In vivo repressed genes ofVibrio choleraereveal inverse requirements of an H+/Cl−transporter along the gastrointestinal passage

Abstract

The facultative human pathogenVibrio choleraechanges its transcriptional profile upon oral ingestion by the host to facilitate survival and colonization fitness. Here, we used a modified version of recombination-based in vivo expression technology to investigate gene silencing during the in vivo passage, which has been understudied. Using a murine model of cholera, we screened aV. choleraetransposon library composed of 10,000 randomly generated reporter fusions and identified 101 in vivo repressed (ivr) genes. Our data indicate that constitutive expression ofivrgenes reduces colonization fitness, highlighting the necessity to down-regulate these genes in vivo. For example, theivrgeneclcA, encoding an H+/Cl−transporter, could be linked to the acid tolerance response against hydrochloric acid. In a chloride-dependent manner, ClcA facilitates survival under low pH (e.g., the stomach), but its presence becomes detrimental under alkaline conditions (e.g., lower gastrointestinal tract). This pH-dependentclcAexpression is controlled by the LysR-type activator AphB, which acts in concert with AphA to initiate the virulence cascade inV. choleraeafter oral ingestion. Thus, transcriptional networks dictating induction of virulence factors and the repression ofivrgenes overlap to regulate in vivo colonization dynamics. Overall, the results presented herein highlight the impact of spatiotemporal gene silencing in vivo. The molecular characterization of the underlying mechanisms can provide important insights into in vivo physiology and virulence network regulation.