Role of oxidative modifications to D-loop region in mTDNA replication in hypoxia

D 环区域氧化修饰在缺氧 mTDNA 复制中的作用

• 批准号: 8191634
• 负责人: Mykhaylo Ruchko
• 金额: $ 18.56万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-15 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8191634
• 关键词: Address; Bacterial Infections; Base Excision Repairs; Binding; Bioenergetics; Biogenesis; Blood Vessels; Cardiopulmonary; Cardiovascular Diseases; Cell Hypoxia; Cell physiology; Cells; Characteristics; Code; Complex; DNA; DNA Maintenance; DNA Repair; DNA Sequence; DNA strand break; Data; Defect; Detection; Development; Diabetes Mellitus; Disease; Endothelial Cells; Epithelial; Event; Exhibits; Funding Mechanisms; Gene Expression; Genes; Genetic Transcription; Health; Human; Hypoxia; Individual; Laboratories; Lead; Lesion; Ligation; Light; Link; Lung; Malignant Neoplasms; Mediating; Mitochondria; Mitochondrial DNA; Mitochondrial Diseases; Modeling; Modification; Molecular; Molecular Conformation; Neurodegenerative Disorders; Nuclear; Nucleic Acid Regulatory Sequences; Nucleosomes; Organelles; Oxidants; Pathology; Pathway interactions; Patients; Physiological; Process; Protein Binding; Reactive Oxygen Species; Reporter Genes; Research; Research Personnel; Response Elements; Risk; Role; Second Messenger Systems; Secondary to; Signal Pathway; Signal Transduction; Stimulus; Testing; Transcriptional Activation; Transcriptional Regulation; Vascular Endothelial Growth Factors; base; flexibility; genetic regulatory protein; high risk; mitochondrial DNA mutation; mitochondrial dysfunction; mitochondrial genome; neoplastic; novel; oxidative DNA damage; oxidative damage; promoter; repaired; response; second messenger; septic; transcription factor

DESCRIPTION (provided by applicant): Bioenergetic defects secondary to mitochondrial dysfunction occur in many severe illnesses, but the mechanisms by which mitochondria respond to, or accommodate the damage are not well understood. Very recently it has been demonstrated that survival in septic patients can be predicted by the strength of their mitochondrial biogenic response. Hypoxia, which complicates many cardiopulmonary, infectious, and neoplastic disorders, is one of multiple stimuli causing mitochondrial biogenesis. The mechanism underlying this adaptive response is unknown, but it is important that many stimuli known to increase mitochondrial biogenesis use reactive oxygen species (ROS) as second messengers. The proposed research will explore, at molecular and functional levels, a novel pathway regulating mitochondrial biogenesis. Traditional concepts hold that maintenance of DNA integrity is required for proper cell function. However, there is emerging evidence that, at least for nuclear genes, controlled DNA damage and repair may be necessary for normal transcriptional regulation. In lung vascular cells, for example, hypoxia causes ROS- dependent base modifications within hypoxic response elements (HREs) of hypoxia inducible genes. Because the lesions are restricted to HREs associated with transcriptionally-active nucleosomes and since mimicking the effect of hypoxia by introducing modified bases in the HRE of the VEGF promoter leads to enhanced DNA flexibility, altered transcription complex assembly and more robust reporter gene expression, it has been proposed that ROS-mediated DNA damage and repair may serve to alter the topology of key DNA sequences to enable regulatory protein binding and facilitate transcription. We propose to test the hypothesis that controlled oxidative DNA damage and repair in the D-loop region facilitates mtDNA transcription and replication. Using established strategies to alter the mtDNA repair efficiency, we will: (1) test the hypothesis that manipulation of hypoxia-induced oxidative damage to the mtDNA D-loop region coordinately regulates mtDNA replication and transcription in hypoxia, and, (2) determine whether formation and repair of hypoxia-caused oxidative base modifications in the D-loop region are required for transcription factor binding. If the concept that controlled DNA damage and repair govern mtDNA transcription and replication in hypoxia is valid, it will represent a significant advance in understanding how mitochondrial gene expression is regulated in health and diseases, including a number of disorders in which hypoxia and mitochondrial dysfunction have been incriminated. It will also contribute to a more detailed appreciation of the link between oxidant signaling and pathways governing mitochondrial adaptation, and thus point to new strategies for correcting mitochondrial bioenergetic defects in many disorders with such abnormalities. PUBLIC HEALTH RELEVANCE: Understanding the mechanisms of mitochondrial genome transcription and replication is very important for the explanation and treatment of a number of pathologies associated with mitochondrial dysfunction. Completion of the proposed studies will reveal a fundamentally new mechanism by which reactive oxygen species regulate replication and transcription of mtDNA. These studies are significant with respect to understanding a molecular link between a normal ROS-dependent process and the mtDNA instability characteristic of a variety of diseases including cancer, cardiovascular disease, diabetes and neurodegenerative diseases.

描述（由申请人提供）：继发于线粒体功能障碍的生物能缺陷发生在许多严重疾病中，但线粒体响应或适应损伤的机制尚不清楚。最近已经证明，脓毒症患者的生存可以通过线粒体生物反应的强度来预测。缺氧使许多心肺疾病、感染性疾病和肿瘤性疾病复杂化，是引起线粒体生物发生的多种刺激之一。这种适应性反应的机制尚不清楚，但重要的是，许多已知可增加线粒体生物发生的刺激都使用活性氧 (ROS) 作为第二信使。拟议的研究将在分子和功能水平上探索调节线粒体生物发生的新途径。传统观念认为，维持 DNA 完整性是细胞正常功能所必需的。然而，越来越多的证据表明，至少对于核基因来说，受控的 DNA 损伤和修复可能是正常转录调控所必需的。例如，在肺血管细胞中，缺氧会导致缺氧诱导基因的缺氧反应元件（HRE）内发生ROS依赖性碱基修饰。由于损伤仅限于与转录活性核小体相关的 HRE，并且通过在 VEGF 启动子的 HRE 中引入修饰碱基来模拟缺氧的影响，可增强 DNA 灵活性、改变转录复合物组装和更稳健的报告基因表达，因此有人提出，ROS 介导的 DNA 损伤和修复可能有助于改变关键 DNA 序列的拓扑结构，从而实现调节蛋白结合并促进转录。我们建议检验以下假设：控制 D 环区域的 DNA 氧化损伤和修复促进 mtDNA 转录和复制。使用已建立的策略来改变 mtDNA 修复效率，我们将：(1) 检验以下假设：操纵缺氧诱导的 mtDNA D 环区域氧化损伤可协调调节缺氧条件下 mtDNA 的复制和转录，(2) 确定是否D 环区域缺氧引起的氧化碱基修饰的形成和修复是转录因子结合所必需的。如果控制 DNA 损伤和修复控制缺氧条件下 mtDNA 转录和复制的概念是正确的，那么这将代表着在理解健康和疾病（包括缺氧和线粒体功能障碍所导致的许多疾病）中线粒体基因表达如何受到调节方面取得了重大进展。被定罪。它还将有助于更详细地了解氧化信号传导和控制线粒体适应的途径之间的联系，从而指出纠正许多具有此类异常的疾病中的线粒体生物能缺陷的新策略。 公共卫生相关性：了解线粒体基因组转录和复制的机制对于解释和治疗与线粒体功能障碍相关的许多病理学非常重要。完成拟议的研究将揭示活性氧调节 mtDNA 复制和转录的全新机制。这些研究对于理解正常 ROS 依赖性过程与多种疾病（包括癌症、心血管疾病、糖尿病和神经退行性疾病）的 mtDNA 不稳定性特征之间的分子联系具有重要意义。