Tumor necrosis factor-induced activation and increased tyrosine phosphorylation of mitogen-activated protein (MAP) kinase in human fibroblasts.
Tumor necrosis factor (TNF) is a pleiotropic cytokine whose many demonstrated actions include effects on cell growth and differentiation. TNF treatment of cells is known to lead to a rapid increase in serine/threonine phosphorylation of many cellular proteins, but the kinases responsible remain largely unidentified. We show that TNF treatment induces a rapid and transient increase in mitogen-activated protein kinase (MAPK) activity in the human diploid FS-4 cell line, for which TNF is known to be mitogenic. TNF-induced activation of MAPK was demonstrated by its enhanced ability to phosphorylate myelin basic protein in vitro and by a characteristic shift in the electrophoretic mobility of MAPK proteins. MAPK activation was accompanied by a significant increase of MAPK phosphorylation on tyrosine residues, which was demonstrated by 32P labeling of cells and isolation of the labeled proteins after immunoprecipitation with antibodies to phosphotyrosine, and by direct immunoblotting of SDS-polyacrylamide gel electrophoresis-fractionated unlabeled cell lysates with antibodies to phosphotyrosine. The pp42 and pp44 MAPK were the only proteins whose tyrosine phosphorylation was demonstrably increased in FS-4 cells after TNF treatment. MAPK activation is likely to represent an important component in the cascade of signals that link TNF receptors to various TNF-elicited cellular responses.
Vietor
I
I
Department of Microbiology, New York University Medical Center, New York 10016.
Schwenger
P
P
Li
W
W
Schlessinger
J
J
Vilcek
J
J
D015153:Blotting, Western; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D005347:Fibroblasts / Q000201:enzymology*; D006801:Humans; D015151:Immunoblotting; D007163:Immunosorbent Techniques; D010766:Phosphorylation; D019000:Phosphotyrosine; D011494:Protein Kinases / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
D014409:Tumor Necrosis Factor-alpha; D019000:Phosphotyrosine; D014443:Tyrosine; D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases