title: Molecular mechanism of the inhibition of estradiol-induced endometrial epithelial cell proliferation by clomiphene citrate.
abstract: We examined the molecular mechanisms of the antiestrogenic effects of clomiphene citrate (CC) in the endometrium using two types of cell lines, Ishikawa and EM-E6/E7/hTERT cells. CC or ICI182780 inhibited 17beta-estradiol (E2)-induced endometrial cell proliferation and transcriptional activation of the estrogen response element (ERE) gene. We directly visualized the ligand-estrogen receptor (ER)alpha interaction using green fluorescent protein (GFP)-tagged ER alpha in a single living cell. Whereas E2 changed the nuclear localization of GFP-ER alpha to a punctate distribution within 5 min, CC or ICI182780 changed the slower and less mobilization of GFP-ER alpha compared with E2. Pretreatment with CC or ICI182780 partly prevented the E2-induced nuclear redistribution of GFP-ER alpha. Fluorescence recovery after photobleaching revealed that GFP-ER alpha mobility treated with E2 was more rapid than that treated by CC or ICI182780. As coactivator recruitment to the ER is essential for ER-dependent transcription, we examined the interaction between ER alpha and steroid receptor coactivator-1 (SRC-1). The complex formation between ER alpha and SRC-1 was significantly increased by E2 but was prevented in the presence of CC or ICI182780 by coimmunoprecipitation. Moreover, the E2-induced colocalization of GFP-ER alpha and SRC-1 was prevented in the presence of CC or ICI182780 according to an immunofluorescence assay. We also observed that the reduction of SRC-1 using small interfering RNA for SRC-1 resulted in the inhibition of E2-induced cell proliferation and transcriptional activation of the ERE gene. Collectively, these results suggest that CC may inhibit E2-induced endometrial epithelial cell proliferation and ERE transactivation by inhibiting the recruitment of SRC-1 to ER alpha.
authors.affiliation: Department of Obstetrics and Gynecology, Yamagata University School of Medicine, Yamagata 990-9585, Japan.
authors.forename: Mitsuyoshi
authors.initials: M
authors.lastname: Amita
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Takahashi
authors.forename: Seiji
authors.initials: S
authors.lastname: Tsutsumi
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Ohta
authors.forename: Keiko
authors.initials: K
authors.lastname: Takata
authors.forename: Noriko
authors.initials: N
authors.lastname: Henmi
authors.forename: Shuichiro
authors.initials: S
authors.lastname: Hara
authors.forename: Hideki
authors.initials: H
authors.lastname: Igarashi
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Takahashi
authors.forename: Hirohisa
authors.initials: H
authors.lastname: Kurachi
mesh_terms: D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D002996:Clomiphene / Q000494:pharmacology*; D015536:Down-Regulation / Q000187:drug effects; D004717:Endometrium / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D004958:Estradiol / Q000494:pharmacology*; D047628:Estrogen Receptor alpha / Q000037:antagonists & inhibitors / Q000378:metabolism; D005260:Female; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D056920:Nuclear Receptor Coactivator 1 / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D034741:RNA, Small Interfering / Q000494:pharmacology; D020218:Response Elements / Q000187:drug effects / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects* / Q000235:genetics; D014018:Tissue Distribution; D015533:Transcriptional Activation / Q000187:drug effects
chemical_list: C506487:ESR1 protein, human; D047628:Estrogen Receptor alpha; D034741:RNA, Small Interfering; D049452:Green Fluorescent Proteins; D002996:Clomiphene; D004958:Estradiol; D056920:Nuclear Receptor Coactivator 1