Dr(a-) polymorphism of decay accelerating factor. Biochemical, functional, and molecular characterization and production of allele-specific transfectants.

Abstract

The Dr' antigen belongs to the Cromer-related blood group system, a series ofantigens on decay accelerating factor (DAF), a glycosyl-phosphatidylinositol-anchored membrane protein that protects host cells from complement-mediated damage. We studied the rare inherited Dr(a-) phenotype to ascertain the associated biochemical and functional changes in DAF and to characterize the basis for this polymorphism. Radioimmunoassay and flow cytometric analysis ofDr(a-) erythrocytes demonstrated 40% of normal surface expression of DAF but normal levels of several other glycosyl-phosphatidylinositol-anchored proteins, distinguishing this phenotype from that of paroxysmal nocturnal hemoglobinuria. Western blots confirmed this reduced DAF expression and indicated a slightly faster mobility of the molecule on SDS-PAGE. Despite the reduced DAF expression, Dr(a-) erythrocytes functioned normally in the complement lysis sensitivity assay. Utilization of the polymerase chain reaction to amplify mononuclear cell genomic DNA from three unrelated Dr(a-) individuals demonstrated that a point mutation underlies the Dr(a-) phenotype: a C to T change in nucleotide 649 resulting in a serine'65 to leucine change. This defines the Drb allele ofDAF, which can be distinguished from Dr' by a Taq I restriction fragment length polymorphism. We created transfected Chinese hamster ovary cell lines expressing either the Dr' or the Dr' allelic form of DAF.These allele-specific transfectants were tested by inhibition of hemagglutination or flow cytometry and confirmed the specificity of anti-Dr' alloantisera. The allele-specific transfectants could form the basis of a new serological approach to immunohematology. (J. Clin. Invest. 1991.87:1945-1952

Short consensus repeat-3 domain of recombinant decay-accelerating factor is recognized by Escherichia coli recombinant Dr adhesin in a model of a cell-cell interaction.

“…4). This shows that our previous finding (12) of a lack of reactivity of the Dr adhesin with Drb erythrocytes (that express only one-third the normal levels of DAF) was due to the absence of an adhesin binding site rather than to the lower expression of DAR The Dr' and Drb change does not appear to affect the complement regulatory function of DAF (30), so the adhesin and complement binding sites may not be the same.…”

“…Thus, the adhesin binding site on DAF is similar, but distinct, for the four adhesins of the Dr adhesin family, and involves SCR3 as well as SCR2 in some cases . We had investigated previously a rare polymorphism of human DAF, designated Dr(a-), and we found that it arose from a point mutation that changed a single amino acid in SCR3: Sertbs i n the high-frequency Dr-allele to Leu in the rare Drb [Dr(a-)] allele (30). We also found that erythrocytes of the Drb phenotype did not bind Dr adhesin (12).…”

“…3). Furthermore, a single-point substitution in SCR3, Sertbs to Leu, corresponding to the Dr' to Drb allelic polymorphism (30), caused complete abolition of adhesin binding to DAF (Fig. 4).…”

Structure-Function Analysis of Decay-Accelerating Factor: Identification of Residues Important for Binding of theEscherichia coliDr Adhesin and Complement Regulation

“…The classical pathway convertase and the Dr adhesin-binding regions are both localized on the SCR-2-SCR-3 interface of DAF, suggesting a possible overlap of binding regions. A rare polymorphism of human DAF containing a point mutation in SCR-3 (Ser165-Leu) (32,39), designated Dr(a Ϫ ), abolishes binding of Dr adhesins (40).…”

Dr^a (Cromer-Related Blood Group Antigen)-Incompatible Renal Transplantation

“…The very rare Dr(a–) phenotype is a partial DAF deficiency phenotype as a result of alternative splicing of DAF mRNA. Despite the reduced DAF expression, Dr(a–) erythrocytes function normally in the complement lysis sensitivity assay [6]. The inherited Dr(a–) phenotype has been found in Israel only in 4 families who immigrated from the area of Bukhara [7, 8].…”