Physical observables to determine the nature of membrane-less cellular sub-compartments

TL;DR

This paper presents a theoretical framework to distinguish between liquid-liquid phase separation and binding site mechanisms in membrane-less cellular compartments using single molecule tracking, aiding understanding of their formation and biological roles.

Contribution

It introduces a novel theoretical approach and observables to empirically differentiate between phase separation and binding site models in cellular foci formation.

Findings

Protein search times are reduced inside liquid droplets.

Binding site aggregates do not facilitate faster protein search.

The framework guides future experimental differentiation of foci mechanisms.

Abstract

The spatial organization of complex biochemical reactions is essential for the regulation of cellular processes. Membrane-less structures called foci containing high concentrations of specific proteins have been reported in a variety of contexts, but the mechanism of their formation is not fully understood. Several competing mechanisms exist that are difficult to distinguish empirically, including liquid-liquid phase separation, and the trapping of molecules by multiple binding sites. Here we propose a theoretical framework and outline observables to differentiate between these scenarios from single molecule tracking experiments. In the binding site model, we derive relations between the distribution of proteins, their diffusion properties, and their radial displacement. We predict that protein search times can be reduced for targets inside a liquid droplet, but not in an aggregate of slowly moving binding sites. These results are applicable to future experiments and suggest different biological roles for liquid droplet and binding site foci.