scholarly journals The Interaction of TRβ1-N Terminus with Steroid Receptor Coactivator-1 (SRC-1) Serves a Full Transcriptional Activation Function of SRC-1

Endocrinology ◽  
2006 ◽  
Vol 147 (3) ◽  
pp. 1452-1457 ◽  
Author(s):  
Toshiharu Iwasaki ◽  
Akira Takeshita ◽  
Wataru Miyazaki ◽  
William W. Chin ◽  
Noriyuki Koibuchi
Keyword(s):  
Thyroid Hormone ◽  
Nuclear Receptor ◽  
Thyroid Hormone Receptor ◽  
Steroid Receptor ◽  
Activation Function ◽  
Binding Domain ◽  
Expression Vectors ◽  
Response Element ◽  
Steroid Receptor Coactivator ◽  
N Terminus

Steroid receptor coactivator-1 (SRC-1) plays a crucial role in nuclear receptor-mediated transcription including thyroid hormone receptor (TR)-dependent gene expression. Interaction of the TR-ligand binding domain and SRC-1 through LXXLL motifs is required for this action. However, potential interactions between the TRβ1-N terminus (N) and SRC-1 have not been explored and thus are examined in this manuscript. Far-Western studies showed that protein construct containing TRβ1-N + DNA binding domain (DBD) bound to nuclear receptor binding domain (NBD)-1 (amino acid residue, aa 595–780) of SRC-1 without ligand. Mammalian two-hybrid studies showed that NBD-1, as well as SRC-1 (aa 595-1440), bound to TRβ1-N+DBD in the absence of ligand in CV-1 cells. However, NBD-2 (aa 1237–1440) did not bind to this protein. Glutathione-S-transferase pull-down studies showed that TRβ1-N (aa 1–105) bound to the broad region of SRC-1-C terminus. Expression vectors encoding a series of truncations and/or point mutations of TRβ1 were used in transient transfection-based reporter assays in CV-1 cells. N-terminal truncated TRβ1 (ΔN-TRβ1) showed lower activity than that of wild-type in both artificial F2-thyroid hormone response element and native malic enzyme response element. These results suggest that there is the interaction between N terminus of TRβ1 and SRC-1, which may serve a full activation of SRC-1, together with activation function-2 on TRβ1-mediated transcription.

scholarly journals Novel Mechanism of Nuclear Receptor Corepressor Interaction Dictated by Activation Function 2 Helix Determinants

2002 ◽  
Vol 22 (19) ◽  
pp. 6831-6841 ◽  
Author(s):  
Anna N. Moraitis ◽  
Vincent Giguère ◽  
Catherine C. Thompson
Keyword(s):  
Retinoic Acid ◽  
Thyroid Hormone ◽  
Ligand Binding ◽  
Nuclear Receptor ◽  
Thyroid Hormone Receptor ◽  
Activation Function ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Cerebellar Development ◽  
Nuclear Receptor Corepressor

ABSTRACT Transcriptional regulation by nuclear receptors is controlled by the concerted action of coactivator and corepressor proteins. The product of the thyroid hormone-regulated mammalian gene hairless (Hr) was recently shown to function as a thyroid hormone receptor corepressor. Here we report that Hr acts as a potent repressor of transcriptional activation by RORα, an orphan nuclear receptor essential for cerebellar development. In contrast to other corepressor-nuclear receptor interactions, Hr binding to RORα is mediated by two LXXLL-containing motifs, a mechanism associated with coactivator interaction. Mutagenesis of conserved amino acids in the ligand binding domain indicates that RORα activity is ligand-dependent, suggesting that corepressor activity is maintained in the presence of ligand. Despite similar recognition helices shared with coactivators, Hr does not compete for the same molecular determinants at the surface of the RORα ligand binding domain, indicating that Hr-mediated repression is not simply through displacement of coactivators. Remarkably, the specificity of Hr corepressor action can be transferred to a retinoic acid receptor by exchanging the activation function 2 (AF-2) helix. Repression of the chimeric receptor is observed in the presence of retinoic acid, demonstrating that in this context, Hr is indeed a ligand-oblivious nuclear receptor corepressor. These results suggest a novel molecular mechanism for corepressor action and demonstrate that the AF-2 helix can play a dynamic role in controlling corepressor as well as coactivator interactions. The interaction of Hr with RORα provides direct evidence for the convergence of thyroid hormone and RORα-mediated pathways in cerebellar development.

