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Volume 148, Issue 12

O-antigen expression in serovar Typhi is regulated by nitrogen availability through RpoN-mediated transcriptional control of the gene Free

Abstract

The authors previously reported increased expression of the serovar Typhi () gene when the bacterial cells reach stationary phase. In this study, using a fusion to the promoter region, they demonstrate that growth-dependent regulation of expression occurs at the level of transcription initiation. It was also observed that production of the lipopolysaccharide (LPS) O-antigen by Ty2 correlated with the differential expression of during bacterial growth. This was probably due to the increased cellular levels of RfaH, since expression of the distal gene in the O-antigen gene cluster of Ty2, , was also increased during stationary growth, as demonstrated by RT-PCR analysis. Examination of the sequences upstream of the coding region revealed homologies to potential binding sites for the RcsB/RcsA dimer of the RcsC/YopJ/RcsB phosphorelay regulatory system and for the RpoN alternative sigma factor. The expression of the gene in and mutants of Ty2 was measured. The results indicate that inactivation of , but not of , suppresses the growth-phase-dependent induction of expression. Furthermore, production of β-galactosidase mediated by the fusion increased approximately fourfold when bacteria were grown in a nitrogen-limited medium. Nitrogen limitation was also shown to increase the expression of the O-antigen by the wild-type Ty2, as demonstrated by a similar electrophoretic profile to that observed during the stationary phase of growth in rich media. It is therefore concluded that the relationship between LPS production and nitrogen limitation parallels the pattern of regulation under the control of RpoN and is consistent with the idea that RpoN modulates LPS formation via its effect on gene expression during bacterial growth.

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Keyword(s): lipopolysaccharide , regulation , sigma factor and transcription
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2002-12-01
2024-04-26
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References

  1. Aguilar A., Merino S., Rubires X., Tomas J. M. 1997; Influence of osmolarity on lipopolysaccharide and virulence of Aeromonas hydrophila serotype O: 34 strains grown at 37 °C. Infect Immun 65:1245–1250
     [Google Scholar]
  2. Al-Hendy A., Toivanen P., Skurnik M. 1991; The effect of growth temperature on the biosynthesis of Yersinia enterocolitica O: 3 lipopolysaccharide: temperature regulates the transcription of the rfb but not the rfa region. Microb Pathog 10:81–86 [CrossRef]
     [Google Scholar]
  3. Arricau N., Hermant D., Waxin H., Echobicon C., Duffey P. S., Popoff M. Y. 1998; The RcsB and RcsC regulatory system of Salmonella typhi differentially regulates the expression of invasion proteins, flagellin and Vi antigen in response to osmolarity. Mol Microbiol 29:835–850 [CrossRef]
     [Google Scholar]
  4. Artsimovitch I., Landick R. 2000; Pausing by bacterial polymerase is mediated by mechanistically distinct classes of signals. Proc Natl Acad Sci USA 97:7090–7095 [CrossRef]
     [Google Scholar]
  5. Artsimovitch I., Landick R. 2002; The transcriptional regulator RfaH stimulates RNA chain elongation after recruitment to elongation complexes by the exposed nontemplate DNA strand. Cell 109:193–203 [CrossRef]
     [Google Scholar]
  6. Bailey M. J. A., Koronakis V., Schmoll T., Hughes C. 1992; Escherichia coli HlyT protein, a transcriptional activator of haemolysin synthesis and secretion, is encoded by the rfaH ( sfrB) locus required for expression of sex factor and lipopolysaccharide genes. Mol Microbiol 6:1003–1012 [CrossRef]
     [Google Scholar]
  7. Bailey M. J. A., Hughes C., Koronakis V. 1996; Increased distal gene transcription by the elongation factor rfaH , a specialized homologue of NusG. Mol Microbiol 22:729–737 [CrossRef]
     [Google Scholar]
  8. Bailey M. J. A., Hughes C., Koronakis V. 1997; RfaH and the ops element, components of a novel system controlling bacterial transcription elongation. Mol Microbiol 26:845–851 [CrossRef]
     [Google Scholar]
  9. Beutin L., Achtman M. 1979; Two Escherichia coli chromosomal cistrons, sfrA and sfrB , which are needed for expression of F factor tra functions. J Bacteriol 139:730–737
     [Google Scholar]
  10. Boucher J. C., Schurr M. J., Deretic V. 2000; Dual regulation of mucoidy in Pseudomonas aeruginosa and sigma factor antagonism. Mol Microbiol 36:341–351 [CrossRef]
     [Google Scholar]
  11. Buck M., Gallegos M.-T., Studholme D. J., Guo Y., Gralla J. D. 2000; The bacterial enhancer-dependent σ54 (σN) transcription factor. J Bacteriol 182:4129–4136 [CrossRef]
     [Google Scholar]
  12. Datsenko K. A., Wanner B. L. 2000; One-step inactivation of chromosomal genes in Escherichia coli K12 using PCR products. Proc Natl Acad Sci USA 97:6640–6645 [CrossRef]
     [Google Scholar]
  13. Ernst R. K., Guina T., Miller S. I. 2001; Salmonella typhimurium outer membrane remodelling: role in resistance to host innate immunity. Microbes Infect 3:1327–1330 [CrossRef]
     [Google Scholar]
  14. Farewell A., Brazas R., Davie E., Mason J., Rothfield L. I. 1991; Suppression of the abnormal phenotype of Salmonella typhimurium rfaH mutants by mutations in the genes for transcription termination factor Rho. J Bacteriol 173:5188–5193
     [Google Scholar]
  15. Gunn J. S., Miller S. I. 1996; PhoP/PhoQ activates transcription of pmrA/B , encoding a two-component system involved in Salmonella typhimurium antimicrobial peptide resistance. J Bacteriol 178:6857–6864
     [Google Scholar]
  16. Gunn J. S., Lim K. B., Krueger J., Kim K., Guo L., Hackett M., Miller S. I. 1998; PmrA-PmrB-regulated genes necessary for 4-aminoarabinose lipid A modification and polymixin resistance. Mol Microbiol 27:1171–1182 [CrossRef]
     [Google Scholar]
  17. Guo L., Lim K. B., Gunn J. S., Bainbridge B., Darveau R. P., Hackett M., Miller S. I. 1997; Regulation of lipid A modifications by Salmonella typhimurium virulence genes phoP-phoQ. Science 276:250–253
     [Google Scholar]
  18. Hitchcock P. J., Brown T. M. 1983; Morphological heterogeneity among Salmonella lipopolysaccharide chemotypes in silver stained polyacrylamide gels. J Bacteriol 154:269–277
     [Google Scholar]
  19. Hobbs M., Reeves P. R. 1994; The JUMPstart sequence: a 39 bp element common to several polysaccharide gene clusters. Mol Microbiol 12:855–856 [CrossRef]
     [Google Scholar]
  20. Ishimoto K. S., Lory S. 1989; Formation of pilin in Pseudomonas aeruginosa requires the alternative σ factor (RpoN) subunit of RNA polymerase. Proc Natl Acad Sci USA 86:1954–1957 [CrossRef]
     [Google Scholar]
  21. Joiner K. A. 1988; Complement evasion by bacteria and parasites. Annu Rev Microbiol 42:201–230 [CrossRef]
     [Google Scholar]
  22. Jones B. D., Falkow S. 1996; Salmonellosis: host immune responses and bacterial virulence determinants. Annu Rev Immunol 14:533–561 [CrossRef]
     [Google Scholar]
  23. Karow M., Raina S., Georgopoulos C., Fayet O. 1991; Complex phenotypes of null mutations in the htr genes, whose products are essential for Escherichia coli growth at elevated temperatures. Res Microbiol 142:289–294 [CrossRef]
     [Google Scholar]
  24. Klimpel K. W., Lesley S. A., Clark V. L. 1989; Identification of subunits of gonococcal RNA polymerase by immunoblot analysis: evidence of multiple sigma factors. J Bacteriol 171:3713–3718
     [Google Scholar]
  25. Klose K. E., Mekalanos J. J. 1998; Distinct roles of an alternative sigma factor during both free-swimming and colonizing phases of the Vibrio cholerae pathogenic cycle. Mol Microbiol 28:501–520 [CrossRef]
     [Google Scholar]
