Fig. 5: In hepatocellular carcinoma cells, USP1 interacts with TAZ and modulates its stability.

A Immunofluorescence imaging of TAZ (green), USP1 (red) and DAPI (blue) in HLF cells, scale bar 40 µm. B Western blot analysis of USP1 and TAZ protein localization in HLF cells. In this study, the cytoplasmic and nuclear fractions were separated using a subcellular protein fractionation kit (Thermo Scientific 78840). The control cytoplasmic and nuclear fractions were analyzed using antibodies specific for tubulin and histone3, respectively. C USP1 and TAZ proteins were found to be associated in HLF cells by a coimmunoprecipitation (Co-IP) assay. D The wild-type and truncated TAZ constructs are shown in a schematic diagram. E An immunoblot demonstrating the interaction between USP1 and WT TAZ or truncated TAZ, as assessed by Co-IP with USP1 (with an anti-Flag antibody). F, I Western blotting were used to measure TAZ and USP1 protein levels in HLF and HEK293 cells treated as indicated. MG132 (10 μM) was applied to the cells for 8 h before western blot analysis was performed. G, H USP1 depletion decreased the TAZ protein half-life in HLF cells. Cells were treated with 100 μmol/L CHX for the indicated time periods before being collected for western blot analysis. H Quantitative analysis of the half-life of the TAZ protein. J, K USP1 prolonged the TAZ protein half-life in HEK293 cells. Cells were treated with 100 μmol/L CHX for the indicated time periods before being collected for western blot analysis. K Quantitative analysis of the half-life of the TAZ protein. All the results are representative of 3 independent experiments. The data are presented as the means ± SDs. **P < 0.01 (Student’s t test).