Involvement of Lyn, src-family tyrosine kinase, in the activation of phospholipase D by anti-Fas monoclonal antibody in A20 cells

초록

The activation of phospholipase D (PLD) is regulated by various factors, such as protein kinase C (PKC), small GTPase, non-receptor tyrosine kinases and other signaling molecules. We previously reported (not published) that D609-sensitive tyrosine phospholylation was involved in Fas-induced PLD activation. Briefly, both PLD activation and tyrosine kinase activation increased by anti-Fas monoclonal antibody (Jo2) were inhibited by D609. Although it was confirmed that D609 blocked both phosphatidyl choline specific-phospholipase C (PC-PLC) and tyrosine kinase, the tyrosine kinase that involved in PLD activation was not identified. In this study, the tyrosine kinase that involved in PLD activation was examined. We selected Src-family tyrosine kinase such as Lyn, Fyn, Lck, Hck, and Src, and extracellular regulatory kinase Erk1/2(p42/p44). To determine the involvement of tyrosine kinase in PLD activation, we used immunoprecipitation, westernblot analysis, and PLD assay. Phosphorylation of each tyrosine kinase was calculated by densitometry analysis. We found that Fas cross-linking increased the tyrosine phosphorylation only in the Lyn among the kinases. Tyrosine phospholylation induced by Fas activation in Lyn was inhibited by pretreatment with PP2 and D609, respectively. Also PP2 and D609 arrested PLD activity increased by Fas cross-linking, respectively. We concluded that Lyn was involved PLD activation by Jo2 in A20 cells.