Molecular cloning of human plasma membrane phospholipid scramblase. A protein mediating transbilayer movement of plasma membrane phospholipids.
The rapid movement of phospholipids (PL) between plasma membrane leaflets in response to increased intracellular Ca2+ is thought to play a key role in expression of platelet procoagulant activity and in clearance of injured or apoptotic cells. We recently reported isolation of a approximately 37-kDa protein in erythrocyte membrane that mediates Ca2+-dependent movement of PL between membrane leaflets, similar to that observed upon elevation of Ca2+ in the cytosol (Bassé, F., Stout, J. G., Sims, P. J., and Wiedmer, T. (1996) J. Biol. Chem. 271, 17205-17210). Based on internal peptide sequence obtained from this protein, a 1,445-base pair cDNA was cloned from a K-562 cDNA library. The deduced "PL scramblase" protein is a proline-rich, type II plasma membrane protein with a single transmembrane segment near the C terminus. Antibody against the deduced C-terminal peptide was found to precipitate the approximately 37-kDa red blood cell protein and absorb PL scramblase activity, confirming the identity of the cloned cDNA to erythrocyte PL scramblase. Ca2+-dependent PL scramblase activity was also demonstrated in recombinant protein expressed from plasmid containing the cDNA. Quantitative immunoblotting revealed an approximately 10-fold higher abundance of PL scramblase in platelet ( approximately 10(4) molecules/cell) than in erythrocyte ( approximately 10(3) molecules/cell), consistent with apparent increased PL scramblase activity of the platelet plasma membrane. PL scramblase mRNA was found in a variety of hematologic and nonhematologic cells and tissues, suggesting that this protein functions in all cells.
Zhou
Q
Q
Blood Research Institute, The Blood Center of Southeastern Wisconsin, Milwaukee, Wisconsin 53201-2178, USA.
Zhao
J
J
Stout
J G
JG
Luhm
R A
RA
Wiedmer
T
T
Sims
P J
PJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D002352:Carrier Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D003001:Cloning, Molecular / Q000379:methods; D017931:DNA Primers; D004910:Erythrocyte Membrane / Q000378:metabolism*; D018922:HL-60 Cells / Q000201:enzymology; D006367:HeLa Cells / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D008051:Lipid Bilayers / Q000378:metabolism*; D008563:Membrane Lipids / Q000097:blood*; D008565:Membrane Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D049294:Phospholipid Transfer Proteins; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D017931:DNA Primers; D008051:Lipid Bilayers; D008563:Membrane Lipids; D008565:Membrane Proteins; D049294:Phospholipid Transfer Proteins; D011994:Recombinant Proteins