Distinct Binding Preferences between Ras and Raf Family Members and the Impact on Oncogenic Ras Signaling

Mol Cell. 2019 Dec 19;76(6):872-884.e5. doi: 10.1016/j.molcel.2019.09.004. Epub 2019 Oct 9.

Abstract

The Ras GTPases are frequently mutated in human cancer, and, although the Raf kinases are essential effectors of Ras signaling, the tumorigenic properties of specific Ras-Raf complexes are not well characterized. Here, we examine the ability of individual Ras and Raf proteins to interact in live cells using bioluminescence resonance energy transfer (BRET) technology. We find that C-Raf binds all mutant Ras proteins with high affinity, whereas B-Raf exhibits a striking preference for mutant K-Ras. This selectivity is mediated by the acidic, N-terminal segment of B-Raf and requires the K-Ras polybasic region for high-affinity binding. In addition, we find that C-Raf is critical for mutant H-Ras-driven signaling and that events stabilizing B-Raf/C-Raf dimerization, such as Raf inhibitor treatment or certain B-Raf mutations, can allow mutant H-Ras to engage B-Raf with increased affinity to promote tumorigenesis, thus revealing a previously unappreciated role for C-Raf in potentiating B-Raf function.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Cell Proliferation*
  • Cell Transformation, Neoplastic / genetics
  • Cell Transformation, Neoplastic / metabolism*
  • Cell Transformation, Neoplastic / pathology
  • Female
  • Gene Expression Regulation, Neoplastic
  • HEK293 Cells
  • HeLa Cells
  • Humans
  • Male
  • Mice
  • Mutation
  • NIH 3T3 Cells
  • Neoplasms / enzymology*
  • Neoplasms / genetics
  • Neoplasms / pathology
  • Protein Binding
  • Protein Interaction Domains and Motifs
  • Protein Multimerization
  • Proto-Oncogene Proteins B-raf / genetics
  • Proto-Oncogene Proteins B-raf / metabolism
  • Proto-Oncogene Proteins c-raf / genetics
  • Proto-Oncogene Proteins c-raf / metabolism
  • Proto-Oncogene Proteins p21(ras) / genetics
  • Proto-Oncogene Proteins p21(ras) / metabolism
  • Signal Transduction / genetics
  • Spheroids, Cellular
  • raf Kinases / genetics
  • raf Kinases / metabolism*
  • ras Proteins / genetics
  • ras Proteins / metabolism*

Substances

  • KRAS protein, human
  • BRAF protein, human
  • Braf protein, mouse
  • Proto-Oncogene Proteins B-raf
  • Proto-Oncogene Proteins c-raf
  • Raf1 protein, human
  • Raf1 protein, mouse
  • raf Kinases
  • HRAS protein, human
  • Hras protein, mouse
  • Proto-Oncogene Proteins p21(ras)
  • ras Proteins