MODIFYING THE SEQUENCE OF AN IMMUNOGLOBULIN V-GENE ALTERS THE RESULTING PATTERN OF HYPERMUTATION

Affinity maturation of antibodies requires localized hypermutation and antigen selection. Hypermutation Is particularly active in certain re gions (notably the CDRs of light and heavy chains) due to the local ac cumulation of hot spots. We have now analyzed the role of individual n ucleotides in the origin of hot spots and show that mutability is larg ely defined by the nucleotide sequence, We compared the mutability pro file of wild-type and modified kappa transgenes that contain silent mu tations in the CDR1 segment, We found a new hot spot created at the th ird base of Ser-31 when its wild-type AGT codon aas substituted by AGC . Two major hot spots associated with this AGC vanished when Ser-31 wa s encoded by the synonymous TCA, In addition to these, which were the most prominent changes, there were compensatory alterations in mutabil ity of residues not directly related to the introduced silent mutation s, so that the average hypermutation remained constant, Thus, mutation s arising early in the immune response, even silent ones, could affect the mutability of critical residues and alter the pattern of affinity maturation. When analyzing hybridomas, we detected such alterations, but they seemed to better correlate with changes in average rather tha n local mutation rates. Overall, this paper shows how evolution could have optimized the mutability of individual residues to minimize delet erious mutations. Thus, the optimal strategy for affinity maturation m ay involve the incorporation of multiple point mutations before antige n selection of the relevant cells.