Monopalmitoylphosphatidylcholine incorporation into human erythrocyte ghost membranes causes protein and lipid immobilization and cholesterol depletion

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Abstract
The effects of lysophosphatidylcholine (lysoPC) on human erythrocyte (RBC) ghost morphology, transmembrane protein and lipid lateral mobilities, and membrane lipid composition were studied in order to elucidate mechanisms by which lysoPC immobilizes ghost membrane components [Golan, D. E., Brown, C. S., Cianci, C. M. L., Furlong, S. T., and Caulfield, J. P. (1986) J. Cell Biol. 103, 819-828]. Under standardized conditions 1.0-1.5 .mu.g/mL egg lysoPC lysed 50% of RBCs and induced, in some ghosts, the formation of large patches of wrinkled membrane. Patches exhibited complete immobilization of glycophorin and band 3 and partial immobilization of the phospholipid analogue fluorescein phosphatidylethanolamine (Fl-PE), whereas adjacent smooth membrane areas manifested only partial immobilization of proteins and no immobilization of Fl-PE. Supralytic concentrations of lysoPC induced both progressive, homogeneous wrinkling of RBC ghost membranes and concentration-dependent decreases in the lateral mobilities of glycophorin, band 3, and Pl-PE. Complete immobilization of glycophorin and band 3 occurred at 8.4 .mu.g/mL lysoPC and of Fl-PE at 16.8 .mu.g/mL lysoPC. Monopalmitoylphosphatidylcholine (MPPC), the major component of egg lysoPC, induced both membrane wrinkling and a concentration-dependent decrease in Fl-PE mobility, with complete immobilization at 10 .mu.g/mL. Other synthetic lysoPCs did not completely immobilize Fl-PE, although some caused membrane wrinkling. MPPC was incorporated into ghost membranes with a linear dependence (r = 0.97) on MPPC concentration. Relative to total membrane lipid, the lysoPC mole fraction increased from 0.2 .+-. 0.1% at 0 .mu.g/mL MPPC to 25 .+-. 2% at 16 .mu.g/mL MPPC. The molar ratio of cholesterol to phospholipid (exclusive of lysoPC) in MPPC-treated ghosts was inversely dependent on MPPC concentration, decreasing from 1.0 .+-. 0.1 at 0 .mu.g/mL MPPC to 0.7 .+-. 0.2 at 8 .mu.g/mL MPPC. This ratio did not decrease further at 12 and 16 .mu.g/mL MPPC. MPPC treatment did not affect the relative amounts of the major RBC phospholipid classes. These results suggest that MPPC causes concentration-dependent changes in the composition and organization of RBC ghost membranes. LysoPC incorporation and/or cholesterol depletion may induce lipid domain formation, which causes membrane protein and lipid immobilization.