Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1991 Jan;173(2):417–425. doi: 10.1128/jb.173.2.417-425.1991

Identification and cloning of a fur regulatory gene in Yersinia pestis.

T M Staggs 1, R D Perry 1
PMCID: PMC207028  PMID: 1898928

Abstract

Yersinia pestis is one of many microorganisms responding to environmental iron concentrations by regulating the synthesis of proteins and an iron transport system(s). In a number of bacteria, expression of iron uptake systems and other virulence determinants is controlled by the Fur regulatory protein. DNA hybridization analysis revealed that both pigmented and nonpigmented cells of Y. pestis possess a DNA locus homologous to the Escherichia coli fur gene. Introduction of a Fur-regulated beta-galactosidase reporter gene into Y. pestis KIM resulted in iron-responsive beta-galactosidase activity, indicating that Y. pestis KIM expresses a functional Fur regulatory protein. A cloned 1.9-kb ClaI fragment of Y. pestis chromosomal DNA hybridized specifically to the fur gene of E. coli. The coding region of the E. coli fur gene hybridized to a 1.1-kb region at one end of the cloned Y. pestis fragment. The failure of this clone to complement an E. coli fur mutant suggests that the 1.9-kb clone does not contain a functional promoter. Subcloning of this fragment into an inducible expression vector restored Fur regulation in an E. coli fur mutant. In addition, a larger 4.8-kb Y. pestis clone containing the putative promoter region complemented the Fur- phenotype. These results suggest that Y. pestis possesses a functional Fur regulatory protein capable of interacting with the E. coli Fur system. In Y. pestis Fur may regulate the expression of iron transport systems and other virulence factors in response to iron limitation in the environment. Possible candidates for Fur regulation in Y. pestis include genes involved in ferric iron transport as well as hemin, heme/hemopexin, heme/albumin, ferritin, hemoglobin, and hemoglobin/haptoglobin utilization.

