1887

Abstract

The Gram-positive intracellular pathogen is endowed with 17 sets of genes encoding two-component systems. is closely related to the Gram-positive model bacterium , in which we have shown previously that the DegS/DegU system plays a central role in controlling stationary phase adaptive responses, including degradative enzyme synthesis and competence. Although an orthologue of the DegU response regulator is present in , the gene encoding the cognate DegS kinase is conspicuously absent. We have inactivated the gene of and shown that DegU negatively regulates its own synthesis. Direct binding of DegU to its own promoter region was shown by gel mobility shift and DNase I footprinting experiments. DegU was also shown to bind upstream from the operon, which also encodes the GmaR anti-repressor of flagellar synthesis. In contrast to the situation in , DegU was shown to be essential for flagellar synthesis and bacterial motility in and is cotranscribed with the gene located downstream. We also show that DegU is required for growth at high temperatures, adherence to plastic surfaces and the formation of efficient biofilms by . DegU plays a role in virulence of as well: in a murine intravenous infection model, an 11-fold increase in LD was observed for the mutant. Taken together, our results indicate that despite the lack of the DegS kinase, DegU is fully functional as an orphan response regulator, and plays a central role in controlling several crucial adaptive responses in .

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.2008/017590-0
2008-08-01
2024-04-18
Loading full text...

Full text loading...

/deliver/fulltext/micro/154/8/2251.html?itemId=/content/journal/micro/10.1099/mic.0.2008/017590-0&mimeType=html&fmt=ahah

References

  1. Altschul S. F., Madden T. L., Schaffer A. A., Zhang J., Zhang Z., Miller W., Lipman D. J. 1997; Gapped blast and psi-blast: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402
    [Google Scholar]
  2. Amati G., Bisicchia P., Galizzi A. 2004; DegU-P represses expression of the motility fla-che operon in Bacillus subtilis . J Bacteriol 186:6003–6014
    [Google Scholar]
  3. Amrein K. E., Takacs B., Stieger M., Molnos J., Flint N. A., Burn P. 1995; Purification and characterization of recombinant human p50csk protein-tyrosine kinase from an Escherichia coli expression system overproducing the bacterial chaperones GroES and GroEL. Proc Natl Acad Sci U S A 92:1048–1052
    [Google Scholar]
  4. Arnaud M., Chastanet A., Debarbouille M. 2004; New vector for efficient allelic replacement in naturally nontransformable, low-GC-content, Gram-positive bacteria. Appl Environ Microbiol 70:6887–6891
    [Google Scholar]
  5. Bigot A., Pagniez H., Botton E., Frehel C., Dubail I., Jacquet C., Charbit A., Raynaud C. 2005; Role of FliF and FliI of Listeria monocytogenes in flagellar assembly and pathogenicity. Infect Immun 73:5530–5539
    [Google Scholar]
  6. Borezée E., Msadek T., Durant L., Berche P. 2000; Identification in Listeria monocytogenes of MecA, a homologue of the Bacillus subtilis competence regulatory protein. J Bacteriol 182:5931–5934
    [Google Scholar]
  7. Bradford M. M. 1976; A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254
    [Google Scholar]
  8. Branda S. S., Gonzalez-Pastor J. E., Ben-Yehuda S., Losick R., Kolter R. 2001; Fruiting body formation by Bacillus subtilis . Proc Natl Acad Sci U S A 98:11621–11626
    [Google Scholar]
  9. Burkholder P. R., Giles N. H. Jr 1947; Induced biochemical mutations in Bacillus subtilis . Am J Bot 34:345–348
    [Google Scholar]
  10. Chastanet A., Prudhomme M., Claverys J. P., Msadek T. 2001; Regulation of Streptococcus pneumoniae clp genes and their role in competence development and stress survival. J Bacteriol 183:7295–7307
    [Google Scholar]
  11. Chastanet A., Fert J., Msadek T. 2003; Comparative genomics reveal novel heat shock regulatory mechanisms in Staphylococcus aureus and other Gram-positive bacteria. Mol Microbiol 47:1061–1073
    [Google Scholar]
  12. Cossart P., Portnoy D. A. 2000; The cell biology of invasion and intracellular growth by Listeria monocytogenes.. In Gram-Positive Pathogens pp 507–515 Edited by Fischetti V. A., Novick R. P., Feretti J. J., Portnoy D. A., Rood J. I. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  13. Cotter P. D., Guinane C. M., Hill C. 2002; The LisRK signal transduction system determines the sensitivity of Listeria monocytogenes to nisin and cephalosporins. Antimicrob Agents Chemother 46:2784–2790
    [Google Scholar]
  14. Dahl M. K., Msadek T., Kunst F., Rapoport G. 1992; The phosphorylation state of the DegU response regulator acts as a molecular switch allowing either degradative enzyme synthesis or expression of genetic competence in Bacillus subtilis . J Biol Chem 267:14509–14514
    [Google Scholar]
  15. Derré I., Rapoport G., Msadek T. 1999; CtsR, a novel regulator of stress and heat shock response, controls clp and molecular chaperone gene expression in Gram-positive bacteria. Mol Microbiol 31:117–132
    [Google Scholar]
  16. Dons L., Eriksson E., Jin Y., Rottenberg M. E., Kristensson K., Larsen C. N., Bresciani J., Olsen J. E. 2004; Role of flagellin and the two-component CheA/CheY system of Listeria monocytogenes in host cell invasion and virulence. Infect Immun 72:3237–3244
    [Google Scholar]
  17. Dubrac S., Msadek T. 2004; Identification of genes controlled by the essential YycG/YycF two-component system of Staphylococcus aureus . J Bacteriol 186:1175–1181
    [Google Scholar]
  18. Dubrac S., Boneca I. G., Poupel O., Msadek T. 2007; New insights into the WalK/WalR (YycG/YycF) essential signal transduction pathway reveal a major role in controlling cell wall metabolism and biofilm formation in Staphylococcus aureus . J Bacteriol 189:8257–8269
    [Google Scholar]
  19. Glaser P., Frangeul L., Buchrieser C., Rusniok C., Amend A., Baquero F., Berche P., Bloecker H., Brandt P. other authors 2001; Comparative genomics of Listeria species. Science 294:849–852
    [Google Scholar]
  20. Glatron M. F., Rapoport G. 1972; Biosynthesis of the parasporal inclusion of Bacillus thuringiensis: half-life of its corresponding messenger RNA. Biochimie 54:1291–1301
    [Google Scholar]
  21. Grundling A., Burrack L. S., Bouwer H. G., Higgins D. E. 2004; Listeria monocytogenes regulates flagellar motility gene expression through MogR, a transcriptional repressor required for virulence. Proc Natl Acad Sci U S A 101:12318–12323
    [Google Scholar]
