Structure of the Carboxy-Terminal Region of Thyroid Hormone Nuclear Receptor and Its Possible Role in Hormone-Dependent Intermolecular Interactions

Bhat, Manoj Kumar; McPhie, Peter; Ting, Yuan‐Tsang; Zhu, Xuguang; Cheng, Sheue-yann

doi:10.1021/bi00033a034

Cited by 31 publications

(25 citation statements)

References 35 publications

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“…The following plasmids were used in the experiments: pCMX expression vectors containing wt human TR 1 (pCMX-hTR 1), the natural mutants TR 1F451X (Miyoshi et al 1995), R338W (Sasaki et al 1993), K443E (Sasaki et al 1992, G345R (Sakurai et al 1989), E449X (Miyoshi et al 1998 and L456fs (Bhat et al 1995), and the artificially constructed mutants TR 1 L428R (Nagaya & Jameson 1993), R338W/K443E (Andoh et al 1996), R338W/F451X (Andoh et al 1996) and G345R/K443E (Andoh et al 1996). Their T 3 -binding, DNA-binding and transcriptional activities had been reported previously.…”

Section: Plasmidsmentioning

confidence: 99%

Very strong correlation between dominant negative activities of mutant thyroid hormone receptors and their binding avidity for corepressor SMRT

Matsushita

Misawa²,

Andoh³

et al. 2000

Journal of Endocrinology

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The syndrome of resistance to thyroid hormone (RTH) is an inherited disorder involving a mutation of the thyroid hormone receptor (TR) gene. Mutant (m) TR inhibits wild-type (wt) TR functions in a dominant negative manner, and this dominant negative effect (DNE) is a crucial factor in RTH pathogenesis. The molecular mechanism of the DNE is still unclear, although several possibilities (including competition between wt-and mTRs at the T 3 response element (TRE), sequestration of TR-associated protein(s) and titration out of functional TR) have been considered. Here we report that the DNE of mTRs is strongly correlated with their binding avidity for the retinoid X receptor (RXR), and especially for corepressor SMRT (silencing mediator for retinoid and thyroid hormone receptor), but not for the nuclear receptor corepressor, NCoR. The DNE of six natural TRs and four artificially constructed mTRs was assayed using a TR reporter gene containing TRE-DR4 (DR=direct repeat), TRE-pal (pal=palindrome) or TRE-lap (lap=inverted palindrome) in CV1 cells treated with 10 nM T 3 . Of the mTRs examined, F451X (with a carboxy-terminal 11-amino-acid truncation) identified in a patient with RTH exhibited the strongest DNE on all TREs. The binding affinities between mTRs and corepressors SMRT or NCoR were quantified using a two-hybrid interference assay system consisting of VP16-TR(LBD) (LBD=ligand binding domain) and Gal4(DBD)-SMRT (DBD=DNA binding domain), or Gal4(DBD)-NCoR respectively, together with the Gal4 reporter gene. In this assay, VP16-TR(LBD) and Gal4(DBD)-SMRT (or Gal4 (DBD)-NCoR) interact with each other and trans-activate the Gal4 reporter gene. When an equal amount of mTR is coexpressed, it reduces the transcriptional activity of the reporter gene, depending on its binding avidity for a corepressor. A very strong correlation was observed between the SMRT-binding activity and the potency of the DNE among six natural mTRs and also among all mTRs, including four artificially constructed ones. The relationship between NCoR and DNE, however, was not significant. When we assayed the binding avidity of mTRs for RXR by using a two-hybrid assay system consisting of Gal4(DBD)-RXR(LBD) and VP16-TR(LBD), a significant correlation between DNE and binding avidity for the RXR was also observed. These results suggest that a corepressor plays an important role in DNE pathogenesis.

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Section: Plasmidsmentioning

confidence: 99%

Very strong correlation between dominant negative activities of mutant thyroid hormone receptors and their binding avidity for corepressor SMRT

Matsushita

Misawa²,

Andoh³

et al. 2000

Journal of Endocrinology

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“…Therefore, this TR␤ mutant is more likely to produce phenotypic changes in the mouse. Second, unlike the missense mutations or single amino acid deletion of TR␤ found in other patients, this unique frame-shifted mutated sequence is immunogenic, for which high-affinity specific antibodies have been developed (16). These antibodies should allow us to ask questions such as whether there is a correlation of the severity of RTH with the level of expression of mutant protein in affected tissues.…”

mentioning

confidence: 99%

Mice with a targeted mutation in the thyroid hormone β receptor gene exhibit impaired growth and resistance to thyroid hormone

Kaneshige

Zhu

et al. 2000

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

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251

261

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Patients with mutations in the thyroid hormone receptor ␤ (TR␤) gene manifest resistance to thyroid hormone (RTH), resulting in a constellation of variable phenotypic abnormalities. To understand the molecular basis underlying the action of mutant TR␤ in vivo, we generated mice with a targeted mutation in the TR␤ gene (TR␤PV; PV, mutant thyroid hormone receptor kindred PV) by using homologous recombination and the Cre͞loxP system. Mice expressing a single PV allele showed the typical abnormalities of thyroid function found in heterozygous humans with RTH. Homozygous PV mice exhibit severe dysfunction of the pituitary-thyroid axis, impaired weight gains, and abnormal bone development. This phenotype is distinct from that seen in mice with a null mutation in the TR␤ gene. Importantly, we identified abnormal expression patterns of several genes in tissues of TR␤PV mice, demonstrating the interference of the mutant TR with the gene regulatory functions of the wild-type TR in vivo. These results show that the actions of mutant and wild-type TR␤ in vivo are distinct. This model allows further study of the molecular action of mutant TR in vivo, which could lead to better treatment for RTH patients. The thyroid hormone 3,3Ј,5-triiodo-L-thyronine (T3) regulates growth, development, and differentiation. Its actions are mainly mediated through thyroid hormone receptors (TRs), which are ligand-dependent transcription factors (1, 2). Three ligand-activated TR isoforms have been identified, TR␣1, TR␤1, and TR␤2, which are derived from the TR␣ and TR␤ genes, respectively. Each TR isoform has a unique developmental and tissue-specific expression (1, 2). TR binds to specific DNA sequences known as thyroid hormone response elements (TRE) in the promoter regions of T3 target genes. The transcriptional activity of TR depends not only on the types of TRE but also on a host of coregulatory proteins (3).RTH is a syndrome characterized by reduced sensitivity of tissues to the action of thyroid hormone. This condition is characterized by elevated levels of circulating thyroid hormones associated with normal or high levels of serum thyroidstimulating hormone (TSH) (4). The most common form of RTH is familial with autosomal dominant inheritance (4). Patients are usually heterozygotes with only one mutant TR␤ gene, and the symptoms are mild. Moreover, clinical manifestations are variable among families with RTH and also among affected family members. Some of the clinical features that have been reported include goiter, short stature, decreased weight, tachycardia, hearing loss, attention deficit-hyperactivity disorder, decreased IQ, and dyslexia (4). One single patient homozygous for a mutant TR␤ has been reported (5). This patient displayed a complex phenotype of extreme RTH with very high levels of thyroid hormone and TSH. Most TR␤ mutants derived from RTH patients have reduced T3-binding affinities and transcriptional capacities. These TR␤ mutants exhibit a dominant-negative effect when cotransfected with wild-type TRs (6, 7). TR␤ mutan...

