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. 1994 Jul;62(7):2669–2678. doi: 10.1128/iai.62.7.2669-2678.1994

The Vibrio cholerae toxin-coregulated-pilus gene tcpI encodes a homolog of methyl-accepting chemotaxis proteins.

C W Harkey 1, K D Everiss 1, K M Peterson 1
PMCID: PMC302867  PMID: 8005659

Abstract

Virulence gene activation in Vibrio cholerae is under the control of the ToxR-ToxT regulatory cascade. The ToxR regulon consists of genes required for toxin-coregulated-pilus (TCP) biogenesis, accessory colonization factor genes, cholera toxin genes, and ToxR-activated genes (tag) of unknown function. The tagB gene was isolated by using a tagB::TnphoA fusion junction to probe a V. cholerae )395 bacteriophage lambda library. Nucleotide sequence analysis revealed that tagB is identical to tcpI, a gene which encodes a protein that negatively regulates the synthesis of the major pilin subunit of TCP (TcpA). Our results show that the tcpI gene encodes a 620-amino-acid protein that shares extensive sequence similarity with the highly conserved signaling domain in methyl-accepting chemotaxis proteins. Expression of tcpI in Escherichia coli results in the synthesis of a 71-kDa polypeptide that becomes localized to the inner membrane. Similarly, TcpI-PhoA alkaline phosphatase activity is enriched in V. cholerae inner membrane preparations. Colonies of V. cholerae tcpI::TnphoA mutant cells display increased swarming on solid media when compared with those of the parental V. cholerae O395. Taken together, these observations suggest that TcpI may play a dual role in promoting vibrio colonization of the small bowel. In response to the appropriate environmental signal(s), TcpI permits maximum expression of tcpA while simultaneously reducing vibrio chemotaxis-directed motility. We believe coordinate regulation of colonization and motility determinants, in such a fashion, facilitates efficient V. cholerae microcolony formation.

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  1. Alley M. R., Maddock J. R., Shapiro L. Polar localization of a bacterial chemoreceptor. Genes Dev. 1992 May;6(5):825–836. doi: 10.1101/gad.6.5.825. [DOI] [PubMed] [Google Scholar]
  2. Allmeier H., Cresnar B., Greck M., Schmitt R. Complete nucleotide sequence of Tn1721: gene organization and a novel gene product with features of a chemotaxis protein. Gene. 1992 Feb 1;111(1):11–20. doi: 10.1016/0378-1119(92)90597-i. [DOI] [PubMed] [Google Scholar]
  3. Alm R. A., Manning P. A. Characterization of the hlyB gene and its role in the production of the El Tor haemolysin of Vibrio cholerae O1. Mol Microbiol. 1990 Mar;4(3):413–425. doi: 10.1111/j.1365-2958.1990.tb00608.x. [DOI] [PubMed] [Google Scholar]
  4. Baudry B., Fasano A., Ketley J., Kaper J. B. Cloning of a gene (zot) encoding a new toxin produced by Vibrio cholerae. Infect Immun. 1992 Feb;60(2):428–434. doi: 10.1128/iai.60.2.428-434.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bollinger J., Park C., Harayama S., Hazelbauer G. L. Structure of the Trg protein: Homologies with and differences from other sensory transducers of Escherichia coli. Proc Natl Acad Sci U S A. 1984 Jun;81(11):3287–3291. doi: 10.1073/pnas.81.11.3287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Borkovich K. A., Kaplan N., Hess J. F., Simon M. I. Transmembrane signal transduction in bacterial chemotaxis involves ligand-dependent activation of phosphate group transfer. Proc Natl Acad Sci U S A. 1989 Feb;86(4):1208–1212. doi: 10.1073/pnas.86.4.1208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Boyd A., Kendall K., Simon M. I. Structure of the serine chemoreceptor in Escherichia coli. Nature. 1983 Feb 17;301(5901):623–626. doi: 10.1038/301623a0. [DOI] [PubMed] [Google Scholar]
  8. Bradley D. E. A function of Pseudomonas aeruginosa PAO polar pili: twitching motility. Can J Microbiol. 1980 Feb;26(2):146–154. doi: 10.1139/m80-022. [DOI] [PubMed] [Google Scholar]
  9. Darzins A. Characterization of a Pseudomonas aeruginosa gene cluster involved in pilus biosynthesis and twitching motility: sequence similarity to the chemotaxis proteins of enterics and the gliding bacterium Myxococcus xanthus. Mol Microbiol. 1994 Jan;11(1):137–153. doi: 10.1111/j.1365-2958.1994.tb00296.x. [DOI] [PubMed] [Google Scholar]
  10. DiRita V. J., Mekalanos J. J. Periplasmic interaction between two membrane regulatory proteins, ToxR and ToxS, results in signal transduction and transcriptional activation. Cell. 1991 Jan 11;64(1):29–37. doi: 10.1016/0092-8674(91)90206-e. [DOI] [PubMed] [Google Scholar]
  11. DiRita V. J., Parsot C., Jander G., Mekalanos J. J. Regulatory cascade controls virulence in Vibrio cholerae. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5403–5407. doi: 10.1073/pnas.88.12.5403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fasano A., Baudry B., Pumplin D. W., Wasserman S. S., Tall B. D., Ketley J. M., Kaper J. B. Vibrio cholerae produces a second enterotoxin, which affects intestinal tight junctions. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5242–5246. doi: 10.1073/pnas.88.12.5242. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Filip C., Fletcher G., Wulff J. L., Earhart C. F. Solubilization of the cytoplasmic membrane of Escherichia coli by the ionic detergent sodium-lauryl sarcosinate. J Bacteriol. 1973 Sep;115(3):717–722. doi: 10.1128/jb.115.3.717-722.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Freter R., O'Brien P. C., Macsai M. S. Role of chemotaxis in the association of motile bacteria with intestinal mucosa: in vivo studies. Infect Immun. 1981 Oct;34(1):234–240. doi: 10.1128/iai.34.1.234-240.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Freter R., O'Brien P. C. Role of chemotaxis in the association of motile bacteria with intestinal mucosa: fitness and virulence of nonchemotactic Vibrio cholerae mutants in infant mice. Infect Immun. 1981 Oct;34(1):222–233. doi: 10.1128/iai.34.1.222-233.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hazelbauer G. L., Berg H. C., Matsumura P. Bacterial motility and signal transduction. Cell. 1993 Apr 9;73(1):15–22. doi: 10.1016/0092-8674(93)90156-k. [DOI] [PubMed] [Google Scholar]
  17. Hazelbauer G. L., Harayama S. Sensory transduction in bacterial chemotaxis. Int Rev Cytol. 1983;81:33–70. doi: 10.1016/s0074-7696(08)62334-7. [DOI] [PubMed] [Google Scholar]
  18. Henrichsen J. Twitching motility. Annu Rev Microbiol. 1983;37:81–93. doi: 10.1146/annurev.mi.37.100183.000501. [DOI] [PubMed] [Google Scholar]
  19. Herrington D. A., Hall R. H., Losonsky G., Mekalanos J. J., Taylor R. K., Levine M. M. Toxin, toxin-coregulated pili, and the toxR regulon are essential for Vibrio cholerae pathogenesis in humans. J Exp Med. 1988 Oct 1;168(4):1487–1492. doi: 10.1084/jem.168.4.1487. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Higgins D. E., Nazareno E., DiRita V. J. The virulence gene activator ToxT from Vibrio cholerae is a member of the AraC family of transcriptional activators. J Bacteriol. 1992 Nov;174(21):6974–6980. doi: 10.1128/jb.174.21.6974-6980.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kaufman M. R., Seyer J. M., Taylor R. K. Processing of TCP pilin by TcpJ typifies a common step intrinsic to a newly recognized pathway of extracellular protein secretion by gram-negative bacteria. Genes Dev. 1991 Oct;5(10):1834–1846. doi: 10.1101/gad.5.10.1834. [DOI] [PubMed] [Google Scholar]
  22. Kaufman M. R., Shaw C. E., Jones I. D., Taylor R. K. Biogenesis and regulation of the Vibrio cholerae toxin-coregulated pilus: analogies to other virulence factor secretory systems. Gene. 1993 Apr 15;126(1):43–49. doi: 10.1016/0378-1119(93)90588-t. [DOI] [PubMed] [Google Scholar]
