Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1993 Jul;175(14):4475–4484. doi: 10.1128/jb.175.14.4475-4484.1993

A PhoP-repressed gene promotes Salmonella typhimurium invasion of epithelial cells.

I Behlau 1, S I Miller 1
PMCID: PMC204888  PMID: 8392513

Abstract

The Salmonella typhimurium transcriptional regulators, PhoP/PhoQ, induce phoP-activated gene (pag) expression to promote virulence and intracellular survival within macrophages. This response to the macrophage intracellular environment is simulated by phoP/phoQ constitutive mutations (phenotype PhoPc) that increase the expression of pag genes and repress the synthesis of approximately 20 proteins encoded by phoP-repressed genes (prg genes) (S. I. Miller and J. J. Mekalanos, J. Bacteriol. 172:2485-2490, 1990). PhoPc bacteria are attenuated for mouse virulence, suggesting that prg genes are virulence genes. We now report the identification of five unlinked prg loci by use of the transposon TnphoA. In general, medium conditions (i.e., starvation) that activate pag expression repress prg expression. However, variable effects on the PhoP regulon were observed when bacteria were grown under different oxygen tensions (pag and prg genes) or exposed to low pH (prg genes), suggesting heterogenous control of the regulon. One prg locus, prgH, was demonstrated to contribute to mouse virulence by both the oral and the intraperitoneal routes. prgH was located at 59 min on the Salmonella chromosome, a region where other genes essential to invasion of epithelial cells are clustered. The prgH locus was highly linked to one invasion locus, hil (C.A. Lee, B.D. Jones, and S. Falkow, Proc. Natl. Acad. Sci. USA 89:1847-1851, 1992), although transcription of prgH was opposite that of the Tn5B50-encoded promoters that result in a hyperinvasive or hil phenotype. Both PrgH and PhoPc mutant S. typhimurium were found to be defective in induction of endocytosis by Madin-Darby canine kidney (MDCK) epithelial cells. The invasion defect of PrgH but not that of PhoPc mutant bacteria was complemented by plasmids containing prgH (hil) DNA. Therefore, two virulence properties of Salmonella species, induction of endocytosis by epithelial cells and survival within macrophages, are oppositely modulated by the PhoP/PhoQ virulence regulators.

