/biosynthesis Molecular Structure RNA, Leu/*genetics Saccharomyces cerevisiae/*genetics Spectrophotometry, Mass Support, Non-U.S. Gov't Uridine/*analogs & derivatives/analysis/chemistry/genetics, Transfer, Ultraviolet Spectrum Analysis, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Footprinting evidence for close contacts of the yeast tRNA(Asp) anticodon region with aspartyl-tRNA synthetase}, author = {A Garcia and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1497679}, isbn = {1497679}, year = {1992}, date = {1992-01-01}, journal = {Biochem Biophys Res Commun}, volume = {186}, number = {2}, pages = {956-962}, abstract = {Chemical footprinting experiments on brewer's yeast tRNA(Asp) complexed to its cognate aspartyl-tRNA synthetase are reported: they demonstrate that bases of the anticodon loop, including the anticodon itself, are in close proximity with the synthetase. Contacts were determined using dimethylsulfate as the probe for testing reactivity of guanine and cytosine residues in free and complexed tRNA. Results correlate with the decrease in aspartylation activity of yeast tRNA(Asp) molecules mutated at these contact positions and will be compared with other structural data arising from solution and crystallographic studies on the aspartic acid complex.}, note = {0006-291x Journal Article}, keywords = {Alkylation Anticodon/*metabolism Aspartate-tRNA Ligase/*metabolism Base Sequence Molecular Sequence Data Nucleic Acid Conformation Protein Binding RNA, Asp/genetics/isolation & purification/*metabolism Saccharomyces cerevisiae/enzymology/*genetics Sulfuric Acid Esters/metabolism/pharmacology Support, Non-U.S. Gov't, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Anticodon-independent aminoacylation of an RNA minihelix with valine}, author = {M Frugier and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1570324}, isbn = {1570324}, year = {1992}, date = {1992-01-01}, journal = {Proc Natl Acad Sci U S A}, volume = {89}, number = {9}, pages = {3990-3994}, abstract = {Minihelices mimicking the amino acid acceptor and anticodon branches of yeast tRNA(Val) have been synthesized by in vitro transcription of synthetic templates. It is shown that a minihelix corresponding to the amino acid acceptor branch and containing solely a valine-specific identity nucleotide can be aminoacylated by yeast valyl-tRNA synthetase. Its charging ability is lost after mutating this nucleotide. This ability is stimulated somewhat by the addition of a second hairpin helix that mimicks the anticodon arm, which suggests that information originating from the anticodon stem-loop can be transmitted to the active site of the enzyme by the core of the protein.}, note = {0027-8424 Journal Article}, keywords = {*Amino Acid Activation Anticodon Base Sequence Hydrogen Bonding In Vitro Molecular Sequence Data RNA, ERIANI, FLORENTZ, FRUGIER, Non-U.S. Gov't Valine-tRNA Ligase/*metabolism, Transfer, Unité ARN, Val/chemistry/*metabolism Saccharomyces cerevisiae Structure-Activity Relationship Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Crystallization of aspartyl-tRNA synthetase-tRNA(Asp) complex from Escherichia coli and first crystallographic results}, author = {S Eiler and M Boeglin and F Martin and G Eriani and J Gangloff and J C Thierry and D Moras}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1569573}, isbn = {1569573}, year = {1992}, date = {1992-01-01}, journal = {J Mol Biol}, volume = {224}, number = {4}, pages = {1171-1173}, abstract = {Crystals of the dimeric aspartyl-tRNA synthetase from Escherichia coli (molecular mass 132,000 Da) complexed with its cognate tRNA (molecular mass 25,000 Da) have been grown using ammonium sulfate as precipitant. The crystals belong to the orthorhombic space group C222(1) with unit cell parameters a = 102.75 A}, note = {0022-2836 Journal Article}, keywords = {Asp/*ultrastructure X-Ray Diffraction, Aspartate-tRNA Ligase/*ultrastructure Crystallography Escherichia coli/enzymology RNA, ERIANI, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @book{, title = {Crystallization of Nucleic Acids and Proteins: A Practical Approach Series (1nd edition).}, editor = {A Ducruix and R Giege}, url = {http://www.worldcat.org/title/crystallization-of-nucleic-acids-and-proteins-a-practical-approach/oclc/24009434}, issn = {ISSN}, year = {1992}, date = {1992-01-01}, publisher = {Oxford University Press}, address = {USA}, series = {The practical approach series}, keywords = {Unité ARN}, pubstate = {published}, tppubtype = {book} } @article{, title = {Specific valylation of turnip yellow mosaic virus RNA by wheat germ valyl-tRNA synthetase determined by three anticodon loop nucleotides}, author = {T W Dreher and C H Tsai and C Florentz and R Giege}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1390705}, isbn = {1390705}, year = {1992}, date = {1992-01-01}, journal = {Biochemistry}, volume = {31}, number = {38}, pages = {9183-9189}, abstract = {The valylation by wheat germ valyl-tRNA synthetase of anticodon loop mutants of turnip yellow mosaic virus RNA has been studied. RNA substrates 264 nucleotides long were made by T7 RNA polymerase from cDNA encompassing the 3' tRNA-like region of genomic RNA. Substitution singly, or in combination, of three nucleotides in the anticodon loop resulted in very poor valylation (Vmax/KM less than 10(-3) relative to wild type). These nucleotides thus represent the major valine identity determinants recognized by wheat germ valyl-tRNA synthetase; their relative contribution to valine identity, in descending order, was as follows: the middle nucleotide of the anticodon (A56 in TYMV RNA), the 3' anticodon nucleotide (C55), and the 3'-most anticodon loop nucleotide (C53). Substitutions in the wobble position (C57) had no significant effect on valylation kinetics, while substitutions of the discriminator base (A4) resulted in small decreases in Vmax/Km. Mutations in the major identity nucleotides resulted in large increases in KM, suggesting that wheat germ valyl-tRNA synthetase has a lowered affinity for variant substrates with low valine identity. Comparison with other studies using valyl-tRNA synthetases from Escherichia coli and yeast indicates that the anticodon has been phylogenetically conserved as the dominant valine identity region, while the identity contribution of the discriminator base has been less conserved. The mechanism by which anticodon mutations are discriminated also appears to vary, being affinity-based for the wheat germ enzyme, and kinetically-based for the yeast enzyme [Florentz et al. (1991) Eur. J. Biochem. 195, 229-234].}, note = {0006-2960 Journal Article}, keywords = {Anticodon/genetics/*metabolism Bacteriophage T7/enzymology Base Sequence DNA-Directed RNA Polymerases/metabolism Kinetics Molecular Sequence Data Mosaic Viruses/genetics/*metabolism Nucleic Acid Conformation RNA, FLORENTZ, Genetic Triticum/*enzymology Valine-tRNA Ligase/*metabolism Variation (Genetics), Non-P.H.S. Support, Non-U.S. Gov't Support, P.H.S. Transcription, Transfer/chemistry/metabolism RNA, U.S. Gov't, Unité ARN, Viral/chemistry/genetics/*metabolism Seeds/enzymology Support}, pubstate = {published}, tppubtype = {article} } @article{, title = {Binding of the yeast tRNA(Met) anticodon by the cognate methionyl-tRNA synthetase involves at least two independent peptide regions}, author = {L Despons and B Senger and F Fasiolo and P Walter}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1602489}, isbn = {1602489}, year = {1992}, date = {1992-01-01}, journal = {J Mol Biol}, volume = {225}, number = {3}, pages = {897-907}, abstract = {As for Escherichia coli methionine tRNAs, the anticodon triplet of yeast tRNA(Met) plays an important role in the recognition by the yeast methionyl-tRNA synthetase (MetRS), indicating that this determinant for methionine identity is conserved in yeast. Efficient aminoacylation of the E. coli tRNA(Met) transcript by the heterologous yeast methionine enzyme also suggests conservation of the protein determinants that interact with the CAU anticodon sequence. We have analysed by site-directed mutagenesis the peptide region 655 to 663 of the yeast MetRS that is equivalent to the anticodon binding region of the E. coli methionine enzyme. Only one change, converting Leu658 into Ala significantly reduced tRNA aminoacylation. Semi-conservative substitutions of L658 allow a correlation to be drawn between side-chain volume of the hydrophobic residue at this site and activity. The analysis of the L658A mutant shows that Km is mainly affected. This suggests that the peptide region 655 to 663 contributes partially to the binding of the anticodon, since separate mutational analysis of the anticodon bases shows that kcat is the most critical parameter in the recognition of tRNA(Met) by the yeast synthetase. We have analysed the role of peptide region (583-GNLVNR-588) that is spatially close to the region 655 to 663. Replacements of residues N584 and R588 reduces significantly the kcat of aminoacylation. The peptide region 583-GNLVNR-588 is highly conserved in all MetRS so far sequenced. We therefore propose that the hydrogen donor/acceptor amino acid residues within this region are the most critical protein determinants for the positive selection of the methionine tRNAs.}, note = {0022-2836 Journal Article}, keywords = {Amino Acid Sequence Anticodon/*metabolism Binding Sites Kinetics Methionine-tRNA Ligase/*metabolism Models, Met/*metabolism Saccharomyces cerevisiae/enzymology Structure-Activity Relationship Support, Molecular Molecular Sequence Data Mutagenesis, Non-U.S. Gov't, Site-Directed Protein Conformation RNA, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Domains of the Escherichia coli threonyl-tRNA synthetase translational operator and their relation to threonine tRNA isoacceptors}, author = {C Brunel and J Caillet and P Lesage and M Graffe and J Dondon and H Moine and P Romby and C Ehresmann and B Ehresmann and M Grunberg-Manago and M Springer}, url = {http://www.ncbi.nlm.nih.gov/pubmed/1383551}, doi = {10.1016/0022-2836(92)90212-3}, isbn = {1383551}, year = {1992}, date = {1992-01-01}, journal = {J Mol Biol}, volume = {227}, number = {3}, pages = {621-634}, abstract = {The expression of the gene for threonyl-tRNA synthetase (thrS) is negatively autoregulated at the translational level in Escherichia coli. The synthetase binds to a region of the thrS leader mRNA upstream from the ribosomal binding site inhibiting subsequent translation. The leader mRNA consists of four structural domains. The present work shows that mutations in these four domains affect expression and/or regulation in different ways. Domain 1, the 3' end of the leader, contains the ribosomal binding site, which appears not to be essential for synthetase binding. Mutations in this domain probably affect regulation by changing the competition between the ribosome and the synthetase for binding to the leader. Domain 2, 3' from the ribosomal binding site, is a stem and loop with structural similarities to the tRNA(Thr) anticodon arm. In tRNAs the anticodon loop is seven nucleotides long, mutations that increase or decrease the length of the anticodon-like loop of domain 2 from seven nucleotides abolish control. The nucleotides in the second and third positions of the anticodon-like sequence are essential for recognition and the nucleotide in the wobble position is not, again like tRNA(Thr). The effect of mutations in domain 3 indicate that it acts as an articulation between domains 2 and 4. Domain 4 is a stable arm that has similarities to the acceptor arm of tRNA(Thr) and is shown to be necessary for regulation. Based on this mutational analysis and previous footprinting experiments, it appears that domains 2 and 4, those analogous to tRNA(Thr), are involved in binding the synthetase which inhibits translation probably by interfering with ribosome loading at the nearby translation initiation site.}, note = {0022-2836}, keywords = {Bacterial/genetics Gene Expression Regulation, Bacterial/genetics RNA, Base Sequence Escherichia coli/genetics Gene Expression Regulation, Enzymologic/*genetics Molecular Sequence Data Mutagenesis, Genetic/*genetics, Messenger/*genetics/metabolism RNA, Non-U.S. Gov't Threonine-tRNA Ligase/*genetics/metabolism Translation, ROMBY, Site-Directed/genetics Nucleic Acid Conformation RNA, Thr/*genetics/metabolism Recombinant Fusion Proteins/genetics Support, Transfer, Unité ARN}, pubstate = {published}, tppubtype = {article} } @article{, title = {Rapid transfer of small RNAs from a polyacrylamide gel onto a nylon membrane using a gel dryer}, author = { M. L. Wilhelm and W. Baranowski and G. Keith and F. X. Wilhelm}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {15}, pages = {4106}, note = {0305-1048 Journal Article}, keywords = {&, 5S/*isolation, Acrylic, Human, Nuclear/*isolation, Nylons, purification, Resins, Ribosomal, RNA, Small, Transfer/*isolation, Yeasts/genetics}, pubstate = {published}, tppubtype = {article} } @article{, title = {Pleiotropic effect of a point mutation in the yeast SUP4-o tRNA gene: in vivo pre-tRNA processing in S. cerevisiae}, author = { M. L. Wilhelm and G. Keith and C. Fix and F. X. Wilhelm}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {4}, pages = {791-6}, abstract = {The expression of mutant tyrosine-inserting ochre suppressor SUP4-o tRNA genes in vivo in S. cerevisiae was examined as a basis for further studies of tRNA transcription and processing. In vivo yeast precursor tRNAs have been identified by filter hybridization and primer extension analysis. We have previously shown that a mutant SUP4-o tRNA gene with a C52----A52 transversion at positive 52 (C52----A52(+IVS) allele) was transcribed but that the primary transcript was not processed correctly. We show here that 5' and 3' end processing as well as splicing are defective for this mutant but that the 5' end processing is restored when the intron is removed from the gene by oligonucleotide directed mutagenesis (C52----A52(-IVS) allele). Our results imply that the C52----A52 transversion by itself cannot account for the lack of susceptibility to RNase P cleavage but that the overall tertiary structure of the mutant tRNA precursor is destabilized by the intron/anticodon stem. A second consequence of the C52----A52 transversion is to prevent complete maturation of the tRNA precursor at its 3' end since intermediates containing incompletely processed 3' trailers accumulate in the yeast cells transformed with the C52----A52(-IVS) allele. A correct structure of the T stem might therefore define a structural feature required for the recognition of the 3' processing activity.}, note = {0305-1048 Journal Article}, keywords = {Base, Blotting, cerevisiae/*genetics/metabolism, Data, Fungal/*genetics, Fungal/genetics/metabolism, Genes, Introns/genetics, Molecular, Mutation/genetics, Northern, Precursors/*metabolism, RNA, Saccharomyces, Sequence, Suppressor/*genetics, Transfer, Tyr/*genetics/metabolism}, pubstate = {published}, tppubtype = {article} } @article{, title = {The major RNA in prosomes of HeLa cells and duck erythroblasts is tRNA(Lys,3)}, author = { H. G. Nothwang and O. Coux and G. Keith and I. Silva-Pereira and K. Scherrer}, year = {1992}, date = {1992-01-01}, journal = {Nucleic Acids Res}, volume = {20}, number = {8}, pages = {1959-65}, abstract = {Two-dimensional gel electrophoresis of HeLa cell prosomal RNAs, 3'-end labeled by RNA ligase, revealed one prominent spot. Determination of a partial sequence at the 3'-end indicated full homology to the 18 nucleotides at the 3'-end of tRNA(Lys,3) from rabbit, the bovine and the human species. An oligonucleotide complementary to the 3'-end of tRNA(Lys,3) hybridized on Northern blots with prosomal RNA from both HeLa cells and duck erythroblasts. In two-dimensional PAGE, the major pRNA of HeLa cells co-migrated with bovine tRNA(Lys,3). Reconstitution of the CCA 3'-end of RNA from both human and duck prosomes, by tRNA-nucleotidyl-transferase, confirmed the tRNA character of this type of RNA. Furthermore, it revealed at least one additional tRNA band about 85 nt long among the prosomal RNA from both species. Finally, confirming an original property of prosomal RNA, we show that in vitro synthesized tRNA(Lys,3) hybridizes stably to duck globin mRNA, and to poly(A)(+)- and poly(A)(-)-RNA from HeLa cells.}, note = {0305-1048 Journal Article}, keywords = {Animals, Base, Blotting, Cells, Data, Ducks, effects, Electrophoresis, Erythroblasts, Gel, Gov't, Hela, Human, Lys/*analysis/metabolism, Molecular, Non-U.S., Northern, Nucleotidyltransferases/metabolism, Ribonucleoproteins/*chemistry/drug, RNA, Sequence, Support, Transfer, Two-Dimensional, Zinc/pharmacology}, pubstate = {published}, tppubtype = {article} } @article{bulet_insect_1991, title = {Insect immunity. Isolation from a coleopteran insect of a novel inducible antibacterial peptide and of new members of the insect defensin family}, author = {Philippe Bulet and S Cociancich and Jean-Luc Dimarcq and J Lambert and Jean-Marc Reichhart and Danièle Hoffmann and Charles Hetru and Jules A Hoffmann}, issn = {0021-9258}, year = {1991}, date = {1991-12-01}, journal = {J. Biol. Chem.}, volume = {266}, number = {36}, pages = {24520--24525}, abstract = {Injection of heat-killed bacteria into larvae of the large tenebrionid beetle Zophobas atratus (Insecta, Endopterygota, Coleoptera) results in the appearance in the hemolymph of a potent antibacterial activity as evidenced by a plate growth inhibition assay. We have isolated three peptides (A-C) from this immune hemolymph which probably account for most of this activity. Their primary structures were established by a combination of peptide sequencing and molecular mass determination by mass spectrometry. Peptide A, which is bactericidal against Gram-negative cells, is a 74-residue glycine-rich molecule with no sequence homology to known peptides. We propose the name coleoptericin for this novel inducible antibacterial peptide. Peptides B and C are isoforms of a 43-residue peptide which contains 6 cysteines and shows significant sequence homology to insect defensins, initially reported from dipteran insects. This peptide is active against Gram-positive bacteria. The results are discussed in connection with recent studies on inducible antibacterial peptides present in the three other major orders of the endopterygote clade of insects: the Lepidoptera, Diptera, and Hymenoptera.}, keywords = {Animals, Antibody Formation, Beetles, Blood Bactericidal Activity, Blood Proteins, Chromatography, Defensins, Hemolymph, High Pressure Liquid, hoffmann, Insect Hormones, Insect Proteins, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{hipskind_ets-related_1991, title = {Ets-related protein Elk-1 is homologous to the c-fos regulatory factor p62TCF}, author = {R A Hipskind and V N Rao and C G Mueller and E S Reddy and A Nordheim}, doi = {10.1038/354531a0}, issn = {0028-0836}, year = {1991}, date = {1991-12-01}, journal = {Nature}, volume = {354}, number = {6354}, pages = {531--534}, abstract = {A key event in the response of cells to proliferative signals is the rapid, transient induction of the c-fos proto-oncogene, which is mediated through the serum response element (SRE) in the fos promoter. Genomic footprinting and transfection experiments suggest that this activation occurs through a ternary complex that includes the serum response factor (SRF) and the ternary complex factor p62. Interaction of p62TCF with the SRF-SRE binary complex requires a CAGGA tract immediately upstream of the SRE. Proteins of the ets proto-oncogene family bind to similar sequences and we have found that a member of this family, Elk-1, forms SRF-dependent ternary complexes with the SRE. Elk-1 and p62TCF have the same DNA sequence requirements and antibodies against Elk-1 block the binding of both proteins. Furthermore, we show that like p62TCF, Elk-1 forms complexes with the yeast SRF-homologue MCM1 but not with yeast ARG80. But ARG80 mutants that convey interaction with p62TCF can also form complexes with Elk-1. The similarity, or even identity, between Elk-1 and p62TCF suggests a novel regulatory role for Ets proteins that is effected through interaction with other proteins, such as SRF. Furthermore, the possible involvement of an Ets protein in the control of c-fos has interesting implications for proto-oncogene cooperation in cellular growth control.}, keywords = {Animals, Antibodies, Base Sequence, Binding Sites, DNA, DNA-Binding Proteins, Epitopes, Escherichia coli, ets-Domain Protein Elk-1, fos, Genes, Genetic, Immune Sera, Macromolecular Substances, Molecular Sequence Data, Mutagenesis, Nucleic Acid, Oligodeoxyribonucleotides, Oncogenic, Promoter Regions, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-ets, Proto-Oncogene Proteins c-fos, Proto-Oncogenes, Retroviridae Proteins, Saccharomyces cerevisiae, Sequence Homology, Site-Directed, Team-Mueller, Transcription Factors, Transfection}, pubstate = {published}, tppubtype = {article} } @article{mueller_protein_1991, title = {A protein domain conserved between yeast MCM1 and human SRF directs ternary complex formation}, author = {C G Mueller and A Nordheim}, issn = {0261-4189}, year = {1991}, date = {1991-12-01}, journal = {The EMBO journal}, volume = {10}, number = {13}, pages = {4219--4229}, abstract = {MCM1 and SRF bind to the same DNA sequence and form ternary complexes with STE12 and p62TCF, respectively. We show that in gel retardation assays, MCM1 recruits both ternary complex factors whereas SRF interacts only with p62TCF. A protein domain of 90 amino acids, shared by MCM1 and SRF, was found to be sufficient for ternary complex formation. The domain is also required for dimerization and DNA binding. Similar regions are found in other proteins, such as ARG80, Deficiens and Agamous. ARG80 and Agamous exhibit similar DNA binding specificities but do not interact with either STE12 or p62TCF. By exchanging three residues of ARG80 with those of corresponding positions in SRF (residues 198, 200 and 203), the ARG80 protein acquires the ability to recruit p62TCF into a ternary complex. Likewise, the substitution of four SRF amino acids by MCM1-derived residues (amino acids 73, 75, 77 and 78) confers on SRF the ability to interact with STE12. Thus, we have identified specific amino acids in MCM1 and SRF that are critical for ternary complex formation and which map to equivalent positions within the shared domains. Therefore, the structural basis for specific protein-protein interaction appears to be conserved in evolution between a class of transcription factors.