Estrogen-occupied Estrogen Receptor Represses Cyclin G2 Gene Expression and Recruits a Repressor Complex at the Cyclin G2 Promoter

Abstract

Estrogens, acting through their nuclear receptors have a broad impact on target cells, eliciting a transcriptional response program that involves gene repression as well as gene stimulation. While much is known about the mechanisms by which the estrogen-occupied estrogen receptor (ER) stimulates gene expression, the molecular events that lead to gene repression by the hormone-ER complex are largely unknown. Because estradiol represses expression of the cyclin G2 gene, which encodes a negative regulator of the cell cycle, our aim was to understand the mechanism by which cyclin G2 is repressed by estrogen. We show that cyclin G2 is a primary ER target gene in MCF-7 breast cancer cells that is rapidly and robustly down-regulated by estrogen. Promoter analysis reveals a responsive region containing a halfestrogen response element and GC-rich region that interact with ER and Sp1 proteins. Mutation of the half-ERE abrogates hormone-mediated repression. Mutational mapping of receptor reveals a requirement for its N-terminal region and DNA binding domain to support cyclin G2 repression. Following estradiol treatment of cells, chromatin immunoprecipitation analyses reveal recruitment of ER to the cyclin G2 regulatory region, dismissal of RNA polymerase II, and recruitment of a complex containing N-CoR and histone deacetylases, leading to a hypoacetylated chromatin state. Our study provides evidence for a mechanism by which the estrogen-occupied ER is able to actively repress gene expression in vivo and indicates a role for nuclear receptor corepressors and associated histone deacetylase activity in mediating negative gene regulation by this hormone-occupied nuclear receptor.In target cells, estrogen hormones act via their nuclear receptors to elicit a diverse transcriptional response program that involves gene repression as well as gene stimulation (1-3). Estrogen receptors (ERs) 3 share their modular domain structure with the other members of the steroid and nuclear receptor superfamily and are comprised of an N-terminal domain characterized by a ligand-independent activation function (AF-1), a central DNA binding domain (DBD) followed by a hinge region, and a C-terminal ligand binding domain (LBD), which contains a ligand-dependent activation function (AF-2) important for coregulator recruitment (4, 5).In the mammary gland, the actions of estrogens are essential for normal growth and development. In breast cancer, the presence of ERs is associated with likely response to endocrine therapy, most usually treatment with antiestrogens such as tamoxifen, or estrogen depletion by the use of aromatase inhibitors (6). Estrogens enhance the proliferation of ER-positive breast cancer cells, and recent studies have shown that this enhancement of proliferation involves both stimulation of the expression of many genes associated with cell cycle progression, as well as the suppression of genes that block the cell cycle (7). Most previous studies, focusing on the mechanisms that lead to stimulation of gene expression, have demonstrat...

Nodal enhances the activity of FoxO3a and its synergistic interaction with Smads to regulate cyclin G2 transcription in ovarian cancer cells

“…Several studies have shown that CCNG2 mRNA levels and/or protein levels are upregulated by growth inhibitory signals, including TGF-b (Horne et al, 1997) and downregulated by oncogenic pathways, such as PI3K-AKT in murine NIH 3T3 cells (Martinez-Gac et al, 2004), HER2, c-jun NH2-terminal kinase and the mammalian target of rapamycin/p70S6K (Le et al, 2007), and estrogen signaling (Stossi et al, 2006) in breast cancer cells. In immortalized ovarian surface epithelial and ovarian cancer cells, Nodal increased CCNG2 mRNA and protein levels, as well as CCNG2 protein stability (Xu et al, 2008).…”

Membrane-initiated estrogen signaling in hypothalamic neurons

“…However, recently the FASEB steroid signaling work group suggested that "membrane-initiated steroid signaling" and "nuclear-initiated steroid signaling" are more appropriate terminologies (Hammes and Levin, 2007). The nuclear-effects on a variety of tissues that involves gene stimulation as well as gene repression (Herbison, 1998, Couse and Korach, 1999, Nilsson et al, 2001, Stossi et al, 2006, Kininis et al, 2007. In general, this "classical" signaling pathway of estrogen involves steroid-dependent formation of nuclear estrogen receptor homo-or heterodimers and the subsequent binding of this complex with a unique DNA sequence known as an estrogen response element (ERE), in E2-responsive gene promoters (O'Malley and Tsai, 1992, Muramatsu and Inoue, 2000, Gruber et al, 2004.…”

FOXA1 Is Essential for Aryl Hydrocarbon Receptor–Dependent Regulation of Cyclin G2

“…Because FKH sites were found to be important in the TCDD-mediated regulation of the CCNG2 luciferase reporter plasmid, we hypothesized that FOXA1 may have a similar role with AHR-chromatin interactions at CCNG2. To test this hypothesis, we used RNAi-mediated knockdown of FOXA1 and measured mRNA expression, as well as the recruitment patterns of AHR, FOXA1, and ERa; ERa is known to negatively regulate CCNG2 (21). Following transient transfection of 2 distinct siRNA oligos into T-47D cells, we determined that 48 hours posttransfection, FOXA1 protein levels were greatly reduced (Fig.…”

“…14,18). CCNG2 has been shown to be a critical gene in human epidermal growth factor receptor 2 (HER2)-and hormone-dependent breast cancers (18)(19)(20)(21). Moreover, CCNG2 was among the few genes upregulated following anti-HER2 antibody trastuzumab treatment, indicating that CCNG2 might play a role in inhibiting HER2-dependent proliferation (22).…”

“…ERa represses CCNG2 expression in response to estrogen by recruiting a complex containing nuclear corepressor (NCoR) and histone deacetylases to the CCNG2 promoter region resulting in the displacement of RNA polymerase II (21). Our previous ChIP-chip studies identified an AHR-bound AHRE-containing sequence in the upstream regulatory region of CCNG2.…”