Dissecting the Relation between a Nuclear Receptor and GATA: Binding Affinity Studies of Thyroid Hormone Receptor and GATA2 on TSHβ Promoter (original) (raw)
Related papers
Plos One, 2010
Background: Much is known about how genes regulated by nuclear receptors (NRs) are switched on in the presence of a ligand. However, the molecular mechanism for gene down-regulation by liganded NRs remains a conundrum. The interaction between two zinc-finger transcription factors, Nuclear Receptor and GATA, was described almost a decade ago as a strategy adopted by the cell to up-or down-regulate gene expression. More recently, cell-based assays have shown that the Zn-finger region of GATA2 (GATA2-Zf) has an important role in down-regulation of the thyrotropin gene (TSHb) by liganded thyroid hormone receptor (TR).
Regulation of gene transcription by the thyroid hormone receptors
2006
Thyroid hormone (TH) has important effects on postnatal development as well as on adult metabolic homeostasis. TH meditates its effects through different isoforms of the thyroid hormone receptors (TRs) encoded by the TRalpha and TRbeta genes, respectively. TRs belong to the nuclear hormone receptor superfamily. These receptors are ligand-modulated transcription factors that bind to specific DNA elements located in the regulatory regions of target genes. The nature of these elements determines the effect TR will have on transcription. On what is called a positive TH response element (pTRE), the un-liganded receptor binds to DNA and represses transcription. Hormone binding induces a conformational change in the receptor, allowing for activation of transcription. In contrast, on a negative TH response element (nTRE), activation of transcription is mediated by the un-liganded receptor, whereas the liganded receptor represses transcription. The mechanism behind the regulation of a pTRE i...
The Journal of Steroid Biochemistry and Molecular Biology, 2005
Nuclear receptor corepressor (N-CoR) regulates gene expression through interaction with DNA-bound nuclear receptors, recruiting multicomponent repressor complexes to the sites of target genes. We recently reported the presence of an LXXLL motif in N-CoR, and showed that this motif interacts in vitro and in vivo with retinoic acid receptor ␣ (RAR␣) and thyroid hormone receptor  (TR). Transient transfection experiments now suggest that TR and N-CoR act synergistically and may both be required for ligand-induced repression from the negative TR response element in the thyroid stimulating hormone- (TSH) gene promoter. Mutation of the LXXLL motif in N-CoR abolished ligand-induced repression at this response element. Furthermore, in vitro binding of N-CoR to a complex between TR and the negative TR response element was strictly ligand-dependent. We conclude that N-CoR and TR cooperate in the regulation of the TSH gene and that the ligand-dependent repression is mediated by the LXXLL motif in N-CoR.
Molecular Endocrinology, 1998
We have employed a chimeric receptor system in which we cotransfected yeast GAL4 DNA-binding domain/retinoid X receptor β ligand-binding domain chimeric receptor (GAL4RXR), thyroid hormone receptor-β (TRβ), and upstream activating sequence-reporter plasmids into CV-1 cells to study repression, derepression, and transcriptional activation. In the absence of T3, unliganded TR repressed transcription to 20% of basal level, and in the presence of T3, liganded TRβ derepressed transcription to basal level. Using this system and a battery of TRβ mutants, we found that TRβ/RXR heterodimer formation is necessary and sufficient for basal repression and derepression in this system. Additionally, an AF-2 domain mutant (E457A) mediated basal repression but not derepression, suggesting that interaction with a putative coactivator at this site may be critical for derepression. Interestingly, a mutant containing only the TRβ ligand binding domain (LBD) not only mediated derepression, but also stimu...
