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Nucleoli are multicomponent condensates defined by coexisting sub-phases. We identified distinct intrinsically disordered regions (IDRs), including acidic (D/E) tracts and K-blocks interspersed by E-rich regions, as defining features of nucleolar proteins. We show that the localization preferences of nucleolar proteins are determined by their IDRs and the types of RNA or DNA binding domains they encompass. In vitro reconstitutions and studies in cells showed how condensation, which combines binding and complex coacervation of nucleolar components, contributes to nucleolar organization. D/E tracts of nucleolar proteins contribute to lowering the pH of co-condensates formed with nucleolar RNAs in vitro. In cells, this sets up a pH gradient between nucleoli and the nucleoplasm. By contrast, juxta-nucleolar bodies, which have different macromolecular compositions, featuring protein IDRs with very different charge profiles, have pH values that are equivalent to or higher than the nucleoplasm. Our findings show that distinct compositional specificities generate distinct physicochemical properties for condensates., QC 20240418

DNA underwinding (untwisting) is a crucial step in transcriptional activation. DNA underwinding occurs between the site where torque is generated by RNA polymerase (RNAP) and the site where the axial rotation of DNA is constrained. However, what constrains DNA axial rotation in the nucleus is yet unknown. Here, we show that the anchorage to the nuclear protein condensates constrains DNA axial rotation for DNA underwinding in the nucleolus. In situ super-resolution imaging of underwound DNA reveal that underwound DNA accumulates in the nucleolus, a nuclear condensate with a core–shell structure. Specifically, underwound DNA is distributed in the nucleolar core owing to RNA polymerase I (RNAPI) activities. Furthermore, underwound DNA in the core decreases when nucleolar shell components are prevented from binding to their recognition structure, G-quadruplex (G4). Taken together, these results suggest that the nucleolar shell provides anchoring sites that constrain DNA axial rotation for RNAPI-driven DNA underwinding in the core. Our findings will contribute to understanding how nuclear protein condensates make up constraints for the site-specific regulation of DNA underwinding and transcription.

Identifying genetic modifiers of familial amyotrophic lateral sclerosis (ALS) may reveal targets for therapeutic modulation with potential application to sporadic ALS. GGGGCC (G4C2) repeat expansions in the C9orf72 gene underlie the most common form of familial ALS, and generate toxic arginine-containing dipeptide repeats (DPRs), which interfere with membraneless organelles, such as the nucleolus. Here we considered senataxin (SETX), the genetic cause of ALS4, as a modifier of C9orf72 ALS, because SETX is a nuclear helicase that may regulate RNA-protein interactions involved in ALS dysfunction. After documenting that decreased SETX expression enhances arginine-containing DPR toxicity and C9orf72 repeat expansion toxicity in HEK293 cells and primary neurons, we generated SETX fly lines and evaluated the effect of SETX in flies expressing either (G4C2)58 repeats or glycine-arginine-50 [GR(50)] DPRs. We observed dramatic suppression of disease phenotypes in (G4C2)58 and GR(50) Drosophila models, and detected a striking relocalization of GR(50) out of the nucleolus in flies co-expressing SETX. Next-generation GR(1000) fly models, that show age-related motor deficits in climbing and movement assays, were similarly rescued with SETX co-expression. We noted that the physical interaction between SETX and arginine-containing DPRs is partially RNA-dependent. Finally, we directly assessed the nucleolus in cells expressing GR-DPRs, confirmed reduced mobility of proteins trafficking to the nucleolus upon GR-DPR expression, and found that SETX dosage modulated nucleolus liquidity in GR-DPR-expressing cells and motor neurons. These findings reveal a hitherto unknown connection between SETX function and cellular processes contributing to neuron demise in the most common form of familial ALS.

We analyze a legal situation in which a plaintiff suffers the total damage of the cumulative injury that is caused by multiple sequences of tortfeasors’ wrongful acts. This liability situation is modeled by a tuple consisting of a rooted-tree network of causation of the injury, and a list of marginal damages. The problem is to determine how to share the total damage among the tortfeasors in such situations. A rule is a mapping that associates with each liability problem a list of payments to be made by the tortfeasors to pay for the plaintiff’s harm. We axiomatize two rules that are obtained as the Nucleolus and the Shapley value, respectively, of a coalitional game with tortfeasors as players. The axioms involved in these axiomatizations are derived from the legal concept of tort law.

In plant cells, a large pool of iron (Fe) is contained in the nucleolus, as well as in chloroplasts and mitochondria. A central determinant for intracellular distribution of Fe is nicotianamine (NA) generated by NICOTIANAMINE SYNTHASE (NAS). Here, we used Arabidopsis thaliana plants with disrupted NAS genes to study the accumulation of nucleolar iron and understand its role in nucleolar functions and more specifically in rRNA gene expression. We found that nas124 triple mutant plants, which contained lower quantities of the iron ligand NA, also contained less iron in the nucleolus. This was concurrent with the expression of normally silenced rRNA genes from nucleolar organizer regions 2 (NOR2). Notably, in nas234 triple mutant plants, which also contained lower quantities of NA, nucleolar iron and rDNA expression were not affected. In contrast, in both nas124 and nas234, specific RNA modifications were differentially regulated in a genotype dependent manner. Taken together, our results highlight the impact of specific NAS activities in RNA gene expression. We discuss the interplay between NA and nucleolar iron with rDNA functional organization and RNA methylation.

This paper introduces the new class of one-bound core games, where the core can be described by either a lower bound or an upper bound on the payoffs of the players, named lower bound core games and upper bound core games, respectively. We study the relation of the class of one-bound core games with several other classes of games and characterize the new class by the structure of the core and in terms of Davis-Maschler reduced games. We also provide explicit expressions and axiomatic characterizations of the nucleolus for one-bound core games, and show that the nucleolus coincides with the Shapley value when these games are convex.

