Estrogen Receptor α, Fos-Related Antigen-2, and c-Jun Coordinately Regulate Human UDP Glucuronosyltransferase 2B15 and 2B17 Expression in Response to 17β-Estradiol in MCF-7 Cells

Abstract

UDP-glucuronosyltransferase 2B15 and 2B17 expression is up-regulated by 17β-estradiol in MCF-7 breast cancer cells, as assessed by quantitative real-time polymerase chain reaction. Using 5′-deletion mapping and site-directed mutagenesis, we demonstrate that 17β-estradiol activation of UGT2B15 gene transcription is mediated by a 282-base pair fragment positioned -454 to -172 nucleotides from the translation start site. This region contains two putative activator protein-1 (AP-1) elements, one imperfect estrogen response element (ERE), and two consensus ERE half-sites. We propose that these five sites act as an estrogen response unit (ERU), because mutation in any site reduces activation of the UGT2B15 promoter by 17β-estradiol. Despite the presence of two AP-1 elements, the UGT2B15 promoter is not responsive to the AP-1 activator phorbol 12-myristate 13-acetate. Although electrophoretic mobility shift assays (EMSA) indicate that the AP-1 proteins c-Jun and Fos-related antigen 2 (Fra-2) bound to the distal AP-1 site, binding of Jun or Fos family members to the proximal AP-1 site was not detected by EMSA. Chromatin immunoprecipitation assays showed a 17β-estradiol-induced recruitment of estrogen receptor (ER) α, c-Jun, and Fra-2 to the 282-bp ERU. The involvement of these three transcription factors in the stimulation of UGT2B15 gene expression by 17β-estradiol was confirmed by siRNA silencing experiments. Mutagenesis and siRNA experiments indicate that UGT2B17 expression is also regulated by 17β-estradiol via the ERU, which is fully conserved in both promoters. Because UGT2B15 and UGT2B17 inactivate steroid hormones by glucuronidation, the regulation of their genes by 17β-estradiol may maintain steroid hormone homeostasis and prevent excessive estrogen signaling activity.