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Fig. 4 | Cell Communication and Signaling

Fig. 4

From: Flavokawain B targets protein neddylation for enhancing the anti-prostate cancer effect of Bortezomib via Skp2 degradation

Fig. 4

FKB degrades Skp2 protein in a proteasome and ubiquitination dependent manner. a, Western blotting analysis of Myc-Skp2 and endogenous Skp2 protein expression after PC3/Skp2 or PC3/PCDNA cells were treated with FKB, MG132 or their combination for 24 h. b, proteasome activities (i.e. OD values) were measured by the 20S Proteasome Assay Kit after PC3 cells were treated with 0.1% DMSO, FKB or MG132 for 24 h. c, Skp2 ubiquitination was measured by immunoprecipitation with anti-ubiquitin antibody and then Western blotting analysis of immunoprecipitates by anti-Skp2 after PC3 cells were treated with 0.1% DMSO or FKB (left two panels). 293 cells were transiently transfected with Skp2 and ubiquitin expression plasmids. After 24 h of transfection, cells were treated with 0.1% DMSO or FKB for 16 h. FKB increases ubiquitination of Skp2. d, the relative ubiquitination levels were measured by the Ubiquitylation Assay Kit after DU145 cells were treated by 0.1% DMSO, 8.8 μM FKB, 5 μM MG132 for 16 h

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