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The Mechanism of Mitochondrial DNA Proofreading in Human Health and Disease

线粒体 DNA 校对在人类健康和疾病中的机制

基本信息

批准号：
8117517
负责人：
SHERINE S CHAN
金额：
$ 24.9万
依托单位：
MEDICAL UNIVERSITY OF SOUTH CAROLINA
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-07 至 2013-07-31
项目状态：
已结题

项目摘要

ABSTRACT Mitochondria have their own closed circular genome. Mitochondrial DNA (mtDNA) has a 10 times greater rate of mutation than nuclear DNA. As DNA polymerase gamma is the only known DNA polymerase within the mitochondrion, it is essential for faithful replication, proofreading and repair of mtDNA. The overall goal of this proposal is to understand the fundamental mechanisms of exonuclease activity of DNA polymerase gamma in mtDNA metabolism, and to define its role in disease. Based on the following, we hypothesize that altered exonucleolytic activity of DNA polymerase gamma leads to mitochondrial dysfunction and disease. First, mutations in the exonuclease domain of DNA polymerase gamma cause devastating mitochondrial diseases. Second, polymerase domain mutations in DNA polymerase gamma that cause an error-prone phenotype can be ameliorated by functional exonuclease. Third, mouse models of proofreading-deficient DNA polymerase gamma show enhanced mtDNA mutations and accelerated ageing phenotype with reduced lifespan. Fourth, nucleoside reverse transcriptase inhibitors (NRTIs) used to control HIV infection induce mitochondrial dysfunction by inhibiting DNA polymerase gamma. Mitochondrial diseases can be caused by genetic and environmental factors. Unfortunately, there is no cure or no effective long-term treatment. As more diseases show mitochondrial defects, it is imperative that we understand how defects lead to disease. We will investigate the role of the exonuclease domain of DNA polymerase gamma in maintaining the integrity of mtDNA by: AIM 1. To investigate the underiying consequences of disease mutations within the exonuclease domain, and how these give rise to mitochondrial dysfunction and disease. AIM 2. To identify the critical regions within the exonuclease domain that are involved in proofreading, and to determine how efficientiy the exonuclease excises incorporated NRTIs, as well as nucleotides incorrectly incorporated opposite miscoding lesions in the template (such as 8oxoG, a marker of oxidative stress), and environmental DNA lesions (such as those caused by benzo[a]pyrene).

摘要线粒体有自己的闭合环状基因组。线粒体DNA（mtDNA）的突变率是核DNA的10倍。由于DNA聚合酶γ是线粒体中唯一已知的DNA聚合酶，它对线粒体DNA的忠实复制、校对和修复至关重要。该提案的总体目标是了解mtDNA代谢中DNA聚合酶γ的外切核酸酶活性的基本机制，并确定其在疾病中的作用。基于以下，我们假设DNA聚合酶γ的外切核酸溶解活性改变导致线粒体功能障碍和疾病。首先，DNA聚合酶γ的核酸外切酶结构域的突变导致毁灭性的线粒体疾病。第二，DNA聚合酶γ中引起易错表型的聚合酶结构域突变可以通过功能性核酸外切酶来改善。第三，校正缺陷DNA聚合酶γ的小鼠模型显示出增强的mtDNA突变和加速衰老表型，寿命缩短。第四，用于控制HIV感染的核苷逆转录酶抑制剂（NRTI）通过抑制DNA聚合酶γ诱导线粒体功能障碍。线粒体疾病可由遗传和环境因素引起。不幸的是，没有治愈或有效的长期治疗。随着越来越多的疾病显示线粒体缺陷，我们必须了解缺陷如何导致疾病。我们将通过以下方式研究DNA聚合酶γ的外切核酸酶结构域在维持mtDNA完整性中的作用： AIM 1.研究核酸外切酶结构域内疾病突变的潜在后果，以及这些突变如何引起线粒体功能障碍和疾病。 AIM 2.确定核酸外切酶结构域中参与校对的关键区域，并确定核酸外切酶如何有效地切除掺入的NRTI，以及模板中不正确掺入的相反错误编码病变（如8oxoG，氧化应激的标志物）和环境DNA病变（如由苯并[a]芘引起的病变）的核苷酸。