1887

Abstract

forms both catecholate and azotobactin siderophores during iron-limited growth. Azotobactin is repressed by about 3 μM iron, but catecholate siderophore synthesis continues up to a maximum of 10 μM iron. This suggests that catecholate siderophore synthesis is regulated by other factors in addition to the ferric uptake repressor (Fur). In this study the first gene required for catecholate siderophore biosynthesis, which encodes an isochorismate synthase (), was isolated. The region upstream of contained a typical σ promoter, with an iron-box overlapping the −35 sequence and a Sox-box (Box 1) overlapping the −10 sequence. Another Sox-box was found further upstream of the −35 sequence (Box 2). Also upstream, an unidentified gene () was detected which would be transcribed from a divergent promoter, also controlled by an iron-box. The activity of and a :: fusion was negatively regulated by iron availability and upregulated by increased aeration and by superoxide stress. The iron-box in the promoter was 74% identical to the Fur-binding consensus sequence and bound the Fur protein of with relatively high affinity. Both Box 1 and Box 2 were in good agreement with the consensus sequence for binding the SoxS protein of and Box 1 was in very good agreement with the Sox-box found in the promoter of , which is also regulated by superoxide stress. Both Sox-boxes bound a protein found in cell extracts, with Box 1 exhibiting the higher binding affinity. The Sox protein identified in this assay appeared to be constitutive, rather than inducible by superoxide stress. This indicates that the Sox response in is different from that in . These data support the hypothesis that catecholate siderophore biosynthesis is under dual control, repressed by a Fur–iron complex and activated by another DNA-binding protein in response to superoxide stress. The interaction between these regulators is likely to account for the delay in ferric repression of catecholate siderophore production, since these siderophores have an additional role to play in the protection of iron-limited cells against oxidative damage.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-146-7-1617
2000-07-01
2019-10-22
Loading full text...

Full text loading...

/deliver/fulltext/micro/146/7/1461617a.html?itemId=/content/journal/micro/10.1099/00221287-146-7-1617&mimeType=html&fmt=ahah

References

  1. Achenbach, L. A. & Yang, W. ( 1997; ). The fur gene from Klebsiella pneumoniae: characterization, genomic organization and phylogenetic analysis. Gene 185, 201-207.[CrossRef]
    [Google Scholar]
  2. 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.[CrossRef]
    [Google Scholar]
  3. Ansari, A., Bradner, J. & O’Halloran, T. ( 1995; ). DNA-bend modulation in a repressor-to-activator switching mechanism. Nature 374, 371-375.
    [Google Scholar]
  4. Auserwald, E. A., Ludwig, G. & Schaller, H. ( 1981; ). Structural analysis of Tn5. Cold Spring Harb Symp Quant Biol 45, 107-113.[CrossRef]
    [Google Scholar]
  5. Bagg, A. & Neilands, J. B. ( 1987; ). Ferric uptake regulation protein acts as a repressor, employing iron(II) as a cofactor to bind the operator of an iron transport operon in Escherichia coli. Biochemistry 26, 5471-5477.[CrossRef]
    [Google Scholar]
  6. Barnum, D. W. ( 1977; ). Spectrophotometric determination of catechol, epinephrine, DOPA, dopamine and other aromatic vic-diols. Anal Chim Acta 89, 157-166.[CrossRef]
    [Google Scholar]
  7. Bibb, M. J., Findlay, P. R. & Johnson, M. W. ( 1984; ). The relationship between base composition and codon usage in bacterial genes and its use for the simple and reliable identification of protein-coding sequences. Gene 30, 157-166.[CrossRef]
    [Google Scholar]
  8. Corbin, J. L. & Bulen, W. A. ( 1969; ). The isolation and identification of 2,3-dihydroxybenzoic acid and 2-N, 6-N-di-(2,3-dihydroxybenzoyl)-l-lysine formed by iron-deficient Azotobacter vinelandii. Biochemistry 8, 757-762.[CrossRef]
    [Google Scholar]
  9. Cornish, A. S. & Page, W. J. ( 1995; ). Production of the tricatecholate siderophore protochelin by Azotobacter vinelandii. BioMetals 8, 332-338.
    [Google Scholar]
  10. Cornish, A. S. & Page, W. J. ( 1998; ). The catecholate siderophores of Azotobacter vinelandii: their affinity for iron and role in oxygen stress management. Microbiology 144, 1747-1754.[CrossRef]
    [Google Scholar]
  11. Crosa, J. H. ( 1989; ). Genetics and molecular biology of siderophore-mediated iron transport in bacteria. Microbiol Rev 53, 517-530.
    [Google Scholar]
  12. Dalet, K., Gouin, E., Cenatiempo, Y., Cossart, P. & Hechard, Y. ( 1999; ). Characterization of a new operon encoding a Zur-like protein and an associated ABC zinc permease in Listeria monocytogenes. FEMS Microbiol Lett 174, 111-116.[CrossRef]
    [Google Scholar]
  13. Demange, P., Wendenbaum, S., Bateman, A., Dell, A., Meyer, J. M. & Abdallah, M. A. (1986). Bacterial siderophores: structure of pyoverdins and related compounds. In Iron, Siderophores, and Plant Diseases, pp. 131–147. Edited by T. R. Swinburne. New York: Plenum.
  14. Demple, B. ( 1996; ). Redox signaling and gene control in the Escherichia coli soxRS oxidative stress regulon – a review. Gene 179, 53-57.[CrossRef]
    [Google Scholar]
  15. Dittrich, W., Betzler, M. & Schrempf, H. ( 1991; ). An amplifiable and detectable chloramphenicol-resistance determinant of Streptomyces lividans 1326 encodes a putative transmembrane protein. Mol Microbiol 5, 2789-2797.[CrossRef]
    [Google Scholar]
  16. Escolar, L., Perez-Martin, J. & de Lorenzo, V. ( 1999; ). Opening the iron box: transcriptional metalloregulation by the Fur protein. J Bacteriol 181, 6223-6229.
    [Google Scholar]
  17. Farinha, M. A. & Kropinski, A. M. ( 1990; ). Construction of broad host range plasmid vectors for easy visible selection and analysis of promoters. J Bacteriol 172, 3496-3499.
    [Google Scholar]
  18. Fawcett, W. P. & Wolf, R. E. ( 1994; ). Purification of the MalE–SoxS fusion protein and identification of the control sites of Escherichia coli superoxide-inducible genes. Mol Microbiol 14, 669-679.[CrossRef]
    [Google Scholar]
  19. Feinberg, A. P. & Vogelstein, B. ( 1983; ). A technique for radiolabelling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 132, 6-13.[CrossRef]
    [Google Scholar]
  20. Frost, L., Lee, S., Yanchar, N. & Paranchych, W. ( 1989; ). finP and fisO mutations in FinP antisense RNA suggest a model for FinOP action in the repression of bacteria conjugation by the Flac plasmid JCFL0. Mol Gen Genet 218, 152-160.[CrossRef]
    [Google Scholar]
  21. Hantke, K. ( 1984; ). Cloning of the repressor protein gene of iron-regulated systems in Escherichia coli K12. Mol Gen Genet 197, 337-341.[CrossRef]
    [Google Scholar]
  22. Isas, J. M., Yannone, S. M. & Burgess, B. K. ( 1995; ). Azotobacter vinelandii NADPH:ferredoxin reductase cloning, sequencing, and overexpression. J Biol Chem 270, 21258-21263.[CrossRef]
    [Google Scholar]
  23. Ishihama, A. ( 1992; ). Role of the RNA polymerase α subunit in transcription activation. Mol Microbiol 6, 3283-3288.[CrossRef]
    [Google Scholar]
  24. Jobling, M. G. & Holmes, R. K. ( 1990; ). Construction of vectors with the p15a replicon, kanamycin resistance, inducible lacZα and pUC18 or pUC19 multiple cloning sites. Nucleic Acids Res 18, 5315-5316.[CrossRef]
    [Google Scholar]
  25. Korbashi, P., Kohen, R., Katzhendler, J. & Chevion, M. ( 1986; ). Iron mediates paraquat toxicity in Escherichia coli. J Biol Chem 261, 12472-12476.
    [Google Scholar]
  26. Lewin, B. (1994). Genes V. New York: Oxford University Press.
  27. de Lorenzo, V., Wee, S., Herrero, M. & Neilands, J. B. ( 1987; ). Operator sequences of the aerobactin operon of plasmid ColV-K30 binding the ferric uptake regulation (fur) repressor. J Bacteriol 169, 2624-2630.
    [Google Scholar]
  28. de Lorenzo, V., Giovannini, F., Herrero, M. & Neilands, J. B. ( 1988; ). Metal ion regulation of gene expression. Fur repressor–operator interaction at the promoter region of the aerobactin system of pColV-K30. J Mol Biol 203, 875-884.[CrossRef]
    [Google Scholar]
  29. Maniatis, T., Fritsch, E. F. & Sambrook, J. (1982). Molecular Cloning: a Laboratory Manual. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  30. Martin, A. E., Burgess, B. K., Iismaa, S. E., Smartt, C. T., Jacobson, M. R. & Dean, D. R. ( 1989; ). Construction and characterization of an Azotobacter vinelandii strain with mutations in the genes encoding flavodoxin and ferredoxin I. J Bacteriol 171, 3162-3167.
    [Google Scholar]
  31. Massad, G., Arceneaux, J. E. L. & Byers, B. R. ( 1994; ). Diversity of siderophore genes encoding biosynthesis of 2,3-dihydroxybenzoic acid in Aeromonas spp. BioMetals 7, 227-236.
    [Google Scholar]
  32. Mehrotra, M. (1997). Role of iron in the regulation of phenylalanyl-tRNA synthetase activity in Azotobacter vinelandii. PhD thesis, University of Alberta.
  33. Miller, J. H. (1972). Experiments in Molecular Genetics. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  34. Morgan, T. V., Lundell, D. J. & Burgess, B. K. ( 1988; ). Azotobacter vinelandii ferredoxin I: cloning, sequencing, and mutant analysis. J Biol Chem 263, 1370-1375.
    [Google Scholar]
  35. Nunoshiba, T. ( 1996; ). Two-stage gene regulation of the superoxide stress response soxRS system in Escherichia coli. Crit Rev Eukaryot Gene Expression 6, 377-389.[CrossRef]
    [Google Scholar]
  36. Oberley, L. W. & Spitz, D. R. ( 1985; ). Nitroblue tetrazolium. In CRC Handbook of Methods in Oxygen Radical Research, pp. 217-220. Edited by R. A. Greenwald. Boca Raton: CRC Press.
  37. Ozenberger, B. A., Nahlik, M. S. & McIntosh, M. A. ( 1987; ). Genetic organization of multiple fep genes encoding ferric enterobactin transport functions in Escherichia coli. J Bacteriol 169, 3638-3646.
    [Google Scholar]
  38. Page, W. J. ( 1993; ). Growth conditions for the demonstration of siderophores and iron-repressible outer membrane proteins in soil bacteria, with an emphasis on free-living diazotrophs. In Iron Chelation in Plants and Soil Microorganisms, pp. 75-110. Edited by L. L. Barton & B. C. Hemming. New York: Academic Press.
  39. Page, W. J. & Huyer, M. ( 1984; ). Derepression of the Azotobacter vinelandii siderophore system, using iron-containing minerals to limit iron repletion. J Bacteriol 158, 496-502.
    [Google Scholar]
  40. Page, W. J. & Sadoff, H. L. ( 1976; ). Physiological factors affecting transformation of Azotobacter vinelandii. J Bacteriol 125, 1080-1087.
    [Google Scholar]
  41. Page, W. J. & von Tigerstrom, M. ( 1988; ). Aminochelin, a catecholamine siderophore produced by Azotobacter vinelandii. J Gen Microbiol 134, 453-460.
    [Google Scholar]
  42. Prince, R. W., Storey, D. G., Vasil, A. I. & Vasil, M. L. ( 1991; ). Regulation of toxA and regA by the Escherichia coli fur gene and identification of a Fur homologue in Pseudomonas aeruginosa PA103 and PA01. Mol Microbiol 5, 2823-2831.[CrossRef]
    [Google Scholar]
  43. Regnström, K., Sauge-Merle, S., Chen, K. & Burgess, B. K. ( 1999; ). In Azotobacter vinelandii, the E1 subunit of the pyruvate dehydrogenase complex binds fpr promoter region DNA and ferredoxin I. Proc Natl Acad Sci USA 96, 12389-12393.[CrossRef]
    [Google Scholar]
  44. Rhodius, V. & Busby, S. ( 1998; ). Positive activation of gene expression. Curr Opin Microbiol 1, 152-159.[CrossRef]
    [Google Scholar]
  45. Robson, R. L. & Postgate, J. R. ( 1980; ). Oxygen and hydrogen in biological nitrogen fixation. Annu Rev Microbiol 34, 183-207.[CrossRef]
    [Google Scholar]
  46. Sambrook, J., Fritsch, E. F. & Maniatis, T. (1989). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  47. Sevinc, M. S. & Page, W. J. ( 1992; ). Generation of Azotobacter vinelandii strains defective in siderophore production and characterization of a strain unable to produce known siderophores. J Gen Microbiol 138, 587-596.[CrossRef]
    [Google Scholar]
  48. Southern, E. M. ( 1975; ). Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98, 503-517.[CrossRef]
    [Google Scholar]
  49. Staskawicz, D., Dahlbeck, D., Keen, N. & Napoli, C. ( 1987; ). Molecular characterization of cloned avirulence genes from race 0 and race 1 of Pseudomonas syringae pv. glycinae. J Bacteriol 169, 5789-5794.
    [Google Scholar]
  50. Summers, A. ( 1992; ). Untwist and shout – a heavy metal-response transcriptional regulator. J Bacteriol 174, 3097-3101.
    [Google Scholar]
  51. Thomas, P. ( 1980; ). Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci USA 77, 5201-5205.[CrossRef]
    [Google Scholar]
  52. Vieira, J. & Messing, J. ( 1987; ). Production of single stranded plasmid DNA. Methods Enzymol 153, 3-11.
    [Google Scholar]
  53. Yannone, S. M. & Burgess, B. K. ( 1997; ). Identification of a palindromic sequence that is responsible for the up-regulation of NAPDH-ferredoxin reductase in a ferredoxin I deletion strain of Azotobacter vinelandii. J Biol Chem 272, 14454-14458.[CrossRef]
    [Google Scholar]
  54. Yannone, S. M. & Burgess, B. K. ( 1998; ). The seven-iron FdI from Azotobacter vinelandii regulates the expression of NADPH:ferredoxin reductase via an oxidative stress response. J Biol Inorg Chem 3, 253-258.[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-146-7-1617
Loading
/content/journal/micro/10.1099/00221287-146-7-1617
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