}, keywords = {Amino Acid Sequence, Base Sequence, DNA, DNA-Binding Proteins, Fungal, Fungal Proteins, Humans, Minichromosome Maintenance 1 Protein, Molecular Sequence Data, Nuclear Proteins, Nucleic Acid, Plasmids, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Sequence Homology, Serum Response Factor, Team-Mueller, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @article{hetru_isolation_1991, title = {Isolation and structural characterization of an insulin-related molecule, a predominant neuropeptide from Locusta migratoria}, author = {Charles Hetru and K W Li and Philippe Bulet and Marie Lagueux and Jules A Hoffmann}, issn = {0014-2956}, year = {1991}, date = {1991-10-01}, journal = {Eur. J. Biochem.}, volume = {201}, number = {2}, pages = {495--499}, abstract = {Neurohaemal lobes of corpora cardiaca of Locusta migratoria are an established storage site for neurohormones produced by the neurosecretory cells of the brain. As previously reported [Hietter, H., Van Dorsselaer, A., Green, B., Denoroy, L., Hoffmann, J.A. & Luu, B. (1990) Eur. J. Biochem. 187, 241-247], the isolation and characterization of a novel 5-kDa peptide from these lobes served as the basis for oligonucleotide screening of cDNA libraries prepared from poly(A) RNA from neurosecretory cells of the central nervous system. From subsequent cDNA cloning studies [Lagueux, M., Lwoff, L., Meister, M., Goltzené, F. & Hoffmann, J.A. (1990) Eur. J. Biochem. 187, 249-254], the existence of a 145-residue precursor protein was deduced, which contained, in addition to the 5-kDa peptide, amino-acid sequences with homology to the A and B chains of an insulin-related peptide. In the present study we have isolated the native molecule from corpora cardiaca of Locusta and characterized, by Edman degradation and plasma-desorption mass spectrometry, the two chains as follows: A chain, Gly-Val-Phe-Asp-Glu-Cys-Cys-Arg-Lys-Ser-Cys-Ser-Ile-Ser-Glu-Leu-Gln-Thr- Tyr-Cys - Gly (Ile, isoleucine); B chain, Ser-Gly-Ala-Pro-Gln-Pro-Val-Ala-Arg-Tyr-Cys-Gly-Glu-Lys-Leu-Ser-Asn-Ala- Leu-Lys - Leu-Val-Cys-Arg-Gly-Asn-Tyr-Asn-Thr-Met-Phe. Taken in conjunction with the previous cloning studies, our data lead to a clear picture of the processing of Locusta preproinsulin. They indicate that locusta corpora cardiaca contain remarkably large amounts of one single insulin form, in contrast to multiple insulin isoforms of Bombyx mori, the only other insect species from which insulin-related peptides have been isolated and characterized [Nagasawa, H., Kataoka, H., Isogai, A., Tamura, S., Suzuki, A., Mizoguchi, A., Fujiwara, Y., Suzuki, A., Takahashi, S. & Ishizaki, H. (1986) Proc. Natl Acad. Sci. USA 83, 5840-5843].}, keywords = {Animals, Chromatography, DNA, Female, Grasshoppers, High Pressure Liquid, hoffmann, Insect Hormones, Insulin, M3i, Mass Spectrometry, Neuropeptides, Proinsulin, Protein Conformation}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_chinless_1991, title = {Chinless wonders. Molecular Aspects of Invertebrate Hormones. A Jacques Monod Conference sponsored by the Centre National de la Recherche Scientifique, Roscoff, France, October 14-19, 1990}, author = {Jules A Hoffmann and J Joosse}, issn = {1043-4674}, year = {1991}, date = {1991-04-01}, journal = {New Biol.}, volume = {3}, number = {4}, pages = {336--340}, keywords = {Animals, Crustacea, Ecdysone, hoffmann, insects, Invertebrate Hormones, Juvenile Hormones, M3i, Neuropeptides, Receptors, Snails, Steroid}, pubstate = {published}, tppubtype = {article} } @article{lepage_determination_1991, title = {Determination of disulfide bridges in natural and recombinant insect defensin A}, author = {P Lepage and F Bitsch and D Roecklin and E Keppi and Jean-Luc Dimarcq and Jean-Marc Reichhart and Jules A Hoffmann and C Roitsch and Van A Dorsselaer}, issn = {0014-2956}, year = {1991}, date = {1991-01-01}, journal = {Eur. J. Biochem.}, volume = {196}, number = {3}, pages = {735--742}, abstract = {The primary-structure comparison of natural insect defensin A from Phormia terranovae and recombinant insect defensin A from Saccharomyces cerevisiae has been accomplished using a combination of Edman degradation and liquid secondary ion mass spectrometry. The natural and recombinant proteins have the same primary structure with identical disulfide-bond designations (formula; see text) as determined from the peptides obtained after thermolysin digestion. The combined use of Edman degradation and mass spectometry allowed the disulfide-bridge structure to be determined with a total of only 40 micrograms (9.9 nmol) natural peptide. Mass spectrometry provides a rapid means of disulfide-bridge verification, requiring not more than 20 micrograms recombinant insect defensin A, which is compatible with use in batch analysis.}, keywords = {Animals, Blood Proteins, Defensins, Diptera, Disulfides, Hemolymph, hoffmann, M3i, Mass Spectrometry, Recombinant Proteins, reichhart}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_cellular_1990, title = {Cellular and molecular aspects of insect immunity}, author = {Danièle Hoffmann and Jules A Hoffmann}, issn = {0923-2494}, year = {1990}, date = {1990-12-01}, journal = {Res. Immunol.}, volume = {141}, number = {9}, pages = {895--896}, keywords = {Animals, Cellular, hoffmann, Immunity, insects, M3i, Molecular Biology}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_inducible_1990, title = {The inducible antibacterial peptides of dipteran insects}, author = {Jules A Hoffmann and Danièle Hoffmann}, issn = {0923-2494}, year = {1990}, date = {1990-12-01}, journal = {Res. Immunol.}, volume = {141}, number = {9}, pages = {910--918}, keywords = {Animals, Antimicrobial Cationic Peptides, Defensins, Diptera, Gene Expression Regulation, Genetic, hoffmann, Insect Hormones, Insect Proteins, M3i, Nucleic Acid, Proteins, Sequence Homology, Transcription}, pubstate = {published}, tppubtype = {article} } @article{dimarcq_insect_1990, title = {Insect immunity: expression of the two major inducible antibacterial peptides, defensin and diptericin, in Phormia terranovae}, author = {Jean-Luc Dimarcq and Daniel Zachary and Jules A Hoffmann and Danièle Hoffmann and Jean-Marc Reichhart}, issn = {0261-4189}, year = {1990}, date = {1990-08-01}, journal = {EMBO J.}, volume = {9}, number = {8}, pages = {2507--2515}, abstract = {Injections of low doses of bacteria into larvae of Phormia terranovae induce the appearance of potent bactericidal peptides in the blood, among which predominate the anti-Gram positive insect defensins and the anti-Gram negative diptericins. Insect defensins show significant homologies to mammalian (including human) microbicidal peptides present in polymorphonuclear leukocytes and macrophages. We report the molecular cloning of cDNAs and primer extension studies which indicate that insect defensin is produced as a prepro-peptide yielding mature defensin A (40 residues) after cleavage of a putative signal peptide (23 residues) and a prosequence (34 residues). Previous studies have established that diptericin (82 residues) is matured from a pre-peptide by cleavage of a putative signal peptide (19 residues) and C-terminal amidation. Using oligonucleotide probes complementary to the sequences of the mRNAs for defensin and diptericin, we show by in situ hybridization that both antibacterial peptides are concomitantly synthesized by the same cells: thrombocytoids, a specialized blood cell type, and adipocytes. Transcriptional studies based on hybridization of RNAs to cDNAs of defensin and diptericin indicate that the transcription of both genes is induced regardless of the nature of the stimulus (injection of Gram positive or Gram negative bacteria, lipopolysaccharides). Even a sterile injury applied to axenically raised larvae is efficient in inducing the transcription of both genes suggesting that the local disruption of the integument aspecifically initiates a signalling mechanism which the thrombocytoids and the adipocytes are able to interpret. The transcription of immune genes is relatively short lived and a second challenge yields a response similar to that of the first stimulus, indicating that the experimental insects do not keep a 'memory' of their first injection.}, keywords = {Animals, Anti-Bacterial Agents, Base Sequence, Blood Proteins, Cloning, Defensins, Diptera, Gene Expression, hoffmann, Insect Hormones, Insect Proteins, Larva, M3i, Molecular, Nucleic Acid Hybridization, Oligonucleotide Probes, Protein Conformation, reichhart}, pubstate = {published}, tppubtype = {article} } @article{schroter_synergism_1990, title = {Synergism in ternary complex formation between the dimeric glycoprotein p67SRF, polypeptide p62TCF and the c-fos serum response element}, author = {H Schröter and C G Mueller and K Meese and A Nordheim}, issn = {0261-4189}, year = {1990}, date = {1990-04-01}, journal = {The EMBO journal}, volume = {9}, number = {4}, pages = {1123--1130}, abstract = {Transcriptional regulation of the c-fos proto-oncogene requires the serum response element (SRE) which is complexed by a multi-protein assembly observed both in vitro and in vivo. Two protein factors, p67SRF and p62TCF (previously called p62), are required to interact with the SRE for efficient induction of c-fos by serum. By quantitative band shift electrophoresis we measure at least a 50-fold increase in SRE affinity for p67SRF/p62TCF over p67SRF alone. Stoichiometrically we determine that the ternary complex with p62TCF involves p67SRF in dimeric form. We demonstrate that p67SRF is a glycosylated nuclear transcription factor carrying terminal N-acetylglucosamine (GlcNAc) as a post-translational modification. A proteolytic limit digestion product, approximately 13 kd in size, was generated from the p67SRF-SRE complex. This p67SRF-core domain binds SRE, can dimerize with p67SRF and is still able to form a ternary complex with p62TCF. Therefore, three functional activities can be ascribed to this small p67SRF-core domain: specific DNA binding, dimerization and interaction with p62TCF. We demonstrate that these functions map within the p67SRF core fragment containing the region between amino acids 93 and 222.}, keywords = {Base Sequence, Chloroquine, Gene Expression Regulation, Genetic, Glycosylation, HeLa Cells, Humans, Kinetics, Macromolecular Substances, Molecular Sequence Data, Nuclear Proteins, Oligonucleotide Probes, Plasmids, Polymerase Chain Reaction, Protein-Tyrosine Kinases, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-fos, Proto-Oncogenes, Serum Response Factor, Team-Mueller, Transcription, Transcription Factors}, pubstate = {published}, tppubtype = {article} } @incollection{hoffmann_caracterisation_1990, title = {Caractérisation de neurohormones de Locusta migratoria : approches expérimentales, résultats et problèmes.