Transcriptional Activation of Thymidylate Synthase by 17β-Estradiol in MCF-7 Human Breast Cancer Cells1

Thymidylate synthase (TS) catalyzes methylation of deoxyuridine phosphate to give deoxythymidine phosphate, and 17β-estradiol (E2) induces TS gene expression in MCF-7 human breast cancer cells. Analysis of the TS gene promoter showed that E2-responsiveness required the −229 to −140 promoter region containing a G-rich sequence and CACCC box. Subsequent mutational analysis of this region indicated that only the G-rich motif (−150 to −142) was required for E2 action. Results of gel mobility shift and in vitro DNA footprinting assays showed that both estrogen receptor α (ERα) and Sp1 proteins were required for hormone-induced trans-activation that involved ERα/Sp1 binding to the G-rich site in which only Sp1 protein bound DNA. Both proteins also interacted in Drosophila cells in functional assays, confirming the transcriptional activation of TS-involved ERα/Sp1, and this adds to the increasing number of genes that are activated through this pathway in breast cancer cells.