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Cefoperazone-treated Mouse Model of Clinically-relevant Clostridium difficile Strain R20291
Published on: December 10, 2016
CcpA-mediated repression of Clostridium difficile toxin gene expression
Ana Antunes1, Isabelle Martin-Verstraete, Bruno Dupuy
1Laboratoire Pathogenèse des Bactéries Anaérobies, Institut Pasteur, Paris, France.
Molecular Microbiology
|February 9, 2011
Summary
Carbon catabolite repression (CCR) in Clostridium difficile regulates toxin synthesis. Glucose uptake, mediated by CcpA, represses toxin genes, linking nutrient availability to virulence.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Molecular Biology
Background:
- Clostridium difficile toxin production is repressed by glucose.
- Carbon catabolite repression (CCR) is a known regulatory mechanism in Gram-positive bacteria.
- The C. difficile genome contains all necessary CCR components.
Purpose of the Study:
- To investigate the in vivo role of CCR in C. difficile toxin synthesis.
- To identify the specific components of the CCR pathway involved in toxin regulation.
- To elucidate the mechanism by which C. difficile CcpA regulates toxin gene expression.
Main Methods:
- ClosTron gene knockout system used to create mutants in C. difficile strain JIR8094.
- Construction of mutants lacking key CCR pathway proteins: PTS proteins (Enzyme I, HPr), HprK/P, and CcpA.
- Analysis of toxin gene expression in response to glucose in wild-type and mutant strains.
- In vitro and in vivo assays to assess CcpA DNA binding and the role of HPr phosphorylation and FBP.
Main Results:
- Inactivation of ptsI, ptsH, and ccpA genes led to derepression of toxin gene expression in the presence of glucose.
- The hprK mutant still showed repression of toxin production, indicating glucose uptake is necessary but HPr phosphorylation by HprK is not essential for repression.
- C. difficile CcpA binds to the regulatory regions of tcdA and tcdB genes, but not via a canonical cre site.
- HPr-Ser45-P did not enhance CcpA binding, while fructose-1,6-biphosphate (FBP) increased CcpA binding affinity.
Conclusions:
- CcpA directly represses C. difficile toxin expression in response to PTS sugar availability.
- This study establishes a direct link between carbon source utilization and virulence gene expression in C. difficile.
- The mechanism involves CcpA binding to toxin gene regulatory regions, influenced by FBP, independent of HPr phosphorylation.
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