Protein kinase CbetaII activation by 1-beta-D-arabinofuranosylcytosine is antagonistic to stimulation of apoptosis and Bcl-2alpha down-regulation.
1-beta-D-Arabinofuranosylcytosine (ara-C) stimulates the formation of both diglyceride and ceramide in the acute myelogenous leukemia cell line HL-60 (Strum, J. C., Small, G. W., Pauig, S. B., and Daniel, L. W. (1994) J. Biol. Chem 269, 15493-15497). ara-C also causes apoptosis in HL-60 cells which can be mimicked by exogenous ceramide. However, the signaling role for ara-C-induced diacylglycerol (DAG) is not defined. We found that Bcl-2 levels were increased by treatment of HL-60 cells with exogenous DAG or 12-O-tetradecanoylphorbol-13-acetate (TPA). In contrast, exogenous ceramide treatment caused a decrease in cellular Bcl-2 levels. Thus, ara-C stimulates the synthesis of two second messengers with opposing effects on Bcl-2. Since the effects of ara-C-induced DAG could be due to protein kinase C (PKC) activation, we determined the effects of ara-C on PKC isozymes. ara-C caused an increase in membrane-bound PKCbetaII (but not PKCalpha or PKCdelta). ara-C or TPA-induced translocation of PKCbetaII was inhibited by 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3), and ara-C-induced apoptosis was stimulated by pretreatment of the cells with ET-18-OCH3. ET-18-OCH3 also inhibited stimulation of Bcl-2 by TPA and enhanced the decrease in Bcl-2 observed in ara-C-treated cells. These data indicate that ara-C-induced apoptosis is limited by ara-C-stimulated PKCbetaII through effects on Bcl-2. To further determine the role of PKC, we used antisense oligonucleotides directed toward PKCbetaII. The antisense, but not the sense, oligonucleotide inhibited PKCbetaII activation and enhanced ara-C-induced apoptosis. These data demonstrate that the stimulation of apoptosis by ara-C is self-limiting and can be enhanced by inhibition of PKC.
Whitman
S P
SP
Department of Biochemistry, Bowman Gray School of Medicine, Wake Forest University, Winston-Salem, North Carolina 27157-1016, USA.
Civoli
F
F
Daniel
L W
LW
D017209:Apoptosis; D001692:Biological Transport; D003561:Cytarabine / Q000494:pharmacology*; D053938:DNA Fragmentation; D015536:Down-Regulation; D004789:Enzyme Activation; D018922:HL-60 Cells; D006801:Humans; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D010742:Phospholipid Ethers / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D064546:Protein Kinase C beta; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D016376:Oligonucleotides, Antisense; D010742:Phospholipid Ethers; D019253:Proto-Oncogene Proteins c-bcl-2; D003561:Cytarabine; C026659:edelfosine; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D013755:Tetradecanoylphorbol Acetate