Effects of Phosphorylation on the DNA Binding Properties of Estrogen Receptor-alpha

Download or read book Effects of Phosphorylation on the DNA Binding Properties of Estrogen Receptor-alpha written by Kyle Thomas Helzer and published by . This book was released on 2019 with total page 254 pages. Available in PDF, EPUB and Kindle. Book excerpt: Estrogen receptor-[small alpha] (ER) is a major driver of breast cancer growth and development making it a common target for therapies. Understanding the activation and control of the ER signaling pathway is critical for developing new breast cancer treatments as well as combating resistance to current treatments. Activation of ER can occur in a ligand-dependent manner by 17[uppercase beta]-estradiol (E2) or in a ligand-independent manner through various growth factors or other stimuli. A common feature to both signaling pathways is the induction of ER phosphorylation at serine 118 (pS118-ER). Previous work has found that pS118-ER is an important modulator of ER-dependent gene transcription. A crucial step in the ER signaling pathway which allows for the altering of target gene expression is the binding of ER to DNA. While much work has been done to define the genome-wide binding profile (cistrome) of ER, the effect of phosphorylation on the distribution of ER across the genome is not well understood. In the work presented here, I aim to define the cistrome of pS118-ER and identify defining characteristics of sites occupied by the phosphorylated receptor compared to all ER occupancy sites. Using a comprehensive approach with multiple pS118-ER specific antibodies, I find that pS118-ER occupies a subset of ER sites in the human genome, with these sites being enriched for enhancer marks as well as the DNA-binding motif for the transcription factor grainyhead-like 2 (GRHL2). Additionally, analysis of these sites on in vitro DNA-binding arrays revealed an association between pS118-ER and direct DNA-binding. Finally, I developed a mutant MCF-7 cell line expressing ER with an S118A mutation to interrogate the biology of pS118-ER in a cell line which is dependent on ER. Consistent with previous reports, I found that E2-dependent ER downregulation was impaired in the S118A ER mutant. Interestingly, the downregulation of the ESR1 gene in response to E2 was also prevented indicating the control of pS118-ER over ER levels is not limited to post-translational mechanisms. These studies further the knowledge of how phosphorylation affects ER biology and may serve as a target for future breast cancer therapies.