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PMID: 9001227 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Role of diacylglycerol-regulated protein kinase C isotypes in growth factor activation of the Raf-1 protein kinase.

Molecular and cellular biology ·Vol. 17 ·No. 2 ·1997-02-00 ·Pages 732-41

Cai H, Smola U, Wixler V, Eisenmann-Tappe I, Diaz-Meco MT, Moscat J, Rapp U, Cooper GM

Abstract

The Raf protein kinases function downstream of Ras guanine nucleotide-binding proteins to transduce intracellular signals from growth factor receptors. Interaction with Ras recruits Raf to the plasma membrane, but the subsequent mechanism of Raf activation has not been established. Previous studies implicated hydrolysis of phosphatidylcholine (PC) in Raf activation; therefore, we investigated the role of the epsilon isotype of protein kinase C (PKC), which is stimulated by PC-derived diacylglycerol, as a Raf activator. A dominant negative mutant of PKC epsilon inhibited both proliferation of NIH 3T3 cells and activation of Raf in COS cells. Conversely, overexpression of active PKC epsilon stimulated Raf kinase activity in COS cells and overcame the inhibitory effects of dominant negative Ras in NIH 3T3 cells. PKC epsilon also stimulated Raf kinase in baculovirus-infected Spodoptera frugiperda Sf9 cells and was able to directly activate Raf in vitro. Consistent with its previously reported activity as a Raf activator in vitro, PKC alpha functioned similarly to PKC epsilon in both NIH 3T3 and COS cell assays. In addition, constitutively active mutants of both PKC alpha and PKC epsilon overcame the inhibitory effects of dominant negative mutants of the other PKC isotype, indicating that these diacylglycerol-regulated PKCs function as redundant activators of Raf-1 in vivo.

MeSH Terms
3T3 Cells Animals COS Cells Cell Division Cell Line Diglycerides/metabolism Enzyme Activation/drug effects Epidermal Growth Factor/pharmacology Isoenzymes/genetics,isolation & purification,metabolism,pharmacology,physiology Mice Mitogens/pharmacology Mutation Protein Kinase C/genetics,isolation & purification,metabolism,pharmacology,physiology Protein Kinase C-alpha Protein Kinase C-epsilon Protein Serine-Threonine Kinases/metabolism Proto-Oncogene Proteins/metabolism Proto-Oncogene Proteins c-raf Recombinant Fusion Proteins Signal Transduction/physiology Spodoptera Tetradecanoylphorbol Acetate/pharmacology Transfection
Chemicals
Diglycerides Isoenzymes Mitogens Proto-Oncogene Proteins Recombinant Fusion Proteins Epidermal Growth Factor Prkce protein, mouse Protein Serine-Threonine Kinases Proto-Oncogene Proteins c-raf Prkca protein, mouse Protein Kinase C Protein Kinase C-alpha Protein Kinase C-epsilon Tetradecanoylphorbol Acetate
Authors & Affiliations
8 authors, click to expand affiliations / ORCID
Cai H
Division of Molecular Genetics, Dana-Farber Cancer Institute, and Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Smola U
Wixler V
Eisenmann-Tappe I
Diaz-Meco M T
Moscat J
Rapp U
Cooper G M
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.17.2.732
Published
1997-02-00
Pages
732-41
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC231799
Subset
IM
Grants
NCI NIH HHS · R01 CA18689 · United States
External Links
PubMed source DOI: 10.1128/MCB.17.2.732 PMC Full Text View journal details
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