scholarly journals In Vivo Interaction of Steroid Receptor Coactivator (SRC)-1 and the Activation Function-2 Domain of the Thyroid Hormone Receptor (TR) β in TRβ E457A Knock-In and SRC-1 Knockout mice

Endocrinology ◽  
2009 ◽  
Vol 150 (8) ◽  
pp. 3927-3934 ◽  
Author(s):  
Manuela Alonso ◽  
Charles Goodwin ◽  
XiaoHui Liao ◽  
Tania Ortiga-Carvalho ◽  
Danielle S. Machado ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Thyroid Hormone Receptor ◽  
Steroid Receptor ◽  
Activation Function ◽  
Steroid Receptor Coactivator ◽  
Pituitary Thyroid Axis ◽  
Serum T4 ◽  
Thyroid Axis

The activation function-2 (AF-2) domain of the thyroid hormone (TH) receptor (TR)-β is a TH-dependent binding site for nuclear coactivators (NCoA), which modulate TH-dependent gene transcription. In contrast, the putative AF-1 domain is a TH-independent region interacting with NCoA. We determined the specificity of the AF-2 domain and NCoA interaction by evaluating thyroid function in mice with combined disruption of the AF-2 domain in TRβ, due to a point mutation (E457A), and deletion of one of the NCoAs, steroid receptor coactivator (SRC)-1. The E457A mutation was chosen because it abolishes NCoA recruitment in vitro while preserving normal TH binding and corepressor interactions resulting in resistance to TH. At baseline, disruption of SRC-1 in the homozygous knock-in (TRβE457A/E457A) mice worsened the degree of resistance to TH, resulting in increased serum T4 and TSH. During TH deprivation, disruption of AF-2 and SRC-1 resulted in a TSH rise 50% of what was seen when AF-2 alone was removed, suggesting that SRC-1 was interacting outside of the AF-2 domain. Therefore, 1) during TH deprivation, SRC-1 is necessary for activating the hypothalamic-pituitary-thyroid axis; 2) ligand-dependent repression of TSH requires an intact AF-2; and 3) SRC-1 may interact with the another region of the TRβ or the TRα to regulate TH action in the pituitary. This report demonstrates the dual interaction of NCoA in vivo: the TH-independent up-regulation possibly through another domain and TH-dependent down-regulation through the AF-2 domain.

Specificity of thyroid hormone receptor subtype and steroid receptor coactivator-1 on thyroid hormone action

2003 ◽  
Vol 284 (1) ◽  
pp. E36-E46 ◽  
Author(s):  
Peter M. Sadow ◽  
Olivier Chassande ◽  
Karine Gauthier ◽  
Jacques Samarut ◽  
Jianming Xu ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Hormone Receptor ◽  
Thyroid Hormone Receptor ◽  
Steroid Receptor ◽  
Receptor Subtype ◽  
Thyroid Function Tests ◽  
Hormone Action ◽  
Peripheral Tissues ◽  
Steroid Receptor Coactivator ◽  
Thyroid Hormone Action

Isoforms of the thyroid hormone receptor ( TR) α and TRβ genes mediate thyroid hormone action. How TR isoforms modulate tissue-specific thyroid hormone (TH) action remains largely unknown. The steroid receptor coactivator-1 (SRC-1) is among a group of transcriptional coactivator proteins that bind to TRs, along with other members of the nuclear receptor superfamily, and modulate the activity of genes regulated by TH. Mice deficient in SRC-1 possess decreased tissue responsiveness to TH and many steroid hormones; however, it is not known whether or not SRC-1-mediated activation of TH-regulated gene transcription in peripheral tissues, such as heart and liver, is TR isoform specific. We have generated mice deficient in TRα and SRC-1, as well as in TRβ and SRC-1, and investigated thyroid function tests and effects of TH deprivation and TH treatment compared with wild-type (WT) mice or those deficient in either TR or SRC-1 alone. The data show that 1) in the absence of TRα or TRβ, SRC-1 is important for normal growth; 2) SRC-1 modulates TRα and TRβ effects on heart rate; 3) two new TRβ-dependent markers of TH action in the liver have been identified, osteopontin (upregulated) and glutathione S-transferase (downregulated); and 4) SRC-1 may mediate the hypersensitivity to TH seen in liver of TRα-deficient mice.

scholarly journals A Quantitative High-Throughput Screen Identifies Novel Inhibitors of the Interaction of Thyroid Receptor β with a Peptide of Steroid Receptor Coactivator 2

2011 ◽  
Vol 16 (6) ◽  
pp. 618-627 ◽  
Author(s):  
Ronald L. Johnson ◽  
Jong Yeon Hwang ◽  
Leggy A. Arnold ◽  
Ruili Huang ◽  
Jennifer Wichterman ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
High Throughput ◽  
Hormone Receptor ◽  
Mammalian Cells ◽  
Thyroid Hormone Receptor ◽  
Steroid Receptor ◽  
Nuclear Hormone Receptor ◽  
High Throughput Screen ◽  
Data Set ◽  
Steroid Receptor Coactivator

The thyroid hormone receptors (TR) are members of the nuclear hormone receptor (NHR) superfamily that regulate development, growth, and metabolism. Upon ligand binding, TR releases bound corepressors and recruits coactivators to modulate target gene expression. Steroid receptor coactivator 2 (SRC2) is an important coregulator that interacts with TRβ to activate gene transcription. To identify novel inhibitors of the TRβ and SRC2 interaction, the authors performed a quantitative high-throughput screen (qHTS) of a TRβ-SRC2 fluorescence polarization assay against more than 290 000 small molecules. The qHTS assayed compounds at 6 concentrations up to 92 µM to generate titration–response curves and determine the potency and efficacy of all compounds. The qHTS data set enabled the characterization of actives for structure–activity relationships as well as for potential artifacts such as fluorescence interference. Selected qHTS actives were tested in the screening assay using fluoroprobes labeled with Texas Red or fluorescein. The retest identified 19 series and 4 singletons as active in both assays with 40% or greater efficacy, free of compound interference, and not toxic to mammalian cells. Selected compounds were tested as independent samples, and a methylsulfonylnitrobenzoate series inhibited the TRβ-SRC2 interaction with 5 µM IC50. This series represents a new class of thyroid hormone receptor–coactivator modulators.

scholarly journals Modulation by Steroid Receptor Coactivator-1 of Target-Tissue Responsiveness in Resistance to Thyroid Hormone

Endocrinology ◽  
2003 ◽  
Vol 144 (9) ◽  
pp. 4144-4153 ◽  
Author(s):  
Yuji Kamiya ◽  
Xiao-Yong Zhang ◽  
Hao Ying ◽  
Yusuhito Kato ◽  
Mark C. Willingham ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Target Genes ◽  
Thyroid Hormone Receptor ◽  
Expression Patterns ◽  
Steroid Receptor ◽  
Target Tissue ◽  
Dependent Manner ◽  
Resistance To Thyroid Hormone ◽  
Steroid Receptor Coactivator

