Alternative End-Joining in DNA Repair and Chromosomal Translocations

描述（由申请方提供）：DNA双链断裂（DSB）修复不当可导致细胞死亡或染色体重排，从而导致致癌转化。两种主要的哺乳动物DSB修复途径是同源重组（HR），其在复制后（S/G2）细胞中是活跃的，以及经典的非同源末端连接（C-NHEJ），其在复制前（G1）细胞中占主导地位。对于C-NHEJ，Ku 70/Ku 80异二聚体（“Ku”）识别DSB，并且XRCC 4/连接酶4（Lig 4）复合物将它们连接。在发育中的B淋巴细胞中的V（D）J重组期间，RAG核酸内切酶在V、D和J基因区段处产生的DSB仅通过C-NHEJ连接以组装编码抗原受体/抗体可变区的外显子。在活化的成熟B淋巴细胞中免疫球蛋白（IG）重链（IgH）类别转换重组（CSR）期间，IgH转换（S）区中活化诱导的胞苷脱氨酶（AID）启动的DSB主要通过C-NHEJ融合，以交换表达的IgH恒定区外显子。V（D）J或CSR相关DSB的异常连接可导致染色体易位，将抗原受体基因座融合到癌基因，从而促进淋巴瘤发生。在不存在C-NHEJ的情况下，RAG-或AID-起始的IgH DSB可以通过替代末端连接（A-EJ）途径（或多个途径）连接以形成这样的致癌染色体易位。此外，在不存在Lig 4、Ku或两者的情况下，通过A-EJ相对稳健地进行CSR。基于大量的初步数据，我们提出了在Lig 4的情况下的A-EJ不同于在Ku的情况下（或Ku加Lig 4）发生的A-EJ。我们将这两种A-EJ途径分别称为“Lig 4-非依赖性”和“Ku-非依赖性”A-EJ。我们建议阐明DSB的识别和连接组件的两个A-EJ途径，它们在G1细胞周期中的相对活性，以及它们对正常与异常末端连接，促进染色体易位的相对贡献。具体而言，我们建议1）鉴定在CSR期间介导Lig 4-和/或Ku-非依赖性A-EJ的因子，和2）评估Lig 4-和/或Ku-非依赖性A-EJ在G1-停滞的pro-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.