Bombesin, bradykinin, vasopressin, and phorbol esters rapidly and transiently activate Src family tyrosine kinases in Swiss 3T3 cells. Dissociation from tyrosine phosphorylation of p125 focal adhesion kinase.
Treatment of quiescent Swiss 3T3 cells with bombesin induces a rapid (</=40 s) and transient increase in the kinase activity of the Src family of tyrosine kinases, as determined by autophosphorylation in immune complex kinase assays (4.6 +/- 0.2-fold stimulation, n = 44) and phosphorylation of exogenous substrates. Phorbol 12, 13-dibutyrate increased the activity of Src family kinases with similar kinetics but was less effective than bombesin. However, Src family kinase activation by bombesin is not dependent either on protein kinase C or Ca2+. Bombesin stimulation of Src family kinase activity could also be dissociated from p125 focal adhesion kinase tyrosine phosphorylation. Neither treatment with cytochalasin D nor placement of the cells in suspension prevented the stimulation of Src family kinase activity induced by bombesin, but both abolished bombesin-induced tyrosine phosphorylation of p125 focal adhesion kinase. The stimulation of the Src family kinase activity by bombesin was completely prevented by treatment with vanadate, a potent inhibitor of protein-tyrosine phosphatases. Bradykinin and vasopressin also stimulated Src family kinase activity transiently, and this stimulation was also inhibited by vanadate. Our results dissect two separate pathways that lead to protein tyrosine phosphorylation in neuropeptide-stimulated Swiss 3T3 cells.
Imperial Cancer Research Fund, P.O. Box 123, 44 Lincoln's Inn Fields, London WC2A 3PX, United Kingdom.
J L
JL
Rodríguez-Fernández
E
E
Rozengurt
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001839:Bombesin / Q000494:pharmacology*; D001920:Bradykinin / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004789:Enzyme Activation; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D013379:Substrate Specificity; D014638:Vanadates / Q000494:pharmacology; D014667:Vasopressins / Q000494:pharmacology*; D019061:src-Family Kinases / Q000378:metabolism*
D015815:Cell Adhesion Molecules; D010446:Peptide Fragments; D010703:Phorbol Esters; D014667:Vasopressins; D019000:Phosphotyrosine; D014638:Vanadates; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D019061:src-Family Kinases; D011493:Protein Kinase C; D001839:Bombesin; D001920:Bradykinin; D002118:Calcium; D004121:Dimethyl Sulfoxide