Abbreviated Summary: Virulence gene regulation in Vibrio cholerae is highly complex, comprising several transcriptional activators and decision check points. The ToxRS complex is a key regulator that is subjected to regulated intramembrane proteolysis (RIP). The participating interaction partners were characterized, addressing DegS, DegP, ToxS and factors influencing ToxR-RIP, such as bile acids and the cysteine redox state of ToxR. In Vibrio cholerae, virulence gene expression is regulated by a transmembrane-localized transcription factor complex designated as ToxRS. ToxR harbours two cysteines in the periplasmic domain that can form inter- and intramolecular disulfide bonds. In this study, we investigated the σE−dependent inner membrane proteolysis of ToxR, which occurs via the periplasmic-localized proteases DegS and DegP. Both proteases respond to the redox state of the two cysteine thiol groups of ToxR. Interestingly, in the presence of sodium deoxycholate, ToxR proteolysis is blocked independently of ToxS, whereas ToxR activation by bile salts requires ToxS function. From these data, we identified at least two levels of control for ToxR activation by sodium-deoxycholate. First, bile inhibits ToxR degradation under starvation and alkaline pH or under conditions in which DegPS responds to the reduced disulfide bonds of ToxR. The second level links bile to ToxRS complex formation and further activation of its transcription factor activity. Overall, our data suggest a comprehensive bile sensory function for the ToxRS complex during host colonization.

Prokaryotes are unicellular organisms that require sensory networks for their survival in rapidly changing habitats. In the course of evolution, transmembrane signaling systems have evolved to transmit signals from the extracellular environment across the cytoplasmic membrane into the cell. One-component signaling systems represent the oldest and simplest solution for such signal transmission, whereas two-component systems are evolutionarily younger (Ulrich et al., 2005). Although one-component systems are widely distributed among bacteria, only 3% are directly integrated into cytoplasmic membranes (Ulrich et al., 2005). A literature search revealed a non-exhaustive list of signaling molecules that includes ToxRS,

The lifecycle of the causative agent of the severe secretory diarrheal disease cholera, Vibrio cholerae , is characterized by the transition between two dissimilar habitats, i.e., as a natural inhabitant of aquatic ecosystems and as a pathogen in the human gastrointestinal tract. Vibrio cholerae faces diverse stressors along its lifecycle, which require effective adaptation mechanisms to facilitate the survival fitness. Not surprisingly, the pathogen's transcriptome undergoes global changes during the different stages of the lifecycle. Moreover, recent evidence indicates that several of the transcription factors (i.e., ToxR, TcpP, and ToxT) and alternative sigma factors (i.e., FliA, RpoS, and RpoE) involved in transcriptional regulations along the lifecycle are controlled by regulated proteolysis. This post-translational control ensures a fast strategy by the pathogen to control cellular checkpoints and thereby rapidly respond to changing conditions. In this review, we discuss selected targets for regulated proteolysis activated by various stressors, which represent a key feature for fast adaptation of V. cholerae .