  23. Kelley J. T., Parker C. D. Identification and preliminary characterization of Vibrio cholerae outer membrane proteins. J Bacteriol. 1981 Feb;145(2):1018–1024. doi: 10.1128/jb.145.2.1018-1024.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Liu J. D., Parkinson J. S. Genetic evidence for interaction between the CheW and Tsr proteins during chemoreceptor signaling by Escherichia coli. J Bacteriol. 1991 Aug;173(16):4941–4951. doi: 10.1128/jb.173.16.4941-4951.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. McBride M. J., Köhler T., Zusman D. R. Methylation of FrzCD, a methyl-accepting taxis protein of Myxococcus xanthus, is correlated with factors affecting cell behavior. J Bacteriol. 1992 Jul;174(13):4246–4257. doi: 10.1128/jb.174.13.4246-4257.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Mekalanos J. J. Cholera toxin: genetic analysis, regulation, and role in pathogenesis. Curr Top Microbiol Immunol. 1985;118:97–118. doi: 10.1007/978-3-642-70586-1_6. [DOI] [PubMed] [Google Scholar]
  27. Miller J. F., Mekalanos J. J., Falkow S. Coordinate regulation and sensory transduction in the control of bacterial virulence. Science. 1989 Feb 17;243(4893):916–922. doi: 10.1126/science.2537530. [DOI] [PubMed] [Google Scholar]
  28. Miller V. L., Mekalanos J. J. 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. J Bacteriol. 1988 Jun;170(6):2575–2583. doi: 10.1128/jb.170.6.2575-2583.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Miller V. L., Mekalanos J. J. 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. doi: 10.1073/pnas.81.11.3471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Miller V. L., Taylor R. K., Mekalanos J. J. Cholera toxin transcriptional activator toxR is a transmembrane DNA binding protein. Cell. 1987 Jan 30;48(2):271–279. doi: 10.1016/0092-8674(87)90430-2. [DOI] [PubMed] [Google Scholar]
  31. Ogierman M. A., Zabihi S., Mourtzios L., Manning P. A. Genetic organization and sequence of the promoter-distal region of the tcp gene cluster of Vibrio cholerae. Gene. 1993 Apr 15;126(1):51–60. doi: 10.1016/0378-1119(93)90589-u. [DOI] [PubMed] [Google Scholar]
  32. Osborn M. J., Gander J. E., Parisi E., Carson J. Mechanism of assembly of the outer membrane of Salmonella typhimurium. Isolation and characterization of cytoplasmic and outer membrane. J Biol Chem. 1972 Jun 25;247(12):3962–3972. [PubMed] [Google Scholar]
  33. Palva E. T., Hirst T. R., Hardy S. J., Holmgren J., Randall L. Synthesis of a precursor to the B subunit of heat-labile enterotoxin in Escherichia coli. J Bacteriol. 1981 Apr;146(1):325–330. doi: 10.1128/jb.146.1.325-330.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Parkinson J. S., Kofoid E. C. Communication modules in bacterial signaling proteins. Annu Rev Genet. 1992;26:71–112. doi: 10.1146/annurev.ge.26.120192.000443. [DOI] [PubMed] [Google Scholar]
  35. Parkinson J. S. Signal transduction schemes of bacteria. Cell. 1993 Jun 4;73(5):857–871. doi: 10.1016/0092-8674(93)90267-t. [DOI] [PubMed] [Google Scholar]
  36. Parsot C., Mekalanos J. J. Expression of ToxR, the transcriptional activator of the virulence factors in Vibrio cholerae, is modulated by the heat shock response. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9898–9902. doi: 10.1073/pnas.87.24.9898. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Parsot C., Mekalanos J. J. Expression of the Vibrio cholerae gene encoding aldehyde dehydrogenase is under control of ToxR, the cholera toxin transcriptional activator. J Bacteriol. 1991 May;173(9):2842–2851. doi: 10.1128/jb.173.9.2842-2851.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Peek J. A., Taylor R. K. Characterization of a periplasmic thiol:disulfide interchange protein required for the functional maturation of secreted virulence factors of Vibrio cholerae. Proc Natl Acad Sci U S A. 1992 Jul 1;89(13):6210–6214. doi: 10.1073/pnas.89.13.6210. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Peterson K. M., Mekalanos J. J. Characterization of the Vibrio cholerae ToxR regulon: identification of novel genes involved in intestinal colonization. Infect Immun. 1988 Nov;56(11):2822–2829. doi: 10.1128/iai.56.11.2822-2829.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Rice M. S., Dahlquist F. W. Sites of deamidation and methylation in Tsr, a bacterial chemotaxis sensory transducer. J Biol Chem. 1991 May 25;266(15):9746–9753. [PubMed] [Google Scholar]
  41. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Schnaitman C. A. Examination of the protein composition of the cell envelope of Escherichia coli by polyacrylamide gel electrophoresis. J Bacteriol. 1970 Nov;104(2):882–889. doi: 10.1128/jb.104.2.882-889.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Tabor S., Richardson C. C. A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1074–1078. doi: 10.1073/pnas.82.4.1074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Taylor R. K., Manoil C., Mekalanos J. J. Broad-host-range vectors for delivery of TnphoA: use in genetic analysis of secreted virulence determinants of Vibrio cholerae. J Bacteriol. 1989 Apr;171(4):1870–1878. doi: 10.1128/jb.171.4.1870-1878.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Taylor R. K., Miller V. L., Furlong D. B., Mekalanos J. J. 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. doi: 10.1073/pnas.84.9.2833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Trucksis M., Galen J. E., Michalski J., Fasano A., Kaper J. B. Accessory cholera enterotoxin (Ace), the third toxin of a Vibrio cholerae virulence cassette. Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5267–5271. doi: 10.1073/pnas.90.11.5267. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. von Heijne G. A new method for predicting signal sequence cleavage sites. Nucleic Acids Res. 1986 Jun 11;14(11):4683–4690. doi: 10.1093/nar/14.11.4683. [DOI] [PMC free article] [PubMed] [Google Scholar]

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