14-3-3ζ binds to and stabilizes phospho-beclin 1 S295 and induces autophagy in hepatocellular carcinoma cells.

Data from The Cancer Genome Atlas (TCGA) indicate that the expression levels of 14-3-3 zeta and beclin 1 (a key molecule involved in cellular autophagy) are up-regulated and positively correlated with each other (R = .5, P < .05) in HCC tissues. Chemoresistance developed in hepatoma cancer cells is associated with autophagy initiation. This study aimed to explore 14-3-3 zeta's role in regulating autophagy in HCC cells, with a focus on beclin 1. The co-localization of 14-3-3 zeta and beclin 1 was detectable in primary HCC tissues. To simulate in vivo tumour microenvironment (hypoxia), CSQT-2 and HCC-LM3 cells were exposed to 2% oxygen for 24 hours. The protein levels of 14-3-3 zeta and phospho-beclin 1(S295) peaked at 12 hours following hypoxia. Meanwhile, the strongest autophagy flux occurred: LC3II was increased, and p62 was decreased significantly. By sequencing the coding area of BECN 1 gene of CSQT-2 and HCC-LM3 cells, we found that the predicted translational products of BECN 1 gene contained (RLPSVP)-V-295 (R, arginine; L, leucine; P, proline; S, serine; V, valine), a classic 14-3-3 zeta binding motif. CO-IP results confirmed that 14-3-3 zeta bound to beclin 1, and this connection was markedly weakened when S-295 was mutated into A(295) (alanine). Further, 14-3-3 zeta overexpression prevented phospho-beclin 1(S295) from degradation and enhanced its binding to VPS34, whilst its knockdown accelerated the degradation. Additionally, 14-3-3 zeta enhanced the chemoresistance of HCC cells to cis-diammined dichloridoplatium by activating autophagy. Our work reveals that 14-3-3 zeta binds to and stabilizes phospho-beclin 1(S295) and induces autophagy in HCC cells to resist chemotherapy.