: Elucidating the mechanisms by which pathogenic bacteria regulate virulence gene expression in response to environmental stimuli is considerably important since this knowledge is central to understanding the molecular events that lead to the establishment of bacterial infections. Vibrio cholerae is the causative agent of epidemic cholera and possesses two distinct pathogenicity islands required for disease: the Vibrio pathogenicity island (VPI), which encodes an essential colonization factor, toxin-coregulated pilus (TCP), and the CTX element that carries the genes for cholera toxin (CT). We have recently identified two new activators, AphA and AphB, which function at what appears to be the initial regulatory step in the virulence transcriptional cascade. AphA and AphB function synergistically to activate the expression of the membrane bound transcriptional activators, TcpP and TcpH, which are encoded on the VPI. AphA appears to be a novel activator with no known homologs and AphB is a member of the LysR family. Interestingly, these two proteins are not encoded on either the VPI or CTX islands but are located in regions of the chromosome not previously associated with pathogenesis. Since activation of tcpPH expression by AphA and AphB occurs only under certain environmental conditions, these proteins may play a role in normal cellular physiology and function to couple physiological responses to virulence gene expression. The importance of AphA and AphB in pathogenesis is reflected by the dramatic attenuation in the infant mouse cholera model of mutants defective in either one of these genes. Furthermore, activation of tcpPH by AphA and AphB is responsible for the differential regulation of virulence genes between the two disease causing biotypes, classical and El Tor. The proposed research will allow us to gain a better understanding of the roles of AphA and AphB in pathogenesis. We propose: (1) to investigate the mechanism by which AphA and AphB differentially activate the expression of the classical and El Tor tcpPH promoters; (2) to determine the mechanism by which cAMP-CRP antagonizes AphA and AphB to negatively regulate tcpPH expression; (3) to elucidate the molecular mechanisms by which environmental stimuli influence the expression of the tcpPH promoter; and 4) to determine the importance of AphB-mediated differential activation of tcpPH expression for pathogenesis in vivo. These studies will enhance our understanding of the complex mechanisms utilized by pathogenic bacteria to regulate virulence gene expression in response to environmental stimuli so that better strategies can be developed to control and prevent bacterial infections.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI041558-10
Application #
7093003
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Program Officer
Hall, Robert H
Project Start
1997-07-01
Project End
2007-07-31
Budget Start
2006-08-01
Budget End
2007-07-31
Support Year
10
Fiscal Year
2006
Total Cost
$270,003
Indirect Cost
Name
Dartmouth College
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
041027822
City
Hanover
State
NH
Country
United States
Zip Code
03755
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Stonehouse, Emily A; Hulbert, Robin R; Nye, Melinda B et al. (2011) H-NS binding and repression of the ctx promoter in Vibrio cholerae. J Bacteriol 193:979-88
Kovacikova, Gabriela; Lin, Wei; Skorupski, Karen (2010) The LysR-type virulence activator AphB regulates the expression of genes in Vibrio cholerae in response to low pH and anaerobiosis. J Bacteriol 192:4181-91
Lowden, Michael J; Skorupski, Karen; Pellegrini, Maria et al. (2010) Structure of Vibrio cholerae ToxT reveals a mechanism for fatty acid regulation of virulence genes. Proc Natl Acad Sci U S A 107:2860-5
Jude, Brooke A; Martinez, Raquel M; Skorupski, Karen et al. (2009) Levels of the secreted Vibrio cholerae attachment factor GbpA are modulated by quorum-sensing-induced proteolysis. J Bacteriol 191:6911-7