title: Endothelins increase tyrosine phosphorylation of astrocytic focal adhesion kinase and paxillin accompanied by their association with cytoskeletal components.
abstract: Astrocytic endothelin receptors are involved in the appearance of activated astrocytes upon injury of the brain [Ishikawa N. et al. (1997) Eur. J. Neurosci. 9, 895-901; Koyama Y. et al. (1999) Glia 26, 268-271]. To clarify signal transduction triggered by endothelin receptors, we examined the effects of endothelins on protein tyrosine phosphorylation in cultured rat astrocytes. Endothelin-1 (1 nM) increased tyrosine phosphorylation of focal adhesion kinase and paxillin. The tyrosine phosphorylation was also induced by endothelin-1 (1 nM) and Ala(1,3,11,15)-endothelin-1 (10nM), an endothelin-B receptor agonist. BQ788 (100 nM), an endothelin-B receptor antagonist, inhibited the effects of endothelin-3. Orthovanadate (VO(4)(3-)), a tyrosine phosphatase inhibitor, but not bradykinin (1 microM), angiotensin II (100 nM), A23187 (5 microM) and phorbol 12-myristate 13-acetate (100 nM), increased tyrosine phosphorylation of focal adhesion kinase and paxillin. The tyrosine phosphorylation by endothelin-3 was not prevented by pertussis toxin, Ca(2+) chelation, protein kinase C inhibitors (calphostin C and staurosporine) or wortmannin. Immunocytochemical staining showed that endothelin-3 and VO(4)(3-) induced redistribution of focal adhesion kinase and paxillin to focal adhesions concomitant with stress fiber formation in dibutyryl cyclic-AMP-treated astrocytes. Treatment with endothelin-3 and VO(4)(3-) increased focal adhesion kinase and paxillin associated with astrocytic cytoskeletal fraction. In the presence of cytochalasin B, an actin disrupting agent, endothelin-3 and VO(4)(3-) did not phosphorylate focal adhesion kinase and paxillin. Application of cytochalasin B after treatment with endothelin-3 and VO(4)(3-) stimulated dephosphorylation of focal adhesion kinase and paxillin. These results suggest that the associations of focal adhesion kinase and paxillin with cytoskeletal components are required in the endothelin-induced tyrosine phosphorylation of the astrocytic proteins.
authors.affiliation: Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical Sciences, Osaka University, Yamada-Oka 1-6, Suita 565-0871, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Koyama
authors.forename: Y
authors.initials: Y
authors.lastname: Yoshioka
authors.forename: H
authors.initials: H
authors.lastname: Hashimoto
authors.forename: T
authors.initials: T
authors.lastname: Matsuda
authors.forename: A
authors.initials: A
authors.lastname: Baba
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D001920:Bradykinin / Q000494:pharmacology; D003994:Bucladesine / Q000494:pharmacology; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D002614:Chelating Agents / Q000494:pharmacology; D003571:Cytochalasin B / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000187:drug effects* / Q000378:metabolism; D003599:Cytoskeleton / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1 / Q000378:metabolism / Q000494:pharmacology; D019334:Endothelin-3 / Q000378:metabolism / Q000494:pharmacology; D016232:Endothelins / Q000378:metabolism* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D051419:Paxillin; D037342:Pertussis Toxin; D010750:Phosphoproteins / Q000187:drug effects* / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011505:Protein-Tyrosine Kinases / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D017466:Receptors, Endothelin / Q000819:agonists; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014443:Tyrosine / Q000187:drug effects* / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D002614:Chelating Agents; D003598:Cytoskeletal Proteins; D065128:Endothelin Receptor Antagonists; D019332:Endothelin-1; D019334:Endothelin-3; D016232:Endothelins; D004791:Enzyme Inhibitors; D051419:Paxillin; D010750:Phosphoproteins; C496063:Pxn protein, rat; D017466:Receptors, Endothelin; D010566:Virulence Factors, Bordetella; D000804:Angiotensin II; D003571:Cytochalasin B; D014638:Vanadates; D014443:Tyrosine; D003994:Bucladesine; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496052:Ptk2 protein, rat; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D001920:Bradykinin; D000077191:Wortmannin