Reentrant Phase Transitions and Non-Equilibrium Dynamics in Membraneless Organelles

Abstract

Compartmentalization of biochemical components, interactions, and reactions is critical for the function of cells. While intracellular partitioning of molecules via membranes has been extensively studied, there has been an expanding focus in recent years on the critical cellular roles and biophysical mechanisms of action of membraneless organelles (MLOs) such as the nucleolus. In this context, a substantial body of recent work has demonstrated that liquid-liquid phase separation plays a key role in MLO formation. However, less is known about MLO dissociation, with phosphorylation being the primary mechanism demonstrated thus far. In this Perspective, we focus on another mechanism for MLO dissociation that has been described in recent work, namely a reentrant phase transition (RPT). This concept, which emerges from the polymer physics field, provides a mechanistic basis for both formation and dissolution of MLOs by monotonic tuning of RNA concentration, which is an outcome of cellular processes such as transcription. Furthermore, the RPT model also predicts the formation of dynamic substructures (vacuoles) of the kind that have been observed in cellular MLOs. We end with a discussion of future directions in terms of open questions and methods that can be used to answer them, including further exploration of RPTs in vitro, in cells, and in vivo using ensemble and single-molecule methods as well as theory and computation. We anticipate that continued studies will further illuminate the important roles of reentrant phase transitions and associated non-equilibrium dynamics in the spatial patterning of the biochemistry and biology of the cell.

Figure 1

Scheme of reentrant phase transition leads to RNP droplet formation and dissolution. (a) Molecular representation shows the three interaction states in a simple system where RNA initially drives the formation of droplets, but subsequently drives dissolution via a charge inversion mechanism. This results in a change from short range electrostatic interactions to long range interactions. (b) Microscopic view showing that the titration of RNA:RLM ratio results in sequential transitions from a light (Phase I) to a dense (Phase II) to a light (Phase III). - Arginine Rich Linear Motif - ssRNA

Figure 2

Schematic of vacuole formation due to the reentrant phase transition. The left side shows nucleation of Phase II droplets during a transition from Phase I to Phase II. On the right side, vacuole formation results from nucleation of the light Phase III droplets inside the dense Phase II droplets during the transition from Phase II to Phase III. Vacuole lifetimes can further be controlled depending on jumps in RNA concentration with respect to the arginine-rich linear motif (RLM) concentration.

Figure 3

Schematic depicting the potential role of transcription. Transcription, which drives RNA synthesis, can drive droplet formation, charge inversion and subsequently vacuole formation, and complete droplet dissolution.