The Gram-negative bacterium Vibrio cholerae, the causative agent of cholera, is a facultative pathogen that resides in both human and aquatic environments. Extensive in vitro studies have identified a number of virulence factors required to produce disease during infection. However, how V. cholerae alters its gene expression upon transition from its marine ecosystem to the human host and along progression of infection is largely unknown. We have discovered that the transcription factor AphB possesses a cysteine residue that undergoes modification in response to the microoxic conditions of the intestines, leading to activation of virulence. We now have further evidence that diverse cysteine modifications in AphB lead to diverse effects on cell physiology through changes in gene transcription and protein stability. We hypothesize that AphB is the central processor of environmental information relevant to infection for V. cholerae. Specifically, we hypothesize that AphB uses thiol modifications to integrate the presence of reductive, oxidative and nitrosative reactants into the gene expression decisions necessary to guide the organism in and out of the human gut. We will use a mix of biochemical and genetic techniques to define mechanistically how AphB orchestrates the pathogenic life cycle of V. cholerae and what other factors are involved at this cysteine-based sensory hub. We believe that by comprehensively defining the way AphB monitors the chemical microenvironment for V. cholerae, we then may be able to extend the paradigm of cysteine-based environmental sensing to other V. cholerae proteins and other pathogens. We will examine how AphB is modified by reactive nitrogen species (RNS), such as nitric oxide (NO), and oxidative stress from reactive oxygen species (ROS) generated in vivo and how these modifications affect AphB functionality. We will study the broad effects of AphB modification on cell physiology, focusing on ROS/RNS stress management with the following model in mind: upon entry to the gut, reduced AphB activates virulence in response to low oxygen tension;in response to increasing chemical stress in the gut, modified AphB deactivates virulence while up-regulating ROS and RNS detoxification, thus preparing the cell for survival in the aquatic environment.

Public Health Relevance

Cholera is a devastating diarrheal illness currently taking lives in poverty-stricken countries around the world, including Haiti. This study proposed will shed light on importance of V. cholerae's in vivo gene regulations in host-pathogen interactions, with the goal of better understanding V. cholerae pathogenesis and, potentially, discovering novel treatment options for the cholera disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21AI109316-01A1
Application #
8769027
Study Section
Special Emphasis Panel (ZRG1-IDM-B (80))
Program Officer
Hall, Robert H
Project Start
2014-06-05
Project End
2016-05-31
Budget Start
2014-06-05
Budget End
2015-05-31
Support Year
1
Fiscal Year
2014
Total Cost
$240,000
Indirect Cost
$90,000
Name
University of Pennsylvania
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104