Molecular Mechanisms of Axonal Transport and Organelle Dynamics

轴突运输和细胞器动力学的分子机制

• 批准号: 9922337
• 负责人: Erika L Holzbaur
• 金额: $ 66.28万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9922337
• 关键词: Actins; Active Biological Transport; Affect; Afferent Neurons; Aging; Amyotrophic Lateral Sclerosis; Autophagosome; Axon; Axonal Transport; Cells; Cytoskeletal Filaments; Cytoskeletal Modeling; Cytoskeleton; Defect; Disease; Dynein ATPase; Endosomes; Exhibits; Goals; Homeostasis; In Vitro; Kinesin; Lead; Length; Mediating; Membrane; Microfilaments; Microtubule Bundle; Microtubules; Modeling; Modification; Molecular; Molecular Motors; Morphology; Motor; Motor Neurons; Movement; Nerve Degeneration; Neurons; Organelles; Pattern; Phosphoric Monoester Hydrolases; Phosphotransferases; Presynaptic Terminals; Proteins; Regulation; Resolution; Scaffolding Protein; Signal Transduction; Site; Sorting - Cell Movement; Testing; Time; Translations; Treatment Efficacy; Tubulin; Vesicle; base; cell motility; insight; interest; live cell imaging; polarized cell; presynaptic; reconstitution; single molecule; trafficking

Project Summary Molecular motors drive the active transport of organelles along the cellular cytoskeleton. This transport is critically important in neurons, highly polarized cells that extend axons up to 1m. Axons are continuously supplied with newly synthesized proteins and organelles from the cell body; active clearance of aging proteins and dysfunctional organelles is also required to maintain axonal homeostasis. Thus, axonal transport driven by the coordinated activities of cytoplasmic dynein and kinesin motors is essential, and deficits in this transport cause neurodegeneration. Here we focus on the molecular coordination of dynein and kinesin motors during axonal transport by scaffolding proteins and effectors, and the upstream regulatory kinases and phosphatases that maintain a sustained regulatory state over long length- and time-scales. We are also interested in interactions between microtubule- and actin-based motors, which affect both the initiation and termination of motility. Finally, we are interested in the mechanisms by which molecular motors and cytoskeletal dynamics actively remodel organelle membranes, leading to deformation, tubulation, fission and fusion. We will tackle these questions using the synergistic approaches of live cell imaging and in vitro reconstitution with single molecule resolution to understand the mechanisms involved. We will focus on three major goals. Goal 1: Understanding the integrated regulation of organelle transport. Each type of organelle moving along the axon has a distinct pattern of motility that directly relates to its function, but we do not yet fully understand the mechanisms regulating this transport. We will focus on essential axonal cargos, autophagosomes and signaling endosomes, testing the model that the cargo-specific, integrated regulation of motors allows for sustained transport over long time scales and distances. In Goal 2, we seek to understand the localized regulation of organelle dynamics within defined axonal zones, including the axon initial segment, presynaptic sites, and the axon terminal. These zones exhibit distinct trafficking patterns that correspond to differences in cytoskeletal organization: microtubule bundling, plus-end dynamics, post-translation modifications of tubulin, and intersections with actin filaments. We are interested in mechanisms that enhance the rate-limiting step of transport initiation, mediate compartment-specific sorting, and control cargo delivery/retention at specific sites of cellular need. And in Goal 3, we will study organelle remodeling driven by opposing motors and/or cytoskeletal dynamics. While some organelles move through the cell with little evident change in morphology, other cargos are dramatically remodeled, undergoing tubulation, fission or fusion. We hypothesize that molecular motors and cytoskeletal filaments provide an adaptable toolbox that can be specifically tuned to regulate dynamic organelle morphology. Together, these approaches should provide important new insights into organelle dynamics during axonal transport. As deficits in axonal transport lead to neurodegeneration, progress may provide new opportunities for targeted and effective therapeutic approaches.

项目摘要 分子电动机驱动细胞器沿细胞骨骼的主动运输。这种运输是 在神经元中至关重要的是高度极化的细胞，将轴突延伸至1M。轴突连续 由细胞体的新合成蛋白和细胞器提供；衰老蛋白的主动清除率 维持轴突稳态也需要功能失调的细胞器。因此，轴突运输由 细胞质动力蛋白和驱动蛋白电动机的协调活性至关重要，并且在该运输中的缺陷 引起神经变性。在这里，我们着重于在 辅助蛋白质和效应子以及上游调节激酶和磷酸酶的轴突运输 在长度和时间尺度上保持持续的监管状态。我们也对 微管和基于肌动蛋白的电动机之间的相互作用，这既影响 运动。最后，我们对分子电机和细胞骨架动力学的机制感兴趣 积极重塑细胞器膜，导致变形，软化，裂变和融合。我们将解决 这些问题使用活细胞成像的协同方法和单一的体外重构 分子解析以了解所涉及的机制。我们将专注于三个主要目标。目标1： 了解细胞器运输的综合调节。每种类型的细胞器沿着 轴突具有与其功能直接相关的独特运动模式，但我们尚未完全理解 调节这种运输的机制。我们将专注于必需的轴突符号，自噬体和 信号传导内体，测试货物特异性，集成的电动机允许的模型 长时间尺度和距离持续运输。在目标2中，我们试图了解本地化 调节定义的轴突区域内细胞器动力学，包括轴突初始段， 突触前部位和轴突末端。这些区域表现出不同的贩运模式 细胞骨架组织的差异：微管捆绑，加末端动力学，翻译后 小管蛋白的修饰以及与肌动蛋白丝的相交。我们对增强的机制感兴趣 运输启动，中介隔室特定排序和控制货物的限制步骤 在细胞需求的特定部位输送/保留。在目标3中，我们将研究Organelle重塑驱动 通过对立电动机和/或细胞骨架动力学。而某些细胞器几乎没有 形态学的明显变化，其他货物进行了巨大的重塑，经历了软化，裂变或 融合。我们假设分子电机和细胞骨架丝提供了一个适应性的工具箱，可以 专门调节以调节动态细胞器形态。这些方法在一起应该提供 轴突运输过程中对细胞器动力学的重要新见解。由于轴突运输中的缺陷导致 神经变性，进展可能为有效的治疗方法提供新的机会。