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2024
Kehrli Janine, Husser Claire, Ryckelynck Michael
Fluorogenic RNA-Based Biosensors of Small Molecules: Current Developments, Uses, and Perspectives Article de journal
Dans: Biosensors (Basel), vol. 14, no. 8, 2024, ISSN: 2079-6374.
Résumé | Liens | BibTeX | Étiquettes: RYCKELYNCK, Unité ARN
@article{pmid39194605,
title = {Fluorogenic RNA-Based Biosensors of Small Molecules: Current Developments, Uses, and Perspectives},
author = {Janine Kehrli and Claire Husser and Michael Ryckelynck},
doi = {10.3390/bios14080376},
issn = {2079-6374},
year = {2024},
date = {2024-08-01},
urldate = {2024-08-01},
journal = {Biosensors (Basel)},
volume = {14},
number = {8},
abstract = {Small molecules are highly relevant targets for detection and quantification. They are also used to diagnose and monitor the progression of disease and infectious processes and track the presence of contaminants. Fluorogenic RNA-based biosensors (FRBs) represent an appealing solution to the problem of detecting these targets. They combine the portability of molecular systems with the sensitivity and multiplexing capacity of fluorescence, as well as the exquisite ligand selectivity of RNA aptamers. In this review, we first present the different sensing and reporting aptamer modules currently available to design an FRB, together with the main methodologies used to discover modules with new specificities. We next introduce and discuss how both modules can be functionally connected prior to exploring the main applications for which FRB have been used. Finally, we conclude by discussing how using alternative nucleotide chemistries may improve FRB properties and further widen their application scope.},
keywords = {RYCKELYNCK, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
Small molecules are highly relevant targets for detection and quantification. They are also used to diagnose and monitor the progression of disease and infectious processes and track the presence of contaminants. Fluorogenic RNA-based biosensors (FRBs) represent an appealing solution to the problem of detecting these targets. They combine the portability of molecular systems with the sensitivity and multiplexing capacity of fluorescence, as well as the exquisite ligand selectivity of RNA aptamers. In this review, we first present the different sensing and reporting aptamer modules currently available to design an FRB, together with the main methodologies used to discover modules with new specificities. We next introduce and discuss how both modules can be functionally connected prior to exploring the main applications for which FRB have been used. Finally, we conclude by discussing how using alternative nucleotide chemistries may improve FRB properties and further widen their application scope.
Y Fu, F Jiang, X Zhang, Y Pan, R Xu, X Liang, X Wu, X Li, K Lin, R Shi, X Zhang, D Ferrandon, J Liu, D Pei, J Wang, T Wang
Perturbation of METTL1-mediated tRNA N7-methylguanosine modification induces senescence and aging Article de journal
Dans: Nature Communications, vol. 8, iss. 15, no. 1, p. 5713, 2024.
Résumé | Liens | BibTeX | Étiquettes: ferrandon, Gene Expression, M3i, RNA, Senescence
@article{Y2024,
title = {Perturbation of METTL1-mediated tRNA N7-methylguanosine modification induces senescence and aging},
author = {Fu Y AND Jiang F and Zhang X and Pan Y and Xu R AND Liang X AND Wu X AND Li X AND Lin K AND Shi R AND Zhang X AND Ferrandon D AND Liu J AND Pei D AND Wang J AND Wang T},
doi = {10.1038/s41467-024-49796-8},
year = {2024},
date = {2024-07-08},
journal = {Nature Communications},
volume = {8},
number = {1},
issue = {15},
pages = {5713},
abstract = {Cellular senescence is characterized by a decrease in protein synthesis, although the underlying processes are mostly unclear. Chemical modifications to transfer RNAs (tRNAs) frequently influence tRNA activity, which is crucial for translation. We describe how tRNA N7-methylguanosine (m7G46) methylation, catalyzed by METTL1-WDR4, regulates translation and influences senescence phenotypes. Mettl1/Wdr4 and m7G gradually diminish with senescence and aging. A decrease in METTL1 causes a reduction in tRNAs, especially those with the m7G modification, via the rapid tRNA degradation (RTD) pathway. The decreases cause ribosomes to stall at certain codons, impeding the translation of mRNA that is essential in pathways such as Wnt signaling and ribosome biogenesis. Furthermore, chronic ribosome stalling stimulates the ribotoxic and integrative stress responses, which induce senescence-associated secretory phenotype. Moreover, restoring eEF1A protein mitigates senescence phenotypes caused by METTL1 deficiency by reducing RTD. Our findings demonstrate that tRNA m7G modification is essential for preventing premature senescence and aging by enabling efficient mRNA translation.},
keywords = {ferrandon, Gene Expression, M3i, RNA, Senescence},
pubstate = {published},
tppubtype = {article}
}
Cellular senescence is characterized by a decrease in protein synthesis, although the underlying processes are mostly unclear. Chemical modifications to transfer RNAs (tRNAs) frequently influence tRNA activity, which is crucial for translation. We describe how tRNA N7-methylguanosine (m7G46) methylation, catalyzed by METTL1-WDR4, regulates translation and influences senescence phenotypes. Mettl1/Wdr4 and m7G gradually diminish with senescence and aging. A decrease in METTL1 causes a reduction in tRNAs, especially those with the m7G modification, via the rapid tRNA degradation (RTD) pathway. The decreases cause ribosomes to stall at certain codons, impeding the translation of mRNA that is essential in pathways such as Wnt signaling and ribosome biogenesis. Furthermore, chronic ribosome stalling stimulates the ribotoxic and integrative stress responses, which induce senescence-associated secretory phenotype. Moreover, restoring eEF1A protein mitigates senescence phenotypes caused by METTL1 deficiency by reducing RTD. Our findings demonstrate that tRNA m7G modification is essential for preventing premature senescence and aging by enabling efficient mRNA translation.
S Ji, JA Hoffmann
Toll-9 prevents the proliferation of injected oncogenic cells in adult flies Article de journal
Dans: Journal of Genetics and Genomics, p. 1673-8527, 2024.
Liens | BibTeX | Étiquettes: Drosophila, hoffmann, M3i, oncogenic cells, Toll-9
@article{JA2024,
title = {Toll-9 prevents the proliferation of injected oncogenic cells in adult flies},
author = {Ji S AND Hoffmann JA},
doi = {doi: 10.1016/j.jgg.2024.07.002},
year = {2024},
date = {2024-07-05},
urldate = {2024-07-05},
journal = {Journal of Genetics and Genomics},
pages = {1673-8527},
keywords = {Drosophila, hoffmann, M3i, oncogenic cells, Toll-9},
pubstate = {published},
tppubtype = {article}
}
Rios-Delgado Gustavo, McReynolds Aubrey K G, Pagella Emma A, Norambuena Javiera, Briaud Paul, Zheng Vincent, Munneke Matthew J, Kim Jisun, Racine Hugo, Carroll Ronan, Zelzion Ehud, Skaar Eric, Bose Jeffrey L, Parker Dane, Lalaouna David, Boyd Jeffrey M
The small non-coding RNA IsrR regulates TCA cycle activity and virulence Article de journal
Dans: bioRxiv, 2024, ISSN: 2692-8205.
Résumé | Liens | BibTeX | Étiquettes: LALAOUNA, ROMBY, Unité ARN
@article{pmid39005296,
title = {The small non-coding RNA IsrR regulates TCA cycle activity and virulence},
author = {Gustavo Rios-Delgado and Aubrey K G McReynolds and Emma A Pagella and Javiera Norambuena and Paul Briaud and Vincent Zheng and Matthew J Munneke and Jisun Kim and Hugo Racine and Ronan Carroll and Ehud Zelzion and Eric Skaar and Jeffrey L Bose and Dane Parker and David Lalaouna and Jeffrey M Boyd},
doi = {10.1101/2024.07.03.601953},
issn = {2692-8205},
year = {2024},
date = {2024-07-01},
urldate = {2024-07-01},
journal = {bioRxiv},
abstract = { has evolved mechanisms to cope with low iron (Fe) availability in host tissues. uses the ferric uptake transcriptional regulator (Fur) to sense titers of cytosolic Fe. Upon Fe depletion, apo-Fur relieves transcriptional repression of genes utilized for Fe uptake. We demonstrate that an Δ mutant has decreased expression of , which codes for the Fe-dependent enzyme aconitase. Decreased expression prevented the Δ mutant from growing with amino acids as sole carbon and energy sources. Suppressor analysis determined that a mutation in , which produces a regulatory RNA, permitted growth by decreasing transcription. The decreased AcnA activity of the Δ mutant was partially relieved by an Δ mutation. Directed mutation of bases predicted to facilitate the interaction between the transcript and IsrR, decreased the ability of IsrR to control expression and IsrR bound to the transcript . IsrR also bound to the transcripts coding the alternate TCA cycle proteins , , , and . Whole cell metal analyses suggest that IsrR promotes Fe uptake and increases intracellular Fe not ligated by macromolecules. Lastly, we determined that Fur and IsrR promote infection using murine skin and acute pneumonia models.},
keywords = {LALAOUNA, ROMBY, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
has evolved mechanisms to cope with low iron (Fe) availability in host tissues. uses the ferric uptake transcriptional regulator (Fur) to sense titers of cytosolic Fe. Upon Fe depletion, apo-Fur relieves transcriptional repression of genes utilized for Fe uptake. We demonstrate that an Δ mutant has decreased expression of , which codes for the Fe-dependent enzyme aconitase. Decreased expression prevented the Δ mutant from growing with amino acids as sole carbon and energy sources. Suppressor analysis determined that a mutation in , which produces a regulatory RNA, permitted growth by decreasing transcription. The decreased AcnA activity of the Δ mutant was partially relieved by an Δ mutation. Directed mutation of bases predicted to facilitate the interaction between the transcript and IsrR, decreased the ability of IsrR to control expression and IsrR bound to the transcript . IsrR also bound to the transcripts coding the alternate TCA cycle proteins , , , and . Whole cell metal analyses suggest that IsrR promotes Fe uptake and increases intracellular Fe not ligated by macromolecules. Lastly, we determined that Fur and IsrR promote infection using murine skin and acute pneumonia models.
Mao Xue-Ling, Eriani Gilbert, Zhou Xiao-Long
ADATs: roles in tRNA editing and relevance to disease Article de journal
Dans: Acta Biochim Biophys Sin (Shanghai), 2024, ISSN: 1745-7270.
Résumé | Liens | BibTeX | Étiquettes: ERIANI, Unité ARN
@article{pmid39034823,
title = {ADATs: roles in tRNA editing and relevance to disease},
author = {Xue-Ling Mao and Gilbert Eriani and Xiao-Long Zhou},
doi = {10.3724/abbs.2024125},
issn = {1745-7270},
year = {2024},
date = {2024-07-01},
urldate = {2024-07-01},
journal = {Acta Biochim Biophys Sin (Shanghai)},
abstract = {Transfer RNAs (tRNAs) play central roles in protein biosynthesis. Post-transcriptional RNA modifications affect tRNA function and stability. Among these modifications, RNA editing is a widespread RNA modification in three domains of life. Proteins of the adenosine deaminase acting on tRNA (ADAT) family were discovered more than 20 years ago. They catalyze the deamination of adenosine to inosine (A-to-I) or cytidine to uridine (C-to-U) during tRNA maturation. The most studied example is the TadA- or ADAT2/3-mediated A-to-I conversion of the tRNA wobble position in the anticodon of prokaryotic or eukaryotic tRNAs, respectively. This review provides detailed information on A-to-I and C-to-U editing of tRNAs in different domains of life, presents recent new findings on ADATs for DNA editing, and finally comments on the association of mutations in the gene with intellectual disability.},
keywords = {ERIANI, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
Transfer RNAs (tRNAs) play central roles in protein biosynthesis. Post-transcriptional RNA modifications affect tRNA function and stability. Among these modifications, RNA editing is a widespread RNA modification in three domains of life. Proteins of the adenosine deaminase acting on tRNA (ADAT) family were discovered more than 20 years ago. They catalyze the deamination of adenosine to inosine (A-to-I) or cytidine to uridine (C-to-U) during tRNA maturation. The most studied example is the TadA- or ADAT2/3-mediated A-to-I conversion of the tRNA wobble position in the anticodon of prokaryotic or eukaryotic tRNAs, respectively. This review provides detailed information on A-to-I and C-to-U editing of tRNAs in different domains of life, presents recent new findings on ADATs for DNA editing, and finally comments on the association of mutations in the gene with intellectual disability.
