Targeting of RhoE inhibits epithelial-mesenchymal transition during colorectal cancer cell migration

Abstract

Abstract Objective Despite microRNA (miR-200b) being proved to promote the proliferation of colorectal cancer (CRC) cells, the relationship between miR-200b and epithelial-mesenchymal transition (EMT) of CRC cells remains poorly understood. The aim of the study was to investigate the relationship between miR-200b and EMT during CRC cell migration. Methods The effect of miR-200b on EMT-associated markers E-cadherin and vimentin was evaluated by western blot in CRC cells (SW620 and HT-29) by treatment with miR-200b mimics and inhibitors. A luciferase reporter assay was employed to detect downstream targets of miR-200b. Transwell migration assays were used to detect CRC cell migration. Results Western blots revealed that treatment with miR-200b mimics led to up-regulation of E-cadherin and down-regulation of vimentin, metalloproteinase (MMP)-9, and MMP-2, whereas treatment with miR-200b inhibitor exhibited opposite effects on expression of E-cadherin and vimentin. Luciferase reporter assays demonstrated that RhoE (RND3) was targeted by miR-200b. Two predicted target sites of miR-200b were present in the 3’-UTR of RhoE. Predicted target site 1 was from nucleotides 1584 to 1591, and site 2 was from nucleotides 1729 to 1735. RhoE knockdown cell lines were also established to investigate the impact of RhoE and miR-200b on EMT and cell migration. RhoE knockdown enhanced the effect of miR-200b mimics, up-regulating E-cadherin and down-regulating vimentin. RhoE knockdown also inhibited cell migration. Furthermore, miR-200b mimic treatment further promoted the inhibitory effect of RhoE knockdown on cell migration. Conclusion miR-200b inhibited EMT and CRC cell migration partly via inhibiting RhoE expression in CRC. RhoE and miR-200b might therefore be promising target genes in the management of CRC.