Estrogen Receptor Ligands and Regulation of Gene Expression

Abstract

93 p.Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2007.Keratin13 (KRT13) and calcitonin receptor (CALCR) gene expression is differentially regulated by estradiol (E2) and the selective estrogen receptor modulators (SERMs) tamoxifen and raloxifene. KRT13 is a cytoskeleton protein while CALCR mediates the biological effects of calcitonin. These two genes may play important roles in breast cancer growth and metastasis. Analysis of the promoters and regulatory regions of these two genes was performed to explore the mechanism of estradiol and SERM regulation of these genes in breast cancer cells. Using a ChIP scanning approach, we identified a 2.5 kb regulatory region for KRT13 and two enhancer regions for CALCR. The regulatory activities of these regions were confirmed by reporter transactivation assays. The 2.5 kb regulatory region of KRT13 was further analyzed using progressive deletions and point mutations. The results demonstrated that three estrogen responsive elements (EREs) and three Sp1 sites were involved in the ligand dependent up-regulation of KRT13. The ERE at site1 is mainly involved in E2 activity, whereas the EREs at site2, site3 and the three Sp1 sites are involved in both E2 and tamoxifen activities. Next, ChIP assays were performed to examine the time courses of E2 and SERM-induced recruitment of ERalpha and cofactors on KRT13 and CALCR regulatory regions. I found that the differential recruitment of factors to the regulatory regions accompanied the time-dependent differential regulation of these genes. Taken together, my results suggest that: (1) the ligand-differential regulations of KRT13/CALCR are due to ligand-differential recruitment of ER and coactivators; (2) EREs mediate both E2 and tamoxifen effects, while certain EREs preferentially mediate the E2 effect, presumably dependent on the ERE sequence and promoter context. These findings provide new insight about gene regulation by different estrogen receptor ligands and mediation via estrogen receptors.U of I OnlyRestricted to the U of I community idenfinitely during batch ingest of legacy ETD