MULTIPLE SITES OF V-LAMBDA DIVERSIFICATION IN CATTLE

Ig repertoire diversification in cattle was studied in the ileal Peyer 's patch (IPP) follicles of young calves and in the spleens of late fi rst-trimester bovine fetuses, To investigate follicular diversificatio n, individual IPP follicles were isolated by microdissection; V lambda diversity was examined by RT-PCR and subsequent cloning and sequencin g. When 52 intrafollicular sequences from a 4-wk-old calf were determi ned and compared, two major groups, one of 23 members and the other of 25, could he delineated. An examination of these groups revealed clea r genealogic relationships that implicated in situ diversification of V lambda sequences within the confines of an IPP follicle, V lambda ex pression was also examined in early (95 and 110 gestational day fetal bovine spleens, Although earlier studies in cattle and sheep implicate d the IPP as a likely site of Ab diversification, a close investigatio n of V lambda sequences in late first-trimester fetal calves revealed that diversity appears in the early fetal spleen before the establishm ent of a diverse repertoire in the ileum. When the sequences for the f etal spleen were compared with an existing pool of germline sequences, we found evidence of possible gene conversion events and possible unt emplated point mutations occurring in sequences recovered from fetal s pleens. We conclude that IPP is not the sole site of V lambda diversif ication in cattle, Also, as suggested for rabbits, cattle may use both gene conversion and untemplated somatic point mutation to diversify t heir primary V lambda repertoire.