Amplification of Risk Caused by Mis-Routing of DNA Double-Strand Break Repair

DNA 双链断裂修复路径错误导致的风险放大

• 批准号: 8063644
• 负责人: Anna L Malkova
• 金额: $ 26.42万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8063644
• 关键词: Affect; Anaphase; Animal Model; Antineoplastic Agents; Cells; Centromere; Characteristics; Chromatids; Chromosomal Breaks; Chromosomal Rearrangement; Chromosome Breakage; Chromosome abnormality; Chromosomes; Complex; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Sequence Rearrangement; DNA biosynthesis; Data; Development; Double Strand Break Repair; Environmental Risk Factor; Etiology; Event; Exposure to; Frequencies; Gene Amplification; Gene Rearrangement; Genetic; Genetic Recombination; Genome; Genomic Instability; Genomics; Goals; Health; Hereditary Disease; Human; Interruption; Knowledge; Laboratories; Lead; Maize; Malignant Neoplasms; Mammals; Mediating; Methods; Molecular; Mutation; Outcome; Pathway interactions; Pharmaceutical Preparations; Process; Research; Research Personnel; Risk; Role; Saccharomyces cerevisiae; Series; Simulate; Site; Source; Structure; System; Testing; Work; Yeast Model System; Yeasts; assault; base; cancer cell; dimer; expectation; experience; high risk; innovation; mutant; neoplastic cell; prevent; repaired; research study; telomere

DESCRIPTION (provided by applicant): Double-strand DNA breaks (DSBs) are dangerous for human health because imprecise or faulty repair often leads to mutations and chromosome aberrations causing genetic diseases and cancer. The long-term goal of the investigator is to develop ways to minimize genomic instability resulting from DSBs. It is essential for this purpose to establish how DSB repair is executed and regulated, and how it leads to genome destabilization. The aim of this project is to unravel a number of molecular mechanisms capable of amplifying the consequences of DSBs in the model organism Saccharomyces cerevisiae. Firstly, this proposal is focused on chromatid fusions, which amplify the destabilizing effect of a single DSB by channeling it into breakage-fusion- bridge (BFB) cycles that create a series of rearrangement-prone secondary DSBs. Preliminary data allowed the investigator to propose that chromatid fusions can be stimulated by DSBs by allowing inter-molecular single-strand annealing (SSA) between inverted DNA repeats (IRs). Genetic methods and physical analyses of molecular intermediates are proposed to investigate this, as well as other homology-driven pathways of chromatid fusions that are currently poorly understood. Second, this proposal will unravel the mechanisms that allow broken chromosomes to acquire telomeres. Preliminary data suggested that break-induced replication (BIR) is the primary mechanism by which chromosomes undergoing BFBs are stabilized, which makes BIR the primary source of BFB-associated GCRs such as deletions, amplifications, and translocations. This research will specifically investigate the formation of translocations, which is the most deleterious outcome of BIR. Finally, results from genetic studies led to the hypothesis that interruption of BIR or other aberrant processing of BIR intermediates results in new chromosomal breakages that lead to cascades of DNA instability similar to the non-reciprocal translocations (NRTs) pathway known to amplify the number of rearrangements that result from an initial DSB in mammals. Thus, this proposal represents the first yeast model capable of simulating mammalian NRTs and is intended to unravel the molecular mechanisms of this process. In addition, the effects of genetic and environmental factors on channeling BIR repair into the GCR-producing pathways will be investigated. In summary, this research will elucidate the molecular mechanisms by which DSB repair can result in genomic consequences more destructive than the initial breakage. It is proposed that chromatid fusions, BIR, and NRTs are three such processes capable of amplifying the risks caused by a DSB due primarily to triggering BFB cycles. Further, experiments are proposed to test whether the magnification of damage that results from these genome-destabilizing DSB repair processes could be further amplified by cellular exposure to various environmental factors. To this end, experiments are planned to test the effects of various DNA damaging agents, including anti-cancer drugs, to investigate whether these agents might increase the frequency of high-risk repair processes or otherwise alter their outcomes. PUBLIC HEALTH RELEVANCE: This research is aimed to unravel the molecular mechanisms that lead to genomic destabilization by channeling double-strand DNA breaks into chromosomal rearrangements. Because genetic aberrations are a hallmark of cancer cells, this research will further our understanding of the etiology of some cancers.

描述（由申请人提供）：双链 DNA 断裂 (DSB) 对人类健康有害，因为不精确或错误的修复通常会导致突变和染色体畸变，从而导致遗传性疾病和癌症。研究人员的长期目标是开发尽量减少 DSB 导致的基因组不稳定性的方法。为此，有必要确定 DSB 修复是如何执行和调节的，以及它如何导致基因组不稳定。该项目的目的是揭示一些能够放大 DSB 在模式生物酿酒酵母中的影响的分子机制。首先，该提案重点关注染色单体融合，通过将单个 DSB 引导至断裂-融合-桥 (BFB) 循环，从而产生一系列易于重排的次级 DSB，从而放大单个 DSB 的不稳定效应。初步数据使研究人员提出，DSB 可以通过允许反向 DNA 重复序列 (IR) 之间的分子间单链退火 (SSA) 来刺激染色单体融合。提出了分子中间体的遗传方法和物理分析来研究这一点，以及目前知之甚少的染色单体融合的其他同源驱动途径。其次，该提议将揭示断裂染色体获得端粒的机制。初步数据表明，断裂诱导复制（BIR）是经历 BFB 的染色体稳定的主要机制，这使得 BIR 成为 BFB 相关 GCR（例如缺失、扩增和易位）的主要来源。这项研究将专门调查易位的形成，这是 BIR 最有害的结果。最后，遗传学研究的结果得出这样的假设：BIR 的中断或 BIR 中间体的其他异常加工会导致新的染色体断裂，从而导致 DNA 不稳定级联，类似于不可逆易位 (NRT) 途径，已知该途径会放大哺乳动物中初始 DSB 产生的重排数量。因此，该提议代表了第一个能够模拟哺乳动物 NRT 的酵母模型，旨在揭示这一过程的分子机制。此外，还将研究遗传和环境因素对将 BIR 修复引导至 GCR 产生途径的影响。总之，这项研究将阐明 DSB 修复可能导致比初始断裂更具破坏性的基因组后果的分子机制。有人提出，染色单体融合、BIR 和 NRT 是三个这样的过程，它们能够放大 DSB 造成的风险，主要是由于触发 BFB 循环。此外，还提出了实验来测试这些破坏基因组稳定的 DSB 修复过程所造成的损伤是否会因细胞暴露于各种环境因素而进一步放大。为此，计划进行实验来测试各种 DNA 损伤剂（包括抗癌药物）的效果，以研究这些药物是否会增加高风险修复过程的频率或以其他方式改变其结果。 公共健康相关性：本研究旨在通过将双链 DNA 断裂引导至染色体重排来揭示导致基因组不稳定的分子机制。由于遗传畸变是癌细胞的标志，这项研究将进一步加深我们对某些癌症病因学的了解。