SOMATIC HYPERMUTATION OF THE NEW ANTIGEN RECEPTOR GENE (NAR) IN THE NURSE SHARK DOES NOT GENERATE THE REPERTOIRE - POSSIBLE ROLE IN ANTIGEN-DRIVEN REACTIONS IN THE ABSENCE OF GERMINAL-CENTERS

The new antigen receptor (NAR) gene in the nurse shark diversifies ext ensively by somatic hypermutation. It is not known, however, whether N AR somatic hypermutation generates the primary repertoire (like in the sheep) or rather is used in antigen-driven immune responses. To addre ss this issue, the sequences of NAR transmembrane (Tm) and secretory ( Sec) forms, presumed to represent the primary and secondary repertoire s, respectively, were examined from the peripheral blood lymphocytes o f three adult nurse sharks. More than 40% of the Sec clones but fewer than 11% of Tm clones contained five mutations or more. Furthermore, m ore than 75% of the Tm clones had few or no mutations. Mutations in th e Sec clones occurred mostly in the complementarity-determining region s (CDR) with a significant bias toward replacement substitutions in CD R1; in Tm clones there was no significant bias toward replacements and only a low level of targeting to the CDRs, Unlike the Tm clones where the replacement mutational pattern was similar to that seen for synon ymous changes, Sec replacements displayed a distinct pattern of mutati ons. The types of mutations in NAR were similar to those found in mous e Ig genes rather than to the unusual pattern reported for shark and X enopus Ig. Finally, an oligoclonal family of Sec clones revealed a str iking trend toward acquisition of glutamic/aspartic acid, suggesting s ome degree of selection. These data strongly suggest that hypermutatio n of NAR does not generate the repertoire, but instead is involved in antigen driven immune responses.