Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1995 May;63(5):2047–2053. doi: 10.1128/iai.63.5.2047-2053.1995

Listeria monocytogenes p60 supports host cell invasion by and in vivo survival of attenuated Salmonella typhimurium.

J Hess 1, I Gentschev 1, G Szalay 1, C Ladel 1, A Bubert 1, W Goebel 1, S H Kaufmann 1
PMCID: PMC173263  PMID: 7729919

Abstract

The extracellular protein p60 is a major virulence factor of the intracellular bacterium Listeria monocytogenes. Its roles in pathogen survival in vivo and host cell invasion in vitro were studied. To this end, Salmonella typhimurium SL7207 was used as carrier for secreted p60-HlyA fusion protein by Escherichia coli HlyB and HlyD transport proteins. C57BL/6 mice infected intravenously with this strain suffered from increased bacterial numbers in livers and spleens compared with the p60-nonexpressing control strain, but only transiently. In vitro experiments showed that p60 promotes invasion of recombinant S. typhimurium SL7207 p60 into hepatocytes and resting macrophages independent from complement. Moreover, the uptake of wild-type L. monocytogenes EGD and L. monocytogenes BUG 8, an internalin-deficient strain, into hepatocytes was partially blocked by anti-p60 antibodies. The impaired invasion of dissociated bacterial chains of L. monocytogenes RIII, a p60 expression mutant, into hepatocytes and macrophages was partially restored by addition of p60- or p60-HlyA-enriched bacterial supernatants. These data suggest that the L. monocytogenes surface-associated proteins, p60 and internalin, act in concert to achieve optimal uptake into nonprofessional phagocytes and macrophages. Together, these experiments reveal a substantial impact of p60 on cell invasion and virulence and thus emphasize the importance of the intracellular habitat for survival of L. monocytogenes in the host.

Full Text

The Full Text of this article is available as a PDF (247.4 KB).

