Figure 5. Autophagic flux was not required for G₁ phase arrest in ATG4B-silenced cells. (A) Scrambled (-) or ATG4B (+) siRNA (5 nM) was introduced into autophagy-deficient (shATG5 or shATG7) human colorectal cancer HCT116 cells for 66 h. The protein levels of SQSTM1, DVL2, and CCND1 were determined by immunoblotting, and the knockdown efficiency of ATG4B, ATG5, or ATG7 was also examined. (B) The proportion of cells in G₁ phase was analyzed by flow cytometry. (C) Scrambled (-) or ATG4B (+) siRNA was transfected into human colorectal cancer HCT116 cells for 56 h, followed by treatment with CQ (10 μM) or MG132 (1μM) for 16 h prior to harvesting. The protein levels of SQSTM1, DVL2, and CCND1 were determined by immunoblotting. (D) The proportion of the cells as (C) in G₁ phase was analyzed by flow cytometry. The image of the immunoblotting results was quantitated using ACTB as the normalization control. The results are expressed as the mean ± SEM from 3 individual experiments.