膜相关自噬基因lgg-2和atg-9调控内吞底物胞内循环运输的机制研究

细胞自噬和细胞内吞都与胞内膜结构紧密相关，两者共同调控细胞内外底物循环运输过程，提供细胞所需各类生物大分子，维持细胞稳态。然而，人们对自噬基因及自噬通路如何影响内吞过程了解的较少。在线虫肠道细胞中利用内吞底物标记蛋白hTAC::GFP进行RNAi筛选，我们得到了两个与膜结构相关的自噬基因，lgg-2和atg-9。lgg-2是自噬基因atg-8在线虫中同源物之一，被PE修饰的Atg8蛋白能够与自噬小体的膜结构相联系；atg-9编码所有自噬蛋白中唯一一个跨膜蛋白。在本项目中，我们拟首先研究lgg-2和atg-9的功能缺失对于不同类型内吞底物循环运输的影响，并确定lgg-2和atg-9基因发挥作用的具体阶段；其次我们将利用免疫沉淀等方法寻找LGG-2和ATG-9的相互作用蛋白，以期揭示lgg-2和atg-9调控内吞底物胞内循环运输过程的分子机制，进一步阐明细胞自噬与内吞循环过程之间的关系。

Autophagy and endocytosis pathways are closely interconnected with intracellular membrane compartments and both involved in cargo recycling transport regulation, providing nutrients and molecules to maintain cellular homeostasis. How autophagic pathway may affect endocytosis remains poorly understood. RNAi screen using hTAC::GFP as the marker has got two essential autophagy genes, lgg-2 and atg-9, both change the pattern of hTAC::GFP in the intestine of C elegans. LGG-2 and ATG-9 proteins are related to intracellular membrane structures. lgg-2 is one of the homolog of core autophagy gene atg8 in yeast. PE-conjugated Atg8 can associate with autophagosome membrane. ATG-9 is the only known transmembrane protein required for autophagy and is proposed to deliver membrane to the preautophagosome structures. Here we plan to examine the effect of lgg-2 and atg-9 on different types of endocytic cargo recycling, and use epistasis analysis to determine the role of lgg-2 and atg-9 in the hierarchy of endocytic recycling regulation. In addition, by means of immunoprecipitation and the like, we will identify regulator or effector proteins of LGG-2 and ATG-9 in the regulation pathway. The mechanism that lgg-2 and atg-9 regulate endocytic cargo recycling will shed light on the interconnection of the autophagosomal and endocytic trafficking routes.