Role of the Cyclic AMP Response Element Binding Complex and Activation of Mitogen-Activated Protein Kinases in Synergistic Activation of the Glycoprotein Hormone α Subunit Gene by Epidermal Growth Factor and Forskolin

Abstract

The aim of these studies was to elucidate a role for epidermal growth factor (EGF) signaling in the transcriptional regulation of the glycoprotein hormone ␣ subunit gene, a subunit of chorionic gonadotropin. Studies examined the effects of EGF and the adenylate cyclase activator forskolin on the expression of a transfected ␣ subunit reporter gene in a human choriocarcinoma cell line (JEG3). At maximal doses, administration of EGF resulted in a 50% increase in a subunit reporter activity; forskolin administration induced a fivefold activation; the combined actions of EGF and forskolin resulted in synergistic activation (greater than eightfold) of the ␣ subunit reporter. Mutagenesis studies revealed that the cyclic AMP response elements (CRE) were required and sufficient to mediate EGF-forskolin-induced synergistic activation. Chorionic gonadotropin (CG) is a heterodimeric glycoprotein hormone consisting of an ␣ subunit common to other glycoprotein hormone family members noncovalently linked to a CG-specific ␤ subunit (58). CG is synthesized and secreted by placental syncytiotrophoblast cells during the first trimester of pregnancy in women and nonhuman primates. CG is a luteotropin required for maintenance of progesterone production by the ovarian corpus luteum in early gestation. An important factor in the establishment of early pregnancy appears to be the timing and rate of increase in the secretion of CG that is highly correlated with a rise in progesterone levels in peripheral circulation (42). Insufficient progesterone production during early pregnancy is correlated with the potential for early or recurrent pregnancy loss in women (6, 7, 27, 47). Thus, endocrine mechanisms that potentiate the synthesis of CG subunits and CG secretion are essential for the establishment of pregnancy. Despite the clear importance of CG to early pregnancy, the specific ligands and signaling mechanisms that regulate the expression of CG subunit genes in placental cells have not been fully elucidated.The ␣ subunit of the glycoprotein hormones is a unique and useful transcriptional model for the study of tissue-specific gene expression because the ␣ subunit gene is expressed in placental and pituitary cells, albeit by various mechanisms. Analysis of the architecture of the ␣ subunit promoter revealed the presence of multiple promoter elements that are required for transcriptional regulation. These include the pituitary glycoprotein hormone basal element, which binds members of the LIM class of homeobox-containing proteins (67,71,72); the ␣ basal element (32); the gonadotrope-specific element, which binds steroidogenic factor 1 (9, 36); the upstream regulatory element (URE) (12,26,38,59); the GATA element, which binds several GATA factors (75); the dual tandem cyclic AMP (cAMP) response elements (CREs), which bind CRE binding protein (CREB) (5,10,12,13,22,25,32,35,52,59,74); the junctional regulatory element (JRE) (4, 12); and a unique CAAT box (12,40). Extensive mutagenesis studies have begun to unravel the specific requirements f...

Stimulation of extracellular signal-regulated kinase by pituitary adenylate cyclase-activating polypeptide in alpha T3-1 gonadotrophs

“…The concentrations chosen were those shown previously to produce the maximal response for each agonist (Burrin et al 1998, Roberson et al 2000. ERK activity was measured using a commercially available kit (New England Biolabs, Hitchin, UK).…”

Transcription factor activation and mitogenic synergism in airway smooth muscle cells

“…Since c-fos is a component of AP-1 and the c-fos gene contains well-characterised SRE and CRE sequences [27], transcriptional increases in c-fos expression by activation of these factors is a potential explanation for the ability of LPA to activate AP-1, although this remains to be established. EGF has been demonstrated to activate the c-fos SRE [26] and CREB [28] in other cell types and this may also be the mechanism for EGF activation of AP-1. Although EGF is not a classical activator of NFAT, a recent report suggests that EGF may activate NFAT through activation of Vav1 in T-cells [29].…”

Activation of Thyroid Hormone Is Transcriptionally Regulated by Epidermal Growth Factor in Human Placenta-Derived JEG3 Cells

“…As previously reported, EGF stimulates the CREB-target gene ␣-chorionic gonadotropin (␣-CG) in placenta, by increasing c-Jun and c-Fos levels and activity (25). To determine whether a similar mechanism could bring about the mitogen-mediated Dio2 transcription, we first performed Western blot experiments on EGF-treated JEG3 cells.…”

“…Upon stimulation of JEG3 cells with EGF, three MAPKs are mainly activated: ERK, p38, and JNK (25). To study the signal transduction pathway involved in this effect, we started using the inhibitors PD98059 and SB203580 to suppress ERK and p38 kinase activity, respectively.…”