Full text

PDF
4475

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Akerley B. J., Monack D. M., Falkow S., Miller J. F. The bvgAS locus negatively controls motility and synthesis of flagella in Bordetella bronchiseptica. J Bacteriol. 1992 Feb;174(3):980–990. doi: 10.1128/jb.174.3.980-990.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aliabadi Z., Warren F., Mya S., Foster J. W. Oxygen-regulated stimulons of Salmonella typhimurium identified by Mu d(Ap lac) operon fusions. J Bacteriol. 1986 Mar;165(3):780–786. doi: 10.1128/jb.165.3.780-786.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Alpuche Aranda C. M., Swanson J. A., Loomis W. P., Miller S. I. Salmonella typhimurium activates virulence gene transcription within acidified macrophage phagosomes. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10079–10083. doi: 10.1073/pnas.89.21.10079. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beattie D. T., Shahin R., Mekalanos J. J. A vir-repressed gene of Bordetella pertussis is required for virulence. Infect Immun. 1992 Feb;60(2):571–577. doi: 10.1128/iai.60.2.571-577.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Buchmeier N. A., Heffron F. Intracellular survival of wild-type Salmonella typhimurium and macrophage-sensitive mutants in diverse populations of macrophages. Infect Immun. 1989 Jan;57(1):1–7. doi: 10.1128/iai.57.1.1-7.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Carter P. B., Collins F. M. The route of enteric infection in normal mice. J Exp Med. 1974 May 1;139(5):1189–1203. doi: 10.1084/jem.139.5.1189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  8. Fields P. I., Groisman E. A., Heffron F. A Salmonella locus that controls resistance to microbicidal proteins from phagocytic cells. Science. 1989 Feb 24;243(4894 Pt 1):1059–1062. doi: 10.1126/science.2646710. [DOI] [PubMed] [Google Scholar]
  9. Foster J. W., Hall H. K. Adaptive acidification tolerance response of Salmonella typhimurium. J Bacteriol. 1990 Feb;172(2):771–778. doi: 10.1128/jb.172.2.771-778.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Galán J. E., Curtiss R., 3rd Cloning and molecular characterization of genes whose products allow Salmonella typhimurium to penetrate tissue culture cells. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6383–6387. doi: 10.1073/pnas.86.16.6383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Galán J. E., Timoney J. F. Cloning and expression in Escherichia coli of a protective antigen of Erysipelothrix rhusiopathiae. Infect Immun. 1990 Sep;58(9):3116–3121. doi: 10.1128/iai.58.9.3116-3121.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Groisman E. A., Chiao E., Lipps C. J., Heffron F. Salmonella typhimurium phoP virulence gene is a transcriptional regulator. Proc Natl Acad Sci U S A. 1989 Sep;86(18):7077–7081. doi: 10.1073/pnas.86.18.7077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kier L. D., Weppelman R., Ames B. N. Regulation of two phosphatases and a cyclic phosphodiesterase of Salmonella typhimurium. J Bacteriol. 1977 Apr;130(1):420–428. doi: 10.1128/jb.130.1.420-428.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kier L. D., Weppelman R., Ames B. N. Resolution and purification of three periplasmic phosphatases of Salmonella typhimurium. J Bacteriol. 1977 Apr;130(1):399–410. doi: 10.1128/jb.130.1.399-410.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Knapp S., Mekalanos J. J. Two trans-acting regulatory genes (vir and mod) control antigenic modulation in Bordetella pertussis. J Bacteriol. 1988 Nov;170(11):5059–5066. doi: 10.1128/jb.170.11.5059-5066.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kukral A. M., Strauch K. L., Maurer R. A., Miller C. G. Genetic analysis in Salmonella typhimurium with a small collection of randomly spaced insertions of transposon Tn10 delta 16 delta 17. J Bacteriol. 1987 May;169(5):1787–1793. doi: 10.1128/jb.169.5.1787-1793.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Lee C. A., Falkow S. The ability of Salmonella to enter mammalian cells is affected by bacterial growth state. Proc Natl Acad Sci U S A. 1990 Jun;87(11):4304–4308. doi: 10.1073/pnas.87.11.4304. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lee C. A., Jones B. D., Falkow S. Identification of a Salmonella typhimurium invasion locus by selection for hyperinvasive mutants. Proc Natl Acad Sci U S A. 1992 Mar 1;89(5):1847–1851. doi: 10.1073/pnas.89.5.1847. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Liu S. L., Sanderson K. E. A physical map of the Salmonella typhimurium LT2 genome made by using XbaI analysis. J Bacteriol. 1992 Mar;174(5):1662–1672. doi: 10.1128/jb.174.5.1662-1672.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. MacLachlan P. R., Sanderson K. E. Transformation of Salmonella typhimurium with plasmid DNA: differences between rough and smooth strains. J Bacteriol. 1985 Jan;161(1):442–445. doi: 10.1128/jb.161.1.442-445.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mekalanos J. J. Duplication and amplification of toxin genes in Vibrio cholerae. Cell. 1983 Nov;35(1):253–263. doi: 10.1016/0092-8674(83)90228-3. [DOI] [PubMed] [Google Scholar]
