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FUS and TDP-43, two RNA-binding proteins from the heterogeneous nuclear ribonucleoprotein family, have gained significant attention in the field of neurodegenerative diseases due to their association with amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD). They possess folded domains for binding ATP and various nucleic acids including DNA and RNA, as well as substantial intrinsically disordered regions (IDRs) including prion-like domains (PLDs) and RG-/RGG-rich regions. They play vital roles in various cellular processes, including transcription, splicing, microRNA maturation, RNA stability and transport and DNA repair. In particular, they are key components for forming ribonucleoprotein granules and stress granules (SGs) through homotypic or heterotypic liquid-liquid phase separation (LLPS). Strikingly, liquid-like droplets formed by FUS and TDP-43 may undergo aging to transform into less dynamic assemblies such as hydrogels, inclusions, and amyloid fibrils, which are the pathological hallmarks of ALS and FTD. This review aims to synthesize and consolidate the biophysical knowledge of the sequences, structures, stability, dynamics, and interdomain interactions of FUS and TDP-43 domains, so as to shed light on the molecular mechanisms underlying their liquid-liquid phase separation (LLPS) and amyloidosis. The review further delves into the mechanisms through which ALS-causing mutants of the well-folded hPFN1 disrupt the dynamics of LLPS of FUS prion-like domain, providing key insights into a potential mechanism for misfolding/aggregation-prone proteins to cause neurodegenerative diseases and aging by gain of functions. With better understanding of different biophysical aspects of FUS and TDP-43, the ultimate goal is to develop drugs targeting LLPS and amyloidosis, which could mediate protein homeostasis within cells and lead to new treatments for currently intractable diseases, particularly neurodegenerative diseases such as ALS, FTD and aging. However, the study of membrane-less organelles and condensates is still in its infancy and therefore the review also highlights key questions that require future investigation. [ABSTRACT FROM AUTHOR]

Some negative-sense RNA viruses, including measles virus (MeV), share the characteristic that during their infection cycle, cytoplasmic inclusion bodies (IBs) are formed where components of the viral replication machinery are concentrated. As a foci of viral replication, how IBs act to enhance the efficiency of infection by affecting virus-host interactions remains an important topic of investigation. We previously established that upon MeV infection, the epigenetic host protein, WD repeat-containing protein 5 (WDR5), translocates to cytoplasmic viral IBs and facilitates MeV replication. We now show that WDR5 is recruited to IBs by forming a complex with IB-associated MeV phosphoprotein via a conserved binding motif located on the surface of WDR5. Furthermore, we provide evidence that WDR5 promotes viral replication by suppressing a major innate immune response pathway, the double-stranded RNA-mediated activation of protein kinase R and integrated stress response. IMPORTANCE MeV is a pathogen that remains a global concern, with an estimated 9 million measles cases and 128,000 measles deaths in 2022 according to the World Health Organization. A large population of the world still has inadequate access to the effective vaccine against the exceptionally transmissible MeV. Measles disease is characterized by a high morbidity in children and in immunocompromised individuals. An important area of research for negative-sense RNA viruses, including MeV, is the characterization of the complex interactome between virus and host occurring at cytoplasmic IBs where viral replication occurs. Despite the progress made in understanding IB structures, little is known regarding the virus-host interactions within IBs and the role of these interactions in promoting viral replication and antagonizing host innate immunity. Herein we provide evidence suggesting a model by which MeV IBs utilize the host protein WDR5 to suppress the protein kinase R-integrated stress response pathway. [ABSTRACT FROM AUTHOR]

Prion-like proteins (PrLPs) have emerged as beneficial molecules with implications in adaptive responses. These proteins possess a conserved prion-like domain (PrLD) which is an intrinsically disordered region capable of adopting different conformations upon perceiving external stimuli. Owing to changes in protein conformation, functional characteristics of proteins harboring PrLDs get altered thereby, providing a unique mode of protein-based regulation. Since PrLPs are ubiquitous in nature and involved in diverse functions, through this study, we aim to explore the role of such domains in yet another important physiological process viz. plant-microbe interactions to get insights into the mechanisms dictating cross-kingdom interactions. We have evaluated the presence and functions of PrLPs in 18 different plant-associated fungi of agricultural importance to unravel their role in plant-microbe interactions. Of the 241,997 proteins scanned, 3,820 (~ 1.6%) were identified as putative PrLPs with pathogenic fungi showing significantly higher PrLP density than their beneficial counterparts. Further, through GO enrichment analysis, we could predict several PrLPs from pathogenic fungi to be involved in virulence and formation of stress granules. Notably, PrLPs involved in (retro)transposition were observed exclusively in pathogenic fungi. We even analyzed publicly available data for the expression alterations of fungal PrLPs upon their interaction with their respective hosts which revealed perturbation in the levels of some PrLP-encoding genes during interactions with plants. Overall, our work sheds light into the probable role of prion-like candidates in plant-fungi interaction, particularly in context of pathogenesis, paving way for more focused studies for validating their role. [ABSTRACT FROM AUTHOR]

