RAG1 and RAG2 form a stable postcleavage synaptic complex with DNA containing signal ends in V(D)J recombination

Abstract

During V(D)J recombination, RAG1 and RAG2 cleave DNA adjacent to highly conserved recombination signals, but nothing is known about the protein-DNA complexes that exist after cleavage. Using a properly regulated in vitro V(D)J cleavage system, together with nuclease sensitivity, mobility shift, and immunoprecipitation experiments, we provide evidence that a stable complex is formed postcleavage between synapsed recombination signals. This complex includes the proteins RAG1, RAG2, HMG-1 or the closely related HMG-2 protein, and the components of the DNA-dependent protein kinase. The existence of such a stable complex explains a number of in vivo observations and suggests that remodeling of postcleavage synaptic complexes is an important step in the resolution of signal ends in V(D)J recombination.