Skip to main content
Article thumbnail
Location of Repository

BAK activation: a multiple step mechanism

By Kathrin Weber


Although the pro-apoptotic BCL-2 family proteins BAK and BAX play a key role in mitochondrial perturbation their transition from an inactive closed conformation to a membrane permeabilising pore remains unclear. I found that BAK in viable cells existed in a primed state which was characterized by an occluded N-terminus and an exposed BH3 domain. This conformation facilitated binding to the hydrophobic groove of BCL-XL and served as a checkpoint maintaining cell survival by preventing its further activation. Isolation of BAK by immunoprecipitation suggests that only a discrete portion is present in this primed conformation. Reconstitution of the BCL-XL BAK complex into a BAK/BAX null background rendered cells more sensitive to the BAD BH3 mimetic ABT-737 indicating that primed BAK is primarily involved in ABT-737 induced apoptosis. \ud Primed BAK was displaced from BCL-XL by ABT-737 followed by an N-terminal conformational change and subsequent formation of dimers and higher molecular weight complexes. These sequential BAK activation steps occurred independently of cell fate and did not represent the rate limiting steps in BAK activation as a BAK BH3 mutant L78A lost proapoptotic function but still oligomerised as efficiently as wt BAK. \ud Thus the transition from inactive BAK to a membrane permeabilising pore requires an additional activation step. I demonstrate that after 30 min of ABT-737 exposure primed BAK, after its displacement from BCL-XL, interacts with BIMEL reflecting the transient nature of this interaction. This interaction represented an additional step in BAK activation as BAK pro-apoptotic function was enhanced when BIMEL and BAK were co-expressed. However BIMEL did not induce the N-terminal conformational change nor oligomerisation of BAK and its interaction occurred downstream of both these events. In addition the pool of BIMEL involved in the further activation of BAK did not represent that sequestered by the antiapoptotic proteins BCL-2 and BCL-XL. \ud These data suggest that BAK activation occurs in multiple steps in which a further activation event is required after the exposure of the BH3 domain, the N-terminal conformational change and the formation of high molecular weight complexes but prior to cytochrome c release. I propose that this event may be represented by interaction of N-terminal conformational changed/oligomerised BAK with BIMEL

Publisher: University of Leicester
Year: 2011
OAI identifier:

Suggested articles


  1. (2002). A matter of life and death." doi
  2. (1976). A rapid and sensitive for the quantitation of microgram quantitites of protein utilizing the principle of protein-dye binding." doi
  3. (2004). Activation of apoptosis in vivo by a hydrocarbon-stapled BH3 helix." doi
  4. (1999). An APAF-1.cytochrome c multimeric complex is a functional apoptosome that activates procaspase-9." doi
  5. (2005). An inhibitor of Bcl-2 family proteins induces regression of solid tumours." doi
  6. (2000). Apoptosis in development."
  7. (2007). Apoptosis initiated when BH3 ligands engage multiple Bcl-2 homologs, not Bax or Bak." doi
  8. (1972). Apoptosis: a basic biological phenomenon with wideranging implications in tissue kinetics." doi
  9. (2009). Bak activation for apoptosis involves oligomerization of dimers via their alpha6 helices." doi
  10. (2008). BAX activation is initiated at a novel interaction site." doi
  11. (2005). Bax forms multispanning monomers that oligomerize to permeabilize membranes during apoptosis." doi
  12. (2001). Bax is present as a high molecular weight oligomer/complex in the mitochondrial membrane of apoptotic cells." doi
  13. (2002). Bcl-2 and Bcl-xL inhibit CD95-mediated apoptosis by preventing mitochondrial release of Smac/DIABLO and subsequent inactivation of X-linked inhibitor-of-apoptosis protein." doi
  14. (1988). Bcl-2 gene promotes haemopoietic cell survival and cooperates with c-myc to immortalize pre-B cells." doi
  15. (1993). Bcl-2 heterodimerizes in vivo with a conserved homolog, Bax, that accelerates programmed cell death." doi
  16. (2009). Bcl-2 inhibitors: small molecules with a big impact on cancer therapy." doi
  17. (1991). bcl-2 inhibits multiple forms of apoptosis but not negative selection in thymocytes." doi
  18. (1993). Bcl-2 oncoprotein blocks chemotherapy-induced apoptosis in a human leukemia cell line."
  19. (2003). BCL-2 selectively interacts with the BID-induced open conformer of BAK, inhibiting BAK auto-oligomerization." doi
  20. (2005). BH3 domains of BH3-only proteins differentially regulate Bax-mediated mitochondrial membrane permeabilization both directly and indirectly." doi
  21. (2001). BH3-only proteins that bind pro-survival Bcl-2 family members fail to induce apoptosis in the absence of Bax and Bak." Gnes and Dev. doi
  22. (2002). Bid, Bax, and lipids cooperate to form supramolecular openings in the outer mitochondrial membrane." doi
  23. (2001). Bmf: a proapoptotic BH3-only protein regulated by interaction with the myosin V actin motor complex, activated by anoikis." doi
  24. (2001). Breaking the mitochondrial barrier."
  25. (1994). C. elegans cell survival gene ced-9 encodes a functional homolog of the mammalian proto-oncogene bcl-2." doi
  26. (1992). Caenorhabditis elegans gene ced-9 protects cells from programmed cell death." doi
  27. (1999). Caspase Activation Involves the Formation of the Aposome, a Large (~700 kDa) Caspase-activating Complex."
  28. (2008). Caspase-mediated Bak activation and cytochrome c release during intrinsic apoptotic cell death in Jurkat cells." doi
  29. (1997). Caspases: the executioners of apoptosis." doi
  30. (1999). Cell damage-induced conformational changes of the pro-apoptotic protein Bak in vivo precede the onset of apoptosis." doi
  31. (2004). Cell death: critical control points." doi
  32. (2001). Cellular damage signals promote sequential changes at the N-terminus and BH-1 domain of the pro-apoptotic protein Bak." Oncogene doi
  33. (2007). Chronic lymphocytic leukemia requires BCL2 to sequester prodeath BIM, explaining sensitivity to BCL2 antagonist ABT737." doi
  34. (1996). Cleavage of lamin A by Mch2 alpha but not CPP32: multiple interleukin 1 beta-converting enzyme-related proteases with distinct substrate recognition properties are active in apoptosis." doi
  35. (1994). Cleavage of poly(ADP-ribose) polymerase by a proteinase with properties like ICE." doi
  36. (2006). Comparison of chemical inhibitors of antiapoptotic Bcl-2-family proteins." doi
  37. (2001). Damage-induced Bax N-terminal change, translocation to mitochondria and formation of Bax dimers/complexes occur regardless of cell fate." doi
  38. (2009). Different forms of cell death induced by putative BCL2 inhibitors." doi
  39. (2005). Differential targeting of prosurvival Bcl-2 proteins by their BH3-only ligands allows complementary apoptotic function." doi
  40. (2002). Direct addition of BimL to mitochondria does not lead to cytochrome c release." doi
  41. (2003). Discovery, characterization, and structure-activity relationships studies of proapoptotic polyphenols targeting B-cell lymphocyte/leukemia-2 proteins." doi
  42. (2002). Distinct BH3 domains either sensitize or activate mitochondrial apoptosis, serving as prototype cancer therapeutics." doi
  43. (2001). E1B 19K blocks Bax oligomerization and tumor necrosis factor alpha-mediated apoptosis." doi
  44. (1995). Early redistribution of plasma membrane phosphatidylserine is a general feature of apoptosis regardless of the initiating stimulus: inhibition by overexpression of Bcl-2 and Abl." doi
  45. (1992). Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. doi
  46. (2007). Embedded together: the life and death consequences of interaction of the Bcl-2 family with membranes." doi
  47. (1999). Genetic control of programmed cell death in the nematode Caenorhabditis elegans." doi
  48. (2007). Gossypol induces apoptosis in human PC-3 prostate cancer cells by modulating caspase-dependent and caspase-independent cell death pathways." doi
  49. (2007). Gossypol reduction of tumor growth through ROSdependent mitochondria pathway in human colorectal carcinoma cells." doi
  50. (2008). Gossypol, a BH3 mimetic, induces apoptosis in chronic lymphocytic leukemia cells." doi
  51. (1997). Heterodimerization-independent functions of cell death regulatory proteins Bax and Bcl-2 in yeast and mammalian cells." doi
  52. (2008). How do BCL-2 proteins induce mitochondrial outer membrane permeabilization?" doi
  53. (2003). Interaction with a membrane surface triggers a reversible conformational change in Bax normally associated with induction of apoptosis." doi
  54. (2006). Mechanisms of apoptosis sensitivity and resistance to the BH3 mimetic ABT-737 in acute myeloid leukemia." doi
  55. (2008). Membrane binding by tBid initiates an ordered series of events culminating in membrane permeabilization by Bax." doi
  56. (2006). Mitochondria primed by death signals determine cellular addiction to antiapoptotic BCL-2 family members." doi
  57. (2007). Mitochondrial permeabilization relies on BH3 ligands engaging multiple prosurvival Bcl-2 relatives, not Bak." doi
  58. (1984). Molecular cloning of the chromosomal breakpoint of B-cell lymphomas and leukemias with the t(11;14) chromosome translocation." doi
  59. (1997). Nonionic detergents induce dimerization among members of the Bcl-2 family." doi
  60. (2005). Proapoptotic Bak is sequestered by Mcl-1 and Bcl-xL, but not Bcl-2, until displaced by BH3-only proteins." Genes Dev doi
  61. (2001). Proapoptotic BAX and BAK: a requisite gateway to mitochondial dysfunction and death." doi
  62. (2001). Proapoptotic BAX and BAK: a requisite gateway to mitochondrial dysfunction and death." doi
  63. (1965). Programmed Cell Death--I. Cytology of Degeneration in the Intersegmental Muscles of the Pernyi Silkmoth." doi
  64. (2005). Promoting apoptosis as a strategy for cancer drug discovery." doi
  65. (2005). Regulatory phosphorylation of Bim: sorting out the ERK from the JNK." doi
  66. (2007). Small molecule obatoclax (GX15-070) antagonizes MCL-1 and overcomes MCL-1-mediated resistance to apoptosis." doi
  67. (2002). Spatial and temporal association of Bax with mitochondrial fission sites, Drp1, and Mfn2 during apoptosis." doi
  68. (1993). Specific proteolytic cleavage of poly(ADPribose) polymerase: an early marker of chemotherapy-induced apoptosis."
  69. (2009). Stepwise activation of BAX and BAK by tBID, BIM, and PUMA initiates mitochondrial apoptosis." doi
  70. (1988). Structure of a bacterial sensory receptor. A sitedirected sulfhydryl study."
  71. (2000). Structure of Bax: coregulation of dimer formation and intracellular localization."
  72. (1997). Structure of Bcl-xL-Bak peptide complex: recognition between regulators of apoptosis." doi
  73. (1999). Suicidal tendancies: apoptotic cell death by caspase family proteinases." doi
  74. (2004). Survival factor-induced extracellular signal-regulated kinase phosphorylates BIM, inhibiting its association with BAX and proapoptotic activity." doi
  75. (2000). tBID, a membrane-targeted death ligand, oligomerizes BAK to release cytochrome c."
  76. (2007). The Bcl-2 apoptotic switch in cancer development and therapy." doi
  77. (2008). The BCL-2 protein family: opposing activities that mediate cell death." doi
  78. (2006). The BH3 mimetic ABT-737 targets selective Bcl-2 proteins and efficiently induces apoptosis via Bak/Bax if Mcl-1 is neutralized." doi
  79. (1998). The C. elegans protein EGL-1 is required for programmed cell death and interacts with the Bcl-2-like protein CED-9." doi
  80. (2004). The first alpha helix of Bax plays a necessary role in its ligand-induced activation by the BH3-only proteins Bid and PUMA." doi
  81. (2000). The hallmarks of cancer." doi
  82. (2005). The multidomain proapoptotic molecules Bax and Bak are directly activated by heat." doi
  83. (2004). The pathophysiology of mitochondrial cell death." doi
  84. (2001). The role of dynamin-related protein 1, a mediator of mitochondrial fission, in apoptosis." doi
  85. (2006). The X-ray structure of a BAK homodimer reveals an inhibitory zinc binding site." doi
  86. (2008). To trigger apoptosis, Bak exposes its BH3 domain and homodimerizes via BH3:groove interactions." doi
  87. (1991). Two C. elegans genes control the programmed deaths of specific cells in the pharynx."
  88. (2003). VDAC2 inhibits BAK activation and mitochondrial apoptosis." doi
  89. (2009). VDAC2 is required for truncated BID-induced mitochondrial apoptosis by recruiting BAK to the mitochondria." doi
  90. (1996). X-ray and NMR structure of human Bcl-xL, an inhibitor of programmed cell death." doi

To submit an update or takedown request for this paper, please submit an Update/Correction/Removal Request.