The RAG2 acidic hinge regulates pathway choice, maintains genomic stability, and prevents lymphomagenesis (P4398)

Abstract

Abstract VDJ recombination-associated DNA double-stranded breaks are normally repaired by the high fidelity, classical NHEJ machinery. Previous studies have implicated the RAG/DNA post-cleavage complex (PCC) in regulating pathway choice by preventing access to inappropriate alternative repair mechanisms such as HR and alternative NHEJ (aNHEJ). We report that RAG2’s acidic hinge is critical for several steps. Mutations reducing the negative charge in this region destabilize the PCC, disrupt pathway choice, permit repair of RAG-mediated DSBs by the low-fidelity, translocation-prone aNHEJ machinery, and reduce genomic stability in developing lymphocytes. Structural predictions and experimental data support our hypothesis that flexibility of the acidic hinge is necessary for these functions. Similar human RAG2 sequence variants alter the predicted flexibility and permit repair by aNHEJ, indicating the potential for hazardous physiological consequences. Mice bearing similar neutralizing mutations (RAG2Neut), in the absence of p53, have an increased incidence of lymphomagenesis with hallmarks of VDJ recombination errors. Thus, the acidic hinge, along with stabilizing the PCC and ensuring proper pathway choice, aids in the prevention of lymphomagenesis by maintaining genomic integrity. This novel regulatory role that helps to ensure the safe development of a diverse immune repertoire, is to the best of our knowledge, the first function ascribed to this conserved region of RAG2.