IGH DIVERSITY IN AN INDIVIDUAL WITH ONLY ONE MILLION B-LYMPHOCYTES

Immunoglobulin sequences from an individual Xenopus laevis froglet were analyzed for combinatorial and junctional diversity. In an animal with about 10(6) B lymphocytes, at least 26 out of the 56 V(H)1 genes available in a diploid genome were expressed, as were all JH segments. Junctional diversity was similar to that observed in Xenopus tadpole sequences, that is, little or no N diversification was found and the recombination site sometimes occurred in a region of V/D or D/J homology. The froglet IgH diversity is further restricted by the elimination of D-gene participation through direct V to J joining. Of the six complementary-determining regions (CDR) contributing to the structure of the antigen-combining site, CDR3 is the most variable in sequence and structure. Froglet IgH CDR3 are restricted to both aspects. Compared to IgH sequences isolated from a 5-month-old adult, froglet CDR3 were, on the average, two codons shorter; overall, 58% of the froglet Ig sequences isolated carried CDR3 of less-than-or-equal-to 7 codons, compared to 30% of the adult sequences. In addition to being shorter, the tadpole/froglet CDR3 are less variable in sequence, as the absence of N regions also results in the V/D and D/J junctions being derived from germline elements. We therefore suggest that latent anti-adult specificities are not eliminated in situ, in the tadpole, but rather that such germline gene segments, singly or in their combinations thereof, that can potentially react to adult self-epitopes after metamorphosis have been counterselected during the course of evolution.