Fig. 5: ACTL6A stabilizes YAP proteins.

a qRT-PCR analysis of YAP in A172- and U251-NC, -sh-ACTL6A, and U87MG-NC and ACTL6A cells. Expression is normalized to GAPDH mRNA. Data are represented as the mean ± SEM. b Western blot analysis to evaluate YAP levels in A172- and U251-NC, and -sh-ACTL6A cells after MG132 (20 µM) treatment for 8 h. GAPDH was used as loading control. c, d, e Western blot analysis of YAP protein in modified A172, U251 and U87MG cells treated with CHX (25 µg/mL) for the indicated time. Line graph shows YAP levels normalized to GAPDH at the indicated time points (n = 4). Data are represented as the mean ± SEM. f Western blot analysis of YAP IPs performed on lysates prepared from HEK293 cells transfected with HA-ubiquitin and plus or minus Flag-ACTL6A and treated with MG132 (20 µM) for 8 h. g, h Western blot analysis of YAP IPs performed on lysates prepared from U251-NC, -sh-ACTL6A, and U87MG-NC and -ACTL6A cells treated with MG132 (20 µM) for 8 h to examine endogenous YAP ubiquitination. Student’s t-test: n.s. = not significant, *P < 0.05, **P < 0.01