Mitochondrial DNA Damage: Cellular Responses, Aging and Disease

线粒体 DNA 损伤：细胞反应、衰老和疾病

• 批准号: 7930438
• 负责人: Christi A Walter
• 金额: --
• 依托单位: SOUTH TEXAS VETERANS HEALTH CARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7930438
• 关键词: Acute; Address; Adult; Affect; Age; Aging; Cell Nucleus; Cell physiology; Cells; Chronic; Cleaved cell; Code; DNA; DNA Damage; DNA Restriction Enzymes; Deoxyribonuclease EcoRI; Diabetes Mellitus; Disease; Doxycycline; Exposure to; Food; Functional disorder; Gene Expression Regulation; Genes; Genetic; Goals; Homeostasis; Inherited; Knowledge; Left; Long-Term Effects; Longevity; Malignant Neoplasms; Mediating; Mitochondria; Mitochondrial DNA; Mitochondrial Matrix; Mitochondrial Proteins; Modeling; Mus; Mutagens; Neurodegenerative Disorders; Nuclear; Oxidative Phosphorylation; Parkinson Disease; Pathway interactions; Physiological; Production; Proteins; Regulation; Research; Research Design; Role; Stress; System; Testing; Tetracyclines; Tissues; Toxin; Transgenic Mice; Trauma; Veterans; abstracting; analog; biological adaptation to stress; disease-causing mutation; experience; middle age; mouse model; prevent; response; young adult

DESCRIPTION (provided by applicant): Project Summary/Abstract Knowledge gap: The effects that emanate specifically from mitochondrial DNA damage have been difficult to study because agents that damage mitochondrial DNA also damage nuclear DNA. Because nuclear DNA represents approximately 98% of the DNA in a cell, the demonstrated effects of DNA damage have likely been due to nuclear DNA damage. The simultaneous occurrence of nuclear and mitochondrial DNA damage has resulted in a critical barrier to our understanding of how cells respond specifically to mitochondrial DNA damage. Mouse models have been developed for this project that facilitate experimental regulation of mitochondrial DNA damage in the absence of concurrent nuclear DNA damage. These models will be used to delineate the effects specifically emanating from mitochondrial DNA damage. The overall hypothesis that will be tested is that mitochondrial DNA damage results in mitochondrial, cellular and tissue dysfunction. The specific aims are: 1) to determine the cellular responses to acute mitochondrial DNA damage (such as might occur during trauma) across the mouse lifespan, 2) to determine the effects of chronic mitochondrial DNA damage (such as might occur with diabetes or Parkinson's disease) in the mouse, and 3) to determine if the consequences of mitochondrial DNA damage can be reversed throughout the mouse lifespan. Research design: Transgenic mouse models have been produced that express the restriction endonuclease EcoRI under experimental regulation via the tetracycline system of gene regulation. Regulation of expression can be achieved by supplying the tetracycline analogue, doxycycline, in food to turn off the gene and providing normal chow (no doxycycline) to achieve expression of EcoRI. A mitochondrial translocation presequence fused in frame with the EcoRI coding sequences assures that the protein is translocated to the mitochondrial matrix and not to the nucleus. Once in the mitochondria, EcoRI cleaves the mitochondrial DNA creating damage in the form of double-strand breaks. For aim 1, EcoRI will be turned on at young adult, middle-age or old age to determine how age impacts the effects of acute mitochondrial DNA damage. A number of mitochondrial functions will be examined to determine the effects on mitochondrial function. For aim 2, EcoRI will be turned on in young adults and left on for the lifespan of the mice. Effects on mitochondrial function will be examined at defined timepoints in the lifespan to determine how chronic mitochondrial DNA damage affects mitochondrial function. For aim 3, the ability to reverse the consequences of mitochondrial DNA damage will be examined at defined timepoints in the lifespan. The proposed studies are intended to address the goal of defining effects specifically emanating from mitochondrial DNA damage. PUBLIC HEALTH RELEVANCE: Project Narrative Mitochondrial DNA damage threatens the ability of mitochondria to fulfill their role in ATP production. A large portion of the cells's ATP is produced by mitochondria through the process of oxidative phosphorylation. Because mitochondrial DNA encodes 13 subunits required for oxidative phosphorylation, integrity of mitochondrial DNA is important for energy production. Mitochondria have been implicated in stroke, severe sepsis, severe burns, aging and neurodegenerative disorders among others. U.S. veterans are susceptible to these health issues. Therefore, understanding the effects of mitochondrial DNA damage is essential to understanding how mitochondria affect processes such as aging, neurodegenerative disease, stroke, infection, burns and other clinical conditions relevant to veterans.

