Diversification of the primary antibody repertoire begins during early follicle development in the rabbit appendix

Abstract

Rabbits generate a diversified primary antibody repertoire by somatically mutating, in gut-associated lymphoid tissue (GALT), an initial repertoire that is limited by preferential rearrangement of the 3′-most IGVH gene segment. To determine when repertoire diversification begins in GALT, we performed in situ hybridization on neonatal rabbit appendix sections with an activation-induced cytidine deaminase (AID) riboprobe, because AID is required for the mutational processes that diversify the primary antibody repertoire. We first detected AID mRNA expression around 1 week of age, in the basal region of developing follicles. By PCR-amplifying V–D–J genes from AID mRNA+ B cells isolated by laser capture microdissection, we found evidence of somatic hypermutation, and one likely instance of somatic gene conversion. Our results suggest that V–(D)–J gene diversification begins during early postnatal appendix development, in B cells stimulated to enter a proliferative program by signals derived from select intestinal commensals.