scholarly journals The Rat Thyroid Hormone Receptor (TR) Δβ3 Displays Cell-, TR Isoform-, and Thyroid Hormone Response Element-Specific Actions

Endocrinology ◽  
2007 ◽  
Vol 148 (4) ◽  
pp. 1764-1773 ◽  
Author(s):  
Clare B. Harvey ◽  
J. H. Duncan Bassett ◽  
Padma Maruvada ◽  
Paul M. Yen ◽  
Graham R. Williams
Keyword(s):  
Thyroid Hormone ◽  
Target Genes ◽  
Thyroid Hormone Receptor ◽  
Cell Types ◽  
Binding Activity ◽  
Hormone Response ◽  
Hormone Response Elements ◽  
Hormone Response Element ◽  
Single Transcript ◽  
Rat Thyroid

The THRB gene encodes the well-described thyroid hormone (T3) receptor (TR) isoforms TRβ1 and TRβ2 and two additional variants, TRβ3 and TRΔβ3, of unknown physiological significance. TRβ1, TRβ2, and TRβ3 are bona fide T3 receptors that bind DNA and T3 and regulate expression of T3-responsive target genes. TRΔβ3 retains T3 binding activity but lacks a DNA binding domain and does not activate target gene transcription. TRΔβ3 can be translated from a specific TRΔβ3 mRNA or is coexpressed with TRβ3 from a single transcript that contains an internal TRΔβ3 translation start site. In these studies, we provide evidence that the TRβ3/Δβ3 locus is present in rat but not in other vertebrates, including humans. We compared the activity of TRβ3 with other TR isoforms and investigated mechanisms of action of TRΔβ3 at specific thyroid hormone response elements (TREs) in two cell types. TRβ3 was the most potent isoform, but TR potency was TRE dependent. TRΔβ3 acted as a cell-specific and TRE-dependent modulator of TRβ3 when coexpressed at low concentrations. At higher concentrations, TRΔβ3 was a TRE-selective and cell-specific antagonist of TRα1, -β1, and -β3. Both TRβ3 and TRΔβ3 were expressed in the nucleus in the absence and presence of hormone, and their actions were determined by cell type and TRE structure, whereas TRΔβ3 actions were also dependent on the TR isoform with which it interacted. Analysis of these complex responses implicates a range of nuclear corepressors and coactivators as cell-, TR isoform-, and TRE-specific modulators of T3 action.

scholarly journals Identification of thyroid hormone response elements in vivo using mice expressing a tagged thyroid hormone receptor α1

2013 ◽  
Vol 33 (2) ◽  
Author(s):  
Susi Dudazy-Gralla ◽  
Kristina Nordström ◽  
Peter Josef Hofmann ◽  
Dina Abdul Meseh ◽  
Lutz Schomburg ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Nuclear Receptor ◽  
Hormone Receptor ◽  
Target Genes ◽  
Thyroid Hormone Receptor ◽  
Hormone Response ◽  
Protein Fusion ◽  
Response Elements ◽  
Hormone Response Elements

TRα1 (thyroid hormone receptor α1) is well recognized for its importance in brain development. However, due to the difficulties in predicting TREs (thyroid hormone response elements) in silico and the lack of suitable antibodies against TRα1 for ChIP (chromatin immunoprecipitation), only a few direct TRα1 target genes have been identified in the brain. Here we demonstrate that mice expressing a TRα1–GFP (green fluorescent protein) fusion protein from the endogenous TRα locus provide a valuable animal model to identify TRα1 target genes. To this end, we analysed DNA–TRα1 interactions in vivo using ChIP with an anti-GFP antibody. We validated our system using established TREs from neurogranin and hairless, and by verifying additional TREs from known TRα1 target genes in brain and heart. Moreover, our model system enabled the identification of novel TRα1 target genes such as RNF166 (ring finger protein 166). Our results demonstrate that transgenic mice expressing a tagged nuclear receptor constitute a feasible approach to study receptor–DNA interactions in vivo, circumventing the need for specific antibodies. Models like the TRα1–GFP mice may thus pave the way for genome-wide mapping of nuclear receptor-binding sites, and advance the identification of novel target genes in vivo.

scholarly journals Ligand-dependent and -independent transactivation by thyroid hormone receptor beta 2 is determined by the structure of the hormone response element.

1995 ◽  
Vol 15 (9) ◽  
pp. 4718-4726 ◽  
Author(s):  
M Sjöberg ◽  
B Vennström
Keyword(s):  
Thyroid Hormone ◽  
Hormone Receptor ◽  
Thyroid Hormone Receptor ◽  
Hormone Response ◽  
Response Element ◽  
Response Elements ◽  
Hormone Response Elements ◽  
Hormone Response Element ◽  
Beta 2 ◽  
Receptor Beta

Chicken thyroid hormone receptor beta 2 (cTR beta 2) is likely to serve specific functions in gene regulation since it possesses a unique N-terminal domain and is expressed in very few tissues. We demonstrate here that TR beta 2 exhibits distinct transactivation properties which are dependent on the availability of ligand and on the structure of the hormone response element. First, a strong ligand-independent transactivation was observed with hormone response elements composed of direct repeats and everted repeats. Second, TR beta 2 was induced by triiodothyronine to transactivate more efficiently than TR beta 0 on palindromic and everted-repeat types of hormone response elements. However, coexpression of the retinoid X receptor reduced the strong transactivation by TR beta 2 but not by TR beta 0 via palindromic response elements, suggesting that TR beta 2 can transactivate as a homodimer. Finally, the N terminus of TR beta 2 contains two distinct transactivation regions rich in tyrosines, which are essential for transactivation. Our results thus show that the activity of the novel transactivating region of TR beta 2 is dependent on the organization of the half-sites in the response element.

The functional relationship between co-repressor N-CoR and SMRT in mediating transcriptional repression by thyroid hormone receptor α

2008 ◽  
Vol 411 (1) ◽  
pp. 19-26 ◽  
Author(s):  
Kyung-Chul Choi ◽  
So-Young Oh ◽  
Hee-Bum Kang ◽  
Yoo-Hyun Lee ◽  
Seungjoo Haam ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Hormone Receptor ◽  
Transcriptional Repression ◽  
Fatty Acid Synthase ◽  
Hormone Receptors ◽  
Target Genes ◽  
Functional Relationship ◽  
Thyroid Hormone Receptor ◽  
B Cell Lymphoma ◽  
Hormone Response Elements

A central issue in mediating repression by nuclear hormone receptors is the distinct or redundant function between co-repressors N-CoR (nuclear receptor co-repressor) and SMRT (silencing mediator of retinoid and thyroid hormone receptor). To address the functional relationship between SMRT and N-CoR in TR (thyroid hormone receptor)-mediated repression, we have identified multiple TR target genes, including BCL3 (B-cell lymphoma 3-encoded protein), Spot14 (thyroid hormone-inducible hepatic protein), FAS (fatty acid synthase), and ADRB2 (β-adrenergic receptor 2). We demonstrated that siRNA (small interfering RNA) treatment against either N-CoR or SMRT is sufficient for the de-repression of multiple TR target genes. By the combination of sequence mining and physical association as determined by ChIP (chromatin immunoprecipitation) assays, we mapped the putative TREs (thyroid hormone response elements) in BCL3, Spot14, FAS and ADRB2 genes. Our data clearly show that SMRT and N-CoR are independently recruited to various TR target genes. We also present evidence that overexpression of N-CoR can restore repression of endogenous genes after knocking down SMRT. Finally, unliganded, co-repressor-free TR is defective in repression and interacts with a co-activator, p300. Collectively, these results suggest that both SMRT and N-CoR are limited in cells and that knocking down either of them results in co-repressor-free TR and consequently de-repression of TR target genes.

scholarly journals Transcriptional silencing by unliganded thyroid hormone receptor beta requires a soluble corepressor that interacts with the ligand-binding domain of the receptor.

1996 ◽  
Vol 16 (5) ◽  
pp. 1909-1920 ◽  
Author(s):  
G X Tong ◽  
M Jeyakumar ◽  
M R Tanen ◽  
M K Bagchi
Keyword(s):  
Thyroid Hormone ◽  
Ligand Binding ◽  
Thyroid Hormone Receptor ◽  
Gene Activation ◽  
Transcriptional Silencing ◽  
Binding Domain ◽  
Hormone Response ◽  
Glutathione S Transferase ◽  
Hormone Response Element ◽  
Nuclear Extracts

Unliganded thyroid hormone receptor (TR) functions as a transcriptional repressor of genes bearing thyroid hormone response elements in their promoters. Binding of hormonal ligand to the receptor releases the transcriptional silencing and leads to gene activation. Previous studies showed that the silencing activity of TR is located within the C-terminal ligand-binding domain (LBD) of the receptor. To dissect the role of the LBD in receptor-mediated silencing, we used a cell-free transcription system containing HeLa nuclear extracts in which exogenously added unliganded TRbeta repressed the basal level of RNA polymerase II-driven transcription from a thyroid hormone response element-linked template. We designed competition experiments with a peptide fragment containing the entire LBD (positions 145 to 456) of TRbeta. This peptide, which lacks the DNA-binding domain, did not affect basal RNA synthesis from the thyroid hormone response element-linked promoter when added to a cell-free transcription reaction mixture. However, the addition of the LBD peptide to a reaction mixture containing TRbeta led to a complete reversal of receptor-mediated transcriptional silencing in the absence of thyroid hormone. An LBD peptide harboring point mutations, which severely impair receptor dimerization, also inhibited efficiently the silencing activity of TR, indicating that the relief of repression by the LBD was not due to the sequestration of TR or its heterodimeric partner retinoid X receptor into inactive homo- or heterodimers. We postulate that the LBD peptide competed with TR for a regulatory molecule, termed a corepressor, that exists in the HeLa nuclear extracts and is essential for efficient receptor-mediated gene repression. We have identified the region from positions 145 to 260 (the D domain) of the LBD as a potential binding site of the putative corepressor. We observed further that a peptide containing the LBD of retinoic acid receptor (RAR) competed for TR-mediated silencing, suggesting that the RAR LBD may bind to the same corepressor activity as the TR LBD. Interestingly, the RAR LBD complexed with its cognate ligand, all-trans retinoic acid, failed to compete for transcriptional silencing by TRbeta, indicating that the association of the LBD with the corepressor is ligand dependent. Finally, we provide strong biochemical evidence supporting the existence of the corepressor activity in the HeLa nuclear extracts. Our studies demonstrated that the silencing activity of TR was greatly reduced in the nuclear extracts preincubated with immobilized, hormone-free glutathione S-transferase-LBD fusion proteins, indicating that the corepressor activity was depleted from these extracts through protein-protein interactions with the LBD. Similar treatment with immobilized, hormone-bound glutathione S-transferase-LBD, on the other hand, failed to deplete the corepressor activity from the nuclear extracts, indicating that ligand binding to the LBD disrupts its interaction with the corepressor. From these results, we propose that a corepressor binds to the LBD of unliganded TR and critically influences the interaction of the receptor with the basal transcription machinery to promote silencing. Ligand binding to TR results in the release of the corepressor from the LBD and triggers the reversal of silencing by allowing the events leading to gene activation to proceed.

Regulation of regional expression in rat brain PC2 by thyroid hormone/characterization of novel negative thyroid hormone response elements in the PC2 promoter

2005 ◽  
Vol 288 (1) ◽  
pp. E236-E245 ◽  
Author(s):  
Xiaoxiong Shen ◽  
Qiao-Ling Li ◽  
Gregory A. Brent ◽  
Theodore C. Friedman
Keyword(s):  
Retinoic Acid ◽  
Thyroid Hormone ◽  
Thyroid Hormone Receptor ◽  
Specific Binding ◽  
Gh3 Cells ◽  
Mrna Levels ◽  
Hormone Response ◽  
Thyroid Status ◽  
Response Elements ◽  
Hormone Response Elements

The prohormone convertases (PCs) PC1 and PC2 are involved in the tissue-specific endoproteolytic processing of neuropeptide precursors within the secretory pathway. We previously showed that changes in thyroid status altered pituitary PC2 mRNA and that this regulation was due to triiodothyronine-dependent interaction of the thyroid hormone receptor (TR) with negative thyroid hormone response elements (nTREs) contained in a large proximal region of the human PC2 promoter. In the current study, we examined the in vivo regulation of brain PC2 mRNA by thyroid status and found that 6- n-propyl-2-thiouracil-induced hypothyroidism stimulated, whereas thyroxine-induced hyperthyroidism suppressed, PC2 mRNA levels in the rat hypothalamus and cerebral cortex. To address the mechanism of T3 regulation of the PC2 gene, we used human PC2 (hPC2) promoter constructs transiently transfected into GH3 cells and found that triiodothyronine negatively and 9- cis-retinoic acid positively regulated hPC2 promoter activity. EMSAs, using purified TRα1 and retinoid X receptor-β (RXRβ) proteins demonstrated that TRα bound the distal putative nTRE-containing oligonucleotide in the PC2 promoter, and RXR bound to both nTRE-containing oligonucleotides. EMSAs with oligonucleotides containing deletion mutations of the nTREs demonstrated that the binding to TR and RXR separately is reduced, but specific binding to TR and RXR together persists even with deletion of each putative nTRE. We conclude that there are two novel TRE-like sequences in the hPC2 promoter and that these regions act in concert in a unique manner to facilitate the effects of thyroid hormone and 9- cis-retinoic acid on PC2.

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