The dynamic and elusive membrane estrogen receptor-α
Introduction
We have previously used the properties of multiple antibodies (Abs) to identify the membrane form of estrogen receptor-α (mERα) (summarized in Fig. 1) and probe its membrane orientation and function. More recently, we have developed Ab-based techniques and experimental designs to quantify different subcellularly located forms of estrogen receptor-α (membrane and nuclear). We have focused our studies on the expression of mERα in cells in which they are naturally expressed, unaided by transfection of ERα cDNAs into recipient receptorless cells.
The ability to separate populations of cells into those enriched and depleted for membrane steroid receptor expression [1], [2], [3], [4] has been essential to demonstrating affiliated functions, because only a subset of cells in clonal populations have mERα. Once selected, the drift of clonal cell populations back to mERα-negative status constantly threatens studies designed to demonstrate functional correlates. This intriguing transitory presence of mERα caused us to review our past data for clues to mERα regulation and provided guidelines for specific study of the regulatory influences on mERα in order to better control our functional studies. As we now use these measurement tools to ask questions about the comparative levels and regulatory influences on membrane vs. intracellular estrogen receptors, we may discover more about the cooperative nature of these forms and their separate functions.
Section snippets
Cell culture
GH3/B6/F10 (F10) cells were subcloned from GH3/B6 cells (a gift from Dr. Bernard Dufy) by limiting dilution cloning, as previously described [1]. These cells were routinely cultured at 37°C in serum-supplemented medium consisting of Ham’s F10 medium (GIBCO-BRL, Grand Island, NY), 12.5% heat-inactivated horse serum, 2.5% heat-inactivated defined-supplemented calf serum, and 1.5% heat-inactivated fetal calf serum (FCS) (all sera supplied by Hyclone, Logan, UT). In preparation for these
The influence of fixation method on cell permeability and resulting distinctions between receptor forms
Cell types vary tremendously in the lipid and protein composition of their membranes, and so are variably penetrated by detergents and the components of various fixative mixtures. In working with a protein which is present both in the membrane and in the nucleus, careful attention to the ingredients and concentrations of fixatives, and the length of treatment times is necessary to delineate receptor populations in specific subcellular locations. In the case of ERα, this is particularly
Acknowledgments
C.H.C. was supported by the Zelda Zinn Casper Fellowship Fund, the NIEHS Toxicology Training Grant 5232ES07254, and the UTMB Centennial Center for Environmental Toxicology. The authors wish to thank Dr. Darrell Carney for use of his plate reader and microscope and Dr. David Konkel for critical reading of the manuscript. Both are from the Univ. of Texas Medical Branch, Galveston, TX.
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Is the membrane estrogen receptor, GPER1, a promiscuous receptor that modulates nuclear estrogen receptor-mediated functions in the brain?
2018, Hormones and BehaviorCitation Excerpt :The identity of this receptor has been controversial since different cell types appear to express different mERs. For example, small amounts of ERα or truncated ERα variant receptors have been shown in the plasma membrane in hypothalamic neurons (Dominguez and Micevych, 2010), hypothalamic cell lines (Dominguez et al., 2013), in presynaptic boutons of CA1 hippocampal neurons (Hart et al., 2007) and in MCF-7 breast cancer cells (Watson et al., 2002). Both ERα and ERβ have been demonstrated at the plasma membrane in caveolae in endothelial cells (Chambliss et al., 2002; Chambliss et al., 2000) and in oligodendrocytes (Hirahara et al., 2009).
The structures and inhibitory effects of Buame [N-(3-hydroxy-1,3,5(10)- estratrien-17β-yl)-butylamine] and Diebud [N,N′-bis-(3-hydroxy-1,3, 5(10)-estratrien-17β-yl)-1,4-butanediamine] on platelet aggregation
2012, SteroidsCitation Excerpt :ERβ has a lower affinity for 17β-estradiol than ERα [17]. The membrane ER shares some structural characteristics with the nuclear ER, particularly with ERα [18–20]. In addition, the 17β-estradiol has already been shown to bind to nuclear estrogen receptors that are tethered to the plasma membrane, activating signaling kinases [21].
Functional interaction of fibroblast growth factor-8, bone morphogenetic protein and estrogen receptor in breast cancer cell proliferation
2011, Molecular and Cellular EndocrinologyCitation Excerpt :Based on the findings that estrogen induced rapid activation of MAPKs, including ERK, p38 and SAPK/JNK pathways, and that BSA-conjugated estradiol, which cannot bind to nuclear ERs (Taguchi et al., 2004), induced modest but significant cell mitosis, nongenomic effects are also involved, at least in part, in the induction of cell proliferation by estrogen. Immunoreactive ER antigen was reportedly detected on the surface of both naturally ER-positive cells and in cells transfected with ER expression constructs (Watson et al., 2002). ER-transfected cells also resulted in detectable membrane ER, in which estradiol mediates MAPK action, cAMP-PKA activation and the inositol-triphosphate (PI3K) pathway (Razandi et al., 1999).
GPRC6A mediates the non-genomic effects of steroids
2010, Journal of Biological ChemistryCitation Excerpt :The serum FSH was not changed before or after administration of testosterone between wild type and GPRC6A−/− mice (Fig. 7D, right panel). Several mechanisms have been proposed to explain the rapid cellular responses to steroids (5, 9), such as translocation of steroid nuclear receptors to the cell surface membrane (12–14), nonspecific effects of steroids on the fluidity of lipids in the plasma membrane, direct allosteric modification of ligand-gated ion channels (5, 48, 49), and GPCRs (9, 15, 16, 50, 51). GPRC6A is an amino acid, calcium, and osteocalcin sensing G-protein coupled receptor that appears to coordinate anabolic responses in multiple tissues (25).
Estrogenic Endocrine Disruptors: Molecular Characteristics and Human Impacts
2010, Comprehensive Toxicology, Second Edition