Phenolic antioxidants, such as tert-butyl-hydroquinone (tBHQ), induce metallothionein (Mt1) gene expression. Induction of Mt1 mRNA correlates with oxidation-reduction functions of the antioxidants. Here we analyzed the biochemical pathway of the induction. Induction depends upon the presence of MTF-1, a transcription factor that is required for metal-induced transcription of Mt1, but does not require Nrf2, a tBHQ-activated CNC bZip protein that is responsible for regulating genes encoding phase II drug-metabolizing enzymes. Moreover, tBHQ induces the expression of MRE-bGeo, a reporter gene driven by five metal regulator elements (MREs) that constitute an optimal MTF-1 binding site. Reconstitution of Mtf1-null cells with MTF-1 restores induction by both zinc and tBHQ; however, reconstitution with only the DNA-binding domain of MTF-1 fused to VP16 activation domain allows in induction by zinc but not by tBHQ. Unlike activation of phase II genes by tBHQ, induction of Mt1 expression does not occur in the presence of EDTA, when cells are cultured in zinc-depleted medium, or in cells with reduced intracellular "free" zinc due to overexpression of ZnT1, a zinc-efflux transporter, indicating that induction requires zinc. These findings establish that phenolic antioxidants activate Mt1 transcription by a zinc-dependent mechanism that involves MTF-1 binding to MREs.
The Toxicologist. Society of Toxicology 43nd Annual Meeting and ToxExpo, March 21-25, 2004, Baltimore, Maryland