title: Mutual regulation of Bcl-2 proteins independent of the BH3 domain as shown by the BH3-lacking protein Bcl-x(AK).
abstract: The BH3 domain of Bcl-2 proteins was regarded as indispensable for apoptosis induction and for mutual regulation of family members. We recently described Bcl-x(AK), a proapoptotic splice product of the bcl-x gene, which lacks BH3 but encloses BH2, BH4 and a transmembrane domain. It remained however unclear, how Bcl-x(AK) may trigger apoptosis.For efficient overexpression, Bcl-x(AK) was subcloned in an adenoviral vector under Tet-OFF control. The construct resulted in significant apoptosis induction in melanoma and nonmelanoma cell lines with up to 50% apoptotic cells as well as decreased cell proliferation and survival. Disruption of mitochondrial membrane potential, and cytochrome c release clearly indicated activation of the mitochondrial apoptosis pathways. Both Bax and Bak were activated as shown by clustering and conformation analysis. Mitochondrial translocation of Bcl-x(AK) appeared as an essential and initial step. Bcl-x(AK) was critically dependent on either Bax or Bak, and apoptosis was abrogated in Bax/Bak double knockout conditions as well by overexpression of Bcl-2 or Bcl-x(L). A direct interaction with Bcl-2, Bax, Bad, Noxa or Puma was however not seen by immunoprecipitation. Thus besides BH3-mediated interactions, there exists an additional way for mutual regulation of Bcl-2 proteins, which is independent of the BH3. This pathway appears to play a supplementary role also for other proapoptotic family members, and its unraveling may help to overcome therapy resistance in cancer.
authors.affiliation: Department of Dermatology and Allergy, Skin Cancer Center, University Medical Center Charité, Berlin, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Plötz
authors.forename: Amir M
authors.initials: AM
authors.lastname: Hossini
authors.forename: Bernhard
authors.initials: B
authors.lastname: Gillissen
authors.forename: Peter T
authors.initials: PT
authors.lastname: Daniel
authors.forename: Eggert
authors.initials: E
authors.lastname: Stockfleth
authors.forename: Jürgen
authors.initials: J
authors.lastname: Eberle
mesh_terms: D017209:Apoptosis / Q000235:genetics; D020169:Caspases / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D045304:Cytochromes c / Q000235:genetics / Q000378:metabolism; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics* / Q000378:metabolism*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000235:genetics* / Q000378:metabolism*; D051020:bcl-X Protein / Q000235:genetics* / Q000378:metabolism*
chemical_list: C494772:BAK1 protein, human; C474007:Bax protein (53-86); D008565:Membrane Proteins; D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051020:bcl-X Protein; D045304:Cytochromes c; D020169:Caspases