Alternative End-Joining in DNA Repair and Chromosomal Translocations

DNA 修复和染色体易位中的选择性末端连接

• 批准号: 9099801
• 负责人: Vipul Kumar
• 金额: $ 3.16万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9099801
• 关键词: Antibodies; Antigen Receptors; B-Cell Lymphomas; B-Lymphocytes; BCL2 gene; Biological Assay; CRISPR/Cas technology; Cell Cycle; Cell Death; Cell Line; Cell Survival; Cell division; Cells; Chemicals; Chimeric Proteins; Chromosomal Breaks; Chromosomal Rearrangement; Chromosomal translocation; Chromosome Deletion; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Cytogenetics; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Sequence Rearrangement; Data; Detection; Double Strand Break Repair; Embryo; Exons; G1 Arrest; G22P1 gene; Gene Mutation; Gene Silencing; Generations; Genome; Genome Stability; Glucocorticoid Receptor; Health; Heavy-Chain Immunoglobulins; Immunoglobulin Class Switching; Immunoglobulin Constant Region; Immunoglobulin Switch Recombination; Immunoglobulin Variable Region; J segment gene; Lead; Ligation; Lymphocyte Activation; Lymphomagenesis; Malignant Neoplasms; Mature B-Lymphocyte; Mediating; Mus; Mutant Strains Mice; Nonhomologous DNA End Joining; Oncogenes; Oncogenic; Pathway interactions; Phase; Phosphotransferases; Poly(ADP-ribose) Polymerases; Proteins; Publishing; Reporter; Role; Site; Southern Blotting; System; Testing; Transgenes; V(D)J Recombination; XRCC1 gene; XRCC4 gene; XRCC5 gene; abl Oncogene; activation-induced cytidine deaminase; ataxia telangiectasia mutated protein; base; endodeoxyribonuclease SceI; endonuclease; genome-wide; homologous recombination; insight; interest; nuclease; p53-binding protein 1; repaired; research study; sensor

DESCRIPTION (provided by applicant): Improperly repaired DNA double-strand breaks (DSBs) can lead to cell death or chromosomal rearrangements that contribute to oncogenic transformation. The two major mammalian DSB repair pathways are homologous recombination (HR), which is active in post-replicative (S/G2) cells, and classical non-homologous end joining (C-NHEJ), which predominates in pre-replicative (G1) cells. For C-NHEJ, the Ku70/Ku80 heterodimer ("Ku") recognizes DSBs and the XRCC4/Ligase4 (Lig4) complex joins them. During V(D)J recombination in developing B lymphocytes, RAG endonuclease generated DSBs at V, D, and J gene segments are joined exclusively by C-NHEJ to assemble exons encoding antigen receptor/antibody variable regions. During immunoglobulin (Ig) heavy chain (IgH) class switch recombination (CSR) in activated mature B lymphocytes, activation-induced cytidine deaminase (AID)-initiated DSBs in IgH switch (S) regions are fused, predominantly by C-NHEJ, to exchange expressed IgH constant region exons. Aberrant joining of V(D)J- or CSR-associated DSBs can lead to chromosomal translocations that fuse antigen receptor loci to oncogenes, and, thereby, contribute to lymphomagenesis. In the absence of C-NHEJ, RAG- or AID-initiated IgH DSBs can be joined to form such oncogenic chromosomal translocations by an alternative end-joining (A-EJ) pathway (or pathways). In addition, CSR is carried out relatively robustly by A-EJ in the absence of Lig4, Ku, or both. Based on substantial preliminary data, we propose the A-EJ in the absence of Lig4 is distinct from that which occurs in the absence of Ku (or Ku plus Lig4). We refer to these two A-EJ pathways as "Lig4-independent" and "Ku- independent" A-EJ, respectively. We propose to elucidate DSB recognition and joining components of the two A-EJ pathways, their relative activity in the G1 cell cycle, and their relative contributions to normal versus aberrant end-joining that promotes chromosomal translocations. Specifically, we propose to 1) Identify factors that mediate Lig4- and/or Ku-independent A-EJ during CSR and 2) Assess potential roles of Lig4- and/or Ku- independent A-EJ in G1-arrested pro-B lines.

描述（由申请人提供）：修复不当的DNA双链断裂（DSB）会导致细胞死亡或染色体重排，从而导致致癌性转化。两个主要的哺乳动物DSB修复途径是同源重组（HR），它在复制后（S/G2）细胞中活跃，经典的非同源末端连接（C-NHEJ），占重复的（G1）细胞。对于C-NHEJ，ku70/ku80异二聚体（“ ku”）识别DSB和XRCC4/Ligase4（Lig4）复合物加入了它们。在V（d）J重组中，在发生B淋巴细胞中，C-NHEJ专门将抹布内切核酸酶在V，D和J基因段上产生的DSB，以组装编码抗原受体/抗体可变区域的外显子。在活化成熟的B淋巴细胞中的免疫球蛋白（IG）重链（IG）重链（CSR）中，激活诱导的胞苷Deaminase（AID）在IGH SWITCON中受IGH Switch的胞质DEAMINASE（AID）引起的DSB，主要由C-NHEJ融合，以C-NHEJ融合，以交换expresseans Expanded Consplacted Standart Expanted eigh eigh eigh eigh eigh eigh eigh。 V（d）J-或CSR相关的DSB的异常连接会导致染色体易位，将抗原受体基因座融合到癌基因中，从而有助于淋巴瘤。在没有C-NHEJ的情况下，可以通过替代的端连接（A-ej）途径（或途径）连接rag或AID引入的IGH DSB，以形成这种致癌染色体易位。另外，在没有Lig4，Ku或两者的情况下，A-EJ对CSR进行了相对牢固的稳定性。基于大量的初步数据，我们在没有LIG4的情况下提出A-EJ与在没有KU（或KU Plus Lig4）的情况下发生的A-EJ不同。我们将这两个A-EJ途径分别称为“ Lig4独立”和“ Ku-Independent” A-EJ。我们建议阐明DSB识别和连接两种A-EJ途径的组件，它们在G1细胞周期中的相对活性以及它们对促进染色体易位的正常和异常最终结合的相对贡献。具体而言，我们建议1）确定在CSR期间介导Lig4-和/或Ku独立的A-EJ的因素，以及2）评估Lig4-和/或Ku-ku-独立a-ej在G1删除Pro-B线中的潜在作用。