RNA Droplets

Abstract

Liquid-liquid phase separation is emerging as the universal mechanism by which membraneless cellular granules form. Despite many previous studies on condensation of intrinsically disordered proteins and low complexity domains, we lack understanding about the role of RNA, which is the essential component of all ribonucleoprotein (RNP) granules. RNA, as an anionic polymer, is inherently an excellent platform for achieving multivalency and can accommodate many RNA binding proteins. Recent findings have highlighted the diverse function of RNA in tuning phase-separation propensity up or down, altering viscoelastic properties and thereby driving immiscibility between different condensates. In addition to contributing to the biophysical properties of droplets, RNA is a functionally critical constituent that defines the identity of cellular condensates and controls the temporal and spatial distribution of specific RNP granules. In this review, we summarize what we have learned so far about such roles of RNA in the context of in vitro and in vivo studies.

Keywords: RNA; RNA binding protein; aberrant aggregation; liquid–liquid phase separation; multivalency; polymer.

Figure 1

RNA droplets at every stage of formation and maturation. (a) RNA–RNA binding protein (RBP) contacts underlie the formation of ribonucleoprotein (RNP) complexes that contribute to droplet formation. Interactions are varied, and they can occur with any combination of structured and unstructured protein domains and RNA sequences. (b) Multivalent RNA–RBP interactions support the formation of phase-separated droplets. (c) Combination of different intrinsically disordered region (IDR) proteins and RNA allow for the formation of specific RNP granules with different cellular functions. (d) Accumulation of aberrant RNAs such as repeat expansion RNAs gives rise to aggregations via improper RNA–RNA and protein–RNA interactions that can lead to cellular toxicity and neurological disorders.

Figure 2

RNA is uniquely poised for liquid–liquid phase separation (LLPS). (a) Single-stranded RNA has a mixture of structured and unstructured regions, along with varied facets for charge–charge and other molecular interactions. (b) DNA has a protected duplex that limits interactions to the grooved or charged surface. (c) Proteins, like RNA, can be structured or unstructured and have varied opportunities to bind RNA.

Figure 3

RNA controls the physical properties of phase-separated droplets. (a) RNA seeds droplet formation through multivalent contacts with RNA binding protein (RBP) partners. After additional RNA–RBP interactions, droplet condensation can occur. (b) The viscoelastic properties of droplets can be tuned by the RNAs incorporated into the droplet. These RNAs can affect the kinetics of the droplet fusion event (viscosity) or the return to a normal circular morphology (elasticity). (c) Droplet size and circularity (also referred to as the aspect ratio) can be controlled by RNA content. (d) Heterotypic RNA droplets can have widely divergent viscoelastic properties that prevent mixing (miscibility) or lead to formation of biphasic droplets. (e) RNA exchange rates with the surrounding environment can vary depending on the RNA contacts within the droplet and the viscoelastic properties of the droplet.

Figure 4

RNA contributes to ribonucleoprotein (RNP) granule formation and function. RNA can assist in the formation of RNP granules in many different ways. (a) RNA serves as a scaffold for RNA–RNA and protein–RNA interactions to seed the formation of RNP granules. (b) Specific RNAs can tune the elasticity or fluidity of RNP granules, forming distinct RNP granules. (c) The targeted localization of RNAs can control the spatial and temporal formation of RNP granules. (d) RNAs such as micro RNAs and piwi-interacting RNAs can assist in the recruitment of messenger RNAs to RNP granules for translational control and proper cell specification and function.

Figure 5

Role of RNA in aberrant phase separation. Expansion repeat RNAs with abnormal structures can cause accumulation and aggregation of RNA and intrinsically disordered proteins in RNP granules to promote cellular toxicity and neurological disease. Abbreviations: IDR, intrinsically disordered region; RNP, ribonucleoprotein.