Spectral revelations of mitochondrial Ca2+ flux interactome

描述（由申请人提供）：线粒体是细胞质[Ca2+]流的多元信号处理器。线粒体[Ca2+]通过激活线粒体酶复合物参与细胞能量产生，但也可以促进不同模式的细胞死亡。线粒体通过多种途径精细地维持胞质Ca 2+梯度。线粒体Ca 2+摄取的失调与许多细胞功能障碍有关，包括慢性氧化负荷、自噬和细胞死亡敏化。高细胞质[Ca2+]通过一个直到2011年才发现的单向转运体促进线粒体Ca2+积累。最近，我们发现线粒体Ca 2+摄取1（MICU1）和线粒体Ca 2+单向转运体调节因子1（MCUR1）在静息和活动状态下分别负性和正性调控线粒体Ca 2+单向转运体孔提交（MCU）活性。MCU分别与MICU1和MCUR1相互作用，在线粒体内膜形成不同的MCU复合物。鉴于线粒体Ca2+对MCU相互作用组的至关重要性，我们假设MCU是MCU复合物的主要“枢纽”，并与蛋白质网络相互作用以形成MCU复合物。此外，MCU相互作用组件对于MCU复合物组装至关重要，并且将取决于细胞溶质Ca2+水平。该提案的目标是通过应用最先进的蛋白质组学技术和超分辨率成像来识别和验证哺乳动物MCU结合伴侣。为了完成功能性线粒体Ca2+通量相互作用组的鉴定，招募了四名高素质的研究人员。Muniswamy和Merali实验室将确定MCU相互作用的伙伴，并专注于与MCU分子相互作用的身份和功能。此外，将在重构系统（Mancia实验室）中测试MCU相互作用组的物理相互作用。最后，Caplan实验室将通过超分辨率成像可视化MCU相互作用组复合体。线粒体生物学的关键下一步需要这四个研究人员和技术的合作，从而为未来大分子相互作用的合作提供理想的策略。这些结果将大大提高我们对MCU复合物及其在细胞正常和活跃状态下线粒体Ca2+内流中的作用的理解。

DESCRIPTION (provided by applicant): Mitochondria are multivariate signal processors of cytoplasmic [Ca2+] flux. Mitochondrial [Ca2+] participates in cellular energy production via activation of mitochondrial enzymatic complexes yet can also promote different modes of cell death. Mitochondria exquisitely maintain cytosolic Ca2+ gradient via numerous pathways. Dysregulation of mitochondrial Ca2+ uptake has been linked to numerous cellular dysfunctions including chronic oxidative burden, autophagy and sensitization for cell death. High cytoplasmic [Ca2+] facilitates mitochondrial Ca2+ accumulation via an unknown until 2011 uniporter. Recently, we demonstrated that mitochondrial Ca2+ uptake 1 (MICU1) and mitochondrial Ca2+ uniporter regulator 1 (MCUR1) negatively and positively control the mitochondrial Ca2+ uniporter pore submit (MCU) activity under resting and active state. MCU interacts with MICU1 and MCUR1 separately and forms different MCU complexes at the mitochondrial inner membrane. Given the critical importance of mitochondrial Ca2+ for the MCU interactome, we hypothesize that MCU is a major "hub" of the MCU complex and interacts with a network of proteins to form the MCU complex. Further, MCU interacting components are crucial for MCU complex assembly and will be dependent on cytosolic Ca2+ levels. The goal of the proposal is to identify and validate mammalian MCU binding partners by the application of state-of-the-art proteomic technologies and super-resolution imaging. To accomplish the identification of the functional mitochondrial Ca2+ flux interactome, four highly qualified investigators are recruited. Muniswamy and Merali laboratories will identify the MCU interacting partners and focus on the identity and function of molecular interactions with MCU. Further, the physical interaction of the MCU interactome will be tested in a reconstituted system (Mancia lab). Finally, the Caplan lab will visualize the MCU interactome complex with super-resolution imaging. The critical next step in mitochondrial biology requires the collaboration of these four investigators and technologies thus providing an ideal strategy for future collaborations of macromolecular interactions. The results will greatly enhance our understanding of the MCU complex and its role in mitochondrial Ca2+ influx during the normal and active states of the cell.