Expression of the ACE2 virus entry protein in the nervus terminalis reveals the potential for an alternative route to brain infection in COVID-19 Previous studies suggested that the SARS-CoV-2 virus may gain access to the brain by using a route along the olfactory nerve. However, there is a general consensus that the obligatory virus entry receptor, angiotensin converting enzyme 2 (ACE2), is not expressed in olfactory receptor neurons, and the timing of arrival of the virus in brain targets is inconsistent with a neuronal transfer along olfactory projections. We determined whether nervus terminalis neurons and their peripheral and central projections should be considered as a potential alternative route from the nose to the brain. Nervus terminalis neurons in postnatal mice were double-labeled with antibodies against ACE2 and two nervus terminalis markers, gonadotropin-releasing hormone (GnRH) and choline acetyltransferase (CHAT). We show that a small fraction of CHAT-labeled nervus terminalis neurons, and the large majority of GnRH-labeled nervus terminalis neurons with cell bodies in the region between the olfactory epithelium and the olfactory bulb express ACE2 and cathepsins B and L. Nervus terminalis neurons therefore may provide a direct route for the virus from the nasal epithelium, possibly via innervation of Bowmans glands, to brain targets, including the telencephalon and diencephalon. This possibility needs to be examined in suitable animal models and in human tissues.
A Practical Guide to Sparse k-means Clustering for Studying Molecular Development of the Human Brain Studying the molecular development of the human brain presents unique challenges for selecting a data analysis approach. The rare and valuable nature of human postmortem brain tissue, especially for developmental studies, means the sample sizes are small (n), but the use of high throughput genomic and proteomic methods measure the expression levels for hundreds or thousands of variables (e.g. genes or proteins (p)) for each sample. This leads to a data structure that is high dimensional (p >> n) and introduces the curse of dimensionality, which poses a challenge for traditional statistical approaches. In contrast, high dimensional analyses, especially cluster analyses developed for sparse data, have worked well for analyzing genomic datasets where p >> n. Here we explore applying a lasso-based clustering method developed for high dimensional genomic data with small sample sizes. Using protein and gene data from the developing human visual cortex, we compared clustering methods. We identified an application of sparse K-means clustering (Robust Sparse K-means Clustering (RSKC)) that partitioned samples into age-related clusters that reflect lifespan stages from birth to aging. RSKC adaptively selects a subset of the genes or proteins contributing to partitioning samples into age-related clusters that progress across the lifespan. This approach addresses a problem in current studies that could not identify multiple postnatal clusters. Moreover, clusters encompassed a range of ages like a series of overlapping waves illustrating that chronological- and brain-age have a complex relationship. In addition, a recently developed workflow to create plasticity phenotypes (Balsor et al., 2020) was applied to the clusters and revealed neurobiologically relevant features that identified how the human visual cortex changes across the lifespan. These methods can help address the growing demand for multimodal integration, from molecular machinery to brain imaging signals, to understand the human brains development.
Loss of Resf1 reduces the efficiency of embryonic stem cell self-renewal and germline entry Retroelement silencing factor 1 (RESF1) interacts with the key regulators of mouse embryonic stem cells (ESCs) OCT4 and NANOG, and its absence results in sterility of mice. However, the function of RESF1 in ESCs and germ line specification is poorly understood. In this study, we used Resf1 knockout cell lines to determine the requirements of RESF1 for ESCs self-renewal and for in vitro specification of ESCs into primordial germ cell-like cells (PGCLCs). We found that deletion of Resf1 in ESCs cultured in serum and LIF reduces self-renewal potential whereas episomal expression of RESF1 has a modest positive effect on ESC self-renewal. In addition, RESF1 is not required for the capacity of NANOG and its downstream target ESRRB to drive self-renewal in the absence of LIF. However, Resf1 deletion reduces efficiency of PGCLC differentiation in vitro. These results identify Resf1 as a novel player in the regulation of pluripotent stem cells and germ cell specification. Cell-specific Bioorthogonal Tagging of Glycoproteins Altered glycosylation is an undisputed corollary of cancer development. Understanding these alterations is paramount but hampered by limitations underlying cellular model systems. For instance, the intricate interactions between tumour and host cannot be adequately recapitulated in monoculture of tumour-derived cell lines. More complex co-culture models usually rely on sorting procedures for proteome analyses and rarely capture the details of protein glycosylation. Here, we report a strategy termed Bio-Orthogonal Cell line-specific Tagging of Glycoproteins (BOCTAG). Cells are equipped by transfection with an artificial biosynthetic pathway that transforms bioorthogonally tagged sugars into the corresponding nucleotide-sugars. Only transfected cells incorporate bioorthogonal tags into glycoproteins in the presence of non-transfected cells. We employ BOCTAG as an imaging technique and to annotate cell-specific glycosylation sites in mass spectrometry-glycoproteomics. We demonstrate application in co-culture and mouse