Integrated RAS signaling defined by parallel NMR detection of effectors and regulators

Nat Chem Biol. 2014 Mar;10(3):223-30. doi: 10.1038/nchembio.1435. Epub 2014 Jan 19.


The RAS GTPase directs cell proliferation and survival by selectively relaying signals amid a dynamic network of regulatory enzymes and protein interactions. Oncogenic mutation of RAS alters cell growth by deleteriously controlling output to RAS-binding effectors. Mechanisms underlying multieffector interactions for both wild-type and oncogenic RAS are poorly understood owing to challenges in quantifying outputs to multiple pathways in parallel. Using highly selective NMR probes for wild-type and oncogenic (G12V) RAS, we develop a systematic approach that quantitatively measures RAS output in composite mixtures of GEF, GAP and effector RAS-binding domains (RBDs). We derive effector signaling hierarchies and establish how oscillating concentrations generate effector 'switching'. The G12V mutation highly perturbs this system, specifically altering interactions with RAL GTPase-specific GEFs and RAF kinases. We further reveal that RAS-RBD complexes show extensive feedback to full-length regulatory proteins. Our approach quantifies output from signaling hubs, here providing an integrated view of the RAS network.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation
  • GTPase-Activating Proteins / chemistry
  • GTPase-Activating Proteins / physiology*
  • Humans
  • Magnetic Resonance Spectroscopy*
  • Protein Structure, Tertiary
  • Proto-Oncogene Proteins c-raf / chemistry
  • Signal Transduction*
  • ras Proteins / chemistry
  • ras Proteins / metabolism
  • ras Proteins / physiology*


  • GTPase-Activating Proteins
  • Proto-Oncogene Proteins c-raf
  • ras Proteins