Systems level-based RNAi screening by high content analysis identifies UBR5 as a regulator of estrogen receptor-α protein levels and activity

Oncogene. 2015 Jan 8;34(2):154-64. doi: 10.1038/onc.2013.550. Epub 2014 Jan 20.

Abstract

Estrogen receptor-α (ERα) is a central transcription factor that regulates mammary gland physiology and a key driver in breast cancer. In the present study, we aimed to identify novel modulators of ERα-mediated transcriptional regulation via a custom-built siRNA library screen. This screen was directed against a variety of coregulators, transcription modifiers, signaling molecules and DNA damage response proteins. By utilizing a microscopy-based, multi-end point, estrogen responsive biosensor cell line platform, the primary screen identified a wide range of factors that altered ERα protein levels, chromatin remodeling and mRNA output. We then focused on UBR5, a ubiquitin ligase and known oncogene that modulates ERα protein levels and transcriptional output. Finally, we demonstrated that UBR5 also affects endogenous ERα target genes and E2-mediated cell proliferation in breast cancer cells. In conclusion, our multi-end point RNAi screen identified novel modulators of ERα levels and activity, and provided a robust systems level view of factors involved in mechanisms of nuclear receptor action and pathophysiology. Utilizing a high throughput RNAi screening approach we identified UBR5, a protein commonly amplified in breast cancer, as a novel regulator of ERα protein levels and transcriptional activity.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Breast Neoplasms / enzymology
  • Breast Neoplasms / genetics
  • Breast Neoplasms / metabolism*
  • Estrogen Receptor alpha / genetics
  • Estrogen Receptor alpha / metabolism*
  • Female
  • Gene Expression Regulation, Neoplastic
  • Gene Knockdown Techniques
  • HeLa Cells
  • Humans
  • MCF-7 Cells
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • RNA Interference
  • Signal Transduction
  • Transcriptional Elongation Factors / genetics
  • Transcriptional Elongation Factors / metabolism
  • Ubiquitin-Protein Ligases / genetics
  • Ubiquitin-Protein Ligases / metabolism*

Substances

  • Estrogen Receptor alpha
  • Nuclear Proteins
  • SUPT5H protein, human
  • Transcriptional Elongation Factors
  • estrogen receptor alpha, human
  • UBR5 protein, human
  • Ubiquitin-Protein Ligases