Ca2+ signalling mechanisms of the beta(2) integrin on neutrophils: Involvement of phospholipase C gamma 2 and Ins(1,4,5)P-3

Engagement of beta(2) integrins triggers a tyrosine kinase-dependent intracellular mobilization and influx of Ca2+ in human neutrophils. However, the transduction pathway involved in generating this Ca2+ signal is obscure. In the present study we identified phospholipase C gamma 2 (PLC gamma 2) as one of the major proteins that was phosphorylated on tyrosine in response to beta(2) integrin activation. This beta(2) integrin-induced phosphorylation of PLC gamma 2 occurred in parallel with an increased accumulation of Ins(1,4,5)P-3. The relevance of these observations for the beta(2) integrin-induced Ca2+ signal was investigated using an inhibitor of PLC signalling pathways, 1-(6-{[17 beta 3-methoxyoestra-1,3,5(10)-trien-17-yl]amino}hexy) (U73122). U73122 dose-dependently (IC50, approx. 0.15 mu M) inhibited both the beta(2) integrin-induced release af Ca2+ from intracellular stores and the subsequent influx of Ca2+ across the plasma membrane. These effects were not observed with the inactive analogue 1-(6-{[17 beta-3-methoxyoestra-1,3,5(10)-trien yl]amino}hexyl)-pyrrolidine-2,5-dione (U73343). To gain further support for an involvement of PLC-induced Ins(1,4,5)P-3 formation in the beta(2) integrin-induced Ca2+ signal, we searched for the molecular event(s) underlying the effects of U73122. Our experiments revealed that U73122 had no effect on either beta(2) integrin-induced tyrosine phosphorylation of PLC gamma 2 (or any of the other proteins) or on the formation of Ins(1,4,5)P-3, but it reduced the Ins(1,4,5)P-3-induced release of Ca-45(2+) from intracellular stores of electropermeabilized cells. Taken together, the present data suggest that the beta(2) integrin-induced Ca2+ signal in human neutrophils is generated through activation of a PLC gamma 2-dependent pathway.