ROLE OF QUINONE-MEDIATED GENERATION OF HYDROXYL RADICALS IN THE INDUCTION OF GLUTATHIONE-S-TRANSFERASE GENE-EXPRESSION

Induction of glutathione S-transferase (GST) Ya gene expression by a variety of chemical agents is mediated by a regulatory element composed of two adjacent AP-1-like binding sites and activated by the Fos/Jun heterodimeric complex (AP-1). We have previously shown that the induction of GST Ya gene expression and of AP-1 binding, activity is regulated by intracellular glutathione (GSH) levels. To study the role of reactive oxygen species in the induction of AP-1 activity and GST Ya gene expression and their effect on intracellular GSH levels, we have exposed hepatoma cells to adriamycin and. two synthetic quinones, Q(c)(b) and Q(n), with different capacities to generate oxygen radicals. The kinetics of quinone-mediated generation of hydroxyl radicals were monitored in intact cells by a spin trapping technique and EPR spectral measurements. We find that quinones which can chelate Fe(III) ions, adriamycin and Q(c)(b), are more effective in hydroxyl radical production than the nonchelating quinone Q(n). Furthermore we show that the induction of AP-1 binding activity and GST Ya gene expression by these quinones correlates with their oxygen radical production, adriamycin and Q(c)(b) being stronger inducers that Q(n). The present study indicates that the AP-1-mediated induction of GST Ya gene expression is part of the response to oxidative stress. A transient increase by 2.5-fold in the intracellular GSH level was observed 30 min after exposure of cells to quinone and was followed by a rapid depletion of GSH. This increase in the GSH level represents an induction of GSH synthesis since it was blocked by buthionine sulfoximine, an inhibitor of gamma-glutamylcysteine synthetase. The oxygen radical-mediated induction of gamma-glutamylcysteine synthetase, the first and rate-limiting step in GSH synthesis, appears therefore to be an early event in cells exposed-to quinone metabolism. These findings therefore indicate a correlation between production of oxygen radicals, GSH level, and induction of AP-1-mediated GST Ya gene expression.