The regulation by phosphorylation of ‘priming’ of phospholipase A 2 activity in the neutrophil model system, differentiated HL60 cells

1 Differentiated HL60 cells have been utilized as a model system to examine the ‘priming’ of neutrophil phospholipase A 2 activity. In control cells activation of phospholipase A 2 by a 5 min stimulation with the chemotactic peptide formyl‐methionyl‐leucyl‐phenylalanine (100 n M ) was essentially undetectable. When cells were primed by preincubation with 5 μ M cytochalasin B for 5 min arachidonate release, a measure of phospholipase A 2 activation, was observed within 20 s. 2 Priming by cytochalasin B did not involve or require a change in intracellular free calcium concentration. 3 Priming was associated with an increase in general protein tyrosine phosphorylation and could also be induced by the receptor tyrosine kinase agonist granulocyte macrophage colony‐stimulating factor (GM‐CSF, 20 ng ml −1 ) and be mimicked by treatment with the phosphotyrosine phosphatase inhibitor perhydrovanadate (0.5 m M ). However, an increase in MAP kinase activity was not involved in the priming process. 4 Western blot analysis demonstrated that phospholipase A 2 was phosphorylated in both control and primed cells, but that an increase in the amount of membrane associated enzyme was found in the primed cells. 5 Thus priming appears to be due to membrane association of the phospholipase and this may be regulated by tyrosine kinase activities.British Journal of Pharmacology (1997) 122 , 13–20; doi: 10.1038/sj.bjp.0701323