Silva Elisabete Cruz Da, Gaki Paraskevi, Flieg Fabien, Messmer Melanie, Gucciardi Floriane, Markovska Yevheniia, Reisch Andreas, Fafi-Kremer Samira, Pfeffer Sébastien, Klymchenko Andrey S
Direct Zeptomole Detection of RNA Biomarkers by Ultrabright Fluorescent Nanoparticles on Magnetic Beads Article de journal
Dans: Small, p. e2404167, 2024, ISSN: 1613-6829.
Résumé | Liens | BibTeX | Étiquettes: PFEFFER, Unité ARN
@article{pmid39011971,
title = {Direct Zeptomole Detection of RNA Biomarkers by Ultrabright Fluorescent Nanoparticles on Magnetic Beads},
author = {Elisabete Cruz Da Silva and Paraskevi Gaki and Fabien Flieg and Melanie Messmer and Floriane Gucciardi and Yevheniia Markovska and Andreas Reisch and Samira Fafi-Kremer and Sébastien Pfeffer and Andrey S Klymchenko},
doi = {10.1002/smll.202404167},
issn = {1613-6829},
year = {2024},
date = {2024-07-01},
urldate = {2024-07-01},
journal = {Small},
pages = {e2404167},
abstract = {Nucleic acids are important biomarkers in cancer and viral diseases. However, their ultralow concentration in biological/clinical samples makes direct target detection challenging, because it leads to slow hybridization kinetics with the probe and its insufficient signal-to-noise ratio. Therefore, RNA target detection is done by molecular (target) amplification, notably by RT-PCR, which is a tedious multistep method that includes nucleic acid extraction and reverse transcription. Here, a direct method based on ultrabright dye-loaded polymeric nanoparticles in a sandwich-like hybridization assay with magnetic beads is reported. The ultrabright DNA-functionalized nanoparticle, equivalent to ≈10 000 strongly emissive rhodamine dyes, is hybridized with the magnetic bead to the RNA target, providing the signal amplification for the detection. This concept (magneto-fluorescent sandwich) enables high-throughput detection of DNA and RNA sequences of varied lengths from 48 to 1362 nt with the limit of detection down to 0.3 fm using a plate reader (15 zeptomoles), among the best reported for optical sandwich assays. Moreover, it allows semi-quantitative detection of SARS-CoV-2 viral RNA directly in clinical samples without a dedicated RNA extraction step. The developed technology, combining ultrabright nanoparticles with magnetic beads, addresses fundamental challenges in RNA detection; it is expected to accelerate molecular diagnostics of diseases.},
keywords = {PFEFFER, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
Nucleic acids are important biomarkers in cancer and viral diseases. However, their ultralow concentration in biological/clinical samples makes direct target detection challenging, because it leads to slow hybridization kinetics with the probe and its insufficient signal-to-noise ratio. Therefore, RNA target detection is done by molecular (target) amplification, notably by RT-PCR, which is a tedious multistep method that includes nucleic acid extraction and reverse transcription. Here, a direct method based on ultrabright dye-loaded polymeric nanoparticles in a sandwich-like hybridization assay with magnetic beads is reported. The ultrabright DNA-functionalized nanoparticle, equivalent to ≈10 000 strongly emissive rhodamine dyes, is hybridized with the magnetic bead to the RNA target, providing the signal amplification for the detection. This concept (magneto-fluorescent sandwich) enables high-throughput detection of DNA and RNA sequences of varied lengths from 48 to 1362 nt with the limit of detection down to 0.3 fm using a plate reader (15 zeptomoles), among the best reported for optical sandwich assays. Moreover, it allows semi-quantitative detection of SARS-CoV-2 viral RNA directly in clinical samples without a dedicated RNA extraction step. The developed technology, combining ultrabright nanoparticles with magnetic beads, addresses fundamental challenges in RNA detection; it is expected to accelerate molecular diagnostics of diseases.
Pitolli Martina, Cela Marta, Kapps Delphine, Chicher Johana, Despons Laurence, Frugier Magali
Comparative proteomics uncovers low asparagine content in Plasmodium tRip-KO proteins Article de journal
Dans: IUBMB Life, 2024, ISSN: 1521-6551.
Résumé | Liens | BibTeX | Étiquettes: PPSE, RYCKELYNCK, Unité ARN
@article{pmid38963319,
title = {Comparative proteomics uncovers low asparagine content in Plasmodium tRip-KO proteins},
author = {Martina Pitolli and Marta Cela and Delphine Kapps and Johana Chicher and Laurence Despons and Magali Frugier},
doi = {10.1002/iub.2891},
issn = {1521-6551},
year = {2024},
date = {2024-07-01},
urldate = {2024-07-01},
journal = {IUBMB Life},
abstract = {tRNAs are not only essential for decoding the genetic code, but their abundance also has a strong impact on the rate of protein production, folding, and on the stability of the translated messenger RNAs. Plasmodium expresses a unique surface protein called tRip, involved in the import of exogenous tRNAs into the parasite. Comparative proteomic analysis of the blood stage of wild-type and tRip-KO variant of P. berghei parasites revealed that downregulated proteins in the mutant parasite are distinguished by a bias in their asparagine content. Furthermore, the demonstration of the possibility of charging host tRNAs with Plasmodium aminoacyl-tRNA synthetases led us to propose that imported host tRNAs participate in parasite protein synthesis. These results also suggest a novel mechanism of translational control in which import of host tRNAs emerge as regulators of gene expression in the Plasmodium developmental cycle and pathogenesis, by enabling the synthesis of asparagine-rich regulatory proteins that efficiently and selectively control the parasite infectivity.},
keywords = {PPSE, RYCKELYNCK, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
tRNAs are not only essential for decoding the genetic code, but their abundance also has a strong impact on the rate of protein production, folding, and on the stability of the translated messenger RNAs. Plasmodium expresses a unique surface protein called tRip, involved in the import of exogenous tRNAs into the parasite. Comparative proteomic analysis of the blood stage of wild-type and tRip-KO variant of P. berghei parasites revealed that downregulated proteins in the mutant parasite are distinguished by a bias in their asparagine content. Furthermore, the demonstration of the possibility of charging host tRNAs with Plasmodium aminoacyl-tRNA synthetases led us to propose that imported host tRNAs participate in parasite protein synthesis. These results also suggest a novel mechanism of translational control in which import of host tRNAs emerge as regulators of gene expression in the Plasmodium developmental cycle and pathogenesis, by enabling the synthesis of asparagine-rich regulatory proteins that efficiently and selectively control the parasite infectivity.
Tajer L., Paillart J. -C., Dib H., Sabatier J. -M., Fajloun Z., Khattar Z. Abi
Molecular Mechanisms of Bacterial Resistance to Antimicrobial Peptides in the Modern Era: An Updated Review Article de journal
Dans: Microorganisms, vol. 12, p. 1259, 2024.
Résumé | Liens | BibTeX | Étiquettes: marquet Paillart, PAILLART, Unité ARN
@article{nokey,
title = {Molecular Mechanisms of Bacterial Resistance to Antimicrobial Peptides in the Modern Era: An Updated Review},
author = {L. Tajer and J.-C. Paillart and H. Dib and J.-M. Sabatier and Z. Fajloun and Z. Abi Khattar},
url = {https://www.mdpi.com/2076-2607/12/7/1259},
doi = {10.3390/ microorganisms12071259},
year = {2024},
date = {2024-06-21},
urldate = {2024-06-21},
journal = {Microorganisms},
volume = {12},
pages = {1259},
abstract = {Antimicrobial resistance (AMR) poses a serious global health concern, resulting in a
significant number of deaths annually due to infections that are resistant to treatment. Amidst
this crisis, antimicrobial peptides (AMPs) have emerged as promising alternatives to conventional
antibiotics (ATBs). These cationic peptides, naturally produced by all kingdoms of life, play a
crucial role in the innate immune system of multicellular organisms and in bacterial interspecies
competition by exhibiting broad-spectrum activity against bacteria, fungi, viruses, and parasites.
AMPs target bacterial pathogens through multiple mechanisms, most importantly by disrupting
their membranes, leading to cell lysis. However, bacterial resistance to host AMPs has emerged
due to a slow co-evolutionary process between microorganisms and their hosts. Alarmingly, the
development of resistance to last-resort AMPs in the treatment of MDR infections, such as colistin, is
attributed to the misuse of this and the high rate of horizontal genetic transfer of the corresponding
resistance genes. AMP-resistant bacteria employ diverse mechanisms, including but not limited to
proteolytic degradation, extracellular trapping and inactivation, active efflux, as well as complex
modifications in bacterial cell wall and membrane structures. This review comprehensively examines
all constitutive and inducible molecular resistance mechanisms to AMPs supported by experimental
evidence described to date in bacterial pathogens. We also explore the specificity of these mechanisms
toward structurally diverse AMPs to broaden and enhance their potential in developing and applying
them as therapeutics for MDR bacteria. Additionally, we provide insights into the significance of
AMP resistance within the context of host–pathogen interactions.},
keywords = {marquet Paillart, PAILLART, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
Antimicrobial resistance (AMR) poses a serious global health concern, resulting in a
significant number of deaths annually due to infections that are resistant to treatment. Amidst
this crisis, antimicrobial peptides (AMPs) have emerged as promising alternatives to conventional
antibiotics (ATBs). These cationic peptides, naturally produced by all kingdoms of life, play a
crucial role in the innate immune system of multicellular organisms and in bacterial interspecies
competition by exhibiting broad-spectrum activity against bacteria, fungi, viruses, and parasites.
AMPs target bacterial pathogens through multiple mechanisms, most importantly by disrupting
their membranes, leading to cell lysis. However, bacterial resistance to host AMPs has emerged
due to a slow co-evolutionary process between microorganisms and their hosts. Alarmingly, the
development of resistance to last-resort AMPs in the treatment of MDR infections, such as colistin, is
attributed to the misuse of this and the high rate of horizontal genetic transfer of the corresponding
resistance genes. AMP-resistant bacteria employ diverse mechanisms, including but not limited to
proteolytic degradation, extracellular trapping and inactivation, active efflux, as well as complex
modifications in bacterial cell wall and membrane structures. This review comprehensively examines
all constitutive and inducible molecular resistance mechanisms to AMPs supported by experimental
evidence described to date in bacterial pathogens. We also explore the specificity of these mechanisms
toward structurally diverse AMPs to broaden and enhance their potential in developing and applying
them as therapeutics for MDR bacteria. Additionally, we provide insights into the significance of
AMP resistance within the context of host–pathogen interactions.
significant number of deaths annually due to infections that are resistant to treatment. Amidst
this crisis, antimicrobial peptides (AMPs) have emerged as promising alternatives to conventional
antibiotics (ATBs). These cationic peptides, naturally produced by all kingdoms of life, play a
crucial role in the innate immune system of multicellular organisms and in bacterial interspecies
competition by exhibiting broad-spectrum activity against bacteria, fungi, viruses, and parasites.
AMPs target bacterial pathogens through multiple mechanisms, most importantly by disrupting
their membranes, leading to cell lysis. However, bacterial resistance to host AMPs has emerged
due to a slow co-evolutionary process between microorganisms and their hosts. Alarmingly, the
development of resistance to last-resort AMPs in the treatment of MDR infections, such as colistin, is
attributed to the misuse of this and the high rate of horizontal genetic transfer of the corresponding
resistance genes. AMP-resistant bacteria employ diverse mechanisms, including but not limited to
proteolytic degradation, extracellular trapping and inactivation, active efflux, as well as complex
modifications in bacterial cell wall and membrane structures. This review comprehensively examines
all constitutive and inducible molecular resistance mechanisms to AMPs supported by experimental
evidence described to date in bacterial pathogens. We also explore the specificity of these mechanisms
toward structurally diverse AMPs to broaden and enhance their potential in developing and applying
them as therapeutics for MDR bacteria. Additionally, we provide insights into the significance of
AMP resistance within the context of host–pathogen interactions.
Imbernon Julia Revillo, Weibel Jean‐Marc, Ennifar Eric, Prévost Gilles, Kellenberger Esther
Structural analysis of neomycin B and kanamycin A binding Aminoglycosides Modifying Enzymes (AME) and bacterial ribosomal RNA Article de journal
Dans: Molecular Informatics, 2024, ISSN: 1868-1751.
Résumé | Liens | BibTeX | Étiquettes: ENNIFAR, Unité ARN
@article{RevilloImbernon2024,
title = {Structural analysis of neomycin B and kanamycin A binding Aminoglycosides Modifying Enzymes (AME) and bacterial ribosomal RNA},
author = {Julia Revillo Imbernon and Jean‐Marc Weibel and Eric Ennifar and Gilles Prévost and Esther Kellenberger},
doi = {10.1002/minf.202300339},
issn = {1868-1751},
year = {2024},
date = {2024-06-10},
urldate = {2024-06-10},
journal = {Molecular Informatics},
publisher = {Wiley},
abstract = {<jats:title>Abstract</jats:title><jats:p>Aminoglycosides are crucial antibiotics facing challenges from bacterial resistance. This study addresses the importance of aminoglycoside modifying enzymes in the context of escalating resistance. Drawing upon over two decades of structural data in the Protein Data Bank, we focused on two key antibiotics, neomycin B and kanamycin A, to explore how the aminoglycoside structure is exploited by this family of enzymes. A systematic comparison across diverse enzymes and the RNA A‐site target identified common characteristics in the recognition mode, while assessing the adaptability of neomycin B and kanamycin A in various environments.</jats:p>},
keywords = {ENNIFAR, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
<jats:title>Abstract</jats:title><jats:p>Aminoglycosides are crucial antibiotics facing challenges from bacterial resistance. This study addresses the importance of aminoglycoside modifying enzymes in the context of escalating resistance. Drawing upon over two decades of structural data in the Protein Data Bank, we focused on two key antibiotics, neomycin B and kanamycin A, to explore how the aminoglycoside structure is exploited by this family of enzymes. A systematic comparison across diverse enzymes and the RNA A‐site target identified common characteristics in the recognition mode, while assessing the adaptability of neomycin B and kanamycin A in various environments.</jats:p>
J Chen, G Lin, K Ma, Z Li, S Liégeois, D Ferrandon
A specific innate immune response silences the virulence of Pseudomonas aeruginosa in a latent infection model in the Drosophila melanogaster host Article de journal
Dans: PLoS Pathog., 2024.
Résumé | Liens | BibTeX | Étiquettes: Drosophila melanogaster, ferrandon, Innate immune response, M3i, Pseudomonas aeruginosa
@article{J2024,
title = {A specific innate immune response silences the virulence of Pseudomonas aeruginosa in a latent infection model in the Drosophila melanogaster host},
author = {Chen J and Lin G and Ma K and Li Z and Liégeois S and Ferrandon D},
editor = {. . PLoS Pathog. 2024 Jun 4;20(6):e1012252. . eCollection 2024 Ju},
doi = {doi: 10.1371/journal.ppat.1012252},
year = {2024},
date = {2024-06-04},
urldate = {2024-06-04},
journal = {PLoS Pathog.},
abstract = {Microbial pathogenicity often depends on the route of infection. For instance, P. aeruginosa or S. marcescens cause acute systemic infections when low numbers of bacteria are injected into D. melanogaster flies whereas flies succumb much slower to the continuous ingestion of these pathogens, even though both manage to escape from the gut compartment and reach the hemocoel. Here, we have developed a latent P. aeruginosa infection model by feeding flies on the bacteria for a short period. The bacteria stably colonize internal tissues yet hardly cause any damage since latently-infected flies live almost as long as noninfected control flies. The apparently dormant bacteria display particular characteristics in terms of bacterial colony morphology, composition of the outer cell wall, and motility. The virulence of these bacteria can however be reactivated upon wounding the host. We show that melanization but not the cellular or the systemic humoral response is the predominant host defense that establishes latency and may coerce the bacteria to a dormant state. In addition, the lasting activation of the melanization responses in latently-infected flies provides a degree of protection to the host against a secondary fungal infection. Latent infection by an ingested pathogen protects against a variety of homologous or heterologous systemic secondary infectious challenges, a situation previously described for the endosymbiotic Wolbachia bacteria, a guard against viral infections.},
keywords = {Drosophila melanogaster, ferrandon, Innate immune response, M3i, Pseudomonas aeruginosa},
pubstate = {published},
tppubtype = {article}
}
Microbial pathogenicity often depends on the route of infection. For instance, P. aeruginosa or S. marcescens cause acute systemic infections when low numbers of bacteria are injected into D. melanogaster flies whereas flies succumb much slower to the continuous ingestion of these pathogens, even though both manage to escape from the gut compartment and reach the hemocoel. Here, we have developed a latent P. aeruginosa infection model by feeding flies on the bacteria for a short period. The bacteria stably colonize internal tissues yet hardly cause any damage since latently-infected flies live almost as long as noninfected control flies. The apparently dormant bacteria display particular characteristics in terms of bacterial colony morphology, composition of the outer cell wall, and motility. The virulence of these bacteria can however be reactivated upon wounding the host. We show that melanization but not the cellular or the systemic humoral response is the predominant host defense that establishes latency and may coerce the bacteria to a dormant state. In addition, the lasting activation of the melanization responses in latently-infected flies provides a degree of protection to the host against a secondary fungal infection. Latent infection by an ingested pathogen protects against a variety of homologous or heterologous systemic secondary infectious challenges, a situation previously described for the endosymbiotic Wolbachia bacteria, a guard against viral infections.
Han Min-Jin, Luo Chaorui, Hu Hai, Lin Meixing, Lu Kunpeng, Shen Jianghong, Ren Jianyu, Ye Yanzhuo, Westhof Eric, Tong Xiaoling, Dai Fangyin
Multiple independent origins of the female W chromosome in moths and butterflies Article de journal
Dans: Sci Adv, vol. 10, no. 25, p. eadm9851, 2024, ISSN: 2375-2548.
Résumé | Liens | BibTeX | Étiquettes: Unité ARN, WESTHOF
@article{pmid38896616,
title = {Multiple independent origins of the female W chromosome in moths and butterflies},
author = {Min-Jin Han and Chaorui Luo and Hai Hu and Meixing Lin and Kunpeng Lu and Jianghong Shen and Jianyu Ren and Yanzhuo Ye and Eric Westhof and Xiaoling Tong and Fangyin Dai},
doi = {10.1126/sciadv.adm9851},
issn = {2375-2548},
year = {2024},
date = {2024-06-01},
urldate = {2024-06-01},
journal = {Sci Adv},
volume = {10},
number = {25},
pages = {eadm9851},
abstract = {Lepidoptera, the most diverse group of insects, exhibit female heterogamy (Z0 or ZW), which is different from most other insects (male heterogamy, XY). Previous studies suggest a single origin of the Z chromosome. However, the origin of the lepidopteran W chromosome remains poorly understood. Here, we assemble the genome from females down to the chromosome level of a model insect () and identify a W chromosome of approximately 10.1 megabase using a newly developed tool. In addition, we identify 3593 genes that were not previously annotated in the genomes of . Comparisons of 21 lepidopteran species (including 17 ZW and four Z0 systems) and three trichopteran species (Z0 system) reveal that the formation of Ditrysia W involves multiple mechanisms, including previously proposed canonical and noncanonical models, as well as a newly proposed mechanism called single-Z turnover. We conclude that there are multiple independent origins of the W chromosome in the Ditrysia (most moths and all butterflies) of Lepidoptera.},
keywords = {Unité ARN, WESTHOF},
pubstate = {published},
tppubtype = {article}
}
Lepidoptera, the most diverse group of insects, exhibit female heterogamy (Z0 or ZW), which is different from most other insects (male heterogamy, XY). Previous studies suggest a single origin of the Z chromosome. However, the origin of the lepidopteran W chromosome remains poorly understood. Here, we assemble the genome from females down to the chromosome level of a model insect () and identify a W chromosome of approximately 10.1 megabase using a newly developed tool. In addition, we identify 3593 genes that were not previously annotated in the genomes of . Comparisons of 21 lepidopteran species (including 17 ZW and four Z0 systems) and three trichopteran species (Z0 system) reveal that the formation of Ditrysia W involves multiple mechanisms, including previously proposed canonical and noncanonical models, as well as a newly proposed mechanism called single-Z turnover. We conclude that there are multiple independent origins of the W chromosome in the Ditrysia (most moths and all butterflies) of Lepidoptera.
Holvec Samuel, Barchet Charles, Lechner Antony, Fréchin Léo, Silva S Nimali T De, Hazemann Isabelle, Wolff Philippe, von Loeffelholz Ottilie, Klaholz Bruno P
The structure of the human 80S ribosome at 1.9 Å resolution reveals the molecular role of chemical modifications and ions in RNA Article de journal
Dans: Nat Struct Mol Biol, 2024, ISSN: 1545-9985.
Résumé | Liens | BibTeX | Étiquettes: ARN-MS, Unité ARN
@article{pmid38844527,
title = {The structure of the human 80S ribosome at 1.9 Å resolution reveals the molecular role of chemical modifications and ions in RNA},
author = {Samuel Holvec and Charles Barchet and Antony Lechner and Léo Fréchin and S Nimali T De Silva and Isabelle Hazemann and Philippe Wolff and Ottilie von Loeffelholz and Bruno P Klaholz},
doi = {10.1038/s41594-024-01274-x},
issn = {1545-9985},
year = {2024},
date = {2024-06-01},
urldate = {2024-06-01},
journal = {Nat Struct Mol Biol},
abstract = {The ribosomal RNA of the human protein synthesis machinery comprises numerous chemical modifications that are introduced during ribosome biogenesis. Here we present the 1.9 Å resolution cryo electron microscopy structure of the 80S human ribosome resolving numerous new ribosomal RNA modifications and functionally important ions such as Zn, K and Mg, including their associated individual water molecules. The 2'-O-methylation, pseudo-uridine and base modifications were confirmed by mass spectrometry, resulting in a complete investigation of the >230 sites, many of which could not be addressed previously. They choreograph key interactions within the RNA and at the interface with proteins, including at the ribosomal subunit interfaces of the fully assembled 80S ribosome. Uridine isomerization turns out to be a key mechanism for U-A base pair stabilization in RNA in general. The structural environment of chemical modifications and ions is primordial for the RNA architecture of the mature human ribosome, hence providing a structural framework to address their role in healthy states and in human diseases.},
keywords = {ARN-MS, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
The ribosomal RNA of the human protein synthesis machinery comprises numerous chemical modifications that are introduced during ribosome biogenesis. Here we present the 1.9 Å resolution cryo electron microscopy structure of the 80S human ribosome resolving numerous new ribosomal RNA modifications and functionally important ions such as Zn, K and Mg, including their associated individual water molecules. The 2'-O-methylation, pseudo-uridine and base modifications were confirmed by mass spectrometry, resulting in a complete investigation of the >230 sites, many of which could not be addressed previously. They choreograph key interactions within the RNA and at the interface with proteins, including at the ribosomal subunit interfaces of the fully assembled 80S ribosome. Uridine isomerization turns out to be a key mechanism for U-A base pair stabilization in RNA in general. The structural environment of chemical modifications and ions is primordial for the RNA architecture of the mature human ribosome, hence providing a structural framework to address their role in healthy states and in human diseases.
Urzhumtseva Ludmila, Barchet Charles, Klaholz Bruno P, Urzhumtsev Alexandre G
Program : analysing distributions of cryo-EM projections using uniform spherical grids Article de journal
Dans: J Appl Crystallogr, vol. 57, no. Pt 3, p. 865–876, 2024, ISSN: 0021-8898.
Résumé | Liens | BibTeX | Étiquettes: Unité ARN
@article{pmid38846771,
title = {Program : analysing distributions of cryo-EM projections using uniform spherical grids},
author = {Ludmila Urzhumtseva and Charles Barchet and Bruno P Klaholz and Alexandre G Urzhumtsev},
doi = {10.1107/S1600576724002383},
issn = {0021-8898},
year = {2024},
date = {2024-06-01},
urldate = {2024-06-01},
journal = {J Appl Crystallogr},
volume = {57},
number = {Pt 3},
pages = {865--876},
abstract = {Three-dimensional cryo electron microscopy reconstructions are obtained by extracting information from a large number of projections of the object. These projections correspond to different 'views' or 'orientations', directions in which these projections show the reconstructed object. Uneven distribution of these views and the presence of dominating preferred orientations may distort the reconstructed spatial images. This work describes the program (views on uniform grids for cryo electron microscopy), designed to study such distributions. Its algorithms, based on uniform virtual grids on a sphere, allow an easy calculation and accurate quantitative analysis of the frequency distribution of the views. The key computational element is the Lambert azimuthal equal-area projection of a spherical uniform grid onto a disc. This projection keeps the surface area constant and represents the frequency distribution with no visual bias. Since it has multiple tunable parameters, the program is easily adaptable to individual needs, and to the features of a particular project or of the figure to be produced. It can help identify problems related to an uneven distribution of views. Optionally, it can modify the list of projections, distributing the views more uniformly. The program can also be used as a teaching tool.},
keywords = {Unité ARN},
pubstate = {published},
tppubtype = {article}
}
Three-dimensional cryo electron microscopy reconstructions are obtained by extracting information from a large number of projections of the object. These projections correspond to different 'views' or 'orientations', directions in which these projections show the reconstructed object. Uneven distribution of these views and the presence of dominating preferred orientations may distort the reconstructed spatial images. This work describes the program (views on uniform grids for cryo electron microscopy), designed to study such distributions. Its algorithms, based on uniform virtual grids on a sphere, allow an easy calculation and accurate quantitative analysis of the frequency distribution of the views. The key computational element is the Lambert azimuthal equal-area projection of a spherical uniform grid onto a disc. This projection keeps the surface area constant and represents the frequency distribution with no visual bias. Since it has multiple tunable parameters, the program is easily adaptable to individual needs, and to the features of a particular project or of the figure to be produced. It can help identify problems related to an uneven distribution of views. Optionally, it can modify the list of projections, distributing the views more uniformly. The program can also be used as a teaching tool.
Lim Shey-Li, Liu Jinhong, Dupouy Gilles, Singh Gaurav, Baudrey Stéphanie, Yang Lang, Zhong Jia Yi, Chabouté Marie-Edith, Lim Boon Leong
In planta imaging of pyridine nucleotides using second-generation fluorescent protein biosensors Article de journal
Dans: Plant J, 2024, ISSN: 1365-313X.
Résumé | Liens | BibTeX | Étiquettes: RYCKELYNCK, Unité ARN
@article{pmid38761168,
title = {In planta imaging of pyridine nucleotides using second-generation fluorescent protein biosensors},
author = {Shey-Li Lim and Jinhong Liu and Gilles Dupouy and Gaurav Singh and Stéphanie Baudrey and Lang Yang and Jia Yi Zhong and Marie-Edith Chabouté and Boon Leong Lim},
doi = {10.1111/tpj.16796},
issn = {1365-313X},
year = {2024},
date = {2024-05-01},
urldate = {2024-05-01},
journal = {Plant J},
abstract = {Redox changes of pyridine nucleotides in cellular compartments are highly dynamic and their equilibria are under the influence of various reducing and oxidizing reactions. To obtain spatiotemporal data on pyridine nucleotides in living plant cells, typical biochemical approaches require cell destruction. To date, genetically encoded fluorescent biosensors are considered to be the best option to bridge the existing technology gap, as they provide a fast, accurate, and real-time readout. However, the existing pyridine nucleotides genetically encoded fluorescent biosensors are either sensitive to pH change or slow in dissociation rate. Herein, we employed the biosensors which generate readouts that are pH stable for in planta measurement of NADH/NAD ratio and NADPH level. We generated transgenic Arabidopsis lines that express these biosensors in plastid stroma and cytosol of whole plants and pollen tubes under the control of CaMV 35S and LAT52 promoters, respectively. These transgenic biosensor lines allow us to monitor real-time dynamic changes in NADH/NAD ratio and NADPH level in the plastids and cytosol of various plant tissues, including pollen tubes, root hairs, and mesophyll cells, using a variety of fluorescent instruments. We anticipate that these valuable transgenic lines may allow improvements in plant redox biology studies.},
keywords = {RYCKELYNCK, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
Redox changes of pyridine nucleotides in cellular compartments are highly dynamic and their equilibria are under the influence of various reducing and oxidizing reactions. To obtain spatiotemporal data on pyridine nucleotides in living plant cells, typical biochemical approaches require cell destruction. To date, genetically encoded fluorescent biosensors are considered to be the best option to bridge the existing technology gap, as they provide a fast, accurate, and real-time readout. However, the existing pyridine nucleotides genetically encoded fluorescent biosensors are either sensitive to pH change or slow in dissociation rate. Herein, we employed the biosensors which generate readouts that are pH stable for in planta measurement of NADH/NAD ratio and NADPH level. We generated transgenic Arabidopsis lines that express these biosensors in plastid stroma and cytosol of whole plants and pollen tubes under the control of CaMV 35S and LAT52 promoters, respectively. These transgenic biosensor lines allow us to monitor real-time dynamic changes in NADH/NAD ratio and NADPH level in the plastids and cytosol of various plant tissues, including pollen tubes, root hairs, and mesophyll cells, using a variety of fluorescent instruments. We anticipate that these valuable transgenic lines may allow improvements in plant redox biology studies.
Omar Reine El, Abdellaoui Naoill, Coulibaly Safiatou T, Fontenille Laura, Lanza François, Gachet Christian, Freund Jean-Noel, Negroni Matteo, Kissa Karima, Tavian Manuela
Macrophage depletion overcomes human hematopoietic cell engraftment failure in zebrafish embryo Article de journal
Dans: Cell Death Dis, vol. 15, no. 5, p. 305, 2024, ISSN: 2041-4889.
Résumé | Liens | BibTeX | Étiquettes: NEGRONI, Unité ARN
@article{pmid38693109,
title = {Macrophage depletion overcomes human hematopoietic cell engraftment failure in zebrafish embryo},
author = {Reine El Omar and Naoill Abdellaoui and Safiatou T Coulibaly and Laura Fontenille and François Lanza and Christian Gachet and Jean-Noel Freund and Matteo Negroni and Karima Kissa and Manuela Tavian},
doi = {10.1038/s41419-024-06682-x},
issn = {2041-4889},
year = {2024},
date = {2024-05-01},
urldate = {2024-05-01},
journal = {Cell Death Dis},
volume = {15},
number = {5},
pages = {305},
abstract = {Zebrafish is widely adopted as a grafting model for studying human development and diseases. Current zebrafish xenotransplantations are performed using embryo recipients, as the adaptive immune system, responsible for host versus graft rejection, only reaches maturity at juvenile stage. However, transplanted primary human hematopoietic stem/progenitor cells (HSC) rapidly disappear even in zebrafish embryos, suggesting that another barrier to transplantation exists before the onset of adaptive immunity. Here, using a labelled macrophage zebrafish line, we demonstrated that engraftment of human HSC induces a massive recruitment of macrophages which rapidly phagocyte transplanted cells. Macrophages depletion, by chemical or pharmacological treatments, significantly improved the uptake and survival of transplanted cells, demonstrating the crucial implication of these innate immune cells for the successful engraftment of human cells in zebrafish. Beyond identifying the reasons for human hematopoietic cell engraftment failure, this work images the fate of human cells in real time over several days in macrophage-depleted zebrafish embryos.},
keywords = {NEGRONI, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
Zebrafish is widely adopted as a grafting model for studying human development and diseases. Current zebrafish xenotransplantations are performed using embryo recipients, as the adaptive immune system, responsible for host versus graft rejection, only reaches maturity at juvenile stage. However, transplanted primary human hematopoietic stem/progenitor cells (HSC) rapidly disappear even in zebrafish embryos, suggesting that another barrier to transplantation exists before the onset of adaptive immunity. Here, using a labelled macrophage zebrafish line, we demonstrated that engraftment of human HSC induces a massive recruitment of macrophages which rapidly phagocyte transplanted cells. Macrophages depletion, by chemical or pharmacological treatments, significantly improved the uptake and survival of transplanted cells, demonstrating the crucial implication of these innate immune cells for the successful engraftment of human cells in zebrafish. Beyond identifying the reasons for human hematopoietic cell engraftment failure, this work images the fate of human cells in real time over several days in macrophage-depleted zebrafish embryos.
PJ Thiele, R Mela-Lopez, SA Blandin, D Klug
Let it glow: genetically encoded fluorescent reporters in Plasmodium Article de journal
Dans: Malaria Journal, vol. 23, no. 114, 2024.
Résumé | Liens | BibTeX | Étiquettes: blandin, fluorescent reporters, M3i, Plasmodium
@article{PJ2024,
title = {Let it glow: genetically encoded fluorescent reporters in Plasmodium},
author = {Thiele PJ and Mela-Lopez R and Blandin SA and Klug D},
editor = {Springer Nature},
doi = {10.1186/s12936-024-04936-9},
year = {2024},
date = {2024-04-20},
urldate = {2024-04-20},
journal = {Malaria Journal},
volume = {23},
number = {114},
abstract = {The use of fluorescent proteins (FPs) in Plasmodium parasites has been key to understand the biology of this obligate intracellular protozoon. FPs like the green fluorescent protein (GFP) enabled to explore protein localization, promoter activity as well as dynamic processes like protein export and endocytosis. Furthermore, FP biosensors have provided detailed information on physiological parameters at the subcellular level, and fluorescent reporter lines greatly extended the malariology toolbox. Still, in order to achieve optimal results, it is crucial to know exactly the properties of the FP of choice and the genetic scenario in which it will be used. This review highlights advantages and disadvantages of available landing sites and promoters that have been successfully applied for the ectopic expression of FPs in Plasmodium berghei and Plasmodium falciparum. Furthermore, the properties of newly developed FPs beyond DsRed and EGFP, in the visualization of cells and cellular structures as well as in the sensing of small molecules are discussed.},
keywords = {blandin, fluorescent reporters, M3i, Plasmodium},
pubstate = {published},
tppubtype = {article}
}
The use of fluorescent proteins (FPs) in Plasmodium parasites has been key to understand the biology of this obligate intracellular protozoon. FPs like the green fluorescent protein (GFP) enabled to explore protein localization, promoter activity as well as dynamic processes like protein export and endocytosis. Furthermore, FP biosensors have provided detailed information on physiological parameters at the subcellular level, and fluorescent reporter lines greatly extended the malariology toolbox. Still, in order to achieve optimal results, it is crucial to know exactly the properties of the FP of choice and the genetic scenario in which it will be used. This review highlights advantages and disadvantages of available landing sites and promoters that have been successfully applied for the ectopic expression of FPs in Plasmodium berghei and Plasmodium falciparum. Furthermore, the properties of newly developed FPs beyond DsRed and EGFP, in the visualization of cells and cellular structures as well as in the sensing of small molecules are discussed.