models, allowing for profiling of the glycoproteome as an important modulator of cellular function. Functional binding dynamics relevant to the evolution of zoonotic spillovers in endemic and emergent Betacoronavirus strains Comparative functional analysis of the dynamic interactions between various Betacoronavirus mutant strains and broadly utilized target proteins such as ACE2 and CD26, is crucial for a more complete understanding of zoonotic spillovers of viruses that cause diseases such as COVID-19. Here, we employ machine learning to replicated sets of nanosecond scale GPU accelerated molecular dynamics simulations to statistically compare and classify atom motions of these target proteins in both the presence and absence of different endemic and emergent strains of the viral receptor binding domain (RBD) of the S spike glycoprotein. Machine learning was used to identify functional binding dynamics that are evolutionarily conserved from bat CoV-HKU4 to human endemic/emergent strains. Conserved dynamics regions of ACE2 involve both the N-terminal helices, as well as a region of more transient dynamics encompassing K353, Q325 and a novel motif AAQPFLL 386-92 that appears to coordinate their dynamic interactions with the viral RBD at N501. We also demonstrate that the functional evolution of Betacoronavirus zoonotic spillovers involving ACE2 interaction dynamics are likely pre-adapted from two precise and stable binding sites involving the viral bat progenitor strains interaction with CD26 at SAMLI 291-5 and SS 333-334. Our analyses further indicate that the human endemic strains hCoV-HKU1 and hCoV-OC43 have evolved more stable N-terminal helix interactions through enhancement of an interfacing loop region on the viral RBD, whereas the highly transmissible SARS-CoV-2 variants (B.1.1.7, B.1.351 and P.1) have evolved more stable viral binding via more focused interactions between the viral N501 and ACE2 K353 alone. Intervene or wait? A model evaluating the timing of intervention in conservation conflicts adaptive management under uncertainty. The timing of biodiversity managers interventions can be critical to the success of conservation, especially in situations of conflict between conservation objectives and human livelihood, i.e., conservation conflicts. Given the uncertainty associated with complex social-ecological systems and the potentially irreversible consequences of delayed action for biodiversity and livelihoods, managers tend to simply intervene as soon as possible by precaution. However, refraining from intervening when the situation allows it can be beneficial, notably by saving critical management resources. Here, we introduce a strategy for managers to decide, based on monitoring, whether intervention is required or if waiting is possible. This study evaluates the performance of this waiting strategy compared to a strategy of unconditional intervention at every opportunity. We built an individual-based model of conservation conflict between a manager aiming to conserve an animal population and farmers aiming to maximize yield by protecting their crop from wildlife damage. We then simulated a budget-constrained adaptive management over time applying each strategy, while accounting for uncertainty around population dynamics and around decision-making of managers and farmers. Our results showed that when the decision for the manager to intervene was based on a prediction of population size trajectory, the waiting strategy performed at least as well as unconditional intervention while also allowing managers to save resources by avoiding unnecessary interventions. Under difficult budgetary constraints on managers, this waiting strategy ensured as high yields as unconditional intervention while significantly improving conservation outcomes by compensating managers lack of resources with the benefits accrued over waiting periods. This suggests that waiting strategies are worth considering in conservation conflicts, as they can facilitate equitable management with a more efficient use of management resources, which are often limiting in biodiversity conservation. SERCA function is impaired in skeletal and cardiac muscles from young DBA/2J mdx mice The DBA/2J (D2) mdx mouse has emerged as a more severe model of Duchenne muscular dystrophy when compared to the traditional C57BL/10 (C57) mdx mouse. Here, we questioned whether sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) function would differ in muscles from young D2 and C57 mdx mice. In gastrocnemius muscles, both D2- and C57 mdx mice exhibited signs of impaired Ca2+ uptake, however, this was more severe in D2 mdx mice. Maximal SERCA activity was lowered only in D2 mdx gastrocnemius muscles and not C57 mdx muscles. Furthermore, in the left ventricle and diaphragm, Ca2+ uptake was impaired in C57 mdx muscles with lowered rates of Ca2+ uptake compared with C57 WT mice, whereas in muscles from D2 mdx mice, rates of Ca2+ uptake were unattainable due to the severe impairments in their ability to transport Ca2+. Overall, our study demonstrates that SERCA function is drastically impaired in young D2 mdx mice. Intermittent Bulk Release of Human Cytomegalovirus Human Cytomegalovirus (HCMV) can infect a variety of cell types by using virions of varying glycoprotein compositions. It is still unclear how this diversity is generated, but spatio-temporally separated envelopment and egress pathways might play a role. So far, one egress pathway has been described in which HCMV particles are individually enveloped into small vesicles and are subsequently exocytosed continuously. However, some studies have also found enveloped virus particles inside multivesicular structures but could not link them to productive egress or degradation pathways.

We used a novel 3D-CLEM workflow allowing us to investigate these structures in HCMV morphogenesis and egress at high spatio-temporal resolution. We found that multiple envelopment events occurred at individual vesicles leading to multiviral bodies (MViBs), which subsequently traversed the cytoplasm to release virions as intermittent bulk pulses at the plasma membrane to form extracellular virus accumulations (EVAs). Our data support the existence of a novel bona fide HCMV egress pathway, which opens the gate to evaluate divergent egress pathways in generating virion diversity. Bovine neutrophils release extracellular traps and cooperate with macrophages in Mycobacterium avium subsp. paratuberculosis clearance in vitro Mycobacterium avium subsp. paratuberculosis (Map) is the underlying pathogen causing bovine paratuberculosis (PTB), an enteric granulomatous disease that mainly affects ruminants and for which an effective treatment is needed. Macrophages are the primary target cells for Map, which survives and replicates intracellularly by inhibiting phagosome maturation. Neutrophils are present at disease sites during the early stages of the infection, but seem to be absent in the late stage, in contrast to healthy tissue. Although neutrophil activity has been reported to be impaired following Map infection, their role in PTB pathogenesis has not been fully defined. Neutrophils are capable of releasing extracellular traps consisting of extruded DNA and proteins that immobilize and kill microorganisms, but this mechanism has not been evaluated against Map. Our main objective was to study the interaction of neutrophils with macrophages during an in vitro mycobacterial infection. For this purpose, neutrophils and macrophages from the same animal were cultured alone or together in the presence of Map or Mycobacterium bovis Bacillus-Calmette-Guerin (BCG). Extracellular trap release, mycobacteria killing as well as IL-1{beta} and IL-8 release were assessed. Neutrophils released extracellular traps against mycobacteria when cultured alone and in the presence of macrophages without direct cell contact, but resulted inhibited in direct contact. Macrophages were extremely efficient at killing BCG, but ineffective at killing Map. In contrast, neutrophils showed similar killing rates for both mycobacteria. Co-cultures infected with Map showed the expected killing effect of combining both cell types, whereas co-cultures infected with BCG showed a potentiated killing effect beyond the expected one, indicating a potential synergistic cooperation. In both cases, IL-1{beta} and IL-8 levels were lower in co-cultures, suggestive of a reduced inflammatory reaction. These data indicate that cooperation of both cell types can be beneficial in terms of decreasing the inflammatory reaction while the effective elimination of Map can be compromised. These results suggest that neutrophils are effective at Map killing and can exert protective mechanisms against Map that seem to fail during PTB disease after the arrival of macrophages at the infection site. Hematopoietic differentiation is characterized by a transient peak of entropy at a single cell level. Hematopoietic differentiation has been metaphorically represented as linear trajectories with discrete steps from hematopoietic stem cells to mature cells. While the transcriptional state of cells at the beginning or at the end of these trajectories are well described from bulk analysis, what happens in the intermediate states has remained elusive until the use of single cell approaches. Applying Shannon entropy to measure cell-to-cell variability among cells at the same stage of differentiation, we observed a transient peak of gene expression variability in all the hematopoietic differentiation pathways. Strikingly, genes with the highest entropy variation in a given differentiation pathway matched genes known as pathway-specific, whereas genes with the highest expression variation were common to all pathways. Finally, we showed that the level of cell-to-cell variation is increased in the most immature compartment of hematopoiesis in myelodysplastic syndromes. These data suggest that differentiation could be better conceptualized as a dynamical stochastic process with a transient stage of cellular indetermination. Population differences in the length and early-life dynamics of telomeres among European pied flycatchers Telomere length and shortening rate are increasingly used as biomarkers for long-term costs in ecological and evolutionary studies because of their relationships with survival and fitness. Both early-life conditions and growth, and later-life stressors can create variation in telomere shortening rate. Studies on between- population telomere length and dynamics are scarce, despite the expectation that populations exposed to varying environmental constraints would present divergent telomere length patterns. The pied flycatcher (Ficedula hypoleuca) is a passerine bird breeding across Eurasia (from Spain to western Siberia) and migrating through the Iberian Peninsula to spend the non-breeding period in sub-Saharan Africa. Thus, different populations show marked differences in migration distance. We studied the large-scale variation of telomere length and early-life dynamics in the pied flycatcher by comparing six European populations across a north- south gradient (Finland, Estonia, England, and Spain) predicting negative effect of migration distance on adult telomere length, and of nestling growth on nestling telomere dynamics. There were clear population differences in telomere length, with English birds from mid-latitudes having the longest telomeres. Telomere length did not thus show consistent latitudinal variation and was not linearly linked to differences in migration distance. Early-life telomere shortening rate tended to vary between populations. Fast growth was associated with shorter telomeres in the early life, but faster nestling growth affected telomeres more negatively in northern than southern populations. While the sources of between-population differences in telomere-related biology remain to be more intensively studied, our study illustrates the need to expand telomere studies at the between-population level. 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. Citation needed? Wikipedia and the COVID-19 pandemic BackgroundWith the COVID-19 pandemics outbreak, millions flocked to Wikipedia for updated information. Amid growing concerns regarding an "infodemic", ensuring the quality of information is a crucial vector of public health. Investigating if and how Wikipedia remained up to date and in line with science is key to formulating strategies to counter misinformation. Using citation analyses, we asked: which sources informed Wikipedias COVID-19-related articles before and during the pandemics first wave (January-May 2020).

ResultsWe found that coronavirus-related articles referenced trusted media sources and high-quality academic research. Moreover, despite a surge in COVID-19 preprints, Wikipedia had a clear preference for open-access studies published in respected journals and made little use of preprints. Building a timeline of English COVID-19 articles from 2001-2020 revealed a nuanced trade-off between quality and timeliness. It further showed how preexisting articles on key topics related to the virus created a framework for integrating new knowledge. Supported by a rigid sourcing policy, this "scientific infrastructure" facilitated contextualization and regulated the influx of new information. Lastly, we constructed a network of DOI-Wikipedia articles, which showed the shifting landscape of pandemic-related knowledge on Wikipedia and how academic citations create a web of shared knowledge supporting topics like COVID-19 vaccine development.

ConclusionsUnderstanding how scientific research interacts with the digital knowledge-sphere during the pandemic provides insight into how Wikipedia can facilitate access to science. It also reveals how, aided by what we term its "citizen encyclopedists", it successfully fended off COVID-19 disinformation and how this unique model may be deployed in other contexts. Hierarchical clustering optimizes the tradeoff between compositionality and expressivity of task structures for flexible reinforcement learning A hallmark of human intelligence, but challenging for reinforcement learning (RL) agents, is the ability to compositionally generalise, that is, to recompose familiar knowledge components in novel ways to solve new problems. For instance, when navigating in a city, one needs to know the location of the destination and how to operate a vehicle to get there, whether it be pedalling a bike or operating a car. In RL, these correspond to the reward function and transition function, respectively. To compositionally generalize, these two components need to be transferable independently of each other: multiple modes of transport can reach the same goal, and any given mode can be used to reach multiple destinations. Yet there are also instances where it can be helpful to learn and transfer entire structures, jointly representing goals and transitions, particularly whenever these recur in natural tasks (e.g., given a suggestion to get ice cream, one might prefer to bike, even in new towns). Prior theoretical work has explored how, in model-based RL, agents can learn and generalize task components (transition and reward functions). But a satisfactory account for how a single agent can simultaneously satisfy the two competing demands is still lacking. Here, we propose a hierarchical RL agent that learns and transfers individual task components as well as entire structures (particular compositions of components) by inferring both through a non-parametric Bayesian model of the task. It maintains a factorised representation of task components through a hierarchical Dirichlet process, but it also represents different possible covariances between these components through a standard Dirichlet process. We