Cell type-specific regulation of B-Raf kinase by cAMP and 14-3-3 proteins.

Cyclic AMP can either activate or inhibit the mitogen-activated protein kinase (MAPK) pathway in different cell types; MAPK activation has been observed in B-Raf-expressing cells and has been attributed to Rap1 activation with subsequent B-Raf activation, whereas MAPK inhibition has been observed in cells lacking B-Raf and has been attributed ...
to cAMP-dependent protein kinase (protein kinase A)-mediated phosphorylation and inhibition of Raf-1 kinase. We found that cAMP stimulated MAPK activity in CHO-K1 and PC12 cells but inhibited MAPK activity in C6 and NB2A cells. In all four cell types, cAMP activated Rap1, and the 95- and 68-kDa isoforms of B-Raf were expressed. cAMP activation or inhibition of MAPK correlated with activation or inhibition of endogenous and transfected B-Raf kinase. Although all cell types expressed similar amounts of 14-3-3 proteins, approximately 5-fold less 14-3-3 was associated with B-Raf in cells in which cAMP was inhibitory than in cells in which cAMP was stimulatory. We found that the cell type-specific inhibition of B-Raf could be completely prevented by overexpression of 14-3-3 isoforms, whereas expression of a dominant negative 14-3-3 mutant resulted in partial loss of B-Raf activity. Our data suggest that 14-3-3 bound to B-Raf protects the enzyme from protein kinase A-mediated inhibition; the amount of 14-3-3 associated with B-Raf may explain the tissue-specific effects of cAMP on B-Raf kinase activity.
Mesh Terms:
14-3-3 Proteins, Animals, Cell Line, Cricetinae, Cyclic AMP, Cyclic AMP-Dependent Protein Kinases, Enzyme Activation, Guanosine Triphosphate, Isoenzymes, MAP Kinase Signaling System, Mitogen-Activated Protein Kinases, Mutation, Organ Specificity, Phosphorylation, Protein Binding, Proto-Oncogene Proteins c-raf, Rats, Thionucleotides, Transfection, Tyrosine 3-Monooxygenase
J. Biol. Chem.
Date: Oct. 13, 2000
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