The Molecular Basis for Myosin Regulation

摘要 对于所有活着的有机体来说，一个关键的功能是在需要的时候移动的能力。这些运动--细胞内 贩运、细胞分裂、肌肉收缩和细胞运动--都是由分子机器驱动的，这些分子机器施加了一种 考虑到它们的直径只有几纳米，力量的大小令人惊讶。考虑到电机的多样性 细胞中的蛋白质，一个关键的问题是马达在移动货物和应用时如何合作和竞争 力量。一种新兴的范例是特殊马达的概念，即微调的马达 特定的功能。尽管这些马达蛋白很重要，但人们对它们的个体知之甚少。 适应以及这些与细胞中发现的运动模式的关系。这项工作的重点是肌球蛋白-6和 肌球蛋白-10及其独特的细胞调节形式。肌球蛋白-6在细胞器中发挥重要作用 形态、细胞形态、胞质分裂、自噬和内吞作用，而肌球蛋白-10提供必要的 将整合素、钙粘附素和网状蛋白受体等物质运送到细胞前缘的丝足细胞。两者都是 在肿瘤中过度表达，因为它们在膜运输、细胞迁移和转移中发挥作用。这个 这项工作将开发新的方法来控制肌球蛋白-6并分离其在细胞中的活性。其中一个主要的 方法将是用光隔离肌球蛋白-6，使细胞具有光学可切换的斯内尔 Waltzer(肌球蛋白-6缺失)表型。实验的目的是确定肌球蛋白-6何时作为乘客， 锚或单元中的传送器，并且如果需要从一个货物适配器直接切换到另一个 功能。拟议的工作还将调查货物结合和肌球蛋白四元结构如何调整 肌球蛋白-10‘S运动。发展了一种综合的方法，将结构研究与功能研究相结合 重组和单分子运动分析。这项提议将检验以下假设：货物和 细胞外配体结合都是激活肌球蛋白-10所必需的，而且这种物质可以引导肌球蛋白-10从 一种类型的肌动蛋白网络连接到另一种类型。这项研究的完成将全面了解如何 细胞骨架马达蛋白被激活和调节，以完成细胞中的不同任务。运动蛋白的调节是 这是一个具有基本生物学意义的过程，但人们对此知之甚少。这项工作将指导未来的努力 了解多个环境中的激活。

Abstract A critical function for all living organisms is the ability to move when needed. These movements--intracellular trafficking, cell division, muscle contraction, and cell motility-- are driven by molecular machines that exert an amazing amount of force considering that they are only a few nanometers across. Given the variety of motor proteins in the cell, a key question is how motors cooperate and compete while moving cargoes and applying forces. An emerging paradigm is the notion of specialized motors, or motors that are fine-tuned to perform a specific function. Despite the importance of these motor proteins, relatively little is known about their individual adaptations and how these relate to the motility patterns found in the cell. This work focuses on myosin-6 and myosin-10 and their unique forms of cellular regulation. Myosin-6 plays essential roles in organelle morphology, cell morphology, cytokinesis, autophagy, and endocytosis, while myosin-10 delivers essential cargoes such as integrins, cadherins and netrin receptors to filopodia at the leading edge of the cell. Both are overexpressed in tumors, owing to their roles in membrane trafficking, cell migration, and metastasis. The work will develop new approaches to control myosin-6 and isolate its activity in cells. One of the main approaches will be to sequester myosin-6 with light, making cells that have an optically switchable Snell's waltzer (myosin-6 null) phenotype. Experiments are designed to identify when myosin-6 acts as a passenger, an anchor, or a transporter in the cells, and if direct handoff from one cargo adaptor to another is required for function. The proposed work will also investigate how cargo binding and myosin quaternary structure tune myosin-10's motility. An integrated approach is developed, combining structural studies with functional reconstitution and single molecule motility assays. This proposal will test the hypotheses that cargo and extracellular ligand binding are both required for myosin-10 activation, and that cargo can steer myosin-10 from one type of actin network to another. Completion of this study will yield a comprehensive view of how cytoskeletal motor proteins are activated and regulated for distinct tasks in the cell. Motor protein regulation is a process of fundamental biological importance, but is poorly understood. This work will direct future efforts to understand activation in multiple contexts.