Identification and Analysis of Two Novel Sites of Rat GnRH Receptor Gene Promoter Activity: The Pineal Gland and Retina

2012 ◽  
Vol 97 (2) ◽  
pp. 115-131 ◽  
Author(s):  
Anne-Laure Schang ◽  
Christian Bleux ◽  
Marie-Claude Chenut ◽  
Valérie Ngô-Muller ◽  
Bruno Quérat ◽  
...  
Keyword(s):  
Pineal Gland ◽  
Promoter Activity ◽  
Receptor Gene ◽  
Gene Promoter ◽  
Gnrh Receptor

scholarly journals GATA2-Induced Silencing and LIM-Homeodomain Protein-Induced Activation Are Mediated by a Bi-Functional Response Element in the Rat GnRH Receptor Gene

2013 ◽  
Vol 27 (1) ◽  
pp. 74-91 ◽  
Author(s):  
Anne-Laure Schang ◽  
Anne Granger ◽  
Bruno Quérat ◽  
Christian Bleux ◽  
Joëlle Cohen-Tannoudji ◽  
...  
Keyword(s):  
Cell Fate ◽  
Promoter Activity ◽  
Receptor Gene ◽  
Gnrh Receptor ◽  
Expression Vectors ◽  
Homeodomain Protein ◽  
Homeodomain Proteins ◽  
Enhancer Activity ◽  
Response Element ◽  
Lim Homeodomain

GATA2 transcription factor and LIM homeodomain proteins Islet1 (ISL1) and LIM homeobox 3 (LHX3) are suspected to be involved in gonadotrope cell fate and maintenance. The GnRH receptor gene (Gnrhr), crucial for gonadotrope function, is expressed in the pituitary gland from embryonic day 13.5 onward, well before LH and FSH β-subunits. This expression pattern together with the presence of WGATAR and TAAT motifs in Gnrhr promoter sequences suggests the involvement of early transcription factors in promoter activation. In this study, using a well-characterized transgenic mouse model, GATA2 was found colocalized with Gnrhr promoter activity in the pituitary. Transient transfection of Gnrhr promoter luciferase fusion constructs together with either GATA2 expression vectors or small interfering RNA in gonadotrope cell lines indicated that GATA2, which typically acts as a trans-activator, unexpectedly repressed Gnrhr promoter activity. Using DNA chromatography affinity and EMSA, we demonstrated that GATA2 operates via a response element containing a peculiar palindromic GATA motif that overlaps a critical TAAT motif involved in LHX3/ISL1 trans-activation. Indeed, despite the inhibitory action of GATA2, this element displayed a clear-cut enhancer activity in gonadotrope cells. Chromatin immunoprecipitation assays indicated that GATA2, LHX3, and ISL1 interact with a Gnrhr promoter fragment encompassing this element. The trans-repressive action of GATA2 on Gnrhr promoter activity is likely balanced or even hindered by trans-activating effects of LIM homeodomain proteins via this novel bifunctional LIM/GATA response element. Such a hierarchical interplay may contribute to finely adjust Gnrhr gene expression in gonadotrope cell lineage during pituitary development as well as in the adult animal.

FUNCTIONAL ACTIVITY OF THE PORCINE TYPE II GNRH RECEPTOR GENE PROMOTER

2007 ◽  
Vol 77 (Suppl_1) ◽  
pp. 227-227
Author(s):  
Jocelyn Wiarda ◽  
Rebecca Cederberg ◽  
Brett White
Keyword(s):  
Functional Activity ◽  
Receptor Gene ◽  
Gene Promoter ◽  
Gnrh Receptor ◽  
Type Ii

scholarly journals The Tripartite Basal Enhancer of the Gonadotropin-Releasing Hormone (GnRH) Receptor Gene Promoter Regulates Cell-Specific Expression Through a Novel GnRH Receptor Activating Sequence

1997 ◽  
Vol 11 (12) ◽  
pp. 1814-1821 ◽  
Author(s):  
Dawn L. Duval ◽  
Scott E. Nelson ◽  
Colin M. Clay
Keyword(s):  
Transcriptional Activation ◽  
Promoter Activity ◽  
Molecular Mechanisms ◽  
Mutational Analysis ◽  
Receptor Gene ◽  
Gnrh Receptor ◽  
Luciferase Expression ◽  
Functional Requirements ◽  
Specific Expression ◽  
Core Motif

Abstract The molecular mechanisms regulating restricted expression of GnRH receptor and gonadotropin subunit genes to gonadotrope cells have been the focus of intense interest. Using deletion and mutational analysis we have identified a tripartite enhancer that regulates cell-specific expression of the GnRH receptor gene in the gonadotrope-derived αT3–1 cell line. Individual elements of this enhancer include binding sites for steroidogenic factor-1; activator protein 1 (AP-1); and a novel element referred to as the GnRH receptor activating sequence (GRAS). Mutation of each element alone results in loss of approximately 60% of promoter activity. Combinatorial mutations of any two elements decreases promoter activity by approximately 80%. Finally, mutation of all three elements reduces promoter activity to a level not different from promoterless vector. Using 2-bp mutations, we have defined the functional requirements for transcriptional activation by GRAS. The core motif of GRAS is at −391 to −380 bp relative to the start site of translation and has the sequence 5′-CTAGTCACAACA-3′. Three copies of GRAS or GRAS with a 2-bp mutation (μGRAS) were cloned into a luciferase expression vector immediately upstream of the thymidine kinase minimal promoter (TK) and tested for expression in αT3–1 cells. When compared with TK promoter alone, activity of 3xGRAS-TKLUC was increased by more than 5-fold while activity of 3xμGRAS-TKLUC was unchanged. When 3xGRAS-TKLUC was transfected into a variety of nongo-nadotrope cell lines, it did not increase activity of the TK promoter. We propose that basal activity of the GnRH receptor gene is regulated by a tripartite enhancer, and the key component of this enhancer is an element, GRAS, that activates transcription in a cell-specific fashion.

scholarly journals The LIM-Homeodomain Proteins Isl-1 and Lhx3 Act with Steroidogenic Factor 1 to Enhance Gonadotrope-Specific Activity of the Gonadotropin-Releasing Hormone Receptor Gene Promoter

2006 ◽  
Vol 20 (9) ◽  
pp. 2093-2108 ◽  
Author(s):  
Anne Granger ◽  
Christian Bleux ◽  
Marie-Laure Kottler ◽  
Simon J. Rhodes ◽  
Raymond Counis ◽  
...  
Keyword(s):  
Promoter Activity ◽  
Specific Activity ◽  
Receptor Gene ◽  
Marker Gene ◽  
Gene Promoter ◽  
Dominant Negative ◽  
R Gene ◽  
Specific Expression ◽  
Steroidogenic Factor 1 ◽  
Lim Homeodomain

Abstract The GnRH receptor (GnRH-R) plays a central role in mammalian reproductive function throughout adulthood. It also appears as an early marker gene of the presumptive gonadotrope lineage in developing pituitary. Here, using transient transfections combined with DNA/protein interaction assays, we have delineated cis-acting elements within the rat GnRH-R gene promoter that represent targets for the LIM-homeodomain (LIM-HD) proteins, Isl-1 and Lhx3. These factors, critical in early pituitary development, are thus also crucial for gonadotrope-specific expression of the GnRH-R gene. In heterologous cells, the expression of Isl-1 and Lhx3, together with steroidogenic factor 1 (SF-1), culminates in the activation of both the rat as well as human GnRH-R promoter, suggesting that this combination is evolutionarily conserved among mammals. The specificity of these LIM-HD factors is attested by the inefficiency of related proteins, including Lhx5 and Lhx9, to activate the GnRH-R gene promoter, as well as by the repressive capacity of a dominant-negative derivative of Lhx3. Accordingly, targeted deletion of the LIM response element decreases promoter activity. In addition, experiments with Gal4-SF-1 fusion proteins suggest that LIM-HD protein activity in gonadotrope cells is dependent upon SF-1 binding. Finally, using a transgenic model that allows monitoring of in vivo promoter activity, we show that the overlapping expression of Isl-1 and Lhx3 in the developing pituitary correlates with promoter activity. Collectively, these data suggest the occurrence of a specific LIM-HD pituitary code and designate the GnRH-R gene as the first identified transcriptional target of Isl-1 in the anterior pituitary.

scholarly journals Fatty Acid Synthase Confers Tamoxifen Resistance to ER+/HER2+ Breast Cancer

Cancers ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 1132
Author(s):  
Javier A. Menendez ◽  
Adriana Papadimitropoulou ◽  
Travis Vander Steen ◽  
Elisabet Cuyàs ◽  
Bharvi P. Oza-Gajera ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Fatty Acid ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Promoter Activity ◽  
Fatty Acid Synthase ◽  
Endocrine Resistance ◽  
Gene Promoter ◽  
Her2 Positive ◽  
Her2 Breast Cancer

The identification of clinically important molecular mechanisms driving endocrine resistance is a priority in estrogen receptor-positive (ER+) breast cancer. Although both genomic and non-genomic cross-talk between the ER and growth factor receptors such as human epidermal growth factor receptor 2 (HER2) has frequently been associated with both experimental and clinical endocrine therapy resistance, combined targeting of ER and HER2 has failed to improve overall survival in endocrine non-responsive disease. Herein, we questioned the role of fatty acid synthase (FASN), a lipogenic enzyme linked to HER2-driven breast cancer aggressiveness, in the development and maintenance of hormone-independent growth and resistance to anti-estrogens in ER/HER2-positive (ER+/HER2+) breast cancer. The stimulatory effects of estradiol on FASN gene promoter activity and protein expression were blunted by anti-estrogens in endocrine-responsive breast cancer cells. Conversely, an AKT/MAPK-related constitutive hyperactivation of FASN gene promoter activity was unaltered in response to estradiol in non-endocrine responsive ER+/HER2+ breast cancer cells, and could be further enhanced by tamoxifen. Pharmacological blockade with structurally and mechanistically unrelated FASN inhibitors fully impeded the strong stimulatory activity of tamoxifen on the soft-agar colony forming capacity—an in vitro metric of tumorigenicity—of ER+/HER2+ breast cancer cells. In vivo treatment with a FASN inhibitor completely prevented the agonistic tumor-promoting activity of tamoxifen and fully restored its estrogen antagonist properties against ER/HER2-positive xenograft tumors in mice. Functional cancer proteomic data from The Cancer Proteome Atlas (TCPA) revealed that the ER+/HER2+ subtype was the highest FASN protein expressor compared to basal-like, HER2-enriched, and ER+/HER2-negative breast cancer groups. FASN is a biological determinant of HER2-driven endocrine resistance in ER+ breast cancer. Next-generation, clinical-grade FASN inhibitors may be therapeutically relevant to countering resistance to tamoxifen in FASN-overexpressing ER+/HER2+ breast carcinomas.

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