The C-terminus of Raf-1 acts as a 14-3-3-dependent activation switch

Amardeep S. Dhillon, Yan Y. Yip, G. Joan Grindlay, Julian L. Pakay, Marc Dangers, Meike Hillmann, William Clark, Andrew Pitt, Harald Mischak, Walter Kolch

Research output: Contribution to journalArticle

Abstract

The Raf-1 protein kinase is a major activator of the ERK MAPK pathway, which links signaling by a variety of cell surface receptors to the regulation of cell proliferation, survival, differentiation and migration. Signaling by Raf-1 is regulated by a complex and poorly understood interplay between phosphorylation events and protein-protein interactions. One important mode of Raf-1 regulation involves the phosphorylation-dependent binding of 14-3-3 proteins. Here, we have examined the mechanism whereby the C-terminal 14-3-3 binding site of Raf-1, S621, controls the activation of MEK-ERK signaling. We show that phosphorylation of S621 turns over rapidly and is enriched in the activated pool of endogenous Raf-1. The phosphorylation on this site can be mediated by Raf-1 itself but also by other kinase(s). Mutations that prevent the binding of 14-3-3 proteins to S621 render Raf-1 inactive by specifically disrupting its capacity to bind to ATP, and not by gross conformational alteration as indicated by intact MEK binding. Phosphorylation of S621 correlates with the inhibition of Raf-1 catalytic activity in vitro, but 14-3-3 proteins can completely reverse this inhibition. Our findings suggest that 14-3-3 proteins function as critical cofactors in Raf-1 activation, which induce and maintain the protein in a state that is competent for both ATP binding and MEK phosphorylation.
LanguageEnglish
Pages1645-1651
Number of pages7
JournalCellular Signalling
Volume21
Issue number11
DOIs
Publication statusPublished - Nov 2009

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14-3-3 Proteins
Phosphorylation
Mitogen-Activated Protein Kinase Kinases
Adenosine Triphosphate
Proto-Oncogene Proteins c-raf
Proteins
MAP Kinase Signaling System
Cell Surface Receptors
Protein Kinases
Cell Movement
Cell Survival
Phosphotransferases
Binding Sites
Cell Proliferation
Mutation

Keywords

  • 14-3-3 proteins
  • amino acid substitution
  • animals
  • binding sites
  • COS cells
  • cell line
  • cercopithecus aethiops
  • extracellular signal-regulated MAP kinases
  • humans
  • MAP kinase kinase kinases
  • mice
  • mutation
  • phosphorylation
  • tertiary protein structure
  • proto-oncogene proteins c-raf
  • signal transduction

Cite this

Dhillon, A. S., Yip, Y. Y., Grindlay, G. J., Pakay, J. L., Dangers, M., Hillmann, M., ... Kolch, W. (2009). The C-terminus of Raf-1 acts as a 14-3-3-dependent activation switch. Cellular Signalling, 21(11), 1645-1651. https://doi.org/10.1016/j.cellsig.2009.07.001
Dhillon, Amardeep S. ; Yip, Yan Y. ; Grindlay, G. Joan ; Pakay, Julian L. ; Dangers, Marc ; Hillmann, Meike ; Clark, William ; Pitt, Andrew ; Mischak, Harald ; Kolch, Walter. / The C-terminus of Raf-1 acts as a 14-3-3-dependent activation switch. In: Cellular Signalling. 2009 ; Vol. 21, No. 11. pp. 1645-1651.
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Dhillon, AS, Yip, YY, Grindlay, GJ, Pakay, JL, Dangers, M, Hillmann, M, Clark, W, Pitt, A, Mischak, H & Kolch, W 2009, 'The C-terminus of Raf-1 acts as a 14-3-3-dependent activation switch' Cellular Signalling, vol. 21, no. 11, pp. 1645-1651. https://doi.org/10.1016/j.cellsig.2009.07.001

The C-terminus of Raf-1 acts as a 14-3-3-dependent activation switch. / Dhillon, Amardeep S.; Yip, Yan Y.; Grindlay, G. Joan; Pakay, Julian L.; Dangers, Marc; Hillmann, Meike; Clark, William; Pitt, Andrew; Mischak, Harald; Kolch, Walter.

In: Cellular Signalling, Vol. 21, No. 11, 11.2009, p. 1645-1651.

Research output: Contribution to journalArticle

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AU - Dhillon, Amardeep S.

AU - Yip, Yan Y.

AU - Grindlay, G. Joan

AU - Pakay, Julian L.

AU - Dangers, Marc

AU - Hillmann, Meike

AU - Clark, William

AU - Pitt, Andrew

AU - Mischak, Harald

AU - Kolch, Walter

PY - 2009/11

Y1 - 2009/11

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KW - COS cells

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KW - cercopithecus aethiops

KW - extracellular signal-regulated MAP kinases

KW - humans

KW - MAP kinase kinase kinases

KW - mice

KW - mutation

KW - phosphorylation

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KW - proto-oncogene proteins c-raf

KW - signal transduction

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Dhillon AS, Yip YY, Grindlay GJ, Pakay JL, Dangers M, Hillmann M et al. The C-terminus of Raf-1 acts as a 14-3-3-dependent activation switch. Cellular Signalling. 2009 Nov;21(11):1645-1651. https://doi.org/10.1016/j.cellsig.2009.07.001