title: Epidermal growth factor receptor downregulation in cultured bovine cumulus cells: reconstitution of calcium signaling and stimulated membrane permeabilization.
abstract: Cumulus cell-oocyte complexes (COCs), cultured in vitro, are competent for maturation and fertilization. Inclusion of epidermal growth factor (EGF) in the COC culture medium enhances in vitro maturation and subsequent embryonic development. It has been shown that isolated COCs exposed to EGF respond with a prolonged and pulsatile release of Ca2+ into the extra-cellular medium and that cumulus cells (CCs) of complexes exhibit both a slow rise in intracellular [Ca2+] ([Ca2+]i) and plasma membrane permeabilization in response to EGF. These unusual signaling responses were examined in isolated, cultured bovine CCs. Few individual CCs showed [Ca2+]i increases; the lack of response was found to be due to decrease of expression of endogenous EGF receptors after dissociation. CCs transfected with a human EGF receptor-GFP fusion protein showed robust, prolonged, EGF-stimulated [Ca2+]i elevations characteristic of CC responses in intact COCs. Many CCs that responded to EGF stimulation with a [Ca2+]i rise also released entrapped fura-2 dye at the peak of the [Ca2+]i response, suggesting that CC permeabilization and death follows activation of the EGF receptor. The [Ca2+]i elevation due to EGF stimulation and subsequent membrane permeabilization was shown to be mediated by the inositol triphosphate signaling pathway.
authors.affiliation: Molecular and Cellular Biology Program and Department of Biochemistry and Molecular Biology, University of Massachusetts, 710 N. Pleasant St, Amherst, Massachusetts 01003, USA.
authors.forename: Zhong
authors.initials: Z
authors.lastname: Zhao
authors.forename: Damien
authors.initials: D
authors.lastname: Garbett
authors.forename: Julia L
authors.initials: JL
authors.lastname: Hill
authors.forename: David J
authors.initials: DJ
authors.lastname: Gross
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling; D002417:Cattle; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D018920:Coculture Techniques; D015536:Down-Regulation; D004815:Epidermal Growth Factor / Q000378:metabolism / Q000494:pharmacology*; D066246:ErbB Receptors / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D005260:Female; D005455:Fluorescent Antibody Technique; D016257:Fura-2; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D009865:Oocytes / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis; D013268:Stimulation, Chemical; D014162:Transfection / Q000379:methods
chemical_list: D007295:Inositol Phosphates; D012333:RNA, Messenger; D049452:Green Fluorescent Proteins; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D002118:Calcium; D016257:Fura-2