Expression of C antigen in transduced K562 cells

BACKGROUND: The Rh blood group system is involved in HDN and transfusion re actions. A retrovirus-expression system was previously used to show that po lypeptides carrying the Rh blood group antigens are encoded by the RHD and RHCE genes. This study investigated the structure of the C antigen. STUDY DESIGN AND METHODS: K562 cells were transduced with full-length cDNA encoding Ce and CE antigens, and the expression of C, e, and E antigens was examined by flow cytometry using MoAbs. The importance of Cys16 in C antig en expression was examined by utilizing site-directed mutagenesis to conver t Cys16 to Trp in cDNA encoding Ce and CE before expression in K562 cells. RESULTS: When K562 cells were transduced with cDNA that was predicted to en code Ce antigens, clear reactivity with anti-e and anti-C was obtained. In contrast, K562 cells transduced with cDNA that was predicted to encode CE a ntigens gave strong reactivity with anti-E but failed to react with two exa mples of anti-C. A third example of anti-C gave weak reactivity. When cDNA encoding Ce antigens was mutated to encode Trp16, one example of anti-C had the same reactivity with the mutated polypeptide as with the wild-type mol ecule, but reactivity with two other anti-C examples was reduced by 50 perc ent. CONCLUSIONS: The nature of polymorphic residue 226 (proline when E is expre ssed, alanine when e is expressed) has a marked effect on the epitopes reco gnized by the three C MoAbs studied. The presence of Cys16 in Ce polypeptid es influences the presentation of the C epitope recognized by two of the th ree MoAbs. These experiments provide the first direct demonstration that C and E/e antigens can be expressed on the same polypeptide.