Regulation of Dynein-Mediated Transport

项目总结 组织是生命的一个基本和决定性的特征，从亚细胞到所有的尺度 一直到生物体的水平。在细胞水平上，分子和细胞器必须位于 具有适当的时间和空间精度，以便进程可以根据需要进行 手机。类似地，细胞被安排成适当的层以定义基本的组织结构，这提供 器官功能的基础，并支持有机体的健康。亚细胞和细胞的主要决定因素 组织是在整个细胞环境中运输不同货物的分子马达。这样的一个 马达是细胞质动力蛋白，它在所有细胞中沿着微管轨迹运输各种类型的货物 周期阶段，在许多类型的细胞中。例如，动力蛋白是主要的逆行微管马达 将许多囊泡和蛋白质运送到神经元的细胞体。除了精心策划 适当的亚细胞组织，动力蛋白在组织的建立和维持中起着重要作用 建筑。例如，细胞命运和随之而来的组织结构的一个主要决定因素是取向 以及有丝分裂纺锤体相对于细胞边界的位置。除了本地化到 小泡状货物的膜-从那里影响它们的运输-动力蛋白马达被锚定在 质膜通过精确调谐的相互作用来定向和定位纺锤体 微管。在生物体发育和组织动态平衡等过程中，纺锤体定向 和位置决定了细胞分裂的平面，从而决定了细胞是对称分裂还是不对称分裂。 对称的干细胞分裂产生两个完全相同的干细胞，而切换到不对称分裂的结果是 在一个干细胞和一个分化细胞中，这促进了组织层化。因此，动力蛋白是一种至关重要的 在许多级别上决定生物组织的分子。动力蛋白的精确机制 通过适当的空间和时间控制来执行所有这些不同的功能是不清楚的。缺乏这样的能力 信息阻碍了可能预防或逆转的有效疗法的发展 细胞和组织结构的缺陷，可导致各种破坏性的疾病(例如 皮质发育、运动神经元疾病)。在拟议的研究中，我们将使用体外和 细胞生物学方法确定动力蛋白被调节以执行其货物的机制 运输功能。具体地说，我们将：(1)解决无脑相关蛋白的机制 Lis1启动动力蛋白介导的货物运输；(2)确定动力蛋白如何影响纺锤运动 定向精确度；(3)确定各种关键调节因子影响动力蛋白的方式和分子基础 以及，(4)研究细胞周期状态和动力蛋白活性之间的协调和相互作用。我们的 研究将为指导大量货物运输的基本机制提供重要的见解 空间和时间的精确度，从而建立和维持细胞和最终的生物体健康。

PROJECT SUMMARY Organization is a fundamental and defining feature of life at all degrees of scale, from the subcellular level all the way up to the level of the organism. On the cellular level, molecules and organelles must be situated with appropriate temporal and spatial precision such that processes may proceed according to the needs of the cell. Similarly, cells are arranged into appropriate layers to define underlying tissue organization, which provides the basis for organ function, and to support health of the organism. Major determinants of subcellular and cellular organization are molecular motors that transport diverse cargoes throughout the cellular environment. One such motor is cytoplasmic dynein, which transports numerous types of cargoes along microtubule tracks during all cell cycle stages, and within many cell types. For instance, dynein is the major retrograde microtubule motor that transports many vesicular and protein cargoes toward the cell body of neurons. In addition to orchestrating appropriate subcellular organization, dynein plays a major role in the establishment and maintenance of tissue architecture. For instance, a major determinant of cell fate and consequent tissue organization is the orientation and position of the mitotic spindle with respect to the boundaries of the cell. In addition to localizing to the membrane of small vesicular cargoes – from where it effects their transport – dynein motors are anchored at the plasma membrane from where they orient and position the spindle through precisely tuned interactions with microtubules. During such processes as organismal development and tissue homeostasis, spindle orientation and position dictate the plane of cell division, and thus whether a cell divides symmetrically or asymmetrically. Symmetric stem cell divisions result in two identical stem cells, whereas a switch to asymmetric division results in one stem cell and a differentiated cell, which promotes tissue stratification. Thus, dynein is a critically important molecule that dictates biological organization on many levels of scale. The precise mechanisms by which dynein performs all these disparate functions with appropriate spatial and temporal control are unclear. The lack of such information presents an impediment towards the development of effective therapies that may prevent or reverse defects in cellular and tissue organization that can lead to various devastating disorders (e.g., malformations of cortical development, motor neuron diseases). In the proposed studies, we will use a combination of in vitro and cell biological approaches to determine the mechanisms by which dynein is regulated to perform its cargo transport functions. Specifically, we will: (1) resolve the mechanism by which the lissencephaly-related protein LIS1 initiates dynein-mediated cargo transport; (2) determine how dynein effects spindle movements with precise directional precision; (3) determine how, and the molecular basis by which various critical regulators affect dynein activity; and, (4) investigate the coordination and interplay between cell cycle state and dynein activity. Our studies will provide critical insight into fundamental mechanisms that dictate transport of numerous cargoes with spatial and temporal precision such that cellular and ultimately organismal health is established and maintained.