Molecular Details of RNA Interactions With FUS Condensates Altering Assembly, Dynamics, and Aggregation

Fused in sarcoma (FUS) is an RNA-binding protein that undergoes phase separation with RNA and other cellular components, forming ribonucleoprotein (RNP) granules. While recent advances in the study of biomolecular phase separation have focused on protein-protein interactions, information on the molecular details of protein-RNA interactions within condensates remains limited. Here, we demonstrate how RNA modulates the phase separation of the low-complexity (LC) and arginine-glycine-glycine motif (RGG1) domains of FUS-low RNA concentrations enhance protein phase separation and excess RNA disrupts it. Integrating biochemical assays, NMR spectroscopy, and molecular dynamics simulations, we show that RNA incorporates into FUS condensates, reducing condensate density, drawing in water and ions, and enhancing local and diffusional motion. Whereas RNA binding in the dispersed phase primarily involves RGG1, within the condensed phase both LC and RGG1 contribute to RNA interactions. A diverse set of interactions between amino acids and RNA moieties, including prominent glutamine contacts, contributes to FUS-RNA co-condensates. RNA displaces RGG-mediated protein contacts while enhancing LC-LC interactions, providing a molecular basis for RNA-driven condensate interaction network remodeling. Furthermore, RNA accelerates the liquid-to-solid transition of FUS LC-RGG1 condensates. Together, these results provide mechanistic insight into how RNA regulates condensate assembly, dynamics, and maturation.