A growing body of evidence concerning estrogen effects cannot be explained by the classic model of hormone action, which involves the binding to estrogen receptors (ERs) α and ERβ and the interaction of the steroid-receptor complex with specific DNA sequences associated with target genes. Using c-fos proto-oncogene expression as an early molecular sensor of estrogen action in ERα-positive MCF7 and ER-negative SKBR3 breast cancer cells, we have discovered that 17β-estradiol (E2), and the two major phytoestrogens, genistein and quercetin, stimulate c-fos expression through ERα as well as through an ER-independent manner via the G protein-coupled receptor homologue GPR30. The c-fos response is repressed in GPR30-expressing SKBR3 cells transfected with an antisense oligonucleotide against GPR30 and reconstituted in GPR30-deficient MDA-MB 231 and BT-20 breast cancer cells transfected with a GPR30 expression vector. GPR30-dependent activation of ERK1/2 by E2 and phytoestrogens occurs via a Gβγ-associated pertussis toxin-sensitive pathway that requires both Src-related and EGF receptor tyrosine kinase activities. The ability of E2 and phytoestrogens to regulate the expression of growth-related genes such as c-fos even in the absence of ER has interesting implications for understanding breast cancer progression.

Estrogens play a crucial role in the development of ovarian tumors; however, the signal transduction pathways involved in hormone action are still poorly defined. The orphan G protein–coupled receptor 30 (GPR30) mediates the nongenomic signaling of 17β-estradiol (E2) in a variety of estrogen-sensitive cancer cells through activation of the epidermal growth factor receptor (EGFR) pathway. Whether estrogen receptor α (ERα) also contributes to GPR30/EGFR signaling is less understood. Here, we show that, in ERα-positive BG-1 ovarian cancer cells, both E2 and the GPR30-selective ligand G-1 induced c-fos expression and estrogen-responsive element (ERE)-independent activity of a c-fos reporter gene, whereas only E2 stimulated an ERE-responsive reporter gene, indicating that GPR30 signaling does not activate ERα-mediated transcription. Similarly, both ligands up-regulated cyclin D1, cyclin E, and cyclin A, whereas only E2 enhanced progesterone receptor expression. Moreover, both GPR30 and ERα expression are required for c-fos stimulation and extracellular signal-regulated kinase (ERK) activation in response to either E2 or G-1. Inhibition of the EGFR transduction pathway inhibited c-fos stimulation and ERK activation by either ligand, suggesting that in ovarian cancer cells GPR30/EGFR signaling relays on ERα expression. Interestingly, we show that both GPR30 and ERα expression along with active EGFR signaling are required for E2-stimulated and G-1–stimulated proliferation of ovarian cancer cells. Because G-1 was able to induce both c-fos expression and proliferation in the ERα-negative/GPR30-positive SKBR3 breast cancer cells, the requirement for ERα expression in GPR30/EGFR signaling may depend on the specific cellular context of different tumor types. [Cancer Res 2007;67(4):1859–66]

Steroid hormones such as estrogens are known to signal through ligand-regulated transcription factors of the nuclear receptor superfamily. In addition, they elicit rapid nongenomic responses from membrane-associated receptors. One of these receptors belongs to an entirely different family of proteins. The G protein-coupled and seven-transmembrane receptor, GPR30, is now widely recognized as an estrogen receptor (ER), hence its official new acronym GPER. It appears to mediate a wide range of responses to estrogen in a large variety of cell types. Its functions are clearly distinct from those of the classical nuclear ERs, although these pathways may overlap and interact in some cases. Here, we review the history of the discovery of this new ER, the evidence for the claim that it is an ER, its signal transduction, and its potential functions in physiology and disease.

The growth of both normal and transformed epithelial cells of the female reproductive system is stimulated by estrogens, mainly through the activation of estrogen receptor α (ERα), which is a ligand-regulated transcription factor. The selective ER modulator tamoxifen (TAM) has been widely used as an ER antagonist in breast tumor; however, long-term treatment is associated with an increased risk of endometrial cancer. To provide new insights into the potential mechanisms involved in the agonistic activity exerted by TAM in the uterus, we evaluated the potential of 4-hydroxytamoxifen (OHT), the active metabolite of TAM, to transactivate wild-type ERα and its splice variant expressed in Ishikawa and HEC1A endometrial tumor cells, respectively. OHT was able to antagonize only the activation of ERα by 17β-estradiol (E2) in Ishikawa cells, whereas it up-regulated c-fos expression in a rapid manner similar to E2 and independently of ERα in both cell lines. This stimulation occurred through the G protein-coupled receptor named GPR30 and required Src-related and epidermal growth factor receptor tyrosine kinase activities, along with the activation of both ERK1/2 and phosphatidylinositol 3-kinase/AKT pathways. Most importantly, OHT, like E2, stimulated the proliferation of Ishikawa as well as HEC1A cells. Transfecting a GPR30 antisense expression vector in both endometrial cancer cell lines, OHT was no longer able to induce growth effects, whereas the proliferative response to E2 was completely abrogated only in HEC1A cells. Furthermore, in the presence of the inhibitors of MAPK and phosphatidylinositol 3-kinase pathways, PD 98059 and wortmannin, respectively, E2 and OHT did not elicit growth stimulation. Our data demonstrate a new mode of action of E2 and OHT in endometrial cancer cells, contributing to a better understanding of the molecular mechanisms involved in their uterine agonistic activity.