ER-mitochondrial communication in calcium signaling, energy metabolism and liver disease

钙信号传导、能量代谢和肝脏疾病中的内质网线粒体通讯

• 批准号: 10785141
• 负责人: GYORGY CSORDAS
• 金额: $ 14.59万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10785141
• 关键词: 3-Dimensional; Affinity; Autophagocytosis; BCL2L1 gene; Binding; Calcium; Calcium Signaling; Cell Line; Cell Respiration; Cell Survival; Cells; Cellular biology; Clustered Regularly Interspaced Short Palindromic Repeats; Communication; Communication Research; Competence; Complex; Computer software; Coupling; Cytoplasm; Data; Data Set; Dependence; Descriptor; Dimensions; Disease; Electron Microscopy; Elements; Endoplasmic Reticulum; Energy Metabolism; Evaluation; Fatty Liver; Genetic; Geometry; Health; Hela Cells; Hepatic; Hepatocyte; Hormones; Image; Impairment; Laboratories; Linear Models; Link; Liver; Liver diseases; Mammalian Cell; Measures; Mediating; Medicine; Membrane; Metabolic; Metabolism; Methods; Mitochondria; Modeling; Molecular; Molecular Analysis; Molecular Target; Morphology; Movement; Neurons; Nucleotides; Obesity; Organ; Organelles; Outer Mitochondrial Membrane; Pharmaceutical Preparations; Phenotype; Physiological; Process; Production; Protein Isoforms; Proteins; Proteomics; Quantitative Evaluations; Reactive Oxygen Species; Reagent; Regulation; Research Personnel; Role; Scheme; Scientist; Signal Transduction; Site; Specificity; Structure; Structure-Activity Relationship; Techniques; Testing; Tissues; Vesicle; Work; cell type; functional disability; imaging approach; immune function; in vivo imaging; lipid biosynthesis; lipid metabolism; microscopic imaging; mitochondrial dysfunction; mitochondrial membrane; mouse model; nano; nanoscale; non-alcoholic fatty liver; non-alcoholic fatty liver disease; novel; protein complex; receptor; scaffold; software development; tool; uptake

ABSTRACT The effects of various hormones on oxidative metabolism and other mitochondrial functions, in the liver and other tissues, are mediated by cytoplasmic [Ca2+] oscillations propagated to the mitochondria. Ca2+ is released to the cytoplasm from the endoplasmic reticulum (ER) through the IP3 receptors (IP3Rs), which, based on findings from us and others, expose mitochondria at ER-mitochondria contact sites (ERMCs) to high [Ca2+] nanodomains to attain activation of the low-affinity mitochondrial Ca2+ uptake sites. ERMCs were also recognized in other processes including lipid metabolism, organelle dynamics and autophagy. Our work has revealed the physical support of ERMCs by tethering proteins. We have created synthetic membrane linkers to measure and perturb ERMCs and local inter-organelle communication in live cells, and provided clues to local calcium and reactive oxygen species (ROS) signaling. We have also provided these reagents to several hundred laboratories worldwide, which together have showed the role of interorganellar contacts in a range of paradigms including metabolism, vesicle dynamics, neuronal and immune functions and linked structural or functional impairments of the ER-mitochondrial coupling to an array of disorders across organs including the liver (e.g. fatty liver). However, fundamental questions remain unanswered. ERMCs are dynamically restructured to meet the continuously changing demands of the cell, but how ERMCs are formed and dissolved is yet to be determined. IP3R-mediated fluctuations in [Ca2+] might provide a means to control contact formation, given that elevations of cytoplasmic [Ca2+] stop mitochondrial movements close to the ER through the Ca2+-sensing Miro proteins, and both the IP3R and Miro proteins, have been implicated as components of interorganellar complexes. However, the interaction partners and mechanisms are elusive. We hypothesize that IP3Rs and Miros mediate ERMC formation in an isoform-specific and Ca2+-dependent manner to regulate physiological functions of hepatocytes. Aims#1&2 will test this hypothesis using novel genetic and microscopic imaging toolkits that will enable us to specifically and systematically measure and perturb ERMC forming elements. The effect of genetic perturbations in the liver will be tested by novel, in vivo imaging approaches. The interactomes of IP3R and Miro will be evaluated primarily by unbiased proteomics, but Bcl-xL will also be specifically tested as a tether forming partner for the IP3R. Finally, a major limitation in the field of inter-organellar communication research is that quantitative evaluation of the geometry of nanometer scale membrane contacts remains difficult. In Aim#3 we will develop and characterize methods of measuring and describing organelle interface geometry in 2-and 3D electron microscopy data and uncover the structural features most relevant to calcium transfer. The proposed work will explore the molecular mechanisms of ERMC dynamics and their physiological relevance and will continue our efforts in creating molecular tools and methods that allow many investigators to explore the local communications of ER and mitochondria or other organelles.

抽象的 各种激素对肝脏和其他线粒体功能的氧化代谢和其他线粒体功能的影响 组织是由传播到线粒体传播的细胞质[Ca2+]振荡介导的。 Ca2+已发布到 通过IP3受体（IP3RS）来自内质网（ER）的细胞质，该发现基于发现 从我们和其他 获得低亲和力线粒体Ca2+摄取位点的激活。在其他方面也认识到ERMC 包括脂质代谢，细胞器动力学和自噬在内的过程。我们的工作揭示了身体 通过绑定蛋白质来支持ERMC。我们创建了合成膜接头来测量和扰动 ERMC和活细胞中的局部轨道间通信，并为局部钙和反应性提供了线索 氧气（ROS）信号传导。我们还向数百个实验室提供了这些试剂 在全球范围内，它共同展示了在包括 代谢，囊泡动力学，神经元和免疫功能以及连接的结构或功能障碍 与包括肝脏在内的器官（例如脂肪肝）的一系列疾病的ER-线粒体耦合。 但是，基本问题仍未得到解决。 ERMC被动态重组以满足 细胞的需求不断变化，但是如何形成和溶解ERMC 决定。 [Ca2+]中的IP3R介导的波动可能提供控制接触形成的方法，鉴于 细胞质[Ca2+]的升高通过Ca2+sensing miro靠近ER的线粒体运动 蛋白质以及IP3R和miro蛋白都被视为跨加工的组成部分 复合物。但是，互动伙伴和机制难以捉摸。我们假设IP3RS和 miros以同种特异性和Ca2+依赖性方式介导ERMC形成，以调节 肝细胞的生理功能。目标＃1和2将使用新的遗传和 微观成像工具包将使我们能够专门和系统地测量和扰动ERMC 形成元素。遗传扰动在肝脏中的影响将通过新颖的体内成像来测试 方法。 IP3R和MIRO的相互作用将主要通过无偏的蛋白质组学评估，但是BCl-XL 还将被专门作为IP3R的系绳组成合作伙伴进行测试。最后，在该领域的主要限制 掌管间通信研究是对纳米量表几何形状的定量评估 膜接触仍然很困难。在AIM＃3中，我们将开发并表征测量方法和 描述2和3D电子显微镜数据中的细胞器界面几何形状并发现结构 与钙转移最相关的特征。拟议的工作将探索ERMC的分子机制 动态及其生理相关性，并将继续我们在创建分子工具和方法方面的努力 这使许多研究人员能够探索ER和线粒体或其他细胞器的当地通信。