SAFA promotes RAG1 aggregation in nucleoli and regulates expression of RAG and DNA-repair-associated factors to impair V(D)J recombination

B cell receptor (BCR) and T cell receptor (TCR) diversity results from V(D)J recombination in developing B and T lymphocytes. V(D)J recombination is mediated by RAG, and includes breaking and repairing DNA. To explore regulation mechanism of V(D)J recombination, we used proximity dependent biotin identification (BioID) to identify biomolecules associated with RAG. We found scaffold attachment factor A (SAFA) is biotinylated by RAG1-BirA*, which implies that SAFA may combine with RAG. GST Pull-down and co-IP assays show that SAFA interacts directly with RAG1 rather than RAG2. SAFA promotes nucleolar aggregation of RAG1, and impairs RAG-mediated plasmid recombination. In chromatin immunoprecipitation-seq assays, SAFA is revealed to enrich genes such as RAG1, X-ray repair cross complementing 5 (XRCC5) and LIG4, etc. Through upregulating expression of these genes, decreased SAFA boosts Gleevec-induced V(D)J recombination in preB cells. Taken together, these results suggest that SAFA can enhance nucleolar condensation of RAG1 and regulate expression of RAG and DSB repair-associated factors to down-regulate V(D)J recombination.