Regulation of Cytoplasmic Dynein

细胞质动力蛋白的调节

• 批准号: 8630495
• 负责人: Andres Leschziner
• 金额: $ 48.66万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-06-10 至 2018-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8630495
• 关键词: ATP Hydrolysis; Address; Affect; Affinity; Beryllium; Binding; Biochemistry; Biological Assay; Biophysics; Cell physiology; Cells; Cellular biology; Complex; Cryoelectron Microscopy; Defect; Development; Disease; Dynein ATPase; Elements; Fluorescence Microscopy; Goals; Grant; Human; In Vitro; Kinesin; Laboratories; Lead; Mechanics; Mediating; Messenger RNA; Methods; Microtubules; Mitotic spindle; Modeling; Molecular; Molecular Motors; Motor; Motor Activity; Movement; Mutate; Mutation; Myosin ATPase; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Property; Proteins; Regulation; Research; Research Project Grants; Resolution; Role; Structure; Testing; Walking; base; cell motility; dimer; dynactin; in vivo; insight; macromolecule; monomer; mutant; polarized cell; protein complex; public health relevance; research study; single molecule; three dimensional structure

PROJECT SUMMARY The long-term goal of this research project is to understand how the microtubule (MT)-based motor cytoplasmic dynein ("dynein") is regulated. Dynein is the largest and most complex of all cytoskeletal motors; this >1 MDa dimeric complex contains numerous mechanical elements whose movements must be coordinated over strikingly long molecular distances to achieve processive motility along MTs. In addition to its enormous size, dynein is also the most versatile of the molecular motors; in sharp contrast to the 45 kinesins and 39 myosins present in humans, a single dynein gene product is responsible for transporting macromolecules within neurons, constructing the mitotic spindle, polarizing cells, and anchoring mRNAs during development. To give dynein the functional plasticity necessary for carrying out its many roles, several ubiquitous co-factors interact with dynein, including Lis1 and the dynactin complex. This project will apply our combined expertise in biochemistry, single-molecule biophysics and cryo-electron microscopy to address the structural and mechanistic bases of dynein's interaction with MTs and its regulation by Lis1 and dynactin. We recently showed that binding of dynein to MTs is accompanied by conformational changes in its MT- binding domain and that Lis1 acts as a "clutch" to uncouple MT binding and release from ATP hydrolysis, promoting a strongly MT-attached state. Dynactin, a 1.2 MDa complex, enhances dynein's processivity and is required for nearly all dynein functions in cells, but its mechanism of action is poorly understood. This grant will address major mechanistic questions about dynein and its regulation. What are the structural and mechanistic bases of MT binding (Aim 1), and of regulation by Lis1 (Aim 2) and dynactin (Aim 3)?

项目概要 该研究项目的长期目标是了解基于微管（MT）的电机如何 细胞质动力蛋白（“动力蛋白”）受到调节。动力蛋白是所有细胞骨架马达中最大、最复杂的； 这个 >1 MDa 二聚体复合物包含许多机械元件，其运动必须是 在惊人的长分子距离上进行协调，以实现沿 MT 的加工运动。除了它的 动力蛋白尺寸巨大，也是用途最广泛的分子马达；与 45 个驱动蛋白形成鲜明对比 和人类中存在的 39 种肌球蛋白一样，单个动力蛋白基因产物负责运输 神经元内的大分子，构建有丝分裂纺锤体，极化细胞，并在有丝分裂过程中锚定 mRNA 发展。为了赋予动力蛋白发挥其多种作用所需的功能可塑性，一些 普遍存在的辅助因子与动力蛋白相互作用，包括 Lis1 和动力蛋白复合物。该项目将应用我们的 结合生物化学、单分子生物物理学和冷冻电子显微镜方面的专业知识来解决 动力蛋白与 MT 相互作用的结构和机制基础及其受 Lis1 和 dynactin 的调节。 我们最近表明，动力蛋白与 MT 的结合伴随着其 MT 的构象变化。 Lis1 充当“离合器”，解开 MT 结合并从 ATP 水解中释放， 促进强烈的 MT 依附状态。 Dynactin 是一种 1.2 MDa 复合物，可增强动力蛋白的持续合成能力，并且 细胞中几乎所有动力蛋白功能都需要它，但其作用机制尚不清楚。这笔赠款将 解决有关动力蛋白及其调节的主要机制问题。结构和机械原理是什么 MT 结合的碱基（目标 1），以及 Lis1（目标 2）和 dynactin（目标 3）调节的碱基？