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J Bacteriol. Jun 1988; 170(6): 2575–2583.
PMCID: PMC211174

A novel suicide vector and its use in construction of insertion mutations: osmoregulation of outer membrane proteins and virulence determinants in Vibrio cholerae requires toxR.

Abstract

The toxR gene of Vibrio cholerae encodes a transmembrane, DNA-binding protein that activates transcription of the cholera toxin operon and a gene (tcpA) for the major subunit of a pilus colonization factor. We constructed site-directed insertion mutations in the toxR gene by a novel method employing the chromosomal integration of a mobilizable suicide plasmid containing a portion of the toxR coding sequence. Mutants containing these new toxR alleles had an altered outer membrane protein profile, suggesting that two major outer membrane proteins (OmpT and OmpU) might be under the control of toxR. Physiological studies indicated that varying the concentration of the amino acids asparagine, arginine, glutamate, and serine caused coordinate changes in the expression of cholera toxin, TcpA, OmpT, and OmpU. Changes in the osmolarity of a tryptone-based medium also produced coordinate changes in the expression of these proteins. Other environmental signals (temperature and pH) had a more pronounced effect on the expression of cholera toxin and TcpA than they did on the outer membrane proteins. These results suggest that certain environmental signals (i.e., osmolarity and the presence of amino acids) are tightly coupled to the expression of toxR-regulated proteins and therefore may be signals that are directly sensed by the ToxR protein.

