Chapter 15 : Mechanisms of Entry into Mammalian Cells

MyBook is a cheap paperback edition of the original book and will be sold at uniform, low price.

Preview this chapter:
Zoom in

Mechanisms of Entry into Mammalian Cells, Page 1 of 2

| /docserver/preview/fulltext/10.1128/9781555818340/9781555810825_Chap15-1.gif /docserver/preview/fulltext/10.1128/9781555818340/9781555810825_Chap15-2.gif


To begin to examine the process whereby the enteric pathogenic yersiniae enter the host, in 1981 Devenish and Schiemann developed the now famous invasion assay, an in vitro model for the entry of into mammalian cells. The introduced sequences are expressed, and Ail protein can be detected on the bacterial cell surface. One explanation for the absence of Ail-mediated entry in these nonpathogenic strains containing the gene is that other factors, in addition to Ail, that affect yersinia entry are required. Two different virulence factors of have been identified as being homologous to Ail. All three genes , , described have been identified as being able to promote bacterial entry into mammalian cells in vitro. isolates do not enter mammalian cells in vitro, however. However, it is known that when the gene from is introduced into , the resulting strain is able to enter mammalian cells. Both Ail and YadA have been shown to mediate other phenotypes in spp., including resistance to serum killing. This apparent redundancy in function may yet turn out to be an in vitro artifact or may instead have evolved to allow the bacterium to survive in the myriad of environments it encounters during the normal course of infection.

Citation: Pierson D. 1994. Mechanisms of Entry into Mammalian Cells, p 235-247. In Miller V, Kaper J, Portnoy D, Isberg R (ed), Molecular Genetics of Bacterial Pathogenesis. ASM Press, Washington, DC. doi: 10.1128/9781555818340.ch15
Highlighted Text: Show | Hide
Loading full text...

Full text loading...


