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Molecular basis of NMDA receptor-coupled ion channel modulation by S-nitrosylation

Abstract

Several ion channels are thought to be directly modulated by nitric oxide (NO), but the molecular basis of this regulation is unclear. Here we show that the NMDA receptor (NMDAR)-associated ion channel was modulated not only by exogenous NO but also by endogenous NO. Site-directed mutagenesis identified a critical cysteine residue (Cys 399) on the NR2A subunit whose S-nitrosylation (NO+ transfer) under physiological conditions underlies this modulation. In cell systems expressing NMDARs with mutant NR2A subunits in which this single cysteine was replaced by an alanine, the effect of endogenous NO was lost. Thus endogenous S-nitrosylation can regulate ion channel activity.

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Figure 1: NO species generated by S-nitrosocysteine (SNOC) inhibit NMDA-evoked currents in NR1/NR2A receptors, and the cysteine modifying agent (2-aminoethyl) methanethiosulfonate (MTSEA) occludes NO inhibition.
Figure 2: Voltage independence of NO-induced inhibition of NMDA-evoked currents.
Figure 3: Effect of NR2A(C399A) on NMDA-evoked responses.
Figure 4: Physiological effects of endogenous NO on NMDAR responses of HEK293 cells transfected with NMDAR subunits and nNOS.
Figure 5: RP-HPLC analysis with UV detection of S-nitrosylation products of wild-type (or mutant) decapeptide containing the N-terminal cysteine (or alanine substitution) at residue 399 of the NR2A subunit.

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Acknowledgements

We thank J. M. Sullivan and E. Liman for sharing cDNA clones or vectors and T. Lishnak for technical assistance. This work was supported in part by National Institutes of Health grants P01 HD29587, R01 EY05477 and R01 EY09024.

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Correspondence to Stuart A. Lipton.

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Choi, YB., Tenneti, L., Le, D. et al. Molecular basis of NMDA receptor-coupled ion channel modulation by S-nitrosylation. Nat Neurosci 3, 15–21 (2000). https://doi.org/10.1038/71090

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