Characterization and expression of a metallothionein gene in the aquatic fern Azolla filiculoides under heavy metal stress

A cDNA encoding a type 2 metallothionein (MT) was isolated from Azolla filiculoides, termed AzMT2, accession no. AF482470. The AzMT2 transcript was expressed in sterile A. filiculoides that were free of the cyanobiont Anabaena azollae after erythromycin treatment, proving that AzMT2 is encoded by the fern genome. AzMT2 RNA expression was enhanced by the addition of Cd⁺², Cu⁺², Zn⁺² and Ni⁺² to the growth medium. The transcript level of AzMT2 correlated with the metal content in the plants. Temporal analysis of AzMT2 expression demonstrated that Cd²⁺ and Ni²⁺ induction of AzMT2 RNA expression occurred within 48 h. AzMT2-enhanced expression responded more intensely to the toxic Cd and Ni ions in A. filiculoides suggesting that AzMT2 may participate in detoxification mechanism. The more moderate response of AzMT2 to Zn and Cu ions, which are essential micronutrients, suggest a role for AzMT2 in metal homeostasis.