Summary: This review focuses on liquid-liquid phase separation (LLPS) in the formation of ribonucleoprotein (RNP) condensates, which are membrane-less organelles crucial for cellular functions like RNA processing and transport. It highlights the role of multivalent interactions between RNA-binding proteins (RBPs), including FUS and TDP-43, and RNA in forming these condensates, citing examples such as stress granules and nucleoli. The paper specifically examines how mutations in RBPs, particularly those linked to ALS and frontotemporal dementia (FTD), alter phase separation properties, leading to aberrant condensates with detrimental effects on mRNA stability and localization. It also discusses the potential of these aberrant condensates to nucleate pathogenic aggregates and explores cellular mechanisms for their resolution and potential therapeutic strategies.

Summary: This review article examines the liquid-liquid phase separation (LLPS) of RNA-binding proteins, specifically TDP-43 and FUS, which are implicated in the formation of membraneless organelles and are pathologically linked to neurodegenerative diseases such as ALS and FTD. It discusses how these proteins' domain structures drive their phase separation, both in vitro and in cellular environments, and reviews factors regulating this process. The paper highlights the connection between LLPS, protein aggregation, and disrupted cellular function, suggesting that restoring functional phase separation of TDP-43 and FUS could be therapeutically beneficial for neuronal cells, while also exploring potential mechanisms for aberrant phase transition and aggregation and current therapeutic strategies.

Summary: This review examines how the prion-like domains of FUS and TDP-43 modulate phase transitions, forming condensates with diverse material states implicated in health and disease. It discusses how sequence, structure, post-translational modifications, and RNA influence the self-assembly of these RNA-binding proteins, and explores how understanding their liquid-liquid phase separation and aggregation could lead to new therapies for neurodegenerative diseases such as ALS, FTD, and LATE.

Summary: This review synthesizes biophysical knowledge on the sequences, structures, stability, dynamics, and inter-domain interactions of FUS and TDP-43, two RNA-binding proteins associated with ALS and FTD. It explains how these proteins, possessing intrinsically disordered regions, form ribonucleoprotein granules and stress granules through liquid-liquid phase separation (LLPS). The review highlights that liquid-like droplets can transform into pathological assemblies like amyloid fibrils, which are hallmarks of ALS and FTD, and discusses how ALS-causing mutants disrupt LLPS dynamics. The ultimate goal is to understand these mechanisms for developing drugs targeting LLPS and amyloidosis to treat neurodegenerative diseases.

Summary: This study investigates how the α-helical structure of the RNA-binding protein TDP-43 influences its liquid-liquid phase separation (LLPS) and function, particularly in the context of amyotrophic lateral sclerosis (ALS) pathogenesis. Using molecular simulation and NMR spectroscopy, the researchers found that specific glycine residues (G335 and G338) inhibit helical extension and helix-helix interactions, and that mutations at these sites, including the ALS-associated G335D, can enhance TDP-43 assembly. Specifically, substituting these glycines with alanine (G335A) significantly boosted phase separation in vitro and reduced the fluidity of TDP-43 compartments in cells, while also improving TDP-43's splicing function in a minigene assay, demonstrating that the helical region is a tunable module for controlling LLPS and function.

Summary: This review paper focuses on the fused in sarcoma protein (FUS) and its involvement in liquid-liquid phase separation (LLPS). It summarizes how LLPS of FUS is regulated by various factors, including physical stimuli, biochemical modulators, and changes in protein structure, detailing the mechanisms and strategies for modulating these processes.

Summary: This review examines the role of liquid-liquid phase separation (LLPS) in the formation of dynamic membrane-less organelles (MLOs) and its connection to the pathogenesis of neurodegenerative diseases. It highlights how aberrant LLPS leads to the accumulation of pathological protein aggregates, transforming from liquid-like condensates into insoluble inclusions. The paper specifically focuses on five key disease-associated proteins—tau, TDP-43, FUS, α-Syn, and HTT—and discusses their mechanisms of aggregation, offering insights into potential therapeutic strategies and future research directions.

