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A low concentration of genistein induces estrogen receptor-alpha and insulin-like growth factor-I receptor interactions and proliferation in uterine leiomyoma cells.

ABSTRACT: Previously, we found that genistein at low concentrations stimulates the growth of human uterine leiomyoma (LM) cells, but not uterine smooth muscle (myometrial) cells (SMC). The aim of this study was to understand the molecular mechanism whereby genistein causes hyperproliferation of LM cells.The effects of genistein at 1 microg/ml on LM cells and SMC were evaluated using estrogen response element gene reporter, real-time RT-PCR, western blot, immunoprecipitation and cell proliferation assays.Elevated estrogen receptor (ER) transactivation, increased mRNA expression of early estrogen-responsive genes, progesterone receptor and insulin-like growth factor-I (IGF-I), and decreased protein levels of ER-alpha (ER alpha) were found in genistein-treated LM cells, but not SMC. Additionally, extracellular regulated kinase (ERK), Src homology/collagen (Shc) and ER alpha were transiently activated, and interactions between ER alpha and IGF-I receptor (IGF-IR) were rapidly induced by genistein in LM cells. Using ER antagonist ICI 182,780 and MAPK/ERK kinase (MEK) inhibitor PD98059, we found that these early events were inhibited and the proliferative effect of genistein on LM cells was abrogated.ER alpha is involved in the transient activation of ERK/mitogen activated protein kinase (MAPK) by genistein via its early association with IGF-IR, leading to hyper-responsiveness of LM cells and confirming that ER signaling is enhanced by activation of ERK/MAPK in LM cells.

SUBMITTER: Di X

PROVIDER: S-EPMC2733822 | biostudies-other | 2008 Aug

REPOSITORIES: biostudies-other

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A low concentration of genistein induces estrogen receptor-alpha and insulin-like growth factor-I receptor interactions and proliferation in uterine leiomyoma cells.

Di X X Yu L L Moore A B AB Castro L L Zheng X X Hermon T T Dixon D D

Human reproduction (Oxford, England) 20080520 8

<h4>Background</h4>Previously, we found that genistein at low concentrations stimulates the growth of human uterine leiomyoma (LM) cells, but not uterine smooth muscle (myometrial) cells (SMC). The aim of this study was to understand the molecular mechanism whereby genistein causes hyperproliferation of LM cells.<h4>Methods</h4>The effects of genistein at 1 microg/ml on LM cells and SMC were evaluated using estrogen response element gene reporter, real-time RT-PCR, western blot, immunoprecipitat ...[more]

PMID: 18492705

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Project description:ER?36 is a naturally occurring, membrane-associated, isoform of estrogen receptor ?. The expression of ER?36 is due to alternative splicing and different promoter usage. ER?36 is a dominant-negative effector of ER?66-mediated transactivational activities and has the potential to trigger membrane-initiated mitogenic, nongenomic, estrogen signaling; however, the subcellular localization of ER?36 remains controversial. To determine the cellular localization of ER?36 in estrogen-responsive human uterine smooth muscle (ht-UtSMC) and leiomyoma (fibroid; ht-UtLM) cells, we conducted systematic confocal microscopy and subcellular fractionation analysis using ER?36 antibodies. With Image J colocalizaton analysis plugin, confocal images were analyzed to obtain a Pearson's Correlation Coefficient (PCC) to quantify signal colocalization of ER?36 with mitochondrial, endoplasmic reticulum, and cytoskeletal components in both cell lines. When cells were double-stained with an ER?36 antibody and a mitochondrial-specific dye, MitoTracker, the PCC for the two channel signals were both greater than 0.75, indicating strong correlation between ER?36 and mitochondrial signals in the two cell lines. A blocking peptide competition assay confirmed that the mitochondria-associated ER?36 signal detected by confocal analysis was specific for ER?36. In contrast, confocal images double-stained with an ER?36 antibody and endoplasmic reticulum or cytoskeletal markers, had PCCs that were all less than 0.4, indicating no or very weak signal correlation. Fractionation studies showed that ER?36 existed predominantly in membrane fractions, with minimal or undetected amounts in the cytosol, nuclear, chromatin, and cytoskeletal fractions. With isolated mitochondrial preparations, we confirmed that a known mitochondrial protein, prohibitin, was present in mitochondria, and by co-immunoprecipitation analysis that ER?36 was associated with prohibitin in ht-UtLM cells. The distinctive colocalization pattern of ER?36 with mitochondria in ht-UtSMC and ht-UtLM cells, and the association of ER?36 with a mitochondrial-specific protein suggest that ER?36 is localized primarily in mitochondria and may play a pivotal role in non-genomic signaling and mitochondrial functions.

Dataset Information

A low concentration of genistein induces estrogen receptor-alpha and insulin-like growth factor-I receptor interactions and proliferation in uterine leiomyoma cells.

Publications

A low concentration of genistein induces estrogen receptor-alpha and insulin-like growth factor-I receptor interactions and proliferation in uterine leiomyoma cells.

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