Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1994 Jan 1;13(1):138–146. doi: 10.1002/j.1460-2075.1994.tb06243.x

An iron-sulfur center essential for transcriptional activation by the redox-sensing SoxR protein.

E Hidalgo 1, B Demple 1
PMCID: PMC394787  PMID: 8306957

Abstract

The soxRS oxidative stress regulon of Escherichia coli is triggered by superoxide (O2.-) generating agents or by nitric oxide through two consecutive steps of gene activation. SoxR protein has been proposed as the redox sensing gene activator that triggers this cascade of gene expression. We have now characterized two forms of SoxR: Fe-SoxR contained non-heme iron (up to 1.6 atoms per monomer); apo-SoxR was devoid of Fe or other metals. The spectroscopic properties of Fe-SoxR indicated that it contains a redox active iron-sulfur (FeS) cluster that is oxidized upon extraction from E. coli. Fe-SoxR and apo-SoxR bound the in vivo target, the soxS promoter, with equal affinities and protected the same region from DNase I in vitro. However, only Fe-SoxR stimulated transcription initiation at soxS in vitro > 100-fold, similar to the activation of soxS expression in vivo. This stimulation occurred at a step after the binding of RNAP and indicates a conformational effect of oxidized Fe-SoxR on the soxS promoter. The variable redox state of the SoxR FeS cluster may thus be employed in vivo to modulate the transcriptional activity of this protein in response to specific types of oxidative stress.

Full text

PDF
138

Images in this article

139Image
on p.139
141Image
on p.141
142Image
on p.142
142Image
on p.142
143Image
on p.143

Selected References

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

  1. Ames B. N. Endogenous oxidative DNA damage, aging, and cancer. Free Radic Res Commun. 1989;7(3-6):121–128. doi: 10.3109/10715768909087933. [DOI] [PubMed] [Google Scholar]
  2. Amábile-Cuevas C. F., Demple B. Molecular characterization of the soxRS genes of Escherichia coli: two genes control a superoxide stress regulon. Nucleic Acids Res. 1991 Aug 25;19(16):4479–4484. doi: 10.1093/nar/19.16.4479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ansari A. Z., Chael M. L., O'Halloran T. V. Allosteric underwinding of DNA is a critical step in positive control of transcription by Hg-MerR. Nature. 1992 Jan 2;355(6355):87–89. doi: 10.1038/355087a0. [DOI] [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. Beinert H. Recent developments in the field of iron-sulfur proteins. FASEB J. 1990 May;4(8):2483–2491. doi: 10.1096/fasebj.4.8.2185975. [DOI] [PubMed] [Google Scholar]
  6. 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]
  7. Cerutti P. A. Prooxidant states and tumor promotion. Science. 1985 Jan 25;227(4685):375–381. doi: 10.1126/science.2981433. [DOI] [PubMed] [Google Scholar]
  8. Chance B., Sies H., Boveris A. Hydroperoxide metabolism in mammalian organs. Physiol Rev. 1979 Jul;59(3):527–605. doi: 10.1152/physrev.1979.59.3.527. [DOI] [PubMed] [Google Scholar]
  9. Christman M. F., Morgan R. W., Jacobson F. S., Ames B. N. Positive control of a regulon for defenses against oxidative stress and some heat-shock proteins in Salmonella typhimurium. Cell. 1985 Jul;41(3):753–762. doi: 10.1016/s0092-8674(85)80056-8. [DOI] [PubMed] [Google Scholar]
  10. Demple B., Amábile-Cuevas C. F. Redox redux: the control of oxidative stress responses. Cell. 1991 Nov 29;67(5):837–839. doi: 10.1016/0092-8674(91)90355-3. [DOI] [PubMed] [Google Scholar]
  11. Demple B., Halbrook J. Inducible repair of oxidative DNA damage in Escherichia coli. Nature. 1983 Aug 4;304(5925):466–468. doi: 10.1038/304466a0. [DOI] [PubMed] [Google Scholar]
  12. Demple B. Regulation of bacterial oxidative stress genes. Annu Rev Genet. 1991;25:315–337. doi: 10.1146/annurev.ge.25.120191.001531. [DOI] [PubMed] [Google Scholar]
  13. Devary Y., Gottlieb R. A., Smeal T., Karin M. The mammalian ultraviolet response is triggered by activation of Src tyrosine kinases. Cell. 1992 Dec 24;71(7):1081–1091. doi: 10.1016/s0092-8674(05)80058-3. [DOI] [PubMed] [Google Scholar]
  14. Drapier J. C., Pellat C., Henry Y. Generation of EPR-detectable nitrosyl-iron complexes in tumor target cells cocultured with activated macrophages. J Biol Chem. 1991 Jun 5;266(16):10162–10167. [PubMed] [Google Scholar]
  15. Farr S. B., Kogoma T. Oxidative stress responses in Escherichia coli and Salmonella typhimurium. Microbiol Rev. 1991 Dec;55(4):561–585. doi: 10.1128/mr.55.4.561-585.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Flattery-O'Brien J., Collinson L. P., Dawes I. W. Saccharomyces cerevisiae has an inducible response to menadione which differs from that to hydrogen peroxide. J Gen Microbiol. 1993 Mar;139(3):501–507. doi: 10.1099/00221287-139-3-501. [DOI] [PubMed] [Google Scholar]
  17. Frei B., England L., Ames B. N. Ascorbate is an outstanding antioxidant in human blood plasma. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6377–6381. doi: 10.1073/pnas.86.16.6377. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gardner P. R., Fridovich I. Superoxide sensitivity of the Escherichia coli 6-phosphogluconate dehydratase. J Biol Chem. 1991 Jan 25;266(3):1478–1483. [PubMed] [Google Scholar]
  19. Georgiadis M. M., Komiya H., Chakrabarti P., Woo D., Kornuc J. J., Rees D. C. Crystallographic structure of the nitrogenase iron protein from Azotobacter vinelandii. Science. 1992 Sep 18;257(5077):1653–1659. doi: 10.1126/science.1529353. [DOI] [PubMed] [Google Scholar]
  20. Green J., Trageser M., Six S., Unden G., Guest J. R. Characterization of the FNR protein of Escherichia coli, an iron-binding transcriptional regulator. Proc Biol Sci. 1991 May 22;244(1310):137–144. doi: 10.1098/rspb.1991.0062. [DOI] [PubMed] [Google Scholar]
  21. Greenberg J. T., Demple B. A global response induced in Escherichia coli by redox-cycling agents overlaps with that induced by peroxide stress. J Bacteriol. 1989 Jul;171(7):3933–3939. doi: 10.1128/jb.171.7.3933-3939.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Greenberg J. T., Monach P., Chou J. H., Josephy P. D., Demple B. Positive control of a global antioxidant defense regulon activated by superoxide-generating agents in Escherichia coli. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6181–6185. doi: 10.1073/pnas.87.16.6181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Harman D. The aging process: major risk factor for disease and death. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5360–5363. doi: 10.1073/pnas.88.12.5360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Helmann J. D., Ballard B. T., Walsh C. T. The MerR metalloregulatory protein binds mercuric ion as a tricoordinate, metal-bridged dimer. Science. 1990 Feb 23;247(4945):946–948. doi: 10.1126/science.2305262. [DOI] [PubMed] [Google Scholar]
  25. Jamieson D. J. Saccharomyces cerevisiae has distinct adaptive responses to both hydrogen peroxide and menadione. J Bacteriol. 1992 Oct;174(20):6678–6681. doi: 10.1128/jb.174.20.6678-6681.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kappus H., Sies H. Toxic drug effects associated with oxygen metabolism: redox cycling and lipid peroxidation. Experientia. 1981 Dec 15;37(12):1233–1241. doi: 10.1007/BF01948335. [DOI] [PubMed] [Google Scholar]
  27. Karin M., Smeal T. Control of transcription factors by signal transduction pathways: the beginning of the end. Trends Biochem Sci. 1992 Oct;17(10):418–422. doi: 10.1016/0968-0004(92)90012-x. [DOI] [PubMed] [Google Scholar]
  28. Keyse S. M., Emslie E. A. Oxidative stress and heat shock induce a human gene encoding a protein-tyrosine phosphatase. Nature. 1992 Oct 15;359(6396):644–647. doi: 10.1038/359644a0. [DOI] [PubMed] [Google Scholar]
  29. Keyse S. M., Tyrrell R. M. Heme oxygenase is the major 32-kDa stress protein induced in human skin fibroblasts by UVA radiation, hydrogen peroxide, and sodium arsenite. Proc Natl Acad Sci U S A. 1989 Jan;86(1):99–103. doi: 10.1073/pnas.86.1.99. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Klausner R. D., Rouault T. A., Harford J. B. Regulating the fate of mRNA: the control of cellular iron metabolism. Cell. 1993 Jan 15;72(1):19–28. doi: 10.1016/0092-8674(93)90046-s. [DOI] [PubMed] [Google Scholar]
  31. Kolb A., Busby S., Buc H., Garges S., Adhya S. Transcriptional regulation by cAMP and its receptor protein. Annu Rev Biochem. 1993;62:749–795. doi: 10.1146/annurev.bi.62.070193.003533. [DOI] [PubMed] [Google Scholar]
  32. Kuo C. F., McRee D. E., Fisher C. L., O'Handley S. F., Cunningham R. P., Tainer J. A. Atomic structure of the DNA repair [4Fe-4S] enzyme endonuclease III. Science. 1992 Oct 16;258(5081):434–440. doi: 10.1126/science.1411536. [DOI] [PubMed] [Google Scholar]
  33. Larson C. J., Verdine G. L. A high-capacity column for affinity purification of sequence-specific DNA-binding proteins. Nucleic Acids Res. 1992 Jul 11;20(13):3525–3525. doi: 10.1093/nar/20.13.3525. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Liochev S. I., Fridovich I. Fumarase C, the stable fumarase of Escherichia coli, is controlled by the soxRS regulon. Proc Natl Acad Sci U S A. 1992 Jul 1;89(13):5892–5896. doi: 10.1073/pnas.89.13.5892. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Meyer M., Schreck R., Baeuerle P. A. H2O2 and antioxidants have opposite effects on activation of NF-kappa B and AP-1 in intact cells: AP-1 as secondary antioxidant-responsive factor. EMBO J. 1993 May;12(5):2005–2015. doi: 10.1002/j.1460-2075.1993.tb05850.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Mulliez E., Fontecave M., Gaillard J., Reichard P. An iron-sulfur center and a free radical in the active anaerobic ribonucleotide reductase of Escherichia coli. J Biol Chem. 1993 Feb 5;268(4):2296–2299. [PubMed] [Google Scholar]
  37. Nunoshiba T., Demple B. Potent intracellular oxidative stress exerted by the carcinogen 4-nitroquinoline-N-oxide. Cancer Res. 1993 Jul 15;53(14):3250–3252. [PubMed] [Google Scholar]
  38. Nunoshiba T., Hidalgo E., Amábile Cuevas C. F., Demple B. Two-stage control of an oxidative stress regulon: the Escherichia coli SoxR protein triggers redox-inducible expression of the soxS regulatory gene. J Bacteriol. 1992 Oct;174(19):6054–6060. doi: 10.1128/jb.174.19.6054-6060.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. O'Halloran T. V., Frantz B., Shin M. K., Ralston D. M., Wright J. G. The MerR heavy metal receptor mediates positive activation in a topologically novel transcription complex. Cell. 1989 Jan 13;56(1):119–129. doi: 10.1016/0092-8674(89)90990-2. [DOI] [PubMed] [Google Scholar]
  40. Parkhill J., Brown N. L. Site-specific insertion and deletion mutants in the mer promoter-operator region of Tn501; the nineteen base-pair spacer is essential for normal induction of the promoter by MerR. Nucleic Acids Res. 1990 Sep 11;18(17):5157–5162. doi: 10.1093/nar/18.17.5157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Parkinson J. S., Kofoid E. C. Communication modules in bacterial signaling proteins. Annu Rev Genet. 1992;26:71–112. doi: 10.1146/annurev.ge.26.120192.000443. [DOI] [PubMed] [Google Scholar]
  42. Reznikoff W. S., Siegele D. A., Cowing D. W., Gross C. A. The regulation of transcription initiation in bacteria. Annu Rev Genet. 1985;19:355–387. doi: 10.1146/annurev.ge.19.120185.002035. [DOI] [PubMed] [Google Scholar]
  43. Rosen D. R., Siddique T., Patterson D., Figlewicz D. A., Sapp P., Hentati A., Donaldson D., Goto J., O'Regan J. P., Deng H. X. Mutations in Cu/Zn superoxide dismutase gene are associated with familial amyotrophic lateral sclerosis. Nature. 1993 Mar 4;362(6415):59–62. doi: 10.1038/362059a0. [DOI] [PubMed] [Google Scholar]
  44. Ross W., Park S. J., Summers A. O. Genetic analysis of transcriptional activation and repression in the Tn21 mer operon. J Bacteriol. 1989 Jul;171(7):4009–4018. doi: 10.1128/jb.171.7.4009-4018.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Schreck R., Albermann K., Baeuerle P. A. Nuclear factor kappa B: an oxidative stress-responsive transcription factor of eukaryotic cells (a review). Free Radic Res Commun. 1992;17(4):221–237. doi: 10.3109/10715769209079515. [DOI] [PubMed] [Google Scholar]
  46. Storz G., Tartaglia L. A., Ames B. N. Transcriptional regulator of oxidative stress-inducible genes: direct activation by oxidation. Science. 1990 Apr 13;248(4952):189–194. doi: 10.1126/science.2183352. [DOI] [PubMed] [Google Scholar]
  47. Summers A. O. Untwist and shout: a heavy metal-responsive transcriptional regulator. J Bacteriol. 1992 May;174(10):3097–3101. doi: 10.1128/jb.174.10.3097-3101.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Tsaneva I. R., Weiss B. soxR, a locus governing a superoxide response regulon in Escherichia coli K-12. J Bacteriol. 1990 Aug;172(8):4197–4205. doi: 10.1128/jb.172.8.4197-4205.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Wu J., Weiss B. Two divergently transcribed genes, soxR and soxS, control a superoxide response regulon of Escherichia coli. J Bacteriol. 1991 May;173(9):2864–2871. doi: 10.1128/jb.173.9.2864-2871.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Wu J., Weiss B. Two-stage induction of the soxRS (superoxide response) regulon of Escherichia coli. J Bacteriol. 1992 Jun;174(12):3915–3920. doi: 10.1128/jb.174.12.3915-3920.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Yanofsky C. Comparison of regulatory and structural regions of genes of tryptophan metabolism. Mol Biol Evol. 1984 Feb;1(2):143–161. doi: 10.1093/oxfordjournals.molbev.a040307. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES