Rose F. Kennedy Center, Dominick P. Purpura Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
J Neurosci. 2010 Jan 6;30(1):242-54. doi: 10.1523/JNEUROSCI.4933-08.2010.
Protein kinase C (PKC) enhances NMDA receptor (NMDAR)-mediated currents and promotes NMDAR delivery to the cell surface via SNARE-dependent exocytosis. Although the mechanisms of PKC potentiation are established, the molecular target of PKC is unclear. Here we show that synaptosomal-associated protein of 25 kDa (SNAP-25), a SNARE protein, is functionally relevant to PKC-dependent NMDAR insertion, and identify serine residue-187 as the molecular target of PKC phosphorylation. Constitutively active PKC delivered via the patch pipette potentiated NMDA (but not AMPA) whole-cell currents in hippocampal neurons. Expression of RNAi targeting SNAP-25 or mutant SNAP-25(S187A) and/or acute disruption of the SNARE complex by treatment with BoNT A, BoNT B or SNAP-25 C-terminal blocking peptide abolished NMDAR potentiation. A SNAP-25 peptide and function-blocking antibody suppressed PKC potentiation of NMDA EPSCs at mossy fiber-CA3 synapses. These findings identify SNAP-25 as the target of PKC phosphorylation critical to PKC-dependent incorporation of synaptic NMDARs and document a postsynaptic action of this major SNARE protein relevant to synaptic plasticity.
蛋白激酶 C（PKC）增强 NMDA 受体（NMDAR）介导的电流，并通过 SNARE 依赖性胞吐作用促进 NMDAR 向细胞表面的转运。虽然 PKC 增强的机制已经确立，但 PKC 的分子靶标尚不清楚。在这里，我们表明突触相关蛋白 25kDa（SNAP-25），一种 SNARE 蛋白，与 PKC 依赖性 NMDAR 插入功能相关，并确定丝氨酸残基-187 是 PKC 磷酸化的分子靶标。通过膜片钳传递的组成型激活的 PKC 增强了海马神经元中的 NMDA（但不是 AMPA）全细胞电流。靶向 SNAP-25 的 RNAi 表达或突变 SNAP-25（S187A）和/或用 BoNT A、BoNT B 或 SNAP-25 C 端阻断肽急性破坏 SNARE 复合物，消除了 NMDAR 的增强作用。SNAP-25 肽和功能阻断抗体抑制了在苔藓纤维-CA3 突触处 PKC 对 NMDA EPSC 的增强作用。这些发现确定了 SNAP-25 是 PKC 磷酸化的靶标，对于 PKC 依赖性突触 NMDAR 掺入至关重要，并证明了这种主要 SNARE 蛋白在突触可塑性方面的突触后作用。
