Bombesin and epidermal growth factor stimulate the mitogen-activated protein kinase through different pathways in Swiss 3T3 cells.
Both bombesin and epidermal growth factor (EGF) are potent mitogens in Swiss 3T3 cells that nonetheless have dissimilar receptor structures. To explore possible common intracellular events involved in the stimulation of cellular growth by these two peptides, we have evaluated the regulation of the mitogen-activated protein (MAP) kinase. Exposure of Swiss 3T3 cells to bombesin, EGF or the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) causes the rapid and transient stimulation of the enzyme activity. Pretreatment of cells with the protein kinase inhibitor H-7, or down-regulation of cellular protein kinase C by prolonged exposure to PMA, causes a decrease of over 90% in the activation of MAP kinase by bombesin. In contrast, these treatments have no effect on the stimulation of MAP kinase by EGF. The stimulation of MAP kinase activity by bombesin is dose-dependent, occurring over a narrow concentration range of the peptide. Both EGF and bombesin stimulate the phosphorylation of an immunoprecipitable MAP kinase protein migrating at 42 kDa on SDS/PAGE. Phosphoamino acid analysis of this phosphorylated protein reveals that EGF and bombesin stimulate phosphorylation on tyrosine, threonine and serine residues. Tyrosine phosphorylation of the enzyme, as evaluated by antiphosphotyrosine blotting of the immunoprecipitated protein, reveals that the time course of phosphorylation by both mitogens correlates with stimulation of enzyme activity. These results provide further evidence for the convergence of discrete pathways emanating from tyrosine kinase and G-protein-linked receptors in the regulation of MAP kinase.
Department of Physiology, University of Michigan, School of Medicine, Ann Arbor 48109.
L
L
Pang
S J
SJ
Decker
A R
AR
Saltiel
D016475:3T3 Cells; D000818:Animals; D001839:Bombesin / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D015536:Down-Regulation; D004789:Enzyme Activation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D051379:Mice; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013912:Threonine / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D013912:Threonine; D014443:Tyrosine; D012694:Serine; D004815:Epidermal Growth Factor; D011494:Protein Kinases; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D013755:Tetradecanoylphorbol Acetate; D001839:Bombesin