Inhibition of expression of protein kinase C alpha by antisense cDNA inhibits phorbol ester-mediated arachidonate release.
A major unresolved issue in the area of signal transduction relates to the role of particular isoforms of protein kinase C (PKC) in mediating cellular responses subsequent to activation of that enzyme. We have addressed this issue by the use of antisense technology. We have stably transfected Madin-Darby canine kidney cells with antisense PKC alpha, PKC beta, or both PKC alpha and -beta cDNAs. The transfected cDNA was integrated and expressed. We have isolated cells in which expression of PKC alpha is inhibited. In cells transfected with antisense PKC alpha or both PKC alpha and -beta, phorbol ester-stimulated release of arachidonate and its metabolites was inhibited, whereas in cells transfected with antisense PKC beta cDNA alone, phorbol ester-stimulated arachidonate release was not significantly different from control cells. We thus demonstrate the use of a novel technique to inhibit PKC isoform expression. We show that inhibition of expression of PKC alpha causes a loss in phospholipase A2-mediated arachidonate release. Antisense-inhibited expression of PKC isoforms may provide a useful approach to define additional functions of particular PKC isoforms.
Godson
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C
Department of Pharmacology, University of California, San Diego, La Jolla 92093.
Bell
K S
KS
Insel
P A
PA
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D015152:Blotting, Northern; D015139:Blotting, Southern; D015153:Blotting, Western; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D016373:DNA, Antisense / Q000235:genetics / Q000494:pharmacology*; D004285:Dogs; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism*; D007668:Kidney; D007700:Kinetics; D011493:Protein Kinase C / Q000235:genetics* / Q000378:metabolism*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014162:Transfection
D016373:DNA, Antisense; D007527:Isoenzymes; D016718:Arachidonic Acid; D012313:RNA; D004247:DNA; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate