Induction of cell death by the BH3-only Bcl-2 homolog Nbk/Bik is mediated by an entirely Bax-dependent mitochondrial pathway.
Nbk/Bik (natural born killer/Bcl-2-interacting killer) is a tissue-specific BH3-only protein whose molecular function is still largely unknown. To investigate the mechanism of Nbk action, we established a single- vector adenoviral system based on the Tet-off conditional expression of Nbk. Upon Nbk expression, only Bax-positive, but not Bax-deficient cells were found to undergo apoptosis. Interestingly, Nbk failed to induce apoptosis in the absence of Bax, even despite expression of the related molecule Bak. Re-expression of Bax restored the sensitivity to Nbk. Similarly, Bax wild-type HCT116 cells were highly susceptible, whereas HCT116 Bax knock-out cells remained resistant to Nbk-induced apoptosis. In Bax-positive cells, Nbk induced a conformational switch in the Bax N-terminus coinciding with cytochrome c release, mitochondrial permeability transition and caspase-9 processing. Immunoprecipitation studies revealed that Nbk interacts with Bcl-x(L) and Bcl-2 but not with Bax. Since, in addition, Nbk did not localize to the mitochondria, our data suggest a model in which Nbk acts as an indirect killer to trigger Bax-dependent apoptosis, whereas Bak is not sufficient to confer sensitivity to Nbk.
Gillissen
Bernhard
B
Department of Hematology, Charité-Campus Berlin-Buch, Humboldt University, D-13125 Berlin-Buch, Germany.
Essmann
Frank
F
Graupner
Vilma
V
Stärck
Lilian
L
Radetzki
Silke
S
Dörken
Bernd
B
Schulze-Osthoff
Klaus
K
Daniel
Peter T
PT
D000256:Adenoviridae / Q000235:genetics; D000595:Amino Acid Sequence; D017209:Apoptosis / Q000235:genetics / Q000502:physiology; D051017:Apoptosis Regulatory Proteins; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D003574:Cytochrome c Group / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D015967:Gene Expression Regulation, Viral; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins; D011487:Protein Conformation; D020033:Protein Isoforms; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D014407:Tumor Cells, Cultured; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein
D051017:Apoptosis Regulatory Proteins; C494772:BAK1 protein, human; C494826:BAX protein, human; C096820:BIK protein, human; D003574:Cytochrome c Group; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D020033:Protein Isoforms; D011506:Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein; D020169:Caspases