Dept. Regulation in infection (REGI)Leiter: Frau Prof. Dr. Emmanuelle Charpentierhttp://hdl.handle.net/10033/2646942024-03-29T10:11:31Z2024-03-29T10:11:31ZA Two-Component Regulatory System Impacts Extracellular Membrane-Derived Vesicle Production in Group A Streptococcus.Resch, UlrikeTsatsaronis, James AnthonyLe Rhun, AnaïsStübiger, GeraldRohde, MKasvandik, SergoHolzmeister, SusanneTinnefeld, PhilipWai, Sun NyuntCharpentier, Emmanuellehttp://hdl.handle.net/10033/6213172019-08-30T11:28:51Z2016-01-01T00:00:00ZA Two-Component Regulatory System Impacts Extracellular Membrane-Derived Vesicle Production in Group A Streptococcus.
Resch, Ulrike; Tsatsaronis, James Anthony; Le Rhun, Anaïs; Stübiger, Gerald; Rohde, M; Kasvandik, Sergo; Holzmeister, Susanne; Tinnefeld, Philip; Wai, Sun Nyunt; Charpentier, Emmanuelle
Export of macromolecules via extracellular membrane-derived vesicles (MVs) plays an important role in the biology of Gram-negative bacteria. Gram-positive bacteria have also recently been reported to produce MVs; however, the composition and mechanisms governing vesiculogenesis in Gram-positive bacteria remain undefined. Here, we describe MV production in the Gram-positive human pathogen group A streptococcus (GAS), the etiological agent of necrotizing fasciitis and streptococcal toxic shock syndrome. M1 serotype GAS isolates in culture exhibit MV structures both on the cell wall surface and in the near vicinity of bacterial cells. A comprehensive analysis of MV proteins identified both virulence-associated protein substrates of the general secretory pathway in addition to "anchorless surface proteins." Characteristic differences in the contents, distributions, and fatty acid compositions of specific lipids between MVs and GAS cell membrane were also observed. Furthermore, deep RNA sequencing of vesicular RNAs revealed that GAS MVs contained differentially abundant RNA species relative to bacterial cellular RNA. MV production by GAS strains varied in a manner dependent on an intact two-component system, CovRS, with MV production negatively regulated by the system. Modulation of MV production through CovRS was found to be independent of both GAS cysteine protease SpeB and capsule biosynthesis. Our data provide an explanation for GAS secretion of macromolecules, including RNAs, lipids, and proteins, and illustrate a regulatory mechanism coordinating this secretory response.
2016-01-01T00:00:00ZIdentification of endoribonuclease specific cleavage positions reveals novel targets of RNase III in Streptococcus pyogenes.Le Rhun, AnaïsLécrivain, Anne-LaureReimegård, JohanProux-Wéra, EstelleBroglia, LauraDella Beffa, CristinaCharpentier, Emmanuellehttp://hdl.handle.net/10033/6210912019-08-30T11:32:41Z2017-03-17T00:00:00ZIdentification of endoribonuclease specific cleavage positions reveals novel targets of RNase III in Streptococcus pyogenes.
Le Rhun, Anaïs; Lécrivain, Anne-Laure; Reimegård, Johan; Proux-Wéra, Estelle; Broglia, Laura; Della Beffa, Cristina; Charpentier, Emmanuelle
A better understanding of transcriptional and post-transcriptional regulation of gene expression in bacteria relies on studying their transcriptome. RNA sequencing methods are used not only to assess RNA abundance but also the exact boundaries of primary and processed transcripts. Here, we developed a method, called identification of specific cleavage position (ISCP), which enables the identification of direct endoribonuclease targets in vivo by comparing the 5΄ and 3΄ ends of processed transcripts between wild type and RNase deficient strains. To demonstrate the ISCP method, we used as a model the double-stranded specific RNase III in the human pathogen Streptococcus pyogenes. We mapped 92 specific cleavage positions (SCPs) among which, 48 were previously described and 44 are new, with the characteristic 2 nucleotides 3΄ overhang of RNase III. Most SCPs were located in untranslated regions of RNAs. We screened for RNase III targets using transcriptomic differential expression analysis (DEA) and compared those with the RNase III targets identified using the ISCP method. Our study shows that in S. pyogenes, under standard growth conditions, RNase III has a limited impact both on antisense transcripts and on global gene expression with the expression of most of the affected genes being downregulated in an RNase III deletion mutant.
2017-03-17T00:00:00ZA flagellum-specific chaperone facilitates assembly of the core type III export apparatus of the bacterial flagellum.Fabiani, Florian DRenault, Thibaud TPeters, BrittaDietsche, TobiasGálvez, Eric J CGuse, AlinaFreier, KarenCharpentier, EmmanuelleStrowig, TillFranz-Wachtel, MiritaMacek, BorisWagner, SamuelHensel, MichaelErhardt, Marchttp://hdl.handle.net/10033/6210602019-08-30T11:36:05Z2017-08-01T00:00:00ZA flagellum-specific chaperone facilitates assembly of the core type III export apparatus of the bacterial flagellum.
Fabiani, Florian D; Renault, Thibaud T; Peters, Britta; Dietsche, Tobias; Gálvez, Eric J C; Guse, Alina; Freier, Karen; Charpentier, Emmanuelle; Strowig, Till; Franz-Wachtel, Mirita; Macek, Boris; Wagner, Samuel; Hensel, Michael; Erhardt, Marc
Many bacteria move using a complex, self-assembling nanomachine, the bacterial flagellum. Biosynthesis of the flagellum depends on a flagellar-specific type III secretion system (T3SS), a protein export machine homologous to the export machinery of the virulence-associated injectisome. Six cytoplasmic (FliH/I/J/G/M/N) and seven integral-membrane proteins (FlhA/B FliF/O/P/Q/R) form the flagellar basal body and are involved in the transport of flagellar building blocks across the inner membrane in a proton motive force-dependent manner. However, how the large, multi-component transmembrane export gate complex assembles in a coordinated manner remains enigmatic. Specific for most flagellar T3SSs is the presence of FliO, a small bitopic membrane protein with a large cytoplasmic domain. The function of FliO is unknown, but homologs of FliO are found in >80% of all flagellated bacteria. Here, we demonstrate that FliO protects FliP from proteolytic degradation and promotes the formation of a stable FliP-FliR complex required for the assembly of a functional core export apparatus. We further reveal the subcellular localization of FliO by super-resolution microscopy and show that FliO is not part of the assembled flagellar basal body. In summary, our results suggest that FliO functions as a novel, flagellar T3SS-specific chaperone, which facilitates quality control and productive assembly of the core T3SS export machinery.
2017-08-01T00:00:00ZToward Whole-Transcriptome Editing with CRISPR-Cas9.Heckl, DirkCharpentier, Emmanuellehttp://hdl.handle.net/10033/6205572019-08-30T11:25:43Z2015-05-21T00:00:00ZToward Whole-Transcriptome Editing with CRISPR-Cas9.
Heckl, Dirk; Charpentier, Emmanuelle
Targeted regulation of gene expression holds huge promise for biomedical research. In a series of recent publications (Gilbert et al., 2014; Konermann et al., 2015; Zalatan et al., 2015), sophisticated, multiplex-compatible transcriptional activator systems based on the CRISPR-Cas9 technology and genome-scale libraries advance the field toward whole-transcriptome control.
2015-05-21T00:00:00Z