The transactivation properties of the two estrogen receptors, ERα and ERβ, were examined with different ligands in context an response element AP1 element. ERβ shown to signal opposite ways when complexed natural hormone estradiol from site: ERα, 17β-estradiol activated transcription, whereas inhibited transcription. Moreover, antiestrogens tamoxifen, raloxifene, Imperial Chemical Industries 164384 potent transcriptional activators at site. Thus, ERs depending on ligand This suggests that...

Combinatorial regulation of transcription implies flexible yet precise assembly multiprotein regulatory complexes in response to signals. Biochemical and crystallographic analyses revealed that hormone binding leads the formation a hydrophobic groove within ligand domain (LBD) thyroid receptor interacts with an LxxLL motif-containing α-helix from GRIP1, coactivator. Residues immediately adjacent motif modulate affinity interaction; sequences are employed different extents receptors. Such...

We find that tamoxifen is a potent activator of estrogen receptor (ER)- mediated induction promoters regulated by AP-1 sites including the human collagenase gene promoter and constructs in which an site fused to herpes thymidine kinase promoter. This contrasts with inability activate otherwise identical bearing classical response elements. Tamoxifen agonism at cell type specific, occurring lines uterine, but not breast, origin. It thus parallels vivo. proteins such as Jun or Jun/Fos are...

The ligand-binding domain of nuclear receptors contains a transcriptional activation function (AF-2) that mediates hormone-dependent binding coactivator proteins. Scanning surface mutagenesis on the human thyroid hormone receptor was performed to define site binds coactivators, glucocorticoid receptor–interacting protein 1 (GRIP1) and steroid (SRC-1). residues involved encircle small hydrophobic cleft. Ligand transcription involves formation this by folding carboxyl-terminal α helix against...

Induction of cyclin D1 gene transcription by estrogen receptor alpha (ERalpha) plays an important role in estrogen-mediated proliferation. There is no classical response element the promoter, and induction ERalpha has been mapped to alternative element, a cyclic AMP-response at -57, with possible participation activating protein-1 site -954. The action ERbeta promoter unknown, although evidence suggests that may inhibit proliferative ERalpha. We examined constructs luciferase assay...

The activity of the AF-2 transcriptional activation function nuclear receptors (NR) is mediated by partially homologous coactivators, glucocorticoid receptor interacting protein 1 (GRIP1)/transcriptional intermediary factor 2 (TIF2) and steroid coactivator (SRC-1). GRIP1 SRC-1 bound nine different NRs exhibited similar, but not identical, NR binding preferences. most striking difference was seen with androgen receptor, which well to poorly SRC-1. contain three copies motif LXXLL (called an...

Estrogen receptors (ERs α and β) enhance transcription in response to estrogens by binding estrogen elements (EREs) within target genes utilizing transactivation functions (AF-1 AF-2) recruit p160 coactivator proteins. The ERs also antiestrogens modulating the activity of AP-1 protein complex. Here, we examine role AF-1 AF-2 ER action at sites. responses sites require integrity ERα activation surfaces complementary on GRIP1 (glucocorticoid receptor interacting 1), NID/AF-1 region, NR boxes....

Estrogen receptor-α contains two transactivation functions, a weak constitutive activation function (AF-1) and hormone-dependent (AF-2). AF-2 works by recruiting large coactivator complex, composed of one or more p160s, CREB-binding protein (CBP)/p300, P/CAF (p300 CBP-associated factor), via direct contacts with the p160s. We report here that independent AF-1 activity also requires p160 contacts. Unlike AF-2, which binds signature NR boxes in center molecule, to sequences near C terminus....

Regulation of nuclear receptor gene expression involves dynamic and coordinated interactions with histone acetyl transferase (HAT) deacetylase complexes. The estrogen (ERα) contains two transactivation domains regulating ligand-independent -dependent transcription (AF-1 AF-2 (activation functions 1 2)). ERα-regulated cointegrators (e.g.p300/CBP, P/CAF) that have the capacity to modify core groups. Here we show ERα is acetylated in vivo.p300, but not P/CAF, selectively directly at lysine...

Members of the 160-kDa nuclear receptor coactivator family (p160 coactivators) bind to conserved AF-2 activation function found in hormone binding domains receptors (NR) and are potent transcriptional coactivators for NRs. Here we report that C-terminal region p160 glucocorticoid interacting protein 1 (GRIP1), steroid (SRC-1a), SRC-1e binds N-terminal AF-1 androgen (AR), can thereby enhance by AR. While they all interact efficiently with AR AF-1, these same have vastly different strengths...

The tumor necrosis factor-alpha (TNF-alpha) promoter was used to explore the molecular mechanisms of estradiol (E(2))-dependent repression gene transcription. E(2) inhibited basal activity and abolished TNF-alpha activation promoter. E(2)-inhibitory element mapped -125 -82 region promoter, known as TNF-responsive (TNF-RE). An AP-1-like site in TNF-RE is essential for activity. Estrogen receptor (ER) beta more potent than ERalpha at repressing -1044 upstream herpes simplex virus thymidine...

We investigated how overexpression of human TATA-box-binding protein (TBP) affects the action estrogen receptor (ER) and compared response with that other activators. When ER activates a simple promoter, consisting element either collagenase or tk TATA box, TBP potentiates transcription. only estrogen-induced not basal transcription does so independent spacing between box. also reduces autoinhibition by overexpressed ER, suggesting one target may be itself. Both AF-1 AF-2 domains are...

Estrogens and glucocorticoids often act in opposition to regulate physiological responses. We investigated whether this might reflect the opposing actions of hormone-bound receptors on target genes regulated by AP-1 response element. performed a series transfection experiments which transcriptional activation, mediated element, was reflected reporter gene activity. As previously described, we found that estrogens stimulate, whereas glucocorticoid dexamethasone (Dex) inhibits, transcription...

Promoter-bound steroid receptors activate gene expression by recruiting members of the p160 family coactivators. Many receptors, most notably progesterone and estrogen are regulated both cognate hormone independently growth factors. Here we show that epidermal factor regulates activities GRIP1 through extracellular signal-regulated kinase (ERK) mitogen-activated protein kinases. ERKs phosphorylate at a specific site, Ser-736, integrity which is required for full induction transcriptional...

To prepare large amounts of the human estrogen receptor (ER) for biochemical and biophysical studies we have employed cloned ER sequences to construct Chinese hamster ovary (CHO) cell line derivatives that overexpress receptor. We an efficient expression vector (SV40 enhancer, metallothionein IIA promoter) a new system gene amplification based on stepwise selection in cadmium. Cells from initial transfected pools, before amplification, had as much or more than MCF7 cells responded subsequent...

Selective estrogen receptor modulators (SERMs) show differential effects upon ERα activation function 1 (AF-1). Tamoxifen allows strong AF-1 activity, whereas raloxifene less and ICI 182,780 (ICI) none. Here, we that blockade of corepressor histone de-acetylase (HDAC) activity reverses the inhibitory effect SERMs in MCF-7 cells. This suggests SERM repression involves HDAC-dependent corepressors. Consistent with this, are more potent than tamoxifen promoting ERα-dependent sequestration...

Unliganded thyroid hormone receptors (TRs) repress transcription through recruitment of corepressors, including nuclear receptor corepressor (N-CoR). We find that N-CoR contains three interaction domains (IDs) bind to TR, rather than the previously reported two. The hitherto unrecognized ID (ID3) serves as a fully functional TR binding site, both in vivo and vitro, may be most important for binding. Each motif conserved hydrophobic core (I/LXXII) resembles boxes (LXXLL), which mediates p160...

While steroid response is generally restricted by the availability of receptors, theoretical limits are not known. We have constructed a series cell lines that stably express estrogen receptor (ER) at levels up to 5,000,000 ERs per and employed these cells explore response. Several reporter genes with elements upstream herpes thymidine kinase promoter showed hyperbolic saturation kinetics increasing ER. Maximum was 10 times seen in titers comparable physiological levels. Half-maximal...