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RNA helicases in virulence expression in Staphylococcus aureus

English title RNA helicases in virulence expression in Staphylococcus aureus
Applicant Linder Patrick
Number 149228
Funding scheme Project funding (Div. I-III)
Research institution Dépt Microbiologie et Médecine Moléculaire Faculté de Médecine Université de Genève
Institution of higher education University of Geneva - GE
Main discipline Medical Microbiology
Start/End 01.10.2013 - 30.09.2016
Approved amount 625'906.00
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All Disciplines (2)

Discipline
Medical Microbiology
Molecular Biology

Keywords (4)

Staphylococcus aureus; gene expression; RNA degradation; RNA helicase

Lay Summary (French)

Lead
Le Staphylocoque doré est un pathogène opportuniste, présent principalement dans les narines d’environ un tiers de la population. En cas de déficience immunitaire ou de présence de brèches dans la peau, ce pathogène peut faire des infections graves. Récemment des jeunes personnes en bonne santé ont aussi développé des infections graves à ce pathogène. Ce changement de la bactérie peu offensive à un pathogène très virulent, est accompagné d’un changement d’expression de gènes de virulence.
Lay summary

Contenu et objectif du travail de recherche

Nous nous intéressons depuis de nombreuses années aux hélicases à ARN qui déroulent des duplex d’ARN. Chez le Staphylocoque doré 2 protéines de cette famille ont été trouvée. Une est nécessaire pour la dégradation de certains ARN. Pour l’autre, si elle est absente ou n’est pas exprimée, les cellules possèdent un métabolisme réduit et forment après un choc acide des petites colonies typiques des bactéries causant des infections persistantes. Dans les deux cas, l’absence de l’hélicase influence seulement une partie des ARN. Il est donc nécessaire de définir pour les deux hélicases les partenaires protéiques et les cibles ARN, afin de mieux comprendre leur régulation et leur spécificité. Nous utilisons une combinaison de méthodes classiques, comme la génétique bactérienne et la biochimie, et des méthodes plus récentes comme la spectrométrie de masse pour identifier les protéines partenaires et le séquençage à haut débit pour identifier les substrats ARN.

 

Contexte scientifique et médical du projet

Cette recherche a pour but d’élucider le rôle des hélicases à ARN dans l’expression des facteurs de virulence chez le Staphylocoque doré. A l’heure où  les antibiotiques commencent à faire défaut à cause des souches multi-résistantes, la compréhension de l’expression de facteurs de virulence est essentielle. On peut s’imaginer combattre la virulence ou persistance d’une bactérie plutôt que d’utiliser des traitements de longue durée aux antibiotiques.

Direct link to Lay Summary Last update: 29.09.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
How does sub-cellular localization affect the fate of bacterial mRNA?
Redder Peter (2016), How does sub-cellular localization affect the fate of bacterial mRNA?, in Current Genetics, 62(4), 687-690.
RNA helicases in bacteria
Khemici Vanessa, Linder Patrick (2016), RNA helicases in bacteria, in Current Opinion in Microbiology, 30, 58-66.
TSS-EMOTE, a refined protocol for a more complete and less biased global mapping of transcription start sites in bacterial pathogens.
Prados J Linder P Redder P. (2016), TSS-EMOTE, a refined protocol for a more complete and less biased global mapping of transcription start sites in bacterial pathogens., in BMC Genomics., 17(1), 849.
Bacterial versatility requires DEAD-box RNA helicases
Redder P., Hausmann S., Khemici V., Yasrebi H., Linder P. (2015), Bacterial versatility requires DEAD-box RNA helicases, in FEMS Microbiology Reviews, 39(3), 392-412.
Decay-Initiating Endoribonucleolytic Cleavage by RNase Y Is Kept under Tight Control via Sequence Preference and Sub-cellular Localisation
Khemici Vanessa, Prados Julien, Linder Patrick, Redder Peter (2015), Decay-Initiating Endoribonucleolytic Cleavage by RNase Y Is Kept under Tight Control via Sequence Preference and Sub-cellular Localisation, in PLOS Genetics, 11(10), e1005577-e1005577.
Happy birthday: 25 years of DEAD-box protein
Linder Patrick, Fuller-Pace Frances (2015), Happy birthday: 25 years of DEAD-box protein, Springer New York, New York, NY.
The C-terminal region of the RNA helicase CshA is required for the interaction with the degradosome and turnover of bulk RNA in the opportunistic pathogen Staphylococcus aureus
Giraud Caroline, Hausmann Stéphane, Lemeille Sylvain, Prados Julien, Redder Peter, Linder Patrick (2015), The C-terminal region of the RNA helicase CshA is required for the interaction with the degradosome and turnover of bulk RNA in the opportunistic pathogen Staphylococcus aureus, in RNA Biology, 12(6), 658-674.
Using EMOTE to map the exact 5'-ends of processed RNA on a transcriptome-wide scale
Redder Peter (2015), Using EMOTE to map the exact 5'-ends of processed RNA on a transcriptome-wide scale, Springer New York, New York, NY.
Transcriptome-Wide Analyses of 5′-Ends in RNase J Mutants of a Gram-Positive Pathogen Reveal a Role in RNA Maturation, Regulation and Degradation
Linder Patrick, Lemeille Sylvain, Redder Peter (2014), Transcriptome-Wide Analyses of 5′-Ends in RNase J Mutants of a Gram-Positive Pathogen Reveal a Role in RNA Maturation, Regulation and Degradation, in PLoS Genetics, 10(2), e1004207-e1004207.

Collaboration

Group / person Country
Types of collaboration
Prof Jérôme Pugin, CMU & HUG Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results

Associated projects

Number Title Start Funding scheme
170207 RNA metabolism in virulence expression in Staphylococcus aureus 01.10.2016 Project funding (Div. I-III)
157760 Qualitative-Quantitative Omics’ Mass Spectrometry for Biomarker Verification 01.12.2014 R'EQUIP
132638 Acquisition and regulation of virulence factors in Staphylococcus aureus 01.10.2010 Project funding (Div. I-III)
170207 RNA metabolism in virulence expression in Staphylococcus aureus 01.10.2016 Project funding (Div. I-III)

Abstract

Staphylococcus aureus is an opportunistic pathogen that can cause a variety of life threatening diseases. Many S. aureus strains harbor antibiotic resistance genes, but can also develop persistent infections that are phenotypically antibiotic resistant and very difficult to eradicate. The formation of biofilms and the phenotypic switching to small colony variants are two important strategies to cause persistent infections. Both, biofilm formation and intracellular survival of small colony variants, depend on the expression control of genes involved in these processes. The regulation of gene expression in bacteria can occur at several levels. In recent years small regulatory RNAs and the role of secondary structures have been recognized as important players in controlling gene expression. It is therefore likely that RNA helicases of the DEAD-box protein family function in local modulation of secondary structures or RNA-RNA unwinding to allow regulation of gene expression. Our published and preliminary data show that CshA, one of the two DEAD-box protein from S. aureus, participates in the turnover of RNA and thereby influences biofilm formation and hemolysis. Although this suggests that CshA participates together with RNases in the degradosome, the role of the helicase in this complex is unknown. Moreover, it is possible that CshA, like other bacterial DEAD-box proteins, performs additional functions in ribosome biogenesis or translation initiation, explaining some of the observed phenotypes in a cshA mutant. Preliminary results in the laboratory have shown that inactivation of the second RNA helicase, CshB, results in increased survival after an acid treatment and in a neutrophil killing assay. In addition, cshB mutant cells show a small colony variant phenotype after acid shock treatment. Finally, cshB mutants, but not wild type or ?cshA cells, are deficient for growth on a defined medium derived from a cell-culture medium. These different characteristics indicate an important role of CshB in gene expression. To learn more about the function of these two RNA helicases, we will search for their target molecules by cross-linking and immunoprecipitation of RNAs and polysome analysis, followed by RNA sequence analysis. To elucidate further the context in which these RNA helicases function and eventually how partner proteins influence their activity, we will also define interacting proteins. Finally, we will use genetic approaches to better define the processes in which these RNA helicase function. We will use suppressor analyses to identify genes that allow growth under non-permissive conditions and we will use saturated transposon mutagenesis to identify genes that - when mutated - enhance the helicase mutant phenotypes.
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