title: Functional regulatory variants implicate distinct transcriptional networks in dementia

abstract:
Predicting functionality of noncoding variation is one of the major challenges in modern genetics. We employed massively parallel reporter assays to screen 5,706 variants from genome-wide association studies for both Alzheimers disease (AD) and Progressive Supranuclear Palsy (PSP). We identified 320 functional regulatory polymorphisms (SigVars) comprising 27 of 34 unique tested loci, including multiple independent signals across the complex 17q21.31 region. We identify novel risk genes including PLEKHM1 in PSP and APOC1 in AD, and perform gene-editing to validate four distinct causal loci, confirming complement 4 (C4A) as a novel genetic risk factor for AD. Moreover, functional variants preferentially disrupt transcription factor binding sites that converge on enhancers with differential cell-type specific activity in PSP and AD, implicating a neuronal SP1-driven regulatory network in PSP pathogenesis. These analyses support a novel mechanism underlying noncoding genetic risk, whereby common genetic variants drive disease risk via their aggregate activity on specific transcriptional programs.

One Sentence SummaryHigh-throughput functional analysis of GWAS loci reveals cell-type specific regulatory networks that mediate genetic risk for dementia.


title: Surprise-induced enhancements in the associability of Pavlovian cues facilitate learning across behavior systems

abstract: Surprising violations of outcome expectancies have long been known to enhance the associability of Pavlovian cues; that is, the rate at which the cue enters into further associations. The adaptive value of such enhancements resides in promoting new learning in the face of uncertainty. However, it is unclear whether associability enhancements reflect increased associative plasticity within a particular behavior system, or whether they can facilitate learning between a cue and any arbitrary outcome, as suggested by attentional models of conditioning. Here, we show evidence consistent with the latter hypothesis. Violating the outcome expectancies generated by a cue in an appetitive setting (feeding behavior system) facilitated subsequent learning about the cue in an aversive setting (defense behavior system). In addition to shedding light on the nature of associability enhancements, our findings offer the neuroscientist a behavioral tool to dissociate their neural substrates from those of other, behavior system- or valence-specific changes. Moreover, our results present an opportunity to utilize associability enhancements to the advantage of counterconditioning procedures in therapeutic contexts.

category: animal behavior and cognition


title: Comprehensive analysis of lectin-glycan interactions reveals determinants of lectin specificity

abstract:
Lectin-glycan interactions facilitate inter- and intracellular communication in many processes including protein trafficking, host-pathogen recognition, and tumorigenesis promotion. Specific recognition of glycans by lectins is also the basis for a wide range of applications in areas including glycobiology research, cancer screening, and antiviral therapeutics. To provide a better understanding of the determinants of lectin-glycan interaction specificity and support such applications, this study comprehensively investigates specificity-conferring features of all available lectin-glycan complex structures. Systematic characterization, comparison, and predictive modeling of a set of 221 complementary physicochemical and geometric features representing these interactions highlighted specificity-conferring features with potential mechanistic insight. Univariable comparative analyses with weighted Wilcoxon-Mann-Whitney tests revealed strong statistical associations between binding site features and specificity that are conserved across unrelated lectin binding sites. Multivariable modeling with random forests demonstrated the utility of these features for predicting the identity of bound glycans based on generalized patterns learned from non-homologous lectins. These analyses revealed global determinants of lectin specificity, such as sialic acid glycan recognition in deep, concave binding sites enriched for positively charged residues, in contrast to high mannose glycan recognition in fairly shallow but well-defined pockets enriched for non-polar residues. Focused analysis of hemagglutinin interactions with human-like and avian-like glycans uncovered features representing both known and novel mutations related to shifts in influenza tropism from avian to human tissues. The presented systematic characterization of lectin binding sites provides a novel approach to studying lectin specificity and is a step towards confidently predicting new lectin-glycan interactions.

AUTHOR SUMMARYGlycans are sugar molecules found attached to many proteins and coating the outsides of cells from most organisms. Specific recognition of glycans by proteins called lectins facilitates many biological processes, for example enabling influenza to gain access to cells, helping the immune system recognize pathogens, and sorting newly built proteins for transport to appropriate cellular regions. Understanding what makes a particular lectin consider a particular glycan "sweeter" than the vast set of other glycans can help us better understand these processes and how to monitor and control them. To that end, we systematically characterized the sites on lectin structures where glycans are bound, breaking down molecular structures into a comprehensive set of biochemical and geometric features summarizing the sites. This enabled us to discover statistical relationships between binding site features and the glycans recognized by the sites, and further to be able to predict, from a lectin structure, which glycans it recognizes. For the first time, we are able to demonstrate that there are general features of lectin binding sites correlated with and predictive of their specificities, even in unrelated lectins. Ultimately, these findings can help us discover and engineer new lectins for use in research, diagnostics, or even therapeutics.

category: bioinformatics


title: Protein turnover in the developing Triticum aestivum grain

abstract: Protein abundance in cereal grains is determined by the relative rates of protein synthesis and protein degradation during grain development. Through combining in vivo stable isotope labelling and in-depth quantitative proteomics, we have measured the turnover of 1400 different types of proteins during wheat grain development. We demonstrate that there is a spatiotemporal pattern to protein turnover rates which explain part of the variation in protein abundances that is not attributable to differences in wheat gene expression. We show that approximately 20% of total grain ATP production is used for grain proteome biogenesis and maintenance, and nearly half of this budget is invested exclusively in storage protein synthesis. We calculate that 25% of newly synthesized storage proteins are turned over during grain development rather than stored. This approach to measure protein turnover rates at proteome scale reveals how different functional categories of grain proteins accumulate, calculates the costs of protein turnover during wheat grain development and identifies the most and the least stable proteins in the developing wheat grain.

category: plant biology


title: Computational fluid dynamics provide evidence for a compensatory suction feeding like effect in the predatory strike of dragonfly larvae

abstract: Most fast-moving aquatic predators face the challenge of bow wave formation. Water in front of the predator alarms or even displaces the prey. To mitigate the formation of such a bow wave, a strategy aiming at pressure reduction via suction has evolved convergently in several animal groups: compensatory suction feeding. The aquatic larvae of dragonflies and damselflies (Insecta: Odonata) are likely to face this challenge as well. They capture prey underwater using a fast-moving raptorial appendage, the so-called prehensile labial mask. Within dragonflies (Odonata: Anisoptera) two basic shapes of the prehensile labial mask have evolved, with an either flat and slender or concave distal segment. While the former is a pure grasping device, the latter is also capable of scooping up smaller prey and retaining it inside the cavity by arrays of bristle-like structures. The hydrodynamics of the prehensile labial mask was previously unknown. We used computational fluid dynamic (CFD) simulations of the distal segment of the mask, to investigate for the first time how different shapes of the mask impact their function. Our results suggest that both shapes are highly streamlined and generate a low-pressure area, likely leading to an effect analogous to compensatory suction feeding. This presents a vivid example of how convergent evolution enables very different animal groups to successfully deal with the challenges of their environment. It may also be an interesting concept for technical application in small scale grasping devices; e.g. for simple sampling mechanisms in small-sized autonomous underwater vehicles (AUVs).

category: zoology


title: statSuma: automated selection and performance of statistical comparisons for microbiome studies

abstract: There is a reproducibility crisis in scientific studies. Some of these crises arise from incorrect application of statistical tests to data that follow inappropriate distributions, have inconsistent equivariance, or have very small sample sizes. As determining which test is most appropriate for all data in a multicategorical study (such as comparing taxa between sites in microbiome studies), we present statsSuma, an interactive Python notebook (which can be run from any desktop computer using the Google Colaboratory web service) and does not require a user to have any programming experience. This software assesses underlying data structures in a given dataset to advise what pairwise or listwise statistical procedure would be best suited for all data. As some users may be interested in further mining specific trends, statSuma performs 5 different two-tailed pairwise tests (Students t-test, Welchs t-test, Mann-Whitney U-test, Brunner-Munzel test, and a pairwise Kruskal-Wallis H-test) and advises the best test for each comparison. This software also advises whether ANOVA or a multicategorical Kruskal-Wallis H-test is most appropriate for a given dataset and performs both procedures. A data distribution-vs-Gaussian distribution plot is produced for each taxon at each site and a variance plot between all combinations of 2 taxa at each site are produced so Gaussian tests and variance tests can be visually confirmed alongside associated statistical determinants.

category: microbiology


title: X-ray induced bio-acoustic emissions from cultured cells

abstract:
PurposeWe characterise for the first time the emission of bio-acoustic waves from cultured cells irradiated at doses of X-ray photon radiation relevant to medical and accidental exposure.

Methods and materialsHuman cancer cell lines (MCF-7, HL-60) and control cell-free media were exposed to 1 Gy X-ray photons while recording the sound generated before, during and after irradiation. Cellular cytotoxicity following photon irradiation was determined by extracellular LDH levels, and irradiated cell conditioned media were tested for their ability to elicit a bystander effect in reporter cells.

ResultsWe report the first recorded acoustic signals captured from a collective pressure wave response to ionising irradiation. The signature of the collective acoustic peaks was temporally wider and with higher acoustic power for irradiated HL-60 than for irradiated MCF-7.

