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"Obesity affects the population worldwide. A hypercaloric diet associated with a sedentary life, stress, and genetic background, triggers various metabolic disorders, such as metabolic syndrome, diabetes mellitus, cancer, cardiovascular diseases, non-alcoholic fatty liver disease, and cognitive impairment. A healthy diet correlated with physical activity, not smoking, and moderate alcohol consumption reduces the risk of developing metabolic diseases. The Mediterranean diet contains antioxidants, fiber, polyunsaturated fats, and compounds with anti-inflammatory, anti-oxidant, anti-cancer, and anti-obesity properties. In a wide variety of species including humans, the reduction of calories between 20-40% significantly improves health, increaseslongevity, and delaysthe development of various pathologies. The main aim of this review is to present the comparative effects of the Mediterranean diet versus the hypercaloric diet on insulin sensitivity. "

The RIMFAX ground penetrating radar instrument on board the Mars 2020 Perseverance Rover has been continuously sounding the subsurface alongthe Rover traverse. In the data of the first 379 mission days on the Jezero Crater Floor we are able to identify hyperbolic patterns, likely caused by buried scatterers such as boulders or cavities in the upper 5 m of the subsurface. We present the first detailed estimates of radar wave propagation velocity by matching theoretical traveltime hyperbolas to the patterns generated by scatterers. The average dielectric permittivities are derived from these velocities and, consequently, the bulk rock densities for material above the scattering source. Simultaneously, we investigate the surface reflectivity to retrieve permittivity and density of the uppermost centimeter. Finally, we assess the radar attenuation by a constant-Q approach.The results are consistent with a solid rock, mafic interpretation of the Jezero Crater subsurface. The talk is based on the respective two most recent publications as well as work in progress.

Superabsorbent hydrogels continue to be very important materials due to their applications in several technologies. Unfortunately, most superabsorbent hydrogels currently on the market are acrylate-based products that are non-biodegradable, and, most importantly, some concerns exist about their toxicity for use in agriculture. This study aimed at synthesizing and characterizing biocompatible superabsorbent hydrogel derived from lemon juice. The process involved polymerizing lemon juice (LJ) with glycerol (G) monomers to form polymeric material (HLG-1). HLG-1 was then converted to HLG-2 by crosslinking with maleic acid. Characterization of the hydrogels was done using Fourier transform infrared (FT-IR) spectroscopy, scanning electron microscope (SEM), and X-ray diffraction (XRD). The synthesis conditions producing optimal swelling capacity were studied by varying contact time and dosage of both lemon juice and the cross-linker. The FT-IR results showed a peak at 1528 cm-1 and 1591.34 cm-1 associated with –COO- stretching indicating successful polymerization and crosslinking processes. XRD analysis showed conversion from amorphous to crystalline phases upon crosslinking. SEM micrographs showed clear pores with large surface area in HLG-2 compared with the rigid and constricted surface of HLG-1 hydrogel. A maximum swelling capacity of 910% was obtained upon synthesizing hydrogel HLG-2 with lemon juice, glycerol, and maleic acid of a volume ratio of 5.4: 3.75: 3.75 respectively. Crosslinking the hydrogel with maleic acid was found to improve the water absorption capacity of the hydrogel. The superabsorbent hydrogel with such high swelling and water absorption ability has the potential of being applied in arid and semi-arid regions to boost agricultural production.

The assembly of membraneless compartments by phase separation has recently been recognized as a mechanism for spatial and temporal organization of biomolecules within the cell. The functions of such mesoscale assemblies, termed biomolecular condensates, depend on networks of multivalent interactions between proteins, their structured and disordered domains, and commonly also include nucleic acids. Cryo-electron tomography is an ideal tool to investigate the three-dimensional architecture of such pleomorphic interaction networks at nanometer resolution and thus form inferences about function. However, preparation of suitable cryo-electron microscopy samples of condensates may be prone to protein denaturation, low retention of material on the sample carrier, and contamination associated with cryo-sample preparation and transfers. Here, we describe a series of protocols designed to obtain high-quality cryo-electron tomography data of biomolecular condensates reconstituted in vitro. These include critical screening by light microscopy, cryo-fixation by plunge freezing, sample loading into an electron microscope operated at liquid nitrogen temperature, data collection, processing of the data into three-dimensional tomograms, and their interpretation.

The assembly of membraneless compartments by phase separation has recently been recognized as a mechanism for spatial and temporal organization of biomolecules within the cell. The functions of such mesoscale assemblies, termed biomolecular condensates, depend on networks of multivalent interactions between proteins, their structured and disordered domains, and commonly also include nucleic acids. Cryo-electron tomography is an ideal tool to investigate the three-dimensional architecture of such pleomorphic interaction networks at nanometer resolution and thus form inferences about function. However, preparation of suitable cryo-electron microscopy samples of condensates may be prone to protein denaturation, low retention of material on the sample carrier, and contamination associated with cryo-sample preparation and transfers. Here, we describe a series of protocols designed to obtain high-quality cryo-electron tomography data of biomolecular condensates reconstituted in vitro. These include critical screening by light microscopy, cryo-fixation by plunge freezing, sample loading into an electron microscope operated at liquid nitrogen temperature, data collection, processing of the data into three-dimensional tomograms, and their interpretation.

Heat shock promotes the assembly of translation factors into condensates to facilitate the production of stress-protective proteins. How translation factors detect heat and assemble into condensates is not well understood. Here, we investigate heat-induced condensate assembly by the translation factor Ded1p from five different fungi, including Ded1p fromSaccharomyces cerevisiae. Using targeted mutagenesis andin vitroreconstitution biochemistry, we find that heat-induced Ded1p assembly is driven by a conformational rearrangement of the folded helicase domain. This rearrangement determines the assembly temperature and the assembly of Ded1p into nanometer-sized particles, while the flanking intrinsically disordered regions engage in intermolecular interactions to promote assembly into micron-sized condensates. Using protein engineering, we identify six amino acid substitutions that determine most of the thermostability of a thermophilic Ded1p ortholog, thereby providing a molecular understanding underlying the adaptation of the Ded1p assembly temperature to the specific growth temperature of the species. We conclude that heat-induced assembly of Ded1p into translation factor condensates is regulated by a complex interplay of the structured domain and intrinsically disordered regions which is subject to evolutionary tuning.

Newer mobile applications are increasingly being defined using Internet Protocol, resulting in increased use of Internet Protocol and subsequent upsurge of smartphones. However, many communication service provider core networks continue to use classical routing protocols and single controller-based networks if deployed. Controller-based networks built on the foundation of software-defined networks include centralization and separation of control plane and data plane, which can address the challenges experienced with the classical routing protocols. When single controllers are used, they tend to get overloaded with traffic. The ability to use multi-controller-based network architecture to improve quality of service in the mobile IP core network is still an open issue. This paper presents a performance evaluation of multi-controller-based network architecture, running OpenFlow and Open Shortest Path First protocol. The long-term evolution simulated network architecture is created using well-known network simulator Objective Modular Network Testbed running OpenFlow and simuLTE add-on. We test and analyze data traffic for Packet data ratio and Jitter and their associated effects on a multi-controller-based network running OpenFlow versus OSPF on a mobile core network. The experiment created two topologies; multi controller-based and Open Shortest path first network. Video and ping traffic is tested by the generation of traffic from User Equipment to the network-based server in the data center and back, and traffic metrics recorded on an inbuilt integrated development environment. The simulation setup consisted of an OpenFlow controller, HyperFlow algorithm, OpenFlow switches, and Open Shortest Path First routers. The multi-controller-based network improved Jitter by 10 ms. The Open Shortest Path first showed packet data ratio values of 89% gain while the controller-based network registered a value of 86%. A standard deviation test revealed 0.7%, which shows that the difference is not significant when testing for Packet data ratio. We provided insight into the performance of multi-controller-based architecture and Open Shortest Path First protocol in the communication service provider's core network.

African Divine Church (ADC) is an African Instituted Church affiliated to the Organisation of the African Instituted Churches, which shapes African Instituted Churches’ (AICs’) theology through theological education. ADC has both lived, and sung-narrated theology and it originated from the Pentecostal Assemblies of Canada, hence Pentecostal with experiential undertones. Selected leaders of ADC in Vihiga County contributed to the study. The de-emphasised place of Christ was investigated to establish his place in ADC’s Trinitarian and missional Theology. The study articulated theological principles regarding Trinitarian ecclesiology, to assist the ADC in reworking its Christology. The historical approach of the early church, medieval, reformation, and modern periods regarding the place of Christ as the second person of the Triune God were investigated, to formulate guidelines for a paradigm shift in understanding the place of Christ in Trinitarian ecclesiology and the missional community that ADC may use in reworking its Trinitarian Mission. The study employed a mixed-method that followed three steps, namely interviews, questionnaires and observation. The study population was determined through purposive sampling. A coding method safeguarded the identity and confidentiality of participants. The results were classified as historical, practical and theological. ADC derived its name from John 15:1–17 through meditation, vision and revelation. The colours red, white and green symbolise its mission and growth, while its experiential and charismatic liturgy was derived from 1 Samuel 6:1–4, with evidence of hermeneutical deficiency. Theological findings included the undeveloped relationship between God and ecclesial mission, emphasis on the Holy Spirit, healing, and prophecy with a lessened place of Christ in the Trinitarian mission, as well as the blend of ancestorology with Christology that threatens the hypostatic Christology and Trinitarian economy. The study implied a call for reworking hermeneutics, Trinitarian theology, and reconstructing the relationship between God’s mission and that of the Church and a reworked Christology, distinctive from ancestorology.Contribution: African Divine Church’s narrative theology is exposed especially the Trinitarian mission to a constructive critique and further study. It has advanced mixed-method in progressing narrative inquiry as an alternative methodology.

The Radar Imager for Mars Subsurface Experiment instrument has conducted the first rover-mounted ground-penetrating radar survey of the Martian subsurface. A continuous radar image acquired over the Perseverance rover’s initial ~3-kilometer traverse reveals electromagnetic properties and bedrock stratigraphy of the Jezero crater floor to depths of ~15 meters below the surface. The radar image reveals the presence of ubiquitous strongly reflecting layered sequences that dip downward at angles of up to 15 degrees from horizontal in directions normal to the curvilinear boundary of and away from the exposed section of the Séitah formation. The observed slopes, thicknesses, and internal morphology of the inclined stratigraphic sections can be interpreted either as magmatic layering formed in a differentiated igneous body or as sedimentary layering commonly formed in aqueous environments on Earth. The discovery of buried structures on the Jezero crater floor is potentially compatible with a history of igneous activity and a history of multiple aqueous episodes.

CAPRIN1 is a ubiquitously expressed protein, abundant in the brain, where it regulates the transport and translation of mRNAs of genes involved in synaptic plasticity. Here we describe two unrelated children, who developed early-onset ataxia, dysarthria, cognitive decline and muscular weakness. Trio exome sequencing unraveled the identical de novo c.1535C>T (p.Pro512Leu) missense variant in CAPRIN1, affecting a highly conserved residue. In silico analyses predict an increased aggregation propensity of the mutated protein. Indeed, overexpressed CAPRIN1P512L forms insoluble ubiquitinated aggregates, sequestrating proteins associated with neurodegenerative disorders (ATXN2, GEMIN5, SNRNP200 and SNCA). Moreover, the CAPRIN1P512L mutation in isogenic iPSC-derived cortical neurons causes reduced neuronal activity and altered stress granule dynamics. Furthermore, nano-differential scanning fluorimetry reveals that CAPRIN1P512L aggregation is strongly enhanced by RNA in vitro. These findings associate the gain-of-function Pro512Leu mutation to early-onset ataxia and neurodegeneration, unveiling a critical residue of CAPRIN1 and a key role of RNA-protein interactions.

Macromolecular phase separation is thought to be one of the processes that drives the formation of membraneless biomolecular condensates in cells. The dynamics of phase separation are thought to follow the tenets of classical nucleation theory, and, therefore, subsaturated solutions should be devoid of clusters with more than a few molecules. We tested this prediction using in vitro biophysical studies to characterize subsaturated solutions of phase-separating RNA-binding proteins with intrinsically disordered prion-like domains and RNA-binding domains. Surprisingly, and in direct contradiction to expectations from classical nucleation theory, we find that subsaturated solutions are characterized by the presence of heterogeneous distributions of clusters. The distributions of cluster sizes, which are dominated by small species, shift continuously toward larger sizes as protein concentrations increase and approach the saturation concentration. As a result, many of the clusters encompass tens to hundreds of molecules, while less than 1% of the solutions are mesoscale species that are several hundred nanometers in diameter. We find that cluster formation in subsaturated solutions and phase separation in supersaturated solutions are strongly coupled via sequence-encoded interactions. We also find that cluster formation and phase separation can be decoupled using solutes as well as specific sets of mutations. Our findings, which are concordant with predictions for associative polymers, implicate an interplay between networks of sequence-specific and solubility-determining interactions that, respectively, govern cluster formation in subsaturated solutions and the saturation concentrations above which phase separation occurs.

CAPRIN1 is a ubiquitously expressed protein, abundant in the brain, where it regulates the transport and translation of mRNAs of genes involved in synaptic plasticity. Here we describe two unrelated children, who developed early-onset ataxia, dysarthria, cognitive decline and muscle weakness. Trio exome sequencing unraveled the identical de novo c.1535C T (p.Pro512Leu) missense variant in CAPRIN1, affecting a highly conserved residue. In silico analyses predict an increased aggregation propensity of the mutated protein. Indeed, overexpressed CAPRIN1

Liquid–liquid phase separation of proteins underpins the formation of membraneless compartments in living cells. Elucidating the molecular driving forces underlying protein phase transitions is therefore a key objective for understanding biological function and malfunction. Here we show that cellular proteins, which form condensates at low salt concentrations, including FUS, TDP-43, Brd4, Sox2, and Annexin A11, can reenter a phase-separated regime at high salt concentrations. By bringing together experiments and simulations, we demonstrate that this reentrant phase transition in the high-salt regime is driven by hydrophobic and non-ionic interactions, and is mechanistically distinct from the low-salt regime, where condensates are additionally stabilized by electrostatic forces. Our work thus sheds light on the cooperation of hydrophobic and non-ionic interactions as general driving forces in the condensation process, with important implications for aberrant function, druggability, and material properties of biomolecular condensates., Elucidating the molecular driving forces underlying liquid–liquid phase separation is a key objective for understanding biological function and malfunction. Here the authors show that a wide range of cellular proteins, including FUS, TDP-43, Brd4, Sox2, and Annexin A11, which form condensates at low salt concentrations, can reenter a phase-separated regime at high salt concentrations.

Macromolecular phase separation is thought to be one of the processes that drives the formation of membraneless biomolecular condensates in cells. The dynamics of phase separation, especially at low endogenous concentrations found in cells, are thought to follow the tenets of classical nucleation theory describing a sharp transition between a dense phase and a dilute phase characterized by dispersed monomers. Here, we usedin vitrobiophysical studies to study subsaturated solutions of phase separating RNA binding proteins with intrinsically disordered prion like domains (PLDs) and RNA binding domains (RBDs). Surprisingly, we find that subsaturated solutions are characterized by heterogeneous distributions of clusters comprising tens to hundreds of molecules. These clusters also include low abundance mesoscale species that are several hundreds of nanometers in diameter. Our results show that cluster formation in subsaturated solutions and phase separation in supersaturated solutions are strongly coupled via sequence-encoded interactions. Interestingly, however, cluster formation and phase separation can be decoupled from one another using solutes that impact the solubilities of phase separating proteins. They can also be decoupled by specific types of mutations. Overall, our findings implicate the presence of distinct, sequence-specific energy scales that contribute to the overall phase behaviors of RNA binding proteins. We discuss our findings in the context of theories of associative polymers.Significance StatementMembraneless biomolecular condensates are molecular communities with distinct compositional preferences and functions. Considerable attention has focused on phase separation as the process that gives rise to condensates. Here, we show that subsaturated solutions of RNA binding proteins form heterogeneous distributions of clusters in subsaturated solutions. The formation of clusters in subsaturated solutions and condensates in supersaturated solution are coupled through sequence-specific interactions. Given the low endogenous concentrations of phase separating proteins, our findings suggest that clusters in subsaturated conditions might be of functional relevance in cells.

The purpose of this study was to ascertain the extent of current e-learning uptake in Tanzanian universities. The quantitative approach involving survey design was adopted in the collection of data. Data were collected through a questionnaire survey of 400 respondents, with a rate of return 85.5%. The average reliability of variables 0.949 was determined using Cronbach's Alpha. Fuzzy Logic model and t-test were adopted for data analysis. The findings revealed that the average extent of current e-learning uptake among students and academicians were less than half of threshold amounting to 50% (i.e. level of awareness was 16%, availability was 20.6%, accessibility was 17%, attitude was 15% as variables used). There was no statistically significant difference in e-learning uptake among students and academicians as the value of p > 0.05. The findings of this study established a base ground and guidelines to inform the e-learning stakeholders and policymakers to find and establish suitable policy as well as mechanism to adopt and encourage sustainable use of e-learning systems for life-long teaching and learning. The originality of this study is based on the addition of new variables and methodologies employed as empirical evidence based on the extent of e-learning uptake in Tanzanian universities. Keyword: ICTs, e-learning uptake, fuzzy logic model, students, academicians

Event Horizon Telescope (EHT) observations at 230 GHz have now imaged polarized emission around the supermassive black hole in M87 on event-horizon scales. This polarized synchrotron radiation probes the structure of magnetic fields and the plasma properties near the black hole. Here we compare the resolved polarization structure observed by the EHT, along with simultaneous unresolved observations with the Atacama Large Millimeter/submillimeter Array, to expectations from theoretical models. The low fractional linear polarization in the resolved image suggests that the polarization is scrambled on scales smaller than the EHT beam, which we attribute to Faraday rotation internal to the emission region. We estimate the average density n e ∼ 104-7 cm-3, magnetic field strength B ∼ 1-30 G, and electron temperature T e ∼ (1-12) 1010 K of the radiating plasma in a simple one-zone emission model. We show that the net azimuthal linear polarization pattern may result from organized, poloidal magnetic fields in the emission region. In a quantitative comparison with a large library of simulated polarimetric images from general relativistic magnetohydrodynamic (GRMHD) simulations, we identify a subset of physical models that can explain critical features of the polarimetric EHT observations while producing a relativistic jet of sufficient power. The consistent GRMHD models are all of magnetically arrested accretion disks, where near-horizon magnetic fields are dynamically important. We use the models to infer a mass accretion rate onto the black hole in M87 of (3-20) 10-4 M o˙ yr-1. © 2021. The Author(s). Published by the American Astronomical Society..

In 2017, the Event Horizon Telescope (EHT) Collaboration succeeded in capturing the first direct image of the center of the M87 galaxy. The asymmetric ring morphology and size are consistent with theoretical expectations for a weakly accreting supermassive black hole of mass ∼6.5 × 109 M o˙. The EHTC also partnered with several international facilities in space and on the ground, to arrange an extensive, quasi-simultaneous multi-wavelength campaign. This Letter presents the results and analysis of this campaign, as well as the multi-wavelength data as a legacy data repository. We captured M87 in a historically low state, and the core flux dominates over HST-1 at high energies, making it possible to combine core flux constraints with the more spatially precise very long baseline interferometry data. We present the most complete simultaneous multi-wavelength spectrum of the active nucleus to date, and discuss the complexity and caveats of combining data from different spatial scales into one broadband spectrum. We apply two heuristic, isotropic leptonic single-zone models to provide insight into the basic source properties, but conclude that a structured jet is necessary to explain M87's spectrum. We can exclude that the simultaneous γ-ray emission is produced via inverse Compton emission in the same region producing the EHT mm-band emission, and further conclude that the γ-rays can only be produced in the inner jets (inward of HST-1) if there are strongly particle-dominated regions. Direct synchrotron emission from accelerated protons and secondaries cannot yet be excluded. © 2021. The Author(s). Published by the American Astronomical Society..

Our understanding of strong gravity near supermassive compact objects has recently improved thanks to the measurements made by the Event Horizon Telescope (EHT). We use here the M87∗ shadow size to infer constraints on the physical charges of a large variety of nonrotating or rotating black holes. For example, we show that the quality of the measurements is already sufficient to rule out that M87∗ is a highly charged dilaton black hole. Similarly, when considering black holes with two physical and independent charges, we are able to exclude considerable regions of the space of parameters for the doubly-charged dilaton and the Sen black holes. © 2021 authors. Published by the American Physical Society.

Very-long-baseline interferometry (VLBI) observations of active galactic nuclei at millimetre wavelengths have the power to reveal the launching and initial collimation region of extragalactic radio jets, down to 10–100 gravitational radii (rg ≡ GM/c2) scales in nearby sources1. Centaurus A is the closest radio-loud source to Earth2. It bridges the gap in mass and accretion rate between the supermassive black holes (SMBHs) in Messier 87 and our Galactic Centre. A large southern declination of −43° has, however, prevented VLBI imaging of Centaurus A below a wavelength of 1 cm thus far. Here we show the millimetre VLBI image of the source, which we obtained with the Event Horizon Telescope at 228 GHz. Compared with previous observations3, we image the jet of Centaurus A at a tenfold higher frequency and sixteen times sharper resolution and thereby probe sub-lightday structures. We reveal a highly collimated, asymmetrically edge-brightened jet as well as the fainter counterjet. We find that the source structure of Centaurus A resembles the jet in Messier 87 on ~500 rg scales remarkably well. Furthermore, we identify the location of Centaurus A’s SMBH with respect to its resolved jet core at a wavelength of 1.3 mm and conclude that the source’s event horizon shadow4 should be visible at terahertz frequencies. This location further supports the universal scale invariance of black holes over a wide range of masses5,6. © 2021, The Author(s).

We present the results from a full polarization study carried out with the Atacama Large Millimeter/submillimeter Array (ALMA) during the first Very Long Baseline Interferometry (VLBI) campaign, which was conducted in 2017 April in the λ3 mm and λ1.3 mm bands, in concert with the Global mm-VLBI Array (GMVA) and the Event Horizon Telescope (EHT), respectively. We determine the polarization and Faraday properties of all VLBI targets, including Sgr A∗, M87, and a dozen radio-loud active galactic nuclei (AGNs), in the two bands at several epochs in a time window of 10 days. We detect high linear polarization fractions (2%-15%) and large rotation measures (RM > 103.3-105.5 rad m-2), confirming the trends of previous AGN studies at millimeter wavelengths. We find that blazars are more strongly polarized than other AGNs in the sample, while exhibiting (on average) order-of-magnitude lower RM values, consistent with the AGN viewing angle unification scheme. For Sgr A∗ we report a mean RM of (-4.2 0.3) 105 rad m-2 at 1.3 mm, consistent with measurements over the past decade and, for the first time, an RM of (-2.1 0.1) 105 rad m-2 at 3 mm, suggesting that about half of the Faraday rotation at 1.3 mm may occur between the 3 mm photosphere and the 1.3 mm source. We also report the first unambiguous measurement of RM toward the M87 nucleus at millimeter wavelengths, which undergoes significant changes in magnitude and sign reversals on a one year timescale, spanning the range from -1.2 to 0.3 105 rad m-2 at 3 mm and -4.1 to 1.5 105 rad m-2 at 1.3 mm. Given this time variability, we argue that, unlike the case of Sgr A∗, the RM in M87 does not provide an accurate estimate of the mass accretion rate onto the black hole. We put forward a two-component model, comprised of a variable compact region and a static extended region, that can simultaneously explain the polarimetric properties observed by both the EHT (on horizon scales) and ALMA (which observes the combined emission from both components). These measurements provide critical constraints for the calibration, analysis, and interpretation of simultaneously obtained VLBI data with the EHT and GMVA. © 2021. The American Astronomical Society.

In stressed cells, proteins and RNA molecules cluster together to form stress granules. It emerges that the small protein modifier ubiquitin is needed to disassemble stress granules in recovering cells. Disassembly of stress granules is enabled by ubiquitination.

The assembly of intracellular proteins into biomolecular condensates via liquid-liquid phase separation (LLPS) has emerged as a fundamental process underlying the organisation and regulation of cellular space and function. Physicochemical characterisation of the parameters that control and modulate phase separation is therefore essential for an improved understanding of protein phase behaviour, not least to inform efforts for the therapeutic modulation of LLPS phenomena. Given the rapidly increasing number of biologically and disease-relevant condensate systems, experimental techniques that enable high-throughput analysis of protein phase behaviour are required. Here, we present a droplet microfluidic platform, termed PhaseScan, for the rapid generation of protein phase diagrams, a fundamental measure with which to characterise protein phase behaviour in chemical space. Using this platform, we demonstrate characterisation of the phase behaviour of a pathologically relevant mutant of the protein fused in sarcoma (FUS) in a highly parallelised manner, with significantly improved assay throughout and reduced sample consumption with respect to conventional experiments. We find that the phase boundary at which FUS transitions from a one-phase to a two-phase state is modulated by the small molecule 1,6-hexanediol, and estimate the free-energy landscape of this system using Flory-Huggins theory. Our study thus provides a basis for the rapid acquisition of phase diagrams through the application of microdroplet techniques and paves the way for a wide range of applications, enabling swift characterisation of the effect of environmental conditions and coacervate species on the thermodynamics of phase separation.

The invention relates to a method for producing an amine by reacting an alcohol, aldehyde and/or ketone with hydrogen and nitrogen-containing compound in the presence of a silver and/or ruthenium promoted cobalt catalyst. The nitrogen-containing compound is selected from ammonia, primary amines and secondary amines.; L'invention concerne un procédé de production d'une amine par réaction d'un alcool, d'un aldéhyde et/ou d'une cétone avec de l'hydrogène et un composé contenant de l'azote en présence d'un catalyseur au cobalt activé par l'argent et/ou le ruthénium. Le composé contenant de l'azote est choisi parmi l'ammoniac, les amines primaires et les amines secondaires.

Cells exposed to starvation have to adjust their metabolism to conserve energy and protect themselves. Protein synthesis is one of the major energy-consuming processes and as such has to be tightly controlled. Many mechanistic details about how starved cells regulate the process of protein synthesis are still unknown. Here, we report that the essential translation initiation factor eIF2B forms filaments in starved budding yeast cells. We demonstrate that filamentation is triggered by starvation-induced acidification of the cytosol, which is caused by an influx of protons from the extracellular environment. We show that filament assembly by eIF2B is necessary for rapid and efficient downregulation of translation. Importantly, this mechanism does not require the kinase Gcn2. Furthermore, analysis of site-specific variants suggests that eIF2B assembly results in enzymatically inactive filaments that promote stress survival and fast recovery of cells from starvation. We propose that translation regulation through filament assembly is an efficient mechanism that allows yeast cells to adapt to fluctuating environments., Summary: Filament formation by eIF2B is an adaptive mechanism that allows for efficient shutdown of translation in starved yeast cells.

