Sck interacts with KDR and Flt-1 via its SH2 domain.
Vascular endothelial growth factor (VEGF) is one of the major angiogenesis regulators. It binds to its tyrosine kinase receptors, KDR and Flt-1. However, little is known about their downstream signal transduction properties. We screened human brain cDNA library using the yeast two-hybrid system with the KDR cytoplasmic region as bait to find KDR binding proteins. After 6.2 x 10(6) clones were screened, we identified Sck, one of the Shc homologues, as a KDR binding protein. Sck also binds to Flt-1 and their binding is dependent on the kinase activities of KDR and Flt-1. Extensive site-directed mutagenesis of KDR revealed that Y1175 of KDR is a major binding site for Sck. As Sck contains the SH2 domain and PTB domain, we tested whether they bind to KDR and Flt-1. The SH2 domain of Sck binds to both of them. Deletion of the SH2 domain from Sck resulted in the complete loss of binding. On the other hand, the PTB domain of Sck does not bind to KDR and Flt-1. These results indicate that Sck binds to KDR and Flt-1 via its SH2 domain and might play an important role in VEGF signal transduction.
Igarashi
K
K
Advanced Technology Research Laboratories, Nippon Steel Corporation, 3-35-1 Ida, Kawasaki, Nakahara-Ku, 211-0035, Japan. igarashi@lab1.nsc.co.jp
Shigeta
K
K
Isohara
T
T
Yamano
T
T
Uno
I
I
D048868:Adaptor Proteins, Signal Transducing; D001923:Brain Chemistry; D002460:Cell Line; D003001:Cloning, Molecular; D004730:Endothelium, Vascular / Q000378:metabolism; D006801:Humans; D016297:Mutagenesis, Site-Directed; D019000:Phosphotyrosine / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D017978:Receptors, Growth Factor / Q000235:genetics / Q000378:metabolism*; D040262:Receptors, Vascular Endothelial Growth Factor; D055765:Shc Signaling Adaptor Proteins; D000071426:Src Homology 2 Domain-Containing, Transforming Protein 2; D040281:Vascular Endothelial Growth Factor Receptor-1; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D011506:Proteins; D011518:Proto-Oncogene Proteins; D017978:Receptors, Growth Factor; C099021:SHC2 protein, human; D055765:Shc Signaling Adaptor Proteins; D000071426:Src Homology 2 Domain-Containing, Transforming Protein 2; D019000:Phosphotyrosine; D020794:Receptor Protein-Tyrosine Kinases; D040262:Receptors, Vascular Endothelial Growth Factor; D040281:Vascular Endothelial Growth Factor Receptor-1