Functional Characterization of the 5′-upstream Region of MTT5 Metallothionein Gene from Tetrahymena thermophila

Abstract

Metallothioneins are ubiquitous small, cysteine-rich, metal-binding proteins that play important roles in intracellular metal homeostasis and detoxification. Very few data are available on the promoter region and the mechanism of metallothionein transcription in Protozoa. In this study, we focused on Tetrahymena thermophila MTT5 5'-flanking region. To define the sequence elements underlying the metal-responsiveness of this promoter, we constructed a series of deletions and mutations starting with a 1777 bp fragment immediately upstream of the start codon of MTT5. As a reporter gene we used the previously tested IAG52B surface antigen from the protozoan fish parasite Ichthyophthirius multifiliis. The results suggest that a region spanning between -300 bp and -274 bp, dubbed Tetrahymena thermophila Cadmium-Response-Element (TtCdRE), is necessary to elicit high-level expression of the transgene following induction with cadmium. This is the first demonstration by in vivo analyses of a regulatory element essential for Cd-mediated control of protozoan metallothionein gene expression, where the sequence GATA appears to be involved.