BACKGROUND: It is hypothesised that cotargeting the androgen receptor (AR) and paracrine androgen biosynthesis with enzalutamide and abiraterone acetate in metastatic castration-resistant prostate cancer (mCRPC) will dissipate adaptive feedback loops observed with either agent alone. OBJECTIVE: To assess the safety, efficacy, androgen signalling/metabolome, and drug-drug interactions (DDIs) of enzalutamide with abiraterone acetate in progressive bone mCRPC (bmCRPC). DESIGN, SETTING, AND PARTICIPANTS: This open-label, single-centre interventional study was conducted in bmCRPC patients. INTERVENTION: Enzalutamide 160mg and abiraterone acetate 1000mg once daily; prednisone 5mg twice daily. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Adverse events (AEs), prostate-specific antigen (PSA) response, progression-free survival (PFS), tumour biomarker/metabolite expression, and Cmin plasma concentrations were evaluated. RESULTS AND LIMITATIONS: Sixty patients were enrolled. Common AEs independent of grade/causality included fatigue (72%), hyperglycaemia (67%), alkaline phosphatase (ALP) elevation (53%), and hot flush (43%). Grade 3 AEs included hypertension (17%), alanine aminotransferase elevation (12%), ALP elevation (5%), and arthralgia (5%). No treatment-related grade 4 AEs or deaths were reported. Median treatment-discontinuation time was 312d (95% confidence interval [CI] 196.0-483.0). Maximal PSA decline ≥50% and ≥90% occurred in 46 (77%) and 29 (48%) patients, respectively. Median PFS was 251d (95% CI 147-337). At week 9, median tumour microenvironment androgens, precursors, and nuclear AR expression decreased (p

Realistic synthetic observations of theoretical source models are essential for our understanding of real observational data. In using synthetic data, one can verify the extent to which source parameters can be recovered and evaluate how various data corruption effects can be calibrated. These studies are important when proposing observations of new sources, in the characterization of the capabilities of new or upgraded instruments, and when verifying model-based theoretical predictions in a comparison with observational data. We present the SYnthetic Measurement creator for long Baseline Arrays (SYMBA), a novel synthetic data generation pipeline for Very Long Baseline Interferometry (VLBI) observations. SYMBA takes into account several realistic atmospheric, instrumental, and calibration effects. We used SYMBA to create synthetic observations for the Event Horizon Telescope (EHT), a mm VLBI array, which has recently captured the first image of a black hole shadow. After testing SYMBA with simple source and corruption models, we study the importance of including all corruption and calibration effects. Based on two example general relativistic magnetohydrodynamics (GRMHD) model images of M87, we performed case studies to assess the attainable image quality with the current and future EHT array for different weather conditions. The results show that the effects of atmospheric and instrumental corruptions on the measured visibilities are significant. Despite these effects, we demonstrate how the overall structure of the input models can be recovered robustly after performing calibration steps. With the planned addition of new stations to the EHT array, images could be reconstructed with higher angular resolution and dynamic range. In our case study, these improvements allowed for a distinction between a thermal and a non-thermal GRMHD model based on salient features in reconstructed images., Comment: 20 pages, 15 figures, accepted for publication in A&A

The objective of this study was to establish the influence of strategy implementation on sustainability of community based tourism in the coast region of Kenya. The target population was 193 members of community based tourism units in coastal Kenya as a representative sample. Stratified random sampling method was used to classify the thirteen community groups into a single stratum. The study used structured questionnaires and documented literature as the main tools for both primary and secondary data collection. The findings indicated that strategy implementation had a positive relationship with sustainability of community based tourism. The variables were found to be normally distributed and hence no significant differences at 95% confidence level. The study established a strong linear relationship existed between strategy implementation and sustainability of community based tourism. Information from the study confirms enterprises require strategy implementation for continued sustainability of their enterprises.

How cells adapt to varying environmental conditions is largely unknown. Here, we show that, in budding yeast, the RNA-binding and stress granule protein Pub1 has an intrinsic property to form condensates upon starvation or heat stress and that condensate formation is associated with cell-cycle arrest. Release from arrest coincides with condensate dissolution, which takes minutes (starvation) or hours (heat shock). In vitro reconstitution reveals that the different dissolution rates of starvation- and heat-induced condensates are due to their different material properties: starvation-induced Pub1 condensates form by liquid-liquid demixing and subsequently convert into reversible gel-like particles; heat-induced condensates are more solid-like and require chaperones for disaggregation. Our data suggest that different physiological stresses, as well as stress durations and intensities, induce condensates with distinct physical properties and thereby define different modes of stress adaptation and rates of recovery., Cell Reports, 23 (11), ISSN:2666-3864, ISSN:2211-1247

Purpose: The purpose of the study was to determine the environmental challenges facing health sector NGOs in Kenya and the strategic responses that health sector NGOs adopt to come with challenges posed by the external environment.Methodology: The research design was descriptive survey study in nature since it focused on all NGOs working in health sector in Nairobi. The target population was 1065 NGOs. A sample of 41 NGOs was selected using the recommended formula for calculating sample size given proportions as recommended by Israel. The respondents were the managing directors of the NGOs. Both qualitative and quantitative data was collected using a questionnaire that consisted of both open ended and close ended questions. The data was analyzed in terms of descriptive statistics like frequencies, means and percentages. The findings were presented in form of tables.Results: The study found that competitive rivarly, threats of new entrants , bargaining power of suppliers, bargaining power of clients, threat of substitutes, changes in law and advancement in information technology contribute to environmental challenges of the organization.Unique contribution to theory, practice and policy: It is recommended that the NGOs train their employees on strategic management and the identification of the environmental factors that challenge NGOs. Specifically, the management of NGOs should be trained on PESTEL and SWOT analysis to facilitate proper environmental scanning.

Objective The present study reviews a series of patients who underwent thyroid surgery in Eldoret, Kenya, to demonstrate the feasibility of conducting long-term (>1 year) outcomes research in a resource-limited setting, impact on the quality of life of the recipient population, and inform future humanitarian collaborations. Study Design Case series with chart review. Setting Tertiary public referral hospital in Eldoret, Kenya. Subjects and Methods Twenty-one patients were enrolled during the study period. A retrospective chart review was performed for all adult patients who underwent thyroid surgery during humanitarian trips (2010-2015). Patients were contacted by mobile telephone. Medical history and physical examination, including laryngoscopy, were performed, and the SF-36 was administered (a quality-of-life questionnaire). Laboratory measurements of thyroid function and neck ultrasound were obtained. Results The mean follow-up was 33.6 ± 20.2 months after surgery: 37.5% of subtotal thyroidectomy patients and 15.4% of lobectomy patients were hypothyroid postoperatively according to serologic studies. There were no cases of goiter recurrence or malignancy. All patients reported postoperative symptomatic improvement and collectively showed positive pre- and postoperative score differences on the SF-36. Conclusion Although limited by a small sample size and the retrospective nature, our study demonstrates the feasibility of long-term surgical and quality-of-life outcomes research in a resource-limited setting. The low complication rates suggest minimal adverse effects of performing surgery in this context. Despite a considerable rate of postoperative hypothyroidism, it is in accordance with prior studies and emphasizes the need for individualized, longitudinal, and multidisciplinary care. Quality-of-life score improvements suggest benefit to the recipient population.

The acquisition of practical and applied skills as well as the basic scientific knowledge that would facilitate efficient occupational training requires good manipulation of skills oriented instructional facilities in a conducive learning situation. Thus, the provision and effective utilization of functional instructional materials are essential for adequate practical training in vocational education for the development of employable skills in the students. However, there are many factors that hinder effective production and utilization of instructional materials in vocational education. Hence appropriate recommendations are made to remedy these ugly situations facing our educational system.

