Mutation at amino acid position 133 of H-2D^d prevents β₂m association and immune recognition but not surface expression

The histocompatibility loss mutation H-2^dm6 was derived from a mouse treated with the chemical mutagen ethylnitrosourea, and previous mapping studies implicated D^d as the affected locus. Southern blot analyses of DNA from H-2^dm6 cells did not detect major deletions in the D^dm6 gene, suggesting that H-2^dm6 was different from the previously characterized D region mutants H-2^dm1 and H-2^dm2. RNA blot analysis identified D^dm6 transcripts of appropriate size and a D^dm6 protein was immunoprecipitated from biosynthetically labeled H-2^dm6 cells. Interestingly, the D^dm6 protein showed no β₂m association and was only precipitated by a mAb to the α3 domain. Furthermore, oligosaccharide maturation and low levels of surface expression of D^dm6 molecules were detected. However, the surface D^dm6 was nonfunctional as a target Ag in in vitro cytotoxicity assays, consistent with its original in vivo detection as a loss mutation. Sequence analyses of D^dm6 cDNA identified a single nucleotide base difference from wild-type, resulting in the substitution of a Trp to Arg at position 133. The significance of this substitution is discussed in the context of other class I expression variants.