Fig. 6

FAM83H binds to β-catenin and protects it from degradation to induce the expression of the effector genes of Wnt/β-catenin signaling. A RT-qPCR results showing that the β-catenin mRNA level was not altered by FAM83H knockdown in either PANC-1 or SW 1990 cells. B Western blot showing that the β-catenin protein level was synchronously changed upon FAM83H overexpression and knockdown. C FAM83H binds to exogenously expressed (left panel) or endogenous β-catenin (right panel). HEK293T cells were transfected with the indicated plasmids, followed by IP with an anti-FLAG antibody and IB with the indicated antibodies. WCL: whole-cell lysate. D FAM83H affected β-catenin protein stability, which was assessed by determining the effect of FAM83H overexpression on the β-catenin degradation rate after adding cycloheximide (100 μg/mL). E Western blot results showing that the expression of β-catenin in the FAM83H-downregulated PDAC cells treated with the proteasome inhibitor MG132 (10 μM) for 6 h was recovered compared with that in the FAM83H-downregulated PDAC cells. F Western blot results showing the change in the ubiquitination of β-catenin. HEK293T cells were cotransfected with HA-β-catenin, sh-FAM83H#1/#2 and MYC-Ub and then treated with 10 μM MG132 for 6 h. Polyubiquitinated proteins were immunoprecipitated with anti-HA beads, followed by immunoblotting with antibodies against MYC and HA. G The expression of the effector gene Wnt/β-catenin signaling upon FAM83H knockdown was determined by RT-qPCR; *P < 0.05, **P < 0.01 and ***P < 0.001