Cell-surface receptors frequently use scaffold proteins to recruit cytoplasmic targets, but the rationale for this is uncertain. Activated receptor tyrosine kinases, for example, engage scaffolds such as Shc1 that contain phosphotyrosine (pTyr)-binding (PTB) domains. Using quantitative mass spectrometry, here we show that mammalian Shc1 responds to epidermal growth factor (EGF) stimulation through multiple waves of distinct phosphorylation events and protein interactions. After stimulation, Shc1 rapidly binds a group of proteins that activate pro-mitogenic or survival pathways dependent on recruitment of the Grb2 adaptor to Shc1 pTyr sites. Akt-mediated feedback phosphorylation of Shc1 Ser 29 then recruits the Ptpn12 tyrosine phosphatase. This is followed by a sub-network of proteins involved in cytoskeletal reorganization, trafficking and signal termination that binds Shc1 with delayed kinetics, largely through the SgK269 pseudokinase/adaptor protein. Ptpn12 acts as a switch to convert Shc1 from pTyr/Grb2-based signalling to SgK269-mediated pathways that regulate cell invasion and morphogenesis. The Shc1 scaffold therefore directs the temporal flow of signalling information after EGF stimulation.
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We thank J. Moffat for shRNA lentiviruses and K. Shokat for PI3Kp110 isoform-specific inhibitors. We thank C. Jorgensen, R. Williams, I. Louria-Hayon, R. Tian, and E. Petsalaki for critical input, A. James, V. Nguyen, and B. Larsen for technical assistance and M. M. Stacey, C. Chen and J. Jin for comments on the manuscript. Supported by Genome Canada through the Ontario Genomics Institute, the Ontario Research Fund from the Ontario Ministry of Research and Innovation, a Terry Fox Foundation team grant, the Canadian Institutes of Health Research (MOP-13466-6849), and the Canada Foundation for Innovation. M.A.S. is supported by a Vanier Canada Graduate Studentship and R.B. is supported by a CIHR postdoctoral fellowship. Support for R.J.D. was from the National Health and Medical Research Council of Australia and Cancer Council New South Wales (NSW), and for D.R.C. from Cancer Institute NSW.
S.A.T. is an employee of AB SCIEX. AB SCIEX has provided support for the Ontario Research Fund grant (awarded to T.P.).
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Zheng, Y., Zhang, C., Croucher, D. et al. Temporal regulation of EGF signalling networks by the scaffold protein Shc1. Nature 499, 166–171 (2013). https://doi.org/10.1038/nature12308
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