title: Delivery of a monomeric p53 subdomain with mitochondrial targeting signals from pro-apoptotic Bak or Bax.
abstract: OBJECTIVE
  p53 targeted to the mitochondria is the fastest and most direct pathway for executing p53 death signaling. The purpose of this work was to determine if mitochondrial targeting signals (MTSs) from pro-apoptotic Bak and Bax are capable of targeting p53 to the mitochondria and inducing rapid apoptosis.
  
  
  METHODS
  p53 and its DNA-binding domain (DBD) were fused to MTSs from Bak (p53-BakMTS, DBD-BakMTS) or Bax (p53-BaxMTS, DBD-BaxMTS). Mitochondrial localization was tested via fluorescence microscopy in 1471.1 cells, and apoptosis was detected via 7-AAD in breast (T47D), non-small cell lung (H1373), ovarian (SKOV-3) and cervical (HeLa) cancer cells. To determine that apoptosis is via the intrinsic apoptotic pathway, TMRE and caspase-9 assays were conducted. Finally, the involvement of p53/Bak specific pathway was tested.
  
  
  RESULTS
  MTSs from Bak and Bax are capable of targeting p53 to the mitochondria, and p53-BakMTS and p53-BaxMTS cause apoptosis through the intrinsic apoptotic pathway. Additionally, p53-BakMTS, DBD-BakMTS, p53-BaxMTS and DBD-BaxMTS caused apoptosis in T47D, H1373, SKOV-3 and HeLa cells. The apoptotic mechanism of p53-BakMTS and DBD-BakMTS was Bak dependent.
  
  
  CONCLUSIONS
  Our data demonstrates that p53-BakMTS (or BaxMTS) and DBD-BakMTS (or BaxMTS) cause apoptosis at the mitochondria and can be used as a potential gene therapeutic in cancer.
authors.affiliation: Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, Utah, USA.
authors.forename: Karina J
authors.initials: KJ
authors.lastname: Matissek
authors.forename: Abood
authors.initials: A
authors.lastname: Okal
authors.forename: Mohanad
authors.initials: M
authors.lastname: Mossalam
authors.forename: Carol S
authors.initials: CS
authors.lastname: Lim
mesh_terms: D000595:Amino Acid Sequence; D017209:Apoptosis; D045744:Cell Line, Tumor; D015316:Genetic Therapy; D006367:HeLa Cells; D006801:Humans; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism / Q000628:therapy*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics*; D015398:Signal Transduction; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000235:genetics*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000737:chemistry / Q000235:genetics*; D051028:bcl-2-Associated X Protein / Q000737:chemistry / Q000235:genetics*
chemical_list: D011993:Recombinant Fusion Proteins; D016159:Tumor Suppressor Protein p53; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein