CRAC钙通道的功能及调控机制探究

内质网-质膜间的膜接触位点不仅是两者之间信息和物质交流的场所，还负责胞内信号的向外传递以及胞外信号的向内传递，有着独特的沟通细胞内外环境的作用。内质网钙池释放激活（CRAC）钙通道的开关元件STIM位于内质网膜，通道元件Orai1位于质膜，两者经物理耦联介导内质网-质膜互作而产生钙池操纵钙内流（SOCE）。SOCE是众多细胞内的一种钙稳态维持及钙信号产生机制，其异常与心血管疾病和奥兹海默症等神经退行性疾病密切相关，但其在神经细胞中起作用的分子机制还需进一步的解析。为此，申请人在前期工作的基础上，拟采用基因敲入、邻近生物素标记、超高分辨率成像等手段，系统描绘神经元中内质网-质膜之间的膜接触位点的动态变化以及相应的蛋白质组图谱；从中鉴定CRAC钙通道的调控因子并阐释其机制；运用光遗传学和光化学工具结合RNA-seq及ChIP-seq等手段，探究其在神经元中的生理功能及在疾病治疗中的潜在应用。

Membrane contact sites (MCS) between endoplasmic reticulum (ER) and plasm membrane (PM) are places not only crucial for ER-PM communications, but also essential for inside-out and outside-in signal transduction. Calcium release activated calcium (CRAC) channels are localized within ER-PM MCS, mediate interactions between ER and PM, resulting calcium influxes called store operated calcium entry (SOCE). SOCE is a ubiquitous process crucial for calcium signalling and calcium homeostasis, closely linked with various cardiovascular diseases, Alzheimer’s diseases and other neurodegenerative diseases. However, molecular mechanisms underlying its function and regulation in neurons still remain elusive. We plan to combine the usage of novel techniques like gene knock-in with CRIS-PITCh system, proximity protein labelling, super-resolution imaging and etc., to map the dynamic behaviors of ER-PM MCSs of neurons under various physiological conditions, identify proteomics that interact with CRAC channels within ER-PM MCSs. We then will dissect out the mechanisms by which those interactors regulate CRAC channel activity. We will also develop new optogenetic tools and light-switchable compounds to examine the physiological roles and therapeutic potentials of CRAC channels.