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A dimeric 14-3-3 protein is an essential cofactor for Raf kinase activity.

cRaf-1 is a mitogen-activated protein kinase that is the main effector recruited by GTP-bound Ras in order to activate the MAP kinase pathway. Inactive Raf is found in the cytosol in a complex with Hsp90, Hsp50 (Cdc37) and the 14-3-3 proteins. GTP-bound Ras binds Raf and is necessary but not sufficient for the stable activation of Raf that occurs in response to serum, epidermal growth factor, platelet-derived growth factor or insulin. These agents cause a two- to threefold increase in overall phosphorylation of Raf on serine/threonine residues, and treatment of cRaf-1 with protein (serine/threonine) phosphatases can deactivate it, at least partially. The role of 14-3-3 proteins in the regulation of Raf's kinase activity is uncertain and is investigated here. Active Raf can be almost completely deactivated in vitro by displacement of 14-3-3 using synthetic phosphopeptides. Deactivation can be substantially reversed by addition of purified recombinant bacterial 14-3-3; however, Raf must have been previously activated in vivo to be reactivated by 14-3-3 in vitro. The ability of 14-3-3 to support Raf activity is dependent on phosphorylation of serine residues on Raf and on the integrity of the 14-3-3 dimer; mutant monomeric forms of 14-3-3, although able to bind Raf in vivo, do not enable Raf to be activated in vivo or restore Raf activity after displacement of 14-3-3 in vitro. The 14-3-3 protein is not required to induce dimerization of Raf. We propose that dimeric 14-3-3 is needed both to maintain Raf in an inactive state in the absence of GTP-bound Ras and to stabilize an active conformation of Raf produced during activation in vivo.

Pubmed ID: 9665134


  • Tzivion G
  • Luo Z
  • Avruch J



Publication Data

July 2, 1998

Associated Grants


Mesh Terms

  • 14-3-3 Proteins
  • Animals
  • COS Cells
  • Dimerization
  • Enzyme Activation
  • Mutation
  • Phosphopeptides
  • Phosphorylation
  • Protein Kinase C
  • Proteins
  • Proto-Oncogene Proteins c-myc
  • Proto-Oncogene Proteins c-raf
  • Recombinant Proteins
  • Serine
  • Transfection
  • Tyrosine 3-Monooxygenase