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: 8397515
• 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/8397515
• 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.

描述（由申请人提供）： 项目摘要/抽象知识差距：很难研究从线粒体DNA损伤发出的效果，因为损害线粒体DNA的药物也损害了核DNA。由于核DNA约占细胞中DNA的98％，因此DNA损伤的作用可能是由于核DNA损伤引起的。核和线粒体DNA损伤的同时出现导致我们对细胞如何对线粒体DNA损伤的反应的关键障碍。在没有并发核DNA损伤的情况下，已经为该项目开发了小鼠模型，以促进线粒体DNA损伤的实验调节。这些模型将用于描述从线粒体DNA损伤中特异性产生的效果。将要测试的总体假设是线粒体DNA损伤导致线粒体，细胞和组织功能障碍。具体目的是：1）确定整个小鼠寿命的细胞对急性线粒体DNA损伤（例如在创伤期间可能发生的），2）确定慢性线粒体DNA损伤的影响（例如，糖尿病或帕金森氏病可能发生的可能发生的小鼠中的糖尿病或帕金森氏病），以及3）是否会在整个过程中造成鼠标dna损伤。研究设计：已经产生了转基因小鼠模型，该模型通过基因调控的四环素系统在实验调控下表达了限制性核酸内切酶ECORI。可以通过在食物中提供四环素类似物（强力霉素）来关闭基因并提供正常的Chow（无强力环素）来实现ECORI的表达来实现表达的调节。与ECORI编码序列融合在框架中的线粒体易位，可确保该蛋白转移到线粒体基质中，而不是核。一旦进入线粒体，ECORI就以双链断裂的形式裂解线粒体DNA。对于AIM 1，将在年轻人，中年或老年开始开启ECORI，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AIM 2，将在年轻人中打开ECORI，并留在老鼠的寿命中。将检查对线粒体功能的影响，以确定寿命的定义时间点，以确定慢性线粒体DNA损伤如何影响线粒体功能。对于AIM 3，将在寿命中定义的时间点上检查线粒体DNA损伤的后果的能力。拟议的研究旨在解决定义从线粒体DNA损伤中特异性作用的目标。