Specific Histone Lysine 4 Methylation Patterns Define TR-Binding Capacity and Differentiate Direct T 3 Responses (original) (raw)
Related papers
Association of thyroid hormone receptors with chromatin
Molecular and Cellular Biochemistry, 1983
A large body of circumstantial evidence indicates that receptors located in nuclei of T 3 responsive tissues represent a site of initiation of thyroid hormone action at the cellular level. Partial characterization of T 3 receptors indicates that these proteins are monomeric structures in nuclei and are chromatin-associated non-histone proteins. Treatment of rat liver nuclei with either pancreatic DNase I or micrococcal nuclease releases T 3 receptors from nuclei in two forms: a predominant (95 400 Mr; 5.5-6.0S) and a minor (265 000-365 000 Mr; 12.5S) nucleoprotein complex. Similar structures are excised from rat kidney, brain, and heart nuclei and from GH~ pituitary cell nuclei by micrococcal nuclease digestion. These endonucleaseexcised receptor-containing complexes are significantly larger than the salt-extracted receptor (50 000 Mr; 3.5S). The presence of DNA and other non-receptor proteins in these structures indicates that T 3 receptors probably function within multimeric complexes in vivo. Although T 3 receptors appear to be associated with DNA between nucleosomes, i.e. linker DNA, it is not entirely clear whether all or only a fraction of T 3 receptors interact with nucleosomal components. The 12.5S receptor-containing nucleoprotein complex may represent T 3 receptors in association with linker DNA and nucleosomal components. T 3 receptors do not appear to be uniformly distributed to all chromatin fractions, but are associated with structures having characteristics of transcriptionally active chromatin. They are found in a region of chromatin which is enriched in RNA polymerase activity, rapidly labeled RNA and non-histone proteins, and depleted of histone H I. This region is also highly sensitive to both micrococcal nuclease and pancreatic DNase I digestion. The association of receptors with transcriptionally active chromatin, however, must be considered provisional until additional details of the precise receptor-chromatin interaction have been established. The recent demonstration of a 20-fold increase in a specific hepatic mRNA four hours following administration ofT 3 to hypothyroid rats indicates that thyroid hormone potentially has very rapid effects on hepatic gene expression. However, significant changes in nuclear protein phosphorylation, nuclear protein composition, and chromatin structure have not been detected within this four-hour period. Thus, effects of T 3 on hepatic gene expression are brought about by local and presumably subtle changes in nuclear function.
Proceedings of The National Academy of Sciences, 1979
Thyroid hormone receptors lose their capability for high-affinity binding of the biologically active triiodothyronine after solubilization and separation from other chromatin proteins. The high-affinity triiodothyronine-binding capacity can be reconstituted by addition of a histone-containing extract of chromatin or purified core histones (H2A, H2B, H3, and H4); a number of other acidic or basic proteins tested were ineffective. The data support a model of the receptor in which a ``core'' receptor subunit that contains a thyroid hormone-binding site interacts with a regulatory subunit, which is possibly a histone or histone-like species. This interaction with the ``core'' subunit enables the resulting ``holo'' receptor to bind biologically active hormones. These data also suggest that histones or related proteins can modulate the activity of nonhistone chromosomal proteins that are involved in regulating the expression of specific genes.
Molecular Endocrinology, 2003
Transcriptional regulation by heterodimers of thyroid hormone receptor (TR) and the 9-cis retinoid X receptor (RXR) is a highly complex process involving a large number of accessory factors, as well as chromatin remodeling. We have used a biochemical approach, including an in vitro chromatin assembly and transcription system that accurately recapitulates ligand-and activation function (AF)-2-dependent transcriptional activation by TR/ RXR␣ heterodimers, as well as in vitro chromatin immunoprecipitation assays, to study the mechanisms of TR-mediated transcription with chromatin templates. Using this approach, we show that chromatin is required for robust ligand-dependent activation by TR. We also show that the binding of liganded TR to chromatin induces promoterproximal chromatin remodeling and histone acetylation, and that histone acetylation is correlated with increased TRdependent transcription. Additionally, we find that steroid receptor coactiva-tors (SRCs) and p300 function synergistically to stimulate TR-dependent transcription, with multiple functional domains of p300 contributing to its coactivator activity with TR. A major conclusion from our experiments is that the primary role of the SRC proteins is to recruit p300/cAMP response element binding protein-binding protein to hormone-regulated promoters. Together, our results suggest a multiple step pathway for transcriptional regulation by liganded TR, including chromatin remodeling, recruitment of coactivators, targeted histone acetylation, and recruitment of the RNA polymerase II transcriptional machinery. Our studies highlight the functional importance of chromatin in transcriptional control and further define the molecular mechanisms by which the SRC and p300 coactivators facilitate transcriptional activation by liganded TR. (Molecular Endocrinology 17: 908-922, 2003) T HE MOLECULAR ACTIONS of thyroid hormone (T 3) are mediated through thyroid hormone receptors (TR␣ and TR). TRs belong to the nuclear receptor (NR) superfamily and play important roles in development, differentiation, homeostasis, and tumorigenesis through their ability to regulate gene expression (1). TRs function as heterodimers with the 9-cis retinoic acid receptor (RXR) and, in the absence of hormone, the heterodimers bind to thyroid hormone response elements (TREs) and actively repress transcription (2, 3). In contrast to unliganded TRs, ligand-bound TRs function as transcriptional activators. Ligand-dependent activation by TR/RXR heterodimers requires an intact activation domain residing at the carboxyl terminus of the TR ligand-binding domain [known as activation function (AF)-2], as well as cellular coactivators (4). Many coactivators have been implicated in T 3dependent activation, including the steroid receptor coactivator (SRC) family of proteins, p300/cAMP response element binding protein-binding protein (CBP), p300/CBP associated factor (PCAF), and the mediatorlike TR associated proteins (TRAP)/vitamin D receptorinteracting proteins (DRIP)/SRB/mediator-containing cofactor complex (SMCC) (hereafter referred to as the TRAP complex; for reviews, see Refs. 5-9). The SRC family contains three highly related and possibly functionally redundant proteins referred to herein under the unified nomenclature SRC-1, SRC-2, and SRC-3 (5).
