Nature of Functional Estrogen Receptors at the Plasma Membrane

Abstract

Although rapid signaling by estrogen at the plasma membrane is established, it is controversial as to the nature of the receptor protein. Estrogen may bind membrane proteins comparable to classical nuclear estrogen receptors (ERs), but some studies identify nonclassical receptors, such as G protein-coupled receptor (GPR)30. We took several approaches to define membrane-localized estrogen-binding proteins. In endothelial cells (ECs) from ERalpha/ERbeta combined-deleted mice, estradiol (E2) failed to specifically bind, and did not activate cAMP, ERK, or phosphatidyinositol 3-kinase or stimulate DNA synthesis. This is in contrast to wild-type ECs, indicating the lack of any functional estrogen-binding proteins in ERalpha/ERbeta combined-deleted ECs. To directly determine the identity of membrane and nuclear-localized ER, we isolated subcellular receptor pools from MCF7 cells. Putative ER proteins were trypsin digested and subjected to tandem array mass spectrometry. The output analysis identified membrane and nuclear E2-binding proteins as classical human ERalpha. We also determined whether GPR30 plays any role in E2 rapid actions. MCF7 (ER and GPR30 positive) and SKBR-3 (ER negative, GPR30 positive) cells were incubated with E2. Only MCF7 responded with significantly increased signaling. In MCF7, the response to E2 was not different in cells transfected with small interfering RNA to green fluorescent protein or GPR30. In contrast, interfering RNA to ERalpha or ER inhibition prevented rapid signaling and resulting biology in MCF7. In breast cancer and ECs, nuclear and membrane ERs are the same proteins. Furthermore, classical ERs mediate rapid signals induced by E2 in these cells.

Membrane-initiated estrogen signaling in hypothalamic neurons

“…However, it is now widely accepted that some of the actions of estrogen are quite rapid and cannot be attributed to the classical nuclear-initiated steroid signaling of ERα or ERβ. One view is that both nuclear and plasma membrane-associated ERs might be products of the same genes (Razandi et al, 1999, Boulware et al, 2005, Pedram et al, 2006, Szegõ et al, 2006, Dewing et al, 2007. This belief stems primarily from the fact that many of the rapid effects of E2 can be induced by selective ERα or ERβ ligands, antagonized by the ER antagonist, ICI 182,780, or are lost in animals bearing mutations in ERα and/or ERβ genes (Couse and Korach, 1999, Singer et al, 1999, Dubal et al, 2001, Wade et al, 2001, Abraham et al, 2003, Boulware et al, 2005.…”

Calmodulin, a regulatory partner of the estrogen receptor alpha in breast cancer cells

“…This finding does not mean that ERα is sequestered within the nucleus since it has been shown to undergo a dynamic intracellular shuttling (Maruvada et al, 2003;Nonclercq et al, 2007;Stenoien et al, 2000) which seems to determine the type of response to estrogen signaling (Leclercq et al, 2006 (Acconcia, Kumar, 2006;Pedram et al, 2006;Song, 2007). In contrast, nuclear form(s) of ERα directly regulate gene expression (Nuclear Initiated Steroid Signaling, NISS), the receptor functioning in this case as an estrogen-dependent transcription factor (or as a coregulator of other transcription factors).…”

Activation of G protein-coupled estrogen receptor 1 mimics, but does not mediate, the anti-proliferative action of estradiol on pituitary lactotrophs in primary culture

“…One of these is G protein-coupled estrogen receptor 1 (Gper1), which upon binding to estrogens stimulates adenylyl cyclase and EGFR transactivation leading to Ca 2+ mobilization and activation of the mitogen-activated protein kinase Erk1/2 and PI3K/Akt signaling pathways [14,15]. However, it is still controversial that Gper1 is a membrane ER that mediates the non-genomic, rapid effects of estrogens because Pedram et al [16] have shown that E2 fails to activate the rapid multiple pathways in cells that lack classical ERα and ERβ even when Gper1 is present.…”