  23. Mekalanos J. J. Environmental signals controlling expression of virulence determinants in bacteria. J Bacteriol. 1992 Jan;174(1):1–7. doi: 10.1128/jb.174.1.1-7.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Michaelis S., Inouye H., Oliver D., Beckwith J. Mutations that alter the signal sequence of alkaline phosphatase in Escherichia coli. J Bacteriol. 1983 Apr;154(1):366–374. doi: 10.1128/jb.154.1.366-374.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Miller J. F., Johnson S. A., Black W. J., Beattie D. T., Mekalanos J. J., Falkow S. Constitutive sensory transduction mutations in the Bordetella pertussis bvgS gene. J Bacteriol. 1992 Feb;174(3):970–979. doi: 10.1128/jb.174.3.970-979.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. 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]
  27. Miller S. I., Kukral A. M., Mekalanos J. J. A two-component regulatory system (phoP phoQ) controls Salmonella typhimurium virulence. Proc Natl Acad Sci U S A. 1989 Jul;86(13):5054–5058. doi: 10.1073/pnas.86.13.5054. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Miller S. I., Mekalanos J. J. Constitutive expression of the phoP regulon attenuates Salmonella virulence and survival within macrophages. J Bacteriol. 1990 May;172(5):2485–2490. doi: 10.1128/jb.172.5.2485-2490.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Miller S. I. PhoP/PhoQ: macrophage-specific modulators of Salmonella virulence? Mol Microbiol. 1991 Sep;5(9):2073–2078. doi: 10.1111/j.1365-2958.1991.tb02135.x. [DOI] [PubMed] [Google Scholar]
  30. Miller S. I., Pulkkinen W. S., Selsted M. E., Mekalanos J. J. Characterization of defensin resistance phenotypes associated with mutations in the phoP virulence regulon of Salmonella typhimurium. Infect Immun. 1990 Nov;58(11):3706–3710. doi: 10.1128/iai.58.11.3706-3710.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Miller V. L., Beer K. B., Loomis W. P., Olson J. A., Miller S. I. An unusual pagC::TnphoA mutation leads to an invasion- and virulence-defective phenotype in Salmonellae. Infect Immun. 1992 Sep;60(9):3763–3770. doi: 10.1128/iai.60.9.3763-3770.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. 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]
  33. Pulkkinen W. S., Miller S. I. A Salmonella typhimurium virulence protein is similar to a Yersinia enterocolitica invasion protein and a bacteriophage lambda outer membrane protein. J Bacteriol. 1991 Jan;173(1):86–93. doi: 10.1128/jb.173.1.86-93.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Ralph P., Prichard J., Cohn M. Reticulum cell sarcoma: an effector cell in antibody-dependent cell-mediated immunity. J Immunol. 1975 Feb;114(2 Pt 2):898–905. [PubMed] [Google Scholar]
  35. Sanderson K. E., Roth J. R. Linkage map of Salmonella typhimurium, edition VII. Microbiol Rev. 1988 Dec;52(4):485–532. doi: 10.1128/mr.52.4.485-532.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Selsted M. E., Brown D. M., DeLange R. J., Harwig S. S., Lehrer R. I. Primary structures of six antimicrobial peptides of rabbit peritoneal neutrophils. J Biol Chem. 1985 Apr 25;260(8):4579–4584. [PubMed] [Google Scholar]
  37. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  38. Stone B. J., Garcia C. M., Badger J. L., Hassett T., Smith R. I., Miller V. L. Identification of novel loci affecting entry of Salmonella enteritidis into eukaryotic cells. J Bacteriol. 1992 Jun;174(12):3945–3952. doi: 10.1128/jb.174.12.3945-3952.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Straley S. C., Harmon P. A. Yersinia pestis grows within phagolysosomes in mouse peritoneal macrophages. Infect Immun. 1984 Sep;45(3):655–659. doi: 10.1128/iai.45.3.655-659.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Swanson J. A. Phorbol esters stimulate macropinocytosis and solute flow through macrophages. J Cell Sci. 1989 Sep;94(Pt 1):135–142. doi: 10.1242/jcs.94.1.135. [DOI] [PubMed] [Google Scholar]
  41. Taub M., Saier M. H., Jr An established but differentiated kidney epithelial cell line (MDCK). Methods Enzymol. 1979;58:552–560. doi: 10.1016/s0076-6879(79)58170-1. [DOI] [PubMed] [Google Scholar]
  42. Tautz D., Renz M. An optimized freeze-squeeze method for the recovery of DNA fragments from agarose gels. Anal Biochem. 1983 Jul 1;132(1):14–19. doi: 10.1016/0003-2697(83)90419-0. [DOI] [PubMed] [Google Scholar]
  43. 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]
  44. Vaudaux P., Waldvogel F. A. Gentamicin antibacterial activity in the presence of human polymorphonuclear leukocytes. Antimicrob Agents Chemother. 1979 Dec;16(6):743–749. doi: 10.1128/aac.16.6.743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Wang R. F., Kushner S. R. Construction of versatile low-copy-number vectors for cloning, sequencing and gene expression in Escherichia coli. Gene. 1991 Apr;100:195–199. [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES