Defining role of Fanconi anemia complementation group A protein in DNA repair

范可尼贫血补充 A 组蛋白在 DNA 修复中的定义作用

基本信息

批准号：
9173233
负责人：
Yanbin Zhang
金额：
$ 38.99万
依托单位：
UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-07-01 至 2020-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9173233
关键词：
Affect Binding Biochemical Biological Biological Assay Cells Child Collaborations Complex Core Protein Critical Pathways Cruciform DNA DNA DNA Damage DNA Double Strand Break DNA Interstrand Crosslinking DNA Maintenance DNA Repair DNA annealing DNA replication fork Data Defect Deoxyribonuclease I Disease Double Strand Break Repair Etiology Fanconi Anemia Complementation Group A Protein Fanconi anemia protein Fanconi&apos s Anemia Genes Genetic Genome Stability Goals Health Hereditary Disease Human Hypersensitivity In Vitro Leucine Zippers Life Link Malignant Neoplasms Measures Mediating Methods Molecular Monitor Monoubiquitination Mutate Mutation Pancytopenia Pathway interactions Patients Phosphorylation Physiological Play Predisposition Process Proteins Research Role Single-Stranded DNA Site Structure Surgical incisions Syndrome System Testing Ubiquitination base crosslink cytotoxic endodeoxyribonuclease SceI homologous recombination imaging system in vitro activity in vivo innovation insight live cell imaging member migration mutant novel patient population recombinase repaired stoichiometry tool

项目摘要

Interstrand crosslinks (ICLs) are deleterious DNA damages in which both strands are covalently bound together, generating a stalled replication state cytotoxic to cells. Defective repair of ICLs is associated with Fanconi Anemia syndrome (FA) and predisposition to cancer. At least seventeen FA genes have been identified up to date. This proposal focuses on a specific FA protein, FANCA, a member of the FA core complex that is mutated in 64% of the entire FA patient population and does not have a clear function. Our preliminary study shows that FANCA destabilizes (unwinds) DNA helix and recognizes DNA interstrand crosslink damage (ICL) in a replication fork. Intriguingly, FANCA anneals single-strand DNA and catalyzes strand exchange as well. Furthermore, FANCA synergistically interacts with Rad51, the major recombinase in double strand break (DSB) repair. Based on these data, we hypothesize that FANCA directly participates in repair of interstrand crosslinks through its DNA-destabilizing (`unwinding') activity and facilitates subsequent repair of double strand DNA breaks by catalyzing strand annealing and exchange. In order to delineate the role of FANCA in DNA repair, we will use a biochemically defined in vitro system, a cell-based DSB repair study system, and a living cell imaging system to accomplish four aims: 1) Delineate the molecular mechanism of how FANCA catalyzes DNA `unwinding', single-strand annealing, and strand exchange. 2) Determine the role of FANCA in recognition and incision of DNA interstrand crosslinks. 3) Study the biological role of FANCA in double strand break repair. 4) Examine the physiological role of the DNA `unwinding', single- strand annealing, and strand exchange activities of FANCA in the etiology of Fanconi anemia. Understanding the role of FANCA in DNA repair will not only contribute to the overall clarification of the ICL repair process, but also provide novel insights into the etiology of Fanconi anemia and its associated cancer.

链间交叉链接（ICL）是有害的DNA损害，这两个链都是共价结合的一起，对细胞产生停滞的复制态细胞毒性。 ICL的维修有缺陷与 Fanconi贫血综合征（FA）和癌症易感性。至少有17个FA基因已经确定最新。该提案重点介绍特定的FA蛋白Fanca，FA核心成员复合物在64％的整个FA患者人群中被突变，没有明确的功能。我们的初步研究表明，Fanca破坏了（Unwinds）DNA螺旋并识别DNA 复制叉中的链间交联损伤（ICL）。有趣的是，Fanca退火单链DNA和也催化链交换。此外，Fanca协同与Rad51协同相互作用双链断裂（DSB）修复中的重组酶。基于这些数据，我们假设Fanca直接通过其DNA-DESTABILIZE（“放松”）活动的活动进行维修，并促进随后通过催化链退火和交换来修复双链DNA断裂。为了描述了粉丝在DNA修复中的作用，我们将使用基于细胞的生化定义的体外系统定义 DSB维修研究系统和一个实现四个目标的活细胞成像系统：1）描述分子 Fanca如何催化DNA“放松”，单链退火和链交换的机制。 2）确定FANCA在识别和切口DNA间链交联中的作用。 3）研究生物学 Fanca在双链断裂修复中的作用。 4）检查DNA“放松”，单一的生理作用链退火和范卡的链交换活动在范科尼贫血的病因中。理解 FANCA在DNA修复中的作用不仅有助于ICL维修过程的总体澄清，还会有助于还提供有关范科尼贫血及其相关癌症病因的新见解。