title: Stimulation of mitogen-activated protein kinase activity by different secretory stimuli in rat basophilic leukemia cells.
abstract: Ag-induced cross-linking of IgE bound to its high affinity receptor (Fc epsilon RI) at the surface of basophils or mast cells triggers a number of biochemical events culminating in the release of several inflammatory mediators. In rat basophilic leukemia (RBL-2H3) cells expressing the G protein-coupled m1 muscarinic receptor, Ag/IgE-induced cross-linking of Fc epsilon RI, calcium ionophore A23187, and carbachol through M1 receptors stimulated tyrosine phosphorylation of several proteins, including two of 42 and 44 kDa. Proteins of identical molecular masses were recognized by anti-MAP-kinase antibodies, and these immunoreactive proteins exhibited in part a slightly increased molecular mass on SDS polyacrylamide gels after incubation of cells with secretory stimuli. All stimuli led to the activation of MAP kinase, which co-purified on Mono Q chromatography with 42- and 44-kDa proteins, which were tyrosine phosphorylated in response to secretory stimuli and reacted with anti-(MAP kinase) antibodies. Finally, 42- and 44-kDa proteins immunoprecipitated by anti-MAP-kinase antibodies and anti-phosphotyrosine antibodies were recognized by anti-phosphotyrosine and anti-MAP-kinase antibodies, respectively. Primarily threonine and tyrosine residues were found to be phosphorylated in 42- and 44-kDa proteins immunoprecipitated from [32P]phosphate-labeled cells that had been treated with secretory stimuli. The dose dependence of secretagogue-induced MAP kinase activation correlated with that of increases in serotonin release from activated cells, and the maximum of MAP kinase activation coincided with the maximum rate of secretion. Down-regulation or inhibition of protein kinase C as well as incubation of cells with the tyrosine kinase inhibitor genistein markedly inhibited MAP kinase activation in parallel with serotonin release. Taken together, these findings demonstrate that 42- and 44-kDa MAP kinases are activated in response to secretory stimuli and provide some evidence for a functional link between MAP kinase activation and signaling events leading to mediator release in RBL cells.
authors.affiliation: Institut für Pharmakologie, Freie Universität Berlin, FRG.
authors.forename: S
authors.initials: S
authors.lastname: Offermanns
authors.forename: S V
authors.initials: SV
authors.lastname: Jones
authors.forename: E
authors.initials: E
authors.lastname: Bombien
authors.forename: G
authors.initials: G
authors.lastname: Schultz
mesh_terms: D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D015550:Cell Degranulation / Q000502:physiology*; D004789:Enzyme Activation; D015471:Leukemia, Basophilic, Acute; D019950:Mitogen-Activated Protein Kinase 1; D008934:Mitogens / Q000494:pharmacology; D008970:Molecular Weight; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D051381:Rats; D017455:Receptors, IgE / Q000502:physiology; D012701:Serotonin / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000378:metabolism
chemical_list: D008934:Mitogens; D017455:Receptors, IgE; D012701:Serotonin; D000001:Calcimycin; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1