BAK multimerization for apoptosis, but not bid binding, is inhibited by negatively charged residue in the BAK hydrophobic groove

Mol Cancer. 2013 Jun 19;12:65. doi: 10.1186/1476-4598-12-65.

Abstract

Background: BCL-2 family proteins BAK and BAX orchestrate outer mitochondrial membrane permeabilization (MOMP) during apoptosis by forming pores in the membrane to release apoptogenic factors that commits a cell to death. BAK and BAX therefore function as a 'point of no return' in the apoptotic cascade. BAK activation is a multi-step process involving conformational changes, mediated by BH3-only proteins or p53, which lead eventually to oligomerization and pore formation. Further, recent reports show that BAK activation is also linked to and dependent upon dephosphorylation of both tyrosine and serine residues.

Findings: We hypothesized that phosphorylation of BAK at tyrosine residue 110 (Y110) was functionally important during the BAK activation process. BAK/BAX double knockout HCT116 cells expressing a phosphor-mimetic BAK mutant (BAK Y110E), showed impaired dimerization and multimerization capacity when treated with either UV irradiation or etoposide when compared to cells reconstituted to express wild-type BAK. The Y110E mutant also showed decreased release of cytochrome c from isolated mitochondria challenged with tBid protein, resulting in a failure to activate caspase 3. Interestingly, co-immunoprecipitation experiments suggest that a negative charge at this residue may be important for the recruitment of Bid to BAK, but conversely that this also impairs BAK:BAK interactions.

Conclusion: These findings implicate dephosphorylation of Y110 as having an important mechanistic role in BAK activation, and underscores how post-translational modifications are intimately linked and coupled to the protein-protein interactions required for BAK activation during apoptosis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acids* / chemistry
  • Apoptosis*
  • BH3 Interacting Domain Death Agonist Protein / metabolism
  • Caspase 3 / metabolism
  • Cell Line, Tumor
  • Cytochromes c / metabolism
  • Humans
  • Hydrophobic and Hydrophilic Interactions*
  • Mitochondria / metabolism
  • Models, Molecular
  • Mutation
  • Phosphorylation
  • Protein Binding
  • Protein Conformation
  • Protein Multimerization*
  • Protein Transport
  • Tumor Suppressor Protein p53 / metabolism
  • Tyrosine / chemistry
  • bcl-2 Homologous Antagonist-Killer Protein / chemistry*
  • bcl-2 Homologous Antagonist-Killer Protein / genetics
  • bcl-2 Homologous Antagonist-Killer Protein / metabolism

Substances

  • Amino Acids
  • BH3 Interacting Domain Death Agonist Protein
  • Tumor Suppressor Protein p53
  • bcl-2 Homologous Antagonist-Killer Protein
  • Tyrosine
  • Cytochromes c
  • Caspase 3