E3泛素连接酶SIAH1调控肝细胞脂质代谢的作用及机制研究

Non-alcoholic fatty liver disease (NAFLD) has become the first major cause of abnormal biochemical indexes of liver in China. The disorder of liver lipid metabolism is the key factor leading to the development of NAFLD. In recent years, ubiquitin enzymes have become important targets for the development of new drugs to treat NAFLD. As an E3 ubiquitin ligase, the function and mechanism of SIAH1 regulating liver lipid metabolism have not been elucidated. Our previous studies have shown that SIAH1 can be upregulated in NAFLD and promote the development of NAFLD, and knockout of SIAH1 displayed an attenuated hepatic lipid deposition, and overexpression of HNF4a or SULT2B1 can inhibit the role of SIAH1 on promoting lipid deposition. In this study, we construct high expressed SIAH1 mice in C57BL/6J mice, SIAH1 knockout mice and mouse primary hepatocytes, and put forward the hypothesis that lipids deposition in hepatocytes can increase SIAH1 expression, which promoting the ubiquification degradation of HNF4a, and the decrease of HNF4a protein level inhibited the transcription level of SULT2B1, while the low expression of SULT2B1 promoted the increase the expression of promoting lipid synthesis genes, and finally the degree of lipids deposition in hepatocytes is aggravated. This project will elucidate the role and molecular mechanism of SIAH1 in NAFLD at animal, tissue, cell and molecular levels, so as to provide new therapeutic targets for NAFLD.

非酒精性脂肪性肝病（NAFLD）已成为我国第一大慢性肝病和健康体检肝生物化学指标异常的首要原因，肝脂质代谢紊乱是导致NAFLD发生发展的关键因素。近年来泛素化酶已成为研究、开发治疗NAFLD新药的重要靶点。SIAH1作为一种E3泛素连接酶，其调控肝脂质代谢的作用和机制尚未阐明。我们前期研究发现，SIAH1在NAFLD中可被诱导表达并促进NAFLD的发展，SIAH1基因敲除可显著改善小鼠的肝脂质沉积，且高表达HNF4a或SULT2B1可抑制SIAH1的促脂质沉积作用。本课题提出研究假说：肝细胞脂质淤积可诱导SIAH1表达升高，进而促进HNF4a泛素化降解，进而抑制SULT2B1的转录水平，而低表达SULT2B1则可上调促脂质合成基因表达，最终加剧肝细胞的脂质淤积。本项目将从动物、组织、细胞、分子水平多层次阐明SIAH1在NAFLD中的作用及分子机制，以期为治疗NAFLD提供新的作用靶点。

非酒精性脂肪肝病（NAFLD）是最常见的肝脏疾病之一，并伴有多种代谢功能障碍。尽管肝细胞脂质淤积已被确定为NAFLD发展的最重要肝脏表现，但其分子机制复杂，尚未完全阐明。本项目中，我们通过体内、外实验发现E3泛素连接酶Siah1（Seven in absentia homolog 1, Siah1）的表达水平与NAFLD的进展呈显著正相关，Siah1表达水平改变可影响NAFLD的进展。Siah1表达下调后，利用液相色谱-质谱（LC-MS/MS）分析蛋白组和泛素化组学变化，发现被其修饰的泛素化的蛋白质广泛分布在细胞质、细胞核和质膜中，并参与多种疾病的发生发展，为Siah1的生物学功能提供了新的见解，为相关功能和分子机制研究打下基础。通过分析筛选出固醇载体蛋白2（Scp2）可能是Siah1影响NAFLD的目标靶基因。进一步的分子机制研究发现，Siah1与Scp2相互作用，并促进其泛素化和降解，最终影响肝细胞内脂质淤积程度。综上研究结果，本课题通过高脂饮食喂养构建NAFLD动物模型，首次明确E3泛素连接酶Siah1在NAFLD中可被诱导表达并促进NAFLD病程发展，为今后NAFLD的研究和治疗提供新的研究方向；其次，本课题首次发现Siah1可作为Scp2一个新的泛素化酶，能够介导Scp2的泛素化降解；最后本课题从分子、细胞、组织、动物整体水平和临床样本等多方面进行实验设计，首次阐明肝细胞脂质淤积时可诱导Siah1表达升高，促进Scp2的泛素化降解，Scp2蛋白水平降低抑制肝细胞脂质转运，最终加剧肝细胞的脂质淤积程度，促进NAFLD的进展。目前为止，我们的研究结果首次报告了Siah1与NAFLD的病程进展呈正相关，Siah1通过Scp2泛素化参与肝脏脂质代谢。Siah1有望为NAFLD的药物治疗策略提供新的潜在靶点。