Tyrosine iodination and iodotyrosyl coupling of the N-terminal thyroid hormone forming site of human thyroglobulin modulate its binding to auto- and monoclonal antibodies

Mallet, Bernard; Lejeune, Pierre-Jean; Ruf, Jean; Piechaczyk, Martine; Marriq, Claudine; Carayon, Pierre

doi:10.1016/0303-7207(92)90012-u

Cited by 15 publications

(14 citation statements)

References 20 publications

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“…Therefore, it was presumably insulin and insulin-like growth factor-1 that was driving Tg production in the TSHR-KO mice. Estimating the amount of Tg produced by the different genotypes was not performed because iodinated Tg has been shown to be more immunogenic than noniodinated Tg (42)(43)(44). By using the TSHR-KO mouse as a model, we were able to demonstrate that thyroid hormone synthesis in mice is divided into a TSH-independent phase consisting of Tg synthesis, and a TSH-dependent phase consisting of Tg iodination ending in hormone production and release.…”

Section: Discussionmentioning

confidence: 99%

Defining thyrotropin-dependent and -independent steps of thyroid hormone synthesis by using thyrotropin receptor-null mice

Marians

Blair

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

193

120

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The thyrotropin (TSH) receptor (TSHR) is a member of the heterotrimeric G protein-coupled family of receptors whose main function is to regulate thyroid cell proliferation as well as thyroid hormone synthesis and release. In this study, we generated a TSHR knockout (TSHR-KO) mouse by homologous recombination for use as a model to study TSHR function. TSHR-KO mice presented with developmental and growth delays and were profoundly hypothyroid, with no detectable thyroid hormone and elevated TSH. Heterozygotes were apparently unaffected. Knockout mice died within 1 week of weaning unless fed a diet supplemented with thyroid powder. Mature mice were fertile on the thyroid-supplemented diet. Thyroid glands of TSHR-KO mice produced uniodinated thyroglobulin, but the ability to concentrate and organify iodide could be restored to TSHR-KO thyroids when cultured in the presence of the adenylate cyclase agonist forskolin. Consistent with this observation was the lack of detectable sodium-iodide symporter expression in TSHR-KO thyroid glands. Hence, by using the TSHR-KO mouse, we provided in vivo evidence, demonstrating that TSHR expression was required for expression of sodium-iodide symporter but was not required for thyroglobulin expression, suggesting that the thyroid hormone synthetic pathway of the mouse could be dissociated into TSHR-dependent and -independent steps.

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Section: Discussionmentioning

confidence: 99%

Defining thyrotropin-dependent and -independent steps of thyroid hormone synthesis by using thyrotropin receptor-null mice

Marians

Blair

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

193

120

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“…This domain was related, in subsequent studies (Henry et al, 1992), to domain 11, previously depicted by Piechaczyk et al (1987). Antigenic mapping was utilized to probe thyroglobulin determinants involved in thyroid-hormone synthesis (Mallet et al, 1992;Debaets et al, 1987;Chan et al, 1986) and to study the recognition of thyroglobulin by its receptor on FRTL5 cells (Mziaut et al, 1996). These latter studies prompted us to construct a topographically oriented antigenic map of thyroglobulin making possible more precise structure/function investigations.…”

Section: Discussionmentioning

confidence: 99%

“…mAb 7, 8 and 9 were initially characterized by Piechaczyk et al (1987). mAb 8 recognizes native thyroglobulin and, more specifically, the N-terminal hormone-forming site, which corresponds to a 25-kDa peptide (Asnl -Met171) obtained after CNBr treatment of the native molecule (Mallet et al, 1992). This antigenic region, identified as cluster 11, was shown to be the major immunodominant thyroglobulin domain in autoimmune thyroid diseases (Piechaczyk et al, 1987).…”

