Co-expression of gamma-glutamylcysteine synthetase sub-units in response to cisplatin and doxorubicin in human cancer cells.

Glutathione (gamma-glutamylcysteinyl glycine, GSH) is believed to be important in the acquisition of resistance to anti-cancer drugs such as cisplatin (CDDP) and doxorubicin (DOX). gamma-Glutamylcysteine synthetase (gamma-GCS) is a key enzyme for maintaining intracellular GSH levels; it is composed of a catalytic heavy (gamma-GCSh) and a regulatory light (gamma-GCSl) sub-unit. In the present study, the expression of gamma-GCS sub-units was examined in human cancer cell lines. The levels of GSH, the expression of gamma-GCSh and gamma-GCSl mRNAs and proteins in human cancer cells were higher in CDDP-resistant cells and DOX-resistant cells than in the drug-sensitive cells. Treatment of CDDP/DOX-resistant human colonic-cancer cells (HCT8DDP) with CDDP and DOX caused simultaneous induction of the expression of gamma-GCSh and gamma-GCSl mRNAs. The transcriptional regulation of gamma-GCS was studied and it was observed that the DNA-binding activity of activator protein 1 (AP-1) is induced by CDDP and DOX using an electrophoretic mobility shift assay. We constructed chimeric genes containing various regions of the gamma-GCSh-promoter gene and the coding region for luciferase. The HCT8DDP cells transiently transfected with a plasmid containing an AP-1 site of the gamma-GCSh-promoter-luciferase construct showed increased luciferase activity when treated with CDDP and DOX. These results suggest that stimulation of the expression of gamma-GCSh by CDDP and DOX is mediated by AP-1, which relates to the resistance of cancer cells to the drug.