MOLECULAR-BASIS OF BOVINE RED-CELL PROTEIN 4.2 POLYMORPHISM IN JAPANESE BLACK CATTLE

Cattle were divided into three groups according to the red cell-protei n 4.2 (P4.2) phenotypes P4.2(76), P4.2(75) and P4.2(76/75), whose red cells contained M-r 76000 (P4.2/76), 75000 (P4.2/75) and both 76000 an d 75000 isoforms respectively. To elucidate the molecular basis that u nderlies the diversity of P4.2, the gene structures of bovine P4.2/76 and P4.2/75 were investigated. Two P4.2 cDNA clones were isolated from bone-marrow cDNAs of the animal with the P4.2(76/75) phenotype. These were identical in size (2.2 kb), encoding major erythroid P4.2 with 6 87 amino acids, but were different in three nucleotides, resulting in changes of amino acids at the 599th, 601st and 627th residues. Analysi s of genomic DNA from the three phenotypes demonstrated that these two clones were derived from gene transcripts by which P4.2/76 and/or P4. 2/75 were produced. In vitro transcription and translation of P4.2/76 and P4.2/75 cDNAs indeed generated P4.2/76 and P4.2/75 identical in si ze to the red-cell proteins. These findings demonstrated that polymorp hism of the P4.2 gene at codons 599, 601 and 627 of P4.2 cDNA was the cause of the molecular diversity of bovine red-cell P4.2. Although dis tinct electrophoretic mobilities suggested a structural difference in the two isoforms, this polymorphism appeared to have little effect at least on P4.2 association with band 3, since no significant difference was observed in the amount of P4.2 relative to total membrane protein s despite the phenotype difference for P4.2.