Potential autoimmunity resulting from molecular mimicry between SARS-CoV-2 Spike and human proteins SARS-CoV-2 causes COVID-19, a disease curiously resulting in varied symptoms and outcomes, ranging from asymptomatic to fatal. Autoimmunity due to cross-reacting antibodies resulting from molecular mimicry between viral antigens and host proteins may provide an explanation. We computationally investigated molecular mimicry between SARS-CoV-2 Spike and known epitopes. We discovered molecular mimicry hotspots in Spike and highlight two examples with tentative autoimmune potential and implications for understanding COVID-19 complications. We show that a TQLPP motif in Spike and thrombopoietin shares similar antibody binding properties. Antibodies cross-reacting with thrombopoietin may induce thrombocytopenia, a condition observed in COVID-19 patients. Another motif, ELDKY, is shared in multiple human proteins such as PRKG1 and tropomyosin. Antibodies cross-reacting with PRKG1 and tropomyosin may cause known COVID-19 complications such as blood-clotting disorders and cardiac disease, respectively. Our findings illuminate COVID-19 pathogenesis and highlight the importance of considering autoimmune potential when developing therapeutic interventions to reduce adverse reactions.
LinearTurboFold: Linear-Time Global Prediction of Conserved Structures for RNA Homologs with Applications to SARS-CoV-2 The constant emergence of COVID-19 variants reduces the effectiveness of existing vaccines and test kits. Therefore, it is critical to identify conserved structures in SARS-CoV-2 genomes as potential targets for variant-proof diagnostics and therapeutics. However, the algorithms to predict these conserved structures, which simultaneously fold and align multiple RNA homologs, scale at best cubically with sequence length, and are thus infeasible for coronaviruses, which possess the longest genomes ([~]30,000 nt) among RNA viruses. As a result, existing efforts on modeling SARS-CoV-2 structures resort to single sequence folding as well as local folding methods with short window sizes, which inevitably neglect long-range interactions that are crucial in RNA functions. Here we present LinearTurboFold, an efficient algorithm for folding RNA homologs that scales linearly with sequence length, enabling unprecedented global structural analysis on SARS-CoV-2. Surprisingly, on a group of SARS-CoV-2 and SARS-related genomes, LinearTurbo-Folds purely in silico prediction not only is close to experimentally-guided models for local structures, but also goes far beyond them by capturing the end-to-end pairs between 5 and 3 UTRs ([~]29,800 nt apart) that match perfectly with a purely experimental work. Furthermore, LinearTurboFold identifies novel conserved structures and conserved accessible regions as potential targets for designing efficient and mutation-insensitive small-molecule drugs, antisense oligonucleotides, siRNAs, CRISPR-Cas13 guide RNAs and RT-PCR primers. LinearTurboFold is a general technique that can also be applied to other RNA viruses and full-length genome studies, and will be a useful tool in fighting the current and future pandemics.

Significance StatementConserved RNA structures are critical for designing diagnostic and therapeutic tools for many diseases including COVID-19. However, existing algorithms are much too slow to model the global structures of full-length RNA viral genomes. We present LinearTurboFold, a linear-time algorithm that is orders of magnitude faster, making it the first method to simultaneously fold and align whole genomes of SARS-CoV-2 variants, the longest known RNA virus ([~]30 kilobases). Our work enables unprecedented global structural analysis and captures long-range interactions that are out of reach for existing algorithms but crucial for RNA functions. LinearTurboFold is a general technique for full-length genome studies and can help fight the current and future pandemics.
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. 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. Sufu regulation of Hedgehog signaling in P19 cells is required for proper astrocyte differentiation Hedgehog signaling is essential for vertebrate development, however, less is known about the negative regulators that influence this pathway during the differentiation of cell fates. Using the mouse P19 embryonal carcinoma cell model, Suppressor of Fused (SUFU), a negative regulator of the Hedgehog pathway, was investigated during retinoic acid-induced neural differentiation. We found Hedgehog signaling was activated in the early phase of neural differentiation but was inactive during terminal differentiation of neurons and astrocytes. This early activation was required for neural differentiation however, it alone was not sufficient to induce neural lineages. SUFU, which regulates signaling at the level of GLI, remained relatively unchanged during the differentiation process, but its loss through CRISPR-Cas9 gene editing resulted in ectopic expression of Hedgehog target genes. Interestingly, these SUFU-deficient cells were unable to differentiate in the absence of retinoic acid, but when differentiated using retinoic acid, they showed delayed and decreased astrocyte differentiation; neuron differentiation was unaffected. Ectopic activation of Hh target genes in SUFU-deficient cells remained throughout retinoic acid-induced differentiation and this phenotype was accompanied by the loss of GLI3, despite the presence of the Gli3 message. Thus, the study indicates the proper timing and proportion of astrocyte differentiation requires SUFU, and its normal regulation of GLI3 to maintain Hh signaling in an inactive state. Changes in Behavior and Salivary Serum Amyloid A in cows with Subclinical Mastitis The aim of this study was to identify detailed changes in behavior, and in salivary serum amyloid A (SAA), associated with subclinical mastitis. This included standard sickness behaviors (which form part of an adaptive response to conserve energy, minimize heat loss and facilitate recovery following infection and inflammation) and lesser-studied behaviors, that are biologically predicted to change at low-level infection, and therefore particularly relevant for detecting subclinical mastitis (here labelled  luxury behaviors). SAA is a physiological marker of systemic inflammation, with levels in milk and serum already known to increase during subclinical mastitis. We investigated whether the same was true for SAA in cow saliva. Data were collected for n = 34 commercial barn-housed dairy cows, tested in 17 matched-pairs. Each pair comprised a cow with subclinical mastitis (SCM) and a healthy control (CTRL), identified using somatic cell count (SCC) (SCM: SCC >200 x1000 cells/ml; CTRL: SCC <100 x1000 cells/ml). SCM cows were selected for study ad-hoc, at which point they were paired with a CTRL cow, based upon parity and calving date; consequently, the full data set was accrued over several months. Data were collected for each pair over 3 days: SCC (l4:00-l5:00h) Day 1; behavior (24h from 00:00h) Day 2; salivary serum amyloid-A (SAA) Day 3. We report, for the first time, that an increase in salivary SAA occurs during subclinical mastitis; SAA was higher in SCM cows and demonstrated a positive (weak) correlation with SCC. The behavioral comparisons revealed that SCM cows had reductions in activity (behavioral transitions and distance moved), social exploration, social reactivity (here: likelihood to move away/be displaced following receipt of agonism), performance of social grooming and head butts, and the receipt of agonistic non-contact challenges. In addition, SCM cows received more head swipes, and spent a greater proportion of time lying with their head on their flank than CTRL cows. SCM cows also displayed an altered feeding pattern; they spent a greater proportion of feeding time in direct contact with two conspecifics, and a lower proportion of feeding time at self-locking feed barriers, than CTRL cows. Behavioral measures were found to correlate, albeit loosely, with serum SAA in a direction consistent with predictions for sickness behavior. These included positive correlations with both lying duration and the receipt of all agonistic behavior, and negative correlations with feeding, drinking, the performance of all social and all agonistic behavior, and social reactivity. We conclude that changes in salivary SAA, social behavior, and activity offer potential in the detection of subclinical mastitis and recommend further investigation to substantiate and refine our findings. Low-level Action Schemas Support Gaze Guidance Behavior for Action Planning and Execution in Novel Tasks AO_SCPLOWBSTRACTC_SCPLOWEye movements in the natural environment have primarily been studied for over-learned activities such as tea-making, sandwich-making, and hand-washing, which have a fixed sequence of actions associated with them. These studies indicate an interplay of low-level cognitive schemas that facilitate task completion. However, it is unclear if this strategy is also in play when the task is novel and a sequence of actions must be planned. Here, to study attention mechanisms, we recorded gaze and body movements in a naturalistic sorting task. In a virtual environment, subjects moved objects on a life-size shelf to achieve a given order. To study the action planning and execution related gaze guidance behavior we also controlled the complexity of the tasks, i.e. the objects were to be sorted based on a single or two different object features. We show that subjects are close to optimal while performing the easy tasks and are more sub-optimal while performing the more complex tasks. Fixations aligned with action onset showed gaze is tightly coupled with the action sequence and less so with task complexity. Task complexity elicited differences at specific time points where oculomotor behavior could be categorized into searching, guiding, checking fixations. Task complexity also affected the scan-paths on the task-relevant regions of interest during action planning and execution where subjects exhibit a greater search and action monitoring behavior in more complex tasks. Furthermore, task complexity also affected the timing of first fixations on the task-relevant regions of interest for action planning but not for action execution. Finally, the scan paths and the temporal sequence of first fixations suggest a just-in-time strategy of gaze allocation. Based on the spatio-temporal features of gaze control, we can conclude that eye movements during novel tasks correspond closely to low-level cognitive schemas that support the production and execution of actions.

Author SummaryEye movements in the natural environment have primarily been studied for over-learned and habitual everyday activities (tea-making, sandwich-making, hand-washing) which have a fixed sequence of actions associated with them. These studies typically categorize eye movements related to low level action plans of locating the object of interest, directing the body or hand to the object, and monitoring the action execution. In this study, we were interested in generalizing these task-oriented eye movements for novel tasks that are not associated with an inherent action sequence. To that end, we asked subjects to sort objects based on object features on a life-size shelf in a virtual environment as we recorded their eye and body movements. We then analyzed gaze features relative to the action onset i.e. when the hand made contact with an object. By only studying the sequence of fixations based on past, current and next actions, we aimed to elucidate the general characteristics of gaze behavior relative to action planning and execution and independent of object identity. With our analysis, we could categorize fixations into searching, guiding, monitoring fixations in a data-driven manner. Based on the spatio-temporal features of gaze control, we can conclude that novel tasks predominantly elicit eye movements corresponding to low-level action plans that support the production and execution of actions. Obesity affects mitochondrial metabolism and proliferative potential of equine endometrial progenitor cells. The study aimed to investigate the influence of obesity on cellular features of equine endometrial progenitor cells (Eca EPCs), including viability, proliferation capacity, mitochondrial metabolism, and oxidative homeostasis. Eca EPCs derived from non-obese (Non-OB) and obese (OB) mares were characterized by cellular phenotype and multipotency. Obesity-induced changes in the activity of Eca EPCs include the decline of their proliferative activity, clonogenic potential, mitochondrial metabolism and enhanced oxidative stress. Eca EPCs isolated from obese mares were characterized by an increased occurrence of early apoptosis, loss of mitochondrial dynamics, and senescence-associated phenotype. Attenuated metabolism of Eca EPCs OB was related to increased expression of pro-apoptotic markers (CASP9, BAX, P53, P21), enhanced expression of OPN, PI3K and AKT, simultaneously with decreased signaling stabilizing cellular homeostasis (including mitofusin, SIRT1, FOXP3). Obesity alters functional features and the self-renewal potential of endometrial progenitor cells. The impaired cytophysiology of progenitor cells from obese endometrium predict lower regenerative capacity if used as autologous transplants. Distributed brain co-processor for tracking electrophysiology and behavior during electrical brain stimulation Early implantable epilepsy therapy devices provided open-loop electrical stimulation without brain sensing, computing, or an interface for synchronized behavioral inputs from patients. Recent epilepsy stimulation devices provide brain sensing but have not yet developed analytics for accurately tracking and quantifying behavior and seizures. Here we describe a distributed brain co-processor providing an intuitive bi-directional interface between patient, implanted neural stimulation and sensing device, and local and distributed computing resources. Automated analysis of continuous streaming electrophysiology is synchronized with patient reports using a hand-held device and integrated with distributed cloud computing resources for quantifying seizures, interictal epileptiform spikes, and patient symptoms during therapeutic electrical brain stimulation. The classification algorithms for interictal epileptiform spikes and seizures were developed and parameterized using long-term ambulatory data from 9 humans and 8 canines with epilepsy, and then implemented prospectively in out-of-sample testing in 2 pet canines and 4 humans with drug resistant epilepsy living in their natural environments. Territory-level temperature influences breeding phenology and reproductive output in three forest passerine birds Temperature plays an important role in determining breeding phenology of temperate birds, with higher spring temperatures associated with earlier breeding. However, the effect of localised territory-scale temperature variations is poorly understood, with relationships between temperature and breeding phenology mostly studied using coarse-grained climatic indices. Here, we interpolate spring temperatures recorded at 150 m2 grid intersections encompassing 417 ha of forest to examine the influence of territory-scale temperature, and its interaction with mean annual temperature, on territory selection, breeding phenology, clutch size and fledgling success for three co-occurring single-brooded passerine birds using data from 672 nests over four years. All species exhibited significant trends in reproductive traits associated with territory-scale temperature. Pied flycatchers Ficedula hypoleuca settled in cooler territories first, where they raised more fledglings. Blue tits Cyanistes caeruleus laid larger clutches in warmer territories in warm years and always laid earlier at warmer territories irrespective of annual temperature variation. Contrastingly, pied flycatcher and wood warbler Phylloscopus sibilatrix breeding phenology was earlier at warmer territories in cool years and cooler territories in warm years, with wood warbler clutch size responding similarly to this interaction. Greater previous breeding experience and increased higher rates of historical territory occupancy (territory quality) also predicted earlier breeding phenology and higher fledging success for pied flycatchers. We suggest that the migratory pied flycatcher and wood warbler are best synchronised with their prey availability in cooler years at a local population level while resident blue tits match local phenology across all years, which is potentially advantageous under warmer predicted climate change scenarios. We conclude that temperature at the territory scale can be an important driver of settlement and breeding phenology and influence reproductive traits. Crowdsourced identification of multi-target kinase inhibitors for RET- and TAU- based disease: the Multi-Targeting Drug DREAM Challenge A continuing challenge in modern medicine is the identification of safer and more efficacious drugs. Precision therapeutics, which have one molecular target, have been long promised to be safer and more effective than traditional therapies. This approach has proven to be challenging for multiple reasons including lack of efficacy, rapidly acquired drug resistance, and narrow patient eligibility criteria. An alternative approach is the development of drugs that address the overall disease network by targeting multiple biological targets ( polypharmacology). Rational development of these molecules will require improved methods for predicting single chemical structures that target multiple drug targets. To address this need, we developed the Multi-Targeting Drug DREAM Challenge, in which we challenged participants to predict single chemical entities that target pro-targets but avoid anti-targets for two unrelated diseases: RET-based tumors and a common form of inherited Tauopathy. Here, we report the results of this DREAM Challenge and the development of two neural network-based machine learning approaches that were applied to the challenge of rational polypharmacology. Together, these platforms provide a potentially useful first step towards developing lead therapeutic compounds that address disease complexity through rational polypharmacology.

Author SummaryMany modern drugs are developed with the goal of modulating a single cellular pathway or target. However, many drugs are, in fact,  dirty; they target multiple cellular pathways or targets. This phenomenon is known as multi-targeting or polypharmacology. While some strive to develop  cleaner therapeutics that eliminate secondary targets, recent work has shown that multi-targeting therapeutics have key advantages for a variety of diseases. However, while multi-targeting drugs that affect a precisely-defined profile of targets may be more effective, it is difficult to computationally predict which molecules have desirable target profiles. Here, we report the results of a competitive crowdsourcing project (the Multi-Targeting Drug DREAM Challenge), where we challenged participants to predict chemicals that have desired target profiles for cancer and neurodegenerative disease. GCN5L1 promotes diastolic dysfunction by inhibiting cardiac pyruvate oxidation Left ventricular diastolic dysfunction is a structural and functional condition that precedes the development of heart failure with preserved ejection fraction (HFpEF). The etiology of diastolic dysfunction includes alterations in fuel substrate metabolism that negatively impact cardiac bioenergetics, and may precipitate the eventual transition to heart failure. To date, the molecular mechanisms that regulate early changes in fuel metabolism leading to diastolic dysfunction remain unclear. In this report, we use a diet-induced obesity model and quantitative acetylproteomics in aged mice to show that inhibitory lysine acetylation of the pyruvate dehydrogenase (PDH) complex promotes energetic deficits and diastolic dysfunction in mouse hearts. Cardiomyocyte-specific deletion of the mitochondrial lysine acetylation regulatory protein GCN5L1 prevented hyperacetylation of the PDH complex subunit PDHA1, allowing aged obese mice to continue using pyruvate as a bioenergetic substrate in the heart. Our findings suggest that changes in mitochondrial protein lysine acetylation represent a key metabolic component of diastolic dysfunction that precedes the development of heart failure. Estrogen-related receptor alpha and Rplp1-dependent translation coordinately regulate starvation response and decrease NASH progression Although general translation declines during fasting, maintaining the translation of a subset or proteins is necessary for metabolic homeostasis and cell viability. Using unbiased proteome analysis of hepatic cells during starvation, we identified a novel pathway in which Esrra-mediated transcription of Rplp1-dependent translation of lysosomal proteins declined during early starvation and recovered after prolonged starvation to restore autophagy-lysosome function. Interestingly, hepatic Esrra-Rplp1-dependent translation rate of lysosomal proteins also was impaired in patients and mice with non-alcoholic steatohepatitis (NASH), and translational response to starvation was dysregulated in mice with NASH. Remarkably, activation of Esrra pharmacologically, genetically, or by alternate day fasting restored protein translation, increased expression of lysosomal proteins, induced autophagy, and reduced lipotoxicity, inflammation, and fibrosis in cell culture and in vivo models of NASH. Thus, hepatic Esrra is essential for ribosome-dependent translation of lysosomal proteins during starvation, and prevention of lipotoxicity and progression in NASH.

Lay summaryFasting for weight loss improves non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH); however, the mechanism is not well understood. Our study shows that a nuclear protein, estrogen related receptor alpha (Esrra), increases ribosome-mediated translation of autophagy and lysosome proteins during chronic starvation to maintain essential metabolic pathways for cell survival. Surprisingly, this translational pathway is impaired during NASH with reduced lysosome-autophagy activity accompanied by increased inflammation and fibrosis gene expression in the liver. Pharmacologic, genetic and dietary activation of Esrra decreases lipid-mediated toxicity in liver cells as well as inflammation and fibrosis in livers from mice with NASH. These findings suggest that the Esrra-ribosome-lysosome pathway is important for liver response to fasting and NASH and thus may be a good therapeutic target for the treatment of NASH. Regulatory perturbations of ribosome allocation optimize the growth proteome with a trade-off in adaptation capacity Bacteria regulate their cellular resource allocation to enable fast growth-adaptation to a variety of environmental niches. We studied the ribosomal allocation, growth and expression profiles of two sets of fast-growing mutants of Escherichia coli K-12 MG1655 in glucose minimal medium. Mutants with only 3 of the seven copies of ribosomal RNA operons grew faster than the wild-type strain in minimal media and show similar phenotype to previously studied rpoB mutants. Higher growth rates due to increased ribosome content affected resource allocation. Expression profiles of fast-growing mutants shared downregulation of hedging functions and upregulated growth functions. Mutants showed longer diauxic shifts and reduced activity of gluconeogenic promoters during glucose-acetate shifts, suggesting reduced availability of the RNA Polymerase for expressing hedging proteome. These results show that the regulation of ribosomal allocation underlies the growth/hedging phenotypes obtained from laboratory evolution experiments. We show how two different regulatory perturbations (rRNA promoters or rpoB mutations) reshape the proteome for growth with a concomitant fitness cost

HighlightsMutants with only 3 ribosomal operons grow faster than wild-type in minimal medium

 {Delta}4 rrn and rpoB mutants share phenotypic traits

Faster growth of mutants is achieved by increased ribosome content

Fast-growing mutants display reduced hedging expression and adaptation trade-offs

Despite similar ribosomal content in rich medium the mutants present growth defects Bacteria swarm-mediated phage transportation disrupts a biofilm inherently protected from phage penetration The treatment of chronic bacterial infections by phages has shown promise in combating antimicrobial resistance. A typical phage particle is at least an order of magnitude larger than an antibiotic molecule. Hence, phages diffuse slower than antibiotics, and can also get trapped in the polymeric mesh of biofilm matrix. By tracking fluorescently labeled lambda phages that do not infect Capnocytophaga gingivalis, a bacterium abundant in the human oral microbiota, we demonstrate active phage transportation by a C. gingivalis swarm. As a result, the rate of disruption of the prey of lambda phage i.e., an Escherichia coli colony, increases 10 times. C. gingivalis drills tunnels within a curli fiber containing E. coli biofilm and increase the efficiency of phage penetration. This provides evidence for a novel mechanism of phage-bacterial warfare. Structure of Infective Getah Virus at 2.8 A-resolution Determined by Cryo-EM Getah virus (GETV), a member of genus alphavirus, is a mosquito-borne pathogen that can cause pyrexia and reproductive losses in animals. Although antibodies to GETV have been found in over 10% of healthy people, there are no reports of clinical symptom associated with GETV. The biological and pathological properties of GETV are largely unknown. Here, we present the structure of infective GETV at a resolution of 2.8 [A] with the capsid protein and the envelope glycoproteins E1 and E2. We have identified numerous glycosylation and S-acylation sites in E1 and E2. The surface-exposed glycans revealed their impact on the viral immune evasion and host cell invasion. The S-acylation sites involve in stabilizing the transmembrane assembly. In addition, a cholesterol and a phospholipid molecule are observed in a transmembrane hydrophobic pocket, together with two more cholesterols surrounding the pocket. The structural information will assist structure-based antiviral and vaccine design. Context-dependence in the symbiosis between Dictyostelium discoideum and Paraburkholderia Symbiotic interactions change with environmental context. Measuring these context-dependent effects in hosts and symbionts is critical to determining the nature of symbiotic interactions. We investigated context-dependence in the symbiosis between social amoeba hosts and their inedible Paraburkholderia bacterial symbionts, where the context is the abundance of host food bacteria. Paraburkholderia have been shown to harm hosts dispersed to food-rich environments, but aid hosts dispersed to food-poor environments by allowing hosts to carry food bacteria. Through measuring symbiont density and host spore production, we show that this food context matters in three other ways. First, it matters for symbionts, who suffer a greater cost from competition with food bacteria in the food-rich context. Second, it matters for host-symbiont conflict, changing how symbiont density negatively impacts host spore production. Third, data-based simulations show that symbiosis often provides a long-term fitness advantage for hosts after rounds of growth and dispersal in variable food-contexts, especially when conditions are harsh with little food. These results show how food context can have many consequences for the Dictyostelium-Paraburkholderia symbiosis and that both sides can frequently benefit.

Impact StatementMany organisms form symbiotic relationships with other species. These symbioses often exhibit context-dependence, where the sign or magnitude of one partners effect on the other will change in different environments. Context-dependent effects make it difficult to assign interactions to categories like mutualisms or antagonisms because they involve both benefits and costs depending on the environment. However, in some cases, accounting for context-dependence can clarify an interaction so that it more easily fits a mutualism or antagonism. We investigated context-dependence using the symbiosis between Dictyostelium discoideum and two symbiotic Paraburkholderia species. In this symbiosis, Paraburkholderia bacteria allow hosts to carry food bacteria to food-poor contexts, where hosts rarely survive without food, but reduce host fitness in the more hospitable food-rich contexts. The effect of food context on Paraburkholderia symbionts is unknown. We show that Paraburkholderia symbionts are also affected by this context, through facing reduced competition after being dispersed by hosts to food-poor contexts. We also identify a new way that symbionts affect hosts, where symbiont density reduces host fitness, but less so in food-poor contexts. Finally, we use simulations to show that infected hosts benefit in the long-term across variable food contexts, especially in the harshest environments with little food. These results show that context-dependence in symbiosis can have many consequences for hosts and symbionts, though in general for D. discoideum and Paraburkholderia, both are likely to benefit. Omicron BA.2 specifically evades broad sarbecovirus neutralizing antibodies Omicron sub-lineage BA.2 has rapidly surged globally, accounting for over 60% of recent SARS-CoV-2 infections. Newly acquired RBD mutations and high transmission advantage over BA.1 urge the investigation of BA.2s immune evasion capability. Here, we show that BA.2 causes strong neutralization resistance, comparable to BA.1, in vaccinated individuals plasma. However, BA.2 displays more severe antibody evasion in BA.1 convalescents, and most prominently, in vaccinated SARS convalescents plasma, suggesting a substantial antigenicity difference between BA.2 and BA.1. To specify, we determined the escaping mutation profiles1,2 of 714 SARS-CoV-2 RBD neutralizing antibodies, including 241 broad sarbecovirus neutralizing antibodies isolated from SARS convalescents, and measured their neutralization efficacy against BA.1, BA.1.1, BA.2. Importantly, BA.2 specifically induces large-scale escape of BA.1/BA.1.1-effective broad sarbecovirus neutralizing antibodies via novel mutations T376A, D405N, and R408S. These sites were highly conserved across sarbecoviruses, suggesting that Omicron BA.2 arose from immune pressure selection instead of zoonotic spillover. Moreover, BA.2 reduces the efficacy of S309 (Sotrovimab)3,4 and broad sarbecovirus neutralizing antibodies targeting the similar epitope region, including BD55-5840. Structural comparisons of BD55-5840 in complexes with BA.1 and BA.2 spike suggest that BA.2 could hinder antibody binding through S371F-induced N343-glycan displacement. Intriguingly, the absence of G446S mutation in BA.2 enabled a proportion of 440-449 linear epitope targeting antibodies to retain neutralizing efficacy, including COV2-2130 (Cilgavimab)5. Together, we showed that BA.2 exhibits distinct antigenicity compared to BA.1 and provided a comprehensive profile of SARS-CoV-2 antibody escaping mutations. Our study offers critical insights into the humoral immune evading mechanism of current and future variants. Engineered bacteria detect tumor DNA Advances in bacterial engineering have catalysed the development of living cell diagnostics and therapeutics1-3, including microbes that respond to diseases such as gut inflammation4, intestinal bleeding5, pathogens6 and hypoxic tumors7. Bacteria can easily access the entire gastrointestinal tract via oral administration8, and they can produce outputs that can be noninvasively measured in stool4 or urine7. Cellular memory, such as bistable switches4,9,10 or genomic rearrangement11, has been used to allow bacteria to store information over time. However, living biosensors have not yet been engineered to detect specific DNA sequences or mutations from outside the cell. Here, we engineer naturally competent Acinetobacter baylyi to detect donor DNA from the genomes of colorectal cancer (CRC) cells and organoids. We characterize the functionality of the biosensors in vitro with co-culture assays and then validate in vivo with sensor bacteria delivered orally or rectally into mice injected with orthotopic donor CRC organoids. We observe horizontal gene transfer from the tumor to the sensor bacteria in vivo, allowing their detection in stool. The sensor bacteria achieved 100% discrimination between mice with and without CRC using both delivery methods. Our findings establish a framework for biosensing applications that require the detection of mutations or organisms within environments that are difficult to sample. In addition, the platform can be readily expanded to include in situ production and delivery of therapeutic payloads at the detection site. The first sheep graph-based pan-genome reveals the spectrum of structural variations and their effects on tail phenotypes Structural variations (SVs) are a major contributor to genetic diversity and phenotypic variations, but their prevalence and functions in domestic animals are largely unexplored. Here, we assembled 26 haplotype-resolved genome assemblies from 13 genetically diverse sheep using PacBio HiFi sequencing. We constructed a graph-based ovine pan-genome and discovered 142,422 biallelic insertions and deletions, 7,028 divergent alleles and 13,419 multiallelic variations. We then used a graph-based approach to genotype the biallelic SVs in 684 individuals from 45 domestic breeds and two wild species. Integration with RNA-seq data allows to identify candidate expression-associated SVs. We demonstrate a direct link of SVs and phenotypes by localizing the putative causative insertion in HOXB13 gene responsible for the long-tail trait and identifying multiple large SVs associated with the fat-tail. Beyond generating a benchmark resource for ovine structural variants, our study highlights that animal genetic research will greatly benefit from using a pan-genome graph rather than a single reference genome. Genetic analysis of mitochondrial DNA copy number and associated traits identifies loci implicated in nucleotide metabolism, platelet activation, and megakaryocyte proliferation, and reveals a causal association of mitochondrial function with mortality Mitochondrial DNA copy number (mtDNA-CN) measured from blood specimens is a minimally invasive marker of mitochondrial function that exhibits both inter-individual and intercellular variation. To identify genes involved in regulating mitochondrial function, we performed a genome-wide association study (GWAS) in 465,809 White individuals from the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium and the UK Biobank (UKB). We identified 133 SNPs with statistically significant, independent effects associated with mtDNA-CN across 100 loci. A combination of fine-mapping, variant annotation, and co-localization analyses were used to prioritize genes within each of the 133 independent sites. Putative causal genes were enriched for known mitochondrial DNA depletion syndromes (p = 3.09 x 10-15) and the gene ontology (GO) terms for mtDNA metabolism (p = 1.43 x 10-8) and mtDNA replication (p = 1.2 x 10-7). A clustering approach leveraged pleiotropy between mtDNA-CN associated SNPs and 41 mtDNA-CN associated phenotypes to identify functional domains, revealing three distinct groups, including platelet activation, megakaryocyte proliferation, and mtDNA metabolism. Finally, using mitochondrial SNPs, we establish causal relationships between mitochondrial function and a variety of blood cell related traits, kidney function, liver function and overall (p = 0.044) and non-cancer mortality (p = 6.56 x 10-4). Temporal and Spatial Dynamics of Hepatic Stellate Cell Activation in Intrahepatic Cholangiocarcinoma Intrahepatic cholangiocarcinoma (iCCA) is an aggressive type of primary liver cancer characterized by its highly desmoplastic stroma. Compared to the ample knowledge on cancer-associated fibroblast residing within iCCA tumor mass, little is known about the fibrotic response from the tumor-surrounding liver and its role in iCCA development. In this study, we find a significant accumulation of peritumoral myofibroblasts (pMFs) in both patient iCCA and tumors from an iCCA orthotopic allograft mouse model. Using tumor and liver spheroid coculture we show that iCCA-liver interaction induces rapid pMF accumulation at the interface. We find pMFs placed around iCCA spheroids exert a strong suppressive effect on tumor cells growth, in contrast to the pro-proliferative effect of MFs mixed within tumor spheroids. However, prolonged iCCA-pMF interaction elicits tumor cell dissemination in vitro. We find an upregulation of Vcam1 in tumor cells in the early phase of iCCA-pMF interaction both in vitro and in vivo which is downregulated when tumor cells disseminate. Blocking Vcam1 activity in iCCA allograft mouse models slows primary tumor growth but lead to increased tumor metastasis. Our data suggest that pMFs are beyond simple pro- or anti-tumorigenic in iCCA, with the ability to suppress tumor growth but elicit tumor cell dissemination. Library-free BoxCarDIA solves the missing value problem in label-free quantitative proteomics The last decade has seen significant advances in the application of quantitative mass spectrometry-based proteomics technologies to tackle important questions in plant biology. The current standard for quantitative proteomics in plants is the use of data-dependent acquisition (DDA) analysis with or without the use of chemical labels. However, the DDA approach preferentially measures higher abundant proteins, and often requires data imputation due to quantification inconsistency between samples. In this study we systematically benchmarked a recently developed library-free data-independent acquisition (directDIA) method against a state-of-the-art DDA label-free quantitative proteomics workflow for plants. We next developed a novel acquisition approach combining MS1-level BoxCar acquisition with MS2-level directDIA analysis that we call BoxCarDIA. DirectDIA achieves a 33% increase in protein quantification over traditional DDA, and BoxCarDIA a further 8%, without any changes in instrumentation, offline fractionation, or increases in mass-spectrometer acquisition time. BoxCarDIA, especially, offers wholly reproducible quantification of proteins between replicate injections, thereby addressing the long-standing missing-value problem in label-free quantitative proteomics. Further, we find that the gains in dynamic range sampling by directDIA and BoxCarDIA translate to deeper quantification of key, low abundant, functional protein classes (e.g., protein kinases and transcription factors) that are underrepresented in data acquired using DDA. We applied these methods to perform a quantitative proteomic comparison of dark and light grown Arabidopsis cell cultures, providing a critical resource for future plant interactome studies. Our results establish BoxCarDIA as the new method of choice in quantitative proteomics using Orbitrap-type mass-spectrometers, particularly for proteomes with large dynamic range such as that of plants. 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. LPS-TLR4 pathway exaggerates alcohol-induced acute-on-chronic liver failure via provoking NETs formation Backgrounds: Intrahepatic infiltration of neutrophils is a character of alcoholic acute-on-chronic liver failure (AACLF) and neutrophil extracellular traps (NETs) are an important strategy for neutrophils to fix and kill invading microorganisms. Intestinal bacteria and the gut-liver axis have been thought to play a key role in many liver diseases also including AACLF. However, whether NETs appear in AACLF and play a role in AACLF is still unsure. Methods: WT, NE KO, and TLR4 KO mice were used to build the AACLF model, and the intestinal bacteria were eliminated at the same time and LPS was given. Then the formation of NETs and AACLF related markers were detected. Results: The serum MPO-DNA and LPS concentration was increased in AACLF patients and a correlation was revealed between these two indexes. More intrahepatic NETs formed in AACLF mice by testing MPO-DNA, Cit H3, and NE. These markers decreased with gut detergent and restored markers with gut detergent plus LPS supplement. While NETs formation failed to change with gut microbiome or combine LPS supplement in TLR4 KO mice. As we tested AACLF related characters, liver injury, intrahepatic fat deposition, inflammation, and fibrosis alleviated with depletion of NE. These related marks were also attenuated with gut sterilization by antibiotics and recovered with combined treatment with antibiotics plus LPS. But the liver injury, intrahepatic fat, fibro deposition, and liver inflammation-related markers did show a significant difference in TLR4 KO mice when they received the same treatment. Conclusion: Intestinal-derived LPS promotes NETs formation in AACLF through the TLR4 pathway and further accelerates the AACLF process by NETs. Pseudomonas aeruginosa C-terminal processing protease CtpA assembles into a hexameric structure that requires activation by a spiral-shaped lipoprotein binding partner Pseudomonas aeruginosa CtpA is a carboxyl terminal-processing protease that partners with the outer membrane lipoprotein LbcA to degrade at least five cell wall-associated proteins, four of which are cell wall hydrolases. This activity plays an important role in supporting P. aeruginosa virulence in a mouse model of acute pneumonia. However, almost nothing is known about the molecular mechanisms underlying CtpA and LbcA function. Here, we used structural analysis to show that CtpA alone assembles into an inactive hexamer comprising a trimer of dimers, which limits its substrate access and prevents nonspecific degradation. The adaptor protein LbcA is a right-handed open spiral with 11 tetratricopeptide repeats, which might wrap around a substrate to deliver it to CtpA for degradation. By structure-guided mutagenesis and functional assays, we also showed that the interfaces of the CtpA trimer-of-dimers, and an N-terminal helix of LbcA, are important for LbcA-mediated substrate degradation by CtpA both in vitro and in vivo. This work improves our understanding of the molecular mechanism of a CTP within the C-terminal processing peptidase-3 group.

IMPORTANCECarboxyl-terminal processing proteases (CTPs) are found in all three domains of life. In bacteria, some CTPs have been associated with virulence, raising the possibility that they could be theraputic targets. However, relatively little is known about their molecular mechanisms of action. In Pseudomonas aeruginosa, CtpA supports virulence by working in complex with the outer membrane lipoprotein LbcA to degrade cell wall hydrolases. Here, we report structure-function analyses of CtpA and LbcA, which reveals that CtpA assembles into an inactive hexamer comprising a trimer of dimers. LbcA is monomeric, with an N-terminal region important for binding to and activating CtpA, followed by a spiral structure composed of 11 tetratricopetide repeats, which could wrap around a substrate for delivery to CtpA. This work provides the first structure of a CTP-3 group member, revealing a unique mutimeric arrangement and insight into how this important proteolytic system functions.