Summary: This study employs a chemically accurate coarse-grained model to investigate how RNA influences the transport properties and stability of phase-separated protein condensates. It examines the phase behavior of RNA-binding proteins such as FUS, hnRNPA1, and TDP-43, along with their prion-like domains and RNA recognition motifs, across varying RNA concentrations. The findings reveal that RNA can enhance phase separation at low concentrations but inhibit it at high concentrations, while also modulating condensate viscosity based on RNA chain length. The research rationalizes how RNA regulates phase separation dynamics and highlights the link between aberrant condensate properties and neuropathologies.

Summary: This review examines how RNA modulates the physiological and neuropathological phase transitions of RNA-binding proteins (RBPs), which are implicated in neurodegenerative disorders like ALS and FTD. It highlights the role of liquid-liquid phase separation (LLPS) in the formation of pathological inclusions and functional membraneless organelles (MLOs) containing RBPs such as TDP-43 and FUS. The paper will describe the mechanisms of RNA-mediated RBP phase transitions, focusing on how RNA properties like length, sequence, and structure influence both physiological and pathological LLPS.

Summary: This manuscript reviews how protein post-translational modifications (PTMs) influence liquid-liquid phase separation (LLPS) driven by low-sequence complexity domains (LCDs) in proteins associated with amyotrophic lateral sclerosis (ALS). LLPS is implicated in the formation of membraneless organelles and the aggregation of proteins like TDP-43 and FUS, which are frequently mutated in ALS and exhibit prion-like behavior. PTMs can modulate these proteinopathies by altering multivalent interactions or recruiting/excluding other molecules within condensates, thereby favoring or counteracting neurodegeneration in ALS.

Summary: This review article explores the role of liquid-liquid phase separation (LLPS) in C9orf72-linked amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). It details how the C9orf72 repeat expansion mutation leads to toxic dipeptide repeat proteins (DPRs) and repeat RNA, both of which undergo phase separation and disrupt the physiological LLPS of membraneless organelles like stress granules and the nucleolus. The paper discusses the biophysical principles of phase separation, the formation and function of these organelles, and how their aberrant LLPS contributes to cellular pathology, including TDP-43 proteinopathy, and suggests LLPS as a therapeutic target.

Summary: This study investigates the role of the RNA-binding protein FUS in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), focusing on how mutations affect its phase separation behavior and RNA interactions. The research demonstrates that wild-type FUS forms dynamic, fluid condensates through stoichiometric, length-dependent RNA binding, facilitated by multimerization. In contrast, specific FUS mutations, particularly those affecting arginine residues, lead to altered conformations, static RNA binding, and the formation of large, less fluid condensates, highlighting arginine's importance in proper RNA interaction. Glycine mutations are shown to cause a rapid loss of fluidity. Notably, the nuclear import receptor Karyopherin-β2 can reverse these mutant-induced defects, restoring wild-type FUS behavior, suggesting distinct pathogenic mechanisms for different mutation types.

Summary: This paper examines FUS (fused in sarcoma), an RNA-binding protein crucial for RNA metabolism and nuclear localization, which is identified as a causative gene for amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). It highlights that FUS undergoes liquid-liquid phase separation (LLPS), a phenomenon linked to membraneless organelle formation, and that abnormal LLPS can lead to irreversible FUS aggregation, suggesting that restoring normal LLPS could be a therapeutic strategy for ALS and FTLD, with potential interventions involving small molecules like 1,6-hexanediol or RNA molecules that modulate FUS complex formation.

Summary: This paper investigates the role of post-translational modifications on the aggregation and liquid-phase separation of the FUS protein, which is implicated in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. It highlights that FUS, an RNA-binding protein, can form liquid-phase droplets and that aberrant interactions within these structures may lead to the formation of disease-causing solid aggregates. The study suggests that post-translational modifications significantly influence FUS localization and aggregation propensity, presenting them as potential therapeutic targets for FUS-linked pathologies.

