Rag mutations reveal robust alternative end joining

Mammalian cells repair DNA double-strand breaks (DSBs) through either homologous recombination or non-homologous end joining (NHEJ). V(D)J recombination, a cut-and-paste mechanism for generating diversity in antigen receptors, relies on NHEJ for repairing DSBs introduced by the Rag1-Rag2 protein complex. Animals lacking any of the seven known NHEJ factors are therefore immunodeficient(1). Nevertheless, DSB repair is not eliminated entirely in these animals: evidence of a third mechanism, 'alternative NHEJ', appears in the form of extremely rare V(D)J junctions(2-4) and a higher rate of chromosomal translocations(5,6). The paucity of these V(D)J events has suggested that alternative NHEJ contributes little to a cell's overall repair capacity, being operative only (and inefficiently) when classical NHEJ fails. Here we find that removing certain portions of murine Rag proteins reveals robust alternative NHEJ activity in NHEJ-deficient cells and some alternative joining activity even in wildtype cells. We propose a two-tier model in which the Rag proteins collaborate with NHEJ factors to preserve genomic integrity during V(D)J recombination.