Jardin Isaac, Lopez José J, Salido Gines M, Rosado Juan A
Department of Physiology, Cellular Physiology Research Group, University of Extremadura, 10071 Caceres, Spain.
Department of Physiology, Cellular Physiology Research Group, University of Extremadura, 10071 Caceres, Spain.
J Biol Chem. 2008 Sep 12;283(37):25296-25304. doi: 10.1074/jbc.M802904200. Epub 2008 Jul 20.
Orai1 and hTRPC1 have been presented as essential components of store-operated channels mediating highly Ca(2+) selective I(CRAC) and relatively Ca(2+) selective I(SOC), respectively. STIM1 has been proposed to communicate the Ca(2+) content of the intracellular Ca(2+) stores to the plasma membrane store-operated Ca(2+) channels. Here we present evidence for the dynamic interaction between endogenously expressed Orai1 and both STIM1 and hTRPC1 regulated by depletion of the intracellular Ca(2+) stores, using the pharmacological tools thapsigargin plus ionomycin, or by the physiological agonist thrombin, independently of extracellular Ca(2+). In addition we report that Orai1 mediates the communication between STIM1 and hTRPC1, which is essential for the mode of activation of hTRPC1-forming Ca(2+) permeable channels. Electrotransjection of cells with anti-Orai1 antibody, directed toward the C-terminal region that mediates the interaction with STIM1, and stabilization of an actin cortical barrier with jasplakinolide prevented the interaction between STIM1 and hTRPC1. Under these conditions hTRPC1 was no longer involved in store-operated calcium entry but in diacylglycerol-activated non-capacitative Ca(2+) entry. These findings support the functional role of the STIM1-Orai1-hTRPC1 complex in the activation of store-operated Ca(2+) entry.
Orai1和hTRPC1分别被认为是储存操纵性通道的重要组成部分，它们介导高度Ca(2+)选择性的I(CRAC)和相对Ca(2+)选择性的I(SOC)。有人提出STIM1可将细胞内Ca(2+)储存的Ca(2+)含量传递至质膜储存操纵性Ca(2+)通道。在此，我们提供证据表明，利用药理学工具毒胡萝卜素加离子霉素或生理激动剂凝血酶，在细胞内Ca(2+)储存耗竭的情况下，内源性表达的Orai1与STIM1和hTRPC1之间存在动态相互作用，且这种相互作用独立于细胞外Ca(2+)。此外，我们报告Orai1介导STIM1与hTRPC1之间的通讯，这对于hTRPC1形成Ca(2+)可渗透通道的激活模式至关重要。用针对介导与STIM1相互作用的C末端区域的抗Orai1抗体电转染细胞，并用茉莉酮酸甲酯稳定肌动蛋白皮质屏障，可阻止STIM1与hTRPC1之间的相互作用。在这些条件下，hTRPC1不再参与储存操纵性钙内流，而是参与二酰基甘油激活的非容量性Ca(2+)内流。这些发现支持了STIM1 - Orai1 - hTRPC1复合物在储存操纵性Ca(2+)内流激活中的功能作用。
