Upregulation of the expression of endogenous Mdr1 P-glycoprotein enhances lipid translocation in MDCK cells transfected with human MRP2

Various ABC transporters can translocate lipid molecules from the cytoplasmic into the exoplasmic leaflet of the plasma membrane bilayer. Two of these, MDR1 P-glycoprotein (Pgp) and MRP1, are multidrug transporters responsible for the resistance of various cancers against chemotherapy. We wanted to study whether MRP2, an ABC transporter of the bile canalicular membrane with a substrate specificity very similar to that of MRP1, is capable of translocating lipids. The translocation of short-chain lipids across the apical membrane of MDCK cells transfected with MRP2 was significantly higher than that in untransfected controls. However, the characteristics of the lipid translocation were similar to substrate transport by MDR1 and not MRP2: transport was strongly inhibited by classic MDR1 Pgp inhibitors, was independent of cellular glutathione, and was insensitive to a drug known to inhibit MRP2 activity. When tested by immunoblot, the MRP2-transfected cells expressed high levels of MRP2 but also of endogenous Mdr1. The expression of Mdr1 was unstable during maintenance of the cell line and correlated with the rate of lipid translocation across the apical membrane. We conclude that the observed increase in lipid transport in the MDCK cells transfected with MRP2 is the consequence of the upregulation of the expression of endogenous Mdr1 and that careful characterization of endogenous Mdr1 expression is needed in studies aimed to identify substrates of plasma membrane transporters.