Different Effects of Enzyme-generated Ceramides and Diacylglycerols in Phospholipid Membrane Fusion and Leakage

When large unilamellar vesicles consisting of sphingomyelin:phosphatidylethanolamine:cholesterol (2:1:1 molar ratio) are treated with sphingomyelinase, production of ceramides in the bilayer is accompanied by leakage of vesicle aqueous contents and by vesicle aggregation in the absence of lipid mixing or vesicle fusion. This is in contrast to the situation of phosphatidylcholine:phosphatidylethanolamine:cholesterol (2:1:1 molar ratio) liposomes when treated with phospholipase C. In that case, in situ generation of diacylglycerol leads to vesicle aggregation followed by vesicle fusion in the absence of leakage (Nieva, J. L., Goñi, F. M., and Alonso, A. (1989) Biochemistry 28, 7364-7367). Moreover, when ceramides (5-10 mol %) are included in the formulation of the phosphatidylcholine-containing vesicles, they reduce the lag time of phospholipase C-induced fusion, although they are less active than diacylglycerols in this respect. 31 P NMR studies of aqueous lipid dispersions show that diacylglycerols as well as ceramides induce a thermotropic lamellar to non-lamellar phase transition in both phospholipid:cholesterol mixtures under study although sphingomyelin-containing bilayers are more stable than those containing phosphatidylcholine, and ceramide is less active than diacylglycerol in promoting non-lamellar phase formation. These observations are relevant to both the physiological role of ceramides and the current views on the mechanism of membrane fusion.