REDUCTION BY INHIBITORS OF MONO(ADP-RIBOSYL)TRANSFERASE OF CHEMOTAXISIN HUMAN NEUTROPHIL LEUKOCYTES BY INHIBITION OF THE ASSEMBLY OF FILAMENTOUS ACTIN

1 Chemotaxis of human neutrophils is mediated by numerous agents [e.g. N-formyl-methionyl-leucyl-phenylalanine (FMLP) and platelet activatin g factor (PAF)] whose receptors are coupled to phospholipase C. Howeve r, the subsequent transduction pathway mediating cell movement remains obscure. We now propose involvement of mono(ADP-ribosyl)transferase a ctivity in receptor-dependent chemotaxis. 2 Human neutrophils were iso lated from whole blood and measurements were made of FMLP or PAF-depen dent actin polymerization and chemotaxis. The activity of cell surface Arg-specific mono(ADP-ribosyl)transferase was also measured. Each of these activities was inhibited by vitamin K-3 and similar IC50 values obtained (4.67 +/- 1.46 mu M, 2.0 +/- 0.1 mu M and 4.7 +/- 0.1 mu M re spectively).3 There were similar close correlations between inhibition of (a) enzyme activity and (b) actin polymerization or chemotaxis by other known inhibitors of mono(ADP-ribosyl)transferase, namely vitamin K-1, novobiocin, nicotinamide and the efficient pseudosubstrate, diet hylamino(benzylidineamino)guanidine (DEA-BAG). 4 Intracellular Ca2+ wa s measured by laser scanning confocal microscopy with two fluorescent dyes (Fluo-3 and Fura-Red). Exposure of human neutrophils to FMLP or P AF was followed by transient increases in intracellular Ca2+ concentra tion, but the inhibitors of mono(ADP-ribosyl)transferase listed above had no effect on the magnitude of the response. 5 A panel of selective inhibitors of protein kinase C, tyrosine kinase, protein kinases A an d G or phosphatases 1 and 2A showed no consistent inhibition of FMLP-d ependent polymerization of actin. 6 We conclude that eukaryotic Arg-sp ecific mono(ADP-ribosyl)transferase activity may be implicated in the transduction pathway mediating chemotaxis of human neutrophils, with i nvolvement in the assembly of actin-containing cytoskeletal microfilam ents.