Synaptic multiprotein complexes associated with 5-HT2C receptors:: a proteomic approach
Synaptic multiprotein complexes associated with 5-HT2C receptors:: a proteomic approach
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DOI：
10.1093/emboj/21.10.2332
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发表时间：
2002-05-15
期刊：
影响因子：
11.4
通讯作者：
Marin, P
中科院分区：
文献类型：
--
作者：
Bécamel, C;Alonso, G;Marin, P
膜结合受体，如酪氨酸激酶和嗜离子受体，与由涉及多结构域蛋白的蛋白-蛋白相互作用构成的大型蛋白质网络有关。尽管这些网络已作为细胞信号传导的一般机制出现，但对与g蛋白偶联受体（gpcr）相关的蛋白质复合物知之甚少。利用基于肽亲和层析、质谱和免疫印迹的蛋白质组学方法，我们鉴定了15个与5-羟色胺2C （5-HT2C）受体c端尾部相互作用的蛋白质。这些蛋白包括几种含有一个或多个PDZ结构域的突触多结构域蛋白（PSD95和三方复合物Veli3-CASK-Mint1蛋白），肌动蛋白/谱蛋白细胞骨架蛋白和信号蛋白。共免疫沉淀实验表明，体内5-HT2C受体与PSD95和Veli3-CASK-Mint1复合物相互作用。电镜观察还发现Veli3和5-HT2C受体突触富集，并在脉络膜细胞微绒毛中共定位。这些结果表明，5-HT2C受体与蛋白质网络相关，这对其突触定位和信号机制的耦合很重要。
Membrane-bound receptors such as tyrosine kinases and ionotropic receptors are associated with large protein networks structured by protein-protein interactions involving multidomain proteins. Although these networks have emerged as a general mechanism of cellular signalling, much less is known about the protein complexes associated with G-protein-coupled receptors (GPCRs). Using a proteomic approach based on peptide affinity chromatography followed by mass spectrometry and immunoblotting, we have identified 15 proteins that interact with the C-terminal tail of the 5-hydroxytryptamine 2C (5-HT2C) receptor, a GPCR. These proteins include several synaptic multidomain proteins containing one or several PDZ domains (PSD95 and the proteins of the tripartite complex Veli3-CASK-Mint1), proteins of the actin/spectrin cytoskeleton and signalling proteins. Coimmunoprecipitation experiments showed that 5-HT2C receptors interact with PSD95 and the Veli3-CASK-Mint1 complex in vivo. Electron microscopy also indicated a synaptic enrichment of Veli3 and 5-HT2C receptors and their colocalization in microvilli of choroidal cells. These results indicate that the 5-HT2C receptor is associated with protein networks that are important for its synaptic localization and its coupling to the signalling machinery.