  26. Koop A. H., Hartley M. E., Bourgeois S. 1987; A low-copy-number vector utilizing β-galactosidase for the analysis of gene control elements. Gene 52:245–256 [CrossRef]
     [Google Scholar]
  27. Kossack R. E., Guerrant R. L., Densen P., Schadelin J., Mandell G. L. 1981; Diminished neutrophil oxidative metabolism after phagocytocis of virulent Salmonella typhi . Infect Immun 31:674–678
     [Google Scholar]
  28. Leeds J. A., Welch R. A. 1996; RfaH enhances elongation of Escherichia coli hlyCABD mRNA. J Bacteriol 178:1850–1857
     [Google Scholar]
  29. Leeds J. A., Welch R. A. 1997; Enhancing transcription through the Escherichia coli hemolysin operon, hlyCABD : RfaH and upstream JUMP-start DNA sequence function together via a postinitiation mechanism. J Bacteriol 179:3519–3527
     [Google Scholar]
  30. Lesse A. J., Campagnari A. A., Bittner W. E., Apicella M. A. 1990; Increased resolution of lipopolysaccharides and lipooligosaccharides utilizing tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis. J Immunol Methods 126:109–117 [CrossRef]
     [Google Scholar]
  31. Lyczak J. B., Zaidi T. S., Grout M., Bittner W. M., Contreras I., Pier G. B. 2001; Epithelial cell contact-induced alterations in Salmonella enterica serovar Typhi lipopolysaccharide are critical for bacterial internalization. Cell Microbiol 3:763–772 [CrossRef]
     [Google Scholar]
  32. Marolda C. L., Valvano M. A. 1998; Promoter region of the Escherichia coli O: 7-specific lipopolysaccharide gene cluster: structural and functional characterization of an upstream untranslated mRNA sequence. J Bacteriol 180:3070–3079
     [Google Scholar]
  33. Merrick J. M. 1993; In a class of its own – the RNA polymerase sigma factor σN (σ54 . Mol Microbiol 10:903–909 [CrossRef]
     [Google Scholar]
  34. Miller J. H. 1972 Experiments in Molecular Genetics Cold Spring Harbor, NY: . Cold Spring Harbor Laboratory;
     [Google Scholar]
  35. Nagy G., Dobrindt U., Kupfer M., Emody L., Karch H., Hacker J. 2001; Expression of hemin receptor molecule ChuA is influenced by RfaH in uropathogenic Escherichia coli 536. Infect Immun 69:1924–1928 [CrossRef]
     [Google Scholar]
  36. Nieto J. M., Bailey J. A. M., Hughes C., Koronakis V. 1996; Suppression of transcription polarity in the Escherichia coli haemolysin operon by a short upstream element shared by polysaccharide and DNA transfer determinants. Mol Microbiol 19:705–713 [CrossRef]
     [Google Scholar]
  37. Pang T., Levine M. M., Ivanoff B., Wain J., Finlay B. B. 1998; Typhoid fever – important issues still remain. Trends Microbiol 6:131–133 [CrossRef]
     [Google Scholar]
  38. Parkhill J., Dougan G., James K. D. 38 other authors 2001; The complete genome sequence of a multidrug resistant Salmonella enterica serovar Typhi CT18. Nature 413:848–852 [CrossRef]
     [Google Scholar]
  39. Pier G., Grout M., Zaidi T., Meluleni G., Mueschenborn S. S., Banting G., Ratcliff R., Evans M. J., Colledge W. H. 1998; Salmonella typhi uses CFTR to enter intestinal epithelial cells. Nature 393:79–82 [CrossRef]
     [Google Scholar]
  40. Pradel E., Schnaitman C. A. 1991; Effect of rfaH ( sfrB) and temperature on expression of rfa genes of Escherichia coli K-12. J Bacteriol 173:6428–6431
     [Google Scholar]
  41. Raetz C. R. others 1996; Bacterial lipopolysaccharides: a remarkable family of bioactive macroamphiphiles. In Escherichia coli and Salmonella typhimurium: Cellular and Molecular Biology pp 1035–1063 Edited by Neidhardt F. C. Washington, DC: American Society for Microbiology;
     [Google Scholar]
  42. Reeves P. R. 1993; Evolution of Salmonella O antigen variation by interspecific gene transfer on a large scale. Trends Genet 9:17–22 [CrossRef]
     [Google Scholar]
  43. Rojas G., Saldı́as S., Bittner M., Zaldı́var M., Contreras I. 2001; The rfaH gene, which affects lipopolysaccharide synthesis in Salmonella enterica Serovar Typhi, is differentially expressed during the bacterial growth phase. FEMS Microbiol Lett 204:123–128 [CrossRef]
     [Google Scholar]
  44. Rosenberger C. M., Scott M. G., Gold M. R., Hancock R. E. W., Finlay B. B. 2000; Salmonella typhimurium infection and lipopolysaccharide stimulation induce similar changes in macrophage gene expression. J Immunol 164:5894–5904 [CrossRef]
     [Google Scholar]
  45. Schnaitman C. A., Klena J. D. 1993; Genetics of lipopolysaccharide biosynthesis in enteric bacteria. Microbiol Rev 57:655–682
     [Google Scholar]
  46. Shingler V. 1996; Signal sensing by sigma 54-dependent regulators: derepression as a control mechanism. Mol Microbiol 19:409–416 [CrossRef]
     [Google Scholar]
  47. Sledjeski D. D., Gottesman S. 1996; Osmotic shock induction of capsule synthesis in Escherichia coli K-12. J Bacteriol 178:1204–1206
     [Google Scholar]
  48. Stevens M., Clarke B., Roberts I. 1997; Regulation of the Escherichia coli K5 capsule gene cluster by transcription antitermination. Mol Microbiol 24:1001–1012 [CrossRef]
     [Google Scholar]
  49. Stout V., Gottesman S. 1990; RcsB and RcsC: a two component regulator of capsular synthesis in Escherichia coli . J Bacteriol 172:659–669
     [Google Scholar]
  50. Takeda S., Fujisawa Y., Matsubara M., Aiba H., Mizuno T. 2001; A novel feature of the multistep phosphorelay in Escherichia coli : a revised model of the RcsC→YojN→RcsB signalling pathway implicated in capsular synthesis and swarming behaviour. Mol Microbiol 40:440–450 [CrossRef]
     [Google Scholar]
  51. Tsai C. M., Frasch C. E. 1982; A sensitive silver stain for detecting lipopolysaccharide in polyacrylamide gels. Anal Biochem 119:115–119 [CrossRef]
     [Google Scholar]
  52. Virlogeux I., Waxin H., Ecobichon C., Lee J. O., Popoff M. Y. 1996; Characterization of the rcsA and rcsB genes from Salmonella typhi : rcsB through tviA is involved in regulation of Vi antigen synthesis. J Bacteriol 178:1691–1698
     [Google Scholar]
  53. Wang L., Gralla J. D. 1998; Multiple in vivo roles for the −12-region elements of sigma 54 promoters. J Bacteriol 180:5626–5631
     [Google Scholar]
  54. Wang L., Reeves P. R. 1994; Involvement of the galactosyl-1-phosphate transferase encoded by the Salmonella enterica rfbP gene in O-antigen subunit processing. J Bacteriol 176:4348–4356
     [Google Scholar]
  55. Wang L., Liu D., Reeves P. R. 1996; C-terminal half of Salmonella enterica WbaP (RfbP) is the galactosyl-1-phosphate transferase domain catalyzing the first step of O-antigen synthesis. J Bacteriol 178:2598–2604
     [Google Scholar]
  56. Wang L., Jensen S., Hallman R., Reeves P. R. 1998; Expression of the O antigen gene cluster is regulated by RfaH through the JUMPstart sequence. FEMS Microbiol Lett 165:201–206 [CrossRef]
     [Google Scholar]
  57. Wehland M., Bernhard F. 2000; The RcsAB box. Characterization of a new operator essential for the regulation of exopolysaccharide biosynthesis in enteric bacteria. J Biol Chem 275:7013–7020 [CrossRef]
     [Google Scholar]
  58. Whitfield C., Roberts I. S. 1999; Structure, assembly and regulation of expression of capsules in Escherichia coli . Mol Microbiol 31:1307–1319 [CrossRef]
     [Google Scholar]
  59. Zhao H., Li X., Johnson D. E., Mobley L. T. 1999; Identification of protease and rpoN -associated genes of uropathogenic Proteus mirabilis by negative selection in a mouse model of ascending urinary tract infection. Microbiology 145:185–195 [CrossRef]
     [Google Scholar]
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O-antigen expression in Salmonella enterica serovar Typhi is regulated by nitrogen availability through RpoN-mediated transcriptional control of the rfaH gene
Microbiology 148, 3789 (2002); https://doi.org/10.1099/00221287-148-12-3789
/content/journal/micro/10.1099/00221287-148-12-3789
/content/journal/micro/10.1099/00221287-148-12-3789
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