Summary: This review discusses how the biophysics of proteins with intrinsically disordered domains, capable of forming biological condensates, has advanced the understanding of neurodegenerative disorders. It uses FUS, TDP-43, and A11 proteins as examples to explain how mutations and post-translational modifications in these proteins lead to amyotrophic lateral sclerosis (ALS) and fronto-temporal lobar degeneration (FTLD).

Summary: This review examines the roles of intrinsically disordered proteins in liquid-liquid phase transitions (LLPTs) that lead to the formation of proteinaceous membrane-less organelles (PMLOs), both in normal physiological states and in pathological conditions associated with amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). It discusses key proteins implicated in these diseases, such as TDP-43 and FUS, their structural properties, intrinsic disorder status, and their involvement in the formation of normal and pathological PMLOs, aiming to elucidate the "Dr. Jekyll-Mr. Hyde" behavior of these proteins driven by LLPTs and intrinsic disorder.

Summary: This paper reviews how liquid-liquid phase separation (LLPS) is a key principle for organizing proteins and RNAs into biomolecular condensates, which are crucial for cellular responses, particularly in neurons and glia. It highlights that in neurodegenerative diseases like Amyotrophic Lateral Sclerosis (ALS), the misregulation of these condensates leads to insoluble aggregates, a hallmark of ALS. The review connects the emerging understanding of LLPS involving ALS-related proteins with their aggregation mechanisms, suggesting that understanding these processes could lead to new drug development strategies.

Summary: This review discusses liquid-liquid phase separation (LLPS) as a fundamental mechanism for intracellular organization, regulating the assembly and composition of numerous membraneless organelles and biomolecular condensates. It highlights how aberrant LLPS, driven by altered physiological conditions or genetic mutations, can lead to disease onset and progression, particularly in neurodegenerative disorders and cancers. The paper aims to summarize the properties of these organelles, discuss phase separation-regulated biological processes, and provide examples of how dysregulated LLPS and mutations in key proteins contribute to human diseases.

Summary: This review focuses on RNA-binding proteins (RBPs) and their central role in triggering liquid-liquid phase separation (LLPS), which underlies the formation of membraneless organelles (MLOs). It explains that RBPs-driven LLPS is primarily mediated by interactions between RNA recognition motifs (RRMs) and mRNA, as well as heterotypic multivalent interactions involving intrinsically disordered regions (IDRs) or prion-like domains (PLDs). The review covers the involvement of RBPs-triggered LLPS in various intracellular processes, its significance in cellular physiology and pathology, and its potential as a therapeutic target for human diseases.

Summary: This review discusses amyotrophic lateral sclerosis (ALS), a neurodegenerative disease, and highlights the role of dysfunctional liquid-liquid phase separation (LLPS) as a mechanism linking ALS-related proteins to pathogenesis. It notes that proteins like TDP-43 and FUS, which possess low-complexity domains (LCDs) and are intrinsically disordered, form liquid droplets in vitro through weak interactions. The paper reviews current knowledge on ALS-related gene products associated with proteinopathy, their status as LLPS proteins, and explores the therapeutic potential of targeting LLPS for ALS treatment.

Summary: This review discusses the critical role of conformational dynamics of intrinsically disordered proteins (IDPs) within biomolecular condensates, which are formed via liquid-liquid phase separation (LLPS). It highlights how LLPS modulates the internal motions of IDPs across various scales and emphasizes the importance of intermolecular interactions in driving LLPS, influencing biomolecular motions, and contributing to condensate aging in human diseases, proposing a framework for future research.

Summary: This review explores the significance of biological phase separation in cellular organization, focusing on liquid-liquid phase separation (LLPS) as the mechanism forming membraneless organelles. It aims to elucidate the molecular determinants driving LLPS, particularly the roles of disorder, sequence, posttranslational modifications, and regulatory stimuli in protein LLPS, with an emphasis on insights from simulation and theory. The review also discusses how these molecular forces influence multicomponent phase separation and selectivity.

Summary: This review consolidates research findings on the pathological involvement of protein phase separation and aggregation in neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's disease. It highlights how cellular phase separation, leading to biomolecular condensates, is crucial for biological processes like neuronal development and synaptic signaling, and how its aberrant forms can cause protein aggregation, a common phenomenon in affected neuronal cells. The paper also discusses therapeutic strategies targeting protein aggregation.

