Interaction networks within biomolecular condensates reveal structural and dynamic inhomogeneities

Abstract

Biomolecular condensates are membraneless organelles inside living cells that primarily comprise proteins and nucleic acids. The thermodynamic process of liquid-liquid phase separation has been proposed as a primary driver of biomolecular condensation, and it is recognized that phase separation is maintained by networks of biomolecular interactions within these liquid droplets. Canonical examples of condensing biomolecules include prion-like low-complexity domains (LCDs) of proteins, and simulations of single-component LCD condensates have predicted the presence of small-world topologies in the interaction networks underlying condensate stability. Recent experimental and theoretical works have also demonstrated inhomogeneities in single-molecule conformation, orientation, and dynamics within biomolecular condensates. Here, we systematically characterize the molecular networks underlying both LCD condensates and condensates formed by generic associative heteropolymers. Further, we investigate the relationship between network topologies and single-molecule properties within condensates. To probe LCD condensates, we employ a chemically specific, coarse-grained model of disordered proteins designed to reproduce phase separation statistics. We generalize our findings by varying sequence hydrophobicities using a generic binary model of associative heteropolymers, dubbed the “hydrophobic–polar” (HP) model. In both model systems, we find persistent small-world topologies underlying single-component condensates. These topologies feature molecular “hubs” with high network betweenness centrality and molecular “cliques” that represent densely interacting clusters of biomolecules; distal cliques in condensate volumes all localize to phase interfaces and are bridged by elongated hubs that remain near condensate centers. Strikingly, we find that relationships between network connectivity and biomolecular structure and dynamics are governed by power laws. Our work demonstrates that inhomogeneous single-molecule behaviors within biomolecular condensates can be well predicted from condensate network connectivities. Furthermore, we find that network cliques have substantially longer lifetimes than molecular hubs, and that the motion of molecules within cliques is spatially constrained. Together, these results reveal a dynamic hub-clique architecture underlying condensates and suggest that the physicochemical characteristics and material properties of phase interfaces are critical to pathological gelation and fibrillization processes observed in condensate aging.