1887

Abstract

needs to synthesize haem in order to replicate intracellularly and to produce virulence in mice. Thus, to gain insight into the pathogenesis of the bacterium, regulatory proteins of the haem biosynthetic pathway were sought. An iron response regulator (Irr) from , which is a close relative of , was previously described as being involved in the coordination of haem biosynthesis and iron availability. The genome was searched for an orthologue gene, and the gene was cloned, sequenced and disrupted. A null mutant was constructed that accumulated protoporphyrin IX under conditions of iron deprivation. This phenotype was overcome by a complementing plasmid carrying the wild-type . Purified recombinant Irr behaved as a stable dimer and bound haem. Interestingly, , Irr was only detected in cells obtained from iron-limited cultures and the protein downregulated its own transcription. Through fusion, it was demonstrated that iron did not regulate transcription. The data reported show that Irr is a homodimeric protein that is accumulated in iron-limited cells, controls its own transcription and downregulates the biosynthesis of haem precursors.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.28213-0
2005-10-01
2020-05-31
Loading full text...

Full text loading...

/deliver/fulltext/micro/151/10/3427.html?itemId=/content/journal/micro/10.1099/mic.0.28213-0&mimeType=html&fmt=ahah

References

  1. Almiron M., Martinez M., Sanjuan N., Ugalde R. A. 2001; Ferrochelatase is present in Brucella abortus and is critical for its intracellular survival and virulence. Infect Immun69:6225–6230[CrossRef]
    [Google Scholar]
  2. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. 1990; Basic local alignment search tool. J Mol Biol215:403–410[CrossRef]
    [Google Scholar]
  3. Becker A., Schmidt M., Jager W., Puhler A. 1995; New gentamicin-resistance and lacZ promoter-probe cassettes suitable for insertion mutagenesis and generation of transcriptional fusions. Gene162:37–39[CrossRef]
    [Google Scholar]
  4. Bellaire B. H., Elzer P. H., Hagius S., Walker J., Baldwin C. L., Roop R. M.. 2nd (2003; Genetic organization and iron-responsive regulation of the Brucella abortus 2,3-dihydroxybenzoic acid biosynthesis operon, a cluster of genes required for wild-type virulence in pregnant cattle. Infect Immun71:1794–1803[CrossRef]
    [Google Scholar]
  5. Cox R., Charles H. P. 1973; Porphyrin-accumulating mutants of Escherichia coli . J Bacteriol113:122–132
    [Google Scholar]
  6. Escolar L., Perez-Martin J., de Lorenzo V. 1999; Opening the iron box: transcriptional metalloregulation by the Fur protein. J Bacteriol181:6223–6229
    [Google Scholar]
  7. Frustaci J. M., O'Brian M. R. 1992; Characterization of a Bradyrhizobium japonicum ferrochelatase mutant and isolation of the hemH gene. J Bacteriol174:4223–4229
    [Google Scholar]
  8. Guo B., Phillips J. D., Yu Y., Leibold E. A. 1995; Iron regulates the intracellular degradation of iron-regulatory protein 2 by the proteasome. J Biol Chem270:21645–21651[CrossRef]
    [Google Scholar]
  9. Hamza I., Chauhan S., Hassett R., O'Brian M. R. 1998; The bacterial Irr protein is required for coordination of heme biosynthesis with iron availability. J Biol Chem273:21669–21674[CrossRef]
    [Google Scholar]
  10. Hamza I., Qi Z., King N. D., O'Brian M. R. 2000; Fur-independent regulation of iron metabolism by Irr in Bradyrhizobium japonicum . Microbiology146:669–676
    [Google Scholar]
  11. Kovach M. E., Elzer P. H., Hill D. S., Robertson G. T., Farris M. A., Roop R. M. 2nd, Peterson K. M. 1995; Four new derivatives of the broad-host-range cloning vector pBBR1MCS, carrying different antibiotic-resistance cassettes. Gene166:175–176[CrossRef]
    [Google Scholar]
  12. Lestrate P., Delrue R. M., Danese I. 7 other authors 2000; Identification and characterization of in vivo attenuated mutants of Brucella melitensis . Mol Microbiol38:543–551[CrossRef]
