sRNAs and Vibrio cholerae Virulence
The bacterium Vibrio cholerae is the causative agent of Asiatic cholera, a severe secretory diarrhea that can be rapidly fatal if left untreated. V. cholerae persists in fresh and estuarine aquatic environments in temperate zones throughout the world. It undergoes dramatic changes in gene expression when switching between the aquatic environment and the human intestinal tract. In the human host, it expresses numerous virulence factors including cholera toxin and the Toxin-Coregulated Pilus (TCP). While many of the critical regulators of virulence in this organism have been determined, it has recently been appreciated that dynamic gene expression changes occur during infection. These lead to subpopulations of bacteria with different gene expression profiles and thus different phenotypes such as populations involved in tissue colonization, spreading to other sites in the intestine, toxin delivery to epithelial cells and dissemination out of the host in the diarrhea. The mediators of these dynamic gene expression changes during infection remain to be determined.
Small Regulatory RNAs (sRNAs) are a relatively recently discovered class of regulators in bacteria that mediate changes in gene expression via sRNA-mRNA (messenger RNA) interactions. Such interactions can change the stability of the mRNA and/or regulate its ability to be translated efficiently. To determine if sRNAs contribute to gene regulation in V. cholerae during infection, Evan Bradley, an MD-PhD student completing his PhD in Molecular Microbiology, sought to identify new sRNAs under the control of the major virulence gene regulator, ToxT.
ToxT is a transcription factor that controls expression of cholera toxin, TCP and many other virulence factors. Evan used high throughput DNA sequencing to identify sRNAs that were expressed at higher levels after artificial induction of ToxT during in vitro growth. In a subsequent experiment, he determined the subset of these sRNAs that were under the direct control of ToxT by performing a genome-wide binding assay for sRNA gene promoters that were bound by ToxT. These experiments revealed two sRNAs within the Vibrio Pathogenicity Island, a locus that encodes ToxT, TCP and other virulence factors. One of these sRNAs, which we named TarB, was found to repress an essential virulence factor, TcpF. Because TarB is activated by ToxT and yet negatively regulates synthesis of an essential virulence factor, it seems logical that TarB may be involved in dynamic gene expression changes that occur during infection.
Some of Evan's work on this topic was published last summer in PLoS Pathogen (Bradley E, et al, 2011 PLoS Pathog. 7: e1002126 
Evan is currently investigating the temporal and spatial patterns of TarB expression during infection using fluorescent protein reporters. Through this analysis we hope to elucidate TarB’s role in generating subpopulations of V. cholerae that are specialized for different roles during infection.