I Iacobucci, V Monaco, A Hovasse, B Dupouy, R Keumoe, B Cichocki, M Elhabiri, B Meunier, JM Strub, M Monti, S Cianférani, SA Blandin, C Schaeffer-Reiss, E Davioud-Charvet
Proteomic Profiling of Antimalarial Plasmodione Using 3-Benz(o)ylmenadione Affinity-Based Probes Article de journal
Dans: Chembiochem, 2024.
Résumé | Liens | BibTeX | Étiquettes: Activity-Based Protein Profiling, Antiprotozoal agents, Biological Chemistry, blandin, Chemical Biology, M3i, Photoaffinity Labeling, proteomics
@article{I2024,
title = {Proteomic Profiling of Antimalarial Plasmodione Using 3-Benz(o)ylmenadione Affinity-Based Probes},
author = {Iacobucci I and Monaco V and Hovasse A AND Dupouy B AND Keumoe R AND Cichocki B AND Elhabiri M AND Meunier B AND Strub JM AND Monti M AND Cianférani S AND Blandin SA AND Schaeffer-Reiss C AND Davioud-Charvet E},
editor = {Wiley-VCH GmbH},
url = {https://pubmed.ncbi.nlm.nih.gov/38639212/},
doi = {10.1002/cbic.202400187},
year = {2024},
date = {2024-04-19},
journal = {Chembiochem},
abstract = {Understanding the mechanisms of drug action in malarial parasites is crucial for the development of new drugs to combat infection and to counteract drug resistance. Proteomics is a widely used approach to study host-pathogen systems and to identify drug protein targets. Plasmodione is an antiplasmodial early-lead drug exerting potent activities against young asexual and sexual blood stages in vitro with low toxicity to host cells. To elucidate its molecular mechanisms, an affinity-based protein profiling (AfBPP) approach was applied to yeast and P. falciparum proteomes. New (pro-) AfBPP probes based on the 3-benz(o)yl-6-fluoro-menadione scaffold were synthesized. With optimized conditions of both photoaffinity labeling and click reaction steps, the AfBPP protocol was then applied to a yeast proteome, yielding 11 putative drug-protein targets. Among these, we found four proteins associated with oxidoreductase activities, the hypothesized type of targets for plasmodione and its metabolites, and other proteins associated with the mitochondria. In Plasmodium parasites, the MS analysis revealed 44 potential plasmodione targets that need to be validated in further studies. Finally, the localization of a 3-benzyl-6-fluoromenadione AfBPP probe was studied in the subcellular structures of the parasite at the trophozoite stage. },
keywords = {Activity-Based Protein Profiling, Antiprotozoal agents, Biological Chemistry, blandin, Chemical Biology, M3i, Photoaffinity Labeling, proteomics},
pubstate = {published},
tppubtype = {article}
}
Understanding the mechanisms of drug action in malarial parasites is crucial for the development of new drugs to combat infection and to counteract drug resistance. Proteomics is a widely used approach to study host-pathogen systems and to identify drug protein targets. Plasmodione is an antiplasmodial early-lead drug exerting potent activities against young asexual and sexual blood stages in vitro with low toxicity to host cells. To elucidate its molecular mechanisms, an affinity-based protein profiling (AfBPP) approach was applied to yeast and P. falciparum proteomes. New (pro-) AfBPP probes based on the 3-benz(o)yl-6-fluoro-menadione scaffold were synthesized. With optimized conditions of both photoaffinity labeling and click reaction steps, the AfBPP protocol was then applied to a yeast proteome, yielding 11 putative drug-protein targets. Among these, we found four proteins associated with oxidoreductase activities, the hypothesized type of targets for plasmodione and its metabolites, and other proteins associated with the mitochondria. In Plasmodium parasites, the MS analysis revealed 44 potential plasmodione targets that need to be validated in further studies. Finally, the localization of a 3-benzyl-6-fluoromenadione AfBPP probe was studied in the subcellular structures of the parasite at the trophozoite stage.
Niedner-Boblenz A., Monecke T., Hennig J., Klostermann M., Hofweber M., Gerber A. P., Anosova I., Mayer W., Müller M., Heym R., Janowski R., Paillart J. -C., Dormann D., Zarnack K., Sattler M., Niessing D.
Intrinsically disordered RNA-binding motifs cooperate to catalyze RNA folding and drive phase separation Article de journal
Dans: bioRxiv, 2024.
Résumé | Liens | BibTeX | Étiquettes: MARQUET, PAILLART, Unité ARN
@article{A.2024,
title = {Intrinsically disordered RNA-binding motifs cooperate to catalyze RNA folding and drive phase separation},
author = {A. Niedner-Boblenz and T. Monecke and J. Hennig and M. Klostermann and M. Hofweber and A.P. Gerber and I. Anosova and W. Mayer and M. Müller and R. Heym and R. Janowski and J.-C. Paillart and D. Dormann and K. Zarnack and M. Sattler and D. Niessing},
url = {https://www.biorxiv.org/content/10.1101/2024.03.27.586925v2},
doi = {10.1101/2024.03.27.586925},
year = {2024},
date = {2024-03-29},
urldate = {2024-03-29},
journal = {bioRxiv},
abstract = {RNA-binding proteins are essential for gene regulation and the spatial organization of cells. Here, we report that the yeast ribosome biogenesis factor Loc1p is an intrinsically disordered RNA-binding protein with eight repeating positively charged, unstructured nucleic acid binding (PUN) motifs. While a single of these previously undefined motifs stabilizes folded RNAs, multiple copies strongly cooperate to catalyze RNA folding. In the presence of RNA, these multivalent PUN motifs drive phase separation. Proteome-wide searches in pro-and eukaryotes for proteins with similar arrays of PUN motifs reveal a strong enrichment in RNA-mediated processes and DNA remodeling. Thus, PUN motifs are potentially involved in a large variety of RNA-and DNA-related processes by concentrating them in membrane-less organelles. The general function and wide distribution of PUN motifs across species suggests that in an ancient “RNA world” PUN-like motifs may have supported the correct folding of early ribozymes.},
keywords = {MARQUET, PAILLART, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
RNA-binding proteins are essential for gene regulation and the spatial organization of cells. Here, we report that the yeast ribosome biogenesis factor Loc1p is an intrinsically disordered RNA-binding protein with eight repeating positively charged, unstructured nucleic acid binding (PUN) motifs. While a single of these previously undefined motifs stabilizes folded RNAs, multiple copies strongly cooperate to catalyze RNA folding. In the presence of RNA, these multivalent PUN motifs drive phase separation. Proteome-wide searches in pro-and eukaryotes for proteins with similar arrays of PUN motifs reveal a strong enrichment in RNA-mediated processes and DNA remodeling. Thus, PUN motifs are potentially involved in a large variety of RNA-and DNA-related processes by concentrating them in membrane-less organelles. The general function and wide distribution of PUN motifs across species suggests that in an ancient “RNA world” PUN-like motifs may have supported the correct folding of early ribozymes.
Imler Jean-Luc, Cai Hua, Meignin Carine, Martins Nelson
Evolutionary immunology to explore original antiviral strategies Article de journal
Dans: Philosophical Transactions B, vol. 379, iss. 1901, 2024.
Résumé | Liens | BibTeX | Étiquettes: antiviral, imler, M3i, meignin
@article{Martins2024,
title = {Evolutionary immunology to explore original antiviral strategies},
author = {Jean-Luc Imler AND Hua Cai AND Carine Meignin AND Nelson Martins},
url = {https://royalsocietypublishing.org/doi/10.1098/rstb.2023.0068},
doi = {10.1098/rstb.2023.0068},
year = {2024},
date = {2024-03-18},
urldate = {2024-03-18},
journal = {Philosophical Transactions B},
volume = {379},
issue = {1901},
abstract = {Over the past 25 years, the field of evolutionary developmental biology (evo–devo) has used genomics and genetics to gain insight on the developmental mechanisms underlying the evolution of morphological diversity of animals. Evo–devo exploits the key insight that conserved toolkits of development (e.g. Hox genes) are used in animals to produce genetic novelties that provide adaptation to a new environment. Like development, immunity is forged by interactions with the environment, namely the microbial world. Yet, when it comes to the study of immune defence mechanisms in invertebrates, interest primarily focuses on evolutionarily conserved molecules also present in humans. Here, focusing on antiviral immunity, we argue that immune genes not conserved in humans represent an unexplored resource for the discovery of new antiviral strategies. We review recent findings on the cGAS-STING pathway and explain how cyclic dinucleotides produced by cGAS-like receptors may be used to investigate the portfolio of antiviral genes in a broad range of species. This will set the stage for evo–immuno approaches, exploiting the investment in antiviral defences made by metazoans over hundreds of millions of years of evolution.},
keywords = {antiviral, imler, M3i, meignin},
pubstate = {published},
tppubtype = {article}
}
Over the past 25 years, the field of evolutionary developmental biology (evo–devo) has used genomics and genetics to gain insight on the developmental mechanisms underlying the evolution of morphological diversity of animals. Evo–devo exploits the key insight that conserved toolkits of development (e.g. Hox genes) are used in animals to produce genetic novelties that provide adaptation to a new environment. Like development, immunity is forged by interactions with the environment, namely the microbial world. Yet, when it comes to the study of immune defence mechanisms in invertebrates, interest primarily focuses on evolutionarily conserved molecules also present in humans. Here, focusing on antiviral immunity, we argue that immune genes not conserved in humans represent an unexplored resource for the discovery of new antiviral strategies. We review recent findings on the cGAS-STING pathway and explain how cyclic dinucleotides produced by cGAS-like receptors may be used to investigate the portfolio of antiviral genes in a broad range of species. This will set the stage for evo–immuno approaches, exploiting the investment in antiviral defences made by metazoans over hundreds of millions of years of evolution.
D'Agostino Mattia, Simonetti Angelita, Motta Stefano, Wolff Philippe, Romagnoli Alice, Piccinini Astra, Spinozzi Francesco, Marino Daniele Di, Teana Anna La, Ennifar Eric
Crystal structure of archaeal IF5A-DHS complex reveals insights into the hypusination mechanism Article de journal
Dans: Structure, 2024, ISSN: 1878-4186.
