Networking and Dynamic Switches in Biological Condensates.

Cellular liquid-liquid phase separation (LLPS) plays a key role in the dynamics and function of RNA-protein condensates like stress granules. In this issue of Cell, Yang et al., Guillén-Boixet et al., and Sanders et al. use a combination of experiment and modeling to provide an exciting mechanistic insight into the relationship between stress granules and LLPS, for example, in the context of protein disorder, switchable interactions, graph theory, and multiple interacting dense phases.

Glutamine-rich regions of the disordered CREB transactivation domain mediate dynamic intra- and intermolecular interactions.

The cyclic AMP response element (CRE) binding protein (CREB) is a transcription factor that contains a 280-residue N-terminal transactivation domain and a basic leucine zipper that mediates interaction with DNA. The transactivation domain comprises three subdomains, the glutamine-rich domains Q1 and Q2 and the kinase inducible activation domain (KID). NMR chemical shifts show that the isolated subdomains are intrinsically disordered but have a propensity to populate local elements of secondary structure. The Q1 and Q2 domains exhibit a propensity for formation of short ß-hairpin motifs that function as binding sites for glutamine-rich sequences. These motifs mediate intramolecular interactions between the CREB Q1 and Q2 domains as well as intermolecular interactions with the glutamine-rich Q1 domain of the TATA-box binding protein associated factor 4 (TAF4) subunit of transcription factor IID (TFIID). Using small-angle X-ray scattering, NMR, and single-molecule Förster resonance energy transfer, we show that the Q1, Q2, and KID regions remain dynamically disordered in a full-length CREB transactivation domain (CREBTAD) construct. The CREBTAD polypeptide chain is largely extended although some compaction is evident in the KID and Q2 domains. Paramagnetic relaxation enhancement reveals transient long-range contacts both within and between the Q1 and Q2 domains while the intervening KID domain is largely devoid of intramolecular interactions. Phosphorylation results in expansion of the KID domain, presumably making it more accessible for binding the CBP/p300 transcriptional coactivators. Our study reveals the complex nature of the interactions within the intrinsically disordered transactivation domain of CREB and provides molecular-level insights into dynamic and transient interactions mediated by the glutamine-rich domains.

Serine deamination by human serine racemase synergizes with antibiotics to curtail the replication of Chlamydia trachomatis.

The obligate intracellular bacterium, Chlamydia trachomatis, has evolved to depend on its human host for many metabolites, including most amino acids and three of the four nucleotides. Given this, it is not surprising that depletion of a single amino acid in the host cell growth medium blocks chlamydial replication. Paradoxically, supra-normal levels of some amino acids also block productive replication of Chlamydia. Here, we have determined how elevated serine levels, generated by exogenous supplementation, impede chlamydial inclusion development and reduce the generation of infectious progeny. Our findings reveal that human serine racemase, which is broadly expressed in multiple tissues, potentiates the anti-chlamydial effect of elevated serine concentrations. In addition to reversibly converting l-serine to d-serine, serine racemase also deaminates serine via ß-elimination. We have determined that d-serine does not directly impact Chlamydia; rather, ammonia generated by serine deamination limits the productive chlamydial replication. Our findings imply that ammonia produced within host cells can traverse the chlamydial inclusion membrane. Further, this property of serine deaminase can be exploited to sensitize Chlamydia to concentrations of doxycycline that are otherwise not bactericidal. Because exogenously elevated levels of serine can be tolerated over extended periods, the broad expression pattern of serine racemase indicates it to be a host enzyme whose activity can be directed against multiple intracellular bacterial pathogens. From a therapeutic perspective, demonstrating host metabolism can be skewed to generate an anti-bacterial metabolite that synergizes with antibiotics, we believe our results provide a new approach to target intracellular pathogens.

Temporal evolution of chest radiographic appearances in COVID-19 with clinicoradiological associations: a multicentre United Kingdom resident-led study.

