Quinone Reduction by the Na<sup>+</sup>-Translocating NADH Dehydrogenase Promotes Extracellular Superoxide Production in<i>Vibrio cholerae</i>

Lin, Po-Chi; Türk, Karin; Häse, Claudia C.; Fritz, Günter; Steuber, Julia

doi:10.1128/jb.01651-06

Cited by 37 publications

(22 citation statements)

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“…We favor another explanation for the observed stabilization of catecholates by V. cholerae growing in serum-SAPI. Distinct conversion products of NE (eluting at 5.4 min) and of E (eluting at 5.8 min) were detected in the V. cholerae cultures grown for 48 h. Most likely, these compounds are formed by oxidation of NE or E by O 2 or, in a faster reaction, by an extracellular superoxide, O 2 Ϫ , formed by V. cholerae during respiration (42). In the case of E, a possible candidate is leucochrome, which further reacts to adrenochrome (28).…”

Section: Discussionmentioning

confidence: 99%

“…We propose that compared to bacteria in the cell-free medium, growing bacteria provide a more reducing environment that slows down the oxidative degradation of NE or E. First, O 2 tension is lowered due to the metabolic activity of the bacteria; second, reactive oxygen species (ROS) like superoxide, which accelerate the oxidative degradation of NE or E (28) and which are formed during the initial oxidation step (44), are eliminated by ROS-protective enzymes like the periplasmic superoxide dismutase of V. cholerae (42). To the best of our knowledge, qseC expression levels in bacteria exposed to NE or E during growth have not yet been correlated with the actual concentrations of these signaling compounds in the medium.…”

Section: Discussionmentioning

confidence: 99%

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Response of Vibrio cholerae to the Catecholamine Hormones Epinephrine and Norepinephrine

et al. 2015

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In Escherichia coli or Salmonella enterica, the stress-associated mammalian hormones epinephrine (E) and norepinephrine (NE) trigger a signaling cascade by interacting with the QseC sensor protein. Here we show that Vibrio cholerae, the causative agent of cholera, exhibits a specific response to E and NE. These catecholates (0.1 mM) enhanced the growth and swimming motility of V. cholerae strain O395 on soft agar in a medium containing calf serum, which simulated the environment within the host. During growth, the hormones were converted to degradation products, including adrenochrome formed by autooxidation with O 2 or superoxide. In E. coli, the QseC sensor kinase, which detects the autoinducer AI-3, also senses E or NE. The genome of V. cholerae O395 comprises an open reading frame coding for a putative protein with 29% identity to E. coli QseC. Quantitative reverse transcriptase PCR (qRT-PCR) experiments revealed increased transcript levels of the qseC-like gene and of pomB, a gene encoding a structural component of the flagellar motor complex, under the influence of E or NE. Phentolamine blocks the response of E. coli QseC to E or NE. A V. cholerae mutant devoid of the qseC-like gene retained the phentolamine-sensitive motility in the presence of E, whereas NE-stimulated motility was no longer inhibited by phentolamine. Our study demonstrates that V. cholerae senses the stress hormones E and NE. A sensor related to the histidine kinase QseC from E. coli is identified and is proposed to participate in the sensing of NE. IMPORTANCEVibrio cholerae is a Gram-negative bacterium that may cause cholera, a severe illness with high mortality due to acute dehydration caused by diarrhea and vomiting. Pathogenic V. cholerae strains possess virulence factors like the cholera toxin (CTX) and the toxin-coregulated pilus (TCP) produced in response to signals provided by the host. In pathogenic enterobacteria, the stressassociated hormones epinephrine (E) and norepinephrine (NE) of the human host act as signal molecules for the production of virulence factors and promote bacterial growth by the sequestration of iron from the host. Here we show that V. cholerae, like some enterobacteria, benefits from these stress hormones and possesses a sensor to recognize them. Stress enhances the susceptibility of a mammalian host to infection by bacteria. The stress-associated mammalian hormones epinephrine (E) and norepinephrine (NE), for example, support the growth (1-3) and motility (4, 5) of the enterobacteria Escherichia coli, Salmonella enterica serovar Typhimurium, and Klebsiella pneumoniae (6), of Pseudomonas aeruginosa (7,8), and of some Vibrio species (9) in a serum-based bacteriostatic medium. As E and NE share structural similarities to the bacterial catechol siderophores-a benzene ring with two adjacent hydroxyl groups-it was suggested that these stress hormones may be involved in the binding and uptake of iron by bacteria, thus promoting growth or motility (1-5). Recently, the mechanism by which catecholamine hormo...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Response of Vibrio cholerae to the Catecholamine Hormones Epinephrine and Norepinephrine

