Fig. 7

PPP1R26 promotes EMT in HCC cells by forming the PPP1R26-pSer37 PKM2-TGIF2 complex. A Huh7 cells were transfected with Flag or Flag-PPP1R26 and cultured under serum starvation. Immunoprecipitation was performed with an anti-Flag antibody on the cellular fractionation. The Flag-PPP1R26 and p-Ser37-PKM2 in the precipitates were determined by Western blot. B Huh7 cells were transfected with Flag or Flag-PPP1R26 and cultured under serum starvation. Immunoprecipitation was performed with an anti-Flag antibody. The Flag-PPP1R26, p-Ser37-PKM2 and TGIF2 in the precipitates were determined by Western blot (left). Schematic diagram showing the positions of primers designed to cover the promoter region of the CDH1 gene (right, upper). Huh7 cells were transfected with Flag or Flag-PPP1R26. ChIP assays were performed with IgG or anti-TGIF2 antibody followed by qPCR to amplify the CDH1 promoter region (right, lower). C Huh7 cells was transfected with Flag or Flag-PPP1R26. ChIP assays were performed with IgG and anti-acetylated H3K9 antibody followed by qPCR as described in B. D Huh7-PPP1R26-shRNA cells were subcutaneously implanted into nude mice. Tumors were dissected at the end of the experiment (left). Tumor weights were evaluated and shown in the middle. Tumor volumes were measured every 5 days during the experiments as shown in the right. E Proteins extracted from the xenografts were subjected to Western blotting probed with anti-PPP1R26, anti-PKM2 and anti-E-cadherin antibodies. Beta-actin was used as a loading control. F IHC was performed with anti-PPP1R26, anti-PKM2 and anti-E-cadherin antibodies on the paraffin-embedded tissue sections of xenografts (right). Data information: In (A-D), data are presented as mean ± SD. Statistical significance was assessed using two-tailed t-tests (B & C), one-way ANOVA with post hoc analysis LSD test or Kruskal-Wallis test (D). *P < 0.05, **P < 0.01 and ****P < 0.0001