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. 1991 Feb;173(4):1406–1413. doi: 10.1128/jb.173.4.1406-1413.1991

Pseudomonas aeruginosa AlgB, a two-component response regulator of the NtrC family, is required for algD transcription.

D J Wozniak 1, D E Ohman 1
PMCID: PMC207277  PMID: 1899859

Abstract

Most strains of Pseudomonas aeruginosa isolated from the respiratory tracts of cystic fibrosis patients have a mucoid colony morphology due to the synthesis of an expolysaccharide called alginate. The algB gene product (AlgB) is necessary for the high-level production of alginate in mucoid P. aeruginosa. In this study, AlgB was shown to be involved in the transcription of algD, a gene previously demonstrated to be activated in mucoid P. aeruginosa. In vitro and in vivo expression studies reveal that algB encodes a protein with a molecular size of 49 kDa. The DNA sequence of a 2.2-kb P. aeruginosa fragment containing algB was also determined. The amino-terminal domain of AlgB was found to be conserved with the amino-terminal domains of the response regulator class of two-component regulatory proteins. The central domain of AlgB has sequences highly conserved with those in the NtrC subfamily of transcriptional activators (NtrC, NifA, HydG, DctD, FlbD, TyrR, and PgtA). The central domain of AlgB also contains a potential nucleotide binding site. AlgB is the first NtrC homolog described from P. aeruginosa. At the carboxy terminus of AlgB, a helix-turn-helix motif was observed, suggesting that AlgB is a DNA-binding protein. The strongly conserved NtrC-like central domain of AlgB is not present in AlgR, another alginate response regulator. This study therefore identifies and characterizes the second of at least two unique response regulators used by P. aeruginosa to control alginate gene expression.

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Selected References

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  1. Baltimore R. S., Mitchell M. Immunologic investigations of mucoid strains of Pseudomonas aeruginosa: comparison of susceptibility to opsonic antibody in mucoid and nonmucoid strains. J Infect Dis. 1980 Feb;141(2):238–247. doi: 10.1093/infdis/141.2.238. [DOI] [PubMed] [Google Scholar]
  2. Berry A., DeVault J. D., Chakrabarty A. M. High osmolarity is a signal for enhanced algD transcription in mucoid and nonmucoid Pseudomonas aeruginosa strains. J Bacteriol. 1989 May;171(5):2312–2317. doi: 10.1128/jb.171.5.2312-2317.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Buck M. Deletion analysis of the Klebsiella pneumoniae nitrogenase promoter: importance of spacing between conserved sequences around positions -12 and -24 for activation by the nifA and ntrC (glnG) products. J Bacteriol. 1986 May;166(2):545–551. doi: 10.1128/jb.166.2.545-551.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chitnis C. E., Ohman D. E. Cloning of Pseudomonas aeruginosa algG, which controls alginate structure. J Bacteriol. 1990 Jun;172(6):2894–2900. doi: 10.1128/jb.172.6.2894-2900.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Darzins A., Chakrabarty A. M. Cloning of genes controlling alginate biosynthesis from a mucoid cystic fibrosis isolate of Pseudomonas aeruginosa. J Bacteriol. 1984 Jul;159(1):9–18. doi: 10.1128/jb.159.1.9-18.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Darzins A., Frantz B., Vanags R. I., Chakrabarty A. M. Nucleotide sequence analysis of the phosphomannose isomerase gene (pmi) of Pseudomonas aeruginosa and comparison with the corresponding Escherichia coli gene manA. Gene. 1986;42(3):293–302. doi: 10.1016/0378-1119(86)90233-7. [DOI] [PubMed] [Google Scholar]
  9. Darzins A., Wang S. K., Vanags R. I., Chakrabarty A. M. Clustering of mutations affecting alginic acid biosynthesis in mucoid Pseudomonas aeruginosa. J Bacteriol. 1985 Nov;164(2):516–524. doi: 10.1128/jb.164.2.516-524.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. DeVault J. D., Kimbara K., Chakrabarty A. M. Pulmonary dehydration and infection in cystic fibrosis: evidence that ethanol activates alginate gene expression and induction of mucoidy in Pseudomonas aeruginosa. Mol Microbiol. 1990 May;4(5):737–745. doi: 10.1111/j.1365-2958.1990.tb00644.x. [DOI] [PubMed] [Google Scholar]
  11. Deretic V., Dikshit R., Konyecsni W. M., Chakrabarty A. M., Misra T. K. The algR gene, which regulates mucoidy in Pseudomonas aeruginosa, belongs to a class of environmentally responsive genes. J Bacteriol. 1989 Mar;171(3):1278–1283. doi: 10.1128/jb.171.3.1278-1283.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Deretic V., Gill J. F., Chakrabarty A. M. Gene algD coding for GDPmannose dehydrogenase is transcriptionally activated in mucoid Pseudomonas aeruginosa. J Bacteriol. 1987 Jan;169(1):351–358. doi: 10.1128/jb.169.1.351-358.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Deretic V., Gill J. F., Chakrabarty A. M. Pseudomonas aeruginosa infection in cystic fibrosis: nucleotide sequence and transcriptional regulation of the algD gene. Nucleic Acids Res. 1987 Jun 11;15(11):4567–4581. doi: 10.1093/nar/15.11.4567. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Deretic V., Govan J. R., Konyecsni W. M., Martin D. W. Mucoid Pseudomonas aeruginosa in cystic fibrosis: mutations in the muc loci affect transcription of the algR and algD genes in response to environmental stimuli. Mol Microbiol. 1990 Feb;4(2):189–196. doi: 10.1111/j.1365-2958.1990.tb00586.x. [DOI] [PubMed] [Google Scholar]
  15. Deretic V., Konyecsni W. M. Control of mucoidy in Pseudomonas aeruginosa: transcriptional regulation of algR and identification of the second regulatory gene, algQ. J Bacteriol. 1989 Jul;171(7):3680–3688. doi: 10.1128/jb.171.7.3680-3688.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ditta G., Schmidhauser T., Yakobson E., Lu P., Liang X. W., Finlay D. R., Guiney D., Helinski D. R. Plasmids related to the broad host range vector, pRK290, useful for gene cloning and for monitoring gene expression. Plasmid. 1985 Mar;13(2):149–153. doi: 10.1016/0147-619x(85)90068-x. [DOI] [PubMed] [Google Scholar]
  17. Drummond M., Whitty P., Wootton J. Sequence and domain relationships of ntrC and nifA from Klebsiella pneumoniae: homologies to other regulatory proteins. EMBO J. 1986 Feb;5(2):441–447. doi: 10.1002/j.1460-2075.1986.tb04230.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Evans L. R., Linker A. Production and characterization of the slime polysaccharide of Pseudomonas aeruginosa. J Bacteriol. 1973 Nov;116(2):915–924. doi: 10.1128/jb.116.2.915-924.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Figurski D. H., Helinski D. R. Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1648–1652. doi: 10.1073/pnas.76.4.1648. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Flynn J. L., Ohman D. E. Cloning of genes from mucoid Pseudomonas aeruginosa which control spontaneous conversion to the alginate production phenotype. J Bacteriol. 1988 Apr;170(4):1452–1460. doi: 10.1128/jb.170.4.1452-1460.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Flynn J. L., Ohman D. E. Use of a gene replacement cosmid vector for cloning alginate conversion genes from mucoid and nonmucoid Pseudomonas aeruginosa strains: algS controls expression of algT. J Bacteriol. 1988 Jul;170(7):3228–3236. doi: 10.1128/jb.170.7.3228-3236.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Friedman A. M., Long S. R., Brown S. E., Buikema W. J., Ausubel F. M. Construction of a broad host range cosmid cloning vector and its use in the genetic analysis of Rhizobium mutants. Gene. 1982 Jun;18(3):289–296. doi: 10.1016/0378-1119(82)90167-6. [DOI] [PubMed] [Google Scholar]
  23. Fry D. C., Kuby S. A., Mildvan A. S. ATP-binding site of adenylate kinase: mechanistic implications of its homology with ras-encoded p21, F1-ATPase, and other nucleotide-binding proteins. Proc Natl Acad Sci U S A. 1986 Feb;83(4):907–911. doi: 10.1073/pnas.83.4.907. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. George R. H. Pseudomonas infections in cystic fibrosis. Arch Dis Child. 1987 May;62(5):438–439. doi: 10.1136/adc.62.5.438. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Goldberg J. B., Ohman D. E. Cloning and expression in Pseudomonas aeruginosa of a gene involved in the production of alginate. J Bacteriol. 1984 Jun;158(3):1115–1121. doi: 10.1128/jb.158.3.1115-1121.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Goldberg J. B., Ohman D. E. Construction and characterization of Pseudomonas aeruginosa algB mutants: role of algB in high-level production of alginate. J Bacteriol. 1987 Apr;169(4):1593–1602. doi: 10.1128/jb.169.4.1593-1602.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Govan J. R., Harris G. S. Pseudomonas aeruginosa and cystic fibrosis: unusual bacterial adaptation and pathogenesis. Microbiol Sci. 1986 Oct;3(10):302–308. [PubMed] [Google Scholar]
  28. Gray G. L., Smith D. H., Baldridge J. S., Harkins R. N., Vasil M. L., Chen E. Y., Heyneker H. L. Cloning, nucleotide sequence, and expression in Escherichia coli of the exotoxin A structural gene of Pseudomonas aeruginosa. Proc Natl Acad Sci U S A. 1984 May;81(9):2645–2649. doi: 10.1073/pnas.81.9.2645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ishimoto K. S., Lory S. Formation of pilin in Pseudomonas aeruginosa requires the alternative sigma factor (RpoN) of RNA polymerase. Proc Natl Acad Sci U S A. 1989 Mar;86(6):1954–1957. doi: 10.1073/pnas.86.6.1954. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kato J., Chu L., Kitano K., DeVault J. D., Kimbara K., Chakrabarty A. M., Misra T. K. Nucleotide sequence of a regulatory region controlling alginate synthesis in Pseudomonas aeruginosa: characterization of the algR2 gene. Gene. 1989 Dec 7;84(1):31–38. doi: 10.1016/0378-1119(89)90136-4. [DOI] [PubMed] [Google Scholar]
  31. Kato J., Misra T. K., Chakrabarty A. M. AlgR3, a protein resembling eukaryotic histone H1, regulates alginate synthesis in Pseudomonas aeruginosa. Proc Natl Acad Sci U S A. 1990 Apr;87(8):2887–2891. doi: 10.1073/pnas.87.8.2887. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Kimbara K., Chakrabarty A. M. Control of alginate synthesis in Pseudomonas aeruginosa: regulation of the algR1 gene. Biochem Biophys Res Commun. 1989 Oct 31;164(2):601–608. doi: 10.1016/0006-291x(89)91502-7. [DOI] [PubMed] [Google Scholar]
  33. Konyecsni W. M., Deretic V. DNA sequence and expression analysis of algP and algQ, components of the multigene system transcriptionally regulating mucoidy in Pseudomonas aeruginosa: algP contains multiple direct repeats. J Bacteriol. 1990 May;172(5):2511–2520. doi: 10.1128/jb.172.5.2511-2520.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Kustu S., Santero E., Keener J., Popham D., Weiss D. Expression of sigma 54 (ntrA)-dependent genes is probably united by a common mechanism. Microbiol Rev. 1989 Sep;53(3):367–376. doi: 10.1128/mr.53.3.367-376.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  36. Linker A., Jones R. S. A new polysaccharide resembling alginic acid isolated from pseudomonads. J Biol Chem. 1966 Aug 25;241(16):3845–3851. [PubMed] [Google Scholar]
  37. Marcus H., Baker N. R. Quantitation of adherence of mucoid and nonmucoid Pseudomonas aeruginosa to hamster tracheal epithelium. Infect Immun. 1985 Mar;47(3):723–729. doi: 10.1128/iai.47.3.723-729.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Miller J. F., Mekalanos J. J., Falkow S. Coordinate regulation and sensory transduction in the control of bacterial virulence. Science. 1989 Feb 17;243(4893):916–922. doi: 10.1126/science.2537530. [DOI] [PubMed] [Google Scholar]
  39. Mizusawa S., Nishimura S., Seela F. Improvement of the dideoxy chain termination method of DNA sequencing by use of deoxy-7-deazaguanosine triphosphate in place of dGTP. Nucleic Acids Res. 1986 Feb 11;14(3):1319–1324. doi: 10.1093/nar/14.3.1319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Pabo C. O., Sauer R. T. Protein-DNA recognition. Annu Rev Biochem. 1984;53:293–321. doi: 10.1146/annurev.bi.53.070184.001453. [DOI] [PubMed] [Google Scholar]
  41. Ramphal R., Pier G. B. Role of Pseudomonas aeruginosa mucoid exopolysaccharide in adherence to tracheal cells. Infect Immun. 1985 Jan;47(1):1–4. doi: 10.1128/iai.47.1.1-4.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Ronson C. W., Astwood P. M., Nixon B. T., Ausubel F. M. Deduced products of C4-dicarboxylate transport regulatory genes of Rhizobium leguminosarum are homologous to nitrogen regulatory gene products. Nucleic Acids Res. 1987 Oct 12;15(19):7921–7934. doi: 10.1093/nar/15.19.7921. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Ronson C. W., Nixon B. T., Ausubel F. M. Conserved domains in bacterial regulatory proteins that respond to environmental stimuli. Cell. 1987 Jun 5;49(5):579–581. doi: 10.1016/0092-8674(87)90530-7. [DOI] [PubMed] [Google Scholar]
  44. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Schwarzmann S., Boring J. R. Antiphagocytic Effect of Slime from a Mucoid Strain of Pseudomonas aeruginosa. Infect Immun. 1971 Jun;3(6):762–767. doi: 10.1128/iai.3.6.762-767.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Stock A., Koshland D. E., Jr, Stock J. Homologies between the Salmonella typhimurium CheY protein and proteins involved in the regulation of chemotaxis, membrane protein synthesis, and sporulation. Proc Natl Acad Sci U S A. 1985 Dec;82(23):7989–7993. doi: 10.1073/pnas.82.23.7989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Stock J. B., Ninfa A. J., Stock A. M. Protein phosphorylation and regulation of adaptive responses in bacteria. Microbiol Rev. 1989 Dec;53(4):450–490. doi: 10.1128/mr.53.4.450-490.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Totten P. A., Lara J. C., Lory S. The rpoN gene product of Pseudomonas aeruginosa is required for expression of diverse genes, including the flagellin gene. J Bacteriol. 1990 Jan;172(1):389–396. doi: 10.1128/jb.172.1.389-396.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. VOGEL H. J., BONNER D. M. Acetylornithinase of Escherichia coli: partial purification and some properties. J Biol Chem. 1956 Jan;218(1):97–106. [PubMed] [Google Scholar]
  50. West S. E., Iglewski B. H. Codon usage in Pseudomonas aeruginosa. Nucleic Acids Res. 1988 Oct 11;16(19):9323–9335. doi: 10.1093/nar/16.19.9323. [DOI] [PMC free article] [PubMed] [Google Scholar]

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