CLAC钙通道调控内质网-线粒体互作和线粒体自噬的新机制

Endoplasmic reticulum (ER) and mitochondria are two types of important organelles in cell, both of them are calcium (Ca2+) stores. The ER-mitochondrial adjoining contact sites are the locus where the two important organelles are structurally and functionally interacted. The ER-mitochondrial adjoining contacts play critical roles in a variety of important physiological functions such as Ca2+ signaling, homeostatic maintenance of Ca2+ stores, ATP production, mitochondrial biogenesis and mobility, ROS stress, ER stress, cell death etc. The molecular mechanisms for dynamic ER-mitochondrial adjoining interaction remain elusive. Last year we have found and named a Ca2+-Load-Activated-Ca2+ (CLAC) channel which is a main player in the maintenance of ER Ca2+ homeostasis. Our recent preliminary data further showed a CLAC-associated-mitochondrial-membrane protein 1 (CAMMP1) that strongly and dynamically bound to CLAC channel, suggesting that the dynamic interaction between CLAC and CAMMP1 constitutes a novel mechanism for the establishment of ER-mitochondrial adjoining interaction. We are planning to use multiple approaches involving molecular and cellular, biochemistry, cellular organelle Ca2+ imaging, super resolution microscopy, electron microscopy, disease mouse model, histopathology, and quantitative proteomic analysis, to elucidate the molecular mechanisms of how the CLAC-CAMMP1 builds up, maintains and dynamically regulates the functional contact/interaction between ER and mitochondrion; to determine the importance of CLAC-CAMMP1 in mitochondrial function and mitophagy; and to unravel the pathogenic mechanisms of congenital lactic acidosis and cardiac disorders caused by human CAMMP1 mutations and deranged ER-mitochondrial contact/interaction. The expected results could reveal a novel mechanism for dynamic ER-mitochondrial adjoining interaction mediated by CLAC-CAMMP1 in health and disease, and broaden our views on hereditary causing factors of congenital lactic acidosis and cardiac disorders.

内质网和线粒体是细胞内的两个重要细胞器，二者也是细胞内的钙库。内质网-线粒体之间的偶联接触(ER-mitochondria adjoining contact)是二者在结构和功能上发生互作的基础，其调控了多个重要的细胞生命活动过程。内质网-线粒体动态互作的分子机制目前还很不明确。我们前期发现并命名了参与内质网钙稳态调控的保守离子通道CLAC，最近进一步发现了与CLAC动态结合的线粒体膜蛋白CAMMP1，提示CLAC-CAMMP1蛋白互作是内质网-线粒体之间互作建立的新机制。本项目拟综合利用分子生化和细胞、亚细胞钙影像、超分辨、电镜、动物模型、蛋白组学等多学科交叉技术，系统研究CLAC-CAMMP1调控内质网-线粒体互作的建立、维持和动态变化的新机制，解析其在线粒体功能调节和线粒体自噬中的重要作用，并阐明CAMMP1突变导致内质网-线粒体互作紊乱进而引发先天性乳酸中毒和心脏疾病的病理机制。

内质网和线粒体是细胞内的两个重要细胞器，二者也是细胞内的钙库。内质网-线粒体在结构和功能上的互作调控了钙信号、代谢等多个重要细胞生命活动过程。我们前期发现并命名了由TMCO1四聚体化构成的内质网钙离子通道CLAC，但其在内质网-线粒体互作以及疾病发生中的作用还不清楚。本项目主要研究内容是：筛选鉴定内质网CLAC与线粒体互作因子；研究CLAC及其互作因子对线粒体代谢和线粒体自噬过程的调控；研究内质网-线粒体偶联互作方面的缺陷与人类疾病之间的关系。通过本项目的实施，我们首先通过蛋白质组学分析，发现多个CLAC互作的线粒体蛋白，并对部分互作因子的功能进行了阐释；发现CLAC介导的内质网-线粒体互作在维持内质网-线粒体钙稳态和钙信号轴、线粒体代谢和功能、线粒体氧化应激过程中的重要作用；发现CLAC互作因子Miro2在损伤线粒体的选择性识别和自噬清除过程中的关键作用，并进一步阐明了Miro2介导自噬的具体分子机制；揭示了由CLAC维持的内质网-线粒体钙稳态和钙信号轴在卵泡发育、雌性生育力维持中的重要作用，发现内质网-线粒体钙稳态失衡是导致卵泡耗竭和卵巢早衰的一个新机制；发现内质网钙库-线粒体钙稳态在调控脂滴生成、骨骼发育过程中发挥重要作用；发现CLAC互作因子Miro2在心肌细胞线粒体稳态维持中的重要作用，其功能缺陷导致小鼠心肌细胞衰老样病理变化和心衰，并且人类的先天性高乳酸症和心脏病婴儿中也存在该基因功能性缺失突变；合作研究发现了内质网-线粒体钙稳态与肿瘤细胞耐药相关，可作为肿瘤治疗的新靶标。相关结果使我们对CLAC介导的内质网-线粒体互作在维持内质网-线粒体钙稳态、钙信号、线粒体代谢、线粒体选择性自噬等重要细胞生命活动过程中的重要作用有了新的认识和理解，也为相关疾病机制的阐释和治疗提供了新的靶标和理论基础。发表相关SCI论文11篇，包括Cell Death Differ (2018)、J Cell Biol (2018)、Sci Bull (2019)、Autophagy(2021)、Proc Natl Acad Sci (2022)等；申请国内专利2项，国际PCT专利1项。