钙信号转导相关新型膜离子通道的结构-功能研究

As an essential secondary messenger in cellular signal transduction pathways, calcium ions play a vital role in many important cellular functions including muscle contraction, cell growth, cell death, and etc. Excessive entry of calcium into a cell may cause cell to undergo apoptosis. Therefore, concentrations of intracellular calcium are tightly regulated under the cooperation of different Ca2+-activated proteins and ion channel proteins. The sarcoplasmic reticulum (SR) or endoplasmic reticulum (ER) is a complex intracellular Ca2+-storage system, while Ryanodine receptor (RyR) and Inositol 1,4,5-triphosphoate receptors (IP3R) acting as channels release the flow of Ca2+ from the SR/ER. The efflux/influx of Ca2+hence leads to the accumulation of a transient negative/positive potential within the SR/ER lumen, which would be expected to inhibit the process of the subsequent release/uptake of Ca2+. Therefore, robust counter-ion currents are vital to balance potential across the SR/ER membrane and to maintain efficient Ca2+ release and re-uptake. Recently identified TRIC channels may function as counter-ion channels for Ca2+ handling, evidenced by the fact that their genetic ablation led to compromised K+-permeability and Ca2+ release across the SR/ER membrane. Recently, some associated papers identified TRIC-A expression levels in linkage with hypertension and muscular diseases, while the mutants of TRIC-B lead to bone and pulmonary diseases. The bioinformatics analysis elucidated that TRIC family began to bifurcate into two subtypes only at the evolutional stage of vertebrates. In this project, we will work on the structural and functional studies of TRIC-A and TRIC-B. The findings will enrich the knowledge of the novel TRIC family, and also provide important information for understanding of TRIC-related diseases.

钙离子作为第二信使，在细胞生命活动中发挥着广泛而重要的作用。胞内的肌浆网/内质网作为钙离子储库，当相关信号来临时，肌浆网/内质网膜上的钙离子通道RyR或IP3R被激活迅速释放钙离子，进而激活钙离子敏感的生物学过程。研究表明，肌浆网/内质网膜上的TRIC新型离子通道在钙离子释放过程中提供反向的阳离子电流，以保证钙离子顺利转运。相关文献报道TRIC-A的表达水平与高血压相关，而TRIC-B的突变与骨发育不全相关。生物信息分析表明TRIC家族在脊椎动物阶段才进一步进化出TRIC-A和TRIC-B两大亚型。调控/协调的蛋白不相同，分布区域也不相同，其相关的突变或缺失造成的疾病差异大，分子作用机理也很可能不相同，因此解析TRIC家族两大亚型结构将丰富和完善TRIC离子通道的结构-功能研究，进一步深化理解复杂的钙离子浓度调控体系，也将为TRIC离子通道在人类健康和疾病上的生理功能提供结构基础和研究方向

TRIC-A的单核苷多态性与高血压相关，它是治疗恶性高血压潜在的药物靶点，TRIC-A 通道的激活剂可能直接产生超极化信号，从而降低血管平滑肌的静息电压；而TRIC-B基因突变会破坏了该离子通道的功能，从而导致骨发育不全病。因此，深入了解TRIC-A和TRIC-B的结构-功能将为相关疾病的发病机制和药物开发提供重要信息和新线索。我们在本课题研究中建立了酵母的真核生物表达体系，获得了脊椎动物来源的TRIC-A和TRIC-B蛋白样品，并解析了一系列高分辨率的三维结构，包括结合钙离子和未结合钙离子的状态。TRIC是对称的三聚体，其中每个单体都有一个跨膜孔道，中间是由高度保守赖氨酸残基形成的H键网络。在静息状态下，Ca2+结合在TRIC三重轴上的的ER腔侧，以稳定了赖氨酸残基形成的H键网络。结合生化实验和电生理实验，我们提出了TRIC调节细胞内Ca2+释放的分子机制。静止状态下，SR/ER腔内高浓度的Ca2+离子抑制了TRIC的活性；当Ca2+离子从RyRs/IP3Rs释放时，Ca2+的耗竭导致Ca2+与RYRs/IP3Rs的解离，并释放出对Ca2+的抑制，同时，SR/ER腔内积累的负电荷引起膜电位变化，随后被TRIC 的TM4跨膜螺旋感应到。这些过程协同调控TRIC离子通道的开放，促进胞内钙离子顺利释放。这些研究结果揭示了胞内钙离子释放的分子机制，进一步深化了我们对钙离子信号转导的理解。