Phosphorylation of cytoskeleton-associated proteins, pp58 and pp60, in tumouricidal murine peritoneal macrophages.
Two highly phosphorylated vimentin-like proteins, pp58 and pp60, are expressed in macrophages activated in vivo to tumouricidal activity. Resident and elicited, non-tumouricidal peritoneal macrophages displayed low and intermediate levels of phosphorylated pp58 and pp60, respectively. C3H/HeN macrophages became tumouricidal after incubation with 0.1 micrograms/mL A23187 plus 10 nmol/L 12-phorbol 13-myristate acetate (PMA), or 0.1 micrograms/mL A23187 plus 100 ng/mL lipopolysaccharide (LPS), and displayed increased phosphorylation of pp58 and pp60. LPS non-responder C3H/HeJ macrophages were not tumouricidal nor did they show increased phosphorylation of pp58 and pp60 after incubation with LPS plus A23187 in vitro. C3H/HeJ macrophages, however, did become tumouricidal and expressed increased phosphorylation of pp58 and pp60 after incubation with A23187 and PMA. Addition of PGE2 (10(-8) mol/L), resulted in down-regulation of macrophage tumouricidal activity and decreased pp58 and pp60 phosphorylation, which was reversed by addition of indomethacin (10(-6) mol/L) to cultures with PGE2. Phosphorylation increased within 5 min after adding activating stimuli while incorporation of [35S]-methionine into a 58 kD protein did not occur until 6 h later. No 60 kD protein synthesis was detected during the first 8 h after adding activating stimuli, indicating that previously synthesized proteins were phosphorylated during macrophage activation. These results signify a physiological role for the phosphorylation of cytoskeleton-associated pp58 and pp60 during macrophage activation to tumour cytotoxicity.
Didier
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ES
Department of Veterinary Pathobiology, College of Veterinary Medicine, University of Illinois, Urbana 61801.
Fan
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S
Wheeler
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Tompkins
W A
WA
D000818:Animals; D002461:Cell Line, Transformed; D003598:Cytoskeletal Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D003601:Cytotoxicity Tests, Immunologic; D003720:Densitometry; D015232:Dinoprostone / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D007700:Kinetics; D008214:Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008809:Mice, Inbred C3H; D010529:Peritoneal Cavity / Q000166:cytology; D010766:Phosphorylation; D014616:Vaccinia virus / Q000276:immunology
D003598:Cytoskeletal Proteins; D015232:Dinoprostone