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Liquid phase separation

Tools are needed to study phase behaviors of membraneless compartments in living cells.

Our everyday experience with water and oil droplets illustrates a simple liquid–liquid phase separation (LLPS). In biology, cells resemble liquid droplets that maintain membraneless compartments, important for concentrating certain molecules and facilitating spatiotemporal regulation of cellular functions. Such structures originate via active LLPS, and this process is ubiquitous in both cytoplasm and nucleoplasm.

Intracellular liquid–liquid phase separation. Credit: Marina Corral Spence/Springer Nature

To understand the LLPS mechanism, scientists have applied classical concepts from polymer physics to enlighten the molecular driving forces behind the assembly behaviors (Nat. Phys. 11, 899–904; 2015). Experimental approaches that generate liquid droplets containing purified proteins of interest have also emerged. For example, a microfluidic device offers a microenvironment for examining the intrinsic associations of protein sequences with LLPS, as well as the extrinsic effects of protein concentration, salt concentration, and temperature (Nat. Chem. 9, 509–515; 2017).

To date, it has been a challenge to translate the knowledge gained from purified proteins into the cellular milieu, which is crowded with macromolecules, small molecules, and complex structures. For measurement of the mesoscopic properties of cellular liquid droplets, current methods rely mainly on fluorescence microscopy to visualize the size and shape of aggregates. Although tools capable of reversibly triggering phase transitions in cells remain sparse, a promising optogenetic platform has been described (Cell 168, 159–171; 2017). Another mesoscopic property, viscosity, plays a critical role in molecule diffusion within and around liquid droplets, and ultrafast-scanning fluorescence correlation spectroscopy has been used to determine droplet viscosity (Nat. Chem. 9, 1118–1125; 2017).

Beyond the phase behavior in the cytoplasm, LLPS is also of interest in nuclear condensates. Two studies have demonstrated that heterochromatin protein 1 has the ability to form phase-separated droplets that mediate the compaction of DNA into a repressive structure (Nature 547, 236–240, 2017; Nature 547, 241–245, 2017).

The combination of methods used to study LLPS and 3D genome structure (such as Hi-C) could be powerful. The collective perspective might shed light on the connection between nuclear condensates and gene regulation.

We anticipate that more tools drawing from polymer physics to genomics will emerge to elucidate the molecular mechanism for liquid phase separation and its functional consequences in biology.

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Correspondence to Lei Tang.

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Tang, L. Liquid phase separation. Nat Methods 16, 18 (2019). https://doi.org/10.1038/s41592-018-0269-7

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