Tyrosine Residues 239 and 240 of Shc Are Phosphatidylinositol 4,5-Bisphosphate-Dependent Phosphorylation Sites by c-Src

• Published in: Biochemical and biophysical research communications Vol. 240; no. 2; pp. 399 - 404
• Main Authors: Sato, Ken-ichi, Gotoh, Noriko, Otsuki, Tetsuji, Kakumoto, Miki, Aoto, Mamoru, Tokmakov, Alexander A., Shibuya, Masabumi, Fukami, Yasuo
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 17.11.1997
• Subjects: Adaptor Proteins, Signal Transducing; Adaptor Proteins, Vesicular Transport; Amino Acid Substitution; Animals; Brain - metabolism; Carcinoma, Squamous Cell; Cattle; Cell Line, Transformed; ErbB Receptors - metabolism; Glutathione Transferase; Humans; Mice; Mutagenesis, Site-Directed; Oncogene Protein pp60(v-src) - metabolism; Phosphatidylinositol 4,5-Diphosphate - metabolism; Phosphatidylinositol 4,5-Diphosphate - pharmacology; Protein Kinases - metabolism; Proteins - chemistry; Proteins - metabolism; Proto-Oncogene Proteins pp60(c-src) - metabolism; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - metabolism; Shc Signaling Adaptor Proteins; Src Homology 2 Domain-Containing, Transforming Protein 1; Tumor Cells, Cultured; Tyrosine
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1006/bbrc.1997.7667

In the previous study (Sato K.-I. et al.(1997) FEBS Lett.410, 136–140), we showed that the phosphorylation of Shc protein by c-Src is dependent on the binding of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) to the PTB domain of Shc. In this study, we demonstrate that, in contrast to c-Src, v-Src and epidermal growth factor (EGF) receptor can phosphorylate Shc in a PtdIns(4,5)P2-independent manner and at different phosphorylation sites. To determine the phosphorylation sites in Shc, we used mutant Shc proteins in which tyrosine residues (Y) 317 and/or 239 and 240 were replaced by phenylalanine residues (F). We found that Y317F Shc but not Y239/240F or Y239/240/317F Shc was phosphorylated by c-Src. The reaction was PtdIns(4,5)P2-dependent and inhibited by the addition of PTB domain of Shc. On the other hand, v-Src and EGF receptor were able to phosphorylate both Y317F and Y239/240F but not Y239/240/317F Shc in a PtdIns(4,5)P2-independent manner. These results highlight the difference between c-Src and v-Src or EGF receptor and suggest that c-Src can phosphorylate predominantly on Tyr239/240 of Shc only when Shc PTB domain is bound to PtdIns(4,5)P2.