Rab32调控猪脂肪细胞脂滴代谢的脂质组学基础

The meat quality of pork is closely related to adipose cells, and the lipid droplet metabolism in adipose cells is quite important. In the early study, applicant found that Rab32, the member of the Rab family, is important in the regulation of mice lipid droplet metabolism, and the mechanism of Rab32 involving in lipid metabolism is not clear. There is no report of Rab32 in pigs and pig fat metabolism has its species specificity. Therefore, this project will detect the biological function of Rab32 in porcine adipocytes by functional proteomics for the first time. The molecular detection, genome editing technique and electron microscopy will be used to reveal the tissue expression profile of Rab32, the function and the distribution of organelles in the course of adipogenic differentiation of porcine subcutaneous fat cells. In order to analyze the mechanism of Rab32 regulating lipid droplet metabolism, we will establish the technology platform of separation lipid droplets, and use this to purify lipid droplets, the purified lipid droplets will be detected their lipidomics and the related data will be used to predict which lipid regulated by Rab32. The study on the molecular mechanism of regulating of lipid droplets in pigs by Rab32 will provide reference for the functional studies of other lipid metabolism related genes, it can also provide theoretical basis for the improvement of pork quality，and the prevention and treatment of diseases with lipid metabolism disorders.

猪肉品质与脂肪细胞密切相关，而脂肪细胞中脂滴代谢机制十分重要。申请人前期研究显示Rab家族成员Rab32在小鼠脂滴代谢中起着关键作用，但机制不明，猪中尚未见报道，且猪脂肪代谢具有其物种特异性。因此，本项目将首次引入可以大规模检测生物体内脂质特性及功能的脂质组学研究Rab32在猪脂肪细胞内的生物学作用。拟通过分子检测、基因编辑和透射电镜等技术阐明Rab32在猪皮下脂肪细胞成脂分化过程的表达规律、作用及其对细胞器分布的影响。为全面解析Rab32调控脂滴代谢的机理，拟建立脂滴分离技术平台，获取高纯度脂滴样本，对其进行规模系统的脂质组学检测并挖掘出差异显著的脂质(量及结构)，再检测该脂质的关键调控因子以揭示Rab32在成脂过程的调控机理。研究清楚Rab32调控猪脂滴代谢分子机制将为其他脂代谢相关基因的功能研究提供借鉴，也可为猪肉品质的改良和脂滴代谢相关疾病防治的研究提供理论依据。

猪肉品质与脂肪细胞密切相关，而脂肪细胞中脂代谢机制十分重要。本项目发现Rab32在巴马猪皮下脂肪组织SVF细胞成脂分化过程中的作用具有剂量效应，在超高表达Rab32后会抑制PPARγ及CEBPβ基因的表达，干扰组shRab32-3会抑制PPARγ及CEBPβ基因的表达，但其他两个干扰组没有这种变化。干扰后对这两成脂关键基因的调控趋势不一致，引起这种不同变化的机制还需深入研究。从成脂分化基因动态表达情况来看Rab1a、Rab5a、Rab6a、Rab7a、Rab8a、Rab14、Rab32、Rab18、Rab38在SVF细胞诱导成脂分化过程中均可能发挥着重要的作用。另外，猪SVF细胞成脂分化前后的转录组及脂质组都发现了极显著的变化，结合两组学的结果显示PPAR 信号通路, 脂肪细胞脂解的调节通路上的差异基因与差异脂质组存在正相关，特别是Mmp3，Adipoq等基因与大部分差异脂质相关。综上Rab1a、Rab5a、Rab6a、Rab7a、Rab8a、Rab14、Rab18、Rab38、Mmp3、Adipoq等可以作为后续继续深入研究猪脂代谢调控的候选基因。这可为后续的相关研究提供思路。