Repair of oxidative DNA damage in mammalian cells

修复哺乳动物细胞中的氧化DNA损伤

• 批准号: 8063044
• 负责人: Tadahide Izumi
• 金额: $ 23.48万
• 依托单位: LSU HEALTH SCIENCES CENTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-03-10 至 2011-10-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8063044
• 关键词: Adjuvant; Affect; Affinity; Alkylation; Base Excision Repairs; Cancer cell line; Cell Death; Cells; DNA; DNA Binding; DNA Damage; Defense Mechanisms; Disease; Double Minutes; Effectiveness; Functional disorder; Genes; Genomic Instability; Genotoxic Stress; Healed; Hormones; Human; Kinetics; Lead; Left; Louisiana; Malignant Neoplasms; Mammalian Cell; Mammals; Methodology; Modality; Monoubiquitination; Mus; Mutagenesis; Mutation; Normal Cell; Oncogene Proteins; Organ Preservation; Oxidative Stress; Parathyroid gland; Pathway interactions; Pharmaceutical Preparations; Play; Predisposition; Protein p53; Proteins; Radiation therapy; Reactive Oxygen Species; Reagent; Regulation; Regulator Genes; Reporting; Resistance; Roentgen Rays; Role; Signal Transduction; Smoke; Suppressor Genes; Testing; Therapeutic; Toxic Environmental Substances; Ubiquitination; age related; anticancer research; cancer cell; cancer therapy; carcinogenesis; cell type; chemotherapy; domain mapping; endonuclease; healing; human APEX1 protein; human HDAC1 protein; improved; in vivo; insight; neoplastic cell; oxidation; oxidative DNA damage; polypeptide; public health relevance; repaired; research facility; response; transcription factor

DESCRIPTION (provided by applicant): DNA base excision repair (BER) is a ubiquitous and indispensable defense mechanism against cell death, mutagenesis, and carcinogenesis caused by spontaneously formed DNA damage, such as oxidation and alkylation. Apurinic/apyrimidinic endonuclease (APE1 in mammals) arguably determines the BER rate in cells and thus plays a central role in the repair pathway. Recent findings indicate that APE1 is essential for cellular viability, and that endogenous DNA damage is lethal if left unrepaired. Independently from its repair activity, APE1 interacts with histone deacetylase 1 (HDAC1) to suppress expression of many genes, including the parathyroid hormone gene. APE1 also regulates cJun/cFos and other crucial transcription factors. A high of APE1 is often in are expression observed malignant tumor cells, which more resistant to radio- and chemotherapeutic treatments, suggesting that APE1's repair function increases the chance of survival for cancer cells as well. Thus, APE1 may be either beneficial or harmful to humans, depending on the type of cells in which it is expressed. Understanding the mechanism for regulation of APE1 activity is therefore crucial for evaluating BER efficiency and its consequences in normal and cancer cells. Indeed, cancer predisposition by APE1 deficiency is known and p53 is an upstream regulator of APE1. We have discovered that APE1 is ubiquitinated by mouse double minute 2 (MDM2), and that the ubiquitination is induced by genotoxic stress in a p53-dependent manner. Ubiquitination occurs at specific Lys residues in the APE1 polypeptide, and appears to have significant effects on the functions of APE1 and other cellular proteins, including XRCC1 (X-ray cross-complementation 1), p53, and HDAC1, that interact directly with APE1. The central hypothesis of this project is that APE1 ubiquitination, regulated by MDM2 and p53, modulates the APE1 level and functions in DNA damage response. In order to elucidate the cellular defense mechanism involving this remarkably multifunctional protein, we will: (1) determine the effect of APE1 ubiquitination on its enzymatic and DNA binding activities; (2) unravel interaction of ubiquitinated APE1 with factors in BER and p53 signaling network; and (3) establish the regulation of APE1 ubiquitination in vivo. These parameters are crucial to predict susceptibility of cancer cells against therapeutic drugs that generates genomic instability. With the research facilities provided by Louisiana Cancer Research Consortium (LCRC), this project in the long-term should provide insights for developing new modalities for adjuvant cancer therapies and organ preservation, and for improving the management of other age-related pathophysiology. PUBLIC HEALTH RELEVANCE: DNA is continuously damaged by environmental toxicants like smoke, and by reactive oxygen radicals generated inside of our bodies. Damaged DNA causes mutations and may ultimately lead to cancer and age- related diseases. This project will help us understand how to enhance natural healing of DNA damage, and will help to improve the current methodology for non-surgical cancer treatments, including radiotherapy and chemotherapy.

描述（由申请人提供）：DNA碱基切除修复（BER）是一种无处不在且不可或缺的防御机制，可抵抗细胞死亡，诱变和由自发形成的DNA损伤引起的癌变，例如氧化和烷基化。可以说是细胞中的apurinic/apyrimidin核酸内切酶（哺乳动物中的APE1）确定细胞的BER速率，因此在修复途径中起着核心作用。最近的发现表明，APE1对于细胞活力至关重要，如果未修复，则内源性DNA损伤是致命的。 APE1独立于其修复活性，与组蛋白脱乙酰基酶1（HDAC1）相互作用，以抑制许多基因的表达，包括甲状旁腺激素基因。 APE1还调节CJUN/CFO和其他关键转录因子。通常在表达的APE1中，观察到的恶性肿瘤细胞对射线治疗和化学治疗更具耐药性，这表明APE1的修复功能也增加了癌细胞生存的机会。因此，取决于表达的细胞的类型，APE1可能对人类有益或有害。因此，了解调节APE1活性的机制对于评估BER效率及其在正常细胞和癌细胞中的后果至关重要。实际上，APE1缺乏症的癌症易感性是已知的，p53是APE1的上游调节剂。我们发现APE1被小鼠双分钟2（MDM2）泛素化，并且泛素化是由p53依赖性方式由遗传毒性应激诱导的。泛素化发生在APE1多肽中的特定LYS残基上，并且似乎对APE1和其他细胞蛋白的功能具有重大影响，包括XRCC1（X-ray交叉汇编1），p53和HDAC1与APE1直接相互作用。该项目的中心假设是，由MDM2和p53调节的APE1泛素化调节了DNA损伤响应中的APE1水平和功能。为了阐明涉及这种非常多功能蛋白的细胞防御机制，我们将：（1）确定APE1泛素化对其酶促和DNA结合活性的影响； （2）泛素化APE1与BER和p53信号网络中的因子的相互作用； （3）在体内建立APE1泛素化的调节。这些参数对于预测癌细胞对产生基因组不稳定性的治疗药物的敏感性至关重要。通过路易斯安那州癌症研究联盟（LCRC）提供的研究设施，该项目在长期内应为开发辅助癌症疗法和器官保存的新方式提供见解，并改善其他与年龄相关的病理生理学的管理。公共卫生相关性：DNA不断受到烟雾（例如烟雾）和人体内部产生的活性氧自由基的损害。受损的DNA会导致突变，并最终导致癌症和与年龄相关的疾病。该项目将有助于我们了解如何增强DNA损伤的自然愈合，并有助于改善当前非手术癌症治疗的方法，包括放疗和化学疗法。