Structure and Mechanism of Non-Homologous End Joining

非同源末端连接的结构和机制

• 批准号: 10331036
• 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/10331036
• 关键词: Adaptive Immune System; Address; Apoptosis; Architecture; Biochemical; Cancer Etiology; Cancerous; Catalytic Domain; Cell Cycle; Cells; Chemicals; Chromatin; Chromosomes; Complex; Coupling; Cryoelectron Microscopy; 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; Lead; Ligase; Ligation; Malignant Neoplasms; Mass Spectrum Analysis; Modeling; Molecular; Molecular Machines; Nonhomologous DNA End Joining; Nucleosomes; Pathway interactions; Phosphorylation; 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; XRCC4 gene; cancer therapy; crosslink; dimer; ds-DNA; improved; interdisciplinary approach; mechanical signal; novel; operation; particle; reconstitution; recruit; repaired; response; senescence

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.

总结 在各种类型的DNA损伤中，双链断裂被认为是毒性最大的DNA损伤 导致基因组不稳定。它们是由内源性物质如活性氧 物种，或外源性电离辐射和化学品。未修复的DSB驱动细胞凋亡和衰老， 不正确的DSB修复可导致不希望的基因组重排，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末端直接连接到DNA末端。 在不涉及同源模板的情况下连接，是保持活性的主要DSB修复途径 在整个细胞周期中。NHEJ还负责V（D）J重组中基因片段的组装， 其中通过免疫细胞中的外显子重组产生各种免疫球蛋白基因。NHEJ已初始化 通过识别DSB末端的Ku异二聚体（Ku 70/80）。在识别dsDNA断裂末端后，Ku 70/80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）的组分， 连接酶IV、XRCC 4和XLF）然后被募集到末端修复位点。通过NHEJ成功修复DSB 依赖于两个断裂的DNA末端的有效桥接，该提议旨在研究 NHEJ通过使用单粒子冷冻EM直接可视化修复的关键步骤。一张更精致的照片， 专门识别和纠正争端解决机构的系统将提供前所未有的全面视角， 这些基本的分子机器在运作过程中，并可能导致新的治疗方法的发展， 用于治疗各种人类癌症。