ATP-dependent glutathione disulphide transport mediated by the MRP gene-encoded conjugate export pump

Abstract

We have previously shown that the multidrug resistance protein (MRP) mediates the ATP-dependent membrane transport of the endogenous glutathione conjugate leukotriene C4 (LTC4) and of structurally related anionic conjugates of lipophilic compounds [Jedlitschky, Leier, Buchholz, Center and Keppler (1994) Cancer Res. 54, 4833-4836; Leier, Jedlitschky, Buchholz, Cole, Deeley and Keppler (1994) J. Biol. Chem. 269, 27807-27810]. We demonstrate in the present study that MRP also mediates the ATP-dependent transport of GSSG, as shown in membrane vesicles from human leukaemia cells overexpressing MRP (HL60/ADR cells) or HeLa cells transfected with an MRP expression vector (HeLa T5 cells) in comparison with the respective parental or control cells. The Km value for ATP-dependent transport of GSSG was 93 +/- 26 microM (mean value +/- S.D., n=5) in membrane vesicles from HeLa T5 cells. GSH, at a concentration of 100 microM, was not a substrate for any significant ATP-dependent MRP-mediated transport. The transport of GSSG was competitively inhibited by LTC4, by the leukotriene D4 receptor antagonist 3-([{3-(2-[7-chloro-2-quinolinyl]ethenyl)phenyl}-{(3-dimethylamino-3- oxopropyl)-thio}-methyl]thio)propanoic acid (MK 571) and by S-decylglutathione, with K1 values of 0.3, 0.6 and 0.7 microM respectively. These studies identify MRP as the membrane glycoprotein which mediates the ATP-dependent export of GSSG from these cells.