Probing structural and biophysical mechanisms of mitochondrial membrane ultrastructure

探究线粒体膜超微结构的结构和生物物理机制

• 批准号: 10661778
• 负责人: Luke H. Chao
• 金额: $ 40.98万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-15 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10661778
• 关键词: Apoptosis; Autosomal Dominant Optic Atrophy; Beds; Biophysical Process; Calcium Signaling; Cardiac; Cardiac health; Cell physiology; Cells; Complex; Computer Analysis; Crista ampullaris; Cryo-electron tomography; Cryoelectron Microscopy; Dependence; Disease; Elements; Encephalopathies; Environment; Family; Foundations; Goals; Heart Mitochondria; Heterogeneity; Homeostasis; In Vitro; Inner mitochondrial membrane; Knowledge; Leigh Disease; Lipids; Liposomes; Liver diseases; Malignant Neoplasms; Membrane; Membrane Proteins; Metabolic; Metabolism; Mitochondria; Modeling; Molecular Conformation; Morphology; Mutate; Nature; Nerve Degeneration; OPA1 gene; Organelles; Physiological; Physiology; Play; Production; Property; Protein Conformation; Proteins; Regulation; Resolution; Role; Shapes; Structure; System; Testing; Tissues; Visualization; biophysical techniques; computational pipelines; early onset; interest; member; membrane reconstitution; mitochondrial membrane; nervous system disorder; particle; programs; protein complex; protein protein interaction; protein structure; reconstitution; solute; targeted treatment; therapeutic development; tool

PROJECT SUMMARY/ABSTRACT Organelle morphology is specialized for the highly adapted functions found in complex tissues. Mitochondrial ultrastructure is exquisitely tuned to metabolic and physiological state. Abnormal morphology is a hallmark of neurological disorders, cardiac conditions and cancer. With the cryo-EM `resolution revolution', we have developed dramatic new understanding of membrane protein structure and regulation, but our knowledge of organelle structure lags behind. This is due to the pleomorphic nature of organelles, and the challenge of assigning specific morphological features to protein states. The overarching goals of my lab are to understand, at a mechanistic level, how protein conformational change is influenced by subcellular context, how membrane ultrastructure is regulated by protein factors, and the functional interplay of these elements in physiology and disease. Over the next five years, my group will develop a technical platform that combines electron cryo- microscopy (cryo-EM) and biophysical methods to study mitochondrial ultrastructure and its regulation. We will apply our recent developed in vitro reconstitution systems to visualize reconstituted membrane proteins in liposomes and bilayers by single-particle cryo-EM. We will mature our newly established electron cryo- tomography (cryo-ET) pipeline for computational analyses of membrane properties to understand their dependence on protein-protein interactions. We shall develop new structural and biophysical methods to characterize organelle lipid heterogeneity. And finally, we will explore assembly or protein complexes in native contexts. Together these approaches will advance mechanistic understanding of protein conformational state and help us identify the fundamental determinants of organelle shape. We are broadly interested in questions of membrane spacing, composition and curvature. We will develop tools precisely tailored for these questions using mitochondria as a test bed, exploring the structure and function of candidate factors that regulate mitochondrial membrane morphology, which play causal roles in neurodegenerative conditions. Opa1 is the inner-membrane fusogen and cristae remodeler mutated in Dominant Optic Atrophy. SLC25A46 is an outer- membrane member of the solute transporter family that plays important roles in coordinating lipid homeostasis in Leigh Syndrome. MICOS is the stabilizer and regulator of cristae junctions (the `choke-point' to the mitochondrial inner-membrane folds) whose loss results in early-onset fatal mitochondrial encephalopathy with liver disease. This project's immediate impacts include sharing new models to understand mitochondrial shape with cell biologists, equipping pharmacologists with new, highly specific conformational targets for therapeutic development, and providing physiologists with fundamental rules for understanding tissue specialization. The long-term goal is to build an extensible approach generalizable to other organelles, and a foundation for rational control of organelle morphology from first principles.

项目概要/摘要 细胞器形态专门用于复杂组织中发现的高度适应的功能。线粒体 超微结构根据代谢和生理状态进行精确调整。形态异常是其标志 神经系统疾病、心脏病和癌症。随着冷冻电镜“分辨率革命”的到来，我们已经 对膜蛋白结构和调节产生了戏剧性的新理解，但我们的知识 细胞器结构落后。这是由于细胞器的多形性，以及 将特定的形态特征分配给蛋白质状态。我实验室的首要目标是了解， 在机制水平上，蛋白质构象变化如何受到亚细胞环境的影响，膜如何 超微结构受蛋白质因子调节，这些元素在生理和功能上的功能相互作用 疾病。未来五年，我的团队将开发一个结合电子冷冻技术的技术平台 显微镜（冷冻电镜）和生物物理方法研究线粒体超微结构及其调节。我们将 应用我们最近开发的体外重建系统来可视化重建的膜蛋白 通过单颗粒冷冻电镜观察脂质体和双层。我们将使我们新建立的电子冷冻技术成熟起来 断层扫描 (cryo-ET) 管道，用于对膜特性进行计算分析，以了解其特性 依赖于蛋白质-蛋白质相互作用。我们将开发新的结构和生物物理方法 表征细胞器脂质异质性。最后，我们将探索天然的组装或蛋白质复合物 上下文。这些方法将共同促进对蛋白质构象状态的机械理解 并帮助我们确定细胞器形状的基本决定因素。我们对问题广泛感兴趣 膜间距、成分和曲率。我们将开发专门针对这些问题定制的工具 使用线粒体作为试验台，探索调节候选因子的结构和功能 线粒体膜形态，在神经退行性疾病中发挥因果作用。 Opa1 是 显性视神经萎缩中内膜融合剂和嵴重塑剂发生突变。 SLC25A46是一个外 溶质转运蛋白家族的膜成员，在协调脂质稳态中发挥重要作用 在利氏综合症中。 MICOS 是嵴连接处的稳定器和调节器（“阻塞点”） 线粒体内膜褶皱），其缺失会导致早发性致命性线粒体脑病 肝脏疾病。该项目的直接影响包括共享新模型以了解线粒体形状 与细胞生物学家合作，为药理学家提供新的、高度特异性的治疗构象靶标 发育，并为生理学家提供理解组织特化的基本规则。这 长期目标是建立一种可推广到其他细胞器的可扩展方法，并为 从第一原理合理控制细胞器形态。