Featured Application: Study explores the application of LLMs, specifically ChatGPT4 and Gemini for biophysical research, with a focus on liquid–liquid phase separation (LLPS). Our findings suggest that while both models show promise in facilitating detailed scientific discussions, further refinements are necessary to improve their accuracy and reliability. Recent advancements in artificial intelligence (AI), notably through generative pretrained transformers, such as ChatGPT and Google's Gemini, have broadened the scope of research across various domains. Particularly, the role of AI in understanding complex biophysical phenomena like liquid–liquid phase separation (LLPS) is promising yet underexplored. In this study, we focus on assessing the application of these AI chatbots in understating LLPS by conducting various interactive sessions. We evaluated their performance based on the accuracy, response time, response length, and cosine similarity index (CSI) of their responses. Our findings show that Gemini consistently delivered more accurate responses to LLPS-related questions than ChatGPT. However, neither model delivered correct answers to all questions posed. Detailed analysis showed that Gemini required longer response times, averaging 272 words per response compared to ChatGPT's 351. Additionally, the average CSI between the models was 0.62, highlighting moderate similarity. Despite both models showing potential to enhance scientific education in complex domains, our findings highlight a critical need for further refinement of these AI tools to improve their accuracy and reliability in specialized academic settings. [ABSTRACT FROM AUTHOR]

Ribosome biogenesis is initiated in the nucleolus, a multiphase biomolecular condensate formed by liquid-liquid phase separation. The nucleolus is a powerful disease biomarker and stress biosensor whose morphology reflects function. Here we have used digital holographic microscopy (DHM), a label-free quantitative phase contrast microscopy technique, to detect nucleoli in adherent and suspension human cells. We trained convolutional neural networks to detect and quantify nucleoli automatically on DHM images. Holograms containing cell optical thickness information allowed us to define a novel index which we used to distinguish nucleoli whose material state had been modulated optogenetically by blue-light-induced protein aggregation. Nucleoli whose function had been impacted by drug treatment or depletion of ribosomal proteins could also be distinguished. We explored the potential of the technology to detect other natural and pathological condensates, such as those formed upon overexpression of a mutant form of huntingtin, ataxin-3, or TDP-43, and also other cell assemblies (lipid droplets). We conclude that DHM is a powerful tool for quantitatively characterizing nucleoli and other cell assemblies, including their material state, without any staining. Synopsis: Digital Holographic Microscopy (DHM) detects membraneless biomolecular condensates and membrane-bound organelles without staining. Correlative Fluorescence - Digital Holographic Microscopy (DHM), a label-free quantitative interferometric technique, detects nucleoli, a biomolecular condensate, and membrane-bound organelles without staining. Convolutional neural networks automatically segment holograms and quantify the nucleolus. Definition of the nucleolar optical thickness as a novel index for characterizing the material state of the nucleolus. Digital Holographic Microscopy (DHM) detects membraneless biomolecular condensates and membrane-bound organelles without staining. [ABSTRACT FROM AUTHOR]

