Abstract
Experiments were performed to examine how human granulocytes, stimulated by N‐formyl‐chemotactic peptides, process the N‐formyl peptide receptor. One percent of the surface N‐formyl‐chemotactic peptide receptors of purified human granulocytes were covalently, specifically, and radioactively labeled at 4°C using the photochemically reactive N‐formyl‐chemotactic hexapeptide CHO‐Nle‐Leu‐Phe‐Nle‐[^l25^I] Tyr‐N^°^(6‐(4′‐azido‐2′‐nitrophenyl‐amino)hexanoyl)‐Lys. After incubation in the presence of 500 nM of N‐formyl‐Met‐Leu‐Phe at 37°C, the cells were lysed and fractionated by isopycnic surcrose density gradient sedimentation. Receptor‐associated radioactivity cosedimented with plasma membrane in fractions from cells kept at 4°C or incubated at 37°C for 2 min or less. Fractionation of cells incubated at 37°C for longer times revealed that the radioactivity sedi‐mented to lower densities coincident with Golgi markers and the site of noncovalently bound and internalized formyl‐chemotactic peptide. To follow the redistribution of unoccupied receptors, human granulocytes were stimulated with 500 nM N‐formyl‐Met‐Leu‐Phe at 37°C for 5 min, washed, lysed by N~2~ cavitation, and fractionated by rate zonal sucrose density gradient sedimentation. Compared to unstimulated controls the specific binding of N‐formyl‐Met‐Leu‐[^3^H]Phe decreased 76% ± 9% in plasma membrane fractions. N‐formyl‐Met‐Leu‐[^3^H]Phe‐binding activity associated with an intracellular pool cosedimenting with specific granules remained unchanged. Approximately 20% of the activity lost in the plasma membrane could be accounted for by a redistribution of specific N‐formyl‐Met‐Leu‐Phe binding to fractions enriched in azurophil granules. We conclude that the receptor is the carrier in the internalization of the N‐formyl‐chemotactic peptides to a Golgi‐enriched fraction and hypothesize that after a short residency in this fraction, the receptor may dissociate from the ligand and pass onto a fraction consedimenting with dense granules.