Onset of Antibody Repertoire Diversification in the Rabbit Appendix (81.17)

Abstract

Abstract Unlike mice and humans, rabbits generate their primary antibody repertoire by somatically diversifying V(D)J genes in gut-associated lymphoid tissues (GALT). Rabbit V(D)J genes are diversified by somatic hyper-point mutation and somatic gene conversion, both of which require activation-induced cytidine deaminase (AID). To determine when repertoire diversification begins, we examined AID expression in neonatal rabbit appendix by in situ hybridization. We first detected AID mRNA expression at one week of age, in B cells in the basolateral region of appendix follicles. We asked whether AID+ B cells were undergoing V(D)J gene diversification by using laser capture microdissection (LCM) to isolate AID+ and AID- B cells from 1-week-old appendix follicles. We RT-PCR-amplified and cloned VDJ genes from the two B cell populations, and searched for evidence of somatic diversification by comparing their nucleotide sequences to those of the appropriate germline genes. We found somatic gene conversion events in the VDJ genes from AID+ B cells, but not in those from AID- B cells. These results demonstrate that diversification of the rabbit primary antibody repertoire begins around 1 week of age, in B cells residing at the basolateral region of appendix follicles. This work was supported by NIH Grant 1 RO1 AI49458-01A1.