Glutathione and Redox Control in the Mitochondrial Intermembrane Space

线粒体膜间空间中的谷胱甘肽和氧化还原控制

基本信息

批准号：
8601188
负责人：
Caryn E Outten
金额：
$ 25.27万
依托单位：
UNIVERSITY OF SOUTH CAROLINA AT COLUMBIA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-03-01 至 2015-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8601188
关键词：
Aging Alcoholic Liver Diseases Anabolism Biochemical Biogenesis Biological Assay Biological Models Cell Respiration Cell physiology Cysteine Cytosol Data Defect Disease Disulfides Environment Equilibrium Fluorescence Functional disorder Glutathione Glutathione Disulfide Goals Homeostasis Knowledge Light Link Malignant Neoplasms Measurement Mediating Membrane Metabolism Methods Mitochondria Mitochondrial Diseases Mitochondrial Matrix Molecular Molecular Genetics Monitor Neurodegenerative Disorders Organelles Organism Oxidation-Reduction Oxidative Phosphorylation Oxidoreductase Pathway interactions Play Production Protein Import Proteins Publishing Reaction Reactive Oxygen Species Regulatory Pathway Research Role Saccharomyces cerevisiae Source Stress Sulfhydryl Compounds System Testing Thioredoxin Yeasts base disulfide bond glutaredoxin glutathione transporter human disease in vivo mitochondrial dysfunction mitochondrion intermembrane space mutant oxidation programs public health relevance receptor research study screening sensor sulfhydryl oxidase therapy development

Aging Alcoholic Liver Diseases Anabolism Biochemical Biogenesis Biological Assay Biological Models Cell Respiration Cell physiology Cysteine Cytosol Data Defect Disease Disulfides Environment Equilibrium Fluorescence Functional disorder Glutathione Glutathione Disulfide Goals Homeostasis Knowledge Light Link Malignant Neoplasms Measurement Mediating Membrane Metabolism Methods Mitochondria Mitochondrial Diseases Mitochondrial Matrix Molecular Molecular Genetics Monitor Neurodegenerative Disorders Organelles Organism Oxidation-Reduction Oxidative Phosphorylation Oxidoreductase Pathway interactions Play Production Protein Import Proteins Publishing Reaction Reactive Oxygen Species Regulatory Pathway Research Role Saccharomyces cerevisiae Source Stress Sulfhydryl Compounds System Testing Thioredoxin Yeasts base disulfide bond glutaredoxin glutathione transporter human disease in vivo mitochondrial dysfunction mitochondrion intermembrane space mutant oxidation programs public health relevance receptor research study screening sensor sulfhydryl oxidase therapy development

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项目摘要

DESCRIPTION (provided by applicant): Intracellular thiol-disulfide balance is critical for the activity of proteins with functionally important cysteine residues. The tripeptide glutathione (GSH) and oxidoreductases like glutaredoxins (GRXs) and thioredoxins (TRX) help maintain thiol-disulfide balance by catalyzing disulfide exchange reactions and protecting cysteines residues from oxidation by reactive oxygen species (ROS). Mitochondrial GSH metabolism is a key component of cellular thiol redox homeostasis since this organelle is the main source and target of ROS produced from aerobic metabolism. Essential mitochondrial functions such as oxidative phosphorylation, protein import, and Fe-S cluster biogenesis are directly dependent on thiol-disulfide balance. Consequently, disruption of mitochondrial thiol-disulfide balance has been linked to cancer, neurodegenerative diseases, and aging. The long-term objective of this research program is to characterize mitochondrial thiol redox homeostasis using the yeast Saccharomyces cerevisiae as a model system. The mitochondrion is divided by a double membrane into two distinct compartments: the matrix and the intermembrane space (IMS). Redox regulatory systems that govern thiol-disulfide balance in the matrix are well-characterized. However, the mechanisms for thiol-disulfide redox control in the IMS and the role of GSH metabolism in IMS thiol redox pathways represent key gaps in our knowledge of mitochondrial redox systems. The goal of this proposal is to determine the mechanisms for maintaining thiol-disulfide balance in the IMS. GFP-based redox sensors that are targeted to the IMS and matrix have been developed to allow localized thiol redox monitoring via in vivo fluorescence measurements. These sensors equilibrate with the local reduced/oxidized glutathione (GSH:GSSG) pool and register thiol redox changes via disulfide bond formation. To determine how GSH metabolism influences the IMS redox environment, GSH:GSSG exchange between the IMS, matrix, and cytosol under redox stress will be characterized using these sensors and mitochondrial GSH transporters will be identified (Aim 1). Furthermore, the connection between GSH metabolism and thiol- dependent protein import into the IMS will be deciphered by determining how alterations in GSH:GSSG or the IMS protein import machinery influence each other (Aim 2). Finally, additional thiol redox systems/pathways that help maintain IMS thiol redox balance will be identified (Aim 3). By using a molecular genetics approach in a highly tractable organism, these studies will shed new light on thiol redox control pathways in the IMS, with strong implications for disorders involving mitochondrial dysfunction.

描述（由申请人提供）：细胞内硫醇 - 硫化物平衡对于具有功能重要的半胱氨酸残基的蛋白质活性至关重要。三肽谷胱甘肽（GSH）和氧化还原酶（如谷蛋白（GRXS）和硫氧还蛋白（TRX）（TRX），通过催化二硫化物交换反应并保护囊肿的残基来帮助维持硫醇 - 二硫化物的平衡，并通过反应性氧气（ROS）保护囊肿残基。线粒体GSH代谢是细胞硫代氧化还原稳态的关键组成部分，因为该细胞器是有氧代谢产生的ROS的主要来源和靶标。必需的线粒体功能，例如氧化磷酸化，蛋白质进口和Fe-S簇生物发生直接取决于硫醇 - 硫化物平衡。因此，线粒体硫醇 - 硫化物平衡的破坏与癌症，神经退行性疾病和衰老有关。该研究计划的长期目标是使用酵母糖酵母作为模型系统来表征线粒体硫醇氧化还原稳态。线粒体除以双膜分为两个不同的隔室：矩阵和膜间空间（IMS）。在基质中控制硫醇 - 二硫化物平衡的氧化还原调节系统经过很好的特征。但是，IMS中硫代二硫化物氧化还原控制的机制以及GSH代谢在IMS硫代氧化还原途径中的作用代表了我们对线粒体氧化还原系统的了解中的关键差距。该建议的目的是确定维持IMS中硫醇二硫化物平衡的机制。已经开发了针对IMS和基质的基于GFP的氧化还原传感器，以允许通过体内荧光测量值局部的硫醇氧化还原监测。这些传感器与局部还原/氧化的谷胱甘肽（GSH：GSSG）库平衡，并通过二硫键键形成登记硫代氧化还原变化。为了确定GSH代谢如何影响IMS，IMS，基质和细胞质之间的GSH：GSSG在氧化还原胁迫下的交换，将使用这些传感器和线粒体GSH转运蛋白进行表征（AIM 1）。此外，通过确定GSH：GSH：GSSG或IMS蛋白蛋白导入机械的变化如何相互影响的GSH代谢与硫醇依赖性蛋白之间的联系将被解密（AIM 2）。最后，将确定有助于维持IMS硫代氧化还原平衡的其他硫醇氧化还原系统/途径（AIM 3）。通过在高度易于处理的生物中使用分子遗传学方法，这些研究将对IMS中的硫代氧化还原控制途径进行新的启示，对涉及线粒体功能障碍的疾病产生了强烈的影响。