Summary: This abstract describes liquid-liquid phase separation (LLPS) as a biological process where proteins form membraneless condensates, influenced by factors like protein concentration, post-translational modifications, solution conditions, and assisting molecules such as RNAs. LLPS enables cells to assemble functional units, modulate cellular functions, and respond rapidly to cues, thereby regulating biology and physiology. The abstract also notes that excessive LLPS can lead to the formation of rigid aggregates, disrupting organelle function and contributing to human disorders, particularly neurodegenerative diseases, suggesting that targeting these abnormal aggregates is a promising therapeutic strategy.

Summary: This review discusses membraneless organelles (MLOs) and their formation via liquid-liquid phase separation (LLPS), highlighting their broad influence on human health and disease, including neurodegenerative disorders. It covers the underlying mechanisms and biophysical properties driving LLPS, the role of the physicochemical environment, molecular interactions, and post-translational modifications in regulating MLO dynamics, and explores the potential of targeting LLPS therapeutically.

Summary: This essay provides a brief and subjective outline of recent developments in the interlinked fields of intrinsically disordered proteins (IDPs), liquid-liquid phase separation (LLPS), and membrane-less organelles (MLOs). It highlights that many proteins crucial for biological functions lack unique structures and that biological processes are compartmentalized into liquid-like biomolecular condensates formed via LLPS, which are not membrane-bound. The abstract emphasizes that intrinsic disorder is a key requirement for proteins to undergo LLPS, driving the biogenesis of numerous MLOs, and positions these phenomena as a bridge between molecular/cellular biology and soft matter physics.

Summary: This study uses coarse-grained Langevin dynamics simulations to investigate the physical basis for structural diversity in condensed phases of multi-domain RNA-binding proteins (RBPs). It reveals a cooperative first-order transition between disordered and ordered phases, where prion-like domains (PLDs) form amyloid-like fibrils with high nematic order. The interplay between PLD-PLD and PLD-RBD interactions dictates various structures and their dynamics, with ordered phases exhibiting significantly lower protein diffusion. This transition's cooperativity suggests malleability to mutations or post-translational modifications, offering a mechanistic understanding of how multi-domain RBPs form assemblies with distinct material properties.

Summary: This review discusses how low-complexity (LC) domains, previously thought to be unstructured, are now understood to drive cellular organization through phase separation. It highlights evidence showing that the phase separation of LC domains organizes cellular assemblies and facilitates biological functions, while also implicating altered phase separation dynamics in human diseases, including neurodegenerative conditions, and suggests potential therapeutic avenues.

Summary: This review focuses on the concepts of protein intrinsic disorder and proteinaceous membrane-less organelles (PMLOs), noting a recent surge in scientific interest. It posits that merging these ideas through intrinsic disorder-based liquid-liquid phase separation offers a foundation for understanding the molecular mechanisms behind PMLO biogenesis.

Summary: This review discusses liquid-liquid phase separation (LLPS) of intracellular proteins, highlighting its role in forming membraneless condensates that regulate gene transcription and cellular responses to environmental changes. It emphasizes that LLPS is modulated by cytosol conditions and, importantly, by post-translational modifications (PTMs) which affect protein multivalency, solubility, and charge interactions. The paper notes that aberrant protein aggregation linked to LLPS is implicated in human diseases, including neurodegenerative conditions, and aims to summarize LLPS functions, its relationship with PTMs, and their implications in human diseases.

Summary: This review discusses liquid-liquid phase separation (LLPS) as a process forming distinct cellular phases, creating condensates found in organelles like nucleoli and stress granules, and playing critical roles in cellular functions. It covers the concept, thermodynamic and biochemical principles, functions (e.g., adjusting reaction rates, regulating protein folding, structural support), and its link to diseases such as cancer and neurodegeneration. The paper also reviews advanced detection methods for LLPS and discusses future directions for precise detection and potential applications.

