The Tetrahymena metallothionein gene family: twenty-one new cDNAs, molecular characterization, phylogenetic study and comparative analysis of the gene expression under different abiotic stressors.

Background: Ciliate metallothioneins (MTs) are included in family 7 of the MT superfamily. This family has been divided into two main subfamilies: 7a or CdMTs and 7b or CuMTs. All ciliate MTs reported have been isolated from different Tetrahymena species and present unique features with regard to standard MTs. Likewise, an expression analysis has been carried out on some of MT genes under metal stress, corroborating their classification into two subfamilies.

Results: We isolated 21 new cDNAs from different Tetrahymena species to obtain a wider view of the biodiversity of these conserved genes. Structural analysis (cysteine patterns) and an updated phylogenetic study both corroborated the previous classification into two subfamilies. A new CuMT from a Tetrahymena-related species Ichthyophthirius multifiliis was also included in this general analysis. We detected a certain tendency towards the presentation of a CdMT tri-modular structure in Borealis group species with respect to Australis group. We report for the first time a semi-complete paralog duplication of a CdMT gene originating a new CdMT gene isoform in T. malaccensis. An asymmetry of the codon usage for glutamine residues was detected between Cd- and CuMTs, and the phylogenetic implications are discussed. A comparative gene expression analysis of several MT genes by qRT-PCR revealed differential behavior among them under different abiotic stressors in the same Tetrahymena species.

Conclusions: The Tetrahymena metallothionein family represents a quite conserved protein structure group with unique features with respect to standard MTs. Both Cd- and CuMT subfamilies present very defined and differentiated characteristics at several levels: cysteine patterns, modular structure, glutamine codon usage and gene expression under metal stress, among others. Gene duplication through evolution seems to be the major genetic mechanism for creating new MT gene isoforms and increasing their functional diversity.