RAG-induced DNA damage: mechanisms and responses

项目摘要 V（D）J重组事件如何参与致癌染色体的产生 白血病和淋巴瘤的潜在易位许多提案已被 先进一些事件似乎涉及将RAG生成的DSB连接到 伴侣基因座的其他机制。几年前，我们提出错误的选择， 修复DSB的修复途径可能导致异常连接事件。我们最近的数据 揭示了RAG 1或RAG 2中的特定突变废除了途径选择， 在体外使切割后复合物不稳定。我们的初步数据显示 鼠淋巴瘤中RAG诱导的致癌易位的突变。这些相同 突变与体内淋巴细胞中增加的替代NHEJ相关。在一起， 这些数据支持以下假设，我们将在所提出的实验中检验这些假设。 假设1：“路径选择控制”在维持基因组稳定性方面起着关键作用， 在V（D）J重组中通过RAG切割后复合物维持。稳定性降低 RAG切割后复合物的增加（由许多原因中的任何一种引起）允许 编码和/或信号末端对不适当的连接途径的可用性， 形成异常V（D）J重组产物，包括致癌染色体 易位假设2：废除途径选择控制允许替代NHEJ 作为一种诱变修复途径出现。理解路径选择的控制和 在V（D）J重组的背景下，禁用这种调节机制的后果将 阐明了一个近30年来一直没有答案的问题： 发生在淋巴细胞发育过程中的易位我们所获得的知识 拟议的研究也可能有助于我们在其他情况下处理这一重要问题。如果， 在拟议的研究中，我们的RAG突变体解除了对通路选择的调节，使我们能够观察到 在完整的经典通路中频繁的致癌重排，我们将 能够建立替代NHEJ确实可以与经典加入竞争 机制等这一结果意味着，对途径选择的控制是由某些人强加的， 在不涉及RAG蛋白的其他DSB修复情况下，

Project Summary How are V(D)J recombination events involved in generating oncogenic chromosome translocations underlying some leukemias and lymphomas? Many proposals have been advanced. Some events appear to involve joining of RAG-generated DSBs to breaks made by other mechanisms at the partner loci. A few years ago, we proposed that errors the choice of repair pathway to heal the DSB could lead to aberrant joining events. Our recent data now reveal that specific mutations in either RAG1 or RAG2 abrogate pathway choice and also destabilize the post-cleavage complex in vitro. Our preliminary data implicate some of these mutations in RAG-induced oncogenic translocations in murine lymphomas. These same mutations are associated with increased alternative NHEJ in lymphocytes in vivo. Together, these data support the following hypotheses, which we will test in the proposed experiments. Hypothesis 1: "Pathway choice control" plays a critical role in maintaining genomic stability, and is maintained in V(D)J recombination by the RAG post-cleavage complex. Decreased stability of the RAG postcleavage complex (resulting from any of a number of causes) allows increased availability of the coding and/or signal ends to inappropriate joining pathways, facilitating formation of aberrant V(D)J recombination products, including oncogenic chromosome translocations. Hypothesis 2: Abrogating pathway choice control allows alternative NHEJ to emerge as a mutagenic repair pathway. Understanding control of pathway choice and the consequences of disabling this regulatory mechanism in the context of V(D)J recombination will illuminate a question that has remained unanswered for almost 30 years: how do the oncogenic translocations that occur in developing lymphocytes arise? The knowledge we gain from the proposed studies is also likely to help us approach this important question in other contexts. If, in the proposed studies, our RAG mutants which deregulate pathway choice allow us to observe frequent oncogenic rearrangements in the context of an intact classical pathway, we will be able to establish that alternative NHEJ can indeed compete with the classical joining mechanisms. This result would imply that control of pathway choice is imposed, by some mechanism, in other DSB repair situations that do not involve the RAG proteins.