内质网介导的自噬小体的膜生成和调控机制

自噬是细胞的一种自我保护机制，它介导胞内受损蛋白和细胞器以及外源侵染物等进入溶酶体进行降解，具有高度的保守性。自噬信号途径的失调与多种疾病如神经退行性疾病、癌症和免疫相关疾病的发生密切相关。自噬作为一个细胞内的动态过程，其膜起源和膜延伸调控机制是本领域亟待解决一个基础问题。研究表明内质网是自噬小体膜的重要供体。本项目将从三个不同的角度，对内质网依赖的自噬小体膜起源做系统的分析。我们将首先利用APEX临近标记系统和抓捕ULK1磷酸化底物的方法，寻找自噬流中参与自噬小体膜延伸的因子，之后对这些候选因子如COPII囊泡组装相关蛋白和Annexin家族蛋白进行功能分析和疾病相关突变体的机制研究。我们也将利用CRISPR/CAS9系统并整合HaloTag标记系统，筛选在病原菌自噬条件下参与自噬小体膜延伸的基因，以找到内质网介导的自噬小体膜调控机制，并分析细胞如何维持内质网内平衡以应对自噬的发生。

Autophagy is a central cellular mechanism for the elimination of damaged proteins, protein complexes, and organelles. This evolutionarily conserved process plays a crucial role in the cellular response to nutrient deprivation as well as defense against pathogens. Dysregulation of autophagy has been implicated in the pathogenesis of various human diseases, including cancers, neurodegenerative diseases, and immune related diseases. Thus, understanding the molecular mechanism of autophagosome biogenesis is pivotal for the development of future autophagy-targeting therapeutics. Numerous studies suggested the involvement of Endoplasmic Reticulum (ER) in the autophagosome biogenesis. However, the detail mechanism of how the process is controlled is not clearly identified. This proposal is aimed to elucidate the mechanism of how ER mediates autophagosome biogenesis and membrane extension. We will combine multiple strategies including APEX proximity labeling approach, ULK1 substrate identification, as well as CRISPR/CAS9 screen plus HaloTag labeling system to identify the genes involved in ER mediated autophagosome biogenesis. We will then use biochemistry and super-resolution imaging technologies to decipher the mechanism of how ER helps autophgosome biogenesis and at the same time keeps its homeostasis.