Molecular and physiological analysis of mitochondrial calcium uptake

线粒体钙摄取的分子和生理分析

• 批准号: 9036744
• 负责人: Julia Chang Liu
• 金额: --
• 依托单位: U.S. NATIONAL HEART LUNG AND BLOOD INST
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-10-01 至 2018-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9036744
• 关键词: Affect; Animal Model; Animals; Ataxia; Autophagocytosis; Behavior; Binding Proteins; Biochemical; Bioenergetics; Biological; Biological Assay; Biological Models; Body Composition; Body Weight; CRISPR/Cas technology; Calcium; Cell Death; Cell Line; Cell physiology; Cells; Co-Immunoprecipitations; Complex; Confocal Microscopy; Cultured Cells; Disease; EF Hand Motifs; Embryo; Energy Metabolism; Exhibits; Fibroblasts; Functional disorder; Gatekeeping; Generations; Genetic; Genus Hippocampus; Health; Homeostasis; Human; Innovative Therapy; Knock-out; Knockout Mice; Lead; Learning Disorders; Link; Measurement; Measures; Membrane Proteins; Metabolism; Mitochondria; Mitochondrial Diseases; Mitochondrial Matrix; Modeling; Molecular; Molecular Weight; Mus; Myopathy; Names; Necrosis; Patients; Permeability; Phenocopy; Phenotype; Physiological; Physiology; Play; Production; Property; Proteins; Regulation; Resources; Respiration; Role; Signaling Molecule; Site; Stimulus; Structure; Swelling; Testing; Tissues; Tremor; VDAC1 gene; Wild Type Mouse; calcium uniporter; clinically relevant; extracellular; human disease; loss of function mutation; mitochondrial permeability transition pore; motor disorder; mouse model; mutant; paralogous gene; protein complex; public health relevance; response; uptake

 DESCRIPTION (provided by applicant): Popularly known as "the powerhouse of the cell," mitochondria are not only the site of metabolism and energy generation but also a hub for other cellular processes, including the initiation of cell death. Mitochondrial uptake of the signaling molecule calcium plays a role in the stimulation of ATP production, but too much calcium can lead to opening of the mitochondrial permeability transition pore (mPTP), triggering necrosis. The recent identification of the molecule forming the pore through which calcium can rapidly enter the mitochondria, the mitochondrial calcium uniporter (MCU), has provided a genetic means to directly test the functional importance of calcium uptake. In particular, MCU is part of a large multi-protein complex including other protein components. EMRE and MICU1 are two of these proteins that in cell lines have been shown to play critical roles in regulation of calcium uptake. EMRE was found to be necessary for MCU activity, and its deletion blocked calcium from entering mitochondria. Though its mechanism is controversial, MICU1 has been characterized as a gatekeeper of MCU, inhibiting MCU activity at low levels of extramitochondrial calcium and stimulating MCU when calcium levels rise. This project began with the generation of the first animal models of EMRE and MICU1 deletion. The aims of this project are to determine the impact of EMRE and MICU1 deletion on isolated mitochondria, on primary cells, and on the physiology of the whole animal. The new EMRE and MICU1 knockout mice also make it possible to determine the topology of the calcium uniporter complex, measure effects on basal bioenergetics, and elucidate the role of EMRE and MICU1 respectively in the regulation of mitochondrial calcium uptake and homeostasis. Furthermore, mouse models will reveal the consequences of mitochondrial calcium deregulation on cell death responses, physiological phenotypes including body weight and composition, and disease pathophysiology. Interestingly, human patients with loss-of-function mutations in MICU1 present with conditions such as proximal myopathy and extrapyramidal motor disorder. MICU1 knockout mice are preliminarily observed to exhibit ataxia and tremors, suggesting that this model may mirror human disease features and thus potentially motivate innovative therapies to treat mitochondrial disorder. Altogether, EMRE and MICU1 knockout mice are valuable resources for answering biological questions with both basic and clinical relevance.

 描述（由申请人提供）：线粒体通常被称为“细胞的发电站”，不仅是代谢和能量产生的场所，而且是其他细胞过程的中心，包括细胞死亡的启动。线粒体摄取信号分子钙在刺激ATP产生中起作用，但过多的钙可导致线粒体通透性转换孔（mPTP）打开，引发坏死。最近鉴定的分子形成的孔，通过它钙可以迅速进入线粒体，线粒体钙单向转运体（MCU），提供了一种遗传手段，直接测试钙摄取的功能重要性。特别是，MCU是 包括其它蛋白质组分的大的多蛋白质复合物。EMRE和MICU1是这些蛋白质中的两种，在细胞系中已被证明在钙摄取的调节中发挥关键作用。EMRE被发现是MCU活动所必需的，其缺失阻止钙进入线粒体。尽管其机制存在争议，但MICU1已被表征为MCU的看门人，在低水平的线粒体外钙时抑制MCU活性，并在钙水平升高时刺激MCU。 该项目始于EMRE和MICU1缺失的第一个动物模型的产生。该项目的目的是确定EMRE和MICU1缺失对分离的线粒体、原代细胞和整个动物的生理学的影响。新的EMRE和MICU1基因敲除小鼠还可以确定钙单向转运体复合物的拓扑结构，测量对基础生物能量学的影响，并分别阐明EMRE和MICU1在线粒体钙摄取和稳态调节中的作用。此外，小鼠模型将揭示线粒体钙失调对细胞死亡反应、生理表型（包括体重和组成）以及疾病病理生理学的影响。有趣的是，MICU1功能缺失突变的人类患者会出现近端肌病和锥体外系运动障碍等疾病。初步观察到MICU1基因敲除小鼠表现出共济失调和震颤，这表明该模型可能反映了人类疾病特征，从而可能激发治疗线粒体疾病的创新疗法。总之，EMRE和MICU1基因敲除小鼠是回答基础和临床相关生物学问题的宝贵资源。