Molecular Characterization of Avian Metallothionein (MT) 1 and 2 Isoforms: mRNA Expression, Transactivation Potency, and Detoxification Potential Associated with Element Exposure

To assess the inducibility of avian MT genes and potential tolerability of their proteins to element exposure, we investigated the expression profiles of MT isoforms by metal accumulation in wild cormorants (Phalacrocorax carbo) and mallards (Anas platyrhynchos). We also measured transcriptional activities of avian MT genes by elements and detoxification potencies of their in vitro-expressed MT proteins. Expression levels of MT1 and MT2 in cormorant livers showed significantly positive correlations with Cu and Zn concentrations, suggesting a similar mechanism for transcriptional activation of both MT isoforms by these elements. Results of an in vitro reporter gene assay that both MT promoter/enhancer regions from cormorant were dose-dependently transactivated by treatments with Cu and Zn agreed well with these observations in the wild population. In contrast, no significant correlation was observed between hepatic Cd levels and MT mRNAs, whereas Cd was the most potent inducer of cormorant MT1/2 promoters by in vitro reporter gene assay. This may be due to the masking effect of multiple element exposures, probably by Zn and/or Cu that were highly accumulated in wild cormorants. In vitro transactivation potency by co-treatment of Zn with Cd supported that Zn could conceal Cd-induced MT expression in the wild cormorant population. In mallard, MT1 had no correlation with any metal while MT2 expression was positively correlated only with Cu, even though hepatic Cu and Zn concentrations in mallard were much higher than in cormorant. This indicates that cormorant is more susceptible to elemental exposure than mallard in terms of MT induction. MT1 and 2 proteins of cormorant and mallard endowed E. coli with significant higher growth rate than control to Cd exposure (500–1000 μM), implying that both avian MTs could be involved in the detoxification of intracellular Cd. This study provides the first evidence on the inducibility of avian MT isoforms by specific elements and functional significance of each avian MT isoform in detoxifying intracellular heavy metals.