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: 10376299
• 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 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10376299
• 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; Sumoylation Pathway; 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.

摘要 甲硫氨酸腺苷转移酶（MAT）是一种重要的细胞酶，催化S-腺苷酸的形成。 腺苷甲硫氨酸（SAMe），主要的生物甲基供体，在肝脏中，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，MAT 1A和MAT 2A，编码两个同源MAT 催化亚基，分别为α1和α2。MAT 1A主要在正常肝脏中表达。大多数患者 慢性肝病患者的MAT 1A编码同工酶的表达和活性降低。我们发现 酒精性肝炎患者的MAT 1A表达和肝脏SAMe水平降低。当MAT 同工酶被广泛认为是催化细胞溶质SAMe生物合成的酶，我们最近的工作 发现了其功能的一些非常新颖的方面。除了细胞质和细胞核，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 因为我们发现肝细胞缺乏线粒体SAMe转运蛋白SLC 25 A26。使用免疫沉淀（IP） 通过质谱分析，我们鉴定了许多线粒体蛋白和细胞色素P450 2 E1 （CYP 2 E1）作为MATα1相互作用蛋白。我们的初步数据表明MATα1负调节 CYP 2 E1主要通过甲基化在蛋白质水平表达，目前尚无报道。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，我们提出 这两种新的机制可能共同导致这种损害。目前的提案考验着中央 假设MATα1在肝细胞线粒体基质内提供SAMe来源， MATα1线粒体靶向功能障碍在ALD发病机制中起关键作用。的 一个推论是MATα1部分通过抑制肝细胞线粒体功能， CYP 2 E1表达。在这里，我们按照这些新的发现，在三个具体的目标：1）检查如何MATα1 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中受损的靶向机制。我们将测试新的假设，这是 由于1）MATα1类小泛素化增加和2）MATα1与PIN 1（肽基脯氨酰顺式- 识别特定磷酸化基序的反式异构酶），2）检测MATα1如何调节CYP 2 E1 蛋白质表达我们将研究MATα1如何在分子水平上调节CYP 2 E1蛋白的稳定性 并确定CYP 2 E1残基和参与这一过程的相互作用蛋白质，3）检查 线粒体MATα1和MATα1调节的CYP 2 E1。我们将检验一个新的假设， 阻止MATα1类小泛素化或与PIN 1的相互作用将防止线粒体MATα1减少 含量和损伤。我们还将研究MATα1是否能通过甲基化 CYP 2 E1在R379上的作用，以增强其降解。如果成功完成，这些研究将改变现有的 MATα1生物学、CYP 2 E1调节的范例，并可能揭示ALD的可药用靶点。