1. Balligand, G.,, Y. Laroche,, and G. Cornells. 1985. Genetic analysis of virulence plasmid from a serogroup 9 Yersinia enterocolitica strain: role of outer membrane protein PI in resistance to human serum and autoagglutination. Infect. Immun. 48: 782 786.
2. Barondess, J. J.,, and J. Beckwith. 1990. A bacterial virulence determinant encoded by lysogenic coliphage λ. Nature (London) 346: 871 874.
3. Beer, K. B.,, and V. L. Miller. 1992. Amino acid substitutions in naturally occurring variants of Ail result in altered invasion activity. J. Bacteriol. 174: 1360 1369.
4. Bercovier, H.,, and H. H. Mollaret,. 1984. Genus XIV. Yersinia. In N. R. Krieg, and J. G. Holt (ed.), Bergey's Manual of Systematic Bacteriology. The Williams & Wilkins Co., Baltimore.
5. Bliska, J.,, and S. Falkow. 1992. Bacterial resistance to complement killing mediated by the Ail protein of Yersinia enterocolitica. Proc. Natl. Acad. Sci. USA 89: 3561 3565.
6. Bliska, J. B.,, M. C. Copass,, and S. Falkow. 1993. The Yersinia pseudotuberculosis adhesin yadA mediates intimate bacterial attachment to and entry into HEp-2 cells. Infect. Immun. 61: 3914 3921.
7. Bolin, I.,, L. Norlander,, and H. Wolf-Watz. 1982. Temperature-inducible outer membrane protein of Yersinia pseudotuberculosis and Yersinia enterocolitica is associated with the virulence plasmid. Infect. Immun. 37: 506 512.
8. Bolin, I.,, and H. Wolf-Watz. 1984. Molecular cloning of the temperature-inducible outer membrane protein 1 of Yersinia pseudotuberculosis. Infect. Immun. 43: 72 78.
9. Bovallius, A.,, and G. Nilsson. 1975. Ingestion and survival of Y. pseudotuberculosis in HeLa cells. Can. J. Microbiol. 21: 1997 2007.
10. Cornells, G.,, Y. Larouche,, G. Balligand,, M.-P. Sory,, and G. Wauters. 1987. Yersinia enterocolitica, a primary model for bacterial invasiveness. Rev. Infect. Dis. 9: 64 87.
11. Cornells, G.,, C. Sluiters,, D. LambertdeRouvroit,, and T. Michiels. 1989. Homology between VirF, the transcriptional activator of the Yersinia virulence regulon, and AraC, the Escherichia coli arabinose operon regulator. J. Bacteriol. 171: 254 262.
12. Cornells, G. R.,, T. Blot,, C. LambertdeRouvroit,, T. Michiels,, B. Mulder,, C. Sluiters,, M. Sory,, M. VanBouchaute,, and J. Vanooteghem. 1989. The Yersinia yop regulon. Mol. Microbiol. 3: 1455 1459.
13. Devenish, J. A.,, and D. A. Schiemann. 1981. HeLa cell infection by Yersinia enterocolitica: evidence for lack of intracellular multiplication and development of a new procedure for quantitative expression of infectivity. Infect. Immun. 32: 48 55.
14. Entody, L.,, J. Heesemann,, H. Wolf-Watz,, M. Skurnik,, G. Kapperud,, P. O'Toole,, and T. Wadstrom. 1989. Binding to collagen by Yersinia enterocolitica and Yersinia pseudotuberculosis: evidence for yopA-mediated and chromosomally encoded mechanisms. J. Bacteriol. 171: 6674 6679.
15. Goguen, J. D.,, W. S. Walker,, T. P. Hatch,, and J. Yother. 1986. Plasmid-determined cytotoxicity in Yersinia pestis and Yersinia pseudotuberculosis. Infect. Immun. 51: 788 794.
16. Grutzkau, S.,, C. Hanski, H, Hahn, and E. O. Riecken. 1990. Involvement of M cells in the bacterial invasion of Peyer's patches: a common mechanism shared by Yersinia enterocolitica and other enteroinvasive bacteria. Gut 31: 1011 1015.
17. Heffernan, E. J.,, J. Harwood,, J. Fierer,, and D. Guiney. 1992. The Salmonella typhimurium virulence plasmid complement resistance gene rck is homologous to a family of virulence-related outer membrane protein genes, including pagC and ail. J. Bacteriol. 174: 84 91.
18. Hynes, R. O. 1987. Integrins: a family of cell surface receptors. Cell 48: 549 554.
19. Isberg, R. R. 1991. Discrimination between intracellular uptake and surface adhesion of bacterial pathogens. Science 252: 934 938.
20. Isberg, R. R.,, and S. Falkow. 1985. A single genetic locus encoded by Yersinia pseudotuberculosis permits invasion of cultured animal cells by Escherichia coli K-12. Nature (London) 317: 262 264.
21. Isberg, R. R.,, and J. M. Leong. 1990. Multiple β1 chain integrins are receptors for invasin, a protein that promotes bacterial penetration into mammalian cells. Cell 60: 861 871.
22. Isberg, R. R.,, A. Swain,, and S. Falkow. 1988. Analysis of expression and thermoregulation of the Yersinia pseudotuberculosis inv gene with hybrid proteins. Infect. Immun. 56: 2133 2138.
23. Isberg, R. R.,, D. L. Voorhis,, and S. Falkow. 1987. Identification of invasin: a protein that allows enteric bacteria to penetrate cultured mammalian cells. Cell 50: 769 778.
24. Kapperud, G.,, E. Namork,, M. Skurnik, andT. Nesbakken. 1987. Plasmid-mediated surfacefibrillae of Yersinia pseudotuberculosis and Yersinia enterocolitica: relationship to the outer membrane protein YOP1 and possible importance for pathogenesis. Infect. Immun. 55: 2247 2254.
25. Lee, W. H.,, P. P. McGrath,, P. H. Carter,, and E. L. Eide. 1977. The ability of some Yersinia enterocolitica strains to invade HeLa cells. Can. J. Microbiol. 23: 1714 1722.
26. Leong, J. M.,, R. S. Fournier,, and R. R. Isberg. 1990. Identification of the integrin-binding domain of the Yersinia pseudotuberculosis invasin protein. EMBO J. 9: 1979 1989.
27. Leong, J. M.,, P. E. Morrissey,, and R. R. Isberg. 1993. A 76-amino-acid disulfide loop in the Yersinia pseudotuberculosis invasin protein is required for integrin receptor recognition. J. Biol. Chem. 268: 20524 20532.
28. Miller, J. H. 1972. Experiments in Molecular Genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y..
29. Miller, S. I.,, A. M. Kukral,, and J. J. Mekalanos. 1989. A two-component regulatory system (phoP phoQ) controls Salmonella typhimurium virulence. Proc. Natl. Acad. Sci. USA 86: 5054 5058.
30. Miller, V. L.,, K. B. Beer,, W. P. Loomis,, J. A. Olson,, and S. I. Miller. 1992. An unusual pagC::Tn phoA mutation leads to an invasion- and virulence-defective phenotype in salmonellae. Infect. Immun. 60: 3763 3770.
31. Miller, V. L.,, J. B. Bliska,, and S. Falkow. 1990. Nucleotide sequence of the Yersinia enterocolitica ail gene and characterization of the Ail protein product. J. Bacteriol. 172: 1062 1069.
32. Miller, V. L.,, and S. Falkow. 1988. Evidence for two genetic loci in Yersinia enterocolitica that can promote invasion of epithelial cells. Infect. Immun. 56: 1242 1248.
33. Miller, V. L.,, J. J. Fanner III,, W. E. Hill,, and S. Falkow. 1989. The ail locus is found uniquely in Yersinia enterocolitica serotypes commonly associated with disease. Infect. Immun. 57: 121 131.
34. Paerregaard, A.,, F. Espersen,, O. M. Jensen,, and M. Skurnik. 1991. Interactions between Yersinia enterocolitica and rabbit ileal mucus: growth, adhesion, penetration, and subsequent changes in surface hydrophobicity and ability to adhere to ileal brush border membrane vesicles. Infect. Immun. 59: 253 260.
35. Pepe, J. C.,, J. L. Badger,, and V. L. Miller. 1994. Growth phase and low pH affect the thermal regulation of the Yersinia enterocolitica inv gene. Mol. Microbiol. 11: 123 135.
36. Pepe, J. C.,, and V. L. Miller. 1993. Yersinia enterocolitica invasin: a primary role in the initiation of infection. Proc. Natl. Acad. Sci. USA 90: 6473 6477.
37. Pierson, D. E. 1994. Mutations affecting lipopolysaccharide enhance Ail-mediated entry of Yersinia enterocolitica into mammalian cells. J. Bacteriol. 176: 4043 4051.
38. Pierson, D. E.,, and S. Falkow. 1990. Nonpathogenic isolates of Yersinia enterocolitica do not contain functional mv-homologous sequences. Infect. Immun. 58: 1059 1064.
39. Pierson, D. E.,, and S. Falkow. 1993. The ail gene of Yersinia enterocolitica has a role in the ability of this organism to survive serum killing. Infect. Immun. 61: 1846 1852.
40. Portnoy, D. A.,, S. L. Moseley,, and S. Falkow. 1981. Characterization of plasmids and plasmid-associated determinants of Yersinia enterocolitica pathogenesis. Infect. Immun. 31: 775 782.
41. Pulkkinen, W. S.,, and S. I. Miller. 1991. A Salmonella typhimurium virulence protein is similar to a Yersinia enterocolitica invasion protein and a bacteriophage lambda outer membrane protein. J. Bacteriol. 173: 86 93.
42. Rankin, S.,, R. R. Isberg,, and J. M. Leong. 1992. The integrin-binding domain of invasin is sufficient to allow bacterial entry into mammalian cells. Infect. Immun. 60: 3909 3912.
43. Rosqvist, R.,, I. Bolin,, and H. Wolf-Watz. 1988. Inhibition of phagocytosis in Yersinia pseudotuberculosis: a virulence plasmid-encoded ability involving the Yop2b protein. Infect. Immun. 56: 2139 2143.
44. Rosqvist, R.,, A. Forsberg,, M. Rimpilainen,, T. Bergman,, and H. Wolf-Watz. 1990. The cytotoxic protein YopE of Yersinia obstructs the primary host defence. Mol. Microbiol. 4: 657 667.
45. Rosqvist, R.,, M. Skurnik,, and H. Wolf-Watz. 1988. Increased virulence of Yersinia pseudotuberculosis by two independent mutations. Nature (London) 334: 522 525.
46. Schiemann, D. A.,, and J. A. Devenish. 1982. Relationship of HeLa cell infectivity to biochemical, serological, and virulence characteristics of Yersinia enterocolitica. Infect. Immun. 35: 497 506.
47. Schulze-Koops, H.,, H. Burkhardt,, J. Heesemann,, K. von der Mark,, and F. Emmrich. 1992. Plasmid-encoded outer membrane protein YadA mediates specific binding of enteropathogenic yersiniae to various types of collagen. Infect. Immun. 60: 2153 2159.
48. Sikkema, D. J.,, and R. R. Brubaker. 1987. Resistance to pesticin, storage of iron, and invasion of HeLa cells by yersiniae. Infect. Immun. 55: 572 578.
49. Skurnik, M.,, and P. Toivanen. 1992. LcrF is the temperature-regulated activator of the yadA gene of Yersinia enterocolitica and Yersinia pseudotuberculosis. J. Bacteriol. 174: 2047 2051.
50. Skurnik, M.,, and P. Toivanen. 1993. Yersinia enterocolitica lipopolysaccharide: genetics and virulence. Trends Microbiol. 1: 148 152.
51. Skurnik, M.,, and H. Wolf-Watz. 1989. Analysis of the yopA gene encoding the Yop1 virulence determinants of Yersinia spp. Mot. Microbiol. 3: 517 529.
52. Tertti, R.,, M. Skurnik,, T. Vartio,, and P. Kuusela. 1992. Adhesion protein YadA of Yersinia species mediates binding of bacteria to fibronectin. Infect. Immun. 60: 3021 3024.
53. TranVanNhieu, G.,, and R. R. Isberg. 1991. The Yersinia pseudotuberculosis invasin protein and human fibronectin bind to mutually exclusive sites on the α5β1 integrin receptor. J. Biol. Chem. 266: 24367 24375.
54. TranVanNhieu, G.,, and R. R. Isberg. 1993. Bacterial internalization mediated by β1 chain integrins is determined by ligand affinity and receptor density. EMBO J. 12: 1887 1895.
55. Wolf-Watz, H.,, D. A. Portnoy,, I. Bolin,, and S. Falkow. 1985. Transfer of the virulence plasmid of Yersinia pestis to Yersinia pseudotuberculosis. Infect. Immun. 48: 241 243.
56. Yang, Y.,, and R. Isberg. 1993. Cellular internalization in the absence of invasin expression is promoted by the Yersinia pseudotuberculosis yadA product. Infect. Immun. 61: 3907 3913.
57. Young, V. B.,, S. Falkow,, and G. K. Schoolnik. 1992. The invasin protein of Yersinia enterocolitica: internalization of invasin-bearing bacteria by eukaryotic cells is associated with reorganization of the cytoskeleton. J. Cell Biol. 116: 197 207.
58. Young, V. B.,, V. L. Miller,, S. Falkow,, and G. K. Schoolnik. 1990. Sequence, localization and function of the invasin protein of Yersinia enterocolitica. Mol. Microbiol. 4: 1119 1128.


Generic image for table
Table 1

Enrichment procedure yields strains that invade cell culture monolayers

From Isberg, R. R., and S. Falkow. 1985. (London) 317:262-264 ( ). Reprinted with permission from (London) 317:262−264. Copyright 1985 Macmillan Magazines Limited. Saturated cultures of bacteria, grown at 28°C, were washed twice in phosphate-buffered saline (PBS) and resuspended to a concentration of 3 × 10 bacteria ml. Aliquots of 50 μl of each strain were added to monolayer cultures of HEp-2 cells seeded at a concentration of 2 × 10 animal cells per microtiter well (24-well Falcon 3047 microliter dishes) in RPMI 1640 medium. Bacteria were centrifuged onto the monolayer at 600 × as described previously ( ), and the infected cultures were incubated at 36°C for 3 h in a 5% CO atmosphere, to allow binding and invasion of bacteria. Nonadherent bacteria were removed from the monolayer by washing three times with sterile PBS, and RPMI 1640 medium containing 40 μg of gentamicin (Sigma) ml was added to each microtiter well. The incubation was continued at 36°C for 2 h in the presence of the antibiotic before washing of the monolayers in PBS twice more. Internalized bacteria were then released from the monolayers by the addition of 1% Triton X-100, and titers were determined on L-agar plates ( ).

Percentage of bacteria added to HEp-2 monolayers that resist treatment by gentamicin.

strain ( ).

K-12 strain HB101.

HB101 harboring cosmids that are denoted in parentheses.

Citation: Pierson D. 1994. Mechanisms of Entry into Mammalian Cells, p 235-247. In Miller V, Kaper J, Portnoy D, Isberg R (ed), Molecular Genetics of Bacterial Pathogenesis. ASM Press, Washington, DC. doi: 10.1128/9781555818340.ch15
Generic image for table
Table 2

Relative entry of clones into HEp-2 cells

Strains were used to infect a monolayer of the human larynx epithelial cell line HEp-2. Percent invasion is as described in footnote of Table 1 . Adapted from Miller, V. L., and S. Falkow. 1988. 56:1242-1248 ( ).

Citation: Pierson D. 1994. Mechanisms of Entry into Mammalian Cells, p 235-247. In Miller V, Kaper J, Portnoy D, Isberg R (ed), Molecular Genetics of Bacterial Pathogenesis. ASM Press, Washington, DC. doi: 10.1128/9781555818340.ch15
Generic image for table
Table 3

YadA-mediated entry of into HEp-2 cells

Percent invasion was determined as described in footnote of Table 1 . Adapted from Yang, Y., and R. Isberg. 1993. 61:3907−3913 ( ).

Citation: Pierson D. 1994. Mechanisms of Entry into Mammalian Cells, p 235-247. In Miller V, Kaper J, Portnoy D, Isberg R (ed), Molecular Genetics of Bacterial Pathogenesis. ASM Press, Washington, DC. doi: 10.1128/9781555818340.ch15

This is a required field
Please enter a valid email address
Please check the format of the address you have entered.
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error