Summary: This paper explores the biophysical mechanisms of liquid-liquid phase separation (LLPS) in biological systems, focusing on its role in forming membrane-less organelles and modulating cellular activities. It delves into the underlying biophysical principles, such as multivalent interactions and entropy-driven processes, and examines how LLPS affects cellular organization and physiological roles, including stress response and gene expression regulation. The study also investigates dysregulated LLPS in human disorders, highlighting potential therapeutic targets and approaches.

Summary: This paper reviews recent studies on liquid-liquid phase separation (LLPS) in biology, focusing on the assembly of non-membrane bound organelles. It highlights that LLPS is driven by multivalent protein-protein and/or protein-nucleic acid interactions, which can be either stoichiometric or mediated by intrinsically disordered regions (IDRs). While IDR-driven condensate formation is currently dominant, the authors emphasize the importance of specific biomolecular interactions for the function of various physiological condensates and propose that a combination of specific and promiscuous IDR-driven interactions is a general feature of biological condensation.

Summary: This review explores the molecular mechanisms behind protein aggregation in Amyotrophic Lateral Sclerosis (ALS) and Frontotemporal Dementia (FTD), highlighting TDP-43 as a primary aggregated protein. It notes that genetic mutations in ALS-FTD disrupt protein stability, phase separation, and interaction networks, leading to misfolding and aggregation. The paper also discusses cellular protective responses, such as molecular chaperones and post-translational modifications, aimed at preventing protein aggregation and mitigating cellular toxicity.

Summary: This review discusses the significant role of intracellular liquid-liquid phase separation (LLPS) in forming membraneless organelles, which are droplets of proteins and RNA. It highlights recent advancements in understanding the molecular interactions that govern the properties and behavior of these condensates, and explores how their metastability might be implicated in protein aggregation diseases.

Summary: This review paper discusses the role of liquid-liquid phase separation (LLPS) in the formation and plasticity of membraneless organelles (MLOs), which are crucial for cellular complexity and function. It highlights that MLOs are constructed through multivalent biomolecular interactions via phase separation, and their dynamic regulation is essential for cell homeostasis. The paper notes that aberrant phase separation is implicated in various diseases, including neurodegenerative disorders and cancer, and aims to cover the mechanistic understanding of phase separation, unifying structural and mechanistic principles, cellular regulatory mechanisms, and potential therapeutic applications.

Summary: This review discusses how phase separation enables the formation of subcompartmentalized structures, such as the nucleolus, which is presented as an example of a highly organized membraneless organelle. It highlights how chemical modifications of proteins, RNA, and lipids influence the molecular environment to facilitate enzymatic reactions within specific cellular regions.

Summary: This review summarizes recent biophysical studies, particularly those using single-molecule fluorescence detection, that investigate the intramolecular conformational changes of intrinsically disordered proteins (IDPs) during liquid-liquid phase separation (LLPS) and their intermolecular clustering within biomolecular condensates. It highlights that while macroscopic properties of condensates have been well-characterized, the molecular-level conformations and interactions within them are less understood. The paper aims to link these microscopic features to the macroscopic phase transitions relevant to the physiological and pathological roles of condensates, noting that IDPs are key components involved in LLPS and that condensates are implicated in human diseases.

Summary: This narrative review comprehensively elucidates the formation of membraneless organelles (MLOs) driven by liquid-liquid phase separation (LLPS) in eukaryotic cells, highlighting their pivotal roles in biological processes such as gene transcription, RNA metabolism, and signal transduction. It discusses the relationship between MLO functions and associated diseases, including neurodegenerative disorders, and examines the underlying mechanisms of phase separation influenced by protein concentration and valency, suggesting potential for novel therapeutic strategies.

Summary: This review discusses biological liquid-liquid phase separation (LLPS) as a driving force for the assembly of membraneless organelles (MLOs), highlighting recent work on rapid, reversible protein condensation in response to osmotic changes, driven by molecular crowding. It reviews methods for visualizing and modulating intracellular condensates and proposes a cloud formation metaphor for understanding phase separation. The paper traces the historical terminology for MLOs, from early observations to the modern concept of biomolecular condensates, emphasizing their dynamic fluid properties and roles in cellular organization and response to fluctuations. It also delves into the molecular features driving LLPS, such as multivalency and specific interactions, and discusses how post-translational modifications and concentration changes, including those from osmotic perturbations, modulate phase behavior, with a brief mention of LLPS in pathological contexts like neurodegeneration involving proteins like TDP-43/FUS.

