Molecular mechanisms that lead to reduced expression of Duffy antigens

BACKGROUND: In the Duffy blood group system, the null phenotype Fy(a-b-) has been classically associated with a mutated GATA box, while the Fy(x) phenotype weak Fy^b is associated with Arg89Cys and Ala100Thr mutations. This report assesses the prevalence of the Duffy GATA box and the Fy(x)-associated mutations in white and African American (black) donors and investigates the molecular mechanism underlying the Fy(x) phenotype. STUDY DESIGN AND METHODS: PCR RFLP Duffy genotyping was performed on blood samples from blacks and whites. Duffy antigen expression (Fy^a Fy^b, Fy6, Fy3) on RBCs was measured by flow cytometry. By site-directed mutagenesis, the relevance of each Fy(x)- associated mutation to Duffy (mRNA, antigen, and protein) expression was analyzed in transfectants by Northern blotting, flow cytometry, and immunoblotting. RESULTS: The mutated GATA box occurred at a high allele frequency (0.8) in blacks and was rare among whites. Conversely, the Fy(x)- associated mutations were absent in blacks, but present in 3.5 percent of whites. By flow cytometry, Duffy antigens (Fy^a or Fy^b, Fy6 and Fy3) showed a dosage effect in RBC samples that were transcriptionally silenced by the GATA box mutation in one allele. By contrast, the reduced (10%) Duffy protein in Fy(x) RBCs was shown by heterologous expression analysis not to be due to reduced RNA levels, but to protein instability caused by Arg89Cys. CONCLUSIONS: Reduced Duffy expression can result from mutations affecting transcription (mutated GATA box in one allele) or instability of the translated protein (Arg89Cys). The frequencies of these mutations vary among populations.