The proposed research is a study of an inherited hemolytic anemia, """"""""xerocytosis."""""""" Red blood cells from patients with this disease exhibit an accelerated passive K ion efflux resulting in cellular dehydration. Cellular 2,3-diphosphoglycerate levels are low despite the anemia. Intracellular pH and hemoglobin are normal, and no defect in metabolism has been established. Xerocyte membranes will be characterized to define the molecular basis for abnormal function. Parameters of characterization will be high-resolution analytic fractionation of membrane proteins, accessibility to surface labeling reagents, pattern of protein cross-linking, content of membrane-associated enzymes, susceptibility to peroxidative damage, and capacity to reseal. Aspects of membrane-cytoplasm interaction will be studied: The effects of normal and xerocyte cytoplasm on K ion passive efflux in resealed ghosts will be determined using exchange hemolysis. The role of hemoglobin in mediating spectrin complex formation in cells under peroxidative stress will be analyzed using cell-free mixtures of the purified proteins. Studies of membrane biogenesis in cultured erythroblasts and affinity-purified reticulocytes will emphasize the timing of synthesis of components 4.1a and 4.1b in relation to other membrane proteins. Patterns of membrane protein synthesis in erythroblasts derived from patients with xerocytosis will be compared to normal controls.
