重组激活基因2(RAG2)在V(D)J重排规则建立中的功能与机制

在V(D)J重排中，小鼠RAG1/2（mmRAG1/2）介导非对称12/23RSS底物有序重排，此为“12/23规则”。ProtoRAG转座子（编码bbRAG1L和bbRAG2L）的发现直接支持V(D)J重排的RAG转座子起源假说，但“12/23重排规则”起源未明。类似V(D)J重排，bbRAG1L/2L介导TIR底物重排。前期研究提示mmRAG2维持RSS底物不对称性促进“12/23规则”建立，预实验却表明bbRAG1L/2L可介导对称TIR底物精确重排，分析bbRAG2L结构发现其与mmRAG2差异较大，突变后影响TIR底物重排。因此我们推测RAG2结构演化导致非对称RSS底物使用偏好而建立“12/23规则”。本项目拟研究RAG2在重排规则建立中发挥功能的结构基础，分析RAG2不同结构域相互嫁接后对“12/23规则”建立的影响，从而揭示RAG2在V(D)J重排规则建立中的功能与机制。

V(D)J rearrangement mediated by the mammal mouse RAG1/2 (mmRAG1/2) is characterized by orderly recognizing the asymmetric 12/23 RSS substrates, which is the “12/23 rule”. The discovery of ProtoRAG transposon (Encoding the bbRAG2L and bbRAG2L transposases) in lancelet Branchiostoma belcheri directly supports the RAG transposon origin hypothesis of V(D)J rearrangement. However, the origin of the keynote V(D)J rearrangement rule (“12/23 rule”) remains to be elucidated. .Being similar to V(D)J rearrangement, the bbRAG1L/2L mediates TIR substrates-dependent recombination. Previous study has suggested that the mmRAG2 plays a key role in the establishment of the “12/23 rule” by helping to maintain the preference for an asymmetrical pair of RSSs during the V(D)J rearrangement. However, our preliminary experiment suggested that, different from the “12/23 rule” of V(D)J rearrangement, the bbRAG1L/2L can precisely mediate symmetric TIR substrates-dependent recombination. Further structure analysis of the bbRAG2L suggested it is big different from that of mmRAG2. Also, bbRAG2L mutation would greatly affect the TIR substrates-dependent recombination mediated by bbRAG1L/2L. Based on those results, we speculate that the domain evolution of RAG2 leads to the usage preference of the asymmetric RSS substrates in V(D)J rearrangement, and the establishment of the “12/23 rule”. .Here, our project aims to identify the functional structures of RAG2 using the symmetric and asymmetric substrates, and their roles in the establishment of substrates recombination rules; and research on the influence of RAG2 domains grafting to the establishment of V(D)J recombination rules. This project is expected to reveal the roles and mechanisms of RAG2 in the establishment of V(D)J rearrangement rules.