Molecular analysis of the human immunoglobulin V lambda II gene family and its relation to 8.12 idiotypic specificity

Abstract

The 8.12 idiotype characterizes a subpopulation of anti-DNA antibodies in patients with systemic lupus erythematosus (SLE). The idiotype is present exclusively on various light chains encoded by members of the V{dollar}\lambda{dollar}II gene family. Southern blot analysis has shown that the V{dollar}\lambda{dollar}II family consists of 10 to 15 members, of which only a few germline genes have been reported. To identify the V{dollar}\lambda{dollar}II genes that encode positive antibodies and to further characterize the V{dollar}\lambda{dollar}II family, we constructed two libraries from a patient with SLE. We have identified 13 germline genes, where 7 are novel, two of which are pseudogenes. We observed that the V{dollar}\lambda{dollar}II family can be subdivided in three groups, two encode both autoantibodies and antibodies to foreign antigens, the third may represent pseudogenes.;A comparison of V{dollar}\lambda{dollar}II germline genes to sequences of 8.12 positive light chains produced by EBV transformed B cell lines, shows that all 8.12 light chains are expressed by five highly homologous members of the V{dollar}\lambda{dollar}II family. 8.12 negative V{dollar}\lambda{dollar}II light chains derive from the same limited subset of V{dollar}\lambda{dollar}II genes, suggesting that only a small number of the available V{dollar}\lambda{dollar}II genes are commonly expressed.;We have utilized an eukaryotic expression system to identify the structural basis for 8.12 expression. Reversion of the 8.12{dollar}\sp+{dollar} DSC light chain to the hslv215.23/DPL11 germline gene sequence identifies the presence of the 8.12 idiotype in the germline. To localize important amino acids in the 8.12 idiotype, point mutations were performed in CDR1, CDR2, FR3 and CDR3, in positions where the 8.12{dollar}\sp+{dollar} DSC differs from the 8.12{dollar}\sp-{dollar} AS17 (V{dollar}\lambda{dollar}II light chain). Mutational analysis indicates that amino acids in CDR1 and the CDR2 proximal region of FR3, but not CDR3, play a crucial role in 8.12 reactivity. Molecular modelling reveals an exterior surface where those residues involved in 8.12 specificity are contiguous.