N-methyl-D-aspartate receptor-dependent denitrosylation of neuronal nitric oxide synthase increase the enzyme activity.
Our laboratory once reported that neuronal nitric oxide synthase (nNOS) S-nitrosylation was decreased in rat hippocampus during cerebral ischemia-reperfusion, but the underlying mechanism was unclear. In this study, we show that nNOS activity is dynamically regulated by S-nitrosylation. We found that overexpressed nNOS in HEK293 (human embryonic kidney) cells could be S-nitrosylated by exogenous NO donor GSNO and which is associated with the enzyme activity decrease. Cys(331), one of the zinc-tetrathiolate cysteines, was identified as the key site of nNOS S-nitrosylation. In addition, we also found that nNOS is highly S-nitrosylated in resting rat hippocampal neurons and the enzyme undergos denitrosylation during the process of rat brain ischemia/reperfusion. Intrestingly, the process of nNOS denitrosylation is coupling with the decrease of nNOS phosphorylation at Ser(847), a site associated with nNOS activation. Further more, we document that nNOS denitrosylation could be suppressed by pretreatment of neurons with MK801, an antagonist of NMDAR, GSNO, EGTA, BAPTA, W-7, an inhibitor of calmodulin as well as TrxR1 antisense oligonucleotide (AS-ODN) respectively. Taken together, our data demonstrate that the denitrosylation of nNOS induced by calcium ion influx is a NMDAR-dependent process during the early stage of ischemia/reperfusion, which is majorly mediated by thioredoxin-1 (Trx1) system. nNOS dephosphorylation may be induced by the enzyme denitrosylation, which suggest that S-nitrosylation/denitrosylation of nNOS may be an important mechanism in regulating the enzyme activity.
Duke Scholars
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Related Subject Headings
- Thioredoxins
- Reperfusion Injury
- Receptors, N-Methyl-D-Aspartate
- Rats
- Phosphorylation
- Nitric Oxide Synthase Type I
- Nitric Oxide
- Neurons
- Male
- Humans
Citation
Published In
DOI
EISSN
ISSN
Publication Date
Volume
Issue
Start / End Page
Related Subject Headings
- Thioredoxins
- Reperfusion Injury
- Receptors, N-Methyl-D-Aspartate
- Rats
- Phosphorylation
- Nitric Oxide Synthase Type I
- Nitric Oxide
- Neurons
- Male
- Humans