Regulatory functions of a non-ligand-binding thyroid hormone receptor isoform
Cell regulation, 1991
Gene regulation by thyroid hormones is mediated through multiple nuclear receptors. Only some of these thyroid hormone receptor (TR) isoforms become transcriptional enhancers in the presence of the thyroid hormone T3. Here we analyze the regulatory function of the human TRa2 isoform. This protein does not bind T3 and is not a transcriptional activator of thyroid hormone-responsive elements (TRE). Transfected TRa2 functions as a constitutive repressor of the transcriptional activators TRal and TR,81 but also represses heterologous receptors, including the retinoic acid receptor and the estrogen receptor, which can activate TRE-controlled genes. TRa2 protein showed strongly reduced DNA binding to a palindromic TRE when compared with the active TRs. Hybrid receptor analysis revealed that the special properties of the TRa2 protein, including its repressor function and DNA binding characteristics, are intrinsic properties of its carboxyterminus and can be transferred to other receptors. Although it has been shown that the active TRs can act as repressors and silencers due to their strong DNA binding in the absence of hormone, our data show that TRa2 is unlikely to inhibit TRs and other receptors through a competitive DNA binding mechanism. Antibody gel shift experiments suggest that repression by TRa2 might result from interaction with active receptors. Thus, the receptorlike TRa2 isoform differs from typical nuclear receptors in its DNA-binding and ligand-binding properties and appears to regulate the activity of other receptors via protein-protein interaction.
Molecular Endocrinology, 2012
Thyroid hormones have a profound influence on human development and disease. The hypothalamic-pituitary-thyroid axis involves finely tuned feedback mechanisms to maintain thyroid hormone (TH) levels. Despite the important role of TH-negative feedback in regulating this axis, the mechanism by which this occurs is not clearly defined. Previous in vivo studies suggest separate roles for the two thyroid hormone receptor isoforms, THRA and THRB, in this axis. We performed studies using a unique pituitary thyrotroph cell line (T␣T1.1) to determine the relative roles of THRA and THRB in the regulation of Tshb. Using chromatin immunoprecipitation assays, we found that THRB, not THRA, bound to the Tshb promoter. By selectively depleting THRB, THRA, or both THRA and THRB in T␣T1.1 cells, we found that simultaneous knockdown of both THRB and THRA abolished T 3-mediated down-regulation of Tshb at concentrations as high as 100 nM T 3. In contrast, THRA knockdown alone had no effect on T 3-negative regulation, whereas THRB knockdown alone abolished T 3-mediated down-regulation of Tshb mRNA levels at 10 nM but not 100 nM T 3 concentrations. Interestingly, chromatin immunoprecipitation assays showed that THRA becomes enriched on the Tshb promoter after knockdown of THRB. Thus, a likely mechanism for the differential effects of THR isoforms on Tshb may be based on their differential DNA-binding affinity to the promoter.
Molecular and Cellular Biology, 1997
A group of transcriptional cofactors referred to as corepressors (CoRs) were recently shown to play a central role in basal silencing of genes that contain positive triiodothyronine (T3) response elements. In a reciprocal manner, negatively regulated genes are stimulated by unliganded thyroid hormone receptor (TR) and repressed upon the addition of T3. We used a TR beta mutant, called P214R, which fails to interact with CoRs, to examine whether CoRs also play a role in the control of genes that are negatively regulated in response to T3. In studies of three negatively regulated genes (the pituitary thyroid-stimulating hormone alpha-subunit [TSH alpha], TSH beta, and hypothalamic thyrotropin-releasing hormone [TRH] genes), stimulation of basal promoter activity by unliganded TR beta was impaired by introducing the P214R CoR mutation. Coexpression of each of the CoRs SMRT (silencing mediator for retinoid receptors and TRs) and NCoR (nuclear receptor CoR) enhanced basal stimulation of ...
Journal of Biological …, 1996
Recently, a family of nuclear co-repressor proteins (TRACs) have been identified that interact with thyroid hormone (TR) and retinoic acid receptors to mediate ligand-independent repression of gene transcription. In this report, we have cloned and characterized a human TRAC, which when expressed as a truncated protein lacking its repressing domains, can abolish endogenous cellular TRAC activity. Use of this inhibitor has uncovered a differential function of TRACs on negative versus positive thyroid hormone response elements and has demonstrated the importance of the TR A/B domain in modulating TRAC function. Thus, isoform-specific functions of the TR may be mediated by their functional interaction with co-repressor proteins.