Clostridium difficile is the principal causative agent of antibiotic-associated colitis and the only known cause of pseudomembranous colitis, a potentially lethal disease. Two large toxin proteins, encoded by the toxA and toxB genes, which lie within a 19 kb pathogenicity islet, are the primary virulence factors. Previous work from the applicants'laboratories has shown that transcription of the tox genes depends entirely on TxeR, a novel RNA polymerase sigma factor encoded within the same pathogenicity locus. The level of expression is strongly modulated, however, by the growth state of the cells and by environmental conditions. For instance, for cells growing in broth medium, tox gene transcription is restricted to stationary phase cells and is repressed by rapidly metabolizable carbon sources, such as glucose. Other factors that influence toxin synthesis are the availability of biotin and certain amino acids and the temperature of cultivation. The present proposal seeks to determine the molecular mechanisms that control toxin synthesis in response to environmental signals. Candidate regulatory proteins, based on comparative genomics, will be specifically tested for their participation in such regulation. These proteins include CodY, CcpA and VirR, whose homologs in Bacillus subtilis or Clostridium perfringens have been shown to mediate the types of regulatory effects in question. In addition, general, unbiased searches for the relevant regulatory proteins will be carried out based on affinity chromatography. A fourth protein, TcdC, will be tested for its potential activity as an antagonist of TxeR. The project makes use of the expertise in C. difficile biochemistry, genetics and physiology of the three collaborating research groups and recent advances made by each of the groups in making this experimental system amenable to detailed molecular genetic analysis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
3R01AI057637-05S1
Application #
8071802
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Program Officer
Wachtel, Marian R
Project Start
2010-06-01
Project End
2010-09-30
Budget Start
2010-06-01
Budget End
2010-09-30
Support Year
5
Fiscal Year
2010
Total Cost
$16,883
Indirect Cost
Name
Tufts University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
039318308
City
Boston
State
MA
Country
United States
Zip Code
02111
Martin-Verstraete, Isabelle; Peltier, Johann; Dupuy, Bruno (2016) The Regulatory Networks That Control Clostridium difficile Toxin Synthesis. Toxins (Basel) 8:
Suarez, Jose M; Edwards, Adrianne N; McBride, Shonna M (2013) The Clostridium difficile cpr locus is regulated by a noncontiguous two-component system in response to type A and B lantibiotics. J Bacteriol 195:2621-31
Liu, Hualan; Bouillaut, Laurent; Sonenshein, Abraham L et al. (2013) Use of a mariner-based transposon mutagenesis system to isolate Clostridium perfringens mutants deficient in gliding motility. J Bacteriol 195:629-36
Govind, Revathi; Dupuy, Bruno (2012) Secretion of Clostridium difficile toxins A and B requires the holin-like protein TcdE. PLoS Pathog 8:e1002727
Antunes, Ana; Camiade, Emilie; Monot, Marc et al. (2012) Global transcriptional control by glucose and carbon regulator CcpA in Clostridium difficile. Nucleic Acids Res 40:10701-18
McBride, Shonna M; Sonenshein, Abraham L (2011) The dlt operon confers resistance to cationic antimicrobial peptides in Clostridium difficile. Microbiology 157:1457-65
Bouillaut, Laurent; McBride, Shonna M; Sorg, Joseph A (2011) Genetic manipulation of Clostridium difficile. Curr Protoc Microbiol Chapter 9:Unit 9A.2
Carter, Glen P; Douce, Gillian R; Govind, Revathi et al. (2011) The anti-sigma factor TcdC modulates hypervirulence in an epidemic BI/NAP1/027 clinical isolate of Clostridium difficile. PLoS Pathog 7:e1002317
Antunes, Ana; Martin-Verstraete, Isabelle; Dupuy, Bruno (2011) CcpA-mediated repression of Clostridium difficile toxin gene expression. Mol Microbiol 79:882-99
McBride, Shonna M; Sonenshein, Abraham L (2011) Identification of a genetic locus responsible for antimicrobial peptide resistance in Clostridium difficile. Infect Immun 79:167-76

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