Transcriptional activation of the ovine follicle-stimulating hormone-beta gene by gonadotropin-releasing hormone involves multiple signal transduction pathways.
GnRH regulates gonadotrope cells through GnRH receptor activation of the PKC-, MAPK-, and calcium-activated signaling cascades. Due to the paucity of homologous model systems expressing FSHbeta, little is known about the specific mechanisms involved in transcriptional regulation of this gene by GnRH. Previous studies from our laboratory demonstrated that the gonadotrope-derived LbetaT2 cell line expresses FSHbeta mRNA. In the present study we characterized the mechanisms involved in GnRH regulation of the FSHbeta promoter using this cell model. Using transfection assays, we show that GnRH regulation of the ovine FSHbeta promoter involves at least two elements, present between -4152/-2878 and -2550/-1089 bp, in association with one or several elements within the proximal region of the promoter. Surprisingly, the two activating protein-1 sites previously shown to be involved in the FSHbeta response to GnRH in heterologous cells do not play a role in GnRH responsiveness in the gonadotrope cell model. Here we demonstrate that calcium influx itself is not sufficient to confer the response, but it is necessary for both 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and GnRH induction of the FSHbeta gene. Moreover, we show that GnRH regulation of FSHbeta gene expression is mediated by PKC and establish the presence of multiple PKC isozymes in LbetaT2 cells. Interestingly, GnRH and TPA induce activity of the FSHbeta promoter through different, although possibly overlapping, pools of PKC isoforms. This is further supported by the use of a MAPK inhibitor, which abolishes the induction of FSHbeta by GnRH, but not by TPA. In conclusion, we have demonstrated that calcium, PKC, and MAPK signaling systems are all involved in the induction of FSHbeta gene expression by GnRH in the LbetaT2 mouse gonadotrope cell model.
Vasilyev
Vyacheslav V
VV
Department of Reproductive Medicine, University of California-San Diego, 9500 Gilman Drive, La Jolla, California 92093-0674, USA.
Pernasetti
Flavia
F
Rosenberg
Suzanne B
SB
Barsoum
Mark J
MJ
Austin
Darrell A
DA
Webster
Nicholas J G
NJ
Mellon
Pamela L
PL
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005640:Follicle Stimulating Hormone / Q000235:genetics*; D037201:Follicle Stimulating Hormone, beta Subunit; D005786:Gene Expression Regulation / Q000187:drug effects; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology*; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism / Q000502:physiology; D010957:Plasmids / Q000235:genetics; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology; D020218:Response Elements / Q000235:genetics; D012756:Sheep; D015398:Signal Transduction / Q000187:drug effects* / Q000235:genetics*; D013268:Stimulation, Chemical; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000235:genetics; D015533:Transcriptional Activation / Q000187:drug effects*; D014162:Transfection
D004791:Enzyme Inhibitors; D037201:Follicle Stimulating Hormone, beta Subunit; D007527:Isoenzymes; D018808:Transcription Factor AP-1; D007987:Gonadotropin-Releasing Hormone; D005640:Follicle Stimulating Hormone; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium