Fig. 5

circPVT1 blocks the binding of β-TrCP to c-Myc and inhibits the ubiquitination of c-Myc. A. The interaction between β-TrCP and c-Myc proteins was examined in CNE2 and 5-8F cells after overexpression or knockdown of circPVT1 by immunoprecipitation using anti-β-TrCP antibody. B. The interaction between WD40 repeat domain of β-TrCP and c-Myc proteins was examined in CNE2 and 5-8F cells after overexpression of circPVT1 by immunoprecipitation using anti-Flag antibody. C. The expression of c-Myc proteins was examined by western blotting in CNE2 and 5-8F cells after transfected with c-Myc siRNA. D. Binding of circPVT1 and WD40 repeat domain of β-TrCP protein was detected in CNE2 and 5-8F cells after knockdown of c-Myc by RNA pull-down assays with biotin-labeled circPVT1 probe. E. Degradation of c-Myc was detected in CNE2 and 5-8F cells after overexpression or knockdown of circPVT1 and treatment with 50 μg/mL cycloheximide (CHX). F. The ubiquitination level of c-Myc protein was determined in CNE2 and 5-8F cells after overexpression or knockdown of circPVT1, immunoprecipitation with anti-c-Myc antibody, and followed by western blotting with anti-ubiquitin antibody