scholarly journals Evaluation of in vitro screening system for estrogenicity: comparison of stably transfected human estrogen receptor-α transcriptional activation (OECD TG455) assay and estrogen receptor (ER) binding assay

2012 ◽  
Vol 37 (2) ◽  
pp. 431-437 ◽  
Author(s):  
Hae Kyung Lee ◽  
Tae Sung Kim ◽  
Chang Yeong Kim ◽  
Il Hyun Kang ◽  
Mi Gyeong Kim ◽  
...  
Keyword(s):  
Estrogen Receptor ◽  
Transcriptional Activation ◽  
Binding Assay ◽  
Estrogen Receptor Α ◽  
In Vitro Screening ◽  
Screening System ◽  
Human Estrogen Receptor

scholarly journals Mediator and p300/CBP-Steroid Receptor Coactivator Complexes Have Distinct Roles, but Function Synergistically, during Estrogen Receptor α-Dependent Transcription with Chromatin Templates

2003 ◽  
Vol 23 (1) ◽  
pp. 335-348 ◽  
Author(s):  
Mari Luz Acevedo ◽  
W. Lee Kraus
Keyword(s):  
Estrogen Receptor ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Steroid Receptor ◽  
Estrogen Receptor Α ◽  
Preinitiation Complex ◽  
Transcription System ◽  
Steroid Receptor Coactivator ◽  
Transcription Reinitiation

ABSTRACT Ligand-dependent transcriptional activation by nuclear receptors involves the recruitment of various coactivators to the promoters of hormone-regulated genes assembled into chromatin. Nuclear receptor coactivators include histone acetyltransferase complexes, such as p300/CBP-steroid receptor coactivator (SRC), as well as the multisubunit mediator complexes (“Mediator”), which may help recruit RNA polymerase II to the promoter. We have used a biochemical approach, including an in vitro chromatin assembly and transcription system, to examine the functional role for Mediator in the transcriptional activity of estrogen receptor α (ERα) with chromatin templates, as well as functional interplay between Mediator and p300/CBP during ERα-dependent transcription. Using three different approaches to functionally inactivate Mediator (immunoneutralization, immunodepletion, and inhibitory polypeptides), we find that Mediator is required for maximal transcriptional activation by ligand-activated ERα. In addition, we demonstrate synergism between Mediator and p300/CBP-SRC during ERα-dependent transcription with chromatin templates, but not with naked DNA. This synergism is important for promoting the formation of a stable transcription preinitiation complex leading to the initiation of transcription. Interestingly, we find that Mediator has an additional distinct role during ERα-dependent transcription not shared by p300/CBP-SRC: namely, to promote preinitiation complex formation for subsequent rounds of transcription reinitiation. These results suggest that one functional consequence of Mediator-ERα interactions is the stimulation of multiple cycles of transcription reinitiation. Collectively, our results indicate an important role for Mediator, as well as its functional interplay with p300/CBP-SRC, in the enhancement of ERα-dependent transcription with chromatin templates.

scholarly journals Epigenetics meets estrogen receptor: regulation of estrogen receptor by direct lysine methylation

2009 ◽  
Vol 16 (2) ◽  
pp. 319-323 ◽  
Author(s):  
Qun Zhou ◽  
Patrick G Shaw ◽  
Nancy E Davidson
Keyword(s):  
Estrogen Receptor ◽  
Transcriptional Activation ◽  
Target Genes ◽  
Estrogen Receptor Α ◽  
Nuclear Hormone Receptor ◽  
Cellular Growth ◽  
Breast Cancers ◽  
Lysine Methylation ◽  
Estrogen Response

The nuclear hormone receptor estrogen receptor α (ERα) promotes cellular growth through ligand-dependent activation of specific target genes, a process which is targeted in the treatment of ERα-expressing breast cancers. ERα activity is regulated at the protein level by post-translational modifications including phosphorylation and acetylation. A study now shows that ERα can also be directly methylated at lysine 302 (K302) by SET7, a histone methyltransferase that is known to monomethylate H3K4 and is associated with transcriptional activation. It was shown that K302 methylation stabilizes ERα protein and is suggested to increase sensitivity of ERα to estrogens, enhancing transcription of estrogen response elements. Furthermore, SET7 methylation of K302 is enhanced by a breast cancer-associated mutation at K303 (K303R) in vitro. These findings provide an additional mechanism of SET7 mediated transcriptional activation, as well as potential insight into the complex regulation of ERα stability and ligand sensitivity.

scholarly journals pp90rsk1 Regulates Estrogen Receptor-Mediated Transcription through Phosphorylation of Ser-167

1998 ◽  
Vol 18 (4) ◽  
pp. 1978-1984 ◽  
Author(s):  
Peteranne B. Joel ◽  
Jeffrey Smith ◽  
Thomas W. Sturgill ◽  
Tracey L. Fisher ◽  
John Blenis ◽  
...  
Keyword(s):  
Estrogen Receptor ◽  
Transcriptional Activation ◽  
Ectopic Expression ◽  
Activation Function ◽  
Estrogen Receptor Α ◽  
Kinase Domain ◽  
Stable Complex ◽  
Wild Type

ABSTRACT The estrogen receptor α (ER), a member of the steroid receptor superfamily, contains an N-terminal hormone-independent transcriptional activation function (AF-1) and a C-terminal hormone-dependent transcriptional activation function (AF-2). Here, we used in-gel kinase assays to determine that pp90rsk1 activated by either epidermal growth factor (EGF) or phorbol myristate acetate specifically phosphorylates Ser-167 within AF-1. In vitro kinase assays demonstrated that pp90rsk1 phosphorylates the N terminus of the wild-type ER but not of a mutant ER in which Ser-167 was replaced by Ala. In vivo, EGF stimulated phosphorylation of Ser-167 as well as Ser-118. Ectopic expression of active pp90rsk1increased the level of phosphorylation of Ser-167 compared to that of either a mutant pp90rsk1, which is catalytically inactive in the N-terminal kinase domain, or to that of vector control. The ER formed a stable complex with the mutant pp90rsk1in vivo. Transfection of the mutant pp90rsk1 depressed ER-dependent transcription of both a wild-type ER and a mutant ER that had a defective AF-2 domain (ER TAF-1). Furthermore, replacing either Ser-118 or Ser-167 with Ala in ER TAF-1 showed similar decreases in transcription levels. A double mutant in which both Ser-118 and Ser-167 were replaced with Ala demonstrated a further decrease in transcription compared to either of the single mutations. Taken together, our results strongly suggest that pp90rsk1 phosphorylates Ser-167 of the human ER in vivo and that Ser-167 aids in regulating the transcriptional activity of AF-1 in the ER.

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