Full text

PDF
425

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Archibald F. S., DeVoe I. W. Iron acquisition by Neisseria meningitidis in vitro. Infect Immun. 1980 Feb;27(2):322–334. doi: 10.1128/iai.27.2.322-334.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. BURROWS T. W., JACKSON S. The pigmentation of Pasteurella pestis on a defined medium containing haemin. Br J Exp Pathol. 1956 Dec;37(6):570–576. [PMC free article] [PubMed] [Google Scholar]
  3. BURROWS T. W., JACKSON S. The virulence-enhancing effect of iron on nonpigmented mutants of virulent strains of Pasteurella pestis. Br J Exp Pathol. 1956 Dec;37(6):577–583. [PMC free article] [PubMed] [Google Scholar]
  4. Bagg A., Neilands J. B. 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. 1987 Aug 25;26(17):5471–5477. doi: 10.1021/bi00391a039. [DOI] [PubMed] [Google Scholar]
  5. Beesley E. D., Brubaker R. R., Janssen W. A., Surgalla M. J. Pesticins. 3. Expression of coagulase and mechanism of fibrinolysis. J Bacteriol. 1967 Jul;94(1):19–26. doi: 10.1128/jb.94.1.19-26.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bolivar F., Backman K. Plasmids of Escherichia coli as cloning vectors. Methods Enzymol. 1979;68:245–267. doi: 10.1016/0076-6879(79)68018-7. [DOI] [PubMed] [Google Scholar]
  8. Boyd J., Oza M. N., Murphy J. R. Molecular cloning and DNA sequence analysis of a diphtheria tox iron-dependent regulatory element (dtxR) from Corynebacterium diphtheriae. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5968–5972. doi: 10.1073/pnas.87.15.5968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Brosius J., Holy A. Regulation of ribosomal RNA promoters with a synthetic lac operator. Proc Natl Acad Sci U S A. 1984 Nov;81(22):6929–6933. doi: 10.1073/pnas.81.22.6929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Brubaker R. R. The Vwa+ virulence factor of yersiniae: the molecular basis of the attendant nutritional requirement for Ca++. Rev Infect Dis. 1983 Sep-Oct;5 (Suppl 4):S748–S758. doi: 10.1093/clinids/5.supplement_4.s748. [DOI] [PubMed] [Google Scholar]
  11. Bullen J. J. The significance of iron in infection. Rev Infect Dis. 1981 Nov-Dec;3(6):1127–1138. doi: 10.1093/clinids/3.6.1127. [DOI] [PubMed] [Google Scholar]
  12. Calderwood S. B., Mekalanos J. J. Confirmation of the Fur operator site by insertion of a synthetic oligonucleotide into an operon fusion plasmid. J Bacteriol. 1988 Feb;170(2):1015–1017. doi: 10.1128/jb.170.2.1015-1017.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Calderwood S. B., Mekalanos J. J. Iron regulation of Shiga-like toxin expression in Escherichia coli is mediated by the fur locus. J Bacteriol. 1987 Oct;169(10):4759–4764. doi: 10.1128/jb.169.10.4759-4764.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Carniel E., Mazigh D., Mollaret H. H. Expression of iron-regulated proteins in Yersinia species and their relation to virulence. Infect Immun. 1987 Jan;55(1):277–280. doi: 10.1128/iai.55.1.277-280.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Cryz S. J., Jr, Russell L. M., Holmes R. K. Regulation of toxinogenesis in Corynebacterium diphtheriae: mutations in the bacterial genome that alter the effects of iron on toxin production. J Bacteriol. 1983 Apr;154(1):245–252. doi: 10.1128/jb.154.1.245-252.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Dyer D. W., West E. P., Sparling P. F. Effects of serum carrier proteins on the growth of pathogenic neisseriae with heme-bound iron. Infect Immun. 1987 Sep;55(9):2171–2175. doi: 10.1128/iai.55.9.2171-2175.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Ernst J. F., Bennett R. L., Rothfield L. I. Constitutive expression of the iron-enterochelin and ferrichrome uptake systems in a mutant strain of Salmonella typhimurium. J Bacteriol. 1978 Sep;135(3):928–934. doi: 10.1128/jb.135.3.928-934.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Expert D., Toussaint A. Bacteriocin-resistant mutants of Erwinia chrysanthemi: possible involvement of iron acquisition in phytopathogenicity. J Bacteriol. 1985 Jul;163(1):221–227. doi: 10.1128/jb.163.1.221-227.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Goguen J. D., Yother J., Straley S. C. Genetic analysis of the low calcium response in Yersinia pestis mu d1(Ap lac) insertion mutants. J Bacteriol. 1984 Dec;160(3):842–848. doi: 10.1128/jb.160.3.842-848.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  21. Hantke K. Cloning of the repressor protein gene of iron-regulated systems in Escherichia coli K12. Mol Gen Genet. 1984;197(2):337–341. doi: 10.1007/BF00330982. [DOI] [PubMed] [Google Scholar]
  22. Hantke K. Regulation of ferric iron transport in Escherichia coli K12: isolation of a constitutive mutant. Mol Gen Genet. 1981;182(2):288–292. doi: 10.1007/BF00269672. [DOI] [PubMed] [Google Scholar]
  23. Hantke K. Selection procedure for deregulated iron transport mutants (fur) in Escherichia coli K 12: fur not only affects iron metabolism. Mol Gen Genet. 1987 Nov;210(1):135–139. doi: 10.1007/BF00337769. [DOI] [PubMed] [Google Scholar]
  24. Kado C. I., Liu S. T. Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol. 1981 Mar;145(3):1365–1373. doi: 10.1128/jb.145.3.1365-1373.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Konopka K., Bindereif A., Neilands J. B. Aerobactin-mediated utilization of transferrin iron. Biochemistry. 1982 Dec 7;21(25):6503–6508. doi: 10.1021/bi00268a028. [DOI] [PubMed] [Google Scholar]
  26. Mazurier J., Spik G. Comparative study of the iron-binding properties of human transferrins. I. Complete and sequential iron saturation and desaturation of the lactotransferrin. Biochim Biophys Acta. 1980 May 7;629(2):399–408. doi: 10.1016/0304-4165(80)90112-9. [DOI] [PubMed] [Google Scholar]
  27. McIntosh M. A., Earhart C. F. Effect of iron of the relative abundance of two large polypeptides of the Escherichia coli outer membrane. Biochem Biophys Res Commun. 1976 May 3;70(1):315–322. doi: 10.1016/0006-291x(76)91144-x. [DOI] [PubMed] [Google Scholar]
  28. Nagel R. L., Gibson Q. H. The binding of hemoglobin to haptoglobin and its relation to subunit dissociation of hemoglobin. J Biol Chem. 1971 Jan 10;246(1):69–73. [PubMed] [Google Scholar]
  29. Neilands J. B. Microbial envelope proteins related to iron. Annu Rev Microbiol. 1982;36:285–309. doi: 10.1146/annurev.mi.36.100182.001441. [DOI] [PubMed] [Google Scholar]
  30. Oudega B., Oldenziel-Werner W. J., Klaasen-Boor P., Rezee A., Glas J., de Graaf F. K. Purification and characterization of cloacin DF13 receptor from Enterobacter cloacae and its interaction with cloacin DF13 in vitro. J Bacteriol. 1979 Apr;138(1):7–16. doi: 10.1128/jb.138.1.7-16.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Pasternack R. F., Gibbs E. J., Hoeflin E., Kosar W. P., Kubera G., Skowronek C. A., Wong N. M., Muller-Eberhard U. Hemin binding to serum proteins and the catalysis of interprotein transfer. Biochemistry. 1983 Apr 12;22(8):1753–1758. doi: 10.1021/bi00277a002. [DOI] [PubMed] [Google Scholar]
  32. Payne S. M. Iron and virulence in the family Enterobacteriaceae. Crit Rev Microbiol. 1988;16(2):81–111. doi: 10.3109/10408418809104468. [DOI] [PubMed] [Google Scholar]
  33. Payne S. M., Niesel D. W., Peixotto S. S., Lawlor K. M. Expression of hydroxamate and phenolate siderophores by Shigella flexneri. J Bacteriol. 1983 Sep;155(3):949–955. doi: 10.1128/jb.155.3.949-955.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Perry R. D., Brubaker R. R. Accumulation of iron by yersiniae. J Bacteriol. 1979 Mar;137(3):1290–1298. doi: 10.1128/jb.137.3.1290-1298.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Perry R. D., Brubaker R. R. Vwa+ phenotype of Yersinia enterocolitica. Infect Immun. 1983 Apr;40(1):166–171. doi: 10.1128/iai.40.1.166-171.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Perry R. D., Harmon P. A., Bowmer W. S., Straley S. C. A low-Ca2+ response operon encodes the V antigen of Yersinia pestis. Infect Immun. 1986 Nov;54(2):428–434. doi: 10.1128/iai.54.2.428-434.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Perry R. D., Pendrak M. L., Schuetze P. Identification and cloning of a hemin storage locus involved in the pigmentation phenotype of Yersinia pestis. J Bacteriol. 1990 Oct;172(10):5929–5937. doi: 10.1128/jb.172.10.5929-5937.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  39. Rogers H. J. Iron-Binding Catechols and Virulence in Escherichia coli. Infect Immun. 1973 Mar;7(3):445–456. doi: 10.1128/iai.7.3.445-456.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Salinas P. C., Tolmasky M. E., Crosa J. H. Regulation of the iron uptake system in Vibrio anguillarum: evidence for a cooperative effect between two transcriptional activators. Proc Natl Acad Sci U S A. 1989 May;86(10):3529–3533. doi: 10.1073/pnas.86.10.3529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Schäffer S., Hantke K., Braun V. Nucleotide sequence of the iron regulatory gene fur. Mol Gen Genet. 1985;200(1):110–113. doi: 10.1007/BF00383321. [DOI] [PubMed] [Google Scholar]
  42. Sikkema D. J., Brubaker R. R. Outer membrane peptides of Yersinia pestis mediating siderophore-independent assimilation of iron. Biol Met. 1989;2(3):174–184. doi: 10.1007/BF01142557. [DOI] [PubMed] [Google Scholar]
  43. Sikkema D. J., Brubaker R. R. Resistance to pesticin, storage of iron, and invasion of HeLa cells by Yersiniae. Infect Immun. 1987 Mar;55(3):572–578. doi: 10.1128/iai.55.3.572-578.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  45. Stoebner J. A., Payne S. M. Iron-regulated hemolysin production and utilization of heme and hemoglobin by Vibrio cholerae. Infect Immun. 1988 Nov;56(11):2891–2895. doi: 10.1128/iai.56.11.2891-2895.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Stoker N. G., Fairweather N. F., Spratt B. G. Versatile low-copy-number plasmid vectors for cloning in Escherichia coli. Gene. 1982 Jun;18(3):335–341. doi: 10.1016/0378-1119(82)90172-x. [DOI] [PubMed] [Google Scholar]
  47. Straley S. C., Bowmer W. S. Virulence genes regulated at the transcriptional level by Ca2+ in Yersinia pestis include structural genes for outer membrane proteins. Infect Immun. 1986 Feb;51(2):445–454. doi: 10.1128/iai.51.2.445-454.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Straley S. C., Brubaker R. R. Localization in Yersinia pestis of peptides associated with virulence. Infect Immun. 1982 Apr;36(1):129–135. doi: 10.1128/iai.36.1.129-135.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Stull T. L. Protein sources of heme for Haemophilus influenzae. Infect Immun. 1987 Jan;55(1):148–153. doi: 10.1128/iai.55.1.148-153.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Surgalla M. J., Beesley E. D. Congo red-agar plating medium for detecting pigmentation in Pasteurella pestis. Appl Microbiol. 1969 Nov;18(5):834–837. doi: 10.1128/am.18.5.834-837.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Twigg A. J., Sherratt D. Trans-complementable copy-number mutants of plasmid ColE1. Nature. 1980 Jan 10;283(5743):216–218. doi: 10.1038/283216a0. [DOI] [PubMed] [Google Scholar]
  52. Wee S., Neilands J. B., Bittner M. L., Hemming B. C., Haymore B. L., Seetharam R. Expression, isolation and properties of Fur (ferric uptake regulation) protein of Escherichia coli K 12. Biol Met. 1988;1(1):62–68. doi: 10.1007/BF01128019. [DOI] [PubMed] [Google Scholar]
  53. Weinberg E. D. Iron withholding: a defense against infection and neoplasia. Physiol Rev. 1984 Jan;64(1):65–102. doi: 10.1152/physrev.1984.64.1.65. [DOI] [PubMed] [Google Scholar]
  54. West S. E., Sparling P. F. Response of Neisseria gonorrhoeae to iron limitation: alterations in expression of membrane proteins without apparent siderophore production. Infect Immun. 1985 Feb;47(2):388–394. doi: 10.1128/iai.47.2.388-394.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Williams P., Brown M. R., Lambert P. A. Effect of iron deprivation on the production of siderophores and outer membrane proteins in Klebsiella aerogenes. J Gen Microbiol. 1984 Sep;130(9):2357–2365. doi: 10.1099/00221287-130-9-2357. [DOI] [PubMed] [Google Scholar]
  56. de Lorenzo V., Wee S., Herrero M., Neilands J. B. Operator sequences of the aerobactin operon of plasmid ColV-K30 binding the ferric uptake regulation (fur) repressor. J Bacteriol. 1987 Jun;169(6):2624–2630. doi: 10.1128/jb.169.6.2624-2630.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES