Internalization of N-formyl peptide chemotactic receptor-ligand complex by human neutrophils. The role of the receptor's 2-kDa external domain and carbohydrates.
We treated human neutrophils with papain to remove the external 2-kDa domain and along with it the two oligosaccharide side chains of the N-formyl peptide chemotactic receptor and investigated what effect their absence has on the ligand-receptor complex internalization. After prelabeling of the cells with 125I-hexapeptide for 5 min at 22 degrees C, about 95% of the bound radioactivity was located on the cell surface. During the first 5-min incubation at 37 degrees C both the control and papain-treated cells internalized 73% of the receptor-ligand complexes suggesting that internalization is very rapid in human neutrophils and that removal of the external domain and the carbohydrates of the receptor does not affect the rate. However, the truncated receptor-ligand complexes were degraded at a faster rate because the radioactivity released into the medium was significantly higher and correspondingly the acid-resistant radioactivity significantly lower in the papain-treated neutrophils than in control cells already at 5 min and all subsequent time points. The radioactivity accumulated in the medium of the control and papain-treated neutrophils represented inactivated 125I-hexapeptide as less than 5% of it at 5, 30 and 120 min were capable of rebinding. No receptor recycling was detected in either cells. These results indicate that removal of the 2-kDa external domain and the carbohydrates of the N-formyl chemotactic receptors has little effect on the internalization rate of the receptor-ligand complexes but accelerates markedly their intracellular degradation.
Remes
J
J
Biocenter, University of Oulu, Finland.
Petäjä-Repo
U
U
Rajaniemi
H
H
D000595:Amino Acid Sequence; D006801:Humans; D008969:Molecular Sequence Data; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000378:metabolism*; D009504:Neutrophils / Q000378:metabolism*; D009844:Oligosaccharides / Q000737:chemistry*; D010206:Papain; D017434:Protein Structure, Tertiary; D011869:Radioligand Assay; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic / Q000378:metabolism*; D018000:Receptors, Peptide / Q000378:metabolism*
D009844:Oligosaccharides; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic; D018000:Receptors, Peptide; D009240:N-Formylmethionine Leucyl-Phenylalanine; D010206:Papain