Membrane estrogen receptors identified by multiple antibody labeling and impeded-ligand binding

T. C. Pappas, B. Gametchu, C. S. Watson

    Research output: Contribution to journalArticle

    455 Citations (Scopus)

    Abstract

    GH3/B6 rat pituitary tumor cells exhibit rapid prolactin release (within 5 min) when treated with nanomolar amounts of estrogen. However, the putative protein mediator of this nongenomic action has not been described. Using antibodies directed against a peptide representing the hinge region of the intracellular estrogen receptor (iER), we have demonstrated that these cells contain a membrane ER (mER). We now report that confocal scanning laser microscopy of cells labeled live with the anti-peptide antibody further supports a membrane localization of ER. The monoclonal antibodies H226 and H222 and a polyclonal antibody, ER21, each recognizing a unique epitope on iER (NH2 terminal to the DNA-binding region, within the steroid binding region, and the NH2-terminal end, respectively), also immunohistochemically label membrane proteins of immuno-selected GH3/B6 cells. These cells also specifically bind a fluorescent estrogen-BSA conjugate. Coincubation of cells with anti-ER antibody and the fluorescent estrogen-BSA conjugate reveals that these labels colocalize on cells. These results suggest that mER may be structurally similar to iER.

    Original languageEnglish (US)
    Pages (from-to)404-410
    Number of pages7
    JournalFASEB Journal
    Volume9
    Issue number5
    StatePublished - 1995

    Fingerprint

    Estrogen Receptors
    Labeling
    Ligands
    Membranes
    conjugated estrogens
    Estrogens
    antibodies
    Antibodies
    Labels
    cells
    Peptides
    peptides
    Hinges
    Anti-Idiotypic Antibodies
    Prolactin
    confocal laser scanning microscopy
    Rats
    Tumors
    Epitopes
    Microscopic examination

    Keywords

    • confocal microscopy
    • E
    • GH/B6
    • monoclonal antibody

    ASJC Scopus subject areas

    • Agricultural and Biological Sciences (miscellaneous)
    • Biochemistry, Genetics and Molecular Biology(all)
    • Biochemistry
    • Cell Biology

    Cite this

    Pappas, T. C., Gametchu, B., & Watson, C. S. (1995). Membrane estrogen receptors identified by multiple antibody labeling and impeded-ligand binding. FASEB Journal, 9(5), 404-410.

    Membrane estrogen receptors identified by multiple antibody labeling and impeded-ligand binding. / Pappas, T. C.; Gametchu, B.; Watson, C. S.

    In: FASEB Journal, Vol. 9, No. 5, 1995, p. 404-410.

    Research output: Contribution to journalArticle

    Pappas, TC, Gametchu, B & Watson, CS 1995, 'Membrane estrogen receptors identified by multiple antibody labeling and impeded-ligand binding', FASEB Journal, vol. 9, no. 5, pp. 404-410.
    Pappas, T. C. ; Gametchu, B. ; Watson, C. S. / Membrane estrogen receptors identified by multiple antibody labeling and impeded-ligand binding. In: FASEB Journal. 1995 ; Vol. 9, No. 5. pp. 404-410.
    @article{a9c9d8c9c9294fadab9e000da042eb26,
    title = "Membrane estrogen receptors identified by multiple antibody labeling and impeded-ligand binding",
    abstract = "GH3/B6 rat pituitary tumor cells exhibit rapid prolactin release (within 5 min) when treated with nanomolar amounts of estrogen. However, the putative protein mediator of this nongenomic action has not been described. Using antibodies directed against a peptide representing the hinge region of the intracellular estrogen receptor (iER), we have demonstrated that these cells contain a membrane ER (mER). We now report that confocal scanning laser microscopy of cells labeled live with the anti-peptide antibody further supports a membrane localization of ER. The monoclonal antibodies H226 and H222 and a polyclonal antibody, ER21, each recognizing a unique epitope on iER (NH2 terminal to the DNA-binding region, within the steroid binding region, and the NH2-terminal end, respectively), also immunohistochemically label membrane proteins of immuno-selected GH3/B6 cells. These cells also specifically bind a fluorescent estrogen-BSA conjugate. Coincubation of cells with anti-ER antibody and the fluorescent estrogen-BSA conjugate reveals that these labels colocalize on cells. These results suggest that mER may be structurally similar to iER.",
    keywords = "confocal microscopy, E, GH/B6, monoclonal antibody",
    author = "Pappas, {T. C.} and B. Gametchu and Watson, {C. S.}",
    year = "1995",
    language = "English (US)",
    volume = "9",
    pages = "404--410",
    journal = "FASEB Journal",
    issn = "0892-6638",
    publisher = "FASEB",
    number = "5",

    }

    TY - JOUR

    T1 - Membrane estrogen receptors identified by multiple antibody labeling and impeded-ligand binding

    AU - Pappas, T. C.

    AU - Gametchu, B.

    AU - Watson, C. S.

    PY - 1995

    Y1 - 1995

    N2 - GH3/B6 rat pituitary tumor cells exhibit rapid prolactin release (within 5 min) when treated with nanomolar amounts of estrogen. However, the putative protein mediator of this nongenomic action has not been described. Using antibodies directed against a peptide representing the hinge region of the intracellular estrogen receptor (iER), we have demonstrated that these cells contain a membrane ER (mER). We now report that confocal scanning laser microscopy of cells labeled live with the anti-peptide antibody further supports a membrane localization of ER. The monoclonal antibodies H226 and H222 and a polyclonal antibody, ER21, each recognizing a unique epitope on iER (NH2 terminal to the DNA-binding region, within the steroid binding region, and the NH2-terminal end, respectively), also immunohistochemically label membrane proteins of immuno-selected GH3/B6 cells. These cells also specifically bind a fluorescent estrogen-BSA conjugate. Coincubation of cells with anti-ER antibody and the fluorescent estrogen-BSA conjugate reveals that these labels colocalize on cells. These results suggest that mER may be structurally similar to iER.

    AB - GH3/B6 rat pituitary tumor cells exhibit rapid prolactin release (within 5 min) when treated with nanomolar amounts of estrogen. However, the putative protein mediator of this nongenomic action has not been described. Using antibodies directed against a peptide representing the hinge region of the intracellular estrogen receptor (iER), we have demonstrated that these cells contain a membrane ER (mER). We now report that confocal scanning laser microscopy of cells labeled live with the anti-peptide antibody further supports a membrane localization of ER. The monoclonal antibodies H226 and H222 and a polyclonal antibody, ER21, each recognizing a unique epitope on iER (NH2 terminal to the DNA-binding region, within the steroid binding region, and the NH2-terminal end, respectively), also immunohistochemically label membrane proteins of immuno-selected GH3/B6 cells. These cells also specifically bind a fluorescent estrogen-BSA conjugate. Coincubation of cells with anti-ER antibody and the fluorescent estrogen-BSA conjugate reveals that these labels colocalize on cells. These results suggest that mER may be structurally similar to iER.

    KW - confocal microscopy

    KW - E

    KW - GH/B6

    KW - monoclonal antibody

    UR - http://www.scopus.com/inward/record.url?scp=0028925081&partnerID=8YFLogxK

    UR - http://www.scopus.com/inward/citedby.url?scp=0028925081&partnerID=8YFLogxK

    M3 - Article

    C2 - 7896011

    AN - SCOPUS:0028925081

    VL - 9

    SP - 404

    EP - 410

    JO - FASEB Journal

    JF - FASEB Journal

    SN - 0892-6638

    IS - 5

    ER -