    [Google Scholar]
  13. Lopez-Goñi I., Moriyón I., Neilands J. B. 1992; Identification of 2,3-dihydroxybenzoic acid as a Brucella abortus siderophore. Infect Immun60:4496–4503
    [Google Scholar]
  14. Miller J. H. 1992; A Short Course in Bacterial Genetics Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  15. O'Brian M. R., Thony-Meyer L. 2002; Biochemistry, regulation and genomics of haem biosynthesis in prokaryotes. Adv Microb Physiol46:257–318
    [Google Scholar]
  16. Ogawa K., Sun J., Taketani S. & 8 other authors. 2001; Heme mediates derepression of Maf recognition element through direct binding to transcription repressor Bach1. EMBO J20:2835–2843[CrossRef]
    [Google Scholar]
  17. Oka A., Sugisaki H., Takanami M. 1981; Nucleotide sequence of the kanamycin resistance transposon Tn903. J Mol Biol147:217–226[CrossRef]
    [Google Scholar]
  18. Paulsen I. T., Seshadri R., Nelson K. E. & 28 other authors. 2002; The Brucella suis genome reveals fundamental similarities between animal and plant pathogens and symbionts. Proc Natl Acad Sci U S A99:13148–13153[CrossRef]
    [Google Scholar]
  19. Pfeifer K., Kim K. S., Kogan S., Guarente L. 1989; Functional dissection and sequence of yeast HAP1 activator. Cell56:291–301[CrossRef]
    [Google Scholar]
  20. Pohl E., Haller J. C., Mijovilovich A., Meyer-Klaucke W., Garman E., Vasil M. L. 2003; Architecture of a protein central to iron homeostasis: crystal structure and spectroscopic analysis of the ferric uptake regulator. Mol Microbiol47:903–915[CrossRef]
    [Google Scholar]
  21. Qi Z., O'Brian M. R. 2002; Interaction between the bacterial iron response regulator and ferrochelatase mediates genetic control of heme biosynthesis. Mol Cell9:155–162[CrossRef]
    [Google Scholar]
  22. Qi Z., Hamza I., O'Brian M. R. 1999; Heme is an effector molecule for iron-dependent degradation of the bacterial iron response regulator (Irr) protein. Proc Natl Acad Sci U S A96:13056–13061[CrossRef]
    [Google Scholar]
  23. Sambrook J., Fritsch E. F., Maniatis T. 1989; Molecular Cloning: a Laboratory Manual , 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  24. Smith L. D., Ficht T. A. 1990; Pathogenesis of Brucella . Crit Rev Microbiol17:209–230[CrossRef]
    [Google Scholar]
  25. Stojiljkovic I., Hantke K. 1995; Functional domains of the Escherichia coli ferric uptake regulator protein (Fur. Mol Gen Genet247:199–205[CrossRef]
    [Google Scholar]
  26. Ugalde R. A. 1999; Intracellular lifestyle of Brucella spp. Common genes with other animal pathogens, plant pathogens, and endosymbionts. Microbes Infect1:1211–1219[CrossRef]
    [Google Scholar]
  27. Wexler M., Todd J. D., Kolade O., Bellini D., Hemmings A. M., Sawers G., Johnston A. W. 2003; Fur is not the global regulator of iron uptake genes in Rhizobium leguminosarum . Microbiology149:1357–1365[CrossRef]
    [Google Scholar]
  28. Woodcock D. M., Crowther P. J., Doherty J., Jefferson S., DeCruz E., Noyer-Weidner M., Smith S. S., Michael M. Z., Graham M. W. 1989; Quantitative evaluation of Escherichia coli host strains for tolerance to cytosine methylation in plasmid and phage recombinants. Nucleic Acids Res17:3469–3478[CrossRef]
    [Google Scholar]
  29. Yang J., Ishimori K., O'Brian M. R. 2005; Two heme binding sites are involved in the regulated degradation of the bacterial iron response regulator (Irr) protein. J Biol Chem280:7671–7676[CrossRef]
    [Google Scholar]
  30. Zhang L., Guarente L. 1995; Heme binds to a short sequence that serves a regulatory function in diverse proteins. EMBO J14:313–320
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.28213-0
Loading
/content/journal/micro/10.1099/mic.0.28213-0
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

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