Spectral revelations of mitochondrial Ca2+ flux interactome

线粒体 Ca2 通量相互作用组的光谱揭示

• 批准号: 8668369
• 负责人: Jeffrey L Caplan
• 金额: $ 36.14万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-15 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8668369
• 关键词: Affinity; Amino Acids; Antibodies; Autophagocytosis; Binding; Bioenergetics; Biology; Cell Culture Techniques; Cell Death; Cells; Chronic; Collaborations; Complex; Data; Electron Transport; Equipment and supply inventories; Functional disorder; Future; Goals; Image; Inner mitochondrial membrane; Ions; Label; Laboratories; Link; Maps; Metabolic; Metabolic Diseases; Metabolism; Mitochondria; Molecular; Monitor; Names; Nature; Oxidoreductase; Pathway interactions; Permeability; Porifera; Process; Production; Proteins; Proteomics; Proton Pump; Qualifying; Reaction; Recruitment Activity; Regulation; Research Personnel; Resolution; Rest; Role; Signal Transduction; Specificity; Stable Isotope Labeling; System; Technology; Testing; Validation; Variant; in vivo; protein complex; public health relevance; reconstitution; response; small hairpin RNA; stable cell line; uptake

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.

描述（由申请人提供）：线粒体是细胞质[Ca2+]通量的多元信号处理器。线粒体[Ca2+]通过激活线粒体酶复合物参与细胞能量的产生，但也可以促进不同的细胞死亡模式。线粒体通过众多途径精确地维持胞质Ca2+梯度。线粒体Ca2+摄取的失调与许多细胞功能障碍有关，包括慢性氧化负担，自噬和对细胞死亡的敏感性。高细胞质[Ca2+]促进了线粒体Ca2+通过未知直到2011年Uniporter的积累。最近，我们证明了线粒体Ca2+摄取1（MICU1）和线粒体Ca2+ Uniporter调节剂1（MCUR1）负面控制线粒体Ca2+ Uniporter孔隙孔（MCU）在静止状态下的活性。 MCU分别与MICU1和MCUR1相互作用，并在线粒体内膜形成不同的MCU复合物。鉴于线粒体Ca2+对MCU Interactome的重要性至关重要，因此我们假设MCU是MCU复合物的主要“集线器”，并且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复合物组件至关重要，将取决于胞质Ca2+水平。该提案的目的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结合伙伴。为了确定功能性线粒体Ca2+通量相互作用组的识别，招募了四名高素质的研究者。 Muniswamy和Merali实验室将确定MCU相互作用的伙伴，并专注于与MCU分子相互作用的同一性和功能。此外，将在重构系统（Mancia Lab）中测试MCU Intercontome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 Intervormem络合物。线粒体生物学的关键下一步要求这四个研究者和技术的合作，因此为未来的大分子相互作用的合作提供了理想的策略。结果将大大增强我们对MCU复合物的理解及其在细胞正常状态下的线粒体Ca2+涌入中的作用。