VPS35: a novel Ras interacting partner and signaling regulator

Protein product of proto-oncogene RAS undergoes multi-step post-translational modifications, and such processing contributes to its localization on the plasma membrane, where most of RAS signaling originates. There are three major RAS isoforms in human (N-, H-, and K-RAS), and subcellular distribution of N-RAS has been generally overlooked due to historical reasons. We revisited this topic and unexpectedly discovered endogenous N-RAS is largely cytosolic, in stark contrast to other Ras isoforms that are primarily on the plasma membrane. Gel filtration analysis of cytosolic RAS revealed a Ras-prenylation-dependent complex with a significantly higher molecular weight than RAS itself. Mass spectrometry analysis uncovered that VPS35, a subunit of retromer complex, binds specifically to prenylated N-RAS in the cytosol. We confirmed the interaction between N-RAS and VPS35 and its dependence of prenylation by co-immunoprecipitation, and revealed attenuated RAS signaling with VPS35 silencing, reflected by reduced level of activation in RAS and its downstream effectors. Such signaling deficiency leads to decreased proliferation in NRAS-dependent melanoma cells. In summary, our study reported VPS35 for the first time as a novel interacting partner and signaling regulator of RAS, and marks its potential as a therapeutic target for RAS-driven cancers.