Selected References

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

  1. Alvarez-Dominguez C., Carrasco-Marin E., Leyva-Cobian F. Role of complement component C1q in phagocytosis of Listeria monocytogenes by murine macrophage-like cell lines. Infect Immun. 1993 Sep;61(9):3664–3672. doi: 10.1128/iai.61.9.3664-3672.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bliska J. B., Galán J. E., Falkow S. Signal transduction in the mammalian cell during bacterial attachment and entry. Cell. 1993 Jun 4;73(5):903–920. doi: 10.1016/0092-8674(93)90270-z. [DOI] [PubMed] [Google Scholar]
  3. Bubert A., Kuhn M., Goebel W., Köhler S. Structural and functional properties of the p60 proteins from different Listeria species. J Bacteriol. 1992 Dec;174(24):8166–8171. doi: 10.1128/jb.174.24.8166-8171.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chatfield S., Roberts M., Londono P., Cropley I., Douce G., Dougan G. The development of oral vaccines based on live attenuated Salmonella strains. FEMS Immunol Med Microbiol. 1993 Jun;7(1):1–7. doi: 10.1111/j.1574-695X.1993.tb00374.x. [DOI] [PubMed] [Google Scholar]
  5. Conlan J. W., North R. J. Early pathogenesis of infection in the liver with the facultative intracellular bacteria Listeria monocytogenes, Francisella tularensis, and Salmonella typhimurium involves lysis of infected hepatocytes by leukocytes. Infect Immun. 1992 Dec;60(12):5164–5171. doi: 10.1128/iai.60.12.5164-5171.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dramsi S., Kocks C., Forestier C., Cossart P. Internalin-mediated invasion of epithelial cells by Listeria monocytogenes is regulated by the bacterial growth state, temperature and the pleiotropic activator prfA. Mol Microbiol. 1993 Sep;9(5):931–941. doi: 10.1111/j.1365-2958.1993.tb01223.x. [DOI] [PubMed] [Google Scholar]
  7. Drevets D. A., Campbell P. A. Roles of complement and complement receptor type 3 in phagocytosis of Listeria monocytogenes by inflammatory mouse peritoneal macrophages. Infect Immun. 1991 Aug;59(8):2645–2652. doi: 10.1128/iai.59.8.2645-2652.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Drevets D. A., Canono B. P., Leenen P. J., Campbell P. A. Gentamicin kills intracellular Listeria monocytogenes. Infect Immun. 1994 Jun;62(6):2222–2228. doi: 10.1128/iai.62.6.2222-2228.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Finlay B. B., Falkow S. Salmonella as an intracellular parasite. Mol Microbiol. 1989 Dec;3(12):1833–1841. doi: 10.1111/j.1365-2958.1989.tb00170.x. [DOI] [PubMed] [Google Scholar]
  10. Flesch I., Früh A., Ferber E. Functional comparison of bone marrow-derived macrophages obtained by cultivation in serum-free or serum-supplemented medium. Immunobiology. 1986 Oct;173(1):72–81. doi: 10.1016/S0171-2985(86)80091-2. [DOI] [PubMed] [Google Scholar]
  11. Flesch I., Kaufmann S. H. Mycobacterial growth inhibition by interferon-gamma-activated bone marrow macrophages and differential susceptibility among strains of Mycobacterium tuberculosis. J Immunol. 1987 Jun 15;138(12):4408–4413. [PubMed] [Google Scholar]
  12. Fürst P., Mösch H. U., Solioz M. A protein of unusual composition from Enterococcus faecium. Nucleic Acids Res. 1989 Aug 25;17(16):6724–6724. doi: 10.1093/nar/17.16.6724. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gaillard J. L., Berche P., Frehel C., Gouin E., Cossart P. Entry of L. monocytogenes into cells is mediated by internalin, a repeat protein reminiscent of surface antigens from gram-positive cocci. Cell. 1991 Jun 28;65(7):1127–1141. doi: 10.1016/0092-8674(91)90009-n. [DOI] [PubMed] [Google Scholar]
  14. Galán J. E., Pace J., Hayman M. J. Involvement of the epidermal growth factor receptor in the invasion of cultured mammalian cells by Salmonella typhimurium. Nature. 1992 Jun 18;357(6379):588–589. doi: 10.1038/357588a0. [DOI] [PubMed] [Google Scholar]
  15. Gentschev I., Hess J., Goebel W. Change in the cellular localization of alkaline phosphatase by alteration of its carboxy-terminal sequence. Mol Gen Genet. 1990 Jul;222(2-3):211–216. doi: 10.1007/BF00633820. [DOI] [PubMed] [Google Scholar]
  16. Gentschev I., Sokolovic Z., Köhler S., Krohne G. F., Hof H., Wagner J., Goebel W. Identification of p60 antibodies in human sera and presentation of this listerial antigen on the surface of attenuated salmonellae by the HlyB-HlyD secretion system. Infect Immun. 1992 Dec;60(12):5091–5098. doi: 10.1128/iai.60.12.5091-5098.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hess J., Gentschev I., Goebel W., Jarchau T. Analysis of the haemolysin secretion system by PhoA-HlyA fusion proteins. Mol Gen Genet. 1990 Nov;224(2):201–208. doi: 10.1007/BF00271553. [DOI] [PubMed] [Google Scholar]
  18. Hess J., Kaufmann S. H. Vaccination strategies against intracellular microbes. FEMS Immunol Med Microbiol. 1993 Aug;7(2):95–103. doi: 10.1111/j.1574-695X.1993.tb00387.x. [DOI] [PubMed] [Google Scholar]
