Inhibition of the protein kinase C pathway promotes anti-CD95-induced apoptosis in Jurkat T cells.
The role of the basal activity of the serine/threonine protein kinase, protein kinase C (PKC) in the regulation of anti-CD95-induced apoptosis in Jurkat T cells was investigated. The PKC-specific inhibitor GF 109203X and the proposed cPKC-specific inhibitor Go 6976, in a concentration-dependent manner, increased the percentage of cells undergoing apoptosis induced by anti-CD95 mAb as demonstrated by propidium iodide (PI) staining, TUNEL assay and DNA fragmentation by gel electrophoresis. Furthermore, Go 6976 and GF 109203X abrogated phorbol myristate acetate-induced inhibition of anti-CD95-induced apoptosis. To examine the molecular mechanism by which PKC modulates anti-CD95-induced apoptosis, the effects of Go 6976 on known effector and regulatory molecules of cell death were studied. Increased recruitment of cells undergoing apoptosis was associated with enhanced anti-CD95-induced proteolytic cleavage of the most receptor-proximal cysteine protease caspase-8, subsequent cleavage and activation of the machinery protease caspase-3, and cleavage of the caspase substrates DNA-dependent protein kinase catalytic subunit, poly-(ADP-ribose) polymerase and lamin B1. CD95 and FADD protein levels in Jurkat T cells were not altered by Go 6976 treatment. In addition, Go 6976 did not alter protein levels and subcellular distribution of the anti-apoptotic molecules Bcl-2 and Bcl-xL. These data suggest indirectly that basal PKC activity acts at an early stage in the anti-CD95-induced caspase pathway to attenuate subsequent activation of downstream effector molecules and associated apoptosis in Jurkat T cells.
Department of Medicine, University of California, Irvine 92697, USA.
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Drew
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Kumar
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Bandyopadhyay
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S
Gupta
D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002227:Carbazoles / Q000494:pharmacology; D053148:Caspase 3; D020169:Caspases; D002478:Cells, Cultured; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000378:metabolism; D004790:Enzyme Induction; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012680:Sensitivity and Specificity; D013347:Subcellular Fractions / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D051020:bcl-X Protein; D019014:fas Receptor / Q000096:biosynthesis / Q000276:immunology*
D000911:Antibodies, Monoclonal; C494811:BCL2L1 protein, human; D002227:Carbazoles; D004791:Enzyme Inhibitors; D007211:Indoles; D008301:Maleimides; D019253:Proto-Oncogene Proteins c-bcl-2; D051020:bcl-X Protein; D019014:fas Receptor; C081021:Go 6976; D011493:Protein Kinase C; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; C070515:bisindolylmaleimide I