A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination

ABSTRACT Clostridium difficile is a Gram-positive spore-forming obligate anaerobe that is a leading cause of antibiotic-associated diarrhea worldwide. In order for C. difficile to initiate infection, its aerotolerant spore form must germinate in the gut of mammalian hosts. While almost all spore-for...

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Autores principales:	M. Lauren Donnelly, William Li, Yong-qing Li, Lauren Hinkel, Peter Setlow, Aimee Shen
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2017
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/669be8ded87a4df08a25b07440ce0d91
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spelling	oai:doaj.org-article:669be8ded87a4df08a25b07440ce0d912021-11-15T15:51:07ZA <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination10.1128/mBio.02085-162150-7511https://doaj.org/article/669be8ded87a4df08a25b07440ce0d912017-03-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02085-16https://doaj.org/toc/2150-7511ABSTRACT Clostridium difficile is a Gram-positive spore-forming obligate anaerobe that is a leading cause of antibiotic-associated diarrhea worldwide. In order for C. difficile to initiate infection, its aerotolerant spore form must germinate in the gut of mammalian hosts. While almost all spore-forming organisms use transmembrane germinant receptors to trigger germination, C. difficile uses the pseudoprotease CspC to sense bile salt germinants. CspC activates the related subtilisin-like protease CspB, which then proteolytically activates the cortex hydrolase SleC. Activated SleC degrades the protective spore cortex layer, a step that is essential for germination to proceed. Since CspC incorporation into spores also depends on CspA, a related pseudoprotease domain, Csp family proteins play a critical role in germination. However, how Csps are incorporated into spores remains unknown. In this study, we demonstrate that incorporation of the CspC, CspB, and CspA germination regulators into spores depends on CD0311 (renamed GerG), a previously uncharacterized hypothetical protein. The reduced levels of Csps in gerG spores correlate with reduced responsiveness to bile salt germinants and increased germination heterogeneity in single-spore germination assays. Interestingly, asparagine-rich repeat sequences in GerG’s central region facilitate spontaneous gel formation in vitro even though they are dispensable for GerG-mediated control of germination. Since GerG is found exclusively in C. difficile, our results suggest that exploiting GerG function could represent a promising avenue for developing C. difficile-specific anti-infective therapies. IMPORTANCE The spore-forming bacterium Clostridium difficile is a leading cause of health care-associated infections. While a subset of antibiotics can treat C. difficile infections (CDIs), the primary determinant of CDI disease susceptibility is prior antibiotic exposure, since it reduces the colonization resistance conferred by a diverse microflora. Thus, therapies that minimize perturbations to the gut microbiome should be more effective at reducing CDIs and their recurrence, the main source of disease complications. Given that spore germination is essential for C. difficile to initiate infection and that C. difficile uses a unique pathway to initiate germination, methods that inhibit distinct elements of germination could selectively prevent C. difficile disease recurrence. Here, we identify GerG as a C. difficile-specific protein that controls the incorporation of germinant signaling proteins into spores. Since gerG mutant spores exhibit germination defects and are less responsive to germinant, GerG may represent a promising target for developing therapeutics against CDI.M. Lauren DonnellyWilliam LiYong-qing LiLauren HinkelPeter SetlowAimee ShenAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 8, Iss 1 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 M. Lauren Donnelly William Li Yong-qing Li Lauren Hinkel Peter Setlow Aimee Shen A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination
description	ABSTRACT Clostridium difficile is a Gram-positive spore-forming obligate anaerobe that is a leading cause of antibiotic-associated diarrhea worldwide. In order for C. difficile to initiate infection, its aerotolerant spore form must germinate in the gut of mammalian hosts. While almost all spore-forming organisms use transmembrane germinant receptors to trigger germination, C. difficile uses the pseudoprotease CspC to sense bile salt germinants. CspC activates the related subtilisin-like protease CspB, which then proteolytically activates the cortex hydrolase SleC. Activated SleC degrades the protective spore cortex layer, a step that is essential for germination to proceed. Since CspC incorporation into spores also depends on CspA, a related pseudoprotease domain, Csp family proteins play a critical role in germination. However, how Csps are incorporated into spores remains unknown. In this study, we demonstrate that incorporation of the CspC, CspB, and CspA germination regulators into spores depends on CD0311 (renamed GerG), a previously uncharacterized hypothetical protein. The reduced levels of Csps in gerG spores correlate with reduced responsiveness to bile salt germinants and increased germination heterogeneity in single-spore germination assays. Interestingly, asparagine-rich repeat sequences in GerG’s central region facilitate spontaneous gel formation in vitro even though they are dispensable for GerG-mediated control of germination. Since GerG is found exclusively in C. difficile, our results suggest that exploiting GerG function could represent a promising avenue for developing C. difficile-specific anti-infective therapies. IMPORTANCE The spore-forming bacterium Clostridium difficile is a leading cause of health care-associated infections. While a subset of antibiotics can treat C. difficile infections (CDIs), the primary determinant of CDI disease susceptibility is prior antibiotic exposure, since it reduces the colonization resistance conferred by a diverse microflora. Thus, therapies that minimize perturbations to the gut microbiome should be more effective at reducing CDIs and their recurrence, the main source of disease complications. Given that spore germination is essential for C. difficile to initiate infection and that C. difficile uses a unique pathway to initiate germination, methods that inhibit distinct elements of germination could selectively prevent C. difficile disease recurrence. Here, we identify GerG as a C. difficile-specific protein that controls the incorporation of germinant signaling proteins into spores. Since gerG mutant spores exhibit germination defects and are less responsive to germinant, GerG may represent a promising target for developing therapeutics against CDI.
format	article
author	M. Lauren Donnelly William Li Yong-qing Li Lauren Hinkel Peter Setlow Aimee Shen
author_facet	M. Lauren Donnelly William Li Yong-qing Li Lauren Hinkel Peter Setlow Aimee Shen
author_sort	M. Lauren Donnelly
title	A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination
title_short	A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination
title_full	A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination
title_fullStr	A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination
title_full_unstemmed	A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination
title_sort	<italic toggle="yes">clostridium difficile</italic>-specific, gel-forming protein required for optimal spore germination
publisher	American Society for Microbiology
publishDate	2017
url	https://doaj.org/article/669be8ded87a4df08a25b07440ce0d91
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A <italic toggle="yes">Clostridium difficile</italic>-Specific, Gel-Forming Protein Required for Optimal Spore Germination

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