Immunoglobulin gene diversification and B-cell ontogeny in cattle
The anatomical compartment within which the B-cell develops is different in different animals. This study attempts to trace B-cell ontogeny in cattle by investigating the expression profile of B-cell specific genes in cattle. Bovine fetal spleen and liver are the site of lymphopoeisis as both RAG1 and RAG2 could be detected at 60 days of gestation by RT-PCR. VpreB and heavy chain genes were also detected at 60 days of gestation, but no light chain expression could be found. TdT expression could not be detected and analysis of expressed light chain genes revealed lack of any junctional diversity in the expressed sequences, suggesting that N-nucleotide addition has no role in diversification of the primary repertoire in cattle.
Most mammals express two Ig light chain isotypes—lambda and kappa. Cattle has long been known to produce only the lambda light chain. This study attempts to characterize the kappa light chain expression in cattle. While kappa light chain could be detected at both the transcript and protein levels, the number of B-cells expressing kappa light chain in the periphery was limited and varied from animal to animal. The diversity of the expressed kappa light chain was determined by RT-PCR cloning and sequencing. The expressed kappa light chain genes show very limited—about half the diversity as compared to the lambda light chain. The complexity of kappa light chain genes present in the cattle genome is comparable to that of the lambda light chain as determined by southern blot analysis of genomic DNA indicating similar number of genes in both the light chain loci. The cloning of kappa germline genes revealed that the nanomer sequence of the RSS elements consistently have a substitution at the fourth base position and the heptamer sequence is also not always conserved. The kappa J/C intronic region was also cloned and sequenced. The conserved enhancer sequence, found in most mammals, was partly missing as only two of the three enhancer elements could be detected.