Alternative End-Joining in DNA Repair and Chromosomal Translocations

描述（由申请人提供）：修复不当的DNA双链断裂（DSBs）可导致细胞死亡或染色体重排，从而导致致癌转化。哺乳动物DSB的两种主要修复途径是同源重组（homologous recombination， HR）和经典非同源末端连接（non-homologous end joining, C-NHEJ），前者活跃于复制后（S/G2）细胞，后者主要存在于复制前（G1）细胞。对于C-NHEJ， Ku70/Ku80异源二聚体（“Ku”）识别dsb， XRCC4/Ligase4 （Lig4）复合物连接它们。在发育中的B淋巴细胞中，在V(D)J基因重组过程中，RAG内切酶在V、D和J基因片段上生成dsb，由C-NHEJ单独连接，组装编码抗原受体/抗体可变区的外显子。在活化的成熟B淋巴细胞的免疫球蛋白（Ig）重链（IgH）类开关重组（CSR）过程中，激活诱导的胞苷脱氨酶（AID）启动的IgH开关(S)区域的dsb主要由C-NHEJ融合，以交换表达的IgH恒定区外显子。V(D)J-或csr相关的dsb的异常连接可导致染色体易位，使抗原受体位点与癌基因融合，从而导致淋巴瘤发生。在缺乏C-NHEJ的情况下，RAG-或aids启动的IgH dsb可以通过另一种末端连接（A-EJ）途径（或多种途径）连接形成这种致癌染色体易位。此外，在没有Lig4、Ku或两者同时存在的情况下，A-EJ可以相对稳健地执行CSR。基于大量的初步数据，我们提出不含Lig4时的A-EJ与不含Ku（或Ku + Lig4）时的A-EJ不同。我们将这两种A-EJ途径分别称为“lig4独立”和“Ku独立”的A-EJ。我们建议阐明两种A-EJ通路的DSB识别和连接成分，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末端连接的相对贡献。具体来说，我们建议1)确定在CSR过程中介导Lig4-和/或Ku-不依赖的A-EJ的因子；2)评估Lig4-和/或Ku-不依赖的A-EJ在g1捕获的前b系中的潜在作用。

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.