New Segmentation and Feature Extraction Algorithm for Classification of White Blood Cells in Peripheral Smear Images This article addresses a new method for the classification of white blood cells (WBCs) using image processing techniques and machine learning methods. The proposed method consists of three steps: detecting the nucleus and cytoplasm, extracting features, and classification. At first, a new algorithm is designed to segment the nucleus. For the cytoplasm to be detected, only a part of it located inside the convex hull of the nucleus is involved in the process. This attitude helps us overcome the difficulties of segmenting the cytoplasm. In the second phase, three shapes and four novel color features are devised and extracted. Finally, by using an SVM model, the WBCs are classified. The segmentation algorithm can detect the nucleus with a dice similarity coefficient of 0.9675. The proposed method can categorize WBCs in Raabin-WBC, LISC, and BCCD datasets with accuracies of 94.65 %, 92.21 %, and 94.20 %, respectively. It is worth mentioning that the hyperparameters of the classifier are fixed only with the Raabin-WBC dataset, and these parameters are not readjusted for LISC and BCCD datasets. The obtained results demonstrate that the proposed method is robust, fast, and accurate.
NOMA: a high-throughput microchip for robust, sequential measurements of secretions from the same single-cells Despite recent advances in single-cell analysis technologies, lacking simple methods to keep the live single-cells traceable for longitudinal detection reliably poses a significant obstacle in single-cell secretion analysis. Here we reported the high-density NOMA (narrow-opening microwell array) microchip that realized the retention of[&ge;] 97% of trapped single cells in dedicated spatial locations during repetitive detection procedures, verified with both adherent and suspension cells by two researchers independently. We applied it in monitoring single-cell protein secretions sequentially from the same single cells, and we found the digital protein secretion patterns dominate the protein secretion. We also demonstrated the microchip for longitudinally tracking IL-8 and the CD81+EV secretions from the same single-cells over days, which revealed the presence of "super secretors" within the cell population be more persistent to secrete protein or extracellular vesicle for an extended period. The NOMA platform reported here is simple, robust, and easy to operate for tracking sequential measurements from the same single cells, representing a novel and informative tool to inspire new observations in biomedical research.

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If Research is Not the Evidence, What is it? The Lack of Research Citations in Health Vlogs Doctor-influencers have the knowledge needed to understand scientific research, and they have the social media popularity to share it with large audiences. In this article, I explore the treatment of evidence in vlogs by Egyptian doctor-influencers: how they present it and what they use instead of it. I answer the following questions: 1) How do doctor-influencers present evidence on social media? 2) If evidence is not the research, then what is it? The data was collected through in-depth interviews with 12 Egyptian doctor-influencers from different specializations; two focus groups; and a critical discourse analysis of 48 of the influencers most popular and most engaging videos. I found that the doctor-influencers cited academic research only in the videos about medical controversies, new treatments, misinformation, or common medical mistakes. In most of their videos, the Egyptian doctor-influencers scarcely referred to medical research. This is because they believe that their audiences would not understand evidence due to their low educational attainment, weak research skills, and the language barrier (as most medical research is published in English rather than Arabic). These findings highlight the challenges of communicating science in societies with a low level of education. In these settings, communicators may need to move beyond mediating research studies on social media and instead focus on simplifying, translating, and connecting evidence-based information to the cultural context. Co-infection of Candidatus Piscichlamydia trichopodus (order Chlamydiales) and Henneguya sp (Myxosporea, Myxobolidae) in snakeskin gourami Trichopodus pectoralis (Regan 1910) The present study describes a simultaneous infection of a novel Chlamydia-like organism (CLO) with a Myxozoa parasite, Henneguya sp. in snakeskin gourami Trichopodus pectoralis in Thailand. A new CLO is proposed "Candidatus Piscichlamydia trichopodus" (CPT) based on 16S rRNA phylogenetic analysis. Systemic intracellular CPT infection was confirmed by histological examination, in situ hybridization, PCR assay, and sequencing of 16S rRNA. This novel pathogen belongs to the order Chlamydiales but differs in certain aspects from other species. The histopathological changes associated with CPT infection were different from the typical pathological lesions of epitheliocystis caused by previously known CLO. Unlike other CLO, CPT localized in the connective tissue rather than in the epithelial cells and formed smaller clumps of intracellular bacteria that stained dark blue with hematoxylin. On the other hand, typical myxospores of the genus Henneguya with tails were observed in the gill sections. Infection with Henneguya sp. resulted in extensive destruction of the gill filaments, most likely leading to respiratory distress. Due to the frequency of co-infections and the unavailability of culture methods for CLO and Henneguya sp, it was difficult to determine which pathogens were directly responsible for the associated mortality. However, co-infections may increase the negative impact on the host and the severity of the disease. Given the commercial importance of the snakeskin gourami and its significant aquaculture potential, the findings of this study are important for further studies on disease prevention. Towards a cumulative science of vocal markers of autism: a cross-linguistic meta-analysis-based investigation of acoustic markers in American and Danish autistic children Acoustic atypicalities in speech production are argued to be potential markers of clinical features in Autism Spectrum Disorder (ASD). A recent meta-analysis highlighted shortcomings in the field, in particular small sample sizes and study heterogeneity (Fusaroli et al., 2017). We showcase a cumulative (i.e., explicitly building on previous studies both conceptually and statistically) yet self-correcting (i.e., critically assessing the impact of cumulative statistical techniques) approach to prosody in ASD to overcome these issues.

