共查询到20条相似文献,搜索用时 15 毫秒
1.
《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2020,1867(12):118831
Viruses reshape the organization of the cell interior to achieve different steps of their cellular cycle. Particularly, viral replication and assembly often take place in viral factories where specific viral and cellular proteins as well as nucleic acids concentrate. Viral factories can be either membrane-delimited or devoid of any cellular membranes. In the latter case, they are referred as membrane-less replication compartments. The most emblematic ones are the Negri bodies, which are inclusion bodies that constitute the hallmark of rabies virus infection. Interestingly, Negri bodies and several other viral replication compartments have been shown to arise from a liquid-liquid phase separation process and, thus, constitute a new class of liquid organelles. This is a paradigm shift in the field of virus replication. Here, we review the different aspects of membrane-less virus replication compartments with a focus on the Mononegavirales order and discuss their interactions with the host cell machineries and the cytoskeleton. We particularly examine the interplay between viral factories and the cellular innate immune response, of which several components also form membrane-less condensates in infected cells. 相似文献
2.
Stefan Reber Daniel Jutzi Helen Lindsay Anny Devoy Jonas Mechtersheimer Brunno Rocha Levone Michal Domanski Eva Bentmann Dorothee Dormann Oliver Mühlemann Silvia M L Barabino Marc-David Ruepp 《Nucleic acids research》2021,49(13):7713
Liquid–liquid phase separation (LLPS) of proteins and RNAs has emerged as the driving force underlying the formation of membrane-less organelles. Such biomolecular condensates have various biological functions and have been linked to disease. The protein Fused in Sarcoma (FUS) undergoes LLPS and mutations in FUS have been causally linked to the motor neuron disease Amyotrophic Lateral Sclerosis (ALS-FUS). LLPS followed by aggregation of cytoplasmic FUS has been proposed to be a crucial disease mechanism. However, it is currently unclear how LLPS impacts the behaviour of FUS in cells, e.g. its interactome. Hence, we developed a method allowing for the purification of LLPS FUS-containing droplets from cell lysates. We observe substantial alterations in the interactome, depending on its biophysical state. While non-LLPS FUS interacts mainly with factors involved in pre-mRNA processing, LLPS FUS predominantly binds to proteins involved in chromatin remodelling and DNA damage repair. Interestingly, also mitochondrial factors are strongly enriched with LLPS FUS, providing a potential explanation for the observed changes in mitochondrial gene expression in mouse models of ALS-FUS. In summary, we present a methodology to investigate the interactomes of phase separating proteins and provide evidence that LLPS shapes the FUS interactome with implications for function and disease. 相似文献
3.
Semen V. Nesterov Nikolay S. Ilyinsky Vladimir N. Uversky 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2021,1868(11):119102
This work is devoted to the phenomenon of liquid-liquid phase separation (LLPS), which has come to be recognized as fundamental organizing principle of living cells. We distinguish separation processes with different dimensions. Well-known 3D-condensation occurs in aqueous solution and leads to membraneless organelle (MLOs) formation. 2D-films may be formed near membrane surfaces and lateral phase separation (membrane rafts) occurs within the membranes themselves. LLPS may also occur on 1D structures like DNA and the cyto- and nucleoskeleton. Phase separation provides efficient transport and sorting of proteins and metabolites, accelerates the assembly of metabolic and signaling complexes, and mediates stress responses. In this work, we propose a model in which the processes of polymerization (1D structures), phase separation in membranes (2D structures), and LLPS in the volume (3D structures) influence each other. Disordered proteins and whole condensates may provide membrane raft separation or polymerization of specific proteins. On the other hand, 1D and 2D structures with special composition or embedded IDRs can nucleate condensates. We hypothesized that environmental change may trigger a LLPS which can propagate within the cell interior moving along the cytoskeleton or as an autowave. New phase propagation quickly and using a low amount of energy adjusts cell signaling and metabolic systems to new demands. Cumulatively, the interconnected phase separation phenomena in different dimensions represent a previously unexplored system of intracellular communication and regulation which cannot be ignored when considering both physiological and pathological cell processes. 相似文献
4.
Olga Vladimirova Alessandra De Leo Zhong Deng Andreas Wiedmer James Hayden Paul M. Lieberman 《PLoS pathogens》2021,17(1)
Liquid-liquid phase separation (LLPS) can drive formation of diverse and essential macromolecular structures, including those specified by viruses. Kaposi’s Sarcoma-Associated Herpesvirus (KSHV) genomes associate with the viral encoded Latency-Associated Nuclear Antigen (LANA) to form stable nuclear bodies (NBs) during latent infection. Here, we show that LANA-NB formation and KSHV genome conformation involves LLPS. Using LLPS disrupting solvents, we show that LANA-NBs are partially disrupted, while DAXX and PML foci are highly resistant. LLPS disruption altered the LANA-dependent KSHV chromosome conformation but did not stimulate lytic reactivation. We found that LANA-NBs undergo major morphological transformation during KSHV lytic reactivation to form LANA-associated replication compartments encompassing KSHV DNA. DAXX colocalizes with the LANA-NBs during latency but is evicted from the LANA-associated lytic replication compartments. These findings indicate the LANA-NBs are dynamic super-molecular nuclear structures that partly depend on LLPS and undergo morphological transitions corresponding to the different modes of viral replication. 相似文献
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6.
《中国科学:生命科学英文版》2020,(7)
Cells are compartmentalized by numerous membrane-enclosed organelles and membraneless compartments to ensure that a wide variety of cellular activities occur in a spatially and temporally controlled manner. The molecular mechanisms underlying the dynamics of membrane-bound organelles, such as their fusion and fission, vesicle-mediated trafficking and membrane contactmediated inter-organelle interactions, have been extensively characterized. However, the molecular details of the assembly and functions of membraneless compartments remain elusive. Mounting evidence has emerged recently that a large number of membraneless compartments, collectively called biomacromolecular condensates, are assembled via liquid-liquid phase separation(LLPS). Phase-separated condensates participate in various biological activities, including higher-order chromatin organization,gene expression, triage of misfolded or unwanted proteins for autophagic degradation, assembly of signaling clusters and actin-and microtubule-based cytoskeletal networks, asymmetric segregations of cell fate determinants and formation of pre-and post-synaptic density signaling assemblies. Biomacromolecular condensates can transition into different material states such as gel-like structures and solid aggregates. The material properties of condensates are crucial for fulfilment of their distinct functions, such as biochemical reaction centers, signaling hubs and supporting architectures. Cells have evolved multiple mechanisms to ensure that biomacromolecular condensates are assembled and disassembled in a tightly controlled manner. Aberrant phase separation and transition are causatively associated with a variety of human diseases such as neurodegenerative diseases and cancers. This review summarizes recent major progress in elucidating the roles of LLPS in various biological pathways and diseases. 相似文献
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8.
Simon Alberti Shambaditya Saha Jeffrey B. Woodruff Titus M. Franzmann Jie Wang Anthony A. Hyman 《Journal of molecular biology》2018,430(23):4806-4820
The formation of membrane-less organelles and compartments by protein phase separation is an important way in which cells organize their cytoplasm and nucleoplasm. In vitro phase separation assays with purified proteins have become the standard way to investigate proteins that form membrane-less compartments. By now, various proteins have been purified and tested for their ability to phase separate and form liquid condensates in vitro. However, phase-separating proteins are often aggregation-prone and difficult to purify and handle. As a consequence, the results from phase separation assays often differ between labs and are not easily reproduced. Thus, there is an urgent need for high-quality proteins, standardized procedures, and generally agreed-upon practices for protein purification and conducting phase separation assays. This paper provides protocols for protein purification and guides the user through the practicalities of in vitro protein phase separation assays, including best-practice approaches and pitfalls to avoid. We believe that this compendium of protocols and practices will provide a useful resource for scientists studying the phase behavior of proteins. 相似文献
9.
Thuy P. Dao Carlos A. Castañeda 《BioEssays : news and reviews in molecular, cellular and developmental biology》2020,42(11):2000036
Liquid-liquid phase separation (LLPS) has recently emerged as a possible mechanism that enables ubiquitin-binding shuttle proteins to facilitate the degradation of ubiquitinated substrates via distinct protein quality control (PQC) pathways. Shuttle protein LLPS is modulated by multivalent interactions among their various domains as well as heterotypic interactions with polyubiquitin chains. Here, the properties of three different shuttle proteins (hHR23B, p62, and UBQLN2) are closely examined, unifying principles for the molecular determinants of their LLPS are identified, and how LLPS is connected to their functions is discussed. Evidence supporting LLPS of other shuttle proteins is also found. In this review, it is proposed that shuttle protein LLPS leads to spatiotemporal regulation of PQC activities by mediating the recruitment of PQC machinery (including proteasomes or autophagic components) to biomolecular condensates, assembly/disassembly of condensates, selective enrichment of client proteins, and extraction of ubiquitinated proteins from condensates in cells. 相似文献
10.
