Regulation of ER homeostasis by TCA cycle activity: mechanisms and consequences

TCA 循环活动调节 ER 稳态：机制和后果

基本信息

批准号：
10442767
负责人：
David Thomas Rutkowski
金额：
$ 31.83万
依托单位：
UNIVERSITY OF IOWA
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10442767
关键词：
Acetyl Coenzyme A Adipocytes Adult Affect Amino Acids Atherosclerosis Biosensor Catabolism Cells Citric Acid Cycle Client Communication Coupled Cytosol Data Diabetes Mellitus Disease Electron Transport Endoplasmic Reticulum Enzymes Genetic Glucose Glutathione Glutathione Disulfide Glutathione Reductase Glycolysis Goals Hepatocyte Homeostasis Human Isocitrate Dehydrogenase Knowledge Lead Link Lipids Maintenance Mediating Metabolic Metabolic Diseases Metabolism Mitochondria Modeling Molecular Biology Molecular Chaperones Monitor Muscle Cells NADP Nutrient Nutritional Obesity Outcome Oxidation-Reduction Pathway interactions Pharmacology Process Production Proteins Pyruvate Quality Control Regulation Steatohepatitis Stress Sulfhydryl Compounds Testing Time Work base building materials burden of illness cell type cofactor comorbidity design endoplasmic reticulum stress fatty acid oxidation genetic manipulation improved insight knock-down lipid biosynthesis malic enzyme novel oxidation prevent protein folding pyruvate carrier response tool

项目摘要

Project Summary Disruption of protein folding in the endoplasmic reticulum—“ER stress”—is associated with many different metabolic diseases, particularly those associated with obesity that affect between 22 and 30 percent of adults in the U.S. Because of this exceptional disease burden, it is important to understand the factors that cause ER stress during metabolic dysregulation. Yet the pathways by which metabolic activity and ER homeostasis are coupled are poorly understood. Mitochondria are central to metabolism, and the TCA cycle is the hub of this activity, accepting substrates from glycolysis and fatty acid oxidation for catabolism, generating reducing equivalents for electron transport and for the maintenance of cellular redox homeostasis, and providing building materials for the reductive biosynthesis of lipids, glucose, and amino acids. Because of its centrality to so many processes, flux through the TCA cycle is likely to affect many diverse cellular pathways, even those with no obvious direct connection. This includes ER protein processing, which is sensitive to changes in redox state, amino acid availability, and cellular lipid content. In this proposal, we provide evidence for a previously unknown functional relationship between TCA cycle activity and ER homeostasis in metabolically active cells, including hepatocytes, myocytes, and adipocytes, that depends on production of NADPH by the TCA cycle and redox regulation of glutathione. This proposal is designed to identify the basic mechanisms linking TCA-dependent NADPH production in the mitochondria to homeostasis in the ER. Toward that end, we propose three specific aims: (1) Determine how NADPH production and compartmentalization link nutrient flow to ER stress; (2) Determine how changes to mitochondrial and cytosolic glutathione redox promote ER oxidation; and (3) Determine how TCA activity and glutathione redox alter ER function. We will achieve these aims using a combination of genetic and pharmacological tools to manipulate TCA cycle activity; cutting-edge biosensors to monitor changes in cellular redox status; manipulation and analysis of ER-mitochondrial contacts; and molecular biology approaches to manipulate and assess ER functionality. The outcome of this work will be a mechanistic understanding of how metabolic activity alters ER function to contribute to disease.

项目概要内质网中蛋白质折叠的破坏（“ER 应激”）与许多不同的因素有关。代谢疾病，特别是与肥胖相关的疾病，影响 22% 至 30% 的成年人在美国，由于这种特殊的疾病负担，了解导致 ER 的因素非常重要代谢失调期间的压力然而代谢活动和内质网稳态的途径。耦合的理解甚少。线粒体是新陈代谢的核心，TCA 循环是该活动的中心，接受底物来自糖酵解和脂肪酸氧化的分解代谢，产生电子传输的还原当量维持细胞氧化还原稳态，并为还原提供建筑材料脂质、葡萄糖和氨基酸的生物合成由于其在许多过程中的中心地位，因此不断变化。 TCA 循环可能会影响许多不同的细胞途径，甚至是那些没有明显直接联系的途径。这包括 ER 蛋白质加工，它对氧化还原状态、氨基酸可用性和细胞脂质含量。在本提案中，我们为 TCA 循环之间以前未知的功能关系提供了证据代谢活跃细胞（包括肝细胞、肌细胞和脂肪细胞）的活性和 ER 稳态，这取决于 TCA 循环产生的 NADPH 和谷胱甘肽的氧化还原调节。旨在确定线粒体中 TCA 依赖性 NADPH 产生与为此，我们提出了三个具体目标：（1）确定 NADPH 如何实现生产和区划将养分流与内质网应激联系起来；(2) 确定如何变化线粒体和细胞质谷胱甘肽氧化还原促进 ER 氧化；以及 (3) 确定 TCA 活性和谷胱甘肽氧化还原改变内质网功能，我们将结合遗传和基因来实现这些目标。操纵 TCA 循环活性的药理学工具；监测细胞变化的尖端生物传感器；氧化还原状态；内质网-线粒体接触的操作和分析；以及分子生物学方法这项工作的结果将是对如何操作和评估 ER 功能的机械理解。代谢活动改变内质网功能从而导致疾病。