Mechanism of centrosome regulation from the Golgi

描述（由申请人提供）：该项目的目的是了解哺乳动物细胞中调节相邻高尔基体中心体的控制机制。虽然高尔基体和中心体的接近是哺乳动物细胞所独有的，并且在酵母、植物或果蝇细胞中没有观察到，但其功能意义一直难以捉摸。我们实验室的工作已经确定了一条中心体调节途径，其中高尔基体蛋白 GM130 会导致称为 Tuba 的鸟嘌呤核苷酸交换因子激活高尔基体的 Cdc42。该项目旨在确定高尔基体的这些事件如何控制中心体的组织和功能，以及它们是否依赖于高尔基体-中心体的邻近性。 目标 1 将确定 GM130 是否通过增加 Tuba 对 Cdc42 的结合亲和力或通过将 Cdc42 招募到高尔基体来激活高尔基体的 Cdc42。该目标还将通过测定 GM130 是否控制高尔基体中已知受 Cdc42 调节的其他细胞过程（例如高尔基体到内质网的运输和高尔基体的局部肌动蛋白组装）来检查 GM130 是否是高尔基体中 Cdc42 的主要调节者。还将研究高尔基体相关 Cdc42 的潜在负调节因子。 目标 2 将研究 GM130-Cdc42 通路如何对中心体发挥作用。将研究 Cdc42 效应器 Par6？，以确定它是否是该调节途径的下游组件，以及它是否通过运动蛋白动力蛋白通过与动力蛋白亚基 p150Glued 相互作用转运到中心体。 PCM-1 是一种将额外蛋白质招募到中心体的蛋白质，将对其进行研究以确定它是否在 Par6 下游发挥作用？通过 GM130-Cdc42 途径介导中心体组织的控制。 目标 3 将通过检查 Cdc42 是否可以从高尔基体动员到中心体从而调节中心体组织来研究高尔基体-中心体接近的功能意义。高尔基体的中心粒周围位置将被破坏，作为测试高尔基体中心体接近是否对于 GM130-Cdc42 途径调节中心体组织所必需的另一种方法。

DESCRIPTION (provided by applicant): The objective of this project is to understand a control mechanism that regulates the centrosome from the adjacent Golgi apparatus in mammalian cells. While the proximity of the Golgi and centrosome is unique to mammalian cells, and not observed in yeast, plant or fly cells, its functional significance has been elusive. Work in our laboratory has identified a centrosome regulatory pathway in which a Golgi protein, GM130, causes a guanine nucleotide exchange factor called Tuba to activate Cdc42 at the Golgi. This project seeks to determine how these events at the Golgi control centrosome organization and function and whether they depend on Golgi-centrosome proximity. Aim 1 will determine whether GM130 activates Cdc42 at the Golgi by increasing the binding affinity of Tuba for Cdc42 or by recruiting Cdc42 to the Golgi. This aim will also examine if GM130 is the major regulator of Cdc42 at the Golgi by assaying whether GM130 controls additional cellular processes at the Golgi that are known to be regulated by Cdc42, such as Golgi to ER transport and local actin assembly at the Golgi. A potential negative regulator of Golgi-associated Cdc42 will also be studied. Aim 2 will examine how the GM130-Cdc42 pathway exerts its effect on the centrosome. A Cdc42 effector, Par6?, will be studied to determine if it is a downstream component of this regulatory pathway and if it is transported to the centrosome by the motor protein, dynein, through interactions with the dynactin subunit, p150Glued. PCM-1, a protein that recruits additional proteins to the centrosome, will be studied to determine if it acts downstream of Par6? to mediate the control of centrosome organization by the GM130-Cdc42 pathway. Aim 3 will study the functional significance of Golgi-centrosome proximity by examining if Cdc42 can be mobilized from the Golgi to the centrosome so that it can regulate centrosome organization. The pericentriolar location of the Golgi will be disrupted as another means of testing whether Golgi centrosome proximity is necessary for regulation of centrosome organization by the GM130-Cdc42 pathway.