Interplay between Estrogen Response Element Sequence and Ligands Controls in Vivo Binding of Estrogen Receptor to Regulated Genes

Adam J. Krieg, Sacha A. Krieg, Bonnie S. Ahn, David J. Shapiro

Research output: Contribution to journalArticle

43 Scopus citations

Abstract

To examine the role of the estrogen response element (ERE) sequence in binding of liganded estrogen receptor (ER) to promoters, we analyzed in vivo interaction of liganded ER with the imperfect ERE in the p82 gene and the composite estrogen-responsive unit (ERU) in the proteinase inhibitor 9 (PI-9) gene. In transient transfections of ER-positive HepG2-ER7 cells, PI-9 was strongly induced by estrogen, moxestrol (MOX), and 4-hydroxytamoxifen (OHT). PI-9 was not induced by raloxifene or ICI 182,780. Quantitative reverse transcriptase-PCR showed that moxestrol strongly induced cellular PI-9 and pS2 mRNAs, whereas OHT moderately induced PI-9 mRNA and weakly induced pS2 mRNA. Chromatin immunoprecipitation experiments demonstrated strong and similar association of 17β-estradiol-hERα and MOX-hERα with the PI-9 ERU and with the pS2 ERE. Binding of MOX-hERα to the PI-9 ERU and the pS2 ERE was rapid and continuous. Although MOX-hERα bound strongly to the PI-9 ERU and less well to the pS2 ERE in chromatin immunoprecipitation, gel shift assays showed that estrogen-hERα binds with higher affinity to the deproteinized pS2 ERE than to the PI-9 ERU. Across a broad range of OHT concentrations, OHT-hERα associated strongly with the pS2 ERE and weakly with the PI-9 ERU. ICI-hERα bound poorly to the PI-9 ERU and effectively to the pS2 ERE. Raloxifene-hERα and MOX-hERα exhibited similar binding to the PI-9 ERU and the pS2 ERE. These studies demonstrate that ER ligand and ERE sequence work together to regulate in vivo binding of ER to estrogen-responsive promoters.

Original languageEnglish (US)
Pages (from-to)5025-5034
Number of pages10
JournalJournal of Biological Chemistry
Volume279
Issue number6
DOIs
StatePublished - Feb 6 2004

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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