《Trends in biochemical sciences》2023,48(3):229-243
Viruses compartmentalize their replication and assembly machinery to both evade detection and concentrate the viral proteins and nucleic acids necessary for genome replication and virion production. Accumulating evidence suggests that diverse RNA and DNA viruses form replication organelles and nucleocapsid assembly sites using phase separation. In general, the biogenesis of these compartments is regulated by two types of viral protein, collectively known as antiterminators and nucleocapsid proteins, respectively. Herein, we discuss how RNA viruses establish replication organelles and nucleocapsid assembly sites, and the evidence that these compartments form through phase separation. While this review focuses on RNA viruses, accumulating evidence suggests that all viruses rely on phase separation and form biomolecular condensates important for completing the infectious cycle. 相似文献
11.
Clustering is a prominent feature of receptors at the plasma membrane (PM). It plays an important role in signaling. Liquid–liquid phase separation (LLPS) of proteins is emerging as a novel mechanism underlying the observed clustering. Receptors/transmembrane signaling proteins can be core components essential for LLPS (such as LAT or nephrin) or clients enriched at the phase-separated condensates (for example, at the postsynaptic density or at tight junctions). Condensate formation has been shown to regulate signaling in multiple ways, including by increasing protein binding avidity and by modulating the local biochemical environment. In moving forward, it is important to study protein LLPS at the PM of living cells, its interplay with other factors underlying receptor clustering, and its signaling and functional consequences. 相似文献
12.
《Biophysical journal》2022,121(1):157-171
The assembly of functional biomolecular condensates often involves liquid-liquid phase separation (LLPS) of proteins with multiple modular domains, which can be folded or conformationally disordered to various degrees. To understand the LLPS-driving domain-domain interactions, a fundamental question is how readily the interactions in the condensed phase can be inferred from interdomain interactions in dilute solutions. In particular, are the interactions leading to LLPS exclusively those underlying the formation of discrete interdomain complexes in homogeneous solutions? We address this question by developing a mean-field LLPS theory of two stoichiometrically constrained solute species. The theory is applied to the neuronal proteins SynGAP and PSD-95, whose complex coacervate serves as a rudimentary model for neuronal postsynaptic densities (PSDs). The predicted phase behaviors are compared with experiments. Previously, a three SynGAP/two PSD-95 ratio was determined for SynGAP/PSD-95 complexes in dilute solutions. However, when this 3:2 stoichiometry is uniformly imposed in our theory encompassing both dilute and condensed phases, the tie-line pattern of the predicted SynGAP/PSD-95 phase diagram differs drastically from that obtained experimentally. In contrast, theories embodying alternate scenarios postulating auxiliary SynGAP-PSD-95 as well as SynGAP-SynGAP and PSD-95-PSD-95 interactions, in addition to those responsible for stoichiometric SynGAP/PSD-95 complexes, produce tie-line patterns consistent with experiment. Hence, our combined theoretical-experimental analysis indicates that weaker interactions or higher-order complexes beyond the 3:2 stoichiometry, but not yet documented, are involved in the formation of SynGAP/PSD-95 condensates, imploring future efforts to ascertain the nature of these auxiliary interactions in PSD-like LLPS and underscoring a likely general synergy between stoichiometric, structurally specific binding and stochastic, multivalent “fuzzy” interactions in the assembly of functional biomolecular condensates. 相似文献
13.
Zhen Lei Ling Wang Eun Yu Kim Jungnam Cho 《The Plant journal : for cell and molecular biology》2021,108(5):1256-1265
Gene expression can be modulated by epigenetic mechanisms, including chromatin modifications and small regulatory RNAs. These pathways are unevenly distributed within a cell and usually take place in specific intracellular regions. Unfortunately, the fundamental driving force and biological relevance of such spatial differentiation is largely unknown. Liquid–liquid phase separation (LLPS) is a natural propensity of demixing liquid phases and has been recently suggested to mediate the formation of biomolecular condensates that are relevant to diverse cellular processes. LLPS provides a mechanistic explanation for the self-assembly of subcellular structures by which the efficiency and specificity of certain cellular reactions are achieved. In plants, LLPS has been observed for several key factors in the chromatin and small RNA pathways. For example, the formation of facultative and obligate heterochromatin involves the LLPS of multiple relevant factors. In addition, phase separation is observed in a set of proteins acting in microRNA biogenesis and the small interfering RNA pathway. In this Focused Review, we highlight and discuss the recent findings regarding phase separation in the epigenetic mechanisms of plants. 相似文献
14.
