Abstract
Binding of ligands to the extracellular region of the erythrocyte transmembrane protein glycophorin A induces a decrease in membrane deformability. Since the property of membrane deformability is regulated by the skeletal proteins on the cytoplasmic side of the membrane, this suggests that ligand binding may initiate a transmembrane signal. To further study this process, we examined which domains of the extracellular region of glycophorin are involved in signal transduction and whether the cytoplasmic domain of the molecule is necessary for transmitting the signal. Using the ektacytometer, we compared the effect on deformability of four monoclonal antibodies that detect different epitopes on glycophorin A. We found that 9A3 (which recognized the amino terminus of glycophorin) caused a 5.8-fold increase in rigidity, R-10 and 10F7 (which recognized epitopes in the mid-region of the extracellular domain) caused a 10.8-fold increase in rigidity and B14 (which binds to glycophorin close to the membrane) caused a 18-fold increase in rigidity. Further, a direct relationship was observed between the degree of antibody-induced rigidity and the amount of glycophorin A that became associated with the skeletal proteins in a Triton shell assay. In Miltenberger V erythrocytes, which contain a hybrid sialoglycoprotein with no cytoplasmic domain, antibody binding did not induce an increase in rigidity. These results imply that glycophorin A is capable of a modulatable form of transmembrane signaling that is determined by the extracellular domain to which the ligand binds, and the cytoplasmic domain of glycophorin A is crucial for this process.