The nucleolus is a subnuclear compartment critical in ribosome biogenesis and cellular stress responses. These mechanisms are governed by a complex interplay of proteins, including NOC1, a member of the NOC family of nucleolar proteins responsible for controlling rRNA processing and ribosomal maturation. This study reveals a novel relationship between NOC1 and MYC transcription factor, known for its crucial role in controlling ribosomal biogenesis, cell growth, and proliferation. Here, we demonstrate that NOC1 functions as a direct target of MYC, as it is transcriptionally induced through a functional MYC-binding E-box sequence in the NOC1 promoter region. Furthermore, protein interactome analysis reveals that NOC1-complex includes the nucleolar proteins NOC2 and NOC3 and other nucleolar components such as Nucleostemin1 Ns1 transporters of ribosomal subunits and components involved in rRNA processing and maturation. In response to MYC, NOC1 expression and localization within the nucleolus significantly increase, suggesting a direct functional link between MYC activity and NOC1 function. Notably, NOC1 over-expression leads to the formation of large nuclear granules and enlarged nucleoli, which co-localize with nucleolar fibrillarin and Ns1. Additionally, we demonstrate that NOC1 expression is necessary for Ns1 nucleolar localization, suggesting a role for NOC1 in maintaining nucleolar structure. Finally, the co-expression of NOC1 and MYC enhances nucleolus size and maintains their co-localization, outlining another aspect of the cooperation between NOC1 and MYC in nucleolar dynamics. This study also reveals an enrichment with NOC1 with few proteins involved in RNA processing, modification, and splicing. Moreover, proteins such as Ythdc1, Flacc, and splenito are known to mediate N6-methyladenosine (m6A) methylation of mRNAs in nuclear export, revealing NOC1's potential involvement in coordinating RNA splicing and nuclear mRNA export. In summary, we uncovered novel rol

Influenza A virus double-helical ribonucleoprotein complex (RNP) performs transcription and replication of viral genomic RNA (vRNA). Although RNP formation occurs in the nuclei of virus-infected cells, the nuclear domains involved in this process remain unclear. Here, we show that the nucleolus is an essential site for functional RNP formation. Viral nucleoprotein (NP), a major RNP component, temporarily localized to the nucleoli of virus-infected cells. Mutations in a nucleolar localization signal (NoLS) on NP abolished double-helical RNP formation, resulting in a loss of viral RNA synthesis ability, whereas ectopic fusion of the NoLS enabled the NP mutant to form functional double-helical RNPs. Furthermore, nucleolar disruption of virus-infected cells inhibited NP assembly into double-helical RNPs, resulting in decreased viral RNA synthesis. Collectively, our findings demonstrate that NP migration into the nucleolus is a critical step for functional RNP formation, showing the importance of the nucleolus in the influenza virus life cycle.

Dynamic morphological changes of intracellular organelles are often regulated by protein phosphorylation or dephosphorylation. Phosphorylation modulates stereospecific interactions among structured proteins, but how it controls molecular interactions among unstructured proteins and regulates their macroscopic behaviours remains unknown. Here we determined the cell cycle-specific behaviour of Ki-67, which localizes to the nucleoli during interphase and relocates to the chromosome periphery during mitosis. Mitotic hyperphosphorylation of disordered repeat domains of Ki-67 generates alternating charge blocks in these domains and increases their propensity for liquid–liquid phase separation (LLPS). A phosphomimetic sequence and the sequences with enhanced charge blockiness underwent strong LLPS in vitro and induced chromosome periphery formation in vivo. Conversely, mitotic hyperphosphorylation of NPM1 diminished a charge block and suppressed LLPS, resulting in nucleolar dissolution. Cell cycle-specific phase separation can be modulated via phosphorylation by enhancing or reducing the charge blockiness of disordered regions, rather than by attaching phosphate groups to specific sites.

We present fluorogenic cationic organo chalcogens that are highly selective to RNA. We have demonstrated that the conformational dynamics and subsequently the optical properties of these dyes can be controlled to facilitate efficient bioimaging. We report the application of organoselenium and organosulfur-based cell-permeable red-emissive probes bearing a favorable cyclic sidearm for selective and high contrast imaging of cell nucleoli. The probes exhibit high quantum yield upon interacting with RNA in an aqueous solution. An in-depth multiscale simulation study reveals that the prominent rotational freezing of the electron-donating sidearm of the probes in the microenvironment of RNA helps in attaining more planar conformation when compared to DNA. It exerts a greater extent of intramolecular charge transfer and hence leads to enhanced fluorescence emission. A systematic structure-interaction relationship study highlighted the impact of heavy-chalcogens toward the improved emissive properties of the probes., QC 20221109

Platinum (Pt) compounds such as oxaliplatin are among the most commonly prescribed anti-cancer drugs. Despite their considerable clinical impact, the molecular basis of platinum cytotoxicity and cancer specificity remain unclear. Here we show that oxaliplatin, a backbone for the treatment of colorectal cancer, causes liquid-liquid demixing of nucleoli at clinically relevant concentrations. Our data suggest that this biophysical defect leads to cell-cycle arrest, shutdown of Pol I-mediated transcription, and ultimately cell death. We propose that instead of targeting a single molecule, oxaliplatin preferentially partitions into nucleoli, where it modifies nucleolar RNA and proteins. This mechanism provides a general approach for drugging the increasing number of cellular processes linked to biomolecular condensates.

In truckload transportation procurement networks, shippers purchasing the services of truckload carriers seek to form multi-company tours which consist of regularly scheduled shipments with minimal empty truck movements with the aim of getting better rates from the carriers in return. Identifying the minimum cost collaborative solution and sharing the costs in a fair manner are two interrelated and critical tasks for the success of such networks. The restrictions on the tours which can be formed make both of these tasks very challenging. In this paper, we take a cooperative game theory approach and study the resulting constrained lane covering game. We first show the set of conditions which must be satisfied for the game to have a non-empty core. We propose nucleolus based cost allocation methods. We also develop column and row generation methods for solving these mathematical models. We compare our proposed cost allocation models with alternative cost allocation methods and each other using a set of core stability metrics through computational experiments on randomly generated instances from the literature. © 2022 Elsevier Ltd, Türkiye Bilimsel ve Teknolojik Araştırma Kurumu, TÜBİTAK: 112M227, The research presented in this paper was partially supported by the Scientific and Technological Research Council of Turkey (TUBITAK) through grant 112M227 .

