Regulation of Dynein-Mediated Transport

项目摘要 组织是生命的一个基本的和决定性的特征，从亚细胞到细胞， 一路上升到有机体的水平。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，使得过程可以根据用户的需要进行。 cell.类似地，细胞被排列成适当的层以限定下面的组织结构，这提供了 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 组织是分子马达，在整个细胞环境中运输各种货物。一个这样 马达是细胞质动力蛋白，它在所有细胞周期中沿着沿着微管轨道运输多种类型的货物。 周期阶段，并在许多细胞类型中。例如，动力蛋白是主要的逆行微管马达， 向神经元的细胞体运输许多囊泡和蛋白质货物。除了精心策划 动力蛋白是一种适当的亚细胞组织，在组织的建立和维持中起着重要作用。 架构例如，细胞命运和随后的组织组织结构的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-从那里它影响他们的运输-动力蛋白马达锚定在 通过精确调节的相互作用， 微管在生物体发育和组织稳态的过程中， 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相同的干细胞，而转换到不对称分裂产生 在一个干细胞和一个分化的细胞中，这促进了组织分层。因此，动力蛋白是一个至关重要的 在许多尺度上决定生物组织的分子。动力蛋白的精确机制 在适当的空间和时间控制下执行所有这些不同的功能是不清楚的。缺乏这种 信息阻碍了有效疗法的发展， 可导致各种破坏性疾病的细胞和组织组织的缺陷（例如，畸形 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 细胞生物学方法来确定动力蛋白被调节以执行其货物的机制 运输功能。具体而言，我们将：（1）解决脑缺血相关蛋白的机制， LIS 1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体运动， 方向精确性;（3）确定各种关键调节剂如何影响动力蛋白以及其分子基础 （4）研究细胞周期状态与动力蛋白活性之间的协调和相互作用。我们 研究将提供关键的洞察力的基本机制，决定运输的许多货物， 空间和时间的精确性，从而建立和维持细胞和最终生物体的健康。

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.