Role of DNA Damage Responses in Immune Development and Function

项目摘要 淋巴细胞发育被精确控制，以使克隆扩增和产生多样化的细胞。 免疫球蛋白受体库，其通过DNA双链断裂（DSB）进行。这两 二分的，但相互依存的过程，是通过不同的细胞信号的合作， 阻止具有DSB的细胞进入细胞周期，在细胞周期中它们可以作为易位被异常修复。期间 在早期B细胞发育中，前B细胞受体（pre-BCR）通过激活SYK激酶， 前B细胞的增殖扩增和免疫球蛋白受体基因的组装。负 需要调节前BCR以强制细胞周期停滞并限制产生的DNA断裂的数量 免疫球蛋白受体基因组装过程中。事实上，无对抗的前BCR信号，特别是增加 SYK活性驱动增殖和白血病转化。我们发现了一种新的细胞类型特异性 由来自DSB的信号激活的检查点通路，其抑制前BCR信号传导。生理DSB 免疫球蛋白受体基因重排过程中产生的DNA损伤反应， 前BCR下游的SYK激酶活性。令人惊讶的是，这种信号网络不是由遗传毒性引发的。 DNA损伤，因此对正常B细胞分化过程中产生的DSB是特异性的。在早期的B细胞中， 对生理和遗传毒性DSB的不同细胞反应对于确保正常的B细胞是必需的 分化和抑制白血病转化。我们的目标是确定来自DSBs的信号 与发育程序整合以协调B细胞成熟。我们认为早期B细胞 在来自前BCR和DSB的信号之间切换以安排免疫球蛋白受体基因组装， 通过防止具有DSB的细胞增殖来维持基因组稳定性。利用创新的实验 的方法，允许隔离的DSB信号和表面受体信号，我们建议：1）定义 转录抑制因子复合物，调节DSB介导的检查点，2）确定DSB依赖的 抑制前BCR信号传导的翻译后途径，和3）确定DSB特异性途径， 决定了对生理性与遗传毒性损伤的独特细胞反应。完成这些研究将 描述一组抑制前BCR信号的关键机制，将为细胞- 类型特定的检查点DSB，并将定义离散细胞反应的基本原则， 生理性与遗传毒性DSB。

PROJECT SUMMARY Lymphocyte development is precisely controlled to enable clonal expansion and generation of a diverse immunoglobulin receptor repertoire, which proceeds through DNA double-stranded breaks (DSBs). These two dichotomous, but interdependent processes, are managed through the cooperation of diverse cellular signals to prevent cells with DSBs from entering cell cycle where they could be aberrantly repaired as translocations. During early B cell development, the pre-B cell receptor (pre-BCR), through activation of the SYK kinase, coordinates both the proliferative expansion of pre-B cells and the assembly of immunoglobulin receptor genes. Negative regulation of the pre-BCR is required to enforce cell cycle arrest and limit the number of DNA breaks generated during immunoglobulin receptor gene assembly. Indeed, unopposed pre-BCR signaling, particularly increased SYK activity, drives proliferation and leukemic transformation. We have identified a novel cell-type specific checkpoint pathway activated by signals from DSBs that inhibits pre-BCR signaling. The physiologic DSBs generated during immunoglobulin receptor gene rearrangement trigger DNA damage responses that suppress SYK kinase activity downstream of the pre-BCR. Surprisingly, this signaling network is not triggered by genotoxic DNA injury and, thus, is specific to the DSBs generated during normal B cell differentiation. In early B cells, distinct cellular responses to physiologic and genotoxic DSBs are essential for ensuring normal B cell differentiation and inhibiting leukemic transformation. Our goal is to determine how signals from DSBs integrate with developmental programs to coordinate B cell maturation. We propose that early B cells toggle between signals from the pre-BCR and DSBs to order immunoglobulin receptor gene assembly and maintain genomic stability by preventing proliferation of cells with DSBs. Utilizing an innovative experimental approach that permits isolation of DSB signals and surface receptor signals, we propose to: 1) define the transcriptional repressor complex that regulates DSB-mediated checkpoints, 2) identify the DSB-dependent post-translational pathways that suppress pre-BCR signaling, and 3) determine the DSB-specific pathways that dictate the unique cellular responses to physiologic versus genotoxic insults. Completion of these studies will delineate a set of mechanisms critical for dampening pre-BCR signals, will establish novel paradigms for cell- type specific checkpoints to DSBs, and will define the underlying principles for discrete cellular responses to physiologic versus genotoxic DSBs.