De novo triiodothyronine formation from thyrocytes activated by thyroid-stimulating hormone

Cintia E Citterio,Balaji Veluswamy,Sarah J Morgan,Valerie A Galton,J Paul Banga,Stephen Atkins,Yoshiaki Morishita,Susanne Neumann,Rauf Latif,Marvin C Gershengorn,Terry J Smith,Peter Arvan

doi:10.1074/jbc.m117.784447

Abstract

The thyroid gland secretes primarily tetraiodothyronine (T4), and some triiodothyronine (T3). Under normal physiological circumstances, only one-fifth of circulating T3 is directly released by the thyroid, but in states of hyperactivation of thyroid-stimulating hormone receptors (TSHRs), patients develop a syndrome of relative T3 toxicosis. Thyroidal T4 production results from iodination of thyroglobulin (TG) at residues Tyr5 and Tyr130, whereas thyroidal T3 production may originate in several different ways. In this study, the data demonstrate that within the carboxyl-terminal portion of mouse TG, T3 is formed de novo independently of deiodination from T4 We found that upon iodination in vitro, de novo T3 formation in TG was decreased in mice lacking TSHRs. Conversely, de novo T3 that can be formed upon iodination of TG secreted from PCCL3 (rat thyrocyte) cells was augmented from cells previously exposed to increased TSH, a TSHR agonist, a cAMP analog, or a TSHR-stimulating antibody. We present data suggesting that TSH-stimulated TG phosphorylation contributes to enhanced de novo T3 formation. These effects were reversed within a few days after removal of the hyperstimulating conditions. Indeed, direct exposure of PCCL3 cells to human serum from two patients with Graves' disease, but not control sera, led to secretion of TG with an increased intrinsic ability to form T3 upon in vitro iodination. Furthermore, TG secreted from human thyrocyte cultures hyperstimulated with TSH also showed an increased intrinsic ability to form T3 Our data support the hypothesis that TG processing in the secretory pathway of TSHR-hyperstimulated thyrocytes alters the structure of the iodination substrate in a way that enhances de novo T3 formation, contributing to the relative T3 toxicosis of Graves' disease.

Highlights

The thyroid gland secretes primarily tetraiodothyronine (T4), and some triiodothyronine (T3)
De novo T3 that can be formed upon iodination of TG secreted from PCCL3 cells was augmented from cells previously exposed to increased TSH, a thyroid-stimulating hormone receptors (TSHRs) agonist, a cAMP analog, or a TSHRstimulating antibody
As Dunn et al [18] reported that the majority of T3 synthesized within TG is located in its C-terminal portion, we extracted mouse thyroid tissue and found by immunoblotting that a single band of intact Tg was cleaved to two fragments of the expected molecular mass after incubation with Factor Xa (Fig. 1C, left)

Summary

To whom correspondence should be addressed

Iodination is catalyzed by thyroid peroxidase, which provides the necessary oxidation to form diiodotyrosine (DIT) and monoiodotyrosine (MIT) within TG. Favored by these same oxidizing conditions, a coupling reaction involving a DIT acceptor residue and a corresponding DIT donor residue allows for the formation of T4 within the TG polypeptide; coupling of an MIT donor with a DIT acceptor allows for de novo T3 formation [15, 16]. During its complex trafficking through the intracellular transport pathway of thyrocytes, TG undergoes considerable post-translational processing prior to its secretion and iodination [20]. We directly demonstrate de novo T3 formation in TG and establish that this ability is directly related to the degree to which thyrocytes have been exposed to prior TSHR stimulation

Results

Discussion

Experimental procedures