Cooperative binding of steroid hormone receptors contributes to transcriptional synergism at target enhancer elements

doi:10.1016/0092-8674(89)90919-7

Cell

Volume 57, Issue 3, 5 May 1989, Pages 443-448

https://doi.org/10.1016/0092-8674(89)90919-7 Get rights and content

Abstract

We demonstrated previously that two molecules of steroid hormone receptor bound efficiently to a single hormone response element ( $GRE PRE$ ) of the tyrosine aminotransferase gene (Tsai et al., 1988). Here, we show that two tandemly linked $GRE PREs$ conferred progesterone inducibility synergistically to a heterologous TK-CAT fusion gene. Binding studies demonstrated that occupation of one $GRE PRE$ site by a progesterone receptor dimer increased the binding affinity of receptors for the second $GRE PRE$ site 100-fold. Thus, the observed synergistic induction of TK-CAT may result from cooperative binding of receptor dimers to the two $GRE PRE$ sites.

References (36)

A. Bender et al.
MATα1 protein, a yeast transcription activator, binds synergistically with a second protein to a set of cell-type-specific genes
Cell
(1987)
M. Bienz et al.
Mechanism of heat shock gene activation in higher eukaryotes
Adv. Genet.
(1987)
M.S. Bradshaw et al.
A far upstream ovalbumin enhancer binds nuclear factor-1-like factor
J. Biol. Chem.
(1988)
G. Chalepakis et al.
Differential gene activation by glucocorticoids and progestins through the hormone regulatory element of mouse mammary tumor virus
Cell
(1988)
L.A. Chodosh et al.
Human CCAAT-binding proteins have heterologous subunits
Cell
(1988)
T. Curran et al.
Fos and Jun: the AP-1 connection
Cell
(1988)
C. Fromental et al.
Cooperativity and hierarchical levels of functional organization in the SV40 enhancer
Cell
(1988)
C.K. Glass et al.
The thyroid hormone receptor binds with opposite transcriptional effects to a common sequence motif in thyroid hormone and estrogen response elements
Cell
(1988)
W. Herr et al.
The SV40 enhancer is composed of multiple functional elements that can compensate for one another
Cell
(1986)
M. Horikoshi et al.
Transcription factor ATF interacts with the TATA factor to facilitate establishment of a preinitiation complex
Cell
(1988)

H.-M. Jantzen et al.

Cooperativity of glucocorticoid response elements located far upstream of the tyrosine aminotransferase gene

Cell

(1987)

L. Klein-Hitpaß et al.

An estrogen-responsive element derived from the 5′-flanking region of the Xenopus vitellogenin A2 gene functions in transfected human cells

Cell

(1986)

R. Renkawitz et al.

Sequences in the promoter region of the chicken lysozyme gene required for steroid regulation and receptor binding

Cell

(1984)

J. Topol et al.

Sequences required for in vitro transcriptional activation of a Drosophila hsp 70 gene

Cell

(1985)

S.Y. Tsai et al.

Interactions between a DNA-binding transciption factor (COUP) and a non-DNA binding factor (S300-II)

Cell

(1987)

S.Y. Tsai et al.

Molecular interactions of steroid hormone receptor with its enhancer element: evidence for receptor dimer formation

Cell

(1988)

W. Ankenbaule et al.

Synergistic action of glucocorticoid and estradiol responsive elements

O.M. Conneely et al.

Sequence and expression of a functional chicken progesterone receptor

J. Mol. Endo.

(1987)

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Histone deacetylase inhibitors down-regulate G-protein-coupled estrogen receptor and the GPER-antagonist G-15 inhibits proliferation in endometriotic cells
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To investigate whether histone deacetylase inhibitors reduce the expression of the G-protein-coupled estrogen receptor (GPER) and whether the functional inhibition of GPER by the antagonist G-15 decreases the proliferation of endometriotic cells.
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Immortalized epithelial endometriotic cells.
Treatment with the histone deacetylase inhibitor romidepsin or suberoylanilide hydroxamic acid (SAHA), or with the GPER antagonist G-15.
Western blot analysis and quantitative real-time polymerase chain reaction (PCR) were used to monitor the expression of GPER in response to drug treatment. Effects of GPER stimulation and inhibition on cell proliferation were investigated by the 93-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide (Sigma) (MTT) assay.
Our results demonstrate that romidepsin and SAHA reduce GPER expression in a concentration-dependent manner. This reduction correlated with the accumulation of acetylated histones. No decreased expression of the estrogen receptor (ER)-α and ERβ was found under comparable experimental conditions. Pretreatment of endometriotic cells with the GPER agonist G-1 stimulated cell proliferation accompanied by rapid Akt phosphorylation. G-15 reversed this stimulation and inhibited cell proliferation, which was accompanied by Akt dephosphorylation.
G-protein-coupled estrogen receptor is proposed as a potential therapeutic target in endometriosis. The down-regulation of GPER and/or the impairment of its function may reduce the estrogen responsiveness in endometriosis, and therefore might be considered a possible treatment option of endometriosis.
17β-Estradiol promotes the invasion and migration of nuclear estrogen receptor-negative breast cancer cells through cross-talk between GPER1 and CXCR1
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40–70% of breast cancer growth is related to estrogen [1]. Traditionally, estrogen modulates the expression of downstream genes by binding to nuclear estrogen receptors (ER), ERα and ERβ, which function as ligand-activated transcription factors [2–4]. In this regard, the clinical application of ER in breast cancers is a successful paradigm, such as tamoxifen (TAM), an endocrine therapeutic agent targeting ERα, has reduced recurrence and improved survival in patients with different stages of breast cancer [5,6].
G protein-coupled estrogen receptor 1 (GPER1) is widely expressed in human breast cancers correlating with increased tumor size and malignancy. Although estrogen signaling via GPER1 was extensively studied in recent years, the underlying molecular mechanism of GPER1-associated metastasis of breast cancer still remains unclear. In this study, the main aims were focused on the potential role of GPER1 in regulating migration and invasion of nuclear estrogen receptor (ER)-negative breast cancer cells upon 17β-estradiol (E2) stimulation and the involved signaling pathway. Key events in estrogen signaling were chosen for our studies, such as the activation of ERK and AKT, nuclear translocation of NF-κB and secretion of Interleukin-8 (IL-8). The migration and invasion activities upon E2 stimulation were also examined in ER-negative SKBR3 and BT-20 breast cancer cells. Compared with ER-positive MCF-7 breast cancer cells, both SKBR3 and BT-20 cells had very similar expression of GPER1, but relatively high expression of CXC receptor-1 (CXCR1), which is considered as an active regulator for cancer metastasis upon binding IL-8. Results showed that E2 facilitated the activation of ERK, AKT and NF-κB, which could be significantly attenuated by GPER1 blockage or knock-down in both SKBR3 and BT-20 cells. Moreover, increased secretion of IL-8 induced by E2 was also inhibited either by specific inhibitors for GPER1, ERK, AKT, and NF-κB, or by knock-down for GPER1. Furthermore, E2 could activate the migration and invasion of both SKBR3 and BT-20 cells, which in turn could also be inhibited by blocking GPER1, ERK, AKT, NF-κB, and CXCR1, respectively, or knock-down for GPER1 and CXCR1. In conclusion, we demonstrated that estrogen signaling via GPER1 associated with the metastasis of breast cancer, which might be through GPER1/ERK&AKT/NF-κB/IL-8/CXCR1 cascade. The cross-talk between GPER1 and CXCR1 could be another potential target for the therapy of metastatic breast cancer.
G protein-coupled estrogen receptor mediates the Up-regulation of fatty acid synthase induced by 17β-estradiol in cancer cells and cancer-associated fibroblasts
2012, Journal of Biological Chemistry
Activation of lipid metabolism is an early event in carcinogenesis and a central hallmark of many tumors. Fatty acid synthase (FASN) is a key lipogenic enzyme catalyzing the terminal steps in the de novo biogenesis of fatty acids. In cancer cells, FASN may act as a metabolic oncogene, given that it confers growth and survival advantages to these cells, whereas its inhibition effectively and selectively kills tumor cells. Hormones such as estrogens and growth factors contribute to the transcriptional regulation of FASN expression also through the activation of downstream signaling and a cross-talk among diverse transduction pathways. In this study, we demonstrate for the first time that 17β-estradiol (E2) and the selective GPER ligand G-1 regulate FASN expression and activity through the GPER-mediated signaling, which involved the EGF receptor/ERK/c-Fos/AP1 transduction pathway, as ascertained by using specific pharmacological inhibitors, performing gene-silencing experiments and ChIP assays in breast SkBr3, colorectal LoVo, hepatocarcinoma HepG2 cancer cells, and breast cancer-associated fibroblasts. In addition, the proliferative effects induced by E2 and G-1 in these cells involved FASN as the inhibitor of its activity, named cerulenin, abolished the growth response to both ligands. Our data suggest that GPER may be included among the transduction mediators involved by estrogens in regulating FASN expression and activity in cancer cells and cancer-associated fibroblasts that strongly contribute to cancer progression.
Chapter 2 Using Thermodynamics to Understand Progesterone Receptor function. Method and Theory
2009, Methods in Enzymology
Citation Excerpt :
