Genetic analysis of the interaction between Vibrio cholerae transcription activator ToxR and toxT promoter DNA

Expression of many virulence genes in Vibrio cholerae is under the control of the ToxT protein, These include genes whose products are required for the biogenesis of the toxin-coregulated pilus, accessory colonization factor, and cholera toxin, ToxT is a member of the AraC family of transcriptional activators and is part of the ToxR regulatory cascade, ToxR is a transmembrane DNA-binding protein that is required for transcription of toxT and also can directly activate transcription of the cholera toxin operon (ctxAB), The sequences upstream of ctxAB and toxT to which ToxR binds show no obvious similarity, which implies that ToxR may be recognizing a degenerate sequence or, alternatively, a common structural motif within both binding sites, Data presented in this report demonstrate that nucleotides within the upstream half-site of an inverted repeat element in the toxT promoter are critical for ToxR-regulated activation of transcription in V, choler ae. In addition, gene fusion and DNA-binding studies with mutant ToxR proteins indicate that residues of ToxR required for binding to the ctx promoter are also required for binding to the toxT promoter. These data suggest that ToxR is not recognizing an inverted repeat sequence per se in the activation of toxT but, rather, some motif composed in part of sequences within the upstream half-site of the inverted repeat and that ToxR recognizes similar motifs within the ctxAB and toxT promoters.