An Activation-Induced Cytidine Deaminase-Independent Mechanism of Secondary VH Gene Rearrangement in Preimmune Human B Cells

Abstract

VH replacement is a form of IgH chain receptor editing that is believed to be mediated by recombinase cleavage at cryptic recombination signal sequences (cRSS) embedded in VH genes. Whereas there are several reports of VH replacement in primary and transformed human B cells and murine models, it remains unclear whether VH replacement contributes to the normal human B cell repertoire. We identified VH→VH(D)JH compound rearrangements from fetal liver, fetal bone marrow, and naive peripheral blood, all of which involved invading and recipient VH4 genes that contain a cryptic heptamer, a 13-bp spacer, and nonamer in the 5′ portion of framework region 3. Surprisingly, all pseudohybrid joins lacked the molecular processing associated with typical VH(D)JH recombination or nonhomologous end joining. Although inefficient compared with a canonical recombination signal sequences, the VH4 cRSS was a significantly better substrate for in vitro RAG-mediated cleavage than the VH3 cRSS. It has been suggested that activation-induced cytidine deamination (AICDA) may contribute to VH replacement. However, we found similar secondary rearrangements using VH4 genes in AICDA-deficient human B cells. The data suggest that VH4 replacement in preimmune human B cells is mediated by an AICDA-independent mechanism resulting from inefficient but selective RAG activity.