Viruses must efficiently and specifically package their genomes while excluding cellular nucleic acids and viral subgenomic fragments. Some viruses use specific packaging signals, which are conserved sequence or structure motifs present only in the full-length genome. Recent work has shown that viral proteins important for packaging can undergo liquid-liquid phase separation (LLPS), in which one or two viral nucleic acid binding proteins condense with the genome. The compositional simplicity of viral components lends itself well to theoretical modeling compared with more complex cellular organelles. Viral LLPS can be limited to one or two viral proteins and a single genome that is enriched in LLPS-promoting features. In our previous study, we observed that LLPS-promoting sequences of severe acute respiratory syndrome coronavirus 2 are located at the 5′ and 3′ ends of the genome, whereas the middle of the genome is predicted to consist mostly of solubilizing elements. Is this arrangement sufficient to drive single genome packaging, genome compaction, and genome cyclization? We addressed these questions using a coarse-grained polymer model, LASSI, to study the LLPS of nucleocapsid protein with RNA sequences that either promote LLPS or solubilization. With respect to genome cyclization, we find the most optimal arrangement restricts LLPS-promoting elements to the 5′ and 3′ ends of the genome, consistent with the native spatial patterning. Genome compaction is enhanced by clustered LLPS-promoting binding sites, whereas single genome packaging is most efficient when binding sites are distributed throughout the genome. These results suggest that many and variably positioned LLPS-promoting signals can support packaging in the absence of a singular packaging signal which argues against necessity of such a feature. We hypothesize that this model should be generalizable to multiple viruses as well as cellular organelles such as paraspeckles, which enrich specific long RNA sequences in a defined arrangement. 相似文献
15.
Liquid–liquid phase separation (LLPS) facilitates the formation of membraneless compartments in a cell and allows the spatiotemporal organization of biochemical reactions by concentrating macromolecules locally. In plants, LLPS defines cellular reaction hotspots, and stimulus‐responsive LLPS is tightly linked to a variety of cellular and biological functions triggered by exposure to various internal and external stimuli, such as stress responses, hormone signaling, and temperature sensing. Here, we provide an overview of the current understanding of physicochemical forces and molecular factors that drive LLPS in plant cells. We illustrate how the biochemical features of cellular condensates contribute to their biological functions. Additionally, we highlight major challenges for the comprehensive understanding of biological LLPS, especially in view of the dynamic and robust organization of biochemical reactions underlying plastic responses to environmental fluctuations in plants. 相似文献
16.
《Journal of molecular biology》2022,434(1):167292
Liquid-liquid phase separation (LLPS) is an important mechanism that mediates the formation of biomolecular condensates. Despite the immense interest in LLPS, phase-separated proteins verified by experiments are still limited, and identification of phase-separated proteins at proteome-scale is a challenging task. Multivalent interaction among macromolecules is the driving force of LLPS, which suggests that phase-separated proteins may harbor distinct biological characteristics in protein–protein interactions (PPIs). In this study, we constructed an integrated human PPI network (HPIN) and mapped phase-separated proteins into it. Analysis of the network parameters revealed differences of network topology between phase-separated proteins and others. The results further suggested the efficiency when applying topological similarities in distinguishing components of MLOs. Furthermore, we found that affinity purification mass spectrometry (AP-MS) detects PPIs more effectively than yeast-two hybrid system (Y2H) in phase separation-driven condensates. Our work provides the first global view of the distinct network topology of phase-separated proteins in human interactome, suggesting incorporation of PPI network for LLPS prediction in further studies. 相似文献
17.
Roshan Mammen Regy Gregory L Dignon Wenwei Zheng Young C Kim Jeetain Mittal 《Nucleic acids research》2020,48(22):12593
Ribonucleoprotein (RNP) granules are membraneless organelles (MLOs), which majorly consist of RNA and RNA-binding proteins and are formed via liquid–liquid phase separation (LLPS). Experimental studies investigating the drivers of LLPS have shown that intrinsically disordered proteins (IDPs) and nucleic acids like RNA and other polynucleotides play a key role in modulating protein phase separation. There is currently a dearth of modelling techniques which allow one to delve deeper into how polynucleotides play the role of a modulator/promoter of LLPS in cells using computational methods. Here, we present a coarse-grained polynucleotide model developed to fill this gap, which together with our recently developed HPS model for protein LLPS, allows us to capture the factors driving protein-polynucleotide phase separation. We explore the capabilities of the modelling framework with the LAF-1 RGG system which has been well studied in experiments and also with the HPS model previously. Further taking advantage of the fact that the HPS model maintains sequence specificity we explore the role of charge patterning on controlling polynucleotide incorporation into condensates. With increased charge patterning we observe formation of structured or patterned condensates which suggests the possible roles of polynucleotides in not only shifting the phase boundaries but also introducing microscopic organization in MLOs. 相似文献
18.
