Quantitative measurement of estrogen-induced ERK 1 and 2 activation via multiple membrane-initiated signaling pathways

Nataliya N. Bulayeva, Bahiru Gametchu, Cheryl S. Watson

    Research output: Contribution to journalArticle

    95 Citations (Scopus)

    Abstract

    Estradiol (E2) and other steroids have recently been shown to initiate various intracellular signaling cascades from the plasma membrane, including those stimulating mitogen-activated protein kinases (MAPKs), and particularly extracellular-regulated kinases (ERKs). In this study we demonstrated the ability of E2 to activate ERKs in the GH3/B6/F10 pituitary tumor cell line, originally selected for its enhanced expression of membrane estrogen receptor-α (mERα). We compared E2 to its cell-impermeable analog (E2 conjugated to peroxidase, E 2-P), and to the synthetic estrogen diethylstilbestrol (DES). Time-dependent ERK activation was quantified with a novel fixed cell-based immunoassay developed to efficiently determine activation by multiple compounds over multiple parameters. Both E2 and DES produced bimodal responses, but with distinctly different time courses of enzyme phosphorylation (activation) and inactivation; E2-P induced a monophasic ERK activation. E2 also phosphorylated ERKs in concentration-dependent manner with two concentration optima (10-14 and 10-8M). Inhibitors were employed to determine pathway (ER, EGFR, membrane organization, PI3 kinase, Src kinase, Ca2+) involvement and timing of pathway activations; all affected ERK activation as early as 3-6min, suggesting simultaneous, not sequential, activation. Therefore, E2 and other estrogenic compounds can produce rapid ERK phosphorylations via nongenomic pathways, using more than one pathway for signal generation.

    Original languageEnglish (US)
    Pages (from-to)181-192
    Number of pages12
    JournalSteroids
    Volume69
    Issue number3
    DOIs
    StatePublished - Mar 2004

    Fingerprint

    Estrogens
    Phosphotransferases
    Chemical activation
    Membranes
    Phosphorylation
    Diethylstilbestrol
    Estradiol Congeners
    Enzyme Activation
    src-Family Kinases
    Pituitary Neoplasms
    Cell membranes
    Mitogen-Activated Protein Kinases
    Tumor Cell Line
    Phosphatidylinositol 3-Kinases
    Immunoassay
    Estrogen Receptors
    Peroxidase
    Tumors
    Estradiol
    Signal Transduction

    Keywords

    • Ca
    • EGF receptor
    • Membrane estrogen receptor
    • Non-genomic
    • PI3K
    • Signaling pathway inhibitors
    • Src kinase

    ASJC Scopus subject areas

    • Biochemistry
    • Endocrinology
    • Molecular Biology

    Cite this

    Quantitative measurement of estrogen-induced ERK 1 and 2 activation via multiple membrane-initiated signaling pathways. / Bulayeva, Nataliya N.; Gametchu, Bahiru; Watson, Cheryl S.

    In: Steroids, Vol. 69, No. 3, 03.2004, p. 181-192.

    Research output: Contribution to journalArticle

    Bulayeva, Nataliya N. ; Gametchu, Bahiru ; Watson, Cheryl S. / Quantitative measurement of estrogen-induced ERK 1 and 2 activation via multiple membrane-initiated signaling pathways. In: Steroids. 2004 ; Vol. 69, No. 3. pp. 181-192.
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