Assembly of Hierarchical Multiphase Condensates Using Designer Surfactant Proteins

Biomolecular condensates are complex fluids formed by liquid-liquid phase separation of macromolecules. Similar to other types of soft matter, they feature a range of biophysical properties that distinguish them from the cellular milieu. As a separate phase, they have an identifiable interface that dictates their interaction with the cytoplasm and other membraneless organelles. In this work, we engineer the interface of condensates to build novel hierarchical mesoscale structures from two immiscible disordered proteins: the RGG domain of LAF-1, a RNA-processing protein involved in germ granule assembly and the low complexity domain (LC) of FUS, an RNA binding protein whose aggregation is implicated in age-related neurodegeneration. RGG and FUS LC do not co-partition with one another and instead form discrete protein-rich condensed phases. Despite their apparent immiscibility, we identified conditions that can promote hierarchical assembly, either kinetically trapping one phase in the other, or using a designer surfactant protein that reduces the interfacial tension between the two phases. In addition, we studied factors that impact condensate miscibility and structure formation, including surface properties and viscoelasticity. This study probes the principles that underlie formation and assembly of complex structures from biomolecular condensates and provides a strategy for designing synthetic multiphasic materials capable of spatial partitioning.