描述（由申请人提供）： 项目摘要/摘要 知识差距：线粒体 DNA 损伤所产生的影响一直难以研究，因为损伤线粒体 DNA 的药物也会损伤核 DNA。由于核 DNA 约占细胞中 DNA 的 98%，因此所证实的 DNA 损伤效应很可能是由于核 DNA 损伤造成的。细胞核和线粒体 DNA 损伤的同时发生，给我们理解细胞如何对线粒体 DNA 损伤做出特异性反应造成了严重障碍。该项目开发了小鼠模型，可在不存在并发核 DNA 损伤的情况下促进线粒体 DNA 损伤的实验调节。这些模型将用于描述线粒体 DNA 损伤所产生的具体影响。将要测试的总体假设是线粒体 DNA 损伤会导致线粒体、细胞和组织功能障碍。具体目标是：1) 确定小鼠整个生命周期中细胞对急性线粒体 DNA 损伤（例如可能在创伤期间发生）的反应，2) 确定慢性线粒体 DNA 损伤（例如糖尿病或帕金森病可能发生的情况）的影响疾病）在小鼠中，以及3）确定线粒体DNA损伤的后果是否可以在小鼠的整个生命周期中逆转。研究设计：已经产生了转基因小鼠模型，该模型在通过四环素基因调控系统的实验调控下表达限制性核酸内切酶EcoRI。可以通过在食物中提供四环素类似物多西环素来关闭该基因并提供正常食物（无多西环素）来实现 EcoRI 的表达，从而实现表达调节。线粒体易位前序列与 EcoRI 编码序列框内融合，确保蛋白质易位至线粒体基质而不是细胞核。一旦进入线粒体，EcoRI 就会切割线粒体 DNA，以双链断裂的形式造成损伤。对于目标 1，EcoRI 将在年轻人、中年或老年时启动，以确定年龄如何影响急性线粒体 DNA 损伤的影响。将检查许多线粒体功能以确定对线粒体功能的影响。对于目标 2，EcoRI 将在年轻成年小鼠中开启，并在小鼠的整个生命周期中保持开启状态。将在生命周期的特定时间点检查对线粒体功能的影响，以确定慢性线粒体 DNA 损伤如何影响线粒体功能。对于目标 3，将在生命周期的特定时间点检查逆转线粒体 DNA 损伤后果的能力。拟议的研究旨在解决明确线粒体 DNA 损伤所产生的影响的目标。 公共卫生相关性： 项目叙述线粒体 DNA 损伤威胁着线粒体发挥其在 ATP 生产中的作用的能力。细胞的 ATP 的很大一部分是由线粒体通过氧化磷酸化过程产生的。由于线粒体 DNA 编码氧化磷酸化所需的 13 个亚基，因此线粒体 DNA 的完整性对于能量产生非常重要。线粒体与中风、严重败血症、严重烧伤、衰老和神经退行性疾病等有关。美国退伍军人很容易受到这些健康问题的影响。因此，了解线粒体 DNA 损伤的影响对于了解线粒体如何影响衰老、神经退行性疾病、中风、感染、烧伤和其他与退伍军人相关的临床病症等过程至关重要。