ConclusionsWe show that at doses of X-ray irradiation capable of producing the release of bystander effect-inducing activity, both cell types emit a characteristic acoustic signal for the duration of the radiation pulse. The rapid signal decay is consistent with a passive rather than an active acoustic signal generation. This preliminary study suggests that further work on the potential role of radiation induced acoustic emission (RIAE) in the cellular bystander effect is merited.

category: cell biology


title: Targeting a proteolytic neo-epitope of CUB-domain containing protein 1 in RAS-driven cancer

abstract:
A central challenge for any therapeutic is targeting diseased over normal cells. Proteolysis is frequently upregulated in disease and can generate proteoforms with unique neo-epitopes. We hypothesize that targeting proteolytic neo-epitopes can enable more effective and safer treatments, reflecting a conditional layer of disease-specific regulation. Here, we characterized the precise proteolytic isoforms of CUB domain containing protein 1 (CDCP1), a protein overexpressed and specifically cleaved in RAS-driven cancers. We validated that the N-terminal and C-terminal fragments of CDCP1 remain associated after proteolysis in vitro and on the surface of pancreatic cancer cells. Using a differential phage display strategy, we generated exquisitely selective recombinant antibodies that target cells harboring cleaved CDCP1 and not the full-length form using antibody-drug conjugates or a bi-specific T-cell engagers. We show tumor-specific localization and anti-tumor activity in a syngeneic pancreatic tumor model having superior safety profiles compared to a pan-CDCP1-targeting antibody. Our studies show proteolytic neo-epitopes can provide an orthogonal "AND" gate for disease-specific targeting.

One-Sentence SummaryAntibody-based targeting of neo-epitopes generated by disease-associated proteolysis improves the therapeutic index

category: biochemistry


title: FlyPhoneDB: An integrated web-based resource for cell-cell communication prediction in Drosophila

abstract: Multicellular organisms rely on cell-cell communication to exchange information necessary for developmental processes and metabolic homeostasis. Cell-cell communication pathways can be inferred from transcriptomic datasets based on ligand-receptor (L-R) expression. Recently, data generated from single cell RNA sequencing (scRNA-seq) have enabled L-R interaction predictions at an unprecedented resolution. While computational methods are available to infer cell-cell communication in vertebrates such a tool does not yet exist for Drosophila. Here, we generated a high confidence list of L-R pairs for the major fly signaling pathways and developed FlyPhoneDB, a quantification algorithm that calculates interaction scores to predict L-R interactions between cells. At the FlyPhoneDB user interface, results are presented in a variety of tabular and graphical formats to facilitate biological interpretation. To demonstrate that FlyPhoneDB can effectively identify active ligands and receptors to uncover cell-cell communication events, we applied FlyPhoneDB to Drosophila scRNA-seq data sets from adult midgut, abdomen, and blood, and demonstrate that FlyPhoneDB can readily identify previously characterized cell-cell communication pathways. Altogether, FlyPhoneDB is an easy-to-use framework that can be used to predict cell-cell communication between cell types from scRNA-seq data in Drosophila.


title: seqgra: Principled Selection of Neural Network Architectures for Genomics Prediction Tasks

abstract: Sequence models based on deep neural networks have achieved state-of-the-art performance on regulatory genomics prediction tasks, such as chromatin accessibility and transcription factor binding. But despite their high accuracy, their contributions to a mechanistic understanding of the biology of regulatory elements is often hindered by the complexity of the predictive model and thus poor interpretability of its decision boundaries. To address this, we introduce seqgra, a deep learning pipeline that incorporates the rule-based simulation of biological sequence data and the training and evaluation of models, whose decision boundaries mirror the rules from the simulation process. The method can be used to (1) generate data under the assumption of a hypothesized model of genome regulation, (2) identify neural network architectures capable of recovering the rules of said model, and (3) analyze a models predictive performance as a function of training set size, noise level, and the complexity of the rules behind the simulated data.


title: Marginally compact phase and ordered ground states in a model polymer with side spheres

abstract: We present the results of a quantitative study of the phase behavior of a model polymer chain with side spheres using two independent computer simulation techniques. We find that the mere addition of side spheres results in key modifications of standard polymer behavior. One obtains a novel marginally compact phase at low temperatures, the structures in this phase are reduced in dimensionality and are ordered, they include strands assembled into sheets and a variety of helices, and at least one of the transitions on lowering the temperature to access these ordered states is found to be first order. Our model serves to partially bridge conventional polymer phases with biomolecular phases.

category: biophysics


title: Cross-species metabolomic analysis of DDT and Alzheimer's disease-associated tau toxicity

abstract:
BackgroundThe formation of hyperphosphorylated tau (p-tau) protein tangles in neurons is a pathological marker of Alzheimers disease (AD). Exposure to the pesticide dichlorodiphenyltrichloroethane (DDT) has been associated with increased risk of AD.

ObjectivesTo determine if there was a connection between DDT exposure and tau toxicity we investigated whether exposure to DDT can exacerbate tau protein toxicity in C. elegans. In addition, we examined the association between p-tau protein and metabolism in a human population study and in a transgenic C. elegans strain neuronally expressing a mutant tau protein fragment that is prone to aggregation.

MethodsIn the human population study, we used a metabolome-wide association framework to determine the association between p-tau measured in the cerebrospinal fluid (CSF) and metabolomic features measured in both plasma (n = 142) and CSF (n = 78) using high-resolution metabolomics (HRM). Using the same HRM method, we determined changes in metabolomic features in the transgenic C. elegans strain compared to its control strain. Metabolites associated with p-tau in both species were analyzed for overlap. We also examined the effect of DDT and aggregating tau protein on growth, swim behavior, mitochondrial function, metabolism, learning, and lifespan in C. elegans.

ResultsPlasma and CSF-derived features associated with p-tau level were related to drug, amino acid, fatty acid and mitochondrial metabolism pathways. Five metabolites overlapped between plasma and C. elegans, and 4 between CSF and C. elegans. DDT exacerbated the inhibitory effect of aggregating tau protein on growth and basal respiration. In the presence of aggregating tau protein, DDT induced more curling and was associated with reduced levels of amino acids but increased levels of uric acid and adenosylselenohomocysteine. Developmental exposure to DDT blunted the lifespan reduction caused by aggregating tau protein.

ConclusionThe model organism C. elegans can complement human studies by providing a means to study mechanisms of environmental toxicants. Specifically, our C. elegans data show that DDT exposure and tau protein aggregation both inhibit mitochondrial function and DDT exposure can exacerbate the mitochondrial inhibitory effects of tau protein aggregation providing a plausible explanation for the observed human associations.

category: pharmacology and toxicology


title: Gui Shao Di Huang Wan promotes angiogenesis and regulates oestrogen receptor α and progesterone receptor β in in vitro bioassays

abstract:
Background and aimThe formula Gui Shao Di Huang Wan (GSDW) is used frequently to treat female infertility. This study aims to investigate some of the possible mechanisms of action of GSDW using in vitro bioassays of angiogenesis in Human Uterine Microvascular Endothelial Cells (HUVEC) and Human Uterine Microvascular Endothelial Cells (HUtMEC) and ovarian steroid receptor expression in a human endometrial cell line (Ishikawa).

Experimental procedureAqueous extracts of GSDW and its component herbs were tested for pro-angiogenic activity using both HUVEC and HUtMEC 2D differentiation assays performed on Matrigel and effects on HUVEC proliferation using the MTT assay. Effects on the expression of Estrogen Receptor  (ER) and Progesterone Receptor {beta} (PR{beta}) in Ishikawa cells were determined using immunoblotting.

Results and ConclusionAll analysed parameters of differentiation were increased by GSDW in both the HUVEC and HUtMEC mesh. Furthermore, measures of total length, segment number, junction number were affected by some but not all component herbs.

The MTT assay showed an increase in proliferation of HUVECs at concentrations of GSDW between 0.68 and 5.47 g/mL at 48 and 72 hours.

In Ishikawa cells downregulation of ER and upregulation of PR{beta} was seen after 48 hours incubation with 4 of the 8 herbs in the formula.

The findings in this study demonstrate that GSDW has the potential to affect key parameters (vascular, sex hormone receptor expression) in vitro. This offers a mechanism by which these herbs may enhance fertility through improved endometrial receptivity and pregnancy rates.


title: The Prediction of Active Ingredients and Polypharmacological Mechanisms of Orthosiphon stamineus Benth. (Lamiaceae) of Dai medicine based on Network Pharmacology

abstract:
BackgroundOrthosiphon stamineus Benth. was a dietary supplement or a traditional folk herb with widespread clinical application, but it was lack of comprehensive understanding of its active ingredients and polypharmacological mechanisms. The aim of this work was to systematically study its natural compounds and the molecular mechanisms of the O. stamineus via network pharmacology.

MethodsCompounds from O. stamineus were collected by literature retrieval and evaluated by SwissADME with the physicochemical property (ADMET model) and the likelihood for a natural medicine. The connection of active ingredients and target genes was built and confirmed by Cytoscape and AutoDock vina. Then, Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were separately performed to obtain a more in-depth understanding of O. stamineus. Finally, the relationship among active ingredients, targets, and diseases was built to clarify the polypharmacological mechanisms and found relevant active substances for further drug discovery.

ResultsA total of 159 compounds of O. stamineus were collected, and 22 potentially active ingredients were screened out. The Ingredient-Target Interaction Network was built with 12 flavonoids, 3 diterpenes, 3 phenols, and 4 volatile oils, and 65 targets. The Docking analysis indicated that the ingredient-target interaction network was reliable; most ligand-receptor had a strong binding affinity (lowest binding energy: -6.9 kcal/mol). After pathway analysis, 185 significant biological processes and 36 signal pathways were found, and the ingredient-target-disease network of O. stamineus was constructed for polypharmacological mechanisms.

