Repair Mechanisms For Oxidative DNA Damage

DNA 氧化损伤的修复机制

基本信息

批准号：
7592049
负责人：
David M Wilson
金额：
$ 74.94万
依托单位：
NATIONAL INSTITUTE ON AGING
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7592049
关键词：
8-oxoguanine APTX gene ATP phosphohydrolase Aging Base Excision Repairs Biochemical C-terminal Cell Aging Cell Death Cell Respiration Cell Survival Cell physiology Cells Chromosomal Stability Chronic Cleaved cell Cockayne Syndrome Complex Coupled Cultured Cells Cutaneous DNA DNA Damage DNA Polymerase beta DNA Repair DNA Structure DNA strand break Defect Disease Disease susceptibility Dominant-Negative Mutation Enzymes Failure Functional disorder Future Genetic Genetic Transcription Growth Human Incidence Individual Interphase Cell Investigation Laboratories Ligation Link Lipids Malignant Neoplasms Mechanics Mediating Methods Modeling Molecular Mus Nerve Degeneration Neurodegenerative Disorders Neurologic Neurons Nucleic Acids Numbers PCNA gene Pathology Pathway interactions Photosensitivity Population Premature aging syndrome Process Property Proteins RNA RNA Interference Range Reactive Oxygen Species Role SWI2/SNF2 Scaffolding Protein Single Strand Break Repair Site Specificity Spinocerebellar Ataxias Structure Surgical incisions Symptoms System Telomere Maintenance Testing Therapeutic Therapeutic Agents Thinking Variant Work XRCC1 gene age related cancer cell cancer therapy cell age chemotherapeutic agent chromatin remodeling cytotoxic design disorder risk helicase homologous recombination novel oxidative DNA damage protein function protein protein interaction repaired response scaffold tyrosyl-DNA phosphodiesterase

项目摘要

Much of our effort has been to define the structure-function mechanisms and biochemical properties of Ape1, the major protein for repairing abasic sites and certain 3-damages in DNA. Our work has revealed that Ape1 cleaves at AP sites in single-stranded regions of complex, biologically-relevant DNA structures, such as bubble and fork intermediates. These findings expand the known repertoire of substrates processed by this enzyme, and suggest novel functions for Ape1 likely coupled to transcription and/or replication. We recently found that the protein defective in the human segmental progeroid, Cockayne Syndrome B (CSB), physically and functionally interacts with Ape1. We are now determining the precise molecular contributions of CSB to BER and testing more exhaustively for possible helicase- and remodeling-type activities for CSB on a variety of DNA and RNA substrates. These studies in total will determine the molecular functions of CSB and how certain activities contribute to the associated disease manifestation. Last, we are designing methods to strategically regulate Ape1 repair activity using a dominant-negative approach in the hopes of developing more effective anti-cancer treatment paradigms. In addition to the investigations above, we are elucidating the biochemical and cellular contributions of XRCC1, a major SSB repair (SSBR) factor. This protein functions primarily as a scaffold component, orchestrating specific protein-protein interactions required for efficient DNA repair. Our studies (i) suggest a link of XRCC1 to replication via an interaction with PCNA, (ii) argue against a role for XRCC1 in the early steps of BER, and (iii) indicate a biologically-relevant role for its interaction with DNA polymerase beta and in the subsequent repair step, nick ligation. Recent work has identified associations of XRCC1 with proteins defective in human neurodegenerative disorders AOA1 (Aprataxin) and SCAN1 (TDP1). We are assessing the role of XRCC1 in non-dividing (neuronal) cells and age-related pathologies (namely neurodegeneration) using cell culture models and heterozygous mice. The contributions of XRCC1 to DNA damage responses, chromosome stability, and telomere maintenance are concurrently being evaluated in dividing human cells using chronic RNAi knockdown strategies. The ongoing investigations will delineate the contribution of SSBs (i.e. an XRCC1 deficiency), a common DNA intermediate, to cancer promotion (genetic stability) and neurodegenerative disease (neuronal cell viability).

我们的大部分努力是定义APE1的结构功能机理和生化特性，APE1是修复无碱性位点的主要蛋白质和DNA中某些3损伤的蛋白质。我们的工作表明，APE1在复杂的，生物学上与生物学的DNA结构（例如气泡和叉子中间体）的单链区域的AP位点裂解。这些发现扩大了该酶处理的底物的已知曲目，并提出了可能与转录和/或复制结合的APE1的新功能。我们最近发现，人类节段后代综合征B（CSB）的蛋白质缺陷在物理和功能上与APE1相互作用。现在，我们正在确定CSB对BER的精确分子贡献，并更详尽地测试CSB可能在多种DNA和RNA底物上的CSB的解旋酶和重塑型活性。这些研究总共将确定CSB的分子功能以及某些活动如何促进相关疾病表现。最后，我们正在设计使用优势阴性方法来战略性地调节APE1修复活动的方法，以期开发更有效的抗癌治疗范例。除了上述研究外，我们还阐明了XRCC1的生化和细胞贡献，XRCC1是主要的SSB修复（SSBR）因子。该蛋白质主要是脚手架成分，策划了有效的DNA修复所需的特定蛋白质 - 蛋白质相互作用。我们的研究（i）提出了XRCC1通过与PCNA的相互作用进行复制的联系，（ii）反对XRCC1在BER的早期步骤中的作用，（III）表明其与DNA聚合酶β的相互作用与DNA聚合酶β的相互作用以及随后的维修步骤，NICK结扎。最近的工作已经确定了XRCC1与人神经退行性疾病中有缺陷的AOA1（Aprataxin）和Scan1（TDP1）中有缺陷的蛋白的关联。我们正在使用细胞培养模型和杂合小鼠评估XRCC1在非分裂（神经元）细胞和年龄相关病理学（即神经变性）中的作用。 XRCC1对DNA损伤反应，染色体稳定性和端粒维持的贡献是在使用慢性RNAi敲低策略分裂的。正在进行的研究将描述SSB（即XRCC1缺乏症），一种常见的DNA中间体对癌症促进（遗传稳定性）和神经退行性疾病（神经元细胞活力）的贡献。