Chemotactic factors induced vimentin phosphorylation in rabbit peritoneal neutrophil.
Rabbit peritoneal neutrophils were reacted for 5-10 s with the chemotactic factors, fMet-Leu-Phe or (5S),(12R)-dihydroxy-6,8,11,14-(cis,trans,trans,cis)-eicosatetraenoic acid and then lysed with a solution of Triton X-100 and [gamma-32P]ATP. They showed an enhanced incorporation of 32P in Mr = 60,000- and 67,000-dalton polypeptides compared to control cells treated similarly. Another chemotactic factor, C5a, produced a similar but much lesser effect. The enhancement was not affected by the addition of the purified catalytic subunit of cAMP-dependent protein kinase, the inhibitor of cAMP-dependent protein kinase, or CaCl2, suggesting that the effect was not mediated by a cAMP-dependent or a Ca2+-dependent protein kinase. When analyzed by two-dimensional gel electrophoresis, the Mr = 60,000 phosphoprotein contained several phosphoproteins with different isoelectric points. The isoelectric point and molecular weight of one of them was similar to those of the intermediate filament protein, vimentin, purified from Chinese hamster ovary cells. Addition of the purified Chinese hamster ovary vimentin and [gamma-32P]ATP to the Triton X-100 lysate of fMet-Leu-Phe-treated neutrophils resulted as an enhanced incorporation of 32P into the vimentin. Addition of fMet-Leu-Phe to 32P-labeled intact neutrophils also enhanced incorporation of 32P into the vimentin of neutrophils. The results suggest that chemotactic factors stimulate vimentin phosphorylation in rabbit neutrophils.
D000818:Animals; D002460:Cell Line; D003182:Complement C5 / Q000494:pharmacology*; D015936:Complement C5a; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D007381:Intermediate Filament Proteins / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000378:metabolism*; D010053:Ovary; D010750:Phosphoproteins / Q000302:isolation & purification; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D011817:Rabbits; D014746:Vimentin
D003182:Complement C5; D007381:Intermediate Filament Proteins; D010750:Phosphoproteins; D014746:Vimentin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015936:Complement C5a; D011494:Protein Kinases