细胞自噬是真核生物中降解非正常物质或细胞器的一种保守机制，参与多种重要的生理过程。有报道参与囊泡运输的Rab小G蛋白及相关调控因子也会影响细胞自噬，但具体机制不明确。我们最近发现酵母Rab小G蛋白Ypt51及其上游激活因子Vps9和下游效应物Vps3、Vps8（属于CORVET复合体专一性亚基）均参与了细胞自噬，缺失后细胞内都产生香蕉状结构。本项目在这些新发现的基础上，探讨囊泡运输与细胞自噬这两种生理过程在早期内体和后期内体之间如何偶联。本研究将采用活体荧光和电镜观察、免疫印迹检测及体外离体实验等方法探讨上述四个蛋白：缺失对细胞自噬的影响；过量表达Ypt51对其它三个蛋白缺失所产生自噬表型的恢复情况；与自噬蛋白的互作；激活的Ypt51通过与CORVET复合体互作结合自噬体的情况。本研究结果将揭示酵母中参与囊泡运输的Ypt51调控细胞自噬的机制，为细胞自噬调控机制研究提供重要理论基础。

Autophagy is a conserved process for degrading abnormal materials or organells in eukaryotic cells, and is required in multiple physiological processes, including growth, development, proliferation and stress resistance. It was reported that some Rab GTPases and their related regulators and effectors, whose main functions are in vesicle trafficking, are also involved in autophagy, but the mechanisms are still unclear. We recently found that Rab GTPase Ypt51, its guanine excahnge factor (GEF) Vps9, and its effectors Vps3 or Vps8 (specific-subunits of CORVET, class C core vacuole-endosome transport) are all involved in autophagy. Dysfunctioning one of these four proteins resulted in accumulation of autophagical protein Atg8 between the nucleus and the vacuole as banana-shaped structures, significantly different from the multiple Atg8 dots in the dysfunction of Ypt7, or its effector proteins Vps39 or Vps41 (specific-subunits of HOPS, homotypic vacuolar fusion and protein sorting). Based on our new finding of the functions of these four proteins in autophagy, we intent to explore the coordination of vesicle trafficking and autophagy between early endsome and late endosome. Fluorescence microscopy, electron microscopy, western blot and in vivo experiments would be used to examine: 1) the effect of either protein dysfucntion on autophagy; 2) suppression effect of Ypt51 on the defective autophagy of the dysfunction of other three proteins; 3) the direct interaction between these four proteins and autophagical proteins, such as Atg8, Atg9 and Ape1 etc.; 4) the binding activity on autophagosomes through the interaction between active Ypt51 and CORVET complex. These results will uncover the regulatory role of Ypt51 on autophagy, and provide mechanistic insight on how Rab small GTPases affect autophagy. This project will provide important theory base for mechanism study of autophagy.