Madhu Ramesh Chenikkayala Balachandra Prayasee Baruah Thimmaiah Govindaraju 《Journal of peptide science》2023,29(5):e3465
Liquid–liquid phase separation (LLPS) is a complex physicochemical phenomenon mediated by multivalent transient weak interactions among macromolecules like polymers, proteins, and nucleic acids. It has implications in cellular physiology and disease conditions like cancer and neurodegenerative disorders. Many proteins associated with neurodegenerative disorders like RNA binding protein FUS (FUsed in Sarcoma), alpha-synuclein (α-Syn), TAR DNA binding protein 43 (TDP-43), and tau are shown to undergo LLPS. Recently, the tau protein responsible for Alzheimer's disease (AD) and other tauopathies is shown to phase separate into condensates in vitro and in vivo. The diverse noncovalent interactions among the biomolecules dictate the complex LLPS phenomenon. There are limited chemical tools to modulate protein LLPS which has therapeutic potential for neurodegenerative disorders. We have rationally designed cyclic dipeptide (CDP)-based small-molecule modulators (SMMs) by integrating multiple chemical groups that offer diverse chemical interactions to modulate tau LLPS. Among them, compound 1c effectively inhibits and dissolves Zn-mediated tau LLPS condensates. The SMM also inhibits tau condensate-to-fibril transition (tau aggregation through LLPS). This approach of designing SMMs of LLPS establishes a novel platform that has potential implication for the development of therapeutics for neurodegenerative disorders. 相似文献
19.
Sandeep K. Rai Adriana Savastano Priyanka Singh Samrat Mukhopadhyay Markus Zweckstetter 《Protein science : a publication of the Protein Society》2021,30(7):1294
Biomolecular condensation via liquid–liquid phase separation (LLPS) of intrinsically disordered proteins/regions (IDPs/IDRs), with and without nucleic acids, has drawn widespread interest due to the rapidly unfolding role of phase‐separated condensates in a diverse range of cellular functions and human diseases. Biomolecular condensates form via transient and multivalent intermolecular forces that sequester proteins and nucleic acids into liquid‐like membrane‐less compartments. However, aberrant phase transitions into gel‐like or solid‐like aggregates might play an important role in neurodegenerative and other diseases. Tau, a microtubule‐associated neuronal IDP, is involved in microtubule stabilization, regulates axonal outgrowth and transport in neurons. A growing body of evidence indicates that tau can accomplish some of its cellular activities via LLPS. However, liquid‐to‐solid transition resulting in the abnormal aggregation of tau is associated with neurodegenerative diseases. The physical chemistry of tau is crucial for governing its propensity for biomolecular condensation which is governed by various intermolecular and intramolecular interactions leading to simple one‐component and complex multi‐component condensates. In this review, we aim at capturing the current scientific state in unveiling the intriguing molecular mechanism of phase separation of tau. We particularly focus on the amalgamation of existing and emerging biophysical tools that offer unique spatiotemporal resolutions on a wide range of length‐ and time‐scales. We also discuss the link between quantitative biophysical measurements and novel biological insights into biomolecular condensation of tau. We believe that this account will provide a broad and multidisciplinary view of phase separation of tau and its association with physiology and disease. 相似文献
20.
The stoichiometric interaction model for mesoscopic MD simulations of liquid-liquid phase separation
《Biophysical journal》2022,121(22):4382-4393
Liquid-liquid phase separation (LLPS) has received considerable attention in recent years for explaining the formation of cellular biomolecular condensates. The fluidity and the complexity of their components make molecular simulation approaches indispensable for gaining structural insights. Domain-resolution mesoscopic model simulations have been explored for cases in which condensates are formed by multivalent proteins with tandem domains. One problem with this approach is that interdomain pairwise interactions cannot regulate the valency of the binding domains. To overcome this problem, we propose a new potential, the stoichiometric interaction (SI) potential. First, we verified that the SI potential maintained the valency of the interacting domains for the test systems. We then examined a well-studied LLPS model system containing tandem repeats of SH3 domains and proline-rich motifs. We found that the SI potential alone cannot reproduce the phase diagram of LLPS quantitatively. We had to combine the SI and a pairwise interaction; the former and the latter represent the specific and nonspecific interactions, respectively. Biomolecular condensates with the mixed SI and pairwise interaction exhibited fluidity, whereas those with the pairwise interaction alone showed no detectable diffusion. We also compared the phase diagrams of the systems containing different numbers of tandem domains with those obtained from the experiments and found quantitative agreement in all but one case. 相似文献