Molecular and Biochemical Studies of Several Novel Estrogen Receptor Alpha-interacting Proteins in Breast Cancer Cells

Download or read book Molecular and Biochemical Studies of Several Novel Estrogen Receptor Alpha-interacting Proteins in Breast Cancer Cells written by Ahmed Edan Dhamad and published by . This book was released on 2017 with total page 402 pages. Available in PDF, EPUB and Kindle. Book excerpt: Breast cancer is the second leading cause of cancer-related death in women, and approximately 70% of incidences are estrogen receptor (ER)-positive breast cancer. ERÜ and its interacting proteins play a key role in the development and progression of breast cancer. However, how ERÜ regulates its target gene expression and hence cell proliferation is not fully understood. To enhance our understanding of the molecular mechanism by which ERÜ regulates gene expression, we used a quantitative proteomic method to identify cellular proteins that interact with ERÜ. The first group of proteins that were identified to associate with ERÜ are heat shock proteins (Hsps). We identified 21 Hsps and 3 Hsp cochaperones that were associated with ERÜ. Co-immunoprecipitation assay demonstrated that Hsp70-1 and Hsc70, the two most abundant ERÜ-associated proteins, interacted with ERÜ in both transcriptionally active and inactive chromatin of MCF7 cells. A novel of protein that was identified to interact with ERÜ is histone acetyltransferase 1 (HAT1). We showed that HAT1 physically binds ERÜ through the E domain of ERÜ, and silencing HAT1 by shRNA significantly increased the ERÜ-mediated transcription in MCF7 cells. Importantly, our data suggest that HAT1 regulates ERÜ transcriptional activity through affecting the interactions of ERÜ with histone proteins around the promoter region of ERÜ target genes in breast cancer cells. We also identified and confirmed that protein arginine methyltransferase 5 (PRMT5) is a new ERÜ interacting partner, and PRMT5 interacts with ERÜ preferentially in the cytoplasm of MCF7 cells. Functionally, we found that overexpression of PRMT5 in MCF7 cells significantly decreased ERÜ transcriptional activity. Finally, we demonstrated that chromatin target of PRMT1 (CHTOP) directly binds to ERÜ through the E domain of ERÜ. We found that knockout of CHTOP by CRISPR-Cas9 significantly decreased ERÜ transcriptional activity, and the effect is potentially through decreasing protein levels of MEP50, an ERÜ coactivator. In summary, we identified and characterized several novel ERÜ-interacting proteins that play significant roles in regulating ERÜ transcriptional activities. Our results provide new insight into the molecular mechanisms by which ERÜ controls its target gene expression and regulates cell proliferation in ERÜ-positive cells.