Bombesin, vasopressin, and endothelin stimulation of tyrosine phosphorylation in Swiss 3T3 cells. Identification of a novel tyrosine kinase as a major substrate.
Neuropeptide-stimulated tyrosine phosphorylation of specific components in Swiss 3T3 cells was investigated using monoclonal antibodies directed against the src transformation-associated substrates p125 focal adhesion kinase (FAK), a novel type of cytosolic tyrosine kinase, and p130. Treatment of Swiss 3T3 cells with the mitogenic peptides bombesin, vasopressin, and endothelin caused a striking increase in the tyrosine phosphorylation of p125FAK, as judged either by anti-phosphotyrosine (anti-Tyr(P)) Western blots of anti-p125FAK immunoprecipitates, or by anti-p125FAK immunoblots of anti-Tyr(P) immunoprecipitates. Bombesin-stimulated tyrosine phosphorylation of p125FAK was detectable within seconds and concentration-dependent (half-maximum effect of 0.3 nM). Neuropeptides also stimulated the tyrosine phosphorylation of a second component of M(r) 130,000, previously identified as the major p130 phosphotyrosyl protein in src-transformed cells. Bombesin stimulated p130 tyrosine phosphorylation with kinetics and concentration dependence similar to those observed for p125FAK. This is the first report to identify substrates for neuropeptide-stimulated tyrosine phosphorylation; the finding that one of these substrates is a tyrosine kinase suggests the existence of a novel signal transduction pathway in the action of mitogenic neuropeptides.
Zachary
I
I
Imperial Cancer Research Fund, London, United Kingdom.
Sinnett-Smith
J
J
Rozengurt
E
E
D016475:3T3 Cells; D000818:Animals; D000911:Antibodies, Monoclonal; D001839:Bombesin / Q000494:pharmacology*; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D016232:Endothelins / Q000494:pharmacology*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D051379:Mice; D008970:Molecular Weight; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism; D014667:Vasopressins / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D015815:Cell Adhesion Molecules; D016232:Endothelins; D010750:Phosphoproteins; D014667:Vasopressins; D019000:Phosphotyrosine; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D001839:Bombesin