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PMID: 7882972 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Regulation of Raf-1 kinase activity by the 14-3-3 family of proteins.

The EMBO journal ·Vol. 14 ·No. 4 ·1995-02-15 ·Pages 685-96

Li S, Janosch P, Tanji M, Rosenfeld GC, Waymire JC, Mischak H, Kolch W, Sedivy JM

Abstract

We have identified the beta (beta) isoform of the 14-3-3 family of proteins as an activator of the Raf-1 protein kinase. 14-3-3 was isolated in a yeast two-hybrid screen for Raf-1 kinase domain binding proteins. Purified bovine brain 14-3-3 interacted specifically with both c-Raf-1 and the isolated Raf-1 kinase domain. Association was sensitive to the activation status of Raf-1; 14-3-3 bound to unactivated Raf-1, but not Raf-1 activated by protein kinase C alpha or Ras and Lck. The significance of these interactions under physiological conditions was demonstrated by co-immunoprecipitation of Raf-1 and 14-3-3 from extracts of quiescent, but not mitogen-stimulated, NIH 3T3 cells. 14-3-3 was not a preferred Raf-1 substrate in vitro and did not significantly affect Raf-1 kinase activity in a purified system. However, in cell-free extracts 14-3-3 acted as a Ras-independent activator of both c-Raf-1 and the Raf-1 kinase domain. The same results were obtained in vivo using transfection assays; 14-3-3 enhanced both c-Raf-1- and Raf-1 kinase domain-stimulated expression of AP-1- and NF-kappa B-dependent reporter genes and accelerated Raf-1 kinase domain-triggered differentiation of PC12 cells. We conclude that 14-3-3 is a latent co-activator bound to unactivated Raf-1 in quiescent cells and mediates mitogen-triggered but Ras-independent regulatory effects aimed directly at the kinase domain.

MeSH Terms
14-3-3 Proteins 3T3 Cells Animals Base Sequence Cell-Free System Cloning, Molecular Enzyme Activation Growth Substances/blood Mice Molecular Sequence Data Protein Binding Protein Serine-Threonine Kinases/metabolism Proteins/metabolism Proto-Oncogene Proteins/metabolism Proto-Oncogene Proteins c-raf Tyrosine 3-Monooxygenase
Chemicals
14-3-3 Proteins Growth Substances Proteins Proto-Oncogene Proteins Tyrosine 3-Monooxygenase Protein Serine-Threonine Kinases Proto-Oncogene Proteins c-raf
Authors & Affiliations
8 authors, click to expand affiliations / ORCID
Li S
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, CT 06520.
Janosch P
Tanji M
Rosenfeld G C
Waymire J C
Mischak H
Kolch W
Sedivy J M
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
Published
1995-02-15
Pages
685-96
Language
English
Region
England
NLM ID
8208664
PMCID
PMC398132
Subset
IM
Grants
NIGMS NIH HHS · GM-R01-41690 · United States
External Links
PubMed source PMC Full Text View journal details
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