CrkII regulates focal adhesion kinase activation by making a complex with Crk-associated substrate, p130Cas |
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Authors: | Iwahara Toshinori Akagi Tsuyoshi Fujitsuka Yuki Hanafusa Hidesaburo |
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Affiliation: | Laboratory of Molecular Oncology, Osaka Bioscience Institute, 6-2-4 Furuedai, Suita, Osaka 565-0874, Japan. |
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Abstract: | CrkII is an adaptor protein possessing oncogenic potential despite the lack of an enzymatic domain. We investigated here the physiological functions of CrkII by studying its ability to induce anchorage-independent cell growth. We found that inhibition or null mutation of focal adhesion kinase (FAK) blocked the anchorage-independent growth induced by CrkII overexpression, indicating that FAK is a critical determinant of the transforming activity of CrkII. CrkII overexpression enhanced the autophosphorylation of FAK at Tyr-397 and tyrosine phosphorylation of p130(Cas) (Crk-associated substrate, Cas) upon stimulation of integrin by fibronectin. Moreover, the constitutive phosphorylation of FAK and Cas was observed in CrkII-overexpressing cells, even when they were in the suspended condition, consistent with the ability of CrkII to induce anchorage-independent growth. Using Cas-deficient cells, we showed Cas function to be essential for both the CrkII-induced phosphorylation of FAK (Tyr-397) and anchorage-independent cell growth. The CrkII-induced FAK autophosphorylation depended upon CrkII-Cas complex formation. Furthermore, we showed that CrkII knockdown resulted in defects in integrin-mediated events, such as cell spreading, haptotactic migration, and FAK autophosphorylation. The integrin-mediated FAK autophosphorylation was also reduced in Cas-deficient cells. These results suggest that the CrkII-Cas complex functions in integrin-mediated FAK activation signaling. Our findings show the importance of CrkII in integrin-mediated events, acting upstream of FAK to affect the activation of this kinase, which appears to have a central role in this pathway. |
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Keywords: | anchorage-independent growth integrin tyrosine phosphorylation |
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