title: Estrogen receptor beta as a novel target of androgen receptor action in breast cancer cell lines.
abstract: BACKGROUND
  The two isoforms of estrogen receptor (ER) alpha and beta play opposite roles in regulating proliferation and differentiation of breast cancers, with ER-alpha mediating mitogenic effects and ER-beta acting as a tumor suppressor. Emerging data have reported that androgen receptor (AR) activation inhibits ER-positive breast cancer progression mainly by antagonizing ER-alpha signaling. However, to date no studies have specifically evaluated a potential involvement of ER-beta in the inhibitory effects of androgens.
  
  
  METHODS
  ER-beta expression was examined in human breast cancer cell lines using real-time PCR, Western blotting and small interfering RNA (siRNA) assays. Mutagenesis studies, electromobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) analysis were performed to assess the effects of mibolerone/AR on ER-beta promoter activity and binding.
  
  
  RESULTS
  In this study, we demonstrate that mibolerone, a synthetic androgen ligand, up-regulates ER-beta mRNA and protein levels in ER-positive breast cancer cells. Transient transfection experiments, using a vector containing the human ER-beta promoter region, show that mibolerone increases basal ER-beta promoter activity. Site-directed mutagenesis and deletion analysis reveal that an androgen response element (ARE), TGTTCT motif located at positions -383 and -377, is critical for mibolerone-induced ER-beta up-regulation in breast cancer cells. This occurs through an increased recruitment of AR to the ARE site within the ER-beta promoter region, along with an enhanced occupancy of RNA polymerase II. Finally, silencing of ER-beta gene expression by RNA interference is able to partially reverse the effects of mibolerone on cell proliferation, p21 and cyclin D1 expression.
  
  
  CONCLUSIONS
  Collectively, these data provide evidence for a novel mechanism by which activated AR, through an up-regulation of ER-beta gene expression, inhibits breast cancer cell growth.
authors.forename: Pietro
authors.initials: P
authors.lastname: Rizza
authors.forename: Ines
authors.initials: I
authors.lastname: Barone
authors.forename: Domenico
authors.initials: D
authors.lastname: Zito
authors.forename: Francesca
authors.initials: F
authors.lastname: Giordano
authors.forename: Marilena
authors.initials: M
authors.lastname: Lanzino
authors.forename: Francesca
authors.initials: F
authors.lastname: De Amicis
authors.forename: Loredana
authors.initials: L
authors.lastname: Mauro
authors.forename: Diego
authors.initials: D
authors.lastname: Sisci
authors.forename: Stefania
authors.initials: S
authors.lastname: Catalano
authors.forename: Karin
authors.initials: K
authors.lastname: Dahlman Wright
authors.forename: Jan-Ake
authors.initials: JA
authors.lastname: Gustafsson
authors.forename: Sebastiano
authors.initials: S
authors.lastname: Andò
mesh_terms: D001943:Breast Neoplasms / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000235:genetics; D019938:Cyclin D1 / Q000096:biosynthesis; D004789:Enzyme Activation; D047628:Estrogen Receptor alpha / Q000378:metabolism; D047629:Estrogen Receptor beta / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D005260:Female; D006801:Humans; D061986:MCF-7 Cells; D016297:Mutagenesis, Site-Directed; D009277:Nandrolone / Q000031:analogs & derivatives / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000187:drug effects; D011485:Protein Binding; D034622:RNA Interference; D012319:RNA Polymerase II / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D034741:RNA, Small Interfering; D011944:Receptors, Androgen / Q000378:metabolism*; D045165:Testosterone Congeners / Q000494:pharmacology; D015854:Up-Regulation; D020741:rho GTP-Binding Proteins / Q000096:biosynthesis
chemical_list: C506487:ESR1 protein, human; D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011944:Receptors, Androgen; D045165:Testosterone Congeners; D019938:Cyclin D1; D009277:Nandrolone; C100075:mibolerone; D012319:RNA Polymerase II; D020741:rho GTP-Binding Proteins