Heterotypic electrostatic interactions control complex phase separation of tau and prion into multiphasic condensates and co-aggregates

Significance

Biological phase separation stands at the crossroads between physiology and disease. Proteins and nucleic acids undergo intracellular phase separation to form dynamic, liquid-like, multicomponent, membraneless compartments that offer spatiotemporal control of cellular functions. However, aberrant phase transitions are associated with deadly human diseases. Here we show that two neuronal proteins, namely tau and prion proteins, can commingle into multicomponent liquid-like condensates via electrostatic crosslinks. Properties of these complex condensates can be modulated by RNA leading to a diverse range of morphologies. We also demonstrate that liquid-like condensates of tau and prion can gradually convert into solid-like amyloid species reminiscent of pathological aggregates. Our findings provide unique mechanistic insights into multicomponent macromolecular phase separation associated with physiology and overlapping neuropathological features.

Abstract

Biomolecular condensates formed via phase separation of proteins and nucleic acids are thought to perform a wide range of critical cellular functions by maintaining spatiotemporal regulation and organizing intracellular biochemistry. However, aberrant phase transitions are implicated in a multitude of human diseases. Here, we demonstrate that two neuronal proteins, namely tau and prion, undergo complex coacervation driven by domain-specific electrostatic interactions to yield highly dynamic, mesoscopic liquid-like droplets. The acidic N-terminal segment of tau interacts electrostatically with the polybasic N-terminal intrinsically disordered segment of the prion protein (PrP). We employed a unique combination of time-resolved tools that encompass several orders of magnitude of timescales ranging from nanoseconds to seconds. These studies unveil an intriguing symphony of molecular events associated with the formation of heterotypic condensates comprising ephemeral, domain-specific, short-range electrostatic nanoclusters. Our results reveal that these heterotypic condensates can be tuned by RNA in a stoichiometry-dependent manner resulting in reversible, multiphasic, immiscible, and ternary condensates of different morphologies ranging from core-shell to nested droplets. This ternary system exhibits a typical three-regime phase behavior reminiscent of other membraneless organelles including nucleolar condensates. We also show that upon aging, tau:PrP droplets gradually convert into solid-like co-assemblies by sequestration of persistent intermolecular interactions. Our vibrational Raman results in conjunction with atomic force microscopy and multi-color fluorescence imaging reveal the presence of amorphous and amyloid-like co-aggregates upon maturation. Our findings provide mechanistic underpinnings of overlapping neuropathology involving tau and PrP and highlight a broader biological role of complex phase transitions in physiology and disease.