We review the experimental evidence accumulated within the past decade regarding the physiologic, biochemical, and molecular characterization of iodothyronine deiodinases (IDs) in piscine species. Agnathans, chondrichthyes, and teleosts express the three isotypes of IDs: ID1, ID2, and ID3, which are responsible for the peripheral fine-tuning of thyroid hormone (TH) bioactivity. At the molecular and operational level, fish IDs share properties with their corresponding vertebrate counterparts. However, fish IDs also exhibit discrete features that seem to be distinctive for piscine species. Indeed, teleostean ID1 is conspicuously resistant to propylthiouracil (PTU) inhibition, and its response to thyroidal status differs from that exhibited by other ID1s. Moreover, both the high level of ID2 activity and its expression in the liver of teleosts are unique among vertebrates. The physiologic role of iodothyronine deiodination in functions regulated by TH in fish is not entirely clear. Nevertheless, current experimental evidence suggests that IDs may coordinate and facilitate, in a tissue-specific fashion, the action of iodothyronines and other hormones involved in such processes.

From an evolutionary perspective, deiodinases may be considered pivotal players in the emergence and functional diversification of both thyroidal systems (TS) and their iodinated messengers. To better understand the evolutionary pathway and the concomitant functional diversification of vertebrate deiodinases, in the present review we summarized the highlights of the available information regarding this ubiquitous enzymatic component that represents the final, common physiological link of TS. The information reviewed here suggests that deiodination of tyrosine metabolites is an ancient feature of all chordates studied to date and consequently, that it precedes the integration of the TS that characterize vertebrates. Phylogenetic analysis presented here points to D1 as the oldest vertebrate deiodinase and to D2 as the most recent deiodinase gene, a hypothesis that agrees with the notion that D2 is the most specialized and finely regulated member of the family and plays a key role in vertebrate neurogenesis. Thus, deiodinases seem to be major participants in the evolution and functional expansion of the complex regulatory network of TS found in vertebrates.

Recent studies in our laboratory have shown that in some teleosts, 3,5-di-iodothyronine (T 2 or 3,5-T 2 ) is as bioactive as 3,5,3 0 -tri-iodothyronine (T 3 ) and that its effects are in part mediated by a TRb1 (THRB) isoform that contains a 9-amino acid insert in its ligand-binding domain (long TRb1 (L-TRb1)), whereas T 3 binds preferentially to a short TRb1 (S-TRb1) isoform that lacks this insert. To further understand the functional relevance of T 2 bioactivity and its mechanism of action, we used in vivo and ex vivo (organotypic liver cultures) approaches and analyzed whether T 3 and T 2 differentially regulate the S-TRb1 and L-TRb1s during a physiological demand such as growth. In vivo, T 3 and T 2 treatment induced body weight gain in tilapia. The expression of L-TRb1 and S-TRb1 was specifically regulated by T 2 and T 3 respectively both in vivo and ex vivo. The TR antagonist 1-850 effectively blocked thyroid hormone-dependent gene expression; however, T 3 or T 2 reversed 1-850 effects only on S-TRb1 or L-TRb1 expression, respectively. Together, our results support the notion that both T 3 and T 2 participate in the growth process; however, their effects are mediated by different, specific TRb1 isoforms.

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Recent molecular cloning studies in mammals and amphibians have demonstrated that the types I, II, and III deiodinases constitute a family of selenoproteins of critical importance in metabolizing T4 to active (i.e. T3) and inactive (i.e. rT3) metabolites. In several tissues of teleost fish, various deiodinase processes have been described, but the structural and functional characteristics of these enzymes and their relationship to the deiodinases present in higher vertebrates remains uncertain. Using a complementary DNA library derived from the liver of the teleost Fundulus heteroclitus, we have identified a complementary DNA that codes for a deiodinase with functional characteristics virtually identical to those of the mammalian and amphibian type II deiodinase. Sequence analysis demonstrates a high degree of homology at both the nucleotide and predicted amino acid levels between the Fundulus clone and these previously characterized type II enzymes, including the presence of an in-frame TGA codon that codes for selenocysteine. These findings demonstrate that the deiodinase family of selenoproteins has been highly conserved during vertebrate evolution and underscores their importance in the regulation of thyroid hormone action.

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Until recently,3, has been considered an inactive by-product of triiodothyronine (T3) deiodination. However, studies from several laboratories have shown that 3,5-T2 has specific, nongenomic effects on mitochondrial oxidative capacity and respiration rate that are distinct from those due to T3. Nevertheless, little is known about the putative genomic effects of 3,5-T2. We have previously shown that hyperthyroidism induced by supraphysiological doses of 3,5-T2 inhibits hepatic iodothyronine deiodinase type 2 (D2) activity and lowers mRNA levels in the killifish in the same manner as T3 and T4, suggesting a pretranslational effect of 3,5-T2 (Garcia-G C, Jeziorski MC, Valverde-R C, Orozco A. Gen Comp Endocrinol 135: 201-209, 2004). The question remains as to whether 3,5-T2 would have effects under conditions similar to those that are physiological for T3. To this end, intact killifish were rendered hypothyroid by administering methimazole. Groups of hypothyroid animals simultaneously received 30 nM of either T3, reverse T3, or 3,5-T2. Under these conditions, we expected that, if it were bioactive, 3,5-T2 would mimic T3 and thus reverse the compensatory upregulation of D2 and tyroid receptor ␤1 and downregulation of growth hormone that characterize hypothyroidism. Our results demonstrate that 3,5-T2 is indeed bioactive, reversing both hepatic D2 and growth hormone responses during a hypothyroidal state. Furthermore, we observed that 3,5-T2 and T3 recruit two distinct populations of transcription factors to typical palindromic and DR4 thyroid hormone response elements. Taken together, these results add further evidence to support the notion that 3,5-T 2 is a bioactive iodothyronine. deiodinase type 2; thyroid hormone receptor ␤1; thyroid hormone response element; killifish IODOTHYRONINES OR THYROID hormones (TH) are essential in regulating energy expenditure and development. Triiodothyronine (T 3 ) is the bioactive TH, which modulates gene expression in virtually every vertebrate tissue through ligand-dependent transcription factors, the TH receptors (TR). Sequential deiodination of thyroxine (T 4 ) generates T 3 as well as other iodothyronines that have been considered inactive by-products, but, recently, interest has grown in identifying bioactive iodothyronines in addition to T 4 and T 3 . Studies from several laboratories have suggested that 3,5-diiodothyronine (3,5-T 2 ), a putative product of the deiodination pathway involved in T 3 metabolism, could be a peripheral mediator of some effects of TH on mitochondrial oxidative capacity and respiration rate. To date, results in mammals suggest that 3,5-T 2 has specific actions on oxygen consumption that are distinct from those of T 3 : they are not attenuated by inhibition of protein synthesis and are more rapid than those due to T 3 (for review, see Ref. 12). Genomic effects of 3,5-T 2 have been analyzed in only a few classic iodothyronine-dependent genes, such as thyroid stimulating hormone (TSH), thyroid receptor ␤2 (TR␤2), iodothyronine deiodinase type 1 (D1)...