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

Estrogen receptor-alpha (ER alpha) 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 ER alpha-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 ER alpha protein levels, chromatin remodeling and mRNA output. We then focused on UBR5, a ubiquitin ligase and known oncogene that modulates ER alpha protein levels and transcriptional output. Finally, we demonstrated that UBR5 also affects endogenous ER alpha target genes and E2-mediated cell proliferation in breast cancer cells. In conclusion, our multi-end point RNAi screen identified novel modulators of ER alpha 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 ER alpha protein levels and transcriptional activity.