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Abstract：
Non-alcoholic fatty liver disease (NAFLD) has emerged as the leading cause of chronic liver disease worldwide. Caspase 8 and FADD-like apoptosis regulator (CFLAR) has been identified as a potent factor in mitigating non-alcoholic steatohepatitis (NASH) by inhibiting the N-terminal dimerization of apoptosis signal-regulating kinase 1 (ASK1). While arginine methyltransferase 1 (PRMT1) was previously reported to be associated with increased hepatic glucose production, its involvement in hepatic lipid metabolism remains largely unexplored. The interaction between PRMT1 and CFLAR and the methylation of CFLAR were verified by Co-IP and immunoblotting assays. Recombinant adenoviruses were generated for overexpression or knockdown of PRMT1 in hepatocytes. The role of PRMT1 in NAFLD was investigated in normal and high-fat diet-induced obese mice. In this study, we found a significant upregulation of PRMT1 and downregulation of CFLAR after 48h of fasting, while the latter significantly rebounded after 12h of refeeding. The expression of PRMT1 increased in the livers of mice fed a methionine choline-deficient (MCD) diet and in hepatocytes challenged with oleic acid (OA)/palmitic acid (PA). Overexpression of PRMT1 not only inhibited the expression of genes involved in fatty acid oxidation (FAO) and promoted the expression of genes involved in fatty acid synthesis (FAS), resulting in increased triglyceride accumulation in primary hepatocytes, but also enhanced the gluconeogenesis of primary hepatocytes. Conversely, knockdown of hepatic PRMT1 significantly alleviated MCD diet-induced hepatic lipid metabolism abnormalities and liver injury in vivo, possibly through the upregulation of CFLAR protein levels. Knockdown of PRMT1 suppressed the expression of genes related to FAS and enhanced the expression of genes involved in FAO, causing decreased triglyceride accumulation in OA/PA-treated primary hepatocytes in vitro. Although short-term overexpression of PRMT1 had no significant effect on hepatic triglyceride levels under physiological conditions, it resulted in increased serum triglyceride and fasting blood glucose levels in normal C57BL/6J mice. More importantly, PRMT1 was observed to interact with and methylate CFLAR, ultimately leading to its ubiquitination-mediated protein degradation. This process subsequently triggered the activation of c-Jun N-terminal kinase 1 (JNK1) and lipid deposition in primary hepatocytes. Together, these results suggested that PRMT1-mediated methylation of CFLAR plays a critical role in hepatic lipid metabolism. Targeting PRMT1 for drug design may represent a promising strategy for the treatment of NAFLD.
摘要：
非酒精性脂肪性肝病（NAFLD）已成为全球慢性肝病的主要原因。半胱天冬酶8和FADD样细胞凋亡调节因子（CFLAR）已被确定为通过抑制细胞凋亡信号调节激酶1（ASK1）的N末端二聚化减轻非酒精性脂肪性肝炎（NASH）的有效因子。虽然精氨酸甲基转移酶1（PRMT1）先前报道与肝葡萄糖产生增加有关，它在肝脏脂质代谢中的参与仍未被探索。通过Co-IP和免疫印迹法验证了PRMT1和CFLAR之间的相互作用以及CFLAR的甲基化。产生重组腺病毒用于在肝细胞中过表达或敲低PRMT1。在正常和高脂饮食诱导的肥胖小鼠中研究了PRMT1在NAFLD中的作用。在这项研究中,我们发现在禁食48h后PRMT1的显著上调和CFLAR的下调,而后者在补食12h后明显反弹。在饲喂甲硫氨酸胆碱缺乏（MCD）饮食的小鼠的肝脏和用油酸（OA）/棕榈酸（PA）攻击的肝细胞中，PRMT1的表达增加。PRMT1的过表达不仅抑制脂肪酸氧化相关基因(FAO)的表达,而且促进脂肪酸合成相关基因(FAS)的表达,导致原代肝细胞中甘油三酯积累增加，而且还增强了原代肝细胞的糖异生。相反，抑制肝脏PRMT1显著减轻MCD饮食诱导的肝脏脂质代谢异常和体内肝损伤,可能是通过CFLAR蛋白水平的上调。敲除PRMT1抑制FAS相关基因的表达，增强FAO相关基因的表达，导致在体外OA/PA处理的原代肝细胞中甘油三酯积累减少。尽管在生理条件下，短期过表达PRMT1对肝脏甘油三酯水平没有显著影响，它导致正常C57BL/6J小鼠的血清甘油三酯和空腹血糖水平升高。更重要的是,观察到PRMT1与CFLAR相互作用并甲基化，最终导致其泛素化介导的蛋白质降解。该过程随后触发了c-JunN末端激酶1（JNK1）的激活和原代肝细胞中的脂质沉积。一起,这些结果表明PRMT1介导的CFLAR甲基化在肝脏脂质代谢中起关键作用。靶向PRMT1用于药物设计可能代表治疗NAFLD的有希望的策略。
