We investigated the role of protein synthesis in human polymorphonuclear leukocyte (PMN) chemotaxis and luminol-dependent chemiluminescence (CL). We used cycloheximide and puromycin to inhibit protein synthesis, and determined the extent of synthesis by measurement of 14C-amino acid incorporation. With 2-hour incubations both puromycin at 9.0 X 10(-6) M and cycloheximide at 1.8 X 10(-6) M inhibited PMN protein synthesis. At concentrations of 2-4 X 10(-5) M both cycloheximide and puromycin inhibited PMN-chemotaxis 40 and 55%, respectively. However, inhibition was observed only when using zymosan-activated serum and not formyl-methionyl-phenylalanine as a chemoattractant. Using 2-hour incubations, PMN-CL was also suppressed by puromycin and cycloheximide, at 20 and 40%, respectively. The data demonstrated that protein synthesis had an important role in chemotaxis that was dependent on the chemoattractant and perhaps the cellular receptor involved in that process. CL did not require de novo protein synthesis but appeared to depend on protein(s) with a relatively rapid turnover.