The virulence gene activator ToxT from Vibrio cholerae is a member of the AraC family of transcriptional activators

Abstract

Virulence gene expression in Vibrio cholerae is postulated to involve ToxR-dependent activation of the toxT gene followed by ToxT activation of virulence genes, including several of those involved in biogenesis of the toxin-coregulated pilus. ToxR is a transmembrane, DNA-binding protein which is a member of the OmpR subclass of two-component activator systems in bacteria. Data presented in this report demonstrate that ToxT is similar to the AraC family of transcriptional activators identified in a variety of gram-negative bacteria. The toxT open reading frame begins approximately 200 nucleotides from the end of the tcpF gene, which is part of a cluster of genes responsible for production of the toxin-coregulated pilus. Accumulation of toxT specific mRNA is ToxR dependent and is modulated by environmental conditions that modulate expression of the regulon. Within the intergenic region between tcpF and toxT is a potential stem-loop structure of an unusual nature which may play a role in regulating expression of toxT mRNA. Experiments with tcpF and toxT cloned behind a strong, constitutive promoter suggest that the two genes can be cotranscribed, but Northern (RNA) blot analysis of V. cholerae suggests that if they are, steady-state levels of their messages may be controlled by a posttranscriptional mechanism. Possible mechanisms for ToxR-dependent expression of toxT are discussed.

Phase variation in tcpH modulates expression of the ToxR regulon in Vibrio cholerae

“…The toxT alleles from O395 and KTT42 were also sequenced from the polymerase chain reaction (PCR) products, starting at bp 1070 within the tcpF gene upstream of the toxT promoter (Higgins and DiRita, 1994) and extending to bp 2132 downstream of the stop codon in toxT. Both sequences differed in one nucleotide from the published sequence of toxT from strain 569B (Higgins et al, 1992), a G to a T at bp 1478, resulting in an alanine to serine substitution, but results were identical between strain O395 and KTT42 (not shown). This same sequence change was found in toxT from another V. cholerae strain reported by Ogierman and Manning (1992).…”

“…The gene encoding ToxT is located within the tcp gene cluster, between tcpF and tcpJ (Higgins et al, 1992;Kaufman et al, 1993;Ogierman et al, 1993). Just downstream of the gene for tcpF, there is a relatively strong transcriptional terminator (approximately 80% effective) (Higgins and DiRita, 1994).…”

A branch in the ToxR regulatory cascade of Vibrio cholerae revealed by characterization of toxT mutant strains

“…Its carboxyl terminal domain has a helix-turn-helix motif characteristic of this family of activators, and, like other AraC-like proteins, its amino terminus does not share significant similarity to other proteins in the database (Higgins et al, 1992). Transcription of toxT in V. cholerae is, in part, due to a ToxR-dependent promoter present upstream of the initiation of toxT transcription.…”

“…Sequences within this promoter are bound by ToxR in gel-shift experiments (Higgins and DiRita, 1994). Interaction between ToxR and the toxT promoter requires the upstream half-site of an inverted repeat element for activation and DNA binding by ToxR (Higgins et al, 1992;Higgins and DiRita, 1996). The ToxR-binding site in the toxT promoter differs in primary sequence from its binding site in the ctxAB promoter, which is characterized by a heptad repeat, TTTTGAT, as well as sequences downstream of the heptad repeat Pfau and Taylor, 1996).…”

“…ToxT is an AraC-like protein that activates several genes in the ToxR regulon (Higgins et al, 1992;DiRita et al, 1991;Ogierman and Manning, 1992). Its carboxyl terminal domain has a helix-turn-helix motif characteristic of this family of activators, and, like other AraC-like proteins, its amino terminus does not share significant similarity to other proteins in the database (Higgins et al, 1992).…”