Identification of Estrogen-Responsive Genes by Complementary Deoxyribonucleic Acid Microarray and Characterization of a Novel Early Estrogen-Induced Gene:EEIG1

Abstract

Estrogen receptors (ERs) are nuclear transcription factors that regulate gene expression in response to estrogen and estrogen-like compounds. Identification of estrogen-regulated genes in target cells is an essential step toward understanding the molecular mechanisms of estrogen action. Using cDNA microarray examinations, 19 genes were identified as induced by 17 beta-estradiol in MCF-7 cells, 10 of which have been reported previously to be estrogen responsive or to be linked with ER status. Five known estrogen-regulated genes, E2IG4, IGFBP4, SLC2A1, XBP1 and B4GALT1, and AFG3L1, responded quickly to estrogen treatment. A novel estrogen-responsive gene was identified and named EEIG1for early estrogen-induced gene 1. EEIG1 was clearly induced by 17 beta-estradiol within 2 h of treatment, and was widely responsive to a group of estrogenic compounds including natural and synthetic estrogens and estrogenic environmental compounds. EEIG1 was expressed in ER-positive but not in ER-negative breast cancer cell lines. EEIG1 expression was repressed by antiestrogens 4-OH-tamoxifen and ICI 182,780 but not by protein synthesis inhibitors cycloheximide and puromycin. These results provide evidence that some estrogenic compounds differentially enhance the transcription of estrogen-regulated genes and suggest a role for EEIG1 in estrogen action.

Convergent transcriptional profiles induced by endogenous estrogen and distinct xenoestrogens in breast cancer cells

“…Estrogen-responsive transcripts involved in growth stimulation, transcription and cell adhesion are listed in Tables III and IV. In agreement with former DNA microarray studies (21)(22)(23)(24)(25)(26)(27)(28)31), the production of TFF1, TFF3 (trefoil factor 3), IGFBP4 (insulin-like growth factor binding protein 4), SDF1, STC2 (stanniocalcin 2), AREG, OLFM1 and OLFML3 (olfactomedin-like 3) was induced upon estrogen treatment. THBS1 (thrombospondin 1) is also upregulated in an estrogendependent manner in T47D cells.…”

“…In view of the steep increase of luciferase induction observed after 24 h estrogen exposures, this time point was used to compare in detail early transcriptional changes in response to ER stimuli. The 24 h period also corresponds to the time of maximal induction of most estrogen-regulated genes according to previous experiments with 17b-estradiol (21), although transcripts that may be subjected to upregulation exclusively during a very early phase of the estrogenic response would be under-represented (27).…”

“…With few exceptions (21,22) these previous studies were performed at high 17b-estradiol concentrations of 1 nM (23) or even 10 nM (24-31), thereby exceeding the peak effect level, observed around a concentration of 0.1 nM (22), by one or more orders of magnitude. Previous reports also described transcriptional patterns induced by phytoestrogens, including genistein, at concentrations of 10 mM (29) or 100 mM (27,28), again exceeding by far the saturation level, which for genistein is reached at a concentration of $1 mM (32). However, molecular insight into the effects of xenoestrogens in a lower subsaturating concentration would be important to assess health hazards or benefits at dose ranges that are more relevant for human exposure.…”

“…Other reports concluded that there are significant differences between the transcriptional effects of 17b-estradiol and xenoestrogens in the uteri of immature mice (34) or in the reproductive tract of adult rats (35). Several studies came to the conclusion that there is only limited overlap between the expression patterns elicited by 17b-estradiol and xenoestrogens in human breast cancer cells (27)(28)(29)31). These contrasting results led us to undertake a large-scale comparative analysis of early gene expression changes in human MCF7 and T47D breast cancer cells treated with equipotent concentrations of 17b-estradiol, genistein, bisphenol-A and the polychlorinated biphenyl congener 54 (PCB54).…”

Interactions between the estrogen receptor, its cofactors and microRNAs in breast cancer

“…It must first be metabolized in the liver by the cytochrome P450 isoform, CYP2D6, into the active metabolites [42]. These metabolites compete with estrogen for the ligand-binding domain (LBD) of the ER, blocking the potential for estrogen stimulation and preventing conformational changes in the receptor critical for the association of cofactors and the transcription of estrogen-responsive genes [43]. Variations in this isoform, whether genetic (e.g., wild-type variant CYP2D61*) or as a result of inhibition by selective serotonin reuptake inhibitors (e.g., fluoxetine and paroxetine) [44], may alter the metabolism of tamoxifen, blunting its antiestrogenic activity [45].…”