Disruption of the VDAC2-Bak interaction by Bcl-x(S) mediates efficient induction of apoptosis in melanoma cells.
The proapoptotic B-cell lymphoma (Bcl)-2 protein Bcl-x(S) encloses the Bcl-2 homology (BH) domains BH3 and BH4 and triggers apoptosis via the multidomain protein Bak, however, the mechanism remained elusive. For investigating Bcl-x(S) efficacy and pathways, an adenoviral vector was constructed with its cDNA under tetracycline-off control. Bcl-x(S) overexpression resulted in efficient apoptosis induction and caspase activation in melanoma cells. Indicative of mitochondrial apoptosis pathways, Bcl-x(S) translocated to the mitochondria, disrupted the mitochondrial membrane potential and induced release of cytochrome c, apoptosis-inducing factor and second mitochondria-derived activator of caspases. In melanoma cells, Bcl-x(S) resulted in significant Bak activation, and Bak knockdown as well as Bcl-x(L) overexpression abrogated Bcl-x(S)-induced apoptosis, whereas Mcl-1 (myeloid cell leukemia-1) knockdown resulted in a sensitization. With regard to the particular role of voltage-dependent anion channel 2 (VDAC2) for inhibition of Bak, we identified here a notable interaction between Bcl-x(S) and VDAC2 in melanoma cells, which was proven in reciprocal coimmunoprecipitation analyses. On the other hand, Bcl-x(S) showed no direct interaction with Bak, and its binding to VDAC2 appeared as also independent of Bak expression. Suggesting a new proapoptotic mechanism, Bcl-x(S) overexpression resulted in disruption of the VDAC2-Bak interaction leading to release of Bak. Further supporting this pathway, overexpression of VDAC2 strongly decreased apoptosis by Bcl-x(S). New proapoptotic pathways are of principle interest for overcoming apoptosis deficiency of melanoma cells.
Department of Dermatology and Allergy, Skin Cancer Center Charité, Charité-Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany.
M
M
Plötz
B
B
Gillissen
A M
AM
Hossini
P T
PT
Daniel
J
J
Eberle
D017209:Apoptosis; D045744:Cell Line, Tumor; D045325:HCT116 Cells; D006801:Humans; D008545:Melanoma / Q000378:metabolism / Q000473:pathology; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D011485:Protein Binding; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D050996:Voltage-Dependent Anion Channel 2 / Q000378:metabolism*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051020:bcl-X Protein / Q000378:metabolism*
D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019253:Proto-Oncogene Proteins c-bcl-2; D034741:RNA, Small Interfering; C494765:VDAC2 protein, human; D050996:Voltage-Dependent Anion Channel 2; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051020:bcl-X Protein