Activation of the type 3 secretion system of enteropathogenic E. coli leads to remodeling of its membrane composition and function The cell envelope of gram-negative bacteria is a complex structure, essential for bacterial survival and for resistance to many antibiotics. Channels that cross the bacterial envelope and the host cell membrane form secretion systems that are activated upon attachment to host, enabling bacteria to inject effector molecules into the host cell, required for bacterial-host interaction. The type III secretion system (T3SS) is critical for the virulence of several pathogenic bacteria, including enteropathogenic E. coli (EPEC). The EPEC T3SS activation is associated with repression of carbon storage regulator (CsrA), resulting in gene expression remodeling, which is known to affect EPEC central carbon metabolism and contributes to the adaptation to a cell-adherent lifestyle in a poorly understood manner. We reasoned that the changes in bacterial envelope upon attachment to host and the activation of a secretion system may involve a modification of the lipid composition of bacterial envelope. Accordingly, we performed a lipidomics analysis on mutant strains that simulate T3SS activation. We saw a shift in glycerophospholipid metabolism towards the formation of lysophospholipids, attributed to corresponding upregulation of the phospholipase pldA and the acyltransferase ygiH upon T3SS activation in EPEC. We also detected a shift from menaquinones and ubiquinones to undecaprenyl lipids, concomitant to abnormal synthesis of O-antigen. The remodeling of lipid metabolism is mediated by CsrA and associated with increased bacteria cell size and Zeta potential, and a corresponding alteration in EPEC permeability to vancomycin, increasing the sensitivity of T3SS-activated strains and of adherent wild type EPEC to the antibiotic.

ImportanceThe characterization of EPEC membrane lipid metabolism upon attachment to host is an important step towards a better understanding the shift of EPEC, a notable human pathogen, from a planktonic to adherent life style. It may also apply to other pathogenic bacteria that use this secretion system. We predict that upon attachment to host cells the lipid remodeling upon T3SS activation contributes to bacterial fitness and promotes host colonization, and show that it is associated with increased cell permeability and higher sensitivity to vancomycin.

To the best of our knowledge, this is the first demonstration of a bacterial lipid remodeling due to activation of a secretion system.
Glycolysis/gluconeogenesis specialization in microbes is driven by biochemical constraints of flux sensing Central carbon metabolism is highly conserved across microbial species, but can catalyze very different pathways depending on the organism and their ecological niche. Here, we study the dynamic re-organization of central metabolism after switches between the two major opposing pathway configurations of central carbon metabolism, glycolysis and gluconeogenesis in Escherichia coli, Pseudomonas aeruginosa and Pseudomonas putida. We combined growth dynamics and dynamic changes of intracellular metabolite levels with a coarse-grained model that integrates fluxes, regulation, protein synthesis and growth and uncovered fundamental limitations of the regulatory network: after nutrient shifts, metabolite concentrations collapse to their equilibrium, rendering the cell unable to sense which direction the flux is supposed to flow through the metabolic network. The cell can partially alleviate this by picking a preferred direction of regulation at the expense of increasing lag times in the opposite direction. Moreover, decreasing both lag times simultaneously comes at the cost of reduced growth rate or higher futile cycling between metabolic enzymes. These three trade-offs can explain why microorganisms specialize for either glycolytic or gluconeogenic substrates and can help elucidate the complex growth patterns exhibited by different microbial species.

Graphical synopsis O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

Standfirst textMicrobes face a series of fundamental trade-offs that limit their ability to optimize simultaneously for both glycolytic and gluconeogenic growth.

Bullet pointsO_LILag times between glycolysis and gluconeogenesis show asymmetry in many microbes: A long lag in one direction, but a short lag in the other.
C_LIO_LILong lag times are caused by an inability to sense fluxes after nutrient shifts.
C_LIO_LIWith existing regulation, lag time asymmetry can only be overcome by reducing either growth rate or increasing futile cycling in metabolism.
C_LI
OncoLoop: A network-based precision cancer medicine framework At present, prioritizing cancer treatments at the individual patient level remains challenging, and performing co-clinical studies using patient-derived models in real-time is often not feasible. To circumvent these challenges, we introduce OncoLoop, a precision medicine framework to predict and validate drug sensitivity in human tumors and their pre-existing high-fidelity (cognate) model(s) by leveraging perturbational profiles of clinically-relevant oncology drugs. As proof-of-concept, we applied OncoLoop to prostate cancer (PCa) using a series of genetically-engineered mouse models (GEMMs) that recapitulate a broad spectrum of disease states, including castration-resistant, metastatic, and neuroendocrine prostate cancer. Interrogation of published cohorts using Master Regulator (MR) conservation analysis revealed that most patients were represented by at least one cognate GEMM-derived tumor (GEMM-DT). Drugs predicted to invert MR activity in patients and their cognate GEMM-DTs were successfully validated, including in two cognate allografts and one cognate patient-derived xenograft (PDX). OncoLoop is a highly generalizable framework that can be extended to other cancers and potentially other diseases.

Significance StatementOncoLoop is a transcriptomic-based experimental and computational framework that can support rapid-turnaround co-clinical studies to identify and validate drugs for individual patients, which can then be readily adapted to clinical practice. This framework should be applicable in many cancer contexts for which appropriate models and drug perturbation data are available. An integrative approach leads to the discovery of a novel anti-leukemic peptide from human milk Chemotherapy in childhood leukemia is associated with late morbidity in leukemic survivors, while certain patient subsets are relatively resistant to standard chemotherapy. It is therefore important to identify new agents with sensitivity and selectivity towards leukemic cells, while having less systemic toxicity. Peptide-based therapeutics has gained much attention during the last few years. Here, we used an integrative workflow combining mass spectrometric peptide library construction, in silico anticancer peptide screening, and in vitro leukemic cell studies to discover a novel anti-leukemic peptide having 3+charges and alpha-helical structure, namely HMP-S7, from human breast milk. HMP-S7 showed cytotoxic activity against four distinct leukemic cell lines in a dose-dependent manner but had no effect on solid malignancies or representative normal cells. HMP-S7 induced leukemic cell death by penetrating the plasma membrane to enter the cytoplasm and cause leakage of lactate dehydrogenase, thus acting in a membranolytic manner. Importantly, HMP-S7 exhibited anti-leukemic effect against patient-derived leukemic cells ex vivo. In conclusion, HMP-S7 is a selective anti-leukemic peptide with promise which requires further validation in preclinical and clinical studies.

TeaserIn silico screening of naturally occurring human milk peptides discovers a new anticancer peptide that kills leukemic cells in vitro and ex vivo. Translation of upstream open reading frames does not trigger nonsense-mediated decay in Arabidopsis Upstream ORFs (uORFs) are widespread cis-regulatory elements in the 5 untranslated regions of eukaryotic genes. In addition to repressing main ORF translation, uORF translation in animals also reduces mRNA stability through nonsense-mediated decay (NMD). In contrast, the roles of uORFs in plants are less understood. One limitation of most prior studies is the inclusion of both translated and untranslated uORFs for analysis. Here, we identified actively translated uORFs (TuORFs) in Arabidopsis through ribosome profiling and examined their roles in gene expression by integrating multiple genome-wide datasets. Similar to animal systems, TuORFs are associated with 38%, 14%, and 43% reductions in translation efficiency, mRNA stability, and protein levels, respectively. Surprisingly, TuORF-containing mRNAs in plants have 51% higher transcript levels, which is opposite to the observations in animals. We present multiple lines of evidence showing that uORF translation generally does not trigger NMD in Arabidopsis. First, TuORF-containing mRNAs are not upregulated in various NMD mutants. Second, TuORF- containing mRNAs and known NMD targets have distinct expression patterns, and they are translationally repressed via different mechanisms. Finally, TuORF-containing mRNAs and NMD targets are degraded through separate pathways. Together, our results highlight fundamental differences in gene regulation mediated by TuORFs in plants and animals. Disruption of the microphysiological niche alters matrix deposition and causes loss of mitochondrial Ca2+ control in skeletal muscle fibers Cells rapidly lose their physiological phenotype upon disruption of their extracellular matrix (ECM)-intracellular cytoskeleton interactions. Here, we investigated acute effects of ECM disruption on cellular and mitochondrial morphology, transcriptomic signatures, and Ca2+ handling in adult mouse skeletal muscle fibers. Adult skeletal muscle fibers were isolated from mouse toe muscle either by collagenase-induced dissociation of the ECM or by mechanical dissection that leaves the contiguous ECM intact. Experiments were generally performed four hours after cell isolation. At this time, there were striking differences in the gene expression patterns between fibers isolated with the two methods; 24h after cell isolation, enzymatically dissociated fibers had transcriptomic signatures resembling dystrophic phenotypes. Mitochondrial appearance was grossly similar in the two groups, but 3D electron microscopy revealed shorter and less branched mitochondria in enzymatically dissociated than in mechanically dissected fibers. Similar increases in free cytosolic [Ca2+] during repeated tetanic stimulation were accompanied by marked mitochondrial Ca2+ uptake only in enzymatically dissociated muscle fibers. The aberrant mitochondrial Ca2+ uptake was partially prevented by the mitochondrial Ca2+ uniporter inhibitor Ru360 and by cyclosporine A and NV556, which inhibit the mitochondrial protein Ppif (also called cyclophilin D). Importantly, inhibition of Ppif with NV556 significantly improved survival of mice with mitochondrial myopathy in which muscle mitochondria take up excessive amounts of Ca2+ even with an intact ECM. In conclusion, skeletal muscle fibers isolated by collagenase-induced dissociation of the ECM display aberrant mitochondrial Ca2+ uptake, which involves a Ppif-dependent mitochondrial Ca2+ influx resembling that observed in mitochondrial myopathies. Transcriptomic analysis of temporal shifts in berry development between two grapevine cultivars of the Pinot family reveals potential genes controlling ripening time BackgroundGrapevine cultivars of the Pinot family represent clonally propagated mutants with major phenotypic and physiological differences, such as different colour or shifted ripening time, as well as changes in important viticultural traits. Specifically, the cultivars  Pinot Noir (PN) and  Pinot Noir Precoce (PNP, early ripening) flower at the same time, but vary in the beginning of berry ripening (veraison) and, consequently, harvest time. In addition to genotype, seasonal climatic conditions (i.e. high temperatures) also affect ripening times. To reveal possible regulatory genes that affect the timing of veraison onset, we investigated differences in gene expression profiles between PN and PNP throughout berry development with a closely meshed time series and over two separate years.

ResultsThe difference in the duration of berry formation between PN and PNP was quantified to be approximately two weeks under the growth conditions applied, using plant material with a proven PN and PNP clonal relationship. Clusters of co-expressed genes and differentially expressed genes (DEGs) were detected which reflect the shift in the timing of veraison onset. Functional annotation of these DEGs fit to observed phenotypic and physiological changes during berry development. In total, we observed 3,342 DEGs in 2014 and 2,745 DEGs in 2017 between PN and PNP, with 1,923 DEGs across both years. Among these, 388 DEGs were identified as veraison-specific and 12 were considered as berry ripening time regulatory candidates. The expression profiles revealed two candidate genes for ripening time control which we designated VviRTIC1 and VviRTIC2 (VIT_210s0071g01145 and VIT_200s0366g00020, respectively). These genes likely contribute the phenotypic differences observed between PN and PNP.

ConclusionsMany of the 1,923 DEGs show highly similar expression profiles in both cultivars if the patterns are aligned according to developmental stage. In our work, putative genes differentially expressed between PNP and PN which could control ripening time as well as veraison-specific genes were identified. We point out connections of these genes to molecular events during berry development and discuss potential candidate genes which may control ripening time. Two of these candidates were observed to be differentially expressed in the early berry development phase. Several down-regulated genes during berry ripening are annotated as auxin response factors / ARFs. Conceivably, general changes in auxin signaling may cause the earlier ripening phenotype of PNP. Frameshift and wild-type proteins are highly similar because the genetic code and genomes were optimized for frameshift tolerance Frameshift protein sequences encoded by alternative reading frames of coding genes have been considered meaningless, and frameshift mutations have been considered of little importance for the molecular evolution of coding genes and proteins. However, functional frameshifts have been found widely existing. It was puzzling how a frameshift protein kept its structure and functionality while its amino-acid sequence was changed substantially. Here we show that frame similarities between frameshifts and wild types are higher than random similarities and are defined at the genetic code, gene, and genome levels. In the standard genetic code, frameshift codon substitutions are more conservative than random substitutions. The frameshift tolerability of the standard genetic code ranks in the top 2.0-3.5% of alternative genetic codes, showing that the genetic code is nearly optimal for frameshift tolerance. Furthermore, frameshift-resistant codons (codon pairs) appear more frequently than expected in many genes and certain genomes, showing that the frameshift optimality is reflected not only in the genetic code but more importantly, in its allowance of further optimizing the frameshift tolerance of a particular gene or genome, which shed light on the role of frameshift mutations in molecular and genomic evolution. Overexpression-based detection of translatable circular RNAs is vulnerable to coexistent linear RNA byproducts In RNA field, the demarcation between coding and non-coding has been negotiated by the recent discovery of occasionally translated circular RNAs (circRNAs). Although absent of 5 cap structure, circRNAs can be translated cap-independently. Complementary intron-mediated overexpression is one of the most utilized methodologies for circRNA research but not without bearing echoing skepticism for its poorly defined mechanism and latent coexistent side products. In this study, leveraging such circRNA overexpression system, we have interrogated the protein-coding potential of 30 human circRNAs containing infinite open reading frames in HEK293T cells. Surprisingly, pervasive translation signals are detected by immunoblotting. However, intensive mutagenesis reveals that numerous translation signals are generated independently of circRNA synthesis. We have developed a dual tag strategy to isolate translation noise and directly demonstrate that the fallacious translation signals originate from cryptically spliced linear transcripts. The concomitant linear RNA byproducts, presumably concatemers, can be translated to allow pseudo rolling circle translation signals, and can involve backsplicing junction (BSJ) to disqualify the BSJ-based evidence for circRNA translation. We also find non-AUG start codons may engage in the translation initiation of circRNAs. Taken together, our systematic evaluation sheds light on heterogeneous translational outputs from circRNA overexpression vector and comes with a caveat that ectopic overexpression technique necessitates extremely rigorous control setup in circRNA translation and functional investigation. Updated vaccine protects from infection with SARS-CoV-2 variants, prevents transmission and is immunogenic against Omicron in hamsters Current first-generation COVID-19 vaccines are based on prototypic spike sequences from ancestral 2019 SARS-CoV-2 strains. However, the ongoing pandemic is fueled by variants of concern (VOC) that threaten to escape vaccine-mediated protection. Here we show in a stringent hamster model that immunization using prototypic spike expressed from a potent YF17D viral vector (1) provides vigorous protection against infection with ancestral virus (B lineage) and VOC Alpha (B.1.1.7), however, is insufficient to provide maximum protection against the Beta (B.1.351) variant. To improve vaccine efficacy, we created a revised vaccine candidate that carries an evolved spike antigen. Vaccination of hamsters with this updated vaccine candidate provides full protection against intranasal challenge with all four VOCs Alpha, Beta, Gamma (P.1) and Delta (B.1.617.2) resulting in complete elimination of infectious virus from the lungs and a marked improvement in lung pathology. Vaccinated hamsters did also no longer transmit the Delta variant to non-vaccinated sentinels. Hamsters immunized with our modified vaccine candidate also mounted marked neutralizing antibody responses against the recently emerged Omicron (B.1.1.529) variant, whereas the old vaccine employing prototypic spike failed to induce immunity to this antigenically distant virus. Overall, our data indicate that current first-generation COVID-19 vaccines need to be urgently updated to cover newly emerging VOCs to maintain vaccine efficacy and to impede virus spread at the community level.

Significance StatementSARS-CoV-2 keeps mutating rapidly, and the ongoing COVID-19 pandemic is fueled by new variants escaping immunity induced by current first-generation vaccines. There is hence an urgent need for universal vaccines that cover variants of concern (VOC). In this paper we show that an adapted version of our vaccine candidate YF-S0* provides full protection from infection, virus transmission and disease by VOCs Alpha, Beta, Gamma and Delta, and also results in markedly increased levels of neutralizing antibodies against recently emerged Omicron VOC in a stringent hamster model. Our findings underline the necessity to update COVID-19 vaccines to curb the pandemic, providing experimental proof on how to maintain vaccine efficacy in view of an evolving SARS-CoV-2 diversity. Sablefish (Anoplopoma fimbria Pallas, 1814) plasma biochemistry and hematology reference intervals including blood cell morphology Plasma biochemistry and hematology reference intervals are integral health assessment tools in all medical fields, including aquatic animal health. As sablefish (Anoplopoma fimbria) are becoming aquaculturally and economically more important, this manuscript provides essential reference intervals (RI) for their plasma biochemistry and hematology along with reference photomicrographs of blood cells in healthy, fasted sablefish. Blood cell morphology can differ between fish species. In addition, blood cell counts and blood chemistry can vary between fish species, demographics, water conditions, seasons, diets, and culture systems, which precludes the use of RIs from other fish species. For this study, blood was collected for plasma biochemistry and hematology analysis between June 20 and July 18, 2019, from healthy, yearling sablefish, hatched and reared in captivity on a commercial diet. Overnight fast of 16-18 hours did not sufficiently reduce lipids in the blood, which led to visible lipemia and frequent rupture of blood cells during analysis. Therefore, sablefish should be fasted for 24 to 36 hours before blood is collected to reduce hematology artifacts or possible reagent interference in plasma biochemistry analysis. Lymphocytes were the most dominant leukocytes (98%), while eosinophils were rare, and basophils were not detected in sablefish. Neutrophils were very large cells with Dohle bodies. In mammals and avian species, Dohle bodies are usually signs of toxic change from inflammation, but no such association was found in these fish. In conclusion, lipemia can interfere with sablefish blood analysis, and available removal methods should be evaluated as fasting for up to 36 h might not always be feasible. Also, more studies are required to establish RI for different developmental stages and rearing conditions. Behavioral and postural analyses establish sleep-like states for mosquitoes that can impact host landing and blood feeding Sleep is an evolutionarily conserved process that has been described in different animal systems. For insects, sleep characterization has been primarily achieved using behavioral and electrophysiological correlates in a few systems. Sleep in mosquitoes, which are important vectors of disease-causing pathogens, has not been directly examined. This is surprising as circadian rhythms, which have been well studied in mosquitoes, influence sleep in other systems. In this study, we characterized sleep in mosquitoes using body posture analysis and behavioral correlates, and quantified the effect of sleep deprivation on sleep rebound, host landing and blood-feeding propensity. Body and appendage position metrics revealed a clear distinction between the posture of mosquitoes in their putative sleep and awake states for multiple species, which correlate with a reduction in responsiveness to host cues. Sleep assessment informed by these posture analyses indicated significantly more sleep during periods of low activity. Nighttime and daytime sleep deprivation resulting from the delivery of vibration stimuli induced sleep rebound in the subsequent phase in day and night active mosquitoes, respectively. Lastly, sleep deprivation suppressed host landing in both laboratory and field settings and also impaired blood feeding of a human host when mosquitoes would normally be active. These results suggest that quantifiable sleep states occur in mosquitoes, and highlight the potential epidemiological importance of mosquito sleep. The Impact of Neuron Morphology on Cortical Network Architecture It has become increasingly clear that the neurons in the cerebral cortex are not randomly interconnected. This wiring specificity can result from synapse formation mechanisms that interconnect neurons depending on their activity or genetically defined identity. Here we report that in addition to these synapse formation mechanisms, the structural composition of the neuropil provides a third prominent source by which wiring specificity emerges in cortical networks. This structurally determined wiring specificity reflects the packing density, morphological diversity and similarity of the dendritic and axonal processes. The higher these three factors are, the more recurrent the topology of the networks. Conversely, low density, diversity and similarity yield feedforward networks. These principles predict connectivity patterns from subcellular to network scales that are remarkably consistent with empirical observations from a rich body of literature. Thus, cortical network architectures reflect the specific morphological properties of their constituents to much larger degrees than previously thought. Hydrophobic mismatch is a key factor in protein transport on the Tat pathway The twin-arginine translocation (Tat) pathway transports folded proteins across membranes in bacteria, thylakoid, plant mitochondria, and archaea. In most species, the active Tat machinery consists of three independent subunits, TatA, TatB and TatC. TatA and TatB from all bacterial species possess short transmembrane alpha-helices (TMHs), both of which are only fifteen residues long in E. coli. Such short TMHs cause a hydrophobic mismatch between Tat subunits and the membrane bilayer. Here, by modifying the length of the TMHs of E. coli TatA and TatB, we assess the functional importance of the hydrophobic mismatch in the Tat transport mechanism. Surprisingly, both TatA and TatB with as few as 11 residues in their respective TMHs are still able to insert into the membrane bilayer, albeit with a decline in membrane integrity. Three different assays, both qualitative and quantitative, were conducted to evaluate the Tat activity of the TMH length mutants. Our experiments indicate that the TMHs of TatA and TatB appear to be evolutionarily tuned to 15 amino acids, with activity dropping off with any modification of this length. We believe our study supports a model of Tat transport utilizing localized toroidal pores that form when the membrane bilayer is thinned to a critical threshold. In this context, the 15-residue length of the TatA and TatB TMHs can be seen as a compromise between the need for some hydrophobic mismatch to allow the membrane to reversibly reach the threshold thinness required for toroidal pore formation, and the permanently destabilizing effect of placing even shorter helices into these energy-transducing membranes. 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. Cryo-EM Structures of the N501Y SARS-CoV-2 Spike Protein in Complex with ACE2 and Two Potent Neutralizing Antibodies The recently reported "UK variant" of SARS-CoV-2 is thought to be more infectious than previously circulating strains as a result of several changes, including the N501Y mutation. We present a 2.9-[A] resolution cryo-EM structure of the complex between the ACE2 receptor and N501Y spike protein ectodomains that shows Y501 inserted into a cavity at the binding interface near Y41 of ACE2. The additional interactions result in increased affinity of ACE2 for the N501Y mutant, accounting for its increased infectivity. However, this mutation does not result in large structural changes, enabling important neutralization epitopes to be retained in the spike receptor binding domain. We confirmed this through biophysical assays and by determining cryo-EM structures of spike protein ectodomains bound to two representative potent neutralizing antibody fragments.

Short summaryThe N501Y mutation found in the coronavirus UK variant increases infectivity but some neutralizing antibodies can still bind. The cyclic dinucleotide 2'3'-cGAMP induces a broad anti-bacterial and anti-viral response in the sea anemone Nematostella vectensis In mammals, cyclic dinucleotides (CDNs) bind and activate STING to initiate an anti-viral type I interferon response. CDNs and STING originated in bacteria and are present in most animals. By contrast, interferons are believed to have emerged in vertebrates; thus, the function of CDN signaling in invertebrates is unclear. Here, we use a CDN, 23-cGAMP, to activate immune responses in a model cnidarian invertebrate, the starlet sea anemone Nematostella vectensis. Using RNA-Seq, we found that 23-cGAMP induces robust transcription of both anti-viral and anti-bacterial genes in N. vectensis. Many of the anti-viral genes induced by 23-cGAMP are homologs of vertebrate interferon stimulated genes, implying that the interferon response predates the evolution of interferons. Knockdown experiments identified a role for NF-{kappa}B in specifically inducing anti-bacterial genes downstream of 23-cGAMP. Some of these putative anti-bacterial genes were also found to be induced during Pseudomonas aeruginosa infection. We characterized the protein product of one of the putative anti-bacterial genes, the N. vectensis homolog of Dae4, and found that it has conserved anti-bacterial activity. This work suggests that a broad anti-bacterial and anti-viral transcriptional response is an evolutionarily ancestral output of 23-cGAMP signaling in animals.

Significance statementCyclic dinucleotides are signaling molecules that originated in bacteria and were subsequently acquired and co-opted by animals for immune signaling. The major cyclic dinucleotide signaling pathway in mammals results in the production of anti-viral molecules called interferons. Invertebrates such as sea anemones lack interferons, and thus it was unclear whether cyclic dinucleotide signaling would play a role in immunity in these animals. Here we report that in the anemone Nematostella vectensis, cyclic dinucleotides activate both anti-viral and anti-bacterial immune responses, and do so through a conserved pathway. These results provide insights into the evolutionary origins of innate immunity, and suggest a broader ancestral role for cyclic dinucleotide signaling that evolved toward more specialized anti-viral functions in mammals. SARS-CoV-2 infection induces soluble platelet activation markers and PAI-1 in the early moderate stage of COVID-19 IntroductionCoagulation dysfunction and thromboembolism emerge as strong comorbidity factors in severe COVID-19. However, it is unclear when particularly platelet activation markers and coagulation factors dysregulated during the pathogenesis of COVID-19. Here, we sought to assess the levels of coagulation and platelet activation markers at moderate and severe stages of COVID-19 to understand the pathogenesis.

MethodsTo understand this, hospitalized COVID-19 patients with (severe cases that required intensive care) or without pneumonia (moderate cases) were recruited. Phenotypic and molecular characterizations were performed employing basic coagulation tests including PT, APTT, D-Dimer and TFPI. The flow cytometry-based multiplex assays were performed to assess FXI, anti-thrombin, prothrombin, fibrinogen, FXIII, P-selectin, sCD40L, plasminogen, tissue-plasminogen activator (tPA), plasminogen activator inhibitor-1 (PAI-1) and D-Dimer.

ResultsThe investigations revealed induction of plasma P-selectin and CD40 ligand (sCD40L) in moderate COVID-19 cases which were significantly abolished with the progression of COVID-19 severity. Moreover, a profound reduction in plasma tissue factor pathway inhibitor (TFPI) and FXIII were identified particularly in the severe COVID-19. Further analysis revealed fibrinogen induction in both moderate and severe patients. Interestingly, an elevated PAI-1 more prominently in moderate, and tPA particularly in severe COVID-19 cases were observed. Particularly, the levels of fibrinogen and tPA directly correlated with the severity of the disease.

ConclusionsIn summary, induction of soluble P-selectin, sCD40L, fibrinogen and PAI-1 in moderate COVID-19 cases suggests the activation of platelets and coagulation system before patients require intensive care. These findings would help in designing better thromboprophylaxis to limit the COVID-19 severity. An End-to-End Deep Learning Speech Coding and Denoising Strategy for Cochlear Implants Cochlear implant (CI) users struggle to understand speech in noisy conditions. To address this problem, we propose a deep learning speech denoising sound coding strategy that estimates the CI electric stimulation patterns out of the raw audio data captured by the micro-phone, performing end-to-end CI processing. To estimate the relative denoising performance differences between various approaches, we compared this technique to a classic Wiener filter and to a convTasNet. Speech enhancement performance was assessed by means of signal-to-noise-ratio improvement and the short-time objective speech intelligibility measure. Additionally, 5 CI users were evaluated for speech intelligibility in noise to assess the potential benefits of each algorithm. Our results show that the proposed method is capable of replacing a CI sound coding strategy while preserving its general use for every listener and performing speech enhancement in noisy environments, without sacrificing algorithmic latency. An Evolutionary Process Without Variation and Selection Natural selection successfully explains how organisms accumulate adaptive change despite that traits acquired over a lifetime are eliminated at the end of each generation. However, in some domains that exhibit cumulative, adaptive change--e.g., cultural evolution, and earliest life--acquired traits are retained; these domains do not face the problem that Darwins theory was designed to solve. Lack of transmission of acquired traits occurs when germ cells are protected from environmental change, due to a self-assembly code used in two distinct ways: (i) actively interpreted during development to generate a soma, and (ii) passively copied without interpretation during reproduction to generate germ cells. Early life and cultural evolution appear not to involve a self-assembly code used in these two ways. We suggest that cumulative, adaptive change in these domains is due to a lower-fidelity evolutionary process, and model it using Reflexively Autocatalytic and Foodset-generated networks. We refer to this more primitive evolutionary process as Self-Other Reorganisation (SOR) because it involves internal self-organising and self-maintaining processes within entities, as well as interaction between entities. SOR encompasses learning but in general operates across groups. We discuss the relationship between SOR and Lamarckism, and illustrate a special case of SOR without variation. Social integration predicts survival in female white-faced capuchin monkeys Across multiple species of social mammals, a growing number of studies have found that that individual sociality is associated with survival. In long-lived species, like primates, lifespan is one of the main components of fitness. We used 18 years of data from the Lomas Barbudal Monkey project to quantify social integration in 11 capuchin (Cebus capucinus) groups and tested whether female survivorship was associated with females tendencies to interact with three types of partners: (1) all group members, (2) adult females, (3) adult males. We found strong evidence that females who engaged more with other females in affiliative interactions and foraged in close proximity experienced increased survivorship. We found some weak evidence that females might also benefit from engaging in more support in agonistic contexts with other females. These benefits were evident in models that account for the females rank and group size. Female interactions with all group members also increased survival, but the estimates of the effects were more uncertain. In interactions with adult males, only females who provided more grooming to males survived longer. The results presented here suggest that social integration may result in survival-related benefits. Females might enjoy these benefits through exchanging grooming for other currencies, such as coalitionary support or tolerance. MT-MAG: Accurate and interpretable machine learning for complete or partial taxonomic assignments of metagenome-assembled genomes We propose MT-MAG, a novel machine learning-based software tool for the complete or partial hierarchically-structured taxonomic classification of metagenome-assembled genomes (MAGs). MT-MAG is capable of classifying large and diverse metagenomic datasets: a total of 245.68 Gbp in the training sets, and 9.6 Gbp in the test sets analyzed in this study. MT-MAG is, to the best of our knowledge, the first machine learning method for taxonomic assignment of metagenomic data that offers a "partial classification" option. MT-MAG outputs complete or partial classification paths, and interpretable numerical classification confidences of its classifications, at all taxonomic ranks. To assess the performance of MT-MAG, we define a "weighted classification accuracy," with a weighting scheme reflecting the fact that partial classifications at different ranks are not equally informative. For the two benchmarking datasets analyzed (genomes from human gut microbiome species, and bacterial and archaeal genomes assembled from cow rumen metagenomic sequences), MT-MAG achieves an average of 80.13% in weighted classification accuracy. At the Species level, MT-MAG outperforms DeepMicrobes, the only other comparable software tool, by an average of 35.75% in weighted classification accuracy. In addition, MT-MAG is able to completely classify an average of 67.7% of the sequences at the Species level, compared with DeepMicrobes which only classifies 47.45%. Moreover, MT-MAG provides additional information for sequences that are not completely classified at the Species level, resulting in the partial or complete classification of 95.15%, of the genomes in the datasets analyzed. In particular, MT-MAG classifies, on average, 88.84% of the test sequences to the Phylum level, 88.39% to the Class level, 86.81% to the Order level, 81.17% to the Family level, and 71.13% to the Genus level. Multi-omic analysis of the Arabidopsis clock activator mutant rve 4 6 8 reveals connections to carbohydrate metabolism and proteasome regulation Plants are able to sense changes in their light environments, such as the onset of day and night, as well as anticipate these changes in order to adapt and survive. Central to this ability is the plant circadian clock, a molecular circuit that precisely orchestrates plant cell processes over the course of a day. REVEILLE proteins (RVEs) are recently discovered members of the plant circadian circuitry that activate the evening complex and PRR genes to maintain regular circadian oscillation. The RVE 8 protein and its two homologs, RVE 4 and 6, have been shown to limit the length of the circadian period, with rve 4 6 8 triple-knockout plants possessing an elongated period along with increased leaf surface area, biomass, cell size and delayed flowering relative to wild-type Col-0 plants. Here, using a multi-omics approach consisting of phenomics, transcriptomics, proteomics, and metabolomics we draw novel connections between RVE8-like proteins and a number of core plant cell processes. In particular, we reveal that loss of RVE8-like proteins results in altered carbohydrate, organic acid and lipid metabolism, including a starch excess phenotype at dawn. We further demonstrate that rve 4 6 8 plants have lower levels of 20S proteasome subunits and possess significantly reduced proteasome activity, potentially explaining the increase in cell-size observed in RVE8-like mutants. Overall, this robust, multi-omic dataset, provides substantial new insights into the far reaching impact RVE8-like proteins have on the diel plant cell environment. 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. A tool for assessing alignment of biomedical data repositories with open, FAIR, citation and trustworthy principles Increasing attention is being paid to the operation of biomedical data repositories in light of efforts to improve how scientific data is handled and made available for the long term. Multiple groups have produced recommendations for functions that biomedical repositories should support, with many using requirements of the FAIR data principles as guidelines. However, FAIR is but one set of principles that has arisen out of the open science community. They are joined by principles governing open science, data citation and trustworthiness, all of which are important aspects for biomedical data repositories to support. Together, these define a framework for data repositories that we call OFCT: Open, FAIR, Citable and Trustworthy. Here we developed an instrument using the open source PolicyModels toolkit that attempts to operationalize key aspects of OFCT principles and piloted the instrument by evaluating eight biomedical community repositories listed by the NIDDK Information Network (dkNET.org). Repositories included both specialist repositories that focused on a particular data type or domain, in this case diabetes and metabolomics, and generalist repositories that accept all data types and domains. The goal of this work was both to obtain a sense of how much the design of current biomedical data repositories align with these principles and to augment the dkNET listing with additional information that may be important to investigators trying to choose a repository, e.g., does the repository fully support data citation? The evaluation was performed from March to November 2020 through inspection of documentation and interaction with the sites by the authors. Overall, although there was little explicit acknowledgement of any of the OFCT principles in our sample, the majority of repositories provided at least some support for their tenets. Priority conservation areas and a global population estimate for the Critically Endangered Philippine Eagle derived from modelled range metrics using remote sensing habitat characteristics Many range-restricted taxa are currently experiencing population declines yet lack fundamental information regarding distribution and population size. Establishing baseline estimates for both these key biological parameters is however critical for directing conservation planning for at-risk range-restricted species. The International Union for the Conservation of Nature (IUCN) Red List uses three range metrics that define species distributions and inform extinction risk assessments: extent of occurrence (EOO), area of occupancy (AOO) and area of habitat (AOH). However, calculating all three metrics using standard IUCN approaches relies on a geographically representative sample of locations, which for rare species is often spatially biased. Here, we apply model-based interpolation using Species Distribution Models (SDMs), correlating occurrences with remote-sensing covariates, to calculate IUCN range metrics, protected area coverage and a global population estimate for the Critically Endangered Philippine Eagle (Pithecophaga jefferyi). Our final range wide continuous SDM had high predictive accuracy (Continuous Boyce Index = 0.927) and when converted to a binary model estimated an AOH = 23,185 km2, a maximum EOO = 605,759 km2, a minimum EOO = 272,272 km2, with an AOO = 53,867 km2. Based on inferred habitat from the AOH metric, we estimate a global population of 318 breeding pairs (range: 258-362 pairs), or 636 mature individuals, across the Philippine Eagle global range. Protected areas covered 34 % of AOH, 15 % less than the target representation, with the continuous model identifying key habitat as priority conservation areas. We demonstrate that even when occurrences are geographically biased, robust habitat models can be built that enable quantification of baseline IUCN range metrics, protected area coverage, and a population size estimate. In the absence of adequate location data for many rare and threatened taxa, our method is a promising spatial modelling tool with widespread applications, in particular for island endemics facing high extinction risk.