Protein kinase Calpha-dependent increase in Ca2+-independent phospholipase A2 in membranes and arachidonic acid liberation in zymosan-stimulated macrophage-like P388D1 cells.
We previously reported that zymosan-stimulated, protein kinase C (PKC)-dependent arachidonic acid liberation occurs with association of Ca2+-independent phospholipase A2 (iPLA2) with the membranes of macrophage-like P388D1 cells. In the present study, the possible involvement of PKC isoforms (alpha, beta, delta, and epsilon) on the increase in iPLA2 was examined. Stimulation of P388D1 cells with zymosan induced increases in iPLA2 activity and protein in the membranes and liberation of arachidonic acid. In the stimulated cells, PKCalpha, PKCdelta, and PKCepsilon, but not PKCbeta, were increased in the membranes. The zymosan-induced increase in iPLA2 activity was suppressed by pretreatment with 4beta-phorbol 12-myristate 13-acetate for 10 hr, by which PKCalpha and PKCdelta, but not PKCbeta and PKCepsilon, were depleted, and by Gö6976, a PKCalpha inhibitor, but not rottlerin, a PKCdelta inhibitor. The zymosan-induced release of arachidonic acid was also reduced by the PKC depletion and Gö6976. However, stimulation with 4beta-phorbol 12-myristate 13-acetate alone did not increase iPLA2 activity in the membranes. Furthermore, the depletion of intracellular Ca2+ also impaired the zymosan-induced increase in iPLA2 activity in the membranes. However, no increase in iPLA2 activity was observed upon stimulation with Ca2+-mobilizing agents (ionomycin or thapsigargin). Cytochalasin D, an inhibitor of actin polymerization, suppressed the zymosan-induced increases in iPLA2 activity and protein in the membranes and the release of arachidonic acid. These results suggest that zymosan stimulates an increase in iPLA2 in the membranes of P388D1 cells probably through activation of PKCalpha in concert with cytochalasin D-sensitive events.
Akiba
Satoshi
S
Department of Pathological Biochemistry, Kyoto Pharmaceutical University, Misasagi, Yamashina-ku, 607-8414, Kyoto, Japan. akiba@mb.kyoto-phu.ac.jp
Ohno
Satoshi
S
Chiba
Masahiro
M
Kume
Keisuke
K
Hayama
Misako
M
Sato
Takashi
T
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D015638:Cytochalasin D / Q000494:pharmacology; D007527:Isoenzymes / Q000378:metabolism*; D008264:Macrophages / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D051379:Mice; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D015054:Zymosan / Q000494:pharmacology*
D007527:Isoenzymes; D019384:Nucleic Acid Synthesis Inhibitors; D015638:Cytochalasin D; D016718:Arachidonic Acid; D015054:Zymosan; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium