CNOT2 promotes degradation of p62/SQSTM1 as a negative regulator in ATG5 dependent autophagy.

Though CNOT2 is involved in regulation of adipogenic differentiation, apoptotic cell death and metastasis, the underlying autophagic mechanism of CNOT2 was unknown until now. Thus, in the present study, the critical role of CNOT2 in autophagy was elucidated in association with p62/SQSTM1 signaling. CNOT2 depletion induced p62/SQSTM1 accumulation and LC3B-II conversion, and also increased the number of puncta with impaired autophagic flux. In contrast, CNOT2 overexpression induced downregulation and ubiquitination of p62/SQSTM1 in HEK293 QBI. Furthermore, ubiquitination of p62/SQSTM1 was blocked by autophagy inhibition. Interestingly, CNOT2 was correlated with p62/SQSTM1 in HEK293 QBI cells and also was colocalized with p62/SQSTM1 in H1299 cells. Additionally, ATG5 was upregulated in CNOT2-depleted H1299 cells, while degradation of p62/SQSTM1 by CNOT2 was detected in ATG5+/+ MEF cells but not in ATG5-/- MEF cells. Of note, CNOT2 induced degradation of p62/SQSTM1 in HEK293 QBI cells co-transfected with Myc-ΔLIR/KIR or Myc-ΔUBA, but not with Myc-ΔPB1. Sub G1 population was increased in CNOT2-depleted H1299 cells by late autophagy inhibitors, ammonium chloride and chloroquine compared to 3-methyladenine. Overall, these findings provide novel insight into the critical role of CNOT2 as a negative regulator in ATG5 dependent autophagy.