  22. Hamoen L. W., Van Werkhoven A. F., Bijlsma J. J., Dubnau D., Venema G. 1998; The competence transcription factor of Bacillus subtilis recognizes short A/T-rich sequences arranged in a unique, flexible pattern along the DNA helix. Genes Dev 12:1539–1550
    [Google Scholar]
  23. Hamoen L. W., Van Werkhoven A. F., Venema G., Dubnau D. 2000; The pleiotropic response regulator DegU functions as a priming protein in competence development in Bacillus subtilis . Proc Natl Acad Sci U S A 97:9246–9251
    [Google Scholar]
  24. Helmann J. D. 1991; Alternative sigma factors and the regulation of flagellar gene expression. Mol Microbiol 5:2875–2882
    [Google Scholar]
  25. Henner D. J., Yang M., Ferrari E. 1988; Localization of Bacillus subtilis sacU(Hy) mutations to two linked genes with similarities to the conserved procaryotic family of two-component signalling systems. J Bacteriol 170:5102–5109
    [Google Scholar]
  26. Hoch J. A., Silhavy T. J. 1995 Two-Component Signal Transduction Washington, DC: American Society for Microbiology;
    [Google Scholar]
  27. Kallipolitis B. H., Ingmer H., Gahan C. G., Hill C., Sogaard-Andersen L. 2003; CesRK, a two-component signal transduction system in Listeria monocytogenes, responds to the presence of cell wall-acting antibiotics and affects β-lactam resistance. Antimicrob Agents Chemother 47:3421–3429
    [Google Scholar]
  28. Karatzas K. A., Wouters J. A., Gahan C. G., Hill C., Abee T., Bennik M. H. 2003; The CtsR regulator of Listeria monocytogenes contains a variant glycine repeat region that affects piezotolerance, stress resistance, motility and virulence. Mol Microbiol 49:1227–1238
    [Google Scholar]
  29. Kathariou S., Kanenaka R., Allen R. D., Fok A. K., Mizumoto C. 1995; Repression of motility and flagellin production at 37 degrees C is stronger in Listeria monocytogenes than in the nonpathogenic species Listeria innocua . Can J Microbiol 41:572–577
    [Google Scholar]
  30. Knudsen G. M., Olsen J. E., Dons L. 2004; Characterization of DegU, a response regulator in Listeria monocytogenes, involved in regulation of motility and contributes to virulence. FEMS Microbiol Lett 240:171–179
    [Google Scholar]
  31. Kobayashi K. 2007a; Gradual activation of the response regulator DegU controls serial expression of genes for flagellum formation and biofilm formation in Bacillus subtilis . Mol Microbiol 66:395–409
    [Google Scholar]
  32. Kobayashi K. 2007b; Bacillus subtilis pellicle formation proceeds through genetically defined morphological changes. J Bacteriol 189:4920–4931
    [Google Scholar]
  33. Kunst F., Debarbouille M., Msadek T., Young M., Mauel C., Karamata D., Klier A., Rapoport G., Dedonder R. 1988; Deduced polypeptides encoded by the Bacillus subtilis sacU locus share homology with two-component sensor-regulator systems. J Bacteriol 170:5093–5101
    [Google Scholar]
  34. Lemon K. P., Higgins D. E., Kolter R. 2007; Flagellar motility is critical for Listeria monocytogenes biofilm formation. J Bacteriol 189:4418–4424
    [Google Scholar]
  35. Mandin P., Fsihi H., Dussurget O., Vergassola M., Milohanic E., Toledo-Arana A., Lasa I., Johansson J., Cossart P. 2005; VirR, a response regulator critical for Listeria monocytogenes virulence. Mol Microbiol 57:1367–1380
    [Google Scholar]
  36. Michel E., Mengaud J., Galsworthy S., Cossart P. 1998; Characterization of a large motility gene cluster containing the cheR, motAB genes of Listeria monocytogenes and evidence that PrfA downregulates motility genes. FEMS Microbiol Lett 169:341–347
    [Google Scholar]
  37. Moszer I., Jones L. M., Moreira S., Fabry C., Danchin A. 2002; SubtiList: the reference database for the Bacillus subtilis genome. Nucleic Acids Res 30:62–65
    [Google Scholar]
  38. Msadek T. 1999; When the going gets tough: survival strategies and environmental signaling networks in Bacillus subtilis . Trends Microbiol 7:201–207
    [Google Scholar]
  39. Msadek T., Kunst F., Henner D., Klier A., Rapoport G., Dedonder R. 1990; Signal transduction pathway controlling synthesis of a class of degradative enzymes in Bacillus subtilis: expression of the regulatory genes and analysis of mutations in degS and degU . J Bacteriol 172:824–834
    [Google Scholar]
  40. Msadek T., Kunst F., Rapoport G. 1993; Two-component regulatory systems. In Bacillus subtilis and other Gram-Positive Bacteria: Biochemistry, Physiology, and Molecular Genetics pp 729–745 Edited by Sonenshein A. L., Hoch J. A., Losick R. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  41. Msadek T., Kunst F., Rapoport G. 1995; A signal transduction network in Bacillus subtilis includes the DegS/DegU and ComP/ComA two-component systems. In Two-Component Signal Transduction pp 447–471 Edited by Hoch J. A., Silhavy T. J. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  42. Msadek T., Dartois V., Kunst F., Herbaud M. L., Denizot F., Rapoport G. 1998; ClpP of Bacillus subtilis is required for competence development, motility, degradative enzyme synthesis, growth at high temperature and sporulation. Mol Microbiol 27:899–914
