Mechanisms of aberrant V(D)J recombination and its suppression

V(D)J recombination, the lymphocyte-exclusive process responsible for the diverse antigen receptor repertoires of vertebrates, occurs in two stages. In the first, the lymphoid specific RAG 1/2 complex recognizes and cleaves at conserved recombination signal sequences, producing double strand break (DSB) intermediates. In the second stage, the intermediates are joined by ubiquitously expressed nonhomologous end joining factors.; I look at three means of regulating V(D)J rearrangement that may be important for suppressing infrequent but dangerous aberrant V(D)J recombination. First, I show that lysine 4 on histone 3 is trimethylated (tri-Me-H3-K4) at Jx signal flanks destined for recombination, suggesting a role for histone modifications in accurately targeting RAG accessibility. Second, I observe the localization of the checkpoint protein ATM to DSB intermediates immediately after cleavage takes place. I also see enrichment for Serine 18-phosphorylated p53, a substrate of ATM, at DSB intermediates in cells deficient for nonhomologous end joining, supporting a model in which ATM triggers a checkpoint response only when intermediates are not destined to be resolved quickly. Third, I use an in vivo transposition assay to show transposition accompanies cellular V(D)J recombination, but may be redirected back to existing V(D)J recombination DSB intermediates. Thus, potentially oncogenic interchromosomal transposition events may be averted when the RAGS transpose DNA back into antigen receptor loci.