Estrogen Receptor β Activates the Human Retinoic Acid Receptorα -1 Promoter in Response to Tamoxifen and Other Estrogen Receptor Antagonists, but Not in Response to Estrogen

Human estrogen receptor-α (hERα) or -β (hERβ) transfected into Hep G2 or COS1 cells each responded to estrogen to increase transcription from an estrogen-responsive element (ERE)-driven reporter vector with similar fold induction through a classical mechanism involving direct receptor binding to DNA. ER antagonists inhibited this estrogen induction through both hERα and hERβ, although raloxifene was more potent through ERα than ERβ, and tamoxifen was more potent via ERβ than ERα. We have shown previously that estrogen stimulated the human retinoic acid receptor-α-1 (hRARα-1) promoter through nonclassical EREs by a mechanism that was ERα dependent, but that did not involve direct receptor binding to DNA. We show here that in contrast to hERα, hERβ did not induce reporter activity driven by the hRARα-1 promoter in the presence of estrogen. While hERβ did not confer estrogen responsiveness on this promoter, it did elicit transcriptional activation in the presence of 4-hydroxytamoxifen (4-OH-Tam). Additionally, this 4-OH-Tam agonist activity via ERβ was completely blocked by estrogen. Like ERα, transcriptional activation of this promoter by ERβ was not mediated by direct receptor binding to DNA. While hERα was shown to act through two estrogen-responsive sequences within the promoter, hERβ acted only at the 3′-region, through two Sp1 sites, in response to 4-OH-Tam. Other ER antagonists including raloxifene, ICI-164,384 and ICI-182,780 also acted as agonists through ERβ via the hRARα-1 promoter. Through the use of mutant and chimeric receptors, it was shown that the 4-OH-Tam activity via ERβ from the hRARα-1 promoter in Hep G2 cells required the amino-terminal region of ERβ, a region that was not necessary for estrogen-induced ERβ activity from an ERE in Hep G2 cells. Additionally, the progesterone receptor (PR) antagonist RU486 acted as a weak (IC50 >1 μm) antagonist via hERα and as a fairly potent (IC50 ∼200 nm) antagonist via hERβ from an ERE-driven reporter in cells that do not express PR. Although RU486 bound only weakly to ERα or ERβ in vitro, it did bind to ERβ in whole-cell binding assays, and therefore, it is likely metabolized to an ERβ-interacting compound in the cell. Interestingly, RU486 acted as an agonist through ERβ to stimulate the hRARα-1 promoter in Hep G2 cells. These findings may have ramifications in breast cancer treatment regimens utilizing tamoxifen or other ER antagonists and may explain some of the known estrogenic or antiestrogenic biological actions of RU486.