Liquid–liquid phase separation (LLPS) is associated with the spatiotemporal organization of the autophagic machinery, cell migration signaling, and stress granule dynamics. The proven functional spectrum of Unc-51-like kinase (ULK)/autophagy-related 1 (Atg1) in LLPS events includes autophagy, stress granule disassembly, and focal adhesion assembly. Phase separation underlies aspects of other cellular process that are regulated by ULK/Atg1 (e.g., cell proliferation and cell death pathways, endoplasmic reticulum-to-Golgi trafficking), but whether ULK/Atg1 activity influences LLPS in these contexts remains to be determined. Autophagy – a highly regulated intracellular degradation process – is pivotal in maintaining cellular homeostasis. Liquid–liquid phase separation (LLPS) is a fundamental mechanism regulating the formation and function of membrane-less compartments. Recent research has unveiled connections between LLPS and autophagy, suggesting that phase separation events may orchestrate the spatiotemporal organization of autophagic machinery and cargo sequestration. The Unc-51-like kinase (ULK)/autophagy-related 1 (Atg1) family of proteins is best known for its regulatory role in initiating autophagy, but there is growing evidence that the functional spectrum of ULK/Atg1 extends beyond autophagy regulation. In this review, we explore the spatial and temporal regulation of the ULK/Atg1 family of kinases, focusing on their recruitment to LLPS-driven compartments, and highlighting their multifaceted functions beyond their traditional role. [ABSTRACT FROM AUTHOR]

Adenosine triphosphate (ATP) acts as the universal energy currency that drives various biological processes, while nucleic acids function to store and transmit genetic information for all living organisms. Liquid–liquid phase separation (LLPS) represents the common principle for the formation of membrane-less organelles (MLOs) composed of proteins rich in intrinsically disordered regions (IDRs) and nucleic acids. Currently, while IDRs are well recognized to facilitate LLPS through dynamic and multivalent interactions, the precise mechanisms by which ATP and nucleic acids affect LLPS still remain elusive. This review summarizes recent NMR results on the LLPS of human FUS, TDP-43, and the viral nucleocapsid (N) protein of SARS-CoV-2, as modulated by ATP and nucleic acids, revealing the following: (1) ATP binds to folded domains overlapping with nucleic-acid-binding interfaces; (2) ATP and nucleic acids interplay to biphasically modulate LLPS by competitively binding to overlapping pockets of folded domains and Arg/Lys within IDRs; (3) ATP energy-independently induces protein folding with the highest efficiency known so far. As ATP likely emerged in the prebiotic monomeric world, while LLPS represents a pivotal mechanism to concentrate and compartmentalize rare molecules for forming primordial cells, ATP appears to control protein homeostasis and shape genome–proteome interfaces throughout the evolutionary trajectory, from prebiotic origins to modern cells. [ABSTRACT FROM AUTHOR]

Inclusion bodies (IBs) of respiratory syncytial virus (RSV) are formed by liquid-liquid phase separation (LLPS) and contain internal structures termed "IB-associated granules" (IBAGs), where anti-termination factor M2-1 and viral mRNAs are concentrated. However, the mechanism of IBAG formation and the physiological function of IBAGs are unclear. Here, we found that the internal structures of RSV IBs are actual M2-1-free viral messenger ribonucleoprotein (mRNP) condensates formed by secondary LLPS. Mechanistically, the RSV nucleoprotein (N) and M2-1 interact with and recruit PABP to IBs, promoting PABP to bind viral mRNAs transcribed in IBs by RNA-recognition motif and drive secondary phase separation. Furthermore, PABP-eIF4G1 interaction regulates viral mRNP condensate composition, thereby recruiting specific translation initiation factors (eIF4G1, eIF4E, eIF4A, eIF4B and eIF4H) into the secondary condensed phase to activate viral mRNAs for ribosomal recruitment. Our study proposes a novel LLPS-regulated translation mechanism during viral infection and a novel antiviral strategy via targeting on secondary condensed phase. [ABSTRACT FROM AUTHOR]

Stress conditions can cause the relocalization of proteasomes to condensates in yeast and mammalian cells. The interactions that facilitate the formation of proteasome condensates, however, are unclear. Here, we show that the formation of proteasome condensates in yeast depends on ubiquitin chains together with the proteasome shuttle factors Rad23 and Dsk2. These shuttle factors colocalize to these condensates. Strains deleted for the third shuttle factor gene, DDI1, show proteasome condensates in the absence of cellular stress, consistent with the accumulation of substrates with long K48-linked ubiquitin chains that accumulate in this mutant. We propose a model where the long K48-linked ubiquitin chains function as a scaffold for the ubiquitin-binding domains of the shuttle factors and the proteasome, allowing for the multivalent interactions that further drive condensate formation. Indeed, we determined different intrinsic ubiquitin receptors of the proteasome--Rpn1, Rpn10, and Rpn13--and the Ubl domains of Rad23 and Dsk2 are critical under different condensate-inducing conditions. In all, our data support a model where the cellular accumulation of substrates with long ubiquitin chains, potentially due to reduced cellular energy, allows for proteasome condensate formation. This suggests that proteasome condensates are not simply for proteasome storage, but function to sequester soluble ubiquitinated substrates together with inactive proteasomes. [ABSTRACT FROM AUTHOR]

Abstract Liquid–liquid phase separation (LLPS) in biology describes a process by which proteins form membraneless condensates within a cellular compartment when conditions are met, including the concentration and posttranslational modifications of the protein components, the condition of the aqueous solution (pH, ionic strength, pressure, and temperature), and the existence of assisting factors (such as RNAs or other proteins). In these supramolecular liquid droplet-like inclusion bodies, molecules are held together through weak intermolecular and/or intramolecular interactions. With the aid of LLPS, cells can assemble functional sub-units within a given cellular compartment by enriching or excluding specific factors, modulating cellular function, and rapidly responding to environmental or physiological cues. Hence, LLPS is emerging as an important means to regulate biology and physiology. Yet, excessive inclusion body formation by, for instance, higher-than-normal concentrations or mutant forms of the protein components could result in the conversion from dynamic liquid condensates into more rigid gel- or solid-like aggregates, leading to the disruption of the organelle’s function followed by the development of human disorders like neurodegenerative diseases. In summary, well-controlled formation and de-formation of LLPS is critical for normal biology and physiology from single cells to individual organisms, whereas abnormal LLPS is involved in the pathophysiology of human diseases. In turn, targeting these aggregates or their formation represents a promising approach in treating diseases driven by abnormal LLPS including those neurodegenerative diseases that lack effective therapies.

Liquid–liquid phase separation (LLPS) in biology describes a process by which proteins form membraneless condensates within a cellular compartment when conditions are met, including the concentration and posttranslational modifications of the protein components, the condition of the aqueous solution (pH, ionic strength, pressure, and temperature), and the existence of assisting factors (such as RNAs or other proteins). In these supramolecular liquid droplet-like inclusion bodies, molecules are held together through weak intermolecular and/or intramolecular interactions. With the aid of LLPS, cells can assemble functional sub-units within a given cellular compartment by enriching or excluding specific factors, modulating cellular function, and rapidly responding to environmental or physiological cues. Hence, LLPS is emerging as an important means to regulate biology and physiology. Yet, excessive inclusion body formation by, for instance, higher-than-normal concentrations or mutant forms of the protein components could result in the conversion from dynamic liquid condensates into more rigid gel- or solid-like aggregates, leading to the disruption of the organelle's function followed by the development of human disorders like neurodegenerative diseases. In summary, well-controlled formation and de-formation of LLPS is critical for normal biology and physiology from single cells to individual organisms, whereas abnormal LLPS is involved in the pathophysiology of human diseases. In turn, targeting these aggregates or their formation represents a promising approach in treating diseases driven by abnormal LLPS including those neurodegenerative diseases that lack effective therapies. [ABSTRACT FROM AUTHOR]

Global climate change has caused severe abiotic and biotic stresses, affecting plant growth and food security. The mechanical understanding of plant stress responses is critical for achieving sustainable agriculture. Intrinsically disordered proteins (IDPs) are a group of proteins without unique three-dimensional structures. The environmental sensitivity and structural flexibility of IDPs contribute to the growth and developmental plasticity for sessile plants to deal with environmental challenges. This article discusses the roles of various disordered proteins in plant stress tolerance and resistance, describes the current mechanistic insights into unstructured proteins such as the disorder-to-order transition for adopting secondary structures to interact with specific partners (i.e., cellular membranes, membrane proteins, metal ions, and DNA), and elucidates the roles of liquid–liquid phase separation driven by protein disorder in stress responses. By comparing IDP studies in animal systems, this article provides conceptual principles of plant protein disorder in stress adaptation, reveals the current research gaps, and advises on the future research direction. The highlighting of relevant unanswered questions in plant protein disorder research aims to encourage more studies on these emerging topics to understand the mechanisms of action behind their stress resistance phenotypes. [ABSTRACT FROM AUTHOR]

In recent years, the role of liquid–liquid phase separation (LLPS) and intrinsically disordered proteins (IDPs) in cellular molecular processes has received increasing attention from researchers. One such intrinsically disordered protein is TSPYL5, considered both as a marker and a potential therapeutic target for various oncological diseases. However, the role of TSPYL5 in intracellular processes remains unknown, and there is no clarity even in its intracellular localization. In this study, we characterized the intracellular localization and exchange dynamics with intracellular contents of TSPYL5 and its parts, utilizing TSPYL5 fusion proteins with EGFP. Our findings reveal that TSPYL5 can be localized in both the cytoplasm and nucleoplasm, including the nucleolus. The nuclear (nucleolar) localization of TSPYL5 is mediated by the nuclear/nucleolar localization sequences (NLS/NoLS) identified in the N-terminal intrinsically disordered region (4–27 aa), while its cytoplasmic localization is regulated by the ordered NAP-like domain (198–382 aa). Furthermore, our results underscore the significant role of the TSPYL5 N-terminal disordered region (1–198 aa) in the exchange dynamics with the nucleoplasm and its potential ability for phase separation. Bioinformatics analysis of the TSPYL5 interactome indicates its potential function as a histone and ribosomal protein chaperone. Taken together, these findings suggest a significant contribution of liquid–liquid phase separation to the processes involving TSPYL5, providing new insights into the role of this protein in the cell's molecular life. [ABSTRACT FROM AUTHOR]

Recent advancements in artificial intelligence (AI), notably through generative pretrained transformers, such as ChatGPT and Google’s Gemini, have broadened the scope of research across various domains. Particularly, the role of AI in understanding complex biophysical phenomena like liquid–liquid phase separation (LLPS) is promising yet underexplored. In this study, we focus on assessing the application of these AI chatbots in understating LLPS by conducting various interactive sessions. We evaluated their performance based on the accuracy, response time, response length, and cosine similarity index (CSI) of their responses. Our findings show that Gemini consistently delivered more accurate responses to LLPS-related questions than ChatGPT. However, neither model delivered correct answers to all questions posed. Detailed analysis showed that Gemini required longer response times, averaging 272 words per response compared to ChatGPT’s 351. Additionally, the average CSI between the models was 0.62, highlighting moderate similarity. Despite both models showing potential to enhance scientific education in complex domains, our findings highlight a critical need for further refinement of these AI tools to improve their accuracy and reliability in specialized academic settings.

Adenosine triphosphate (ATP) acts as the universal energy currency that drives various biological processes, while nucleic acids function to store and transmit genetic information for all living organisms. Liquid–liquid phase separation (LLPS) represents the common principle for the formation of membrane-less organelles (MLOs) composed of proteins rich in intrinsically disordered regions (IDRs) and nucleic acids. Currently, while IDRs are well recognized to facilitate LLPS through dynamic and multivalent interactions, the precise mechanisms by which ATP and nucleic acids affect LLPS still remain elusive. This review summarizes recent NMR results on the LLPS of human FUS, TDP-43, and the viral nucleocapsid (N) protein of SARS-CoV-2, as modulated by ATP and nucleic acids, revealing the following: (1) ATP binds to folded domains overlapping with nucleic-acid-binding interfaces; (2) ATP and nucleic acids interplay to biphasically modulate LLPS by competitively binding to overlapping pockets of folded domains and Arg/Lys within IDRs; (3) ATP energy-independently induces protein folding with the highest efficiency known so far. As ATP likely emerged in the prebiotic monomeric world, while LLPS represents a pivotal mechanism to concentrate and compartmentalize rare molecules for forming primordial cells, ATP appears to control protein homeostasis and shape genome–proteome interfaces throughout the evolutionary trajectory, from prebiotic origins to modern cells.

Global climate change has caused severe abiotic and biotic stresses, affecting plant growth and food security. The mechanical understanding of plant stress responses is critical for achieving sustainable agriculture. Intrinsically disordered proteins (IDPs) are a group of proteins without unique three-dimensional structures. The environmental sensitivity and structural flexibility of IDPs contribute to the growth and developmental plasticity for sessile plants to deal with environmental challenges. This article discusses the roles of various disordered proteins in plant stress tolerance and resistance, describes the current mechanistic insights into unstructured proteins such as the disorder-to-order transition for adopting secondary structures to interact with specific partners (i.e., cellular membranes, membrane proteins, metal ions, and DNA), and elucidates the roles of liquid–liquid phase separation driven by protein disorder in stress responses. By comparing IDP studies in animal systems, this article provides conceptual principles of plant protein disorder in stress adaptation, reveals the current research gaps, and advises on the future research direction. The highlighting of relevant unanswered questions in plant protein disorder research aims to encourage more studies on these emerging topics to understand the mechanisms of action behind their stress resistance phenotypes.

