Relationship of phospholipid distribution to shape change in Ca(2+)-crenated and recovered human erythrocytes

Echinocytosis induced by elevation of intracellular Ca2+ in human erythrocytes can be reversed by removal of the cation. Using back-extraction of radiolabeled dilauroyl phospholipid analogs which had been incorporated into the cell membrane, we examined the relationship between this reversible shape transformation and phospholipid distribution. Upon Ca2+ crenation of cells, surface exposure of phosphatidylserine and phosphatidylethanolamine was observed simultaneously with inward diffusion of phosphatidylcholine. Removal of Ca2+ allowed resequestration of exposed phosphatidylserine to the membrane inner monolayer, but randomized phosphatidylethanolamine and phosphatidylcholine were not redistributed to their original states. Both shape reversion and retranslocation of phosphatidylserine were reversibly inhibited vanadate. On the other hand, the cell shape recovery was found to be independent of membrane skeleton and phosphoinositide metabolism and was supported by ATP resynthesis only under conditions where the aminophospholipid translocator is active. Other Ca(2+)-mediated biochemical changes, such as generation of diacylglycerol and fatty acids, were found to have no effect on Ca2+ crenation or its reversal, or upon transbilayer distribution of any phospholipid. These findings suggest that Ca2+ induces phospholipid redistribution, possibly by direct interaction with the lipid bilayer and, further, that metabolic recovery from Ca2+ crenation reflects selective retransport of phosphatidylserine to the membrane inner monolayer.