• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jbacterPermissionsJournals.ASM.orgJournalJB ArticleJournal InfoAuthorsReviewers
J Bacteriol. Jul 1992; 174(13): 4338–4349.
PMCID: PMC206218

Molecular and functional characterization of the Salmonella invasion gene invA: homology of InvA to members of a new protein family.

Abstract

One of the earliest steps in the pathogenic cycle of the facultative intracellular pathogen Salmonella spp. is the invasion of the cells of the intestinal epithelium. We have previously identified a genetic locus, inv, that allows Salmonella spp. to enter cultured epithelial cells. invA is a member of this locus, and it is the first gene of an operon consisting of at least two additional invasion genes. We have constructed strains carrying nonpolar mutations in invA and examined the individual contribution of this gene to the invasion phenotype of Salmonella typhimurium. Nonpolar S. typhimurium invA mutants were deficient in invasion of cultured epithelial cells although they were fully capable of attaching to the same cells. In addition, unlike wild-type S. typhimurium, invA mutants did not alter the normal architecture of the microvilli of polarized epithelial cells nor did they cause any alterations in the distribution of actin microfilaments of infected cells. The invasion phenotype of invA mutants was readily rescued by wild-type S. typhimurium when cultured epithelial cells were simultaneously infected with both strains. On the contrary, in a similar experiment, the adherent Escherichia coli strain RDEC-1 was not internalized into cultured cells when coinfected with wild-type S. typhimurium. The invA locus was found to be located at about 59 min on the Salmonella chromosome, 7% linked to mutS. The nucleotide sequence of invA showed an open reading frame capable of encoding a polypeptide of 686 amino acids with eight possible membrane-spanning regions and a predicted molecular weight of 75,974. A protein of this size was visualized when invA was expressed in a bacteriophage T7 RNA polymerase-based expression system. The predicted sequence of InvA was found to be homologous to Caulobacter crescentus FlbF, Yersinia LcrD, Shigella flexneri VirH, and E. coli FlhA proteins. These proteins may form part of a family of proteins with a common function, quite possibly the translocation of specific proteins across the bacterial cell membrane.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (3.3M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Bernardini ML, Mounier J, d'Hauteville H, Coquis-Rondon M, Sansonetti PJ. Identification of icsA, a plasmid locus of Shigella flexneri that governs bacterial intra- and intercellular spread through interaction with F-actin. Proc Natl Acad Sci U S A. 1989 May;86(10):3867–3871. [PMC free article] [PubMed]
  • Chang AC, Cohen SN. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. [PMC free article] [PubMed]
  • Cheney CP, Schad PA, Formal SB, Boedeker EC. Species specificity of in vitro Escherichia coli adherence to host intestinal cell membranes and its correlation with in vivo colonization and infectivity. Infect Immun. 1980 Jun;28(3):1019–1027. [PMC free article] [PubMed]
  • Clerc P, Sansonetti PJ. Entry of Shigella flexneri into HeLa cells: evidence for directed phagocytosis involving actin polymerization and myosin accumulation. Infect Immun. 1987 Nov;55(11):2681–2688. [PMC free article] [PubMed]
  • 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. [PMC free article] [PubMed]
  • Elsinghorst EA, Baron LS, Kopecko DJ. Penetration of human intestinal epithelial cells by Salmonella: molecular cloning and expression of Salmonella typhi invasion determinants in Escherichia coli. Proc Natl Acad Sci U S A. 1989 Jul;86(13):5173–5177. [PMC free article] [PubMed]
  • Ernst RK, Dombroski DM, Merrick JM. Anaerobiosis, type 1 fimbriae, and growth phase are factors that affect invasion of HEp-2 cells by Salmonella typhimurium. Infect Immun. 1990 Jun;58(6):2014–2016. [PMC free article] [PubMed]
  • Falkow S. Bacterial entry into eukaryotic cells. Cell. 1991 Jun 28;65(7):1099–1102. [PubMed]
  • Fields PI, Groisman EA, Heffron F. A Salmonella locus that controls resistance to microbicidal proteins from phagocytic cells. Science. 1989 Feb 24;243(4894 Pt 1):1059–1062. [PubMed]
  • Finlay BB, Falkow S. Salmonella interactions with polarized human intestinal Caco-2 epithelial cells. J Infect Dis. 1990 Nov;162(5):1096–1106. [PubMed]
  • Finlay BB, Ruschkowski S, Dedhar S. Cytoskeletal rearrangements accompanying salmonella entry into epithelial cells. J Cell Sci. 1991 Jun;99(Pt 2):283–296. [PubMed]
  • Finlay BB, Starnbach MN, Francis CL, Stocker BA, Chatfield S, Dougan G, Falkow S. Identification and characterization of TnphoA mutants of Salmonella that are unable to pass through a polarized MDCK epithelial cell monolayer. Mol Microbiol. 1988 Nov;2(6):757–766. [PubMed]
  • Forsberg A, Viitanen AM, Skurnik M, Wolf-Watz H. The surface-located YopN protein is involved in calcium signal transduction in Yersinia pseudotuberculosis. Mol Microbiol. 1991 Apr;5(4):977–986. [PubMed]
  • Gahring LC, Heffron F, Finlay BB, Falkow S. Invasion and replication of Salmonella typhimurium in animal cells. Infect Immun. 1990 Feb;58(2):443–448. [PMC free article] [PubMed]
  • Gaillard JL, 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. [PubMed]
  • Galán JE, 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. [PMC free article] [PubMed]
  • Galán JE, Curtiss R., 3rd Expression of Salmonella typhimurium genes required for invasion is regulated by changes in DNA supercoiling. Infect Immun. 1990 Jun;58(6):1879–1885. [PMC free article] [PubMed]
  • Galán JE, Curtiss R., 3rd Distribution of the invA, -B, -C, and -D genes of Salmonella typhimurium among other Salmonella serovars: invA mutants of Salmonella typhi are deficient for entry into mammalian cells. Infect Immun. 1991 Sep;59(9):2901–2908. [PMC free article] [PubMed]
  • Henikoff S. Unidirectional digestion with exonuclease III in DNA sequence analysis. Methods Enzymol. 1987;155:156–165. [PubMed]
  • Isberg RR. Determinants for thermoinducible cell binding and plasmid-encoded cellular penetration detected in the absence of the Yersinia pseudotuberculosis invasin protein. Infect Immun. 1989 Jul;57(7):1998–2005. [PMC free article] [PubMed]
  • Isberg RR. Discrimination between intracellular uptake and surface adhesion of bacterial pathogens. Science. 1991 May 17;252(5008):934–938. [PubMed]
  • Isberg RR, Falkow S. A single genetic locus encoded by Yersinia pseudotuberculosis permits invasion of cultured animal cells by Escherichia coli K-12. Nature. 1985 Sep 19;317(6034):262–264. [PubMed]
  • Isberg RR, Leong JM. Multiple beta 1 chain integrins are receptors for invasin, a protein that promotes bacterial penetration into mammalian cells. Cell. 1990 Mar 9;60(5):861–871. [PubMed]
  • Isberg RR, Voorhis DL, Falkow S. Identification of invasin: a protein that allows enteric bacteria to penetrate cultured mammalian cells. Cell. 1987 Aug 28;50(5):769–778. [PubMed]
  • Kihlström E, Nilsson L. Endocytosis of Salmonella typhimurium 395 MS and MR10 by HeLa cells. Acta Pathol Microbiol Scand B. 1977 Oct;85B(5):322–328. [PubMed]
  • Kukral AM, Strauch KL, Maurer RA, Miller CG. 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. [PMC free article] [PubMed]
  • Kyte J, Doolittle RF. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. [PubMed]
  • Lee CA, 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. [PMC free article] [PubMed]
  • LENNOX ES. Transduction of linked genetic characters of the host by bacteriophage P1. Virology. 1955 Jul;1(2):190–206. [PubMed]
  • Liu SL, Ezaki T, Miura H, Matsui K, Yabuuchi E. Intact motility as a Salmonella typhi invasion-related factor. Infect Immun. 1988 Aug;56(8):1967–1973. [PMC free article] [PubMed]
  • MacLachlan PR, Sanderson KE. Transformation of Salmonella typhimurium with plasmid DNA: differences between rough and smooth strains. J Bacteriol. 1985 Jan;161(1):442–445. [PMC free article] [PubMed]
  • Makino S, Sasakawa C, Kamata K, Kurata T, Yoshikawa M. A genetic determinant required for continuous reinfection of adjacent cells on large plasmid in S. flexneri 2a. Cell. 1986 Aug 15;46(4):551–555. [PubMed]
  • Manoil C, Beckwith J. TnphoA: a transposon probe for protein export signals. Proc Natl Acad Sci U S A. 1985 Dec;82(23):8129–8133. [PMC free article] [PubMed]
  • MARMUR J, DOTY P. Determination of the base composition of deoxyribonucleic acid from its thermal denaturation temperature. J Mol Biol. 1962 Jul;5:109–118. [PubMed]
  • Michiels T, Vanooteghem JC, Lambert de Rouvroit C, China B, Gustin A, Boudry P, Cornelis GR. Analysis of virC, an operon involved in the secretion of Yop proteins by Yersinia enterocolitica. J Bacteriol. 1991 Aug;173(16):4994–5009. [PMC free article] [PubMed]
  • Miller VL, Falkow S. Evidence for two genetic loci in Yersinia enterocolitica that can promote invasion of epithelial cells. Infect Immun. 1988 May;56(5):1242–1248. [PMC free article] [PubMed]
  • Miller VL, Finlay BB, Falkow S. Factors essential for the penetration of mammalian cells by Yersinia. Curr Top Microbiol Immunol. 1988;138:15–39. [PubMed]
  • Moulder JW. Comparative biology of intracellular parasitism. Microbiol Rev. 1985 Sep;49(3):298–337. [PMC free article] [PubMed]
  • Mroczenski-Wildey MJ, Di Fabio JL, Cabello FC. Invasion and lysis of HeLa cell monolayers by Salmonella typhi: the role of lipopolysaccharide. Microb Pathog. 1989 Feb;6(2):143–152. [PubMed]
  • Newton A, Ohta N, Ramakrishnan G, Mullin D, Raymond G. Genetic switching in the flagellar gene hierarchy of Caulobacter requires negative as well as positive regulation of transcription. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6651–6655. [PMC free article] [PubMed]
  • Oka A, Sugisaki H, Takanami M. Nucleotide sequence of the kanamycin resistance transposon Tn903. J Mol Biol. 1981 Apr 5;147(2):217–226. [PubMed]
  • Ortega-Barria E, Pereira ME. A novel T. cruzi heparin-binding protein promotes fibroblast adhesion and penetration of engineered bacteria and trypanosomes into mammalian cells. Cell. 1991 Oct 18;67(2):411–421. [PubMed]
  • Plano GV, Barve SS, Straley SC. LcrD, a membrane-bound regulator of the Yersinia pestis low-calcium response. J Bacteriol. 1991 Nov;173(22):7293–7303. [PMC free article] [PubMed]
  • Ramakrishnan G, Zhao JL, Newton A. The cell cycle-regulated flagellar gene flbF of Caulobacter crescentus is homologous to a virulence locus (lcrD) of Yersinia pestis. J Bacteriol. 1991 Nov;173(22):7283–7292. [PMC free article] [PubMed]
  • Russell CB, Thaler DS, Dahlquist FW. Chromosomal transformation of Escherichia coli recD strains with linearized plasmids. J Bacteriol. 1989 May;171(5):2609–2613. [PMC free article] [PubMed]
  • Russel M, Kidd S, Kelley MR. An improved filamentous helper phage for generating single-stranded plasmid DNA. Gene. 1986;45(3):333–338. [PubMed]
  • Sanderson KE, Ross H, Ziegler L, Mäkelä PH. F + , Hfr, and F' strains of Salmonella typhimurium and Salmonella abony. Bacteriol Rev. 1972 Dec;36(4):608–637. [PMC free article] [PubMed]
  • Sanderson KE, Roth JR. Linkage map of Salmonella typhimurium, edition VII. Microbiol Rev. 1988 Dec;52(4):485–532. [PMC free article] [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PMC free article] [PubMed]
  • Sansonetti PJ. Genetic and molecular basis of epithelial cell invasion by Shigella species. Rev Infect Dis. 1991 Mar-Apr;13 (Suppl 4):S285–S292. [PubMed]
  • Schenkman S, Jiang MS, Hart GW, Nussenzweig V. A novel cell surface trans-sialidase of Trypanosoma cruzi generates a stage-specific epitope required for invasion of mammalian cells. Cell. 1991 Jun 28;65(7):1117–1125. [PubMed]
  • SCHNEIDER HA, ZINDER ND. Nutrition of the host and natural resistance to infection. V. An improved assay employing genetic markers in the double strain inoculation test. J Exp Med. 1956 Feb 1;103(2):207–223. [PMC free article] [PubMed]
  • Small PL, Isberg RR, Falkow S. Comparison of the ability of enteroinvasive Escherichia coli, Salmonella typhimurium, Yersinia pseudotuberculosis, and Yersinia enterocolitica to enter and replicate within HEp-2 cells. Infect Immun. 1987 Jul;55(7):1674–1679. [PMC free article] [PubMed]
  • Straley SC, Bowmer WS. Virulence genes regulated at the transcriptional level by Ca2+ in Yersinia pestis include structural genes for outer membrane proteins. Infect Immun. 1986 Feb;51(2):445–454. [PMC free article] [PubMed]
  • Studier FW, Moffatt BA. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. [PubMed]
  • Tabor S, Richardson CC. 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. [PMC free article] [PubMed]
  • Takeuchi A. Electron microscope studies of experimental Salmonella infection. I. Penetration into the intestinal epithelium by Salmonella typhimurium. Am J Pathol. 1967 Jan;50(1):109–136. [PMC free article] [PubMed]
  • Tilney LG, Portnoy DA. Actin filaments and the growth, movement, and spread of the intracellular bacterial parasite, Listeria monocytogenes. J Cell Biol. 1989 Oct;109(4 Pt 1):1597–1608. [PMC free article] [PubMed]
  • Vieira J, Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. [PubMed]
  • Viitanen AM, Toivanen P, Skurnik M. The lcrE gene is part of an operon in the lcr region of Yersinia enterocolitica O:3. J Bacteriol. 1990 Jun;172(6):3152–3162. [PMC free article] [PubMed]
  • Vogler AP, Homma M, Irikura VM, Macnab RM. Salmonella typhimurium mutants defective in flagellar filament regrowth and sequence similarity of FliI to F0F1, vacuolar, and archaebacterial ATPase subunits. J Bacteriol. 1991 Jun;173(11):3564–3572. [PMC free article] [PubMed]
  • Wang K, Feramisco JR, Ash JF. Fluorescent localization of contractile proteins in tissue culture cells. Methods Enzymol. 1982;85(Pt B):514–562. [PubMed]

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

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...