In truckload transportation procurement networks, shippers purchasing the services of truckload carriers seek to form multi-company tours which consist of regularly scheduled shipments with minimal empty truck movements with the aim of getting better rates from the carriers in return. Identifying the minimum cost collaborative solution and sharing the costs in a fair manner are two interrelated and critical tasks for the success of such networks. The restrictions on the tours which can be formed make both of these tasks very challenging. In this paper, we take a cooperative game theory approach and study the resulting constrained lane covering game. We first show the set of conditions which must be satisfied for the game to have a non-empty core. We propose nucleolus based cost allocation methods. We also develop column and row generation methods for solving these mathematical models. We compare our proposed cost allocation models with alternative cost allocation methods and each other using a set of core stability metrics through computational experiments on randomly generated instances from the literature. © 2022 Elsevier Ltd, Türkiye Bilimsel ve Teknolojik Araştırma Kurumu, TÜBİTAK: 112M227, The research presented in this paper was partially supported by the Scientific and Technological Research Council of Turkey (TUBITAK) through grant 112M227 .

This work deals with the weighted excesses of players in cooperative games which are obtained by summing up all the weighted excesses of all coalitions to which they belong. We first show that the resulting payoff vector is the corresponding least square value by lexicographically minimizing the individual weighted excesses of players over the preimputation set, and thus give an alternative characterization of the least square values. Second, we show that these results give rise to lower and upper bounds for the core payoff vectors and, using these bounds, we show that the least square values can be seen as the center of a polyhedron defined by these bounds. This provides a second new characterization of the least square values. Third, we show that the individually rational least square value is the solution that lexicographically minimizes the individual weighted excesses of players over the imputation set.

Microalgae are used in food and feed, and they are considered a potential feedstock for sustainably produced chemicals and biofuel. However, production of microalgal-derived chemicals is not yet economically feasible. Genetic engineering could bridge the gap to industrial application and facilitate the production of novel products from microalgae. Here, we report the discovery of a novel gene expression system in the oleaginous microalga Nannochloropsis that exploits the highly efficient transcriptional activity of RNA polymerase I and an internal ribosome entry site for translation. We identified the nucleolus as a genomic safe harbor for Pol I transcription and used it to construct transformant strains with consistently strong transgene expression. The new expression system provides an outstanding tool for genetic and metabolic engineering of microalgae and thus will probably make substantial contributions to microalgal research.

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This paper introduces the new class of two-bound core games, where the core can be described by a lower bound and an upper bound on the payoffs of the players. Many classes of games turn out to be two-bound core games. We show that the core of each two-bound core game can be described equivalently by the pair of exact core bounds, and study to what extent the exact core bounds can be stretched while retaining the core description. We provide explicit expressions of the nucleolus for two-bound core games in terms of all pairs of bounds describing the core, using the Talmud rule for bankruptcy problems, and study to what extent these expressions are robust against game changes.

This thesis focuses on the spatiotemporal mapping of proteins at a subcellular level. In other words, determining the cellular location of proteins over time. From a biological point of view, knowledge about protein location is fundamental to understand protein function. In the longer run, this also means better understanding of cells in the context of health and disease, since protein malfunction and mislocalization are two important factors during disease development. Using an antibody-based imaging approach, Paper I contains a subcellular map of 12 003 protein in 30 different cellular structures, freely accessible as part of the Human Protein Atlas (www.proteinatlas.org). Apart from enabling exploration of the organellar proteomes, we conclude that half of the human proteins localize to multiple compartments, and that almost one fifth display cell-to-cell variations in terms of protein expression. Paper II aimed to decrease the cumbersome work of manual protein location annotations by leveraging the power of the crowd through citizen science. By integrating the image-classification task into a video game with a massive player base, EVE online, protein location labels could be efficiently and rapidly assessed compared to manual curation from a few experts. To compare the performance of the players, a deep learning classifier was developed. The algorithm was capable of classifying protein location in images containing several challenging localization problems, such as multilocalizing proteins, cell line variations and rare classes. Using the protein location data from Paper I and Paper II, Paper III presents an image-based characterization of the nucleolar proteome. In total, 1 318 nucleolar proteins are included, of which 157 localizes to a fourth nucleolar compartment, the nucleolar rim. Additionally, 65 proteins were detected on the chromosomal periphery during mitosis, and these could be further divided into two recruitment phenotypes with different temporal p, Den här avhandlingen fokuserar på spatiotemporal karakterisering av proteiner på subcellulär nivå. Med andra ord beskrivs proteiners lokalisation i cellen över tid. Kunskap om proteiners lokalisation är grundläggande för att bättre förstå proteiners funktion. I ett längre perspektiv ger det också bättre förståelse av både friska och sjuka celler, eftersom felfungerande proteiner och felaktig proteinlokalisation är två faktorer som driver sjukdomsutveckling framåt. Som en del av Human Protein Atlas-projektet (www.proteinatlas.org), innehåller Artikel I en subcellulär karta över 12 003 proteiner i 30 stycken olika organeller som kartlagts med hjälp av antikroppsbaserad mikroskopi. Utöver att ge en överblick över de olika organellproteomen, kunde vi också konstatera att hälften av alla proteiner lokaliserar till mer än en cellstruktur, samt att nästan en femtedel uppvisar skillnader i uttryck mellan celler. Målet i Artikel II var att minska arbetstiden för manuell annotering av proteinlokalisation genom att låta tusentals datorspelare i ett medborgarforskningsprojekt bidra. En bildklassificeringuppgift integrerades i datorspelet EVE online. Tack vare detta kunde proteinernas lokalisation snabbt och effektivt bestämmas, till skillnad från manuell annotering utförd av ett fåtal experter. För att utvärdera hur spelarna presterade, utvecklade vi samtidigt en klassificeringsalgoritm med hjälp av djup maskininlärning. Algoritmen visade sig kunna predicera proteinlokalisation i bilder trots att de innehöll flera försvårande omständigheter, såsom multilokaliserande proteiner, cellinjevariationer och ovanliga klasser. Med hjälp av proteinlokalisationerna från Artikel I och Artikel II, presenterar vi i Artikel III en bildbaserad karakterisering av nukleolens proteom. Sammanlagt visar vi data för 1 318 nukleolproteiner, varav 157 lokaliserar till en fjärde substruktur i gränsen mellan nukleolen och nukleoplasman. Vi visar även att 65 nukleolproteiner omlokaliseras till kromosomer, QC 2021-10-12

1-convex games have been introduced by Theo Driessen in his 1985 PhD dissertation. They form an interesting class of games for at least one reason: the core of a 1-convex n-player game is a regular simplex of dimension n – 1 or a single point. As a consequence, its nucleolus is the center of gravity of the core. We recall and extend the results obtained by Driessen and provide examples and applications.

