Fig. 5.

ARRDC3 regulates Hippo signaling independently of its function on PAR1 trafficking and degradation. (A) Illustration of ARRDC3 and ALIX requirement for facilitating lysosomal degradation and hypothesis of the effect of ARRDC3 on Hippo signaling. Illustration created with BioRender.com. (B–E) MDA-MB-231 HA–ARRDC3 pSLIK cells were transfected with the indicated siRNAs (NS, non-specific control siRNA) and treated with doxycycline (DOX) for ARRDC3 overexpression. Cells were then stimulated with 10 nM α-thrombin (α-Th) for the indicated times, and samples were immunoblotted (IB) using antibodies against the indicated proteins. Results are the fold-increase in JNK phosphorylation (C), CTGF expression (D) and ANKRD1 expression (E) relative to 0 min −DOX, NS-transfected control. Data are mean±s.d., n=3. Statistical significance was determined by two-way ANOVA with Tukey′s post hoc test. **P<0.01; ****P<0.0001.