MicroRNA-145, a p70 S6 kinase-activated microRNA, regulates N-cadherin expression and epithelial-mesenchymal transition

Abstract

BACKGROUND: Ovarian cancer is highly metastatic with a poor prognosis (5-year survival <25%), and understanding the underlying mechanisms is of obvious importance. Epithelial–mesenchymal transition (EMT) has been identified as of potential importance in ovarian cancer metastasis. N-cadherin is critically involved in the EMT. Yet, very little is known about the factors regulating N-cadherin expression. Recently, we demonstrated that p70 S6 kinase (p70S6K), a downstream effector of phosphatidylinositol 3-kinase/Akt, which is frequently activated in ovarian carcinoma, plays a key role as intracellular signaling mediator for the effects of multiple growth factors in the tumor microenvironment. However, the molecular details of how p70S6Kcontrols the malignant properties are still poor understood. MATERIAL AND METHODS: Expression of Twist, SOX9, and N-cadherin expression was measured by reverse transcription-PCR. The functional effects of p70S6K were examined further using overexpression and knockdown approaches. The putative microRNA (miRNA) was investigated by in silico prediction. A luciferase reporter assay was conducted to confirm miRNA-target interaction. RESULTS: Here we show that p70S6Kis a pivotal regulator of N-cadherin, which was mediated through activation of the transcription factors Twist and Sox9. Interestingly, we found that upregulation of Twist and Sox9 was due to increased mRNA stabilities. Using multiple in silico algorithms, we showed that miR-145 may play a critical role in regulating Twist and Sox9. We demonstrated direct targeting of miR-145 to the 3’-untranslated regions of Twist and Sox9. Notably, overexpression of miR-145 significantly suppressed, whereas inhibition of miR-145 enhanced N-cadherin expression and EMT, suggesting that inappropriate expression of miR-145 contributes to the metastatic phenotype. On exploring how p70S6K might regulate miR-145, we found that p70S6K could regulate miRNA biogenesis. CONCLUSIONS: These results illustrate a novel regulatory role of p70S6Kdependent miR-145-Twist/Sox9 signaling axis in the regulation of N-cadherin and EMT properties in ovarian cancer and could have important clinical implications.