Fig. 6: CK2α-mediated phosphorylation of DUB3 promotes YAP1 stability and oncogenic function.

A Control (Ctrl) or DUB3-depleted A2780 cells were subjected to DMSO or Silmitasertib for 24 h. Protein levels of YAP1 and DUB3 were examined by immunoblotting. B Vector (V) or Flag-DUB3 was transfected into A2780 cells and then subjected to DMSO or Silmitasertib for 24 h. The protein level of YAP1 was examined by immunoblotting. C Indicated constructs were transfected into cells followed by the treatment with DMSO or Silmitasertib for 24 h. MG-132 (10 μM) was then administered for 10 h. HA-protein agarose was used to immunoprecipitate YAP1, and immunoblotting was performed to measure the polyubiquitylated YAP1. D Vector (V) or Flag-DUB3 WT was transfected into cells and then treated with DMSO or Silmitasertib. MG-132 (10 μM) was then administered for 10 h. Anti-Flag affinity gel was used to immunoprecipitate Flag-DUB3 and the interaction between DUB3 and YAP1 was measured by immunoblotting. E Indicated constructs were transfected into Control (Ctrl) or DUB3-depleted A2780 cells. Cells were subjected to the treatment with DMSO or Silmitasertib. The protein level of YAP1 was examined by immunoblotting. F Indicated constructs were transfected into cells, and MG-132 (10 μM) was then administered for 10 h. HA-protein agarose was used to immunoprecipitate YAP1, and immunoblotting was performed to measure the polyubiquitylated YAP1. G Indicated constructs were transfected into cells. Anti-Flag affinity gel was used to immunoprecipitate Flag-DUB3, and the interaction of DUB3-YAP1 was measured by immunoblotting. H Indicated constructs were co-transfected into cells followed by treatment of MG-132 (10 μM) for 10 h. Anti-Flag affinity gel was used to immunoprecipitate YAP1, and immunoblotting was used to measure the ubiquitination of YAP1. I Indicated constructs were transfected into cells. Anti-Flag affinity gel was used to immunoprecipitate Flag-DUB3, and the interaction of DUB3-YAP1 was measured by immunoblotting. J Indicated constructed were transfected into DUB3-depleted A2780 cells and then subjected to the treatment with Silmitasertib. The protein level of YAP1 was examined by immunoblotting. Cell proliferation (K) and survival at indicated concentrations of Cisplatin were measured (L).