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Protein kinase C alpha mediates phospholipase D activation by nucleotides and phorbol ester in Madin-Darby canine kidney cells. Stimulation of phospholipase D is independent of activation of polyphosphoinositide-specific phospholipase C and phospholipase A2
Protein kinase C (PKC) has been implicated in the activation of phospholipase D (PLD) in a number of systems. By antisense technology, we have "knocked out" alpha and beta isoforms of PKC to study the role of these isoforms in PLD activation in Madin-Darby canine kidney (MDCK) cells. To th...
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Published in: | The Journal of biological chemistry 1994-04, Vol.269 (14), p.10511-10516 |
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Main Authors: | , , , , |
Format: | Article |
Language: | English |
Subjects: | |
Citations: | Items that this one cites Items that cite this one |
Online Access: | Get full text |
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Summary: | Protein kinase C (PKC) has been implicated in the activation of phospholipase D (PLD) in a number of systems. By antisense
technology, we have "knocked out" alpha and beta isoforms of PKC to study the role of these isoforms in PLD activation in
Madin-Darby canine kidney (MDCK) cells. To this end, we have studied PLD activation by phorbol 12-myristate 13-acetate (PMA),
ATP, UTP, and 2-methylthio-ATP in cells labeled with [3H]palmitic acid. [3H]Phosphatidylethanol (PEt) production catalyzed
by PLD in the presence of ethanol was time- and concentration-dependent in PMA- and nucleotide-stimulated cells. In Ca(2+)-free
medium, [3H]PEt accumulation was diminished for all stimuli assayed. Treatment of cells with chelerythrine, an inhibitor of
PKC, and phorbol ester down-regulation of PKC inhibited [3H]PEt production by both PMA and nucleotides. In cells transfected
with antisense PKC alpha or both PKC alpha and PKC beta, PLD activation was inhibited by both PMA and nucleotides, whereas
in cells transfected with antisense PKC beta, PLD activation was similar to that of control cells. Moreover, inhibition of
polyphosphoinositide-specific PLC (by neomycin) or of release of arachidonic acid and arachidonic acid metabolites (by nordihydroguaiaretic
acid or by indomethacin) failed to decrease [3H]PEt accumulation in PMA- and nucleotide-stimulated MDCK-D1 cells. From these
data, we conclude that in MDCK-D1 cells PMA and nucleotide receptors utilize PKC alpha to regulate PLD activity and that PLD
activation is independent of the activation of polyphosphoinositide-specific PLC and phospholipase A2-mediated release of
arachidonic acid or arachidonic acid metabolites. |
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ISSN: | 0021-9258 1083-351X |
DOI: | 10.1016/s0021-9258(17)34089-9 |