Abstract
The purpose of this study was to analyze the mechanism of transcriptional activation of human chorionic gonadotropin-alpha (hCGalpha) gene by epidermal growth factor (EGF) in trophoblast cells. We stably transfected hCGalpha promoter-chloramphenicol acetyltransferase constructs into Rcho-1 trophoblast cells and monitored the promoter activities. -290-base pair hCGalpha promoter containing a tandem repeat of cAMP response element (CRE) was activated by EGF in a dose- and time-dependent manner. Deletion analysis of hCGalpha promoter suggested an involvement of CRE in EGF-induced hCGalpha transcriptional activation. Moreover, the hCGalpha promoter, of which both CREs were mutated, did not respond to EGF. These results indicate that EGF activates the hCGalpha gene transcription through CRE. Although EGF did not alter the amount of CRE-binding protein (CREB), EGF induced CREB phosphorylation. We next examined the mechanism of CREB phosphorylation by EGF. Protein kinase C inhibitors (H7, staurosporin, and chelerythrine) inhibited EGF-induced CREB phosphorylation, whereas either mitogen-activated protein kinase kinase-1 inhibitor (PD98059) or protein kinase A inhibitor (H8) showed no effect. Furthermore, H7 and staurosporin but not H8 inhibited hCGalpha promoter activation by EGF. In conclusion, EGF promotes hCGalpha gene transcription via the CRE region probably by phosphorylating CREB mainly through the protein kinase C pathway in trophoblast cells.
Highlights
Epidermal growth factor (EGF),1 which consists of 53 amino acids, stimulates proliferation and differentiation in many kinds of cells and tissues [1]
It has been known that EGF stimulates human chorionic gonadotropin (hCG) secretion by trophoblast cells [33]
Cao et al showed that EGF increased hCG␣ and hCG mRNA level by stabilizing them in JEG-3 cells [16]; it has not been known whether or not EGF promotes hCG gene transcription
Summary
Epidermal growth factor (EGF), which consists of 53 amino acids, stimulates proliferation and differentiation in many kinds of cells and tissues [1]. EGF has been shown to promote functional differentiation of human trophoblast cells [10]; EGF treatment of choriocarci-. EGF has been shown to increase hCG␣ and hCG mRNA level and their stability [16]; it remains unclear whether or not EGF increases the transcriptional activity of hCG genes. In this study we analyzed the effect of EGF on the hCG␣ gene. We used Rcho-1 cells to examine the molecular mechanism of EGF effect on the hCG␣ gene transcription. Several genes have been shown to be transcriptionally activated during Rcho-1 cell differentiation [22, 23]. Using Rcho-1 cells stably transfected with hCG␣ promoter-CAT construct, we studied the EGF effect on the transcriptional activity of hCG␣ gene and analyzed the mechanism of EGF action
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