670 nm light inhibits AMPK-mediated YAP phosphorylation and the Hippo pathway. (A) MG were treated with or without 670 nm light and then immunostained 24 h after light treatment with anti-YAP and anti-SOX9 antibodies. Hoechst dye served as a counterstain. Percentages of YAP/SOX9 double-positive cells is shown in the right panel. Scale bars, 100 μm. (B) Western blot analysis for expression of phospho-LATS1, LATS1, phospho-YAP, and YAP from MG cultures described in A. (C) Quantitative analysis for the ratio of phospho-YAP to total YAP and of phospho-LATS1 to LATS1 from B, respectively. (D) Western blot analysis of phospho-AMPKα and AMPKα expression from MG cultures described in A. Quantitative analysis are shown in right panel. (E) MG exposed to AICAR or DMSO were treated with or without 670 nm light and subjected to western blotting analysis 24 h after light treatment. (F) Relative LATS1 and YAP phosphorylation levels were quantified by the ratio of phosph-LATS1 to total LATS1 and of phospho-YAP to total YAP from E. For quantification, levels are given as mean ± SD (n = 3 independent samples for each group). Student’s t test (*P < 0.05, **P < 0.01).
