GENETIC VARIATION IN HUMAN NHEJ DNA REPAIR GENES

人类 NHEJ DNA 修复基因的遗传变异

• 批准号: 6228824
• 负责人: CARL W ANDERSON
• 金额: $ 40.41万
• 依托单位: BROOKHAVEN SCIENCE ASSOC-BROOKHAVEN LAB
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6228824
• 关键词: DNA repair; breast neoplasms; gene mutation; gene targeting; genetic polymorphism; genetic recombination; high performance liquid chromatography; human genetic material tag; immunogenetics; ionizing radiation; neoplasm /cancer genetics; polymerase chain reaction; protein kinase; radiation genetics; severe combined immunodeficiency

DESCRIPTION: (Applicant's Description) DNA double-strand breaks (DSBs) occur spontaneously and are caused by exposures to a variety of environmental insults including ionizing radiation, oxidative stress, metals, and natural and man-made genotoxic substances. Accurate repair of DSBs is essential for cell survival and for the prevention of genome rearrangements that may lead to cancer. Human somatic cells have several mechanisms for repairing DSBs, but the most commonly used mechanism appears to be the error prone non-homologous end-joining pathway (NHEJ). This pathway includes at least five proteins that appear to be unique to the NHEJ mechanism, the three components of DNA-PK (DNA-PKcs, Ku70, Ku80), the double-stranded DNA activated protein kinase, and DNA ligase IV and XRCC4. Additional NHEJ components (e.g. MRE11, polymerase b, exonuclease FEN-1) may be shared with other repair systems, and undoubtedly other NHEJ components remain to be discovered. Together the genes for the five unique NHEJ components, which must work in concert, span about one megabase of the human genome, 20,145 bp of which code for amino acids. Natural genetic variation is expected to yield about20 single nucleotide variants within coding regions of this gene set, of which about half might be expected to alter amino acid sequences. Half of these will occur within the very large DNA-PK catalytic subunit, DNA-PKcs. We propose to use denaturing high-performance liquid chromatography and direct nucleotide sequencing of PCR products to discover the common polymorphisms in all ( about160) of the exons of the five NHEJ genes from about 90 different reference individuals. Then we will examine the pattern of polymorphisms in patients with medical conditions that might suggest a deficiency in the activity of one or more of these genes. Finally, we will characterize individual reactions within the NHEJ pathway in cell lines from individuals with different NHEJ polymorphims to determine if the efficiency or accuracy of NHEJ in humans is affected by the natural genetic variations within these genes in a way that might contribute to human disease. This study also will provide a moderately dense set of mapped polymorphic markers within the five know NHEJ genes for subsequence linkage studies that could reveal subtle effects and more complex allele interactions.

描述：(申请人描述)发生DNA双链断裂 自发性的，由暴露在各种环境侮辱中引起的 包括电离辐射、氧化应激、金属、天然和 人为的遗传毒性物质。双链断裂的准确修复对细胞至关重要 生存和防止基因组重排可能导致 癌症。人类体细胞有几种修复双链断裂的机制，但 最常用的机制似乎是容易出错的非同源 末端连接途径(NHEJ)。该途径包括至少五种蛋白质，它们 似乎是NHEJ机制所独有的，DNA-PK的三个组成部分 (DNA-PKcs，Ku70，Ku80)，双链DNA激活的蛋白激酶，以及 DNA连接酶IV和XRCC4。额外的NHEJ成分(例如，Mre11，聚合酶b， 核酸外切酶Fen-1)可以与其他修复系统共享，并且毫无疑问 NHEJ的其他成分仍有待发现。把五个人的基因放在一起 独特的NHEJ组件必须协同工作，跨越大约1兆数据库 人类基因组，其中20,145个核苷酸编码氨基酸。自然遗传 预计变异将在编码范围内产生约20个单核苷酸变体 这个基因组的区域，其中大约一半可能会改变氨基 酸序列。其中一半将发生在非常大的DNA-PK催化剂中 亚基，DNA-PKcs。我们建议使用变性高性能液体 对PCR产物进行层析和直接核苷酸测序以发现 五个NHEJ基因全部外显子(约160个)的常见多态 来自大约90个不同的参考个体。然后我们将检查该模式 在有疾病的患者中的多态可能表明 缺失这些基因中的一个或多个的活性缺失最后，我们会 表征NHEJ通路中的单个反应 具有不同NHEJ多态的个体来确定效率或 NHEJ在人类中的准确性受到体内自然遗传变异的影响 这些基因在某种程度上可能会导致人类疾病。这项研究还 中将提供一组中等密度的多态标记 5个已知NHEJ基因用于后续连锁研究，可能揭示微妙之处 影响和更复杂的等位基因相互作用。