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• W4255830188 abstract "Review29 June 2016free access Droplet organelles? Edward M Courchaine Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA Search for more papers by this author Alice Lu Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA Search for more papers by this author Karla M Neugebauer Corresponding Author Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA Search for more papers by this author Edward M Courchaine Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA Search for more papers by this author Alice Lu Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA Search for more papers by this author Karla M Neugebauer Corresponding Author Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA Search for more papers by this author Author Information Edward M Courchaine1, Alice Lu1 and Karla M Neugebauer 1 1Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA *Corresponding author. Tel: +1 203 785 3322; E-mail: [email protected] EMBO J (2016)35:1603-1612https://doi.org/10.15252/embj.201593517 See the Glossary for abbreviations used in this article. PDFDownload PDF of article text and main figures. ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract Cells contain numerous, molecularly distinct cellular compartments that are not enclosed by lipid bilayers. These compartments are implicated in a wide range of cellular activities, and they have been variously described as bodies, granules, or organelles. Recent evidence suggests that a liquid–liquid phase separation (LLPS) process may drive their formation, possibly justifying the unifying term “droplet organelle”. A veritable deluge of recent publications points to the importance of low-complexity proteins and RNA in determining the physical properties of phase-separated structures. Many of the proteins linked to such structures are implicated in human diseases, such as amyotrophic lateral sclerosis (ALS). We provide an overview of the organizational principles that characterize putative “droplet organelles” in healthy and diseased cells, connecting protein biochemistry with cell physiology. Glossary Many of the terms used in the recent literature have meanings that are overlapping or refer to subtle differences between concepts. Here, we provide the classical definitions for these terms and comment on their usage. Liquid–liquid phase separation (LLPS) It is the phenomenon in which solutes spontaneously separate into a demixed liquid phase suspended within the bulk solvent. Conventionally, the solute is a flexible chain polymer, but the term is also applied to biological macromolecules that may not have a flexible chain-like tertiary structure. Low-complexity domain (LCD) It is a region within a protein that contains an overrepresentation of a subset of amino acids in the primary sequence. Often this occurs as a repeat motif, but repeats are not a requirement. Intrinsically disordered regions (IDRs) These are the protein domains, often containing low-complexity sequences that appear to lack well-defined secondary and tertiary structure. Some IDRs have been determined experimentally, while others are inferred and may be structured in certain contexts. Droplet It is the spherical fluid morphology adopted by phase-separated macromolecules in solution. Droplets have measureable surface tension and viscosity. Molecular constituents diffuse within them and can exchange with the bulk solvent. Hydrogel It is the hydrated matrix formed by cross-linked protein polymers. These polymers are best thought of as a stable colloidal solid suspended in water. Aggregate It is a solid formation composed of proteins that have precipitated from solution. Precipitation occurs because water is excluded from macromolecular interactions to the extent that the protein mass is no longer stably suspended Amyloid It is a class of protein aggregate characterized by a semi-regular structure formed by the stacking of β sheets among protein monomers in trans. They are experimentally identified by characteristic X-ray diffraction patterns and staining with the dye, thioflavin T. Prion-like domain It is a protein region characterized by sequence similarity to that of prototypical yeast prion proteins domains. These can be thought of as a special case of low-complexity domain Introduction Cells spatially organize biochemical reactions, a characteristic that is fundamental to life but often evades analysis. Lipid membranes create discrete chemical environments within canonical organelles and achieve separation of constituents from the bulk cytoplasm. Enclosing membrane-bound compartments requires dedicated machinery to construct and maintain the lipid bilayer and transport substances across them, thus expending energy (Grossman et al, 2012; Neupert, 2015). Many organelles lack lipid bilayers, circumventing these issues and introducing the potential for greater dynamics and alternative mechanisms of regulation. Nuclear structures, such as nucleoli and Cajal bodies, regulate ribonucleoprotein (RNP) assembly in this manner (Mao et al, 2011; Machyna et al, 2013). In the cytoplasm, stress granules and P bodies regulate RNA stability and protein translation in response to cellular stimuli (Buchan & Parker, 2009). Accumulating evidence suggests that such non-membrane-bound organelles behave as fluid droplets, which undergo LLPS (see Glossary). The molecular mechanisms that underlie their form and function are currently under intense investigation, and this review focuses on the emergent common principles. We survey the literature on systems that exhibit LLPS and related behavior in order to bridge the in vivo studies of cellular structures and the molecular understanding afforded by in vitro studies of proteins, RNAs, and their interactions. Our focus will be on the most recent work that has shed light on the molecular mechanisms that lead to LLPS, but we also reference previous key studies to provide a prelude to recent developments. The review closes by returning to the biological consequences of competing models for LLPS systems and discusses how they may relate to diseases characterized by the formation of aberrant protein aggregates. Our goal is to synthesize an outlook for the relevance of LLPS at both molecular and cellular scales—a number of excellent recent reviews provide a more detailed analysis of the individual topics covered (Weber & Brangwynne, 2012; Malinovska et al, 2013; Hyman et al, 2014; Toretsky & Wright, 2014; Uversky, 2015). We emphasize several unresolved issues in the field and attempt to address controversy where it arises. Nuclear and cytoplasmic bodies are the sites of RNP biogenesis Many non-membrane-bound organelles can be found in the nucleus and were first observed there over a century ago (Gall, 2000). Figure 1 illustrates how a few of these organelles are positioned. Nuclear bodies have been investigated extensively regarding the exchange of constituents with surrounding nucleoplasm. For most components, residence times are on the order of seconds with the most stable constituents exchanging within tens of seconds (Phair & Misteli, 2000; Dundr et al, 2004). The dynamic nature of nuclear bodies likely underlies their function, which the most recent studies of LLPS strive to explain. The most prominent nuclear body is the nucleolus, which forms on active rDNA loci and is the site of pre-rRNA processing and pre-ribosomal subunit assembly (Boisvert et al, 2007; Pederson, 2011; Falahati et al, 2016). Similarly, the Cajal body forms on active snRNA loci and is the site of snRNA processing, snRNP assembly, and snRNP surveillance (Frey et al, 1999; Stanek & Neugebauer, 2004; Machyna et al, 2013; Novotny et al, 2015). Nuclear speckles and paraspeckles are more granular in morphology, contain mRNAs and their binding proteins, and form on two long non-coding RNAs (lncRNAs) MALAT1 and NEAT1, respectively (Mao et al, 2011). Many of the specific functions of these subdomains are still poorly characterized, but at the descriptive level, they are consistent with phase-separated systems. Figure 1. In homeostatic cellular conditions, dynamic fluid droplets demix from surrounding nucleoplasm(A) Nucleoli, Cajal bodies (CBs), histone locus bodies (HLBs), speckles, and paraspeckles participate in RNA and RNP biogenesis in the nucleus. Associated with chromosomal loci, these nuclear bodies contain specific RNAs and proteins that pass in and out of nuclear bodies during RNP assembly. Unstable RNAs concentrate in P bodies in the cytoplasm, where mRNA decay factors co-localize. (B) Analogous dynamics and fluid properties are obtained when a purified RNA-binding protein with a low-complexity region is incubated in with RNA and observed over time in vitro. (C) Electron micrograph of a droplet showing overall spherical shape with an irregular outline. Micrographs reproduced from Li et al (2012). Download figure Download PowerPoint Cytoplasmic bodies are more granular in morphology and have functions often related to translational control and/or mRNA stability. The processing body (P body) falls into this second category, in which translation is stalled and transcripts are targeted for degradation by exonucleases (Parker & Sheth, 2007) or selective reactivation of translation (Arribere et al, 2011). Stress granules are related to P bodies, in that they contain translationally repressed mRNA, but form in response to heat, osmotic, and chemical stress stimuli. Figure 2 illustrates changes seen in several bodies during cellular stress including the altered structure of the nucleolus. Stress granules are an example of a phase-separated structure where the physical properties change in response to a stimulus (Boulon et al, 2010). Others including nuclear speckles, histone locus bodies, and paraspeckles also respond to stress with a variety of morphological and biochemical changes (Carmo-Fonseca et al, 1992; Lamond & Spector, 2003; Bongiorno-Borbone et al, 2010; Boulon et al, 2010). Stress granules contain factors that stall translation, as well as a number of RNA-binding proteins, associated with ALS and other diseases. The normal physiological roles of many stress granule proteins are still unclear (Anderson & Kedersha, 2008; Li et al, 2013). Figure 2. During cellular stress, hydrogel-like assemblies form(A) Stressors like heat shock, UV light, and transcription inhibition can cause cellular bodies to disassemble (CBs), change morphology (nucleoli, nuclear speckles), or appear de novo (stress bodies). (B) These granules contain proteins that form hydrogels and ultimately form an intricate proteinaceous network or aggregate (C). Micrographs reproduced from Hennig et al (2015). Download figure Download PowerPoint Some cytoplasmic RNA granules are developmentally important. In C. elegans, the germ cell lineage is specified by the asymmetric inheritance of P granules during mitosis. The exact function of these RNP granules is unknown, but they have some similarity to P bodies and are functionally implicated in translational control of the germ line (Seydoux & Braun, 2006). In mammals, germ line specification follows a different mechanism, but germ cells still display characteristic RNP structures known as nuage or the chromatoid body. Nuage contains many RNAs as well as several helicases, endonucleases, and proteins involved in miRNA-mediated degradation of RNA (Kotaja & Sassone-Corsi, 2007). RNP bodies and granules display characteristics of phase-separated liquids Even as progress has been made on elucidating the biological functions of these nuclear and cytoplasmic bodies, their physical properties have only recently come to light through high-resolution live-cell imaging. Like nucleoli, Cajal bodies fuse and split (Platani et al, 2000). In 2005, Gall and colleagues speculated that Cajal bodies were actually “semi-fluid spheres suspended in semi-fluid nucleoplasm” (Handwerger et al, 2005). This conclusion was based on observed shape, permeability, and differential protein concentrations between the nucleoplasm and the large Cajal bodies present in Xenopus germinal vesicles. In 2009, seminal work on the P granules of C. elegans demonstrated their liquid-like properties and that they localize to the future germ cell cytoplasm by dissolving and condensing rather than by moving as discrete objects through the cytoplasm (Brangwynne et al, 2009). Extrachromosomal nucleoli of Xenopus laevis were also shown to exhibit properties expected of liquid droplets, including surface tension and fluidity (Brangwynne et al, 2011). Advanced microscopy techniques, including tracking of subcellular structures over time, turned the conjecture that nuclear and other organelles undergo LLPS into a well-characterized phenomenon. These apparent manifestations of LLPS in vivo were striking and harken back to studies of other biomolecules that undergo phase separation in vitro. In 2006, Görlich and colleagues showed that the yeast FG (phenylalanine–glycine) repeat proteins of the nuclear pore complex are capable of condensing into a hydrated gel matrix (Frey et al, 2006). These gels showed no fluorescence recovery after photobleaching (FRAP), suggesting that the hydrogel is not fluid and instead may trap constituent molecules in an immobile meshwork. Cross-β amyloid-like interactions are likely to underlie hydrogel formation (see Glossary) and are disrupted by nuclear transport factors. This “melting” of the gel matrix by some proteins but not others effectively constitutes a selectivity filter (Ader et al, 2010; Schmidt & Görlich, 2015). Species other than fungi are less likely to have FG nucleoporins that show strong amyloid character, but all rely on hydrophobic interactions in their formation (Labokha et al, 2013; Schmidt & Görlich, 2015). This in vitro molecular behavior is distinct from that observed for constituents of Cajal bodies and nucleoli in vivo. Specifically, the GFP florescence recovery of coilin and SMN, two core components of Cajal bodies, indicates that they exchange rapidly with bulk nucleoplasm (Phair & Misteli, 2000; Dundr et al, 2004; Brangwynne et al, 2011), which would not be expected of hydrogels. Comparing the findings of in vitro studies of the nuclear pore components and of nuclear bodies in vivo, it would seem that there are two different ways that biological systems can separate from the bulk solvent: either as solid hydrogels or as LLPS droplets. In vitro systems show fundamental properties of phase separation New assays have been devised to probe the underlying physical and molecular nature of phase-separating systems. Using purified protein domains derived from signaling proteins of the N-WASP pathway, Rosen and colleagues were able to show that multivalency is sufficient to drive the phase separation of concatenated SH3 