CHARACTERIZATION OF THE GENE ENCODING THE HUMAN KIDD BLOOD-GROUP UREATRANSPORTER PROTEIN - EVIDENCE FOR SPLICE-SITE MUTATIONS IN JK(NULL) INDIVIDUALS

The Kidd (JK) blood group is carried by an integral membrane glycoprot ein which transports urea through the red cell membrane and is also pr esent on endothelial cells of the vasa recta in the kidney. The exon-i ntron structure of the human blood group Kidd/urea transporter gene ha s been determined, It is organized into 11 exons distributed over 30 k ilobase pairs. The mature protein is encoded by exons 4-11, The transc ription initiation site was identified by 5'-rapid amplification of cD NA ends-polymerase chain reaction at 335 base pairs upstream of the tr anslation start point located in exon 4, The 5'-flanking region, from nucleotide -837 to -336, contains TATA and inverted CAAT boxes as well as GATA-1/SP1 erythroid-specific cis-acting regulatory elements. Anal ysis of the S'-untranslated region reveals that the two equally abunda nt erythroid transcripts of 4.4 and 2.0 kilobase pairs arise from usag e of different alternative polyadenylation signals. No obvious abnorma lity of the Kidd/urea transporter gene, including the 5'- and 3'-untra nslated regions, has been detected by Southern blot analysis of the bl ood of two unrelated Jk(null) individuals (B.S. and L.P.), which lacks all Jk antigens and Jk proteins on red cells, but was genotyped as ho mozygous for a ''silent'' Jk(b) allele, Further analysis indicated tha t different splice site mutations occurred in each variant. The first mutation affected the invariant G residue of the 3'-acceptor splice si te of intron 5 (variant B.S.), while the second mutation affected the invariant G residue of the 5'-donor splice site of intron 7 (variant L .P.), These mutations caused the skipping of exon 6 and 7, respectivel y, as seen by sequence analysis of the Jk transcripts present in retic ulocytes, Expression studies in Xenopus oocytes demonstrated that the truncated proteins encoded by the spliced transcripts did not mediate a facilitated urea transport compared with the wild type Kidd/urea tra nsporter protein and mere not expressed on the oocyte's plasma membran e. These findings provide a rational explanation for the lack of Kidd/ urea transporter protein and defect in urea transport of Jk(null) cell s.