title: Estrogen-induced activation of Erk-1 and Erk-2 requires the G protein-coupled receptor homolog, GPR30, and occurs via trans-activation of the epidermal growth factor receptor through release of HB-EGF.
abstract: Estrogen rapidly activates the mitogen-activated protein kinases, Erk-1 and Erk-2, via an as yet unknown mechanism. Here, evidence is provided that estrogen-induced Erk-1/-2 activation occurs independently of known estrogen receptors, but requires the expression of the G protein-coupled receptor homolog, GPR30. We show that 17beta-estradiol activates Erk-1/-2 not only in MCF-7 cells, which express both estrogen receptor alpha (ER alpha) and ER beta, but also in SKBR3 breast cancer cells, which fail to express either receptor. Immunoblot analysis using GPR30 peptide antibodies showed that this estrogen response was associated with the presence of GPR30 protein in these cells. MDA-MB-231 breast cancer cells (ER alpha-, ER beta+) are GPR30 deficient and insensitive to Erk-1/-2 activation by 17beta-estradiol. Transfection of MDA-MB-231 cells with a GPR30 complementary DNA resulted in overexpression of GPR30 protein and conversion to an estrogen-responsive phenotype. In addition, GPR30-dependent Erk-1/-2 activation was triggered by ER antagonists, including ICI 182,780, yet not by 17alpha-estradiol or progesterone. Consistent with acting through a G protein-coupled receptor, estradiol signaling to Erk-1/-2 occurred via a Gbetagamma-dependent, pertussis toxin-sensitive pathway that required Src-related tyrosine kinase activity and tyrosine phosphorylation of tyrosine 317 of the Shc adapter protein. Reinforcing this idea, estradiol signaling to Erk-1/-2 was dependent upon trans-activation of the epidermal growth factor (EGF) receptor via release of heparan-bound EGF (HB-EGF). Estradiol signaling to Erk-1/-2 could be blocked by: 1) inhibiting EGF-receptor tyrosine kinase activity, 2) neutralizing HB-EGF with antibodies, or 3) down-modulating HB-EGF from the cell surface with the diphtheria toxin mutant, CRM-197. Our data imply that ER-negative breast tumors that continue to express GPR30 may use estrogen to drive growth factor-dependent cellular responses.
authors.affiliation: Department of Medicine and Surgery, Rhode Island Hospital and Brown University, Providence 02903, USA. edward_filardo@brown.edu
authors.forename: E J
authors.initials: EJ
authors.lastname: Filardo
authors.forename: J A
authors.initials: JA
authors.lastname: Quinn
authors.forename: K I
authors.initials: KI
authors.lastname: Bland
authors.forename: A R
authors.initials: AR
authors.lastname: Frackelton
mesh_terms: D001943:Breast Neoplasms / Q000737:chemistry / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000378:metabolism*; D066246:ErbB Receptors / Q000502:physiology*; D004958:Estradiol / Q000031:analogs & derivatives* / Q000494:pharmacology*; D000077267:Fulvestrant; D006493:Heparin / Q000378:metabolism; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011956:Receptors, Cell Surface / Q000235:genetics / Q000502:physiology*; D011960:Receptors, Estrogen / Q000032:analysis / Q000037:antagonists & inhibitors; D043562:Receptors, G-Protein-Coupled; D015398:Signal Transduction; D015533:Transcriptional Activation; D014162:Transfection; D014407:Tumor Cells, Cultured; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: D004791:Enzyme Inhibitors; C109584:GPER1 protein, human; D011956:Receptors, Cell Surface; D011960:Receptors, Estrogen; D043562:Receptors, G-Protein-Coupled; D010566:Virulence Factors, Bordetella; D000077267:Fulvestrant; D004958:Estradiol; D004815:Epidermal Growth Factor; D006493:Heparin; D037342:Pertussis Toxin; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases