School of Basic Medical Sciences, Hubei University of Medicine, Shiyan, 442000, China.
Department of Genetics and Cell Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China.
Biochemistry (Mosc). 2023 Sep;88(9):1296-1303. doi: 10.1134/S0006297923090092.
Store-operated calcium entry (SOCE) is the main mechanism for the Ca2+ influx in non-excitable cells. The two major components of SOCE are stromal interaction molecule 1 (STIM1) in the endoplasmic reticulum and Ca2+ release-activated Ca2+ channel (CRAC) Orai on the plasma membrane. SOCE requires interaction between STIM1 and Orai. Mammals have three Orai homologs: Orai1, Orai2, and Orai3. Although Orai1 has been widely studied and proven to essential for numerous cellular processes, Orai3 has also attracted a significant attention recently. The gating and activation mechanisms of Orai3 have yet to be fully elucidated. Here, we expressed, purified, and reconstituted Orai3 protein into liposomes and investigated its orientation and oligomeric state in the resulting proteoliposomes. STIM1 interacted with the Orai3-containing proteoliposomes and mediated calcium release from the them, suggesting that the Orai3 channel was functional and that recombinant STIM1 could directly open the Orai3 channel in vitro. The developed in vitro calcium release system could be used to study the structure, function, and pharmacology of Orai3 channel.
钙库操纵性钙内流（SOCE）是无兴奋细胞钙离子内流的主要机制。SOCE 的两个主要组成部分是内质网中的基质相互作用分子 1（STIM1）和质膜上的钙释放激活钙通道（CRAC）Orai。SOCE 需要 STIM1 和 Orai 之间的相互作用。哺乳动物有三种 Orai 同源物：Orai1、Orai2 和 Orai3。尽管 Orai1 已被广泛研究并被证明对许多细胞过程至关重要，但 Orai3 最近也引起了广泛关注。Orai3 的门控和激活机制尚未完全阐明。在这里，我们表达、纯化并将 Orai3 蛋白重构成脂质体，并研究了其在所得蛋白脂质体中的取向和寡聚状态。STIM1 与含有 Orai3 的蛋白脂质体相互作用并介导钙从其中释放，表明 Orai3 通道是功能性的，重组 STIM1 可以在体外直接打开 Orai3 通道。开发的体外钙释放系统可用于研究 Orai3 通道的结构、功能和药理学。
