title: Serum glycoprotein markers in non-alcoholic steatohepatitis and hepatocellular carcinoma

abstract:
Fatty liver disease progresses through stages of fat accumulation and inflammation to non-alcoholic steatohepatitis (NASH), fibrosis and cirrhosis and eventually hepatocellular carcinoma (HCC). Currently available diagnostic tools for HCC lack sensitivity and specificity and deliver little value to patients. In this study, we investigated the use of circulating serum glycoproteins to identify a panel of potential prognostic markers that may be indicative of progression from the healthy state to NASH and further to HCC. Serum samples were processed using a standard pre-analytical sample preparation protocol and were analyzed using a novel high throughput glycoproteomics platform. We analyzed 413 glycopeptides, representing 57 abundant serum proteins and compared among the three phenotypes. Our initial dataset contained healthy, NASH, and HCC serum samples. We analyzed normalized abundance of common glycoforms and found 40 glycopeptides with statistically significant differences in abundances in NASH and HCC compared to controls. Summary level relative abundance of core-fucosylated, sialylated and branched glycans containing glycopeptides were higher in NASH and HCC as compared to controls. We replicated some of our findings in an independent set of samples of individuals with benign liver conditions and HCC, respectively. Our results may be of value in the management of liver diseases.

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category: biochemistry


title: Structural mapping of antibody landscapes to human betacoronavirus spike proteins

abstract:
Preexisting immunity against seasonal coronaviruses (CoV) represents an important variable in predicting antibody responses and disease severity to Severe Acute Respiratory Syndrome CoV-2 (SARS-2) infections. We used electron microscopy based polyclonal epitope mapping (EMPEM) to characterize the antibody specificities against {beta}-CoV spike proteins in sera from healthy donors (HDs) or SARS-2 convalescent donors (CDs). We observed that most HDs possessed antibodies specific to seasonal human CoVs (HCoVs) OC43 and HKU1 spike proteins while the CDs showed reactivity across all human {beta}-CoVs. Detailed molecular mapping of spike-antibody complexes revealed epitopes that were differentially targeted by antibodies in preexisting and convalescent serum. Our studies provide an antigenic landscape to {beta}-HCoV spikes in the general population serving as a basis for cross-reactive epitope analyses in SARS-2 -infected individuals.

One-Sentence summaryWe present the epitope mapping of polyclonal antibodies against beta-coronavirus spike proteins in human sera.

category: immunology


title: A spike-sorting method based on hierarchical clustering to discriminate extracellularly recorded simple spikes and complex spikes from cerebellar Purkinje cells

abstract:
BackgroundIn the cerebellar cortex, Purkinje cells are the only output neurons and exhibit two types of discharge. Most Purkinje cell discharges are simple spikes, which are commonly appearing action potentials exhibiting a rich variety of firing patterns with a rate of up to 400 Hz. More infrequent discharges are complex spikes, which consist of a short burst of impulses accompanied by a massive increase in dendritic Ca2+ with a firing rate of around 1 Hz. The discrimination of these spikes in extracellular single-unit recordings is not always straightforward, as their waveforms vary depending on recording conditions and intrinsic fluctuations.

New MethodTo discriminate complex spikes from simple spikes in the extracellular single-unit data, we developed a semiautomatic spike-sorting method based on divisive hierarchical clustering.

ResultsQuantitative evaluation using parallel in vivo two-photon Ca2+ imaging of Purkinje cell dendrites indicated that 96.6% of the complex spikes were detected using our spike-sorting method from extracellular single-unit recordings obtained from anesthetized mice.

Comparison with Existing Method(s)No reports have conducted a quantitative evaluation of spike-sorting algorithms used for the classification of extracellular spikes recorded from cerebellar Purkinje cells.

ConclusionsOur method could be expected to contribute to research in information processing in the cerebellar cortex and the development of a fully automatic spike-sorting algorithm by providing ground-truth data useful for deep learning.

HighlightsO_LIA spike-sorting algorithm on hierarchical clustering was developed.
C_LIO_LIIt was applied to extracellular recordings from cerebellar Purkinje cells.
C_LIO_LIComplex and simple spikes were discriminated based on their waveforms.
C_LIO_LISpike-sorting performance was quantified using in vivo Ca2+ imaging data.
C_LIO_LIThe algorithm isolated over 96.6% of complex spikes.
C_LI

category: neuroscience


title: TES-1/Tes protects junctional actin networks under tension from self-injury during epidermal morphogenesis in the C. elegans embryo

abstract: During embryonic morphogenesis, the integrity of epithelial tissues depends on the ability of cells in tissue sheets to undergo rapid changes in cell shape while preventing self-injury to junctional actin networks. LIM domain-containing repeat (LCR) proteins are recruited to sites of strained actin filaments in cultured cells [1-3], and are therefore promising candidates for mediating self-healing of actin networks, but whether they play similar roles in living organisms has not been determined. Here, we establish roles for Caenorhabditis elegans TES-1/Tes, an actin-binding LCR protein present at apical junctions, during epithelial morphogenesis. TES-1::GFP is recruited to apical junctions during embryonic elongation, when junctions are under tension; in embryos in which stochastic failure of cell elongation occurs, TES-1 is only strongly recruited to junctions in cells that successfully elongate, and recruitment is severely compromised in genetic backgrounds in which cell shape changes do not successfully occur. tes-1 mutant embryos display junctional F-actin defects, and loss of TES-1 strongly enhances tension-dependent injury of junctional actin networks in hypomorphic mutant backgrounds for CCC components, suggesting that TES-1 helps to prevent self-injury of junctional actin networks during rapid cell shape change. Consistent with such role, a fragment of TES-1 containing its LIM domains localizes to stress fiber strain sites (SFSS) in cultured vertebrate cells. Together, these data establish TES-1 as a tension-sensitive stabilizer of the junctional actin cytoskeleton during embryonic morphogenesis.

category: developmental biology


title: Relative Risk Assessment of Ecological Areas with the Highest Potential Impact of Underwater Released Exhaust CO2 from Innovative Ships

abstract:
Applying underwater released exhaust gas as  air lubrication along the ships hull to reduce the energy consumption is under development. However, this direct emission to the water could pose a risk to the local marine environment, especially in shipping-dense areas. Specifically, CO2, a dominant component in the exhaust gas, has the potency to enhance algal blooms and cause acidification. This study provides the first relative risk assessment of ships with underwater release exhaust gas systems on a global scale, taking into account local water conditions and shipping intensity. Risk was characterized for 262 marine ecoregions by plotting the expected CO2 emission from ships to water against the estimated vulnerability to acidification and algal blooms. The vulnerability of each ecoregion was assessed based on background dissolved inorganic carbon (DIC) level, chlorophyll-a concentrations and total alkalinity. The results reveal that areas with relatively high vulnerability are mainly located above 30{degrees} N latitude. The Yellow Sea, Southern China Sea, and North Sea come out as relatively high risk areas. Looking in more detail to European high-risk ecoregions, the highest risk levels are found in areas with dense shipping lanes and maritime chokepoints, e.g. the Strait of Dover and the Strait of Gibraltar. This was the first attempt to make such a risk assessment and the outcome is only indicative. In a next phase additional parameters, such as water currents and biological composition of the ecosystem should be included.

Key PointsO_LIThe first global environmental risk assessment was made for innovative ships that apply underwater released exhaust gas as  air lubrication
C_LIO_LIEcoregions with relatively high vulnerability to acidification and algal blooms are mainly located above 30{degrees} N latitude
C_LIO_LIThe risk for the marine ecosystem of the underwater released exhaust CO2 from ships is limited to specific shipping-dense areas
C_LI

category: ecology


title: Simulation-based inference of evolutionary parameters from adaptation dynamics using neural networks

abstract: The rate of adaptive evolution depends on the rate at which beneficial mutations are introduced into a population and the fitness effects of those mutations. The rate of beneficial mutations and their expected fitness effects is often difficult to empirically quantify. As these two parameters determine the pace of evolutionary change in a population, the dynamics of adaptive evolution may enable inference of their values. Copy number variants (CNVs) are a pervasive source of heritable variation that can facilitate rapid adaptive evolution. Previously, we developed a locus-specific fluorescent CNV reporter to quantify CNV dynamics in evolving populations maintained in nutrient-limiting conditions using chemostats. Here, we use the observed CNV adaptation dynamics to estimate the rate at which beneficial CNVs are introduced through de novo mutation and their fitness effects using simulation-based Bayesian likelihood-free inference approaches. We tested the suitability of two evolutionary models: a standard Wright-Fisher model and a chemostat growth model. We evaluated two likelihood-free inference algorithms: the well-established Approximate Bayesian Computation with Sequential Monte Carlo (ABC-SMC) algorithm, and the recently developed Neural Posterior Estimation (NPE) algorithm, which applies an artificial neural network to directly estimate the posterior distribution. By systematically evaluating the suitability of different inference methods and models we show that NPE has several advantages over ABC-SMC and that a Wright-Fisher evolutionary model suffices in most cases. Using our validated inference framework, we estimate the CNV formation rate at the GAP1 locus in yeast as 10-4.7 -10-4 per cell division, and a selection coefficient of 0.04 - 0.1 per generation for GAP1 CNVs in glutamine-limited chemostats. We experimentally validated our estimates using barcode lineage tracking and pairwise fitness assays. Our results are consistent with a high beneficial CNV supply rate that is 10-fold greater than the estimated rates of beneficial single-nucleotide mutations, explaining their outsized importance in rapid adaptive evolution. More generally, our study demonstrates the utility of novel simulation-based likelihood-free inference methods for inferring the rates and effects of evolutionary processes from empirical data.

category: evolutionary biology


title: Tissue engineered organoid co-culture model of the cycling human endometrium in a fully-defined synthetic extracellular matrix

abstract:
The human endometrium is a mucosal barrier that undergoes cycles of growth, differentiation, and breakdown in response to sex hormone fluctuations. Dynamic tissue responses to hormones are primarily driven by epithelial-stromal communication and its dysregulation is linked to myriad gynecological disorders. The lack of robust in vitro models for the long-term 3D co-culture of patient-derived endometrial epithelial and stromal cells hinders dissection of this crosstalk and thus impairs progress in disease treatment. Here, we describe a versatile synthetic extracellular matrix tailored to the endometrium that enables the in vitro modeling of human healthy and disease states across the menstrual cycle. We used a tissue-inspired approach to semi-empirically screen a parameter space that encompasses the biophysical and molecular features of the endometrial microenvironment. Leveraging cell-specific integrin expression profiles, we defined a modular polyethylene glycol (PEG)-based hydrogel that fosters hormone-driven expansion and differentiation of epithelial organoids co-cultured with stromal cells. Characteristic morphological and molecular responses of each cell type to hormone changes were observed when cells were co-encapsulated in hydrogels tuned to a stiffness regime similar to the native tissue and functionalized with a collagen-derived adhesion peptide (GFOGER) and a fibronectin-derived peptide (PHSRN-K-RGD). Using transcriptomic and functional assays, we demonstrate the systems ability to recapitulate menstrual-cycle specific reproductive events and identified that inflammation-induced dysregulation of epithelial proliferation is mediated via the stromal compartment. Altogether, we demonstrate the development of a fully synthetic matrix to sustain the dynamic changes of the endometrial microenvironment and support its applications to understand endometriotic diseases.

