棕榈酰化修饰对肝脏FAT/CD36蛋白定位和功能的影响及其在非酒精性脂肪性肝病发生中的作用机制研究

Fatty acid translocase (FAT/CD36), a double-transmembrane glycoprotein, which is closely related to fatty acid uptake and metabolic inflammation, plays an important role in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Subcellular localization of FAT/CD36 determines its physiological function. Palmitoylation is a kind of protein modification in post-translation level and regulates membrane protein localization and function. Our preliminary studies demonstrated that palmitic acid or high fat diet causes lipid accumulation in hepatic cells with increased palmitoylation of hepatic FAT/CD36, suggesting that FAT/CD36 palmitoylation may be a key factor in the progressing of NAFLD. However, how palmitoylation regulates FAT/CD36 localization and function in hepatic cells is largely unknown. . We propose that increased palmitoylation of hepatic FAT/CD36 promotes more efficient incorporation into membrane rafts (important domains at the cell surface) and enhances FAT/CD36 protein stability and ability of fatty acid uptake resulting in hepatic steatosis. Meanwhile, increased palmitoylation FAT/CD36 activates NF-κB inflammatory signaling pathway by more assembly of heterodimer with toll like receptor (TLR) 4/6 in hepatic plasma membrane causing hepatitis. In the absence of palmitoylation, posttranslational processing is considerably delayed leading to the accumulation of the FAT/CD36 precursor molecule in the endoplasmic reticulum and mitochondria. Abrogation of palmitoylation by mutation or inhibitor also poorly incorporates into plasma membrane rafts and reduces efficiency of uptake of fatty acid and FAT/CD36-mediated inflammation. We will examine this hypothesis in cultured hepatic cells and in mice carrying lentivirus vector with wild-type of FAT/CD36 or mutation of FAT/CD36 palmitoylation site.

脂肪酸转运酶（FAT/CD36）是与脂肪酸摄取和代谢性炎症产生密切相关的膜糖蛋白，它在细胞膜及亚器的定位决定了其功能，在非酒精性脂肪性肝病（NAFLD）中扮演了重要角色。棕榈酰化修饰是蛋白质翻译后脂质共价修饰的一种形式，对蛋白质定位和功能有重要调节作用。我们前期发现：肝FAT/CD36棕榈酰化修饰对NAFLD发生至关重要，然而棕榈酰化修饰如何影响肝FAT/CD36定位和功能、导致NAFLD发生的分子机制尚不清楚。. 本项目拟应用课题组已建立的多种实验技术，探讨棕榈酰化修饰如何调节FAT/CD36在肝细胞膜（脂筏）及内质网、线粒体上定位，进而影响它对脂肪酸摄取和肝脏代谢性炎症的产生，深入分析其与NAFLD发生发展的关系，弄清去棕榈酰化是否可以改善由FAT/CD36介导的脂肪肝，从一全新角度阐释NAFLD的发病机制，为防治NAFLD及针对FAT/CD36靶点的新药开发提供实验依据。

脂肪酸转运酶（FAT/CD36）是与脂肪酸摄取和代谢性炎症产生密切相关的膜糖蛋白，它在细胞膜及亚器的定位决定了其功能，在非酒精性脂肪性肝病（NAFLD）中扮演了重要角色。棕榈酰化修饰是蛋白质翻译后脂质共价修饰的一种形式，对蛋白质定位和功能有重要调节作用。本项目应用课题组已建立的多种实验技术，探讨棕榈酰化修饰如何调节FAT/CD36在肝细胞膜（脂筏）及内质网、线粒体上定位，进而影响它对脂肪酸摄取和肝脏代谢性炎症的产生，深入分析其与NAFLD发生发展的关系。通过一系列体内外实验，我们发现：与正常人和单纯脂肪变性患者相比，NASH患者肝细胞质膜上CD36的定位显着增加。在患有NASH的小鼠肝脏中，还观察到CD36的棕榈酰化增加和CD36在肝细胞质膜上的定位增加。此外，抑制CD36棕榈酰化可保护小鼠免受NASH的侵害。棕榈酰化的抑制降低了CD36蛋白的疏水性，从而减少了其在质膜以及肝细胞脂筏中的定位。抑制棕榈酰化会降低HepG2细胞中的脂肪酸摄取和CD36/Fyn/Lyn复合物的形成。CD36棕榈酰化的抑制不仅可以通过激活AMPK途径改善细胞内脂质的积累，还可以通过抑制JNK信号通路来抑制炎症反应。因此，我们提出了一个重要的创新性理论：CD36转录后蛋白修饰，而不是单纯的蛋白表达，决定了该蛋白功能并参与非酒精性脂肪性肝炎病变的发生；抑制CD36棕榈酰化可能是未来防治非酒精性脂肪性肝炎发生发展的重要靶点。该研究从一全新角度阐释了NAFLD的发病机制，为防治NAFLD及针对FAT/CD36靶点的新药开发提供了实验依据。