title: PKC activation contributes to caspase-mediated eIF2alpha phosphorylation and cell death.
abstract: BACKGROUND
  Stress-induced phosphorylation of the alpha-subunit of eukaryotic initiation factor 2 (eIF2alpha), involved in translation, promotes cell suicide or survival. Since multiple signaling pathways are implicated in cell death, the present study has analyzed the importance of PKC activation in the stress-induced eIF2alpha phosphorylation, caspase activation and cell death in the ovarian cells of Spodoptera frugiperda (Sf9) and in their extracts.
  
  
  METHODS
  Cell death is analyzed by flow cytometry. Caspase activation is measured by Ac-DEVD-AFC hydrolysis and also by the cleavage of purified recombinant PERK, an endoplasmic reticulum-resident eIF2alpha kinase. Status of eIF2alpha phosphorylation and cytochrome c levels are analyzed by western blots.
  
  
  RESULTS
  PMA, an activator of PKC, does not promote cell death or affect eIF2alpha phosphorylation. However, PMA enhances late stages of UV-irradiation or cycloheximide-induced caspase activation, eIF2alpha phosphorylation and apoptosis in Sf9 cells. PMA also enhances cytochrome c-induced caspase activation and eIF2alpha phosphorylation in cell extracts. These changes are mitigated more efficiently by caspase inhibitor, z-VAD-fmk, than by calphostin, an inhibitor of PKC. In contrast, tunicamycin-induced eIF2alpha phosphorylation that does not lead to caspase activation or cell death is unaffected by PMA, z-VAD-fmk or by calphostin.
  
  
  CONCLUSIONS
  While caspase activation is a cause and consequence of eIF2alpha phosphorylation, PKC activation that follows caspase activation further enhances caspase activation, eIF2alpha phosphorylation, and cell death in Sf9 cells.
  
  
  CONCLUSIONS
  Caspases can activate multiple signaling pathways to enhance cell death.
authors.affiliation: Department of Biochemistry, University of Hyderabad, Hyderabad 500 046, Andhra Pradesh, India.
authors.forename: Pendyala
authors.initials: P
authors.lastname: Pushpanjali
authors.forename: Kolluru V A
authors.initials: KV
authors.lastname: Ramaiah
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D020169:Caspases / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects / Q000528:radiation effects; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects / Q000528:radiation effects; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D019476:Insect Proteins / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects / Q000528:radiation effects; D011493:Protein Kinase C / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D018411:Spodoptera; D013312:Stress, Physiological; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014415:Tunicamycin / Q000494:pharmacology; D014466:Ultraviolet Rays / Q000009:adverse effects
chemical_list: D000900:Anti-Bacterial Agents; D002273:Carcinogens; D015852:Eukaryotic Initiation Factor-2; D019476:Insect Proteins; D011500:Protein Synthesis Inhibitors; D014415:Tunicamycin; D045304:Cytochromes c; D003513:Cycloheximide; D011493:Protein Kinase C; D020169:Caspases; D013755:Tetradecanoylphorbol Acetate