et al. 2015

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“…V. cholerae operates a redox-driven Na ϩ pump in the NADH:quinone segment of its respiratory chain, but it does not possess a NADH dehydrogenase belonging to the complex I family of enzymes (2). The Na ϩ -translocating NADH:quinone oxidoreductase produced superoxide in vitro (8) and participated in the extracellular formation of superoxide by V. cholerae in vivo (30). We compared the ROS formation activities of membranes from different V. cholerae strains with various amounts of active NQR.…”

Section: Resultsmentioning

confidence: 99%

“…The strong fluorescence of cells suggested that DCF was localized inside the cell, but since V. cholerae is a Gram-negative bacterium, a periplasmic localization of the fluorophore was also considered. If the fluorophore were predominantly accumulated in the periplasm surrounding the cell, formation of DCF fluorescence would reflect extracellular formation of superoxide (30). To address this question, V. cholerae cells loaded with DCFH-DA were treated with lysozyme which results in the formation of spheroplasts due to lysis of the peptidoglycan layer.…”

Section: Resultsmentioning

confidence: 99%

“…With the exception of membranes from V. cholerae ⌬nqr transformed with pNqr1, which were diluted to 0.25 mg protein ml Ϫ1 , all membranes were adjusted to 1 mg protein ml Ϫ1 with 50 mM sodium phosphate (pH 8.0), 0.3 M NaCl, 5% (wt/vol) glycerol. The oxidation of NADH (disodium salt; Sigma-Aldrich) by membranes in the presence of ubiquinone-1 (Q1) (MCAT GmbH) was accompanied by the formation of superoxide which reacted with oxidized cytochrome c (from bovine heart; Sigma-Aldrich) to yield reduced cytochrome c in equimolar amounts (30). The activity was determined in 20 mM Tris-H 2 SO 4 (pH 8.0) with 50 mM Na 2 SO 4 , 50 g BSA ml Ϫ1 , 5% (wt/vol) glycerol, 100 M Q1, 40 M cytochrome c, and 100 M NADH.…”

Section: Strains and Plasmids Vibrio Cholerae O395n1 (17) And Its Vamentioning

confidence: 99%

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The Na ⁺ -Translocating NADH:Quinone Oxidoreductase Enhances Oxidative Stress in the Cytoplasm of Vibrio cholerae