ConclusionOur study clarified the polypharmacological mechanisms via the relationship among active ingredients, targets, and diseases and provided better guidance for subsequent experiments and potential active ingredients for drug discovery or health promotion.

category: systems biology


title: An integrative analysis to predict the Active Ingredients and explore Polypharmacological Mechanisms of Orthosiphon stamineus Benth. for human health


title: Neuronal Activity in Dorsal Anterior Cingulate Cortex during EconomicChoices under Variable Action Costs

abstract: The role of the dorsal anterior cingulate cortex (ACCd) in decision making has often been discussed but remains somewhat unclear. On the one hand, numerous studies implicated this area in decisions driven by effort or action cost. On the other hand, work on economic choices between goods (under fixed action costs) found that neurons in ACCd encoded only post-decision variables. To advance our understanding of the role played by this area in decision making, we trained monkeys to choose between different goods (juice types) offered in variable amounts and with different action costs. Importantly, the task design dissociated computation of the action cost from planning of any particular action. Neurons in ACCd encoded the chosen value and the binary choice outcome in several reference frames (chosen juice, chosen cost, chosen action). Thus, this area provided a rich representation of post-decision variables. In contrast, neurons in ACCd did not represent pre-decision variables such as individual offer values in any reference frame. Hence, ongoing decisions are unlikely guided by ACCd. Conversely, neuronal activity in this area might inform subsequent choices.

category: neuroscience


title: Neuronal Activity in Dorsal Anterior Cingulate Cortex during Economic Choices under Variable Action Costs


title: Enhanced processing of aversive stimuli on embodied artificial limbs by the human amygdala

abstract: Body perception has been extensively investigated, with one particular focus being the integration of vision and touch within a neuronal body representation. Previous studies have implicated a distributed network comprising the extrastriate body area (EBA), posterior parietal cortex (PPC) and ventral premotor cortex (PMv) during illusory self-attribution of a rubber hand. Here, we set up an fMRI paradigm in virtual reality (VR) to study whether and how threatening (artificial) body parts affects their self-attribution. Participants (N=30) saw a spider (aversive stimulus) or a toy-car (neutral stimulus) moving along a 3D-rendered virtual forearm positioned like their real forearm, while tactile stimulation was applied on the real arm in the same (congruent) or opposite (incongruent) direction. We found that the PPC was more activated during congruent stimulation; higher visual areas and the anterior insula (aIns) showed increased activation during aversive stimulus presentation; and the amygdala was more strongly activated for aversive stimuli when there was stronger multisensory integration of body-related information (interaction of aversiveness and congruency). Together, these findings suggest an enhanced processing of aversive stimuli within the amygdala when they represent a bodily threat.


title: Sniffing Out New Friends: Similarity in Body-Odor Predicts the Quality of Same-Sex None-Romantic Dyadic Interactions

abstract: Most are familiar with the notion of socially "clicking" with someone, namely sensing an immediate bond that can lead to strong and often long-lasting friendships. The mechanisms underlying such rapid bonding remain unclear. Given that body-odor similarity is a critical cue for social interaction in non-human mammals, we tested the hypothesis that body-odor similarly contributes to bonding in same-sex non-romantic human dyads. We observed that objective ratings obtained with an electronic nose, and subjective ratings obtained from human smellers, converged to suggest that click-friends smell more similar to each other than random dyads. Remarkably, we then found that we could use the electronic nose to predict which strangers would later form better dyadic interactions. Thus, humans may literally sniff-out new friends based on similarities in body-odor.


title: Sniffing Out New Friends: Similarity in Body-Odor Predicts the Quality of Same-Sex Non-Romantic Dyadic Interactions


title: Longitudinal developmental trajectories do not follow cross-sectional age associations in hippocampal subfield and memory development

abstract:
Many cross-sectional findings suggest that volumes of specific hippocampal subfields increase in middle childhood and early adolescence. In contrast, a small number of available longitudinal studies observed decreased volumes in most subfields over this age range. Further, it remains unknown whether structural changes in development are associated with corresponding gains in childrens memory. Here we report cross-sectional age differences in childrens hippocampal subfield volumes together with longitudinal developmental trajectories and their relationships with memory performance. In two waves, 109 healthy participants aged 6 to 10 years (wave 1: MAge=7.25, wave 2: MAge=9.27) underwent high-resolution magnetic resonance imaging to assess hippocampal subfield volumes, and completed cognitive tasks assessing hippocampus dependent memory processes. We found that cross-sectional age-associations and longitudinal developmental trends in hippocampal subfield volumes were highly discrepant, both by subfields and in direction. Further, volumetric changes were largely unrelated to changes in memory, with the exception that increase in subiculum volume was associated with gains in spatial memory. Importantly, the observed longitudinal patterns of brain-cognition coupling could not be inferred from cross-sectional findings. We discuss potential sources of these discrepancies. This study underscores that childrens structural brain development and its relationship to cognition cannot be inferred from cross-sectional age comparisons.

HighlightsO_LIThe subiculum undergoes volumetric increase between 6-10 years of age
C_LIO_LIChange across two years in CA1-2 and DG-CA3 was not observed in this age window
C_LIO_LIChange across two years did not reflect age differences spanning two years
C_LIO_LICross-sectional and longitudinal slopes in stark contrast for hippocampal subfields
C_LIO_LILongitudinal brain-cognition coupling cannot be inferred from cross-sectional data
C_LI


title: Dynamic autonomic nervous system patterns differentiate human emotions and manifest in resting physiology

abstract:
Whether activity in the autonomic nervous system differs during distinct emotions remains controversial. We obtained continuous multichannel recordings of autonomic nervous system activity in healthy adults during a video-based emotional reactivity task. Dimensionality reduction revealed five principal components in the autonomic time series data, and these modes of covariation differentiated periods of baseline from those of video-viewing. Unsupervised clustering of the principal component time series data uncovered separable autonomic states that distinguished among the five emotion-inducing trials. These autonomic states were also detected in baseline physiology but were intermittent and of smaller magnitude. Our results suggest the autonomic nervous system assembles dynamic activity patterns during emotions that are similar across people and are present even during undirected moments of rest.

One Sentence SummaryDynamic autonomic patterns distinguish among emotions and are evident in resting physiology.


title: Transcriptomic analysis of dorsal and ventral subiculum after induction of acute seizures by electric stimulation of the perforant pathway in rats

abstract: Preconditioning is a mechanism in which injuries induced by non-lethal hypoxia or seizures trigger cellular resistance to subsequent events. Norwood et al., in a 2010 study, showed that an 8-hour-long period of electrical stimulation of the perforant pathway in rats is required for the induction of hippocampal sclerosis. However, in order to avoid generalized seizures, status epilepticus (SE), and death, a state of resistance to seizures must be induced in the hippocampus by a preconditioning paradigm consisting of 2 daily 30-minute stimulation periods. Due to the importance of the subiculum in the hippocampal formation, this study aims to investigate differential gene expression patterns in the dorsal and ventral subiculum using RNA-sequencing, after induction of a preconditioning protocol by electrical stimulation of the perforant pathway. The dorsal (dSub) and ventral (vSub) subiculum regions were collected by laser-microdissection 24 hours after preconditioning protocol induction in rats. RNA sequencing was performed in a Hiseq 4000 platform, reads were aligned using the STAR and DESEq2 statistics package was used to estimate gene expression. We identified 1176 differentially expressed genes comparing control to preconditioned subiculum regions, 204 genes were differentially expressed in dSub and 972 in vSub. The gene ontology enrichment analysis showed that the most significant common enrichment pathway considering up-regulated genes in dSub and vSub was Cholesterol Biosynthesis. In contrast, the most significant enrichment pathway considering down-regulated genes in vSub was Axon guidance. Our results indicate that preconditioning induces synaptic reorganization, increased cholesterol metabolism, and astrogliosis in both dSub and vSub. Both regions also presented a decrease in glutamatergic transmission, an increase in complement system activation, and increased in GABAergic transmission. The down-regulation of proapoptotic and axon guidance genes in the ventral subiculum suggests that preconditioning induces a neuroprotective environment in this region.


title: MorphVAE: Generating Neural Morphologies from 3D-Walks using a Variational Autoencoder with Spherical Latent Space

abstract: For the past century, the anatomy of a neuron has been considered one of its defining features: The shape of a neurons dendrites and axon fundamentally determines what other neurons it can connect to. These neurites have been described using mathematical tools e.g. in the context of cell type classification, but generative models of these structures have only rarely been proposed and are often computationally inefficient. Here we propose MO_SCPLOWORPHC_SCPLOWVAE, a sequence-to-sequence variational autoencoder with spherical latent space as a generative model for neural morphologies. The model operates on walks within the tree structure of a neuron and can incorporate expert annotations on a subset of the data using semi-supervised learning. We develop our model on artificially generated toy data and evaluate its performance on dendrites of excitatory cells and axons of inhibitory cells of mouse motor cortex (M1) and dendrites of retinal ganglion cells. We show that the learned latent feature space allows for better cell type discrimination than other commonly used features. By sampling new walks from the latent space we can easily construct new morphologies with a specified degree of similarity to their reference neuron, providing an efficient generative model for neural morphologies.


title: Single-synapse analyses of Alzheimers disease implicate pathologic tau, DJ1, CD47, and ApoE

abstract: Synaptic molecular characterization is limited for Alzheimers disease (AD). We used mass cytometry to quantify 38 probes in approximately 17 million single synaptic events from human brains without pathologic change or with pure AD or Lewy body disease (LBD), non-human primates (NHP), and PS/APP mice. Synaptic molecular integrity in humans and NHP was similar. Although not detected in human synapses, A{beta} was in PS/APP mice synaptic events. Clustering and pattern identification of human synapses showed expected disease-specific differences, like increased hippocampal pathologic tau in AD and reduced caudate dopamine transporter in LBD, and revealed novel findings including increased hippocampal CD47 and lowered DJ1 in AD and higher ApoE in AD dementia. Our results were independently supported by multiplex ion beam imaging of intact tissue.


title: Value-based decision-making between affective memories

abstract: Affective biases can influence how past events are recalled from memory. However, the mechanisms underlying how discrete affective events shape memory formation and subsequent recall are not well understood. Further understanding this is important given the central role of negative biases in affective memory recall in depression and antidepressant drug action. In order to capture cognitive processes associated with affective memory formation and recall, we studied value-based decision-making between affective memories in two within-subject experiments (n=45 and n=74). Our findings suggest that discrete affective events, created by large magnitude Wheel of Fortune (WoF) outcomes, influence affective memory formation processes during reinforcement-learning (RL). After 24 hours, we show that healthy volunteers display stable preferences during value-based recall of affective memories in a binary decision-making task. Computational modelling of these preferences demonstrated a positive bias during value-based recall, induced by previously winning in the WoF. We further showed that value-based decision-making between affective memories engages the pupil-linked central arousal systems, leading to pupil constriction prior to, and differential pupil dilation after the decision onset depending on the valence of the chosen options. Taken together, we demonstrate that mechanisms underlying human affective memory systems can be described by RL and probability weighting models. This approach could be used as a translational assay to study the effects of novel antidepressants.


title: Distributional reinforcement learning in prefrontal cortex

abstract: Prefrontal cortex is crucial for learning and decision-making. Classic reinforcement learning (RL) theories centre on learning the expectation of potential rewarding outcomes and explain a wealth of neural data in prefrontal cortex. Distributional RL, on the other hand, learns the full distribution of rewarding outcomes and better explains dopamine responses. Here we show distributional RL also better explains prefrontal cortical responses, suggesting it is a ubiquitous mechanism for reward-guided learning.


title: The IL-33-ILC2 pathway protects from amebic colitis

abstract: Entamoeba histolytica is a pathogenic protozoan parasite that causes intestinal colitis, diarrhea, and in some cases, liver abscess. Through transcriptomics analysis, we observed that E. histolytica infection was associated with increased expression of IL-33 mRNA in both the human and murine colon. IL-33, the IL-1 family cytokine, is released after cell injury to alert the immune system of tissue damage during infection. Treatment with recombinant IL-33 protected mice from amebic infection and colonic tissue damage; moreover, blocking IL-33 signaling made mice more susceptible to infection and weight loss. IL-33 limited the recruitment of inflammatory immune cells and decreased the pro-inflammatory cytokine IL-6 in the colon. Type 2 immune responses, which are known to be involved in tissue repair, were upregulated by IL-33 treatment during amebic infection. Interestingly, administration of IL-33 protected RAG2-/- mice but not RAG2-/-{gamma}c-/- mice, demonstrating that IL-33 mediated protection occurred in the absence of T or B cells but required the presence of innate lymphoid cells (ILCs). IL-33 induced recruitment of ILC2 but not ILC1 and ILC3 in RAG2-/- mice. Adoptive transfer of ILC2s to RAG2-/-{gamma}c-/- mice restored IL-33 mediated protection. These data reveal that the IL-33-ILC2 pathway is an important host defense mechanism against amebic colitis.

category: immunology


title: Profiling the in vitro and in vivo activity of streptothricin-F against carbapenem-resistant Enterobacterales: a historic scaffold with a novel mechanism of action

abstract:
Streptothricins are components of the natural product, nourseothricin; each containing identical streptolidine and gulosamine aminosugar moieties attached to varying numbers of linked {beta}-lysines. Nourseothricin was discovered by Waksman and colleagues in the early 1940s, generating intense interest because of excellent Gram-negative activity. However, the natural product mixture was associated with toxicity, and subsequent exploration was limited. Here, we establish the activity spectrum of nourseothricin and its main components, streptothricin-F (S-F, one lysine) and streptothricin D (S-D, three lysines), purified to homogeneity, against highly drug-resistant, carbapenem-resistant Enterobacterales (CRE). The MIC50 and MIC90 for S-F and S-D were 2 and 4 {micro}M, and 0.25 and 0.5 {micro}M, respectively. S-F and nourseothricin showed rapid, bactericidal activity. S-F and S-D both showed approximately 40-fold greater selectivity for prokaryotic than eukaryotic ribosomes in in vitro translation assays. There was >10-fold in vitro selectivity of S-F compared with S-D on LLC-PK1 and J774 cell lines. In vivo, delayed renal toxicity occurred at >10-fold higher doses of S-F compared with S-D. Substantial treatment effect of S-F in the murine thigh model was observed against the otherwise pandrug-resistant, NDM-1-expressing Klebsiella pneumoniae Nevada strain at dosing levels without observable or minimal toxicity. Resistance mutations obtained in single ribosomal operon E. coli identify novel interactions with 16S rRNA helix 34, i.e., C1504A and A1196G/C conferred high level resistance to nourseothricin. Based on promising, unique activity, we suggest that the streptothricin scaffold deserves further pre-clinical exploration as a potential therapeutic for the treatment of CRE and potentially other multidrug-resistant, gram-negative pathogens.

IMPORTANCEStreptothricins are a historic class of antibiotics discovered by Waksman and colleagues in the 1940s. Toxicities associated with the streptothricin natural product mixture, also known as nourseothricin, discouraged further development. However, we found that a component of nourseothricin, streptothricin-F, retained potent activity against contemporary carbapenem-resistant Enterobacterales with significant selectivity in in vitro and in vivo assays. This included demonstration of rapid bactericidal activity in vitro and substantial therapeutic effect in the murine thigh model against a pandrug-resistant Klebsiella pneumoniae isolate at non-toxic concentrations. Through resistance mutation analysis, we identified helix 34 of 16S rRNA in the prokaryotic ribosome, and specifically bases C1054 and A1196, as critical for streptothricins activity. The mechanism of action is distinct from other known translation inhibitors. Based on promising and unique activity, we believe the streptothricin scaffold deserves further pre-clinical exploration as a potential therapeutic for the treatment of CRE and potentially other multidrug-resistant, Gram-negative pathogens.


title: Phase-separated stress granules and processing bodies are compromised in Myotonic Dystrophy Type 1

abstract: RNA regulation in mammalian cells requires complex physical compartmentalisation using structures thought to be formed by liquid-liquid phase separation. Disruption of these structures is implicated in numerous degenerative diseases. Myotonic Dystrophy Type 1 (DM1) is a multi-systemic trinucleotide repeat disorder resulting from a CTG expansion in the dystonia myotonica protein kinase gene (DMPK). The cellular hall-mark of DM1 is the formation of nuclear foci containing expanded DMPK RNA (CUGexp). We report here the deregulation of stress granules and processing bodies (P-bodies), two cytoplasmic structures key for mRNA regulation, in cell culture models of DM1. Alterations to the rates of formation and dispersal of stress granules suggest an altered ability to respond to stress associated with DM1, while changes to the structure and dynamics of stress granules and P-bodies suggest that a more widespread alteration to the biophysical properties of cellular structures may be a consequence of the presence of CUGexp RNA.


title: Here be dragons: important spatial uncertainty driven by climate data in forecasted distribution of an endangered insular reptile

abstract: The effect of future climate change is poorly documented in the tropics, especially in mountainous areas. Yet, species living in these environments are predicted to be strongly affected. Newly available high-resolution environmental data and statistical methods enable the development of forecasting models. Nevertheless, the uncertainty related to climate models can be strong, which can lead to ineffective conservation actions. Predicted studies aimed at providing conservation guidelines often account for a range of future climate predictions (climate scenarios and global circulation models). However, very few studies considered potential differences related to baseline climate data and/or did not account for spatial information (overlap) in uncertainty assessments. We modelled the environmental suitability for Phelsuma borbonica, an endangered reptile native to Reunion Island. Using two metrics of species range change (difference in overall suitability and spatial overlap), we quantified the uncertainty related to the modelling technique (n = 10), sample bias correction, climate change scenario, global circulation models (GCM) and baseline climate (CHELSA versus Worldclim). Uncertainty was mainly driven by GCMs when considering overall suitability, while for spatial overlap the uncertainty related to baseline climate became more important than that of GCMs. The uncertainty driven by sample bias correction and variable selection was much higher when assessed based on spatial overlap. The modelling technique was a strong driver of uncertainty in both cases. We eventually provide a consensus ensemble prediction map of the environmental suitability of P. borbonica to identify the areas predicted to be the most suitable in the future with the highest certainty. Predictive studies aimed at identifying priority areas for conservation in the face of climate change need to account for a wide panel of modelling techniques, GCMs and baseline climate data. We recommend the use of multiple approaches, including spatial overlap, when assessing uncertainty in species distribution models.

category: ecology


title: miR-486 is an epigenetic modulator of Duchenne muscular dystrophy pathologies

abstract:
Duchenne muscular dystrophy (DMD) is an X-linked progressive muscle disorder resulting in muscle weakness and cardiomyopathy. MicroRNAs have been shown to play essential roles in muscle development, metabolism, and disease pathologies. We demonstrated that miR-486 expression is reduced in DMD muscles and its expression levels correlate with dystrophic disease severity. MicroRNA-486 knockout mice developed disrupted myofiber architecture, decreased myofiber size, decreased locomotor activity, increased cardiac fibrosis, and metabolic defects that were exacerbated on the dystrophic mdx5cv background. We integrated RNA-sequencing and chimeric eCLIP-sequencing data to identify direct in vivo targets of miR-486 and associated dysregulated gene signatures in skeletal muscle. In comparison to our DMD mouse muscle transcriptomes, we identified several of these miR-486 muscle targets including known modulators of dystrophinopathy disease symptoms. Together, our studies identify miR-486 as a driver of muscle remodeling in DMD, a useful biomarker for dystrophic disease progression, and highlight chimeric eCLIP-sequencing as a useful tool to identify direct in vivo microRNA target transcripts.

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category: genetics


title: Regulome-wide association study identifies enhancer properties associated with risk for schizophrenia

abstract:
Genetic risk for complex traits is strongly enriched in non-coding genomic regions involved in gene regulation, especially enhancers. However, we lack adequate tools to connect the characteristics of these disruptions to genetic risk. Here, we propose RWAS (Regulome Wide Association Study), a new framework to identify the characteristics of enhancers that contribute to genetic risk for disease. Applying our technique to interrogate genetic risk for schizophrenia, we found that risk-associated enhancers in this disease are predominantly active in the brain, evolutionarily conserved, and AT-rich. The association between AT percentage and risk corresponds to an overrepresentation in risk-associated enhancers for the binding sites of transcription factors that recognize AT-rich cis-regulatory motifs. Several of the TFs identified in our model as being overrepresented in risk-associated enhancers, including MEF2C, are master regulators of neuronal development. The genes that encode several of these TFs are themselves located at genetic risk loci for schizophrenia. This list also includes brain-expressed TFs that have not previously been linked to schizophrenia. In summary, we developed a generalizable approach that integrates GWAS summary statistics with enhancer characteristics to identify risk factors in tissue-specific regulatory regions.