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  • Betley MJ, Miller VL, Mekalanos JJ. Genetics of bacterial enterotoxins. Annu Rev Microbiol. 1986;40:577–605. [PubMed]
  • Calderwood SB, Mekalanos JJ. Iron regulation of Shiga-like toxin expression in Escherichia coli is mediated by the fur locus. J Bacteriol. 1987 Oct;169(10):4759–4764. [PMC free article] [PubMed]
  • Callahan LT, 3rd, Richardson SH. Biochemistry of Vibrio cholerae virulence. 3. Nutritional requirements for toxin production and the effects of pH on toxin elaboration in chemically defined media. Infect Immun. 1973 Apr;7(4):567–572. [PMC free article] [PubMed]
  • Hall MN, Silhavy TJ. Genetic analysis of the major outer membrane proteins of Escherichia coli. Annu Rev Genet. 1981;15:91–142. [PubMed]
  • Hall MN, Silhavy TJ. Genetic analysis of the ompB locus in Escherichia coli K-12. J Mol Biol. 1981 Sep 5;151(1):1–15. [PubMed]
  • Holmes RK, Baine WB, Vasil ML. Quantitative measurements of cholera enterotoxin in cultures of toxinogenic wild-type and nontoxinogenic mutant strains of Vibrio cholerae by using a sensitive and specific reversed passive hemagglutination assay for cholera enerotoxin. Infect Immun. 1978 Jan;19(1):101–106. [PMC free article] [PubMed]
  • Holmgren J. Comparison of the tissue receptors for Vibrio cholerae and Escherichia coli enterotoxins by means of gangliosides and natural cholera toxoid. Infect Immun. 1973 Dec;8(6):851–859. [PMC free article] [PubMed]
  • Huq A, West PA, Small EB, Huq MI, Colwell RR. Influence of water temperature, salinity, and pH on survival and growth of toxigenic Vibrio cholerae serovar 01 associated with live copepods in laboratory microcosms. Appl Environ Microbiol. 1984 Aug;48(2):420–424. [PMC free article] [PubMed]
  • Kelley JT, Parker CD. Identification and preliminary characterization of Vibrio cholerae outer membrane proteins. J Bacteriol. 1981 Feb;145(2):1018–1024. [PMC free article] [PubMed]
  • Kolter R, Inuzuka M, Helinski DR. Trans-complementation-dependent replication of a low molecular weight origin fragment from plasmid R6K. Cell. 1978 Dec;15(4):1199–1208. [PubMed]
  • Laimins LA, Rhoads DB, Epstein W. Osmotic control of kdp operon expression in Escherichia coli. Proc Natl Acad Sci U S A. 1981 Jan;78(1):464–468. [PMC free article] [PubMed]
  • Le Rudulier D, Strom AR, Dandekar AM, Smith LT, Valentine RC. Molecular biology of osmoregulation. Science. 1984 Jun 8;224(4653):1064–1068. [PubMed]
  • Maurelli AT, Blackmon B, Curtiss R., 3rd Temperature-dependent expression of virulence genes in Shigella species. Infect Immun. 1984 Jan;43(1):195–201. [PMC free article] [PubMed]
  • Mekalanos JJ. Duplication and amplification of toxin genes in Vibrio cholerae. Cell. 1983 Nov;35(1):253–263. [PubMed]
  • Mekalanos JJ, Collier RJ, Romig WR. Affinity filters, a new approach to the isolation of tox mutants of Vibrio cholerae. Proc Natl Acad Sci U S A. 1978 Feb;75(2):941–945. [PMC free article] [PubMed]
  • Mekalanos JJ, Moseley SL, Murphy JR, Falkow S. Isolation of enterotoxin structural gene deletion mutations in Vibrio cholerae induced by two mutagenic vibriophages. Proc Natl Acad Sci U S A. 1982 Jan;79(1):151–155. [PMC free article] [PubMed]
  • Mekalanos JJ, Swartz DJ, Pearson GD, Harford N, Groyne F, de Wilde M. Cholera toxin genes: nucleotide sequence, deletion analysis and vaccine development. Nature. 1983 Dec 8;306(5943):551–557. [PubMed]
  • Miller VL, Mekalanos JJ. Synthesis of cholera toxin is positively regulated at the transcriptional level by toxR. Proc Natl Acad Sci U S A. 1984 Jun;81(11):3471–3475. [PMC free article] [PubMed]
  • Miller VL, Mekalanos JJ. Genetic analysis of the cholera toxin-positive regulatory gene toxR. J Bacteriol. 1985 Aug;163(2):580–585. [PMC free article] [PubMed]
  • Miller VL, Taylor RK, Mekalanos JJ. Cholera toxin transcriptional activator toxR is a transmembrane DNA binding protein. Cell. 1987 Jan 30;48(2):271–279. [PubMed]
  • Murphy JR, Skiver J, McBride G. Isolation and partial characterization of a corynebacteriophage beta, tox operator constitutive-like mutant lysogen of Corynebacterium diphtheriae. J Virol. 1976 Apr;18(1):235–244. [PMC free article] [PubMed]
  • Sagar IK, Nagesha CN, Bhat JV. Effect of metal ions on the production of vascular permeability factor by 569B strain of Vibrio cholerae. Indian J Med Res. 1979 Jan;69:18–25. [PubMed]
  • Sagar IK, Nagesha CN, Bhat JV. The role of trace elements and phosphates in the synthesis of vascular-permeability factor by Vibrio cholerae. J Med Microbiol. 1981 Aug;14(3):243–250. [PubMed]
  • Shimamura T, Watanabe S, Sasaki S. Enhancement of enterotoxin production by carbon dioxide in Vibrio cholerae. Infect Immun. 1985 Aug;49(2):455–456. [PMC free article] [PubMed]
  • Straley SC, Bowmer WS. Virulence genes regulated at the transcriptional level by Ca2+ in Yersinia pestis include structural genes for outer membrane proteins. Infect Immun. 1986 Feb;51(2):445–454. [PMC free article] [PubMed]
  • Tamplin ML, Colwell RR. Effects of microcosm salinity and organic substrate concentration on production of Vibrio cholerae enterotoxin. Appl Environ Microbiol. 1986 Aug;52(2):297–301. [PMC free article] [PubMed]
  • Taylor RK, Miller VL, Furlong DB, Mekalanos JJ. Use of phoA gene fusions to identify a pilus colonization factor coordinately regulated with cholera toxin. Proc Natl Acad Sci U S A. 1987 May;84(9):2833–2837. [PMC free article] [PubMed]
  • Thuring RW, Sanders JP, Borst P. A freeze-squeeze method for recovering long DNA from agarose gels. Anal Biochem. 1975 May 26;66(1):213–220. [PubMed]
  • Weiss AA, Hewlett EL. Virulence factors of Bordetella pertussis. Annu Rev Microbiol. 1986;40:661–686. [PubMed]

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