Possible involvement of a 95-kDa protein phosphorylation in phorbol 12-myristate 13-acetate-induced suppression of zymosan phagocytosis in guinea pig macrophages.
Recently, we characterized a surface antigen (Z-1) of guinea pig macrophages by monoclonal anti-Z-1 antibody. The Z-1 antigen consists of two different polypeptide chains; alpha (140 kDa) and beta (95 kDa). This antigen is closely correlated with the phagocytic activity of the cells for zymosan and presumably functions as a receptor for zymosan. In the present study, the effect of phorbol 12-myristate 13-acetate (PMA) on the function of Z-1 was examined. Incubation of ortho-[32P]phosphate-labeled macrophages with PMA greatly increased the phosphorylation of the beta subunit of Z-1 but not that of the alpha subunit. Optimal phosphorylation was observed when cells were incubated with 300 ng/ml of PMA for 60-120 min. The PMA-induced phosphorylation was markedly suppressed by treatment of the macrophages with H-7, an inhibitor of protein kinase C. A chemotactic peptide, N-formyl-Met-Leu-Phe (fMLP) also caused phosphorylation of the beta subunit. Unlike PMA, fMLP maximized the phosphorylation within 30 s. Purified Z-1 was an excellent substrate for the exogenously added protein kinase C only in the presence of both Ca2+ and phosphatidylserine. H-7 completely inhibited the in vitro phosphorylation. These data suggest that the beta subunit of Z-1 is phosphorylated by protein kinase C. The phosphorylation of Z-1 by PMA and fMLP coincided with inhibition of zymosan phagocytosis. A linear relationship was obtained between the level of phosphorylation of Z-1 and the degree of inhibition of zymosan phagocytosis induced by PMA. Thus, the results suggest that zymosan uptake is negatively regulated by protein kinase C-mediated phosphorylation of the beta subunit of Z-1.
Hazeki
K
K
Department of Microbiology, Faculty of Pharmaceutical Sciences, Higashi-Nippon-Gakuen University, Ishikari-Tobetsu, Japan.
Tamoto
K
K
Tada
M
M
Mori
Y
Y
D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis / Q000276:immunology*; D000937:Antigen-Antibody Reactions; D000954:Antigens, Surface / Q000032:analysis / Q000276:immunology*; D001345:Autoradiography; D002462:Cell Membrane / Q000187:drug effects; D003720:Densitometry; D006168:Guinea Pigs; D066298:In Vitro Techniques; D008264:Macrophages / Q000187:drug effects* / Q000502:physiology; D008565:Membrane Proteins / Q000032:analysis; D010587:Phagocytosis / Q000187:drug effects*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D015054:Zymosan / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D008565:Membrane Proteins; D015054:Zymosan; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate