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: 10397408
• 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/10397408
• 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; Tubulin; Vesicle; base; cell motility; insight; interest; live cell imaging; polarized cell; presynaptic; reconstitution; single molecule; therapeutically effective; 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 分子马达驱动细胞器沿细胞骨架的主动运输。这辆车是 在神经元中至关重要，这是一种高度极化的细胞，将轴突延伸到1M。轴突不断地 从细胞体中提供新合成的蛋白质和细胞器；主动清除老化蛋白质 而功能障碍的细胞器也需要维持轴突的内稳态。因此，轴突运输由 胞浆动力蛋白和运动蛋白马达的协调活动是必不可少的，而这种转运的缺陷 导致神经退行性变。在这里，我们关注的是动力蛋白和运动蛋白马达的分子配位。 支架蛋白和效应物以及上游调节蛋白和磷酸酶的轴突运输 在长期和时间尺度上保持持续的监管状态。我们还感兴趣的还有 微管和基于肌动蛋白的马达之间的相互作用，这既影响启动和终止 能动性。最后，我们对分子马达和细胞骨架动力学的机制感兴趣。 主动重塑细胞膜，导致变形、管状、分裂和融合。我们会解决的 利用活细胞成像和单个细胞体外重建的协同方法解决这些问题 分子分辨率，以了解所涉及的机制。我们将围绕三大目标展开。目标1： 了解细胞器运输的综合调控。每种细胞器都沿着 轴突有一种与其功能直接相关的独特的运动模式，但我们还不完全了解 调节这种运输的机制。我们将重点研究必要的轴突货物、自噬小体和 信号内体，测试货物特有的马达集成调节所允许的模型 在长时间、规模和距离上持续运输。在目标2中，我们试图理解本地化的 确定的轴突区内细胞器动力学的调节，包括轴突起始段， 突触前部位和轴突终末。这些区域显示出不同的贩运模式，符合 细胞骨架组织的差异：微管捆绑、加端动力学、翻译后 微管蛋白的修饰，以及与肌动蛋白细丝的交叉。我们对增强 启动运输、协调特定舱室分拣和控制货物的限速步骤 在细胞需求的特定位置提供/保留。在目标3中，我们将研究细胞器重塑驱动 通过相反的马达和/或细胞骨架动力学。而一些细胞器在细胞中移动时几乎没有 在形态上的明显变化，其他货物被戏剧性地重塑，经历管状，分裂或 核聚变。我们假设，分子马达和细胞骨架细丝提供了一个适应性强的工具箱，可以 被特别调节以调节动态细胞器的形态。总而言之，这些方法应该提供 轴突运输过程中细胞器动力学的重要新见解。由于轴突运输的缺陷导致 神经变性，进展可能为有针对性和有效的治疗方法提供新的机会。