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Nonnative disulfide bond formation activates the ?32-dependent heat shock response in Escherichia coli.

ABSTRACT: Formation of nonnative disulfide bonds in the cytoplasm, so-called disulfide stress, is an integral component of oxidative stress. Quantification of the extent of disulfide bond formation in the cytoplasm of Escherichia coli revealed that disulfide stress is associated with oxidative stress caused by hydrogen peroxide, paraquat, and cadmium. To separate the impact of disulfide bond formation from unrelated effects of these oxidative stressors in subsequent experiments, we worked with two complementary approaches. We triggered disulfide stress either chemically by diamide treatment of cells or genetically in a mutant strain lacking the major disulfide-reducing systems TrxB and Gor. Studying the proteomic response of E. coli exposed to disulfide stress, we found that intracellular disulfide bond formation is a particularly strong inducer of the heat shock response. Real-time quantitative PCR experiments showed that disulfide stress induces the heat shock response in E. coli ?(32) dependently. However, unlike heat shock treatment, which induces these genes transiently, transcripts of ?(32)-dependent genes accumulated over time in disulfide stress-treated cells. Analyzing the stability of ?(32), we found that this constant induction can be attributed to an increase of the half-life of ?(32) upon disulfide stress. This is concomitant with aggregation of E. coli proteins treated with diamide. We conclude that oxidative stress triggers the heat shock response in E. coli ?(32) dependently. The component of oxidative stress responsible for the induction of heat shock genes is disulfide stress. Nonnative disulfide bond formation in the cytoplasm causes protein unfolding. This stabilizes ?(32) by preventing its DnaK- and FtsH-dependent degradation.

SUBMITTER: Muller A

PROVIDER: S-EPMC3697257 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature

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Nonnative disulfide bond formation activates the σ32-dependent heat shock response in Escherichia coli.

Müller Alexandra A Hoffmann Jörg H JH Meyer Helmut E HE Narberhaus Franz F Jakob Ursula U Leichert Lars I LI

Journal of bacteriology 20130412 12

Formation of nonnative disulfide bonds in the cytoplasm, so-called disulfide stress, is an integral component of oxidative stress. Quantification of the extent of disulfide bond formation in the cytoplasm of Escherichia coli revealed that disulfide stress is associated with oxidative stress caused by hydrogen peroxide, paraquat, and cadmium. To separate the impact of disulfide bond formation from unrelated effects of these oxidative stressors in subsequent experiments, we worked with two complem ...[more]

PMID: 23585533

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Project description:The envelope of gram-negative bacteria serves as the first line of defense against environmental insults. Therefore, its integrity is continuously monitored and maintained by several envelope stress response (ESR) systems. Due to its oxidizing environment, the envelope represents an important site for disulfide bond formation. In Escherichia coli, the periplasmic oxidoreductase, DsbA introduces disulfide bonds in substrate proteins and transfers electrons to the inner membrane oxidoreductase, DsbB. Under aerobic conditions, the reduced form of DsbB is re-oxidized by ubiquinone, an electron carrier in the electron transport chain (ETC). Given the critical role of ubiquinone in transferring electrons derived from the oxidation of reduced cofactors, we were intrigued whether metabolic conditions that generate a large number of reduced cofactors render ubiquinone unavailable for disulfide bond formation. To test this, here we investigated the influence of metabolism of long-chain fatty acid (LCFA), an energy-rich carbon source, on the redox state of the envelope. We show that LCFA degradation increases electron flow in the ETC. Further, whereas cells metabolizing LCFAs exhibit characteristics of insufficient disulfide bond formation, these hallmarks are averted in cells exogenously provided with ubiquinone. Importantly, the ESR pathways, Cpx and ?E, are activated by envelope signals generated during LCFA metabolism. Our results argue that Cpx is the primary ESR that senses and maintains envelope redox homeostasis. Amongst the two ESRs, Cpx is induced to a greater extent by LCFAs and senses redox-dependent signal. Further, ubiquinone accumulation during LCFA metabolism is prevented in cells lacking Cpx response, suggesting that Cpx activation helps maintain redox homeostasis by increasing the oxidizing power for disulfide bond formation. Taken together, our results demonstrate an intricate relationship between cellular metabolism and disulfide bond formation dictated by ETC and ESR, and provide the basis for examining whether similar mechanisms control envelope redox status in other gram-negative bacteria.

Dataset Information

Nonnative disulfide bond formation activates the ?32-dependent heat shock response in Escherichia coli.

Publications

Nonnative disulfide bond formation activates the σ32-dependent heat shock response in Escherichia coli.

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