Any space exploration initiative, such as the human presence in the Moon and Mars, must incorporate plants for life support. To enable space plant culture we need to understand how plants respond to extraterrestrial conditions, adapt to them, and compensate their deleterious effects at multiple levels. Gravity is a major difference between the terrestrial and the extraterrestrial environment. Gravitropism is the process of establishing a growth direction for plant organs according to the gravity vector. Gravity signals are sensed at specialized tissues by the motion of amyloplasts called statoliths and transduced to produce a cellular polarization capable of influencing the transport of auxin. Gravity alterations eventually result in changes in the lateral balance of auxin in the root, producing deviations of the growth direction. Under microgravity, auxin changes affect the root meristem causing increased cell proliferation and decreased cell growth. The nucleolus, the nuclear site of production of ribosomes, is a marker of this unbalance, which could alter plant development. At the molecular level, microgravity induces a reprogramming of gene expression that mostly affects plant defense systems against abiotic stresses, indicating that these categories of genes are involved in the adaptation to extraterrestrial habitats. Nevertheless, no specific genes for plant response to gravitational stress have been identified. Despite this stress, plants survive, developing until the adult stage and reproducing under microgravity conditions. A major research challenge is to identify environmental factors, such as light, which could interact, modulate, or balance the impact of gravity, contributing to the tolerance and survival of plants under spaceflight conditions. Understanding the crosstalk between light and gravity sensing will contribute to the success of the next generation agriculture in human settlements outside the Earth.

Game theory based energy sharing schemes emerged in recent years to incentivize efficient management of the increasing amount of distributed energy resources. Among these, cooperative game theoretic schemes provide detailed financial incentives on the individual prosumer level. The nucleolus, a mechanism to allocate these financial incentives, has been proven to guarantee the prosumers’ willingness to participate. However, the computation time of the nucleolus increases exponentially with the number of participants, strictly limiting the size of this scheme. This study proposes to incorporate clustering techniques to estimate the nucleolus at reduced computation times, where a novel marginal contribution profile is used as the clustering features. A stratified random sampling based approach is formulated to evaluate the estimation performance, showing that the proposed method is able to scale up the cooperative energy management scheme from less than 15 players to over 100 players while maintaining high accuracy of the nucleolus estimation.

Unlike in most eukaryotic cells, the genetic information of budding yeast in the exponential growth phase is only present in the form of decondensed chromatin, a configuration that does not allow its visualization in cell nuclei conventionally prepared for transmission electron microscopy. In this work, we studied the distribution of chromatin and its relationships to the nucleolus using different cytochemical and immunocytological approaches applied to yeast cells subjected to hyperosmotic shock. Our results show that osmotic shock induces the formation of heterochromatin patches in the nucleoplasm and intranucleolar regions of the yeast nucleus. In the nucleolus, we further revealed the presence of osmotic shock-resistant DNA in the fibrillar cords which, in places, take on a pinnate appearance reminiscent of ribosomal genes in active transcription as observed after molecular spreading (“Christmas trees”). We also identified chromatin-associated granules whose size, composition and behaviour after osmotic shock are reminiscent of that of mammalian perichromatin granules. Altogether, these data reveal that it is possible to visualize heterochromatin in yeast and suggest that the yeast nucleus displays a less-effective compartmentalized organization than that of mammals., SCOPUS: ar.j, info:eu-repo/semantics/published

The nucleolus is essential for ribosome biogenesis and is involved in many other cellular functions. We performed a systematic spatiotemporal dissection of the human nucleolar proteome using confocal microscopy. In total, 1,318 nucleolar proteins were identified; 287 were localized to fibrillar components, and 157 were enriched along the nucleoplasmic border, indicating a potential fourth nucleolar subcompartment: the nucleoli rim. We found 65 nucleolar proteins (36 uncharacterized) to relocate to the chromosomal periphery during mitosis. Interestingly, we observed temporal partitioning into two recruitment phenotypes: early (prometaphase) and late (after metaphase), suggesting phase-specific functions. We further show that the expression ofMKI67 is critical for this temporal partitioning. We provide the first proteome-wide analysis of intrinsic protein disorder for the human nucleolus and show that nucleolar proteins in general, and mitotic chromosome proteins in particular, have significantly higher intrinsic disorder level compared to cytosolic proteins. In summary, this study provides a comprehensive and essential resource of spatiotemporal expression data for the nucleolar proteome as part of the Human Protein Atlas., QC 20201103

The transcription factor BMAL1 is a core element of the circadian clock that contributes to cyclic control of genes transcribed by RNA polymerase II. By using biochemical cellular fractionation and immunofluorescence analyses we reveal a previously uncharacterized nucleolar localization for BMAL1. We used an unbiased approach to determine the BMAL1 interactome by mass spectrometry and identified NOP58 as a prominent nucleolar interactor. NOP58, a core component of the box C/D small nucleolar ribonucleoprotein complex, associates with Snord118 to control specific pre-ribosomal RNA (pre-rRNA) processing steps. These results suggest a non-canonical role of BMAL1 in ribosomal RNA regulation. Indeed, we show that BMAL1 controls NOP58-associated Snord118 nucleolar levels and cleavage of unique pre-rRNA intermediates. Our findings identify an unsuspected function of BMAL1 in the nucleolus that appears distinct from its canonical role in the circadian clock system.

Circadian rhythms regulate our physiological, metabolic, and behavioral activity in a 24-hour cycle to maintain organismal homeostasis. In virtually all tissues, the intrinsic circadian clock is governed by a canonical molecular transcriptional-translational feedback mechanism that is self-sustained. Whether additional functions for the molecular circadian clock exist remains largely unexplored. Here we investigate an unprecedented role for the circadian clock protein, BMAL1, in the nucleolus. We identified novel BMAL1 interactions with resident nucleolar proteins using biochemical fractionation and pull-down assays. Moreover, through use of molecular assays we reveal a critical role in pre-ribosomal RNA processing and maturation. Instead, environmental-mediated synchronization of peripheral tissues is regulated in part by external cues including light-dark, sleep-wake, and feed-fasting cycles. The degree to which non-photic environmental factors perturb the circadian clock in the brain continues to be a major point of interest. We studied an in vivo model to elucidate the effects of drugs of abuse as stressors on the striatal circadian clock. Mice treated with an acute dose of cocaine were sampled throughout a full circadian cycle. Using high-throughput RNA sequencing, our findings show cocaine drastically reprograms the circadian transcriptional clock in the nucleus accumbens. Furthermore, we reveal dopamine signaling is required for the cocaine-mediated PPARgamma activation that leads to de novo circadian transcriptional rhythmicity. Taken together, in the studies presented, we implement two discrete approaches to further our knowledge on the versatility and reprogrammability of the molecular clock. Our novel findings provide previously unexplored connections between RNA regulation or drugs of abuse and the circadian clock, and uncover new possible therapeutic targets for treatment development for disorders such as addiction.

Background/Purpose : Reducing unnecessary laboratory investigations is a health economic priority. Anti-nuclear antibody (ANA) testing is performed as part of the diagnostic work up for autoimmune disease, or in patients with inflammatory or musculoskeletal symptoms. The value of serial ANA testing is unclear, but anecdotal evidence suggests it is frequent. We sought to evaluate the utility of repeated ANA testing in a large tertiary healthcare network, to determine the evidence base for decision support actions. Methods : The primary endpoint was whether a longitudinal change in ANA resulted in new ANA associated diagnoses. Secondary endpoints included calculation of the total cost associated with repeated testing and the examination of baseline ANA testing behaviours. We retrospectively analysed data from a multi-centre tertiary health network across a 7-year period (March 2011 to July 2018). ANA and other autoimmune test results were obtained from the hospital systems. The laboratory positive ANA cut offof 1:160 was used. Clinical information was sourced from medical records on all patients who had a change in ANA result from negative to positive on repeat testing. The cost of repeated ANA testing was calculated based on the baseline cost to the payer. Descriptive statistics were performed using Stata version 15. Results : A total of 36,715 ANA tests (excluding 980 cancelled same-day requests) were performed in 28,840 patients. Of these, 14,058 (38.3%) were positive, with females accounting for 9,265 (65.9%, p< 0.001). The most frequent ANA patterns were homogenous (47.4%) and nucleolar (23.3%). The distribution of ANA titres was: 1:160 (41.4%), 1:320 (15.3%), 1:640 (13.1%) and 1:1280 (29.2%). 7,875 (21.4%) tests were repeat tests in 4,887 patients, with test frequency in an individual patient ranging from 2-45. Of repeated tests, 79% of results remained the same, while 541 (11.1%) results changed from negative to positive. In the 501 of these in whom medical rec

Nucleophosmin (NPM1) is a mostly nucleolar protein with crucial functions in cell growth and homeostasis, including regulation of ribosome biogenesis and stress response. Such multiple activities rely on its ability to interact with nucleic acids and with hundreds of proteins, as well as on a dynamic subcellular distribution. NPM1 is thus regulated by a complex interplay between localization and interactions, further modulated by post-translational modifications. NPM1 is a homopentamer, with globular domains connected by long, intrinsically disordered linkers. This configuration allows NPM1 to engage in liquid-liquid phase separation phenomena, which could underlie a key role in nucleolar organization. Here, we will discuss NPM1 conformational and functional versatility, emphasizing its emerging, and still largely unexplored, role in DNA damage repair. Since NPM1 is altered in a subtype of acute myeloid leukaemia (AML), we will also present ongoing research on the molecular mechanisms underlying its pathogenic role and potential NPM1-targeting therapeutic strategies.

Upon acute heat stress (HS), overall mRNA transcription, processing, and export are inhibited, leading to cell growth arrest. However, how cells turn off mRNA metabolism is not fully understood. Here, we show that acute HS results in the segregation and aggregation of multiple nuclear and nucleolar proteins into ring-like structures located at the nucleolar periphery (nucleolar rings [NuRs]). NuRs sequester essential factors required for nuclear mRNA metabolism and nuclear pore complex function, as well as cell-cycle regulators. When cells are switched back to growing temperatures, NuRs disaggregate, and their components relocate to their functional environments in an Hsf1- and Hsp104-dependent manner, and concomitantly with the reinitiation of cell growth. These findings highlight the contribution of reversible protein aggregation to the inhibition of overall RNA-related activities in the nucleus and its functional relevance in the maintenance of cellular homeostasis during acute HS.

The Cdc14 phosphatase is a key regulator of mitosis in the budding yeast Saccharomyces cerevisiae. Cdc14 was initially described as playing an essential role in the control of cell cycle progression by promoting mitotic exit on the basis of its capacity to counteract the activity of the cyclin-dependent kinase Cdc28/Cdk1. A compiling body of evidence, however, has later demonstrated that this phosphatase plays other multiple roles in the regulation of mitosis at different cell cycle stages. Here, we summarize our current knowledge about the pivotal role of Cdc14 in cell cycle control, with a special focus in the most recently uncovered functions of the phosphatase.

The development of a murine model of osmotic demyelinating syndrome (ODS) allowed to study changes incurred in extrapontine zones of the CNS and featured neuron and glial cell changes in the relay thalamic ventral posterolateral (VPL) and ventral posteromedial (VPM) nuclei before, during and after ODS induction, and characterized without immune response. There, the neuron Wallerian-type deteriorations were verified with fine structure modifications of the neuron cell body, including some nucleus topology and its nucleolus changes. Morphologic analyses showed a transient stoppage of transcriptional activities while myelinated axons in the surrounding neuropil incurred diverse damages, previously reported. Even though the regional thalamus myelin deterioration was clearly recognized with light microscopy 248 h after osmotic recovery of ODS, ultrastructure analyses demonstrated that, at that time, the same damaged parenchyma regions contained nerve cell bodies that have already reactivated nucleus transcriptions and neuroplasm translations because peculiar accumulations of fibro-granular materials, similar to those detected in restored ODS astrocytes, were revealed in these restructuring nerve cell bodies. Their aspects suggested to be accumulations of ribonucleoproteins. The findings suggested that progressive neural function's recovery in the murine model could imitate some aspects of human ODS recovery cases., SCOPUS: ar.j, info:eu-repo/semantics/published

The Glutamate Receptor Ionotropic NMDA-Associated Protein 1 (GRINA) belongs to the Lifeguard family and is involved in calcium homeostasis, which governs key processes, such as cell survival or the release of neurotransmitters. GRINA is mainly associated with membranes of the endoplasmic reticulum, Golgi, endosome, and the cell surface, but its presence in the nucleus has not been explained yet. Here we dissect, with the help of different software tools, the potential roles of GRINA in the cell and how they may be altered in diseases, such as schizophrenia or celiac disease. We describe for the first time that the cytoplasmic N-terminal half of GRINA (which spans a Proline-rich domain) contains a potential DNA-binding sequence, in addition to cleavage target sites and probable PY-nuclear localization sequences, that may enable it to be released from the rest of the protein and enter the nucleus under suitable conditions, where it could participate in the transcription, alternative splicing, and mRNA export of a subset of genes likely involved in lipid and sterol synthesis, ribosome biogenesis, or cell cycle progression. To support these findings, we include additional evidence based on an exhaustive review of the literature and our preliminary data of the protein–protein interaction network of GRINA.

