Two-dimensional gel comparisons of murine H-2D region-associated antigens of different H-2 haplotypes.

The heterogeneity of histocompatibility antigens encoded within the H-2D region of murine chromosome 17 was investigated in several mouse strains using monoclonal antibodies. Sequential immunoprecipitation and two dimensional gel electrophoresis (2D) were used to determine the number and structural relatedness of antigenically distinct products of the same H-2D region. Strikingly different, haplotype-specific patterns were observed. The three antigenically distinct molecules, Dd, Ld and Rd, all encoded within the Dd region, each showed a unique constellation of spots by 2D analysis, although the Ld and Rd molecules showed more structural similarity to each other than either molecule showed to the Dd molecule. Comparisons of the Ld and Rd molecules using sequential precipitations and a cloned Ld gene suggested that they are not conformation or glycosylation variants of the same polypeptide, implying they have different primary amino acid sequences. Using an anti-Ld antibody, the Ldml molecule was isolated from cells of the Dd region mutant mouse strain B10.D2-H-2dml and was found to have a 2D gel profile clearly distinguishable from the profile of either Ld or Dd molecules isolated from the wild-type strain B10.D2. Of the three antigenically-distinct molecules determined by the Dq region, the 2D gel profile of the Rq molecule was different from that of the Dq and Lq, the latter two molecules being indistinguishable. In contrast to these findings, analysis of the gene products of the Db region failed to resolve either antigenic or molecular heterogeneity and the multiple, antigenically distinct gene products of the Dw16 region were indistinguishable by 2D comparisons. These haplotype specific variations in the structural relatedness and number of antigenically distinct molecules determined by D region genes support the hypothesis that class I MHC genes are in a dynamic evolutionary state of gene expansion and contraction.