FUNCTIONAL FACTORS IN THE RED-CELL MEMBRANE - INTERACTIONS BETWEEN THE MEMBRANE AND ITS UNDERLYING SKELETON

Recent studies involving two abnormal red cell phenotypes (South-east Asian ovalocytosis and Leach phenotype) provide novel information conc erning the nature and significance of interactions of both the anion t ransport protein AE-1 (syn. band 3) and Glycophorins C and D with the underlying skeleton. The location of Wr(a) and Di(a) blood group antig ens to mutations on AE-1 at residues 658 and 854 respectively, togethe r with the availability of monoclonal antibodies recognising epitopes dependent upon the integrity of the third extracellular loop of AE-1, have allowed us to study the organisation of the membrane domain of th e mutant AE-1 found in South-east Asian ovalocytes (AE-1 SAG). The res ults suggest that the organisation of the whole membrane domain of AE- 1 SAO is abnormal and that the organisation of other integral membrane proteins like those involved in expression of Rh blood group antigens may also be affected. Increased homo- and hetero-associations involvi ng AE-1 SAO and other integral proteins may in turn result in reduced membrane flexibility. Purified protein 4.1 binds with 50-fold higher a ffinity to protein 4.1 depleted normal red cell membranes than to prot ein 4.1 depleted red cell membranes of Leach phenotype which lack Glyc ophorin C (GPC) and Glycophorin D (GPD). Experiments using purified pr otein 4.1 and p55 together with synthetic peptides corresponding to di fferent regions of the cytoplasmic domain of Glycophorins C and D (GPC /D) demonstrate that protein 4.1 interacts directly with GPC through r esidues 82-98. They also show that p55 binds to GPC through residues 1 12-128. Since p55 also binds directly to protein 4.1 it is clear that protein 4.1 can bind to GPC through two different sites either directl y through residues 82-98 or indirectly through p55. These results show that GPC and GPD provide major attachment sites for the red cell skel eton via protein 4.1 and that p55 is part of this complex.