CTXphi is a filamentous bacteriophage that encodes cholera toxin. This is the principal virulence factor of Vibrio cholerae, the Gram-negative bacterium that causes the severe diarrheal disease cholera. CTXphi is the first filamentous bacteriophage shown to mediate the horizontal transfer of a virulence gene. CTXphi integrates into the Vibrio cholerae chromosome and, in the lysogenic state, most CTXphi genes are not expressed due to the activity of the CTXphi repressor, RstR. Generally, the integrated form of CTXphi is found as part of tandem arrays of prophage DNA interspersed with the related genetic element RS1. RS1 encodes a protein, RstC, that can counter RstR repression and lead to markedly enhanced expression of CTX prophage genes including ctxAB, the genes encoding cholera toxin. ? ? The long-term goal of this work is to understand the molecular events in the life cycle of CTXphi and the role that this phage plays in the pathogenesis of cholera. The proposed studies will explore 3 processes central to the phage life cycle: i) the site-specific integration of phage DNA into the bacterial chromosome; ii) the repression of most phage gene expression following integration; and iii) the activation of phage gene expression and virion production by environmental and genetic stimuli. Experiments in Aim 1 to identify the mechanism and factors that mediate the integration of CTXphi DNA into the V. cholerae chromosome will reveal how the chromosome encoded recombinases XerC and XerD interact with phage and chromosome sequences to accomplish CTXphi integration. These studies will elucidate a novel mechanism of phage integration and may shed light on the mechanism of ctxAB amplification as well. Experiments in Aim 2: to characterize the regulation and mode of action of RstR will clarify how CTXphi can be maintained in a quiescent state. rstR autoregulation and modulation of RstR levels by environmental factors will be explored. RstR's binding to its unusual operators will also be studied. Experiments in Aim 3 to determine the mode of action of RstC-will explore how RstC can inactivate RstR-mediated repression. RstC's ability to bind to either RstR and/or RstR's binding sites will be investigated and the expression of rstC during infection will be measured. All of these studies will yield insights into fundamental aspects of phage biology. In addition, they may reveal ways in which changes in phage gene expression or copy number can contribute to the pathogenicity of V. cholerae.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
7R01AI042347-12
Application #
7355845
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Program Officer
Hall, Robert H
Project Start
1998-01-01
Project End
2007-12-31
Budget Start
2007-03-01
Budget End
2007-12-31
Support Year
12
Fiscal Year
2007
Total Cost
$214,104
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
030811269
City
Boston
State
MA
Country
United States
Zip Code
02115
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