Y-27632, an inhibitor of Rho-associated kinases, prevents tyrosine phosphorylation of focal adhesion kinase and paxillin induced by bombesin: dissociation from tyrosine phosphorylation of p130(CAS).
A rapid increase in tyrosine phosphorylation of focal adhesion kinase (FAK), paxillin, and Crk-associated substrate (CAS) are prominent early events triggered by many G protein-coupled receptors (GPCRs), but the mechanisms involved remain unclear. Here, we examined whether the Rho-associated protein serine/threonine kinase family (ROCK) is a critical Rho effector in the pathway that links GPCR activation to the tyrosine phosphorylation of FAK, CAS, and paxillin. Treatment of Swiss 3T3 cells with Y-27632, a preferential inhibitor of ROCK, dramatically inhibited the formation of actin stress fibers, the assembly of focal contacts, and the increase in tyrosine phosphorylation of FAK and paxillin induced by bombesin in these cells. Surprisingly, we found that treatment with Y-27632 did not produce any detectable effect on bombesin-elicited CAS tyrosine phosphorylation even at the highest concentrations of Y-27632 tested. HA-1077, a preferential inhibitor of ROCK activity structurally unrelated to Y-27632, also attenuated the increase in the tyrosine phosphorylation of FAK and paxillin but did not affect the tyrosine phosphorylation of CAS induced by bombesin in Swiss 3T3 cells. The results demonstrate that ROCK-dependent tyrosine phosphorylation of FAK and paxillin can be dissociated from a ROCK-independent pathway leading to tyrosine phosphorylation of CAS.
Sinnett-Smith
J
J
Department of Medicine, School of Medicine and Molecular Biology Institute, University of California, Los Angeles, 90095-1786, USA.
Lunn
J A
JA
Leopoldt
D
D
Rozengurt
E
E
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D016475:3T3 Cells; D000577:Amides / Q000494:pharmacology*; D000818:Animals; D001839:Bombesin / Q000494:pharmacology*; D050719:Crk-Associated Substrate Protein; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D022001:Focal Adhesions / Q000187:drug effects / Q000648:ultrastructure; D047908:Intracellular Signaling Peptides and Proteins; D008246:Lysophospholipids / Q000494:pharmacology; D051379:Mice; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins; D011725:Pyridines / Q000494:pharmacology*; D050717:Retinoblastoma-Like Protein p130; D022502:Stress Fibers / Q000187:drug effects / Q000648:ultrastructure; D054460:rho-Associated Kinases
D000577:Amides; C493842:Bcar1 protein, mouse; D050719:Crk-Associated Substrate Protein; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D008246:Lysophospholipids; D051419:Paxillin; D010750:Phosphoproteins; D010982:Platelet-Derived Growth Factor; D011506:Proteins; C496062:Pxn protein, mouse; D011725:Pyridines; D050717:Retinoblastoma-Like Protein p130; C108830:Y 27632; D019000:Phosphotyrosine; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D001839:Bombesin; C049347:fasudil