[EN] This paper introduces a row and column generation algorithm for finding the nucleolus, based on a linear programming model proposed in an earlier research. Since this approach cannot return an allocation for large games, we also propose a heuristic approach, which is based on sampling the coalitions space. Experiments over medium sized games show that the proposed heuristic finds allocations which are close to the true nucleolus, in a reasonable amount of time. Experiments over 100-player games show that the proposed heuristic can be applied to games of large size.

The onset of invasion is a key step towards the development of metastatic cancer. For a cell to invade through interstitial spaces in the tissue requires a reduction in nuclear rigidity as the cell needs to deform to squeeze through small spaces. Heterochromatin Protein 1α (HP1α) is a protein that defines domains of heterochromatin, the highly compact regions of the genome, and is essential for maintaining the appropriate patterns of gene expression and genome stability. Loss or reduction of HP1α has been correlated with an increase in invasive potential in human tumours. Using an established model of Drosophila melanogaster epithelial cell invasion, the causative role HP1α plays in suppressing cellular invasive is confirmed within an epithelial tissue microenvironment. This model also demonstrates that loss of the Drosophila melanogaster HP1 homologue synergistically promotes cellular invasion in conjunction with an activated malignant signalling pathway. Importantly, human HP1α is shown to rescue this highly invasive Drosophila phenotype and demonstrates the relevance of this model to human disease, and its use for exploring protein interactions in a cellular microenvironment. As loss of nuclear integrity has been linked to a reduction in peripheral heterochromatin, the biophysical mechanisms by which HP1α acts as a suppressor of invasive potential were explored in the poorly invasive MCF7 breast cancer cell line with constitutive HP1α knock-down. These cells with reduced HP1α expression had a significant loss of nuclear membrane integrity and stiffness. The underlying nuclear lamina meshwork and associated peripheral heterochromatin was disrupted. This was associated with an increased solubility of lamina proteins, particularly lamin A, as well as the altered localisation of a number of peripheral nuclear proteins. In summary, this work established the important contribution of HP1α to the mechanical integrity of the nucleoskeleton and the role HP1α plays in supp

Production of healthy gametes in meiosis relies on the quality control and proper distribution of both nuclear and cytoplasmic contents. Meiotic differentiation naturally eliminates age-induced cellular damage by an unknown mechanism. Using time-lapse fluorescence microscopy in budding yeast, we found that nuclear senescence factors - including protein aggregates, extrachromosomal ribosomal DNA circles, and abnormal nucleolar material - are sequestered away from chromosomes during meiosis II and subsequently eliminated. A similar sequestration and elimination process occurs for the core subunits of the nuclear pore complex in both young and aged cells. Nuclear envelope remodeling drives the formation of a membranous compartment containing the sequestered material. Importantly, de novo generation of plasma membrane is required for the sequestration event, preventing the inheritance of long-lived nucleoporins and senescence factors into the newly formed gametes. Our study uncovers a new mechanism of nuclear quality control and provides insight into its function in meiotic cellular rejuvenation.

Background: Minimising unnecessary tests is a global health economic priority with multiple initiatives in place to avoid inappropriate healthcare utilisation(1) and harm. Anti-nuclear antibody (ANA) testing is frequently performed as a diagnostic test for autoimmune conditions, such as systemic lupus erythematosus (SLE) or as a screening test in patients with inflammatory or musculoskeletal symptoms. The value of serial testing in the monitoring of such conditions is unclear and false positive tests can lead to unnecessary further investigation and increased patient anxiety(2) Objectives: To evaluate the frequency of repeated ANA testing as a prelude to Electronic Medical Record (EMR) test alert design in an Australian healthcare network. The primary endpoint was calculation of the total cost associated with repeated testing and whether a longitudinal change in ANA resulted in any new ANA associated rheumatological diagnoses. Our secondary endpoint was the examination of baseline ANA testing behaviours. Method(s): We retrospectively analysed data from a multi-centre tertiary health network in Melbourne, Australia across a 7-year period (19 March 2011 to 23 July 2018). ANA and other autoimmune test results were obtained from the hospital pathology system with a positive ANA cut off set at 1:160. Clinical information was sourced from clinical information systems on patients who had a change in ANA result from negative to positive on repeat testing. The associated cost of repeated ANA testing was calculated based on the baseline cost to the public system. Result(s): A total of 36,715 ANA tests (excluding 980 cancelled same-day requests) were performed in 28,840 patients. Of these, 14,058 (38.3%) were positive with females accounting for 9,265 (65.9%, p<0.001). The most frequent ANA patterns were homogenous (47.4%) and nucleolar (23.3%). ANA titres were as follows; 1:160 (41.4%), 1:320 (15.3%), 1:640 (13.1%) and 1:1280 (29.2%). 7,875 (21.4%) of tests were repeat tests.

One of David Schmeidler’s many important contributions in his distinguished career was the introduction of the nucleolus, one of the central single-valued solution concepts in cooperative game theory. This paper is an updated survey on the nucleolus and its two related supersolutions, i.e., the kernel and the bargaining set. As a first approach to these concepts, we refer the reader to the great survey by Maschler (1992); see also the relevant chapters in Peleg and Sudholter (2003). Building on the notes of four lectures on the nucleolus and the kernel delivered by one of the authors at the Hebrew University of Jerusalem in 1999, we have updated Maschler’s survey by adding more recent contributions to the literature. Following a similar structure, we have also added a new section that covers the bargaining set. The nucleolus has a number of desirable properties, including nonemptiness, uniqueness, core selection, and consistency. The first way to understand it is based on an egalitarian principle among coalitions. However, by going over the axioms that characterize it, what comes across as important is its connection with coalitional stability, as formalized in the notion of the core. Indeed, if one likes a single-valued version of core stability that always yields a prediction, one should consider the nucleolus as a recommendation. The kernel, which contains the nucleolus, is based on the idea of “bilateral equilibrium” for every pair of players. And the bargaining set, which contains the kernel, checks for the credibility of objections coming from coalitions. In this paper, section 2 presents preliminaries, section 3 is devoted to the nucleolus, section 4 to the kernel, and section 5 to the bargaining set.

