title: Tissue Factor/ FVIIa prevents the extrinsic pathway of apoptosis by regulation of the tumor suppressor Death-Associated Protein Kinase 1 (DAPK1).
abstract: BACKGROUND
  This study determines the impact of tissue factor (TF)-signaling on the extrinsic pathway of apoptosis in cancer cells and propose death associated protein kinase-1 (DAPK1) as a novel key regulator.
  
  
  METHODS
  In MDA-MB-231 breast and PC3 prostate cancer cells, mRNA levels were analyzed by real-time PCR and protein expressions were assessed by flow cytometry or western blot. Caspase-8 and -3 levels, cell size, and changes in nuclear morphology were recorded using the ArrayScan microscope and 84 apoptosis-related genes were screened with the RT2 Profiler™ PCR Array.
  
  
  RESULTS
  In serum starved MDA-MB-231 cells, a TF/FVIIa-sensitive upregulation of apoptosis markers was recorded. Similarly, TRAIL-induced apoptosis was negatively regulated by TF/FVIIa (10 and 100 nM) and TF/FVIIa/FXa but not by active-site inhibited FVIIa. FVIIa, moreover, decreased the transcription of DAPK1 and thereby diminished the association between DAPK1 and FADD in the caspase-8 activating death-inducing signaling complex (DISC). TF/FVIIa regulation of caspase-8 and DAPK1 was dependent on PI3-kinase/AKT and independent of the protease activated receptors (PAR) 1 and 2. Despite of receptor expression and functional signaling, both PAR-agonist treatment and PAR-blocking antibodies in combination with FVIIa failed to influence the anti-apoptotic signal.
  
  
  CONCLUSIONS
  We hereby report that TF/FVIIa-induced signaling governs the extrinsic pathway of apoptosis by reducing the levels of DAPK1 in the DISC independently of PAR1 and PAR2. This implies the conceivable involvement of cell surface components other than the PARs and entails the search for TF-dependent regulators of DAPK1 transcription.
authors.affiliation: Department of Medical Sciences, Clinical Chemistry, Uppsala University, Sweden. mikael.aberg@medsci.uu.se
authors.forename: M
authors.initials: M
authors.lastname: Aberg
authors.forename: M
authors.initials: M
authors.lastname: Johnell
authors.forename: M
authors.initials: M
authors.lastname: Wickström
authors.forename: A
authors.initials: A
authors.lastname: Siegbahn
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D053181:Caspase 8 / Q000378:metabolism; D061945:Caspase Inhibitors; D045744:Cell Line, Tumor; D064550:Death-Associated Protein Kinases; D004789:Enzyme Activation; D015942:Factor VIIa / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D015951:Factor Xa / Q000494:pharmacology; D053200:Fas-Associated Death Domain Protein / Q000378:metabolism; D005260:Female; D006801:Humans; D008297:Male; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011471:Prostatic Neoplasms / Q000235:genetics / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D044463:Receptor, PAR-1 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D044464:Receptor, PAR-2 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction; D013925:Thromboplastin / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
chemical_list: D051017:Apoptosis Regulatory Proteins; D061945:Caspase Inhibitors; C505533:FADD protein, human; D053200:Fas-Associated Death Domain Protein; D012333:RNA, Messenger; D044463:Receptor, PAR-1; D044464:Receptor, PAR-2; D011994:Recombinant Proteins; D013925:Thromboplastin; C103587:recombinant FVIIa; D019869:Phosphatidylinositol 3-Kinases; C579096:DAPK1 protein, human; D064550:Death-Associated Protein Kinases; D051057:Proto-Oncogene Proteins c-akt; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D015942:Factor VIIa; D015951:Factor Xa; D053181:Caspase 8