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Lifespan extension and paraquat resistance in a ubiquinone-deficient Escherichia coli mutant depend on transcription factors ArcA and TdcA.

ABSTRACT: We recently reported a genome-wide screen for extended stationary phase survival in Escherichia coli. One of the mutants recovered is deleted for ubiG, which encodes a methyltransferase required for the biosynthesis of ubiquinone. The ubiG mutant exhibits longer lifespan, as well as enhanced resistance to thermal and oxidative stress compared to wt at extracellular pH9. The longevity of the mutant, as well as its resistance to the superoxide-generating agent paraquat, is partially dependent on the hypoxia-inducible transcription factor ArcA. A microarray analysis revealed several genes whose expression is either suppressed or enhanced by ArcA in the ubiG mutant. TdcA is a transcription factor involved in the transport and metabolism of amino acids during anaerobic growth. Its enhanced expression in the ubiG mutant is dependent on ArcA. Our data are consistent with the hypothesis that ArcA and TdcA function in the same genetic pathway to increase lifespan and enhance oxidative stress resistance in the ubiG mutant. Our results might be relevant for the elucidation of the mechanism of lifespan extension in mutant mice and worms bearing mutations in ubiquinone biosynthetic genes.

SUBMITTER: Gonidakis S

PROVIDER: S-EPMC3091522 | biostudies-literature | 2011 Mar

REPOSITORIES: biostudies-literature

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Lifespan extension and paraquat resistance in a ubiquinone-deficient Escherichia coli mutant depend on transcription factors ArcA and TdcA.

Gonidakis Stavros S Finkel Steven E SE Longo Valter D VD

Aging 20110301 3

We recently reported a genome-wide screen for extended stationary phase survival in Escherichia coli. One of the mutants recovered is deleted for ubiG, which encodes a methyltransferase required for the biosynthesis of ubiquinone. The ubiG mutant exhibits longer lifespan, as well as enhanced resistance to thermal and oxidative stress compared to wt at extracellular pH9. The longevity of the mutant, as well as its resistance to the superoxide-generating agent paraquat, is partially dependent on t ...[more]

PMID: 21464517

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Project description:1. The function of ubiquinone in Escherichia coli was studied by using whole cells and membrane preparations of normal E. coli and of a mutant lacking ubiquinone. 2. The mutant lacking ubiquinone, strain AN59 (Ubi(-)), when grown under aerobic conditions, gave an anaerobic type of growth yield and produced large quantities of lactic acid, indicating that ubiquinone plays a vital role in electron transport. 3. NADH and lactate oxidase activities in membranes from strain AN59 (Ubi(-)) were greatly impaired and activity was restored by the addition of ubiquinone (Q-1). 4. Comparison of the percentage reduction of flavin, cytochrome b(1) and cytochrome a(2) in the aerobic steady state in membranes from the normal strain (AN62) and strain AN59 (Ubi(-)) and the effect of respiratory inhibitors on these percentages in membranes from strain AN62 suggest that ubiquinone functions at more than one site in the electron-transport chain. 5. Membranes from strain AN62, in the absence of substrate, showed an electron-spin-resonance signal attributed to ubisemiquinone. The amount of reduced ubiquinone (50%) found after rapid solvent extraction is consistent with the existence of ubiquinone in membranes as a stabilized ubisemiquinone. 6. The effects of piericidin A on membranes from strain AN62 suggest that this inhibitor acts at the ubiquinone sites: thus inhibition of electron transport is reversed by ubiquinone (Q-1); the aerobic steady-state oxidation-reduction levels of flavins and cytochrome b(1) in the presence of the inhibitor are raised to values approximating those found in the membranes of strain AN59 (Ubi(-)); the inhibitor rapidly eliminates the electron-spin-resonance signal attributed to ubisemiquinone and allows slow oxidation of endogenous ubiquinol in the absence of substrate and prevents reduction of ubiquinone in the presence of substrate. It is concluded that piericidin A separates ubiquinone from the remainder of the electron-transport chain. 7. A scheme is proposed in which ubisemiquinone, complexed to an electron carrier, functions in at least two positions in the electron-transport sequence.

Project description:There is an urgent need for strategies to discover secondary drugs to prevent or disrupt antimicrobial resistance (AMR), which is causing >700,000 deaths annually. Here, we demonstrate that tetracycline-resistant (TetR) Escherichia coli undergoes global transcriptional and metabolic remodeling, including downregulation of tricarboxylic acid cycle and disruption of redox homeostasis, to support consumption of the proton motive force for tetracycline efflux. Using a pooled genome-wide library of single-gene deletion strains, at least 308 genes, including four transcriptional regulators identified by our network analysis, were confirmed as essential for restoring the fitness of TetR E. coli during treatment with tetracycline. Targeted knockout of ArcA, identified by network analysis as a master regulator of this new compensatory physiological state, significantly compromised fitness of TetR E. coli during tetracycline treatment. A drug, sertraline, which generated a similar metabolome profile as the arcA knockout strain, also resensitized TetR E. coli to tetracycline. We discovered that the potentiating effect of sertraline was eliminated upon knocking out arcA, demonstrating that the mechanism of potential synergy was through action of sertraline on the tetracycline-induced ArcA network in the TetR strain. Our findings demonstrate that therapies that target mechanistic drivers of compensatory physiological states could resensitize AMR pathogens to lost antibiotics. IMPORTANCE Antimicrobial resistance (AMR) is projected to be the cause of >10 million deaths annually by 2050. While efforts to find new potent antibiotics are effective, they are expensive and outpaced by the rate at which new resistant strains emerge. There is desperate need for a rational approach to accelerate the discovery of drugs and drug combinations that effectively clear AMR pathogens and even prevent the emergence of new resistant strains. Using tetracycline-resistant (TetR) Escherichia coli, we demonstrate that gaining resistance is accompanied by loss of fitness, which is restored by compensatory physiological changes. We demonstrate that transcriptional regulators of the compensatory physiologic state are promising drug targets because their disruption increases the susceptibility of TetR E. coli to tetracycline. Thus, we describe a generalizable systems biology approach to identify new vulnerabilities within AMR strains to rationally accelerate the discovery of therapeutics that extend the life span of existing antibiotics.

Dataset Information

Lifespan extension and paraquat resistance in a ubiquinone-deficient Escherichia coli mutant depend on transcription factors ArcA and TdcA.

Publications

Lifespan extension and paraquat resistance in a ubiquinone-deficient Escherichia coli mutant depend on transcription factors ArcA and TdcA.

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