A systematic analysis of the regulation of small G proteins on the Golgi apparatus

高尔基体是分泌途径的中心分选站，也是连接内体系统的关键。囊泡运输途径在高尔基体的多样性是由Arf和Rab家族的小G蛋白，其中每一个本地化到一个子集的高尔基体膜。这些G蛋白的定位由特定的调节剂控制，所述调节剂激活G蛋白以诱导从胞质溶胶到特定膜的易位，或激活G蛋白并导致释放回到胞质溶胶。已经描述了各种调节因子家族，并且它们也与特定的膜外周相关。有趣的是，最近的证据表明，G蛋白可以通过与第二个G蛋白的调节因子相互作用来控制第二个G蛋白的定位。这样的级联反应可以耦合细胞器的身份时间事件的细胞器的成熟，但只有少数G蛋白的关系已经研究到data.We的目的是进行系统的调查，本地化到高尔基体的果蝇培养细胞的所有Rabs和Arfs的调节。昆虫的高尔基体与哺乳动物的高尔基体相似，但果蝇中的基因比哺乳动物中的旁系同源基因更少，因此便于系统研究。我们将选择所有的果蝇Rabs和Arfs及其已知的调节子定位于高尔基体，并系统地研究它们之间的关系。首先，我们将研究RNA干扰介导的敲除单个G蛋白后每个蛋白的定位。其次，我们将测试单个固定化G蛋白和每个调节器之间的直接相互作用。这些数据将被整合到一个网络中，我们希望解释G蛋白如何在高尔基体中进行调节，并解释在这个细胞器中发生的运输过程的准确性。该网络将通过将调节因子重新定位到其他细胞器并测试这些发现是否在哺乳动物细胞中保守来验证。

The Golgi apparatus is the central sorting station of the secretory pathway and the key connection to the endosomal system. The diversity of vesicular transport routes at the Golgi is regulated by the Arf and Rab families of small G proteins each of which localizes to a subset of Golgi membranes. The localization of these G proteins is controlled by specific regulators that either activate G proteins to induce translocation from the cytosol to a specific membrane or inactivate G proteins and cause release back to the cytosol. Various regulator families have been described and they are also peripherally associated with specific membranes. Interestingly, recent evidence has indicated that a G protein can control the localization of a second G protein by interacting with a regulator of that second G protein. Such cascades could couple organelle identity to temporal events as organelles mature, but the relationship of only a few G proteins has been studied to date.We aim to perform a systematic investigation of the regulation of all Rabs and Arfs that localize to the Golgi of Drosophila cultured cells. The insect Golgi is similar to its mammalian counterpart but genes in Drosophila tend to have fewer paralogs than in mammals, thus facilitating systematic studies. We will select all the Drosophila Rabs and Arfs and their known regulators that localize to the Golgi and study systematically their relationship. First, we will examine the localization of each protein upon RNA interference mediated knock-down of individual G proteins. Second, we will test for direct interactions between individual immobilized G proteins and each regulator. The data will be integrated into a network that we expect to explain how G proteins are regulated at the Golgi and account for the accuracy of transport processes that take place at this organelle. The network will be validated by relocating regulators to other organelles and by testing if the findings are conserved in mammalian cells.