Rab8a效应蛋白在Fsp27介导的脂滴融合过程中的作用

The size of lipid droplets (LDs) in adipose tissue indicates body lipid storage capability. The disorder of lipid storage is closely connected to obesity, diabetes, fatty liver and other lipid metabolism diseases. Fsp27 could localize on LD-LD contact site (LDCS) and mediate lipid droplet fusion to form larger lipid droplet in adipocyte. This is crucial for fast increasing lipid storage in adipose tissue to prevent overnutrition caused lipotoxicity by transforming free fatty acid to triacylglycerol and storing in LDs. We found Rab8a could enhance this process, however, its molecular mechanism remains unclear. Purify Rab8a protein as bait, and use affinity chromatography and LC-MS/MS to identify potential Rab8a effectors; then confirm and verify these proteins could function in the process of Fsp27 mediated lipid droplet fusion by co-immunoprecipitation, confocal and live cell imaging, and also investigate its/their mechanisms of regulating lipid droplet fusion by single-molecule imaging, lipid droplet proteomics and lipidomics; in addition, construct gene knock out or transgenic mouse model to study its/their physiological functions in lipid metabolism. This would provide more experimental information and clues for studying Fsp27 mediated lipid droplet fusion and designing cure for obesity and fatty liver.

脂滴的大小反映了脂肪组织储存脂质的能力，并且与肥胖、糖尿病和脂肪肝等代谢疾病的发生密切相关。Fsp27能够介导脂滴融合形成大脂滴，这对于脂肪组织快速增加脂肪储存至关重要，我们发现Rab8a能够促进这一过程，但是其分子机制却缺乏相应的研究。本研究首先以Rab8a为诱饵蛋白，通过亲和层析与质谱联用的方法筛选出潜在的Rab8a效应蛋白；随后在细胞中利用免疫共沉淀，共聚焦显微镜和活细胞成像等方法验证这些潜在的Rab8a效应蛋白能否促进或抑制Fsp27介导的脂滴融合，并利用单分子荧光成像，脂滴蛋白质组学和脂质组学来分析其在脂滴融合过程中的调控机理；最终构建转基因或基因敲除小鼠模型来研究其在机体脂肪代谢过程中的生理功能。通过这些研究能够为研究Fsp27介导的脂滴融合提供更加坚实的实验基础，有助于深入了解肥胖和脂肪肝的发生发展机制，也为该类疾病的干预提供了新的理论基础和药物筛选靶点。

脂滴的大小反映了脂肪组织储存脂质的能力，并且与肥胖、糖尿病和脂肪肝等代谢疾病的发生密切相关。Fsp27能够介导脂滴融合形成大脂滴，这对于脂肪组织快速增加脂 肪储存至关重要，我们发现Rab8a能够促进这一过程，但是其分子机制却缺乏相应的研究。本研究首先以Rab8a为研究节点，通过串联亲和层析与质谱联用的方法筛选出潜在的Rab8a 效应蛋白；并希望在接下来的工作中验证这些蛋白能否在Fsp27介导的脂滴融合过程中发挥功能及探讨其发挥功能的分子机制；我们构建了脂肪组织特异型Rab8a基因敲除小鼠，发现与正常小鼠相比该敲除小鼠能够显著的抵抗高脂饮食引起的肥胖和胰岛素抵抗。通过这些研究能够为研究Fsp27介导的脂滴融合提供更加坚实的实验基础，有助于深入了解肥胖和脂肪肝及胰岛素抵抗的发生发展机制，也为该类疾病的干预提供了新的理论基础和药物筛选靶点。