title: Chk1 activity is required for BAK multimerization in association with PUMA during mitochondrial apoptosis.
abstract: BACKGROUND
  The Bcl-2 protein BAK is a key player in mitochondrial apoptosis and responds to a myriad of different death signals. Activation of BAK is a multistep process that involves a number of conformational changes mediated by BH3-only proteins or p53 which leads to BAK multimerization and pore formation in the mitochondrial outer membrane. We previously reported that BAK activation is dependent upon dephosphorylation of both tyrosine and serine residues. Further, recent reports demonstrated that PP2A activity is required for BAK multimerization. Since Chk1, a checkpoint kinase involved in the activation of G2 checkpoint, is regulated by PP2A, we therefore hypothesized that Chk1 is involved in BAK multimerization during cell cycle arrest upon severe DNA damage.
  
  
  RESULTS
  We now show that treatment of HCT116-WT BAK cells with a Chk1 inhibitor impaired BAK dimerization and mutimerization when treated with the DNA damaging agents UV or etoposide. As a result there is a concomitant decrease of cytochrome c release from isolated mitochondria challenged with tBid protein and failure in the activation of caspase3. Interestingly, co-immunoprecipitation studies suggest that Chk1 is required for recruitment of BH3- only protein PUMA to BAK. We also showed that Chk1 is associated with BAK upon DNA damage.
  
  
  CONCLUSIONS
  These findings novelly demonstrate the involvement of a checkpoint kinase Chk1 is required for BAK activation and underscores the importance of involvement of Chk1 in mitochondrial apoptosis upon severe DNA damage.
authors.affiliation: Department of Oncology, The Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, OX3 9DS, UK. aazad03@gmail.com.
authors.forename: Abul
authors.initials: A
authors.lastname: Azad
authors.affiliation: Department of Oncology, The Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, OX3 9DS, UK. astorey62@gmail.com.
authors.forename: Alan
authors.initials: A
authors.lastname: Storey
mesh_terms: D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism*; D020169:Caspases / Q000378:metabolism; D000071877:Checkpoint Kinase 1; D045304:Cytochromes c / Q000378:metabolism; D004249:DNA Damage; D005047:Etoposide / Q000494:pharmacology; D045325:HCT116 Cells; D006801:Humans; D008928:Mitochondria / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism*; D055503:Protein Multimerization; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D014466:Ultraviolet Rays; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000378:metabolism*
chemical_list: D051017:Apoptosis Regulatory Proteins; C494772:BAK1 protein, human; C438378:BBC3 protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D050998:bcl-2 Homologous Antagonist-Killer Protein; D005047:Etoposide; D045304:Cytochromes c; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D020169:Caspases