Transcriptional Activation of the Ovine Follicle-Stimulating Hormone β-Subunit Gene by Gonadotropin-Releasing Hormone: Involvement of Two Activating Protein-1-Binding Sites and Protein Kinase C**This work was supported by the North Carolina State University Agricultural Research Service, NICHD Grant 34863, and the Mellon Foundation.

• DOI: 10.1210/endo.139.11.6281
• Publication Date: 2014-01-08T16:08:52Z
• Authors: Brian D. Strahl; Huey-Jing Huang; Joseph Sebastian; Basavdutta R. Ghosh; William L. Miller
• Keywords: GNRHR

Abstract FSH is an α/β heterodimeric glycoprotein, the formation of which regulated primarily by expression its β-subunit. Recent studies on transcriptional regulation ovine β-subunit gene (oFSHβ) have defined two functional activating protein-1 (AP-1) enhancers in proximal promoter (located at −120 and −83 bp) that are probably physiologically important for FSHβ expression. As GnRH a major regulator also known to stimulate synthesis Jun Fos family members (AP-1), we investigated possibility oFSHβ transcription may be through AP-1. Here report use vitro cell system involving transient transfection receptors (GnRHR) into HeLa cells define regulatory elements involved GnRH-mediated induction oFSHβ. This was used show luciferase constructs containing either −4741/+759 region (−4741oFSHβ-Luc) or −846/+44 human α (α-Luc; positive control) stimulated 3.1 ± 0.3- 7.7 1.9-fold, respectively, 100 nm GnRH. Another plasmid Rous sarcoma virus (a negative showed no response Similar results with these were obtained COS-7 cells. Studies progressive 5′-deletion site-specific mutations demonstrated this stimulation dependent each AP-1 site Gel shift assays ability GnRHR increase binding activity. Responses (ED50 = 0.5 nm) GnRHR, identified photoaffinity labeling. In addition, GnRHR-expressing exhibited normal GnRH-dependent mobilization intracellular calcium. Finally, as protein kinase C (PKC) target action gonadotropes, role PKC α-subunit genes investigated. Although 12-O-tetradecanoyl 13-acetate α-Luc −215oFSHβ-Luc could completely blocked dose-dependent manner specific inhibitor bisindolylmaleimide I, only 57–65% promoters blocked, demonstrating involvement well other signaling systems induction. These data molecular involves enhancer activation. Furthermore, establish usefulness investigate aspects gonadotropin subunit expression, similar pathways factors activated gonadotropes (such PKC, mitogen-activated kinase, Ca2+, AP-1) exist