Abstract
Cellular proliferation, and differentiation of cells in response to extracellular signals, are controlled by the signal transduction pathway of Ras, Raf and MAP (mitogen-activated protein) kinase. The mechanisms that regulate this pathway are not well known. Here we describe two structurally similar tyrosine kinase substrates, Spred-1 and Spred-2. These two proteins contain a cysteine-rich domain related to Sprouty (the SPR domain) at the carboxy terminus. In Drosophila, Sprouty inhibits the signalling by receptors of fibroblast growth factor (FGF) and epidermal growth factor (EGF) by suppressing the MAP kinase pathway2,3,4,5,6,7. Like Sprouty, Spred inhibited growth-factor-mediated activation of MAP kinase. The Ras–MAP kinase pathway is essential in the differentiation of neuronal cells and myocytes. Expression of a dominant negative form of Spred and Spred-antibody microinjection revealed that endogenous Spred regulates differentiation in these types of cells. Spred constitutively associated with Ras but did not prevent activation of Ras or membrane translocation of Raf. Instead, Spred inhibited the activation of MAP kinase by suppressing phosphorylation and activation of Raf. Spred may represent a class of proteins that modulate Ras–Raf interaction and MAP kinase signalling.
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Acknowledgements
The first two authors contributed almost equally to this work. We thank H. Ohgusu, M. Sasaki and N. Tanaka for technical assistance; T. Sato and Y. Kaziro for wild-type and mutant Ras; H. Miki for the Drosophila cDNA library; Y. Hiromi for Drosophila Sprouty cDNA; P. Chambon, R. Denton, M. Yokouchi and J.-M. Garnier for library construction; Y. Sako and S. Minoguchi for valuable discussions; and M. Ohara for critical comments on the manuscript and for language assistance. This work was supported in part by grants from the Ministry of Education, Science, Technology, Sports, and Culture of Japan, Japan Research Foundation for Clinical Pharmacology, Fukuoka Cancer Society, Uehara Memorial Foundation and the Mitsubishi Foundation. Murine Spred-1 and Spred-2 are deposited in the nucleotide sequence databases under accession numbers AB063495 and AB063496, respectively.
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Wakioka, T., Sasaki, A., Kato, R. et al. Spred is a Sprouty-related suppressor of Ras signalling. Nature 412, 647–651 (2001). https://doi.org/10.1038/35088082
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DOI: https://doi.org/10.1038/35088082