Section: Characteristics Of Mabmentioning

confidence: 99%

Antigenic Mapping of Human Thyroglobulin

Erregragui

Prato

Miquelis

et al. 1997

European Journal of Biochemistry

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We characterized 26 mAb to human thyroglobulin to obtain a topographic map o f the thyroglobulin antigenic surface. Among these mAb, three bind thyroglobulin peptides that are located in the primary sequence of thyroglobulin at either the N terminus or in the middle part of the molecule, three bind thyroglobulin via epitopes comprising the thyroid-hormone moiety, and three bind thyroglobulin through epitopes involved in the recognition of the molecule by its receptor. The 18 remaining mAb bind thyroglobulin through undetermined epitopes ; most of these epitopes are resistant to trypsinization. We used two methods to map the antigenic regions of thyroglobulin: all 26 mAb were grouped, by means of crossinhibition experiments, in 11 clusters corresponding to 11 antigenic regions of the thyroglobulin surface; by means of thyroglobulin peptides of decreasing size, obtained by time-controlled tryptic digestion, we analyzed the relative distance between pairs of epitopes in sandwich immunoassays. By combining these two methods, we organized most of the 11 antigenic regions on a topographic representation of the thyroglobulin surface. This new topographic map of thyroglobulin led us to some unexpected features of the thyroglobulin structure. First, antigenic region 8 located far from the N-terminal region is in close contact with two remote N-terminal antigenic regions (1 and 4), both involved in hormone formation. This antigenic region is likely to play a role in the correct positioning of hormonogenic tyrosines so as to optimize iodination-coupling reactions. Secondly, the domain involved in the binding of thyroglobulin to its receptor, probed by three mAb, is shared by two distinct mid-molecule antigenic regions, one being the main autoantigenic region of thyroglobulin.Keywords: thyroglobulin ; monoclonal antibody ; antigen ; hormone ; receptor.Human thyroglobulin is the precursor of thyroid-hormones and is synthesized exclusively in the thyroid. It is a large dimeric molecule (660 kDa) and the most complex glycoprotein isolated (Malthiery et al., 1989). Post-translational events leading to mature thyroglobulin include disulfide-bond formation, glycosylation, sulfation, phosphorylation, and the formation of thyroidhormones thyroxine and 3,5,3-triiodothyronine. This latter event occurs by iodination of tyrosyl residues to monoiodotyrosyl or diiodotyrosyl, followed by a coupling reaction in which two iodotyrosyl residues form a hormone residue. Both of these reactions are catalyzed by thyroid peroxidase at the apical membrane of the thyrocyte in the follicular lumen (Taurog, 1991). Of the 67 tyrosyl residues of the thyroglobulin monomer, only five undergo iodination and coupling to yield iodothyronines (residues 24, 704, 1309, 2572 and 2765). Nine other tyrosyl residues are iodinated to iodotyrosyls and are assumed to serve either as donor residues for the coupling reaction or as storage sites for organic iodine (Lamas et al., 1989). It is thus assumed that hormone formation is highly dependent on the structural enviro...

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“…In this work, we used the N-terminal domain of human Tg because after separation from the core molecule, this peptide maintains most of its initial three-dimensional conformation, as demonstrated by surface epitope mapping [14]. Furthermore, it remains able to form thyroid hormone according to the type of N-glycan structures borne at Asn 57 and Asn 91 [5].…”

Section: Discussionmentioning

confidence: 99%

Dityrosine bridge formation and thyroid hormone synthesis are tightly linked and are both dependent on N‐glycans

et al. 1996

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Formation of dityrosine bridges is a ubiquitous process mainly attributed to oxidative stress leading to protein degradation and cellular damages. Here we show that dityrosine formation is involved in a physiological process, thyroid hormone synthesis, and is strictly dependent on structural characteristics, namely N-glycans, presented by the protein acting as the prothyroid hormone. We used two isoforms of the N-terminal thyroid hormone forming domain (NTD) of human thyroglobulin: one without N-glyean (19 kDa isoform) and the other with high mannose type structures (25 kDa isoform). Both isoforms were able to form iodotyrosines after in vitro iodination. However, iodotyrosine coupling to form thyroxine did not occur with the unglycosylated 19 kDa NTD. In contrast, the 25 kDa isoform formed thyroxine. Strikingly, thyroxine synthesis was accompanied by dimerization of the 25 kDa isoform and formation of a dityrosine bridge; none of this was observed with the 19 kDa isoform. Taken as a whole, our results indicate that dimerization through dityrosine bridging accompanies and could have a role in thyroid hormone synthesis.Key words: Thyroid; Hormone synthesis; Glycosylation; Dityrosine ecular heterogeneity. This precludes detailed analysis of the various domains in terms of structure-activity relationship. To circumvent this problem, we focused on the N-terminal domain (NTD) of human Tg, which contains the preferential site of hormone synthesis located at Tyr 5 [4]. This peptide, obtained after CNBr treatment of a poorly iodinated Tg, is able to form hormone (T4) by in vitro iodination and iodotyrosine coupling. It has an apparent molecular mass ranging from 19 to 25 kDa according to the number of oligosaccharide side chains linked at Asn 57 and Asn 9x [5]. The large variability of the N-glycan structures borne by the NTD offered an interesting opportunity to determine the role of N-glycans in thyroid hormone synthesis and their possible association with the dimerization process through formation of dityrosine bridges. We selected two types of NTD: the first was unglycosylated and the second was rich in high mannose structures. We observed that dityrosine bridge formation and thyroid hormone synthesis are both dependent on the N-glycan structures and both the results of the same oxidative process under the action of TPO and the H202 generating system. Materials and mcthods

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Tyrosine iodination and iodotyrosyl coupling of the N-terminal thyroid hormone forming site of human thyroglobulin modulate its binding to auto- and monoclonal antibodies

Cited by 15 publications

References 20 publications

Defining thyrotropin-dependent and -independent steps of thyroid hormone synthesis by using thyrotropin receptor-null mice

Defining thyrotropin-dependent and -independent steps of thyroid hormone synthesis by using thyrotropin receptor-null mice

Antigenic Mapping of Human Thyroglobulin

Dityrosine bridge formation and thyroid hormone synthesis are tightly linked and are both dependent on N‐glycans

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