Skip to main content
Infection and Immunity logoLink to Infection and Immunity
. 1997 May;65(5):1615–1625. doi: 10.1128/iai.65.5.1615-1625.1997

Identification of four new members of the internalin multigene family of Listeria monocytogenes EGD.

S Dramsi 1, P Dehoux 1, M Lebrun 1, P L Goossens 1, P Cossart 1
PMCID: PMC175184  PMID: 9125538

Abstract

Listeria monocytogenes is a bacterial pathogen that is able to invade nonphagocytic cells. Two surface proteins, internalin, the inlA gene product, and InlB, play important roles in the entry into cultured mammalian cells. These proteins also have extensive sequence similarities. Previously, Southern hybridization predicted the existence of an internalin multigene family. Recently, InlC, a secreted protein of 30 kDa homologous to InlA and InlB, was identified. In this work, we identified and characterized four new members of the internalin multigene family, inlC2, inlD, inlE, and inlF which encode proteins of 548, 567, 499, and 821 amino acids respectively. inlC2, inlD, and inlE are contiguous on the chromosome of L. monocytogenes EGD, whereas inlF is located in a different chromosomal region. These four inl gene products display the principal features of internalin, namely, a signal sequence, two regions of repeats (or LRR and B repeats), and a putative cell wall anchor sequence containing the sorting motif LPXTG. The four inl genes were maximally expressed albeit at a low level during early exponential growth in bacterial medium at 37 degrees C. The role of these inl genes in L. monocytogenes invasion was assessed by constructing isogenic chromosomal deletion mutants and testing them for entry into various nonphagocytic cells. Unexpectedly, the inlC2, inlD, inlE, and inlF null mutants were not affected for entry into any of the cell lines tested, raising the possibility that these genes are needed for an aspect of pathogenicity other than invasion. The identity of such an aspect remains to be determined.

Full Text

The Full Text of this article is available as a PDF (882.6 KB).

Selected References

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

  1. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  2. Bhat K. S., Gibbs C. P., Barrera O., Morrison S. G., Jähnig F., Stern A., Kupsch E. M., Meyer T. F., Swanson J. The opacity proteins of Neisseria gonorrhoeae strain MS11 are encoded by a family of 11 complete genes. Mol Microbiol. 1991 Aug;5(8):1889–1901. doi: 10.1111/j.1365-2958.1991.tb00813.x. [DOI] [PubMed] [Google Scholar]
  3. Bhat K. S., Gibbs C. P., Barrera O., Morrison S. G., Jähnig F., Stern A., Kupsch E. M., Meyer T. F., Swanson J. The opacity proteins of Neisseria gonorrhoeae strain MS11 are encoded by a family of 11 complete genes. Mol Microbiol. 1992 Apr;6(8):1073–1076. doi: 10.1111/j.1365-2958.1992.tb02172.x. [DOI] [PubMed] [Google Scholar]
  4. Biswas I., Gruss A., Ehrlich S. D., Maguin E. High-efficiency gene inactivation and replacement system for gram-positive bacteria. J Bacteriol. 1993 Jun;175(11):3628–3635. doi: 10.1128/jb.175.11.3628-3635.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Boland A., Sory M. P., Iriarte M., Kerbourch C., Wattiau P., Cornelis G. R. Status of YopM and YopN in the Yersinia Yop virulon: YopM of Y.enterocolitica is internalized inside the cytosol of PU5-1.8 macrophages by the YopB, D, N delivery apparatus. EMBO J. 1996 Oct 1;15(19):5191–5201. [PMC free article] [PubMed] [Google Scholar]
  6. 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]
  7. Buysse J. M., Stover C. K., Oaks E. V., Venkatesan M., Kopecko D. J. Molecular cloning of invasion plasmid antigen (ipa) genes from Shigella flexneri: analysis of ipa gene products and genetic mapping. J Bacteriol. 1987 Jun;169(6):2561–2569. doi: 10.1128/jb.169.6.2561-2569.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Camilli A., Tilney L. G., Portnoy D. A. Dual roles of plcA in Listeria monocytogenes pathogenesis. Mol Microbiol. 1993 Apr;8(1):143–157. doi: 10.1111/j.1365-2958.1993.tb01211.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chen J., Brosch R., Luchansky J. B. Isolation and characterization of Listeria monocytogenes-specific nucleotide sequences. Appl Environ Microbiol. 1993 Dec;59(12):4367–4370. doi: 10.1128/aem.59.12.4367-4370.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Domann E., Zechel S., Lingnau A., Hain T., Darji A., Nichterlein T., Wehland J., Chakraborty T. Identification and characterization of a novel PrfA-regulated gene in Listeria monocytogenes whose product, IrpA, is highly homologous to internalin proteins, which contain leucine-rich repeats. Infect Immun. 1997 Jan;65(1):101–109. doi: 10.1128/iai.65.1.101-109.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Dramsi S., Biswas I., Maguin E., Braun L., Mastroeni P., Cossart P. Entry of Listeria monocytogenes into hepatocytes requires expression of inIB, a surface protein of the internalin multigene family. Mol Microbiol. 1995 Apr;16(2):251–261. doi: 10.1111/j.1365-2958.1995.tb02297.x. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Dramsi S., Lebrun M., Cossart P. Molecular and genetic determinants involved in invasion of mammalian cells by Listeria monocytogenes. Curr Top Microbiol Immunol. 1996;209:61–77. doi: 10.1007/978-3-642-85216-9_4. [DOI] [PubMed] [Google Scholar]
  15. Engelbrecht F., Chun S. K., Ochs C., Hess J., Lottspeich F., Goebel W., Sokolovic Z. A new PrfA-regulated gene of Listeria monocytogenes encoding a small, secreted protein which belongs to the family of internalins. Mol Microbiol. 1996 Aug;21(4):823–837. doi: 10.1046/j.1365-2958.1996.541414.x. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Gaillard J. L., Jaubert F., Berche P. The inlAB locus mediates the entry of Listeria monocytogenes into hepatocytes in vivo. J Exp Med. 1996 Feb 1;183(2):359–369. doi: 10.1084/jem.183.2.359. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gregory S. H., Sagnimeni A. J., Wing E. J. Expression of the inlAB operon by Listeria monocytogenes is not required for entry into hepatic cells in vivo. Infect Immun. 1996 Oct;64(10):3983–3986. doi: 10.1128/iai.64.10.3983-3986.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hartman A. B., Venkatesan M., Oaks E. V., Buysse J. M. Sequence and molecular characterization of a multicopy invasion plasmid antigen gene, ipaH, of Shigella flexneri. J Bacteriol. 1990 Apr;172(4):1905–1915. doi: 10.1128/jb.172.4.1905-1915.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ireton K., Payrastre B., Chap H., Ogawa W., Sakaue H., Kasuga M., Cossart P. A role for phosphoinositide 3-kinase in bacterial invasion. Science. 1996 Nov 1;274(5288):780–782. doi: 10.1126/science.274.5288.780. [DOI] [PubMed] [Google Scholar]
  21. Kobe B., Deisenhofer J. Proteins with leucine-rich repeats. Curr Opin Struct Biol. 1995 Jun;5(3):409–416. doi: 10.1016/0959-440x(95)80105-7. [DOI] [PubMed] [Google Scholar]
  22. Kocks C., Gouin E., Tabouret M., Berche P., Ohayon H., Cossart P. L. monocytogenes-induced actin assembly requires the actA gene product, a surface protein. Cell. 1992 Feb 7;68(3):521–531. doi: 10.1016/0092-8674(92)90188-i. [DOI] [PubMed] [Google Scholar]
  23. Kupsch E. M., Knepper B., Kuroki T., Heuer I., Meyer T. F. Variable opacity (Opa) outer membrane proteins account for the cell tropisms displayed by Neisseria gonorrhoeae for human leukocytes and epithelial cells. EMBO J. 1993 Feb;12(2):641–650. doi: 10.1002/j.1460-2075.1993.tb05697.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. 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]
  26. Leung K. Y., Reisner B. S., Straley S. C. YopM inhibits platelet aggregation and is necessary for virulence of Yersinia pestis in mice. Infect Immun. 1990 Oct;58(10):3262–3271. doi: 10.1128/iai.58.10.3262-3271.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Leung K. Y., Straley S. C. The yopM gene of Yersinia pestis encodes a released protein having homology with the human platelet surface protein GPIb alpha. J Bacteriol. 1989 Sep;171(9):4623–4632. doi: 10.1128/jb.171.9.4623-4632.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lingnau A., Chakraborty T., Niebuhr K., Domann E., Wehland J. Identification and purification of novel internalin-related proteins in Listeria monocytogenes and Listeria ivanovii. Infect Immun. 1996 Mar;64(3):1002–1006. doi: 10.1128/iai.64.3.1002-1006.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lingnau A., Domann E., Hudel M., Bock M., Nichterlein T., Wehland J., Chakraborty T. Expression of the Listeria monocytogenes EGD inlA and inlB genes, whose products mediate bacterial entry into tissue culture cell lines, by PrfA-dependent and -independent mechanisms. Infect Immun. 1995 Oct;63(10):3896–3903. doi: 10.1128/iai.63.10.3896-3903.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. MACKANESS G. B. Cellular resistance to infection. J Exp Med. 1962 Sep 1;116:381–406. doi: 10.1084/jem.116.3.381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Maguin E., Duwat P., Hege T., Ehrlich D., Gruss A. New thermosensitive plasmid for gram-positive bacteria. J Bacteriol. 1992 Sep;174(17):5633–5638. doi: 10.1128/jb.174.17.5633-5638.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Marck C. 'DNA Strider': a 'C' program for the fast analysis of DNA and protein sequences on the Apple Macintosh family of computers. Nucleic Acids Res. 1988 Mar 11;16(5):1829–1836. doi: 10.1093/nar/16.5.1829. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mengaud J., Geoffroy C., Cossart P. Identification of a new operon involved in Listeria monocytogenes virulence: its first gene encodes a protein homologous to bacterial metalloproteases. Infect Immun. 1991 Mar;59(3):1043–1049. doi: 10.1128/iai.59.3.1043-1049.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Mengaud J., Ohayon H., Gounon P., Mege R-M, Cossart P. E-cadherin is the receptor for internalin, a surface protein required for entry of L. monocytogenes into epithelial cells. Cell. 1996 Mar 22;84(6):923–932. doi: 10.1016/s0092-8674(00)81070-3. [DOI] [PubMed] [Google Scholar]
  35. Michel E., Cossart P. Physical map of the Listeria monocytogenes chromosome. J Bacteriol. 1992 Nov;174(22):7098–7103. doi: 10.1128/jb.174.22.7098-7103.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Navarre W. W., Schneewind O. Proteolytic cleavage and cell wall anchoring at the LPXTG motif of surface proteins in gram-positive bacteria. Mol Microbiol. 1994 Oct;14(1):115–121. doi: 10.1111/j.1365-2958.1994.tb01271.x. [DOI] [PubMed] [Google Scholar]
  37. Ohkura H., Yanagida M. S. pombe gene sds22+ essential for a midmitotic transition encodes a leucine-rich repeat protein that positively modulates protein phosphatase-1. Cell. 1991 Jan 11;64(1):149–157. doi: 10.1016/0092-8674(91)90216-l. [DOI] [PubMed] [Google Scholar]
  38. Renouf S., Beullens M., Wera S., Van Eynde A., Sikela J., Stalmans W., Bollen M. Molecular cloning of a human polypeptide related to yeast sds22, a regulator of protein phosphatase-1. FEBS Lett. 1995 Nov 13;375(1-2):75–78. doi: 10.1016/0014-5793(95)01180-m. [DOI] [PubMed] [Google Scholar]
  39. Schneewind O., Fowler A., Faull K. F. Structure of the cell wall anchor of surface proteins in Staphylococcus aureus. Science. 1995 Apr 7;268(5207):103–106. doi: 10.1126/science.7701329. [DOI] [PubMed] [Google Scholar]
  40. Schneewind O., Mihaylova-Petkov D., Model P. Cell wall sorting signals in surface proteins of gram-positive bacteria. EMBO J. 1993 Dec;12(12):4803–4811. doi: 10.1002/j.1460-2075.1993.tb06169.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sheehan B., Kocks C., Dramsi S., Gouin E., Klarsfeld A. D., Mengaud J., Cossart P. Molecular and genetic determinants of the Listeria monocytogenes infectious process. Curr Top Microbiol Immunol. 1994;192:187–216. doi: 10.1007/978-3-642-78624-2_9. [DOI] [PubMed] [Google Scholar]
  42. Smith K., Youngman P. Use of a new integrational vector to investigate compartment-specific expression of the Bacillus subtilis spoIIM gene. Biochimie. 1992 Jul-Aug;74(7-8):705–711. doi: 10.1016/0300-9084(92)90143-3. [DOI] [PubMed] [Google Scholar]
  43. Thompson J. D., Higgins D. G., Gibson T. J. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 1994 Nov 11;22(22):4673–4680. doi: 10.1093/nar/22.22.4673. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. 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