Systems level-based RNAi screening by high content analysis identifies UBR5 as a regulator of estrogen receptor-α protein levels and activity.
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.
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
M J
MJ
Bolt
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
F
F
Stossi
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
A M
AM
Callison
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
M G
MG
Mancini
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
R
R
Dandekar
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, USA.
M A
MA
Mancini
D001943:Breast Neoplasms / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D047628:Estrogen Receptor alpha / Q000235:genetics / Q000378:metabolism*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D061986:MCF-7 Cells; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D015398:Signal Transduction; D035602:Transcriptional Elongation Factors / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*
C506487:ESR1 protein, human; D047628:Estrogen Receptor alpha; D009687:Nuclear Proteins; C112224:SUPT5H protein, human; D035602:Transcriptional Elongation Factors; C117617:UBR5 protein, human; D044767:Ubiquitin-Protein Ligases