A Nutrient-Regulated Cyclic Diguanylate Phosphodiesterase Controls Clostridium difficile Biofilm and Toxin Production during Stationary Phase

• Type of Material: Article
• Authors: Erin B. Purcell, Old Dominion University; Robert W. McKee, University of North Carolina Chapel Hill; David S. Courson, Old Dominion University; Elizabeth M. Garrett, University of North Carolina Chapel Hill; Shonna McBride, Emory University; Richard E. Cheney, University of North Carolina Chapel Hill; Rita Tamayo, University of North Carolina Chapel Hill
• Language: English
• Date: 2017-09-01
• Publisher: American Society for Microbiology
• Publication Version: Final Published Version
• Copyright Statement: © 2017 American Society for Microbiology.
• Final Published Version (URL): http://dx.doi.org/10.1128/IAI.00347-17
• Title of Journal or Parent Work: Infection and Immunity
• ISSN: 0019-9567
• Volume: 85
• Issue: 9
• Start Page: e00347-17
• End Page: e00347-17
• Grant/Funding Information: E.B.P. was supported by T32-DK007737 to the Center for Gastrointestinal Biology and Disease, and D.S.C. was supported by T32-CA009156 to the Lineberger Comprehensive Cancer Center.; This research was supported by NIH grants U54-AI057157 and R01-AI107029 to R.T., R01-DC003299 to R.E.C., and K01-DK087763 to S.M.M.; The contents of this report are solely the responsibility of the authors and do not necessarily represent the official views of the funding bodies.; UPLC-MS was performed at the University of North Carolina Environmental Sciences and Engineering Biomarker Mass Spectrometry Core Facility, which is supported in part by a grant from the National Institute of Environmental Health Sciences (P30ES010126).
• Keywords: nutrient; Immunology; toxin; ALLOSTERIC CONTROL; GRAM-POSITIVE BACTERIA; biofilm; flagellar motility; VIBRIO-CHOLERAE; flagella; EAL DOMAIN PROTEIN; cyclic diguanylate; PSEUDOMONAS-AERUGINOSA; CodY; BACILLUS-SUBTILIS STRAINS; DI-GMP PHOSPHODIESTERASE; INTESTINAL MICROBIOTA; Life Sciences & Biomedicine; BINDING PROTEIN-RHO; VIRULENCE GENE-EXPRESSION; Infectious Diseases; Science & Technology; c-di-GMP
• Research Categories: Health Sciences, Immunology; Biology, Microbiology

Abstract

The signa ling molecule cyclic diguanylate (c-di-GMP) mediates physiological adaptation to extracellular stimuli in a wide range of bacteria. The complex metabolic pathways governing c-di-GMP synthesis and degradation are highly regulated, but the specific cues that impact c-di-GMP signaling are largely unknown. In the intestinal pathogen Clostridium difficile, c-di-GMP inhibits flagellar motility and toxin production and promotes pilus-dependent biofilm formation, but no specific biological functions have been ascribed to any of the individual c-di-GMP synthases or phosphodiesterases (PDEs). Here, we report the functional and biochemical characterization of a c-di-GMP PDE, PdcA, 1 of 37 confirmed or putative c-di-GMP metabolism proteins in C. difficile 630. Our studies reveal that pdcA transcription is controlled by the nutrient-regulated transcriptional regulator CodY and accordingly increases during stationary phase. In addition, PdcA PDE activity is allosterically regulated by GTP, further linking c-di-GMP levels to nutrient availability. Mutation of pdcA increased biofilm formation and reduced toxin biosynthesis without affecting swimming motility or global intracellular c-di-GMP. Analysis of the transcriptional response to pdcA mutation indicates that PdcA-dependent phenotypes manifest during stationary phase, consistent with regulation by CodY. These results demonstrate that inactivation of this single PDE gene is sufficient to impact multiple c-di-GMPdependent phenotypes, including the production of major virulence factors, and suggest a link between c-di-GMP signaling and nutrient availability.