Structure and Mechanism of Non-Homologous End Joining

非同源末端连接的结构和机制

• 批准号: 10546447
• 负责人: Yuan He
• 金额: $ 34.56万
• 依托单位: NORTHWESTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-11 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10546447
• 关键词: Adaptive Immune System; Address; Apoptosis; Architecture; Biochemical; Cancer Etiology; Cancerous; Cell Cycle; Cells; Chemicals; Chromatin; Chromosomes; Complex; Coupling; Cryoelectron Microscopy; Cytoprotection; DNA; DNA Damage; DNA-PKcs; DNA-dependent protein kinase; Development; Double Strand Break Repair; Exons; Filament; G22P1 gene; Genes; Genetic Recombination; Genome; Genome Stability; Genomic Instability; Goals; Holoenzymes; Human; Immune; Immunoglobulin Genes; In Vitro; Ionizing radiation; LIG4 gene; Ligase; Ligation; Malignant Neoplasms; Mass Spectrum Analysis; Modeling; Molecular; Molecular Machines; Nonhomologous DNA End Joining; Nucleosomes; PIK3CG gene; Pathway interactions; Phosphotransferases; Physiological; Process; Reactive Oxygen Species; Regulation; Resolution; Role; Signal Transduction; Site; Structure; Structure-Activity Relationship; Synapses; System; Techniques; Testing; Time; Transcription Initiation; V(D)J Recombination; Visualization; XRCC4 gene; cancer therapy; crosslink; dimer; ds-DNA; improved; interdisciplinary approach; mechanotransduction; novel; operation; particle; presynaptic; reconstitution; recruit; repaired; response; senescence; tumor

Summary Among all different types of DNA damages, double strand breaks (DSBs) are viewed as the most toxic ones that lead to genome instability. They are created by either endogenous agents such as reactive oxygen species, or exogenous ionizing radiation and chemicals. Unrepaired DSBs drive apoptosis and senescence, and incorrect DSB repair can lead to undesired genome rearrangements, such as deletions, translocations, and fusions. Non-homologous end-joining (NHEJ) pathway, in which the two broken DNA ends are directly ligated without referring to a homologous template, is the primary DSB repair pathway that remains active throughout the cell cycle. NHEJ is also responsible for the assembly of gene segments in V(D)J recombination, where various immunoglobulin genes are generated by exon recombination in immune cells. NHEJ is initialized by Ku heterodimer (Ku70/80) recognizing DSB ends. Upon recognizing a dsDNA broken end, Ku70/80 recruits the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and assembles into the so-called DNA-PK holoenzyme. Other evolutionarily conserved NHEJ factors, including components of the ligase complex (DNA ligase IV, XRCC4 and XLF) are then recruited to the end reparation site. Successful DSB repair through NHEJ relies on the efficient bridging of two broken DNA ends, and this proposal aims to investigate the mechanism of NHEJ by directly visualizing the key steps of repair using single-particle cryo-EM. A more refined picture of the system specifically recognizing and correcting DSBs will provide an unprecedented, comprehensive view of these essential molecular machines during operation, and could lead to the development of novel treatments for various types of human cancer.

概括 在所有不同类型的 DNA 损伤中，双链断裂 (DSB) 被认为是毒性最强的损伤 导致基因组不稳定。它们是由内源性物质（例如活性氧）产生的 物种或外源性电离辐射和化学物质。未修复的 DSB 会导致细胞凋亡和衰老， 不正确的 DSB 修复可能导致不希望的基因组重排，例如缺失、易位、 和融合。非同源末端连接（NHEJ）途径，其中两个断裂的DNA末端直接连接 不参考同源模板进行连接，是保持活性的主要 DSB 修复途径 整个细胞周期。 NHEJ 还负责 V(D)J 重组中基因片段的组装， 其中各种免疫球蛋白基因是通过免疫细胞中的外显子重组产生的。 NHEJ 已初始化 通过 Ku 异二聚体 (Ku70/80) 识别 DSB 末端。在识别出 dsDNA 断裂末端后，Ku70/80 重新招募 DNA 依赖性蛋白激酶催化亚基 (DNA-PKcs) 并组装成所谓的 DNA-PK 全酶。其他进化上保守的 NHEJ 因子，包括连接酶复合物（DNA 连接酶 IV、XRCC4 和 XLF) 然后被募集到末端修复位点。通过 NHEJ 成功修复 DSB 依赖于两个断裂 DNA 末端的有效桥接，该提案旨在研究其机制 NHEJ 通过使用单粒子冷冻电镜直接可视化修复的关键步骤。更精致的图片 专门识别和纠正 DSB 的系统将提供前所未有的全面视图 这些在操作过程中必不可少的分子机器，可能会导致新疗法的开发 用于各种类型的人类癌症。