Effect of cholesterol content on lipid microdomains in model membranes and cells.

摘要

Much experimental and theoretical evidence suggests small variations in the mole fraction of sterol can produce significant changes in the lateral organization, which could play a key role in the modulation of membrane function. The initial rate of sterol oxidation in multi-component systems composed of dehydroergosterol(DHE)/1-palmitoyl-2-oleoyl-L-alpha-phosphatidylcholine(POPC)/porcine brain sphingomyelins(pSPMs) is found to change with cholesterol content in an alternating manner, showing local maxima at critical mole fractions (e.g., 20.0, 25.0, 33.3 and 40.0 mol% sterol) predicted for maximal superlattice formation. Thus, this result indicates that the initial rate of sterol oxidation is governed by the extent of cholesterol superlattice and supports the existence of superlattice in multi-component model membranes. In the same system, the initial rate of sterol oxidation was measured in the presence of antioxidants. Lipoic acid and ascorbic acid do not disturb the original membrane lateral organization in spite of being able to reduce sterol oxidation rate. Next, I examined whether superlattice is related to lipid rafts observed in cells and how cholesterol content would affect lipid raft properties and protein behaviors related to lipid rafts. I found the majority of opioid receptors are localized in lipid rafts isolated from rat brain, human placenta, Chinese hamster ovary cells overexpressing FLAG tagged human kappa receptors (FLAG-hKOR) and NG108-15 neuroblastoma x glioma hybrid cells using a non-detergent method. The cholesterol reduction by 2% MCD treatment (∼48%) caused the change in cell shape from spindle to spherical, shift of lipid rafts, caveolin-1 and FLAG-hKOR to higher density fractions and an increase in (-)(trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidiny) cyclohexyl]benzeneacetamide (U50,488H) induced [35S]guanosine 5-(gamma-thio)triphosphate binding. Cholesterol replenishment reversed all the MCD effects. Lipid rafts isolated from multi-component model membranes and from CHO cells also display an alternating variation in raft density with the cholesterol content in original membranes or cells. This biphasic change was shown to be related to protein/lipid ratio and membrane packing in lipid rafts isolated from cells. In addition, the raft density reaches local maxima at critical sterol mole fractions for maximal superlattice formation in model membranes. This new finding suggests that lipid rafts and sterol superlattices may be related.