Estrogen exhibits mitogenic activity in early ovarian carcinogenesis and plays an important role in ovarian tumorigenesis. Due to the increased expression of ERα and decreased expression of the ERβ, the ratio of ERα and ERβ is markedly increased in ovarian cancer. We have recently reported that PES1 regulates the balance of ERα and ERβ at the post-transcriptional level in breast cancer. Here, we report that PES1 inversely regulates the expression of ERα and ERβ in addition to their transcriptional activities in epithelial ovarian cancer. We found that the ablation of PES1 resulted in the significant downregulation of ERα and estrogen-responsive genes such as cylin D1, HIF-1α and VEGF and the up-regulation of ERβ and p21WAF1. Cell proliferation in both tested ovarian cell lines was markedly inhibited and cells were arrested in G2 after PES1 was ablated. Further analysis of clinical samples showed that expression of PES1 correlated positively with ERα expression and negatively with ERβ expression. Our results demonstrate that PES1 may play important role in the progression of ovarian cancer by inversely regulating the ERα and ERβ expression. PES1 may be a new target for ovarian cancer therapy. © 2013 IUBMB Life, 65(12):1017–1025, 2013.