SQSTM1/p62 activates NFE2L2/NRF2 via ULK1-mediated autophagic KEAP1 degradation and protects mouse liver from lipotoxicity

Autophagy. 2020 Nov;16(11):1949-1973. doi: 10.1080/15548627.2020.1712108. Epub 2020 Jan 10.

Abstract

Lipotoxicity, induced by saturated fatty acid (SFA)-mediated cell death, plays an important role in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). The KEAP1 (kelch like ECH associated protein 1)-NFE2L2/NRF2 (nuclear factor, erythroid 2 like 2) pathway is a pivotal defense mechanism against lipotoxicity. We previously reported that SQSTM1/p62 has a cytoprotective role against lipotoxicity through activation of the noncanonical KEAP1- NFE2L2 pathway in hepatocytes. However, the underlying mechanisms and physiological relevance of this pathway have not been clearly defined. Here, we demonstrate that NFE2L2-mediated induction of SQSTM1 activates the noncanonical KEAP1-NFE2L2 pathway under lipotoxic conditions. Furthermore, we identified that SQSTM1 induces ULK1 (unc-51 like autophagy activating kinase 1) phosphorylation by facilitating the interaction between AMPK (AMP-activated protein kinase) and ULK1, leading to macroautophagy/autophagy induction, followed by KEAP1 degradation and NFE2L2 activation. Accordingly, the activity of this SQSTM1-mediated noncanonical KEAP1-NFE2L2 pathway conferred hepatoprotection against lipotoxicity in the livers of conventional sqstm1- and liver-specific sqstm1-knockout mice. Moreover, this pathway activity was evident in the livers of patients with nonalcoholic fatty liver. This axis could thus represent a novel target for NAFLD treatment. Abbreviations: ACACA: acetyl-CoA carboxylase alpha; ACTB: actin beta; BafA1: bafilomycin A1; CM-H2DCFDA:5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate; CQ: chloroquine; CUL3: cullin 3; DMSO: dimethyl sulfoxide; FASN: fatty acid synthase; GSTA1: glutathione S-transferase A1; HA: hemagglutinin; Hepa1c1c7: mouse hepatoma cells; HMOX1/HO-1: heme oxygenase 1; KEAP1: kelch like ECH associated protein 1; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; MTORC1: mechanistic target of rapamycin kinase complex 1; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; NAC: N-acetyl-L-cysteine; NAFLD: nonalcoholic fatty liver disease; NASH: nonalcoholic steatohepatitis; NFE2L2/NRF2: nuclear factor, erythroid 2 like 2; NQO1: NAD(P)H quinone dehydrogenase 1; PA: palmitic acid; PARP: poly (ADP-ribose) polymerase 1; PRKAA1/2: protein kinase AMP-activated catalytic subunits alpha1/2; RBX1: ring-box 1; ROS: reactive oxygen species; SESN2: sestrin 2; SFA: saturated fatty acid; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; SREBF1: sterol regulatory element binding transcription factor 1; TBK1: TANK binding kinase 1; TUNEL: terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling; ULK1: unc-51 like autophagy activating kinase.

Keywords: KEAP1-NFE2L2 pathway; NAFLD; SQSTM1; ULK1; lipotoxicity.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism
  • Animals
  • Autophagy / genetics
  • Autophagy / physiology*
  • Fatty Acids / toxicity*
  • Fibroblasts / metabolism
  • Hepatocytes / drug effects*
  • Hepatocytes / metabolism
  • Humans
  • Kelch-Like ECH-Associated Protein 1 / drug effects
  • Liver / drug effects*
  • Mice, Knockout
  • NF-E2-Related Factor 2 / drug effects
  • Reactive Oxygen Species / metabolism
  • Sequestosome-1 Protein / drug effects

Substances

  • Adaptor Proteins, Signal Transducing
  • Fatty Acids
  • KEAP1 protein, human
  • Keap1 protein, mouse
  • Kelch-Like ECH-Associated Protein 1
  • NF-E2-Related Factor 2
  • NFE2L2 protein, human
  • Nfe2l2 protein, mouse
  • Reactive Oxygen Species
  • SQSTM1 protein, human
  • Sequestosome-1 Protein
  • Sqstm1 protein, mouse

Grants and funding

This work was supported by the National Research Foundation of Korea (NRF-2017R1A2B4007400 to S.H. Bae; NRF-2017R1D1A1B03032808 to J.S. Park) and a Faculty Research Grant from the Yonsei University College of Medicine (6-2019-0068 to S.H. Bae). The study was also supported by a grant from the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI) (HI17C0913 and HI16C0257 to S.H.Bae).