We analyze the question of how to distribute the asset value of an insolvent firm among its creditors and the firm itself. Compared to standard bankruptcy games as studied in the game-theoretic literature, we treat the firm as a player and define a new class of transferable utility games called liability games. We show that the core of a liability game is empty. We analyze the nucleolus of the game. The firm always gets a positive payment, at most equal to half of the asset value. Creditors with higher liabilities receive higher payments, but also suffer from higher deficiencies. We provide conditions under which the nucleolus coincides with a generalized proportional rule.

BPL power lines (Broadband Power Line) run under the HomePlug AV (HPAV) standard which uses the technologies CSMA/CA and TDMA as a mechanism of access to the medium, in which CSMA/CA is intended for the transmission of data packets and TDMA is used for the transmission of voice and video packets, in order to offer adequate levels of QoS. However, notwithstanding that the HPAV can reach high transfer rates, it lacks the adequate bandwidth (AB) allocation mechanism, which in turn interferes significantly with the network’s performance as the number of users rise due to the fact that only one node can transmit at once. In line with the raised above and taking into account that a BPL network can be represented as a cooperative game with transferable utilities (UT), the present paper proposes the use of two equitable bandwidth allocation techniques: nucleolus and max-min fairness, which are part of the cooperative game theory. In the comparison of the nucleolus y max-min fairness techniques as an strategy for resource allocation it was found that the latter produces the best results. Furthermore, it was made evident that game theory can be regarded as a groundbreaking strategy for the optimization of resources in a LAN network on BPL., As redes BPL (Broadband Power Line) operam sob o padrão HomePlug AV (HPAV), o qual utiliza as tecnologias CSMA/CA e TDMA como mecanismo de acesso ao meio, em que CSMA/CA está destinado à transmissão de pacotes de dados e TDMA à transmissão de pacotes de voz e vídeo, com o objetivo de oferecer adequados níveis de QoS. Contudo, embora o HPAV possa atingir altas taxas de transmissão, não conta com um adequado mecanismo de designação de largura de banda (LB), o que afeta consideravelmente o rendimento da rede à medida que aumenta o número de usuários devido a que unicamente um nó pode transmitir ao mesmo tempo. De acordo com o proposto e considerando que uma rede BPL pode ser representada como um jogo cooperativo de utilidade transferível (UT), neste artigo, propõe-se o uso de duas técnicas para a assignação equitativa de largura de banda: nucleolus e max-min fairness, as quais fazem parte da teoria de jogos cooperativos. Ao realizar o comparativo das técnicas nucleolus e max-min fairness como estratégia para a destinação de recursos, verificou-se que esta última gera os melhores resultados. Além disso, pôde-se evidenciar que a teoria de jogos pode ser considerada como uma nova estratégia para otimizar recursos em uma rede LAN sobre BPL., Las redes BPL (Broadband Power Line) operan bajo el estándar HomePlug AV (HPAV) el cual utiliza las tecnologías CSMA/CA y TDMA como mecanismo de acceso al medio, donde CSMA/CA está destinado para la transmisión de paquetes de datos y TDMA se utiliza para la transmisión de paquetes de voz y video, con el fin de ofrecer adecuados niveles de QoS. Sin embargo, pese a que el HPAV puede alcanzar altas tasas de transmisión, no cuenta con un adecuado mecanismo de asignación de ancho de banda (AB), lo cual afecta considerablemente el rendimiento de la red a medida que aumenta el número de usuarios debido a que únicamente un nodo puede transmitir a la vez. De acuerdo a lo planteado y, considerando que una red BPL puede ser representada como un juego cooperativo de utilidad transferible (UT), en el presente artículo se propone el uso de dos técnicas para la asignación equitativa de ancho de banda: nucleolus y max-min fairness, las cuales forman parte de la teoría de juegos cooperativos. Al realizar el comparativo de las técnicas nucleolus y max-min fairness como estrategia para la asignación de recursos, se encontró que esta última genera los mejores resultados. Además, se pudo evidenciar que la teoría de juegos puede ser considerada como una novedosa estrategia para la optimización de recursos en una red LAN sobre BPL.

The three-dimensional organization of the genome contributes to its maintenance and regulation. While chromosomal regions associate with nucleolar ribosomal RNA genes (rDNA), the biological significance of rDNA-genome interactions and whether they are dynamically regulated during disease remain unclear. rDNA chromatin exists in multiple inactive and active states and their transition is regulated by the RNA polymerase I transcription factor UBTF. Here, using a MYC-driven lymphoma model, we demonstrate that during malignant progression the rDNA chromatin converts to the open state, which is required for tumor cell survival. Moreover, this rDNA transition co-occurs with a reorganization of rDNA-genome contacts which correlate with gene expression changes at associated loci, impacting gene ontologies including B-cell differentiation, cell growth and metabolism. We propose that UBTF-mediated conversion to open rDNA chromatin during malignant transformation contributes to the regulation of specific gene pathways that regulate growth and differentiation through reformed long-range physical interactions with the rDNA

In Saccharomyces cerevisiae, more than 250 trans-acting factors are involved in the maturation of 40S and 60S ribosomal subunits. The expression of most of these factors is transcriptionally coregulated to ensure correct ribosome production under a wide variety of environmental and intracellular conditions. Here, we identified the essential nucleolar Pol5 protein as a novel trans-acting factor required for the synthesis of 60S ribosomal subunits. Pol5 weakly and/or transiently associates with early to medium pre-60S ribosomal particles. Depletion of and temperature-sensitive mutations in Pol5 result in a deficiency of 60S ribosomal subunits and accumulation of half-mer polysomes. Both processing of 27SB pre-rRNA to mature 25S rRNA and release of pre-60S ribosomal particles from the nucle(ol)us to the cytoplasm are impaired in the Pol5-depleted strain. Moreover, we identified the genes encoding ribosomal proteins uL23 and eL27A as multicopy suppressors of the slow growth of a temperature-sensitive pol5 mutant. These results suggest that Pol5 could function in ensuring the correct folding of 25S rRNA domain III; thus, favoring the correct assembly of these two ribosomal proteins at their respective binding sites into medium pre-60S ribosomal particles. Pol5 is homologous to the human tumor suppressor Myb-binding protein 1A (MYBBP1A). However, expression of MYBBP1A failed to complement the lethal phenotype of a pol5 null mutant strain though interfered with 60S ribosomal subunit biogenesis.

