Cytoplasmic protein granules organize kinase-mediated RAS signaling

Abstract The spatial organization of a cell includes lipid membrane-based compartments and an emerging class of subcellular structures collectively described as biomolecular condensates1. Lipid membranes act as a biologically active scaffold to concentrate signaling molecules in multiple signal transduction pathways that regulate normal physiology and pathologic conditions such as cancer2. Notably, receptor tyrosine kinase (RTK)-mediated RAS GTPase/MAP kinase (MAPK) pathway signaling is thought to occur exclusively from lipid membrane compartments in mammalian cells3,4. Here, we uncover a membraneless, protein granule-based subcellular structure that can organize RTK/RAS/MAPK signaling. Chimeric (fusion) oncoproteins involving certain RTKs including ALK and RET undergo de novo assembly into cytoplasmic protein granules that locally concentrate the RAS activating complex GRB2/SOS1 and activate RAS in a lipid membrane-independent manner to initiate MAPK signaling. We show that formation of higher-order membraneless protein granules is both necessary and sufficient for RAS/MAPK signaling output in cells. These large-scale protein assemblies are functionally distinct from lower-order oligomerization of cytoplasmic RTK fusion oncoproteins. Our findings reveal membraneless, higher-order cytoplasmic protein assembly as a subcellular platform to activate RAS GTPases and a distinct principle by which cells can organize kinase-mediated oncogenic signaling.