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category: bioengineering


title: MoSBi: Automated signature mining for molecular stratification and subtyping

abstract: The improving access to increasing amounts of biomedical data provides completely new chances for advanced patient stratification and disease subtyping strategies. This requires computational tools that produce uniformly robust results across highly heterogeneous molecular data. Unsupervised machine learning methodologies are able to discover de-novo patterns in such data. Biclustering is especially suited by simultaneously identifying sample groups and corresponding feature sets across heterogeneous omics data. The performance of available biclustering algorithms heavily depends on individual parameterization and varies with their application. Here, we developed MoSBi (Molecular Signature identification using Biclustering), an automated multi-algorithm ensemble approach that integrates results utilizing an error model-supported similarity network. We evaluated the performance of MoSBi on transcriptomics, proteomics and metabolomics data, as well as synthetic datasets covering various data properties. Profiting from multi-algorithm integration, MoSBi identified robust group and disease specific signatures across all scenarios overcoming single algorithm specificities. Furthermore, we developed a scalable network-based visualization of bicluster communities that support biological hypothesis generation. MoSBi is available as an R package and web-service to make automated biclustering analysis accessible for application in molecular sample stratification.

category: bioinformatics


title: interpolatedXY: a two-step strategy to normalise DNA methylation microarray data avoiding sex bias

abstract:
MotivationData normalization is an essential step to reduce technical variation within and between arrays. Due to the different karyotypes and the effects of X chromosome inactivation, females and males exhibit distinct methylation patterns on sex chromosomes, thus it poses a significant challenge to normalise sex chromosome data without introducing bias. Currently, existing methods do not provide unbiased solutions to normalise sex chromosome data, usually, they just process autosomal and sex chromosomes indiscriminately.

ResultsHere, we demonstrate that ignoring this sex difference will lead to introducing artificial sex bias, especially for thousands of autosomal CpGs. We present a novel two-step strategy (interpolatedXY) to address this issue, which is applicable to all quantile-based normalisation methods. By this new strategy, the autosomal CpGs are first normalised independently by conventional methods, such as funnorm or dasen; then the corrected methylation values of sex chromosome linked CpGs are estimated as the weighted average of their nearest neighbours on autosomes. The proposed two-step strategy can also be applied to other non-quantile-based normalisation methods, as well as other array-based data types. Moreover, we propose a useful concept: the sex explained fraction of variance, to quantitatively measure the normalisation effect.

AvailabilityThe proposed methods are available by calling the function  adjustedDasen or  adjustedFunnorm in the latest wateRmelon package (https://github.com/schalkwyk/wateRmelon), with methods compatible with all the major workflows, including minfi.

Contactxzhai@essex.ac.uk; lschal@essex.ac.uk

Supplementary informationSupplementary data are available at...


title: Open Imputation Server provides secure Imputation services with provable genomic privacy

abstract:
SummaryAs DNA sequencing data is available for personal use, genomic privacy is becoming a major challenge. Nevertheless, high-throughput genomic data analysis outsourcing is performed using pipelines that tend to overlook these challenges.

ResultsWe present a client-server-based outsourcing framework for genotype imputation, an important step in genomic data analyses. Genotype data is encrypted by the client and encrypted data are used by the server that never observes the data in plain. Cloud-based framework can benefit from virtually unlimited computational resources while providing provable confidentiality. We demonstrate servers utility from several aspects using genotype dataset from the 1000 Genomes datasets. First, we benchmark the accuracy of common variant imputation in comparison to BEAGLE, a state-of-the-art imputation method. We also provide the detailed time requirements of the server to showcase scaling of time usage in different steps of imputation. We also present a simple correlation metric that can be used to estimate imputation accuracy using only the reference panels. This is important for filtering the variants in downstream analyses. As a further demonstration and a different use case, we performed a simulated genomewide association study (GWAS) using imputed and known genotypes and highlight potential utility of the server for association studies. Overall, our study present multiple lines of evidence for usability of secure imputation service.

AvailabilityServer is publicly available at https://www.secureomics.org/OpenImpute. Users can anonymously test and use imputation server without registration.

ContactArif.O.Harmanci@uth.tmc.edu


title: Deformation-induced actuation of cells in asymmetric periodic flow fields

abstract: Analyzing and sorting particles and/or biological cells in microfluidic devices is a topical problem in soft-matter and biomedical physics. An easy and rapid screening of the deformation of individual cells in constricted microfluidic channels allows, for example, the identification of sick or aberrant cells with altered mechanical properties, even in vast cell ensembles. The subsequently desired softness-specific segregation of cells is, however, still a major challenge. Moreover, aiming at an intrinsic and unsupervised approach raises a very general question: How can one achieve a softnessdependent net migration of particles in a microfluidic channel? Here we show that this is possible by exploiting a deformation-induced actuation of soft cells in asymmetric periodic flow fields in which rigid beads show a vanishing net drift.

category: biophysics


title: Climatic clustering and longitudinal analysis with impacts on food, bioenergy, and pandemics

abstract: Predicted growth in world population will put unparalleled stress on the need for sustainable energy and global food production, as well as increase the likelihood of future pandemics. In this work, we identify high-resolution environmental zones in the context of a changing climate and predict longitudinal processes relevant to these challenges. We do this using exhaustive vector comparison methods that measure the climatic similarity between all locations on earth at high geospatial resolution. The results are captured as networks, in which edges between geolocations are defined if their historical climates exceed a similarity threshold. We then apply Markov clustering and our novel Correlation of Correlations method to the resulting climatic networks, which provides unprecedented agglomerative and longitudinal views of climatic relationships across the globe. The methods performed here resulted in the fastest (9.37 x 1018 operations/sec) and one of the largest (168.7 x 1021 operations) scientific computations ever performed, with more than 100 quadrillion edges considered for a single climatic network. Correlation and network analysis methods of this kind are widely applicable across computational and predictive biology domains, including systems biology, ecology, carbon cycles, biogeochemistry, and zoonosis research.

category: systems biology


title: Dynamical modeling of the H3K27 epigenetic landscape in mouse embryonic stem cells

abstract: The Polycomb system via the methylation of the lysine 27 of histone H3 (H3K27) plays central roles in the silencing of many lineage-specific genes during development. Recent experimental evidence suggested that the recruitment of histone modifying enzymes like the Polycomb repressive complex 2 (PRC2) at specific sites and their spreading capacities from these sites are key to the establishment and maintenance of a proper epigenomic landscape around Polycomb-target genes. Here, to test whether such mechanisms, as a minimal set of qualitative rules, are quantitatively compatible with data, we developed a mathematical model that can predict the locus-specific distributions of H3K27 modifications based on previous biochemical knowledge. Within the biological context of mouse embryonic stem cells, our model showed quantitative agreement with experimental profiles of H3K27 acetylation and methylation around Polycomb-target genes in wild-type and mutants. In particular, we demonstrated the key role of the reader-writer module of PRC2 and of the competition between the binding of activating and repressing enzymes in shaping the H3K27 landscape around transcriptional start sites. The predicted dynamics of establishment and maintenance of the repressive trimethylated H3K27 state suggest a slow accumulation, in perfect agreement with experiments. Our approach represents a first step towards a quantitative description of PcG regulation in various cellular contexts and provides a generic framework to better characterize epigenetic regulation in normal or disease situations.


title: Metacognition and the effect of incentive motivation in two compulsive disorders: gambling disorder and obsessive-compulsive disorder

abstract: Compulsivity is a common phenotype amongst various psychiatric disorders, such as obsessive-compulsive disorder (OCD) and gambling disorder (GD). Deficiencies in metacognition, such as the inability to properly estimate ones own performance via well-calibrated confidence judgments could contribute to pathological decision-making in these psychiatric disorders. Earlier research has indeed suggested that OCD and GD patients reside at opposite ends of the confidence spectrum, with OCD patients exhibiting underconfidence, and GD patients exhibiting overconfidence. Recently, several studies established that motivational states (e.g. monetary incentives) influence metacognition, with gain (respectively loss) prospects increasing (respectively decreasing) confidence judgments. Here, we reasoned that the OCD and GD symptomatology might correspond to an exacerbation of this interaction between metacognition and motivational states. We hypothesized GDs overconfidence to be exaggerated during gain prospects, while OCDs underconfidence to be worsened in loss context, which we expected to see represented in ventromedial prefrontal cortex (VMPFC) blood-oxygen-level-dependent (BOLD) activity. We tested those hypotheses in a task-based functional magnetic resonance imaging (fMRI) design. Our initial analyses showed increased confidence levels for GD versus OCD patients, that could partly be explained by sex and IQ. Although our primary analyses did not support the hypothesized interaction between incentives and groups, exploratory analyses did show increased confidence in GD patients specifically in gain context. fMRI analyses confirmed a central role for VMPFC in the processing of confidence and incentives, but with no differences between the clinical samples. The trial is registered in the Dutch Trial Register (Trial NL6171, registration number: NTR6318) (https://www.trialregister.nl/trial/6171).


title: A morphometric double dissociation: cortical thickness is more related to aging; surface area is more related to cognition

abstract: The thickness and surface area of cortex are genetically distinct aspects of brain structure, and may be affected differently by age. However, their potential to differentially predict age and cognitive abilities has been largely overlooked, likely because they are typically aggregated into the commonly used measure of volume. In a large sample of healthy adults (N=647, aged 18-88), we investigated the brain-age and brain-cognition relationships of thickness, surface area, and volume, plus five additional morphological shape metrics. Cortical thickness was the metric most strongly associated with age cross-sectionally, as well as exhibiting the steepest longitudinal change over time (subsample N=261, aged 25-84). In contrast, surface area was the best single predictor of age-residualized cognitive abilities (fluid intelligence), and changes in surface area were most strongly associated with cognitive change over time. These findings were replicated in an independent dataset (N=1345, aged 18-93). Our results suggest that cortical thickness and surface area make complementary contributions the age-brain-cognition triangle, and highlight the importance of considering these volumetric components separately.


title: Multivariate pattern analysis reveals location specific aftereffects of 10Hz motor cortex transcranial alternating current stimulation

abstract:
BackgroundTranscranial alternating current stimulation (tACS) may induce frequency-specific aftereffects on brain oscillations in the stimulated location, which could serve as evidence for region-specific neuroplasticity. Aftereffects of tACS on the motor system remain unknown.