We relied on the recommendations contained in the meta-analysis to build and analyze a cross-linguistic corpus of multiple speech productions in 77 autistic and 72 neurotypical children and adolescents (>1000 recordings in Danish and US English). We used meta-analytically informed and skeptical priors, with informed priors leading to more generalizable inference. We replicated findings of a minimal cross-linguistically reliable distinctive acoustic profile for ASD (higher pitch and longer pauses) with moderate effect sizes. We identified novel reliable differences between the two groups for normalized amplitude quotient, maxima dispersion quotient, and creakiness. However, the differences were small, and there is likely no one acoustic profile characterizing all autistic individuals. We identified reliable relations of acoustic features with individual differences (age, gender), and clinical features (speech rate and ADOS sub-scores).

Besides cumulatively building our understanding of acoustic atypicalities in ASD, the study shows how to use systematic reviews and meta-analyses to guide the design and analysis of follow-up studies. We indicate future directions: larger and more diverse cross-linguistic datasets, focus on heterogeneity, self-critical cumulative approaches and open science.

Lay SummaryAutistic individuals are reported to speak in distinctive ways. Distinctive vocal production can affect social interactions and social development and could represent a noninvasive way to support the assessment of ASD. We systematically checked whether acoustic atypicalities highlighted in previous articles could be actually found across multiple recordings and two languages. We find a minimal acoustic profile of ASD: higher pitch, longer pauses, increased hoarseness and creakiness of the voice. However, there is much individual variability (by age, sex, language, and clinical characteristics). This suggests that the search for one common "autistic voice" might be naive and more fine-grained approaches are needed. Enhancing CDK4/6 inhibitor therapy for medulloblastoma using nanoparticle delivery and scRNA-seq-guided combination with sapanisertib CDK4/6 inhibitors hold promise for brain tumor treatment, but efficacy has been limited by recurrence in both preclinical models and clinical trials. To address recurrence, we tested a nanoparticle formulation of the CDK4/6 inhibitor palbociclib (POx-palbo) in mice genetically-engineered to develop SHH-driven medulloblastoma. We then analyzed medulloblastomas in mice receiving palbociclib treatment, and compared the efficacy of combining palbociclib with specific inhibitors suggested by our analysis. POx-Palbo showed reduced toxicity compared to conventional palbociclib, was tolerable in parenteral administration, improved CNS pharmacokinetics, and extended survival of mice with medulloblastoma. Recurrence, however, remained problematic as fractions of tumor cells proliferated during therapy. ScRNA-seq identified a gene expression pattern unique to proliferating medulloblastoma cells in POx-Palbo-treated mice, marked by up-regulation of the glutamate transporter Slc1a2 and down-regulation of diverse ribosomal genes. Reduced mTORC1 signaling, suggested by ribosomal suppression in POx-Palbo-treated tumors was confirmed by decreased 4EBP1 phosphorylation (p4EBP1). Further reducing mTORC1 activity by combining POx-Palbo with the mTORC1 inhibitor sapanisertib produced mutually enhancing effects, with increased suppression of both pRB and p4EBP1, and prolonged mouse survival compared to either agent alone. In contrast, targeting cell cycle progression by combining POx-Palbo with the SHH-pathway inhibitor vismodegib, or with the replication-targeting agents gemcitabine or etoposide, failed to enhance efficacy. Our data show the potential of nanoparticle formulation and scRNA-seq analysis of resistance to improve brain tumor treatment, and identify POx-palbo plus sapanisertib as effective combinatorial therapy for SHH medulloblastoma. This combination may be appropriate for testing in patients with recurrence, who need new options. Inter-tissue convergence of gene expression during ageing suggests age-related loss of tissue and cellular identity Developmental trajectories of gene expression may reverse in their direction during ageing, a phenomenon previously linked to cellular identity loss. Our analysis of cerebral cortex, lung, liver and muscle transcriptomes of 16 mice, covering development and ageing intervals, revealed widespread but tissue-specific ageing-associated expression reversals. Cumulatively, these reversals create a unique phenomenon: mammalian tissue transcriptomes diverge from each other during postnatal development, but during ageing, they tend to converge towards similar expression levels, a process we term Divergence followed by Convergence, or DiCo. We found that DiCo was most prevalent among tissue-specific genes and associated with loss of tissue identity, which is confirmed using data from independent mouse and human datasets. Further, using publicly available single-cell transcriptome data, we showed that DiCo could be driven both by alterations in tissue cell type composition and also by cell-autonomous expression changes within particular cell types. Re-distribution of antibiotic within a community is behind the unpredictable change in efficacy observed in classic laboratory assays Microbes associate in nature forming complex communities, but they are often studied in purified form. Here I show that differences in antibiotic sensitivity between co-existing bacterial species can modify the diffusion of antibiotic molecules, and alter the drug efficacy measured in conventional in vitro assays. I developed a model, validated experimentally using an artificial two-species community, showing antibiotic molecules re-distribute evenly across all species if they have similar drug sensitivity. The result is lower drug efficacy with respect to conventional in vitro assays. However, when species have different sensitivities, the re-distribution of antibiotic is uneven and most sensitive species accumulate more drug. The result is increased drug efficacy against these species. Despite this discrepancy with conventional laboratory assays, my data suggests the changes in drug efficacy are predictable and provides insight into how communities can affect the diffusion of other molecules in the environment. Analysis of Drosophila cardiac hypertrophy by micro-computerized tomography for genetic dissection of heart growth mechanisms Heart failure is often preceded by pathological cardiac hypertrophy, a thickening of the heart musculature driven by complex gene regulatory and signaling processes. The Drosophila heart has great potential as a genetic model for deciphering the underlying mechanisms of cardiac hypertrophy. However, current methods for evaluating hypertrophy of the Drosophila heart are laborious and difficult to carry out reproducibly. Here we demonstrate that micro-computerized tomography (microCT) is an accessible, highly reproducible method for non-destructive, quantitative analysis of Drosophila heart morphology and size. To validate our microCT approach for analyzing Drosophila cardiac hypertrophy, we show that expression of constitutively active Ras (Ras85DV12), previously shown to cause hypertrophy of the fly heart, results in significant thickening of both adult and larval heart walls when measured from microCT images. We then show using microCT analysis that genetic upregulation of store-operated Ca2+ entry (SOCE) driven by expression of constitutively active Stim (StimCA) or Orai (OraiCA) proteins also results in significant hypertrophy of the Drosophila heart, through a process that specifically depends on Orai Ca2+ influx channels. Intravital imaging of heart contractility revealed significantly reduced end diastolic dimensions in StimCA and OraiCA expressing hearts, consistent with the hypertrophic phenotype. These results demonstrate that increased SOCE activity is an important driver of hypertrophic cardiomyocyte growth, and demonstrate how microCT analysis combined with tractable genetic tools in Drosophila can be used to delineate molecular signaling processes that underlie cardiac hypertrophy and heart failure.

