Abstract
We examined how clomiphene citrate (CC) reduces estrogen receptor-α (ERα) in a human endometrial cancer cell line. Ishikawa human endometrial cancer cells were treated with ERα ligands such as 17β-estradiol (E2), CC, and the pure antiestrogen, ICI 182,780 (ICI). Thereafter, the expression levels of ERα protein and mRNA were analyzed by western blot and real-time quantitative PCR, respectively, and those of ubiquitinated ERα were analyzed by immunoprecipitation of ERα followed by immunoblotting with an anti-ubiquitin antibody. The expression levels of ERα protein after treatment with E2, CC, and ICI were significantly decreased compared to pre-treatment levels without a corresponding decrease in ERα mRNA. These ligands significantly increased the levels of ubiquitinated ERα compared to vehicle treatment. Co-treatment with the proteasome inhibitor, MG132, abrogated the decrease in ERα levels caused by treatment with the ligands only. We demonstrated, for the first time, a CC-induced decrease in ERα mediated by the ubiquitin-proteasome pathway in human endometrial cancer cells.
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