Positive control of a global antioxidant defense regulon activated by superoxide-generating agents in Escherichia coli.

Greenberg, Jean T.; Monach, Paul A.; Chou, Joseph H.; Josephy, P. David; Demple, Bruce

doi:10.1073/pnas.87.16.6181

Cited by 521 publications

(453 citation statements)

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“…In E. coli, katE is regulated by rpoS, while katG is H 2 O 2 and ascorbate induced and is regulated by oxyR (9,40,43). katX appeared to be regulated by superoxides in a fashion similar to genes in the soxRS regulon of E. coli (10,16). It can be argued that superoxide anions generated by redox-cycling agents would be converted to H 2 O 2 by superoxide dismutase (18).…”

Section: Discussionmentioning

confidence: 99%

Heterologous growth phase- and temperature-dependent expression and H2O2 toxicity protection of a superoxide-inducible monofunctional catalase gene from Xanthomonas oryzae pv. oryzae

et al. 1996

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Catalase is an important protective enzyme against H 2 O 2 toxicity. Here, we report the characterization of a Xanthomonas oryzae pv. oryzae catalase gene (katX). The gene was localized and its nucleotide sequence was determined. The gene codes for a 77-kDa polypeptide. The deduced katX amino acid sequence shares regions of high identity with other monofunctional catalases in a range of organisms from bacteria to eukaryotes. The transcriptional regulation of katX was atypical of bacterial monofunctional kat genes. Northern (RNA) analysis showed that katX transcription was highly induced by treatments with low concentrations of menadione, a superoxide generator, and methyl methanesulfonate, a mutagen. It was only weakly induced by H 2 O 2 . Unlike in other bacteria, a high level of catalase in Xanthomonas spp. provided protection from the growth-inhibitory and killing effects of H 2 O 2 but not from those of organic peroxides and superoxide generators. Unexpectedly, heterologous expression of katX in Escherichia coli was both growth phase and temperature dependent. Catalase activity in E. coli kat mutants harboring katX on an expression vector was detectable only when the cells entered the stationary phase of growth and at 28؇C. The patterns of transcription regulation, heterologous expression, and physiological function of katX are different from previously studied bacterial kat genes.Catalase is a heme-containing enzyme involved in dismutation of H 2 O 2 to oxygen and water. The enzyme plays an important role in detoxifying H 2 O 2 and minimizing oxidative stress caused by highly reactive oxygen species (ROS, i.e., OH⅐) which arise from H 2 O 2 degradation in the Fenton reaction (13). Mutations in kat have always resulted in increased sensitivity to H 2 O 2 stress, and this is an indication of the important physiological role of the enzyme (10, 13). In many bacteria, there are two types of catalase enzyme, namely a monofunctional catalase and a bifunctional catalase/peroxidase. Each enzyme is encoded by a different gene (e.g., in Escherichia coli, katE and katG code for monofunctional and bifunctional catalases, respectively) (35, 45). Monofunctional catalases share regions of an amino acid sequence that is highly conserved among microbial, plant, and mammalian enzymes (5,26,47). In many bacteria, the two kat genes are regulated differently in terms of growth phase and response to oxidative stress, suggesting that they may have different physiological functions (10,23,29).Xanthomonas oryzae pv. oryzae (Xoo) is the causative agent for the most destructive bacterial disease (the bacterial leaf blight) in rice (37). Oxidative stress is an important component of normal aerobic life and in bacterial-plant interactions. Increasing production of ROS, including superoxide, H 2 O 2 , and OH⅐, is associated with an active plant defense response and with aerobic respiration (44). ROS are highly toxic to all cellular components, and their rapid detoxification is essential for microbial survival.Xoo monofunctional catalase ...

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Section: Discussionmentioning

confidence: 99%

Heterologous growth phase- and temperature-dependent expression and H2O2 toxicity protection of a superoxide-inducible monofunctional catalase gene from Xanthomonas oryzae pv. oryzae

et al. 1996

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“…Our initial approach to delineating the cis-acting site in zwf that is involved in the soxRS-mediated response to oxidative stress was based on the observation that a zwf-lacZ operon fusion, DR52 (26), was regulated by soxRS at the transcriptional level (18). This operon fusion carries a 686-bp BclI promoter-containing zwf fragment (positions Ϫ140 to ϩ546) fused to ЈtrpA-ЈlacOZYAЈ (26).…”

Section: Resultsmentioning

confidence: 99%

“…Thus, unlike gnd, zwf expression is transcriptionally activated by SoxS during episodes of oxidative stress induced by exposure of E. coli to superoxide-generating agents such as paraquat (17,18,39). Indeed, E. coli strains deleted for zwf are hypersensitive to oxidative stress, thus making zwf a pivotal member of the soxRS regulon (18,21). In addition, treatment of cells with several antibiotics and aromatic weak acids appears to activate zwf transcription through the action of marA, thus making zwf a member of the multiple antibiotic resistance (mar) regulon (2,6,7,14,16).…”

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Genetic definition of the Escherichia coli zwf "soxbox," the DNA binding site for SoxS-mediated induction of glucose 6-phosphate dehydrogenase in response to superoxide