Résumé | Liens | BibTeX | Étiquettes: ARN-MS, ENNIFAR, Unité ARN
@article{pmid38582076,
title = {Crystal structure of archaeal IF5A-DHS complex reveals insights into the hypusination mechanism},
author = {Mattia D'Agostino and Angelita Simonetti and Stefano Motta and Philippe Wolff and Alice Romagnoli and Astra Piccinini and Francesco Spinozzi and Daniele Di Marino and Anna La Teana and Eric Ennifar},
doi = {10.1016/j.str.2024.03.008},
issn = {1878-4186},
year = {2024},
date = {2024-03-01},
urldate = {2024-03-01},
journal = {Structure},
abstract = {The translation factor IF5A is highly conserved in Eukarya and Archaea and undergoes a unique post-translational hypusine modification by the deoxyhypusine synthase (DHS) enzyme. DHS transfers the butylamine moiety from spermidine to IF5A using NAD as a cofactor, forming a deoxyhypusine intermediate. IF5A is a key player in protein synthesis, preventing ribosome stalling in proline-rich sequences during translation elongation and facilitating translation elongation and termination. Additionally, human eIF5A participates in various essential cellular processes and contributes to cancer metastasis, with inhibiting hypusination showing anti-proliferative effects. The hypusination pathway of IF5A is therefore an attractive new therapeutic target. We elucidated the 2.0 Å X-ray crystal structure of the archaeal DHS-IF5A complex, revealing hetero-octameric architecture and providing a detailed view of the complex active site including the hypusination loop. This structure, along with biophysical data and molecular dynamics simulations, provides new insights into the catalytic mechanism of the hypusination reaction.},
keywords = {ARN-MS, ENNIFAR, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
The translation factor IF5A is highly conserved in Eukarya and Archaea and undergoes a unique post-translational hypusine modification by the deoxyhypusine synthase (DHS) enzyme. DHS transfers the butylamine moiety from spermidine to IF5A using NAD as a cofactor, forming a deoxyhypusine intermediate. IF5A is a key player in protein synthesis, preventing ribosome stalling in proline-rich sequences during translation elongation and facilitating translation elongation and termination. Additionally, human eIF5A participates in various essential cellular processes and contributes to cancer metastasis, with inhibiting hypusination showing anti-proliferative effects. The hypusination pathway of IF5A is therefore an attractive new therapeutic target. We elucidated the 2.0 Å X-ray crystal structure of the archaeal DHS-IF5A complex, revealing hetero-octameric architecture and providing a detailed view of the complex active site including the hypusination loop. This structure, along with biophysical data and molecular dynamics simulations, provides new insights into the catalytic mechanism of the hypusination reaction.
Messmer Mélanie, Pierson Louison, Pasquier Charline, Djordjevic Nikola, Chicher Johana, Hammann Philippe, Pfeffer Sébastien, Girardi Erika
DEAD box RNA helicase 5 is a new pro-viral host factor for Sindbis virus infection Article de journal
Dans: Virol J, vol. 21, no. 1, p. 76, 2024, ISSN: 1743-422X.
Résumé | Liens | BibTeX | Étiquettes: PFEFFER, PPSE, Unité ARN
@article{pmid38553727,
title = {DEAD box RNA helicase 5 is a new pro-viral host factor for Sindbis virus infection},
author = {Mélanie Messmer and Louison Pierson and Charline Pasquier and Nikola Djordjevic and Johana Chicher and Philippe Hammann and Sébastien Pfeffer and Erika Girardi},
doi = {10.1186/s12985-024-02349-3},
issn = {1743-422X},
year = {2024},
date = {2024-03-01},
urldate = {2024-03-01},
journal = {Virol J},
volume = {21},
number = {1},
pages = {76},
abstract = {BACKGROUND: RNA helicases are emerging as key factors regulating host-virus interactions. The DEAD-box ATP-dependent RNA helicase DDX5, which plays an important role in many aspects of cellular RNA biology, was also found to either promote or inhibit viral replication upon infection with several RNA viruses. Here, our aim is to examine the impact of DDX5 on Sindbis virus (SINV) infection.nnMETHODS: We analysed the interaction between DDX5 and the viral RNA using imaging and RNA-immunoprecipitation approaches. The interactome of DDX5 in mock- and SINV-infected cells was determined by mass spectrometry. We validated the interaction between DDX17 and the viral capsid by co- immunoprecipitation in the presence or absence of an RNase treatment. We determined the subcellular localization of DDX5, its cofactor DDX17 and the viral capsid protein by co-immunofluorescence. Finally, we investigated the impact of DDX5 depletion and overexpression on SINV infection at the viral protein, RNA and infectious particle accumulation level. The contribution of DDX17 was also tested by knockdown experiments.nnRESULTS: In this study we demonstrate that DDX5 interacts with the SINV RNA during infection. Furthermore, the proteomic analysis of the DDX5 interactome in mock and SINV-infected HCT116 cells identified new cellular and viral partners and confirmed the interaction between DDX5 and DDX17. Both DDX5 and DDX17 re-localize from the nucleus to the cytoplasm upon SINV infection and interact with the viral capsid protein. We also show that DDX5 depletion negatively impacts the viral replication cycle, while its overexpression has a pro-viral effect. Finally, we observed that DDX17 depletion reduces SINV infection, an effect which is even more pronounced in a DDX5-depleted background, suggesting a synergistic pro-viral effect of the DDX5 and DDX17 proteins on SINV.nnCONCLUSIONS: These results not only shed light on DDX5 as a novel and important host factor to the SINV life cycle, but also expand our understanding of the roles played by DDX5 and DDX17 as regulators of viral infections.},
keywords = {PFEFFER, PPSE, Unité ARN},
pubstate = {published},
tppubtype = {article}
}
BACKGROUND: RNA helicases are emerging as key factors regulating host-virus interactions. The DEAD-box ATP-dependent RNA helicase DDX5, which plays an important role in many aspects of cellular RNA biology, was also found to either promote or inhibit viral replication upon infection with several RNA viruses. Here, our aim is to examine the impact of DDX5 on Sindbis virus (SINV) infection.nnMETHODS: We analysed the interaction between DDX5 and the viral RNA using imaging and RNA-immunoprecipitation approaches. The interactome of DDX5 in mock- and SINV-infected cells was determined by mass spectrometry. We validated the interaction between DDX17 and the viral capsid by co- immunoprecipitation in the presence or absence of an RNase treatment. We determined the subcellular localization of DDX5, its cofactor DDX17 and the viral capsid protein by co-immunofluorescence. Finally, we investigated the impact of DDX5 depletion and overexpression on SINV infection at the viral protein, RNA and infectious particle accumulation level. The contribution of DDX17 was also tested by knockdown experiments.nnRESULTS: In this study we demonstrate that DDX5 interacts with the SINV RNA during infection. Furthermore, the proteomic analysis of the DDX5 interactome in mock and SINV-infected HCT116 cells identified new cellular and viral partners and confirmed the interaction between DDX5 and DDX17. Both DDX5 and DDX17 re-localize from the nucleus to the cytoplasm upon SINV infection and interact with the viral capsid protein. We also show that DDX5 depletion negatively impacts the viral replication cycle, while its overexpression has a pro-viral effect. Finally, we observed that DDX17 depletion reduces SINV infection, an effect which is even more pronounced in a DDX5-depleted background, suggesting a synergistic pro-viral effect of the DDX5 and DDX17 proteins on SINV.nnCONCLUSIONS: These results not only shed light on DDX5 as a novel and important host factor to the SINV life cycle, but also expand our understanding of the roles played by DDX5 and DDX17 as regulators of viral infections.
Pellequer Jean-Luc, Westhof Eric
Marc van Regenmortel, personal recollections on a forward-thinking editor Article de journal
Dans: J Mol Recognit, p. e3080, 2024, ISSN: 1099-1352.
Résumé | Liens | BibTeX | Étiquettes: Unité ARN, WESTHOF
@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}
}
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.
L Hédelin, A Thiebaut, J Huang, X Li, A Lemoine, G Haas, C Meignin, H Cai, R Waterhouse, N Martins, JL Imler
Investigating the Evolution of Drosophila STING-Dependent Antiviral Innate Immunity by Multispecies Comparison of 2′3′-cGAMP Responses Article de journal
Dans: Molecular Biology and Evolution, vol. 41, iss. 3, 2024.
Résumé | Liens | BibTeX | Étiquettes: 2'3'-cGAMP, cGAS/STING, Drosophila, evo-immuno, imler, M3i, meignin, transcriptome
@article{JL2024,
title = {Investigating the Evolution of Drosophila STING-Dependent Antiviral Innate Immunity by Multispecies Comparison of 2′3′-cGAMP Responses},
author = {Hédelin L AND Thiebaut A AND Huang J AND Li X AND Lemoine A AND Haas G AND Meignin C AND Cai H AND Waterhouse R AND Martins N AND Imler JL},
url = {https://academic.oup.com/mbe/article/41/3/msae032/7611634},
doi = {10.1093/molbev/msae032},
year = {2024},
date = {2024-02-20},
urldate = {2024-02-20},
journal = {Molecular Biology and Evolution},
volume = {41},
issue = {3},
abstract = {Viruses represent a major threat to all animals, which defend themselves through induction of a large set of virus-stimulated genes that collectively control the infection. In vertebrates, these genes include interferons that play a critical role in the amplification of the response to infection. Virus- and interferon-stimulated genes include restriction factors targeting the different steps of the viral replication cycle, in addition to molecules associated with inflammation and adaptive immunity. Predictably, antiviral genes evolve dynamically in response to viral pressure. As a result, each animal has a unique arsenal of antiviral genes. Here, we exploit the capacity to experimentally activate the evolutionarily conserved stimulator of IFN genes (STING) signaling pathway by injection of the cyclic dinucleotide 2′3′-cyclic guanosine monophosphate-adenosine monophosphate into flies to define the repertoire of STING-regulated genes in 10 Drosophila species, spanning 40 million years of evolution. Our data reveal a set of conserved STING-regulated factors, including STING itself, a cGAS-like-receptor, the restriction factor pastel, and the antiviral protein Vago, but also 2 key components of the antiviral RNA interference pathway, Dicer-2, and Argonaute2. In addition, we identify unknown species- or lineage-specific genes that have not been previously associated with resistance to viruses. Our data provide insight into the core antiviral response in Drosophila flies and pave the way for the characterization of previously unknown antiviral effectors.},
keywords = {2'3'-cGAMP, cGAS/STING, Drosophila, evo-immuno, imler, M3i, meignin, transcriptome},
pubstate = {published},
tppubtype = {article}
}
Viruses represent a major threat to all animals, which defend themselves through induction of a large set of virus-stimulated genes that collectively control the infection. In vertebrates, these genes include interferons that play a critical role in the amplification of the response to infection. Virus- and interferon-stimulated genes include restriction factors targeting the different steps of the viral replication cycle, in addition to molecules associated with inflammation and adaptive immunity. Predictably, antiviral genes evolve dynamically in response to viral pressure. As a result, each animal has a unique arsenal of antiviral genes. Here, we exploit the capacity to experimentally activate the evolutionarily conserved stimulator of IFN genes (STING) signaling pathway by injection of the cyclic dinucleotide 2′3′-cyclic guanosine monophosphate-adenosine monophosphate into flies to define the repertoire of STING-regulated genes in 10 Drosophila species, spanning 40 million years of evolution. Our data reveal a set of conserved STING-regulated factors, including STING itself, a cGAS-like-receptor, the restriction factor pastel, and the antiviral protein Vago, but also 2 key components of the antiviral RNA interference pathway, Dicer-2, and Argonaute2. In addition, we identify unknown species- or lineage-specific genes that have not been previously associated with resistance to viruses. Our data provide insight into the core antiviral response in Drosophila flies and pave the way for the characterization of previously unknown antiviral effectors.
Compain Guillaume, Monsarrat Clément, Blagojevic Julie, Brillet Karl, Dumas Philippe, Hammann Philippe, Kuhn Lauriane, Martiel Isabelle, Engilberge Sylvain, Oliéric Vincent, Wolff Philippe, Burnouf Dominique Y, Wagner Jérôme, Guichard Gilles
Peptide-Based Covalent Inhibitors Bearing Mild Electrophiles to Target a Conserved His Residue of the Bacterial Sliding Clamp Article de journal
Dans: JACS Au, vol. 4, no. 2, p. 432–440, 2024, ISSN: 2691-3704.
Résumé | Liens | BibTeX | Étiquettes: ARN-MS, ENNIFAR, PPSE, Unité ARN
@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}
}
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.
Soufi G El, Jorio L Di, Gerber Z, Cluzel N, Assche J Van, Delafoy D, Olaso R, Daviaud C, Loustau T, Schwartz C, Trebouet D, Hernalsteens O, Marechal V, Raffestin S, Rousset D, Lint C Van, Deleuze J F, Boni M, and O Rohr, Villain-Gambier M, Wallet C
Highly efficient and sensitive membrane-based concentration process allows quantification, surveillance, and sequencing of viruses in large volumes of wastewater Article de journal
Dans: Water Res, vol. 249, p. 120959, 2024, ISSN: 1879-2448.
Résumé | Liens | BibTeX | Étiquettes: ROHR, Unité ARN
@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}
}
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.
Brillet Karl, Janczuk-Richter Marta, Poon Amanda, Laukart-Bradley Joanne, Ennifar Eric, Lebars Isabelle
Characterization of SLA RNA promoter from dengue virus and its interaction with the viral non-structural NS5 protein Article de journal
Dans: Biochimie, vol. 222, p. 87–100, 2024, ISSN: 1638-6183.
Résumé | Liens | BibTeX | Étiquettes: ENNIFAR, Unité ARN
@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}
}
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.
Zhang Jian-Hui, Eriani Gilbert, Zhou Xiao-Long
Pathophysiology of human mitochondrial tRNA metabolism Article de journal
Dans: Trends Endocrinol Metab, 2024, ISSN: 1879-3061.
Résumé | Liens | BibTeX | Étiquettes: ERIANI, Unité ARN
@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}
}
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.
CF Estevez-Castro, MF Rodrigues, A Babarit, FV Ferreira, de Andrade EG, E Marois, R Cogni, ERGR Aguiar, JT Marques, RP Olmo
Neofunctionalization driven by positive selection led to the retention of the loqs2 gene encoding an Aedes specific dsRNA binding protein Article de journal
Dans: BMC Biol , vol. 22, no. 14, 2024.
Résumé | Liens | BibTeX | Étiquettes: Aedes mosquitoes, Double-stranded RNA (dsRNA), dsRNA binding protein (dsRBP), loqs2, M3i, marois, Marques, Olmo, RNA interference (RNAi)
@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}
}
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.
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.
Green Emily I, Jaouen Etienne, Klug Dennis, Olmo Roenick Proveti, Gautier Amandine, Blandin Stéphanie, Marois Eric
A population modification gene drive targeting both Saglin and Lipophorin disables Plasmodium transmission in Anopheles mosquitoes Article de journal
Dans: Genetics and Genomics, 2024.
Résumé | Liens | BibTeX | Étiquettes: Anopheles, blandin, gene drive, Lipophorin, M3i, marois, marque, Marques, Olmo, Plasmodium, Saglin
@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}
}
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.
Krishnan A., Ali L. M., Prabhu S. G., Pillai V. N., Chameettachal A., Vivet-Boudou V., Bernacchi S., Mustafa F., Marquet R., Rizvi T. A.
Identification of a putative Gag binding site critical for feline immunodeficiency virus genomic RNA packaging Article de journal
Dans: RNA Journal, vol. 30, p. 68-88, 2024.
Résumé | Liens | BibTeX | Étiquettes: bernacchi, MARQUET, Unité ARN, vivet-boudou
@article{nokey,
title = {Identification of a putative Gag binding site critical for feline immunodeficiency virus genomic RNA packaging},
author = {A. Krishnan and L.M. Ali and S.G. Prabhu and V.N. Pillai and A. Chameettachal and V. Vivet-Boudou and S. Bernacchi and F. Mustafa and R. Marquet and T.A. Rizvi},
editor = {cold spring harbor laboratory press},
url = {https://rnajournal.cshlp.org/content/30/1/68.long},
doi = {10.1261/rna.079840.123},
year = {2024},
date = {2024-01-10},
urldate = {2024-01-10},
journal = {RNA Journal},
volume = {30},
pages = {68-88},
abstract = {The retroviral Gag precursor plays a central role in the selection and packaging of viral genomic RNA (gRNA) by binding to
virus-specific packaging signal(s) (psi or ψ). Previously, we mapped the feline immunodeficiency virus (FIV) ψ to two discontinuous
regions within the 5′ end of the gRNA that assumes a higher order structure harboring several structural motifs. To
better define the region and structural elements important for gRNA packaging, we methodically investigated these FIV ψ
sequences using genetic, biochemical, and structure–function relationship approaches. Our mutational analysis revealed
that the unpaired U85CUG88 stretch within FIV ψ is crucial for gRNA encapsidation into nascent virions. High-throughput
selective 2′ hydroxyl acylation analyzed by primer extension (hSHAPE) performed on wild type (WT) and mutant FIV ψ sequences,
with substitutions in the U85CUG88 stretch, revealed that these mutations had limited structural impact and maintained
nucleotides 80–92 unpaired, as in the WT structure. Since these mutations dramatically affected packaging, our
data suggest that the single-stranded U85CUG88 sequence is important during FIV RNA packaging. Filter-binding assays
performed using purified FIV Pr50Gag on WT and mutant U85CUG88 ψ RNAs led to reduced levels of Pr50Gag binding to
mutant U85CUG88 ψ RNAs, indicating that the U85CUG88 stretch is crucial for ψ RNA–Pr50Gag interactions. Delineating
sequences important for FIV gRNA encapsidation should enhance our understanding of both gRNA packaging and virion
assembly, making them potential targets for novel retroviral therapeutic interventions, as well as the development of
FIV-based vectors for human gene therapy.},
keywords = {bernacchi, MARQUET, Unité ARN, vivet-boudou},
pubstate = {published},
tppubtype = {article}
}
The retroviral Gag precursor plays a central role in the selection and packaging of viral genomic RNA (gRNA) by binding to
virus-specific packaging signal(s) (psi or ψ). Previously, we mapped the feline immunodeficiency virus (FIV) ψ to two discontinuous
regions within the 5′ end of the gRNA that assumes a higher order structure harboring several structural motifs. To
better define the region and structural elements important for gRNA packaging, we methodically investigated these FIV ψ
sequences using genetic, biochemical, and structure–function relationship approaches. Our mutational analysis revealed
that the unpaired U85CUG88 stretch within FIV ψ is crucial for gRNA encapsidation into nascent virions. High-throughput
selective 2′ hydroxyl acylation analyzed by primer extension (hSHAPE) performed on wild type (WT) and mutant FIV ψ sequences,
with substitutions in the U85CUG88 stretch, revealed that these mutations had limited structural impact and maintained
nucleotides 80–92 unpaired, as in the WT structure. Since these mutations dramatically affected packaging, our
data suggest that the single-stranded U85CUG88 sequence is important during FIV RNA packaging. Filter-binding assays
performed using purified FIV Pr50Gag on WT and mutant U85CUG88 ψ RNAs led to reduced levels of Pr50Gag binding to
mutant U85CUG88 ψ RNAs, indicating that the U85CUG88 stretch is crucial for ψ RNA–Pr50Gag interactions. Delineating
sequences important for FIV gRNA encapsidation should enhance our understanding of both gRNA packaging and virion
assembly, making them potential targets for novel retroviral therapeutic interventions, as well as the development of
FIV-based vectors for human gene therapy.
virus-specific packaging signal(s) (psi or ψ). Previously, we mapped the feline immunodeficiency virus (FIV) ψ to two discontinuous
regions within the 5′ end of the gRNA that assumes a higher order structure harboring several structural motifs. To
better define the region and structural elements important for gRNA packaging, we methodically investigated these FIV ψ
sequences using genetic, biochemical, and structure–function relationship approaches. Our mutational analysis revealed
that the unpaired U85CUG88 stretch within FIV ψ is crucial for gRNA encapsidation into nascent virions. High-throughput
selective 2′ hydroxyl acylation analyzed by primer extension (hSHAPE) performed on wild type (WT) and mutant FIV ψ sequences,
with substitutions in the U85CUG88 stretch, revealed that these mutations had limited structural impact and maintained
nucleotides 80–92 unpaired, as in the WT structure. Since these mutations dramatically affected packaging, our
data suggest that the single-stranded U85CUG88 sequence is important during FIV RNA packaging. Filter-binding assays
performed using purified FIV Pr50Gag on WT and mutant U85CUG88 ψ RNAs led to reduced levels of Pr50Gag binding to
mutant U85CUG88 ψ RNAs, indicating that the U85CUG88 stretch is crucial for ψ RNA–Pr50Gag interactions. Delineating
sequences important for FIV gRNA encapsidation should enhance our understanding of both gRNA packaging and virion
assembly, making them potential targets for novel retroviral therapeutic interventions, as well as the development of
FIV-based vectors for human gene therapy.
Marois Eric
Using the CRISPR / Cas9 system for genome editing in Anopheles mosquitoes En ligne
2024, Visité: 08.01.2024.
Résumé | Liens | BibTeX | Étiquettes: Anopheles, CRISPR/Cas9, Genome editing, M3i, marois, mosquitoes
@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}
}
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.
Khodr Radi, Husser Claire, Ryckelynck Michael
Direct fluoride monitoring using a fluorogenic RNA-based biosensor Chapitre d'ouvrage
Dans: Stockbridge, Randy B. (Ed.): vol. 696, p. 85–107, Randy B. Stockbridge, 2024, ISSN: 1557-7988.
Résumé | Liens | BibTeX | Étiquettes: RYCKELYNCK, Unité ARN
@inbook{pmid38658090,
title = {Direct fluoride monitoring using a fluorogenic RNA-based biosensor},
author = {Radi Khodr and Claire Husser and Michael Ryckelynck},
editor = {Randy B. Stockbridge},
doi = {10.1016/bs.mie.2023.12.019},
issn = {1557-7988},
year = {2024},
date = {2024-01-01},
urldate = {2024-01-01},
journal = {Methods Enzymol},
volume = {696},
pages = {85--107},
publisher = {Randy B. Stockbridge},
abstract = {Fluorinated compounds, whether naturally occurring or from anthropogenic origin, have been extensively exploited in the last century. Degradation of these compounds by physical or biochemical processes is expected to result in the release of fluoride. Several fluoride detection mechanisms have been previously described. However, most of these methods are not compatible with high- and ultrahigh-throughput screening technologies, lack the ability to real-time monitor the increase of fluoride concentration in solution, or rely on costly reagents (such as cell-free expression systems). Our group recently developed "FluorMango" as the first completely RNA-based and direct fluoride-specific fluorogenic biosensor. To do so, we merged and engineered the Mango-III light-up RNA aptamer and the fluoride-specific aptamer derived from a riboswitch, crcB. In this chapter, we explain how this RNA-based biosensor can be produced in large scale before providing examples of how it can be used to quantitatively detect (end-point measurement) or monitor in real-time fluoride release in complex biological systems by translating it into measurable fluorescent signal.},
keywords = {RYCKELYNCK, Unité ARN},
pubstate = {published},
tppubtype = {inbook}
}
Fluorinated compounds, whether naturally occurring or from anthropogenic origin, have been extensively exploited in the last century. Degradation of these compounds by physical or biochemical processes is expected to result in the release of fluoride. Several fluoride detection mechanisms have been previously described. However, most of these methods are not compatible with high- and ultrahigh-throughput screening technologies, lack the ability to real-time monitor the increase of fluoride concentration in solution, or rely on costly reagents (such as cell-free expression systems). Our group recently developed "FluorMango" as the first completely RNA-based and direct fluoride-specific fluorogenic biosensor. To do so, we merged and engineered the Mango-III light-up RNA aptamer and the fluoride-specific aptamer derived from a riboswitch, crcB. In this chapter, we explain how this RNA-based biosensor can be produced in large scale before providing examples of how it can be used to quantitatively detect (end-point measurement) or monitor in real-time fluoride release in complex biological systems by translating it into measurable fluorescent signal.
Janvier Aurélie, Hayek Hassan, Alghoul Fatima, Gross Lauriane, Allmang Christine, Martin Franck, Eriani Gilbert
Purification of In Vivo or In Vitro-Assembled RNA-Protein Complexes by RNA Centric Methods Chapitre d'ouvrage
Dans: vol. 3234, p. 17–29, M. Cristina Vega, Francisco J. Fernández, Springer, 2024, ISSN: 0065-2598.
Résumé | Liens | BibTeX | Étiquettes: ERIANI, MARTIN, Unité ARN
@inbook{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},
publisher = {M. Cristina Vega, Francisco J. Fernández},
edition = {Springer},
series = {Advanced Technologies for Protein Complex Production and Characterization},
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 = {inbook}
}
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.
Hayek Hassan, Gross Lauriane, Alghoul Fatima, Martin Franck, Eriani Gilbert, Allmang Christine
Immunoprecipitation Methods to Isolate Messenger Ribonucleoprotein Complexes (mRNP) Chapitre d'ouvrage
Dans: vol. 3234, p. 1–15, M. Cristina Vega, Francisco J. Fernández, Springer, 2024, ISSN: 0065-2598.
Résumé | Liens | BibTeX | Étiquettes: ERIANI, MARTIN, PPSE, Unité ARN
@inbook{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},
publisher = {M. Cristina Vega, Francisco J. Fernández},
edition = {Springer},
series = {Advanced Technologies for Protein Complex Production and Characterization},
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 = {inbook}
}
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.
Quignon E., Ferhadian D., Hache A., Vivet-Boudou V., Isel C., Printz-Schweigert A., Donchet A., Crépin T., Marquet R.
Structural Impact of the Interaction of the Influenza A Virus Nucleoprotein with Genomic RNA Segments Article de journal
Dans: Viruses, vol. 16, no. 3, p. 421, 2024, ISBN: 10.3390/v16030421.
Résumé | Liens | BibTeX | Étiquettes: MARQUET, MARQUET influenza A virus NP nucleoprotein vRNA RNA structure RNA chaperon chemical probing, Unité ARN
@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}
}
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.
Nerantzaki Maria, Husser Claire, Ryckelynck Michael, Lutz Jean-François
Exchanging and Releasing Information in Synthetic Digital Polymers Using a Strand-Displacement Strategy Article de journal
Dans: J Am Chem Soc, 2024, ISSN: 1520-5126.
Résumé | Liens | BibTeX | Étiquettes: RYCKELYNCK, Unité ARN
@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}
}
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.
Baldaccini Morgane, Gaucherand Léa, Chane-Woon-Ming Béatrice, Messmer Mélanie, Gucciardi Floriane, Pfeffer Sébastien
The helicase domain of human Dicer prevents RNAi-independent activation of antiviral and inflammatory pathways Article de journal
Dans: EMBO J, 2024, ISSN: 1460-2075.
Résumé | Liens | BibTeX | Étiquettes: PFEFFER, Unité ARN
@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}
}
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.
Kohl Maximilian P, Chane-Woon-Ming Béatrice, Bahena-Ceron Roberto, Jaramillo-Ponce Jose, Antoine Laura, Herrgott Lucas, Romby Pascale, Marzi Stefano
Ribosome Profiling Methods Adapted to the Study of RNA-Dependent Translation Regulation in Staphylococcus aureus Chapitre d'ouvrage
Dans: Claudio ValverdeVéronique Arluison Véronique Arluison, Claudio Valverde (Ed.): vol. 2741, p. 73–100, Véronique Arluison, Claudio Valverde, Springer, 2024, ISSN: 1940-6029.
Résumé | Liens | BibTeX | Étiquettes: MARZI, ROMBY, Unité ARN
@inbook{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},
editor = {Véronique Arluison, Claudio ValverdeVéronique Arluison, Claudio Valverde},
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},
publisher = {Véronique Arluison, Claudio Valverde},
edition = {Springer},
series = {Bacterial Regulatory RNA, Methods and Protocols},
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 = {inbook}
}
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.
Lechner Antony, Wolff Philippe
In-Gel Cyanoethylation for Pseudouridines Mass Spectrometry Detection of Bacterial Regulatory RNA Chapitre d'ouvrage
Dans: Véronique Arluison, Claudio Valverde (Ed.): vol. 2741, p. 273–287, Véronique Arluison, Claudio Valverde, 2024, ISSN: 1940-6029.
Résumé | Liens | BibTeX | Étiquettes: ARN-MS, Unité ARN
@inbook{pmid38217659,
title = {In-Gel Cyanoethylation for Pseudouridines Mass Spectrometry Detection of Bacterial Regulatory RNA},
author = {Antony Lechner and Philippe Wolff},
editor = {Véronique Arluison, Claudio Valverde},
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},
publisher = {Véronique Arluison, Claudio Valverde},
series = {Bacterial Regulatory RNA, Methods and Protocols},
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 = {inbook}
}
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.
2023
Ponce José R Jaramillo, Frugier Magali
Plasmodium, the Apicomplexa Outlier When It Comes to Protein Synthesis Article de journal
Dans: Biomolecules, vol. 14, no. 1, p. 46, 2023, ISSN: 2218-273X.
Résumé | Liens | BibTeX | Étiquettes: FRUGIER, Unité ARN
@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}
}
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.
Bahena-Ceron Roberto, Teixeira Chloe, Ponce Jose R Jaramillo, Wolff Philippe, Couzon Florence, François Pauline, Klaholz Bruno, Vandenesch François, Romby Pascale, Moreau Karen, Marzi Stefano
RlmQ: A Newly Discovered rRNA Modification Enzyme Bridging RNA Modification and Virulence Traits in Article de journal
Dans: RNA, vol. 30, no. 3, p. 200-212, 2023, ISSN: 1469-9001.
Résumé | Liens | BibTeX | Étiquettes: ARN-MS, ROMBY, Unité ARN
@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}
}
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.
Kompatscher Maria, Bartosik Karolina, Erharter Kevin, Plangger Raphael, Juen Fabian Sebastian, Kreutz Christoph, Micura Ronald, Westhof Eric, Erlacher Matthias D
Contribution of tRNA sequence and modifications to the decoding preferences of E. coli and M. mycoides tRNAGlyUCC for synonymous glycine codons Article de journal
Dans: Nucleic Acids Res, 2023, ISSN: 1362-4962.
Résumé | Liens | BibTeX | Étiquettes: Unité ARN, WESTHOF
@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}
}
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.
Skerniskyte Jurate, Mulet Céline, André Antonin C, Anderson Mark C, Injarabian Louise, Buck Achim, Prade Verena M, Sansonetti Philippe J, Reibel-Foisset Sophie, Walch Axel K, Lebel Michel, Lykkesfeldt Jens, Marteyn Benoit S
Ascorbate deficiency increases progression of shigellosis in guinea pigs and mice infection models Article de journal
Dans: Gut Microbes, vol. 15, no. 2, p. 2271597, 2023, ISSN: 1949-0984.
Résumé | Liens | BibTeX | Étiquettes: MARTEYN, Unité ARN
@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}
}
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.
A Hinze, J Pelletier, M Ghaninia, E Marois, Hill, R S Ignell
Knockout of OR39 reveals redundancy in the olfactory pathway regulating the acquisition of host seeking in Anopheles coluzzii Article de journal
Dans: Proc Biol Sci 290(2011):20232092, vol. 290, no. 2011, 2023.
Résumé | Liens | BibTeX | Étiquettes: CRISPR–Cas9, human odour, M3i, marois, mosquito flight, odour valence, SSR
@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}
}
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.
SC Weng, I Antoshechkin, E Marois, O Akbari
Efficient Sex Separation by Exploiting Differential Alternative Splicing of a Dominant Marker in Aedes aegypti Article de journal
Dans: PLoS Genetics, 2023.
Résumé | Liens | BibTeX | Étiquettes: Aedes aegypti, Introns, larvae, M3i, marois, mosquitoes, pupae, rna sequencing, transcriptome analyses
@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}
}
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.
Arrivé Mathilde, Bruggeman Mathieu, Skaltsogiannis Vasileios, Coudray Léna, Quan Yi-Fat, Schelcher Cédric, Cognat Valérie, Hammann Philippe, Chicher Johana, Wolff Philippe, Gobert Anthony, Giegé Philippe
A tRNA-modifying enzyme facilitates RNase P activity in Arabidopsis nuclei Article de journal
Dans: Nat Plants, vol. 9, iss. 12, p. 2031-2041, 2023, ISSN: 2055-0278.
Résumé | Liens | BibTeX | Étiquettes: ARN-MS, PPSE, Unité ARN
@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}
}
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.
Lu Kunpeng, Pan Yifei, Shen Jianghong, Yang Lin, Zhan Chengyu, Liang Shubo, Tai Shuaishuai, Wan Linrong, Li Tian, Cheng Tingcai, Ma Bi, Pan Guoqing, He Ningjia, Lu Cheng, Westhof Eric, Xiang Zhonghuai, Han Min-Jin, Tong Xiaoling, Dai Fangyin
SilkMeta: a comprehensive platform for sharing and exploiting pan-genomic and multi-omic silkworm data Article de journal
Dans: Nucleic Acids Res, vol. 52, iss. D1, p. D1024-D1032, 2023, ISSN: 1362-4962.
Résumé | Liens | BibTeX | Étiquettes: Unité ARN, WESTHOF
@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}
}
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.
Das Rhiju, Kretsch Rachael C, Simpkin Adam J, Mulvaney Thomas, Pham Phillip, Rangan Ramya, Bu Fan, Keegan Ronan M, Topf Maya, Rigden Daniel J, Miao Zhichao, Westhof Eric
Assessment of three-dimensional RNA structure prediction in CASP15 Article de journal
Dans: Proteins, vol. 91, iss. 12, p. 1747-1770, 2023, ISSN: 1097-0134.
Résumé | Liens | BibTeX | Étiquettes: Unité ARN, WESTHOF
@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}
}
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.
Silva P Malaka De, Bennett Rebecca J, Kuhn Lauriane, Ngondo Patryk, Debande Lorine, Njamkepo Elisabeth, Ho Brian, Weill François-Xavier, Marteyn Benoît S, Jenkins Claire, Baker Kate S
Escherichia coli killing by epidemiologically successful sublineages of Shigella sonnei is mediated by colicins Article de journal
Dans: EBioMedicine, vol. 97, p. 104822, 2023, ISSN: 2352-3964.
Résumé | Liens | BibTeX | Étiquettes: MARTEYN, PPSE, Unité ARN
@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}
}
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.
AI Kalita, E Marois, M Kozielska, FJ Weissing, E Jaouen, Möckel MM and Rühle F, F Butter, MF Basilicata, CI Keller Valsecchi
The sex-specific factor SOA controls dosage compensation in Anopheles mosquitoes Article de journal
Dans: Nature, vol. 623, p. 175–182, 2023.
Résumé | Liens | BibTeX | Étiquettes: Anopheles, M3i, marois, SOA
@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}
}
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.
