Evolutionary conserved N-terminal domain of Nrf2 is essential for the Keap1-mediated degradation of the protein by proteasome

Arch Biochem Biophys. 2005 Jan 15;433(2):342-50. doi: 10.1016/j.abb.2004.10.012.

Abstract

Under homeostatic conditions, Nrf2 activity is constitutively repressed. This process is dependent on Keap1, to which Nrf2 binds through the Neh2 domain. Since the N-terminal subdomain of Neh2 (Neh2-NT) contains evolutionarily conserved motifs, we examined the roles they play in the degradation of Nrf2. In Neh2-NT, we defined a novel motif that is distinct from the previously characterized DIDLID motif and designated it DLG motif. Deletion of Neh2-NT or mutation of the DLG motif largely abolished the Keap1-mediated degradation of Nrf2. These mutations were found to enfeeble the binding affinity of Nrf2 to Keap1. The Neh2-NT subdomain directed DLG-dependent, Keap1-independent, degradation of a reporter protein in the nucleus. By contrast, mutation of DLG did not affect the half-life of native Nrf2 protein in the nucleus under oxidative stress conditions. These results thus demonstrate that DLG motif plays essential roles in the Keap1-mediated proteasomal degradation of Nrf2 in the cytoplasm.

Publication types

  • Research Support, Non-U.S. Gov't
  • Review

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism*
  • Amino Acid Motifs
  • Animals
  • COS Cells
  • Cell Nucleus / metabolism
  • Chlorocebus aethiops
  • Conserved Sequence
  • Cytoplasm / metabolism
  • Cytoskeletal Proteins / metabolism*
  • DNA-Binding Proteins / chemistry*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Evolution, Molecular*
  • Genes, Reporter
  • Green Fluorescent Proteins / metabolism
  • Half-Life
  • Humans
  • Kelch-Like ECH-Associated Protein 1
  • Leucine / chemistry
  • Luciferases / metabolism
  • Mice
  • Models, Biological
  • Molecular Sequence Data
  • NF-E2-Related Factor 2
  • NIH 3T3 Cells
  • Oxidative Stress
  • Plasmids
  • Proteasome Endopeptidase Complex / metabolism*
  • Protein Binding
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / metabolism
  • Sequence Deletion
  • Sequence Homology, Amino Acid
  • Trans-Activators / chemistry*
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*
  • Ubiquitins / metabolism

Substances

  • Adaptor Proteins, Signal Transducing
  • Cytoskeletal Proteins
  • DNA-Binding Proteins
  • Keap1 protein, mouse
  • Kelch-Like ECH-Associated Protein 1
  • NF-E2-Related Factor 2
  • NFE2L2 protein, human
  • Nfe2l2 protein, mouse
  • Recombinant Fusion Proteins
  • Trans-Activators
  • Ubiquitins
  • Green Fluorescent Proteins
  • Luciferases
  • Proteasome Endopeptidase Complex
  • Leucine