The theory of cooperative n-person games offers a rigorous approach for analysing multilateral real-world agreements, but its practical application is hindered by the exacting data requirements demanded by the fully specified theoretical models. In this article, we demonstrate how the formal analytic can be made more amenable to application. We utilize our approach to model international climate negotiations as an n-person cooperative game, the solution of which allocates carbon reductions across the grand coalition of nations. Using a simplified game to represent the carbon reduction allocation problem, we obtain theoretical solutions using a game-theoretic concept known as the proportional nucleolus. The solution to the game allows us to ideally determine countries’ relative percentage carbon reductions. These theoretical results are compared against actual commitments established in the Paris Agreement of 2015. The paper discusses the implications of the game-theoretic results, including the significant under-commitment of nations such as the United States. More generally, the approach developed herein provides an illustration of how rigorous game-theoretic methods can be adapted to the practical considerations of policy analysis. © 2018 Journal of Environmental Economics and Policy Ltd.

In Saccharomyces cerevisiae, more than 250 trans-acting factors are involved in the maturation of 40S and 60S ribosomal subunits. The expression of most of these factors is transcriptionally coregulated to ensure correct ribosome production under a wide variety of environmental and intracellular conditions. Here, we identified the essential nucleolar Pol5 protein as a novel trans-acting factor required for the synthesis of 60S ribosomal subunits. Pol5 weakly and/or transiently associates with early to medium pre-60S ribosomal particles. Depletion of and temperature-sensitive mutations in Pol5 result in a deficiency of 60S ribosomal subunits and accumulation of half-mer polysomes. Both processing of 27SB pre-rRNA to mature 25S rRNA and release of pre-60S ribosomal particles from the nucle(ol)us to the cytoplasm are impaired in the Pol5-depleted strain. Moreover, we identified the genes encoding ribosomal proteins uL23 and eL27A as multicopy suppressors of the slow growth of a temperature-sensitive pol5 mutant. These results suggest that Pol5 could function in ensuring the correct folding of 25S rRNA domain III; thus, favoring the correct assembly of these two ribosomal proteins at their respective binding sites into medium pre-60S ribosomal particles. Pol5 is homologous to the human tumor suppressor Myb-binding protein 1A (MYBBP1A). However, expression of MYBBP1A failed to complement the lethal phenotype of a pol5 null mutant strain though interfered with 60S ribosomal subunit biogenesis.

The Glutamate Receptor Ionotropic NMDA-Associated Protein 1 (GRINA) belongs to the Lifeguard family and is involved in calcium homeostasis, which governs key processes, such as cell survival or the release of neurotransmitters. GRINA is mainly associated with membranes of the endoplasmic reticulum, Golgi, endosome, and the cell surface, but its presence in the nucleus has not been explained yet. Here we dissect, with the help of different software tools, the potential roles of GRINA in the cell and how they may be altered in diseases, such as schizophrenia or celiac disease. We describe for the first time that the cytoplasmic N-terminal half of GRINA (which spans a Proline-rich domain) contains a potential DNA-binding sequence, in addition to cleavage target sites and probable PY-nuclear localization sequences, that may enable it to be released from the rest of the protein and enter the nucleus under suitable conditions, where it could participate in the transcription, alternative splicing, and mRNA export of a subset of genes likely involved in lipid and sterol synthesis, ribosome biogenesis, or cell cycle progression. To support these findings, we include additional evidence based on an exhaustive review of the literature and our preliminary data of the protein–protein interaction network of GRINA.

The nucleolus is a prominent nuclear condensate that plays a central role in ribosome biogenesis by facilitating the transcription and processing of nascent ribosomal RNA (rRNA). A number of studies have highlighted the active viscoelastic nature of the nucleolus, whose material properties and phase behavior are a consequence of underlying molecular interactions. However, the ways in which the material properties of the nucleolus impact its function in rRNA biogenesis are not understood. Here we utilize the Cry2olig optogenetic system to modulate the viscoelastic properties of the nucleolus. We show that above a threshold concentration of Cry2olig protein, the nucleolus can be gelled into a tightly linked, low mobility meshwork. Gelled nucleoli no longer coalesce and relax into spheres but nonetheless permit continued internal molecular mobility of small proteins. These changes in nucleolar material properties manifest in specific alterations in rRNA processing steps, including a buildup of larger rRNA precursors and a depletion of smaller rRNA precursors. We propose that the flux of processed rRNA may be actively tuned by the cell through modulating nucleolar material properties, which suggests the potential of materials-based approaches for therapeutic intervention in ribosomopathies., SCOPUS: ar.j, info:eu-repo/semantics/published

Zero-gravity is an environmental challenge unknown to organisms throughout evolution on Earth. Nevertheless, plants are sensitive to altered gravity, as exemplified by changes in meristematic cell proliferation and growth. We found that synchronized Arabidopsis cultured cells exposed to simulated microgravity showed a shortened cell cycle, caused by a shorter G2/M phase and a slightly longer G1 phase. The analysis of selected marker genes and proteins by qPCR and flow cytometry in synchronic G1 and G2 subpopulations indicated changes in gene expression of core cell cycle regulators and chromatin-modifying factors, confirming that microgravity induced misregulation of G2/M and G1/S checkpoints and chromatin remodeling. Changes in chromatin-based regulation included higher DNA methylation and lower histone acetylation, increased chromatin condensation and overall depletion of nuclear transcription. Estimation of ribosome biogenesis rate using nucleolar parameters and selected nucleolar genes and proteins indicated reduced nucleolar activity under simulated microgravity, especially at G2/M. These results expand our knowledge of how meristematic cells are affected by real and simulated microgravity. Counteracting this cellular stress is necessary for plant culture in space exploration.