ATM FUNCTION DURING V(D)J RECOMBINATION

摘要 淋巴细胞抗原受体基因的组装通过V(D)J重组反应进行。这种反应 当形成RAG内切酶的RAG-1和RAG-2蛋白引入两个DNA时启动 两个重组基因片段及其相关的V、D或J片段边界的双链断裂 基因片段。这导致在每个DSB处形成编码端和信号端。编码结束 必须连接以形成编码连接，并且信号末端连接以形成信号连接。此外，作为一名单曲 在发育中的B和T细胞中，每个人一小时可以产生高达6000万克的DSB，途径必须是 防止未修复的碎布DSB被异常修复的地方，形成潜在的变形 染色体缺失和易位。DNA DSB修复的非同源末端连接途径 加入由碎屑切割产生的DNA双链断裂。NHEJ依赖于四个核心NHEJ因子，XRCC4，DNA连接酶 IV、库70和库80。然而，在NHEJ介导的过程中，还有其他非核心的NHEJ因子发挥作用 碎布DSB的修复。例如，编码联接形成依赖于ncNHEJ因子Artemis，它是 打开由碎屑切割产生的发夹密封的编码端的内切酶。不需要Artemis 用于连接信号端。此外，还有其他ncNHEJ因素本身并不是RAG所必需的 DSB修复，但可能影响RAG DSB修复效率或防止RAG异常修复的功能 未有效连接的DSB。确定ncNHEJ因素一直具有挑战性，因为它们的删除 通常不会导致碎石DSB的修复出现明显的缺陷。然而，最近的研究表明 确定假定的ncNHEJ因子的组合缺陷可导致RAG中可证明的缺陷 DSB修复。这是一种强大的基于候选人的方法来确定几个ncNHEJ因素和 阐明它们在NHEJ介导的DSB修复和DNA损伤反应(DDR)中的作用机制。 事实上，使用这种方法，我们已经确定了一种新的ncNHEJ蛋白，它是逆转录病毒的调节器 感染(MRI)，通过其N-末端和C-末端与一组功能不同的NHEJ和DDR结合 蛋白质。我们认为，在NHEJ介导的RAG修复的几个步骤中，MRI起着适配器的作用 DSB和在DDR期间对DSB进行碎裂。在这里，我们提出了一种新的无偏筛选方法来识别 在RAG DSB修复过程中起作用的ncNHEJ因子网络。我们还建议澄清 这些ncNHEJ因子的作用机制最初集中在新的ncNHEJ因子MRI上，即我们 最近发现它在破烂的DSB修复中起作用。这些研究将揭示 NHEJ介导的RAG双链断裂修复。这些活动可能与其他组织中的NHEJ大体相关 以及我们对DNA修复缺陷的理解，这些缺陷导致了癌症的基因组进化。

Abstract Lymphocyte antigen receptor gene assembly occurs through the V(D)J recombination reaction. This reaction is initiated when the RAG-1 and RAG-2 proteins, which form the RAG endonuclease, introduces two DNA double strand breaks (DSBs) at the border of two recombining gene segments and their associated V, D or J gene segments. This results in the formation of a coding end and a signal end at each DSB. The coding ends must be joined to form a coding join and the signal ends joined to form a signal join. Also, as a single individual can make up to 60,000,000 RAG DSBs in an hour in developing B and T cells, pathways must be in place to prevent un-repaired RAG DSBs from being aberrantly repaired, forming potentially transforming chromosomal deletions and translocations. The non-homologous end-joining pathway of DNA DSB repair joins DNA DSBs generated by RAG cleavage. NHEJ relies on four core NHEJ factors, XRCC4, DNA Ligase IV, Ku70 and Ku80. However, there are other non-core NHEJ factors that function during the NHEJ-mediated repair of RAG DSBs. For example, coding join formation relies on the ncNHEJ factor, Artemis, which is endonuclease that opens the hairpin-sealed coding ends generated by RAG cleavage. Artemis is not required for joining signal ends. In addition, there are other ncNHEJ factors that are not themselves required for RAG DSB repair, but that likely effect the efficiency of RAG DSB repair or function to prevent aberrant repair of RAG DSBs that are not joined efficiently. Identifying ncNHEJ factors has been challenging given that their deletion does not often lead to a demonstrable defect in the repair of RAG DSBs. However, recent studies have established that combined deficiencies of putative ncNHEJ factors can lead to demonstrable defects in RAG DSBs repair. This has been a powerful candidate-based approach for identifying several ncNHEJ factors and elucidating their mechanisms of action in NHEJ-mediated DSB repair and DNA damage responses (DDR). Indeed, using this type of approach, we have identified a novel ncNHEJ protein, modulator of retroviral infection (MRI), that binds through its N- and C-terminus to a functionally diverse set of NHEJ and DDR proteins. We believe that MRI functions as an adaptor during several steps of NHEJ-mediated repair of RAG DSBs and during the DDR to RAG DSBs. Here we propose a novel unbiased screening approach to identify the network of ncNHEJ factors that function during RAG DSB repair. We also propose to elucidate the mechanisms of action of these ncNHEJ factors focusing initially on the novel ncNHEJ factor, MRI, that we recently discovered that functions in RAG DSB repair. These studies will reveal novel activities required for the NHEJ-mediated repair of RAG DSBs. These activities will likely be generally relevant to NHEJ in other tissues and to our understanding to DNA repair defects that lead to genome evolution in cancers.