S-glutathionylation uncouples eNOS and regulates its cellular and vascular function

Article

关键词: NITRIC-OXIDE SYNTHASE;    ENDOTHELIUM-DEPENDENT RELAXATION;    SUPEROXIDE GENERATION;    ENDOGENOUS METHYLARGININES;    TETRAHYDROBIOPTERIN;    REDUCTASE;    OXIDATION;    CELLS;    PHOSPHORYLATION;    GLUTATHIOLATION;   

DOI  :  10.1038/nature09599

来源: SCIE

【 摘 要 】

Endothelial nitric oxide synthase (eNOS) is critical in the regulation of vascular function, and can generate both nitric oxide (NO) and superoxide (O-2(center dot-)), which are key mediators of cellular signalling. In the presence of Ca2+/calmodulin, eNOS produces NO, endothelial-derived relaxing factor, from L-arginine (L-Arg) by means of electron transfer from NADPH through a flavin containing reductase domain to oxygen bound at the haem of an oxygenase domain, which also contains binding sites for tetrahydrobiopterin (BH4) and L-Arg(1-3). In the absence of BH4, NO synthesis is abrogated and instead O-2(center dot-) is generated(4-7). While NOS dysfunction occurs in diseases with redox stress, BH4 repletion only partly restores NOS activity and NOS-dependent vasodilation(7). This suggests that there is an as yet unidentified redox-regulated mechanism controlling NOS function. Protein thiols can undergo S-glutathionylation, a reversible protein modification involved in cellular signalling and adaptation(8,9). Under oxidative stress, S-glutathionylation occurs through thiol-disulphide exchange with oxidized glutathione or reaction of oxidant-induced protein thiyl radicals with reduced glutathione(10,11). Cysteine residues are critical for the maintenance of eNOS function(12,13); we therefore speculated that oxidative stress could alter eNOS activity through S-glutathionylation. Here we show that S-glutathionylation of eNOS reversibly decreases NOS activity with an increase in O-2(center dot-) generation primarily from the reductase, in which two highly conserved cysteine residues are identified as sites of S-glutathionylation and found to be critical for redox-regulation of eNOS function. We show that eNOS S-glutathionylation in endothelial cells, with loss of NO and gain of O-2(center dot-) generation, is associated with impaired endothelium-dependent vasodilation. In hypertensive vessels, eNOS S-glutathionylation is increased with impaired endothelium-dependent vasodilation that is restored by thiol-specific reducing agents, which reverse this S-glutathionylation. Thus, S-glutathionylation of eNOS is a pivotal switch providing redox regulation of cellular signalling, endothelial function and vascular tone.

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