Summary: This paper reviews liquid-liquid phase transitions (LLPTs), also known as liquid-liquid phase separation (LLPS), which are responsible for the formation and disintegration of proteinaceous membrane-less organelles (PMLOs) in cells. It discusses the emerging understanding of the physiological functions of these transitions and highlights their recent prominence due to associations with various pathological conditions, including cancers, neurodegenerative diseases, and cardiovascular diseases, stemming from dysregulated biogenesis or loss of dynamics in PMLOs.

Summary: This review discusses membraneless organelles, which are formed through liquid-liquid phase separation driven by weak multivalent interactions. It focuses on transcription regulators that undergo LLPS, highlighting the importance of intrinsically disordered regions in this process, and reviews experimental methods for studying membraneless organelles.

Summary: This article discusses intracellular liquid droplets, also known as proteinaceous membrane-less organelles (PMLOs), found across various cellular compartments in eukaryotes and bacteria. It highlights their role in intracellular compartmentalization, information processing, and rapid environmental response. The abstract emphasizes that these PMLOs exhibit liquid-like properties and are driven by liquid-liquid phase transitions (LLPTs) orchestrated by intrinsically disordered proteins (IDPs) and hybrid proteins with intrinsically disordered protein regions (IDPRs), which facilitate multivalent interactions and contribute to the 'supramolecular fuzziness' essential for PMLO biogenesis.

Summary: This paper addresses the growing interest in liquid-liquid phase separation (LLPS) and biomolecular condensates, which underlie membraneless compartments in cells and are implicated in various biological processes and diseases. It proposes guidelines for rigorous experimental characterization of LLPS in vitro and in cells, discusses common experimental caveats, and identifies gaps in current research methods and theory.

Summary: This paper provides a historical overview of the shift in understanding intracellular space organization, moving from a mechanistic model to one emphasizing dynamic, multifunctional 'soft matter' driven by liquid-liquid phase transitions of biopolymers. It highlights how these self-organizing, membrane-less cellular compartments are dynamic systems crucial for spatio-temporal organization and regulation of intracellular processes, noting that disruptions can lead to aggregation and amyloid formation. The abstract emphasizes the dual requirement for membrane-less organelles to maintain resistance to environmental changes while remaining sensitive to external signals for proper cellular function.

Summary: This review discusses how condensate formation by phase separation is a new principle for understanding cellular organization and its association with various human diseases, including cancer, neurodegeneration, and infectious diseases, highlighting aberrant condensates as drivers of disease mechanisms and potential therapeutic targets.

Summary: This paper reviews the role of macromolecular crowding in liquid-liquid phase separation (LLPS) of biomolecular condensates, which are non-membranous organelles with liquid-like properties. It explores how crowding affects protein interactions, folding, aggregation, and LLPS by influencing phase boundaries and condensate properties, drawing comparisons between in vivo and in vitro studies and suggesting similarities to segregative phase separation.

Summary: This review explores the theoretical physical basis of liquid–liquid phase separation (LLPS) driven by intrinsic disorder in biomolecules and synthetic systems. It covers experimental and computational methods used to study phase-separating proteins and their mechanisms in physiology and disease, and discusses the development of engineered phase-separating polypeptides for controlling self-assembly and reprogramming biological processes.

Summary: This article explores how RNA multimerization, driven by strong and redundant RNA-RNA interactions, acts as a key force in liquid-liquid phase separation (LLPS). It examines the thermodynamic, kinetic, and structural properties of RNA that govern its ability to form monomers, dimers, and higher-order structures, noting these principles are broadly applicable. The paper also speculates on how external conditions, particularly in plants, can influence condensate formation through effects on RNA base-pairing, potentially impacting biological processes.