Liquid-liquid phase separation (LLPS) regulates many biological processes including RNA metabolism, chromatin rearrangement, and signal transduction. Aberrant LLPS potentially leads to serious diseases. Therefore, the identification of the LLPS proteins is crucial. Traditionally, biochemistry-based methods for identifying LLPS proteins are costly, time-consuming, and laborious. In contrast, artificial intelligence-based approaches are fast and cost-effective and can be a better alternative to biochemistry-based methods. Previous research methods employed word2vec in conjunction with machine learning or deep learning algorithms. Although word2vec captures word semantics and relationships, it might not be effective in capturing features relevant to protein classification, like physicochemical properties, evolutionary relationships, or structural features. Additionally, other studies often focused on a limited set of features for model training, including planar π contact frequency, pi-pi, and β-pairing propensities. To overcome such shortcomings, this study first constructed a reliable dataset containing 1206 protein sequences, including 603 LLPS and 603 non-LLPS protein sequences. Then a computational model was proposed to efficiently identify the LLPS proteins by perceiving semantic information of protein sequences directly; using an ESM2-36 pre-trained model based on transformer architecture in conjunction with a convolutional neural network. The model could achieve an accuracy of 85.68% and 89.67%, respectively on training data and test data, surpassing the accuracy of previous studies. The performance demonstrates the potential of our computational methods as efficient alternatives for identifying LLPS proteins. [ABSTRACT FROM AUTHOR]

The phase separation of N 6 -methyladenosine (m 6 A) binding protein YTHDF2 plays a vital role in arsenic-induced skin damage, and YTHDF2 can bind to m 6 A-methylated mRNA of tumor suppressor PTEN. However, whether and how YTHDF2 phase separation regulates PTEN involved in arsenic-induced malignant transformation of keratinocytes remains blank. Here, we established arsenite-induced transformation models with stable expression of wild-type YTHDF2 or mutant YTHDF2 protein in vitro and in vivo. We found that the YTHDF2 protein underwent phase separation during arsenite-induced malignant transformation of keratinocytes, and YTHDF2 phase separation promoted the malignant phenotype of keratinocytes. Mechanically, YTHDF2 phase separation reduced PTEN protein levels, which in turn activated the pro-survival AKT signal. The binding of YTHDF2 to multiple m 6 A sites on PTEN mRNA drove YTHDF2 phase separation, inhibiting PTEN translation initiation and thus reducing PTEN protein levels. YTHDF2 phase separation recruited translation-initiation-factor kinase EIF2AK1 to phosphorylate eIF2α, thereby inhibiting translation initiation of poly-m 6 A-methylated PTEN mRNA. Furthermore, arsenite-induced oxidative stress triggered YTHDF2 phase separation by increasing m 6 A levels of PTEN mRNA. Our results demonstrated that YTHDF2 phase separation promotes arsenite-induced malignant transformation by inhibiting PTEN translation in a poly-m 6 A-dependent manner. This study sheds light on arsenic carcinogenicity from the novel aspect of m 6 A-mediated YTHDF2 phase separation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The SARS-CoV-2 Nucleocapsid protein (N) performs several functions during the viral lifecycle, including transcription regulation and viral genome encapsulation. We hypothesized that N toggles between these functions via phosphorylation-induced conformational change, thereby altering N interactions with membranes and RNA. We found that phosphorylation changes how biomolecular condensates composed of N and RNA interact with membranes: phosphorylated N (pN) condensates form thin films, while condensates with unmodified N are engulfed. This partly results from changes in material properties, with pN forming less viscous and elastic condensates. The weakening of protein-RNA interaction in condensates upon phosphorylation is driven by a decrease in binding between pN and unstructured RNA. We show that phosphorylation induces a conformational change in the serine/arginine-rich region of N that increases interaction between pN monomers and decreases nonspecific interaction with RNA. These findings connect the conformation, material properties, and membrane-associated states of N, with potential implications for COVID-19 treatment., Competing Interests: Declaration of Interests The authors declare no competing interests.