AIM: To describe the (a) frequency of improving, static, and worsening chest radiograph (CXR) appearances; (b) differences in demographic, initial rudimentary haematological and CXR variables and these patterns; and (c) frequency of different trajectories of serial CXR evolution, in COVID-19 patients presenting consecutively. MATERIALS AND METHODS: This multicentre retrospective study included all COVID-19 patients admitted from 1-30 April 2020, meeting the inclusion criteria across 24 (blinded) hospitals. Follow-up CXRs on admission, the subsequent (where available), and at 4-8 weeks were scored for the presence of parenchymal opacities across six zones. Three cohorts were defined: improved, static, and/or worsened. The chi-squared and Kruskal-Wallis tests were used to compare demographic, laboratory, and CXR variables. Trajectories of CXR evolution were assessed when all three CXRs were available (226 patients). RESULTS: Of 452 included patients (median age 66 years, interquartile range 54.3-79, 262 men), 211 (46.7%) improved, 140 (31%) were static, and 101 (22.3%) worsened. Improving patients were more likely younger, with a classic COVID-19 radiograph and higher initial CXR zonal severity scores (both p<0.001), while worsening patients had lower initial lymphocyte counts (p=0.008). The most frequent trajectory was worsened then improved (n=63, 27.9%) followed by static then improved (n=46, 20.4%) and static (n=42, 18.6%). CONCLUSION: Most patients with COVID-19 during the first wave of the pandemic demonstrated radiographic improvement; these patients were more likely younger with classic COVID-19 appearances and initially more extensive abnormality. Conversely, radiographic deterioration was associated with lower lymphocyte counts. The three most common trajectories were worsening then improvement, static then improvement, and static throughout.

Synthesis and study of optical properties of a Gd3+-doped NaYF4 phosphor for phototherapy lamp application.

In recent trends, radiation falls under the narrowband ultraviolet-B region (305-315 nm) widely used in phototherapy lamp applications in the treatment of skin diseases. In this paper, we report a Gd3+-doped NaYF4 luminescent material synthesized for the first time using the low-temperature co-precipitation method. It crystallized into a face-centred cubic structure, as confirmed by X-ray diffraction characterization techniques and Rietveld refinement. The photoluminescence property of the as-prepared sample shows a highly intense, sharp emission band obtained at 311 nm, which belongs to the narrowband ultraviolet-B region and corresponds to the transition of the 6P7/2→8S7/2 level of the Gd3+ ions under 272 nm excitation (8S7/2 to 6IJ). The transitions of the Gd3+ ions are detected entirely with different concentrations of Gd3+ ions. Scanning electron microscopy analysis indicated that the average particle was 288 nm. The critical distance for energy transfer was calculated to be equal to 11.5017 Å. Dipole-dipole interaction is responsible for energy transfer, as analyzed by Dexter theory. These excellent optical characteristics, together with their highly efficient and low-cost synthesis approach, indicate that synthesized NaYF4:Gd3+ phosphors have excessive potential for phototherapeutic lamp applications.

Academic clinical fellows in radiology: how can we improve success?

AIM: To survey past and current radiology academic clinical fellows (ACFs) for feedback on their experiences, academic achievements, challenges faced in balancing academic and clinical responsibilities, and opinion on how to optimise the fellowship programme. MATERIALS & METHODS: A 26-question online survey approved by the Royal College of Radiologists (RCR) Academic Committee was distributed over a 7-month period (June 2021 to January 2022) to current and past radiology ACFs via the National Institute for Health and Care Research (NIHR) integrated academic training imaging leads, radiology training programme directors, and social media. RESULTS: Thirty-five survey responses were received from past or present ACFs. Of the respondents, 42.8% (15/35) entered ACF training from another research post, and most continued their academic interests after ACF training (59.3%, 16/27 that had completed the post). The majority (22/35, 63%) had or were in the process of obtaining a postgraduate research degree. The most common academic outputs were scientific publications and national/international conference presentations. Most (23/35, 66%) would recommend the ACF post to colleagues, although some found it challenging balancing on-call and examination commitments during training. CONCLUSIONS: Entry into the radiology ACF programme is often after a prior academic post. Many ACFs appear to enjoy their fellowship experience and continue academic interests after training, some achieving higher research degrees. Challenges in balancing clinical workload require some flexibility from local clinical and academic supervisors. Suggestions for alternative structuring of the ACF pathway and how to optimise entry into these competitive posts are also outlined.

Synthesis and study of luminescence properties of a deep red-emitting phosphor K2 LiAlF6 :Mn4+ for plant cultivation.

Light is the most important component in plant growth and development. This study synthesised a novel Mn4+ -doped K2 LiAlF6 red-emitting phosphor using the coprecipitation method. We observed that on addition of dopant Mn4+ ions to the host K2 LiAlF6 , its phase changed from rhombohedral to cubic due to the change in the lattice position of the atoms. When the atoms are excited at 468 nm, the K2 LiAlF6 :Mn4+ phosphor exhibited a red emission band ranging from 630 to 700 nm, centred at 638 nm, which matched well with the absorption spectra of phytochrome PR. The critical quenching content of Mn4+ ions was ~3 mol%. The critical distance between Mn4+ ions was determined to be 19.724 Å, and non-radiative energy transfer among the nearest-neighbour Mn4+ ions was the mechanism used for the concentration quenching effect. The Commission International de l'Eclairage (CIE) chromaticity coordinates of the K2 LiAlF6 :0.03 Mn4+ sample were (x = 0.7162, y = 0.2837). The luminescence mean decay time was calculated to be 8.29 ms. These results demonstrated the promising prospect of K2 LiAlF6 :Mn4+ as a red-emitting phosphor for application in red light-emitting diodes for plant cultivation.

Do children in India grow well into adolescents? Longitudinal analysis of growth transitions from Young Lives panel survey in India.

OBJECTIVES: Studies that examined the growth during late childhood and early adolescence beyond 8 years of age are very limited. Further, most studies have used dichotomized classification of stunting, thereby limiting the understanding of moderate stunting in childhood growth trajectory. We aimed to examine the course of stunting from childhood to adolescence by undertaking robust analyses of the Young Lives Survey (YLS) longitudinal data from India using multilevel categorization of stunting. STUDY DESIGN: Retrospective cohort analysis was undertaken from YLS in India among 1827 children from the younger cohort born in 2001-02 with complete follow-up data in all five rounds of YLS collected in 2002, 2006, 2009, 2013, and 2016. METHODS: A three-state multistate Markov model (not stunted, moderate, severe) was performed to estimate annual transition probabilities, mean sojourn-time, and transition-specific risk factors. RESULTS: Between Round-one and Round-five, cross-sectional prevalence of severe stunting decreased from 10.4% (95% confidence interval [CI]: 7.8%, 13.7%) to 5.3% (95% CI: 3.8%, 7.3%), while moderate stunting increased from 19.9% (95% CI: 16.3%,23.9%) to 21.7% (95% CI: 18.4%, 24.9%). Mean Sojourn time estimation indicated a relatively concise state for moderate stunting. The stunting trajectory had shown gender differential where more faltering to severe stunting and lower recovery to the normal state was observed among girls between 8 and 12 years and among boys between 12 and 15 years. Compared with boys, girls had 40% excess likelihood (Hazard Ratio: 1.40; 95% CI 1.00 to 1.95) for moderate-to-severe stunting transition and also had 19% excess likelihood (Hazard Ratio: 1.19; 95% CI 1.01 to 1.40) of favorable transition (moderate-to-non-stunted). CONCLUSIONS: The transition trajectory highlights preadolescence, especially among girls, as an additional window of opportunity to ensure better nutrition in adolescent life. With a fifth of adolescents living in India, study findings call for coordinated, multisectoral, age-appropriate, and gender-responsive approach to take India closer to meeting SDG-2.

Paper-Based Optode Devices (PODs) for Selective Quantification of Potassium in Biological Fluids.

This paper describes the design, fabrication, and feasibility of paper-based optode devices (PODs) for sensing potassium selectively in biological fluids. PODs operate in exhaustive mode and integrate with a handheld, smartphone-connected optical reader. This integrated measuring system provides significant advantages over traditional optode membranes and other paper-based designs, by obtaining a linear optical response to potassium concentration via a simple, stackable design and by harnessing a smartphone to provide an easy-to-use interface, thus enabling remote monitoring of diseases.

Modelling and optimization of process parameters for production of desiccated Chhana-murki (Indian cottage cheese-based dessert).

In the present study, process parameters were optimized for the production of desiccated chhana-murki (Indian cottage cheese-based dessert). Response Surface Methodology (RSM) was employed to explore the mutual effects of coagulation temperature (CT) of milk (70-90 °C), % fat level in milk (3.5%-5.5%), and sugar-to-paneer cube (SP) ratio (0.6-0.9) on instrumental hardness (N), water activity (aw), yield (%), sensory sweetness and overall acceptability (on 100-point intensity scale) of chhana-murki. The resulted responses were evaluated by analysis of variance (ANOVA), and the second-order polynomial response surface equations were fitted using multiple regression analysis. Determination coefficients (R 2) were equal to 80% or higher for individual responses stated that the developed models were well fitted to the experimental results. The optimized product was prepared using CT 79.22 °C, milk fat 4.8%, and SP ratio 0.7. Confirmatory experiment values for instrument hardness, water activity (aw), yield (%), sensory sweetness and overall acceptability were 105.05 N, 0.85, 115.2%, 61.2 and 78.8, respectively.

Transcriptional reprogramming and enhanced photosynthesis drive inducible salt tolerance in sugarcane mutant line M4209.

Sugarcane (Saccharum officinarum) is a globally cultivated cash crop whose yield is negatively affected by soil salinity. In this study, we investigated the molecular basis of inducible salt tolerance in M4209, a sugarcane mutant line generated through radiation-induced mutagenesis. Under salt-contaminated field conditions, M4209 exhibited 32% higher cane yield as compared with its salt-sensitive parent, Co86032. In pot experiments, post-sprouting phenotyping indicated that M4209 had significantly greater leaf biomass compared with Co86032 under treatment with 50 mM and 200 mM NaCl. This was concomitant with M4209 having 1.9-fold and 1.6-fold higher K+/Na+ ratios, and 4-fold and 40-fold higher glutathione reductase activities in 50 mM and 200 mM NaCl, respectively, which suggested that it had better ionic and redox homeostasis than Co86032. Transcriptome profiling using RNA-seq indicated an extensive reprograming of stress-responsive modules associated with photosynthesis, transmembrane transport, and metabolic processes in M4209 under 50 mM NaCl stress. Using ranking analysis, we identified Phenylalanine Ammonia Lyase (PAL), Acyl-Transferase Like (ATL), and Salt-Activated Transcriptional Activator (SATA) as the genes most associated with salt tolerance in M4209. M4209 also exhibited photosynthetic rates that were 3-4-fold higher than those of Co86032 under NaCl stress conditions. Our results highlight the significance of transcriptional reprogramming coupled with improved photosynthetic efficiency in determining salt tolerance in sugarcane.

Assessment of preferable site for temperature measurement using non contact infra-red temperature among pediatric patients.

BACKGROUND: Accurate temperature measurement with little or no discomfort that is safe, without risk of hospital-acquired infections or perforations, is the preferred choice of medical professionals in pediatric settings. The objective was to discover the preferable site for body temperature measurement using non-contact infra-red thermometer (NCIT) among pediatric patients. METHODS: NCIT measurement at mid forehead (F), right temporal region (T), right side of neck- over-carotid artery area (N), jugular notch (J), sternum (S), umbilical region and sublingual region (U) were compared with digital axillary temperature (DAT) in a single attempt in 500 patients, aged between 2 to 5 years with fever. Data was analyzed using Pearson's correlation, paired T-test and Bland-Altman plot to assess the correlation and agreement between the DAT and NCIT sites. RESULTS: The mean temperature of NCIT-T (38.42 ± 0.64 °C) was more agreeable with DAT (38.42 ± 0.63 °C) compared to other body sites. The minimum mean bias of -0.00480 °C was noted for NCIT-F with 95% CI of -0.164-0.15; however, NCIT-F revealed many outliers as compared to NCIT-J. A strong positive correlation existed between DAT and NCIT sites (r value: 0.99-0.98). However, significant difference was found between DAT and NCIT-N, NCIT-F and NCIT-U (p value < 0.0001). CONCLUSIONS: NCIT-J is the most preferable choice for measuring body temperature and can be interchanged with DAT. It will help to deliver fast results with enhanced patient comfort due to its non-invasive nature.

