v-Crk activates the phosphoinositide 3-kinase/AKT pathway by utilizing focal adhesion kinase and H-Ras.
v-Crk, an oncogene product of avian sarcoma virus CT10, efficiently transforms chicken embryo fibroblasts (CEF). We have recently reported that constitutive activation of the phosphoinositide 3-kinase (PI3K)/AKT pathway plays a critical role in the v-Crk-induced transformation of CEF. In the present study we investigated the molecular mechanism by which v-Crk activates the PI3K/AKT pathway. First, we found that v-Crk promotes the association of the p85 regulatory subunit of PI3K with focal adhesion kinase (FAK) by inducing the phosphorylation of the Y397 residue in FAK. This FAK phosphorylation needs activation of the Src family tyrosine kinase(s) for which the v-Crk SH2 domain is responsible. v-Crk was unable to activate the PI3K/AKT pathway in FAK-null cells, indicating the functional importance of FAK. In addition, we found that H-Ras is also required for the activation of the PI3K/AKT pathway. The v-Crk-induced activation of AKT was greatly enhanced by the overexpression of H-Ras or its guanine nucleotide exchange factor mSOS, which binds to the v-Crk SH3 domain, whereas a dominant-negative mutant of H-Ras almost completely suppressed this activation. Furthermore, we showed that v-Crk stimulates the interaction of H-Ras with the Ras binding domain in the PI3K p110 catalytic subunit. Our data indicated that the v-Crk-induced activation of PI3K/AKT pathway was cooperatively achieved by two distinct interactions. One is the interaction of p85 with tyrosine-phosphorylated FAK promoted by the v-Crk SH2 domain, and another is the interaction of p110 with H-Ras dictated by the v-Crk SH3 domain.
Akagi
Tsuyoshi
T
Laboratory of Molecular Oncology, Osaka Bioscience Institute, 6-2-4 Furuedai, Suita, Osaka 565-0874, Japan.
Murata
Kazutaka
K
Shishido
Tomoyuki
T
Hanafusa
Hidesaburo
H
D000818:Animals; D001358:Avian Sarcoma Viruses / Q000378:metabolism; D002642:Chick Embryo; D004789:Enzyme Activation; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D050736:Oncogene Protein v-crk; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015682:Retroviridae Proteins, Oncogenic / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014170:Transformation, Genetic; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism
D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D050736:Oncogene Protein v-crk; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D016283:Proto-Oncogene Proteins p21(ras); D011993:Recombinant Fusion Proteins; D015682:Retroviridae Proteins, Oncogenic; D014443:Tyrosine; D019061:src-Family Kinases; C496050:PTK2 protein, human; C494918:AKT1 protein, human; C501009:HRAS protein, human