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Protein phosphatases 1 and 2A promote Raf-1 activation by regulating 14-3-3 interactions

Oncogene volume 20, pages 3949–3958 (2001)Cite this article

Abstract

Raf-1 activation is a complex process which involves plasma membrane recruitment, phosphorylation, protein-protein and lipid-protein interactions. We now show that PP1 and PP2A serine-threonine phosphatases also have a positive role in Ras dependent Raf-1 activation. General serine-threonine phosphatase inhibitors such sodium fluoride, or ß-glycerophosphate and sodium pyrophosphate, or specific PP1 and PP2A inhibitors including microcystin-LR, protein phosphatase 2A inhibitor I1 or protein phosphatase inhibitor 2 all abrogate H-Ras and K-Ras dependent Raf-1 activation in vitro. A critical Raf-1 target residue for PP1 and PP2A is S259. Serine phosphatase inhibitors block the dephosphorylation of S259, which accompanies Raf-1 activation, and Ras dependent activation of mutant Raf259A is relatively resistant to serine phosphatase inhibitors. Sucrose gradient analysis demonstrates that serine phosphatase inhibition increases the total amount of 14-3-3 and Raf-1 associated with the plasma membrane and significantly alters the distribution of 14-3-3 and Raf-1 across different plasma membrane microdomains. These observations suggest that dephosphorylation of S259 is a critical early step in Ras dependent Raf-1 activation which facilitates 14-3-3 displacement. Inhibition of PP1 and PP2A therefore causes plasma membrane accumulation of Raf-1/14-3-3 complexes which cannot be activated.

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Acknowledgements

We thank Andrey Shaw for supplying the phosphospecific Raf antisera and the Raf259A expression plasmid and Brian Gabrielli, Albert Pol and other members of the Oncology Laboratory for helpful comments on the manuscript. This work was supported by a grant from the National Health and Medical Research Council of Australia. JF Hancock is also supported by the Royal Children's Hospital Foundation, Queensland, Australia.

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  1. Department of Pathology, Laboratory of Experimental Oncology, University of Queensland Medical School, Herston Road, 4006, Queensland, Australia

    Montserrat Jaumot & John F Hancock

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Jaumot, M., Hancock, J. Protein phosphatases 1 and 2A promote Raf-1 activation by regulating 14-3-3 interactions. Oncogene 20, 3949–3958 (2001). https://doi.org/10.1038/sj.onc.1204526

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