A NOVEL MUTATION IN THE ERYTHROCYTE PROTEIN-4.2 GENE OF JAPANESE PATIENTS WITH HEREDITARY SPHEROCYTOSIS (PROTEIN-4.2(FUKUOKA))

Human erythrocyte protein 4.2 (band 4.2; pallidin) is a major membrane protein that comprises 5% of the total weight of the human erythrocyt e membrane. Deficiencies of this protein have been observed in heredit ary spherocytosis with anaemia, suggesting a role of protein 4.2 in er ythrocyte stability and integrity. The molecular basis of this disorde r remains unknown. As a first step in elucidating the pathogenesis of hereditary spherocytosis associated with protein 4.2 deficiency, we cl oned and sequenced the erythrocyte protein 4.2 gene from a normal Japa nese person. We prepared sets of oligonucleotide primers for polymeras e chain reaction (PCR) and determined nucleotide sequences of exons an d exon-intron boundaries of the protein 4.2 gene from three unrelated Japanese patients with hereditary spherocytosis due to a complete defe ct of protein 4.2, using PCR-related techniques. Two patients were hom ozygous for a missense mutation in codon 142 with the Ala (GCT) --> Th r (ACT) amino acid substitution that has been reported previously (pro tein 4.2(NIPPON)), whereas one patient was compound heterozygous for t he same missense mutation in codon 142 and a guanine-adenine transitio n in codon 119 that changes the codon for Trp (TGG) to the termination codon (TGA) (protein 4.2(Fukuoka)). No additional mutation was identi fied in other exons of the protein 4.2 genes. Dot-blot hybridization w ith allele-specific oligonucleotide probes showed that homozygosity fo r the missense mutation in codon 142 and compound heterozygosity for t he codon 142 and the codon 119 mutations were related to protein 4.2 d eficiency in the families. Although two alleles of missense mutation o f the codon 142 were also detected in 100 alleles of healthy Japanese, results obtained in this study indicate that the two mutations descri bed above are closely related to the pathogenesis of hereditary sphero cytosis due to protein 4.2 defect.