Differential regulation of phospholipase D and phospholipase A2 by protein kinase C in P388D1 macrophages.
Activation of P388D1 macrophages by phorbol myristate acetate (PMA) resulted in the translocation of the protein kinase C (PKC) isoforms alpha, delta, and epsilon from the cytosol to membranes. Furthermore, PMA activated phospholipase D (PLD) in these cells, and potentiated the effect of the inflammatory lipid mediator platelet-activating factor (PAF) on PLD activation. PAF also activated phospholipase A2 (PLA2) and enhanced arachidonic acid (AA) release in P388D1 macrophages, and bacterial lipopolysaccharide (LPS) increased the responsiveness of these cells to PAF. In contrast with PLD, PLA2 activation in P388D1 macrophages was found to take place independently of PKC. This was supported by the following evidence: (i) PMA neither induced AA release nor enhanced the PAF response; (ii) inclusion of PMA along with LPS during priming did not have any effect on PAF-stimulated AA release; (iii) down-regulation of PMA-activatable PKC isoforms by chronic treatment with the phorbol ester had no effect on the PAF response; and (iv) the PKC inhibitor staurosporine did not alter the PAF-induced AA release. The present study provides an example of cells in which the direct activation of PKC by phorbol esters does not lead to a primed and/or enhanced AA release. As a unique example in which PKC activation is neither necessary nor sufficient for AA release to occur, this now allows study of the separate and distinct roles for PLD and PLA2 in signal-transduction processes. This has hitherto been difficult to achieve because of the lack of specific inhibitors of these two phospholipases.
Balsinde
J
J
Department of Chemistry, University of California at San Diego, La Jolla 92093-0601, U.S.A.
Balboa
M A
MA
Insel
P A
PA
Dennis
E A
EA
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D015153:Blotting, Western; D015536:Down-Regulation / Q000502:physiology; D004789:Enzyme Activation / Q000187:drug effects; D007527:Isoenzymes / Q000378:metabolism; D007941:Leukemia P388 / Q000201:enzymology*; D008264:Macrophages / Q000201:enzymology*; D051379:Mice; D010704:Phorbols / Q000494:pharmacology; D010739:Phospholipase D / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D010743:Phospholipids / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism* / Q000494:pharmacology
D007527:Isoenzymes; D010704:Phorbols; D010743:Phospholipids; D016718:Arachidonic Acid; D011493:Protein Kinase C; D010741:Phospholipases A; D054467:Phospholipases A2; D010739:Phospholipase D