The Role of Gm4951 in Nonalcoholic Fatty Liver Disease

项目摘要 非酒精性脂肪性肝病（NAFLD）正在成为一个全球性的人类健康问题。我们的长期目标是 了解NAFLD的分子机制，并将这些知识转化为新的治疗策略。 由于人类和小鼠之间的生理相似性，以及小鼠发展疾病的倾向， 当喂食高脂肪饮食（HFD）时，小鼠密切模仿NAFLD，为我们提供了基本的见解， NAFLD发病机制。在人类和小鼠中，遗传变异影响脂肪肝的发生率和严重程度 在特定的环境条件下。为了确定影响这一过程的基因，我们采用了无偏 前向遗传筛选和高度自动化的减数分裂图谱，以确定导致小鼠NAFLD的突变 由HFD致敏。预测基因4951（Gm 4951）的两个半显性错义等位基因，命名为Oily和 石炭纪，在这个屏幕上被检测到。与大多数NAFLD突变体不同， 肥胖，我们的初步数据显示，Gm 4951缺陷小鼠的肝脏脂质显著增加， 在HFD时，没有伴随的体重增加。GM 4951在大肠杆菌中高表达， 肝细胞Gm 4951基因敲除增加脂质含量，Gm 4951基因过表达降低脂质含量 体外原代肝细胞的脂质含量，表明肝细胞对脂质含量的内在调节。Gm4951 敲除的肝脏显示脂质氧化基因的表达降低。的质谱分析 内源性GM 4951相互作用蛋白与脂滴蛋白羟基类固醇17β- 脱氢酶13（HSD 17 B13）和脂质氧化酶。此外，Gm 4951基因在大肠杆菌中的转录水平也高于在大肠杆菌中的转录水平。 肝细胞被γ-干扰素（IFN-γ）激活，这有效地降低了脂质含量，就像当 GM 4951过表达。这些结果导致了我们的中心假设，即GM 4951对于促进 脂质氧化，并限制肝脏脂质蓄积。为了验证这一假设，我们提出了三个具体的 目标。目的1进一步研究GM 4951基因缺陷小鼠NAFLD的发生发展。目标2将决定 GM 4951在调节脂质代谢中的确切机制作用。目的3将研究转录 Gm 4951的调节，包括肝脏特异性表达模式和IFN-γ的诱导表达。 了解如何激活GM 4951以及GM 4951的人类同源物将提供方法， 预防或治疗NAFLD。目标3也将研究这些问题。GM 4951引起的NAFLD表型 这种缺乏从根本上不同于经典的肥胖相关的NAFLD小鼠模型。GM 4951是 在肝脏中特异性表达，并在那里起作用以限制脂质积累。因此，找到激活 GM 4951将提供一种降低肝脏脂质含量的新方法。完成拟议工作将 提出了新的治疗靶点来对抗NAFLD。

PROJECT SUMMARY Nonalcoholic fatty liver disease (NAFLD) is becoming a global human health problem. Our long-term goal is to understand molecular mechanisms of NAFLD, and to translate this knowledge into novel therapeutic strategies. Due to the physiologic similarities between humans and mice, and the propensity of mice to develop a disease closely mimicking NAFLD when fed a high fat diet (HFD), mice have provided us with fundamental insights into NAFLD pathogenesis. In humans and mice, genetic variation influences the rate and severity of hepatosteatosis under a given set of environmental conditions. To identify genes that influence the process, we utilized unbiased forward genetic screening and highly automated meiotic mapping to identify mutations that cause NAFLD in mice sensitized by a HFD. Two semi-dominant missense alleles of predicted gene 4951 (Gm4951), named Oily and Carboniferous, were detected in this screen. As distinct from most NAFLD mutants that are associated with obesity, our preliminary data showed that Gm4951 deficient mice had dramatically increased hepatic lipid accumulation without a concomitant increase of body weight on a HFD. Gm4951 was highly expressed in hepatocytes. Knockout of Gm4951 increased lipid content and overexpression of Gm4951 decreased lipid content of primary hepatocytes in vitro, suggesting hepatocyte-intrinsic regulation of lipid content. Gm4951 knockout livers showed decreased expression of lipid oxidation genes. Mass spectrometry analysis of endogenous GM4951 interacting proteins revealed interaction with lipid droplet protein Hydroxysteroid 17β- dehydrogenase 13 (HSD17B13) and lipid oxidation enzymes. Moreover, the transcription of Gm4951 in hepatocytes was activated by interferon gamma (IFN-γ), which effectively decreased lipid content, much as when GM4951 was overexpressed. These results led to our central hypothesis that GM4951 is critical for promoting lipid oxidation, and limits hepatic lipid accumulation. To test this hypothesis, we propose to pursue three Specific Aims. Aim 1 will further investigate the development of NAFLD in GM4951 deficient mice. Aim 2 will determine the precise mechanistic role of GM4951 in regulating lipid metabolism. Aim 3 will study the transcriptional regulation of Gm4951, including the liver-specific expression pattern and the inducible expression by IFN-γ. Understanding how to activate GM4951 and what’s the human homolog of GM4951 would offer approaches to preventing or treating NAFLD. These will be studied in Aim 3 as well. The NAFLD phenotype caused by GM4951 deficiency is fundamentally distinct from the classic obesity-associated NAFLD mouse models. GM4951 is specifically expressed in the liver and operates there to limit lipid accumulation. Thus, finding ways to activate GM4951 will provide a new means of reducing hepatic lipid content. Completion of the proposed work will suggest new therapeutic targets to combat NAFLD.