Hypoxia Enhances S-Nitrosylation-Mediated NMDA Receptor Inhibition via a Thiol Oxygen Sensor Motif

Abstract

Under ambient air conditions, NO inhibits NMDAR activity by reacting with the NR2A subunit C399 along with two additional cysteine pairs if their disulfide bonds are reduced to free thiol groups [NR1(C744,C798); NR2(C87,C320)]. Here we demonstrate that relative hypoxia enhances S-nitrosylation of NMDARs by a unique mechanism involving an "NO-reactive oxygen sensor motif" whose determinants include C744 and C798 of the NR1 subunit. Redox reactions involving these two thiol groups sensitize other NMDAR sites to S-nitrosylation and consequent receptor inhibition, while their own nitrosylation has little effect on NMDAR activity. The crystal structure of the ligand-binding domain of NR1 reveals a flexible disulfide bond (C744-C798), which may account for its susceptibility to reduction and subsequent reaction with NO that is observed with biochemical techniques. These thiols may be nitrosylated preferentially during increasing hypoxia or stroke conditions, thus preventing excessive activity associated with cytotoxicity while avoiding blockade of physiologically active NMDARs.

Ischemia-elicited Oxidative Modulation of Ca2+/Calmodulin-dependent Protein Kinase II

“…This later modification, which reduces the capacity of CaMKII to undergo autophosphorylation, is a likely cause of the depressed HFS-induced LTP following ischemia/re-oxygenation. It is also possible that impairment of NMDA receptors resulting from S-nitrosylation and disulfide formation (47,48) may also contribute to the attenuated response in LTP.…”

Redox-based regulation of signal transduction: Principles, pitfalls, and promises

“…Indeed, a number of targets such as caspases, receptors (e.g. alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic acid (AMPA) and NMDA), kinases, G proteins and ion channels have been demonstrated to be S-nitrosylated under physiological conditions [15,53,54]. In diverse cell types, S-nitrosylation can be detected in situ under both non-stimulated conditions and following physiological activation (e.g stimulation of GPCRs to dilate blood vessels or stimulate cardiac contractility [55,56]).…”

“…Furthermore, as will be described below, regulatory enzymes important in redox control, such as thioredoxin, glutaredoxin, peroxiredoxin, and protein disulfide isomerase, are themselves affected and regulated by both H 2 O 2 and NO-based cysteine oxidations. It is of interest to note that S-nitrosylation may be criticaly sensitive to oxygen tension, often being potentiated by hypoxia [15,54,57]. Further, it has been shown that oxidation/reduction (of a group) of thiols may regulate S-nitrosylation by an allosteric mechanism [57].…”

S-Nitrosylation and uncompetitive/fast off-rate (UFO) drug therapy in neurodegenerative disorders of protein misfolding

“…Recently, we have found that relatively hypoxic conditions render the NMDA receptor exquisitely sensitive to S-nitrosylation of Cys744 and Cys798 on the NR1 subunit. 65 NO modification of these two thiols in NR1 further enhances S-nitrosylation of the Cys399 site on the NR2A, resulting in the inhibition of receptor activity by very low levels of NO. This finding highlights the physiological relevance of S-nitrosylation of the NMDA receptor because brain oxygen levels are normally low under physiological conditions (pO 2 of 10-20 mmHg) and even lower during hypoxia/ischemia (stroke).…”

“…6,8,44 Moreover, as mentioned earlier, as the oxygen tension is lowered (a pO 2 of 10-20 torr is found in normal brain, and even lower levels under hypoxic/ischemic conditions), the NMDA receptor becomes more sensitive to inhibition by S-nitrosylation. 65 Unfortunately, nitroglycerin itself is not very attractive as a neuroprotective agent. The same cardiovascular vasodilator effect that makes it useful in the treatment of angina could cause dangerously large drops in blood pressure in patients with dementia, stroke, traumatic injury, or glaucoma.…”