Abstract

Many cellular proteins demix spontaneously from solution to form liquid condensates. These phase-separated systems have wide-ranging roles in health and disease. Elucidating the molecular driving forces underlying liquid-liquid phase separation (LLPS) is therefore a key objective for understanding biological function and malfunction. Here we show that proteins implicated in cellular LLPS, including FUS, TDP-43, Brd4, Sox2, and Annexin A11, which form condensates at low salt concentrations, can reenter a phase-separated regime at high salt concentrations. By bringing together experiments and simulations, we demonstrate that phase separation in the high-salt regime is driven by hydrophobic and non-ionic interactions, and is mechanistically distinct from the low-salt regime, where condensates are additionally stabilized by electrostatic forces. Our work thus provides a new view on the cooperation of hydrophobicity and non-ionic interactions as non-specific driving forces for the condensation process, with important implications for aberrant function, druggability, and material properties of biomolecular condensates.