N type Ca2+ channels and rim scaffold protein covary at the presynaptic transmitter release face but are components of independent protein complexes
N type Ca2+ channels and rim scaffold protein covary at the presynaptic transmitter release face but are components of independent protein complexes
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DOI：
10.1016/j.neuroscience.2006.04.053
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发表时间：
2006-01-01
期刊：
影响因子：
3.3
通讯作者：
Stanley, E. F.
中科院分区：
文献类型：
--
作者：
Khanna, R.;Li, Q.;Stanley, E. F.
突触前末梢的快速神经递质释放发生在专门的递质释放位点，其中停靠的分泌囊泡被通过局部N型（CaV2.2）钙通道进入的Ca 2+离子的流入触发与膜融合。因此，神经分泌涉及两个关键过程：囊泡在递质释放位点的对接，这一过程涉及支架蛋白RIM（Rab 3A相互作用分子）及其结合伴侣Munc-13，以及随后通过激活囊泡融合的门控Ca 2+通道。然而，目前尚不清楚囊泡融合复合物及其附着的Ca 2+通道和囊泡对接复合物是否是单个多功能实体的一部分。Ca 2+通道本身和RIM被用作这两个元件的标记来解决这个问题。我们进行了免疫染色在巨大的calyxtype突触的小鸡睫状神经节本地化的蛋白质在一个本地的，不受干扰的突触前神经末梢。采用定量免疫染色（强度相关分析/强度相关商法）检测这两种蛋白在神经末梢递质释放面的关系。CaV2.2和RIM的染色强度变化很大，这与它们都是递质释放位点的组分的预期一致。然后，我们使用免疫沉淀来测试这些蛋白质是否也是共同分子复合物的一部分。然而，CaV2.2的沉淀未能捕获RIM或RIM结合配偶体Munc-13。这些研究结果表明，虽然囊泡融合和囊泡对接机制共存的递质释放面，他们不是一个共同的稳定复合物的一部分。(c)2006年IBRO。由爱思唯尔有限公司出版。保留所有权利。
Fast neurotransmitter release at presynaptic terminals occurs at specialized transmitter release sites where docked secretory vesicles are triggered to fuse with the membrane by the influx of Ca2+ ions that enter through local N type (CaV2.2) calcium channels. Thus, neurosecretion involves two key processes: the docking of vesicles at the transmitter release site, a process that involves the scaffold protein RIM (Rab3A interacting molecule) and its binding partner Munc-13, and the subsequent gating of vesicle fusion by activation of the Ca2+ channels. It is not known, however, whether the vesicle fusion complex with its attached Ca2+ channels and the vesicle docking complex are parts of a single multifunctional entity. The Ca2+ channel itself and RIM were used as markers for these two elements to address this question. We carried out immunostaining at the giant calyxtype synapse of the chick ciliary ganglion to localize the proteins at a native, undisturbed presynaptic nerve terminal. Quantitative immunostaining (intensity correlation analysis/intensity correlation quotient method) was used to test the relationship between these two proteins at the nerve terminal transmitter release face. The staining intensities for CaV2.2 and RIM covary strongly, consistent with the expectation that they are both components of the transmitter release sites. We then used immunoprecipitation to test if these proteins are also parts of a common molecular complex. However, precipitation of CaV2.2 failed to capture either RIM or Munc-13, a RIM binding partner. These findings indicate that although the vesicle fusion and the vesicle docking mechanisms coexist at the transmitter release face they are not parts of a common stable complex. (c) 2006 IBRO. Published by Elsevier Ltd. All rights reserved.