AUTHOR SUMMARYEnhancers are regulatory regions that influence gene expression via the binding of transcription factors. Risk for many heritable diseases is enriched in regulatory regions, including enhancers. In this study, we introduce a novel method of testing for association between enhancer attributes and risk and use this method to determine the enhancer characteristics that are associated with risk for schizophrenia. We found that enhancers associated with schizophrenia risk are both evolutionarily conserved and in physical contact with mutation-intolerant genes, many of which have neurodevelopmental functions. Risk-associated enhancers are also AT-rich and contain binding sites for neurodevelopmental transcription factors.

category: genomics


title: Daily Patterns of Foraging and Aggressive Behaviors in Great-tailed Grackle (Quiscalus mexicanus) at an Urban Patch with Availability or Absence of Resources

abstract: The Great-tailed Grackle (Quiscalus mexicanus) seems to take advantage of inhospitable environments such as cities. However, it is not yet fully understood how these birds exploit hostile environments to their advantage. Casual observation seems to suggest that this species can obtain resources of biological importance such as food or nesting material from the trash. As a first approach to the problem, we located a patch outside a residential building, in a high-density urban area, where the residents left their trash for pick-up, and a group of wild Great-tailed grackles was identified as regular visitors. In total, 25 days were recorded at the site (November 2017 - January 2018). Events such as foraging, number of subjects present at the foraging site, aggressive behaviors between members of the group, and their relation with the presence or absence of the garbage collector truck were registered. The results show a higher number of grackles at the observation site and a higher frequency of foraging behaviors in the presence of garbage collection than in its absence. In its presence, the distribution of foraging during the day follows a normal distribution. In the absence, the distribution shows more variability towards the day. The highest frequency of interactions occurred between two grackles, yet there were records of up to eight subjects. The highest number of aggressions registered took place in the absence of garbage collection than in its presence. Moreover, the focal subject exhibits fewer agonistic behaviors compared to other members of the group, a result expected if the producer-scrounger game literature is considered. The outcome is explained in terms of deprivation and availability of resources. Finally, we conclude that grackles can exploit hazardous environments such as cities due to the highly social behaviors exhibited during foraging.


title: Sequencing uracil in DNA at single-nucleotide resolution

abstract: As an aberrant base in DNA, uracil is generated by dUMP misincorporation or cytosine deamination, and involved in multiple physiological and pathological processes. Current methods for whole-genome mapping of uracil all rely on uracil-DNA N-glycosylase (UNG) and are limited in resolution or specificity. Here, we present a UNG-independent Single-Nucleotide resolution Uracil Sequencing (SNU-seq) method utilizing the UdgX protein which specifically excises the uracil and forms a covalent bond with the resulting deoxyribose. SNU-seq was validated on synthetic DNA and applied to mammalian genomes. We found that the uracil content of pemetrexed-treated cells fluctuated along with DNA replication timing. We also accurately detected uracil introduced through cytosine deamination by the cytosine base editor (nCas9-APOBEC) and verified uracil occurrence in "WRC" motif within Activation-Induced Cytidine Deaminase (AID) hotspot regions in CSR-activated UNG -/- B cells.


title: Embryonic development of fully biocompatible organic light-emitting diodes

abstract: Prototype fully biocompatible organic light-emitting diodes are investigated, with a view to creating a suitable and high-performance light source as a medical implant device. A selection of organic LED materials that have potential suitability for the biological environment are examined. First, the biocompatibility of selected OLED materials was evaluated by the study of cell adhesion and cytotoxicity of HeLa cells cultured on the candidate materials. Thus it was possible to design a device structure composed entirely of biocompatible materials. Second, the characterization of the electroluminescence properties of the prototype OLED is shown and its limitation evaluated. Third, the aqueous stability of the fully biocompatible light source is examined. There is strong evidence that fully biocompatible and stable light-emitting implant devices can be easily constructed. This is the first time a fully biocompatible organic light-emitting diode, albeit embryonic, is reported, with the hope that it may lead to further research to optimize the device performance. Some suggestions on suitable device properties towards in vivo transition are provided.

category: bioengineering


title: Klarigi: Explanations for Semantic Groupings

abstract:
SummarySemantic annotation facilitates the use of background knowledge in analysis. This includes approaches that sort entities into groups, clusters, or assign labels or outcomes that are typically difficult to derive semantic explanations for. We introduce Klarigi, a tool that creates semantic explanations for groups of entities described by ontology terms implemented in a manner that balances multiple scoring heuristics. We demonstrate Klarigi by using it to identify characteristic terms for text-derived phenotypes of emergency admissions for two frequently conflated diagnoses, pulmonary embolism and pneumonia. Klarigi provides a universal method by which entity groups or labels can be explained semantically, and thus contributes to improved explainability of analysis methods.

Availability and ImplementationKlarigi is freely available under an open source licence at http://github.com/reality/klarigi. Supplementary data is available with this article.

Contactl.slater.1@bham.ac.uk


title: Deep learning trained on H&E tumor ROIs predicts HER2 status and Trastuzumab treatment response in HER2+ breast cancer

abstract: The current standard of care for many patients with HER2-positive breast cancer is neoadjuvant chemotherapy in combination with anti-HER2 agents, based on HER2 amplification as detected by in situ hybridization (ISH) or protein immunohistochemistry (IHC). However, hematoxylin & eosin (H&E) tumor stains are more commonly available, and accurate prediction of HER2 status and anti-HER2 treatment response from H&E would reduce costs and increase the speed of treatment selection. Computational algorithms for H&E have been effective in predicting a variety of cancer features and clinical outcomes, including moderate success in predicting HER2 status. In this work, we present a novel convolutional neural network (CNN) approach able to predict HER2 status with increased accuracy over prior methods. We trained a CNN classifier on 188 H&E whole slide images (WSIs) manually annotated for tumor regions of interest (ROIs) by our pathology team. Our classifier achieved an area under the curve (AUC) of 0.90 in cross-validation of slide-level HER2 status and 0.81 on an independent TCGA test set. Within slides, we observed strong agreement between pathologist annotated ROIs and blinded computational predictions of tumor regions / HER2 status. Moreover, we trained our classifier on pre-treatment samples from 187 HER2+ patients that subsequently received trastuzumab therapy. Our classifier achieved an AUC of 0.80 in a five-fold cross validation. Our work provides an H&E-based algorithm that can predict HER2 status and trastuzumab response in breast cancer at an accuracy that is better than IHC and may benefit clinical evaluations.


title: Early mismatch between cancer cells and their tumour microenvironment blocks long term growth

abstract: Cancer cells thrive embedded in a fine-tuned cellular and extracellular environment or tumour microenvironment (TME). There is a general understanding of a co-evolution between cancer cells and their surrounding TME, pointing at a functional connection between cancer cells characteristics and the perturbations induced in their surrounding tissue. However, whether this functional connection needs to be set from the start or if aggressive cancer cells can always be dominating their microenvironment has never been formally proven with a dedicated experimental setting where malignant cells can be challenged to grow in a different TME from the one they would naturally create. Here we generated an experimental setting where we transiently perturb the secretory profile of aggressive breast cancer cells without affecting their intrinsic growth ability. This led to the initial establishment of an atypical TME. Interestingly, even if initially tumours are formed, this atypical TME evolves to impair long term in vivo cancer growth. Using a combination of in vivo transcriptomics, protein arrays and in vitro co-cultures, we found that the atypical TME culminates in the infiltration of macrophages with STAT1high activity. These macrophages show strong anti-tumoural functions which reduce long-term tumour growth, despite lacking canonical M1 markers. Importantly, gene signatures of the mesenchymal compartment of the TME, as well as the anti-tumoural macrophages show striking prognostic power-correlating with less aggressive human breast cancers.

category: cancer biology


title: Early functional mismatch between breast cancer cells and their tumour microenvironment suppresses long term growth


title: Live Imaging Reveals the Cellular Events Downstream of SARM1 Activation

abstract: SARM1 is an inducible NAD+ hydrolase that triggers axon loss and neuronal cell death in the injured and diseased nervous system. While SARM1 activation and enzyme function are well defined, the cellular events downstream of SARM1 activity but prior to axonal demise are much less well understood. Defects in calcium, mitochondria, ATP, and membrane homeostasis occur in injured axons, but the relationships among these events have been difficult to disentangle because prior studies analyzed large collections of axons in which cellular events occur asynchronously. Here we used live imaging with single axon resolution to investigate the cellular events downstream of SARM1 activity. Our studies support a model in which SARM1 NADase activity leads to an ordered sequence of events from loss of cellular ATP, to defects in mitochondrial movement and depolarization, followed by calcium influx, externalization of phosphatidylserine, and loss of membrane permeability prior to catastrophic axonal self-destruction.


title: Regulation of auditory fear discrimination by the novel Kv3 voltage-gated potassium channel modulator AUT00206

abstract: Psychiatric diseases like anxiety-related disorders and schizophrenia are characterized by impaired cognition and emotional regulation linked to corticolimbic disinhibition. Restoring the balance between excitation and inhibition in corticolimbic circuits may therefore ameliorate certain features of these disorders, such as inappropriately attributing affective salience to innocuous cues. Corticolimbic activity is tightly controlled by parvalbumin-expressing GABAergic interneurons, which also regulate fear discrimination. The voltage-gated potassium channels Kv3.1 and Kv3.2 are highly expressed in these neurons, therefore Kv3.1/3.2 modulation may have potential for treating disorders associated with cognitive and emotional dysregulation. We determined the effects of the novel Kv3.1/3.2 positive modulator AUT00206 on fear discrimination. Female rats underwent limited or extended auditory fear discrimination training that we previously showed leads to discrimination or generalization, respectively, based on passive fear responding (i.e. freezing). We also assessed darting as an active fear response. We found that limited training resulted in discrimination based on freezing, which was unaffected by AUT00206. In contrast, we found that extended training resulted in generalization based on freezing and the emergence of discrimination based on darting. Importantly, AUT00206 had dissociable effects on fear discrimination and expression with extended training. While AUT00206 mitigated generalization without affecting expression based on freezing, it reduced expression without affecting discrimination based on darting. Our results indicate that Kv3.1/3.2 modulation regulates the attribution of affective significance to threat- and safety-related cues in a response-specific manner. This suggests that targeting Kv3.1 and Kv3.2 channels may provide a promising avenue for treating cognitive and emotional dysregulation in psychiatric disease.


title: The voltage and spiking responses of subthreshold resonant neurons to structured and fluctuating inputs: resonance, loss of resonance and variability

abstract: We systematically investigate the response of neurons to oscillatory currents and synaptic-like inputs and we extend our investigation to non-structured synaptic-like spiking inputs with more realistic distributions of presynaptic spike times. We use two types of chirp-like inputs consisting of (i) a sequence of cycles with discretely increasing frequencies over time, and (ii) a sequence having the same cycles arranged in an arbitrary order. We develop and use a number of frequency-dependent voltage response metrics to capture the different aspects of the voltage response, including the standard impedance (Z) and the peak-to-trough amplitude envelope (VENV) profiles. We show that Z-resonant cells (cells that exhibit subthreshold resonance in response to sinusoidal inputs) also show VENV-resonance in response to sinusoidal inputs, but generally do not (or do it very mildly) in response to square-wave and synaptic-like inputs. In the latter cases the resonant response using Z is not predictive of the preferred frequencies at which the neurons spike when the input amplitude is increased above subthreshold levels. We also show that responses to conductance-based synaptic-like inputs are attenuated as compared to the response to current-based synaptic-like inputs, thus providing an explanation to previous experimental results. These response patterns were strongly dependent on the intrinsic properties of the participating neurons, in particular whether the unperturbed Z-resonant cells had a stable node or a focus. In addition, we show that variability emerges in response to chirp-like inputs with arbitrarily ordered patterns where all signals (trials) in a given protocol have the same frequency content and the only source of uncertainty is the subset of all possible permutations of cycles chosen for a given protocol. This variability is the result of the multiple different ways in which the autonomous transient dynamics is activated across cycles in each signal (different cycle orderings) and across trials. We extend our results to include high-rate Poisson distributed current- and conductance-based synaptic inputs and compare them with similar results using additive Gaussian white noise. We show that the responses to both Poisson-distributed synaptic inputs are attenuated with respect to the responses to Gaussian white noise. For cells that exhibit oscillatory responses to Gaussian white noise (band-pass filters), the response to conductance-based synaptic inputs are low-pass filters, while the response to current-based synaptic inputs may remain band-pass filters, consistent with experimental findings. Our results shed light on the mechanisms of communication of oscillatory activity among neurons in a network via subthreshold oscillations and resonance and the generation of network resonance.


title: The voltage and spiking responses of subthreshold resonant neurons to structured and fluctuating inputs: persistence and loss of resonance and variability


title: Oscillations and variability in neuronal systems: interplay of autonomous transient dynamics and fast deterministic fluctuations

abstract: Neuronal systems are subject to rapid fluctuations both intrinsically and externally. These fluctuations can be disruptive or constructive. We investigate the dynamic mechanisms underlying the interactions between rapidly fluctuating signals and the intrinsic properties of the target cells to produce variable and/or coherent responses. We use linearized and non-linear conductance-based models and piecewise constant (PWC) inputs with short duration pieces. The amplitude distributions of the constant pieces consist of arbitrary permutations of a baseline PWC function. In each trial within a given protocol we use one of these permutations and each protocol consists of a subset of all possible permutations, which is the only source of uncertainty in the protocol. We show that sustained oscillatory behavior can be generated in response to various forms of PWC inputs independently of whether the stable equilibria of the corresponding unperturbed systems are foci or nodes. The oscillatory voltage responses are amplified by the model nonlinearities and attenuated for conductance-based PWC inputs as compared to current-based PWC inputs, consistent with previous theoretical and experimental work. In addition, the voltage responses to PWC inputs exhibited variability across trials, which is reminiscent of the variability generated by stochastic noise (e.g., Gaussian white noise). Our analysis demonstrates that both oscillations and variability are the result of the interaction between the PWC input and the target cells autonomous transient dynamics with little to no contribution from the dynamics in vicinities of the steady-state, and do not require input stochasticity.


title: Identification of enamel knot gene signature within the developing mouse molar

abstract: In most mammals, the primary teeth develop in utero and the cells capable of contributing to hard surface regeneration are lost before tooth eruption. These cells differentiate through a series of reciprocal induction steps between the epithelium and mesenchyme, initially orchestrated by an epithelial signaling center called the enamel knot. While the factors secreted by this structure are of interest to the dental regeneration and development communities, its small size makes it difficult to isolate for analysis. Here we describe our work to identify the enamel knot from whole E14 molars using publicly available scRNA-seq data. We identified 335 genes differentially expressed in the enamel knot compared to the surrounding tissues, including known enamel knot marker genes. We validated expression of the most highly enriched enamel knot marker genes and identified 42 novel marker genes of the enamel knot which provide excellent targets for future dental regeneration investigations.

category: developmental biology


title: 300 years of change for native fish species in the upper Danube River Basin - historical flow alterations versus future climate change

abstract: River ecosystems belong to the most threatened ecosystems on Earth. Historical anthropogenic alterations have, and future climate change will further affect river ecosystems and the species therein. While many studies assess the potential effects of expected future changes on species, little is known about the severity of these changes compared to historical alterations. Here, we used a unique 300-year time series of hydrological and climate data to assess the vulnerability of 48 native fish species in the upper Danube River Basin. We calculated species-specific vulnerability estimates relative to the reference period (1970-2000) for the periods 1800-1830, 1900-1930, and 2070-2100, including two Representative Concentration Pathways (RCP 4.5 and 8.5) and identified the environmental drivers of vulnerability estimates. Models showed that future changes under RCP 4.5 would result in moderate species vulnerability compared to historical conditions, while under RCP 8.5, the vulnerability for all species increased substantially. In addition, species vulnerability was mainly driven by hydrology in the past and is likely to be driven by temperature in the future. Our results show that future climate change would alter environmental conditions for riverine fish species at a similar magnitude as historical anthropogenic hydrological river alterations have. Shedding light on such long-term historical and possible future anthropogenic alterations provides valuable insights for prioritising conservation actions for riverine fish species.


title: Harnessing the power of metabarcoding in the ecological interpretation of plant-pollinator DNA data: strategies and consequences of reads filtering

abstract: DNA metabarcoding approaches to analyse complex mixtures of pollen has become the standard in pollination biology, especially in the light of recent threats affecting pollination. In spite of the increasing adoption of High Throughput Sequencing (HTS) approaches, these studies generate huge numbers of raw reads, some of which might be associated to false positives or infrequently recorded species with potentially little biological information. If these reads are not discarded (i.e. pruned), they can lead to changes in the ecological findings and lead to biased conclusions. In this study we reviewed 42 papers in the recent pollen DNA metabarcoding literature and focused on the type of pruning applied. We also tested whether the different types of those cut off threshold may leave a mark on the DNA metabarcoding data. To do so, we compared for the first time community composition, species richness and networks of species interactions (i.e. Connectace, Modularity, Connectivity and Shannon entropy) associated with the most relevant ways of treating HTS outputs: no cut (no reads filtering), or cutting levels obtained as proportional 1% of sample total reads, or as fixed amount of 100 reads, or from ROC (Receiver operator characteristic). Results clearly indicated that pruning type shapes species composition and that to apply or not a threshold dramatically impacts ecological indices, potentially increasing the risk of misinterpreting DNA metabarcoding data under an ecological point of view. Given the high methodological heterogeneity from the revised literature, we discuss in what conditions filtering types may be more appropriate, and also recommend to biologically justify the pruning threshold when analysing DNA metabarcoding raw reads, and to develop shared approaches to make future studies more comparable.


title: Managing African Swine Fever: Assessing the Potential of Camera Traps in Monitoring Wild Boar Occupancy Trends in Infected and Noninfected Zones, Using Spatio-temporal Statistical Models

abstract: The recent spreading of African swine fever (ASF) over the Eurasian continent has been acknowledged as a serious economic threat for the pork industry. Consequently, an extensive body of research focuses on the epidemiology and control of ASF. Nevertheless, little information is available on the combined effect of ASF and ASF-related control measures on wild boar (Sus scrofa) population abundances. This is crucial information given the role of the remaining wild boar that act as an important reservoir of the disease. Given the high potential of camera traps as a non-invasive method for ungulate trend estimation, we assess the effectiveness of ASF control measures using a camera trap network. In this study, we focus on a major ASF outbreak in 2018-2020 in the South of Belgium. This outbreak elicited a strong management response, both in terms of fencing off a large infected zone as well as an intensive culling regime. We apply a Bayesian multi-season site-occupancy model to wild boar detection-nondetection data. Our results show that (1) occupancy rates at the onset of our monitoring period reflect the ASF infection status; (2) ASF-induced mortality and culling efforts jointly lead to decreased occupancy over time; and (3) the estimated mean total extinction rate ranges between 22.44% and 91.35%, depending on the ASF infection status. Together, these results confirm the effectiveness of ASF-control measures implemented in Wallonia (Belgium), which has regained its disease-free status in December 2020, as well as the usefulness of a camera trap network to monitor these effects.


title: Application of ATAC-Seq for genome-wide analysis of the chromatin state at single myofiber resolution

abstract: Skeletal myofibers are the main components of skeletal muscle which is the largest tissue in the body. Myofibers are highly adaptive in nature and they can vary in different biological and disease conditions. Therefore, transcriptional and epigenetic studies on myofibers are crucial to discover how chromatin alterations occur in the skeletal muscle under different conditions. However, due to the heterogenous nature of skeletal muscle, studying myofibers in isolation proves to be a challenging task. Single cell sequencing has permitted for the study of the epigenome of isolated myonuclei. While this provides sequencing with high dimensionality, the sequencing depth is lacking, which makes comparisons between different biological conditions difficult. Here we report the first implementation of single myofiber ATAC-Seq, which permits for the sequencing of an individual myofiber at a depth sufficient for peak calling and for comparative analysis of chromatin accessibility under various physiological, physical and disease conditions. Application of this technique revealed significant differences in chromatin accessibility between resting and regenerating myofibers. This technique can lead to wide application in identifying chromatin regulatory elements and epigenetic mechanisms in muscle fibers during development and in muscle-wasting diseases.


title: Engineered small extracellular vesicles as a FGL1/PD-L1 dual-targeting delivery system for alleviating immune rejection

abstract: There is an urgent need for developing new immunosuppressive agents due to the toxicity of long-term use of broad immunosuppressive agents post organ transplantation. Comprehensive sample analysis revealed dysregulation of FGL1/LAG-3 and PD-L1/PD-1 immune checkpoints in allogeneic heart transplantation mice and clinical kidney transplant patients. In order to enhance these two immunosuppressive signal axes, we developed a bioengineering strategy to simultaneously display FGL1/PD-L1 (FP) on the surface of small extracellular vesicles (sEVs). Among various cell sources, FP sEVs derived from mesenchymal stem cells (MSCs) not only enriched FGL1/PD-L1 expression but also maintained the immunomodulatory properties of unmodified MSC sEVs. Next, we confirmed that FGL1 and PD-L1 on sEVs were specifically bound to their receptors LAG-3 and PD-1 on target cells. Importantly, FP sEVs significantly inhibited T cell activation and proliferation in vitro and a heart allograft model. Furthermore, FP sEVs encapsulated with low-dose FK506 (FP sEVs@FK506) exerted stronger effects on inhibiting T cell proliferation, reducing CD8+ T cell density and cytokine production in the spleens and heart grafts, inducing regulatory T cells in lymph nodes, and extending graft survival. Taken together, dual-targeting sEVs have the potential to boost the immune inhibitory signalings in synergy and slow down transplant rejection.


title: OsciDrop: A versatile on-demand droplet generator

abstract: Droplet microfluidics is a powerful tool in many biological and clinical applications. Microfluidic chips, such as flow-focusing droplet generators, have been extensively used to high-throughput encapsulate reactions with single-cell and single-molecular resolutions. However, microfabrication is expensive and precision-demanding, preventing it from widespread use in biomedical laboratories and clinical facilities. Herein, we present a versatile chip-free droplet generator, OsciDrop, for generating size-tunable droplets on demand, with high uniformity. OsciDrop segments the fluid flowing out of the orifice of a micropipette tip into droplets by oscillating the tip under the surface of a continuous oil phase. We investigated the factors influencing droplet generation by examining several control parameters. Results show that flow rate, oscillating amplitude, and frequency are key parameters to generate monodisperse droplets on demand. And OsciDrop is able to generate droplets in a flexible and repeatable manner. Importantly, using an optimal asymmetrical oscillation waveform, OsciDrop can controllably generate monodisperse droplets spanning a wide volume range (200 pL - 2 L). To demonstrate the ability of OsciDrop for chip-free droplet assays, a digital loop-mediated isothermal amplification (dLAMP) was performed to absolutely quantify African swine fever virus (ASFV). The OsciDrop method opens up a feasible and versatile avenue to perform droplet-based assays, exhibiting full accessibility for chip-free droplet microfluidics.


title: Enhancing Sampling of Water Rearrangements on Ligand Binding: A Comparison of Techniques

abstract: 1Water often plays a key role in protein structure, molecular recognition, and mediating protein-ligand interactions. Thus, free energy calculations must adequately sample water motions, which often proves challenging in typical MD simulation timescales. Thus, the accuracy of methods relying on MD simulations ends up limited by slow water sampling. Particularly, as a ligand is removed or modified, bulk water may not have time to fill or rearrange in the binding site. In this work, we focus on several molecular dynamics (MD) simulation-based methods attempting to help rehydrate buried water sites: BLUES, using nonequilibrium candidate Monte Carlo (NCMC); grand, using grand canonical Monte Carlo (GCMC); and normal MD. We assess the accuracy and efficiency of these methods in rehydrating target water sites. We selected a range of systems with varying numbers of waters in the binding site, as well as those where water occupancy is coupled to the identity or binding mode of the ligand. We analyzed rehydration of buried water sites in binding pockets using both clustering of trajectories and direct analysis of electron density maps. Our results suggest both BLUES and grand enhance water sampling relative to normal MD and grand is more robust than BLUES, but also that water sampling remains a major challenge for all of the methods tested. The lessons we learned for these methods and systems are discussed.


title: Enhancing Sampling of Water Rehydration on Ligand Binding: A Comparison of Techniques


title: Modeling progression of single cell populations through the cell cycle as a sequence of switches

abstract: AO_SCPLOWBSTRACTC_SCPLOWCell cycle is the most fundamental biological process underlying the existence and propagation of life in time and space. It has been an object for mathematical modeling for long, with several alternative mechanistic modeling principles suggested, describing in more or less details the known molecular mechanisms. Recently, cell cycle has been investigated at single cell level in snapshots of unsynchronized cell populations, exploiting the new methods for transcriptomic and proteomic molecular profiling. This raises a need for simplified semi-phenomenological cell cycle models, in order to formalize the processes underlying the cell cycle, at a higher abstracted level. Here we suggest a modeling framework, recapitulating the most important properties of the cell cycle as a limit trajectory of a dynamical process characterized by several internal states with switches between them. In the simplest form, this leads to a limit cycle trajectory, composed by linear segments in logarithmic coordinates describing some extensive (depending on system size) cell properties. We prove a theorem connecting the effective embedding dimensionality of the cell cycle trajectory with the number of its linear segments. We also develop a simplified kinetic model with piecewise-constant kinetic rates describing the dynamics of lumps of genes involved in S-phase and G2/M phases. We show how the developed cell cycle models can be applied to analyze the available single cell datasets and simulate certain properties of the observed cell cycle trajectories. Based on our modeling, we can predict with good accuracy the cell line doubling time from the length of cell cycle trajectory.


title: Sensitivity to Error During Visuomotor Adaptation is Similarly Modulated by Abrupt, Gradual and Random Perturbation Schedules

abstract:
It has been suggested that sensorimotor adaptation involves at least two processes (i.e., fast and slow) that differ in retention and error sensitivity. Previous work has shown that repeated exposure to an abrupt force field perturbation results in greater error sensitivity for both the fast and slow processes. While this implies that the faster relearning is associated with increased error sensitivity, it remains unclear what aspects of prior experience modulate error sensitivity. In the present study, we manipulated the initial training using different perturbation schedules, thought to differentially affect fast and slow learning processes based on error magnitude, and then observed what effect prior learning had on subsequent adaptation. During initial training of a visuomotor rotation task, we exposed three groups of participants to either an abrupt, a gradual, or a random perturbation schedule. During a testing session, all three groups were subsequently exposed to an abrupt perturbation schedule. Comparing the two sessions of the control group who experienced repetition of the same perturbation, we found an increased error sensitivity for both processes. We found that the error sensitivity was increased for both the fast and slow processes, with no reliable changes in the retention, for both the gradual and structural learning groups when compared to the first session of the control group. We discuss the findings in the context of how fast and slow learning processes respond to a history of errors.

New & NoteworthyWe investigated what aspects of prior experience modulate error sensitivity, within the framework of a two-state model of short-term sensorimotor adaptation. We manipulated initial training on a visuomotor adaptation reaching task using specific perturbation schedules that are thought to differentially affect fast and slow learning processes, and we tested what effect these had on subsequent adaptation. We found that sensitivity to adaptation error was similarly modulated by abrupt, gradual, and random perturbation schedules.


title: Mouse models of immune dysfunction: Their neuroanatomical differences reflect their anxiety-behavioural phenotype

abstract: Extensive evidence supports the role of the immune system in modulating brain function and behaviour. However, past studies have revealed striking heterogeneity in behavioural phenotypes produced from immune system dysfunction. Using magnetic resonance imaging (MRI), we studied the neuroanatomical differences among 11 distinct genetically-modified mouse lines (n=371), each deficient in a different element of the immune system. We found a significant and heterogeneous effect of immune dysfunction on the brains of both male and female mice. However, by imaging the whole brain and using bayesian hierarchical modelling, we were able to identify patterns within the heterogeneous phenotype. Certain structures -- such as the corpus callosum, midbrain, and thalamus -- were more likely to be affected by immune dysfunction. A notable brain-behaviour relationship was identified with neuroanatomy endophenotypes across mouse models clustering according to anxiety-like behaviour phenotypes reported in literature, such as altered volume in brains regions associated with promoting fear response (e.g., the lateral septum and cerebellum). Interestingly, genes with preferential spatial expression in the most commonly affected regions are also associated with multiple sclerosis and other immune-mediated diseases. In total, our data suggest that the immune system modulates anxiety behaviour through well-established brain networks.


