Abstract
We report the first example of a gene, hmp, encoding a soluble flavohemoglobin in Escherichia coli K-12, which is up-regulated by paraquat in a SoxRS-independent manner. Unlike what is found for other paraquat-inducible genes, high concentrations of paraquat (200 microM) were required to increase the level of hmp expression, and maximal induction was observed only after 20 min of exposure to paraquat. Neither a mutation in soxS nor one in soxR prevented the paraquat-dependent increase in phi(hmp-lacZ) expression, but either mutant allele delayed full expression of phi(hmp-lacZ) activity after paraquat addition. Induction of hmp by paraquat was demonstrated in aerobically grown cultures during exponential growth and the stationary phase, thus revealing two Sox-independent regulatory mechanisms. Induction of hmp by paraquat in the stationary phase was dependent on the global regulator of stationary-phase gene expression, RpoS (sigma S). However, a mutation in rpoS did not prevent an increase in hmp expression by paraquat in exponentially growing cells. Induction of sigma S in the exponential phase by heat shock also induced phi(hmp-lacZ) expression in the presence of paraquat, supporting the role of sigma S in one of the regulatory mechanisms. Mutations in oxyR or rob, known regulators of several stress promoters in E. coli, had no effect on the induction of hmp by paraquat. Other known superoxide-generating agents (plumbagin, menadione, and phenazine methosulfate) were not effective in inducing hmp expression.
Full Text
The Full Text of this article is available as a PDF (182.4 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Altuvia S., Almirón M., Huisman G., Kolter R., Storz G. The dps promoter is activated by OxyR during growth and by IHF and sigma S in stationary phase. Mol Microbiol. 1994 Jul;13(2):265–272. doi: 10.1111/j.1365-2958.1994.tb00421.x. [DOI] [PubMed] [Google Scholar]
- 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]
- Ariza R. R., Li Z., Ringstad N., Demple B. Activation of multiple antibiotic resistance and binding of stress-inducible promoters by Escherichia coli Rob protein. J Bacteriol. 1995 Apr;177(7):1655–1661. doi: 10.1128/jb.177.7.1655-1661.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chung C. T., Niemela S. L., Miller R. H. One-step preparation of competent Escherichia coli: transformation and storage of bacterial cells in the same solution. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2172–2175. doi: 10.1073/pnas.86.7.2172. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Compan I., Touati D. Interaction of six global transcription regulators in expression of manganese superoxide dismutase in Escherichia coli K-12. J Bacteriol. 1993 Mar;175(6):1687–1696. doi: 10.1128/jb.175.6.1687-1696.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cramm R., Siddiqui R. A., Friedrich B. Primary sequence and evidence for a physiological function of the flavohemoprotein of Alcaligenes eutrophus. J Biol Chem. 1994 Mar 11;269(10):7349–7354. [PubMed] [Google Scholar]
- 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]
- Ermler U., Siddiqui R. A., Cramm R., Friedrich B. Crystal structure of the flavohemoglobin from Alcaligenes eutrophus at 1.75 A resolution. EMBO J. 1995 Dec 15;14(24):6067–6077. doi: 10.1002/j.1460-2075.1995.tb00297.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Favey S., Labesse G., Vouille V., Boccara M. Flavohaemoglobin HmpX: a new pathogenicity determinant in Erwinia chrysanthemi strain 3937. Microbiology. 1995 Apr;141(Pt 4):863–871. doi: 10.1099/13500872-141-4-863. [DOI] [PubMed] [Google Scholar]
- Giacomini A., Corich V., Ollero F. J., Squartini A., Nuti M. P. Experimental conditions may affect reproducibility of the beta-galactosidase assay. FEMS Microbiol Lett. 1992 Dec 15;100(1-3):87–90. doi: 10.1111/j.1574-6968.1992.tb14024.x. [DOI] [PubMed] [Google Scholar]
- 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]
- Hengge-Aronis R. Survival of hunger and stress: the role of rpoS in early stationary phase gene regulation in E. coli. Cell. 1993 Jan 29;72(2):165–168. doi: 10.1016/0092-8674(93)90655-a. [DOI] [PubMed] [Google Scholar]
- Imlay J., Fridovich I. Exogenous quinones directly inhibit the respiratory NADH dehydrogenase in Escherichia coli. Arch Biochem Biophys. 1992 Jul;296(1):337–346. doi: 10.1016/0003-9861(92)90581-g. [DOI] [PubMed] [Google Scholar]
- Ioannidis N., Cooper C. E., Poole R. K. Spectroscopic studies on an oxygen-binding haemoglobin-like flavohaemoprotein from Escherichia coli. Biochem J. 1992 Dec 1;288(Pt 2):649–655. doi: 10.1042/bj2880649. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Iuchi S., Lin E. C. arcA (dye), a global regulatory gene in Escherichia coli mediating repression of enzymes in aerobic pathways. Proc Natl Acad Sci U S A. 1988 Mar;85(6):1888–1892. doi: 10.1073/pnas.85.6.1888. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jishage M., Ishihama A. Regulation of RNA polymerase sigma subunit synthesis in Escherichia coli: intracellular levels of sigma 70 and sigma 38. J Bacteriol. 1995 Dec;177(23):6832–6835. doi: 10.1128/jb.177.23.6832-6835.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kakeda M., Ueguchi C., Yamada H., Mizuno T. An Escherichia coli curved DNA-binding protein whose expression is affected by the stationary phase-specific sigma factor sigma S. Mol Gen Genet. 1995 Sep 20;248(5):629–634. doi: 10.1007/BF02423459. [DOI] [PubMed] [Google Scholar]
- Kawamukai M., Utsumi R., Takeda K., Higashi A., Matsuda H., Choi Y. L., Komano T. Nucleotide sequence and characterization of the sfs1 gene: sfs1 is involved in CRP*-dependent mal gene expression in Escherichia coli. J Bacteriol. 1991 Apr;173(8):2644–2648. doi: 10.1128/jb.173.8.2644-2648.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kogoma T., Farr S. B., Joyce K. M., Natvig D. O. Isolation of gene fusions (soi::lacZ) inducible by oxidative stress in Escherichia coli. Proc Natl Acad Sci U S A. 1988 Jul;85(13):4799–4803. doi: 10.1073/pnas.85.13.4799. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Koh Y. S., Roe J. H. Dual regulation of the paraquat-inducible gene pqi-5 by SoxS and RpoS in Escherichia coli. Mol Microbiol. 1996 Oct;22(1):53–61. doi: 10.1111/j.1365-2958.1996.tb02655.x. [DOI] [PubMed] [Google Scholar]
- Koh Y. S., Roe J. H. Isolation of a novel paraquat-inducible (pqi) gene regulated by the soxRS locus in Escherichia coli. J Bacteriol. 1995 May;177(10):2673–2678. doi: 10.1128/jb.177.10.2673-2678.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- LaCelle M., Kumano M., Kurita K., Yamane K., Zuber P., Nakano M. M. Oxygen-controlled regulation of the flavohemoglobin gene in Bacillus subtilis. J Bacteriol. 1996 Jul;178(13):3803–3808. doi: 10.1128/jb.178.13.3803-3808.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Liochev S. I., Fridovich I. Paraquat diaphorases in Escherichia coli. Free Radic Biol Med. 1994 May;16(5):555–559. doi: 10.1016/0891-5849(94)90055-8. [DOI] [PubMed] [Google Scholar]
- Membrillo-Hernández J., Ioannidis N., Poole R. K. The flavohaemoglobin (HMP) of Escherichia coli generates superoxide in vitro and causes oxidative stress in vivo. FEBS Lett. 1996 Mar 11;382(1-2):141–144. doi: 10.1016/0014-5793(96)00154-8. [DOI] [PubMed] [Google Scholar]
- Mito S., Zhang Q. M., Yonei S. Isolation and characterization of Escherichia coli strains containing new gene fusions (soi::lacZ) inducible by superoxide radicals. J Bacteriol. 1993 May;175(9):2645–2651. doi: 10.1128/jb.175.9.2645-2651.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Muffler A., Barth M., Marschall C., Hengge-Aronis R. Heat shock regulation of sigmaS turnover: a role for DnaK and relationship between stress responses mediated by sigmaS and sigma32 in Escherichia coli. J Bacteriol. 1997 Jan;179(2):445–452. doi: 10.1128/jb.179.2.445-452.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Nunoshiba T., deRojas-Walker T., Wishnok J. S., Tannenbaum S. R., Demple B. Activation by nitric oxide of an oxidative-stress response that defends Escherichia coli against activated macrophages. Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):9993–9997. doi: 10.1073/pnas.90.21.9993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Poole R. K., Anjum M. F., Membrillo-Hernández J., Kim S. O., Hughes M. N., Stewart V. Nitric oxide, nitrite, and Fnr regulation of hmp (flavohemoglobin) gene expression in Escherichia coli K-12. J Bacteriol. 1996 Sep;178(18):5487–5492. doi: 10.1128/jb.178.18.5487-5492.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Poole R. K. Oxygen reactions with bacterial oxidases and globins: binding, reduction and regulation. Antonie Van Leeuwenhoek. 1994;65(4):289–310. doi: 10.1007/BF00872215. [DOI] [PubMed] [Google Scholar]
- Simons R. W., Houman F., Kleckner N. Improved single and multicopy lac-based cloning vectors for protein and operon fusions. Gene. 1987;53(1):85–96. doi: 10.1016/0378-1119(87)90095-3. [DOI] [PubMed] [Google Scholar]
- Straus D. B., Walter W. A., Gross C. A. The heat shock response of E. coli is regulated by changes in the concentration of sigma 32. Nature. 1987 Sep 24;329(6137):348–351. doi: 10.1038/329348a0. [DOI] [PubMed] [Google Scholar]
- Touati D., Jacques M., Tardat B., Bouchard L., Despied S. Lethal oxidative damage and mutagenesis are generated by iron in delta fur mutants of Escherichia coli: protective role of superoxide dismutase. J Bacteriol. 1995 May;177(9):2305–2314. doi: 10.1128/jb.177.9.2305-2314.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vasudevan S. G., Armarego W. L., Shaw D. C., Lilley P. E., Dixon N. E., Poole R. K. Isolation and nucleotide sequence of the hmp gene that encodes a haemoglobin-like protein in Escherichia coli K-12. Mol Gen Genet. 1991 Apr;226(1-2):49–58. doi: 10.1007/BF00273586. [DOI] [PubMed] [Google Scholar]
- Walkup L. K., Kogoma T. Escherichia coli proteins inducible by oxidative stress mediated by the superoxide radical. J Bacteriol. 1989 Mar;171(3):1476–1484. doi: 10.1128/jb.171.3.1476-1484.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]