Synergy between activin A and gonadotropin-releasing hormone in transcriptional activation of the rat follicle-stimulating hormone-beta gene.
Both activin and GnRH can independently stimulate expression of the FSHbeta subunit gene. In this study, we used the gonadotrope-derived LbetaT2 cell line to investigate the potential interaction between activin and GnRH in regulating the transcriptional activity of the rat FSHbeta gene promoter. Activin A and GnRH synergistically enhanced rat FSHbeta transcriptional activity. Overexpression of SMAD3 (mediator of decapentaplegic-related protein 3), but not of SMAD2, increased transcriptional activation of the rat (r) FSHbeta gene promoter, which was further enhanced by the combined overexpression of SMAD3 and 4 (3+4). The stimulatory effects of SMAD3 overexpression were localized to -472/-256 of the rFSHbeta gene promoter, and activin- and GnRH-responsive proteins were shown to bind to region -284/-252. Sequence analysis identified a consensus palindromic SMAD-binding site at -266/-259 of the rFSHbeta gene promoter. Mutation of two bases located in the center of this palindrome effectively abrogated SMAD4 binding, markedly reduced SMAD3 and 3+4 stimulation of the rFSHbeta gene promoter, and significantly decreased the synergistic enhancement of promoter activity by both activin A and GnRH, and SMAD3 and GnRH. Blockade of the MAPK-signaling pathway did not significantly affect the response to combined stimulation with activin and GnRH. In contrast, interference with SMAD3 signaling caused a significant reduction in activin and GnRH synergy. The results indicate that SMAD3 plays an important role in the synergistic effects of activin and GnRH and demonstrate that this synergy is mediated by a palindromic cis-element located at -266/-259 of the rFSHbeta gene promoter.
Gregory
Susan J
SJ
Brigham and Women's Hospital and Harvard Medical School, Division of Endocrinology, Diabetes and Hypertension, 221 Longwood Avenue, Boston, Massachusetts 02115, USA.
Lacza
Charlemagne T
CT
Detz
Alissa A
AA
Xu
Shuyun
S
Petrillo
Laura A
LA
Kaiser
Ursula B
UB
D028341:Activins / Q000494:pharmacology*; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D037201:Follicle Stimulating Hormone, beta Subunit / Q000235:genetics* / Q000494:pharmacology; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology*; D006801:Humans; D028322:Inhibin-beta Subunits / Q000494:pharmacology*; D020935:MAP Kinase Signaling System; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D051381:Rats; D020218:Response Elements / Q000235:genetics; D051899:Smad2 Protein; D051900:Smad3 Protein; D051901:Smad4 Protein; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation / Q000187:drug effects* / Q000235:genetics
D004268:DNA-Binding Proteins; D037201:Follicle Stimulating Hormone, beta Subunit; D009687:Nuclear Proteins; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; C497796:SMAD4 protein, human; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; C497784:Smad2 protein, rat; D051900:Smad3 Protein; C497791:Smad3 protein, mouse; C497787:Smad3 protein, rat; D051901:Smad4 Protein; D015534:Trans-Activators; C422405:activin A; D028341:Activins; D007987:Gonadotropin-Releasing Hormone; D028322:Inhibin-beta Subunits