Vanadate-induced activation of cytosolic phospholipase A2alpha in L929 cells: Roles of tyrosine kinase, protein kinase C, and extracellular signal-regulated kinase.
Orthovanadate (Na3VO4), which acts as an inhibitor of protein tyrosine phosphatases, has a various pharmacological effects including the release of arachidonic acid (AA) from cells. We investigated roles of alpha-type cytosolic phospholipase A2 (cPLA2alpha), Src family kinases (Src) and protein kinase C (PKC) in the release of AA induced by Na3VO4 from a murine fibroblast cell line, L929. C12 cells, a variant of L929 that lacks expression of cPLA2alpha, were used along with a clone of C12 cells that are stably expressing cPLA2alpha (C12-cPLA2alpha cells). In the presence of a Ca2+ ionophore (10 microM A23187), 5 and 10mM Na3VO4 synergistically stimulated AA release from L929 and C12-cPLA2alpha cells, and to a much lesser extent from control C12 cells. The release of AA by Na3VO4/A23187 was inhibited by a selective cPLA2alpha inhibitor (3 microM pyrrophenone). The release of AA by Na3VO4/A23187 was significantly inhibited by a PKC inhibitor (10 microM GF109203X), in PKC-depleted cells, by a Src inhibitor (2 microM PP2) and by an inhibitor of extracellular signal-regulated kinase 1/2 (ERK1/2) kinase (10 microM U0126). The phosphorylation of ERK1/2 was stimulated by Na3VO4, and the response was significantly decreased by inhibitors of Src, PKC and ERK1/2 kinase. Our data show that Na3VO4 stimulates AA release largely via cPLA2alpha activation in Ca2+-dependent manner, and the cross-talk between Src and PKC and the ERK-dependent pathways are involved in Na3VO4-induced AA release from L929 cells.
Taniguchi
Tomoko
T
Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba 260-8675, Japan.
Shimizu
Masaya
M
Nakamura
Hiroyuki
H
Hirabayashi
Tetsuya
T
Fujino
Hiromichi
H
Saito
Takeshi
T
Murayama
Toshihiko
T
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D066246:ErbB Receptors / Q000502:physiology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology*; D006861:Hydrogen Peroxide / Q000378:metabolism; D051379:Mice; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D011493:Protein Kinase C / Q000502:physiology*; D021381:Protein Transport / Q000187:drug effects; D015398:Signal Transduction; D014638:Vanadates / Q000494:pharmacology*; D019061:src-Family Kinases / Q000502:physiology*
D016718:Arachidonic Acid; D014638:Vanadates; D006861:Hydrogen Peroxide; D066246:ErbB Receptors; D019061:src-Family Kinases; D011493:Protein Kinase C; D048049:Extracellular Signal-Regulated MAP Kinases; D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium