AbstractHeavy metal‐induced transcriptional activation of the genes coding for metallothionein (MT) is mediated by a cis‐acting DNA element, the metal‐responsive element (MRE). MRE‐binding transcription factor‐1 (MTF‐1) is a highly conserved heavy metal‐induced transcriptional activator. MTF‐1 also activates transcription in response to oxidative stress and regulates the expression of several cytoprotective factor genes, including MT, γ‐glutamylcysteine synthetase, and Cu/Zn‐superoxide dismutase. It is thus thought that MTF‐1 plays a role in cellular stress response. The physiological role of MTF‐1 remains unclear because of the lack of MTF‐1‐specific activators and/or inhibitors. To obtain an MTF‐1‐specific inhibitor, we constructed an MTFΔC (amino acids 1–317), a C‐terminal deletion mutant of MTF‐1. MTFΔC could bind MRE and competed with MTF‐1 for MTF–MRE complex formation. Transient expression of MTFΔC in HepG2 cells reduced MRE‐driven gene expression, demonstrating that MTFΔC is dominant to MTF‐1. HepG2 cells stably expressing MTFΔC showed increased susceptibility to the cytotoxic effects of tert‐butyl hydroperoxide (tBH). Furthermore, we constructed Ad5MTFΔC, a recombinant adenovirus that expresses MTFΔC. Infection with the virus induced MTFΔC expression and increased susceptibility to the cytotoxic effects of tBH. These results indicate that MTF‐1 participates in controlling the cellular redox state. © 2004 Wiley‐Liss, Inc.