All affected patients in four families with autosomal dominant familial renal tubular acidosis (dRTA) were heterozygous for mutations in their red cell HCO 3 Ϫ /Cl Ϫ exchanger, band 3 ( AE1, SLC4A1 ) genes, and these mutations were not found in any of the nine normal family members studied. The mutation Arg 589 → His was present in two families, while Arg 589 → Cys and Ser 613 → Phe changes were found in the other families. Linkage studies confirmed the co-segregation of the disease with a genetic marker close to AE1 . The affected individuals with the Arg 589 mutations had reduced red cell sulfate transport and altered glycosylation of the red cell band 3 N-glycan chain. The red cells of individuals with the Ser 613 → Phe mutation had markedly increased red cell sulfate transport but almost normal red cell iodide transport. The erythroid and kidney isoforms of the mutant band 3 proteins were expressed in Xenopus oocytes and all showed significant chloride transport activity. We conclude that dominantly inherited dRTA is associated with mutations in band 3; but both the disease and its autosomal dominant inheritance are not related simply to the anion transport activity of the mutant proteins. ( J. Clin. Invest. 1997.

IntroductionProtein 4.2 is a major constituent of the red blood cell (RBC) membrane skeletal network, present at about 200 000 copies per RBC. 1 The protein-4.2 gene EPB42 contains 13 exons. 2,3 There are two isoforms of protein 4.2, a minor 74-kDa isoform obtained when all these exons of the gene are expressed and a 72-kDa major isoform of protein 4.2 that lacks 30 of the 33 amino acids that are encoded by exon 1. 4,5 The exact role of protein 4.2 in RBCs has not been elucidated, but protein 4.2 binds to the N-terminal cytoplasmic domain of the band-3 anion exchanger (AE1) and also interacts with ankyrin in RBCs. 6,7 The presence of band 3 is critical for the stable incorporation of protein 4.2 into the RBC membrane, since human, 8 mouse, 9 and cow 10 RBCs deficient in band 3 are also completely deficient in protein 4.2. The functional significance of the association of protein 4.2 with band 3 in RBCs is currently unclear. In liposomes containing reconstituted band 3, anion transport activity decreased in the presence of increasing amounts of protein 4.2, 11 which suggested that protein 4.2 was a negative modulator of band-3 anion exchange activity. However, band-3-mediated anion transport activity has been reported to be unaffected 12 or increased 13 in human RBCs with protein-4.2 deficiency. In contrast, the absence of protein 4.2 slightly decreased anion transport activity in protein-4.2-null mouse RBCs. 14 Protein 4.2 also associates with spectrin, 15,16 ankyrin, 7 and protein 4.1 7 in solution, although the significance of these associations in vivo has yet to be demonstrated. Recently, protein 4.2 has been shown to interact with CD47, which probably contributes to the anchoring of the Rh complex to the RBC skeleton. 17,18 Protein 4.2 can probably interact with the inner leaflet of the lipid bilayer since it is fatty acylated with myristoyl and palmitoyl chains. 19,20 The complete or nearly complete absence of protein 4.2 is associated with an atypical form of hereditary spherocytosis (HS), highlighting the important role 4.2 plays in maintaining the stability and flexibility of RBCs. To date, 9 protein-4.2 mutations have been found associated with HS. Five lead to the premature termination of translation. 17,[21][22][23][24] Other protein-4.2 variants result from missense mutations that yield amino acid substitutions: protein 4.2 Nippon (GCTϾACT; A142T 25 ; occurs sporadically in the Japanese population and has been encountered once in whites), 26 protein 4.2 Tozeur (CGAϾCAA; R310Q), 27 protein 4.2 Shiga (CGCϾTGC; R317C), 28 and protein 4.2 Komatsu (GATϾTAT; D175Y). 29 Although there is a strict correlation between the occurrence of HS and the presence of these mutations (in homozygous or compound heterozygous states), the fact that these mutations are the direct cause of the absence of protein 4.2 has not been formally established. One possibility is that these point mutations affect the binding of protein 4.2 to band 3 and therefore its function. This has proved difficult to study using biochemical We h...

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