Jardin Isaac, Gómez Luis J, Salido Gines M, Rosado Juan A
Department of Physiology, University of Extremadura, Caceres, Spain.
Biochem J. 2009 May 13;420(2):267-76. doi: 10.1042/BJ20082179.
TRPC (canonical transient receptor potential) channel subunits have been shown to assemble into homo- or hetero-meric channel complexes, including different Ca2+-handling proteins, required for the activation of CCE (capacitative Ca2+ entry) or NCCE (non-CCE) pathways. In the present study we found evidence for the dynamic interaction between endogenously expressed hTRPC6 (human TRPC6) with either both Orai1 and STIM1 (stromal interaction molecule 1) or hTRPC3 to participate in CCE or NCCE. Electrotransjection of cells with an anti-hTRPC6 antibody, directed towards the C-terminal region, reduces CCE induced by TPEN [N,N,N',N'-tetrakis-(2-pyridylmethyl)-ethylenediamine], which reduces the intraluminal free Ca2+ concentration. Cell stimulation with thrombin or extensive Ca2+-store depletion by TG (thapsigargin)+ionomycin enhanced the interaction between hTRPC6 and the CCE proteins Orai1 and STIM1. In contrast, stimulation with the diacylglycerol analogue OAG (1-oleoyl-2-acetyl-sn-glycerol) displaces hTRPC6 from Orai1 and STIM1 and enhances the association between hTRPC6 and hTRPC3. The interaction between hTRPC6 and hTRPC3 was abolished by dimethyl-BAPTA [1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid] loading, which indicates that this phenomenon is Ca2+-dependent. These findings support the hypothesis that hTRPC6 participates both in CCE and NCCE through its interaction with the Orai1-STIM1 complex or hTRPC3 respectively.
已证明瞬时受体电位经典型（TRPC）通道亚基可组装成同聚或异聚通道复合物，其中包括激活容量性Ca²⁺内流（CCE）或非CCE（NCCE）途径所需的不同Ca²⁺处理蛋白。在本研究中，我们发现内源性表达的hTRPC6（人类TRPC6）与Orai1和基质相互作用分子1（STIM1）或hTRPC3之间存在动态相互作用的证据，它们参与CCE或NCCE。用针对C末端区域的抗hTRPC6抗体对细胞进行电转染，可降低由N,N,N',N'-四（2-吡啶甲基）乙二胺（TPEN）诱导的CCE，TPEN可降低管腔内游离Ca²⁺浓度。用凝血酶刺激细胞或用毒胡萝卜素（TG）+离子霉素使Ca²⁺储存大量耗竭，可增强hTRPC6与CCE蛋白Orai1和STIM1之间的相互作用。相反，用二酰基甘油类似物1-油酰-2-乙酰基-sn-甘油（OAG）刺激可使hTRPC6与Orai1和STIM1分离，并增强hTRPC6与hTRPC3之间的结合。用二甲基-1,2-双（邻氨基苯氧基）乙烷-N,N,N',N'-四乙酸（dimethyl-BAPTA）负载可消除hTRPC6与hTRPC3之间的相互作用，这表明该现象依赖于Ca²⁺。这些发现支持以下假设：hTRPC6分别通过与Orai1-STIM1复合物或hTRPC3相互作用参与CCE和NCCE。
