THE V(D)J RECOMBINATION REACTION AND ITS IMPACT ON LYMPHOCYTE DEVELOPMENT

描述（由申请人提供）：淋巴细胞发育受多种细胞信号调控，其中许多信号来自细胞表面受体。所有发育中的淋巴细胞必须组装编码B细胞受体（BCR）或T细胞受体（TCR）不同链的基因，并表达一种功能性的、非自身反应性的抗原受体，以离开胸腺（T细胞）或骨髓（B细胞）进入外周。抗原受体基因的组装是通过V(D)J重组过程进行的，当RAG蛋白在抗原受体位点的V、D和J基因片段引入DNA双链断裂（DSBs）时，就会启动V(D)J重组过程。这些DSB被DSB修复NHEJ通路的蛋白加工和连接，附加V、D和J基因片段，完成抗原受体基因的第二外显子。由基因毒性物质（如电离辐射）产生的dsb激活细胞DNA损伤反应，包括修复途径和促进细胞周期阻滞和细胞死亡的途径，如果dsb持续未修复。目前尚不清楚的是，生理性DNA断裂（如RAG dsb）是否也会激活广泛的DNA损伤反应，或者它们是否会被阻止，除非它们持续未修复，从而对细胞构成危害。本研究表明，在发育中的淋巴细胞中，RAG dsb激活了一种名为Atm的激酶，该激酶是DNA损伤反应信号的主要传感器，并通过参与包括STAT3、Foxo1和NF:B在内的多种转录途径，促进了广泛的遗传程序。由RAG dsb调控的几个基因编码在典型DNA损伤反应中起作用的蛋白质。然而，令人惊讶的是，很大一部分在DNA损伤反应中没有已知的功能。相反，这些基因编码的蛋白质在正常淋巴细胞发育过程中发挥重要作用。由此，我们假设RAG dsb提供了调节发育过程的信号，这些信号至少对发育淋巴细胞组装抗原受体基因很重要。在这里，我们将验证这一假设，阐明RAG dsb激活淋巴细胞发育中的转录途径的机制，并确定这些途径的功能如何与其他淋巴细胞发育信号相结合。

DESCRIPTION (provided by applicant): Lymphocyte development is regulated by a variety of cellular cues, many of which emanate from cell surface receptors. All developing lymphocytes must assemble genes encoding the different chains of the B cell receptor (BCR), or T cell receptor (TCR) and express a functional, non-auto-reactive, antigen receptor to exit the thymus (T cells) or bone marrow (B cells) and enter the periphery. Antigen receptor gene assembly occurs through the process of V(D)J recombination, which is initiated when the RAG proteins introduce DNA double strand breaks (DSBs) at V, D and J gene segments in antigen receptor loci. These DSBs are processed and joined by the proteins of the NHEJ pathway of DSB repair, appending the V, D and J gene segments to complete the second exon of antigen receptor genes. DSBs generated by genotoxic agents (such as ionizing radiation) activate cellular DNA damage responses including repair pathways and pathways that promote cell cycle arrest and cell death if the DSBs persist unrepaired. It has been unclear whether physiologic DNA breaks such as RAG DSBs also activate broad DNA damage responses or whether they are prevented from doing so unless they persist un-repaired, thus posing a hazard to the cell. Here, we show that in developing lymphocytes, RAG DSBs activate Atm, a kinase that is a major transducer of DNA damage response signals, and that this promotes a broad genetic program by engaging several transcriptional pathways including STAT3, Foxo1 and NF:B. Several of the genes regulated by RAG DSBs encode proteins that function in canonical DNA damage responses. Surprisingly however, a large fraction has no known function in DNA damage responses. Rather, the genes encode proteins that function in processes that are important for normal lymphocyte development. From this, we hypothesize that RAG DSBs provide signals that regulate developmental processes that are at a minimum, important for developing lymphocytes assembling antigen receptor genes. Here, we will test this hypothesis, elucidate the mechanisms by which RAG DSBs activate transcriptional pathways in developing lymphocytes and determine how the function of these pathways is integrated with other lymphocyte developmental signals.