Molecular and cellular biology, 2002
Previous studies have established an important role of histone acetylation in transcriptional control by nuclear hormone receptors. With chromatin immunoprecipitation assays, we have now investigated whether histone methylation and phosphorylation are also involved in transcriptional regulation by thyroid hormone receptor (TR). We found that repression by unliganded TR is associated with a substantial increase in methylation of H3 lysine 9 (H3-K9) and a decrease in methylation of H3 lysine 4 (H3-K4), methylation of H3 ...
Chromatin remodeling and developmental gene regulation by thyroid hormone receptor
Summary Thyroid hormone (TH) receptors (TRs) are dual function transcription factors. They activate or repress transcription in the presence or absence of TH, respectively. Using the Xenopus laevis oocyte as an in vivo system to assemble TH target promoters into chromatin under conditions mimicking somatic cells, we have shown that transcriptional repression by unliganded TR involves histone deacetylase while transcriptional activation by TH-bound TR leads to chromatin disruption. Using Xenopus laevis development as a developmental model, we have demonstrated that TR is constitutively bound to its target genes in chromatin. Transcriptional activation induced by TH is accompanied by the release of at least one histone deacetylase and increase in local histone acetylation. These studies together with the developmental expression profiles of TR genes suggest that TH-induced changes in chromatin remodeling play an important role in the dual functions of TR in frog development: gene repr...
The EMBO Journal, 1999
We have investigated ligand-dependent negative regulation of the thyroid-stimulating hormone β (TSHβ) gene. Thyroid hormone (T3) markedly repressed activity of the TSHβ promoter that had been stably integrated into GH 3 pituitary cells, through the conserved negative regulatory element (NRE) in the promoter. By DNA affinity binding assay, we show that the NRE constitutively binds to the histone deacetylase 1 (HDAC1) present in GH 3 cells. Significantly, upon addition of T3, the NRE further recruited the thyroid hormone receptor (TRβ) and another deacetylase, HDAC2. This recruitment coincided with an alteration of in vivo chromatin structure, as revealed by changes in restriction site accessibility. Supporting the direct interaction between TR and HDAC, in vitro assays showed that TR, through its DNA binding domain, strongly bound to HDAC2. Consistent with the role for HDACs in negative regulation, an inhibitor of the enzymes, trichostatin A, attenuated T3-dependent promoter repression. We suggest that ligand-dependent histone deacetylase recruitment is a mechanism of the negative-feedback regulation, a critical function of the pituitary-thyroid axis.
Journal of Biological Chemistry, 2007
It is well documented that unliganded thyroid hormone receptor (TR) functions as a transcriptional repressor of specific cellular target genes by acting in concert with a corepressor complex harboring histone deacetylase (HDAC) activity. To fully explore the cofactors that interact with the transcriptionally repressive form of TR, we biochemically isolated a multiprotein complex that assembles on a TR⅐retinoid X receptor (RXR) heterodimer in HeLa nuclear extracts and identified its polypeptide components by mass spectrometry. A subset of TR⅐RXRassociated polypeptides included NCoR, SMRT, TBL1, and HDAC3, which represent the core components of a previously described NCoR/SMRT corepressor complex. We also identified several polypeptides that constitute a DNA-dependent protein kinase (DNA-PK) enzyme complex, a regulator of DNA repair, recombination, and transcription. These polypeptides included the catalytic subunit DNA-PKcs, the regulatory subunits Ku70 and Ku86, and the poly(ADP-ribose) polymerase 1. Density gradient fractionation and immunoprecipitation analyses provided evidence for the existence of a high molecular weight TR⅐RXR⅐corepressor holocomplex containing both NCoR/SMRT and DNA-PK complexes. Chromatin immunoprecipitation studies confirmed that unliganded TR⅐RXR recruits both complexes to the triiodothyronine-responsive region of growth hormone gene in vivo. Interestingly, DNA-PKcs, a member of the phosphatidylinositol 3-kinase family, was found to phosphorylate HDAC3 when the purified TR⅐RXR⅐corepressor holocomplex was incubated with ATP. This phosphorylation was accompanied by a significant enhancement of the HDAC activity of this complex. Collectively, our results indicated that DNA-PK promotes the establishment of a repressive chromatin at a TR target promoter by enhancing the HDAC activity of the receptor-bound NCoR/ SMRT corepressor complex.