Molecular analysis of Rh transcripts and polypeptides from individuals expressing the DVI variant phenotype: an RHD...

The D antigen is a mosaic comprising at least 30 epitopes. Partial Rh D phenotypes occur when there is absence of one or more of these epitopes, with the remainder expressed. The DVI phenotype is the most common of the partial D phenotypes, lacking most D antigen epitopes (ep D) (epD1, 2, 5-8 using the 9-epitope model or epD 1-4, 7-22, 26-29 using the 30-epitope model). DVI mothers may become immunized by transfusion with D-positive blood (if typed as D-positive using polyclonal typing reagents) or by fetuses which have all of the D antigen. This situation can give rise to severe hemolytic disease of the newborn (HDN). The molecular basis of the DVI phenotype has previously been proposed to occur by two different genetic mechanisms, one (in individuals of DVICcee phenotype) where a gene conversion event generates a hybrid RHD-RHCE-RHD gene; the second (in individuals of DVIccEe phenotype) was proposed to be caused by a partial RHD gene deletion. We present evidence that in four DVICcee phenotypes studied, this phenotype is not generated by a partial RHD gene deletion, but occurs by a similar mechanism to the DVICcee phenotypes. In two individuals we have found hybrid RHD-RHCE-RHD transcripts in both DVICe and DVIcE haplotypes. These differ in that the DVICe transcripts are derived from an RHD gene where exons 4-6 have been replaced with RHCE equivalents (encoding Ala226); the DVIcE transcripts are derived from an RHD gene where exons 4 and 5 are replaced by RHCE equivalents (encoding Pro226). We provide direct evidence that Rh DVI polypeptides are expressed at the erythrocyte surface as full-length polypeptide products. We have used immunoprecipitation experiments using anti-D reactive with DVI erythrocytes followed by immunoblotting the immune complexes with rabbit sera immunoreactive to the fourth external and C-terminal domains of all Rh polypeptides. Our results illustrate that these domains are present on all Rh DVI proteins studied, and suggest that Rh DVI polypeptide species studied here exist as full-length Rh proteins.