Fig. 5

WWP2 orchestrates the Hippo-YAP1 pathway by ubiquitinating LATS1 protein. A, B Western blotting assays were performed to detect the protein expression of LATS1/2, YAP1, p-YAP and its downstream target genes CYR61 and CTGF in GC cells upon corresponding treatments as presented. C, D qRT‒PCR analysis of the mRNA expression of WWP2, LATS1, YAP1, CTGF, CYR61 and AREG in GC cells upon WWP2 upregulation or knockdown. E Western blotting analysis of LATS1 protein in modified SGC-7901 cells upon WWP2 silencing and treatment with CHX (25 μg/mL) for specific time points. F The abundance of LATS1 protein was quantified and is shown in line graphs. G, H Western blotting analysis of in vivo ubiquitination. G BGC-823 cells were co-transfected with WWP2 shRNA or scramble shRNA and treated with MG132 for 6 h before harvesting. H SGC-7901 cells were transfected with combinations of plasmids encoding HA-WWP2, Flag-LATS1 and His-Ub, and the cells were treated with MG132 for 6 h before being harvested for further study. Student’s t test: *p < 0.05, **p < 0.01