  19. Hoiseth S. K., Stocker B. A. Aromatic-dependent Salmonella typhimurium are non-virulent and effective as live vaccines. Nature. 1981 May 21;291(5812):238–239. doi: 10.1038/291238a0. [DOI] [PubMed] [Google Scholar]
  20. Hormaeche C. E. Live attenuated Salmonella vaccines and their potential as oral combined vaccines carrying heterologous antigens. J Immunol Methods. 1991 Aug 28;142(1):113–120. doi: 10.1016/0022-1759(91)90298-t. [DOI] [PubMed] [Google Scholar]
  21. Horstmann R. D., Sievertsen H. J., Knobloch J., Fischetti V. A. Antiphagocytic activity of streptococcal M protein: selective binding of complement control protein factor H. Proc Natl Acad Sci U S A. 1988 Mar;85(5):1657–1661. doi: 10.1073/pnas.85.5.1657. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jones B. D., Ghori N., Falkow S. Salmonella typhimurium initiates murine infection by penetrating and destroying the specialized epithelial M cells of the Peyer's patches. J Exp Med. 1994 Jul 1;180(1):15–23. doi: 10.1084/jem.180.1.15. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kaufmann S. H. Immunity to intracellular bacteria. Annu Rev Immunol. 1993;11:129–163. doi: 10.1146/annurev.iy.11.040193.001021. [DOI] [PubMed] [Google Scholar]
  24. Kohbata S., Yokoyama H., Yabuuchi E. Cytopathogenic effect of Salmonella typhi GIFU 10007 on M cells of murine ileal Peyer's patches in ligated ileal loops: an ultrastructural study. Microbiol Immunol. 1986;30(12):1225–1237. doi: 10.1111/j.1348-0421.1986.tb03055.x. [DOI] [PubMed] [Google Scholar]
  25. Kuhn M., Goebel W. Identification of an extracellular protein of Listeria monocytogenes possibly involved in intracellular uptake by mammalian cells. Infect Immun. 1989 Jan;57(1):55–61. doi: 10.1128/iai.57.1.55-61.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kuhn M., Kathariou S., Goebel W. Hemolysin supports survival but not entry of the intracellular bacterium Listeria monocytogenes. Infect Immun. 1988 Jan;56(1):79–82. doi: 10.1128/iai.56.1.79-82.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Köhler S., Bubert A., Vogel M., Goebel W. Expression of the iap gene coding for protein p60 of Listeria monocytogenes is controlled on the posttranscriptional level. J Bacteriol. 1991 Aug;173(15):4668–4674. doi: 10.1128/jb.173.15.4668-4674.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Köhler S., Leimeister-Wächter M., Chakraborty T., Lottspeich F., Goebel W. The gene coding for protein p60 of Listeria monocytogenes and its use as a specific probe for Listeria monocytogenes. Infect Immun. 1990 Jun;58(6):1943–1950. doi: 10.1128/iai.58.6.1943-1950.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. 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]
  30. Marco A. J., Prats N., Ramos J. A., Briones V., Blanco M., Dominguez L., Domingo M. A microbiological, histopathological and immunohistological study of the intragastric inoculation of Listeria monocytogenes in mice. J Comp Pathol. 1992 Jul;107(1):1–9. doi: 10.1016/0021-9975(92)90090-h. [DOI] [PubMed] [Google Scholar]
  31. Portnoy D. A., Chakraborty T., Goebel W., Cossart P. Molecular determinants of Listeria monocytogenes pathogenesis. Infect Immun. 1992 Apr;60(4):1263–1267. doi: 10.1128/iai.60.4.1263-1267.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Potel J., Schulze-Lammers J. Listeria monocytogenes-vaccine: production and control. Zentralbl Bakteriol Mikrobiol Hyg A. 1985 May;259(3):331–340. doi: 10.1016/s0176-6724(85)80035-3. [DOI] [PubMed] [Google Scholar]
  33. Ruhland G. J., Hellwig M., Wanner G., Fiedler F. Cell-surface location of Listeria-specific protein p60--detection of Listeria cells by indirect immunofluorescence. J Gen Microbiol. 1993 Mar;139(3):609–616. doi: 10.1099/00221287-139-3-609. [DOI] [PubMed] [Google Scholar]
  34. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Vogel M., Hess J., Then I., Juarez A., Goebel W. Characterization of a sequence (hlyR) which enhances synthesis and secretion of hemolysin in Escherichia coli. Mol Gen Genet. 1988 Apr;212(1):76–84. doi: 10.1007/BF00322447. [DOI] [PubMed] [Google Scholar]
  36. Wagner W., Vogel M., Goebel W. Transport of hemolysin across the outer membrane of Escherichia coli requires two functions. J Bacteriol. 1983 Apr;154(1):200–210. doi: 10.1128/jb.154.1.200-210.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Wood S., Maroushek N., Czuprynski C. J. Multiplication of Listeria monocytogenes in a murine hepatocyte cell line. Infect Immun. 1993 Jul;61(7):3068–3072. doi: 10.1128/iai.61.7.3068-3072.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Wuenscher M. D., Köhler S., Bubert A., Gerike U., Goebel W. The iap gene of Listeria monocytogenes is essential for cell viability, and its gene product, p60, has bacteriolytic activity. J Bacteriol. 1993 Jun;175(11):3491–3501. doi: 10.1128/jb.175.11.3491-3501.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES