Human DNA Repair Enzymes for Redox and Alkylation Damage

用于氧化还原和烷基化损伤的人类 DNA 修复酶

基本信息

批准号：
7752826
负责人：
Bruce F. Demple
金额：
$ 31.76万
依托单位：
STATE UNIVERSITY NEW YORK STONY BROOK
依托单位国家：
美国
项目类别：
财政年份：
1988
资助国家：
美国
起止时间：
1988-06-01 至 2012-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7752826
关键词：
Address Aging-Related Process Alkylating Agents Alkylation Antioxidants Apoptosis Base Excision Repairs Biochemical Biochemistry Biological Biological Process Cell Line Cell physiology Cells Collaborations Complementary DNA DNA DNA Damage DNA Polymerase beta DNA Repair DNA Repair Enzymes DNA glycosylase DNA lesion DNA-(apurinic or apyrimidinic site) lyase DNA-Directed DNA Polymerase Data Deoxyribose Disease Down-Regulation Doxycycline Embryo Enzymes Excision Excision Repair Family Process Fibroblasts Free Radicals Genes Genetic Genetic screening method Genome Goals Growth Histocompatibility Testing Histone Deacetylase Human Human Cell Line Human Genome In Vitro Kinetics Knock-out Laboratories Lesion Link Longevity Lyase Maintenance Malignant Neoplasms Mammalian Cell Metabolic Mouse Cell Line Mus Mutation Normal Cell OGG1 gene Oxidation-Reduction Pathway interactions Phenotype Physiological Premature aging syndrome Process Progress Reports Promoter Regions Proteins RNA Interference Research Personnel Resistance Role Site Small Interfering RNA Source Specificity System Technology Testing Tetanus Helper Peptide Transcription Coactivator Work Yeasts age related base cell growth cell type endonuclease human APEX1 protein human DNA mutant normal aging novel osteosarcoma oxidative damage programs repaired success

项目摘要

Genetic inheritance, and resistance to age-related diseases such as cancer, depend on the stability of the human genome. Stable genome maintenance in turn depends critically on cellular systems that process "spontaneous" DNA damage. Such damage includes hydrolytic decay lesions, in particular abasic sites, as well as lesions formed by metabolic by-products such as free radicals and alkylating agents. Existing data indicate that the base excision repair (BER) system is most likely to deal with this endogenous DNA damage. In BER, DNA glycosylases remove altered bases to generate abasic sites, which are further processed in several steps dependent on the Ape1 endonuclease and DNA polymerase beta. Our long-term goal has been to ascertain the biological functions of Ape1 protein as a central player in base excision repair (BER) of DNA and perhaps other processes, and to understand the coordination of activities in the various branches of BER. Past efforts have been valuable in defining the biochemistry of Ape1 activity, but only recently have we been able to conduct genetic tests of the protein's cellular function through the use of small-interfering RNA (siRNA). These data clearly support an essential role of Ape1 in maintaining the viability of human cells by processing abasic DNA damage formed by endogenous processes. Other recent work points to a novel role for the sirtuin Sirt6 in modulating BER, possibly through effects on the activity of DNA polymerase beta. Since deletion of the SIRT6 in mice shortens lifespan and generates various phenotypes associated with premature aging, these observations provide the first direct link between BER and the normal aging process. The work proposed here builds on this success in order to define the cellular function of Ape1 and the origins of the DNA damage that it processes, and how the BER can be modulated by a sirtuin. Three specific aims will address our goals: 1. Establish human or mouse cell lines with regulated expression of APE1-specific si-RNA and examine the kinetics of the biological effects accompanying Ape1 depletion; 2. Determine the contribution of DNA glycosylases (in particular UNG2, OGG1, AAG/MPG, and NTH1) and of oxidative damage as sources of the lethal lesions that accumulate in Ape1-deficient cells; 3. Directly test the role of SirtG in DNA repair and the sub-pathways of BER.

遗传遗传以及对年龄相关疾病（例如癌症）的抵抗，取决于人基因组。稳定的基因组维持反过来严重取决于处理的细胞系统 “自发” DNA损伤。这种损害包括水解衰减病变，特别是无碱性部位，例如以及由自由基和烷基化剂等代谢副产品形成的病变。现有数据表明基本切除修复（BER）系统最有可能处理这种内源性DNA损伤。在BER中，DNA糖基酶去除了改变的碱基以产生无碱性位点，这些位点进一步加工几个步骤取决于APE1核酸内切酶和DNA聚合酶β。我们的长期目标为了确定APE1蛋白作为基础切除修复（BER）的核心参与者的生物学功能（BER） DNA以及其他过程，并了解各个分支机构的活动的协调伯。过去的努力在定义APE1活动的生物化学方面非常有价值，但直到最近才我们能够通过使用小型疗法进行蛋白质细胞功能的基因检测 RNA（siRNA）。这些数据显然支持APE1在维持人的可行性中的重要作用通过处理由内源过程形成的无碱性DNA损伤的细胞。其他最近的工作指向 Sirtuin Sirt6在调节BER中的新作用，可能通过对DNA聚合酶活性的影响 beta。由于删除小鼠中的SIRT6会缩短寿命并产生相关的各种表型随着过早的衰老，这些观察结果提供了BER与正常衰老之间的第一个直接联系过程。此处提出的工作是基于此成功的，以定义APE1和它处理的DNA损伤的起源，以及如何通过Sirtuin调节BER。三具体目标将解决我们的目标： 1。建立具有APE1特异性SI-RNA的调节表达的人或小鼠细胞系，并检查伴随APE1耗竭的生物学作用的动力学； 2。确定DNA糖基酶的贡献（尤其是UNG2，OGG1，AAG/MPG和NTH1）和氧化损伤作为积累在APE1缺陷细胞中的致命病变的来源； 3。直接测试SIRTG在DNA修复和BER的子街道中的作用。