Selective involvement of p130Cas/Crk/Pyk2/c-Src in endothelin-1-induced JNK activation.
Both integrin-based focal adhesion complexes and receptor tyrosine kinases have been proposed as scaffolds on which the G protein-coupled receptor (GPCR)-induced signaling complex might assemble. We have recently reported that Ca2+-sensitive tyrosine kinase, Pyk2, and epidermal growth factor receptor (EGFR) act as independently regulated scaffolds in cardiomyocytes. In this report, we investigated the activation and regulation of p130Cas, Crk, Pyk2, and c-Src by a well-known hypertrophic agonist, endothelin-1 (ET), and determined their contributions to the activation of c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) in cardiomyocytes. Like Pyk2, ET-induced tyrosine phosphorylation of p130Cas was significantly inhibited by either chelating intracellular Ca2+ ([Ca2+]i) or a protein kinase C inhibitor, calphostin C. This activation of p130Cas was also abrogated by the tetrapeptide RGDS, which disrupts integrin heterodimerization; cytochalasin D, which depolymerizes the actin cytoskeleton; or a selective Src family kinase inhibitor, PP2, but not by an EGFR inhibitor, AG1478. We also observed ET-induced temporal associations of Pyk2 with active c-Src, followed by p130Cas with Pyk2, c-Src, and Crk. Overexpression of a dominant-negative mutant of p130Cas (CasDeltaSD), Crk (CrkSH2m), Pyk2 (PKM), or C-terminal Src kinase (Csk), but not of a deletion mutant of EGFR (533delEGFR), attenuated ET-induced JNK activation. Similarly, an ET-induced increase in c-jun promoter luciferase activity was inhibited by overexpression of CasDeltaSD, CrkSH2m, PKM, or Csk. In contrast, ET-induced ERK activation and c-fos gene expression were predominantly regulated by EGFR. Collectively, the focal adhesion-dependent p130Cas/Crk/Pyk2/c-Src-mediated pathway is selectively involved in ET-induced JNK activation in cardiomyocytes.
Cardiopulmonary Division, Department of Internal Medicine, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan.
Hiroaki
H
Kodama
Keiichi
K
Fukuda
Eiichi
E
Takahashi
Satoko
S
Tahara
Yuichi
Y
Tomita
Masaki
M
Ieda
Kensuke
K
Kimura
Koji M
KM
Owada
Kristiina
K
Vuori
Satoshi
S
Ogawa
D000818:Animals; D000081247:CSK Tyrosine-Protein Kinase; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D050719:Crk-Associated Substrate Protein; D019332:Endothelin-1 / Q000494:pharmacology*; D004789:Enzyme Activation; D051418:Focal Adhesion Kinase 2; D022001:Focal Adhesions / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases; D020935:MAP Kinase Signaling System; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009154:Mutation; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics / Q000502:physiology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000502:physiology; D011506:Proteins; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology; D050737:Proto-Oncogene Proteins c-crk; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D050717:Retinoblastoma-Like Protein p130; D015533:Transcriptional Activation; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases
C493843:Bcar1 protein, rat; C493872:Crk protein, rat; D050719:Crk-Associated Substrate Protein; D019332:Endothelin-1; D010750:Phosphoproteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; D050737:Proto-Oncogene Proteins c-crk; D050717:Retinoblastoma-Like Protein p130; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D051418:Focal Adhesion Kinase 2; D016392:Proto-Oncogene Proteins pp60(c-src); C496054:Ptk2b protein, rat; D019061:src-Family Kinases; D011493:Protein Kinase C; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium