Fig. 9.

Validation of the interaction between circ-PTK2 and miR-639. A Reverse transcriptase-quantitative PCR validation of level of miR-639 following inhibition of circ-PTK2 expression. B Binding sequences between circ-PTK2 and miR-639 were predicted and wild-type and mutated circ-PTK2 genes were synthesized and cloned into luciferase vector to generate reporter. C Luciferase activity was determined to validate the interaction between circ-PTK2 and miR-639. D Transwell assay with SK-OV-3 cells showing enhanced cell migration and invasion due to overexpression of circ-PTK2, whereas overexpression of miR-639 inhibited those processes. E Transwell assay with OVCAR-3 cells showing enhanced cell migration and invasion due to overexpression of circ-PTK2, whereas overexpression of miR-639 inhibited those processes. F Western blot analysis of effect of overexpression of circ-PTK2 in SK-OV-3 cells: the protein level of E-cadherin was decreased, while the protein levels of N-cadherin, Snail, and vimentin were increased, although overexpression of miR-639 reversed the effect. G Western blot analysis of effect of overexpression of circ-PTK2 in OVCAR-3 cells: the protein level of E-cadherin was decreased, while the protein levels of N-cadherin, Snail, and vimentin were increased, although overexpression of miR-639 reversed the effect. H Overexpression of circ-PTK2 in SK-OV-3 cells decreased the mRNA level of E-cadherin and increased the mRNA level of vimentin and Snail, while overexpression of miR-639 reversed the effect; adjustment of expression of circ-PTK2 and miR-639 did not significantly affect expression of N-cadherin. I Overexpression of circ-PTK2 in OVCAR-3 cells decreased the mRNA level of E-cadherin and increased the mRNA level of N-cadherin and Snail, while that of miR-639 reversed the effect; adjustment of expression of circ-PTK2 and miR-639 did not significantly affect expression of vimentin. Data are presented as the mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001