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

• 批准号: 7916876
• 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
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-07 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7916876
• 关键词: Age; Benzo(a)pyrene; DNA; DNA Replication Proofreading; DNA lesion; DNA polymerase gamma; DNA-Directed DNA Polymerase; Defect; Disease; Environmental Risk Factor; Exonuclease; Genetic; Genome; Goals; HIV Infections; Health; Human; Lead; Lesion; Longevity; Metabolism; Mitochondria; Mitochondrial DNA; Mitochondrial Diseases; Mutation; Nuclear; Nucleosides; Nucleotides; Oxidative Stress; Phenotype; Polymerase; Reverse Transcriptase Inhibitors; Role; Time; abstracting; base; mitochondrial DNA mutation; mitochondrial dysfunction; mouse model; repaired

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).

抽象的 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 是线粒体内唯一已知的 DNA 聚合酶，因此它对于 mtDNA 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 核酸外切酶活性的基本机制，并确定其在疾病中的作用。基于以下内容，我们假设 DNA 聚合酶 γ 的核酸外切活性改变会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会导致毁灭性的线粒体疾病。 其次，功能性核酸外切酶可以改善 DNA 聚合酶 γ 中导致易错表型的聚合酶结构域突变。第三，DNA聚合酶γ校对缺陷的小鼠模型显示出增强的mtDNA突变和加速的衰老表型以及缩短的寿命。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来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，目前尚无治愈方法或有效的长期治疗方法。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 的核酸外切酶结构域在维持 mtDNA 完整性中的作用： 目的 1. 研究核酸外切酶结构域内疾病突变的潜在后果，以及这些突变如何导致线粒体功能障碍和疾病。 目的 2. 鉴定核酸外切酶结构域内参与校对的关键区域，并确定核酸外切酶切除掺入 NRTI 的效率，以及模板中错误掺入相反错误编码损伤的核苷酸（例如 8oxoG，氧化应激标记）和环境 DNA 损伤（例如由苯并[a]芘引起的损伤）。