The complex of phosphatidylinositol 4,5-bisphosphate and calcium ions is not responsible for Ca2+-induced loss of phospholipid asymmetry in the human erythrocyte: a study in Scott syndrome, a disorder of calcium-induced phospholipid scrambling.
Elevation of cytoplasmic Ca2+ levels in human erythrocytes induces a progressive loss of membrane phospholipid asymmetry, a process that is impaired in erythrocytes from a patient with Scott syndrome. We show here that porcine erythrocytes are similarly incapable of Ca2+-induced redistribution of membrane phospholipids. Because a complex of phosphatidylinositol 4,5-bisphosphate (PIP2) and Ca2+ has been proposed as the mediator of enhanced transbilayer movement of lipids (J Biol Chem 269:6347,1994), these cell systems offer a unique opportunity for testing this mechanism. Analysis of both total PIP2 content and the metabolic-resistant pool of PIP2 that remains after incubation with Ca2+ ionophore showed no appreciable differences between normal and Scott erythrocytes. Moreover, porcine erythrocytes were found to have slightly higher levels of both total and metabolic-resistant PIP2 in comparison with normal human erythrocytes. Although loading of normal erythrocytes with exogenously added PIP2 gave rise to a Ca2+-induced increase in prothrombinase activity and apparent transbilayer movement of nitrobenzoxadiazolyl (NBD)-phospholipids, these PIP2-loaded cells were also found to undergo progressive Ca2+-dependent cell lysis, which seriously hampers interpretation of these data. Moreover, loading Scott cells with PIP2 did not abolish their impaired lipid scrambling, even in the presence of a Ca2+-ionophore. Finally, artificial lipid vesicles containing no PIP2 or 1 mole percent of PIP2 were indistinguishable with respect to transbilayer movement of NBD-phosphatidylcholine in the presence of Ca2+. Our findings suggest that Ca2+-induced redistribution of membrane phospholipids cannot simply be attributed to the steady-state concentration of PIP2, and imply that such lipid movement is regulated by other cellular processes.
Bevers
E M
EM
Department of Biochemistry, Cardiovascular Research Institute Maastricht, University of Limburg, The Netherlands.
Wiedmer
T
T
Comfurius
P
P
Zhao
J
J
Smeets
E F
EF
Schlegel
R A
RA
Schroit
A J
AJ
Weiss
H J
HJ
Williamson
P
P
Zwaal
R F
RF
Sims
P J
PJ
D009327:4-Chloro-7-nitrobenzofurazan; D002118:Calcium / Q000097:blood*; D004910:Erythrocyte Membrane / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism*; D006467:Hemophilia A / Q000097:blood*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008051:Lipid Bilayers; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000097:blood*; D010743:Phospholipids / Q000097:blood*; D012016:Reference Values; D013577:Syndrome; D013925:Thromboplastin / Q000096:biosynthesis
D008051:Lipid Bilayers; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010743:Phospholipids; D013925:Thromboplastin; D009327:4-Chloro-7-nitrobenzofurazan; D002118:Calcium