domains that bind to concatenated proline-rich motifs (Li et al, 2012). Figure 1B depicts the fusion of droplets containing concatenated SH3 domains (Li et al, 2012), consistent with the proposal that they possess fluid properties. In the same study, RNA with multiple binding motifs was also able to form droplets in combination with the multivalent protein PTB, which exhibit FRAP and form above a critical concentration threshold. Other work has also shown that the properties of phase-separated proteins depend heavily upon phosphorylation state (Kwon et al, 2013, 2014; Wang et al, 2014; Su et al, 2016), as discussed below. One investigation into these effects on FUS, an RNA- and DNA-binding protein found in both cytoplasm and nucleoplasm, highlighted the importance of post-translational modification as a means of regulating phase separation and implicated low-complexity protein domains (LCDs; see Glossary) as fundamental to body formation (Kato et al, 2012). Proteins with low-complexity domains are often found in bodies The bodies so far discussed almost all contain at least one, if not several, disordered proteins that notably contain characteristic LCDs. Many of these proteins are listed in Table 1. It is worth clarifying some of the terminology that has been used to describe LCDs: Several studies have designated LCD sequences based on their sequence similarity to the yeast prion proteins (see Glossary) (Alberti et al, 2009; Lancaster et al, 2014). These so-called prion-like domains seem to be related to important diseases and have a propensity to aggregate (King et al, 2012); however, this term is too narrow for the present discussion of phase-separating systems. “Low-complexity domain” is currently the most satisfying term available because it includes the prion-like domains as well as disordered domains with other amino acid compositions that might be important for forming phase-separated systems like stress granules and P bodies (Decker et al, 2007; Sun et al, 2011). Table 1. Protein components of cellular compartments that phase separate in vitro Protein (aa) Low-complexity motif(s) Structured domains Cellular compartment In vitro morphology Disease association References DDX4 (724) 19aa R/G-rich 16aa R/G/S 9aa P-rich DEXDc HELICc Nuage RNP granules Droplets* N/A Nott et al (2015) eIF4GII (914) yeast 38aa N/K/Y 21aa T/P 13aa S/R 17aa E/A 29aa N/S MIF4G Stress granules Droplets* N/A Lin et al (2015), Molliex et al (2015) EWS (656) 19aa Y/A/S 82aa A/Q/T 85aa Q/S/Y 33aa R/G 22aa D-rich 60aa R/G/P 86aa R/G/P RRM ZnF_RBZ DNA damage sites Hydrogel* ALS Kwon et al (2013), Altmeyer et al, (2015) Fibrillarin (321) 72aa R/G Methyl-transferase (fibrillarin) Nucleolus Cajal body Droplets Autoimmunity Berry et al (2015) FUS (526) 156aa S/G/Q 55aa RGG 76aa RGG RRM ZnF Paraspeckles stress granules Droplets hydrogel ALS Altmeyer et al (2015), Burke et al (2015), Murakami et al (2015), Patel et al (2015) hnRNP A1/A2 (372/353) 180aa G/S/R/Q 2 RRMs Stress granules Droplets hydrogel ALS, IBM, Paget's, FTLD Kim et al (2013), Lin et al (2015), Molliex et al (2015), Xiang et al (2015) LAF-1 (708) C. elegans 130aa G/R 75aa G/R/Q DEXDc HELICc P granules Droplets N/A Elbaum-Garfinkle et al (2015) Lsm4 (187) yeast 173aa N/R Sm P bodies stress granules Droplets* N/A Lin et al (2015) PTB (557) 13aa S/N/A 18aa A-rich 30aa A-rich 4 RRMs Nuclear speckles peri-nucleolar Droplets N/A Li et al (2012), Lin et al (2015) Pub1 (453) yeast 55aa N/M 33aa Q-rich 3 RRMs Stress granules Droplets* N/A Lin et al (2015) RBM14 (669) 300aa A/R/S/Q/P 2 RRMs Paraspeckles Hydrogel ALS Hennig et al (2015) SRSF2 (221) 113aa R/S RRM Nuclear speckles Hydrogel MDS leukemias Kwon et al (2014) TAF15 (589) 148 S/G/Q/Y 39 RGG 26aa R/G 186aa RGG/YGG RRM ZnF_RBZ DNA damage sites Hydrogel* ALS FTLD Kwon et al (2013), Altmeyer et al (2015) TDP-43 (414) 43aa G/F/N 9aa A-rich 17aa Q/N 37aa S/G 2 RRMs Stress granules Droplets ALS Burke et al (2015), Molliex et al (2015) Tia1 (386) 23aa S/T/Q 3 RRMs Stress granules Droplets* Welander distal myopathy Lin et al (2015) Whi3 (729) A. gossypii Poly-Q RRM Cytoplasmic RNP granules Droplets* N/A Zhang et al (2015) Protein and its amino acid length (aa) are listed. Unless otherwise noted below the protein name, all proteins are human. “Yeast” is shorthand for Saccharomyces cerevisiae. Low-complexity motifs are listed in the order of primary sequence, and amino acid enrichment is indicated. Structured domains are according to the SMART database. “Cellular compartment” refers to the structure in which the protein concentrates in vivo, in the absence of mutation. Morphologies noted with an asterisk have only been determined using a fragment of the protein, rather than full length. Disease association signifies the protein's involvement in human disease, not necessarily a disease of protein aggregation; N/A in this category means not applicable (e.g., for a non-human protein) or none known. Two proteins