Association of c-Src with p52Shc in mitotic NIH3T3 cells as revealed by Src-Shc binding site-specific antibodies.
In a previous study, we presented evidence that the adaptor protein Shc interacts with and activates the tyrosine kinase c-Src without affecting the phosphorylation state of Tyr-527 in c-Src. Here we show that Shc-mediated c-Src activation occurs in mitotic NIH 3T3 cells. Co-immunoprecipitation studies demonstrate that the c-Src-p52Shc complex involves the activation segment/inter-DFG-APE (IDA) region of c-Src and the amino-terminal region of p52Shc. The complex formation contributes to the c-Src activation, because (i) specific activity of c-Src associated with p52Shc is higher than that of the total c-Src, and (ii) a recombinant protein containing the c-Src IDA sequence disrupts the complex and decreases the c-Src activity. Anti-Src IDA antibody can activate c-Src in vitro, and synthetic peptides that cover the carboxyl-terminal half of the Src IDA region interfere with the kinase-activating effect of anti-Src IDA antibody. These results support the idea that dephosphorylation-independent activation of c-Src by Shc is mediated by a molecular interaction involving the c-Src IDA region.
Sato
Ken-ichi
K
Research Center for Environmental Genomics, Kobe University, Nada, Kobe 657-8501, Japan. kksato@kobe-u.ac.jp
Iwasaki
Tetsushi
T
Fukami
Yasuo
Y
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D000906:Antibodies / Q000276:immunology; D001666:Binding Sites, Antibody; D051379:Mice; D046912:Multiprotein Complexes / Q000276:immunology / Q000378:metabolism*; D041681:NIH 3T3 Cells; D011485:Protein Binding; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000276:immunology / Q000378:metabolism*; D011817:Rabbits; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1
D048868:Adaptor Proteins, Signal Transducing; D000906:Antibodies; D046912:Multiprotein Complexes; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D016392:Proto-Oncogene Proteins pp60(c-src)