1995

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In Escherichia coli K-12, transcription of zwf, the gene for glucose 6-phosphate dehydrogenase, is subject to growth rate-dependent regulation and is activated by SoxS in response to superoxide stress. To define genetically the site of SoxS activation, we undertook a detailed deletion analysis of the zwf promoter region. Using specifically targeted 5 and 3 deletions of zwf sequences, we localized the SoxS activation site to a 21-bp region upstream of the zwf promoter. This minimal ''soxbox'' was able to confer paraquat inducibility when placed upstream of a normally unresponsive gnd-lacZ protein fusion. In addition, we used these findings as the basis for resecting unnecessary sequences from the region upstream of the promoters of two other SoxSregulated genes, sodA and nfo. Like the zwf soxbox, the regions required for SoxS activation of sodA and nfo appear to lie just upstream of or overlap the ؊35 hexamers of the corresponding promoters. Importantly, the sequence boundaries established here by deletion analysis agree with the primary SoxS recognition sites of zwf, sodA, and nfo that we previously identified in vitro by gel mobility shift and DNase I protection assays with a purified MalE-SoxS fusion protein.In Escherichia coli, the oxidative branch of the pentose phosphate pathway provides ribose for nucleoside biosynthesis and NADPH for reductive biosynthesis (9, 12). Two of the enzymes of this pathway, glucose 6-phosphate dehydrogenase (encoded by zwf) and 6-phosphogluconate dehydrogenase (encoded by gnd) are subject to growth rate-dependent regulation (40). The specific activities of these enzymes increase in proportion to increased growth rate during steady-state growth on different carbon sources. Studies on the growth rate-dependent regulation of zwf and gnd have shown that at least part of the increase in the specific activities of these enzymes is attributable to an increase in the transcription rates of the respective genes (25,26).Apart from its basic growth rate-dependent regulation, zwf is also a member of at least two other regulons. Thus, unlike gnd, zwf expression is transcriptionally activated by SoxS during episodes of oxidative stress induced by exposure of E. coli to superoxide-generating agents such as paraquat (17,18,39). Indeed, E. coli strains deleted for zwf are hypersensitive to oxidative stress, thus making zwf a pivotal member of the soxRS regulon (18, 21). In addition, treatment of cells with several antibiotics and aromatic weak acids appears to activate zwf transcription through the action of marA, thus making zwf a member of the multiple antibiotic resistance (mar) regulon (2,6,7,14,16).The work presented here has several overlapping long-range objectives. One is to understand the physiological basis for zwf's membership in the soxRS and marRAB regulons. The other is to identify the cis-acting regulatory sites of zwf that are involved in superoxide control for subsequent comparison with the sites required for growth rate control and for induction by antibiotics and aromatic weak...

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“…The closely related compound, 2-methyl-1,4-naphthoquinol-bis-disodium phosphate (synkavit), has been used to enhance the effect of ionizing radiation in patients being treated for cancer (Deeley, 1962). Menadione has also been used as a model compound to investigate the effects of oxidative stress in both prokaryotic and eukaryotic cells (Thor et al, 1982;Greenberg et al, 1990).…”

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Menadione-resistant Chinese hamster ovary cells have an increased capacity for glutathione synthesis

Vallis

Reglinski²,

Garner³

et al. 1997

Br J Cancer

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Summary A cell line (MRc4O) resistant to the model quinone compound, menadione, has been isolated from a parental Chinese hamster ovary cell line (CHO-K1). The known relationship between menadione toxicity and glutathione (GSH) depletion led us to investigate whether the mechanism of resistance of MRc40 was related to alteration in GSH homeostasis. Intracellular concentrations of GSH and cysteine (CySH) were twofold and 3.2-fold greater in MRc40 than in CHO-Kl. Following exposure to menadione, GSH and CySH were depleted, but subsequent recovery of thiols was more rapid and of greater magnitude in MRc40 than in CHO-Kl. Twelve hours after exposure to menadione, the concentrations of GSH and CySH were 9.7-and 4.2-fold greater in MRc40 than in CHO-Kl. Using nuclear magnetic resonance (NMR) spectroscopy, we observed the in situ removal of menadione from cell suspensions of CHO-Kl and MRc40. However, only in CHO-Kl did we observe concomitant depletion of NMR-visible GSH. We conclude that the perturbation of GSH metabolism contributes to the resistant phenotype and is an important characteristic of menadione-resistant CHO cells.

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Positive control of a global antioxidant defense regulon activated by superoxide-generating agents in Escherichia coli.

Abstract: Escherichia coli responds to superoxidegenerating agents by inducing w4O proteins. We have identified a genetic locus, soxR (superoxide response), that positively regulates 9 of these proteins during superoxide stress.

Cited by 521 publications

References 37 publications

Heterologous growth phase- and temperature-dependent expression and H2O2 toxicity protection of a superoxide-inducible monofunctional catalase gene from Xanthomonas oryzae pv. oryzae

Heterologous growth phase- and temperature-dependent expression and H2O2 toxicity protection of a superoxide-inducible monofunctional catalase gene from Xanthomonas oryzae pv. oryzae

Genetic definition of the Escherichia coli zwf "soxbox," the DNA binding site for SoxS-mediated induction of glucose 6-phosphate dehydrogenase in response to superoxide

Menadione-resistant Chinese hamster ovary cells have an increased capacity for glutathione synthesis

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