UC San Diego

A diverse antibody repertoire is the backbone of a successful adaptive immune response. This diversity is created in developing B cells by the rearrangement of the V (variable), D (diversity), and J (joining) gene segments, termed V(D)J recombination, to produce unique antigen specific B cell receptors (BCRs). V(D)J recombination is regulated by an ensemble of transcription factors, co- activators as well as architectural proteins. We know that certain V gene regions are more highly rearranged in wild type cells, yet the mechanism for this bias is not well known. We propose that the enhancer-associated E2A, p300 and BRD4 proteins influence the frequency and pattern of gene rearrangement. Using CRISPR/Cas9 technology, we have successfully deleted E2A as well as BRD4 to determine the effects of each mutation on the frequency and pattern of rearrangement in the immunoglobulin kappa locus (Igκ) locus of an inducible cell line. We have examined the consequences of these deletions on Vκ rearrangements. Notably, our results indicate that E2A and BRD4 are responsible for influencing the pattern and frequency of Vκ gene rearrangement. These data indicate that E2A and BRD4 are key factors in modulating nuclear architecture and the Igκ repertoire.