Background: The growth and metastasis of pancreatic cancer (PC) has been found to be closely associated with liquid-liquid phase separation (LLPS). This study sought to identify LLPS-related biomarkers in PC to construct a robust prognostic model., Methods: Transcriptomic data and clinical information related to PC were retrieved from publicly accessible databases. The PC-related data set was subjected to differential expression, Mendelian randomization (MR), univariate Cox, and least absolute selection and shrinkage operator analyses to identify biomarkers. Using the biomarkers, we subsequently constructed a risk model, identified the independent prognostic factors of PC, established a nomogram, and conducted an immune analysis., Results: The study identified four genes linked with an increased risk of PC; that is, PYGB, ACTR3, CCNA2 , and ITGB1 . Conversely, ATP8A1 , and RAP1GAP2 were found to provide protection against PC. These findings contributed significantly to the development of a highly precise risk model in which risk, age, and pathology N stage were categorized as independent factors in predicting the prognosis of PC patients. Using these factors, a nomogram was established to predict survival outcomes accurately. An immune analysis revealed varying levels of eosinophils, gamma delta T cells, and other immune cells between the distinct risk groups. The high-risk patients exhibited increased potential for immune escape, while the low-risk patients showed a higher response to immunotherapy., Conclusions: Six genes were identified as having potential causal relationships with PC. These genes were integrated into a prognostic risk model, thereby serving as unique prognostic signatures. Our findings provide novel insights into predicting the prognosis of PC patients., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://jgo.amegroups.com/article/view/10.21037/jgo-24-426/coif). The authors have no conflicts of interest to declare., (2024 Journal of Gastrointestinal Oncology. All rights reserved.)

Liquid ‒ liquid phase separation (LLPS) is a general phenomenon recently recognized to be critically involved in the regulation of a variety of cellular biological processes, such as transcriptional regulation, heterochromatin formation and signal transduction, through the compartmentalization of proteins or nucleic acids into droplet-like condensates. These processes are directly or indirectly related to tumor initiation and treatment. Posttranslational modifications (PTMs), which represent a rapid and reversible mechanism involved in the functional regulation of proteins, have emerged as key events in modulating LLPS under physiological or pathophysiological conditions, including tumorigenesis and antitumor therapy. In this review, we introduce the biological functions participated in cancer-associated LLPS, discuss the potential roles of LLPS during tumor onset or therapy, and emphasize the mechanistic characteristics of LLPS regulated by PTMs and its effects on tumor progression. We then provide a perspective on further studies on LLPS and its regulation by PTMs in cancer research. This review aims to broaden the understanding of the functions of LLPS and its regulation by PTMs under normal or aberrant cellular conditions. • Tumor-related LLPS can be finely tuned by PTMs. • The progression of tumors appears to be intricately linked with aberrant LLPS. • PTMs regulated-LLPS play an important role in cancer initiation and clinical tumor treatment. [ABSTRACT FROM AUTHOR]

Inclusion bodies (IBs) of respiratory syncytial virus (RSV) are formed by liquid-liquid phase separation (LLPS) and contain internal structures termed "IB-associated granules" (IBAGs), where anti-termination factor M2-1 and viral mRNAs are concentrated. However, the mechanism of IBAG formation and the physiological function of IBAGs are unclear. Here, we found that the internal structures of RSV IBs are actual M2-1-free viral messenger ribonucleoprotein (mRNP) condensates formed by secondary LLPS. Mechanistically, the RSV nucleoprotein (N) and M2-1 interact with and recruit PABP to IBs, promoting PABP to bind viral mRNAs transcribed in IBs by RNA-recognition motif and drive secondary phase separation. Furthermore, PABP-eIF4G1 interaction regulates viral mRNP condensate composition, thereby recruiting specific translation initiation factors (eIF4G1, eIF4E, eIF4A, eIF4B and eIF4H) into the secondary condensed phase to activate viral mRNAs for ribosomal recruitment. Our study proposes a novel LLPS-regulated translation mechanism during viral infection and a novel antiviral strategy via targeting on secondary condensed phase., Competing Interests: Conflict of interest Prof. Mingzhou Chen is an ​editorial board member ​for ​Virologica Sinica, and was not involved in the editorial review or the decision to publish this article. ​The authors declare that they have no conflict of interest., (Copyright © 2023 The Authors. Publishing services by Elsevier B.V. All rights reserved.)