validate our approach on a variety of navigation tasks covering a wide range of statistical correlations between task components and show that it can also improve generalisation and transfer in more complex, hierarchical tasks with goal/subgoal structures. Finally, we end with a discussion of our work including how this clustering algorithm could conceivably be implemented by cortico-striatal gating circuits in the brain. Algal photosystem I dimer and high resolution model of PSI:plastocyanin complex Photosystem I (PSI) enables photo-electron transfer and regulates photosynthesis in the bioenergetic membranes of cyanobacteria and chloroplasts. Being a multi-subunit complex, its macromolecular organization affects the dynamics of photosynthetic membranes. Here, we reveal a chloroplast PSI from the green alga Chlamydomonas reinhardtii that is organized as a homodimer, comprising 40 protein subunits with 118 transmembrane helices that provide scaffold for 568 pigments. Our cryo-EM structure identifies that the absence of PsaH and Lhca2 gives rise to a head-to-head relative orientation of the PSI-LHCI monomers in a way that is essentially different from the oligomer formation in cyanobacteria. The light-harvesting protein Lhca9 is the key element for mediating this dimerization. The interface between the monomers is lacking PsaH, and thus partially overlaps with the surface area that would bind one of the LHCII complexes in state transitions. We also define the most accurate available PSI-LHCI model at 2.3 [A] resolution, including a flexibly bound electron donor plastocyanin, and assign correct identities and orientations of all the pigments, as well as 621 water molecules that affect energy transfer pathways. FACT regulates pluripotency through distal regulation of gene expression in murine embryonic stem cells The FACT complex is a conserved histone chaperone with essential roles in transcription and histone deposition. FACT is essential in pluripotent and cancer cells, but otherwise dispensable for most mammalian cell types. FACT deletion or inhibition can block induction of pluripotent stem cells, yet the mechanism through which FACT regulates cell fate decisions remains unclear. To determine this mechanism, we used inducible depletion of FACT subunit SPT16 in murine embryonic stem cells paired with genomic factor localization, nascent transcription, and chromatin accessibility analyses. Over a timecourse of SPT16 depletion, nucleosomes invade loci bound by master pluripotency factors and gene-distal DNaseI hypersensitive sites. Simultaneously, transcription of Pou5f1 (OCT4), Sox2, Nanog, and enhancer RNAs produced at the genes' associated enhancers are downregulated, suggesting that FACT regulates expression of the pluripotency factors themselves. We find that FACT maintains cellular pluripotency through a precise nucleosome-based regulatory mechanism for appropriate expression of both coding and non-coding transcripts associated with pluripotency. Cell-type specific circadian bioluminescence rhythms in Dbp reporter mice Circadian rhythms are endogenously generated physiological and molecular rhythms with a cycle length of about 24 h. Bioluminescent reporters have been exceptionally useful for studying circadian rhythms in numerous species. Here, we report development of a reporter mouse generated by modification of a widely expressed and highly rhythmic gene encoding D-site albumin promoter binding protein (Dbp). In this line of mice, firefly luciferase is expressed from the Dbp locus in a Cre-recombinase-dependent manner, allowing assessment of bioluminescence rhythms in specific cellular populations. A mouse line in which luciferase expression was Cre-independent was also generated. The Dbp reporter alleles do not alter Dbp gene expression rhythms in liver or circadian locomotor activity rhythms. In vivo and ex vivo studies show the utility of the reporter alleles for monitoring rhythmicity. Our studies reveal cell-type specific characteristics of rhythms among neuronal populations within the suprachiasmatic nuclei ex vivo. In vivo studies show Dbp-driven bioluminescence rhythms in the liver of Albumin-Cre;DbpKI/+ "liver reporter" mice. After a shift of the lighting schedule, locomotor activity achieved the proper phase relationship with the new lighting cycle more rapidly than hepatic bioluminescence did. As previously shown, restricting food access to the daytime altered the phase of hepatic rhythmicity. Our model allowed assessment of the rate of recovery from misalignment once animals were provided with food ad libitum. These studies confirm the previously demonstrated circadian misalignment following environmental perturbations and reveal the utility of this model for minimally invasive, longitudinal monitoring of rhythmicity from specific mouse tissues. Comparative toxicogenomics of glyphosate and Roundup herbicides by mammalian stem cell-based genotoxicity assays and molecular profiling in Sprague-Dawley rats Whether glyphosate-based