Phosphatidylinositol 4,5-bisphosphate domain inducers promote phospholipid transverse redistribution in biological membranes

Transmembrane phospholipid redistribution (scrambling), leading to exposure of phosphatidylserine on the cell surface, plays a physiological role to induce platelet procoagulant activity and clearance of injured or apoptotic cells. Scrambling is generally attributed to an increase in intracellular Ca2+ and would be mediated by a protein (scramblase), whose activity could be modulated by cofactors. We reported previously that phosphatidylinositol 4,5-bisphosphate (PIP2) is a positive regulator of Ca2+-induced Scrambling, We show here, using inside-out vesicles from erythrocyte membranes, that a pleckstrin homology (PH) domain, which interacts with high affinity with PIP2, inhibited Ca2+-induced scrambling, confirming the role of PIP2. As Ca2+ is known to interact with PIP2 and to promote the formation of lateral domains of acidic phospholipids in membranes, we investigated whether PIP2 domain formation could be involved in scrambling. Spermine, polylysine, and MARCKS (151-175) peptide caused scrambling in parallel to their reported ability to form domains of acidic phospholipids, including PIP2. Similarly, neomycine, another PIP2-interacting polycation, induced scrambling. A PIP2 antibody was also found to induce scrambling, presumably by a similar mechanism, since phospholipid antibodies are known to promote phospholipid capping. In conclusion, Ca2+ is not the sole inducer of scrambling, and formation of PIP2 domains could play a critical role in this process.