Western blot
The tissue and cells were lysed in ice-cold lysis buffer (KeyGEN Biotech, Nanjing, China) followed by centrifugation at 4°C for 15 min. Cell lysates were separated by 8% SDS‒PAGE and then transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, Billerica, USA). After blocking with 5% nonfat milk, the membranes were incubated overnight at 4°C with specific primary antibodies against the following proteins: TP53BP2 (1:1000, Abcam, Cambridge, USA), LC3B (1:1000, Abcam, Cambridge, USA), p62 (1:1000, Abcam, Cambridge, USA), Beclin-1 (1:1000, Abcam, Cambridge, USA), Bcl-2 (1:1000, Abcam, Cambridge, USA), E2F1 (1:1000, Abcam, Cambridge, USA), DNMT1 (1:1000, Abcam, Cambridge, USA), and G9a (1:1000, Abcam, Cambridge, USA). After three washes, the membranes were incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies for 1 h, after which protein expression was detected via a chemiluminescence kit (KeyGEN, Nanjing, China). The optical density of each band was analyzed via densitometry and normalized to that of a β-actin loading control.