Magnetic Levitation in Chemistry, Materials Science, and Biochemistry.

All matter has density. The recorded uses of density to characterize matter date back to as early as ca. 250 BC, when Archimedes was believed to have solved "The Puzzle of The King's Crown" using density.[1] Today, measurements of density are used to separate and characterize a range of materials (including cells and organisms), and their chemical and/or physical changes in time and space. This Review describes a density-based technique-magnetic levitation (which we call "MagLev" for simplicity)-developed and used to solve problems in the fields of chemistry, materials science, and biochemistry. MagLev has two principal characteristics-simplicity, and applicability to a wide range of materials-that make it useful for a number of applications (for example, characterization of materials, quality control of manufactured plastic parts, self-assembly of objects in 3D, separation of different types of biological cells, and bioanalyses). Its simplicity and breadth of applications also enable its use in low-resource settings (for example-in economically developing regions-in evaluating water/food quality, and in diagnosing disease).

Physical Chemistry of Cellular Liquid-Phase Separation.

Compartmentalization of biochemical processes is essential for cell function. Although membrane-bound organelles are well studied in this context, recent work has shown that phase separation is a key contributor to cellular compartmentalization through the formation of liquid-like membraneless organelles (MLOs). In this Minireview, the key mechanistic concepts that underlie MLO dynamics and function are first briefly discussed, including the relevant noncovalent interaction chemistry and polymer physical chemistry. Next, a few examples of MLOs and relevant proteins are given, along with their functions, which highlight the relevance of the above concepts. The developing area of active matter and non-equilibrium systems, which can give rise to unexpected effects in fluctuating cellular conditions, are also discussed. Finally, our thoughts for emerging and future directions in the field are discussed, including in vitro and in vivo studies of MLO physical chemistry and function.

The effects of ketamine on dopaminergic function: meta-analysis and review of the implications for neuropsychiatric disorders.

Ketamine is a non-competitive antagonist at the N-methyl-d-aspartate receptor. It has recently been found to have antidepressant effects and is a drug of abuse, suggesting it may have dopaminergic effects. To examine the effect of ketamine on the dopamine systems, we carried out a systematic review and meta-analysis of dopamine measures in the rodent, human and primate brain following acute and chronic ketamine administration relative to a drug-free baseline or control condition. Systematic search of PubMed and PsychInfo electronic databases yielded 40 original peer-reviewed studies. There were sufficient rodent studies of the acute effects of ketamine at sub-anaesthetic doses for meta-analysis. Acute ketamine administration in rodents is associated with significantly increased dopamine levels in the cortex (Hedge's g= 1.33, P<0.01), striatum (Hedge's g=0.57, P<0.05) and the nucleus accumbens (Hedge's g=1.30, P<0.05) compared to control conditions, and 62-180% increases in dopamine neuron population activity. Sub-analysis indicated elevations were more marked in in vivo (g=1.93) than ex vivo (g=0.50) studies. There were not enough studies for meta-analysis in other brain regions studied (hippocampus, ventral pallidum and cerebellum), or of the effects of chronic ketamine administration, although consistent increases in cortical dopamine levels (from 88 to 180%) were reported in the latter studies. In contrast, no study showed an effect of anaesthetic doses (>100 mg kg-1) of ketamine on dopamine levels ex vivo, although this remains to be tested in vivo. Findings in non-human primates and in human studies using positron emission tomography were not consistent. The studies reviewed here provide evidence that acute ketamine administration leads to dopamine release in the rodent brain. We discuss the inter-species variation in the ketamine induced dopamine release as well as the implications for understanding psychiatric disorders, in particular substance abuse, schizophrenia, and the potential antidepressant properties of ketamine, and comparisons with stimulants and other NMDA antagonists. Finally we identify future research needs.

Modulation of allostery by protein intrinsic disorder.

Allostery is an intrinsic property of many globular proteins and enzymes that is indispensable for cellular regulatory and feedback mechanisms. Recent theoretical and empirical observations indicate that allostery is also manifest in intrinsically disordered proteins, which account for a substantial proportion of the proteome. Many intrinsically disordered proteins are promiscuous binders that interact with multiple partners and frequently function as molecular hubs in protein interaction networks. The adenovirus early region 1A (E1A) oncoprotein is a prime example of a molecular hub intrinsically disordered protein. E1A can induce marked epigenetic reprogramming of the cell within hours after infection, through interactions with a diverse set of partners that include key host regulators such as the general transcriptional coactivator CREB binding protein (CBP), its paralogue p300, and the retinoblastoma protein (pRb; also called RB1). Little is known about the allosteric effects at play in E1A-CBP-pRb interactions, or more generally in hub intrinsically disordered protein interaction networks. Here we used single-molecule fluorescence resonance energy transfer (smFRET) to study coupled binding and folding processes in the ternary E1A system. The low concentrations used in these high-sensitivity experiments proved to be essential for these studies, which are challenging owing to a combination of E1A aggregation propensity and high-affinity binding interactions. Our data revealed that E1A-CBP-pRb interactions have either positive or negative cooperativity, depending on the available E1A interaction sites. This striking cooperativity switch enables fine-tuning of the thermodynamic accessibility of the ternary versus binary E1A complexes, and may permit a context-specific tuning of associated downstream signalling outputs. Such a modulation of allosteric interactions is probably a common mechanism in molecular hub intrinsically disordered protein function.

In vivo effect of two different dietary fiber blends on the milk calcium bioavailability.

Milk is a valuable source of dietary calcium and it becomes important to establish whether incorporation of dietary fiber (DF), a health promoting food constituent, would lead to any undesirable impact on the bioavailability of milk calcium or not. The DF fortified spray dried partly skimmed milk powder with prestandardized fiber Blend-I (psyllium husk, oat fiber, MCC, inulin) and fiber Blend-II (psyllium husk, oat bran, wheat fiber and inulin) was subjected to rat-feeding studies to examine the possible effects on the bioavailability of milk calcium. The differences for calcium absorption and retention among diets containing DF Blend-I, DF Blend-II and cellulose (control) were found to be non-significant. It was evident that the milk calcium bioavailability of the diets containing two fiber formulations tested (at the levels studied) was at par with that of control standard diet containing only cellulose as DF. Therefore, it is reasonable to incorporate these DF blends into dairy products, and thereby add value.

Reentrant Phase Transitions and Non-Equilibrium Dynamics in Membraneless Organelles.

Compartmentalization of biochemical components, interactions, and reactions is critical for the function of cells. While intracellular partitioning of molecules via membranes has been extensively studied, there has been an expanding focus in recent years on the critical cellular roles and biophysical mechanisms of action of membraneless organelles (MLOs) such as the nucleolus. In this context, a substantial body of recent work has demonstrated that liquid-liquid phase separation plays a key role in MLO formation. However, less is known about MLO dissociation, with phosphorylation being the primary mechanism demonstrated thus far. In this Perspective, we focus on another mechanism for MLO dissociation that has been described in recent work, namely a reentrant phase transition (RPT). This concept, which emerges from the polymer physics field, provides a mechanistic basis for both formation and dissolution of MLOs by monotonic tuning of RNA concentration, which is an outcome of cellular processes such as transcription. Furthermore, the RPT model also predicts the formation of dynamic substructures (vacuoles) of the kind that have been observed in cellular MLOs. We end with a discussion of future directions in terms of open questions and methods that can be used to answer them, including further exploration of RPTs in vitro, in cells, and in vivo using ensemble and single-molecule methods as well as theory and computation. We anticipate that continued studies will further illuminate the important roles of reentrant phase transitions and associated non-equilibrium dynamics in the spatial patterning of the biochemistry and biology of the cell.