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

Infect Immun. 2017 Aug 18;85(9):e00347-17. doi: 10.1128/IAI.00347-17. Print 2017 Sep.


The signaling 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-GMP-dependent phenotypes, including the production of major virulence factors, and suggest a link between c-di-GMP signaling and nutrient availability.

Keywords: CodY; biofilm; c-di-GMP; cyclic diguanylate; flagella; flagellar motility; nutrient; toxin.

MeSH terms

  • Bacterial Toxins / metabolism*
  • Biofilms / growth & development*
  • Clostridioides difficile / enzymology*
  • Clostridioides difficile / metabolism
  • Clostridioides difficile / physiology*
  • Cyclic GMP / analogs & derivatives*
  • Cyclic GMP / metabolism
  • Gene Expression Regulation, Bacterial
  • Gene Knockout Techniques
  • Locomotion
  • Phosphoric Diester Hydrolases / genetics
  • Phosphoric Diester Hydrolases / metabolism*


  • Bacterial Toxins
  • bis(3',5')-cyclic diguanylic acid
  • Phosphoric Diester Hydrolases
  • Cyclic GMP