The Phospholipid Flippase in Isolated Hog Gastric Vesicles: Evidence for Translocation of Endogenous Glycerophospholipids

The gastric flippase has been found recently in isolated hog gastric vesicles. We found here that the flippase ATP-dependently translocates endogenous phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine from the outer (cytosolic) to inner (luminal) leaflet of the vesicle membrane bilayer or to the vesicle interior. In the experiments, vesicles were incubated in the presence and absence of ATP, and change in the distribution of endogenous phospholipids between the outer and inner leaflets was determined. Furthermore, the vesicle diameter was measured by a quasi-elastic laser light scattering method. The diameter significantly decreased during the translocation, indicating that the glycerophospholipids are translocated into the vesicle interior. The decrease in the size depended on ATP hydrolysis and was inhibited by flippase inhibitors such as 2-methyl-8(phenylmethoxy)imidazo[1,2-a ]pyridine-3-acetonitrile (SCH 28080) and 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). The gastric flippase, which moves into the apical membrane of the parietal cell upon stimulation, may be a candidate that provides the cytoprotective hydrophobic barrier of adsorbed monolayer on the apical membrane covering the Phospholipid bilayer and the luminal surface of transmembrane proteins.