Mechanism of Activation of the RAF-ERK Signaling Pathway by Oncogenic Mutations of B-RAF

• Published in: Cell 2004-03, Vol.116 (6), p.855-867
• Main Authors: Wan, Paul T.C, Garnett, Mathew J, Roe, S.Mark, Lee, Sharlene, Niculescu-Duvaz, Dan, Good, Valerie M, Project, Cancer Genome, Jones, C.Michael, Marshall, Christopher J, Springer, Caroline J, Barford, David, Marais, Richard
• Format: Article
• Language: English
• Subjects: Allosteric Regulation - genetics; Animals; Catalytic Domain - genetics; Cell Transformation, Neoplastic - genetics; Enzyme Inhibitors - pharmacology; Gene Expression Regulation, Enzymologic - genetics; MAP Kinase Kinase 1; MAP Kinase Signaling System - genetics; Mice; Mitogen-Activated Protein Kinase Kinases - genetics; Mitogen-Activated Protein Kinase Kinases - metabolism; Mitogen-Activated Protein Kinases - genetics; Models, Molecular; Molecular Conformation; Mutation - genetics; Neoplasms - enzymology; Neoplasms - genetics; NIH 3T3 Cells; Oncogenes - genetics; Oocytes; Phosphorylation; Phosphotransferases - genetics; Phosphotransferases - metabolism; Proto-Oncogene Proteins B-raf; Proto-Oncogene Proteins c-raf - antagonists & inhibitors; Proto-Oncogene Proteins c-raf - genetics; Proto-Oncogene Proteins c-raf - metabolism; Up-Regulation - genetics; Xenopus
• ISSN: 0092-8674; 1097-4172
• DOI: 10.1016/S0092-8674(04)00215-6

Over 30 mutations of the B-RAF gene associated with human cancers have been identified, the majority of which are located within the kinase domain. Here we show that of 22 B-RAF mutants analyzed, 18 have elevated kinase activity and signal to ERK in vivo. Surprisingly, three mutants have reduced kinase activity towards MEK in vitro but, by activating C-RAF in vivo, signal to ERK in cells. The structures of wild type and oncogenic V599EB-RAF kinase domains in complex with the RAF inhibitor BAY43-9006 show that the activation segment is held in an inactive conformation by association with the P loop. The clustering of most mutations to these two regions suggests that disruption of this interaction converts B-RAF into its active conformation. The high activity mutants signal to ERK by directly phosphorylating MEK, whereas the impaired activity mutants stimulate MEK by activating endogenous C-RAF, possibly via an allosteric or transphosphorylation mechanism.