herbicides (GBHs) are more potent than glyphosate alone at activating cellular mechanisms, which drive carcinogenesis remains controversial. As GBHs are more cytotoxic that glyphosate, we reasoned they may also be more capable of activating carcinogenic pathways. We tested this hypothesis by comparing the effects of glyphosate with Roundup GBHs both in vitro and in vivo. First, glyphosate was compared with representative GBHs namely MON 52276 (EU), MON 76473 (UK) and MON 76207 (USA) using the mammalian stem cell-based ToxTracker system. Here, MON 52276 and MON 76473, but not glyphosate and MON 76207, activated oxidative stress and unfolded protein responses. Second, molecular profiling of liver was performed in female Sprague-Dawley rats exposed to glyphosate or MON 52276 (both at 0.5, 50, 175 mg/kg bw/day glyphosate) for 90 days. MON 52276 but not glyphosate increased hepatic steatosis and necrosis. MON 52276 and glyphosate altered the expression of genes in liver reflecting TP53 activation by DNA damage and circadian rhythm regulation. Genes most affected in liver were similarly altered in kidneys. Small RNA profiling in liver showed decreased amounts of miR-22 and miR-17 from MON 52276 ingestion. Glyphosate decreased mir-30 while miR-10 levels were increased. DNA methylation profiling of liver revealed 5,727 and 4,496 differentially methylated CpG sites between the control and glyphosate and MON 52276 exposed animals respectively. Apurinic/apyrimidinic DNA damage formation in liver was increased with glyphosate exposure. Altogether, our results show that Roundup formulations cause more biological changes linked with carcinogenesis than glyphosate. Design of Specific Primer Sets for the Detection of B.1.1.7, B.1.351, P.1, B.1.617.2 and B.1.1.519 Variants of SARS-CoV-2 using Artificial Intelligence As the COVID-19 pandemic continues, new SARS-CoV-2 variants with potentially dangerous features have been identified by the scientific community. Variant B.1.1.7 lineage clade GR from Global Initiative on Sharing All Influenza Data (GISAID) was first detected in the UK, and it appears to possess an increased transmissibility. At the same time, South African authorities reported variant B.1.351, that shares several mutations with B.1.1.7, and might also present high transmissibility. Earlier this year, a variant labelled P.1 with 17 non-synonymous mutations was detected in Brazil. Recently the World Health Organization has raised concern for the variants B.1.617.2 mainly detected in India but now exported worldwide. It is paramount to rapidly develop specific molecular tests to uniquely identify new variants. Using a completely automated pipeline built around deep learning and evolutionary algorithms techniques, we designed primer sets specific to variants B.1.1.7, B.1.351, P.1 and respectively. Starting from sequences openly available in the GISAID repository, our pipeline was able to deliver the primer sets for each variant. In-silico tests show that the sequences in the primer sets present high accuracy and are based on 2 mutations or more. In addition, we present an analysis of key mutations for SARS-CoV-2 variants. Finally, we tested the designed primers for B.1.1.7 using RT-PCR. The presented methodology can be exploited to swiftly obtain primer sets for each new variant, that can later be a part of a multiplexed approach for the initial diagnosis of COVID-19 patients. Spatial phylogenetics of Japanese ferns: Patterns, processes, and implications for conservation PremiseBiodiversity is often only measured with species richness. However, this ignores evolutionary history and is not sufficient for making conservation decisions. Here, we characterize multiple facets and drivers of biodiversity to understand how these relate to bioregions and conservation status in the ferns of Japan.

MethodsWe compiled a community dataset of 1,239 20 km x 20 km grid-cells including 672 taxa based on > 300,000 specimen records. We combined this with a phylogeny and functional traits to analyze taxonomic, phylogenetic, and functional diversity, and modeled biodiversity metrics in response to environmental factors and reproductive mode. Hierarchical clustering was used to delimit bioregions. Conservation status and threats were assessed by comparing the overlap of significantly diverse grid-cells with conservation zones and range maps of native Japanese deer.

ResultsTaxonomic richness was highest at mid-latitudes. Phylogenetic and functional diversity and phylogenetic endemism were highest in small southern islands. Relative phylogenetic and functional diversity were high at high and low latitudes, and low at mid-latitudes. Grid-cells were grouped into three (phylogenetic) or four (taxonomic) major bioregions. Temperature and apomixis were identified as drivers of biodiversity patterns. Conservation status was generally high for grid-cells with significantly high biodiversity, but the threat due to herbivory by deer was greater for taxonomic richness than other metrics.

ConclusionsOur integrative approach reveals previously undetected patterns and drivers of biodiversity in the ferns of Japan. Future conservation efforts should recognize that threats can vary by biodiversity metric and consider multiple multiple metrics when establishing conservation priorities. RNA degradation of blood reveals a time diagnostic window in a controlled environment Determining the time since deposition (TSD) of bloodstains would provide forensic scientists with critical information regarding the timeline of the events involving bloodshed. The physicochemical changes occurring to biomolecules as a bloodstain degrades can be used to approximate the TSD of bloodstains. Our study aims to quantify the timewise degradation trends and temperature dependence found in total RNA from bloodstains without the use of amplification, expanding the scope of the RNA TSD research which has previously targeted mRNA molecules. Whole bovine blood was stored in plastic microcentrifuge tubes at 21{degrees}C or 4{degrees}C and tested over different timepoints spanning one week. Nine RNA metrics were visually assessed and quantified using linear and mixed models; the RNA Integrity Number equivalent (RINe) and the DV200 demonstrated strong negative trends over time and statistical independence. The RINe model fit was high (R2 = 0.60), and while including the biological replicate as a random effect increased the fit for all RNA metrics, no significant differences were found between biological replicates stored at the same temperature for the RINe and DV200 metrics. Importantly, this suggests that these standardized metrics can likely be directly compared between scenarios and individuals, with DV200 having an inflection point at [~]28 hrs. This study provides a novel approach for blood TSD estimates, producing metrics that are not affected by inter-individual variation and improving our understanding of the rapid degradation occurring in bloodstains.

HIGHLIGHTSO_LIAmplification-free analysis of total RNA in degrading bloodstains.
C_LIO_LIShort-term RNA degradation assessment using high-resolution size measurements.
C_LIO_LITotal RNA quality and quantity metrics were assessed across a one-week.
C_LIO_LITotal RNA quality metrics demonstrated the strongest timewise trends.
C_LIO_LIBiological replicates produced similar results for RNA quality metrics.
C_LI Hidden dynamics of proteasome autoregulation discovered by cryo-EM data-driven deep learning The cellular functions are executed by macromolecular complexes in nonequilibrium dynamic processes, which exhibit a vast diversity of conformational states. Solving conformational continuum of important complexes at atomic level remains a key challenge in the methodology of computational biophysics. Here we introduce a deep-learning framework, named AlphaCryo4D, which enables atomic-level cryogenic electron microscopy reconstructions of nonequilibrium conformational continuum. AlphaCryo4D integrates 3D deep residual learning with manifold embedding of energy landscapes, which simultaneously improves 3D classification accuracy and reconstruction resolution via an energy-based particle-voting algorithm. Applications of this methodology to six experimental datasets demonstrated its generality in breaking resolution barrier of visualizing dynamic components of functional complexes, in choreographing continuous inter-subunit motions and in exploring their  hidden conformational space. Importantly, AlphaCryo4D identified 64 conformers of the substrate-bound human 26S proteasome and reconstituted single-nucleotide-exchange dynamics of proteasomal AAA-ATPase motor during translocation initiation, which together illuminates a grand hierarchical allostery for proteasome autoregulation. Integrated analysis of cervical squamous cell carcinoma cohorts from three continents reveals conserved subtypes of prognostic significance. Human papillomavirus (HPV)-associated cervical cancer represents one of the leading causes of cancer death worldwide. Although low-middle income countries are disproportionately affected, our knowledge of the disease predominantly originates from populations in high-income countries. Using the largest multi-omic analysis of cervical squamous cell carcinoma (CSCC) to date, totalling 643 tumours and representing patient populations from the USA, Europe and Sub-Saharan Africa, we identify two CSCC subtypes (C1 and C2) with differing prognosis. C1 tumours are largely HPV16-driven, display increased cytotoxic T-lymphocyte infiltration and frequently harbour PIK3CA and EP300 mutations. C2 tumours are associated with shorter overall survival, are frequently driven by HPVs from the HPV18-containing alpha-7 clade, harbour alterations in the Hippo signalling pathway and increased expression of immune checkpoint genes, B7-H3 (also known as CD276) and NT5E (also known as CD73) and PD-L2 (also known as PDCD1LG2). In conclusion, we identify two novel, therapy-relevant CSCC subtypes that share the same defining characteristics across three geographically diverse cohorts. Gain-of-function cardiomyopathic mutations in RBM20 rewire splicing regulation and re-distribute ribonucleoprotein granules within processing bodies RNA binding motif protein 20 (RBM20) is a key regulator of alternative splicing in the heart, and its mutation leads to malignant dilated cardiomyopathy (DCM). To understand the mechanism of RBM20-associated DCM, we engineered isogenic human induced pluripotent stem cells (iPSCs) with heterozygous or homozygous DCM-associated missense mutations in RBM20 (R636S) as well as RBM20 knockout (KO) iPSCs. iPSC-derived engineered heart tissues made from these cell lines recapitulated contractile dysfunction of RBM20-associated DCM and revealed greater dysfunction with missense mutations than KO. Analysis of RBM20 RNA binding by eCLIP revealed a gain-of-function preference of mutant RBM20 for 3' UTR sequences that are shared with amyotrophic lateral sclerosis (ALS) and processing-body associated RNA binding proteins (FUS, DDX6). Deep RNA sequencing revealed that the RBM20 R636S mutant has unique gene, splicing, polyadenylation and circular RNA defects that differ from RBM20 KO, impacting distinct cardiac signaling pathways. Splicing defects specific to KO or R636S mutations were supported by data from R636S gene-edited pig hearts and eCLIP. Super-resolution microscopy verified that mutant RBM20 maintains limited nuclear localization potential; rather, the mutant protein associates with cytoplasmic processing bodies (DDX6) under basal conditions, and with stress granules (G3BP1) following acute stress. Taken together, our results highlight a novel pathogenic mechanism in cardiac disease through splicing-dependent and -independent pathways that are likely to mediate differential contractile phenotypes and stress-associated heart pathology. Anaerobic Thermophilic Co-digestion of Sugarcane Biorefinery Residues Enhances Biogas Production This is a pioneer study evaluating the methane (CH4) production potential from residues of integrated 1st (vinasse and filter cake) and 2nd (deacetylation pretreatment liquor from straw) generation (1G2G) sugarcane biorefinery, providing a fully chemical characterization of them and their relation with the anaerobic digestion (AD) process. Small-scale assays provided fundamentals for basing the co-digestion optimization by assessing the optimal co-substrates synergistic conditions. Biochemical Methane Potential (BMP) tests showed co-digestion enhanced CH4 yield of isolated substrates, reaching up to 605 NmLCH4 gVS-1. The association of vinasse and deacetylation liquor as co-substrates increased the BMP by ~38% mostly by nutritionally benefiting the methanogenic activity. The kinetic analysis confirmed that the deacetylation liquor was the co-substrate responsible for improving the CH4 production in the co-digestion systems due to the highest CH4 conversion rate. The alkaline characteristic of the liquor (pH~12) also prevented alkalizing from being added to the co-digestion, an input that normally makes the process economically unfeasible to implement on an industrial scale due to the large quantities required for buffering the reactor. The filter cake had the lowest BMP (262 NmLCH4 gVS-1) and digestibility ([&le;] 40%), further limited by the required stirring to improve the mass transfer of biochemical reactions. The present study drives towards more sustainable use of vinasse, the most voluminous waste from the sugarcane industry, and lignin-rich residues derived from pre-treatment alkaline methods, aiming at an energy-efficient utilization, by at least 16% when compared to the traditional vinasse AD. The experimental and modeling elements from this work indicated the lignin-rich liquor is the main responsible for putting the co-digestion as a disruptive technological arrangement within the 1G2G sugarcane biorefineries, reinforcing the biogas production as the hub of the bioeconomy in the agroindustrial sector. Genomic insights into longan evolution from a chromosome-level genome assembly and population genomics of longan accessions Longan (Dimocarpus longan) is a subtropical fruit best known for its nutritious fruit and regarded as a precious tonic and traditional medicine since ancient times. High-quality chromosome-scale genome assembly is valuable for functional genomic study and genetic improvement of longan. Here, we report a chromosome-level reference genome sequence for longan cultivar JDB with an assembled genome of 455.5 Mb in size anchored to fifteen chromosomes, representing a significant improvement of contiguity (contig N50=12.1 Mb, scaffold N50= 29.5 Mb) over a previous draft assembly. A total of 40,420 protein-coding genes were predicted in D. longan genome. Synteny analysis suggests longan shares the widespread gamma event with core eudicots, but has no other whole genome duplications. Comparative genomics showed that D. longan genome experienced significant expansions of gene families related to phenylpropanoid biosynthesis and UDP-glucosyltransferase. Deep genome sequencing analysis of longan cultivars identified longan biogeography as a major contributing factor for genetic diversity, and revealed a clear population admixture and introgression among cultivars of different geographic origins, postulating a likely migration trajectory of longan overall confirmed by existing historical records. Finally, genome-wide association studies (GWAS) of longan cultivars identified quantitative trait loci (QTL) for six different fruit quality traits and revealed a shared QTL containing three genes for total soluble solid and seed weight. The chromosome-level reference genome assembly, annotation and population genetic resource for D. longan will facilitate the molecular studies and breeding of desirable longan cultivars in the future.