Regulation of DNA recombination in Saccharomyces cerevisiae

酿酒酵母 DNA 重组的调控

• 批准号: 8667465
• 负责人: Grzegorz A Ira
• 金额: $ 29.55万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-05-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8667465
• 关键词: Aging; Animal Model; Biochemical; Biological Assay; Cell Cycle Regulation; Cells; Chemicals; Chromosomal Breaks; Cleaved cell; DNA; DNA Double Strand Break; DNA Repair; DNA Repair Pathway; DNA Sequence Rearrangement; DNA Structure; DNA biosynthesis; DNA-Directed DNA Polymerase; Data; Defect; Development; Disease; Double Strand Break Repair; Enzymes; Eukaryota; Event; Evolution; Gene Conversion; Genetic; Genetic Recombination; Genome; Genomic Instability; Goals; Human; Immune; Infertility; Invaded; Isotopes; Lead; Lesion; M cell; Malignant Neoplasms; Mediating; Modeling; Molecular; Molecular Genetics; Mutagenesis; Neurodegenerative Disorders; Pathway interactions; Play; Polymerase; Process; Proteins; Rad51 recombinase; Radiation therapy; Reaction; Recruitment Activity; Regulation; Research; Resolvase; Role; Saccharomyces cerevisiae; Sequence Homologs; Site; Telomerase; Telomere Maintenance; Testing; Work; cancer therapy; chemotherapy; chromatin immunoprecipitation; cytotoxic; density; helicase; homologous recombination; in vivo; irradiation; migration; novel; nuclease; prevent; reconstitution; repaired; research study

DNA recombination is an essential process that repairs DNA double-strand breaks (DSBs) and gaps that occur spontaneously, or are induced by chemicals or irradiation. In human, defects in recombination result in immune deficiencies, infertility, neurodegenerative disorders, developmental abnormalities, aging and cancer. DSBs as the most cytotoxic lesions are a fundamental component of the most prevalent cancer treatments, radiotherapy and radiomimetic chemotherapy. Therefore defining the genetic requirements and mechanisms of recombination pathways is of critical importance. A fundamental reaction during repair of broken chromosomes by recombination is DNA synthesis that copies homologous sequences from a template DNA molecule. The goal of this project is to understand the mechanisms and regulation of DNA synthesis during recombination, which remains very poorly understood. Two assays will be utilized to examine repair DNA synthesis that reflect two major recombination pathways with distinct DNA synthesis features. Both pathways play different yet important roles in cells. One is the simple repair of two-ended DSBs by gene conversion where both 3' ends prime DNA synthesis and thus only short leading strands are synthesized. The second assay employs break induced replication (BIR), in which a single DSB end invades a template, followed by extensive leading- and lagging- strand DNA synthesis. BIR is thought to be a mechanism of HR-dependent telomere maintenance in the absence of telomerase found in 10- 15% of all cancers. The specific aims are: (1) To understand the unique and redundant functions of multiple DNA polymerases recruited to DSBs and determine which DNA helicases and other enzymes specifically promote DNA synthesis during homologous recombination. The major focus will be on studying Pif1, the DNA helicase that we propose to be the first eukaryotic nonreplicative helicase that stimulates DNA synthesis during recombination. (2) To understand the mechanism of Break Induced Replication. Using isotope density transfer we will establish the mode of DNA synthesis in BIR and determine the role of DNA helicases and structure specific nucleases in BIR. Together we will provide a comprehensive view of DNA synthesis during homologous recombination.

DNA重组是修复DNA双链断裂（DSB）的重要过程， 自发发生的间隙，或由化学品或辐射引起的间隙。在人类中， 重组导致免疫缺陷、不育、神经退行性疾病， 发育异常衰老和癌症DSB作为最具细胞毒性的病变， 最普遍的癌症治疗的基本组成部分，放射治疗和 拟放射化疗因此，确定遗传要求和机制， 重组途径至关重要。在修复破损的皮肤时， 通过重组复制染色体上的同源序列是DNA合成， 模板DNA分子。该项目的目标是了解机制， 重组过程中DNA合成的调控，这仍然知之甚少。两 将利用分析来检查反映两种主要重组的修复DNA合成 具有独特的DNA合成特征的途径。这两种途径发挥着不同但重要的作用 在细胞中。一种是通过基因转换对两端DSB进行简单修复， 引物DNA合成，因此仅合成短的前导链。第二测定 采用断裂诱导复制（BIR），其中单个DSB末端侵入模板， 随后是大量的前导链和滞后链DNA合成。BIR被认为是 在10- 20岁儿童中发现，在缺乏端粒酶的情况下，HR依赖性端粒维持的机制。 占所有癌症的15%具体目标是：（1）理解唯一性和冗余性 多个DNA聚合酶的功能招募到DSB，并确定哪些DNA解旋酶 和其它酶在同源重组期间特异性地促进DNA合成。的 主要的重点将是研究Pif 1，DNA解旋酶，我们建议是第一个真核生物 在重组过程中刺激DNA合成的非复制性解旋酶。(2)了解 断裂诱导复制的机制。利用同位素密度转移，我们将建立 BIR中DNA合成的模式，并确定DNA解旋酶的作用和结构 BIR中的特异性核酸酶。我们将一起提供一个全面的看法DNA合成 在同源重组过程中。