The plasma membrane of Saccharomyces cerevisiae was studied using the probes trans-parinaric acid (t-PnA) and diphenylhexatriene (DPH). DPH anisotropy is a good reporter of global membrane order. The fluorescence lifetimes of t-PnA are particularly sensitive to the presence and nature of ordered domains, but thus far they have not been measured in yeast cells. A long lifetime typical of the gel phase (> 30 ns) was found in wild-type (wt) cells from two different genetic backgrounds, at 24 and 30 degrees C, providing the first direct evidence for the presence of gel domains in living cells. To understand their nature and location, the study of wt cells was extended to spheroplasts, the isolated plasma membrane and liposomes from total lipid- and plasma membrane lipid-extracts (with or without ergosterol extraction by cyclodextrin). It is concluded that the plasma membrane is mostly constituted by ordered domains, and that the gel domains found in living cells are predominantly at the plasma membrane and are formed by lipids. To understand their composition, strains with mutations in sphingolipid and ergosterol metabolism, and in the GPI-anchor remodeling pathway were also studied. The results strongly indicate that the gel domains are not ergosterol-enriched lipid rafts; they are mainly composed of sphingolipids, possibly inositolphosphorylceramide, and contain GPI-anchored proteins, suggesting an important role in membrane traffic and signaling, and interactions with the cell wall. The abundance of the sphingolipid-enriched gel domains was inversely related to the cellular membrane system global order, suggesting their involvement in the regulation of membrane properties.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Reference||Annotation Extension|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Conditions||Strain||Source||Reference|