Figure 3.
14‐3‐3ζ induces autophagy in HCC‐LM3 and CSQT‐2 cells. (A) HCC‐LM3 and (C) CSQT‐2 cells were infected with n.c. oe‐LV or 14‐3‐3ζ oe‐LV for 72 h, and the mRNA levels of YWHAZ gene were determined with qRT‐PCR and normalized to β‐actin. For knockdown detection, cells were first infected with n.c. sh‐LV or 14‐3‐3ζ sh‐LV for 72 h and then incubated under hypoxia for additional 12 h. The expression mRNA levels of YWHAZ gene in (B) HCC‐LM3 and (D) CSQT‐2 cells were determined with qRT‐PCR and normalized to β‐actin. (E‐H) Western blot was performed to determine the protein levels of 14‐3‐3ζ, p62, beclin 1 and phospho‐beclin 1S295. (F,H) Co‐IP with anti‐beclin 1 for IP, and anti‐beclin 1 and anti‐VPS34 for IB was performed. (I‐L) Immunofluorescence was performed to analyse the formation of LC3 puncta. Data were shown in mean with SD (N = 3). A *P value <.05 was considered significant. nc, negative control; LV, lentivirus; oe, overexpression; shRNA, short hairpin RNA