Abstract
BackgroundAlthough membrane-associated estrogen receptors (mERs) have been known to play important role in steroid-induced signal transmission, we still know little about their function in the estrogen-induced proliferation of breast cancer cells.MethodsIn our current work we tried to separate membrane-initiated estrogen receptor signaling from the overall estrogenic effect in MCF-7 breast carcinoma cells. Re-analyzing expression data from multiple microarray experiments, we selected a set of key regulatory genes involved in proliferation regulation and estrogen signaling to monitor estrogen-induced transcription changes. We then compared these expression changes after 17β-estradiol and a membrane receptor selective estrogen–BSA treatment using quantitative real-time PCR. In order to follow receptor trafficking we used light and electron microscopy.ResultsOur quantitative real-time PCR results confirmed that the selective membrane receptor agonist, estrogen–BSA induces similarly pronounced expression changes regarding these genes as 17β-estradiol. Morphological study revealed that the membrane-bound form of classical estrogen receptor alpha is internalized after ligand binding via dynamin-dependent, caveola-mediated endocytosis. Inhibition of this internalization with dynamin inhibitor, dynasore practically abolished the regulatory effect of E2-BSA, suggesting that interaction and internalization with the scaffold protein is necessary for effective signaling.ConclusionsThe physiological role of plasma membrane estrogen receptor alpha is intensively studied, yet there are still several aspects of it to be resolved. The dynamin-dependent, ligand-mediated internalization of mERs seems to play an important role in estrogen signaling. Our results may serve as another example of how membrane initiated estrogen signaling and nuclear receptor initiated signaling overlap and form an intertwined system.
Highlights
Membrane-associated estrogen receptors have been known to play important role in steroid-induced signal transmission, we still know little about their function in the estrogen-induced proliferation of breast cancer cells
By mapping the probe sets to genes, we have identified 285 unique upregulated and 49 downregulated genes that were differentially expressed. (All results obtained from microarray meta-analysis are available in Additional file 1: Table Sheet 1.) The selected key genes (KCNK5, lysine demethylase 4B (KDM4B), Myc proto-oncogene (MYC), and cyclin D1 (CCND1)) were all significantly upregulated while Erb-B2 Receptor Tyrosine Kinase 2 (ERBB2) (HER2) was downregulated, its expression change was not significant compared to untreated cells
We further focused on the expression of the above-mentioned five genes as their expression has already been extensively studied and all have been confirmed as a target of E2
Summary
Membrane-associated estrogen receptors (mERs) have been known to play important role in steroid-induced signal transmission, we still know little about their function in the estrogen-induced proliferation of breast cancer cells. Marczell et al Eur J Med Res (2018) 23:31 in cellular functions via second messenger pathways through which they contribute to the transcriptional effects of estrogen, including the regulation of proliferation [5], cell migration, and development [6]. These effects are mostly transmitted via the activation of alternative MAP kinase pathways [7,8,9,10]. Membrane-bound ERs show different signaling kinetics and are more exposed to various stimuli such as paracrine, autocrine, or endocrine signals. Caveolin-1 is known to be involved in trafficking of the estrogen receptor alpha to and from the cell surface and in maintaining an environment where coupling between signaling partners is possible [18]
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