Calyculin-A induces focal adhesion assembly and tyrosine phosphorylation of p125(Fak), p130(Cas), and paxillin in Swiss 3T3 cells.
Treatment of intact Swiss 3T3 cells with calyculin-A, an inhibitor of myosin light chain (MLC) phosphatase, induces tyrosine phosphorylation of p125(Fak) in a sharply concentration- and time-dependent manner. Maximal stimulation was 4.2 +/- 2.1-fold (n = 14). The stimulatory effect of calyculin-A was observed at low nanomolar concentrations (<10 nM); at higher concentrations (>10 nM) tyrosine phosphorylation of p125(Fak) was strikingly decreased. Calyculin-A induced tyrosine phosphorylation of p125(Fak) through a protein kinase C- and Ca(2+)-independent pathway. Exposure to either cytochalasin-D or latrunculin-A, which disrupt actin organization by different mechanisms, abolished tyrosine phosphorylation of p125(Fak) in response to calyculin-A. Treatment with high concentrations of platelet-derived growth factor (20 ng/ml) which also disrupt actin stress fibers, completely inhibited tyrosine phosphorylation of p125(Fak) in response to calyculin-A. This agent also induced tyrosine phosphorylation of the focal adhesion-associated proteins p130(Cas) and paxillin. These tyrosine phosphorylation events were associated with a striking increase in the assembly of focal adhesions. The Rho kinase (ROK) inhibitor HA1077 that blocked focal adhesion formation by bombesin, had no effect on the focal adhesion assembly induced by calyculin-A. Thus, calyculin-A induces transient focal adhesion assembly and tyrosine phosphorylation of p125(Fak), p130(Cas), and paxillin, acting downstream of ROK.
Leopoldt
D
D
Department of Medicine, School of Medicine and Molecular Biology Institute, University of California, Los Angeles, CA 90095, USA.
Yee
H F
HF
Rozengurt
E
E
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D016475:3T3 Cells; D000818:Animals; D001839:Bombesin / Q000494:pharmacology; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D050719:Crk-Associated Substrate Protein; D015638:Cytochalasin D / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D003599:Cytoskeleton / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D022001:Focal Adhesions / Q000378:metabolism*; D015151:Immunoblotting; D007211:Indoles / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D008387:Marine Toxins; D051379:Mice; D008856:Microscopy, Fluorescence; D043263:Myosin-Light-Chain Phosphatase; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology*; D051419:Paxillin; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins; D050717:Retinoblastoma-Like Protein p130; D013844:Thiazoles / Q000494:pharmacology; D053778:Thiazolidines; D013997:Time Factors
C493842:Bcar1 protein, mouse; D019086:Bridged Bicyclo Compounds, Heterocyclic; D050719:Crk-Associated Substrate Protein; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D007211:Indoles; D008301:Maleimides; D008387:Marine Toxins; D019384:Nucleic Acid Synthesis Inhibitors; D010080:Oxazoles; D051419:Paxillin; D010750:Phosphoproteins; D011506:Proteins; C496062:Pxn protein, mouse; D050717:Retinoblastoma-Like Protein p130; D013844:Thiazoles; D053778:Thiazolidines; D019000:Phosphotyrosine; D015638:Cytochalasin D; C059041:calyculin A; 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; D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases; D043263:Myosin-Light-Chain Phosphatase; C070515:bisindolylmaleimide I; D001839:Bombesin; C049347:fasudil; C037067:latrunculin A; D002118:Calcium