Interaction proteomics identifies estrogen receptor beta association with multiple chromatin repressive complexes to inhibit cholesterol biosynthesis and exert an oncosuppressive role in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is characterized by poor response to therapy and low overall patient survival. Recently, Estrogen Receptor beta (ERbeta) has been found to be expressed in a fraction of TNBCs where, because of its oncosuppressive actions on the genome, it represents a potential therapeutic target, provided a better understanding of its actions in these tumors becomes available. To this end, the cell lines Hs 578T, MDA-MB-468 and HCC1806, representing the claudin-low, basal-like 1 and 2 TNBC molecular subtypes respectively, were engineered to express ERbeta under the control of a Tetracycline-inducible promoter and used to investigate the effects of this transcription factor on gene activity. The antiproliferative effects of ERbeta in these cells were confirmed by multiple functional approaches, including transcriptome profiling and global mapping of receptor binding sites in the genome, that revealed direct negative regulation by ERbeta of genes, encoding for key components of cellular pathways associated to TNBC aggressiveness representing novel therapeutic targets such as angiogenesis, invasion, metastasis and cholesterol biosynthesis. Supporting these results, interaction proteomics by immunoprecipitation coupled to nano LC-MS/MS mass spectrometry revealed ERbeta association with several potential nuclear protein partners, including key components of regulatory complexes known to control chromatin remodeling, transcriptional and post-transcriptional gene regulation and RNA splicing. Among these, ERbeta association with the Polycomb Repressor Complexes 1 and 2 (PRC1/2), known for their central role in gene regulation in cancer cells, was confirmed in all three TNBC subtypes investigated, suggesting its occurrence independently from the cellular context. These results demonstrate a significant impact of ERbeta in TNBC genome activity mediated by its cooperation with regulatory multiprotein chromatin remodeling complexes, providing novel ground to devise new strategies for the treatment of these diseases based on ligands affecting the activity of this nuclear receptor or some of its protein partners.