NEW AND NOTEWORTHYGenetic analysis of cardiac hypertrophy in Drosophila holds immense potential for the discovery of new therapeutic targets to prevent and treat heart failure. However, this potential has been hindered by a lack of rapid and effective methods for analysis of heart size in flies. Here we demonstrate that analysis of the Drosophila heart with micro-computerized tomography yields accurate and highly reproducible heart size measurements that can be used to efficiently analyze heart growth and cardiac hypertrophy in Drosophila. Bulk-assembly of monodisperse coacervates and giant unilamellar vesicles with programmable hierarchical complexity Compartmentalization is an essential step for the emergence of protocellular life on the early earth and the endeavor to bottom-up construct synthetic cells in the lab. Among manifold natural strategies for compartmentalization, assembly of lipid vesicles or coacervate droplets with resemblance to cell envelope or cytoplasm have emerged as two dominant paradigms. However, the spontaneous assembly of lipid vesicles or coacervates generally results in polydisperse microcompartments and lacks the ability to efficiently integrate functional building blocks towards defined complexity, which give rise to the difficulty to achieve the order and complexity towards the scope of that of life via spontaneous assembly. Herein, we show that the interplay of coacervates and colloidal particles provides a bulk-assembly approach to form monodisperse coacervates and giant unilamellar vesicles (GUVs) with programmable hierarchical complexity. The hierarchical complexity of the synthetic cell microcompartments can be delicately engineered from the control of the spatial or temporal organization of components in entity unit to manifold forms of interconnected synthetic cell consortia, achieving a compartment hierarchy analogous to the organelle-cell-tissue structure. The increase in complexity gives rise to emergent properties, for instance, collective morphology evolution of coacervate assemblages, gated permeability of GUVs without sophisticated protein machinery, and remarkable structural and functional stability of GUVs at extreme conditions. This work paves an unprecedented step to form monodisperse, hierarchical coacervates and GUVs via bulk-assembly, which provides a prebiotically plausible approach for the proper integration of inanimate matter towards the order and complexity of life and monodisperse, hierarchical microdroplets for follow-up scientific and technological applications. Protein cost minimization promotes the emergence of coenzyme redundancy Coenzymes distribute a variety of chemical moieties throughout cellular metabolism, participating in group (e.g., phosphate, acyl) and electron transfer. For a variety of reactions requiring acceptors or donors of specific resources, there often exist degenerate sets of molecules (e.g., NAD(H) and NADP(H)) that carry out similar functions. Although the physiological roles of various coenzyme systems are well established, it is unclear what selective pressures may have driven the emergence of coenzyme redundancy. Here we use genome-wide metabolic modeling approaches to decompose the selective pressures driving enzymatic specificity for either NAD(H) or NADP(H) in the metabolic network of Escherichia coli. We found that few enzymes are thermodynamically constrained to using a single coenzyme, and in principle, a metabolic network relying on only NAD(H) is feasible. However, structural and sequence analyses revealed widespread conservation of residues that retain selectivity for either NAD(H) or NADP(H), suggesting that additional forces may shape specificity. Using a model accounting for the cost of oxidoreductase enzyme expression, we found that coenzyme redundancy universally reduces the minimal amount of protein required to catalyze coenzyme-coupled reactions, inducing individual reactions to strongly prefer one coenzyme over another when reactions are near thermodynamic equilibrium. We propose that protein minimization generically promotes coenzyme redundancy, and that coenzymes typically thought to exist in a single pool (e.g., CoA) may exist in more than one form (e.g., dephospho-CoA).

Significance statementMetabolism relies on a small class of molecules (coenzymes) that serve as universal donors and acceptors of key chemical groups and electrons. Although metabolic networks crucially depend on structurally redundant coenzymes (e.g., NAD(H) and NADP(H)) associated with different enzymes, the criteria that led to the emergence of this redundancy remain poorly understood. Our combination of modeling, and structural and sequence analysis indicates that coenzyme redundancy is not essential for metabolism, but rather an evolved strategy promoting efficient usage of enzymes when biochemical reactions are near equilibrium. Our work suggests that early metabolism may have operated with fewer coenzymes, and that adaptation for metabolic efficiency may have driven the rise of coenzyme diversity in living systems. Inferring mammalian tissue-specific regulatory conservation by predicting tissue-specific differences in open chromatin BackgroundEvolutionary conservation is an invaluable tool for inferring functional significance in the genome, including regions that are crucial across many species and those that have undergone convergent evolution. Computational methods to test for sequence conservation are dominated by algorithms that examine the ability of one or more nucleotides to align across large evolutionary distances. While these nucleotide alignment-based approaches have proven powerful for protein-coding genes and some non-coding elements, they fail to capture conservation at many enhancers, distal regulatory elements that control spatio-temporal patterns of gene expression. The function of enhancers is governed by a complex, often tissue- and cell type-specific, code that links combinations of transcription factor binding sites and other regulation-related sequence patterns to regulatory activity. Thus, function of orthologous enhancer regions can be conserved across large evolutionary distances, even when nucleotide turnover is high.

ResultsWe present a new machine learning-based approach for evaluating enhancer conservation that leverages the combinatorial sequence code of enhancer activity rather than relying on the alignment of individual nucleotides. We first train a convolutional neural network model that is able to predict tissue-specific open chromatin, a proxy for enhancer activity, across mammals. Then, we apply that model to distinguish instances where the genome sequence would predict conserved function versus a loss regulatory activity in that tissue. We present criteria for systematically evaluating model performance for this task and use them to demonstrate that our models accurately predict tissue-specific conservation and divergence in open chromatin between primate and rodent species, vastly out-performing leading nucleotide alignment-based approaches. We then apply our models to predict open chromatin at orthologs of brain and liver open chromatin regions across hundreds of mammals and find that brain enhancers associated with neuron activity and liver enhancers associated with liver regeneration have a stronger tendency than the general population to have predicted lineage-specific open chromatin.

ConclusionThe framework presented here provides a mechanism to annotate tissue-specific regulatory function across hundreds of genomes and to study enhancer evolution using predicted regulatory differences rather than nucleotide-level conservation measurements. TAP: Targeting and analysis pipeline for optimization and verification of coil placement in transcranial magnetic stimulation ObjectiveTranscranial magnetic stimulation (TMS) can modulate brain function via an electric field (E-field) induced in a region of interest (ROI) in the brain. The ROI E-field can be computationally maximized and set to match a specific reference using individualized head models to find the optimal coil placement and stimulus intensity. However, the available software lacks many practical features for prospective planning of TMS interventions and retrospective evaluation of the experimental targeting accuracy.

ApproachThe TMS targeting and analysis pipeline (TAP) software uses an MRI/fMRI-derived brain target to optimize coil placement considering experimental parameters such as the subjects hair thickness and coil placement restrictions. The coil placement optimization is implemented in SimNIBS 3.2, for which an additional graphical user interface (TargetingNavigator) is provided to visualize and adjust procedural parameters. The coil optimization process also computes the E-field at the target, allowing the selection of the TMS device intensity setting to achieve a specific E-field strength. The optimized coil placement information is prepared for neuronavigation software, which supports targeting during the TMS procedure. The neuronavigation system can record the coil placement during the experiment, and these data can be processed in TAP to quantify the accuracy of the experimental TMS coil placement and induced E-field.

Main resultsTAP was demonstrated in a study consisting of three repetitive TMS sessions in five subjects. TMS was delivered by an experienced operator under neuronavigation with the computationally optimized coil placement. Analysis of the experimental accuracy from the recorded neuronavigation data indicated coil location and orientation deviations up to about 2 mm and 2{degrees}, respectively, resulting in an 8% median decrease in the target E-field magnitude compared to the optimal placement.

SignificanceTAP supports navigated TMS with a variety of features for rigorous and reproducible stimulation delivery, including planning and evaluation of coil placement and intensity selection for E-field-based dosing. Fecal immunoglobulin A (IgA) and its subclasses in systemic lupus erythematosus patients are nuclear antigen reactive and this feature correlates with gut permeability marker levels Systemic lupus erythematosus (SLE) is characterized by the production of anti-nuclear autoantibodies. Here, for the first time, we show that the abundances of gut permeability marker Zonulin and IgA1- and IgA2-subclasses are significantly higher in the fecal samples of SLE patients compared to HCs. Importantly, IgA-total, IgA1- and IgA2-subclasses from SLE patients showed higher nAg reactivity titers. Notably, we found that not only the nuclear antigen (nAg) reactive fecal IgA1:IgA2 ratio is higher in SLE patients, but also the abundance and nAg reactivity of fecal IgA and subclasses, IgA1 particularly, correlate with the fecal levels of Zonulin, which is produced primarily in the small intestine. These observations suggest that higher amounts of nAg-reactive IgA are produced, particularly, in the proximal gut as indicated by higher IgA1:IgA2 abundance and nAg reactivity ratios and correlation of these features with the Zonulin levels. Social behavioral profiling by unsupervised deep learning reveals a stimulative effect of dopamine D3 agonists on zebrafish sociality It has been a major challenge to systematically evaluate and compare how pharmacological perturbations influence social behavioral outcomes. Although some pharmacological agents are known to alter social behavior, precise description and quantification of such effects have proven difficult. The complexity of brain functions regulating sociality makes it challenging to predict drug effects on social behavior without testing in live animals, and most existing behavioral assays are low-throughput and provide only unidimensional readouts of social function. To achieve richer characterization of drug effects on sociality, we developed a scalable social behavioral assay for zebrafish named ZeChat based on unsupervised deep learning. High-dimensional and dynamic social behavioral phenotypes are automatically classified using this method. By screening a neuroactive compound library, we found that different classes of chemicals evoke distinct patterns of social behavioral fingerprints. By examining these patterns, we discovered that dopamine D3 agonists possess a social stimulative effect on zebrafish. The D3 agonists pramipexole, piribedil, and 7-hydroxy-DPAT-HBr rescued social deficits in a valproic acid-induced zebrafish autism model. The ZeChat platform provides a promising approach for dissecting the pharmacology of social behavior and discovering novel social-modulatory compounds. Embryonic exposure to cannabidiol disrupts active neural circuits, an effect increased by Δ9-tetrahydrocannabinol and involving CB1R and CB2R in zebrafish In the light of legislative changes and the widespread use of cannabis as a recreational and medicinal drug, delayed effects of cannabis upon brief exposure during embryonic development are of high interest as early pregnancies often go undetected. Here, zebrafish embryos were exposed to cannabidiol (CBD) and {Delta}-9-tetrahydrocannabinol l (THC) until the end of gastrulation (1-10 hours post-fertilization) and analyzed later in development (4-5 days post-fertilization). In order to measure neural activity, we implemented CaMPARI (Calcium-Modulated Photoactivatable Ratiometric Integrator) and optimized the protocol for a 96-well format complemented by locomotor analysis. Our results revealed that neural activity was decreased by CBD more than THC. At higher doses, both cannabinoids could dramatically reduce neural activity and locomotor activity. Interestingly, the decrease was more pronounced when CBD and THC were combined. At the receptor level, CBD-mediated reduction of locomotor activity was partially prevented using cannabinoid type 1 and 2 receptor inhibitors. Overall, we report that CBD toxicity occurs via two cannabinoid receptors and is synergistically enhanced by THC exposure to negatively impact neural activity late in larval development. Future studies are warranted to reveal other cannabinoids and receptors involved in this pathway to understand the subsequent health implications of cannabis consumption on fetal development. Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. SARS-CoV-2 Beta Variant Substitutions Alter Spike Glycoprotein Receptor Binding Domain Structure and Stability The emergence of severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) and the subsequent COVID-19 pandemic has visited a terrible cost on the world in the forms of disease, death, and economic turmoil. The rapid development and deployment of extremely effective vaccines against SARS-CoV-2 have seemingly brought within reach the end of the pandemic. However, the virus has acquired mutations; and emerging variants of concern (VOC) are more infectious and reduce the efficacy of existing vaccines. While promising efforts to combat these variants are underway, the evolutionary pressures leading to these variants are poorly understood. To that end, here we have studied the effects on the structure and function of the SARS-CoV-2 spike glycoprotein receptor-binding domain of three amino-acid substitutions found in several variants of concern, including alpha (B.1.1.7), beta (B.1.351), and gamma (P.1). We found that these substitutions alter the RBD structure, stability, and ability to bind to ACE2, in such a way as to possibly have opposing and compensatory effects. These findings provide new insights into how these VOC may have been selected for infectivity while maintaining the structure and stability of the receptor binding domain. Experimental sexual selection affects the evolution of physiological and life history traits Sexual selection and sexual conflict are expected to affect all aspects of the phenotype, not only traits that are directly involved in reproduction. Here, we show coordinated evolution of multiple physiological and life history traits in response to long-term experimental manipulation of the mating system in populations of Drosophila pseudoobscura. Development time was extended under polyandry relative to monogamy in both sexes, potentially due to higher investment in traits linked to sexual selection and sexual conflict. Individuals (especially males) evolving under polyandry had higher metabolic rates and locomotor activity than those evolving under monogamy. Polyandry individuals also invested more in metabolites associated with increased endurance capacity and efficient energy metabolism and regulation, namely lipid and glycogen. Finally, polyandry males were less desiccation- and starvation-resistant than monogamy males, suggesting trade-offs between resistance and sexually selected traits. Our results provide experimental evidence that mating systems can impose selection that influences the evolution of non-sexual phenotypes such as development, activity, metabolism, and nutrient homeostasis. Live imaging YAP signaling in mouse embryo development YAP protein is a critical regulator of mammalian embryonic development. By generating a near-infrared fusion YAP reporter mouse line, we have achieved high-resolution live imaging of YAP localization during mouse embryonic development. We have validated the reporter by demonstrating its predicted responses to blocking LATs kinase activity or blocking cell polarity. By time lapse imaging preimplantation embryos, we revealed a mitotic reset behavior of YAP nuclear localization. We also demonstrated deep tissue live imaging in post-implantation embryos and revealed an intriguing nuclear YAP pattern in migrating cells. The YAP fusion reporter mice and imaging methods will open new opportunities for understanding dynamic YAP signaling in vivo in many different situations. Association between land use, land cover, plant genera, and pollinator abundance in mixed-use landscapes Pollinators are globally threatened by land-use change, but its effect varies depending on the taxa and the intensity of habitat degradation. However, pollinator-landscape studies typically focus on regions of intensive human activities and on a few focal species. Evaluating pollinator responses in landscapes with moderate land-use changes and on multiple pollinator groups would therefore fill an important knowledge gap. This study aims to determine the predictive capacity and effect of habitat characteristics on the relative abundance of multiple pollinator groups in mixed-use landscapes. To do this, we collected field data on the relative abundance of nectivorous birds, bees, beetles, and butterflies across the Tasman Peninsula (Tasmania, Australia). We then applied Random Forests to resolve the effects of land use (protected areas, plantation, and pasture), land cover at different radii (100 m and 2000 m), and plant genera on pollinator abundance. Overall, land cover and plant genera were more important predictors of pollinator abundance than land use. And the effect of land use, land cover, and plant genera varied depending on the pollinating group. Pollinator groups were associated with a range of plant genera, with the native genera Acacia, Leptospermum, Leucopogon, Melaleuca, Pomaderris, and Pultenaea being among the most important predictors. Our results highlight that one size does not fit all--that is pollinator response to different landscape characteristics vary, emphasise the importance of considering multiple habitat factors to manage and support a dynamic pollinator community, and demonstrates how land management can be informed using predictive modelling. Differential metabolism of arsenicals regulates Fps1-mediated arsenite transport Arsenic is an environmental toxin that exists mainly as pentavalent arsenate and trivalent arsenite. Both forms activate the yeast SAPK Hog1, but with different consequences. We describe a mechanism by which cells distinguish between these arsenicals through one-step metabolism to differentially regulate the bidirectional glycerol channel Fps1, an adventitious port for arsenite. Cells exposed to arsenate reduce it to thiol-reactive arsenite, which modifies a set of cysteine residues in target proteins; whereas cells exposed to arsenite metabolize it to methylarsenite, which modifies an additional set of cysteine residues. Hog1 becomes arsenylated, which prevents it from closing Fps1. However, this block is overcome in cells exposed to arsenite through methylarsenylation of Acr3, an arsenite efflux pump that we found also regulates Fps1 directly. This adaptation allows cells to restrict arsenite entry through Fps1, but also allows its exit when produced from arsenate exposure. These results have broad implications for understanding how SAPKs activated by diverse stressors can drive stress-specific outputs.

Summary (for JCB table of contents)

Jongmin Lee, David E. Levin

Lee and Levin investigate the question of how different stressors can drive divergent outputs from an active SAPK. The work describes the mechanism by which two forms of arsenic that both activate the yeast SAPK Hog1 differentially regulate the glycerol channel Fps1. MTG16 (CBFA2T3) regulates colonic epithelial differentiation, colitis, and tumorigenesis by repressing E protein transcription factors Aberrant epithelial differentiation and regeneration contribute to colon pathologies including inflammatory bowel disease (IBD) and colitis-associated cancer (CAC). MTG16 (CBFA2T3) is a transcriptional corepressor expressed in the colonic epithelium. MTG16 deficiency in mice exacerbates colitis and increases tumor burden in CAC, though the underlying mechanisms remain unclear. Here, we identified MTG16 as a central mediator of epithelial differentiation, promoting goblet and restraining enteroendocrine cell development in homeostasis and enabling regeneration following dextran sulfate sodium (DSS)-induced colitis. Transcriptomic analyses implicated increased E box-binding transcription factor (E protein) activity in MTG16-deficient colon crypts. Using a novel mouse model with a point mutation that disrupts MTG16:E protein complex formation (Mtg16P209T), we established that MTG16 exerts control over colonic epithelial differentiation and regeneration by repressing E protein-mediated transcription. Mimicking murine colitis, MTG16 expression was increased in biopsies from patients with active IBD compared to unaffected controls. Finally, uncoupling MTG16:E protein interactions only partially phenocopied the enhanced tumorigenicity of Mtg16-/- colon in the azoxymethane(AOM)/DSS-induced model of CAC, indicating that MTG16 protects from tumorigenesis through additional mechanisms. Collectively, our results demonstrate that MTG16, via its repression of E protein targets, is a key regulator of cell fate decisions during colon homeostasis, colitis, and cancer.

Graphical Abstract

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org.highwire.dtl.DTLVardef@21948borg.highwire.dtl.DTLVardef@9f60eeorg.highwire.dtl.DTLVardef@194b9fdorg.highwire.dtl.DTLVardef@13eb1a7_HPS_FORMAT_FIGEXP  M_FIG C_FIG Negative DNA supercoiling makes protein-mediated looping deterministic and ergodic within the bacterial doubling time Protein-mediated DNA looping is fundamental to gene regulation and such loops occur stochastically in purified systems. Additional proteins increase the probability of looping, but these probabilities maintain a broad distribution. For example, the probability of lac repressor-mediated looping in individual molecules ranged 0-100%, and individual molecules exhibited representative behavior only in observations lasting an hour or more. Titrating with HU protein progressively compacted the DNA without narrowing the 0-100% distribution. Increased negative supercoiling produced an ensemble of molecules in which all individual molecules more closely resembled the average. Furthermore, in only twelve minutes of observation, well within the doubling time of the bacterium, most molecules exhibited the looping probability of the ensemble. DNA supercoiling, an inherent feature of all genomes, appears to impose time-constrained, emergent behavior on otherwise random molecular activity. First detection of herpesvirus and prevalence of mycoplasma infection in free-ranging Hermann s tortoises (Testudo hermanni), and in potential pet vectors AO_SCPLOWBSTRACTC_SCPLOWTwo types of pathogens cause highly contagious upper respiratory tract diseases (URTD) in Chelonians: testudinid herpesviruses (TeHV) and a mycoplasma (Mycoplasma agassizii). In captivity, these infections are frequent and can provoke outbreaks. Pet trade generates international flow of tortoises, often without sanitary checking; individuals intentionally or accidentally released in the wild may spread pathogens. A better understanding of the transmission of infectious agents from captivity to wild tortoises is needed. Many exotic individuals have been introduced in populations of the endangered western Hermanns tortoise (Testudo hermanni hermanni), notably spur-thighed tortoises (Testudo graeca). We assessed the presence of TeHV and mycoplasma in native western Hermanns tortoises and in potential pet vectors in south-eastern France. Using a large sample (N=572 tortoises), this study revealed, by PCR, the worrying presence of herpesvirus in 7 free-ranging individuals (3 sub-populations). Additionally, Mycoplasma agassizii was detected, by PCR, in 15 of the 18 populations sampled with a frequency ranging from 3.4% (1 of 29 tortoises) to 25% (3 of 12 tortoises). Exotic spur-thighed tortoises showed high frequency of mycoplasma infection in captivity (18.2%) and in individuals (50%) found in native Hermanns tortoise sub-populations, suggesting that this species could be a significant vector. The paucity of information of TeHV on European tortoise URTD in natural settings, especially in combination with mycoplasma, prompts for further studies. Indeed, sick tortoises remain concealed and may not be easily detected in the field. Our results indicate a good health for most infected tortoise but it should be screened in the field as well as in captivity. Isoprenoid biosynthesis regulation in poplars by methylerythritol phosphate and mevalonic acid pathways The isoprenoids found in plants are extremely important to survive with various human applications, such as flavoring, fragrance, dye, pharmaceuticals, and biomass used for biofuels. Methylerythritol phosphate (MEP) and mevalonic acid (MVA) pathways are critical in plants, responsible for isoprenoid biosynthesis. 1-deoxy-D-xylulose5-phosphate synthase (DXS) and 1-deoxy-D-xylulose5-phosphate reductoisomerase (DXR) catalyze the rate-limiting steps in the MEP pathway, while 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) catalyzes the rate-limiting step in the MVA pathway. Here, we showed while PtHMGR overexpressors (OEs) exhibited different MEP- and MVA-related gene expressions compared with non-transgenic poplars (NT), the PtDXR-OEs revealed upregulated MEP-related and downregulated MVA-related gene expressions. PtDXR and PtHMGR overexpressions caused changes in MVA-derived trans-zeatin-riboside, isopentenyl adenosine, castasterone, and 6-deoxocastasterone well as MEP-derived carotenoids and gibberellins. In PtHMGR-OEs, the accumulated geranyl diphosphate synthase (GPS) and geranyl pyrophosphate synthase (GPPS) transcript levels in the MEP pathway led to an accumulation of MEP-derived isoprenoids. In contrast, upregulation of farnesyl diphosphate synthase (FPS) expression in the MVA pathway contributed to increased levels of MVA-derived isoprenoids. In addition, PtHMGR-OEs increased MEP-related GPS and GPPS transcript levels, expanded MEP-derived isoprenoid levels, changed FPS transcript levels, and affected MVA-derived isoprenoid yields. These results demonstrate the contribution of MVA and MEP pathways regulating isoprenoid biosynthesis in poplars.