FIG. 2.

Transcriptional activation of endogenous genes by and functional specificity of EZH2. (A) EZH2 enhanced estrogen-stimulated mRNA expression of c-Myc and cyclin D1. MCF-7 cells were transfected with an empty vector or with an EZH2 expression plasmid. Sixty hours after transfection, the cells were switched to estrogen-deprived medium for 48 h. The cells were then left untreated or treated with 100 nM E2 for different times, as indicated, before the cells were collected for RNA extraction and for gene expression analysis by real-time RT-PCR. (B) EZH1 did not affect the transactivation of c-Myc promoters. MCF-7 cells were grown in the absence of estrogen and were cotransfected with c-Myc-Luc, a Renilla construct, along with different amounts of an EZH1 expression construct. Eighteen hours after transfection, cells were treated with E2 or ethanol (vehicle) for different times and harvested for a luciferase activity assay. Each bar represents the mean ± standard deviation (SD) for triplicate experiments. (C) EZH2 did not affect androgen receptor-regulated gene transcription. LnCAP cells were transfected with a PSA-luc (prostate-specific antigen-luciferase) construct together with 100 ng or 500 ng of SRC-1 or 100 ng or 500 ng of EZH2 expression constructs as indicated. After growing the cells in the absence of steroids for 48 h, cells were treated with 100 nM dihydrotestosterone (DHT) for 12 h, and the reporter activity was measured. Each bar represents the mean ± SD for triplicate experiments.