Inhibitor of Nrf2 (INrf2 or Keap1) protein degrades Bcl-xL via phosphoglycerate mutase 5 and controls cellular apoptosis

J Biol Chem. 2011 Dec 30;286(52):44542-56. doi: 10.1074/jbc.M111.275073. Epub 2011 Nov 9.

Abstract

INrf2 (Keap1) is an adaptor protein that facilitates INrf2-Cul3-Rbx1-mediated ubiquitination/degradation of Nrf2, a master regulator of cytoprotective gene expression. Here, we present evidence that members of the phosphoglycerate mutase family 5 (PGAM5) proteins are involved in the INrf2-mediated ubiquitination/degradation of anti-apoptotic factor Bcl-xL. Mass spectrometry and co-immunoprecipitation assays revealed that INrf2, through its DGR domain, interacts with PGAM5, which in turn interacts with anti-apoptotic Bcl-xL protein. INrf2-Cul3-Rbx1 complex facilitates ubiquitination and degradation of both PGAM5 and Bcl-xL. Overexpression of PGAM5 protein increased INrf2-mediated degradation of Bcl-xL, whereas knocking down PGAM5 by siRNA decreased INrf2 degradation of Bcl-xL, resulting in increased stability of Bcl-xL. Mutation of PGMA5-E79A/S80A abolished INrf2/PGAM5/Bcl-xL interaction. Therefore, PGAM5 protein acts as a bridge between INrf2 and Bcl-xL interaction. Further studies showed that overexpression of INrf2 enhanced degradation of PGAM5-Bcl-xL complex, led to etoposide-mediated accumulation of Bax, increased release of cytochrome c from mitochondria, activated caspase-3/7, and enhanced DNA fragmentation and apoptosis. In addition, antioxidant (tert-butylhydroquinone) treatment destabilized the Nrf2-INrf2-PGAM5-Bcl-xL complex, which resulted in release of Nrf2 in cytosol and mitochondria, release of Bcl-xL in mitochondria, increase in Bcl-xL heterodimerization with Bax in mitochondria, and reduced cellular apoptosis. These data provide the first evidence that INrf2 controls Bcl-xL via PGAM5 and controls cellular apoptosis.

Publication types

  • Research Support, N.I.H., Extramural
  • Retracted Publication

MeSH terms

  • Adaptor Proteins, Signal Transducing / genetics
  • Adaptor Proteins, Signal Transducing / metabolism*
  • Amino Acid Substitution
  • Animals
  • Apoptosis / physiology*
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism
  • Caspase 3 / genetics
  • Caspase 3 / metabolism
  • Caspase 7 / genetics
  • Caspase 7 / metabolism
  • Cell Line, Tumor
  • Cullin Proteins / genetics
  • Cullin Proteins / metabolism
  • Cytochromes c / genetics
  • Cytochromes c / metabolism
  • Cytoskeletal Proteins / genetics
  • Cytoskeletal Proteins / metabolism*
  • Cytosol / metabolism
  • DNA Fragmentation
  • Kelch-Like ECH-Associated Protein 1
  • Mice
  • Mitochondria / genetics
  • Mitochondria / metabolism
  • Multiprotein Complexes / genetics
  • Multiprotein Complexes / metabolism
  • Mutation, Missense
  • Phosphoglycerate Mutase / genetics
  • Phosphoglycerate Mutase / metabolism*
  • Phosphoprotein Phosphatases
  • Phosphoric Monoester Hydrolases / genetics
  • Phosphoric Monoester Hydrolases / metabolism*
  • Protein Stability
  • Protein Structure, Tertiary
  • Proteolysis*
  • Ubiquitination / physiology
  • bcl-X Protein / genetics
  • bcl-X Protein / metabolism*

Substances

  • Adaptor Proteins, Signal Transducing
  • Bcl2l1 protein, mouse
  • Carrier Proteins
  • Cul3 protein, mouse
  • Cullin Proteins
  • Cytoskeletal Proteins
  • Keap1 protein, mouse
  • Kelch-Like ECH-Associated Protein 1
  • Multiprotein Complexes
  • RBX1 protein, mouse
  • bcl-X Protein
  • Cytochromes c
  • PGAM5 protein, mouse
  • Phosphoprotein Phosphatases
  • Phosphoric Monoester Hydrolases
  • Casp3 protein, mouse
  • Casp7 protein, mouse
  • Caspase 3
  • Caspase 7
  • Phosphoglycerate Mutase