containing LCDs have come to the forefront of much recent research due to their relevance to ALS and frontotemporal dementia: FUS and hnRNPA1. FUS is associated with several nuclear and cytoplasmic bodies, including paraspeckles and stress granules. Under normal conditions, FUS is found in the nucleus where it takes part in DNA repair and transcriptional regulation (Wang et al, 2008, 2013). For reasons that are not entirely clear, stress stimuli result in the export of FUS to the cytoplasm where it joins stress granules (Bentmann et al, 2012). These studies have identified the importance of both RNA-binding properties and the presence of the LCD within FUS. Similar to FUS, nuclear hnRNPA1 has two RNA recognition motifs (RRMs) and an LCD, is recruited to stress granules, and phase separates in vitro (Kim et al, 2013; Kwon et al, 2014). Last year, significant inroads were made into understanding how FUS, hnRNPA1, and other LCD proteins (see Table 1) can generate fluid cellular bodies as outlined in several reviews (Malinovska et al, 2013; Toretsky & Wright, 2014; Uversky, 2015). Electrostatic interactions and LLPS What causes an LCD to transition into either a liquid droplet or a solid hydrogel is a matter of debate, and the recent literature considers several contributing factors (Burke et al, 2015; Lin et al, 2015; Xiang et al, 2015). Isolated LCDs from a variety of proteins that scaffold cellular structures are sufficient for phase-separating behavior. Strikingly, the low-complexity sequences of eIF4GII, hnRNPA1, and FUS all form liquid droplets without the addition of any other components (Lin et al, 2015). The integrity of the full low-complexity domain is essential for the formation of paraspeckles, and a tyrosine to serine mutation in the repeat motif in RBM14 clearly alters the morphology of the resulting hydrogel (Hennig et al, 2015). Figure 2B and C shows the hydrogel cross-linking that may occur during droplet maturation, a process described for numerous intrinsically disordered regions (IDRs) in vitro (see Glossary). Typically, cross-linking is induced over time by manipulating protein and/or salt concentration, temperature, and molecular crowding (Lin et al, 2015; Nott et al, 2015). This is in contrast to the fluid droplets shown in Fig 1B and C, which have a notably different structure as seen in electron micrographs. Thus, electrostatic interactions may provide an underlying force for LLPS, though the evidence for this is taken from experiments that were performed above physiological protein concentrations or below physiological salt concentrations. It remains to be seen whether hydrogel formation and stability also depend on polar contacts. By demonstrating that the two-phase state of these systems requires a low ionic strength, these studies allow us to speculate on the importance of charge and polarity in the composition of LCDs as well as the relevance of post-translational modifications that can render amino acids either more or less charged. For example, hydrogel recruitment of the SRSF2 LCD, which is serine and arginine-rich, is blocked by phosphorylation (Kwon et al, 2014). Intriguingly, phosphorylation can also alter the range of structural ensembles and binding interactions for some IDRs (Arai et al, 2015), suggesting that some IDRs may become less disordered or even structured when post-translationally modified. Experiments employing phosphomimetic or alanine replacements could provide decisive evidence here. While existing data indicate that electrostatic interactions between LCDs are important, we have yet to address the fact that LLPS often coincides with the presence of another charged macromolecule: RNA. Roles for RNA in vitro and in vivo Until recently, the role of RNA in droplet formation had been relatively neglected. New studies re-invigorate considerations of RNA's role in vitro and in vivo. In considering how electrostatic interactions might mediate the formation of liquid droplets, recall that RNA is concentrated in most bodies and granules in vivo. Because of its anionic phosphate backbone, RNA is a highly charged molecule that can potentially contribute to electrostatic interactions with positively charged residues in LCDs. Indeed, RNA enhances the fluid properties of droplets formed by the P granule component LAF1, which contains positively charged arginine–glycine–glycine (RGG) repeats (Elbaum-Garfinkle et al, 2015). This increase in fluidity might be related to a general promotion of LLPS by RNA, as is also observed in the formation of fibrillarin droplets (Berry et al, 2015). Poly-(ADP ribose), or PAR, is a polynucleotide synthesized upon DNA damage; PAR acts as a signal in the localization of the DNA repair machinery to sites of DNA damage (Table 1). The chemical structure of PAR strongly resembles RNA and appears to nucleate liquid phase-separated regions through electrostatic interactions with RGG repeats found in FUS, EWS, and