W09D10.1/SMAP在高尔基体至内体的极性胞吐运输中的功能机制研究

Polarized exocytosis is required for the establishment and maintenance of plasma membrane polarity. Previous studies found that the small GTPase RAB-11/Rab11 and phosphatidylinositol homeostasis play a critical regulatory role in the regulation of polarized exocytosis, but the regulatory mechanism of RAB-11 activity and the regulation of related organelle phosphatidylinositol levels are still not clear. Our preliminary studies found that W09D10.1/SMAP is implicated in the transport of cargo proteins from Golgi to the endosomes. Loss of W09D10.1 leads to abnormal accumulation of cargo proteins in the cytoplasm and the loss of RAB-11 membrane localization. Furthermore, PI(3)P level rises and the PI(4)P level decreases in W09D10.1-depleted cells. This current study intends to use the whole animal research system to explore the mechanistic role of W09D10.1 in regulating the activity of RAB-11, and to clarify the synergistic regulation mechanism of W09D10.1 and RAB-11 on the phosphatidylinositol level in Golgi and endosomes, revealing the functional significance of phosphatidylinositol dynamics in polarized exocytosis. The implementation of this study will deepen the understanding of the molecular regulatory network of the polarized exocytic transport pathway under physiological conditions.

极性胞吐运输参与了细胞质膜极性的建立和维持。此前的研究发现小G蛋白RAB-11/Rab11和磷脂酰肌醇稳态维持在极性胞吐运输中扮演重要调控角色，但RAB-11活性调控机制和相关细胞器磷脂酰肌醇水平调控过程尚不明晰。我们的前期工作发现W09D10.1/SMAP参与介导货物膜蛋白从高尔基体至内体的极性胞吐转运，W09D10.1缺失导致细胞质中胞吐货物膜蛋白异常累积、RAB-11膜定位丧失、PI(3)P水平上升和PI(4)P水平降低。本项目拟运用整体动物研究手段探索W09D10.1调控RAB-11活性的机理，阐明W09D10.1和RAB-11对高尔基体、内体上磷脂酰肌醇水平的协同调控机制，揭示磷脂酰肌醇动态在极性胞吐运输中功能意义。本研究的实施将加深对生理状态下极性胞吐运输通路的分子调控网络的理解。

极性胞吐运输参与了细胞质膜极性的建立和维持，反式高尔基体管网状结构TGN是重要分选细胞器。本项目系统性研究W09D10.1/SMAP的亚细胞定位和功能作用，阐明了W09D10.1定位于TGN，调控系列货物膜蛋白的极性分选。衔接蛋白复合体AP-1对W09D10.1的胞内正常定位是必须的。W09D10.1的功能缺失减少了网格蛋白在TGN处的组装，过表达网格蛋白可以缓解突变体中货物膜蛋白的错误定位。我们的工作一方面揭示了AP-1、W09D10.1、网格蛋白在极性细胞胞吐运输中的功能和调控关系，同时在项目开展过程中发现了经由非经典蛋白分泌途径的货物膜蛋白，为后续非经典蛋白分泌研究的开展奠定了基础。