Metabolic instability of type 2 deiodinase is transferable to stable proteins independently of subcellular localization

Thyroid hormone activation is catalyzed by two deiodinases, D1 and D2. Whereas D1 is a stable plasma membrane protein, D2 is resident in the endoplasmic reticulum (ER) and has a 20-min half-life due to selective ubiquitination and proteasomal degradation. Here we have shown that stable retention explains D2 residency in the ER, a mechanism that is nevertheless over-ridden by fusion to the long-lived plasma membrane protein, sodium-iodine symporter. Fusion to D2, but not D1, dramatically shortened sodium-iodine symporter half-life through a mechanism dependent on an 18-amino acid D2-specific instability loop. Similarly, the D2-specific loop-mediated protein destabilization was also observed after D2, but not D1, was fused to the stable ER resident protein SEC62. This indicates that the instability loop in D2, but not its subcellular localization, is the key determinant of D2 susceptibility to ubiquitination and rapid turnover rate. Our data also show that the 6 N-terminal amino acids, but not the 12 C-terminal ones, are the ones required for D2 recognition by WSB-1

The Hedgehog-inducible ubiquitin ligase subunit WSB-1 modulates thyroid hormone activation and PTHrP secretion in the developing growth plate

WSB-1 is a SOCS-box-containing WD-40 protein of unknown function that is induced by Hedgehog signalling in embryonic structures during chicken development. Here we show that WSB-1 is part of an E3 ubiquitin ligase for the thyroid-hormone-activating type 2 iodothyronine deiodinase (D2). The WD-40 propeller of WSB-1 recognizes an 18-amino-acid loop in D2 that confers metabolic instability, whereas the SOCS-box domain mediates its interaction with a ubiquitinating catalytic core complex, modelled as Elongin BC-Cul5-Rbx1 (ECS(WSB-1)). In the developing tibial growth plate, Hedgehog-stimulated D2 ubiquitination via ECS(WSB-1) induces parathyroid hormone-related peptide (PTHrP), thereby regulating chondrocyte differentiation. Thus, ECS(WSB-1) mediates a mechanism by which 'systemic' thyroid hormone can effect local control of the Hedgehog-PTHrP negative feedback loop and thus skeletogenesis

THE THYROID HORMONE INACTIVATING DEIODINASE FUNCTIONS AS HOMODIMER

International audienceThe type 3 deiodinase (D3) inactivates thyroid hormone action by catalyzing tissue-specific inner ring de- iodination, predominantly during embryonic development. D3 has gained much attention as a player in the euthyroid sick syndrome, given its robust reactivation during injury and/or illness. While much of the structure biology of the deiodinases is derived from studies with D2, a dimeric endoplasmic-reticulum obligatory activating deiodinase, little is known about the holostructure of the plasma membrane-resident D3, the deiodinase capable of thyroid hormone inactivation. Here we used fluorescence resonance energy transfer (FRET) in live cells to demonstrate that D3 exists as homodimer. While D3 homodimerized in its native state, minor heterodimerization was also observed between D3:D1 and D3:D2 in intact cells, the significance of which remains elusive. Incubation with 0.5-1.2 M urea resulted in loss of D3 homodimeri- zation as assessed by bioluminescence resonance energy transfer (BRET) and a proportional loss of en- zyme activity, to a maximum of ~50%. Protein modeling using a D2-based scaffold identified potential dimerization surfaces in the transmembrane and globular domains. Truncation of the transmembrane do- main (ΔD3) abrogated dimerization and deiodinase activity except when co-expressed with full-length catalytically inactive deiodinase, thus assembled as ΔD3:D3 dimer; thus the D3 globular domain also ex- hibits dimerization surfaces. In conclusion, the inactivating deiodinase D3 exists as homo- or heterodimer in living intact cells, a feature that is critical for their catalytic activities

Cellular and Molecular Basis of Deiodinase-Regulated Thyroid Hormone Signalinga

The iodothyronine deiodinases initiate or terminate thyroid hormone action and therefore are critical for the biological effects mediated by thyroid hormone. Over the years, research has focused on their role in preserving serum levels of the biologically active molecule T3 during iodine deficiency. More recently, a fascinating new role of these enzymes has been unveiled. The activating deiodinase (D2) and the inactivating deiodinase (D3) can locally increase or decrease thyroid hormone signaling in a tissue- and temporal-specific fashion, independent of changes in thyroid hormone serum concentrations. This mechanism is particularly relevant because deiodinase expression can be modulated by a wide variety of endogenous signaling molecules such as sonic hedgehog, nuclear factor-κB, growth factors, bile acids, hypoxia-inducible factor-1α, as well as a growing number of xenobiotic substances. In light of these findings, it seems clear that deiodinases play a much broader role than once thought, with great ramifications for the control of thyroid hormone signaling during vertebrate development and metamorphosis, as well as injury response, tissue repair, hypothalamic function, and energy homeostasis in adults