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“…previously of a 2-fold increase in the transcriptional activity of the transfected TR␤1 by OA (17). Thus, the induced TR␤1 was functional, and its transcriptional activity was enhanced by phosphorylation.…”

Section: Cell Type-and Tr Isoform-dependent Induction Of Expression Omentioning

confidence: 90%

“…The epitope for mAb J52 is in the middle of the A/B domain (18). To be certain that the band detected in lane 7 was TR␤1, we also used another monoclonal antibody, C4, whose epitope is located at the COOHterminal Glu 457 -Val-Phe-Glu-Asp 461 (17). As shown in lane 10, a protein band with the same electrophoretic mobility as for transfected h-TR␤1 (lanes 12 and 6) was detected which was not seen in the absence of OA (lane 4).…”

Section: Cell Type-and Tr Isoform-dependent Induction Of Expression Omentioning

confidence: 99%

Tissue-specific Stabilization of the Thyroid Hormone β1 Nuclear Receptor by Phosphorylation

Ting

Bhat

Wong

et al. 1997

Journal of Biological Chemistry

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The present study evaluated the expression and regulation of endogenous thyroid hormone receptors (TRs) in cultured cells. In COS-1 cells, the endogenous TR, subtype ␤1 (TR␤1), but not subtype ␤2 or ␣1, was induced to express by okadaic acid (OA) in a concentrationdependent manner. The induced TR␤1 had immunoreactivity and partial V8 proteolytic maps similar to those of the transfected and in vitro translated human TR␤1 (h-TR␤1). The OA-induced expression of endogenous TR␤1 was, however, not observed in a variety of other cultured cell lines tested, indicating that the induction was cell type-dependent. TR␤1 induced by OA was a multisite phosphorylated protein, in which serine and threonine in a ratio of 10:1 were phosphorylated. The induced TR␤1 was functional as it could mediate the thyroid hormone-dependent transcriptional activity via several thyroid hormone response elements. The induction of endogenous TR␤1 expression by OA was not accompanied by an increase in mRNA levels but was the result of an increase in the stability of the TR␤1 protein. This is the first report to indicate that one of the mechanisms by which the TR isoforms are differentially expressed is via the tissue-specific stabilization of the TR isoform proteins. Furthermore, this selective stability of TR␤1 could be conferred by phosphorylation.Thyroid hormone nuclear receptors (TRs) 1 are members of the steroid hormone/retinoic acid receptor superfamily. Two TR genes, TR␣ and TR␤, have been identified which are located on chromosome 17 and chromosome 3, respectively. Four isoforms, ␤1, ␤2, ␣1, and ␣2, are generated from each of two TR genes, ␣ and ␤, by alternative splicing (1, 2). They are transcription factors that regulate the expression of target genes by interacting with specific DNA sequences known as thyroid hormone response elements (TREs) in the promoter region of target genes (1, 2). The transcriptional activity of TRs is not only dependent on thyroid hormone 3,3Ј,5-triiodo-L-thyronine (T 3 ) but also on the type of TRE. Recent studies have indicated that the transcriptional activity of TRs is further modulated via interaction with four groups of cellular proteins: (a) members of the nuclear receptor superfamily, notably the retinoid X receptors (1, 2); (b) corepressors including p270/N-CoR (3), SMRT (4), TRUP (5), SHP (6), and TRACs (7); (c) coactivator SRC-1 (8); and (d) the tumor suppressor p53 (9). It is clear that regulation of the transcriptional activity of TRs is more complex than envisioned previously.One important but less well studied mode of modulation of TR activity is the regulation of TR expression at the protein level. TR␣1 and TR␤1 are differentially expressed at different developmental stages and at different levels in various tissues (10 -12). Moreover, using the specific T 3 binding activity and immunoreactivity as measures of the expressed receptor proteins, it was found that there are marked variations in the TR protein:mRNA ratios in different tissues (11, 12), suggesting isoform-and tissue-dependent posttra...

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Structure of the Carboxy-Terminal Region of Thyroid Hormone Nuclear Receptor and Its Possible Role in Hormone-Dependent Intermolecular Interactions

Cited by 31 publications

References 35 publications

Very strong correlation between dominant negative activities of mutant thyroid hormone receptors and their binding avidity for corepressor SMRT

Very strong correlation between dominant negative activities of mutant thyroid hormone receptors and their binding avidity for corepressor SMRT

Mice with a targeted mutation in the thyroid hormone β receptor gene exhibit impaired growth and resistance to thyroid hormone

Tissue-specific Stabilization of the Thyroid Hormone β1 Nuclear Receptor by Phosphorylation

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