Characterization of a Propylthiouracil-Insensitive Type I Iodothyronine Deiodinase*

Abstract

Mammalian type I iodothyronine deiodinase (D1) activates and inactivates thyroid hormone by outer ring deiodination (ORD) and inner ring deiodination (IRD), respectively, and is potently inhibited by propylthiouracil (PTU). Here we describe the cloning and characterization of a complementary DNA encoding a PTU-insensitive D1 from teleost fish (Oreochromis niloticus, tilapia). This complementary DNA codes for a protein of 248 amino acids, including a putative selenocysteine (Sec) residue, encoded by a TGA triplet, at position 126. The 3' untranslated region contains two putative Sec insertion sequence (SECIS) elements. Recombinant enzyme expressed in COS-1 cells catalyzes both ORD of T4 and rT3 and IRD of T3 and T3 sulfate with the same substrate specificity as native tilapia D1 (tD1), i.e. rT3 >> T4 > T3 sulfate > T3. Native and recombinant tD1 show equally low sensitivities to inhibition by PTU, iodoacetate, and gold thioglucose compared with the potent inhibitions observed with mammalian D1s. Because the residue 2 positions downstream from Sec is Pro in tD1 and in all (PTU-insensitive) type II and type III iodothyronine deiodinases but Ser in all PTU-sensitive D1s, we prepared the Pro128Ser mutant of tD1. The mutant enzyme showed strongly decreased ORD and somewhat increased IRD activity, but was still insensitive to PTU. These results provide new information about the structure-activity relationship of D1 concerning two characteristic properties, i.e. catalysis of both ORD and IRD, and inhibition by PTU.

Low salinity acclimation and thyroid hormone metabolizing enzymes in gilthead seabream (Sparus auratus)

“…Due to the scarcity of data, it is difficult to construct a generalised tissue distribution of deiodinases in fishes. Indeed, the tissue distribution of seabream D1 m RNA we show here, with positive signals only for liver and kidney preparations, differs distinctly from that in Nile tilapia preochromis niloticus) where abundant expression and activity were measured in the kidney, but intermediate levels in the gills and none in the liver (Sanders et al, 1997). The high ORD activities towards T 4, T3 and rT3 measured in seabream gills, that were responsive to a hypoosmotic challenge, are indicative for an im portant role of branchial tissue in thyroid horm one metabolism in seabream.…”

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

“…However, at least in tilapia and trout, fish D1 is insensitive to inhibition by 6-propylthiouracil (PTU), in contrast to the potent inhibition of mammalian (and chicken) D1 by this thyrostatic drug (5, 36 -38). To investigate the molecular basis for this difference in PTU sensitivity between fish and mammalian D1, we have recently cloned and characterized com-plementary DNA (cDNA) coding for D1 in tilapia (20). In contrast to our hypothesis, we found that tilapia D1 contains a Sec residue in a position corresponding to the Sec residue in PTU-sensitive D1s, indicating that differences in PTU sensitivity are determined by other structural elements (20).…”

“…The amino acid identity of tD3 amounts to 62-65% with frog D3 (12,13), 67% with chicken D3 (19), and 57% with mammalian D3 (14,15). The amino acid sequence of tD3 shows 35% identity with tilapia D1 (20), 33% identity with chicken D1 (19) and 30 -33% identity with mammalian D1 (8 -11). The amino acid identity of tD3 with fish (F. heteroclitus) D2 is 36% (18), with frog D2 33% (16) and with mammalian D2 38 -39% (6,17).…”

Developmental toxicity of the PBDE metabolite 6-OH-BDE-47 in zebrafish and the potential role of thyroid receptor β

“…Additionally, positive control chemicals with known modes of action (IOP, PTU) were utilized to further examine their impacts on developmental morphology via thyroid hormone disruption Propylthiouracil targets thyroid peroxidase, and may inhibit DI 2 in fish species (Orozco et al 2000; Sanders et al, 1997; Visser et al1983). Iopanoic acid targets DI 1 and DI 2, inhibiting peripheral conversion of T 4 to T 3 in target tissues (Bouzaffour et al, 2010).…”