TAF15 (Altmeyer et al, 2015; Patel et al, 2015). Like LAF-1, these proteins also contain RNA-binding domains and two regions of RGG repeats, but only the N-terminal LCD is sufficient for LLPS. Synergistic effects between these regions may contribute to in vivo LLPS by binding multiple proteins and RNAs and associating with other LCDs (Chen et al, 2011; Phan et al, 2011; Burke et al, 2015). That said, the potential for non-canonical binding of RNA by low-complexity domains should not be discounted. Indeed, the Cajal body scaffolding protein coilin, which contains RGG repeats as well as other low-complexity sequences, was recently shown to bind RNA in the absence of an annotated RNA-binding domain (Machyna et al, 2014). Thus, RNA may play a role as a generic poly-anion or may regulate LLPS through specific interactions with proteins. A second way that RNA can participate in LLPS is through protein–RNA interactions mediated by canonical RNA-binding domains, such as RNA recognition motifs (RRMs), zinc fingers (ZnFs), and KH domains. Indeed, droplet scaffolding proteins frequently contain one or more RNA-binding domain (Table 1). RNA sequence can afford multivalency to RNA–protein interactions by presenting repeated motifs for protein binding. For example, the highly structured RNA-binding protein, PTB, forms droplets when added to concatemerized RNA target sequences (Li et al, 2012). More recently, a study of Whi3, a poly-Q protein that forms cytoplasmic mRNA granules in the fungus Ashbya gossypii, showed that droplet formation depends on a functional RRM and binding to the CLN3 mRNA (Zhang et al, 2015). Full-length Whi3 never formed fibrous structures in vivo or in vitro, while the LCD alone formed filaments that became less soluble over time. Similar results were obtained in studies of FUS (see below), which was shown to transition from a droplet to filaments over time in vitro (Patel et al, 2015) and to form aggregates in the context of hydrogels (Hennig et al, 2015). The potential role of the FUS RRM in modulating LLPS, aggregate, and/or filament formation has not yet been directly analyzed. Because RNA-rich nuclear bodies—Cajal bodies, nucleoli, histone locus bodies, and paraspeckles—can form at transcription sites, nascent RNA likely plays a role in their nucleation (Bond & Fox, 2009; Mao et al, 2011; Shevtsov & Dundr, 2011; Machyna et al, 2014; Falahati et al, 2016). The combination of an RRM with an LCD should be ideal for scaffolding transcriptionally dependent LLPS structures. Indeed, many proteins within the paraspeckle contain N-terminal RRMs and C-terminal LCDs (Table 1), and both FUS and RBM14 form droplets and aggregates (Hennig et al, 2015). Recent investigations of hnRNP A1 indicate that RNA binding by N-terminal RRMs modulates droplet formation promoted by the protein's C-terminal LCD (Lin et al, 2015; Molliex et al, 2015). Interestingly, fusion of PTB's RRMs to the IDRs of a variety of RNA-binding proteins rendered these IDRs capable of droplet formation in the presence of RNA, and hnRNP A1's RRM was essential for droplet formation. Thus, nucleic acids may serve in the formation of LLPS cellular structures, by providing a multivalent platform to seed phase separation and by buffering the protein–protein contacts to promote fluidization. Different molecular interactions create distinct protein states Why would cells risk the use of systems that promote toxic aggregation? A major focus of the recent work on LLPS has been on determining which LCDs tend to form liquid droplets, hydrogels, or solid amyloid aggregates (Kato et al, 2012; Kwon et al, 2013; Altmeyer et al, 2015; Burke et al, 2015; Courchaine & Neugebauer, 2015; Hennig et al, 2015; Kroschwald et al, 2015; Lin et al, 2015; Molliex et al, 2015; Murakami et al, 2015; Patel et al, 2015). Many neurological diseases include protein aggregation in their pathology. Figure 3 conceptualizes these amyloid-like aggregates and depicts their formation. The solid stress granules of yeast are controlled by the disaggregase machinery and must be disassembled before the cell may resume growth (Cherkasov et al, 2013; Kroschwald et al, 2015). In addition, yeast stress granules can be disassembled by autophagy in response to cellular conditions (Buchan et al, 2013). In higher organisms, there are likely to be more subtle control processes that determine when LCDs remain liquid and when they aggregate. The idea that proteins have a range of states of matter has been raised previously and is an important concept in the regulation of these processes in" @default.
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• W4255830188 date "2016-06-29" @default.
• W4255830188 modified "2023-10-18" @default.
• W4255830188 title "Droplet organelles?" @default.
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• W4255830188 doi "https://doi.org/10.15252/embj.201593517" @default.
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