ObjectiveTo find evidence for aftereffects in short EEG segments following tACS to two critical nodes of the motor network, namely, left motor cortex (lMC) and right cerebellum (rCB). We hypothesized that aftereffects of lMC will be stronger in and around lMC compared to both active stimulation of rCB, as well as inactive ("sham") control conditions.

MethodsTo this end, we employed multivariate pattern analysis (MVPA), and trained a classifier to distinguish between EEG signals following each of the three stimulation protocols. This method accounts for the multitude facets of the EEG signal and thus is more sensitive to subtle modulation of the EEG signal.

ResultsEEG signals in both theta ({theta}, 4-8Hz) and alpha (, 8-13Hz) were better classified to lMC-tACS compared to rCB-tACS/sham, in and around lMC-tACS stimulation locations (electrodes FC3 and CP3). This effect was associated with a decrease in power following tACS. Source reconstruction revealed significant differences in premotor cortex but not in primary motor cortex as the computational model suggested. Correlation between classification accuracies in {theta} and  in lMC-tACS was stronger compared to rCB-tACS/sham, suggesting cross-frequency effects of tACS. Nonetheless, {theta}/ phase-coupling did not differ between stimulation protocols.

ConclusionsSuccessful classification of EEG signals to left motor cortex using MVPA revealed focal tACS aftereffects on the motor cortex, indicative of region-specific neuroplasticity.


title: Cx43 carboxyl terminal domain determines AQP4 and Cx30 endfoot organization and blood brain barrier permeability

abstract: The neurovascular unit (NVU) consists of cells intrinsic to the vessel wall, the endothelial cells and pericytes, and astrocyte endfeet that surround the vessel but are separated from it by basement membrane. Endothelial cells are primarily responsible for creating and maintaining blood-brain-barrier (BBB) tightness, but astrocytes contribute to the barrier through paracrine signaling to the endothelial cells and by forming the glia limitans. Gap junctions (GJs) between astrocyte endfeet are composed of connexin 43 (Cx43) and Cx30, which form plaques between cells. GJ plaques formed of Cx43 do not diffuse laterally in the plasma membrane and thus potentially provide stable organizational features to the endfoot domain, whereas GJ plaques formed of other connexins and of Cx43 lacking a large portion of its cytoplasmic carboxyl terminus are quite mobile. In order to examine the organizational features that immobile GJs impose on the endfoot, we have used super-resolution confocal microscopy to map number and sizes of GJ plaques and aquaporin (AQP)-4 channel clusters in the perivascular endfeet of mice in which astrocyte GJs (Cx30, Cx43) were deleted or the carboxyl terminus of Cx43 was truncated. To determine if blood-brain-barrier integrity was compromised in these transgenic mice, we conducted perfusion studies under elevated hydrostatic pressure using horseradish peroxide as a molecular probe enabling detection of micro-hemorrhages in brain sections. These studies revealed that microhemorrhages were more numerous in mice lacking Cx43 or its carboxyl terminus. In perivascular domains of cerebral vessels, we found that density of Cx43 GJs was higher in the truncation mutant, while GJ size was smaller. Density of perivascular particles formed by AQP4 and its extended isoform AQP4ex was inversely related to the presence of full length Cx43, whereas the ratio of sizes of the particles of the AQP4ex isoform to total AQP4 was directly related to the presence of full length Cx43. Confocal analysis showed that Cx43 and Cx30 were substantially colocalized in astrocyte domains near vasculature of truncation mutant mice. These results showing altered distribution of some astrocyte nexus components (AQP4 and Cx30) in Cx43 null mice and in a truncation mutant, together with leakier cerebral vasculature, support the hypothesis that localization and mobility of gap junction proteins and their binding partners influences organization of astrocyte endfeet which in turn impacts BBB integrity of the NVU.


title: Hippocampal CA1 pyramidal cells do not receive monosynaptic input from thalamic nucleus reuniens

abstract: Midline thalamic nuclei play a critical role in cognitive functions such as memory, decision-making and spatial navigation, by facilitating communication between the many brain regions involved in these processes. One canonical feature of thalamic interactions with the cortex or hippocampus appears to be that the thalamus receives input from, and projects to, excitatory neurons. Thalamic nucleus reuniens (NRe) is located on the midline and is viewed primarily as a relay from prefrontal cortex to hippocampal and entorhinal areas, although these connections are poorly defined at the cellular and synaptic level. Using electrophysiology and monosynaptic circuit-tracing, we found that pyramidal cells in CA1 receive no direct input from NRe. This contrasts starkly with prefrontal cortex, subiculum and entorhinal cortex, and indicates that NRe inputs to CA1 primarily drive local inhibition and not excitation they do in the other regions. The NRe to CA1 projection is thus a unique thalamic projection and as such is raising important questions about the function of NRe-mediated prefrontal control of the hippocampus.


title: Representation of motion concepts in occipitotemporal cortex: fMRI activation, decoding and connectivity analyses

abstract: Embodied theories of semantic cognition predict that brain regions involved in motion perception are engaged when people comprehend motion concepts expressed in language. Left lateral occipitotemporal cortex (LOTC) is implicated in both motion perception and motion concept processing but prior studies have produced mixed findings on which parts of this region are engaged by motion language. We scanned participants performing semantic judgements about sentences describing motion events and static events. We performed univariate analyses, multivariate pattern analyses (MVPA) and psychophysiological interaction (PPI) analyses to investigate the effect of motion on activity and connectivity in different parts of LOTC. In multivariate analyses that decoded whether a sentence described motion or not, the whole of LOTC showed above-chance level performance, with performance exceeding that of other brain regions. Univariate ROI analyses found the middle part of LOTC was more active for motion events than static ones. Finally, PPI analyses found that when processing motion events, the middle and posterior parts of LOTC (overlapping with motion perception regions), increased their connectivity with cognitive control regions. Taken together, these results indicate that the whole of the LOTC responded differently to motion vs. static events, and that these effects are most pronounced in more posterior sites. These findings are consistent with embodiment accounts of semantic processing, and suggest that understanding verbal descriptions of motion engages areas of the occipitotemporal cortex involved in perceiving motion.


title: Knockout of a single Sox gene resurrects an ancestral cell type in the sea anemone Nematostella vectensis

abstract: Cnidocytes are the explosive stinging cells found only in cnidarians (corals, jellyfish, etc). Specialized for prey capture and defense, cnidocytes are morphologically complex and vary widely in form and function across taxa; how such diversity evolved is unknown. Using CRISPR/Cas9-mediated genome editing in the burrowing sea anemone Nematostella vectensis, we show that a single transcription factor (NvSox2) acts as a binary switch between two alternative cnidocyte fates. Knockout of NvSox2 caused a complete transformation of nematocytes (piercing cells) into spirocytes (ensnaring cells). The type of spirocyte induced by NvSox2 knockout (robust spirocyte) is not normally found in N. vectensis but is common in sea anemones from other habitats. Homeotic control of cell fate provides a mechanistic explanation for the discontinuous distribution of cnidocyte types across cnidarians and demonstrates how simple counts of cell types can underestimate biodiversity.


title: The history of body size evolution in termites

abstract: Termites are social cockroaches. Because non-termite cockroaches are larger than basal termite lineages, which themselves include large termite species, it has been proposed that termites experienced a unidirectional body size reduction since they evolved eusociality. However, the validity of this hypothesis remains untested in a phylogenetic framework. Here, we reconstructed termite body size evolution using head width measurements of 1638 modern and fossil termite species. We found that the unidirectional body size reduction model was only supported by analyses excluding fossil species. Analyses including fossil species suggested that body size diversified along with speciation events and estimated that the size of the common ancestor of modern termites was comparable to that of modern species. Our analyses further revealed that body size variability among species, but not body size reduction, is associated with features attributed to advanced termite societies. Our results suggest that miniaturization took place at the origin of termites, while subsequent complexification of termite societies did not lead to further body size reduction.


title: Isotopic evidence for population dynamics in the Central Italian Copper Age and Bronze Age.

abstract: The mobility patterns in the Italian peninsula during prehistory are still relatively unknown. The excavation of the Copper Age and Bronze Age deposits in La Sassa cave (Sonnino, Italy) allowed to broaden the knowledge about some local and regional dynamics. We employed a multi-disciplinary approach, including stable (carbon and nitrogen - C and N, respectively) and radiogenic (strontium-Sr) isotopes analyses and the identification of the cultural traits in the material culture to identify mobility patterns that took place in the region. The Sr isotopic analyses on the human bones show that in the Copper Age and at the beginning of the Bronze Age, the cave was used as a burial place by different villages, perhaps spread in a radius of no more than 5 km. Stable isotopes analyses suggest the introduction of C4 plants in the diet of the Middle Bronze Age (MBA) communities in the area. Remarkably, in the same period, the material culture shows increasing influxes coming from the North. This evidence is consistent with the recent genomic findings tracing the arrival of people carrying a Steppe-related ancestry in Central Italy in MBA.


