Direct interaction of Ras and the amino-terminal region of Raf-1 in vitro (original) (raw)

Nature volume 364, pages 352–355 (1993)Cite this article

Abstract

THE Ras proteins are key regulators of the growth of eukaryotic cells, but their direct target enzymes, or 'effectors', are unknown1. The protein encoded by the c-raf-1 proto-oncogene is thought to function downstream of p21ras because disruption of Raf blocks signalling by Ras in a number of systems2–5. Here we report that the amino-terminal cysteine-rich regulatory region of p74_c-raf-1_ expressed as a glutathione-S-transferase (GST) fusion protein binds directly to Ras with relatively high affinity (50 nM). The binding is strictly dependent on the Ras protein being in the active GTP-bound conformation rather than the inactive GDP-bound state. Raf–GST interacts with wild-type and oncogenic Ras (Val 12) but fails to interact with a biologically inert effector mutant of Ras (Ala 38) and a dominant negative mutant (Asn 17). A peptide based on the effector region of Ras inhibits the interaction. Raf–GST acts as a potent competitive inhibitor of the GTPase-activating proteins p120GAPand neurofibromin. In addition, Raf itself displays weak GTPase-stimulating activity towards Ras. It is therefore likely that Raf is a direct effector of Ras.

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  1. Julian Downward: To whom correspondence should be addressed

Authors and Affiliations

  1. Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London, WC2A 3PX, UK
    Patricia H. Warne, Pablo Rodrigueza Vician & Julian Downward

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  1. Patricia H. Warne
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  2. Pablo Rodrigueza Vician
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  3. Julian Downward
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Warne, P., Vician, P. & Downward, J. Direct interaction of Ras and the amino-terminal region of Raf-1 in vitro.Nature 364, 352–355 (1993). https://doi.org/10.1038/364352a0

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