API

Relationship: 1698

Title

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Peptide Oxidation leads to S-Glutathionylation, eNOS

Upstream event

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Peptide Oxidation

Downstream event

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S-Glutathionylation, eNOS

Key Event Relationship Overview

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AOPs Referencing Relationship

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AOP Name Adjacency Weight of Evidence Quantitative Understanding
Peptide Oxidation Leading to Hypertension adjacent Moderate Low

Taxonomic Applicability

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Term Scientific Term Evidence Link
rat Rattus norvegicus Moderate NCBI
human Homo sapiens High NCBI
mouse Mus musculus High NCBI
cow Bos taurus High NCBI

Sex Applicability

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Sex Evidence
Unspecific High

Life Stage Applicability

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Term Evidence
All life stages High

Key Event Relationship Description

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Oxidation of GSH results in the formation of a disulphide-bridged glutathione dimer (GSSG).  GSSG is either rapidly re-reduced back to GSH by nicotinamide adenine dinucleotide phosphate (NADPH)-dependent GSSG-reductases or extruded from the cell by adenosine triphosphate (ATP)-dependent translocases. However, when these mechanisms become overwhelmed by high local oxidant concentrations, GSSG can interact with protein thiol groups to form protein-GSSG adducts, a process termed S-glutathionylation. Interestingly, glutathione disulfide-protein formation has been suggested to occur with a certain degree of specificity to cellular proteins, since protein thiol groups exhibit a considerable heterogeneity in terms of their individual pKa values and their location in protein structures (Schuppe et al. 1992).  The oxidation of GSH to GSSG elevates levels of GSSG, which then covalently bind to critical serine residues on endothelial nitric oxide synthase (eNOS; Chen et al 2010, Du et al. 2013, De Pascali et al. 2014).

Evidence Supporting this KER

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Biological Plausibility

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When antioxidant defence mechanisms become overwhelmed by high local oxidant concentrations, GSSG can interact with protein thiol groups to form protein-GSSG adducts, a process termed S-glutathionylation (Schuppe et al. 1992).

Hypoxia/reoxygenation-induced oxidative stress (associated with ischaemia-reperfusion injury) was shown to deplete GSH in bovine aortic endothelial cells, which led to S-glutathionylation of eNOS and eNOS uncoupling.  This phenomenon was partially reversible, in bovine aortic endothelial cells and rat aortic rings, by raising intracellular GSH levels upon administration of N-acetylcysteine (Chen et al. 2010, DePascali et al. 2014).

Empirical Evidence

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Chen and colleagues (2010) have shown that eNOS is particularly sensitive to S-glutathionylation at cysteine residues 689 and 908 of the reductase domain, a phenomenon that is dose-dependent with application of exogenous GSSG.  This finding was corroborated by Peng et al. (2015) using mutated eNOS constructs in E. coli, demonstrating that superoxide was produced by the eNOS phosphorylation site in the reductase domain.

Wu et al. (2014) studied responses in human lung microvascular endothelial cells to lipopolysaccharide (LPS) in vitro.  Upon LPS administration, NADPH oxidase 2 (NOX2) expression levels were increased with a subsequent rise in superoxide production, which led to S-glutathionylation of eNOS.  Furthermore, in mice, co-immunoprecipitation studies revealed that NOX2 associated with eNOS, and that S-glutathionylation in response to LPS was much more apparent in elderly animals compared to younger animals.  Similar observations were made by De Pascali et al. (2014) following hypoxia-induced oxidative stress in bovine aortic endothelial cells.

Uncertainties and Inconsistencies

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Quantitative data for humans is very limited.

Quantitative Understanding of the Linkage

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Chen et al. (2013) demonstrated that co-administration of glutaredoxin-1 and GSH reversed GSSG-mediated eNOS S-glutathionylation and restored eNOS-mediated NO production, also in bovine aortic cells.  Interestingly, inhibition of eNOS function occurred when the GSH/GSSG ratio was >0.2 and function was restored at a ratio of <0.1.

Response-response Relationship

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Time-scale

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Known modulating factors

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Known Feedforward/Feedback loops influencing this KER

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Domain of Applicability

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The above evidence demonstrates similar responses to stressors in cows, mice and humans.  A functional response using aortic rings was demonstrated in rats.

References

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Chen CA, Wang TY, Varadharaj, S et al. S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature (2010) 468: 1115–1118.

Chen CA, De Pascali F, Basye A et al. Redox modulation of endothelial nitric oxide synthase by glutaredoxin-1 through reversible oxidative post-translational modification.  (2013)  Biochemistry. 52(38):6712-23

Dhar A, Dhar I, Desai KM et al.  Methylglyoxal scavengers attenuate endothelial dysfunction induced by methylglyoxal and high concentrations of glucose. (2010) Br. J. Pharmacol. 161: 1843–1856.

De Pascali F, Hemann C, Samons, K et al. Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (2014). 53 : 3679–3688.

Du Y, Navab M, Shen M, et al.  Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. (2013) 436 : 462–466.

Peng H, Zhuang Y, Chen Y et al.  The Characteristics and Regulatory Mechanisms of Superoxide Generation from eNOS Reductase Domain.  (2015) PLoS One. 10(10):e0140365

Schuppe I, Moldéus P, and Cotgreave IA. Protein-specific S-thiolation in human endothelial cells during oxidative stress. (1992) Biochem. Pharmacol. 44: 1757–1764.

Wu F, Szczepaniak WS, Shiva S et al.  Nox2-dependent glutathionylation of endothelial NOS leads to uncoupled superoxide production and endothelial barrier dysfunction in acute lung injury.  (2014) Am J Physiol Lung Cell Mol Physiol. 307(12):L987-97.