Nucleoid-associated protein HU controls three regulons that coordinate virulence, response to stress and general physiology in serovar Typhimurium Free

Abstract

The role of the HU nucleoid-associated proteins in gene regulation was examined in serovar Typhimurium. The dimeric HU protein consists of different combinations of its and subunits. Transcriptomic analysis was performed with cultures growing at 37 °C at 1, 4 and 6 h after inoculation with mutants that lack combinations of HU and HU . Distinct but overlapping patterns of gene expression were detected at each time point for each of the three mutants, revealing not one but three regulons of genes controlled by the HU proteins. Mutations in the genes altered the expression of regulatory and structural genes in both the SPI1 and SPI2 pathogenicity islands. The double mutant was defective in invasion of epithelial cell lines and in its ability to survive in macrophages. The double mutant also had defective swarming activity and a competitive fitness disadvantage compared with the wild-type. In contrast, inactivation of just the gene resulted in increased fitness and correlated with the upregulation of members of the RpoS regulon in exponential-phase cultures. Our data show that HU coordinates the expression of genes involved in central metabolism and virulence and contributes to the success of as a pathogen.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.046359-0
2011-04-01
2024-03-29
Loading full text...

Full text loading...

/deliver/fulltext/micro/157/4/1075.html?itemId=/content/journal/micro/10.1099/mic.0.046359-0&mimeType=html&fmt=ahah

References

  1. Azam T. A., Ishihama A. 1999; Twelve species of the nucleoid-associated protein from Escherichia coli . Sequence recognition specificity and DNA binding affinity. J Biol Chem 274:33105–33113
    [Google Scholar]
  2. Balandina A., Claret L., Hengge-Aronis R., Rouvière-Yaniv J. 2001; The Escherichia coli histone-like protein HU regulates rpoS translation. Mol Microbiol 39:1069–1079
    [Google Scholar]
  3. Bobik T. A., Havemann G. D., Busch R. J., Williams D. S., Aldrich H. C. 1999; The propanediol utilization ( pdu ) operon of Salmonella enterica serovar Typhimurium LT2 includes genes necessary for formation of polyhedral organelles involved in coenzyme B12-dependent 1, 2-propanediol degradation. J Bacteriol 181:5967–5975
    [Google Scholar]
  4. 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]
  5. Brown J. D., Saini S., Aldridge C., Herbert J., Rao C. V., Aldridge P. D. 2008; The rate of protein secretion dictates the temporal dynamics of flagellar gene expression. Mol Microbiol 70:924–937
    [Google Scholar]
  6. Champion K., Higgins N. P. 2007; Growth rate toxicity phenotypes and homeostatic supercoil control differentiate Escherichia coli from Salmonella enterica serovar Typhimurium. J Bacteriol 189:5839–5849
    [Google Scholar]
  7. Cheng S., Bobik T. A. 2010; Characterization of the PduS cobalamin reductase of Salmonella enterica and its role in the Pdu microcompartment. J Bacteriol 192:5071–5080
    [Google Scholar]
  8. Cirillo D. M., Valdivia R. H., Monack D. M., Falkow S. 1998; Macrophage-dependent induction of the Salmonella pathogenicity island 2 type III secretion system and its role in intracellular survival. Mol Microbiol 30:175–188
    [Google Scholar]
  9. Claret L., Rouvière-Yaniv J. 1997; Variation in HU composition during growth of Escherichia coli : the heterodimer is required for long term survival. J Mol Biol 273:93–104
    [Google Scholar]
  10. Clarke T. A., Mills P. C., Poock S. R., Butt J. N., Cheesman M. R., Cole J. A., Hinton J. C., Hemmings A. M., Kemp G. other authors 2008; Escherichia coli cytochrome c nitrite reductase NrfA. Methods Enzymol 437:63–77
    [Google Scholar]
  11. Datsenko K. A., Wanner B. L. 2000; One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97:6640–6645
    [Google Scholar]
  12. Davidson A. L., Dassa E., Orelle C., Chen J. 2008; Structure, function, and evolution of bacterial ATP-binding cassette systems. Microbiol Mol Biol Rev 72:317–364
    [Google Scholar]
  13. Dillon S. C., Dorman C. J. 2010; Bacterial nucleoid-associated proteins, nucleoid structure and gene expression. Nat Rev Microbiol 8:185–195
    [Google Scholar]
  14. Dillon S. C., Cameron A. D. S., Hokamp K., Lucchini S., Hinton J. C. D., Dorman C. J. 2010; Genome-wide analysis of the H-NS and Sfh regulatory networks in Salmonella Typhimurium identifies a plasmid-encoded transcription silencing mechanism. Mol Microbiol 76:1250–1265
    [Google Scholar]
  15. Dorman C. J. 2007; H-NS, the genome sentinel. Nat Rev Microbiol 5:157–161
    [Google Scholar]
  16. Dorman C. J., Deighan P. 2003; Regulation of gene expression by histone-like proteins in bacteria. Curr Opin Genet Dev 13:179–184
    [Google Scholar]
  17. Doyle M., Fookes M., Ivens A., Mangan M. W., Wain J., Dorman C. J. 2007; An H-NS-like stealth protein aids horizontal DNA transmission in bacteria. Science 315:251–252
    [Google Scholar]
  18. Eriksson S., Lucchini S., Thompson A., Rhen M., Hinton J. C. 2003; Unravelling the biology of macrophage infection by gene expression profiling of intracellular Salmonella enterica . Mol Microbiol 47:103–118
    [Google Scholar]
  19. Fahlen T. F., Mathur N., Jones B. D. 2000; Identification and characterization of mutants with increased expression of hilA , the invasion gene transcriptional activator of Salmonella typhimurium . FEMS Immunol Med Microbiol 28:25–35
    [Google Scholar]
  20. Finlay B. B., Brumell J. H. 2000; Salmonella interactions with host cells: in vitro to in vivo . Philos Trans R Soc Lond B Biol Sci 355:623–631
    [Google Scholar]
  21. Galán J. E. 2001; Salmonella interactions with host cells: type III secretion at work. Annu Rev Cell Dev Biol 17:53–86
    [Google Scholar]
  22. Grifantini R., Sebastian S., Frigimelica E., Draghi M., Bartolini E., Muzzi A., Rappuoli R., Grandi G., Genco C. A. 2003; Identification of iron-activated and -repressed Fur-dependent genes by transcriptome analysis of Neisseria meningitidis group B. Proc Natl Acad Sci U S A 100:9542–9547
    [Google Scholar]
  23. Groisman E. A., Mouslim C. 2000; Molecular mechanisms of Salmonella pathogenesis. Curr Opin Infect Dis 13:519–522
    [Google Scholar]
  24. Groisman E. A., Ochman H. 1997; How Salmonella became a pathogen. Trends Microbiol 5:343–349
    [Google Scholar]
  25. Guo F., Adhya S. 2007; Spiral structure of Escherichia coli HU αβ provides foundation for DNA supercoiling. Proc Natl Acad Sci U S A 104:4309–4314
    [Google Scholar]
  26. Hardt W. D., Chen L. M., Schuebel K. E., Bustelo X. R., Galán J. E. 1998; S. typhimurium encodes an activator of Rho GTPases that induces membrane ruffling and nuclear responses in host cells. Cell 93:815–826
    [Google Scholar]
  27. Harrison J. A., Pickard D., Higgins C. F., Khan A., Chatfield S. N., Ali T., Dorman C. J., Hormaeche C. E., Dougan G. 1994; Role of hns in the virulence phenotype of pathogenic salmonellae. Mol Microbiol 13:133–140
    [Google Scholar]
  28. Hautefort I., Thompson A., Eriksson-Ygberg S., Parker M. L., Lucchini S., Danino V., Bongaerts R. J., Ahmad N., Rhen M., Hinton J. C. 2008; During infection of epithelial cells Salmonella enterica serovar Typhimurium undergoes a time-dependent transcriptional adaptation that results in simultaneous expression of three type 3 secretion systems. Cell Microbiol 10:958–984
    [Google Scholar]
  29. Hensel M. 2000; Salmonella pathogenicity island 2. Mol Microbiol 36:1015–1023
    [Google Scholar]
  30. Hensel M., Shea J. E., Gleeson C., Jones M. D., Dalton E., Holden D. W. 1995; Simultaneous identification of bacterial virulence genes by negative selection. Science 269:400–403
    [Google Scholar]
  31. Higgins C. F., Dorman C. J., Stirling D. A., Waddell L., Booth I. R., May G., Bremer E. 1988; A physiological role for DNA supercoiling in the osmotic regulation of gene expression in S. typhimurium and E. coli . Cell 52:569–584
    [Google Scholar]
  32. Hinton J. C. D., Hautefort I., Eriksson S., Thompson A., Rhen M. 2004; Benefits and pitfalls of using microarrays to monitor bacterial gene expression during infection. Curr Opin Microbiol 7:277–282
    [Google Scholar]
  33. Hoiseth S. K., Stocker B. A. 1981; Aromatic-dependent Salmonella typhimurium are non-virulent and effective as live vaccines. Nature 291:238–239
    [Google Scholar]
  34. Holden D. W. 2002; Trafficking of the Salmonella vacuole in macrophages. Traffic 3:161–169
    [Google Scholar]
  35. Huisman O., Faelen M., Girard D., Jaffé A., Toussaint A., Rouvière-Yaniv J. 1989; Multiple defects in Escherichia coli mutants lacking HU protein. J Bacteriol 171:3704–3712
    [Google Scholar]
  36. Kamashev D., Balandina A., Rouvière-Yaniv J. 1999; The binding motif recognized by HU on both nicked and cruciform DNA. EMBO J 18:5434–5444
    [Google Scholar]
  37. Kamashev D., Balandina A., Mazur A. K., Arimondo P. B., Rouvière-Yaniv J. 2008; HU binds and folds single-stranded DNA. Nucleic Acids Res 36:1026–1036
    [Google Scholar]
  38. Kar S., Edgar R., Adhya S. 2005; Nucleoid remodeling by an altered HU protein: reorganization of the transcription program. Proc Natl Acad Sci U S A 102:16397–16402
    [Google Scholar]
  39. Kelly A., Goldberg M. D., Carroll R. K., Danino V., Hinton J. C. D., Dorman C. J. 2004; A global role for Fis in the transcriptional control of metabolism and type III secretion in Salmonella enterica serovar Typhimurium. Microbiology 150:2037–2053
    [Google Scholar]
  40. Lacour S., Landini P. 2004; σ S-dependent gene expression at the onset of stationary phase in Escherichia coli : function of σ S-dependent genes and identification of their promoter sequences. J Bacteriol 186:7186–7195
    [Google Scholar]
  41. Lawhon S. D., Frye J. G., Suyemoto M., Porwollik S., McClelland M., Altier C. 2003; Global regulation by CsrA in Salmonella typhimurium . Mol Microbiol 48:1633–1645
    [Google Scholar]
  42. Lewis D. E., Geanacopoulos M., Adhya S. 1999; Role of HU and DNA supercoiling in transcription repression: specialized nucleoprotein repression complex at gal promoters in Escherichia coli . Mol Microbiol 31:451–461
    [Google Scholar]
  43. Lewis D. E. A., Lee S. J., Adhya S. 2010; Role of HU in regulation of gal promoters. In Bacterial Chromatin pp 395–417 Edited by Dame R. T., Dorman C. J. Heidelberg: Springer;
    [Google Scholar]
  44. Li S., Waters R. 1998; Escherichia coli strains lacking protein HU are UV sensitive due to a role for HU in homologous recombination. J Bacteriol 180:3750–3756
    [Google Scholar]
  45. Lucchini S., Rowley G., Goldberg M. D., Hurd D., Harrison M., Hinton J. C. D. 2006; H-NS mediates the silencing of laterally acquired genes in bacteria. PLoS Pathog 2:e81
    [Google Scholar]
  46. Lucchini S., McDermott P., Thompson A., Hinton J. C. 2009; The H-NS-like protein StpA represses the RpoS ( σ 38) regulon during exponential growth of Salmonella Typhimurium. Mol Microbiol 74:1169–1186
    [Google Scholar]
  47. Macnab R. M. 1986; Proton-driven bacterial flagellar motor. Methods Enzymol 125:563–581
    [Google Scholar]
  48. Macnab R. M. 2003; How bacteria assemble flagella. Annu Rev Microbiol 57:77–100
    [Google Scholar]
  49. Mahan M. J., Slauch J. M., Mekalanos J. J. 1993; Selection of bacterial virulence genes that are specifically induced in host tissues. Science 259:686–688
    [Google Scholar]
  50. Mangan M. W., Lucchini S., Danino V., Ó Cróinín T., Hinton D. J. C, Dorman C. J. 2006; The integration host factor (IHF) integrates stationary-phase and virulence gene expression in Salmonella enterica serovar Typhimurium. Mol Microbiol 59:1831–1847
    [Google Scholar]
  51. Marshall D. G., Sheehan B. J., Dorman C. J. 1999; A role for the leucine-responsive regulatory protein and integration host factor in the regulation of the Salmonella plasmid virulence ( spv ) locus in Salmonella typhimurium . Mol Microbiol 34:134–145
    [Google Scholar]
  52. Marshall D. G., Bowe F., Hale C., Dougan G., Dorman C. J. 2000; DNA topology and adaptation of Salmonella typhimurium to an intracellular environment. Philos Trans R Soc Lond B Biol Sci 355:565–574
    [Google Scholar]
  53. Merickel S. K., Johnson R. C. 2004; Topological analysis of Hin-catalysed DNA recombination in vivo and in vitro . Mol Microbiol 51:1143–1154
    [Google Scholar]
  54. Mills D. M., Bajaj V., Lee C. A. 1995; A 40 kb chromosomal fragment encoding Salmonella typhimurium invasion genes is absent from the corresponding region of the Escherichia coli K-12 chromosome. Mol Microbiol 15:749–759
    [Google Scholar]
  55. Navarre W. W., Porwollik S., Wang Y., McClelland M., Rosen H., Libby S. J., Fang F. C. 2006; Selective silencing of foreign DNA with low GC content by the H-NS protein in Salmonella . Science 313:236–238
    [Google Scholar]
  56. O'Byrne C. P., Dorman C. J. 1994a; The spv virulence operon of Salmonella typhimurium LT2 is regulated negatively by the cyclic AMP (cAMP)–cAMP receptor protein system. J Bacteriol 176:905–912
    [Google Scholar]
  57. O'Byrne C. P., Dorman C. J. 1994b; Transcription of the Salmonella typhimurium spv virulence locus is regulated negatively by the nucleoid-associated protein H-NS. FEMS Microbiol Lett 121:99–105
    [Google Scholar]
  58. Oberto J., Nabti S., Jooste V., Mignot H., Rouvière-Yaniv J. 2009; The HU regulon is composed of genes responding to anaerobiosis, acid stress, high osmolarity and SOS induction. PLoS ONE 4:e4367
    [Google Scholar]
  59. Patten C. L., Kirchhof M. G., Schertzberg M. R., Morton R. A., Schellhorn H. E. 2004; Microarray analysis of RpoS-mediated gene expression in Escherichia coli K-12. Mol Genet Genomics 272:580–591
    [Google Scholar]
  60. Perni S., Andrew P. W., Shama G. 2005; Estimating the maximum growth rate from microbial growth curves: definition is everything. Food Microbiol 22:491–495
    [Google Scholar]
  61. Pontiggia A., Negri A., Beltrame M., Bianchi M. E. 1993; Protein HU binds specifically to kinked DNA. Mol Microbiol 7:343–350
    [Google Scholar]
  62. Rhen M., Dorman C. J. 2005; Hierarchical gene regulators adapt Salmonella enterica to its host milieus. Int J Med Microbiol 294:487–502
    [Google Scholar]
  63. Ryan V. T., Grimwade J. E., Nievera C. J., Leonard A. C. 2002; IHF and HU stimulate assembly of pre-replication complexes at Escherichia coli oriC by two different mechanisms. Mol Microbiol 46:113–124
    [Google Scholar]
  64. Sambrook J., Russell D. 2001 Molecular Cloning: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  65. Schechter L. M., Jain S., Akbar S., Lee C. A. 2003; The small nucleoid-binding proteins H-NS, HU, and Fis affect hilA expression in Salmonella enterica serovar Typhimurium. Infect Immun 71:5432–5435
    [Google Scholar]
  66. Semsey S., Tolstorukov M. Y., Virnik K., Zhurkin V. B., Adhya S. 2004; DNA trajectory in the Gal repressosome. Genes Dev 18:1898–1907
    [Google Scholar]
  67. Shanado Y., Kato J., Ikeda H. 1998; Escherichia coli HU protein suppresses DNA-gyrase-mediated illegitimate recombination and SOS induction. Genes Cells 3:511–520
    [Google Scholar]
  68. Signon L., Kleckner N. 1995; Negative and positive regulation of Tn 10 /IS 10 -promoted recombination by IHF: two distinguishable processes inhibit transposition off of multicopy plasmid replicons and activate chromosomal events that favor evolution of new transposons. Genes Dev 9:1123–1136
    [Google Scholar]
  69. Sternberg N. L., Maurer R. 1991; Bacteriophage-mediated generalized transduction in Escherichia coli and Salmonella typhimurium . Methods Enzymol 204:18–43
    [Google Scholar]
  70. Swinger K. K., Rice P. A. 2004; IHF and HU: flexible architects of bent DNA. Curr Opin Struct Biol 14:28–35
    [Google Scholar]
  71. Uzzau S., Figueroa-Bossi N., Rubino S., Bossi L. 2001; Epitope tagging of chromosomal genes in Salmonella . Proc Natl Acad Sci U S A 98:15264–15269
    [Google Scholar]
  72. Valdivia R. H., Falkow S. 1997; Fluorescence-based isolation of bacterial genes expressed within host cells. Science 277:2007–2011
    [Google Scholar]
  73. Waterman S. R., Holden D. W. 2003; Functions and effectors of the Salmonella pathogenicity island 2 type III secretion system. Cell Microbiol 5:501–511
    [Google Scholar]
  74. Weber H., Polen T., Heuveling J., Wendisch V. F., Hengge R. 2005; Genome-wide analysis of the general stress response network in Escherichia coli : σ S-dependent genes, promoters, and sigma factor selectivity. J Bacteriol 187:1591–1603
    [Google Scholar]
  75. Wood M. W., Rosqvist R., Mullan P. B., Edwards M. H., Galyov E. E. 1996; SopE, a secreted protein of Salmonella dublin , is translocated into the target eukaryotic cell via a sip -dependent mechanism and promotes bacterial entry. Mol Microbiol 22:327–338
    [Google Scholar]
  76. Yanisch-Perron C., Vieira J., Messing J. 1985; Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene 33:103–119
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.046359-0
Loading
/content/journal/micro/10.1099/mic.0.046359-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Supplementary material 2

PDF

Supplementary material 3

PDF

Supplementary material 4

PDF

Supplementary material 5

PDF

Supplementary material 6

PDF

Supplementary material 7

PDF

Most cited Most Cited RSS feed