title: Stable transfection of an estrogen receptor beta cDNA isoform into MDA-MB-231 breast cancer cells.
abstract: We previously reported stable transfection of estrogen receptor alpha (ERalpha) into the ER-negative MDA-MB-231 cells (S30) as a tool to examine the mechanism of action of estrogen and antiestrogens [J. Natl. Cancer Inst. 84 (1992) 580]. To examine the mechanism of ERbeta action directly, we have similarly created ERbeta stable transfectants in MDA-MB-231 cells. MDA-MB-231 cells were stably transfected with ERbeta cDNA and clones were screened by estrogen response element (ERE)-luciferase assay and ERbeta mRNA expression was quantified by real-time RT-PCR. Three stable MDA-MB-231/ERbeta clones were compared with S30 cells with respect to their growth properties, ability to activate ERE- and activating protein-1 (AP-1) luciferase reporter constructs, and the ability to activate the endogenous ER-regulated transforming growth factor alpha (TGFalpha) gene. ERbeta6 and ERbeta27 clones express 300-400-fold and the ERbeta41 clone express 1600-fold higher ERbeta mRNA levels compared with untransfected MDA-MB-231 cells. Unlike S30 cells, 17beta-estradiol (E2) does not inhibit ERbeta41 cell growth. ERE-luciferase activity is induced six-fold by E2 whereas neither 4-hydroxytamoxifen (4-OHT) nor ICI 182, 780 activated an AP-1-luciferase reporter. TGFalpha mRNA is induced in response to E2, but not in response to 4-OHT. MDA-MB-231/ERbeta clones exhibit distinct characteristics from S30 cells including growth properties and the ability to induce TGFalpha gene expression. Furthermore, ERbeta, at least in the context of the MDA-MB-231 cellular milieu, does not enhance AP-1 activity in the presence of antiestrogens. In summary, the availability of both ERalpha and ERbeta stable breast cancer cell lines now allows us to compare and contrast the long-term consequences of individual signal transduction pathways.
authors: Department of Biopharmaceutical Sciences, College of Pharmacy M/C 865, University of Illinois at Chicago, 833 S. Wood Street, Chicago, IL 60612, USA. dtonetti@uic.edu
authors: Debra A
authors: DA
authors: Tonetti
authors: Robyn
authors: R
authors: Rubenstein
authors: Michael
authors: M
authors: DeLeon
authors: Huiping
authors: H
authors: Zhao
authors: Sam G
authors: SG
authors: Pappas
authors: David J
authors: DJ
authors: Bentrem
authors: Bin
authors: B
authors: Chen
authors: Andreas
authors: A
authors: Constantinou
authors: V
authors: V
authors: Craig Jordan
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics* / Q000378:metabolism; D002455:Cell Division / Q000235:genetics; D045744:Cell Line, Tumor; D018076:DNA, Complementary / Q000235:genetics; D004958:Estradiol / Q000031:analogs & derivatives / Q000494:pharmacology; D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000502:physiology; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D012333:RNA, Messenger / Q000032:analysis; D011960:Receptors, Estrogen / Q000235:genetics* / Q000378:metabolism; D020218:Response Elements / Q000502:physiology; D013629:Tamoxifen / Q000031:analogs & derivatives* / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology; D014162:Transfection; D016211:Transforming Growth Factor alpha / Q000819:agonists / Q000378:metabolism
chemical_list: D018076:DNA, Complementary; D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D012333:RNA, Messenger; D011960:Receptors, Estrogen; D018808:Transcription Factor AP-1; D016211:Transforming Growth Factor alpha; D013629:Tamoxifen; C016601:afimoxifene; D004958:Estradiol