Intracellular organelles are either membrane-bound vesicles or membrane-less compartments that are made up of proteins and RNA. These organelles play key biological roles, by compartmentalizing the ...cell to enable spatiotemporal control of biological reactions. Recent studies suggest that membrane-less intracellular compartments are multicomponent viscous liquid droplets that form via phase separation. Proteins that have an intrinsic tendency for being conformationally heterogeneous seem to be the main drivers of liquid-liquid phase separation in the cell. These findings highlight the relevance of classical concepts from the physics of polymeric phase transitions for understanding the assembly of intracellular membrane-less compartments. However, applying these concepts is challenging, given the heteropolymeric nature of protein sequences, the complex intracellular environment, and non-equilibrium features intrinsic to cells. This provides new opportunities for adapting established theories and for the emergence of new physics.
Phase transitions are ubiquitous in nonliving matter, and recent discoveries have shown that they also play a key role within living cells. Intracellular liquid-liquid phase separation is thought to ...drive the formation of condensed liquid-like droplets of protein, RNA, and other biomolecules, which form in the absence of a delimiting membrane. Recent studies have elucidated many aspects of the molecular interactions underlying the formation of these remarkable and ubiquitous droplets and the way in which such interactions dictate their material properties, composition, and phase behavior. Here, we review these exciting developments and highlight key remaining challenges, particularly the ability of liquid condensates to both facilitate and respond to biological function and how their metastability may underlie devastating protein aggregation diseases.
The coordinated growth of cells and their organelles is a fundamental and poorly understood problem, with implications for processes ranging from embryonic development to oncogenesis. Recent ...experiments have shed light on the cell size-dependent assembly of membrane-less cytoplasmic and nucleoplasmic structures, including ribonucleoprotein (RNP) granules and other intracellular bodies. Many of these structures behave as condensed liquid-like phases of the cytoplasm/nucleoplasm. The phase transitions that appear to govern their assembly exhibit an intrinsic dependence on cell size, and may explain the size scaling reported for a number of structures. This size scaling could, in turn, play a role in cell growth and size control.
Just as organ size typically increases with body size, the size of intracellular structures changes as cells grow and divide. Indeed, many organelles, such as the nucleus 1, 2, mitochondria 3, ...mitotic spindle 4, 5, and centrosome 6, exhibit size scaling, a phenomenon in which organelle size depends linearly on cell size. However, the mechanisms of organelle size scaling remain unclear. Here, we show that the size of the nucleolus, a membraneless organelle important for cell-size homeostasis 7, is coupled to cell size by an intracellular phase transition. We find that nucleolar size directly scales with cell size in early C. elegans embryos. Surprisingly, however, when embryo size is altered, we observe inverse scaling: nucleolar size increases in small cells and decreases in large cells. We demonstrate that this seemingly contradictory result arises from maternal loading of a fixed number rather than a fixed concentration of nucleolar components, which condense into nucleoli only above a threshold concentration. Our results suggest that the physics of phase transitions can dictate whether an organelle assembles, and, if so, its size, providing a mechanistic link between organelle assembly and cell size. Since the nucleolus is known to play a key role in cell growth, this biophysical readout of cell size could provide a novel feedback mechanism for growth control.
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•Nucleolar size scales directly with cell size during development•Nucleolar size scales inversely with cell size across RNAi conditions•Nucleolar size and assembly depend on the concentration of components•Direct and inverse scaling regimes can be explained by a phase transition model
Weber and Brangwynne show that nucleolar size is determined by the concentration of maternally loaded components, which condense into nucleoli only above a threshold concentration. This work suggests that intracellular phase transitions may provide a general mechanism for organelle assembly that inherently couples organelle size with cell size.
Getting RNA and Protein in Phase Weber, Stephanie C.; Brangwynne, Clifford P.
Cell,
06/2012, Letnik:
149, Številka:
6
Journal Article
Recenzirano
Odprti dostop
Nonmembrane-bound organelles such as RNA granules behave like dynamic droplets, but the molecular details of their assembly are poorly understood. Several recent papers identify structural features ...that drive granule assembly, shedding light on how phase transitions functionally organize the cell and may lead to pathological protein aggregation.
The cell nucleus contains a large number of membrane-less bodies that play important roles in the spatiotemporal regulation of gene expression. Recent work suggests that low complexity/disordered ...protein motifs and repetitive binding domains drive assembly of droplets of nuclear RNA/protein by promoting nucleoplasmic phase separation. Nucleation and maturation of these structures is regulated by, and may in turn affect, factors including post-translational modifications, protein concentration, transcriptional activity, and chromatin state. Here we present a concise review of these exciting recent advances, and discuss current and future challenges in understanding the assembly, regulation, and function of nuclear RNA/protein bodies.
Compartmentalization in cells is central to the spatial and temporal control of biochemistry. In addition to membrane-bound organelles, membrane-less compartments form partitions in cells. Increasing ...evidence suggests that these compartments assemble through liquid-liquid phase separation. However, the spatiotemporal control of their assembly, and how they maintain distinct functional and physical identities, is poorly understood. We have previously shown an RNA-binding protein with a polyQ-expansion called Whi3 is essential for the spatial patterning of cyclin and formin transcripts in cytosol. Here, we show that specific mRNAs that are known physiological targets of Whi3 drive phase separation. mRNA can alter the viscosity of droplets, their propensity to fuse, and the exchange rates of components with bulk solution. Different mRNAs impart distinct biophysical properties of droplets, indicating mRNA can bring individuality to assemblies. Our findings suggest that mRNAs can encode not only genetic information but also the biophysical properties of phase-separated compartments.
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•RNA drives phase transition of Whi3, an RNA-binding protein with a long polyQ tract•RNA alters Whi3 droplet viscosity, dynamics, and their propensity to fuse•Different target mRNAs drive Whi3 to form droplets with distinct properties•Whi3 droplets mature and appear fibrillar over time
Zhang et al. show that RNAs drive an RNA-binding protein with a polyQ expansion to undergo a phase transition to form dynamic droplets. RNA sequences influence droplet biophysical properties and may help keep aggregation-prone proteins in functional liquid-like states and avoid fibrous and potentially toxic aggregate states.
The size of a typical eukaryotic cell is of the order of ∼10 μm. However, some cell types grow to very large sizes, including oocytes (immature eggs) of organisms from humans to starfish. For ...example, oocytes of the frog Xenopus laevis grow to a diameter ≥1 mm. They have a correspondingly large nucleus (germinal vesicle) of ∼450 μm in diameter, which is similar to smaller somatic nuclei, but contains a significantly higher concentration of actin. The form and structure of this nuclear actin remain controversial, and its potential mechanical role within these large nuclei is unknown. Here, we use a microrheology and quantitative imaging approach to show that germinal vesicles contain an elastic F-actin scaffold that mechanically stabilizes these large nuclei against gravitational forces, which are usually considered negligible within cells. We find that on actin disruption, ribonucleoprotein droplets, including nucleoli and histone locus bodies, undergo gravitational sedimentation and fusion. We develop a model that reveals how gravity becomes an increasingly potent force as cells and their nuclei grow larger than ∼10 μm, explaining the requirement for a stabilizing nuclear F-actin scaffold in large Xenopus oocytes. All life forms are subject to gravity, and our results may have broad implications for cell growth and size control.
Phase transitions driven by intrinsically disordered protein regions (IDRs) have emerged as a ubiquitous mechanism for assembling liquid-like RNA/protein (RNP) bodies and other membrane-less ...organelles. However, a lack of tools to control intracellular phase transitions limits our ability to understand their role in cell physiology and disease. Here, we introduce an optogenetic platform that uses light to activate IDR-mediated phase transitions in living cells. We use this “optoDroplet” system to study condensed phases driven by the IDRs of various RNP body proteins, including FUS, DDX4, and HNRNPA1. Above a concentration threshold, these constructs undergo light-activated phase separation, forming spatiotemporally definable liquid optoDroplets. FUS optoDroplet assembly is fully reversible even after multiple activation cycles. However, cells driven deep within the phase boundary form solid-like gels that undergo aging into irreversible aggregates. This system can thus elucidate not only physiological phase transitions but also their link to pathological aggregates.
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•OptoDroplets enable light-activatable control of intracellular phase transitions•Rapid growth and fast inactivation lead to droplet assembly in subcellular regions•Cells driven to deep supersaturation form solid-like gels•Gels are initially reversible but undergo aging into irreversible aggregates
Light-dependent triggering of protein association allows selective temporal and spatial control of droplet and gel formation aimed at understanding the different forms of membrane-less bodies and fibrillar structures within cells.
Cells compartmentalize their intracellular environment to orchestrate countless simultaneous biochemical processes. Many intracellular tasks rely on membrane-less organelles, multicomponent ...condensates that assemble by liquid-liquid phase separation. A decade of intensive research has provided a basic understanding of the biomolecular driving forces underlying the form and function of such organelles. Here we review the technologies enabling these developments, along with approaches to designing spatiotemporally actuated organelles based on multivalent low-affinity interactions. With these recent advances, it is now becoming possible both to modulate the properties of native condensates and to engineer entirely new structures, with the potential for widespread biomedical and biotechnological applications.
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