Different sphingolipids show differential partitioning into sphingolipid/cholesterol-rich domains in lipid bilayers

Two fluorescence-based approaches have been applied to examine the differen tial partitioning of fluorescent phospho- and sphingolipid molecules into s phingolipid-enriched domains modeling membrane "lipid rafts." Fluorescence- quenching measurements reveal that N-(diphenylhexatrienyl)propionyl- (DPH3: 0-)-labeled gluco- and galactocerebroside partition into sphingolipid-enric hed domains in sphingolipid/phosphatidylcholine/cholesterol bilayers with s ubstantially higher affinity than do analogous sphingomyelin, ceramide, or phosphatidylcholine molecules. By contrast, the affinity of sphingomyelin a nd ceramide for such domains is only marginally greater than that of a phos phatidylcholine with similar hydrocarbon chains. By using direct measuremen ts of molecular partitioning between Vesicles of different compositions, we show that the relative affinities of different C-6-NBD- and C-5-Bodipy-lab eled sphingolipids for sphingolipid-enriched domains are quantitatively, an d in most circumstances even qualitatively, quite different from those foun d for species whose N-acyl chains more closely resemble the long saturated chains of cellular sphingolipids. These findings lend support in principle to previous suggestions that differential partitioning of different sphingo lipids into "raft" domains could contribute to the differential trafficking of these species in eukaryotic cells. However, our findings also indicate that short-chain sphingolipid probes previously used to examine this phenom enon are in general ill-suited for such applications.