Fig. 2

Interaction between HDAC5 and p53 induced by RT and activation of HDAC5 mediated p53-dependent exosome secretion pathway in HCC. RT-induced expression of HDAC5, p53, P21, and PUMA in p53^−/− and p53^+/+ HepG2 was confirmed by western blot and quantified using b-actin as an internal control (A). Data are shown as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001 vs. untreated by Student’s t-test. The interaction between HDAC5 and p53 after RT was confirmed using Duolink proximity ligation assay in HepG2 (B). p53^+/+ HepG2 cells were treated with 4 Gy radiation and incubated for 12 h, and were then immunoprecipitated with anti-p53 antibodies. Precipitated proteins were subjected to co-immunoprecipitation (CO-IP) using an anti-HDAC5 antibody and analyzed by immunoblotting (C). The expression of TSAP6 induced by RT in p53^−/− and p53^+/+ HepG2 was verified by western blot. TSAP6 expression was quantified using b-actin as an internal control (D). Data are shown as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001 vs. untreated by Student’s t-test. Twenty-four hours after siHDAC5 was transfected into HepG2, exposed to 4 Gy radiation. After 12 h incubation, HDAC5, p53, and TSAP6 expressions were confirmed by western blot and quantified using b-actin as an internal control (E). Data are shown as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001 vs. untreated by Student’s t-test; #P < 0.05, ##P < 0.01, and ###P < 0.001 vs. 4 Gy treatment by Student’s t-test