Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease characterized by lipid accumulation and endoplasmic reticulum (ER) stress, while effective therapies targeting the specific characteristics of NAFLD are limited. Ufmylation is a newly found post-translational modification process that involves the attachment of the Ubiquitin-fold modifier 1 (UFM1) protein to its substrates via ufmylation modification system. Ufmylation regulates ER stress via modifying UFM1 binding protein 1 (UFBP1), suggesting a potential role for ufmylation in NAFLD pathogenesis. However, the precise role of ufmylation in NAFLD remains unclear. Herein, we aim to elucidate the impact of ufmylation on UFBP1 in NAFLD and explore the underlying mechanisms involved. We observed increased expression of UFM1-conjugated proteins and ufmylation modification system components in livers with steatosis derived from NAFLD patients and NAFLD models. Upregulation of ufmylation on hepatic proteins appeared to be an adaptive response to hepatic ER stress in NAFLD. In vitro, knocking down UFBP1 resulted in increased lipid accumulation and lipogenesis in hepatocytes treated with free fatty acids (FFA), which could be rescued by wild-type UFBP1 (WT UFBP1) but not by a mutant form of UFBP1 lacking the main ufmylation site lys267 (UFBP1 K267R). In vivo, ufmylation on UFBP1 ameliorated obesity, hepatic steatosis, hepatic lipogenesis, dyslipidemia, insulin resistance and liver damage in mice with NAFLD induced by a high fat diet (HFD). We also demonstrated that the downregulation of UFBP1 induced ER stress, whereas the reintroduction or overexpression of UFBP1 alleviated ER stress in a manner dependent on ufmylation in NAFLD. This mechanism could be responsible for the amelioration of aberrant hepatic lipogenesis and insulin resistance in NAFLD. Our data reveal a protective role of ufmylation on UFBP1 against NAFLD and offer a specific target for NAFLD treatment.

非酒精性脂肪肝病（NAFLD）是最常见的肝脏疾病，其特征是脂质堆积和内质网（ER）应激，而针对NAFLD特定特征的有效治疗方法有限。Ufmylation 是一种新发现的翻译后修饰过程，涉及通过 ufmylation 修饰系统将泛素折叠修饰剂 1 (UFM1) 蛋白附着到其底物上。Ufmylation 通过修饰 UFM1 结合蛋白 1 (UFBP1) 调节 ER 应激，这表明 ufmylation 在 NAFLD 发病机制中具有潜在作用。然而，ufmylation 在 NAFLD 中的确切作用仍不清楚。在此，我们的目的是阐明 ufmylation 对 NAFLD 中 UFBP1 的影响，并探讨相关的潜在机制。我们观察到来自 NAFLD 患者和 NAFLD 模型的脂肪变性肝脏中 UFM1 缀合蛋白和 ufmylation 修饰系统成分的表达增加。肝蛋白上 ufmylation 的上调似乎是 NAFLD 中肝脏 ER 应激的适应性反应。在体外，敲低 UFBP1 导致用游离脂肪酸 (FFA) 处理的肝细胞中脂质积累和脂肪生成增加，这可以通过野生型 UFBP1 (WT UFBP1) 来挽救，但不能通过缺乏主要 ufmylation 位点的 UFBP1 突变体来挽救lys267 (UFBP1 K267R)。在体内，UFBP1 上的 ufmylation 可以改善高脂饮食 (HFD) 引起的 NAFLD 小鼠的肥胖、肝脂肪变性、肝脂肪生成、血脂异常、胰岛素抵抗和肝损伤。我们还证明，UFBP1 的下调会诱导内质网应激，而 UFBP1 的重新引入或过度表达则以依赖于 NAFLD 中 ufmylation 的方式减轻内质网应激。这种机制可能是改善 NAFLD 中异常的肝脏脂肪生成和胰岛素抵抗的原因。我们的数据揭示了 UFBP1 上的 ufmylation 对 NAFLD 的保护作用，并为 NAFLD 治疗提供了具体靶点。