Fig 1.

DNA sequence of the V. cholerae classical strain O395 promoter-proximal region of the toxT promoter and ToxR-dependent activation of single-base-pair substitutions. (A) Nucleotides are numbered relative to the toxT transcription start site (5). The region of ToxR-dependent DNase I protection is indicated above the DNA sequence (6). The solid gray arrows above the sequence indicate the position of the putative 5′-TNAAA-N₅-TNAAA-3′ direct repeat motif important for ToxR binding. An inverted repeat sequence (5, 50) is indicated by the black convergent arrows between −93 and −58. A promoter-proximal degenerate ToxR-binding site is indicated by dashed gray arrows from −69 to −56. The boxed nucleotides indicate the pentameric direct repeat motif recognized by TcpP (25). Single-nucleotide substitutions generated within the toxT promoter region from −100 to −57 are indicated on the bottom line in italics. (B) Effects of ToxR-binding site mutations on toxT-lacZ activity in wild-type V. cholerae strain O395. Strains carrying a plasmid-borne wild-type toxT-lacZ fusion (−172 to +45), single-base-pair substitution toxT promoter mutants, promoter deletion derivatives, or empty vector (promoterless lacZ vector, pTG24) were assessed for β-galactosidase activity. The positions of substitutions and endpoints are indicated relative to the toxT transcription start site. Error bars represent the standard deviations for each data set. *, P < 0.005 as assessed using the Student t test, n = 6 or more measurements.