TRANSFUSION MEDICINE Molecular heterogeneity of the Jknull phenotype: expression analysis of the Jk(S291P) mutation found in Finns

Polymerase chain reaction genotyping of 32 unrelated Jknull individuals originating predominantly from Polynesia and Finland indicated that all were homozygous for the JK*B polymorphism and that 17 of 32, including the 14 Polynesians, carried a 3*-acceptor splice site mutation of intron 5 that resulted in the skipping of exon 6 (called mutation JkD6). The remaining 15 Jknull donors from Finland were homozygous for a new T871C transition resulting in a S291P amino acid substitution at a consensus N-glycosylation site of the Jk polypeptide. Transcriptiontranslation assays revealed that the Jk(S291P) mutant was translated into a glycosylated component as efficiently as the wild-type Jk polypeptide (wt Jk)] in the presence of microsomes, thus indicating that the S291P mutation has no effect on the N-glycosylation pattern of the Jk protein. Expression studies in Xenopus oocytes revealed that the Jk(S291P) polypeptide functions as a urea transporter, but the transport activity and the membrane expression level of the mutant protein was reduced to a similar extent. A substantial fraction of the mutant protein was retained intracellularly suggesting that the transit to the plasma membrane was reduced, presumably because of the S3P mutation. After transfection in erythroleukemia K562 cells the wild-type, but not the mutant, protein was efficiently expressed at the cell surface. Because the Jk(S291P) mutant polypeptide was not present in human red cells from Jknull individuals, expression data in the erythroid context clearly indicates that the S3P mutation is the molecular basis of the Finnish Jknull phenotype. (Blood. 2000; 96:1566-1573)