Inhibition of vascular endothelial growth factor expression in HEC1A endometrial cancer cells through interactions of estrogen receptor α and Sp3 proteins

Matthew Stoner, Fan Wang, Mark Wormke, Thu Nguyen, Ismael Samudio, Carrie Vyhlidal, Dieter Marme, Gunter Finkenzeller, Stephen Safe

Research output: Contribution to journalArticle

76 Scopus citations

Abstract

Treatment of HEC1A endometrial cancer cells with 10 nM 17β-estradiol (E2) resulted in decreased vascular endothelial growth factor (VEGF) mRNA expression, and a similar response was observed using a construct, pVEGF1, containing a VEGF gene promoter insert from -2018 to +50. In HEC1A cells transiently transfected with pVEGF1 and a series of deletion plasmids, it was shown that E2-dependent down-regulation was dependent on wild-type estrogen receptor α (ERα) and reversed by the anti-estrogen ICI 182,780, and this response was not affected by progestins. Deletion analysis of the VEGF gene promoter identified an overlapping G/GC-rich site between -66 to -47 that was required for decreased transactivation by E2. Protein-DNA binding studies using electrophoretic mobility shift and DNA footprinting assays showed that both Sp1 and Sp3 proteins bound this region of the VEGF promoter. Coimmunoprecipitation and pull-down assays demonstrated that Sp3 and ERα proteins physically interact, and the interacting domains of both proteins are different from those previously observed for interactions between Sp1 and ERα proteins. Using a dominant negative form of Sp3 and transcriptional activation assays in Schneider SL-2 insect cells, it was confirmed that ERα-Sp3 interactions define a pathway for E2-mediated inhibition of gene expression, and this represents a new mechanism for decreased gene expression by E2.

Original languageEnglish (US)
Pages (from-to)22769-22779
Number of pages11
JournalJournal of Biological Chemistry
Volume275
Issue number30
DOIs
StatePublished - Jul 28 2000

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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