MRP1 (multidrug-resistance-related
protein 1)/ABCC1 (
ATP-binding cassette transporter C1) has been localized in
cholesterol-enriched
lipid rafts, which suggests a role for these
lipid rafts and/or
cholesterol in
MRP1 function. In the present study, we have shown for the first time that nearly complete oxidation of free
cholesterol in the plasma membrane of BHK-MRP1 (
MRP1-expressing baby hamster kidney) cells did not affect
MRP1 localization in
lipid rafts or its efflux function, using
5-carboxyfluorescein diacetate as a substrate. Inhibition of
cholesterol biosynthesis, using
lovastatin in combination with
RO 48-8071, an inhibitor of
oxidosqualene cyclase, resulted in a shift of
MRP1 out of
lipid raft fractions, but did not affect MRP1-mediated efflux in Neuro-2a (
neuroblastoma) cells. Short-term methyl-β-
cyclodextrin treatment was equally effective in removing free
cholesterol from Neuro-2a and BHK-MRP1 cells, but affected
MRP1 function only in the latter. The kinetics of loss of both
MRP1 efflux function and
lipid raft association during long-term methyl-β-
cyclodextrin treatment did not match the kinetics of free
cholesterol removal in both cell lines. Moreover,
MRP1 activity was measured in vesicles consisting of membranes isolated from BHK-MRP1 cells using the substrate
cysteinyl leukotriene C4 and was not changed when the free
cholesterol level of these membranes was either decreased or increased. In conclusion,
MRP1 activity is not correlated with the level of free
cholesterol or with localization in
cholesterol-dependent
lipid rafts.