Localized control of protein phase separation via membrane binding

Phase separation of protein-rich condensates is key for the spatial organization of cells. Multivalent proteins can phase separate in the bulk cytoplasm to form 3D condensates or at the membrane surface to form 2D condensates. How cells control 2D versus 3D phase transitions is not well understood. Here, we combine an in vitro model system of membrane surface phase separation with thermodynamic modelling to explore the relation of 2D and 3D phase transitions. We engineered the phase-separating protein FUS to bind to membranes and quantified 2D and 3D phase separation as a function of total FUS concentration and salt concentration. We find that membrane binding induces the formation of 2D surface condensates far below the bulk saturation concentration. In addition to direct FUS-membrane binding, surface condensates sequester more FUS molecules from the bulk via protein-protein interactions, resulting in a substantial deviation from the classical binding isotherm. By changing the protein-protein interaction strength via salt titration, we find that the saturation concentrations for 2D and 3D phase separation are coupled. By extending the classical theory of condensate wetting via membrane binding, we recapitulate the experimental results and show that tuning the membrane-binding strength of a phase-separating protein provides a robust way to control 2D condensation at the membrane over a wide concentration range without entering the 3D condensation regime. Taken together, our results provide a simple framework to understand how cells tune protein interactions and membrane binding to control 2D and 3D phase transitions.