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 v ariety 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 induct ion of GST Ya gene expression and of AP-1 binding, activity is regulat ed by intracellular glutathione (GSH) levels. To study the role of rea ctive oxygen species in the induction of AP-1 activity and GST Ya gene expression and their effect on intracellular GSH levels, we have expo sed hepatoma cells to adriamycin and. two synthetic quinones, Q(c)(b) and Q(n), with different capacities to generate oxygen radicals. The k inetics of quinone-mediated generation of hydroxyl radicals were monit ored in intact cells by a spin trapping technique and EPR spectral mea surements. We find that quinones which can chelate Fe(III) ions, adria mycin and Q(c)(b), are more effective in hydroxyl radical production t han the nonchelating quinone Q(n). Furthermore we show that the induct ion of AP-1 binding activity and GST Ya gene expression by these quino nes correlates with their oxygen radical production, adriamycin and Q( c)(b) being stronger inducers that Q(n). The present study indicates t hat the AP-1-mediated induction of GST Ya gene expression is part of t he response to oxidative stress. A transient increase by 2.5-fold in t he intracellular GSH level was observed 30 min after exposure of cells to quinone and was followed by a rapid depletion of GSH. This increas e in the GSH level represents an induction of GSH synthesis since it w as blocked by buthionine sulfoximine, an inhibitor of gamma-glutamylcy steine synthetase. The oxygen radical-mediated induction of gamma-glut amylcysteine synthetase, the first and rate-limiting step in GSH synth esis, appears therefore to be an early event in cells exposed-to quino ne metabolism. These findings therefore indicate a correlation between production of oxygen radicals, GSH level, and induction of AP-1-media ted GST Ya gene expression.