Regulation of Cytoplasmic Dynein

描述（由申请人提供）：本研究项目的长期目标是了解基于微管（MT）的运动细胞质动力蛋白（“动力蛋白”）是如何调节的。动力蛋白是所有细胞骨架马达中最大和最复杂的；这种bbb1mda二聚体复合体包含许多机械元件，它们的运动必须在非常长的分子距离上协调才能沿着mt实现进程运动。除了其巨大的尺寸外，动力蛋白也是分子马达中最通用的；与人类中存在的45种动力蛋白和39种肌球蛋白形成鲜明对比的是，一个动力蛋白基因产物负责在神经元内运输大分子，构建有丝分裂纺锤体，使细胞极化，并在发育过程中锚定mrna。为了使动力蛋白具有执行其许多作用所必需的功能可塑性，几种普遍存在的辅助因子与动力蛋白相互作用，包括Lis1

DESCRIPTION (provided by applicant): 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 o 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)?