Abstract Mutations in the thyroid hormone receptor-β gene (TRβ) cause resistance to thyroid hormone. How the action of mutant thyroid hormone nuclear receptors (TRs) is regulated in vivo is not clear. We examined the effect of a TR coactivator, steroid receptor coactivator-1 (SRC-1), on target-tissue responsiveness by using a mouse model of resistance to thyroid hormone, TRβPV knockin mice, in the SRC-1 null background. Lack of SRC-1 intensified the dysfunction of the pituitary-thyroid axis and impaired growth in TRβPV/+ mice but not in TRβPV/PV mice. In TRβPV/PV mice, however, lack of SRC-1 intensified the pathological progression of thyroid follicular cells to papillary hyperplasia, reminiscent of papillary neoplasia. In contrast, lack of SRC-1 did not affect responsiveness in the liver in regulating serum cholesterol in either TRβPV/+ or TRβPV/PV mice. Lack of SRC-1 led to changes in the abnormal expression patterns of several T3 target genes in the pituitary and liver. Thus, the present studies show that a coactivator such as SRC-1 could modulate the in vivo action of TRβ mutants in a tissue-dependent manner.

Mechanisms of androgen receptor signalling via steroid receptor coactivator-1 in prostate.

2004 ◽  
Vol 11 (1) ◽  
pp. 117-130 ◽  
Author(s):  
S M Powell ◽  
V Christiaens ◽  
D Voulgaraki ◽  
J Waxman ◽  
F Claessens ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Nuclear Receptors ◽  
Nuclear Receptor ◽  
Point Mutations ◽  
Steroid Receptor ◽  
Activation Function ◽  
Steroid Receptor Coactivator ◽  
Basal Transcriptional Machinery ◽  
Independent States

The androgen receptor (AR) is a member of the nuclear receptor superfamily. These ligand-activated transcription factors usually contain two activation functions, a ligand-independent activation function 1(AF1) in the divergent N-terminal domain and a ligand-dependent AF2 in the more conserved C-terminal ligand-binding domain. To promote transcription from target promoters, DNA-bound nuclear receptors recruit coactivator proteins that promote transcription by modifying histones within nucleosomes, resulting in altered topology of chromatin to allow access of the basal transcriptional machinery, or stabilising the pre-initiation complex. It is well known that most coactivators interact with AF2 of many nuclear receptors via conserved, helical LxxLL motifs (where L is leucine and x is any amino acid). The AF2 of the AR is very weak, but we were able to demonstrate that its intrinsic ligand-dependent activity is potentiated by steroid receptor coactivator-1 (SRC1) and that this region interacts with coactivators via LxxLL motifs. However, a mutant SRC1 coactivator with no functional LxxLL motifs was still able to potentiate AR activity. We found that SRC1 can also be recruited to (and increase activity of) AF1 of the AR via a conserved, glutamine-rich region. Point mutations within this region abolish SRC1 interaction with AF1 and also abolish or severely impair its ability to potentiate AR activity on all promoters tested. Thus the AR interacts with SRC1 via two different regions and the AF1 interaction is functionally the more important, although the contribution of the two interactions varies in a promoter-dependent fashion. SRC1 then potentiates receptor activity via recruitment of CBP/p300, a histone acetyltranferase. This is important in the context of prostate cancer as SRC1 and other coactivators including CBP are coexpressed with AR in the luminal epithelial cells of the prostate, where over 90% of prostate tumours arise. There is a need for effective second-line prostate cancer therapy aimed at blocking the AR pathway when anti-androgen therapy has failed. Since there is growing evidence that nuclear receptor cofactors may be implicated in the progression of hormone-dependent tumours to hormone-independent states, novel targets could include the interaction of AR with coactivator proteins. We suggest that the N-terminal interaction would be a more specific and effective target in the case of prostate cancer than the LxxLL/AF2 interaction.

CBP-dependent and independent enhancing activity of steroid receptor coactivator-1 in thyroid hormone receptor-mediated transactivation

1999 ◽  
Vol 147 (1-2) ◽  
pp. 103-112 ◽  
Author(s):  
Masato Ikeda ◽  
Akio Kawaguchi ◽  
Akira Takeshita ◽  
William W. Chin ◽  
Toyoshi Endo ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Hormone Receptor ◽  
Thyroid Hormone Receptor ◽  
Steroid Receptor ◽  
Steroid Receptor Coactivator
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