The Role of Toxr Signal Transduction in Vibrio Cholerae Pathogenesis

Vibrio cholerae (Vc) is the causative agent of the severe diarrheal disease cholera, a disease that affects3-5 million people each year. V. cholerae is a native to aquatic ecosystems; its worldwide disruption andepidemic capability makes V. cholerae a significant public health threat. Cholera is acquired by ingestionof Vc-contaminated food or water. Upon entering the gastrointestinal tract, V. cholerae responds tounknown in vivo environmental cues to activate expression of the ToxR regulon, a regulatory cascadethat controls expression of genes that facilitate colonization and virulence factor production. Onceestablished in the small intestine, Vc replicates to a high cell density before exiting the host in thediarrheal purge that is the hallmark of disease. This represents a significant dissemination event thatcontributes to epidemic spread of cholera. Phenotypic analysis has shown that the repression of theToxR regulon occurs late in infection while genes that are important for dissemination and transmissionare activated. The regulatory elements and signals that contribute to late infection phenotypes areunknown. Previous studies in our laboratory characterized the activity of a cyclic dipeptide metabolite,cFP, and showed that cFP inhibits virulence factor production by a ToxR-dependent mechanism. Thefunction of ToxR in the establishment of Vc in the gastrointestinal tract has been well studied; however,the mechanism by which ToxR integrates environmental signals to modulate gene expression remainspoorly understood. The finding that ToxR can function to repress virulence expression in response toenvironmental cues significantly expands the role of ToxR in Vc pathogenesis. This exemplifies theimportance of delineating the mechanisms of ToxR signal transduction to gain to a better understandingof virulence regulation. ToxR is a one-component transcriptional activator that contains a cytoplasmicDNA-binding domain and a periplasmic domain proposed to function in signal sensing; these domainsare linked by a transmembrane domain. Preliminary studies indicate that the periplasmic domain isrequired for ToxR-dependent gene induction in response to certain stimuli. Therefore, we hypothesizethat ToxR functions to transduce signals by directly interacting with environmental cues via itsperiplasmic domain. Two aims are proposed to test this hypothesis. Aim 1 will define the role of theToxR periplasmic domain (PPD) in environmental sensing. This includes characterizing the interactionbetween effector molecules and the PPD and identifying functional regions within the PPD that contributeto ToxR activity. Aim 2 will characterize the mechanism of ToxR-dependent transcription activation.Results from these studies will provide significant insight into ToxR signal transduction pathways.Furthermore, determining the mechanism of action of antivirulence compounds, like cyclic dipeptides, willcontribute to the development of novel approaches to combat cholera.