et al. 2016

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We searched for a source of reactive oxygen species (ROS) in the cytoplasm of the human pathogen Vibrio cholerae and addressed the mechanism of ROS formation using the dye 2=,7=-dichlorofluorescein diacetate (DCFH-DA) in respiring cells. By comparing V. cholerae strains with or without active Na ؉ -translocating NADH:quinone oxidoreductase (Na ؉ -NQR), this respiratory sodium ion redox pump was identified as a producer of ROS in vivo. The amount of cytoplasmic ROS detected in V. cholerae cells producing variants of Na ؉ -NQR correlated well with rates of superoxide formation by the corresponding membrane fractions. Membranes from wild-type V. cholerae showed increased superoxide production activity (9.8 ؎ 0.6 mol superoxide min ؊1 mg ؊1 membrane protein) compared to membranes from the mutant lacking Na ؉ -NQR (0.18 ؎ 0.01 mol min ؊1 mg ؊1 ). Overexpression of plasmid-encoded Na ؉ -NQR in the nqr deletion strain resulted in a drastic increase in the formation of superoxide (42.6 ؎ 2.8 mol min ؊1 mg ؊1 ). By analyzing a variant of Na ؉ -NQR devoid of quinone reduction activity, we identified the reduced flavin adenine dinucleotide (FAD) cofactor of cytoplasmic NqrF subunit as the site for intracellular superoxide formation in V. cholerae. The impact of superoxide formation by the Na ؉ -NQR on the virulence of V. cholerae is discussed. IMPORTANCEIn several studies, it was demonstrated that the Na ؉ -NQR in V. cholerae affects virulence in a yet unknown manner. We identified the reduced FAD cofactor in the NADH-oxidizing NqrF subunit of the Na ؉ -NQR as the site of superoxide formation in the cytoplasm of V. cholerae. Our study provides the framework to understand how reactive oxygen species formed during respiration could participate in the regulated expression of virulence factors during the transition from aerobic to microaerophilic (intestinal) habitats. This hypothesis may turn out to be right for many other pathogens which, like V. cholerae, depend on the Na ؉ -NQR as the sole electrogenic NADH dehydrogenase. Vibrio cholerae is a Gram-negative bacterium and the causative agent of cholera, a devastating diarrheal disease. It is found in estuaries, inhabiting seawater or brackish water where it is commonly associated with biofilms on phyto-or zooplankton (1). The high salinity in these habitats is counteracted by an electrochemical sodium motive force (SMF) generated by the respiratory chain of V. cholerae (2, 3). This SMF drives cellular processes like substrate transport (4) and motility (5). It is generated by the Na ϩ -translocating NADH:quinone oxidoreductase (Na ϩ -NQR), which couples the energy that is released by oxidation of NADH with ubiquinone to the transport of sodium ions out of the cytoplasm with a 1e Ϫ /1Na ϩ stoichiometry (6). The Na ϩ -NQR is a membrane-bound protein complex consisting of six subunits, NqrABCDEF, and contains a variety of different cofactors (3). Electron transfer from NADH to ubiquinone is mediated by a noncovalently bound flavin adenine dinucleotide (FAD) and a [2Fe-2...

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Respiratory Membrane Protein Complexes Convert Chemical Energy

Muras

Toulouse

Fritz

et al. 2019

Subcellular Biochemistry

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Quinone Reduction by the Na⁺-Translocating NADH Dehydrogenase Promotes Extracellular Superoxide Production inVibrio cholerae

Cited by 37 publications

References 37 publications

Response of Vibrio cholerae to the Catecholamine Hormones Epinephrine and Norepinephrine

Response of Vibrio cholerae to the Catecholamine Hormones Epinephrine and Norepinephrine

The Na ⁺ -Translocating NADH:Quinone Oxidoreductase Enhances Oxidative Stress in the Cytoplasm of Vibrio cholerae

Respiratory Membrane Protein Complexes Convert Chemical Energy

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Quinone Reduction by the Na+-Translocating NADH Dehydrogenase Promotes Extracellular Superoxide Production inVibrio cholerae

Cited by 37 publications

References 37 publications

Response of Vibrio cholerae to the Catecholamine Hormones Epinephrine and Norepinephrine

Response of Vibrio cholerae to the Catecholamine Hormones Epinephrine and Norepinephrine

The Na + -Translocating NADH:Quinone Oxidoreductase Enhances Oxidative Stress in the Cytoplasm of Vibrio cholerae

Respiratory Membrane Protein Complexes Convert Chemical Energy

Contact Info

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Quinone Reduction by the Na⁺-Translocating NADH Dehydrogenase Promotes Extracellular Superoxide Production inVibrio cholerae

The Na ⁺ -Translocating NADH:Quinone Oxidoreductase Enhances Oxidative Stress in the Cytoplasm of Vibrio cholerae