Methionine Adenosyltransferase Alpha1 in Alcoholic Liver Disease

酒精性肝病中的蛋氨酸腺苷转移酶 Alpha1

• 批准号: 9902269
• 负责人: Shelly Chi-Loo Lu
• 金额: $ 39.38万
• 依托单位: CEDARS-SINAI MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-15 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9902269
• 关键词: Affect; Alcoholic Hepatitis; Alcoholic Liver Diseases; Anabolism; Antioxidants; Biological; Biology; CYP2E1 gene; Catalytic Domain; Cell Nucleus; Cell physiology; Co-Immunoprecipitations; Competence; Cytosol; Data; Development; Enzymes; Fibrosis; Gene Expression; Genes; Glutathione; Goals; Health; Heat shock proteins; Heat-Shock Proteins 70; Heat-Shock Proteins 90; Hepatic; Hepatocyte; Hepatotoxicity; Human; Immunoprecipitation; Impairment; In Vitro; Injury; Isoenzymes; Knock-out; Knockout Mice; Liver; Mammals; Mass Spectrum Analysis; Mediating; Methylation; Mitochondria; Mitochondrial Matrix; Mitochondrial Proteins; Molecular; Molecular Chaperones; Mus; Normal Cell; Pathogenesis; Patients; Peptidylprolyl Isomerase; Play; Process; Proteins; Recombinants; Regulation; Reporting; Role; S-Adenosylmethionine; Source; Testing; Work; cell type; chronic liver disease; dimer; druggable target; follow-up; improved; in vivo; methionine adenosyltransferase; mitochondrial dysfunction; new therapeutic target; novel; prevent; protein expression; protein folding; transcription factor

ABSTRACT Methionine adenosyltransferase (MAT) is an essential cellular enzyme that catalyzes the formation of S- adenosylmethionine (SAMe), the principal biological methyl donor and in liver, precursor of the key antioxidant glutathione (GSH). In mammals, two different genes, MAT1A and MAT2A, encode for two homologous MAT catalytic subunits, α1 and α2, respectively. MAT1A is primarily expressed in normal liver. Majority of patients with chronic liver disease have decreased expression and activity of MAT1A-encoded isoenzymes. We found MAT1A expression and hepatic SAMe levels are reduced in alcoholic hepatitis patients. While MAT isoenzymes are widely acknowledged for catalyzing cytosolic SAMe biosynthesis, our recent works have uncovered highly novel aspects of their functions. In addition to cytosol and nucleus, our preliminary data indicate MATα1 is also present in the mitochondrial matrix to regulate mitochondrial function. This is important as we found hepatocytes lack the mitochondrial SAMe transporter SLC25A26. Using immunoprecipitation (IP) followed by mass spectrometry, we have identified many mitochondrial proteins and cytochrome P450 2E1 (CYP2E1) as MATα1-interacting proteins. Our preliminary data show that MATα1 negatively regulates CYP2E1 expression mainly at the protein level via methylation, which has not been reported. Importantly, MATα1 mitochondrial targeting is impaired in murine and human alcoholic liver disease (ALD) and we propose two novel mechanisms that may cooperate in causing this impairment. The current proposal tests the central hypothesis that MATα1 provides the SAMe source within the hepatocyte's mitochondrial matrix and impairment in MATα1 mitochondrial targeting in ALD plays a key role in the pathogenesis of ALD. The corollary hypothesis is that MATα1 maintains hepatocyte mitochondrial function in part by suppressing CYP2E1 expression. Here we follow up these novel findings in three specific aims: 1) examine how MATα1 targets the mitochondria and why its targeting is impaired in ALD. We will elucidate how MATα1 targets the mitochondria and mechanisms of its impaired targeting in ALD. We will test the novel hypothesis that this is due to 1) increased MATα1 sumoylation and 2) increased interaction of MATα1 with PIN1 (a peptidyl-prolyl cis- trans isomerase that recognizes a specific phosphorylated motif), 2) examine how MATα1 regulates CYP2E1 protein expression. We will examine how MATα1 regulates CYP2E1 protein stability at the molecular level and identify CYP2E1 residues and interacting proteins that participate in this process, 3) examine the role of mitochondrial MATα1 and MATα1-regulated CYP2E1 in ALD. We will test the novel hypothesis that preventing MATα1 sumoylation or interaction with PIN1 will protect against reduced mitochondrial MATα1 content and injury in ALD. We will also examine whether MATα1 will protect against ALD by methylating CYP2E1 at R379 to enhance its degradation. If successfully accomplished, these studies will change existing paradigms on MATα1 biology, CYP2E1 regulation and may uncover druggable targets for ALD.

抽象的 蛋氨酸腺苷转移酶 (MAT) 是一种重要的细胞酶，可催化 S- 的形成 腺苷甲硫氨酸 (SAMe)，主要的生物甲基供体，在肝脏中是关键抗氧化剂的前体 谷胱甘肽（GSH）。在哺乳动物中，两个不同的基因 MAT1A 和 MAT2A 编码两个同源 MAT 催化亚基分别为α1和α2。 MAT1A 主要在正常肝脏中表达。大多数患者 患有慢性肝病的人 MAT1A 编码的同工酶的表达和活性降低。我们发现 酒精性肝炎患者中 MAT1A 表达和肝脏 SAMe 水平降低。而垫 同工酶被广泛认为可以催化胞质 SAMe 生物合成，我们最近的工作有 揭示了它们功能的高度新颖的方面。除了细胞质和细胞核外，我们的初步数据 表明 MATα1 也存在于线粒体基质中以调节线粒体功能。这很重要 我们发现肝细胞缺乏线粒体 SAMe 转运蛋白 SLC25A26。使用免疫沉淀 (IP) 随后通过质谱分析，我们鉴定出了许多线粒体蛋白和细胞色素 P450 2E1 (CYP2E1) 作为 MATα1 相互作用蛋白。我们的初步数据表明 MATα1 负向调节 CYP2E1主要通过甲基化在蛋白水平表达，目前尚未见报道。重要的是， MATα1 线粒体靶向在小鼠和人类酒精性肝病 (ALD) 中受损，我们建议 两种新机制可能共同导致这种损害。目前的提案是对中央的考验 假设 MATα1 在肝细胞线粒体基质内提供 SAMe 来源，并且 ALD 中 MATα1 线粒体靶向受损在 ALD 的发病机制中起着关键作用。这 推论假设是 MATα1 部分通过抑制肝细胞线粒体功能来维持肝细胞线粒体功能 CYP2E1 表达。在这里，我们以三个具体目标跟进这些新发现：1）研究 MATα1 如何 靶向线粒体以及为什么其靶向在 ALD 中受损。我们将阐明 MATα1 如何靶向 线粒体及其在 ALD 中靶向受损的机制。我们将测试这个新假设 由于 1) MATα1 sumoylation 增加，以及 2) MATα1 与 PIN1（一种肽基-脯氨酰顺式- 识别特定磷酸化基序的反式异构酶），2）检查 MATα1 如何调节 CYP2E1 蛋白质表达。我们将研究 MATα1 如何在分子水平上调节 CYP2E1 蛋白稳定性 并识别参与此过程的 CYP2E1 残基和相互作用蛋白，3) 检查 ALD 中线粒体 MATα1 和 MATα1 调节的 CYP2E1。我们将测试这个新假设 防止 MATα1 sumoylation 或与 PIN1 相互作用将防止线粒体 MATα1 减少 ALD 中的含量和损伤。我们还将研究 MATα1 是否会通过甲基化来预防 ALD R379 处的 CYP2E1 增强其降解。如果成功完成，这些研究将改变现有的 MATα1 生物学、CYP2E1 调控的范例，并可能发现 ALD 的药物靶标。