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Pseudomonas aeruginosa promoters which contain a conserved GG-N10-GC motif but appear to be RpoN-independent

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Abstract

The proC gene of Pseudomonas aeruginosa encodes the constitutive Δ1-pyrroline 5-carboxylate reductase (the third enzyme of proline biosynthesis) and ranks among the numerous Pseudomonas genes which are poorly transcribed in Escherichia coli. The promoters of the proC gene were located by deletion mapping. The 5′ ends of the proC transcripts originating from one promoter were determined by primer extension. This promoter has a GG-N10-GC motif with a 16 by spacing between the GC doublet and the transcription start site. Such spacing is unusually long for σ54-dependent promoters. In rpoN mutants of P. aeruginosa and P. putida a proC′-′lacZ fusion was expressed at wild-type levels, suggesting that σ54 RNA polymerase is not involved in proC transcription. The expression of another P. aeruginosa gene, anr (for anaerobic regulation of nitrate respiration and anaerobic arginine degradation), also appeared to be independent of RpoN in Pseudomonas and occurred at a very low level in E. coli. The proC and anr promoters have sequence similarities in addition to the conserved GG-N10-GC motif and may also be related to some alg (alginate) promoters of P. aeruginosa. We propose that the proC and am promoters are activated by proteins, including perhaps an alternative σ factor, which are present in Pseudomonas but absent from E. coli.

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References

  • Allan B, Kropinski AM (1987) DNA-dependent RNA polymerase from Pseudomonas aeruginosa. Biochem Cell Biol 65:776–782

    Google Scholar 

  • Allan B, Linsemann M, MacDonald LA, Lam JS, Kropinski AM (1988) Heat shock response of Pseudomonas aeruginosa. J Bacteriol 170:3668–3674

    Google Scholar 

  • Bagdasarian MM, Amann E, Lurz R, Rückert B, Bagdasarian M (1983) Activity of the hybrid trp-lac (tac) promoter of Escherichia coli in Pseudomonas putida. Construction of broad-host-range, controlled-expression vectors. Gene 26:273–282

    Google Scholar 

  • Buckel P, Zehelein E (1981) Expression of Pseudomonas fluorescens d-galactose dehydrogenase in E. coli}. Gene 16:149–159

    Google Scholar 

  • Chung CT, Miller RH (1988) A rapid and convenient method for the preparation and storage of competent bacterial cells. Nucleic Acids Res 16:3580

    Google Scholar 

  • Coppard JR, Merrick MJ (1991) Cassette mutagenesis implicates a helix-turn-helix motif in promoter recognition by the novel RNA polymerase sigma factor σ54. Mot Microbiol 5:1309–1317

    Google Scholar 

  • Del Sal G, Manfioletti G, Schneider C (1988) A one tube plasmid DNA mini-preparation suitable for sequencing. Nucleic Acids Res 16:9878

    Google Scholar 

  • Deretic V, Konyecsni WM (1989) Control of mucoidy in Pseudomonas aeruginosa: transcriptional regulation of algR and identification of the second regulatory gene, algQ. J Bacteriol 171:3680–3688

    Google Scholar 

  • Deretic V, Konyecsni WM, Mohr CD, Martin DW, Hibler NS (1989) Common denominators of promoter control in Pseudomonas and other bacteria. Bio/Technology 7:1249–1254

    Google Scholar 

  • Deretic V, Mohr CD, Martin DW (1991) Mucoid Pseudomonas aeruginosa in cystic fibrosis: signal transduction and histone-like elements in the regulation of bacterial virulence. Mol Microbiol 5:1577–1583

    Google Scholar 

  • Fellay R, Frey J, Krisch H (1987) Interposon mutagenesis of soil and water bacteria: a family of DNA fragments designed for in vitro insertional mutagenesis of Gram-negative bacteria. Gene 52:147–154

    Google Scholar 

  • Galimand M, Gamper M, Zimmermann A, Haas D (1991) Positive FNR-like control of anaerobic arginine degradation and nitrate respiration in Pseudomonas aeruginosa. J Bacteriol 173:1598–1606

    Google Scholar 

  • Gao J, Gussin GN (1991) RNA polymerases from Pseudomonas aeruginosa and Pseudomonas syringae respond to Escherichia coli activator proteins. J Bacteriol 173:394–397

    Google Scholar 

  • Gray GL, Smith DH, Baldridge JS, Harkins RN, Vasil ML, Chen EY, Heynecker HL (1984) Cloning, nucleotide sequence, and expression in Escherichia coli of the exotoxin A structural gene of Pseudomonas aeruginosa. Proc Natl Acad Sci USA 81:2645–2649

    Google Scholar 

  • Greener A, Lehman SM, Helinski DR (1992) Promoters of the broad host range plasmid RK2: analysis of transcription (initiation) in five species of gram-negative bacteria. Genetics 130:27–36

    Google Scholar 

  • Guzzo J, Murgier M, Filloux A, Lazdunski A (1990) Cloning of the Pseudomonas aeruginosa alkaline protease gene and secretion of the protease into the medium by Escherichia coli. J Bacteriol 172:942–948

    Google Scholar 

  • Haas D, Holloway BW (1976) R factor variants with enhanced sex factor activity in Pseudomonas aeruginosa. Mot Gen Genet 144:243–251

    Google Scholar 

  • Han C-Y, Crawford IP, Harwood CS (1991) Up-promoter mutations in the trpBA operon of Pseudomonas aeruginosa. J Bacteriol 173:3756–3762

    Google Scholar 

  • Hennecke H, Günther I, Binder F (1982) A novel cloning vector for the direct selection of recombinant DNA in E. coli. Gene 19:231–234

    Google Scholar 

  • Holloway BW (1969) Genetics of Pseudomonas. Bacteriol Rev 33:419–443

    Google Scholar 

  • Hopwood DA, Bibb MJ, Chater KF, Kieser T, Bruton CJ, Kieser HM, Lydiate DJ, Smith CP, Ward JM, Schrempf H (1985) Genetic manipulation of Streptomyces. A laboratory manual. John Innes foundation, Norwich (UK)

    Google Scholar 

  • Ishimoto KS, Lory S (1989) Formation of pilin in Pseudomonas aeruginosa requires the alternative a factor (RpoN) of RNA polymerase. Proc Natl Acad Sci USA 86:1954–1957

    Google Scholar 

  • Itoh Y, Matsumoto H (1992) Mutations affecting regulation of the anabolic argF and the catabolic aru genes in Pseudomonas aeruginosa PAO. Mol Gen Genet 231:417–425

    Google Scholar 

  • Itoh Y, Watson JM, Haas D, Leisinger T (1984) Genetic and molecular characterization of the Pseudomonas plasmid pVSI. Plasmid 11:206–220

    Google Scholar 

  • Itoh Y, Soldati L, Stalon V, Falmagne P, Terawaki Y, Leisinger T, Haas D (1988) Anabolic ornithine carbamoyltransferase of Pseudomonas aeruginosa: nucleotide sequence and transcriptional control of the argF structural gene. J Bacteriol 170:2725–2734

    Google Scholar 

  • Jeenes DJ, Soldati L, Baur H, Watson JM, Mercenier A, Reimmann C, Leisinger T, Haas D (1986) Expression of biosynthetic genes from Pseudomonas aeruginosa and Escherichia coli in the heterologous host. Mol Gen Genet 203:421–429

    Google Scholar 

  • Kimbara K, Chakrabarty AM (1989) Control of alginate synthesis in Pseudomonas aeruginosa: regulation of the algR1 gene. Biochem Biophys Res Commun 164:601–608

    Google Scholar 

  • Köhler T, Harayama S, Ramos J-L, Timmis KN (1989) Involvement of Pseudomonas putida RpoN σ factor in regulation of various metabolic functions. J Bacteriol 171:4326–4333

    Google Scholar 

  • Kornitzer D, Teff D, Altuvia S, Oppenheim AB (1991) Isolation, characterization, and sequence of an Escherichia coli heat shock gene, htpX. J Bacteriol 173:2944–2953

    Google Scholar 

  • Krishna RV, Beilstein V, Leisinger T (1979) Biosynthesis of proline in Pseudomonas aeruginosa. Properties of γ-glutamyl phosphate reductase and 1-pyrroline-5-carboxylate reductase. Biochem J 181:223–230

    Google Scholar 

  • Kustu S, Santero E, Keener J, Popham D, Weiss D (1989) Expression of σ54 (ntrA)-dependent genes is probably united by a common mechanism. Microbiol Rev 53:367–376

    Google Scholar 

  • Lüthi E, Mercenier A, Haas D (1986) The arcABC operon required for fermentative growth of Pseudomonas aeruginosa on arginine Tn5-751-assisted cloning and localization of structural genes. J Gen Microbiol 132:2667–2675

    Google Scholar 

  • Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York

    Google Scholar 

  • Minton NP (1984) Improved plasmid vectors for the isolation of translational lac gene fusions. Gene 31:269–273

    Google Scholar 

  • Mohr CD, Martin DW, Konyecsni WM, Govan JRW, Lory S, Deretic V (1990) Role of the far-upstream sites of the algD promoter and the algR and rpoN genes in environmental modulation of mucoidy in Pseudomonas aeruginosa. J Bacteriol 172:6576–6580

    Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning. A laboratory manual, 2nd edn. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York

    Google Scholar 

  • Savioz A, Jeenes DJ, Kocher HP, Haas D (1990) Comparison of proC and other housekeeping genes of Pseudomonas aeruginosa with their counterparts in Escherichia coli. Gene 86:107–111

    Google Scholar 

  • Sawers RG (1991) Identification and molecular characterization of a transcriptional regulator from Pseudomonas aeruginosa PAO1 exhibiting structural and functional similarity to the FNR protein of Escherichia coli. Mol Microbiol 5:1469–1481

    Google Scholar 

  • Soldati L, Jeenes DJ, Haas D (1987) Effective gene expression in Pseudomonas aeruginosa under the control of the Escherichia coli consensus promoter. FEMS Microbiol Lett 42:163–167

    Google Scholar 

  • Starnbach MN, Lory S (1992) The fliA (rpoF) gene of Pseudomonas aeruginosa encodes an alternative sigma factor required for flagellin synthesis. Mol Microbiol 6:459–469

    Google Scholar 

  • Tanaka K, Takahashi H (1991) Cloning and analysis of the gene (rpoDA) for the principal σ factor of Pseudomonas aeruginosa. Biochim Biophys Acta 1089:113–119

    Google Scholar 

  • Tanaka K, Shiina T, Takahashi H (1988) Multiple principal sigma factor homologs in eubacteria: identification of the “rpoD box”. Science 242:1040–1042

    Google Scholar 

  • Thöny B, Hennecke H (1989) The −24/−12 promoter comes of age. FEMS Microbiol Rev 63:341–358

    Google Scholar 

  • Totten PA, Lara JC, Lory S (1990) The rpoN gene product of Pseudomonas aeruginosa is required for expression of diverse genes, including the flagellin gene. J Bacteriol 172:389–396

    Google Scholar 

  • Vicente M, Kushner SR, Garrido T, Aldea M (1991) The role of the ‘gearbox’ in the transcription of essential genes. Mol Microbiol 5:2085–2091

    Google Scholar 

  • Vfegenthart JS, Ketelaar-van Gaalen PAG, van de Klundert JAM (1991) Nucleotide sequence of the aacC3 gene, a gentamicin resistance determinant encoding aminoglycoside-(3)-N-acetyl-transferase III expressed in Pseudomonas aeruginosa but not in Escherichia coli. Antimicrob Agents Chemother 35:892–897

    Google Scholar 

  • Vögtli M, Hütter R (1987) Characterization of the hydroxystreptomycin phosphotransferase gene (sph) of Streptomyces glaucescens: nucleotide sequence and promoter analysis. Mol Gen Genet 208:195–203

    Google Scholar 

  • Whitchurch CB, Hobbs M, Livingston SP, Krishnapillai V, Mattick JS (1990) Characterisation of a Pseudomonas aeruginosa twitching motility gene and evidence for a specialised protein export system widespread in eubacteria. Gene 101:33–44

    Google Scholar 

  • Wong SC, Abdelal AT (1990) Unorthodox expression of an enzyme: evidence for an untranslated region within carA from Pseudomonas aeruginosa. J Bacteriol 172:630–642

    Google Scholar 

  • 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 

  • Zimmermann A (1992) Anaerobe Regulation des Arginin-Deiminase-Operons von Pseudomonas aeruginosa durch das Protein ANR. Dissertation ETH no. 9679, Zürich

  • Zimmermann A, Reimmann C, Galimand M, Haas D (1991) Anaerobic growth and cyanide synthesis of Pseudomonas aeruginosa depend on anr, a regulatory gene homologous with fnr of Escherichia coli. Mot Microbiol 5:1483–1490

    Google Scholar 

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Communicated by A. Kondorosi

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Savioz, A., Zimmermann, A. & Haas, D. Pseudomonas aeruginosa promoters which contain a conserved GG-N10-GC motif but appear to be RpoN-independent. Molec. Gen. Genet. 238, 74–80 (1993). https://doi.org/10.1007/BF00279533

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  • DOI: https://doi.org/10.1007/BF00279533

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