Rebecca Glynn (Mentor: Craig Bassing, PhD)

“Elucidating RAG DNA Cleavage Signals and Mechanisms that Feedback Inhibit V(D)J Recombination”

Rebecca Glynn, Katharina Hayer, Brittney Allyn and Craig Bassing

The creation of antigen receptors via RAG1/RAG2 (RAG) endonuclease-mediated assembly of antigen receptor genes must be regulated to prevent autoimmunity and oncogenic genomic lesions. We deduced that RAG DNA double strand breaks (DSBs) induced during Vkappa-to-Jkappa recombination in pre-B cells signal via the ATM tumor suppressor kinase to repress Rag1/Rag2 expression, Jkappa accessibility, and RAG cleavage of the other allele. We hypothesize that the most important role of DSB feedback inhibition of V recombination is to block oncogenic IgH and TCRbeta translocations. To test our model, we have begun to study effects of RAG DSBs generated during recombination of these loci. We find that RAG DSBs induced during IgH D-to-J recombination in pro-B cells or TCR rearrangements before Vbeta recombination in pro-T cells signal independent of ATM to decrease Rag1/Rag2 transcripts. TCR DSBs initiate a mainly ATM-independent transcriptional program. One of the transcripts most repressed by ATM is a Vbeta anti-sense long non-coding RNA that our lab discovered is expressed only in pro-T cells where Vbeta segments rearrange. Our findings imply that RAG DSBs feedback inhibit V(D)J recombination through complementary mechanisms, including DSB type-specific signals that repress Rag1/Rag2 transcription and locus-specific mechanisms that suppress synapsis of V and (D)J segments.