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Disruption of the 14-3-3 Binding Site within the B-Raf Kinase Domain Uncouples Catalytic Activity from PC12 Cell Differentiation*
- Source :
- Journal of Biological Chemistry; February 2000, Vol. 275 Issue: 6 p3803-3809, 7p
- Publication Year :
- 2000
-
Abstract
- A number of Raf-associated proteins have recently been identified, including members of the 14-3-3 family of phosphoserine-binding proteins. Although both positive and negative regulatory functions have been ascribed for 14-3-3 interactions with Raf-1, the mechanisms by which 14-3-3 binding modulates Raf activity have not been fully established. We report that mutational disruption of 14-3-3 binding to the B-Raf catalytic domain inhibits B-Raf biological activity. Expression of the isolated B-Raf catalytic domain (B-Rafcat) induces PC12 cell differentiation in the absence of nerve growth factor. By contrast, the B-Rafcat 14-3-3 binding mutant, B-Rafcat S728A, was severely compromised for the induction of PC12 cell differentiation. Interestingly, the B-Rafcat 14-3-3 binding mutant retained significant in vitrocatalytic activity. InXenopusoocytes, the analogous full-length B-Raf 14-3-3 binding mutant blocked progesterone-stimulated maturation and the activation of endogenous mitogen-activated protein kinase kinase and mitogen-activated protein kinase. Similarly, the full-length B-Raf 14-3-3 binding mutant inhibited nerve growth factor-stimulated PC12 cell differentiation. We conclude that 14-3-3 interaction with the catalytic domain is not required for kinase activity per sebut is essential to couple B-Raf catalytic activity to downstream effector activation.
Details
- Language :
- English
- ISSN :
- 00219258 and 1083351X
- Volume :
- 275
- Issue :
- 6
- Database :
- Supplemental Index
- Journal :
- Journal of Biological Chemistry
- Publication Type :
- Periodical
- Accession number :
- ejs55812786
- Full Text :
- https://doi.org/10.1074/jbc.275.6.3803