Trafficking and folding defects in hereditary spherocytosis mutants of thehuman red cell anion exchanger

Hereditary spherocytosis (HS) is a common inherited hemolytic anemia caused by mutations in erythrocyte proteins including the anion exchanger, AE1 (b and 3). This study examined seven missense mutations (L707P, R760Q, R760W, R808C, H834P, T837M, and R870W) located in the membrane domain of the human AE1 that are associated with this disease, The HS mutants, constructed in full-length AE1 cDNA, could be transiently expressed to similar levels in H EK 293 cells. Immunofluorescence, cell surface biotinylation, and pulse cha se labeling showed that the HS mutants all exhibited defective cellular tra fficking from the endoplasmic reticulum to the plasma membrane, Impaired bi nding to an inhibitor affinity matrix indicated that the mutant proteins ha d non-native structures and may be misfolded, Further characterization of t he HS R760Q mutant showed no change in its oligomeric structure or turnover (half-life =15 h) compared to wild-type AE1, suggesting the mutant was not aggregated or targeted for rapid degradation via the proteasome. Intracell ular retention of HS mutant AE1 would lead to destruction of the protein du ring erythroid development and would account for the lack of HS mutant AE1 in the plasma membrane of the mature red cell.