Mitochondrial Response to Oxidative Stress

描述(由申请人提供) 线粒体在细胞生理和对环境胁迫的反应中起着至关重要的作用。许多细胞毒素，包括鱼藤酮、1-甲基-4-苯基吡啶(MPP+)和百草枯，会损害线粒体的电子传递，产生ROS。活性氧物种在砷、淀粉样蛋白和神经酰胺的毒性中也是重要的。线粒体一直被视为ROS的潜在来源，可能导致帕金森氏症、衰老和其他病理状况。由于线粒体只包含一个16.5kb的mtDNA小基因组，只编码13种蛋白质，细胞器依赖于细胞核来获得大多数基因产物，包括DNA复制、表达和修复所需的所有因子。我们实验室和其他实验室最近的研究表明，在线粒体和其他细胞隔间发挥功能的蛋白质集合越来越多。其中许多蛋白质在线粒体DNA氧化损伤的修复中发挥作用。线粒体病理学的一个重要后果是线粒体DNA突变的产生，其中许多突变具有组织特异性，最常见的发生在复制后组织，如神经和肌肉。研究人员建议测试这一假设，即分化细胞中的线粒体可能包含不同于主动分裂细胞的蛋白质补充，这可能使复制后细胞易于发生更高比例的mtDNA突变，或者可能改变细胞进入凋亡的能力。为了实现这一目标，他们将研究氧化应激对活跃分裂的胚胎癌细胞和已被诱导沿神经元途径分化的细胞中线粒体蛋白质组的影响。二维凝胶方法和定量同位素编码亲和标签(ICAT)方法将被用来比较对照细胞和暴露在氧化应激下的细胞中线粒体蛋白质的丰度。广泛的蛋白质组筛选将允许发现以前未知的在线粒体中起作用的新基因产物。将对数据进行分析，以提供对蛋白质网络的新见解，这些蛋白质网络作用于修复线粒体DNA的氧化损伤或解毒ROS。

DESCRIPTION (provided by applicant) Mitochondria play a vital role in cell physiology and the response to environmental stress. A number of cellular toxins, including rotenone, 1-methyl-4-phenylpyridine (MPP+) and paraquat act to impair mitochondrial electron transport, generating ROS. Reactive oxygen species are also important in the toxicity of arsenic, amyloid, and ceramide. Mitochondria have been viewed as a potential source of ROS that may contribute to Parkinson's disease, aging and other pathological conditions. Since mitochondria contain only a small 16.5 kb mtDNA genome, encoding only 13 proteins, the organelle depends on the nucleus for most gene products, including all of the factors required for DNA replication, expression and repair. Recent studies from our laboratory and others have revealed an increasing collection of proteins that function in both mitochondria and other cellular compartments. A number of these proteins function in repair of oxidative damage to mtDNA. One significant consequence of mitochondrial pathology is the generation of mutations in mtDNA, many of which have a tissue-specific incidence, occurring most commonly in postreplicative tissues such as nerve and muscle. The investigators propose to test the hypothesis that mitochondria in differentiated cells may contain a different complement of proteins than actively dividing cells which may predispose post-replicative cells to a higher rate of mtDNA mutations or may alter the ability of cells to enter apoptosis. To accomplish this, they will study the effect of oxidative stress on the mitochondrial proteome in both embryonal carcinoma cells that are actively dividing and in cells that have been induced to differentiate along a neuronal pathway. Both 2-D gel methods and quantitative isotope-coded affinity tag (ICAT) methods will be used to compare the abundance of mitochondrial proteins in control cells and cells exposed to oxidative stress. The broad proteomic screen will permit the discovery of novel gene products not previously known to function in mitochondria. Data will be analyzed to provide new insights into networks of proteins acting to repair oxidative damage to mtDNA or to detoxify ROS.