    [Google Scholar]
  43. Nair S., Derre I., Msadek T., Gaillot O., Berche P. 2000; CtsR controls class III heat shock gene expression in the human pathogen Listeria monocytogenes . Mol Microbiol 35:800–811
    [Google Scholar]
  44. O'Neil H. S., Marquis H. 2006; Listeria monocytogenes flagella are used for motility, not as adhesins, to increase host cell invasion. Infect Immun 74:6675–6681
    [Google Scholar]
  45. Peel M., Donachie W., Shaw A. 1988; Temperature-dependent expression of flagella of Listeria monocytogenes studied by electron microscopy, SDS-PAGE and Western blotting. J Gen Microbiol 134:2171–2178
    [Google Scholar]
  46. Premaratne R. J., Lin W. J., Johnson E. A. 1991; Development of an improved chemically defined minimal medium for Listeria monocytogenes . Appl Environ Microbiol 57:3046–3048
    [Google Scholar]
  47. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual , 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  48. Sanchez-Campillo M., Dramsi S., Gomez-Gomez J. M., Michel E., Dehoux P., Cossart P., Baquero F., Perez-Diaz J. C. 1995; Modulation of DNA topology by flaR, a new gene from Listeria monocytogenes . Mol Microbiol 18:801–811
    [Google Scholar]
  49. Schmittgen T. D., Zakrajsek B. A. 2000; Effect of experimental treatment on housekeeping gene expression: validation by real-time, quantitative RT-PCR. J Biochem Biophys Methods 46:69–81
    [Google Scholar]
  50. Shen A., Higgins D. E. 2006; The MogR transcriptional repressor regulates nonhierarchal expression of flagellar motility genes and virulence in Listeria monocytogenes . PLoS Pathog 2:e30
    [Google Scholar]
  51. Shen A., Kamp H. D., Grundling A., Higgins D. E. 2006; A bifunctional O-GlcNAc transferase governs flagellar motility through anti-repression. Genes Dev 20:3283–3295
    [Google Scholar]
  52. Smits W. K., Eschevins C. C., Susanna K. A., Bron S., Kuipers O. P., Hamoen L. W. 2005; Stripping Bacillus: ComK auto-stimulation is responsible for the bistable response in competence development. Mol Microbiol 56:604–614
    [Google Scholar]
  53. Smits W. K., Hoa T. T., Hamoen L. W., Kuipers O. P., Dubnau D. 2007; Antirepression as a second mechanism of transcriptional activation by a minor groove binding protein. Mol Microbiol 64:368–381
    [Google Scholar]
  54. Stanley N. R., Lazazzera B. A. 2005; Defining the genetic differences between wild and domestic strains of Bacillus subtilis that affect poly- γ-dl-glutamic acid production and biofilm formation. Mol Microbiol 57:1143–1158
    [Google Scholar]
  55. Stock J. B., Ninfa A. J., Stock A. M. 1989; Protein phosphorylation and regulation of adaptive responses in bacteria. Microbiol Rev 53:450–490
    [Google Scholar]
  56. Studier F. W., Moffatt B. A. 1986; Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol 189:113–130
    [Google Scholar]
  57. Sullivan M. A., Yasbin R. E., Young F. E. 1984; New shuttle vectors for Bacillus subtilis and Escherichia coli which allow rapid detection of inserted fragments. Gene 29:21–26
    [Google Scholar]
  58. Tokunaga T., Rashid M. H., Kuroda A., Sekiguchi J. 1994; Effect of degS-degU mutations on the expression of sigD, encoding an alternative sigma factor, and autolysin operon of Bacillus subtilis . J Bacteriol 176:5177–5180
    [Google Scholar]
  59. Tresse O., Lebret V., Benezech T., Faille C. 2006; Comparative evaluation of adhesion, surface properties, and surface protein composition of Listeria monocytogenes strains after cultivation at constant pH of 5 and 7. J Appl Microbiol 101:53–62
    [Google Scholar]
  60. Verhamme D. T., Kiley T. B., Stanley-Wall N. R. 2007; DegU co-ordinates multicellular behaviour exhibited by Bacillus subtilis . Mol Microbiol 65:554–568
    [Google Scholar]
  61. Way S. S., Thompson L. J., Lopes J. E., Hajjar A. M., Kollmann T. R., Freitag N. E., Wilson C. B. 2004; Characterization of flagellin expression and its role in Listeria monocytogenes infection and immunity. Cell Microbiol 6:235–242
    [Google Scholar]
  62. Williams T., Bauer S., Beier D., Kuhn M. 2005a; Construction and characterization of Listeria monocytogenes mutants with in-frame deletions in the response regulator genes identified in the genome sequence. Infect Immun 73:3152–3159
    [Google Scholar]
  63. Williams T., Joseph B., Beier D., Goebel W., Kuhn M. 2005b; Response regulator DegU of Listeria monocytogenes regulates the expression of flagella-specific genes. FEMS Microbiol Lett 252:287–298
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.2008/017590-0
Loading
/content/journal/micro/10.1099/mic.0.2008/017590-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error