Fig. 4

NUCKS silencing induces autophagy and apoptosis in gastric cancer cells. a Cleaved Caspase 3 and Cleaved PARP were detected by Western blot analysis after the downregulation of NUCKS. The drug-resistant cells were selected and continued to culture for 3, 6 and 9 days for Western blot analysis. b Detecting the change of gastric cancer cells morphology by light microscopy. c Cells transfected with the GFP-LC3B plasmid after NUCKS knockdown and restoration were examined using fluorescence microscopy (Scale bars, 10 μm). The quantification of LC3B-positive puncta is presented as a histogram (*P < 0.05, **P < 0.01, ***P < 0.001). d Immunofluorescence staining with a LC3B antibody was performed to confirm the induction of autophagy after NUCKS downregulation and upregulation. Representative LC3B-positive cells are shown. e The level of autophagy was evaluated by p62 and LC3B expression, as determined using Western blot analysis after NUCKS knockdown and restoration. f CQ blocked shNUCKS-induced autophagy. Cells were pretreated with 50 μM CQ and analyzed at indicated time points by Western blot