For comparison, the results of an identical analysis of PR‐B interactions at the PRE2 promoter are also shown (Heneghan et al., 2006). Although visual inspection of the binding curves suggests consistency with earlier biochemical studies (Onate et al., 1994; Tsai et al., 1989), our computational analysis reveals a number of results inconsistent with the traditional model of receptor function. First, noting the values presented in Table 2.2 it should be evident that the 30 nM apparent binding affinity of PR‐A for the PRE1− promoter (as determined by a half‐saturation value; Fig. 2.4B) has little similarity to the −11.4 kcal/mol or 1 nM intrinsic binding affinity for actual dimer binding.
Progesterone receptors (PRs) are members of the nuclear receptor superfamily of ligand‐activated transcription factors. The mechanisms by which receptors such as PR assemble at a promoter and recruit coactivators are well understood at the biochemical level. However, a rigorous and thus quantitatively predictive understanding of function is entirely lacking. This is so in part because the study of receptor function has largely been carried out using semiquantitative or qualitative approaches. These types of analyses are limited in their ability to resolve thermodynamically valid and physically meaningful microscopic interaction parameters. This includes resolution of intrinsic binding constants and cooperativity terms, as well as the mathematical framework for integrating these values into a larger molecular code for function. Here we present our experimental and theoretical approach for dissecting the linked reactions associated with PR and coactivator assembly at complex promoter sequences. We discuss the use of analytical ultracentrifugation and quantitative DNase footprint titration and their coupling to exact theoretical treatments. We then highlight the major findings of these studies and their implications for understanding and reevaluating receptor function.
Thermodynamic Dissection of Progesterone Receptor Interactions at the Mouse Mammary Tumor Virus Promoter: Monomer Binding and Strong Cooperativity Dominate the Assembly Reaction
2008, Journal of Molecular Biology
Citation Excerpt :
This was despite the fact that, at least for GR, the binding transitions at each response element were extremely steep,4 indicating that additional reactions were somehow coupled to DNA binding (e.g., cooperative assembly or solution dimerization). Furthermore, the results were in contrast to concurrent studies of a synthetic PR-regulated promoter, which indicated the presence of highly cooperative interactions between adjacently bound PR dimers.7 This discordance thus left unanswered the mechanism of receptor-mediated transcriptional synergy and raised the broader question of whether cooperativity was a common feature of PR (and GR) function.
Progesterone receptors (PRs) play critical roles in eukaryotic gene regulation, yet the mechanisms by which they assemble at their promoters are poorly understood. One of the few promoters amenable to analysis is the mouse mammary tumor virus gene regulatory sequence. Embedded within this sequence are four progesterone response elements (PREs) corresponding to a palindromic PRE and three half-site PREs. Early mutational studies indicated that the presence of all four sites generated a synergistic and strong transcriptional response. However, DNA binding analyses suggested that receptor assembly at the promoter occurred in the absence of significant cooperativity. Taken together, the results indicated that cooperative interactions among PREs could not account for the observed functional synergy. More broadly, the studies raised the question of whether cooperativity was a common feature of PR-mediated gene regulation. As a step toward obtaining a quantitative and, thus, predictive understanding of receptor function, we have carried out a thermodynamic dissection of PR A-isoform interactions at the mouse mammary tumor virus promoter. Utilizing analytical ultracentrifugation and quantitative footprinting, we have resolved the microscopic energetics of PR A-isoform binding, including cooperativity terms. Our results reveal a model contrary to that inferred from previous biochemical investigations. Specifically, the binding unit at a half-site is not a receptor dimer but is instead a monomer; monomers bound at half-sites are capable of significant pairwise cooperative interactions; occupancy of all three half-sites is required to cooperatively engage the palindromic-bound dimer; and finally, large unfavorable forces accompany assembly. Overall, monomer binding accounts for the majority of the intrinsic binding energetics and cooperativity contributes an approximately 1000-fold increase in receptor–promoter stability. Finally, the partitioning of cooperativity suggests a framework for interpreting in vivo transcriptional synergy. These results highlight the insight available from rigorous analysis and demonstrate that receptor–promoter interactions are considerably more complex than typically envisioned.
Dexamethasone treatment affects nuclear glucocorticoid receptor and glucocorticoid response element binding activity in liver of rats (Rattus norvegicus) during aging
2007, Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
Aging is associated with marked changes in the biochemical processes of many organs. Basal and glucocorticoid induced of liver nuclear glucocorticoid receptor (GR) on the level of protein expression and DNA-binding activity were investigated at different ages (3, 6, 12, 18 and 24 months old) in two groups of rats in: untreated and dexamethasone treated. The results showed a significant decline of GR protein immunopurified from untreated rats of advanced age. In dexamethasone-treated rats, the quantity of GR protein was lower than in controls at all ages. The interactions of liver nuclear proteins with radioactively labelled synthetic oligonucleotide analogue containing consensus GRE sequence were analysed during aging. The results showed that GRE binding activity demonstrated a decrease both in untreated and in dexamethasone treated rats. However, relative to untreated rats, dexamethasone treatment resulted in a significant increase in GRE binding at all ages, except that of three months old animals. In conclusion, the observed alterations in GR protein expression and its DNA binding activity may play a role in the changes of the cell response to glucocorticoid during aging.

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ArticleCooperative binding of steroid hormone receptors contributes to transcriptional synergism at target enhancer elements

Abstract

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Adv. Genet.

J. Biol. Chem.

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Synergistic action of glucocorticoid and estradiol responsive elements

Sequence and expression of a functional chicken progesterone receptor

J. Mol. Endo.

Article
Cooperative binding of steroid hormone receptors contributes to transcriptional synergism at target enhancer elements