LEVEL OF EXPRESSION OF PHOSPHOLIPID SCRAMBLASE REGULATES INDUCED MOVEMENT OF PHOSPHATIDYLSERINE TO THE CELL-SURFACE

We recently identified a 35-kDa erythrocyte membrane protein, phosphol ipid scramblase, that promotes Ca2+-dependent transbilayer movement of phosphatidylserine (PS) and other phospholipids (PL) in reconstituted proteoliposomes (Zhou, Q., Zhao, J., Stout, J. G., Luhm, R. A, Wiedme r, T., and Sims, P. J, (1997) J. Biol. Chen 272, 18240-18244). To dete rmine whether this same protein is responsible for the rapid movement of PS from inner-to-outer plasma membrane leaflets in other cells expo sed to elevated cytosolic calcium concentration ([Ca2+](c)), we analyz ed how induced movement of PS to the cell surface related to expressio n of PL scramblase. Exposure to Ca2+ ionophore A28187 resulted in rapi d PS exposure in those cell lines constitutively high in PL scramblase (HEL, Epstein-Barr virus-transformed B-lymphocytes, and Jurkat), wher eas this response was markedly attenuated in cells expressing low amou nts of this protein (Raji, HL60, and Dami). To confirm this apparent c orrelation between PL scramblase expression and PS egress at elevated [Ca2+](c), Raji cells were transfected with PL scramblase cDNA in pEGF P-C2, and stable transformants expressing various amounts of GFP-PL sc ramblase fusion protein were obtained. Clones expressing GFP-PL scramb lase showed distinctly plasma membrane-localized fluorescence. When co mpared either with untransfected Raji cells or with transformants expr essing GFP alone, clones expressing GFP-PL scramblase fusion protein s howed increased exposure of PS at the cell surface in response to elev ated [Ca2+](c), accompanied by increased expression of membrane cataly tic function for the prothrombinase enzyme complex, These data indicat e that transfection with PL scramblase cDNA promotes movement of PS to cell surfaces and suggest that this protein normally mediates redistr ibution of plasma membrane phospholipids in activated, injured, or apo ptotic cells.