Effects of FSH and 17beta-estradiol on the transactivation of estrogen-regulated promoters and cell proliferation in L cells.
In the present study, we analyzed human follicle-stimulating hormone (FSH)-induced cell proliferation and transactivation of estrogen-sensitive reporter genes-in L cells stably expressing the human FSH receptor [L-(hFSHR(+)) cells]. In order to dissect the signaling pathways involved in this process, L-(hFSHR(+)) cells were transiently transfected with either the 3X-ERE-TAT-Luc or the ERE-VitA2-TK-CAT reporter genes and treated with FSH or PKA activators (cholera toxin, forskolin and 8-Br-cAMP) in the presence or absence of various kinase inhibitors. We found that FSH and all PKA activators, specifically induced transactivation of both reporter genes. Transactivation of estrogen-sensitive genes by FSH or PKA activators were blocked (approximately 90%) by H89 (PKA inhibitor) and LY294002 but not by Wortmannin (PI3-K inhibitors), 4-OH-tamoxifen, ICI182,780 or SB203580 (p38 MAPK inhibitor); PD98059 (ERK1/2 inhibitor) partially (approximately 30%) blocked the FSH-mediated effect. The combination of FSH and estradiol resulted in a synergistic effect on transactivation as well as on cell proliferation, and this enhancement was attenuated by antiestrogens. We additionally analyzed the participation of the coactivators SRC-1 and cAMP response element binding protein (CREB)-binding protein (CBP) in FSH-evoked estrogen receptor (ER)-dependent transactivation; we found that CBP but not SRC-1 potentiated FSH-induced transcriptional activation of both ER-sensitive reporters, being this effect stronger on the ERE-VitA2-TK-CAT than on the 3X-ERE-TAT-Luc reporter. Thus, in L-(hFSHR(+)) cells FSH induces transcriptional activation of estrogen-sensitive genes through an A-kinase-triggered signaling pathway, using also to a lesser extent the ERK1/2 and p38 pathways. PI3-K is not apparently involved in this FSH-mediated process since LY294002, but not Wortmannin, specifically binds ERs and completely blocks estrogen action. Presumably, CBP cooperates with the ER on genes that contain estrogen responsive elements through mechanisms involving the participation of other proteins and/or basal transcription factors (e.g. CREB), which in turn mediate the transcriptional response of estrogen-sensitive reporter genes to FSH stimulation.
Pasapera
Ana María
AM
Research Unit in Reproductive Medicine, Hospital de Gineco-Obstetricia Luis Castelazo Ayala, Instituto Mexicano del Seguro Social, Apdo. Postal 99-065, Unidad Independencia, México D.F. C.P. 10101, Mexico.
Jiménez-Aguilera
María del Pilar
Mdel P
Chauchereau
Anne
A
Milgrom
Edwin
E
Olivares
Aleida
A
Uribe
Aída
A
Gutiérrez-Sagal
Rubén
R
Ulloa-Aguirre
Alfredo
A
D000818:Animals; D001667:Binding, Competitive; D050882:CREB-Binding Protein; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002522:Chlorocebus aethiops; D004357:Drug Synergism; D004958:Estradiol / Q000494:pharmacology*; D005640:Follicle Stimulating Hormone / Q000494:pharmacology*; D017930:Genes, Reporter / Q000187:drug effects*; D006801:Humans; D007739:L Cells / Q000187:drug effects*; D051379:Mice; D009687:Nuclear Proteins / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011960:Receptors, Estrogen / Q000378:metabolism; D011962:Receptors, FSH / Q000378:metabolism; D015534:Trans-Activators / Q000494:pharmacology; D015533:Transcriptional Activation / Q000187:drug effects*
D009687:Nuclear Proteins; D011960:Receptors, Estrogen; D011962:Receptors, FSH; D015534:Trans-Activators; D004958:Estradiol; D005640:Follicle Stimulating Hormone; D050882:CREB-Binding Protein; C494363:CREBBP protein, human; C494364:Crebbp protein, mouse