Related Experiment Video

Updated: Dec 8, 2025

Author Spotlight: Integrating BRET-Based Assays and Rare Mutation Analysis to Decipher RAF Kinase Regulation in Live Cells
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Author Spotlight: Integrating BRET-Based Assays and Rare Mutation Analysis to Decipher RAF Kinase Regulation in Live Cells

Published on: March 1, 2024

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Dimerization Induced by C-Terminal 14-3-3 Binding Is Sufficient for BRAF Kinase Activation

Nicholas P D Liau, Avinashnarayan Venkatanarayan, John G Quinn

    Biochemistry
    |September 24, 2020
    PubMed

    Insights

    14-3-3 protein binding activates BRAF kinase by inducing dimerization, not by requiring a specific BRAF C-terminal region. This BRAF dimerization is crucial for achieving catalytic activity, similar to oncogenic mutations.

    Area of Science:

    • Molecular Biology
    • Cell Signaling
    • Biochemistry

    Background:

    • The Ras-RAF-MEK-ERK pathway is critical in cell signaling and frequently altered in cancers.
    • 14-3-3 proteins are key regulators that modulate RAF kinase activity through specific binding interactions.
    • Previous studies indicated a C-terminal "DTS" region in BRAF is essential for 14-3-3-mediated activation.

    Purpose of the Study:

    • To investigate the role of the BRAF C-terminal "DTS" region in 14-3-3-mediated BRAF activation.
    • To elucidate the mechanism by which 14-3-3 binding leads to BRAF kinase activation.

    Main Methods:

    • Biochemical assays to assess catalytic activity and ATP binding affinity of BRAF:14-3-3 complexes.
    • X-ray crystallography to determine the structural basis of BRAF:14-3-3 interaction.

    Main Results:

    • The catalytic activity and ATP binding affinity of BRAF:14-3-3 complexes are independent of the DTS region.
    • Crystal structure reveals the DTS is not required for the catalytically active BRAF conformation.
    • 14-3-3-induced BRAF dimerization is the primary mechanism for activation.

    Conclusions:

    • BRAF activation by 14-3-3 relies on dimerization, not the DTS region.
    • The BRAF:14-3-3 tetramer achieves catalytic competence comparable to oncogenic BRAF V600E.
    • This finding refines understanding of RAF pathway regulation in normal and cancer cells.

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