Mitochondrial metabolism and the Lon-PDH axis

线粒体代谢和 Lon-PDH 轴

• 批准号: 10379257
• 负责人: CAROLYN K SUZUKI
• 金额: $ 32.17万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10379257
• 关键词: Acetyl Coenzyme A; Acetylation; Animal Model; Architecture; Astrocytes; Atrophic; Brain; Carbohydrates; Catabolism; Cells; Cellular Stress; Cerebrum; Citric Acid Cycle; Complex; Data; Defect; Dietary Intervention; Disease; Energy Metabolism; Energy-Generating Resources; Enzymes; Erythrocytes; Failure; Fatty Acids; Fibroblasts; Functional disorder; Gatekeeping; Generations; Genes; Glucose; Glutamine; Glycolysis; Glycolysis Inhibition; Goals; Heart Diseases; Human; Impairment; Investigation; Ketone Bodies; Knowledge; Leucine; Link; Malignant Neoplasms; Measures; Mediating; Medium chain fatty acid; Metabolic; Metabolism; Missense Mutation; Mitochondria; Modification; Molecular; Molecular Machines; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurologic; Neurologic Dysfunctions; Neurons; Outcome; Oxidative Phosphorylation; Oxides; PDH kinase; Parents; Pathogenicity; Pathway interactions; Patients; Pharmacology; Phosphorylation; Positioning Attribute; Post-Translational Protein Processing; Production; Proline; Proteins; Proteolysis; Proteomics; Pyruvate; Pyruvate Dehydrogenase (Lipoamide)-Phosphatase; Pyruvate Dehydrogenase Complex; Pyruvate Dehydrogenase E1; Reaction; Regulation; Siblings; Skin; Stream; Testing; Transcript; Variant; Work; amino acid metabolism; brain cell; dihydrolipoamide dehydrogenase; dihydrolipoyllysine-residue acetyltransferase; endopeptidase La; experimental study; fatty acid oxidation; high throughput screening; induced pluripotent stem cell; intermolecular interaction; metabolomics; mitochondrial metabolism; mutant; novel; oxidation; proteostasis; pyruvate dehydrogenase; response; stem cell differentiation; therapeutic protein; uptake

The human Lon protease is a master regulator of mitochondrial proteostasis, which is essential for regulating mitochondrial energy metabolism and mitigating cell stress. We recently identified a novel pathogenic variant in the LONP1 gene encoding Lon, in two siblings with profound neurologic impairment, cerebral and cerebellar atrophy, in which proline at position 761 was replaced by leucine (Lon-P761L). Primary skin fibroblasts from these siblings, showed that the activity of pyruvate dehydrogenase (PDH) was substantially reduced. PDH deficiency was caused by the failure of Lon-P761L to degrade the phosphorylated E1a subunit of PDH, which accumulates and inhibits PDH activity. PDH is the central gatekeeper linking glycolysis to the tricarboxylic acid (TCA) cycle, and is also a key regulatory node for glucose and fatty acid catabolism. Our long term goal is to elucidate why homozygous Lon-P761L expression causes severe neurologic dysfunction and neurodegeneration. Glucose is the brain’s principal source of energy. Neurons generate ATP almost exclusively by glucose oxidization, thus fully functional PDH activity is crucial. Astrocytes by contrast, have broader metabolic capacity and supply neurons with lactate, glutamine and ketone bodies, which are used to form acetyl CoA and TCA cycle intermediates required for glucose oxidation. We hypothesize that wild type Lon regulates the architecture and activities of the PDH complex, and modulates upstream and downstream effectors, to calibrate mitochondrial metabolism and energetics. In this project, we will employ patient-and parent-derived fibroblasts, and also fibroblasts that have been reprogrammed to generate induced pluripotent stem cells (iPSCs). These iPSCs will be differentiated into neurons and astrocytes. Using the patient- and parent- derived fibroblasts, Aim 1 will test the hypothesis that Lon-mediated degradation regulates the architecture and activity of the PDH complex. Aim 2 will identify the up- and down-stream modulators of the Lon-PDH axis, which are altered in cells expressing wild type Lon versus Lon-P761L. In Aim 3, we will investigate the regulation of PDH by Lon in iPSCs differentiated into neurons and astrocytes. Our investigation will establish new molecular mechanisms for the Lon-dependent regulation of PDH. The knowledge gained will also help to identify potential therapeutic protein targets (e.g. PDK, PDP, Lon), pharmacologic and dietary interventions for increasing PDH activity and/or for treating PDH deficiency associated with Lon dysfunction. These outcomes have a broader impact for understanding how PDH activity and mitochondrial metabolism can be calibrated in both rare and more common disorders such as heart disease, cancer and neurodegeneration.

人类 Lon 蛋白酶是线粒体蛋白质稳态的主要调节因子，对于维持线粒体功能至关重要。 调节线粒体能量代谢并减轻细胞应激。我们最近发现了一本小说 两个患有严重神经系统疾病的兄弟姐妹中编码 Lon 的 LONP1 基因的致病性变异 损伤、大脑和小脑萎缩，其中 761 位脯氨酸被亮氨酸取代 （Lon-P761L）。来自这些兄弟姐妹的原代皮肤成纤维细胞表明丙酮酸的活性 脱氢酶（PDH）显着降低。 PDH缺乏症是由于Lon-P761L未能降解PDH的磷酸化E1a亚基而引起的，该亚基会积累并抑制PDH 活动。 PDH 是连接糖酵解与三羧酸 (TCA) 循环的中央看门人，并且是 也是葡萄糖和脂肪酸分解代谢的关键调节节点。我们的长期目标是阐明原因 纯合 Lon-P761L 表达会导致严重的神经功能障碍和神经变性。 葡萄糖是大脑的主要能量来源。神经元几乎完全通过葡萄糖产生 ATP 氧化，因此功能齐全的 PDH 活性至关重要。相比之下，星形胶质细胞具有更广泛的代谢 能力并为神经元提供乳酸、谷氨酰胺和酮体，这些物质用于形成乙酰基 葡萄糖氧化所需的 CoA 和 TCA 循环中间体。我们假设野生型 Lon 调节 PDH 复合物的结构和活动，并调节上游和下游 效应器，校准线粒体代谢和能量。在这个项目中，我们将使用患者和父母来源的成纤维细胞，以及经过重新编程以产生诱导性成纤维细胞的成纤维细胞。 多能干细胞（iPSC）。这些 iPSC 将分化为神经元和星形胶质细胞。使用 患者和亲本来源的成纤维细胞，目标 1 将检验 Lon 介导的降解的假设 调节 PDH 复合物的结构和活性。目标 2 将确定上游和下游 Lon-PDH 轴的调节剂，与表达野生型 Lon 的细胞相比，表达 Lon-P761L 的细胞中的调节剂发生改变。 在目标 3 中，我们将研究分化为神经元和神经元的 iPSC 中 Lon 对 PDH 的调节。 星形胶质细胞。我们的研究将为 Lon 依赖性建立新的分子机制 PDH 的调节。获得的知识也将有助于识别潜在的治疗蛋白靶点 （例如 PDK、PDP、Lon）、药物和饮食干预以增加 PDH 活性和/或 治疗与 Lon 功能障碍相关的 PDH 缺乏。这些结果对以下方面产生了更广泛的影响： 了解如何在罕见和更多情况下校准 PDH 活性和线粒体代谢 常见疾病，如心脏病、癌症和神经退行性疾病。