title: Divergent modulation of Rho-kinase and Ca(2+) influx pathways by Src family kinases and focal adhesion kinase in airway smooth muscle.
abstract: OBJECTIVE
  The importance of tyrosine kinases in airway smooth muscle (ASM) contraction is not fully understood. The aim of this study was to investigate the role of Src-family kinases (SrcFK) and focal adhesion kinase (FAK) in GPCR-mediated ASM contraction and associated signalling events.
  
  
  METHODS
  Contraction was recorded in intact or α-toxin permeabilized rat bronchioles. Phosphorylation of SrcFK, FAK, myosin light-chain-20 (MLC20 ) and myosin phosphatase targeting subunit-1 (MYPT-1) was evaluated in cultured human ASM cells (hASMC). [Ca(2+) ]i was evaluated in Fura-2 loaded hASMC. Responses to carbachol (CCh) and bradykinin (BK) and the contribution of SrcFK and FAK to these responses were determined.
  
  
  RESULTS
  Contractile responses in intact bronchioles were inhibited by antagonists of SrcFK, FAK and Rho-kinase, while after α-toxin permeabilization, they were sensitive to inhibition of SrcFK and Rho-kinase, but not FAK. CCh and BK increased phosphorylation of MYPT-1 and MLC20 and auto-phosphorylation of SrcFK and FAK. MYPT-1 phosphorylation was sensitive to inhibition of Rho-kinase and SrcFK, but not FAK. Contraction induced by SR Ca(2+) depletion and equivalent [Ca(2+) ]i responses in hASMC were sensitive to inhibition of both SrcFK and FAK, while depolarization-induced contraction was sensitive to FAK inhibition only. SrcFK auto-phosphorylation was partially FAK-dependent, while FAK auto-phosphorylation was SrcFK-independent.
  
  
  CONCLUSIONS
  SrcFK mediates Ca(2+) -sensitization in ASM, while SrcFK and FAK together and individually influence multiple Ca(2+) influx pathways. Tyrosine phosphorylation is therefore a key upstream signalling event in ASM contraction and may be a viable target for modulating ASM tone in respiratory disease.
authors: Division of Asthma, Allergy and Lung Biology, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Yasin
authors: Y
authors: Shaifta
authors: Division of Asthma, Allergy and Lung Biology, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Nneka
authors: N
authors: Irechukwu
authors: Division of Asthma, Allergy and Lung Biology, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Jesus
authors: J
authors: Prieto-Lloret
authors: Division of Asthma, Allergy and Lung Biology, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Charles E
authors: CE
authors: MacKay
authors: Division of Asthma, Allergy and Lung Biology, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Keisha A
authors: KA
authors: Marchon
authors: Division of Asthma, Allergy and Lung Biology, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Jeremy P T
authors: JP
authors: Ward
authors: Division of Asthma, Allergy and Lung Biology, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Greg A
authors: GA
authors: Knock
mesh_terms: D000328:Adult; D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D055745:Bronchioles / Q000166:cytology / Q000502:physiology*; D016085:Bronchoconstrictor Agents / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D005260:Female; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000502:physiology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000502:physiology; D017208:Rats, Wistar; D014132:Trachea / Q000187:drug effects / Q000502:physiology; D055815:Young Adult; D054460:rho-Associated Kinases / Q000502:physiology*; D019061:src-Family Kinases / Q000502:physiology*
chemical_list: D016085:Bronchoconstrictor Agents; D002217:Carbachol; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019061:src-Family Kinases; D054460:rho-Associated Kinases; D001920:Bradykinin; D002118:Calcium