GS-X pump is functionally overexpressed in cis-diamminedichloroplatinum (II)-resistant human leukemia HL-60 cells and down-regulated by cell differentiation.
The ATP-dependent glutathione S-conjugate export pump, named GS-X pump, has been shown to eliminate a potentially cytotoxic glutathione-platinum (GS.Pt) complex from tumor cells, thereby modulating glutathione (GSH)-associated resistance to cis-diamminedichloroplatinum(II) (cisplatin) (Ishikawa, T., and Ali-Osman, F. (1993) J. Biol. Chem. 268, 20116-20125). The present study provides evidence that the GS-X pump is functionally overexpressed in cisplatin-resistant human promyelocytic leukemia HL-60 (HL-60/R-CP) cells, in which the cellular GSH level was substantially enhanced. Indeed, the rate of ATP-dependent transport of the GS.Pt complex, measured with plasma membrane vesicles, was about 4-fold greater in HL-60/R-CP cells than in HL-60 cells. Three membrane proteins with apparent molecular masses of 200, 110, and 70 kDa were overexpressed in HL-60/R-CP cells, whereas P-glycoprotein (MDR1) was not immunologically detected in the membrane preparations from resistant and sensitive cells. Unlike in HL-60 cells, increased numbers of intracellular vesicles were observed in the cytoplasm of HL-60/R-CP cells. Fluorescence microscopy with syn-(CICH2,CH3)-1,5-diazabicyclo-[3.3.0]-octa-3,6-dione-2,8-dione (monochlorobimane) revealed that the fluorescent glutathione S-conjugate accumulated in intracellular vesicles of the cisplatin-resistant cells in an energy-dependent manner. The GS-X pump is suggested to contribute to vesicle-mediated excretion of GSH-drug conjugates from cells. In addition, both HL-60 and HL-60/R-CP cells underwent cell differentiation in response to 12-O-tetradecanoylphorbol-13-acetate, retinoic acid, and dimethyl sulfoxide, resulting in proliferation arrest as well as a remarkable decrease in the c-myc mRNA levels. After cell differentiation, a significant decrease was observed in the activity of ATP-dependent transport of the GS.Pt complex in membrane vesicles prepared from both HL-60/R-CP and HL-60 cells. These results suggest that the expression of the GS-X pump in both cisplatin-resistant and -sensitive cells is related to cell proliferation.
Department of Experimental Pediatrics, University of Texas, M.D. Anderson Cancer Center, Houston 77030.
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D000255:Adenosine Triphosphate / Q000378:metabolism; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D002454:Cell Differentiation; D002945:Cisplatin / Q000494:pharmacology*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D015536:Down-Regulation; D004351:Drug Resistance; D005978:Glutathione / Q000378:metabolism*; D006801:Humans; D007938:Leukemia / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C086860:glutathione transporter; D014212:Tretinoin; D000255:Adenosine Triphosphate; D005978:Glutathione; D013755:Tetradecanoylphorbol Acetate; D002945:Cisplatin; D004121:Dimethyl Sulfoxide