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Cloning and Characterization of Type III Iodothyronine Deiodinase from the Fish Oreochromis niloticus¹

Department of Internal Medicine III (J.P.S., S.V.d.G., E.K., T.J.V.), Erasmus University Medical School, 3000 DR Rotterdam, The Netherlands; Laboratory of Comparative Endocrinology (S.V.d.G., V.M.D., E.R.K.), K.U. Leuven, 3000 Leuven, Belgium; Department of Nuclear Medicine (J.L.L.), University of Massachusetts Medical Center, Worcester, Massachusetts 01655

Address all correspondence and requests for reprints to: Theo J. Visser, Ph.D., Department of Internal Medicine III, Erasmus University Medical School, Room Bd 234, P.O. Box 1738, 3000 DR Rotterdam, The Netherlands. E-mail: visser{at}inw3.azr.nl

Type III iodothyronine deiodinase (D3) catalyzes the inner ring deiodination (IRD) of T₄ and T₃ to the inactive metabolites rT₃ and 3,3'-diiodothyronine (3,3'-T2), respectively. Here we describe the cloning and characterization of complementary DNA (cDNA) coding for D3 in fish (Oreochromis niloticus, tilapia). This cDNA contains 1478 nucleotides and codes for a protein of 267 amino acids, including a putative selenocysteine (Sec) residue, encoded by a TGA triplet, at position 131. The deduced amino acid sequence shows 57–67% identity with frog, chicken, and mammalian D3, 33–39% identity with frog, fish (Fundulus heteroclitus) and mammalian D2, and 30–35% identity with fish (tilapia), chicken, and mammalian D1. The 3' UTR contains a putative Sec insertion sequence (SECIS) element. Recombinant tilapia D3 (tD3) expressed in COS-1 cells and native tD3 in tilapia brain microsomes show identical catalytic activities, with a strong preference for IRD of T₃ (K_m 20 nM). IRD of [3,5-¹²⁵I]T₃ by native and recombinant tD3 are equally sensitive to inhibition by substrate analogs (T₃ > T₄ >> rT₃) and inhibitors (gold thioglucose >> iodoacetate > propylthiouracil). Northern analysis using a tD3 riboprobe shows high expression of a 1.6-kb messenger RNA in gill and brain, although D3 activity is much higher in brain than in gill. The characterization of tD3 cDNA provides new information about the structure-activity relationship of iodothyronine deiodinases and an important tool to study the regulation of thyroid hormone bioactivity in fish.

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