Mechanism and Regulation of Nonhomologous DNA End Joining

非同源DNA末端连接的机制和调控

• 批准号: 8894424
• 负责人: MICHAEL R LIEBER
• 金额: --
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8894424
• 关键词: Affect; Aging; BRCT Domain; Binding; Cancer Etiology; Cells; Chemicals; Chromosomal translocation; Chromosome Pairing; Complex; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Repair Pathway; DNA Sequence; DNA Structure; DNA ligase IV; DNA-PKcs; DNA-Protein Interaction; Development; Enzymes; Free Radicals; Health; Histones; Human; Ionizing radiation; Ku Protein; Length; Ligase; Ligation; Malignant Neoplasms; Mediating; Modification; Mutation; Nonhomologous DNA End Joining; Pathway interactions; Polymerase; Positioning Attribute; Price; Process; Property; Proteins; Radiation; Radiation-Sensitizing Agents; Regulation; Site; Somatic Cell; System; Tail; Testing; Therapeutic; Time; Variant; Vertebral column; Vertebrates; Work; X-Ray Crystallography; XRCC4 gene; artemis; chemotherapy; flexibility; homologous recombination; improved; in vivo; inhibitor/antagonist; inorganic phosphate; insight; mutant; nuclease; phosphodiester; reconstitution; repaired; small molecule; spatial relationship; stoichiometry

DESCRIPTION (provided by applicant): Double-strand DNA breaks (DSBs) occur in all cells multiple times per day. Homologous recombination (HR) and nonhomologous DNA end-joining (NHEJ) are the two primary pathways for repairing DSBs. However, NHEJ is the more general pathway because it does not require a homologous donor, and NHEJ is sufficiently flexible that it can join almost any pair of incompatible DNA ends. The flexibility of NHEJ is essential because natural causes of DSBs (ionizing radiation, free radicals, enzyme malfunction) generate DNA ends with diverse chemical and structural configurations. Hence, that flexibility is well-suited for the task, but the price that we pay for that flexibility is that NHEJ causes DNA sequence changes at most sites where it functions (in humans and other vertebrates). Hence, NHEJ generates somatic cell mutations that cause cancer and likely contribute to aging. The flexibility of NHEJ represents one of the most sophisticated protein:DNA interaction pathways. Though we know most of the proteins that participate in NHEJ and know, in broad terms, what they do, we do not have a clear picture of how they function together, their spatial configuration, or the temporal relationships. With a clearer mechanistic and structural picture of human NHEJ, we will be in a position to develop small molecule inhibitors that may be useful for treating cancer (as chemotherapy or as a radiation sensitizer). Aims 1 and 2 of this proposal focus precisely on the issues of structural and spatial relationships among the NHEJ proteins and with the DNA ends. Aim 3 tests the hypothesis that the NHEJ ligase complex is important for making key end-to-end contacts during NHEJ. Aim 4 describes development of a more robust NHEJ reconstitution. In Aim 4, that system is then used to identify any remaining NHEJ factors by testing for enhancement of activity. Aim 5 describes analysis of how histone octamers influence the nuclease, polymerase, and ligase activities of NHEJ. These studies will markedly improve our understanding of how NHEJ participates in processes such as chromosomal translocations that are of key importance in cancer etiology and how NHEJ might be made 'druggable' for therapeutic purposes.

描述（由申请人提供）：每天多次发生双链DNA断裂（DSB）。同源重组（HR）和非同源DNA端连接（NHEJ）是修复DSB的两种主要途径。但是，NHEJ是更通用的途径，因为它不需要同源供体，并且NHEJ足够灵活，几乎可以连接任何一对不兼容的DNA末端。 NHEJ的灵活性是必不可少的，因为DSB（电离辐射，自由基，酶故障）的自然原因会以不同的化学和结构构型产生DNA结束。因此，这种灵活性非常适合任务，但是我们为此灵活性付出的价格是NHEJ在其功能（在人类和其他脊椎动物中）的大多数站点都会导致DNA序列发生变化。因此，NHEJ会产生导致癌症并可能导致衰老的体细胞突变。 NHEJ的灵活性代表了最复杂的蛋白质之一：DNA相互作用途径。尽管我们知道大多数参与NHEJ的蛋白质，并以广义的方式知道它们的工作，但我们对它们的运作方式，空间配置或时间关系并不清楚。通过人类NHEJ的机械和结构图，我们将有能力开发出可能有助于治疗癌症（作为化学疗法或作为放射线敏化剂）的小分子抑制剂。该提案的目标1和2精确地关注NHEJ蛋白之间以及与DNA末端之间的结构和空间关系问题。 AIM 3检验了以下假设：NHEJ连接酶复合物对于在NHEJ期间建立关键的端到端接触很重要。 AIM 4描述了更强大的NHEJ重组的发展。在AIM 4中，该系统随后用于通过测试增强活动来识别任何剩余的NHEJ因素。 AIM 5描述了组蛋白八聚体如何影响NHEJ的核酸酶，聚合酶和连接酶活性的分析。这些研究将显着提高我们对NHEJ如何参与诸如染色体易位的过程，这些过程在癌症病因中至关重要，以及如何使NHEJ成为用于治疗目的的“吸毒”。