Vibrio cholerae TolC Is Required for Expression of the ToxR Regulon

Vibrio cholerae is a Gram-negative bacterium that causes the enteric disease cholera. V. cholerae colonization of the human intestine is dependent on the expression of both virulence genes and environmental adaptation genes involved in antimicrobial resistance. The expression of virulence genes, including the genes encoding the main virulence factors cholera toxin (CT) and the toxin-coregulated pilus (TCP), are coordinately regulated by the ToxR regulon. Tripartite transport systems belonging to the ATP binding cassette, major facilitator, and resistance-nodulation-division families are critical for V. cholerae pathogenesis. Transport systems belonging to these families contribute to myriad phenotypes, including protein secretion, antimicrobial resistance, and virulence. TolC plays a central role in bacterial physiology by functioning as the outer membrane pore protein for tripartite transport systems. Consistent with this, V. cholerae tolC was previously found to be required for MARTX toxin secretion and antimicrobial resistance. Here, we investigated the contribution of TolC to V. cholerae virulence. We documented that tolC was required for CT and TCP production in O1 El Tor V. cholerae. This phenotype was linked to repression of the critical ToxR regulon transcription factor aphA. Decreased aphA transcription correlated with increased expression of the LysR-family transcription factor leuO. Deletion of leuO restored aphA expression, and CT and TCP production, in a tolC mutant. The collective results document that tolC is required for ToxR regulon expression and further suggest that tolC participates in an efflux-dependent feedback circuit to regulate virulence gene expression.