Fig. 7

circPDK1 acts as a scaffold to enhance the binding of BIN1 proteins with UBE2O. A RNA pull-down verified the interaction of circPDK1 with BIN1. β-actin was used as a negative control. RIP assay was used to determine the relative expression of circPDK1 with rabbit UBE2O and IgG antibodies in MIA PaCa-2 cells. B Co-IP assays demonstrated the binding of UBE2O with BIN1 in MIA PaCa-2 cells. C Co-localization of circPDK1 (red), BIN1 proteins (green), and UBE2O proteins (orange) in MIA PaCa cells. Scale bar = 5 μm. D UBE2O and BIN1 protein levels were detected after knockdown of UBE2O. E UBE2O and BIN1 protein levels were detected with indicated treatments. F Ubiquitination modification levels of BIN1 in MIA PaCa-2 and PANC-1 cells with UBE2O overexpression and loss of circPDK1. G The interaction between UBE2O and BIN1 proteins were verified by Co-IP assays in MIA PaCa-2 cells. RNase A (10 μg/ml) and RNase R (100 U/ml). H The expression of UBE2O in PC tumor tissues and matched normal tissues verified by IHC assays in tissue microarrays. I The correction of UBE2O and BIN1 protein level was verified by IHC assays in tissue microarrays. Scale bar = 1000 μm. J The luciferase activity of c-myc responsive transcriptional reporter was evaluated with indicated treatments. *P < 0.05; **P < 0.01; ***P < 0.001; ns, no significance