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 mediatin g cellular responses subsequent to activation of that enzyme. We have addressed this issue by the use of antisense technology. We have stabl y transfected Madin-Darby canine kidney cells with antisense PKCalpha, PKCbeta, or both PKCalpha and -beta cDNAs. The transfected cDNA was i ntegrated and expressed. We have isolated cells in which expression of PKCalpha is inhibited. In cells transfected with antisense PKCalpha o r both PKCalpha and -beta, phorbol ester-stimulated release of arachid onate and its metabolites was inhibited, whereas in cells transfected with antisense PKCbeta cDNA alone, phorbol ester-stimulated arachidona te release was not significantly different from control cells. We thus demonstrate the use of a novel technique to inhibit PKC isoform expre ssion. We show that inhibition of expression of PKCalpha causes a loss in phospholipase A2-mediated arachidonate release. Antisense-inhibite d expression of PKC isoforms may provide a useful approach to define a dditional functions of particular PKC isoforms.