pThe mechanisms of NO inhibition of CaMK [Casup2+/sup/CaM (calmodulin)-dependent protein kinase] II activity were studied. In rat pituitary tumour GH3 cells, TRH [thyrotrophin (TSH)-releasing hormone]-stimulated phosphorylation of nNOS [neuronal NOS (NO synthase)] at Sersup847/sup was sensitive to an inhibitor of CaMKs, KN-93, and was enhanced by inhibition of nNOS with 7NI (7-nitroindazole). Enzyme activity of CaMKII following iin situ/i treatment with 7NI was also increased. The iin vitro/i activity of CaMKII was inhibited by co-incubation either with nNOS and L-arginine or with NO donors SNAP (iS/i-nitroso-iN/i-acetyl-DL-penicillamine) and DEA-NONOate [diethylamine-NONOate (diazeniumdiolate)]. Once inhibited by these treatments, CaMKII was observed to undergo full reactivation on the addition of a reducing reagent, DTT (dithiothreitol). In transfected cells expressing CaMKII and nNOS, treatment with the calcium ionophore A23187 further revealed nNOS phosphorylation at Sersup847/sup, which was enhanced by 7NI and CaMKII iS/i-nitrosylation. Mutated CaMKII (C6A), in which Cyssup6/sup was substituted with an alanine residue, was refractory to 7NI-induced enhancement of nNOS phosphorylation or to CaMKII iS/i-nitrosylation. Furthermore, we could identify Cyssup6/sup as a direct target for iS/i-nitrosylation of CaMKII using MS. In addition, treatment with glutamate caused an increase in CaMKII iS/i-nitrosylation in rat hippocampal slices. This glutamate-induced iS/i-nitrosylation was blocked by 7NI. These results suggest that inactivation of CaMKII mediated by iS/i-nitrosylation at Cyssup6/sup may contribute to NO-induced neurotoxicity in the brain./p
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机译：>研究了NO抑制CaMK [Ca 2 + / CaM（钙调蛋白）依赖性蛋白激酶] II活性的机制。在大鼠垂体肿瘤GH3细胞中，TRH [促甲状腺激素（TSH）释放激素]刺激Ser 847 处nNOS [神经型NOS（NO合酶）]的磷酸化对CaMKs抑制剂KN-敏感93，并被7NI（7-硝基吲唑）抑制nNOS增强。用7NI原位处理后，CaMKII的酶活性也增加了。通过与nNOS和L-精氨酸或NO供体SNAP（ S -亚硝基- N 处的nNOS磷酸化，这被7NI和CaMKII S -亚硝基化所增强。 Cys 6 被丙氨酸残基取代的突变的CaMKII（C6A）对7NI诱导的nNOS磷酸化增强或CaMKII S 亚硝基化均具有抵抗力。此外，我们可以确定使用Cys 6 作为CaMKII的 S -亚硝基化的直接目标。另外，用谷氨酸盐处理导致大鼠海马切片中CaMKII S -亚硝基化的增加。谷氨酸诱导的 S -亚硝基化被7NI阻断。这些结果表明，在Cys 6 处由 S -亚硝基化介导的CaMKII失活可能有助于NO诱导的脑神经毒性。
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