}, author = {Jules A Hoffmann and Maurice Charlet and F Goltzené and F Holder and Marie Lagueux and L Lwoff and Marie Meister and C Melchior and Jean-Marc Reichhart and M Schneider and Daniel Zachary and H Hietter and B Luu}, year = {1990}, date = {1990-01-01}, booktitle = {Neurobiology and Endocrinology of Selected Invertebrates}, publisher = {Ed. Loughton BG & Saleuddin ASM}, address = {North York, Ontario}, edition = {Captus Press Inc.}, abstract = {A major objective of our research groups is the structural and functional characterization of neurohormones involved in the control of development and reproduction of Locusta (with emphasis on ecdysiotropic neurohormones). In the absence of reliable and reproducible microscale bioassays we have developed two strategies for the structural characterization of neurohormones. The first approach is based on the generation of a library of monoclonal antibodies directed against peptides of the neurohaemal lobes of the corpora cardiaca, which store the majority of the neurohormones synthesized in the neurosecretory cells of the brain. The antibody library is screened by immunocytochemistry of the neurosecretory cells of the brain : only those antibodies recognizing antigenic determinants within the neurosecretory granules of these cells are retained and serve in turn to secreen cDNA libraries prepared from brain neurosecretory cell mRNA in an expression vector. The second experimental strategy is based on microsequencing of the major peptides contained within the neurohaemal lobes of the corpora cardiaca. From the data obtained by microsequencing, oligonucleotide probes can be devised and used to secreen cDNA libraries from brain cells. The genes coding for the peptide can thus be characterized and their transcription can be followed during development and reproduction. It is obvious that the structural determinations, whether peptide or nucleotide sequences are generated, must be followed by functional studies. The production of the peptides, either by synthetic chemistry or by recombinant DNA technology, together with the corresponding antibodies, are essential tools for such studies.}, keywords = {hoffmann, M3i, reichhart}, pubstate = {published}, tppubtype = {incollection} } @conference{reichhart_expression_1990, title = {Expression and secretion of insect immune peptides in yeast.}, author = {Jean-Marc Reichhart and T Achstetter}, editor = {34th Forum Immunology in Res. in Immunol.}, year = {1990}, date = {1990-01-01}, booktitle = {Cellular and molecular aspects of insect immunity}, volume = {141}, pages = {943--946}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {conference} } @article{lagueux_cdnas_1990, title = {cDNAs from neurosecretory cells of brains of Locusta migratoria (Insecta, Orthoptera) encoding a novel member of the superfamily of insulins}, author = {Marie Lagueux and L Lwoff and Marie Meister and F Goltzené and Jules A Hoffmann}, issn = {0014-2956}, year = {1990}, date = {1990-01-01}, journal = {Eur. J. Biochem.}, volume = {187}, number = {1}, pages = {249--254}, abstract = {From neurohaemal lobes of corpora cardiaca of Locusta migratoria a 5-kDa peptide has been isolated and its sequence established [see the accompanying paper, by Hietter et al. (1990) Eur. J. Biochem. 187, 241-247]. We have designed oligonucleotide probes from the peptide sequence of this molecule and screened a library prepared from mRNA of the neurosecretory cell region of the brain of this insect. Several positive cDNAs were isolated, the combined nucleotide sequences of which predict a large precursor of 145 residues (15770 Da) containing the newly isolated 5-kDa peptide. The peptide is flanked by regions homologous to the A and B chains of the superfamily of insulins. The overall organization of the precursor is as follows: signal peptide/domain homologous to the B chain of insulins/C (connecting)-peptide (corresponding to the newly isolated 5-kDa peptide)/domain homologous to the A chain of insulins. The numbers and relative positions of the cysteines of the Locusta peptide are equivalent to those of the other members of the insulin superfamily and most of the hydrophobic core residues are conserved.}, keywords = {Animals, Base Sequence, DNA, Genes, Grasshoppers, hoffmann, Humans, Insulin, M3i, Multigene Family, Nervous System, Neuropeptides, Neurosecretory Systems, Nucleic Acid, Nucleic Acid Hybridization, Oligonucleotide Probes, RNA, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{wicker_insect_1990, title = {Insect immunity. Characterization of a Drosophila cDNA encoding a novel member of the diptericin family of immune peptides}, author = {C Wicker and Jean-Marc Reichhart and Danièle Hoffmann and D Hultmark and C Samakovlis and Jules A Hoffmann}, issn = {0021-9258}, year = {1990}, date = {1990-01-01}, journal = {J. Biol. Chem.}, volume = {265}, number = {36}, pages = {22493--22498}, abstract = {Drosophila shows an immune response when challenged by injection of low doses of bacteria. To date, the molecules involved in this immune reaction have remained elusive, with the exception of cecropins (4-kDa antibacterial peptides initially isolated from the moth Hyalophora cecropia) for which three closely related genes have been characterized recently. We report the molecular cloning and sequencing of a cDNA from a library of immune Drosophila which encodes a novel member of the family of diptericins (9-kDa antibacterial peptides initially isolated from the fly Phormia terranovae). Transcripts for the Drosophila diptericin are detected 2 h after injection of bacteria. They are apparently derived from a single gene mapping at position 56 A on the right arm of the second chromosome. We discuss the existence of a distant relationship between the diptericins and two other groups of anti-bacterial insect proteins, the attacins, and the sarcotoxins II.}, keywords = {Animals, Anti-Bacterial Agents, Base Sequence, Cloning, Diptera, DNA, Escherichia coli, hoffmann, Insect Hormones, Insect Proteins, M3i, Molecular, Multigene Family, Nucleic Acid, Oligonucleotide Probes, reichhart, Sequence Homology}, pubstate = {published}, tppubtype = {article} } @article{lambert_insect_1989, title = {Insect immunity. Isolation from immune blood of the Dipteran Phormia terranovae of two novel antibacterial peptides with sequence homology to rabbit lung macrophage bactericidal peptides.}, author = {J Lambert and E Keppi and Jean-Luc Dimarcq and C Wicker and Jean-Marc Reichhart and B Dunbar and P Lepage and Van A Dorsselaer and Jules A Hoffmann and J Forthergill and Danièle Hoffmann}, year = {1989}, date = {1989-01-01}, journal = {Proc. Nat. Acad. Sci. USA}, volume = {86}, pages = {262--266}, abstract = {We have isolated from the hemolymph of immunized larvae of the dipteran insect Phormia terranovae two novel peptides that are selectively active against Gram positive bacteria. They are positively charged peptides of 40 residues containing three intramolecular disulfide bridges, and differ from one another by only a single amino acid. These peptides are neither functionally nor structurally related to any known insect immune peptides, but show significant homology to microbicidal cationic peptides from mammalian granulocytes (defensins). We propose the name "insect defensins" for these novel insect antibiotic peptides.}, keywords = {hoffmann, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{reichhart_insect_1989, title = {Insect immunity. Isolation of cDNA clones corresponding to diptericin, an inducible antibacterial peptide from Phormia terranovae (Diptera). Transcriptional profiles during immunization}, author = {Jean-Marc Reichhart and M Essrich and Jean-Luc Dimarcq and Danièle Hoffmann and Jules A Hoffmann and Marie Lagueux}, issn = {0014-2956}, year = {1989}, date = {1989-01-01}, journal = {Eur. J. Biochem.}, volume = {182}, number = {2}, pages = {423--427}, abstract = {We have previously isolated and characterized a family of novel 8-kDa cationic antibacterial peptides synthesized by larvae of Phormia terranovae (Diptera) in response to various injuries. These molecules have been named diptericins. The peptide sequence of diptericin A was used to prepare oligonucleotides for screening cDNA libraries and we report in the present paper the isolation of several cDNA clones encoding diptericin. The analysis of the nucleotide sequences indicates that diptericin is synthesized as a prepeptide which is matured in two steps: (a) cleavage of a signal peptide and (b) amidation of the C-terminal residue. Interestingly, the 3' untranslated region of the mRNA contains a consensus sequence TTATTTAT which is also observed in the mRNA of another insect antibacterial peptide (attacin-related sarcotoxin IIA) and in mRNAs encoding proteins related to the inflammatory response in mammals. Our data illustrate that diptericins form a polymorphic family of immune peptides. The transcription of the diptericin genes is rapidly induced in the fat body after inoculation of bacteria, as evidenced by the transcriptional profile.}, keywords = {Animals, Anti-Bacterial Agents, Bacterial Proteins, Base Sequence, Blotting, Diptera, DNA, Endoribonucleases, Enterobacter, Enterobacteriaceae, Gene Expression Regulation, Genes, Genetic, hoffmann, Insect Hormones, Insect Proteins, M3i, messenger, MHC Class II, Northern, reichhart, Ribonuclease H, RNA, Transcription}, pubstate = {published}, tppubtype = {article} } @article{rosenthal_l-canavanine_1989, title = {L-canavanine incorporation into vitellogenin and macromolecular conformation}, author = {G A Rosenthal and Jean-Marc Reichhart and Jules A Hoffmann}, issn = {0021-9258}, year = {1989}, date = {1989-01-01}, journal = {J. Biol. Chem.}, volume = {264}, number = {23}, pages = {13693--13696}, abstract = {L-Canavanine is a potentially deleterious arginine antimetabolite whose toxicity is expressed in canavanine-sensitive organisms ranging from viruses to humans. Canavanine, a substrate for arginyl-tRNA synthetase, is incorporated into nascent polypeptide chains in place of arginine. This substitution results in the production of structurally aberrant, canavanyl proteins. Chemical, physical, and immunological studies of native and canavanine-containing vitellogenin obtained from female migratory locusts (Locusta migratoria migratorioides (Orthoptera] provide the first experimental evidence that canavanine can disrupt the tertiary and/or quaternary structure that yields the three-dimensional conformation unique to the protein. These findings enhance our understanding of the biochemical basis for canavanine's antimetabolic and potent insecticidal properties.}, keywords = {Animals, Antibodies, Canavanine, fluorescence, Grasshoppers, hoffmann, M3i, Monoclonal, Protein Conformation, reichhart, Spectrometry, Vitellogenins}, pubstate = {published}, tppubtype = {article} } @article{kappler_characterization_1988, title = {Characterization of three hydroxylases involved in the final steps of biosynthesis of the steroid hormone ecdysone in Locusta migratoria (Insecta, Orthoptera)}, author = {Christine Kappler and M Kabbouh and Charles Hetru and F Durst and Jules A Hoffmann}, issn = {0022-4731}, year = {1988}, date = {1988-12-01}, journal = {J. Steroid Biochem.}, volume = {31}, number = {6}, pages = {891--898}, abstract = {It is most generally accepted that the last three enzymatic reactions in the biosynthetic pathway of ecdysone are, in this order, the hydroxylations at positions C-25, C-22 and C-2. Using high specific activity tritiated ecdysone precursors (2,22,25-trideoxyecdysone, 2,22-dideoxyecdysone and 2-deoxyecdysone) we have characterized the hydroxylases involved in these reactions, in the major biosynthetic tissue of ecdysone, i.e. the prothoracic glands. We show that C-2 hydroxylase is a mitochondrial oxygenase which differs from conventional cytochrome P-450-dependent monooxygenases by its relative insensitivity to CO. In contrast, C-22 and C-25 hydroxylases appear as classical cytochrome P-450 monooxygenases; C-22 hydroxylase is a mitochondrial enzyme whereas our data point to a microsomal localization of the C-25 hydroxylase.}, keywords = {Animals, Biological, Ecdysone, Grasshoppers, hoffmann, Kinetics, M3i, Mixed Function Oxygenases, Models, NAD, NADP, Subcellular Fractions}, pubstate = {published}, tppubtype = {article} } @article{lanot_further_1988, title = {Further experimental evidence for the involvement of ecdysone in the control of meiotic reinitiation in oocytes of Locusta migratoria (Insecta, Orthoptera)}, author = {R Lanot and J Thiebold and M F Costet-Corio and P Benveniste and Jules A Hoffmann}, issn = {0012-1606}, year = {1988}, date = {1988-03-01}, journal = {Dev. Biol.}, volume = {126}, number = {1}, pages = {212--214}, abstract = {Ecdysone has recently been shown to be able to trigger meiotic reinitiation in vitro in submature oocytes of Locusta. In the present study we have experimentally depressed (by 60-70%) ecdysone biosynthesis in the ovaries of adult females by rearing them on a diet with a modified sterol profile. Mature oocytes from such females fail to undergo normal reinitiation, but when incubated in vitro, can be induced to break their meiotic arrest by the addition of exogenous ecdysone. These results lend further support to the hypothesis that in Locusta, ovarian ecdysone is involved in the control of meiotic reinitiation.}, keywords = {Animals, Ecdysone, Female, Grasshoppers, hoffmann, M3i, Meiosis, Oocytes, Ovary}, pubstate = {published}, tppubtype = {article} } @article{holder_production_1988, title = {Production of monoclonal antibodies against corpora cardiaca extracts of Locusta migratoria : a potential tool for the isolation of insect neurohormones.}, author = {F C Holder and L Lwoff and C Wicker and F Goltzené and Marie F Meister and Daniel Zachary and Jean-Marc Reichhart}, year = {1988}, date = {1988-01-01}, journal = {Int. J. Invert. Reprod. Develop.}, volume = {14}, pages = {105--118}, abstract = {Corpora cardiaca of Locusta migratoria contain the axon endings of the neurosecretory cells of the brain and store in neurosecretory granules a variety of mostly unidentified neurohormones. Homogenates of corpora cardiaca served to generate a battery of monoclonal antibodies screened by their immunoreactivity to antigenic determinants present in the neurosecretory cells of the pars intercerebralis in the brain. The results are illustrated with three selected monoclonal antibodies which recognize antigenes located within the neurosecretory granules of the pericarya of the pars intercerebralis, the cerebro-cardiac axon tracts and the axonic endings in the neurohaemal part of the corpora cardiaca. The apparent molecular weights of these antigenes were determined by Western blotting. We discuss the potential of these monoclonal antibodies for the isolation and structure determination of neuropeptides.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{dimarcq_insect_1988, title = {Insect immunity. Purification and characterization of a family of novel inducible antibacterial proteins from immunized larvae of the dipteran Phormia terranovae and complete amino-acid sequence of the predominant member, diptericin A}, author = {Jean-Luc Dimarcq and E Keppi and B Dunbar and J Lambert and Jean-Marc Reichhart and Danièle Hoffmann and S M Rankine and J E Fothergill and Jules A Hoffmann}, issn = {0014-2956}, year = {1988}, date = {1988-01-01}, journal = {Eur. J. Biochem.}, volume = {171}, number = {1-2}, pages = {17--22}, abstract = {Injury or injection of live bacteria into third instar larvae of the dipteran insect Phormia terranovae results in the appearance in the haemolymph of at least five groups of heat-stable, more or less basic peptides with antibacterial activity against Escherichia coli. Three of these peptides have been purified. The amino acid sequence has been completely established for one of these and partially (first 40 residues from the N-terminus) for the two others. The sequences show marked homologies indicating that the three peptides belong to a common family. They are not related to other known antibacterial peptides from insects [lysozymes, cecropins (including sarcotoxin I) and attacins]. We propose the name of diptericins for this new family of antibiotic molecules.}, keywords = {Amino Acids, Animals, Anti-Bacterial Agents, Diptera, Escherichia coli, hoffmann, Insect Hormones, Insect Proteins, Isoelectric Point, Larva, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{costet_ecdysteroid_1987, title = {Ecdysteroid biosynthesis and embryonic development are disturbed in insects (Locusta migratoria) reared on plant diet (Triticum sativum) with a selectively modified sterol profile}, author = {M F Costet and El M Achouri and Maurice Charlet and R Lanot and P Benveniste and Jules A Hoffmann}, issn = {0027-8424}, year = {1987}, date = {1987-02-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {84}, number = {3}, pages = {643--647}, abstract = {Wheat seedlings germinating in the presence of the systemic fungicide fenpropimorph accumulate 9beta,19-cyclopropylsterols (95% of total sterols) in place of Delta(5)-sterols, which are normally produced in these plants. Adult females of the phytophagous insect Locusta migratoria show a dramatic decrease in their cholesterol content when reared on fenpropimorph-treated wheat. These females lay eggs with the ecdysteroid concentration reduced by up to 80% as compared to controls. Injection of fenpropimorph to the insects or feeding them on wheat coated with the fungicide (normal sterol composition) does not affect their sterol or ecdysteroid profiles; addition of cholesterol to fenpropimorph-treated wheat prior to feeding restores normal ecdysteroid titers in the insects. The severe reduction of the ecdysteroid content in eggs laid by females reared on fenpropimorph-treated wheat is associated with a series of developmental arrests and/or abnormalities. The results show that the dietary 9beta,19-cyclopropylsterols cannot be used by Locusta in place of Delta(5)-sterols for ecdysteroid biosynthesis. They suggest that the selective inhibition of specific enzymes in the sterol biosynthetic pathway of the plants can be used as a strategy to control insect development.}, keywords = {hoffmann, M3i}, pubstate = {published}, tppubtype = {article} } @article{freund-mercier_pharmacological_1987, title = {Pharmacological characteristics and anatomical distribution of (3H)oxytocin-binding sites in the wistar rat brain studied by autoradiography.}, author = {M J Freund-Mercier and M E Stoekel and J M Palacios and A Pazos and Jean-Marc Reichhart and A Porte and P Richard}, year = {1987}, date = {1987-01-01}, journal = {Neuroscience}, volume = {20}, pages = {599--614}, abstract = {Oxytocin-binding sites were detected by autoradiography on rat brain sections incubated in the presence of the [3H]oxytocin. These sites were characterized pharmacologically using quantitative autoradiography. High pressure liquid chromatography controls of the incubation media indicated that labelling was due to the intact [3H]oxytocin molecule. Pharmacological analysis of different locations (central amygdaloid nucleus, ventral subiculum and ventromedial hypothalamic nucleus) showed that the sites detected had a high affinity for oxytocin and also for arginine-vasopressin. In contrast, some areas known to bind vasopressin intensely, such as suprachiasmatic and lateral septum nuclei, had little or no affinity for oxytocin. Autoradiographs revealed [3H]oxytocin-binding sites in already known brain areas (olfactory centres, ventral subiculum, central amygdaloid nucleus, bed nucleus of the stria terminalis) albeit with more extensive labelling of some of these formations, in particular, the amygdaloid complex. In addition, specific [3H]oxytocin-binding sites were found in areas not yet reported to bind oxytocin, such as the paraventricular thalamic and caudate nuclei. In the hypothalamus, specific binding sites were not detected in the supraoptic and paraventricular nuclei : the only structure labelled was the ventrolateral part of the ventromedial nucleus. Discrepancies between the concentrations of [3H]oxytocin-binding sites, the known distribution of oxytocin-containing endings and electrophysiological data indicate that autoradiography, under our conditions, apparently only reveals some of the oxytocin receptors in the brain. Thus, in the hypothalamus, no relationship can be established between the known effect of oxytocin on oxytocinergic magnocellular neurons and detection of specific [3H]oxytocin-binding sites. Autoradiography may reveal mainly oxytocin-binding sites in areas receiving diverse "parasynaptic" information, where oxytocin might play a modulatory role rather than exerting rapid, short-term effects of the neurotransmitter type.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{debono_a21978c_1987, title = {A21978C, a complex of new acidic peptide antibiotics: isolation, chemistry, and mass spectral structure elucidation}, author = {M Debono and M Barnhart and C B Carrell and Jules A Hoffmann and J L Occolowitz and B J Abbott and D S Fukuda and R L Hamill and K Biemann and W C Herlihy}, issn = {0021-8820}, year = {1987}, date = {1987-01-01}, journal = {J. Antibiot.}, volume = {40}, number = {6}, pages = {761--777}, abstract = {A21978C, produced by Streptomyces roseosporus, NRRL 11379, is a complex of new acidic lipopeptolide antibiotics which inhibits Gram-positive bacteria. HPLC separation of the various components from the purified complex resulted in the isolation of A21978C1, -C2 and -C3 (major components) and -C4, -C5, and -C0 (minor components). Each of these components was fermented with cultures of Actinoplanes utahensis (NRRL 12052) to give the identical inactive peptide ("A21978C nucleus") by removal of the fatty acid acyl groups from the N-terminus. This peptide was composed of 13 amino acids: L-kynurenine, L-threo-3-methylglutamic acid, L-asparagine, L-aspartic acid (3 residues), glycine (2 residues), L-tryptophan, L-ornithine, D-alanine, D-serine and L-threonine. The amino acid sequence was determined using a combination of the Edman degradation and gas chromatography mass spectrum (GC-MS) analysis of appropriately derivatized peptides obtained from partial hydrolysis. Each major component was shown to be acylated with a branched chain fatty acid at the N-terminus and the structure of this fatty acid was determined by 1H NMR and mass spectral methods. A structure for A21978C was assigned on the basis of this degradative and physico-chemical information.}, keywords = {Acylation, Amino Acids, Anti-Bacterial Agents, Chemical Phenomena, Chemistry, Chromatography, Cyclic, Fatty Acids, Gas Chromatography-Mass Spectrometry, High Pressure Liquid, hoffmann, Hydrolysis, M3i, Magnetic Resonance Spectroscopy, Mass Spectrometry, Molecular Conformation, Peptides, Spectrophotometry, Streptomyces}, pubstate = {published}, tppubtype = {article} } @article{keppi_recherches_1986, title = {Recherches sur les mécanismes de défense antibactérienne chez les Insectes : isolement de peptides antibactériens dans l'hémolymphe du Diptère Phormia terranovae.}, author = {E Keppi and Jean-Luc Dimarcq and J Lambert and Daniel Zachary and Jean-Marc Reichhart and Danièle Hoffmann and R Keller and Jules A Hoffmann}, year = {1986}, date = {1986-01-01}, journal = {303}, pages = {155--160}, address = {Paris}, edition = {C. R. Acad. Sc.}, abstract = {Three groups of antibacterial peptides were isolated from immune hemolymph of larvae of the Dipteran Phormia terranovae through a combination of heat-treatment, cation exchange chromatography, gel permeation chromatography and HPLC. Their amino-acid composition was determined and showed marked analogies between the various peptides. They differed in several respects from known antibacterial peptides such as cecropins and attacins of Lepidopterans and from Sarcotoxin I from the Dipteran Sarcophaga peregrina.}, keywords = {hoffmann, M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{reichhart_ecdysiotropic_1986, title = {Ecdysiotropic activity in brains and corpora cardiaca of larvae and adults of Locusta migratoria : in vitro assays.}, author = {Jean-Marc Reichhart and Maurice Charlet}, year = {1986}, date = {1986-01-01}, journal = {Intern. J. of Invertebrate Reprod. and Develop.}, volume = {10}, pages = {17--25}, abstract = {Under in vitro conditions the excised left and right prothoracic glands of a larva of Locusta migratoria produce similar amounts of ecdysone, as monitored by RIA. When brain-corpora cardiaca extracts from last instar larvae of the same species are added to one of the two glands, its ecdysone production is significantly increased over that of the homologous gland. The stimulation is dose dependant. This in vitro system has been used as an assay to probe the presence of ecdysiotropic factors in brain-corpora cardiaca complexes of larvae and adult females of Locusta migratoria. The results show that the apparent titres of ecdysiotropic factors in these extracts fluctuated during larval development and in adult females. These fluctuations are discussed in relation to our present information on the role of brain and corpora cardiaca in the control of ecdysone synthesis in prothoracic glands and follicle cell epithelia in this species.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{meister_conversion_1985, title = {Conversion of a radiolabelled ecdysone precursor, 2,22,25-trideoxyecdysone, by embryonic and larval tissues of Locusta migratoria}, author = {Marie F Meister and Jean-Luc Dimarcq and Christine Kappler and Charles Hetru and Marie Lagueux and R Lanot and B Luu and Jules A Hoffmann}, issn = {0303-7207}, year = {1985}, date = {1985-06-01}, journal = {Mol. Cell. Endocrinol.}, volume = {41}, number = {1}, pages = {27--44}, abstract = {A high specific activity tritiated ecdysone precursor, 2,22,25-trideoxyecdysone, was used to probe the capacity of various embryonic and larval tissues to perform the last 3 hydroxylation steps in ecdysone biosynthesis. Embryos at early stages of development, prior to the differentiation of their endocrine glands and embryonic heads, thoraces and abdomens of later stages, were found to have the capacity to hydroxylate the precursor to ecdysone. Larval epidermis and fat body are also able to transform 2,22,25-trideoxyecdysone into ecdysone; Malpighian tubules and midgut hydroxylate the precursor at C-2 but are apparently unable to hydroxylate both at C-22 and C-25. Larval prothoracic glands convert the precursor to ecdysone at a very efficient rate, which is 1-2 magnitudes higher than that of the other tissues investigated; several data argue for the existence of a privileged sequence of hydroxylations, C-25, C-22, C-2, in the larval prothoracic glands.}, keywords = {Abdomen, Animals, Cholestenones, Chromatography, Ecdysone, Epidermis, Fat Body, Grasshoppers, Head, High Pressure Liquid, hoffmann, Hydroxylation, Larva, M3i, Malpighian Tubules, Thorax}, pubstate = {published}, tppubtype = {article} } @article{grau_growth_1985, title = {Growth stimulation of the immature chick oviduct by androgens : the vagina as a new target tissue.}, author = {Y Grau and Jean-Marc Reichhart and J Thiebold}, year = {1985}, date = {1985-01-01}, volume = {85}, pages = {1--13}, abstract = {Testosterone when injected alone stimulates growth of the vagina but is inactive upon the other segments of the oviduct of the immature chicken. This action of testosterone can already be detected in embryos : it is expressed by the beginning of differenciation of the vaginal mesenchyme cells into smooth muscle cells. In the treated immature chicken, stimulation of growth is considerable and is specifically caused by androgens (testosterone and 5-a-dihydrotestosterone); the vaginal mesenchyme differentiates into ciliated cells and goblets cells. (3H) testosterone binding has been found in the vagina of the immature chicken (data not shown). The characteristics of testosterone binding to cytoplasmic components of the chick vagina are consistent with its identity as a testosterone receptor.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{hetru_ecdysone_1985, title = {Ecdysone conjugates: isolation and identification}, author = {Charles Hetru and B Luu and Jules A Hoffmann}, issn = {0076-6879}, year = {1985}, date = {1985-01-01}, journal = {Meth. Enzymol.}, volume = {111}, pages = {411--419}, keywords = {Animals, Bombyx, Chromatography, Ecdysone, Helix (Snails), High Pressure Liquid, hoffmann, Insect Hormones, M3i, Magnetic Resonance Spectroscopy, Mass Spectrometry, Spectrophotometry, Structure-Activity Relationship}, pubstate = {published}, tppubtype = {article} } @article{tsoupras_cytokinin_1983, title = {A Cytokinin (Isopentenyl-Adenosyl-Mononucleotide) Linked to Ecdysone in Newly Laid Eggs of Locusta migratoria}, author = {G Tsoupras and B Luu and Jules A Hoffmann}, doi = {10.1126/science.220.4596.507}, issn = {0036-8075}, year = {1983}, date = {1983-01-01}, journal = {Science}, volume = {220}, number = {4596}, pages = {507--509}, abstract = {Newly laid eggs of the insect Locusta migratoria contain high concentrations (50 nanomoles per gram) of an ecdysone conjugate of maternal origin; 3 milligrams of this conjugate were isolated by conventional techniques, and the structure was established by mass spectrometry and (1)H, (13)C, and (31)P nuclear magnetic resonance as the 22-N(6)-(isopentenyl)-adenosine monophosphoric ester of ecdysone.}, keywords = {hoffmann, M3i}, pubstate = {published}, tppubtype = {article} } @article{tsoupras_isolation_1982, title = {Isolation and identification of three ecdysteroid conjugates with a C-20 hydroxy group in eggs of Locusta migratoria}, author = {G Tsoupras and B Luu and Jules A Hoffmann}, issn = {0039-128X}, year = {1982}, date = {1982-11-01}, journal = {Steroids}, volume = {40}, number = {5}, pages = {551--560}, abstract = {Three ecdysteroids conjugates with a hydroxy group at C-20 were isolated from developing eggs of locusta migratoria and identified as 22-phosphate conjugates of 2-deoxy-20-hydroxy-ecdysone, 20-hydroxyecdysone and 20-hydroxyecdysone acetate.}, keywords = {Animals, Chromatography, Ecdysone, Ecdysterone, Female, Grasshoppers, High Pressure Liquid, hoffmann, M3i, Magnetic Resonance Spectroscopy, Ovum, Phosphates, Spectrophotometry, Ultraviolet}, pubstate = {published}, tppubtype = {article} } @article{lachaise_ecdysteroids_1982, title = {Ecdysteroids and embryonic development in the shore crab, Carcinus maenas}, author = {F Lachaise and Jules A Hoffmann}, issn = {0018-4888}, year = {1982}, date = {1982-09-01}, journal = {Hoppe-Seyler's Z. Physiol. Chem.}, volume = {363}, number = {9}, pages = {1059--1067}, abstract = {Eggs at various stages of embryonic development of Carcinus maenas contain high concentrations of the ecdysteroid ponasterone A together with lower titres of 20-hydroxyecdysone and ecdysone. Correlative studies on ecdysteroid titres and developmental characteristics of Carcinus embryos indicate that one function of ponasterone A might be related to the control of deposition of an embryonic envelope.}, keywords = {Animals, Brachyura, Ecdysone, Ecdysterone, Embryo, Female, hoffmann, M3i, Nonmammalian, Ovum, Radioimmunoassay}, pubstate = {published}, tppubtype = {article} } @article{hetru_biosynthetic_1982, title = {The biosynthetic pathway of ecdysone: studies with vitellogenic ovaries of Locusta migratoria (Orthoptera)}, author = {Charles Hetru and Christine Kappler and Jules A Hoffmann and R Nearn and Lee Bang and D H Horn}, issn = {0303-7207}, year = {1982}, date = {1982-04-01}, journal = {Mol. Cell. Endocrinol.}, volume = {26}, number = {1-2}, pages = {51--80}, abstract = {Ovaries of adult females of Locusta migratoria synthesize impressive amounts of the steroid hormone ecdysone (and related ecdysteroids) during the late phases of vitellogenesis. The present study, aimed at elucidating the sequence of the biosynthetic steps that lead from cholesterol to ecdysone, has taken benefit of this remarkable biological model by using a double approach: (1) isolation and physico-chemical identification of endogenous biogenetic intermediates; (2) metabolic study of labelled putative precursor molecules. The data presented in this paper lead us to propose the following sequence of events: conversion of cholesterol to 3 beta-hydroxy-5 beta-cholest-7-en-6-one (via several intermediates not identified in this study) followed by 14 beta-hydroxylation to 3 beta, 14 alpha-dihydroxy-5 beta-cholest-7-en-6-one; hydroxylation on the side-chain at C-25 and C-22 (in this order) to 2-deoxyecdysone; hydroxylation at C-2 to ecdysone.}, keywords = {Animals, Chromatography, Ecdysone, Female, Gas Chromatography-Mass Spectrometry, High Pressure Liquid, hoffmann, M3i, Orthoptera, Ovary}, pubstate = {published}, tppubtype = {article} } @article{lagueux_ecdysteroids_1981, title = {Ecdysteroids are bound to vitellin in newly laid eggs of Locusta}, author = {Marie Lagueux and P Harry and Jules A Hoffmann}, issn = {0303-7207}, year = {1981}, date = {1981-12-01}, journal = {Mol. Cell. Endocrinol.}, volume = {24}, number = {3}, pages = {325--338}, abstract = {The follicle cells of vitellogenic ovaries of Locusta migratoria have been reported to synthesize impressive amounts of ecdysteroids which accumulate inside the oöcytes where they persist during egg-laying; these ecdysteroids are conjugated to more than 95%, and it is believed that their hydrolysis in the egg is the source of the peaks of free ecdysone observed in early embryonic development. The present paper shows that, in the eggs, the ecdysteroid conjugates are bound to a 520 000-dalton macromolecule which shares several characteristics with the major yolk protein vitellin and is precipitated by an anti-vitellin antibody. The physiological relevance of the binding of ovarian ecdysteroid conjugates is discussed in respect to the transfer of maternal ecdysteroids to the embryo.}, keywords = {Animals, Chromatography, Ecdysteroids, Female, Grasshoppers, High Pressure Liquid, hoffmann, Invertebrate Hormones, Lipoproteins, M3i, Molecular Weight, Ovum, Peptide Hydrolases, Radioimmunoassay, Vitellogenins}, pubstate = {published}, tppubtype = {article} } @incollection{thiebold_asymetric_1981, title = {The asymetric development of the gonads in the female chick embryo : new experimental data and retrospect.}, author = {J Thiebold and A Petit and Jean-Marc Reichhart}, year = {1981}, date = {1981-01-01}, booktitle = {Development and function of reproductive organs}, pages = {244--249}, publisher = {Byskov & Peters, excerpta Medica, Amsterdam-Oxford-Princeton}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {incollection} } @article{lachaise_ecdysteroids_1981, title = {Ecdysteroids and ovarian development in the shore crab, Carcinus maenas}, author = {F Lachaise and M Goudeau and Charles Hetru and Christine Kappler and Jules A Hoffmann}, issn = {0018-4888}, year = {1981}, date = {1981-01-01}, journal = {Hoppe-Seyler's Z. Physiol. Chem.}, volume = {362}, number = {5}, pages = {521--529}, abstract = {Mature ovaries of the shore crab Carcinus maenas contain large concentrations of three major ecdysteroids which we have identified by physicochemical methods as ecdysone, 20-hydroxyecdysone and ponasterone A. The fluctuations of ovarian and blood ecdysteroid concentrations are presented in relation to the various stages of ovarian development.}, keywords = {aging, Animals, Brachyura, Cross Reactions, Ecdysone, Ecdysterone, Female, hoffmann, M3i, Ovary, Radioimmunoassay, Sexual Maturation}, pubstate = {published}, tppubtype = {article} } @article{zachary_endocrine_1980, title = {Endocrine control of the metamorphosis of the larval muscles in Calliphora erythrocephala (Diptera): in vitro studies of the role of ecdysteroids}, author = {Daniel Zachary and Jules A Hoffmann}, issn = {0012-1606}, year = {1980}, date = {1980-11-01}, journal = {Dev. Biol.}, volume = {80}, number = {1}, pages = {235--247}, keywords = {Animals, Biological, Diptera, Ecdysone, Ecdysterone, hoffmann, Larva, M3i, Metamorphosis, Muscles, Organ Culture Techniques}, pubstate = {published}, tppubtype = {article} } @article{reichhart_responsiveness_1980, title = {Responsiveness of developing chick müllerian ducts to sex steroids.}, author = {Jean-Marc Reichhart and J P Weniger and J Thiebold}, year = {1980}, date = {1980-01-01}, journal = {Gen.Comp.Endocrin.}, volume = {40}, pages = {328}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{goltzene_follicle_1978, title = {The follicle cell epithelium of maturing ovaries of Locusta migratoria: a new biosynthetic tissue for ecdysone}, author = {F Goltzené and Marie Lagueux and Maurice Charlet and Jules A Hoffmann}, issn = {0018-4888}, year = {1978}, date = {1978-10-01}, journal = {Hoppe-Seyler's Z. Physiol. Chem.}, volume = {359}, number = {10}, pages = {1427--1434}, abstract = {Follicle cells of maturing ovaries of Locusta migratoria are demonstrated to synthesize the moulting hormone ecdysone (2beta,3beta,14alpha,22R,25-pentahydroxy-5beta-cholest-7-en-6-one). Studies of secretory kinetics under in vitro conditions show that the intensity of hormone secretion is strictly dependent on the stage of maturation of the excised ovaries.}, keywords = {Animals, Ecdysone, Epithelium, Female, Grasshoppers, hoffmann, Kinetics, M3i, Oocytes, Ovarian Follicle, Ovary, Sexual Maturation}, pubstate = {published}, tppubtype = {article} } @article{hetru_adult_1978, title = {Adult ovaries of Locusta migratoria contain the sequence of biosynthetic intermediates for ecdysone}, author = {Charles Hetru and Marie Lagueux and L Bang and Jules A Hoffmann}, issn = {0024-3205}, year = {1978}, date = {1978-06-01}, journal = {Life Sci.}, volume = {22}, number = {23}, pages = {2141--2154}, keywords = {Animals, Chemical Phenomena, Chemistry, Cholesterol, Ecdysone, Female, Grasshoppers, hoffmann, M3i, Mass Spectrometry, Ovary, Ovum}, pubstate = {published}, tppubtype = {article} } @article{brehelin_comparative_1978, title = {A comparative ultrastructural study of blood cells from nine insect orders}, author = {M Brehélin and Daniel Zachary and Jules A Hoffmann}, issn = {0302-766X}, year = {1978}, date = {1978-01-01}, journal = {Cell Tissue Res.}, volume = {195}, number = {1}, pages = {45--57}, abstract = {An ultrastructural study of hemocytes from 9 different insect orders has led to the identification of 8 cell types: (1) Plasmatocytes, whose cytoplasm is filled with small dense lysosomes and large heterogeneous structures, are phagocytic cells. (2) Granulocytes, filled with uniformly electron dense granules, are involved in capsule formation. (3) Coagulocytes, which contain granules and structured globules and which possess a well developed RER, are involved in phagocytosis. (4) Spherule cells are filled with large spherical inclusions. (5) Oenocytoids are large cells with few cytoplasmic organelles. These 5 hemocyte types represent the majority of insect blood cells. (6) Prohemocytes, blastic cells which are one of the stem cells a hemocytes, are very few in number in each species investigated. (7) Thrombocytoids and (8) Prodocytes are restricted to a small number of insect species. The ultrastructural characteristics of these hemocyte types are discussed.}, keywords = {Animals, Blood Cells, Electron, Granulocytes, Hemocytes, hoffmann, insects, M3i, Microscopy, Species Specificity}, pubstate = {published}, tppubtype = {article} } @article{tilak_excess_1977, title = {Excess azide method of peptide synthesis}, author = {M A Tilak and Jules A Hoffmann}, issn = {0022-3263}, year = {1977}, date = {1977-06-01}, journal = {J. Org. Chem.}, volume = {42}, number = {12}, pages = {2098--2100}, keywords = {Azides, Dipeptides, hoffmann, M3i, methods, Oligopeptides}, pubstate = {published}, tppubtype = {article} } @article{brehelin_comparative_1977, title = {Comparative study of hemocyte capsule formation in Locusta migratoria, Melolontha melolontha and Calliphora erythrocephala}, author = {M Brehelin and Daniel Zachary and Jules A Hoffmann}, issn = {0003-4150}, year = {1977}, date = {1977-02-01}, journal = {Ann Parasitol Hum Comp}, volume = {52}, number = {1}, pages = {66--67}, keywords = {Animals, Beetles, Blood Cells, Cellular, Diptera, Grasshoppers, Hemocytes, hoffmann, Immunity, M3i}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_first_1977, title = {First results of the antibacterial defense reactions of Locusta migratoria larva and imago}, author = {Danièle Hoffmann and M Brehelin and Jules A Hoffmann}, issn = {0003-4150}, year = {1977}, date = {1977-02-01}, journal = {Ann Parasitol Hum Comp}, volume = {52}, number = {1}, pages = {87--88}, keywords = {Animals, Bacillus thuringiensis, bacteria, Cellular, Grasshoppers, Hematopoietic System, Hemocytes, Hemolymph, hoffmann, Immunity, Larva, M3i, Muramidase}, pubstate = {published}, tppubtype = {article} } @article{lagueux_ecdysone_1977, title = {Ecdysone during ovarian development in Locusta migratoria}, author = {Marie Lagueux and M Hirn and Jules A Hoffmann}, issn = {0022-1910}, year = {1977}, date = {1977-01-01}, journal = {J. Insect Physiol.}, volume = {23}, number = {1}, pages = {109--119}, keywords = {Animals, Ecdysone, Female, Grasshoppers, hoffmann, M3i, Male, Oogenesis, Ovary}, pubstate = {published}, tppubtype = {article} } @article{reichhart_nouvelles_1977, title = {Nouvelles données sur la croissance asymétrique du tractus müllérien chez l'embryon de Poulet femelle.}, author = {Jean-Marc Reichhart and J Thiebold}, year = {1977}, date = {1977-01-01}, journal = {C. R. Soc. Biol.}, volume = {171}, pages = {1097--1100}, abstract = {Shape and orientation of the mesothelial cells were examined in the mullerian ducts of 8, 13 and 15 day female chick embryos with the scanning eletron microscope. The observed evolution in the pattern of these cells likely reflects the mechanical conditions to which these organs are subjected during embryonic development : stretching for the left duct, slackening for the right duct. These observations, together with data concerning growth of these organs suggest that topographical relationships between cells, which in this system result from mechanical factors, play an important role in controlling cell proliferation.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{bang_identification_1976, title = {Identification, by gas chromatography and mass spectrometry, of ecdysone synthesized in the ovaries of adult female Locusta migratoria (Insecta, Orthoptera)}, author = {L Bang and Marie Lagueux and M Hirn and Jules A Hoffmann}, year = {1976}, date = {1976-10-01}, journal = {C.R. Hebd. Seances Acad. Sci., Ser. D, Sci. Nat.}, volume = {283}, number = {9}, pages = {1081--1084}, abstract = {Gas chromatographic analysis, coupled to mass spectrometry, demonstrates that the considerable amounts of ecdysteroids produced in the ovaries of female adults of Locusta migratoria at the end of each ovarian cycle, consist predominantly of ecdysone.}, keywords = {Age Factors, Animals, Chromatography, Ecdysone, Female, Gas, Grasshoppers, hoffmann, M3i, Mass Spectrometry, Ovary}, pubstate = {published}, tppubtype = {article} } @article{feyereisen_dynamics_1976, title = {Dynamics of ecdysone metabolism after ingestion and injection in Locusta migratoria}, author = {R Feyereisen and Marie Lagueux and Jules A Hoffmann}, issn = {0016-6480}, year = {1976}, date = {1976-07-01}, journal = {Gen. Comp. Endocrinol.}, volume = {29}, number = {3}, pages = {319--327}, keywords = {Animals, Ecdysone, Grasshoppers, hoffmann, Hydroxylation, Injections, Intestines, Larva, M3i, Malpighian Tubules, Oral}, pubstate = {published}, tppubtype = {article} } @article{lagueux_ecdysteroid_1976, title = {Ecdysteroid levels and ovarian development in adult female Locust migratoria}, author = {Marie Lagueux and M Hirn and M de Reggi and Jules A Hoffmann}, year = {1976}, date = {1976-03-01}, journal = {C.R. Hebd. Seances Acad. Sci., Ser. D, Sci. Nat.}, volume = {282}, number = {12}, pages = {1187--1190}, abstract = {During ovarian development in adult females of Locusta, an important biosynthesis of ecdysteroids is observed at the end of the maturation process of the terminal oocyte. This phenomenon occurs regularly during the different ovarian cycles. No synthesis is observed in allatectomized or ovariectomized female adults.}, keywords = {Adipose Tissue, Animals, Castration, Female, Grasshoppers, Hemolymph, hoffmann, Invertebrate Hormones, M3i, Neurosecretory Systems, Oogenesis, Ovary, Oviposition}, pubstate = {published}, tppubtype = {article} } @article{thiebold_ontogenese_1976, title = {Ontogenèse de la sensibilité des canaux de Müller à l'oestradiol chez le Poulet.}, author = {J Thiebold and Jean-Marc Reichhart}, year = {1976}, date = {1976-01-01}, journal = {Bull. Soc. Zool. France}, volume = {101}, pages = {115--119}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{thiebold_proliferation_1976, title = {Prolifération cellulaire et différenciation des canaux de Müller chez l'embryon de Poulet femelle.}, author = {J Thiebold and Jean-Marc Reichhart}, year = {1976}, date = {1976-01-01}, journal = {Bull. Soc. Zool. France}, volume = {101}, pages = {107--111}, abstract = {A hypothesis is put forward according to which mullerian asymmetry in birds (functional left oviduct, rudimentary right mullerian duct) results from different mechanic conditions for the right and left ducts during embryonic life. The slowing down of growth in the right duct could be correlative to a loss of sensitivity to the ovarian hormone.}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{zachary_role_1975, title = {Role of the “thrombocytoids” in capsule formation in the dipteran Calliphora erythrocephala}, author = {Daniel Zachary and M Bréhelin and Jules A Hoffmann}, issn = {0302-766X}, year = {1975}, date = {1975-10-01}, journal = {Cell Tissue Res.}, volume = {162}, number = {3}, pages = {343--348}, abstract = {Of the three hemocyte types present in the blood of Calliphora, only one participates in capsule formation around implanted cellophane. This hemocyte, the thrombocytoid, shows in the blood a tendency to dissociate into numerous small cytoplasmic fragments, comparable to the mammalian megakaryocyte. This tendency is dramatically increased during the process of encapsulation. Most of the intact thrombocytoids and the numerous fragments participating in capsule formation do not show any particular modifications in their cytoplasm during this process, which corresponds to a mere sequestration of the implant. Dense material, resulting from necrotic cell debris and hemolymph lipoproteins, is often observed between the cellophane and encapsulating thrombocytoids, which apparently participate in the resorption of this material.}, keywords = {Agglutination, Animals, Diptera, Foreign Bodies, Hemolymph, hoffmann, M3i, Phagocytosis}, pubstate = {published}, tppubtype = {article} } @article{feyereisen_hemolymphatic_1975, title = {The hemolymphatic transport of molting hormone during the development of Locusta migratoria L}, author = {R Feyereisen and Marie Lagueux and Jules A Hoffmann}, year = {1975}, date = {1975-04-01}, journal = {C.R. Hebd. Seances Acad. Sci., Ser. D, Sci. Nat.}, volume = {280}, number = {14}, pages = {1709--1712}, abstract = {Shortly after injection of radio-labelled ecdysone into fifth instar larvae of Locusta migratoria, 20-hydroxy-ecdysone (ecdysterone) is the main hormone found in the blood. Some 10% of the circulating hormone are bound to hemolymph macromolecules. The ratio of bound to free hormone is stage-dependent; it decreases considerably after previous injections of non-labelled ecdysone, but increases in insects in which ecdysone biosynthesis has been blocked by extirpation of the prothoracic glands or selective X-ray treatment of the hemocytopoietic tissue.}, keywords = {Age Factors, Animals, Carrier Proteins, Chromatography, Ecdysone, Ecdysterone, Gel, Grasshoppers, Hematopoietic System, hoffmann, Larva, M3i, Neurosecretory Systems, Protein Binding, Time Factors}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_role_1975, title = {The role of the prothoracic glands in the production of ecdysone during the last larval instar of Locusta migratoria L}, author = {Jules A Hoffmann and J Koolman and C Beyler}, year = {1975}, date = {1975-02-01}, journal = {C.R. Hebd. Seances Acad. Sci., Ser. D, Sci. Nat.}, volume = {280}, number = {6}, pages = {733--736}, abstract = {Tritiated cholesterol is rapidly converted to labelled ecdysone in vitro by prothoracic glands from last instar larvae of Locusta at the time of the maximum endogenous hormone increase of the insects. Glands from larvae with low hormone content or fat body fragments do not make similar conversions.}, keywords = {Animals, Cholesterol, Ecdysone, Grasshoppers, hoffmann, Larva, M3i}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_modifications_1974, title = {Modifications of the hemogram and of the hemocytopoietic tissue of male adults of Locusta migratoria (Orthoptera) after injection of Bacillus thuringiensis}, author = {Danièle Hoffmann and M Brehelin and Jules A Hoffmann}, issn = {0022-2011}, year = {1974}, date = {1974-09-01}, journal = {J. Invertebr. Pathol.}, volume = {24}, number = {2}, pages = {238--247}, keywords = {Bacillus, Blood Cell Count, Grasshoppers, Hematopoietic System, hoffmann, M3i}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_prothoracic_1974, title = {Prothoracic glands in the regulation of ecdysone titres and metabolic fate of injected labelled ecdysone in Locusta migratoria}, author = {Jules A Hoffmann and J Koolman}, issn = {0022-1910}, year = {1974}, date = {1974-08-01}, journal = {J. Insect Physiol.}, volume = {20}, number = {8}, pages = {1593--1601}, keywords = {Animals, Ecdysone, Grasshoppers, hoffmann, Injections, M3i}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_active_1974, title = {Active protein synthesis in the prothoracic glands and ecdysone titer in the permanent larvae of Locusta migratoria after selective irradiation of hematopoietic tissue}, author = {Jules A Hoffmann and M J Weins}, issn = {0014-4754}, year = {1974}, date = {1974-07-01}, journal = {Experientia}, volume = {30}, number = {7}, pages = {821--822}, keywords = {Animals, Ecdysone, Grasshoppers, Hematopoietic System, hoffmann, Larva, Leucine, M3i, Protein Biosynthesis, Radiation Effects, Tritium}, pubstate = {published}, tppubtype = {article} } @article{goltzene_control_1974, title = {Control of haemolymph protein synthesis and oocyte maturation by the corpora allata in female adults of locusta migratoria (Orthoptera): role of the blood-forming tissue}, author = {F Goltzené and Jules A Hoffmann}, issn = {0016-6480}, year = {1974}, date = {1974-04-01}, journal = {Gen. Comp. Endocrinol.}, volume = {22}, number = {4}, pages = {489--498}, keywords = {Animals, Disc, Electrophoresis, Female, Grasshoppers, Hematopoietic System, Hemolymph, hoffmann, M3i, Ovum, Protein Biosynthesis, Radiation Effects}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_molting_1974, title = {Molting hormone titer and metabolic fate of injected ecdysone during the fifth larval instar and in adults of Locusta migratoria (Orthoptera)}, author = {Jules A Hoffmann and J Koolman and P Karlson and P Joly}, issn = {0016-6480}, year = {1974}, date = {1974-01-01}, journal = {Gen. Comp. Endocrinol.}, volume = {22}, number = {1}, pages = {90--97}, keywords = {Age Factors, Animals, Chromatography, Ecdysone, Feces, Grasshoppers, hoffmann, Hydroxylation, Invertebrate Hormones, Larva, M3i, Thin Layer, Time Factors, Tritium}, pubstate = {published}, tppubtype = {article} } @article{koolman_sulphage_1973, title = {Sulphage esters as inactivation products of ecdysone in Locusta migratoria}, author = {J Koolman and Jules A Hoffmann and P Karlson}, issn = {0018-4888}, year = {1973}, date = {1973-09-01}, journal = {Hoppe-Seyler's Z. Physiol. Chem.}, volume = {354}, number = {9}, pages = {1043--1048}, keywords = {Animals, Biological, Cattle, Chromatography, Ecdysone, Electrophoresis, Esterases, Glucosidases, Glucuronidase, Grasshoppers, hoffmann, Hydrogen-Ion Concentration, Hydrolysis, Ion Exchange, Isotope Labeling, Kinetics, Larva, Liver, M3i, Metamorphosis, Paper, Plants, Saccharomyces cerevisiae, Snails, Sulfatases, Sulfur Radioisotopes, Sulfuric Acids, Swine, Thin Layer, Time Factors, Tritium}, pubstate = {published}, tppubtype = {article} } @article{zachary_haemocytes_1973, title = {The haemocytes of Calliphora erythrocephala (Meig.) (Diptera)}, author = {Daniel Zachary and Jules A Hoffmann}, issn = {0340-0336}, year = {1973}, date = {1973-07-01}, journal = {Z Zellforsch Mikrosk Anat}, volume = {141}, number = {1}, pages = {55--73}, keywords = {Animals, Cell Count, Cell Nucleus, Diptera, Electron, Endoplasmic Reticulum, Hemolymph, Hemostasis, hoffmann, Inclusion Bodies, Larva, Lipids, Lysosomes, M3i, Microscopy, mitochondria, Phase-Contrast, Pupa, Radiation Effects}, pubstate = {published}, tppubtype = {article} } @article{joly_development_1973, title = {Development of the prothoracic glands of permanent larvae of Locusta migratoria obtained by selective irradiation of the hemocytopoietic tissue}, author = {L Joly and M J Weins and Jules A Hoffmann and A Porte}, issn = {0340-0336}, year = {1973}, date = {1973-02-01}, journal = {Z Zellforsch Mikrosk Anat}, volume = {137}, number = {3}, pages = {387--397}, keywords = {Age Factors, Animals, Biological, Ecdysone, Electron, Endocrine Glands, Golgi Apparatus, Grasshoppers, hoffmann, Larva, M3i, Metamorphosis, Microscopy, Radiation Effects, ribosomes}, pubstate = {published}, tppubtype = {article} } @article{thiebold_inhibition_1973, title = {Inhibition du développement ovarien par la greffe de gonades embryonnaires chez le Poulet.}, author = {J Thiebold and Jean-Marc Reichhart}, year = {1973}, date = {1973-01-01}, journal = {C. R. Soc. Zool. France}, volume = {167}, pages = {1958--1960}, keywords = {M3i, reichhart}, pubstate = {published}, tppubtype = {article} } @article{zachary_new_1972, title = {A new type of blood cell (the thrombocytoid) in Calliphora erythrocephala}, author = {Daniel Zachary and Jules A Hoffmann and A Porte}, year = {1972}, date = {1972-07-01}, journal = {C.R. Hebd. Seances Acad. Sci., Ser. D, Sci. Nat.}, volume = {275}, number = {3}, pages = {393--395}, keywords = {Animals, Blood Cells, Blood Platelets, Cell Membrane, Cell Nucleus, Cytoplasm, Diptera, Electron, Hemolymph, hoffmann, M3i, Microscopy, Phase-Contrast}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_endocrine_1970, title = {Endocrine regulation of production and differentiation of hemocytes in an orthopteran insect: Locusta migratoria migratoroides}, author = {Jules A Hoffmann}, issn = {0016-6480}, year = {1970}, date = {1970-10-01}, journal = {Gen. Comp. Endocrinol.}, volume = {15}, number = {2}, pages = {198--219}, keywords = {Animals, Biological, Blood Cells, Electrocoagulation, Female, Hemolymph, hoffmann, insects, M3i, Male, Metamorphosis, Microscopy, Neurosecretory Systems, Phase-Contrast}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_localization_1970, title = {On the localization of phenol-oxidase activity in coagulocytes of Locusta migratoria L. (Orthoptera)}, author = {Jules A Hoffmann and A Porte and P Joly}, year = {1970}, date = {1970-01-01}, journal = {C.R. Hebd. Seances Acad. Sci., Ser. D, Sci. Nat.}, volume = {270}, number = {4}, pages = {629--631}, keywords = {Animals, Catechol Oxidase, Hemolymph, hoffmann, insects, M3i, Tanning}, pubstate = {published}, tppubtype = {article} } @article{hoffmann_hemopoietic_1970, title = {The hemopoietic organs of the two orthopterans Locusta migratoria and Gryllus bimaculatus}, author = {Jules A Hoffmann}, issn = {0340-0336}, year = {1970}, date = {1970-01-01}, journal = {Z Zellforsch Mikrosk Anat}, volume = {106}, number = {3}, pages = {451--472}, keywords = {Animals, Cell Differentiation, Diptera, Electron, Hematopoiesis, Hematopoietic System, hoffmann, M3i, Microscopy, Species Specificity}, pubstate = {published}, tppubtype = {article} }