Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>

ABSTRACT Clostridium difficile is the major etiologic agent of antibiotic-associated intestinal disease. Pathogenesis of C. difficile is mainly attributed to the production and secretion of toxins A and B. Unlike most clostridial toxins, toxins A and B have no signal peptide, and they are therefore...

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Autores principales:	Sandra Wydau-Dematteis, Imane El Meouche, Pascal Courtin, Audrey Hamiot, René Lai-Kuen, Bruno Saubaméa, François Fenaille, Marie-José Butel, Jean-Louis Pons, Bruno Dupuy, Marie-Pierre Chapot-Chartier, Johann Peltier
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2018
Materias:	Clostridium difficile autolysins lytic transglycosylase peptidoglycan surface proteins toxins Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/981e2cb1b5b04f1781907b9a09bd4184
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spelling	oai:doaj.org-article:981e2cb1b5b04f1781907b9a09bd41842021-11-15T16:00:25ZCwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>10.1128/mBio.00648-182150-7511https://doaj.org/article/981e2cb1b5b04f1781907b9a09bd41842018-07-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00648-18https://doaj.org/toc/2150-7511ABSTRACT Clostridium difficile is the major etiologic agent of antibiotic-associated intestinal disease. Pathogenesis of C. difficile is mainly attributed to the production and secretion of toxins A and B. Unlike most clostridial toxins, toxins A and B have no signal peptide, and they are therefore secreted by unusual mechanisms involving the holin-like TcdE protein and/or autolysis. In this study, we characterized the cell surface protein Cwp19, a newly identified peptidoglycan-degrading enzyme containing a novel catalytic domain. We purified a recombinant His6-tagged Cwp19 protein and showed that it has lytic transglycosylase activity. Moreover, we observed that Cwp19 is involved in cell autolysis and that a C. difficile cwp19 mutant exhibited delayed autolysis in stationary phase compared to the wild type when bacteria were grown in brain heart infusion (BHI) medium. Wild-type cell autolysis is correlated to strong alterations of cell wall thickness and integrity and to release of cytoplasmic material. Furthermore, we demonstrated that toxins were released into the extracellular medium as a result of Cwp19-induced autolysis when cells were grown in BHI medium. In contrast, Cwp19 did not induce autolysis or toxin release when cells were grown in tryptone-yeast extract (TY) medium. These data provide evidence for the first time that TcdE and bacteriolysis are coexisting mechanisms for toxin release, with their relative contributions in vitro depending on growth conditions. Thus, Cwp19 is an important surface protein involved in autolysis of vegetative cells of C. difficile that mediates the release of the toxins from the cell cytosol in response to specific environment conditions. IMPORTANCE Clostridium difficile-associated disease is mainly known as a health care-associated infection. It represents the most problematic hospital-acquired infection in North America and Europe and exerts significant economic pressure on health care systems. Virulent strains of C. difficile generally produce two toxins that have been identified as the major virulence factors. The mechanism for release of these toxins from bacterial cells is not yet fully understood but is thought to be partly mediated by bacteriolysis. Here we identify a novel peptidoglycan hydrolase in C. difficile, Cwp19, exhibiting lytic transglycosylase activity. We show that Cwp19 contributes to C. difficile cell autolysis in the stationary phase and, consequently, to toxin release, most probably as a response to environmental conditions such as nutritional signals. These data highlight that Cwp19 constitutes a promising target for the development of new preventive and curative strategies.Sandra Wydau-DematteisImane El MeouchePascal CourtinAudrey HamiotRené Lai-KuenBruno SaubaméaFrançois FenailleMarie-José ButelJean-Louis PonsBruno DupuyMarie-Pierre Chapot-ChartierJohann PeltierAmerican Society for MicrobiologyarticleClostridium difficileautolysinslytic transglycosylasepeptidoglycansurface proteinstoxinsMicrobiologyQR1-502ENmBio, Vol 9, Iss 3 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Clostridium difficile autolysins lytic transglycosylase peptidoglycan surface proteins toxins Microbiology QR1-502
spellingShingle	Clostridium difficile autolysins lytic transglycosylase peptidoglycan surface proteins toxins Microbiology QR1-502 Sandra Wydau-Dematteis Imane El Meouche Pascal Courtin Audrey Hamiot René Lai-Kuen Bruno Saubaméa François Fenaille Marie-José Butel Jean-Louis Pons Bruno Dupuy Marie-Pierre Chapot-Chartier Johann Peltier Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>
description	ABSTRACT Clostridium difficile is the major etiologic agent of antibiotic-associated intestinal disease. Pathogenesis of C. difficile is mainly attributed to the production and secretion of toxins A and B. Unlike most clostridial toxins, toxins A and B have no signal peptide, and they are therefore secreted by unusual mechanisms involving the holin-like TcdE protein and/or autolysis. In this study, we characterized the cell surface protein Cwp19, a newly identified peptidoglycan-degrading enzyme containing a novel catalytic domain. We purified a recombinant His6-tagged Cwp19 protein and showed that it has lytic transglycosylase activity. Moreover, we observed that Cwp19 is involved in cell autolysis and that a C. difficile cwp19 mutant exhibited delayed autolysis in stationary phase compared to the wild type when bacteria were grown in brain heart infusion (BHI) medium. Wild-type cell autolysis is correlated to strong alterations of cell wall thickness and integrity and to release of cytoplasmic material. Furthermore, we demonstrated that toxins were released into the extracellular medium as a result of Cwp19-induced autolysis when cells were grown in BHI medium. In contrast, Cwp19 did not induce autolysis or toxin release when cells were grown in tryptone-yeast extract (TY) medium. These data provide evidence for the first time that TcdE and bacteriolysis are coexisting mechanisms for toxin release, with their relative contributions in vitro depending on growth conditions. Thus, Cwp19 is an important surface protein involved in autolysis of vegetative cells of C. difficile that mediates the release of the toxins from the cell cytosol in response to specific environment conditions. IMPORTANCE Clostridium difficile-associated disease is mainly known as a health care-associated infection. It represents the most problematic hospital-acquired infection in North America and Europe and exerts significant economic pressure on health care systems. Virulent strains of C. difficile generally produce two toxins that have been identified as the major virulence factors. The mechanism for release of these toxins from bacterial cells is not yet fully understood but is thought to be partly mediated by bacteriolysis. Here we identify a novel peptidoglycan hydrolase in C. difficile, Cwp19, exhibiting lytic transglycosylase activity. We show that Cwp19 contributes to C. difficile cell autolysis in the stationary phase and, consequently, to toxin release, most probably as a response to environmental conditions such as nutritional signals. These data highlight that Cwp19 constitutes a promising target for the development of new preventive and curative strategies.
format	article
author	Sandra Wydau-Dematteis Imane El Meouche Pascal Courtin Audrey Hamiot René Lai-Kuen Bruno Saubaméa François Fenaille Marie-José Butel Jean-Louis Pons Bruno Dupuy Marie-Pierre Chapot-Chartier Johann Peltier
author_facet	Sandra Wydau-Dematteis Imane El Meouche Pascal Courtin Audrey Hamiot René Lai-Kuen Bruno Saubaméa François Fenaille Marie-José Butel Jean-Louis Pons Bruno Dupuy Marie-Pierre Chapot-Chartier Johann Peltier
author_sort	Sandra Wydau-Dematteis
title	Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>
title_short	Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>
title_full	Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>
title_fullStr	Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>
title_full_unstemmed	Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>
title_sort	cwp19 is a novel lytic transglycosylase involved in stationary-phase autolysis resulting in toxin release in <italic toggle="yes">clostridium difficile</italic>
publisher	American Society for Microbiology
publishDate	2018
url	https://doaj.org/article/981e2cb1b5b04f1781907b9a09bd4184
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Cwp19 Is a Novel Lytic Transglycosylase Involved in Stationary-Phase Autolysis Resulting in Toxin Release in <italic toggle="yes">Clostridium difficile</italic>

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