title: Iron-dependent mutualism between Chlorella sorokiniana and Ralstonia pickettii forms the basis for a sustainable bioremediation system

abstract: Mutualism between microalgae and bacteria is ubiquitous, but remains underexplored as a basis for biodegradation of anthropogenic pollutants. In industrial systems, poor iron uptake by microalgae limits growth, bioprocessing efficacy, and bioremediation potential. Iron supplementation is costly and ineffective because iron remains insoluble in aqueous medium and biologically unavailable. In aquatic environments, microalgae develop an association with bacteria that solubilize iron by production of siderophore, which increases the bioavailability of iron as a public good. Algae, in exchange, provides dissolved organic matter to bacteria to sustain such interkingdom associations. Therefore, using a case study of azo dye degradation, we combine environmental isolations and synthetic ecology as a workflow, establishing a microbial community to degrade industrially relevant Acid Black 1 dye. We create a mutualism between previously non-associated chlorophyte alga Chlorella sorokiniana and siderophore-producing bacterium Ralstonia pickettii, based on the eco-evolutionary principle of exchange of iron and carbon. This siderophore-mediated increased iron bioavailability increases reductive iron uptake, growth rate, and azoreductase-mediated dye degradation of microalga. In exchange, C. sorokiniana produces galactose, glucose, and mannose as major extracellular monosaccharides, supporting bacterial growth. We propose a mechanism whereby extracellular ferrireductase is crucial for azoreductase-mediated dye degradation in microalgae. Our work demonstrates that bioavailability of iron, which is often overlooked in industrial bio-designs, governs microalgal growth and enzymatic processes. Our results suggest that algal-bacterial consortia based on the active association are a self-sustainable mechanism to overcome existing challenges of micronutrient availability in bioremediation systems and their industrial translation.


title: Accurate prediction of protein structures and interactions using a 3-track network

abstract:
DeepMind presented remarkably accurate protein structure predictions at the CASP14 conference. We explored network architectures incorporating related ideas and obtained the best performance with a 3-track network in which information at the 1D sequence level, the 2D distance map level, and the 3D coordinate level is successively transformed and integrated. The 3-track network produces structure predictions with accuracies approaching those of DeepMind in CASP14, enables rapid solution of challenging X-ray crystallography and cryo-EM structure modeling problems, and provides insights into the functions of proteins of currently unknown structure. The network also enables rapid generation of accurate models of protein-protein complexes from sequence information alone, short circuiting traditional approaches which require modeling of individual subunits followed by docking. We make the method available to the scientific community to speed biological research.

One-Sentence SummaryAccurate protein structure modeling enables rapid solution of structure determination problems and provides insights into biological function.

category: bioinformatics


title: The TR locus annotation and characteristics of Rhinolophus ferrumequinum

abstract: T cell antigen receptors (TCRs) in vertebrate could be divided into {beta} or {gamma}{delta}, which are encoded by TRA/D, TRG and TRB loci respectively. TCRs play a central role in mammal cellular immunity, which are functionally produced by rearrangement of V gene, D gene, J gene and C gene in the loci. Bat is the only mammal with flying ability, and is considered as the main host of zoonotic virus, which occupies an important position in the field of public health. At present, little is known about the composition of bat TRs gene. According to the whole genome sequencing results of the Rhinolophus ferrumequinum, and referring to the TR/IG annotation rules formulated by IMGT. We make a complete annotation on the TRA/D, TRG and TRB loci of the Rhinolophus ferrumequinum. A total of 128 V segments, 3 D segments, 85 J segments and 6 C segments were annotated, in addition to compared with the known mammalian, the characteristics of the TRs locus and germline genes of the Rhinolophus ferrumequinum were analyzed.

category: molecular biology


title: Quantification of microRNA editing using two-tailed RT-qPCR for improved biomarker discovery

abstract:
Even though microRNAs have been viewed as promising biomarkers for years, their clinical implementation is still lagging far behind. This is in part due to the lack of RT-qPCR technologies that can differentiate between microRNA isoforms. For example, A-to-I editing of microRNAs through adenosine deaminase acting on RNA (ADAR) enzymes can affect their expression levels and functional roles, but editing isoform-specific assays are not commercially available. Here, we describe RT-qPCR assays that are specific for editing isoforms, using microRNA-379 (miR-379) as a model. The assays are based on two-tailed RT-qPCR, and we show them to be compatible both with SYBR Green and hydrolysis-based chemistries, as well as with both qPCR and digital PCR. The assays could readily detect different miR-379 editing isoforms in various human tissues. We found that the miR-379 editing frequency was higher in prostate cancer samples compared to benign prostatic hyperplasia samples. Furthermore, decreased expression of unedited miR-379, but not edited miR-379, was associated with treatment resistance, metastasis and shorter overall survival. Taken together, this study presents the first RT-qPCR assays that were demonstrated to distinguish A-to-I-edited microRNAs, and shows that they can be useful in the identification of biomarkers that previously have been masked by other isoforms.

Graphical abstract

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title: The Fusarium graminearum transporters Abc1 and Abc6 are important for xenobiotic resistance, trichothecene accumulation, and virulence to wheat.

abstract: The plant pathogenic fungus Fusarium graminearum is the causal agent of Fusarium Head Blight (FHB) disease on small grain cereals. F. graminearum produces trichothecene mycotoxins such as deoxynivalenol (DON) that are required for full virulence. DON must be exported outside the cell to cause FHB disease, a process that may require the involvement of membrane-bound transporters. In this study we how the deletion of membrane-bound transporters results in reduced DON accumulation as well as reduced FHB symptoms on wheat. Deletion of the ATP-Binding Cassette (ABC) transporter Abc1 results in the most severe reduction in DON accumulation and virulence. Deletion of another ABC transporter, Abc6, also reduces FHB symptoms to a lesser degree. Combining deletions fails to reduce DON accumulation or virulence in an additive fashion, even when including an{Delta} abc1 deletion. Heterologous expression of F. graminearum transporters in a DON-sensitive strain of yeast confirms Abc1 as a major DON resistance mechanism. Yeast expression further indicates that multiple transporters, including Abc1 play an important role in resistance to the wheat phytoalexin 2-benzoxazolinone (BOA) and other xenobiotics. Thus, Abc1 may contribute to wheat virulence both by allowing export of DON and by providing resistance to the wheat phytoalexin BOA. This research provides useful information which may aid in designing novel management techniques of FHB or other destructive plant diseases.


title: Effect of Substrate Stiffness on Human Intestinal Enteroids Infectivity by Enteroaggregative Escherichia coli

abstract:
Human intestinal enteroids (HIE) models have contributed significantly to our understanding of diarrheal diseases and other intestinal infections, but their routine culture conditions fail to mimic the mechanical environment of the native intestinal wall. Because the mechanical characteristics of the intestine significantly alter how pathogens interact with the intestinal epithelium, we used different concentrations of polyethylene glycol (PEG) to generate soft ([~]2 kPa), medium ([~]10 kPa), and stiff ([~]100 kPa) hydrogel biomaterial scaffolds. The height of HIEs cultured in monolayers atop these hydrogels was 18 {micro}m whereas HIEs grown on rigid tissue culture surfaces (with stiffness in the GPa range) were 10 {micro}m. Substrate stiffness also influenced the amount of enteroaggregative E. coli (EAEC strain 042) adhered to the HIEs. We quantified a striking difference in adherence pattern; on the medium and soft gels, the bacteria formed clusters of >100 and even >1000 on both duodenal and jejunal HIEs (such as would be found in biofilms), but did not on glass slides and stiff hydrogels. All hydrogel cultured HIEs showed significant enrichment for gene and signaling pathways related to epithelial differentiation, cell junctions and adhesions, extracellular matrix, mucins, and cell signaling compared to the HIEs cultured on rigid tissue culture surfaces. Collectively, these results indicate that the HIE monolayers cultured on the hydrogels are primed for a robust engagement with their mechanical environment, and that the soft hydrogels promote the formation of larger EAEC aggregates, likely through an indirect differential effect on mucus.

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title: RNA-Seq data analysis for Planarian with tensor decomposition-based unsupervised feature extraction

abstract: RNA-Seq data analysis of non-model organisms is often difficult because of the lack of a well-annotated genome. In model organisms, after short reads are mapped to the genome, it is possible to focus on the analysis of regions well-annotated regions. However, in non-model organisms, contigs can be generated by de novo assembling. This can result in a large number of transcripts, making it difficult to easily remove redundancy. A large number of transcripts can also lead to difficulty in the recognition of differentially expressed transcripts (DETs) between more than two experimental conditions, because P-values must be corrected by considering multiple comparison corrections whose effect is enhanced as the number of transcripts increases. Heavily corrected P-values often fail to take sufficiently small P-values as significant. In this study, we applied a recently proposed tensor decomposition (TD)-based unsupervised feature extraction (FE) to the RNA-seq data obtained for a non-model organism, Planarian; we successfully obtained a limited number of transcripts whose expression was altered between normal and defective samples as well as during time development. TD-based unsupervised FE is expected to be an effective tool that can identify a limited number of DETs, even when a poorly annotated genome is available.


title: Discovery and mechanism of action of small molecule inhibitors of ceramidases

abstract:
Sphingolipid metabolism is tightly controlled by enzymes to regulate essential processes such as energy utilisation and cell proliferation. The central metabolite is ceramide, a pro-apoptotic lipid catabolized by ceramidase enzymes to ultimately produce pro-proliferative sphingosine-1-phosphate. Human ceramidases can be soluble proteins (acid and neutral ceramidase) or integral membrane proteins (alkaline ceramidases). Increasing ceramide levels to increase apoptosis has shown efficacy as a cancer treatment using small molecules inhibiting a soluble ceramidase. Due to the transmembrane nature of alkaline ceramidases, no specific small molecule inhibitors have been reported. Here, we report novel fluorescent substrates (FRETceramides) of ceramidases that can be used to monitor enzyme activity in real-time. We use FRETceramides to discover the first drug-like inhibitors of alkaline ceramidase 3 (ACER3) which are active in cell-based assays. Biophysical characterization of enzyme:inhibitor interactions reveal a new paradigm for inhibition of lipid metabolising enzymes with non-lipidic small molecules.

Table of contents summaryUse of synthetic fluorescent ceramide molecules allows the discovery of the first selective drug-like small molecule inhibitors for alkaline ceramidase 3, an intra-membrane enzyme involved in sphingolipid metabolism in health and disease.