Activation of the chicken metallothionein promoter by metals and oxidative stress in cultured cells and transgenic mice.

Cis-acting elements in the chicken metallothionein promoter were tested for their ability to direct responses of reporter genes to metal ions and oxidative stress in transfected mouse cells and in transgenic mice. In addition, protein interactions with the promoter were analyzed by the electrophoretic mobility shift assay. In transient transfection assays and in transgenic mice, 107-bp of the chicken MT promoter was sufficient to direct responses to Zn. This promoter region also directed response to oxidative stress in transfected cells and transgenic mice, but in transgenic mice, maximal responsiveness to oxidative stress apparently involved other elements in the proximal promoter region (307-bp). The proximal 200-bp of the promoter contains sequences homologous to a metal response element (-47-bp), Sp1 binding sites (-70-bp and -161-bp), and an antioxidant response element (-189-bp). Electrophoretic mobility shift assay demonstrated that metal response element binding activity was low in control Hepa cell nuclear extracts, but was induced 6-fold after 45 min of H2O2 treatment. In contrast, Sp1 binding remained unchanged, and no evidence for specific binding to the core antioxidant response element consensus sequence was obtained. These studies demonstrate that cis-acting elements mediating induction of metallothionein gene expression by metals and oxidative stress are present in the chicken metallothionein promoter and suggest a role for increased binding of the transcription factor MTF-1 to the metal response element(s).