小G蛋白Rab13调控血管内皮细胞巨自噬的作用及机理研究

血管内皮细胞巨自噬是影响内皮功能的重要因素，但其分子机制并未搞清。本课题组利用新鉴定的小分子自噬诱导剂紫檀芪处理血管内皮细胞，发现小G蛋白Rab13上调显著。阻断Rab13抑制紫檀芪诱导的自噬，而过表达Rab13诱导血管内皮细胞自噬。我们构建了Rab13的三种突变体，发现只有表达Rab13活化突变体能够诱导血管内皮细胞自噬。据此，我们推测：活化态Rab13可能参与调控血管内皮细胞自噬。此外，免疫共沉淀结果显示，Rab13和有丝分裂原诱导基因6（MIG6）互作。在此基础上，本项目拟进一步解析活化态Rab13在血管内皮细胞自噬中的作用；验证活化态Rab13与MIG6的互作；研究Rab13是否通过MIG6调节血管内皮细胞自噬以及二者是否存在调控关系；探索Rab13和mTOR信号的关系以及MIG6在其中的作用，为阐明血管内皮细胞自噬分子机制提供新的实验证据，为心脑血管疾病的防治提供新的线索和靶点。

Macroautophagy is an important factor affecting endothelial function, but its molecular mechanism remains unclear. We used pterostilbene, a newly identified small-molecule inducer of autophagy, to induce vascular endothelial cell autophagy and found that the small GTPase Rab13 is significantly up-regulated. Knockdown of Rab13 suppressed the pterostilbene-induced autophagy, whereas Rab13 overexpression could induce autophagy in vascular endothelial cells. We generated three Rab13 mutants and found that only expression of the constitutively active Rab13 mutant could induce autophagy in vascular endothelial cells. Accordingly, we speculated that the active form of Rab13 might be involved in the regulation of autophagy in vascular endothelial cells. In addition, co-immunoprecipitation analyses showed that Rab13 could interact with mitogen-induced gene 6 (MIG6). Based on these findings, we will further analyze the role of the active form of Rab13 in autophagy in vascular endothelial cells; validate the interaction between the active form of Rab13 and MIG6; determine whether Rab13 regulates vascular endothelial cell autophagy through MIG6 and whether a regulatory relationship exists between them; investigate the relationships between mTOR signaling and Rab13 as well as MIG6. The project will provide new experimental evidence for understanding the mechanism underlying vascular endothelial cell autophagy and provide new clues and targets for the prevention and treatment of cardiovascular diseases.