Androgen Suppression of GnRH-Stimulated Rat LHβ Gene Transcription Occurs Through Sp1 Sites in the Distal GnRH-Responsive Promoter Region

Self Cite

157

A yeast two-hybrid assay was employed to identify androgen receptor (AR) protein partners in gonadotropin-releasing hormone neuronal cells. By using an AR deletion construct (AR-(⌬371-485)) as a bait, ␤-catenin was identified as an AR-interacting protein from a gonadotropin-releasing hormone neuronal cell library. Immunolocalization of co-transfected AR and FLAG-␤-catenin demonstrated that FLAG-␤-catenin was predominantly cytoplasmic in the absence of androgen. In the presence of 5␣-dihydrotestosterone, FLAG-␤-catenin completely co-localized to the nucleus with AR. This effect was specific to AR because liganded progesterone, glucocorticoid, or estrogen ␣ receptors did not translocate FLAG-␤-catenin to the nucleus. Agonist-bound AR was required because the AR antagonists casodex and hydroxyflutamide failed to translocate ␤-catenin. Time course experiments demonstrated that co-translocation occurred with similar kinetics. Nuclear co-localization was independent of the glycogen synthase kinase-3␤, p42/44 ERK mitogen-activated protein kinase, and phosphatidylinositol 3-kinase pathways because inhibitors of these pathways had no effect. Transcription assays demonstrated that liganded AR repressed ␤-catenin/T cell factor-responsive reporter gene activity. Conversely, co-expression of ␤-catenin/T cell factor repressed AR stimulation of AR-responsive reporter gene activity. Our data suggest that liganded AR shuttles ␤-catenin to the nucleus and that nuclear interaction of AR with ␤-catenin may modulate transcriptional activity in androgen target tissues.

Section: Androgen Receptor (Ar)mentioning

confidence: 89%

Liganded Androgen Receptor Interaction with β-Catenin

Pawlowski¹,

Ertel²,

Allen³

et al. 2002

Self Cite

157

“…Crosstalk between transcription factors and nuclear hormone receptors have previously been found to produce either positive (39,40,44) or negative (41,45) regulation.…”

Section: Discussionmentioning

confidence: 99%

Glucocorticoid and Androgen Activation of Monoamine Oxidase A Is Regulated Differently by R1 and Sp1

Chen

Shih

2006

145

123

Monoamine oxidase (MAO)A Monoamine oxidase (MAO)2 is located on outer membranes of mitochondria in neuronal, glial, and other cells. It catalyzes the oxidative deamination of monoamine neurotransmitters such as serotonin, norepinephrine, dopamine, and phenylethylamine (1). MAO exists in two isoforms, MAO A and MAO B. MAO A preferentially oxidizes serotonin (5-hydroxytryptamine), norepinephrine, and dopamine and is irreversibly inhibited by low concentration of clorgyline (2). MAO B preferentially oxidizes phenylethylamine and benzylamine and is irreversibly inactivated by low concentrations of pargyline and deprenyl (3). MAO A and B are coded by different genes (4) closely linked on the X chromosome, Xp11.2-11.4 (5). Both genes consist of 15 exons with identical exonintron organization (6) and both are regulated by Sp1 factors; however, there are some differences in their cis-elements (7-9). The core promoter of MAO A consists of Sp1-binding sites that are activated by Sp1 (7) and repressed by a novel repressor R1 (RAM2/CDCA7L) (8). This core promoter has bi-directional activity (7). In addition, a functional polymorphism of a 30-bp repeat sequence has been identified and located ϳ1.2 kb upstream of the MAO A translation start site (10). The human MAO B promoter contains two clusters of overlapping Sp1 sites separated by a CACCC element (11). An Sp1-like transcription factor, transforming growth factor-␤-inducible early gene, also activates the MAO B promoter via overlapping Sp1 sites (12). The expression of MAO B is regulated by protein kinase C and the MAPK signaling pathway (13).A mutation in the human MAO A gene, which results in no enzymatic activity, is associated with aggressive behavior in males in a Dutch family (14). MAO A knock-out (KO) mice showed aggressive behavior in males (15), but MAO B KO mice did not show aggressive behavior (16). Interestingly, a spontaneous mutation of the MAO A gene in MAO B KO mice resulted in MAO A/B double KO mice, which showed hyperreactivity and anxiety-like behavior (12).Steroid hormones are involved in the regulation of many functions in which MAO also plays an important roles such as response to stress, behavioral adaptation and mood (17). Significant hypersecretion of glucocorticoids has been shown to be associated with depression (18). The synthetic glucocorticoid, dexamethasone, increases MAO A activity in human fibroblasts (19) and rat brain frontal cortex (20). Both anti-glucocorticoid agents (21) and MAO A inhibitors (22, 23) have been used in the treatment of depression. Androgens have also been shown to increase MAO activity in the rat brain (24).The biological action of glucocorticoids and androgens are mediated through their respective receptors, glucocorticoid receptor (GR) and androgen receptor (AR). GR and AR bind to derivatives of a common response element, i.e. the consensus glucocorticoid response element (GRE), 5Ј-GGTACAnnn-

“…For example, Sp1 and NFY cooperatively interact to regulate multiple genes through NFY-GC-rich motifs, and both proteins also physically interact (6 -10). Sp1 also binds estrogen receptor ␣ (ER␣) 1 and other members of the nuclear receptor family of transcription factors (11)(12)(13)(14)(15)(16)(17)(18)(19), and research in our laboratory has focused on the molecular mechanisms of the ligand-dependent activation of ER␣/Sp1 in breast and endometrial cancer cell lines (20 -31). Promoter analysis studies in breast cancer cells have identified GC-rich sites required for hormone activation of several genes including E2F1, DNA polymerase ␣, cyclin D1, insulin-like growth factor-binding protein 4, retinoic acid receptor ␣1, cathepsin D, vascular endothelial growth factor, c-fos, heat shock protein 27, bcl-2, thymidylate synthase, and adenosine deaminase (20 -27, 29 -31).…”

mentioning

confidence: 99%

Small Inhibitory RNA Duplexes for Sp1 mRNA Block Basal and Estrogen-induced Gene Expression and Cell Cycle Progression in MCF-7 Breast Cancer Cells

Abdelrahim

Samudio

Smith

et al. 2002

111

115

Sp1 is a member of the Sp and Krü ppel-like family of transcription factors that bind GC and CACCC boxes to regulate gene expression (1-3). Sp1 is widely expressed in multiple tissues (4), and targeted disruption of Sp1 in mice results in retarded development and embryo-lethality (5). Sp1 interacts with GC-rich Sp1 binding sites in multiple promoters to regulate gene expression, and there are an increasing number of studies showing that Sp1 interacts with other nuclear proteins, including promoter-bound transcription factors, to attenuate tissue-specific expression of selected genes (1-3). For example, Sp1 and NFY cooperatively interact to regulate multiple genes through NFY-GC-rich motifs, and both proteins also physically interact (6 -10). Sp1 also binds estrogen receptor ␣ (ER␣) 1 and other members of the nuclear receptor family of transcription factors (11)(12)(13)(14)(15)(16)(17)(18)(19), and research in our laboratory has focused on the molecular mechanisms of the ligand-dependent activation of ER␣/Sp1 in breast and endometrial cancer cell lines (20 -31). Promoter analysis studies in breast cancer cells have identified GC-rich sites required for hormone activation of several genes including E2F1, DNA polymerase ␣, cyclin D1, insulin-like growth factor-binding protein 4, retinoic acid receptor ␣1, cathepsin D, vascular endothelial growth factor, c-fos, heat shock protein 27, bcl-2, thymidylate synthase, and adenosine deaminase (20 -27, 29 -31). Studies in other cell lines have also demonstrated a role for ER␣/Sp1 activation of the progesterone receptor, epidermal growth factor receptor, telomerase, and receptor for advanced glycation end products (32-35). Activation of ER␣/Sp1 does not require the DNA binding domain of ER␣ (promoter DNA-independent) and is primarily dependent on the activation function-1 (AF1) of ER␣ (30), whereas DNAdependent activation through ER binding to estrogen response elements (EREs) is primarily dependent on AF2 of ER␣. Recent studies have demonstrated that RNA interference through small inhibitory RNAs (iRNAs) targeted to endogenous or heterologous genes can be used to suppress intracellular expression of these genes in mammalian cells, and this technique is well suited for mechanistic studies on gene/protein function (36 -42). This study investigates the role of Sp1 protein in mediating hormone-responsiveness in MCF-7 cells using sequence-specific duplexes of 21 nucleotides targeted to Sp1 mRNA as well as Lamin B1 and the heterologous firefly luciferase gene (GL2) mRNAs. Transfection of iRNA for Sp1 (iSp1) decreases (40 -60%) the expression of nuclear Sp1 protein in ER-positive MCF-7 and ZR-75 human breast cancer cell extracts. In transfected cells, Sp1 protein is barely detectable by immunofluorescence, and both basal and estrogen-inducible transactivation is decreased in cells transfected with iSp1 and a GC-rich construct. These data, combined with results showing that iSp1 inhibits hormone-induced MCF-7 cell cycle progression from G 0 /G 1 to S phase, demonstrate that ER␣/Sp1...