This article describes the development of curriculum as it relates to vocational education in Nigeria Upper Basic Education Curriculum. The definition of Curriculum development was highlighted to reflect contemporary concepts of curriculum integration. Curriculum development was stressed to include the rudiments necessary in its stages of development. Instructional materials production and utilization in broad terms of audio-visual aids were also treated. The work however drew up the role and relevant position of evaluation in curriculum development. The paper concluded with the need to have the development, production and utilization of instructional materials as a communication channel in the teaching-learning processes of vocational education.

Members of the MOAG/SERF protein family have been attributed a neuropathologic significance because of their ability to enhance the proteotoxic polymerization of amyloidogenic proteins such as alpha-Synuclein (aSyn). However, the cellular function remains unknown. Here, we identify SERF1a as an RNA-organizing protein i) with no structural homology to canonical RNA-binding proteins, ii) with an RNA-chaperoning activity which favours the incorporation of RNA into nucleoli and liquid-like artificial RNA-organelles, and iii) with a high degree of conformational disorder in the RNA-bound state. We demonstrate that this type of structural fuzziness determines an undifferentiated interaction of SERF1a with aSyn and RNA. Both molecules bind to one identical, positively charged site of SERF1a by an analogous electrostatic binding mode, with similar binding affinities, and without any observable disorder-to-order transition. The absence of primary or secondary structure discriminants results in SERF1a being unable to distinguish between nucleic acid and amyloidogenic protein, leading the pro-amyloid aSyn:SERF1a interaction to prevail under conditions of cellular stress. We suggest that structural fuzziness of SERF1a accounts for an adverse gain-of-interaction which favours toxic binding to aSyn at the expense of non-toxic RNA binding, therefore promoting a functionally distorted and pathogenic process. Our results provide a direct link between structural disorder, amyloid aggregation and the malfunctioning of RNA-associated cellular processes, three hallmarks of neurodegenerative diseases.

Neoadjuvant therapy with abiraterone acetate + prednisone (AAP) + luteinising hormone-releasing hormone agonist (LHRHa) induced significant cytoreduction associated with a lack of recurrence versus LHRHa, though staging did not differ significantly. This first randomised AAP study in localised high-risk prostate cancer explores response heterogeneity and associated molecular effects to guide further research. © 2019 Optimal therapeutic strategy remains an unmet need in localised high-risk prostate cancer (LHRPC). Androgen biosynthesis inhibition in the preoperative setting may improve outcomes. In this single-centre randomised trial, we looked at therapy outcomes of preoperative treatment with abiraterone acetate + prednisone (AAP) + luteinising hormone-releasing hormone agonist (LHRHa) or LHRHa alone followed by radical prostatectomy in 65 men. We did not see a significant difference of organ-confined carcinoma (p = 0.27). However, tumour volume measures were significantly lower for AAP + LHRHa treatment (p ≤ 0.001). Of note, lower tumour epithelium volume correlated with improved biochemical recurrence-free survival at ≥4-yr follow-up (p = 0.0014). Tumours pretreated with AAP + LHRHa had lower proliferation and androgen signalling expression than LHRHa. On multivariate analysis, glucocorticoid receptor (GR) overexpression correlated with persistent tumours in AAP + LHRHa (p = 0.018). The presence of nuclear androgen receptor splice variant (nARV7) correlated with persistent tumours in both arms. No new safety signals were observed. This is the first study investigating the role of preoperative AAP + LHRHa versus LHRHa alone in LHRPC. We report significant cytoreduction by tumour volume measures inversely correlating with biochemical relapse. Validation of these proposed tumour volume measures is planned. A potential role of GR in resistance to androgen biosynthesis inhibition warrants further study. Patient summary: This is the first study of abiraterone acetate plus leuprolide versus leuprolide alone in high-risk localised prostate cancer followed by prostatectomy. Patients in the combination arm had a significantly smaller tumour size. © 2019

This Chapter introduces the need for general Digital Rights Management (DRM) requirements. Further, it intertwines DRM with its spatial counterpart, Geospatial DRM (GeoDRM). However, unlike DRM, GeoDRM is far much complicated due to issues such as the development of Web Mapping technology among other issues. The Chapter discusses the ability of GeoDRM to mitigate transgression of Intellectual Property Rights (IPR). Highlighting economical and environmental wellbeing and other benefits of Spatial Data Infrastructure (SDI) geared towards global sustainable developments, the Chapter focuses on challenges of National Spatial Data Infrastructures (NSDIs) and Regional SDIs and the need to harmonize their standards for the upward mobility of global SDI (GSDI). Emphasizing the undisputed need for Local, Regional and Global Spatial Data Infrastructures (SDIs), in the presence of various Geo-communities and different GeoDRM models, the Chapter concludes that capacity building need to be urgently but carefully harnessed across all levels in order to develop cohesive GeoDRM policies.

Nuclear protein aggregation has been linked to genome instability and disease. The main source of aggregation‐prone proteins in cells is defective ribosomal products (DRiPs), which are generated by translating ribosomes in the cytoplasm. Here, we report that DRiPs rapidly diffuse into the nucleus and accumulate in nucleoli and PML bodies, two membraneless organelles formed by liquid–liquid phase separation. We show that nucleoli and PML bodies act as dynamic overflow compartments that recruit protein quality control factors and store DRiPs for later clearance. Whereas nucleoli serve as constitutive overflow compartments, PML bodies are stress‐inducible overflow compartments for DRiPs. If DRiPs are not properly cleared by chaperones and proteasomes due to proteostasis impairment, nucleoli undergo amyloidogenesis and PML bodies solidify. Solid PML bodies immobilize 20S proteasomes and limit the recycling of free ubiquitin. Ubiquitin depletion, in turn, compromises the formation of DNA repair compartments at fragile chromosomal sites, ultimately threatening cell survival.

Contemporary biological cells are sophisticated and highly compartmentalized. Compartmentalization is an essential principle of prebiotic life as well as a key feature in bottom-up synthetic biology research. In this review, the dynamic growth of compartments as an essential prerequisite for enabling self-reproduction as a fundamental life process is discussed. The micrometer-sized compartments are focused on due to their cellular dimensions. Two types of compartments are considered, membraneless droplets and membrane-bound microcompartments. Growth mechanisms of aqueous droplets such as protein (condensates) or macromolecule-rich droplets (aqueous two phase systems) and coacervates are discussed, for which growth occurs via Ostwald ripening or coalescence. For membrane-bound compartments, vesicles are considered, which are composed of fatty acids, lipids, or polymers, where directed growth can occur via fusion or uptake of material from the surrounding. The development of novel approaches for growth of biomimetic microcompartments can eventually be utilized to construct new synthetic cells.

Many industrial activities impose a threat on biodiversity, and it is unclear to what extent environmental regulations can reduce the threat of such activities. Bycatch in industrial fisheries is one of the greatest sources of mortality for seabirds, but a threat for which effective mitigation exists. Here we quantify whether the introduction of a new regulation that required the use of bird-scaring lines reduced seabird mortality in two of the most hazardous fisheries in the South Atlantic. The Namibian hake demersal trawl and longline fisheries, estimated to be killing 20,000–30,000 birds/year, have been required to use bird-scaring lines since 2015. We used data from BirdLife International's Albatross Task Force and the Namibian Fisheries Observer Agency to quantify changes in seabird mortality in these fisheries before and after the introduction of these regulations. Our estimated bycatch rates in the longline fleet were 0.468 birds/1000 hooks (95% confidence interval 0.067–1.450) before regulations and 0.004 birds/1000 hooks (0.001–0.013) following their introduction, a 98.4% reduction. Our estimate suggests that 215 (1–751) seabirds were killed across this fleet in 2018 compared to 22,222 (3187–68,786) in 2009. In the trawl fleet, observers recorded seabird mortality resulting from interactions with trawl cables. The average rate of heavy interactions was 1.09 interactions/h (0.81–1.39) before the regulation came into effect, and 0.49 interactions/h (0.23–0.84) since then. Extrapolations based on the number of observed fatal interactions suggest 1452 (0–3865) birds were killed by this fleet in 2017 compared to 7030 (0–16,374) in 2009. The lower mortality reduction in the trawl fleet is likely due to incomplete implementation of regulations and highlights the importance of adequate enforcement for effective bycatch mitigation. Overall, we demonstrate that regulations that mandate that well-tested safeguards are used during industrial operations can have enormous benefits for the conservation of threatened species.

The rapid growth of Botswana's economy since independence in 1966 has brought more tarred roads and vehicles, accompanied by an escalating road crash fatality rate. We tested the hypothesis that motor vehicle crash fatality increases resulted from, rather than just corresponded with, annual gross domestic product (GDP) increases. Data from Zambia, adjacent to Botswana, were used for comparison.Annual social and economic indicators and motor vehicle crash fatality rates in Botswana and Zambia were accessed from 1960 to 2012 and analysed using vector autoregressive analysis and Granger causality tests.In Botswana, annual changes in per capita GDP predicted annual changes in motor vehicle crash fatality rates (Road crash fatalities increased in recent decades in both Zambia and Botswana. But the rapid economic development in Botswana over this time period appears to have driven proportionate road traffic fatality increases. There are opportunities for newly emerging economies such as Zambia, Angola, and others to learn from the Botswana experience. Evidence-based investments in road safety interventions should be concomitant with economic development.La croissance rapide de l’économie du Botswana depuis l’indépendance en 1966 s’est traduite par le développement du nombre de routes goudronnées et de véhicules, accompagnés d’un taux de mortalité due aux accidents de la route qui va s’accélérant. Nous avons testé l’hypothèse selon laquelle les hausses de la mortalité due aux accidents de véhicules motorisés seraient attribuables aux augmentations du produit intérieur brut (PIB), plutôt que d’en être un simple reflet. Des données provenant de Zambie, pays adjacent au Botswana, ont été utilisées pour établir une comparaison.Des indicateurs économiques et sociaux annuels et les taux de mortalité due aux accidents de la route au Botswana et en Zambie ont été examinés sur la période 1960–2012 et analysés en utilisant une analyse vectorielle autorégressive et des tests de causalité au sens de Granger.Au Botswana, les variations annuelles de PIB par habitant ont prédit les variations annuelles des taux de mortalité due aux accidents de véhicule motorisés (p = 0,042). L’inverse n’a pas été observé; les variations annuelles de taux de mortalité due aux accidents de véhicules motorisés ne permettent pas de prédire les variations annuelles de PIB. Ces résultats suggèrent que la croissance du PIB pour une année donnée a causé des décès occasionnés par des accidents de la route au Botswana et qu’en moyenne, chaque augmentation d’un milliard de dollars du PIB a produit une augmentation du taux de décès occasionnés par des accidents de la route. En Zambie, les variations annuelles de PIB ont prédit les variations annuelles du taux de mortalité trois ans plus tard (p = 0,029), mais les variations annuelles des taux de mortalité des accidents de la route ont également prédit les augmentations annuelles de PIB par habitant (p = 0,026) trois ans plus tard, ce qui suggère une corrélation entre les tendances mais pas un effet de causalité du PIB.Les décès occasionnés par les accidents de la route ont augmenté au cours des dernières décennies en Zambie comme au Botswana. Mais le développement économique rapide au Botswana au cours de cette période semble avoir entraîné des augmentations proportionnelles des décès dus aux accidents de la route. Il est possible, pour les nouvelles économies émergentes comme la Zambie, l’Angola, et d’autres, de tirer des leçons de l’expérience du Botswana. Des investissements dans des interventions en matière de sécurité routière, fondés sur des données concrètes, doivent être concomitants au développement économique.

Summary Cells sense elevated temperatures and mount an adaptive heat shock response that involves changes in gene expression, but the underlying mechanisms, particularly on the level of translation, remain unknown. Here we report that, in budding yeast, the essential translation initiation factor Ded1p undergoes heat-induced phase separation into gel-like condensates. Using ribosome profiling and an in vitro translation assay, we reveal that condensate formation inactivates Ded1p and represses translation of housekeeping mRNAs while promoting translation of stress mRNAs. Testing a variant of Ded1p with altered phase behavior as well as Ded1p homologs from diverse species, we demonstrate that Ded1p condensation is adaptive and fine-tuned to the maximum growth temperature of the respective organism. We conclude that Ded1p condensation is an integral part of an extended heat shock response that selectively represses translation of housekeeping mRNAs to promote survival under conditions of severe heat stress.

Stressed cells shut down translation, release mRNA molecules from polysomes, and form stress granules (SGs) via a network of interactions that involve G3BP. Here we focus on the mechanistic underpinnings of SG assembly. We show that, under non-stress conditions, G3BP adopts a compact auto-inhibited state stabilized by electrostatic intramolecular interactions between the intrinsically disordered acidic tracts and the positively charged arginine-rich region. Upon release from polysomes, unfolded mRNAs outcompete G3BP auto-inhibitory interactions, engendering a conformational transition that facilitates clustering of G3BP through protein-RNA interactions. Subsequent physical crosslinking of G3BP clusters drives RNA molecules into networked RNA/protein condensates. We show that G3BP condensates impede RNA entanglement and recruit additional client proteins that promote SG maturation or induce a liquid-to-solid transition that may underlie disease. We propose that condensation coupled to conformational rearrangements and heterotypic multivalent interactions may be a general principle underlying RNP granule assembly.

© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Among the best-performing homogeneous catalysts for the direct amination of activated secondary alcohols with electron-poor amine derivatives, metal triflates, such as aluminum triflate, Al(OTf)3, stand out. Herein we report the extension of this reaction to electron-rich amines and activated primary alcohols. We provide detailed insight into the structure and reactivity of the catalyst under working conditions in both nitromethane and toluene solvent, through experiment (cyclic voltammetry, conductimetry, NMR spectroscopy), and density functional theory (DFT) simulations. Competition between aniline and benzyl alcohol for Al in the two solvents explains the different reactivities. The catalyst structures predicted from the DFT calculations were validated by the experiments. Whereas a SN1-type mechanism was found to be active in nitromethane, we propose a SN2 mechanism in toluene to rationalize the much higher selectivity observed when using this solvent. Also, unlike what is commonly assumed in homogeneous catalysis, we show that different active species may be active instead of only one.

RNA and membraneless organelles Membraneless compartments can form in cells through liquidliquid phase separation (see the Perspective by Polymenidou). But what prevents these cellular condensates from randomly fusing together? Using the RNA-binding protein (RBP) Whi3, Langdon et al. demonstrated that the secondary structure of different RNA components determines the distinct biophysical and biological properties of the two types of condensates that Whi3 forms. Several RBPs, such as FUS and TDP43, contain prion-like domains and are linked to neurodegenerative diseases. These RBPs are usually soluble in the nucleus but can form pathological aggregates in the cytoplasm. Maharana et al. showed that local RNA concentrations determine distinct phase separation behaviors in different subcellular locations. The higher RNA concentrations in the nucleus act as a buffer to prevent phase separation of RBPs; when mislocalized to the cytoplasm, lower RNA concentrations trigger aggregation. Science , this issue p. 922 , p. 918 ; see also p. 859

The formation of membrane-less organelles and compartments by protein phase separation is an important way in which cells organize their cytoplasm and nucleoplasm. In vitro phase separation assays with purified proteins have become the standard way to investigate proteins that form membrane-less compartments. By now, various proteins have been purified and tested for their ability to phase separate and form liquid condensates in vitro. However, phase-separating proteins are often aggregation-prone and difficult to purify and handle. As a consequence, the results from phase separation assays often differ between labs and are not easily reproduced. Thus, there is an urgent need for high-quality proteins, standardized procedures, and generally agreed-upon practices for protein purification and conducting phase separation assays. This paper provides protocols for protein purification and guides the user through the practicalities of in vitro protein phase separation assays, including best-practice approaches and pitfalls to avoid. We believe that this compendium of protocols and practices will provide a useful resource for scientists studying the phase behavior of proteins., Graphical Abstract Unlabelled Image, Highlights • Membrane-less organelles form by phase separation. • Membrane-less organelles can be reconstituted from minimal components. • Phase-separating proteins are difficult to purify and handle. • We provide guidelines and protocols for working with phase-separating proteins.

Provided is a method for producing an alkene. The method comprises a step of converting a compound (I) comprising at least two hydroxyl functions into a compound (II) comprising at least two alkenyl functions with cerium oxide particles as catalyst. It notably permits the conversion of 1,3-butanediol,1,4-butanediol into butadiene.; L'invention concerne un procédé de production d'un alcène. Le procédé comprend une étape de conversion d'un composé (I) comprenant au moins deux fonctions hydroxyle en un composé (II) comprenant au moins deux fonctions alcényle avec des particules d'oxyde de cérium comme catalyseur. Elle permet notamment la conversion de 1,3-butanediol,1,4-butanediol en butadiène.

INTRODUCTION The formation of dynamic, membraneless compartments using intracellular phase transitions such as phase separation and gelation provides an efficient way for cells to respond to environmental changes. Recent work has identified a special class of intrinsically disordered domains enriched for polar amino acids such as glycine, glutamine, serine, or tyrosine as potential drivers of phase separation in cells. However, more traditional work has highlighted the ability of these domains to drive the formation of fibrillar aggregates. Such domains are also known as prion domains. They have first been identified in budding yeast proteins that form amyloid-like aggregates. Because these aggregates are heritable and change the activity of the prion-domain–containing protein, they are thought to be a common mechanism for phenotypic inheritance in fungi and other organisms. However, the aggregation of prion domains has also been associated with neurodegenerative diseases in mammals. Therefore, the relationship between the role of these domains as drivers of phase separation and their ability to form prion-like aggregates is unknown. RATIONALE The budding yeast translation termination factor Sup35 is an archetypal prion-domain–containing protein. Sup35 forms irreversible heritable aggregates, and these aggregates have been proposed to be either a disease or an adaptation that generates heritable phenotypic variation in populations of budding yeast. Despite having been described almost 25 years ago, the physiological functions of the Sup35 prion domain and other prion-like domains remain unclear. Uncovering these functions is a prerequisite for understanding the evolutionary pressures shaping prion-like sequences and how their physiological and pathological transitions affect cellular fitness. RESULTS Here, we show that the prion domain of Sup35 drives the reversible phase separation of the translation termination factor into biomolecular condensates. These condensates are distinct and different from fibrillar amyloid-like prion particles. Combining genetic analysis in cells with in vitro reconstitution protein biochemistry and quantitative biophysical methods, we demonstrate that Sup35 condensates form by pH-induced liquid-like phase separation as a response to sudden stress. The condensates are liquid-like initially but subsequently solidify to form protective protein gels. Cryo–electron tomography demonstrates that these gel-like condensates consist of cross-linked Sup35 molecules forming a porous meshwork. A cluster of negatively charged amino acids functions as a pH sensor and regulates condensate formation. The ability to form biomolecular condensates is shared among distantly related budding yeast and fission yeast. This suggests that condensate formation is a conserved and ancestral function of the prion domain of Sup35. In agreement with an important physiological function of the prion domain, the catalytic guanosine triphosphatase (GTPase) domain of the translation termination factor Sup35 readily forms irreversible aggregates in the absence of the prion domain. Consequently, cells lacking the prion domain exhibit impaired translational activity and a growth defect when recovering from stress. These data demonstrate that the prion domain rescues the essential GTPase domain of Sup35 from irreversible aggregation, thus ensuring that the translation termination factor remains functional during harsh environmental conditions. CONCLUSION The prion domain of Sup35 is a highly regulated molecular device that has the ability to sense and respond to physiochemical changes within cells. The N-terminal prion domain provides the interactions that drive liquid phase separation. Phase separation is regulated by the adjacent stress sensor. The synergy of these two modules enables the essential translation termination factor to rapidly form protective condensates during stress. This suggests that prion domains are protein-specific stress sensors and modifiers of protein phase transitions that allow cells to respond to specific environmental conditions.

SummaryMany proteins contain disordered regions of low-sequence complexity, which cause aging-associated diseases because they are prone to aggregate. Here, we study FUS, a prion-like protein containing intrinsically disordered domains associated with the neurodegenerative disease ALS. We show that, in cells, FUS forms liquid compartments at sites of DNA damage and in the cytoplasm upon stress. We confirm this by reconstituting liquid FUS compartments in vitro. Using an in vitro “aging” experiment, we demonstrate that liquid droplets of FUS protein convert with time from a liquid to an aggregated state, and this conversion is accelerated by patient-derived mutations. We conclude that the physiological role of FUS requires forming dynamic liquid-like compartments. We propose that liquid-like compartments carry the trade-off between functionality and risk of aggregation and that aberrant phase transitions within liquid-like compartments lie at the heart of ALS and, presumably, other age-related diseases.Video Abstract