Molecular cloning of the rabbit deiodinase type I, type II and type III genes

Thyroid hormone is essential for normal development and metabolism in vertebrate species. Three iodothyronine deiodinases have been identified which have a crucial role in controlling the availability of the biologically active thyroid hormone T3. The goal of this study was to sequence the full-length rabbit type I deiodinase (DIO1), type II deiodinase (DIO2) and type III deiodinase (DIO3) cDNAs. Total RNA was isolated from rabbit liver, pituitary and hypothalamic tissue and subsequently used for reverse transcription and cDNA synthesis. Forward and backward primers were designed from the published nucleotide sequences for highly conserved regions of DIO1, DIO2 and DIO3 in human (Homo sapiens) and mouse (Mus musculus). Polymerase chain reaction was carried out with these primers using rabbit cDNA as a template. Fragments of the expected size were cloned into a pGEM-T plasmid vector and sequenced. Full-length sequences of rabbit DIO1 and rabbit DIO2 were obtained, while cloning of DIO3 is in progress. Rabbit DIO1 and DIO2 showed 67-83% and 83-93% amino acid identity with DIO1 and DIO2 from other mammalian species, respectively. Combining these results with previous experimental data and with datamining of the relevant databases has increased the number of full-length vertebrate deiodinase sequences to 28. Alignment of these amino acid sequences reveals some remarkable aspect of the deiodinase family, namely the existence of a functional DIO1 in Xenopus laevis, in contrast to previous conclusions, as well as the existence of two DIO3 subtypes in fish and X. laevis. All deiodinases have a strongly conserved core catalytic sequence including the essential selenocysteine residue.status: accepte