Violation of the 12/23 rule of genomic V(D)J recombination is common in lymphocytes

Abstract

V(D)J genomic recombination joins single gene segments to encode an extensive repertoire of antigen receptor specificities in T and B lymphocytes. This process initiates with double-stranded breaks adjacent to conserved recombination signal sequences that contain either 12-or 23-nucleotide spacer regions. Only recombination between signal sequences with unequal spacers results in productive coding genes, a phenomenon known as the ''12/23 rule.'' Here we present two novel genomic tools that allow the capture and analysis of immune locus rearrangements from whole thymic and splenic tissues using second-generation sequencing. Further, we provide strong evidence that the 12/23 rule of genomic recombination is frequently violated under physiological conditions, resulting in unanticipated hybrid recombinations iñ 10% of Tcra excision circles. Hence, we demonstrate that strict adherence to the 12/23 rule is intrinsic neither to recombination signal sequences nor to the catalytic process of recombination and propose that nonclassical excision circles are liberated during the formation of antigen receptor diversity.

V(DD)J recombination is an important and evolutionarily conserved mechanism for generating antibodies with unusually long CDR3s

“…Although the 12/23 rule explains the mechanism of VDJ recombination, tandem V(DD)J recombination (also known as tandem fusion) represents an exception to this rule (Parkinson et al 2015). The question about the molecular mechanism leading to tandem fusions of D genes has remained opened since the discovery of tandem fusions thirty years ago (Meek et al 1989).…”