title: Microevolution of Pieris butterfly genes involved in host-plant adaptation along a host-plant community cline

abstract: Herbivorous insects have evolved counteradaptations to overcome the chemical defenses of their host plants. Several of these counteradaptations have been elucidated at the molecular level, in particular for insects specialized on cruciferous host plants. While the importance of these counteradaptations for host plant colonization is well established,little is known about their microevolutionary dynamics in the field. In this study, we examine patterns of host plant use and insect counteradaptation in three Pieris butterfly species across Japan. The larvae of these butterflies express nitrile-specifier protein (NSP) and its paralog major allergen (MA) in their gut to overcome the highly diversified glucosinolate-myrosinase defense system of their cruciferous host plants. Pieris napi and Pieris melete colonize wild Brassicaceae whereas Pieris rapae typically uses cultivated Brassica as a host, regardless of the local composition of wild crucifers. As expected, NSP and MA diversity was independent of the local composition of wild Brassicaceae in P. rapae. In contrast, NSP diversity correlated with local host plant diversity in both species that preferred wild Brassicaceae. P. melete and P. napi both revealed two distinct major NSP alleles, which shaped diversity among local populations, albeit with different evolutionary trajectories. In comparison, MA showed no indication for local adaptation. Altogether, MA appeared to be evolutionary more conserved than NSP, suggesting that both genes play different roles in diverting host plant chemical defense.


title: ZBP1 induces inflammatory signaling via RIPK3 and promotes SARS-CoV-2-induced cytokine expression

abstract: COVID-19 caused by the SARS-CoV-2 virus remains a threat to global health. The disease severity is mediated by cell death and inflammation, which regulate both the antiviral and the pathological innate immune responses. ZBP1, an interferon-induced cytosolic nucleic acid sensor, facilitates antiviral responses via RIPK3. Although ZBP1-mediated cell death is widely described, whether and how it promotes inflammatory signaling is unclear. Here, we report a ZBP1-induced inflammatory signaling pathway that depends on ubiquitination and RIPK3s scaffolding ability independently of cell death. In human cells, ZBP1 associates with RIPK1 and RIPK3 as well as ubiquitin ligases cIAP1 and LUBAC. RIPK1 and ZBP1 are ubiquitinated to promote TAK1- and IKK-mediated inflammatory signaling. Additionally, RIPK1 recruits the p43/41-caspase-8-p43-FLIP heterodimer to suppress RIPK3 kinase activity, which otherwise promotes inflammatory signaling in a kinase activity-dependent manner. Lastly, we show that ZBP1 contributes to SARS-CoV-2-induced cytokine production. Taken together, we describe a ZBP1-RIPK1-RIPK3-mediated inflammatory signaling pathway relayed by the scaffolding role of RIPKs and regulated by caspase-8. Our results suggest the ZBP1 pathway contributes to inflammation in response to SARS-CoV-2 infection.


title: Comparison of Plaque Size, Thermal Stability, and Replication Rate among SARS-CoV-2 Variants of Concern

abstract:
SARS-CoV-2, like other RNA viruses, has a propensity for genetic evolution owing to the low fidelity of its viral polymerase. This evolution results in the emergence of novel variants with different characteristics than their ancestral strain. Several recent reports have described a series of novel SARS-CoV-2 variants. Some of these have been identified as variants of concern (VOCs), including alpha (B.1.1.7, Clade GRY), beta (B.1.351, Clade GH), gamma (P.1, Clade GR), and delta (B.1.617.2, Clade G). VOCs are likely to have some effect on transmissibility, antibody evasion, and changes in therapeutic or vaccine effectiveness. However, the physiological and virological understanding of these variants remains poor. We demonstrated that these four VOCs exhibited differences in plaque size, thermal stability at physiological temperature, and replication rates. The mean plaque size of beta was the largest, followed by those of gamma, delta, and alpha. Thermal stability, evaluated by measuring infectivity and half-life after prolonged incubation at physiological temperature, was correlated with plaque size in all variants except alpha. However, despite its relatively high thermal stability, alphas small plaque size resulted in lower replication rates and fewer progeny viruses. Our findings may inform further virological studies of SARS-CoV-2 variant characteristics, VOCs, and variants of interest. These studies are important for the effective management of the COVID-19 pandemic.

IMPORTANCEThe global pandemic caused by SARS-CoV-2 continues to persist, due in part to mutations that have resulted in the emergence of different variants. Many of these variants have become more virulent and infectious than their ancestral strain, resulting in an ever-increasing spread. However, our virological understanding of these variants remains poor. Here, we directly compared the plaque size, stability, and replication kinetics of four SARS-CoV-2 variants of concern following prolonged incubation at physiological temperatures. Our observations may help to characterize each variant in terms of their interactions with host factors and responses to environmental conditions. We also believe that our evaluations will improve understanding of the emergence of new variants and contribute to controlling their spread.

category: microbiology


title: Bioorthogonal Functionalization of Material Surfaces with Bioactive Molecules

abstract: The ability to control and modify the bioactivity of surfaces is pivotal to the success of many medical devices. A biocompatible and bioorthogonal method to functionalize surfaces with a wide variety of bioactive molecules is an important tool for enabling innovative biotechnology and medical applications. We report herein a novel, catecholamine-based surface functionalization method that is chemoselective and free of a metal catalyst. This method utilizes the ligation between a coating formed from the tyrosinase-catalyzed polymerization of a tetrazine-containing catecholamine and one or more strained alkene-containing molecules of interest. The process is achieved under conditions ideal for biological applications. Using this method, we graft surfaces with a variety of active molecules, including a small molecule fluorophore, enzymes, and a cyclic peptide with the RGD motif, and demonstrate the maintenance of their activity on the surface. Additionally, we establish the cytocompatibility of grafted ECM-mimicking surfaces with fibroblasts and show improved cell adhesion and cytoskeletal organization.


title: Optimized Workflow for High Spatial Resolution MALDI Imaging Mass Spectrometry ofFresh-Frozen Bone

abstract: Bone and bone marrow are vital to mammalian structure, movement, and immunity. These tissues are also commonly subjected to molecular alterations giving rise to debilitating diseases like rheumatoid arthritis, osteoporosis, osteomyelitis, and cancer. Technologies such as matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) enable the discovery of spatially resolved chemical information in biological tissue samples to help elucidate the complex molecular processes underlying pathology. Traditionally, preparation of native osseous tissue for MALDI IMS has been difficult due to the mineralized composition and heterogenous morphology of the tissue, and compensation for these challenges with decalcification and fixation protocols can remove or delocalize molecular species. Here, sample preparation methods were advanced to enable multimodal MALDI IMS of undecalcified, fresh-frozen murine femurs allowing the distribution of endogenous lipids to be linked to specific tissue structures and cell types. Adhesive-bound bone sections were mounted onto conductive glass slides with a microscopy-compatible glue and freeze-dried to minimize artificial bone marrow damage. Subliming matrix does not induce further bone marrow cracking, and recrystallizing the deposited matrix improves lipid signal. High spatial resolution (10 m) MALDI IMS was employed to characterize lipid distributions in fresh-frozen bone, and use of complementary microscopy modalities aided tissue and cell assignments. For example, various phosphatidylcholines localize to bone marrow, adipose tissue, marrow adipose tissue, and muscle. Further, sphingomyelin(42:1) was abundant in megakaryocytes, whereas sphingomyelin(42:2) was diminished in this cell type. These data reflect the vast molecular and cellular heterogeneity indicative of the bone marrow and the soft tissue surrounding the femur. Multimodal MALDI IMS has the potential to advance bone-related biomedical research by offering deep molecular coverage with spatial relevance in a preserved native bone microenvironment.


title: High Spatial Resolution MALDI Imaging Mass Spectrometry ofFresh-Frozen Bone


title: Neutrophils contribute to ER stress in lung epithelial cells in the pristane-induced diffuse alveolar hemorrhage mouse model

abstract: Diffuse alveolar hemorrhage (DAH), although rare, is a life-threatening complication of systemic lupus erythematosus (SLE). Little is known about the pathophysiology of DAH in humans, although increasingly neutrophils, NETosis and inflammatory monocytes have been shown to play an important role in the pristane-induced model of SLE which develops lung hemorrhage and recapitulates many of the pathologic features of human DAH. Using this experimental model, we asked whether endoplasmic reticulum (ER) stress played a role in driving the pathology of pulmonary hemorrhage and what role infiltrating neutrophils had in this process. Analysis of lung tissue from pristane-treated mice showed genes associated with ER stress and NETosis were increased in a time-dependent manner and reflected the timing of CD11b+Ly6G+ neutrophil accumulation in the lung. Using precision cut lung slices from untreated mice we observed that neutrophils isolated from the peritoneal cavity of pristane-treated mice could directly induce the expression of genes associated with ER stress, namely Chop and Bip. Mice which had myeloid-specific deletion of PAD4 were generated and treated with pristane to assess the involvement of PAD4 and PAD4-dependent NET formation in pristane-induced lung inflammation. Specific deletion of PAD4 in myeloid cells resulted in decreased expression of ER stress genes in the pristane model, with accompanying reduction in IFN-driven genes and pathology. Lastly, coculture experiments of human neutrophils and human lung epithelial cell line (BEAS-2b) showed neutrophils from SLE patients induced significantly more ER stress and interferon-stimulated genes in epithelial cells compared to healthy control neutrophils. These results support a pathogenic role of neutrophils and NETs in lung injury during pristane-induced DAH through the induction of ER stress response and suggest that overactivation of neutrophils in SLE and NETosis may underlie development of DAH.


title: Efficient methods for generation and expansion of, and gene delivery to Rhesus Macaque plasma B cells

abstract: Engineered long lived plasma cells have the potential to be a new area of cell therapy. A key step in developing this cell therapy is testing in a model with an intact immune system similar to humans. To that end, we have developed methods to purify, expand, and differentiate non-human primate (NHP; rhesus macaque) B cells ex vivo. By comparing several media types and conditions, we consistently achieved 10-fold expansion of NHP B cells using a readily available commercial supplement. After only seven days in culture, large percentages of cells in NHP B cell cultures were differentiated. These cells expressed surface markers found in human antibody secreting cells (CD38 and CD138) and secreted immunoglobulin G. We also identified the serotypes (2.5 and D-J) and conditions necessary for efficient transduction of NHP B cells with AAV vectors for the purposes of producing a secreted protein (BAFF). We hope that this work will accelerate proof-of-concept in vivo studies using engineered protein-secreting B cells in an NHP model.


title: OXPHOS Promotes Apoptotic Resistance and Persistence in TH17 cells

abstract: Apoptotic cell death is a cell-intrinsic, immune tolerance mechanism that regulates the magnitude and resolution of T cell-mediated responses. Evasion of apoptosis is critical for the generation of memory T cells, as well as autoimmune T cells, and knowledge of the mechanisms that enable resistance to apoptosis will provide insight into ways to modulate their activity during protective and pathogenic responses. IL-17-producing CD4 T cells (TH17s) are long-lived, memory cells. These features enable their role in host defense, chronic inflammatory disorders, and anti-tumor immunity. A growing number of reports now indicate that TH17s in vivo require mitochondrial oxidative phosphorylation (OXPHOS), a metabolic phenotype that is poorly induced in vitro. To elucidate the role of OXPHOS in TH17 processes, we developed a system to polarize TH17s that metabolically resembled their in vivo counterparts. We discovered that directing TH17s to use OXPHOS promotes mitochondrial fitness, glutamine anaplerosis, and an anti-apoptotic phenotype marked by high BCL-XL and low BIM. Through competitive co-transfer experiments and tumor studies, we further revealed how OXPHOS protects TH17s from cell death while enhancing their persistence in the periphery and tumor microenvironment. Together, our work demonstrates a non-classical role of metabolism in regulating TH17 cell fate and highlights the potential for therapies that target OXPHOS in TH17-driven diseases.


title: A mycobacterial glycine-rich protein governs actin-based motility

abstract: Mycobacterium marinum, a close relative of the significant human pathogen Mycobacterium tuberculosis, polymerizes host actin at the bacterial surface to drive intracellular movement and cell-to-cell spread during infection. Here, we report the identification and characterization of MirA, the M. marinum actin-based motility factor. MirA is a member of the glycine-rich PE_PGRS family of ESX-5-secreted proteins. MirA uses an amphipathic helix to anchor into the mycobacterial outer membrane and, surprisingly, also the surface of host lipid droplet organelles. The glycine-rich PGRS domain in MirA directly binds and activates host N-WASP to stimulate actin polymerization through the Arp2/3 complex, directing both bacterial and lipid droplet actin- based motility. MirA is dissimilar to known N-WASP activating ligands and may represent a new class of microbial and host actin regulator. Additionally, the MirA-N-WASP interaction represents a model to understand how the enigmatic PE_PGRS proteins contribute to mycobacterial pathogenesis.


title: New insights into the architecture and dynamics of archaella

abstract:
Archaea swim by means of a unique molecular machine called the archaellum. The archaellum consists of an ATP-powered intracellular motor that drives the rotation of an extracellular filament, allowing the cell to rapidly propel itself through liquid media.

The archaellum filament comprises multiple copies of helically organised subunits named archaellins. While in many species several archaellin homologs are encoded in the same operon, structural studies conducted to date have suggested that archaella consist of only one protein species. Thus, the role of the remaining archaellin genes remains elusive.

Here we present the structure of the Methanocaldococcus villosus archaellum filament at 3.08 [A] resolution. We find that the filament is composed of two alternating archaellins - ArlB1 and ArlB2, suggesting that the architecture and assembly of archaella is more complex than previously thought. Moreover, we identify two major structural elements that enable the archaellum filament to move.

Our findings provide new insights into archaeal motility and challenge the current view on the archaellum architecture and assembly.


title: Mechanistic Insights in Biofilm Eradication Potential of Tyrosol Functionalized Chitosan Gold Nanoparticles against Candida spp.

abstract: In the present study, tyrosol functionalized chitosan gold nanoparticles (Chi-TY-AuNPs) were prepared as an alternative treatment strategy to combat fungal infections. Various biophysical techniques were used to characterize the synthesized Chi-TY-AuNPs. The antifungal and antibiofilm activities of Chi-TY-AuNPs were evaluated against C. albicans and C. glabrata and efforts have been made to elucidate the possible mechanism of action. Chi-TY-AuNPs showed a high fungicidal effect against both sessile and planktonic cells of Candida spp. Additionally, Chi-TY-AuNPs completely eradicated (100%) the mature biofilms of both the Candida spp. FESEM analysis highlighted the morphological alterations in Chi-TY-AuNPs treated Candida biofilm cells. Effect of Chi-TY-AuNPs on the ECM components showed significant reduction in protein content in C. glabrata biofilm and substantial decrease in extracellular DNA (eDNA) content of both the Candida spp. ROS generation analysis using DCFDA-PI staining showed high ROS levels in both the Candida spp., whereas pronounced ROS production was observed in Chi-TY-AuNPs treated C. glabrata biofilm. Biochemical analysis revealed decreased ergosterol content in Chi-TY-AuNPs treated C. glabrata cells, while inconsequential changes were observed in C. albicans. Furthermore, the transcriptional expression of selected genes (ergosterol biosynthesis, efflux, sterol importer, and glucan biogenesis) was reduced in C. glabrata in response to Chi-TY-AuNPs except ERG11 and CDR1. Conclusively the result showed the biofilm inhibition and biofilm eradication efficacy of Chi-TY-AuNPs in both the Candida spp. Findings of the present study manifest Chi-TY-AuNPs as a potential therapeutic solution to Candida biofilm-related chronic infections and overcome biofilm antifungal resistance.


title: Plausible Mechanistic Insights in Biofilm Eradication Potential of against Candida spp. using In Situ Synthesized Tyrosol Functionalized Chitosan Gold Nanoparticles as a versatile Antifouling Coating on Implant Surfaces


title: Digital control of c-di-GMP in E. coli balances population-wide developmental transitions and phage sensitivity

abstract: Nucleotide-based signaling molecules (NSMs) are widespread in bacteria and eukaryotes, where they control important physiological and behavioral processes. In bacteria, NSM-based regulatory networks are highly complex, entailing large numbers of enzymes involved in the synthesis and degradation of active signaling molecules. How the converging input from multiple enzymes is transformed into robust and unambiguous cellular responses has remained unclear. Here we show that Escherichia coli converts dynamic changes of c-di-GMP into discrete binary signaling states, thereby generating heterogeneous populations with either high or low c-di-GMP. This is mediated by an ultrasensitive switch protein, PdeL, which senses the prevailing cellular concentration of the signaling molecule and couples this information to c-di-GMP degradation and transcription feedback boosting its own expression. We demonstrate that PdeL acts as a digital filter that facilitates precise developmental transitions, confers cellular memory, and generates functional heterogeneity in bacterial populations to evade phage predation. Based on our findings, we propose that bacteria apply ultrasensitive regulatory switches to convert dynamic changes of NSMs into binary signaling modes to allow robust decision-making and bet-hedging for improved overall population fitness.


title: Stress response in Entamoeba histolytica is associated with robust processing of tRNA to tRNA-halves

abstract:
tRNA-derived fragments have been reported in many different organisms and have diverse cellular roles such as regulating gene expression, inhibiting protein translation, silencing transposable elements and modulating cell proliferation. In particular tRNA halves, a class of tRNA fragments produced by the cleavage of tRNAs in the anti-codon loop, have been widely reported to accumulate under stress and regulate translation in cells. Here we report the presence of tRNA-derived fragments in Entamoeba with tRNA halves being the most abundant. We further established that tRNA halves accumulate in the parasites upon different stress stimuli such as oxidative stress, heat shock, and serum starvation. We also observed differential expression of tRNA halves during developmental changes of trophozoite to cyst conversion with various tRNA halves accumulating during early encystation. In contrast to other systems, the stress response does not appear to be mediated by a few specific tRNA halves as multiple tRNAs appear to be processed during the various stresses. Furthermore, we identified some tRNA-derived fragments are associated with Entamoeba Argonaute proteins, EhAgo2-2, and EhAgo2-3, which have a preference for different tRNA-derived fragment species. Finally, we show that tRNA halves are packaged inside extracellular vesicles secreted by amoeba. The ubiquitous presence of tRNA-derived fragments, their association with the Argonaute proteins, and the accumulation of tRNA halves during multiple different stresses including encystation suggest a nuanced level of gene expression regulation mediated by different tRNA-derived fragments in Entamoeba.

ImportancetRNA-derived fragments are small RNAs formed by the cleavage of tRNAs at specific positions. These have been reported in many organisms to modulate gene expression and thus regulate various cell functions. In the present study, we report for the first time the presence of tRNA-derived fragments in Entamoeba. tRNA-derived fragments were identified by bioinformatics analyses of small RNA sequencing datasets from the parasites and also confirmed experimentally. We found that tRNA halves accumulated in parasites exposed to environmental stress or during developmental process of encystation. We also found that shorter tRNA-derived fragments are bound to Entamoeba Argonaute proteins, indicating that they may have a potential role in the Argonaute-mediated RNA-interference pathway which mediates robust gene silencing in Entamoeba. Our results suggest that tRNA-derived fragments in Entamoeba have a possible role in regulating gene expression during environmental stress.


title: Bacillus subtilis with a 1% population enhances the activity of Enterococcus faecalis with a 99% population

abstract:
Microbes are present as communities in the environment. However, the importance of minor populations has not been well studied experimentally. In this study, we evaluated the role of Bacillus subtilis with a 1% population and its effect on co-incubated Enterococcus faecalis with a 99% population. Here we used an azo dye-decolorizing Enterococcus faecalis strain T6a1 and non-dye-decolorizing Bacillus subtilis strain S4ga. The dye decolorization assay enabled the investigation of the effects of B. subtilis S4ga on the activity of E. faecalis T6a1, even when B. subtilis S4ga was present at only 1% relative abundance or lower. We found that non-decolorizing B. subtilis S4ga enhanced the dye decolorization activity of E. faecalis T6a1, shortened the lag time of E. faecalis T6a1 to start decreasing the dye concentration, and increased the time for E. faecalis T6a1 to continue dye decolorization. These effects were correlated with redox potential values. We compared the extracellular amino acids between each incubation culture of E. faecalis T6a1 and B. subtilis S4ga, which revealed their mutual relationship by cross-feeding of specific amino acids. We also compared the intracellular primary metabolites between co-incubation and sole incubation of E. faecalis T6a1. The arginine deiminase (ADI) pathway in the co-incubated E. faecalis T6a1 was activated compared to that of E. faecalis T6a1 incubated solely. These findings explained that co-incubation with B. subtilis S4ga promoted ATP production in E. faecalis T6a1 cells to a greater extent and enhanced dye-decolorization activity.

IMPORTANCEThis study highlights the importance of minor bacterial populations and their effects on major populations. We used Enterococcus faecalis as the major population and Bacillus subtilis as the minor population. Both species are becoming increasingly important. Some strains of E. faecalis are antibiotic-resistant pathogens, show probiotic effects, or are applicable for textile wastewater treatment. Some strains of B. subtilis are known to produce antimicrobial agents, reduce intestinal inflammation, or restore gut microbiota. We demonstrated that a low abundance of B. subtilis with 1% population increased the amount of energy produced by E. faecalis with 99% population, which appeared as enhanced dye decolorization activity of dye-decolorizing E. faecalis. Metabolomic analysis suggested that E. faecalis and B. subtilis had a mutual relationship by feeding specific amino acids to each other. These results provide new insights into co-existing minor populations in microbial communities and will improve our understanding of bacterial control.

Graphical abstract

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title: Fungal Mineral Weathering Mechanisms Revealed Through Direct Molecular Visualization

abstract: Soil fungi facilitate the translocation of inorganic nutrients from soil minerals to other microorganisms and plants. This ability is particularly advantageous in impoverished soils, because fungal mycelial networks can bridge otherwise spatially disconnected and inaccessible nutrient hotspots. However, the molecular mechanisms underlying fungal mineral weathering and transport through soil remains poorly understood. Here, we addressed this knowledge gap by directly visualizing nutrient acquisition and transport through fungal hyphae in a mineral doped soil micromodel using a multimodal imaging approach. We observed that Fusarium sp. DS 682, a representative of common saprotrophic soil fungi, exhibited a mechanosensory response (thigmotropism) around obstacles and through pore spaces ([~]12 {micro}m) in the presence of minerals. The fungus incorporated and translocated potassium (K) from K-rich mineral interfaces, as evidenced by visualization of mineral derived nutrient transport and unique K chemical moieties following fungal induced mineral weathering. Specific membrane transport proteins were expressed in the presence of minerals, including those involved in oxidative phosphorylation pathways and transmembrane transport of small molecular weight organic acids. This study establishes the significance of fungal biology and nutrient translocation mechanisms in maintaining fungal growth under water and nutrient limitations in a soil-like microenvironment.


title: Fundamentals of Vaping-Associated Pulmonary Injury Leading to Severe Respiratory Distress

abstract: Vaping of flavored liquids has been touted as safe alternative to traditional cigarette smoking with decreased health risks. The popularity of vaping has dramatically increased over the last decade, particularly among teenagers who incorporate vaping into their daily life as a social activity. Despite widespread and increasing adoption of vaping among young adults there is little information on long term consequences of vaping and potential health risks. This study demonstrates Vaping-Induced Pulmonary Injury (VAPI) using commercial JUUL pens with flavored vape juice using an inhalation exposure murine model. Profound pathological changes to upper airway, lung tissue architecture, and cellular structure are evident within 9 weeks of exposure. Marked histologic changes include increased parenchyma tissue density, cellular infiltrates proximal to airway passages, alveolar rarefaction, increased collagen deposition, and bronchial thickening with elastin fiber disruption. Transcriptional reprogramming includes significant changes to gene families coding for xenobiotic response, glycerolipid metabolic processes, and oxidative stress. Cardiac contractile performance for systemic output is moderately but significantly impaired, and the shows severe pulmonary side structural remodeling with chamber enlargement. This VAPI model with pulmonary circuit failure demonstrates mechanistic underpinnings of vaping-related pathologic injury.

category: molecular biology


title: Structural basis of ATP-dependent high-fidelity epigenome maintenance

abstract:
Epigenetic evolution occurs over million-year timescales in Cryptococcus neoformans and is mediated by DNMT5, the first maintenance-type cytosine methyltransferase identified in the fungal or protist kingdoms. DNMT5 requires ATP and displays exquisite hemimethyl-DNA specificity. To understand these novel properties, we solved cryo-EM structures of CnDNMT5 in three states. These studies reveal an elaborate allosteric cascade in which hemimethylated DNA first activates the SNF2 ATPase domain by a large rigid body rotation while the target cytosine partially flips out the DNA duplex. ATP binding then triggers a striking structural reconfiguration of the methyltransferase catalytic pocket that enables cofactor binding, completion of base-flipping, and catalysis. Unmethylated DNA binding fails to open cofactor pocket and subsequent ATP binding triggers its ejection to ensure fidelity. This chaperone-like, enzyme-remodeling role of the SNF2 domain illuminates how energy can be used to enable faithful epigenetic memory.

HighlightsO_LIStructures of DNMT5 reveal mechanism of ATP-dependent DNA methylation
C_LIO_LIHemimethylated CpG recognition triggers partial base flipping of the target cytosine
C_LIO_LIHemimethylated DNA induces rigid body rotation to activate the SNF2 ATPase domain
C_LIO_LIMTase catalytic pocket is remodeled by the SNF2 ATPase to achieve specificity
C_LI


title: CDK5 influences the organization of the Circadian Machinery in peripheral clocks.

abstract:
Circadian rhythms are self-sustained physiological changes that drive rhythmicity within the 24-hours cycles. Posttranslational modifications (PMTs), such as protein phosphorylation, acetylation, sumoylation, and ubiquitination, are biochemical processes that modify protein structure and functions, ensuring circadian rhythm precision. For example, phosphorylation is considered the most important hallmark of rhythmicity from cyanobacteria to mammals. Cyclin-dependent kinase 5 (CDK5) has been shown to regulate the mammalian SCNs circadian clock via phosphorylation of PER2. Here, we show that CDK5 influences the clock machinery assembling, using immortalized mouse embryonic fibroblast as an in vitro model for studying the peripheral clock. In fact, the circadian period at the cellular level is lengthened. Furthermore, the clock-controlled genes expression amplitude is dampened in Cdk5 ko cell lines, while the phase is delayed about 4 hours.

Taken together, we show in vitro that CDK5 is critically involved in regulating the peripheral clocks, influencing their temporal and spatial dynamics.


title: Deep mutational scan of a drug efflux pump reveals its structure-function landscape

abstract: Drug efflux is a common resistance mechanism found in bacteria and cancer cells. Although several structures of drug efflux pumps are available, they provide only limited functional information on the phenomenon of drug efflux. Here, we performed deep mutational scanning (DMS) on the bacterial ATP binding cassette (ABC) transporter EfrCD to determine the drug efflux activity profile of more than 1500 single variants. These systematic measurements revealed that the introduction of negative charges at different locations within the large substrate binding pocket results in strongly increased efflux activity towards positively charged ethidium, while additional aromatic residues did not display the same effect. Data analysis in the context of an inward-facing cryo-EM structure of EfrCD uncovered a high affinity binding site, which releases bound drugs through a peristaltic transport mechanism as the transporter transits to its outward-facing conformation. Finally, we identified substitutions resulting in rapid Hoechst influx without affecting the efflux activity for ethidium and daunorubicin. Hence, single mutations can convert the ABC exporter EfrCD into a drug-specific ABC importer.


title: Structural basis for C-type inactivation in a Shaker family voltage gated K+ channel

abstract: C-type inactivation is a process by which ion flux through a voltage-gated K+ (Kv) channel is regulated at the selectivity filter. While prior studies have indicated that C-type inactivation involves structural changes at the selectivity filter, the nature of the changes have not been resolved. Here we report the crystal structure of the Kv1.2 channel in a C-type inactivated state. The structure shows that C-type inactivation involves changes in the selectivity filter that disrupt the outer two ion binding sites in the filter. The changes at the selectivity filter propagate to the extracellular mouth and the turret regions of the channel pore. The structural changes observed are consistent with the functional hallmarks of C-type inactivation. This study highlights the intricate interplay between K+ occupancy at the ion binding sites and the interactions of the selectivity filter in determining the balance between the conductive and the inactivated conformations of the filter.


title: Role of Ring6 in the function of the E. coli MCE protein LetB

abstract:
LetB is a tunnel-forming protein found in the cell envelope of some double-membraned bacteria, and is thought to be important for the transport of lipids between the inner and outer membranes. In Escherichia coli the LetB tunnel is formed from a stack of seven rings (Ring1 - Ring7), in which each ring is composed of a homo-hexameric assembly of MCE domains. The primary sequence of each MCE domain of the LetB protein is substantially divergent from the others, making each MCE ring unique in nature. The role of each MCE domain and how it contributes to the function of LetB is not well understood. Here we probed the importance of each MCE ring for the function of LetB, using a combination of bacterial growth assays and cryo-EM. Surprisingly, we find that {Delta}Ring3 and {Delta}Ring6 mutants, in which Ring3 and Ring6 have been deleted, confer increased resistance to membrane perturbing agents. Specific mutations in the pore-lining loops of Ring6 similarly confer increased resistance. A cryo-EM structure of the {Delta}Ring6 mutant shows that despite the absence of Ring6, which leads to a shorter assembly, the overall architecture is maintained, highlighting the modular nature of MCE proteins. Previous work has shown that Ring6 is dynamic and in its closed state, may restrict the passage of substrate through the tunnel. Our work suggests that removal of Ring6 may relieve this restriction. The deletion of Ring6 combined with mutations in the pore-lining loops leads to a model for the tunnel gating mechanism of LetB. Together, these results provide insight into the functional roles of individual MCE domains and pore-lining loops in the LetB protein.

HighlightsO_LIDeleting MCE domains 3 or 6 from LetB confers increased resistance to membrane-perturbing agents
C_LIO_LICryo-EM structure of {Delta}Ring6 LetB mutant highlights the modular nature of MCE domains
C_LIO_LIMutations along LetB pore-lining loops modulate detergent resistance
C_LI


title: The structure balance of gene-gene networks beyond pairwise interactions

abstract: Despite its high and direct impact on nearly all biological processes, the underlying structure of gene-gene interaction networks is investigated so far according to pair connections. To address this, we explore the gene interaction networks of the yeast Saccharomyces cerevisiae beyond pairwise interaction using the structural balance theory (SBT). Specifically, we ask whether essential and nonessential gene interaction networks are structurally balanced. We study triadic interactions in the weighted signed undirected gene networks and observe that balanced and unbalanced triads are over and underrepresented in both networks, thus beautifully in line with the strong notion of balance. Moreover, we note that the energy distribution of triads is significantly different in both essential and nonessential networks compared with the shuffled networks. Yet, this difference is greater in the essential network regarding the frequency as well as the energy of triads. Additionally, results demonstrate that triads in the essential gene network are more interconnected through sharing common links, while in the nonessential network they tend to be isolated. Last but not least, we investigate the contribution of all-length signed walks and its impact on the degree of balance. Our findings reveal that interestingly when considering longer cycles the nonessential gene network is more balanced compared to the essential network.


title: Improving the global dimensions of intrinsically disordered proteins in Martini 3

abstract: Coarse-grained molecular dynamics simulations are a useful tool to determine conformational ensembles of proteins. Here, we show that the coarse-grained force field Martini 3 underestimates the global dimensions of intrinsically disordered proteins (IDPs) and multidomain proteins when compared with small angle X-ray scattering (SAXS) data, and that increasing the strength of protein-water interactions favours more expanded conformations. We find that increasing the strength of interactions between protein and water by ca. 10% results in improved agreement with the SAXS data for IDPs and multi-domain proteins. We also show that this correction results in a more accurate description of self-association of IDPs and folded proteins and better agreement with paramagnetic relaxation enhancement data for most IDPs. While simulations with this revised force field still show deviations to experiments for some systems our results suggest that it is overall a substantial improvement for coarse-grained simulations of soluble proteins.


title: Improving Martini 3 for disordered and multidomain proteins


title: Natural and designed proteins inspired by extremotolerant organisms can form condensates and attenuate apoptosis in human cells

abstract: Many organisms can survive extreme conditions and successfully recover to normal life. This extremotolerant behavior has been attributed in part to repetitive, amphipathic, and intrinsically disordered proteins that are upregulated in the protected state. Here, we assemble a library of approximately 300 naturally-occurring and designed extremotolerance-associated proteins to assess their ability to protect human cells from chemically-induced apoptosis. We show that several proteins from tardigrades, nematodes, and the Chinese giant salamander are apoptosis protective. Notably, we identify a region of the human ApoE protein with similarity to extremotolerance-associated proteins that also protects against apoptosis. This region mirrors the phase separation behavior seen with such proteins, like the tardigrade protein CAHS2. Moreover, we identify a synthetic protein, DHR81, that shares this combination of elevated phase separation propensity and apoptosis protection. Finally, we demonstrate that driving protective proteins into the condensate state increases apoptosis protection, and highlight the ability for DHR81 condensates to sequester caspase-7. Taken together, this work draws a link between extremotolerance-associated proteins, condensate formation, and designing human cellular protection.

category: synthetic biology


title: A novel locally c-di-GMP-controlled exopolysaccharide synthase required for N4 phage infection of E. coli

abstract:
A major target of c-di-GMP signaling is the production of biofilm-associated extracellular polymeric substances (EPS), which in Escherichia coli K-12 include amyloid curli fibres, phosphoethanolamine-modified (pEtN-)cellulose and poly-N-acetyl-glucosamine (PGA). However, the characterized c-di-GMP-binding effector systems are largely outnumbered by the 12 diguanylate cyclases (DGCs) and 13 phosphodiesterases (PDEs), which synthetize and degrade c-di-GMP, respectively. E. coli possesses a single protein with a potentially c-di-GMP-binding MshEN domain, NfrB, which - together with the outer membrane protein NfrA - is known to serve as a receptor system for phage N4. Here, we show that NfrB not only binds c-di-GMP with high affinity, but as a novel c-di-GMP-controlled glycosyltransferase synthesizes a secreted EPS, which can impede motility and is required as an initial receptor for phage N4 infection. In addition, a systematic screening of the 12 DGCs of E. coli K-12 revealed that specifically DgcJ is required for the infection with phage N4 and interacts directly with NfrB. This is in line with local signaling models, where specific DGCs and/or PDEs form protein complexes with particular c-di-GMP effector/target systems. Our findings thus provide further evidence that intracellular signaling pathways, which all use the same diffusible second messenger, can act in parallel in a highly specific manner.

ImportanceKey findings in model organisms led to the concept of  local signaling, challenging the dogma of a gradually increasing global intracellular c-di-GMP concentration driving the motile-sessile transition in bacteria. In our current model, bacteria dynamically combine global as well as local signaling modes, in which specific DGCs and/or PDEs team up with effector/target systems in multiprotein complexes. Our present study highlights a novel example of how specificity in c-di-GMP signaling can be achieved by showing NfrB as a novel c-di-GMP binding effector in E. coli, which is controlled in a local manner specifically by DgcJ. We further show that NfrB (which was initially found as a part of a receptor system for phage N4) is involved in the production of a novel exopolysaccharide. Finally, our data shine new light on host interaction of phage N4, which uses this exopolysaccharide as an initial receptor for adsorption.


title: The Human STAT2 Coiled-Coil Domain Contains a Degron for Zika Virus Interferon Evasion

abstract:
The ability of viruses to evade the host antiviral immune system determines their level of replication fitness, species specificity, and pathogenic potential. Flaviviruses rely on the subversion of innate immune barriers including the type I and type III IFN antiviral systems. Zika virus infection induces the degradation of STAT2, an essential component of the IFN stimulated gene transcription factor, ISGF3. The mechanisms that lead to STAT2 degradation by Zika virus are poorly understood, but it is known to be mediated by the viral NS5 protein that binds to STAT2 and targets it for proteasome-mediated destruction. To better understand how NS5 engages and degrades STAT2, functional analysis of the protein interactions that lead to Zika virus and NS5-dependent STAT2 proteolysis were investigated. Data implicate the STAT2 coiled-coil domain as necessary and sufficient for NS5 interaction and proteasome degradation after Zika virus infection. Molecular dissection reveals that the first two -helices of the STAT2 coiled-coil contain a specific targeting region for IFN antagonism. These functional interactions provide a more complete understanding of the essential protein-protein interactions needed for Zika virus evasion of the host antiviral response, and identifies new targets for antiviral therapeutic approaches.

ImportanceZika virus infection can cause mild fever, rash, and muscle pain, and in rare cases lead to brain or nervous system diseases including Guillain-Barre syndrome. Infections in pregnant women can increase the risk of miscarriage or serious birth defects including brain anomalies and microcephaly. There are no drugs or vaccines for Zika disease. Zika virus is known to break down the host antiviral immune response, and this research project reveals how the virus suppresses interferon signaling, and may reveal therapeutic vulnerabilities.


title: Dcp2 C-terminal Cis-Binding Elements Control Selective Targeting of the Decapping Enzyme by Forming Distinct Decapping Complexes

abstract:
A single Dcp1-Dcp2 decapping enzyme targets diverse classes of yeast mRNAs for decapping-dependent 5 to 3 decay, but the molecular mechanisms controlling selective mRNA targeting by the enzyme remain elusive. Through extensive genetic analyses we uncover cis-regulatory elements in the Dcp2 C-terminal domain that control selective targeting of the decapping enzyme by forming distinct decapping complexes. Two Upf1-binding motifs target the decapping enzyme to NMD substrates, and a single Edc3-binding motif targets both Edc3 and Dhh1 substrates. Pat1-binding leucine-rich motifs target Edc3 and Dhh1 substrates under selective conditions. Although it functions as a unique targeting component of specific complexes, Edc3 is a common component of multiple complexes. Xrn1 also has a specific Dcp2 binding site, allowing it to be directly recruited to decapping complexes. Collectively, our results demonstrate that Upf1, Edc3, and Pat1 function as regulatory subunits of the holo-decapping enzyme, controlling both its targeting specificity and enzymatic activation.

HighlightsLoss of Dcp2 cis-binding elements causes selective stabilization of distinct decapping substrates

Dcp2 cis-binding elements promote the assembly of target-specific decapping complexes in vivo

Xrn1 binds to Dcp2, and both Edc3 and Xrn1 are common components of multiple decapping complexes

Upf1, Edc3, and Pat1 function as unique targeting subunits of the yeast holo-decapping enzyme


title: ERK signaling dissolves ERF Repression Condensates in Living Embryos

abstract: Phase separation underlies the organization of the nucleus, including the biogenesis of nucleoli and the packaging of heterochromatin. Here we explore the regulation of transcription factor condensates involved in gene repression by ERK signaling in gastrulating embryos of a simple proto-vertebrate (Ciona). ERK signaling induces nuclear export of the transcriptional repressor ERF, which has been linked to various human developmental disorders. Using high resolution imaging we show that ERF is localized within discrete nuclear condensates that dissolve upon ERK activation. Interestingly, we observe dynamic pulses of assembly and dissociation during the cell cycle, providing the first visualization of a nuclear phase separation process that is regulated by cell signaling. We discuss the implications of these observations for producing sharp on/off switches in gene activity and suppressing noise in cell-cell signaling events.


title: Direct Reprogramming of Non-limb Fibroblasts toCells with Properties of Limb Progenitors

abstract: The early limb bud consists of mesenchymal progenitors (limb progenitors) derived from the lateral plate mesoderm (LPM) that produce most of the tissues of the mature limb bud. The LPM also gives rise to the mesodermal components of the trunk, flank and neck. However, the mesenchymal cells generated at these other axial levels cannot produce the variety of cell types found in the limb bud, nor can they be directed to form a patterned appendage-like structure, even when placed in the context of the signals responsible for organizing the limb bud. Here, by taking advantage of a direct reprogramming approach, we find a set of factors (Prdm16, Zbtb16, and Lin28) normally expressed in the early limb bud, that are capable of imparting limb progenitor-like properties to non-limb fibroblasts. Cells reprogrammed by these factors show similar gene expression profiles, and can differentiate into similar cell types, as endogenous limb progenitors. The further addition of Lin41 potentiates proliferation of the reprogrammed cells while suppressing differentiation. These results suggest that these same four key factors may play pivotal roles in the specification of endogenous limb progenitors.


title: Effects of productive management of an agroforestry system on the associated diversity of parasitoids (Hymenoptera: Braconidae)

abstract: Tropical agroecosystems have emerged from the continuous modification of natural environments, as a sustainability alternative for food production and biodiversity conservation. This work explores how the diversity of parasitoids is modified in environments where plant diversity is limited e.g. crops and when these are adjacent to secondary vegetation, i.e. a scenario fragmented continuously in a limited space. It was found that there is no direct effect of plant diversity in the group of parasitoids studied; but the number of specialist species is high, which indicates that in diversified agroecosystems these probably function as remnants of natural habitat or as a refuge for parasitoids that disperse to different types of management within the agroecosystem. Therefore, it is necessary to consider in future studies the controls exerted by the plant diversity effects bottom-up and consumer top-down. Adding to this the context of the interactions that occur in agroecosystems.


title: ModelMatcher: A scientist-centric online platform to facilitate collaborations between stakeholders of rare and undiagnosed disease research

abstract: Next-generation sequencing is a prevalent diagnostic tool for undiagnosed diseases, and has played a significant role in rare disease gene discovery. While this technology resolves some cases, others are given a list of possibly damaging genetic variants necessitating functional studies. Productive collaborations between scientists, clinicians, and patients can help resolve such medical mysteries, and provide insights into in vivo function of human genes. Furthermore, facilitating interactions between scientists and research funders, including non-profit organizations or commercial entities, can dramatically reduce the time to translate discoveries from bench to bedside. Several systems designed to connect clinicians and researchers with a shared gene of interest have been successful. However, these platforms exclude some stakeholders based on their role or geography. Here we describe ModelMatcher, a global online matchmaking tool designed to facilitate cross-disciplinary collaborations, especially between scientists and other stakeholders of rare and undiagnosed disease research. ModelMatcher is integrated into the Rare Diseases Models and Mechanisms Network and Matchmaker Exchange, allowing users to identify potential collaborators in other registries. This living database decreases the time from when a scientist or clinician is making discoveries regarding their genes of interest, to when they identify collaborators and sponsors to facilitate translational and therapeutic research.

category: genetics


title: IMPROVED SSRs-BASED GENETIC DIVERSITY ASSESSMENT OF COCONUTS (COCOS NUCIFERA L) ALONG THE COAST OF KENYA

abstract:
Coconut is the most important cash crop along the Coast of Kenya, yet its genetic diversity has not been fully established. A genetic diversity study of 48 coconut genotypes that were collected along the Coast of Kenya was conducted with 13 polymorphic short sequence repeats (SSRs) markers. SSR analysis was performed using GeneMapper while data analysis was done with PowerMarker and DARwin softwares.

The results revealed a total of 68 alleles ranging from 2 to 11 per locus with a mean of 5.23 per marker. Gene diversity and polymorphic information content (PIC) ranged between 0.41 to 0.83 and 0.33 to 0.79, respectively. A neighbour-joining dendrogram grouped the genotypes into three major clusters containing distinct sub-clusters. This study underscored that capillary electrophoresis is a more accurate and informative technique for SSRs allele scoring than agarose gels, which was reported in a previous study with the same SSRs markers and coconut genotypes in Kenya. The clusters observed forms the basis to isolate conservation blocks, which are the key to establishing a genebank, since there is no documented coconut genebank for ex-situ conservation in Kenya.


title: Exploration of deep-learning based classification with human SNP image graphs

abstract:
BackgroundWith the advancement of NGS platform, large numbers of human variations and SNPs are discovered in human genomes. It is essential to utilize these massive nucleotide variations for the discovery of disease genes and human phenotypic traits. There are new challenges in utilizing such large numbers of nucleotide variants for polygenic disease studies. In recent years, deep-learning based machine learning approaches have achieved great successes in many areas, especially image classifications. In this preliminary study, we are exploring the deep convolutional neural network algorithm in genome-wide SNP images for the classification of human populations.

ResultsWe have processed the SNP information from more than 2,500 samples of 1000 genome project. Five major human races were used for classification categories. We first generated SNP image graphs of chromosome 22, which contained about one million SNPs. By using the residual network (ResNet 50) pipeline in CNN algorithm, we have successfully obtained classification models to classify the validation dataset. F1 scores of the trained CNN models are 95 to 99%, and validation with additional separate 150 samples indicates a 95.8% accuracy of the CNN model. Misclassification was often observed between the American and European categories, which could attribute to the ancestral origins. We further attempted to use SNP image graphs in reduced color representations or images generated by spiral shapes, which also provided good prediction accuracy. We then tried to use the SNP image graphs from chromosome 20, almost all CNN models failed to classify the human race category successfully, except the African samples.

ConclusionsWe have developed a human race prediction model with deep convolutional neural network. It is feasible to use the SNP image graph for the classification of individual genomes.

category: genomics


title: Evolutionary insights into primate skeletal gene regulation using a comparative cell culture model

abstract:
The evolution of complex skeletal traits in primates was likely influenced by both genetic and environmental factors. Because skeletal tissues are notoriously challenging to study using functional genomic approaches, they remain poorly characterized even in humans, let alone across multiple species. The challenges involved in obtaining functional genomic data from the skeleton, combined with the difficulty of obtaining such tissues from nonhuman apes, motivated us to consider an alternative in vitro system with which to comparatively study gene regulation in skeletal cell types. Specifically, we differentiated six human and six chimpanzee induced pluripotent stem cell lines (iPSCs) into mesenchymal stem cells (MSCs) and subsequently into osteogenic cells (bone cells). We validated differentiation using standard methods and collected single-cell RNA sequencing data from over 100,000 cells across multiple samples and replicates at each stage of differentiation. While most genes that we examined display conserved patterns of expression across species, hundreds of genes are differentially expressed (DE) between humans and chimpanzees within and across stages of osteogenic differentiation. Some of these interspecific DE genes show functional enrichments relevant in skeletal tissue trait development. Moreover, topic modeling indicates that interspecific gene programs become more pronounced as cells mature. Overall, we propose that this in vitro model can be used to identify interspecific regulatory differences that may have contributed to skeletal trait differences between species.

Author SummaryPrimates display a range of skeletal morphologies and susceptibilities to skeletal diseases, but the molecular basis of these phenotypic differences is unclear. Studies of gene expression variation in primate skeletal tissues are extremely restricted due to the ethical and practical challenges associated with collecting samples. Nevertheless, the ability to study gene regulation in primate skeletal tissues is crucial for understanding how the primate skeleton has evolved. We therefore developed a comparative primate skeletal cell culture model that allows us to access a spectrum of human and chimpanzee cell types as they differentiate from stem cells into bone cells. While most gene expression patterns are conserved across species, we also identified hundreds of differentially expressed genes between humans and chimpanzees within and across stages of differentiation. We also classified cells by osteogenic stage and identified additional interspecific differentially expressed genes which may contribute to skeletal trait differences. We anticipate that this model will be extremely useful for exploring questions related to gene regulation variation in primate bone biology and development.


title: RecPD: A Recombination-Aware Measure of Phylogenetic Diversity

abstract:
A critical step in studying biological features (e.g., genetic variants, gene families, metabolic capabilities, or taxa) underlying traits or outcomes of interest is assessing their diversity and distribution. Accurate assessments of these patterns are essential for linking features to traits or outcomes and understanding their functional impact. Consequently, it is of crucial importance that the metrics employed for quantifying feature diversity can perform robustly under any evolutionary scenario. However, the standard metrics used for quantifying and comparing the distribution of features, such as prevalence, phylogenetic diversity, and related approaches, either do not take into consideration evolutionary history, or assume strictly vertical patterns of inheritance. Consequently, these approaches cannot accurately assess diversity for features that have undergone recombination or horizontal transfer. To address this issue, we have devised RecPD, a novel recombination-aware phylogenetic-diversity metric for measuring the distribution and diversity of features under all evolutionary scenarios. RecPD utilizes ancestral-state reconstruction to map the presence / absence of features onto ancestral nodes in a species tree, and then identifies potential recombination events in the evolutionary history of the feature. We also derive a number of related metrics from RecPD that can be used to assess and quantify evolutionary dynamics and correlation of feature evolutionary histories. We used simulation studies to show that RecPD reliably identifies evolutionary histories under diverse recombination and loss scenarios. We then apply RecPD in a real-world scenario in a preliminary study type III effector protein families secreted by the plant pathogenic bacterium Pseudomonas syringae and demonstrate that prevalence is an inadequate metric that obscures the potential impact of recombination. We believe RecPD will have broad utility for revealing and quantifying complex evolutionary processes for features at any biological level.

AUTHOR SUMMARYPhylogenetic diversity is an important concept utilized in evolutionary ecology which has extensive applications in population genetics to help us understand how evolutionary processes have distributed genetic variation among individuals of a species, and how this impacts phenotypic diversification over time. However, existing approaches for studying phylogenetic diversity largely assume that the genetic features follow vertical inheritance, which is frequently violated in the case of microbial genomes due to horizontal transfer. To address this shortcoming, we present RecPD, a recombination-aware phylogenetic diversity metric, which incorporates ancestral state reconstruction to quantify the phylogenetic diversity of genetic features mapped onto a species phylogeny. Through simulation experiments we show that RecPD robustly reconstructs the evolutionary histories of features evolving under various scenarios of recombination and loss. When applied to a real-world example of type III secreted effector protein families from the plant pathogenic bacterium Pseudomonas syringae, RecPD reveals that horizontal transfer has played an important role in shaping the phylogenetic distributions of aa substantial proportion of families across the P. syringae species complex. Furthermore, we demonstrate that the traditional measures of feature prevalence are unsuitable as a metric for comparing feature diversity.