Molecular mechanisms underlying inhibition of STIM1-Orai1-mediated Ca(2+) entry induced by 2-aminoethoxydiphenyl borate.
Molecular mechanisms underlying inhibition of STIM1-Orai1-mediated Ca(2+) entry induced by 2-aminoethoxydiphenyl borate.
复制标题
2-氨基乙氧基二苯硼酸盐诱导的 STIM1-Orai1 介导的 Ca(2) 内流抑制的分子机制。
DOI：
10.1007/s00424-016-1880-z
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发表时间：
2016-11
期刊：
影响因子：
--
通讯作者：
Wang Y
中科院分区：
文献类型：
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作者：
Wei M;Zhou Y;Sun A;Ma G;He L;Zhou L;Zhang S;Liu J;Zhang SL;Gill DL;Wang Y
由STIM 1和Orai 1介导的钙库操纵的钙内流（SOCE）对于大多数细胞类型中的钙信号传导和稳态至关重要。2-氨基乙氧基二苯基硼酸酯（2-APB）是一种已被充分描述的SOCE抑制剂，但其作用在很大程度上仍然是难以捉摸的。在这里，我们表明，2-APB不影响二聚体状态的STIM 1，但增强了卷曲螺旋1（CC 1）和STIM-Orai激活区（SOAR）的STIM 1的分子内耦合，随后减少在形成的STIM 1斑点在Orai 1过表达的情况下。2-APB还抑制Orai 1通道，直接抑制Ca 2+通过组成型活性，STIM 1-独立的Orai 1突变体，Orai 1-P245 T和Orai 1-V102 A进入。当与STIM 1结合时，组成型活性Orai 1-V102 C突变体不受2-APB抑制。因此，我们使用Orai 1-V012 C作为工具来检查2-APB是否也可以抑制STIM 1和Orai 1之间的偶联。我们发现STIM 1和Orai 1-V102 C之间的功能偶联被2-APB抑制。这种偶联抑制作用是间接的，由2-APB对STIM 1的作用引起，并且很可能是通过Orai 1 N末端的门控残基介导的。总的来说，我们发现这种双直接位点抑制可能有助于更好地理解STIM 1激活Orai 1和未来的药物设计。
Store operated Ca2+ entry (SOCE) mediated by STIM1 and Orai1 is crucial for Ca2+ signaling and homeostasis in most cell types. 2-aminoethoxydiphenyl borate (2-APB) is a well described SOCE inhibitor but its action remains largely elusive. Here we show that 2-APB does not affect the dimeric state of STIM1, but enhances the intramolecular coupling between the coiled-coil 1 (CC1) and STIM-Orai Activating Region (SOAR) of STIM1, with subsequent reduction in the formation of STIM1 puncta in the absence of Orai1overexpression. 2-APB also inhibits Orai1 channels, directly inhibiting Ca2+ entry through the constitutively active, STIM1-independent Orai1 mutants, Orai1-P245T and Orai1-V102A. When unbound with STIM1, the constitutively active Orai1-V102C mutant is not inhibited by 2-APB. Thus we used Orai1-V012C as a tool to examine whether 2-APB can also inhibit the coupling between STIM1 and Orai1. We find that the functional coupling between STIM1 and Orai1-V102C is inhibited by 2-APB. This inhibition on coupling is indirect, arising from 2-APB’s action on STIM1, and it is most likely mediated by gating residues in Orai1 N terminus. Overall, our finding of this two-direct-site inhibition may help better understanding of Orai1-activation by STIM1 and future drug design.