title: Optimised multiplex amplicon sequencing for mutation identification using the MinION nanopore sequencer

abstract:
ObjectiveRapid, cost-effective identification of genetic variants in small candiate genomic regions remains a challenge, particularly for less well equipped or lower throughput laboratories. Application of Oxford Nanopore Technologies MinION sequencer has the potential to fulfil this requirement. We have developed a multiplexing assay which pools PCR amplicons for MinION sequencing to enable sequencing of multiple templates from multiple individuals which could be applied to gene-targeted diagnostics.

MethodsA combined strategy of barcoding and sample pooling was developed for simultaneous multiplex MinION sequencing of 100 PCR amplicons, spanning 30 loci in DNA isolated from 82 neurodevelopmental cases and family members. The target regions were chosen for further interegation because a potentially disease-causative variants had been identified in affected individuals by Illumina exome sequencing. The pooled MinION sequences were deconvoluted by aligning to custom references using the guppy aligner software.

ResultsOur multiplexing approach produced interpretable and expected sequence from 29 of the 30 targeted genetic loci. The sequence variant which was not correctly resolved in the MinION sequence was adjacent to a five nucleotide homopolymer. It is already known that homopolymers present a resolution problem with the MinION approach. Interstingly despite equimolar quantities of PCR amplicon pooled for sequencing, significant variation in the depth of coverage (139x - 21,499x; mean = 9,050, std err = 538.21) was observed. We observed independent relationships between depth of coverage and target length, and depth of coverage and GC content. These relationships demonstrate biases of the MinION sequencer for longer templates and those with lower GC content.

ConclusionWe demonstrate an efficient approach for variant discovery or confirmation from short DNA templates using the MinION sequencing device. With less than 140x depth of coverage required for accurate genotyping, the methodology described here allows for rapid highly multiplexed targeted sequencing of large numbers of samples in a minimally equipped laboratory.


title: SCONCE: A method for profiling Copy Number Alterations in Cancer Evolution using Single Cell Whole Genome Sequencing

abstract: Copy number alterations are a significant driver in cancer growth and development, but remain poorly characterized on the single cell level. Although genome evolution in cancer cells is Markovian through evolutionary time, copy number alterations are not Markovian along the genome. However, existing methods call copy number profiles with Hidden Markov Models or change point detection algorithms based on changes in observed read depth, corrected by genome content, and do not account for the stochastic evolutionary process. We present a theoretical framework to use tumor evolutionary history to accurately call copy number alterations in a principled manner. In order to model the tumor evolutionary process and account for technical noise from low coverage single cell whole genome sequencing data, we developed SCONCE, a method based on a Hidden Markov Model to analyze read depth data from tumor cells using matched normal cells as negative controls. Using a combination of public datasets and simulations, we show SCONCE accurately decodes copy number profiles, with broader implications for understanding tumor evolution. SCONCE is implemented in C++11 and is freely available from https://github.com/NielsenBerkeleyLab/sconce.

category: bioinformatics


title: SIMPATI: patient classifier identifies signature pathways as patient similarity networks for the disease prediction

abstract:
BACKGROUNDPathway-based patient classification is a supervised learning task which implies a model learning pathways as features to predict the classes of patients. The counterpart of enrichment tools for the pathway analysis are fundamental methods for clinicians and biomedical scientists. They allow to find signature cellular functions which help to define and annotate a disease phenotype. They provide results which lead human experts to manually classify patients. It is a paradox that pathwaybased classifiers which natively resolve this objective are not strongly developed. They could simulate the human way of thinking, decipher hidden multivariate relationships between the deregulated pathways and the disease phenotype, and provide more information than a probability value. Instead, there are currently only two classifiers of such kind, they require a nontrivial hyperparameter tuning, are difficult to interpret and lack in providing new insights. There is the need of new classifiers which can provide novel perspectives about pathways, be easy to apply with different biological omics and produce new data enabling a further analysis of the patients.

RESULTSWe propose Simpati, an innovative and interpretable patient classifier based on pathway-specific patient similarity networks. The first classifier to adopt ad-hoc novel algorithms for such graph type. It standardizes the biological high-throughput dataset of patients profiles with a propagation algorithm that considers the interconnected nature of the cells molecules for inferring a new activity score. This allows Simpati to classify with dense, sparse, and non-homogenous omic data. Simpati organizes patients molecules in pathways represented by patient similarity networks for being interpretable, handling missing data and preserving the patient privacy. A network represents patients as nodes and a novel similarity measure determines how much every pair act co-ordinately in a pathway. Simpati detects signature biological processes based on how much the topological properties of the related networks separate the patient classes. In this step, it includes a new cohesive subgroup detection algorithm to handle patients not showing the same pathway activity as the other class members. An unknown patient is then classified by a unique recommender system which considers how much is similar to known patients and distant from being an outlier. Simpati outperforms previously published classifiers on five cancer datasets described with two biological omics, classifies well with sparse data, identifies more relevant pathways associated to the patients disease than the competitors and has the lowest computational requirements.

CONCLUSIONSimpati can serve as generic-purpose pathway-based classifier of patient classes. It provides signature pathways to unveil the altered biological mechanisms of a disease phenotype and to classify patients according to the learnt pathway-specific similarities. The signature condition and patient prediction can be deciphered considering the patient similarity networks which must reveal the members of a patient class more cohesive and similar than the non-members. Simpati divides the pathways in up and downinvolved. Upinvolved when the signaling cascades generated by the altered molecules of the disease patients impact stronger the pathway than the ones of the control class. We provide an R implementation, a graphical user interface and a visualization function for the patient similarity networks. The software is available at: https://github.com/LucaGiudice/Simpati


title: Simpati: patient classifier identifies signature pathways based on similarity networks for the disease prediction


title: PharmacoDB 2.0 : Improving scalability and transparency of in vitro pharmacogenomics analysis

abstract:
Cancer pharmacogenomics studies provide valuable insights into disease progression and associations between genomic features and drug response. PharmacoDB integrates multiple cancer pharmacogenomics datasets profiling approved and investigational drugs across cell lines from diverse tissue types. The web-application enables users to efficiently navigate across datasets, view and compare drug dose-response data for a specific drug-cell line pair. In the new version of PharmacoDB (version 2.0, https://pharmacodb.ca/), we present: (i) new datasets such as NCI-60, the Profiling Relative Inhibition Simultaneously in Mixtures (PRISM) dataset, as well as updated data from the Genomics of Drug Sensitivity in Cancer (GDSC) and the Genentech Cell Line Screening Initiative (gCSI); (ii) implementation of FAIR data pipelines using ORCESTRA and PharmacoDI; (iii) enhancements to drug response analysis such as tissue distribution of dose-response metrics and biomarker analysis; (iv) improved connectivity to drug and cell line databases in the community. The web interface has been rewritten using a modern technology stack to ensure scalability and standardization to accommodate growing pharmacogenomics datasets. PharmacoDB 2.0 is a valuable tool for mining pharmacogenomics datasets, comparing and assessing drug response phenotypes of cancer models.

HIGHLIGHTSO_LIPharmacoDB 2.0 includes new and updated large pharmacogenomic datasets. The data processing for PharmacoDB is made fully reproducible through the use of the ORCESTRA platform and automated data ingestion pipelines
C_LIO_LIThe new release contains enriched annotations for drugs and cell lines via connectivity to external databases, as well as new analytical methods for tissue-specific and pan-cancer biomarker discovery
C_LIO_LIThe new version of PharmacoDB incorporates a scalable and reproducible framework that can accelerate the implementation of analytical pipelines including machine learning/AI for biomarker discovery in the future
C_LI


title: Structures of plasmepsin X from P. falciparum reveal a novel inactivation mechanism of the zymogen and molecular basis for binding of inhibitors in mature enzyme

abstract: Plasmodium falciparum plasmepsin X (PfPMX), involved in the invasion and egress of this deadliest malarial parasite, is essential for its survival and hence considered as an important drug target. We report the first crystal structure of PfPMX zymogen containing a novel fold of its prosegment. A unique twisted loop from the prosegment and arginine 244 from the mature enzyme are involved in zymogen inactivation; such mechanism, not previously reported, might be common for apicomplexan proteases similar to PfPMX. The maturation of PfPMX zymogen occurs through cleavage of its prosegment at multiple sites. Our data provide thorough insights into the mode of binding of a substrate and a potent inhibitor 49c to PfPMX. We present molecular details of inactivation, maturation, and inhibition of PfPMX that should aid in the development of potent inhibitors against pepsin-like aspartic proteases from apicomplexan parasites.

category: biophysics


title: Hyperconcentrated Mucus Unifies Submucosal Gland and Superficial Airway Dysfunction in Cystic Fibrosis

abstract: Cystic fibrosis (CF) is characterized by abnormal transepithelial ion transport. However, a description of CF lung disease pathophysiology unifying superficial epithelial and submucosal gland (SMG) dysfunctions has remained elusive. We hypothesized that biophysical abnormalities associated with CF mucus hyperconcentration provide a unifying mechanism. Studies of the anion secretion-inhibited pig airway CF model revealed elevated SMG mucus concentrations, osmotic pressures, and SMG mucus accumulation. Human airway studies revealed hyperconcentrated CF SMG mucus with raised osmotic pressures and cohesive forces predicted to limit SMG mucus secretion/release. Utilizing proline-rich protein 4 (PRR4) as a biomarker of SMG secretion, proteomics analyses of CF sputum revealed markedly lower PRR4 levels compared to healthy and bronchiectasis controls, consistent with a failure of CF SMGs to secrete mucus onto airway surfaces. Raised mucus osmotic/cohesive forces, reflecting mucus hyperconcentration, provide a unifying mechanism that describes disease-initiating mucus accumulation on airway surfaces and within SMGs of the CF lung.

category: pathology


title: Inhibition of Bruton's tyrosine kinase activity attenuates trauma-induced multiple organ dysfunction in rats

abstract:
ObjectiveThe aim of this study was to investigate (a) the potential of the Brutons tyrosine kinase (BTK) inhibitors (BTKi) acalabrutinib and fenebrutinib to reduce multiple organ dysfunction syndrome (MODS) in acute and chronic hemorrhagic shock (HS) rat models and (b) whether treatment with either acalabrutinib or fenebrutinib attenuates BTK, NF-{kappa}B and NLRP3 activation in HS.

BackgroundThe MODS caused by an excessive systemic inflammatory response following trauma is associated with a high morbidity and mortality. The protein BTK is known to play a role in the activation of the NLRP3 inflammasome, which is a key component of the innate inflammatory response. However, its role in trauma-hemorrhage is unknown.

MethodsAcute and chronic HS rat models were performed to determine the influence of acalabrutinib or fenebrutinib on MODS. The activation of BTK, NF-{kappa}B and NLRP3 pathways were analyzed by western blot in the kidney.

ResultsWe demonstrated that (a) HS caused organ injury and/or dysfunction and hypotension (post resuscitation) in rats, while (b) treatment of HS-rats with either acalabrutinib or fenebrutinib attenuated the organ injury and dysfunction in acute and chronic HS models and (c) reduced the activation of BTK, NF-{kappa}B and NLRP3 pathways in the kidney.

ConclusionOur results point to a role of BTK in the pathophysiology of organ injury and dysfunction caused by trauma/hemorrhage and indicate that BTK inhibitors may be repurposed as a potential therapeutic approach for MODS after trauma and/or hemorrhage.

MINI-ABSTRACTThis study evaluated the role of Brutons tyrosine kinase (BTK) in trauma/hemorrhage. Patients with trauma had elevated gene expression of BTK. The BTK inhibitors acalabrutinib (irreversible) and fenebrutinib (reversible) attenuated the trauma-induced multiple organ dysfunction in rats with hemorrhagic shock, indicating that BTK could be a potential therapeutic target.

category: pharmacology and toxicology


title: Silencing alanine transaminase 2 in diabetic liver attenuates hyperglycemia by reducing gluconeogenesis from amino acids

abstract: Hepatic gluconeogenesis from amino acids contributes significantly to diabetic hyperglycemia, but the molecular mechanisms involved are incompletely understood. Alanine transaminases (ALT1 and ALT2) catalyze the interconversion of alanine and pyruvate, which is required for gluconeogenesis from alanine. We found that ALT2 was overexpressed in liver of diet-induced obese and db/db mice and that the expression of the gene encoding ALT2 (GPT2) was downregulated following bariatric surgery in people with obesity. The increased hepatic expression of Gpt2 in db/db liver was mediated by activating transcription factor 4; an endoplasmic reticulum stress-activated transcription factor. Hepatocyte-specific knockout of Gpt2 attenuated incorporation of 13C-alanine into newly synthesized glucose by hepatocytes. In vivo Gpt2 knockdown or knockout in liver had no effect on glucose concentrations in lean mice, but Gpt2 suppression alleviated hyperglycemia in db/db mice. These data suggest that ALT2 plays a significant role in hepatic gluconeogenesis from amino acids in diabetes.

category: physiology


title: Perilacunar bone tissue exhibits sub-micrometer modulus gradation which depends on the recency of osteocyte bone formation

abstract:
Osteocytes are capable of resorbing and replacing bone local to the lacunar-canalicular system (LCS remodeling). However, the impacts of these processes on perilacunar bone quality are not understood. It is well established that aging is associated with reduced whole-bone fracture resistance, reduced osteocyte viability, and truncated LCS geometries, but it remains unclear if aging changes perilacunar bone quality. In this study, we employed atomic force microscopy (AFM) to quantify sub-micrometer gradations from 2D maps surrounding osteocyte lacunae in young (5 mo) and aged (22 mo) female mice. AFM-mapped lacunae were also imaged with confocal laser scanning microscopy to determine which osteocytes had recently deposited bone as determined by the presence of fluorochrome labels. These assays allowed us to quantify gradations in nanoscale mechanical properties of bone-forming/non-bone-forming osteocytes in young and aged mice. This study reports for the first time that there are sub-micrometer gradations in modulus surrounding lacunae and that these gradations are dependent upon recent osteocyte bone formation. Perilacunar bone adjacent to bone-forming osteocytes demonstrated lower peak and bulk modulus values when compared to bone near non-bone-forming osteocytes from the same mouse. Bone-forming osteocytes also showed increased perilacunar modulus variability. Age reduced lacunar size but did not significant effect modulus gradation or variability. In general, lacunar morphology was not a strong predictor of modulus gradation patterns. These findings support the idea that lacunar-canalicular remodeling activity changes the material properties of surrounding bone tissue on a sub-micrometer scale. Therefore, conditions that affect osteocyte health have the potential to impact bone quality.

GRAPHICAL ABSTRACT

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title: Glutathione contributes to plant defense against parasitic cyst nematodes

abstract: Cyst nematodes (CNs) are an important group of root-infecting sedentary endoparasites that severely damage many crop plants worldwide. An infective CN juvenile enters the hosts roots and migrates towards the vascular cylinder, where it induces the formation of syncytial feeding cells, which nourish the CN throughout its parasitic stages. Here, we examined the role of glutathione (L-{gamma}-glutamyl-L-cysteinylglycine, GSH) in Arabidopsis thaliana upon infection with the CN Heterodera schachtii. Arabidopsis lines with mutations pad2, cad2, or zir1 in the glutamate-cysteine ligase (GSH1) gene, which encodes the first enzyme in the glutathione biosynthetic pathway, displayed enhanced CN susceptibility, but susceptibility was reduced for rax1, another GSH1 allele. Biochemical analysis revealed differentially altered thiol levels in these mutants that was independent of nematode infection. All GSH-deficient mutants exhibited impaired activation of defense marker genes as well as genes for biosynthesis of the antimicrobial compound camalexin early in infection. Further analysis revealed a link between glutathione-mediated plant susceptibility to CN infection and the production of camalexin upon nematode infection. These results suggest that GSH levels affects plant susceptibility to CN by fine-tuning the balance between the cellular redox environment and the production of compounds related to defense against infection.

category: plant biology


title: The ectomycorrhizal basidiomycete Laccaria bicolor releases a GH28 polygalacturonase that plays a key role in symbiosis establishment

abstract:
O_LIIn ectomycorrhiza, root penetration and colonization of the intercellular space by symbiotic hyphae is thought to rely on the mechanical force that results from hyphal tip growth, enhanced by the activity of secreted cell-wall-degrading enzymes.
C_LIO_LIHere, we characterize the biochemical properties of the symbiosis-induced polygalacturonase LbGH28A from the ectomycorrhizal fungus Laccaria bicolor. The transcriptional regulation of LbGH28A was measured by qPCR. The biological relevance of LbGH28A was confirmed by generating RNAi-silenced LbGH28A mutants. We localized the LbGH28A protein by immunofluorescence confocal and immunogold cytochemical microscopy in poplar ectomycorrhizal roots.
C_LIO_LIqPCR confirmed the induced expression of LbGH28A during ectomycorrhiza formation. L. bicolor RNAi mutants have a lower ability to establish ectomycorrhiza confirming the key role of this enzyme in symbiosis. The purified recombinant LbGH28A has its highest activity towards pectin and polygalacturonic acid. In situ localization of LbGH28A indicates that this endopolygalacturonase is located in both fungal and plant cell walls at the symbiotic hyphal front.
C_LIO_LIThe present findings suggest that the symbiosis-induced pectinase LbGH28A is involved in the Hartig net formation and is an important determinant for successful symbiotic colonization.
C_LI


title: Dynamics of fibril collagen remodeling by tumor cells using individual cell-based mathematical modeling

abstract: Many solid tumors are characterized by dense extracellular matrix (ECM) composed of various ECM fibril proteins that provide structural support and biological context for the residing cells. The growing tumor cell colonies are capable of remodeling the ECM structure in tumor immediate vicinity to form specific microenvironmental niches. The changes in fibril patterns of the collagen (one of the ECM proteins) surrounding the tumor can be visualized experimentally using both histology and fluorescent imaging. In particular, three diverse tumor associated collagen signatures (TACS) were identified and related to tumor behavior, such as benign growth or invasion. Here we will use an off-lattice hybrid agent-based model (MultiCell-LF) to identify the rules of cell-ECM interactions that guide the development of various patterns of alignment of the ECM fibrils.

category: systems biology


title: Determining cardiac structure by diffuse reflectance of different wavelengths

abstract: Catheter ablation in patients suffering from chronic arrhythmias requires detailed knowledge of the underlying cardiac anatomy; such real-time, high resolution mapping is currently unavailable in a clinical setting. We present here preliminary work towards a novel optical strategy based on diffuse optical reflectance to provide quantitative anatomical measurements of the cardiac structure, including tissue thickness and presence of scar. An in-depth literature search is conducted to collate available experimental data regarding optical parameters in cardiac tissue and scar. Computational simulations of photon movement through cardiac tissue using Monte Carlo modelling are performed, with analysis being focussed on the effects on surface emission profiles of (i) optical parameters; (ii) tissue thickness; (iii) presence of scar. Our results demonstrate (i) sensitivity of the approach to changes in optical parameters within tissue, (ii) difference of results depending on light wavelength. These suggest that this can be used to detect cardiac anatomical structure to a depth of [~] 2 mm, for both thickness of cardiac tissue and presence of scar. This study demonstrates the feasibility of using diffuse optical reflectance to determine cardiac structure, enabling a potential route for high-resolution, real-time structural information to guide catheter ablation and similar surgeries.


title: Multiple molecular events determine stochastic cell fate switching in a eukaryotic bistable system

abstract: Eukaryotic transcriptional networks are often large and contain several levels of feedback regulation. Many of these networks have the ability to generate and maintain several distinct transcriptional states across multiple cell divisions and to switch between them. In certain instances, switching between cell states is stochastic, occurring in a small subset of cells of an isogenic population in a seemingly homogenous environment. Given the scarcity and unpredictability of switching in these cases, investigating the determining molecular events is challenging. White-opaque switching in the fungal species Candida albicans is an example of stably inherited cell states that are determined by a complex transcriptional network and can serve as an experimentally accessible model system to study characteristics important for stochastic cell fate switching in eukaryotes. In standard lab media, genetically identical cells maintain their cellular identity (either "white" or "opaque") through thousands of cell divisions and switching between the states is rare. By isolating populations of white or opaque cells, previous studies have elucidated the many differences between the two stable cell states and identified a set of transcriptional regulators needed for cell type switching. Yet little is known about the molecular events that determine the rare, stochastic switching events that occur in single cells. We use microfluidics combined with fluorescent reporters to directly observe rare switching events between the white and opaque states. We investigate the stochastic nature of switching by beginning with white cells and monitoring the activation of Wor1, a master regulator and marker for the opaque state, in single cells and throughout cell pedigrees. Our results indicate that switching requires two stochastic steps; first an event occurs that predisposes a lineage of cells to switch. In the second step, some but not all, of those predisposed cells rapidly express high levels of Wor1 and commit to the opaque state. To further understand the rapid rise in Wor1, we used a synthetic inducible system in Saccharomyces cerevisiae into which a controllable C. albicans Wor1 and a reporter for its transcriptional control region have been introduced. We document that Wor1 positive autoregulation is highly cooperative (Hill coefficient > 3), leading to rapid activation and producing an "all or none" rather than a graded response. Taken together, our results suggest that reaching a threshold level of a master regulator is sufficient to drive cell type switching in single cells and that an earlier molecular event increases the probability of reaching that threshold in certain small lineages of cells. Quantitative molecular analysis of the white-opaque circuit can serve as a model for the general understanding of complex circuits.


title: Multiple molecular events underlie stochastic switching between two heritable cell states in a eukaryotic system


title: Integrated phylogenomics and fossil data illuminate the evolution of beetles

abstract: With over 380,000 described species and possibly several million more yet unnamed, beetles represent the most biodiverse animal order. Recent phylogenomic studies have arrived at considerably incongruent topologies and widely varying estimates of divergence dates for major beetle clades. Here we use a dataset of 68 single-copy nuclear protein coding genes sampling 129 out of the 194 recognized extant families as well as the first comprehensive set of fully-justified fossil calibrations to recover a refined timescale of beetle evolution. Using phylogenetic methods that counter the effects of compositional and rate heterogeneity we recover a topology congruent with morphological studies, which we use, combined with other recent phylogenomic studies, to propose several formal changes in the classification of Coleoptera: Scirtiformia and Scirtoidea sensu nov., Clambiformia ser. nov. and Clamboidea sensu nov., Rhinorhipiformia ser. nov., Byrrhoidea sensu nov., Dryopoidea stat. res., Nosodendriformia ser. nov., and Staphyliniformia sensu nov., alongside changes below the superfamily level. The heterogeneous former superfamily Cucujoidea is divided into three monophyletic groups: Erotyloidea stat. nov., Nitiduloidea stat. nov., and Cucujoidea sensu nov. Our divergence time analysis recovered an evolutionary timescale congruent with the fossil record: a late Carboniferous origin of Coleoptera, a late Paleozoic origin of all modern beetle suborders, and a Triassic-Jurassic origin of most extant families. While fundamental divergences within beetle phylogeny did not coincide with the hypothesis of a Cretaceous Terrestrial Revolution, many polyphagan superfamilies exhibited increases in richness with Cretaceous flowering plants.

category: zoology


title: Revisiting tolerance to ocean acidification: insights from a new framework combining physiological and molecular tipping points of Pacific oyster

abstract: Studies on the impact of ocean acidification on marine organisms involve exposing organisms to future acidification scenarios as projected for open ocean, which has limited relevance for coastal calcifiers. Characterization of reaction norms across a range of pH and identification of tipping points beyond which detrimental effects are observed has been limited and focus on only a few macro-physiological traits. Here we filled this knowledge gap by developing a framework to analyze the broad macro-physiological and molecular responses over a wide pH range of juvenile Pacific oyster, a model species for which the tolerance threshold to acidification remains unknown. We identify low tipping points for physiological traits at pH 7.3-6.9 that coincide with a major reshuffling in membrane lipids and transcriptome. In contrast, shell parameters exhibit effects with pH drop well before tipping points, likely impacting animal fitness. These findings were made possible by the development of an innovative methodology to synthesize and identify the main patterns of variations in large -omic datasets, fit them to pH and identify molecular tipping-points. We propose the application of our framework broadly to the assessment of effects of global change on other organisms.


title: Novel microRNA-455-3p mouse models to study Alzheimers disease pathogenesis

abstract: MicroRNA-455-3p is one of the highly conserved miRNAs involved in several human diseases but newly explored by our lab in Alzheimers disease (AD). Our past studies unveiled the biomarker and therapeutic potentials of miR-455-3p in AD. Our in vitro study exhibited the protective role of miR-455-3p against AD toxicities in reducing full-length APP and amyloid-{beta} (A{beta}) protein levels, and also reducing defective mitochondrial biogenesis, impaired mitochondrial dynamics and synaptic deficiencies. Next, we sought to determine the essential roles of miR-455-3p in AD using mouse models. Therefore, for the first time we generated both transgenic (TG) and knockout (KO) mouse models of miR-455-3p. We determined the positive and negative effects of miR-455-3p on mice cognitive function, mitochondrial biogenesis, mitochondrial dynamics, mitochondrial number & length, dendritic spine density, synapse numbers and synaptic activity in 12-month-old miR-455-3p TG and KO mice. MiR-455-3p TG mice lived 5 months longer than wild-type (WT) mice, whereas KO mice lived 4 months shorter than their WT counter parts. Morris water maze test showed improved cognitive behavior, spatial learning and memory in miR-455-3p TG mice relative to age-matched WT mice and miR-455-3p KO mice. Further, mitochondrial biogenesis, dynamics and synaptic activities were enhanced in miR-455-3p TG mice, while these were reduced in KO mice. Overall, miR-455-3p TG mice displayed protective effects and miR-455-3p KO mice exhibited deleterious effects in relation to AD pathogenesis. Both mouse models could be ideal research tools to understand the molecular mechanism of miR-455-3p in AD and other human diseases.

category: neuroscience


title: Oscillatory calcium release and sustained store-operated oscillatory calcium signaling prevents differentiation of human oligodendrocyte progenitor cells

abstract:
Endogenous remyelination in demyelinating diseases such as multiple sclerosis is contingent upon the successful differentiation of oligodendrocyte progenitor cells (OPCs). Signaling via the Gq-coupled muscarinic receptor (M1/3R) inhibits human OPC differentiation and impairs endogenous remyelination in experimental models. We hypothesized that calcium release following Gq-coupled receptor (GqR) activation directly regulates human OPC (hOPC) cell fate. In this study, we show that specific GqR agonists activating muscarinic and metabotropic glutamate receptors induce characteristic oscillatory calcium release in hOPCs and that these agonists similarly block hOPC maturation in vitro. Both agonists induce calcium release from endoplasmic reticulum (ER) stores and store operated calcium entry (SOCE) likely via STIM/ORAI-based channels. siRNA mediated knockdown (KD) of obligate calcium sensors STIM1 and STIM2 decreased the magnitude of muscarinic agonist induced oscillatory calcium release and attenuated SOCE in hOPCs. In addition, STIM2 expression was necessary to maintain the frequency of calcium oscillations and STIM2 KD reduced spontaneous OPC differentiation. Furthermore, STIM2 siRNA prevented the effects of muscarinic agonist treatment on OPC differentiation suggesting that SOCE is necessary for the anti-differentiative action of muscarinic receptor-dependent signaling. Finally, using a gain-of-function approach with an optogenetic STIM lentivirus, we demonstrate that independent activation of SOCE was sufficient to significantly block hOPC differentiation and this occurred in a frequency dependent manner while increasing hOPC proliferation. These findings suggest that intracellular calcium oscillations directly regulate hOPC fate and that modulation of calcium oscillation frequency may overcome inhibitory Gq-coupled signaling that impairs myelin repair.

Significance StatementIn this study, Seidman et al. show that SOCE is a common component of ligand-based Gq-coupled signaling in hOPCs and that SOCE alone is sufficient to block hOPC differentiation and drive proliferation. Therefore, SOCE blocks differentiation and pathological SOCE could contribute to myelin disease.


title: Assessing the relationships between neurological and psychiatric diseases with astrocyte subtypes and psychotropic medications

abstract: Astrocytes have many important functions in the brain, but their roles in CNS disorders and their responses to psychotropic medications are still being elucidated. In this study, we used gene enrichment analysis to assess the relationships between different astrocyte subtypes, neurological and psychiatric diseases, and psychotropic medications. We also carried out qPCR analyses and "look-up" studies to further assess the chronic effects of these drugs on astrocyte marker gene expression. Our bioinformatic analysis identified differential gene enrichment of different astrocyte subtypes in CNS disorders. The "common" astrocyte subtype was the most frequently enriched across disorders, but the highest level of enrichment was found in depression, supporting a role for astrocytes in this disorder. We also identified common enrichment of metabolic and signal transduction-related biological processes in astrocyte subtypes and CNS disorders. However, enrichment of different psychotropic medications, including antipsychotics, antidepressants, and mood stabilizers, was limited in astrocyte subtypes. These results were confirmed by "look-up" studies and qPCR analysis, which also reported little effect of common psychotropic medications on astrocyte marker gene expression, suggesting that astrocytes are not a primary target of these medications. Overall, this study provides a unique view of astrocyte subtypes and the effect of medications on astrocytes in disease, which will contribute to our understanding of their role in CNS disorders and offers insights into targeting astrocytes therapeutically.


title: State-dependent linearisation of mixture representations by the olfactory bulb

abstract: Sensory systems are often tasked to analyse complex signals from the environment, separating relevant from irrelevant parts. This process of decomposing signals is challenging when a mixture of signals does not equal the sum of its parts, leading to an unpredictable corruption of signal patterns. In olfaction, nonlinear summation is prevalent at various stages of sensory processing. Here, we investigate how the olfactory system deals with binary mixtures of odours under different brain states, using two-photon imaging of olfactory bulb (OB) output neurons. Unlike previous studies using anaesthetised animals, we found that mixture summation is more linear in the early phase of evoked responses in awake, head-fixed mice performing an odour detection task, due to dampened responses. Despite this, and responses being more variable, decoding analyses indicated that the data from behaving mice was well discriminable. Curiously, the time course of decoding accuracy did not correlate strictly with the linearity of summation. Further, a comparison with naive mice indicated that learning to accurately perform the mixture detection task is not accompanied by more linear mixture summation. Finally, using a simulation, we demonstrate that, while saturating sublinearity tends to degrade the discriminability, the extent of the impairment may depend on other factors, including pattern decorrelation. Altogether, our results demonstrate that the mixture representation in the primary olfactory area is state-dependent, but the analytical perception may not strictly correlate with linearity in summation.


title: State-dependent representation of mixtures by the olfactory bulb


title: A Comparison of Methods to Harmonize Cortical Thickness Measurements Across Scanners and Sites

abstract: Results of neuroimaging datasets aggregated from multiple sites may be biased by site- specific profiles in participants demographic and clinical characteristics, as well as MRI acquisition protocols and scanning platforms. We compared the impact of four different harmonization methods on results obtained from analyses of cortical thickness data: (1) linear mixed-effects model (LME) that models site-specific random intercepts (LMEINT), (2) LME that models both site-specific random intercepts and age-related random slopes (LMEINT+SLP), (3) ComBat, and (4) ComBat with a generalized additive model (ComBat-GAM). Our test case for comparing harmonization methods was cortical thickness data aggregated from 29 sites, which included 1,343 cases with posttraumatic stress disorder (PTSD) (6.2-81.8 years old) and 2,067 trauma-exposed controls without PTSD (6.3-85.2 years old). We found that, compared to the other data harmonization methods, data processed with ComBat-GAM were more sensitive to the detection of significant case-control differences in regional cortical thickness (X2(3) = 34.339, p < 0.001), and case-control differences in age-related cortical thinning (X2(3) = 15.128, p = 0.002). Specifically, ComBat-GAM led to larger effect size estimates of cortical thickness reductions (corrected p-values < 0.001), smaller age-appropriate declines (corrected p-values < 0.001), and lower female to male contrast (corrected p-values < 0.001) in cases compared to controls relative to other harmonization methods. Harmonization with ComBat-GAM also led to greater estimates of age-related declines in cortical thickness (corrected p-values < 0.001) in both cases and controls compared to other harmonization methods. Our results support the use of ComBat-GAM for harmonizing cortical thickness data aggregated from multiple sites and scanners to minimize confounds and increase statistical power.


title: The impact of genetic risk for Alzheimers disease on the structural brain networks of young adults

abstract:
To facilitate pre-symptomatic diagnosis of late-onset Alzheimers disease, non-invasive imaging biomarkers could be combined with genetic risk information. In this work, we investigated the structural brain networks of young adults in relation to polygenic risk for Alzheimers disease, using magnetic resonance imaging (MRI) and genotype data for 562 19-year-old participants from the Avon Longitudinal Study of Parents and Children. Diffusion MRI data were acquired on a 3T scanner, and the data were used to perform whole-brain tractography. The resulting tractograms were used to generate structural brain networks, using the number of streamlines and the diffusion tensor fractional anisotropy as edge weights. This was done for the whole-brain connectome, and for the default mode, limbic and visual subnetworks. The mean clustering coefficient, mean betweenness centrality, characteristic path length, global efficiency and mean nodal strength were calculated for these networks, for each participant. The hubs of the networks were also identified, and the connectivity of the rich-club, feeder and local connections was also calculated. Polygenic risk scores (PRS), estimating the burden of genetic risk carried by an individual, were calculated both at genome-wide level and for nine specific disease pathways. The correlation coefficients were calculated between the PRS and a) the graph theoretical metrics of the structural networks and b) the rich-club, feeder and local connectivity of the whole-brain networks.

In the visual subnetwork, the mean nodal strength was negatively correlated with the genome-wide PRS including the APOE locus (r = -0.19, p = 1.3x10-5), the mean betweenness centrality was positively correlated with the pathway-specific PRS for plasma lipoprotein particle assembly including the APOE locus (r = 0.16, p = 9.2x10-4), and the mean clustering coefficient was negatively correlated with the tau protein binding PRS including the APOE locus (r = 0.16, p = 9.2x10-4). In the default mode network, the mean nodal strength was negatively correlated with the genome-wide PRS, including the APOE locus (r = -0.14, p=1.5x10-3). The rich-club and feeder connectivities were negatively correlated with the genome-wide PRS including the APOE locus (r = -0.16, p = 3.7x10-4; r = -0.15, p 8.8x10-4, respectively). Our results indicate small changes in the brain connectome of young adults at risk of developing Alzheimers disease in later life.


title: Altered synaptic connectivity in an in vitro human model of STXBP1 encephalopathy

abstract: Early infantile developmental and epileptic encephalopathies are devastating conditions, generally of genetic origin, but the pathological mechanisms often remain obscure. A major obstacle in this field of research is the difficulty of studying cortical brain development in humans, in utero. To address this, we established an in vitro assay to study the impact of gene variants on the developing human brain, using living organotypic cultures of the human subplate and neighbouring cortical regions, prepared from ethically sourced, 14-17 post conception week brain tissue (www.hdbr.org). We were able to maintain cultures for several months, during which time, the gross anatomical structures of the cortical plate, subplate and marginal zone persisted, while neurons continued to develop morphologically, and form new synaptic networks. This preparation thus permits the study of genetic manipulations, and their downstream effects upon an intact developing human cortical network. We focused upon STXBP1 haploinsufficiency, which is among the most common genetic causes of developmental and epileptic encephalopathy. This was induced using shRNA interference, leading to impaired synaptic function and a drop in the number of glutamatergic synapses. We thereby provide a critical proof-of-principle for how to study the impact of any gene of interest on the development of the human cortex.


title: Targeting of Cdc42 GTPase in regulatory T cells unleashes anti-tumor T cell immunity

abstract:
Regulatory T (Treg) cells play an important role in maintaining immune tolerance through inhibiting effector T cell function. In the tumor microenvironment, Treg cells are utilized by tumor cells to counteract effector T cell-mediated tumor killing. Targeting Treg cells may thus unleash the anti-tumor activity of effector T cells. While systemic depletion of Treg cells can cause excessive effector T cell responses and subsequent autoimmune diseases, controlled targeting of Treg cells may benefit cancer patients. Here we show that Treg cell-specific heterozygous deletion or pharmacological targeting of Cdc42 GTPase does not affect Treg cell numbers but induces Treg cell plasticity, leading to anti-tumor T cell immunity without detectable autoimmune reactions. Cdc42 targeting potentiates an immune checkpoint blocker anti-PD-1 antibody-mediated T cell response against mouse and human tumors. Mechanistically, Cdc42 targeting induces Treg cell plasticity and unleashes antitumor T cell immunity through carbonic anhydrase I-mediated pH changes. Thus, rational targeting of Cdc42 in Treg cells holds therapeutic promises in cancer immunotherapy.

SignificanceEffector T lymphocytes promote autoimmune diseases but have potential to kill tumor cells. However, cancer cells can evade T cell-mediated killing in part by utilizing regulatory T (Treg) cells to inhibit effector T cell function. Here we show that Treg cell-specific heterozygous deletion of Cdc42 gene that encodes Cdc42 GTPase dampens Treg cell fitness through carbonic anhydrase I-mediated pH changes, leading to anti-tumor T cell immunity. Pharmacological targeting of Cdc42 mimics genetic deletion of Cdc42 in impairing Treg cell fitness and evoking anti-tumor T cell immunity. Importantly, Cdc42 targeting does not appear to cause systemic autoimmunity. Given that current cancer immunotherapies only demonstrate limited clinical efficacies, our findings may open a new avenue for cancer immunotherapy.

category: immunology


title: Structural Mechanics of the Alpha-2-Macroglobulin Transformation

abstract: Alpha-2-Macroglobulin (A2M) is the critical pan-protease inhibitor of the innate immune system. When proteases cleave the A2M bait region, global structural transformation of the A2M tetramer is triggered to entrap the protease. The structural basis behind the cleavage-induced transformation and the protease entrapment remains unclear. Here, we report cryo-EM structures of native- and intermediate-forms of the Xenopus laevis egg A2M homolog (A2Moo or ovomacroglobulin) tetramer at 3.7-4.1 [A] and 6.4 [A] resolution, respectively. In the native A2Moo tetramer, two pairs of dimers arrange into a cross-like configuration with four 60 [A]-wide bait-exposing grooves. Each bait in the native form threads into an aperture formed by three macroglobulin domains (MG2, MG3, MG6). The bait is released from the narrowed aperture in the induced protomer of the intermediate form. We propose that the intact bait region works as a "latch-lock" to block futile A2M transformation until its protease-mediated cleavage.


title: Engineering CAR-NK cells to secrete IL15 sustains their anti-AML functionality, but is associated with systemic toxicities

abstract:
BackgroundThe prognosis of patients with recurrent/refractory acute myelogenous leukemia (AML) remains poor and cell-based immunotherapies hold promise to improve outcomes. NK cells can elicit an anti-leukemic response via a repertoire of activating receptors that bind AML surface ligands. NK cell adoptive transfer is safe but thus far has shown limited anti-AML efficacy. Here, we aimed to overcome this limitation by engineering NK cells to express chimeric antigen receptors (CARs) to boost their anti-AML activity, and interleukin-15 (IL15) to enhance their persistence.

MethodsWe characterized in detail NK cell populations expressing a panel of AML (CD123)-specific CARs and/or IL15 in vitro and in AML xenograft models.

ResultsCARs with 2B4.{zeta} or 4-1BB.{zeta} signaling domains demonstrated greater cell surface expression and endowed NK cells with improved anti-AML activity in vitro. Initial in vivo testing revealed that only 2B4.{zeta} CAR-NK cells had improved anti-AML activity in comparison to untransduced (UTD) and 4-1BB.{zeta} CAR-NK cells. However, the benefit was transient due to limited CAR-NK cell persistence. Transgenic expression of secretory (s)IL15 in 2B4.{zeta} CAR and UTD NK cells improved their effector function in the setting of chronic antigen simulation in vitro. Multiparameter flow analysis after chronic antigen exposure identified the expansion of unique NK cell subsets. 2B4.{zeta}/sIL15 CAR and sIL15 NK cells maintained an overall activated NK cell phenotype. This was confirmed by transcriptomic analysis, which revealed a highly proliferative and activated signature in these NK cell groups. In vivo, 2B4.{zeta}/sIL15 CAR-NK cells had potent anti-AML activity in one model, while 2B4.{zeta}/sIL15 CAR and sIL15 NK cells induced lethal toxicity in a second model.

ConclusionTransgenic expression of CD123-CARs and sIL15 enabled NK cells to function in the setting of chronic antigen exposure but was associated with systemic toxicities. Thus, our study provides the impetus to explore inducible and controllable expression systems to provide cytokine signals to AML-specific CAR-NK cells before embarking on early phase clinical testing.

O_FIG O_LINKSMALLFIG WIDTH=161 HEIGHT=200 SRC="FIGDIR/small/461509v1_ufig1.gif" ALT="Figure 1">
View larger version (42K):
org.highwire.dtl.DTLVardef@4d09eaorg.highwire.dtl.DTLVardef@1c61411org.highwire.dtl.DTLVardef@14f8097org.highwire.dtl.DTLVardef@8375ac_HPS_FORMAT_FIGEXP  M_FIG C_FIG In BriefSecretion of IL15 from anti-CD123.2B4.CD3{zeta} CAR-NK cells enhances their activation and killing potential against Acute Myelogenous Leukemia, but carries the risk of severe systemic toxicity.


title: Identification of Immunodominant T-lymphocyte Epitope Peptides in HPV 1, 2 and 3 L1 Protein for Novel Cutaneous Wart Vaccine

abstract:
BackgroundCommon warts and flat warts are caused by the human papillomavirus (HPV). Peak incidence of wart infection occurs in schoolchildren aged 12-16, where prevalence can be as high as 20%. Traditional treatments aimed at destruction of wart tissue have low clearance rates and high recurrence rates. Occasional reports have even shown warts becoming malignant and progressing into verrucous carcinoma. Current licensed HPV vaccines largely target higher-risk oncogenic HPV types, but do not provide coverage of low-risk types associated with warts. To date, little attention has been given to the development of effective, anti-viral wart treatments.

ObjectiveThis study aims to identify immunodominant T-lymphocyte epitopes from the L1 major capsid protein of HPV 1, 2 and 3, a foundational step in bioengineering a peptide-based vaccine for warts.

MethodsCytotoxic T-cell and helper T-cell epitopes were predicted using an array of immunoinformatic tools against a reference panel of frequently observed MHC-I and MHC-II alleles. Predicted peptides were ranked based on IC50 and IFN-{gamma} Inducer Scores, respectively, and top performing epitopes were synthesized and subjected to in vitro screening by IFN-{gamma} enzyme-linked immunosorbent spot assay (ELISpot). Independent trials were conducted using PBMCs of healthy volunteers. Final chosen peptides were fused with flexible GS linkers in silico to design a novel polypeptide vaccine.

ResultsSeven immunodominant peptides screened from 44 predicted peptides were included in the vaccine design, selected to elicit specific immune responses across MHC class I and class II, and across HPV types. Evaluation of the vaccines properties suggest that the vaccine is stable, non-allergenic, and provides near complete global population coverage (>99%). Solubility prediction and rare codon analysis indicate that the DNA sequence encoding the vaccine is suitable for high level expression in Escherichia coli.

ConclusionsIn sum, this study demonstrates the potential and lays the framework for the development of a peptide-based vaccine against warts.


title: Active Translation Control of CD4 T Cell Activation by Regulatory T Cells

abstract: Increased protein synthesis is a hallmark of lymphocyte activation. Regulatory T cells (Tregs) suppress the activation and subsequent effector functions of CD4 effector T cells (Teff). Molecular mechanisms that enforce suppression on CD4 Teff cell function are unclear. Control of CD4 Teff cell activation by Tregs has largely been defined at the transcriptional level, which does not reflect changes in post-transcriptional control. We found that Tregs suppressed activation-induced global protein synthesis in CD4 Teff cells prior to cell division. We analyzed genome-wide changes in the transcriptome and translatome of activated CD4 Teff cells using two independent approaches. We show that mRNAs encoding for the protein synthesis machinery are regulated at the level of translation in activated Teff cells. Strikingly, Tregs suppressed global protein synthesis of CD4 Teff cells by specifically inhibiting mRNAs of the translation machinery at the level of mTORC1-mediated translation control. Lastly, we found that the RNA helicase eIF4A inhibitor rocaglamide A (RocA) can suppress CD4 Teff activation in vitro to alleviate inflammatory CD4 Teff activation caused by acute Treg depletion in vivo. These data provide evidence that peripheral tolerance is enforced by Tregs, mediated by IL-10, through mRNA translational control in CD4 Teff cells. Therefore, therapeutic targeting of the protein synthesis machinery can mitigate inflammatory responses invoked by Treg loss of function.


title: Immune checkpoint expression on HIV-specific CD4+ T cells and response to their blockade are dependent on lineage and function

abstract:
BackgroundAntigen-specific T cell impairment is observed in chronic infections. CD4+ T cells are diverse in phenotype and function; how their different lineages are impacted by inhibitory immune checkpoints (IC) is unknown.

MethodsWe examined IC expression and function in HIV-specific CD4+ T cells of viremic individuals prior to ART initiation and persons with spontaneous or therapy-induced viral suppression. We investigated IC patterns associated with exhaustion-related transcription factors and chemokine receptors using cytokine-independent activation-induced marker assays. We determined effector functions representative of TFH, TH1 and TH17/TH22 using ultra-sensitive RNA flow cytometric fluorescence in situ hybridization (FISH), and their response to IC blockade.

FindingsThe dysfunction-related transcription factor TOX was elevated in HIV-specific CD4+ T cells of viremic patients, and its expression was associated with lineage differentiation. We observed a hierarchy of PD-1, TIGIT and CD200 expression associated with both infection status and effector profile. In vitro responsiveness to PD-L1 blockade varied with defined CD4+ T cell functions rather than IC expression levels: frequencies of cells with TH1- and TH17/TH22-, but not TFH-related functions, increased. Response to PD-L1 blockade was strongest in viremic participants and reduced after ART initiation.

InterpretationOur data highlight a polarization-specific regulation of IC expression and differing sensitivities of antigen-specific Thelper subsets to PD-1-mediated inhibition. This heterogeneity may direct ICB efficacy on CD4+ T cells in HIV infection.

FundingThis work was supported by the National Institutes of Health, the Canadian Institutes for Health Research, the Canada Foundation for Innovation and the Fonds de Recherche du Quebec-Sante.

Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSCombination antiretroviral therapy (ART) is highly effective in controlling HIV but requires life-long medication due to the latent viral reservoir, and does not restore suppressive immune responses. In particular, there is no generation of effective HIV-specific T cell responses, which are thought to play an important role in controlling HIV in the rare individuals who can spontaneously control the virus. Inhibitory immune checkpoints (IC) such as PD-1 contribute to T cell dysfunction and failure to control viral infections, including HIV, and IC blockade (ICB) represents a potential adjuvant to ART through restoration of T cell functions. While most studies have focused on CD8+ T cells, increasing evidence shows that the remarkable impact of ICB therapy in a subset of cancer patients is enhanced by functional CD4+ T cell help, which can be directly affected by ICB. While effective virus-specific CD4+ T cell responses are also thought to be important for immune control of HIV, these cells are highly heterogenous. How IC expression and function differs across CD4+ T cell lineages and the consequences of this diversity for IC blockade (ICB) strategies are still poorly understood.

Added value of this studyTo compare various stages of immune dysfunction, we examined people living with HIV (PLWH) with different levels of viral control pre-ART (including elite controllers who spontaneously control virus) and followed a cohort longitudinally post-ART. We used a panel of assays to characterize HIV-specific CD4+ T cell subsets, including activation-induced marker (AIM) assays and flow cytometric detection of mRNAs coding for a wider variety of HIV-specific CD4+ T cell functions than what is detected by standard procedures. Our experiments indicate a hierarchy of IC (PD-1, TIGIT, CD200) expression on blood HIV-specific CD4+ T cells that depends not only on the persons infection status but also on expression of lineage differentiation markers and effector functions representative of CD4+ T cell subsets critical for antiviral responses (TFH, TH1 and TH17/TH22 cells). This hierarchy was also present in the putatively functional cells of elite controllers. We characterized the expression of the dysfunction-related transcription factor TOX, and saw that its association with the key IC PD-1 in the setting of viremia varied across CD4+ T cell polarizations. Response to blockade of the PD-1 pathway resulted in increased antiviral and mucosal-protective functions, but did not affect TFH-related functions. Response to ICB was most prominent in viremic patients, and subdued but not fully abrogated in the setting of viral suppression.

Implications of all available evidenceThese results highlight a previously unrecognized impact of ICB on mucosal immunity-related CD4+ functions, which are known to be depleted upon HIV infection and not restored by ART, and strong links between IC expression patterns and HIV-specific CD4+ T cell differentiation. The impact of ICB on CD4+ T cells in HIV infection has primarily been studied in the context of viral reservoir reactivation, which are preferentially harbored in IC+ cells. Our work emphasizes the importance of considering the differentiation profile of the virus-specific CD4+ T cells in studies of ICB blockade, as it may direct ICB efficacy in HIV infection. This data may also have implications for CD4+ T cell help in other infectious and non-infectious chronic human diseases.


title: The mosquito microbiome includes habitat-specific but rare symbionts

abstract: Microbial communities are known to influence mosquito lifestyles by modifying essential metabolic and behavioral processes that affect reproduction, development, immunity, digestion, egg survival, and ability to transmit pathogens. Many studies have used 16S rRNA gene amplicons to characterize mosquito microbiota and investigate factors that influence host-microbiota dynamics. However, a relatively low taxonomic resolution due to clustering methods based on arbitrary threshold and the overall dominance of Wolbachia or Asaia populations obscured the investigation of rare members of mosquito microbiota in previous studies. Here, we used high resolution Shannon entropy-based oligotyping approaches to analyze the microbiota of Culex pipiens, Culex quinquefasciatus and Aedes individuals from continental and overseas regions in Southern France and Guadeloupe as well as from laboratories with or without antibiotics treatment. Our experimental design that resulted in a series of mosquito samples with a gradient of Wolbachia density and relative abundance along with high-resolution analyses of amplicon sequences enabled the recovery of a robust signal from typically less accessible bacterial taxa. Our data confirm species-specific mosquito-bacteria associations with geography as a primary factor that influences bacterial community structure. But interestingly, they also reveal co-occurring symbiotic bacterial variants within single individuals for both Elizabethkingia and Erwinia genera, distinct and specific Asaia and Chryseobacterium in continental and overseas territories and a putative rare Wolbachia variant. Overall, our study reveals the presence of previously-overlooked microdiversity and multiple closely related symbiotic strains within mosquito individuals with a remarkable habitat-specificity.

category: microbiology


title: Alkaliphilus flagellata sp. nov., Butyricicoccus intestinisimiae sp. nov., Clostridium mobile sp. nov., Clostridium simiarum sp. nov., Dysosmobacter acutus sp. nov., Paenibacillus brevis sp. nov., Peptoniphilus ovalis sp. nov., and Tissierella simiarum sp. nov., isolated from monkey feces

abstract: Non-human primates harbor diverse microbiomes in their guts. As a part of China Microbiome Initiatives, we cultivated and characterized the gut microbiome of cynomolgus monkeys (Macaca fascicularis). In this report, we communicate the characterization and taxonomy of 8 bacterial strains that were obtained from fecal samples of captive cynomolgus monkeys. The results revealed that they represented 8 novel bacterial species. The proposed names of the 8 novel species are Alkaliphilus flagellate (type strain MSJ-5T =CGMCC 1.45007T=KCTC 15974T), Butyricicoccus intestinisimiae MSJd-7T (type strain MSJd-7T =CGMCC 1.45013T =KCTC 25112T), Clostridium mobile (type strain MSJ-11T =CGMCC 1.45009T=KCTC 25065T), Clostridium simiarum (type strain MSJ-4T =CGMCC 1.45006T =KCTC 15975T), Dysosmobacter acutus (type strain MSJ-2T =CGMCC 1.32896T=KCTC 15976T), Paenibacillus brevis MSJ-6T (type strain MSJ-6T =CGMCC 1.45008T=KCTC 15973T), Peptoniphilus ovalis (type strain MSJ-1T =CGMCC 1.31770T=KCTC 15977T), and Tissierella simiarum (type strain MSJ-40T =CGMCC 1.45012T=KCTC 25071T).


title: How Our Cells Become Our Selves: The Cellular Phylodynamic Biology of Growth and Development

abstract: Our lives begin with 1 cell, then 2, then 4, then the trillion cell adult, comprised of cell lineages, tissues, organs. How does this occur? Examination in numbers of cells, N, Cellular Phylodynamics, revealed two previously unappreciated processes: UNI-GROWTH, the slowing of growth that occurs as we become larger, caused by fewer cells dividing, captured by the Universal Mitotic Fraction and Universal Growth Equations, with accuracy confirmed for 13 species, including nematodes, mollusks, and vertebrates; and ALLO-GROWTH, the creation of body parts from Founder Cells, captured by the Cellular Allometric Growth Equation, which describes mitotic expansion by Cell-Heritable change in the Cell Cycle Time. These equations can generate cell lineage approximations, bringing the power of coalescent theory to developmental biology.

category: developmental biology


title: CHEK2 SIGNALING IS THE KEY REGULATOR OF OOCYTE SURVIVAL AFTER CHEMOTHERAPY

abstract:
Radiation and chemotherapy can damage the primordial follicle reserve in female cancer patients leading to ovarian failure and infertility. Preservation of ovarian function requires treatment strategies that prevent loss of immature oocytes in primordial follicles during cancer therapy. Checkpoint kinase 2 (CHEK2) inhibition prevents loss of primordial oocytes caused by DNA damage and thus is a promising target for ovoprotective treatment against genotoxic agents. To determine which cancer treatments could benefit from ovoprotective activity of CHEK2 inhibition we investigated oocyte survival in Chek2-/- mice exposed to different chemotherapy drugs. Here, we show that loss of CHEK2 function prevents elimination of primordial oocytes damaged by cisplatin, cyclophosphamide, mafosfamide, doxorubicin, and etoposide, suggesting it could be used to reduce ovarian damage caused by wide range of drugs. Using genetic knockouts we reveal a critical role for TRP53 in oocyte response to chemotherapy drugs and show that both targets of CHEK2--TAp63 and TRP53-- are activated by cisplatin and cyclophosphamide. Furthermore, we show that checkpoint kinase inhibitor and radiation- and chemotherapy sensitizer AZD7762 reduces oocyte elimination after radiation and chemotherapy treatments, despite its cytotoxic effect on ovarian somatic cells. Altogether, these findings demonstrate the role for CHEK2 as the master regulator of primordial oocyte survival or death and credential its targeting for ovoprotective treatments.

SIGNIFICANCEChemotherapy and radiation are ovotoxic and increase the risk of premature ovarian insufficiency and infertility in women cancer survivors. Development of treatment strategies preserving ovarian function and ensuring future reproductive health of female cancer patients depends on better understanding of the mechanisms underlying ovarian toxicity caused by different chemotherapy treatments. Preservation of long-term ovarian function can only be achieved by preventing the loss of immature oocytes in primordial follicles during toxic cancer therapies. Checkpoint kinase 2 (CHEK2) inhibition is an attractive strategy for protecting ovarian reserve with a potential additional benefit of sensitizing cancer cells to radiation and chemotherapy. Using a genetic approach, we show that blocking CHEK2 function is sufficient to prevent elimination of primordial oocytes damaged by chemotherapy drugs such as cisplatin, cyclophosphamide, mafosfamide, doxorubicin and etoposide. Many chemotherapy drugs are used in combination (e.g. cyclophosphamide with doxorubicin), thus the protective effect of CHEK2 inhibition is likely to be beneficial for a broad spectrum of patient treatments.


title: Early anteroposterior regionalisation of human neural crest is shaped by a pro-mesodermal factor

abstract: The neural crest (NC) is an important multipotent embryonic cell population and its impaired specification leads to various developmental defects, often in an anteroposterior (A-P) axial level-specific manner. The mechanisms underlying the correct A-P regionalisation of human NC cells remain elusive. Recent studies have indicated that trunk NC cells, the presumed precursors of the childhood tumour neuroblastoma, are derived from neuromesodermal-potent progenitors of the postcranial body (NMPs). Here we employ human embryonic stem cell differentiation to define how NMP-derived NC cells acquire a posterior axial identity. We show that TBXT, a pro-mesodermal transcription factor, mediates early posterior NC regionalisation together with WNT signalling effectors. This occurs by TBXT-driven chromatin remodelling via its binding in key enhancers within HOX gene clusters and other posterior regulator-associated loci. In contrast, posteriorisation of NMP-derived spinal cord cells is TBXT/WNT-independent and takes place under the influence of FGF signalling. Our work reveals a previously unknown role of TBXT in influencing posterior NC fate and points to the existence of temporally discrete, cell type-dependent modes of posterior axial identity control.


title: Towards a system-level causative knowledge of pollinator communities

abstract: Pollination plays a central role both in the maintenance of biodiversity and in crop production. However, habitat loss, pesticides, invasive species, and larger environmental fluctuations are contributing to a dramatic decline of numerous pollinators world-wide. This has increased the need for interventions to protect the composition, functioning, and dynamics of pollinator communities. Yet, how to make these interventions successful at the system level remains extremely challenging due to the complex nature of species interactions and the various unknown or unmeasured confounding ecological factors. Here, we propose that this knowledge can be derived by following a probabilistic causal analysis of pollinator communities. This analysis implies the inference of interventional expectations from the integration of observational and synthetic data. We propose that such synthetic data can be generated using theoretical models that can enable the tractability and scalability of unseen confounding ecological factors affecting the behavior of pollinator communities. We discuss a road map for how this probabilistic causal analysis can be accomplished to increase our system-level causative knowledge of natural communities.

category: ecology


title: A global meta-analysis reveals more variable life histories in urban birds compared to their non-urban neighbours

abstract: Cities pose a major ecological challenge for wildlife worldwide. Phenotypic variation, which can result from underlying genetic variation or plasticity, is an important metric to understand eco-evolutionary responses to environmental change. Recent work suggests that urban populations might have higher levels of phenotypic variation than non-urban counterparts. This prediction, however, has never been tested across taxa nor over a broad geographical range. Here, we conduct a meta-analysis of the avian literature to compare urban versus non-urban means and variances in breeding phenology (i.e., lay date) and reproductive effort (i.e., clutch size, number of fledglings). First, we show that urban populations reproduce earlier and have smaller broods than non-urban conspecifics. Second, we show that urban bird populations have higher phenotypic variation in laying date than non-urban populations. This result arises from differences in phenological variation between populations within breeding seasons, conceivably due to higher landscape heterogeneity in urban habitats. Higher variation in reproductive timing in birds subject to anthropogenic disturbance might be a consequence of plastic responses or higher genetic variation in phenology in urban populations. Our results reveal a novel effect of urbanisation on animal life-history traits with important implications for our understanding of how species adapt to urban environments.

category: evolutionary biology


title: A global meta-analysis reveals higher life-history phenotypic variation in urban birds than in their non-urban neighbours


title: Negative regulation of IMD contributes to disease tolerance during systemic bacterial infection in Drosophila

abstract: Disease tolerance is an infection phenotype where hosts show relatively high health despite harbouring elevated pathogen loads. Compared to the mechanisms of immune clearance our knowledge of the mechanisms underlying increased tolerance remains incomplete. Variation in the ability to reduce immunopathology may explain why some hosts can tolerate higher pathogen burdens with reduced pathology. Negative immune regulation would therefore appear to be a clear candidate for a mechanism underlying disease tolerance but this has not been tested directly for bacterial infections. Here, we examined how the negative regulation of the immune deficiency (IMD) pathway affects disease tolerance in Drosophila melanogaster when infected with the gram-negative bacterial pathogen Pseudomonas entomophila. We find that UASRNAi-mediated reduced expression of the negative regulators of IMD (pirk and caudal) severely reduced the ability to tolerate infection in both males and females across a wide range of infectious doses. While flies unable to regulate the IMD response exhibited higher expression of antimicrobial peptides and lower bacterial loads as expected, this was not accompanied by a proportional reduction in mortality. Instead, tolerance (measured as fly survival relative to its microbe load) was drastically reduced, likely due to the combination of increased immunopathology and cytotoxicity of elevated AMP expression. Our results therefore highlight that in addition to regulating an efficient pathogen clearance response, negative regulators of IMD also contribute to disease tolerance.


title: Regionalization, constraints, and the ancestral ossification patterns in the vertebral column of amniotes

abstract: The development of the vertebral column has been studied extensively in modern amniotes, yet many aspects of its evolutionary history remain enigmatic. Here we expand the existing data on four major vertebral developmental patterns in amniotes based on exceptionally well-preserved specimens of the early Permian mesosaurid reptile Stereosternum: (i) centrum ossification, (ii) neural arch ossification, (iii) neural arch fusion, and (iv) neurocentral fusion. We retrace the evolutionary history of each pattern and reconstruct the ancestral condition in amniotes. Despite 300 million years of evolutionary history, vertebral development patterns show a surprisingly stability in amniotes since their common ancestor. We propose that this conservatism may be linked to constraints posed by underlying developmental processes across amniotes. However, we also point out that mammals and birds differ more strongly from the ancestral condition than other clades, which might be linked to a stronger regionalization of the column in these two clades.


title: Genetic studies of human-chimpanzee divergence using stem cell fusions

abstract:
Complete genome sequencing has identified millions of DNA changes that differ between humans and chimpanzees. Although a subset of these changes likely underlies important phenotypic differences between humans and chimpanzees, it is currently difficult to distinguish causal from incidental changes and to map specific phenotypes to particular genome locations. To facilitate further genetic study of human-chimpanzee divergence, we have generated human and chimpanzee auto-tetraploids and allo-tetraploids by fusing induced pluripotent stem cells (iPSCs) of each species. The resulting tetraploid iPSCs can be stably maintained and retain the ability to differentiate along ectoderm, mesoderm, and endoderm lineages. RNA sequencing identifies thousands of genes whose expression differs between humans and chimpanzees when assessed in single-species diploid or auto-tetraploid iPSCs. Analysis of gene expression patterns in inter-specific allo-tetraploid iPSCs shows that human-chimpanzee expression differences arise from substantial contributions of both cis-acting changes linked to the genes themselves, and trans-acting changes elsewhere in the genome. To enable further genetic mapping of species differences, we tested chemical treatments for stimulating genome-wide mitotic recombination between human and chimpanzee chromosomes, and CRISPR methods for inducing species-specific changes on particular chromosomes in allo-tetraploid cells. We successfully generated derivative cells with nested deletions or inter-specific recombination on the X chromosome. These studies identify a long distance cis-regulatory domain of the Fragile X-associated gene (FMR1), confirm an important role for the X chromosome in trans-regulation of other expression differences, and illustrate the potential of this system for more detailed mapping of the molecular basis of human and chimpanzee evolution.

Significance StatementComparative studies of humans and chimpanzees have revealed many anatomical, physiological, behavioral, and molecular differences. However, it has been challenging to map these differences to particular chromosome regions. Here, we develop a genetic approach in fused stem cell lines that makes it possible to map human-chimpanzee molecular and cellular differences to specific regions of the genome. We illustrate this approach by mapping chromosome regions responsible for species-specific gene expression differences in fused tetraploid cells. This approach is general, and could be used in the future to map the genomic changes that control many other humanchimpanzee differences in various cell types or organoids in vitro.


title: The Jak/Stat pathway mediates disease tolerance during systemic bacterial infection in Drosophila

abstract: Disease tolerance describes a hosts ability to maintain health independently of the ability to clear microbe loads. However, we currently know little about the mechanisms that underlie disease tolerance or how known mechanisms of tissue damage signalling and repair may contribute to variation in tolerance. The Jak/Stat pathway plays a pivotal role in Drosophila humoral innate immunity, signalling tissue damage and triggering cellular renewal, making it a potential mechanism underlying the disease tolerance phenotype. Here, we show that disrupting the Jak/Stat pathway in Drosophila melanogaster alters disease tolerance during Pseudomonas entomophila systemic infection. Overall, flies with disrupted Jak/Stat show variation in survival that is not explained by variation in pathogen loads. For instance, mutations disrupting the function of ROS-producing dual oxidase (duox) or the negative regulator of Jak/Stat, Socs36E render males less tolerant to systemic bacterial infection but not females. We also investigated whether the negative regulator of Jak/Stat, G9a -which has previously been associated with tolerance of viral infections - is also implicated in tolerance of bacterial infection. While female flies lacking G9a showed higher mortality and reduced bacterial clearance, disease tolerance did not differ between G9a mutants and the wildtype. This suggests that G9a does not affect tolerance during systemic bacterial infection as it appears to do with viral infection. Overall, our findings highlight that Jak/Stat signalling mediates disease tolerance during systemic bacterial infection and that this response differs between males and females. Our work therefore suggests that differences in Jak/Stat mediated disease tolerance may be a potential source of sexually dimorphic response to infection in Drosophila.


title: Tyr is Responsible for the Cctq1a QTL and Links Developmental Environment to Central Corneal Thickness Determination

abstract: Central corneal thickness is a quantitative trait with important associations to human health. In a phenotype-driven approach studying corneal thickness of congenic derivatives of C57BLKS/J and SJL/J mice, the critical region for a quantitative trait locus influencing corneal thickness, Cctq1a, was delimited to a 10-gene interval. Exome sequencing, RNAseq, and studying independent mutations eliminated multiple candidate genes and confirmed one. Though the causative gene, Tyr, has no obvious direct function in the transparent cornea, studies with multiple alleles on matched genetic backgrounds, both in isolation and genetic complementation crosses, confirmed allelism of Tyr-Cctq1a; albino mice lacking Tyr function had thin corneas. Albino mice also had increased axial length. Because albinism exposes eyes to increased light, the effect of dark-rearing was tested and found to rescue central corneal thickness. In sum, the results point to an epiphenomenon; developmental light exposure interacts with genotype as an important determinate of adult corneal thickness.

category: genetics


title: Dominant negative effects of SCN5A missense variants

abstract:
IntroductionUp to 30% of patients with Brugada Syndrome (BrS) carry loss-of-function (LoF) variants in the cardiac sodium channel gene SCN5A. Recent studies have suggested that the SCN5A protein product NaV1.5 can form dimers and exert dominant negative effects.

MethodsWe identified 35 LoF variants (<10% peak current compared to wild type (WT)) and 15 partial LoF variants (10-50% peak current compared to WT) that we assessed for dominant negative behavior. SCN5A variants were studied in HEK293T cells alone or in heterozygous co-expression with WT SCN5A using automated patch clamp. To assess clinical risk, we compared the prevalence of dominant negative vs. putative haploinsufficient (frameshift/splice site) variants in a BrS case consortium and the gnomAD population database.

ResultsIn heterozygous expression with WT, 32/35 LoF variants and 6/15 partial LoF showed reduction to <75% of WT-alone peak INa, demonstrating a dominant negative effect. Carriers of dominant negative LoF missense variants had an enriched disease burden compared to putative haploinsufficient variant carriers (2.7-fold enrichment in BrS cases, p=0.019).

ConclusionsMost SCN5A missense LoF variants exert a dominant negative effect. Cohort analyses reveal that this class of variant confers an especially high burden of BrS.


title: Genetic effect modification of cis-acting C-reactive protein variants in cardiometabolic disease status

abstract: Mendelian randomization (MR) studies carried out among patients with a particular health condition should establish the genetic instrument influences the exposure in that subgroup, however this is normally investigated in the general population. Here, we investigated whether the genetic associations of four cis-acting C-reactive protein (CRP) variants differed between participants with and without three cardiometabolic conditions: obesity, type 2 diabetes, and cardiovascular disease. Associations of cis-genetic variants with CRP differed between obese and non-obese individuals. A multivariable analysis suggested strong independent associations of the gene-by-body mass index (BMI) interaction on CRP (P<1.18x10-8 for the CRP variants). Applying MR, we observed strong causal effect of BMI on CRP (P=2.14x10-65). In summary, our study indicates that genetic associations with CRP differ across disease sub-groups, with evidence to suggest that BMI is an effect modifier. MR studies of disease progression should report on the genetic instrument-exposure association in the disease subgroup under investigation.


title: Intraspecific de novo gene birth revealed by presence absence variant genes in Caenorhabditis elegans

abstract: Genes embed their evolutionary history in the form of various alleles. Presence-absence variants (PAVs) are extreme cases of such alleles, where a gene present in one haplotype does not exist in another. Since PAVs may result from either birth or death of a gene, PAV genes and their alternative alleles, if available, can represent a basis for rapid intraspecific gene evolution. Here, we traced a possible evolution of PAV genes in the PD1074 and CB4856 C. elegans strains as well as their alternative alleles found in other 14 wild strains, using long-read sequencing technologies. We updated the CB4856 genome by filling 18 gaps and identified 50 novel genes and 7,460 novel isoforms from both strains. We verified 328 PAV genes, out of which 48 were C. elegans-specific. Among these possible newly-born genes, 13 had alternative alleles in other wild strains and, in particular, alternative alleles of three genes showed signatures active transposons. Alternative alleles of four other genes showed another type of signature reflected in accumulation of small insertions or deletions. Our results exemplify that research on gene evolution using both species-specific PAV genes and their alternative alleles is expected to provide new perspectives for how genes evolve.

category: genomics


title: Internal oligo(dT) priming in bulk and single cell RNA sequencing

abstract: Significant advances in RNA sequencing have been recently made possible by the use of oligo(dT) primers for simultaneous mRNA enrichment and reverse transcription priming. The associated increase in efficiency has enabled more economical bulk RNA sequencing methods as well as the advent of high throughput single cell RNA sequencing, now already one of the most widely adopted new methods in the study of transcriptomics. However, the effects of off-target oligo(dT) priming on gene expression quantification have not been fully appreciated. In the present study, we describe the extent, the possible causes, and the consequences of internal oligo(dT) priming across multiple publicly available datasets obtained from a variety of bulk and single cell RNA sequencing platforms. In order to explore and address this issue, we developed a computational algorithm for identification of sequencing read alignments that likely resulted from internal oligo(dT) priming and their subsequent removal from the data. Directly comparing filtered datasets to those obtained by an alternative method reveals significant improvements in gene expression measurement. Finally, we infer a list of genes whose expression quantification is most likely to be affected by internal oligo(dT) priming.


title: A Comparative Genome-wide Transcriptome Analysis of Glucocorticoid Responder and Non-Responder Primary Human Trabecular Meshwork Cells

abstract: The genome-wide gene expression analysis of primary human trabecular meshwork (HTM) cells with known glucocorticoid (GC) responsiveness was not reported earlier. Therefore, the purpose of this study was to investigate genes and pathways involved in the GC responsiveness in human trabecular meshwork (HTM) cells using RNA sequencing. A perfusion cultured human anterior segment ex vivo model was utilized to identify the induction of GC-induced ocular hypertension in one eye of a paired eyes after dexamethasone treatment based on the maximum intraocular pressure response and in the contralateral eye, HTM cells were isolated to classify GC-responder and non-responder cells. Some previously reported and unique genes and their associated pathways were identified in HTM cells in response to dexamethasone treatment versus vehicle control and more significantly in GC-responder and non-responder cells. This study will open up the possibility of identifying suitable molecular targets which have the potential to treat GC-induced ocular hypertension/glaucoma.


title: Neural network modeling of differential binding between wild-type and mutant CTCF reveals putative binding preferences for zinc fingers 1-2

abstract:
BackgroundMany transcription factors (TFs), such as multi zinc-finger (ZF) TFs, have multiple DNA binding domains (DBDs) with multiple components, and deciphering the DNA binding motifs of individual components is a major challenge. One example of such a TF is CCCTC-binding factor (CTCF), a TF with eleven ZFs that plays a variety of roles in transcriptional regulation, most notably anchoring DNA loops. Previous studies found that CTCF zinc fingers (ZFs) 3-7 bind CTCFs core motif and ZFs 9-11 bind a specific upstream motif, but the motifs of ZFs 1-2 have yet to be identified.

ResultsWe developed a new approach to identifying the binding motifs of individual DBDs of a TF through analyzing chromatin immunoprecipitation sequencing (ChIP-seq) experiments in which a single DBD is mutated: we train a deep convolutional neural network to predict whether wild-type TF binding sites are preserved in the mutant TF dataset and interpret the model. We applied this approach to mouse CTCF ChIP-seq data and, in addition to identifying the known binding preferences of CTCF ZFs 3-11, we identified a GAG binding motif for ZF1 and a weak ATT binding motif for ZF2. We analyzed other CTCF datasets to provide additional evidence that ZFs 1-2 interact with the motifs we identified, and we found that the presence of the motif for ZF1 is associated with Ctcf peak strength.

ConclusionsOur approach can be applied to any TF for which in vivo binding data from both the wild-type and mutated versions of the TF are available, and our findings provide an unprecedently comprehensive understanding of the binding preferences of CTCFs DBDs.


title: A Discrete Formulation of Two Alternative Forced Choice Decision Dynamics Derived from a Double-Well Quantum Landscape

abstract: Tools from quantum theory have been effectively leveraged in modeling otherwise poorly understood effects in decision-making such as apparent fallacies in probability judgments and context effects. This approach has described the dynamics of two alternative forced choice (2AFC) decisions in terms of the path of a single quantum particle evolving in a single potential well. Here, we present a variant on that approach, which we name the Multi-Particle and Multi-Well (MPMW) quantum cognitive framework, in which decisions among N alternatives are treated by the sum of positional measurements of many independent quantum particles representing stimulus information, acted on by an N-well landscape that defines the decision alternatives. In this article, we apply the MPMW model to the simplest and most common case of N-alternative decision making, 2AFC dynamics. This application calls for a multi-particle double-well implementation, which allows us to construct a simple, analytically tractable discrete drift diffusion model (DDM), in the form of a Markov chain, wherein the parameters of the attractor wells reflect bottom-up (automatic) and top-down (control-dependent) influences on the integration of external information. We first analyze this Markov chain in its simplest form, as a single integrator with a generative process arising from a static quantum landscape and fixed thresholds, and then consider the case of multi-integrator processing under the same conditions. Within this system, stochasticity arises directly from the double-well quantum attractor landscape as a function of the dimensions of its wells, rather than as an external parameter requiring independent fitting. The simplicity of the Markov chain component of this model allows for easy analytical computation of closed forms for response time distributions and response probabilities that match qualitative properties of the accuracies and reaction times of humans performing 2AFC tasks. The MPMW framework produces response time distributions following inverse gaussian curves familiar from previous DDM models and empirical data, including the common observation that mean response times are faster for incorrect than for correct responses. The work presented in this paper serves as a proof of concept, based on which the MPMW framework can be extended to address more complex decision-making processes, (e.g., N-alternative forced choice, dynamic control allocation, and nesting quantum landscapes to allow for modeling at both the task and stimulus levels of processing) that we discuss as future directions.

category: animal behavior and cognition


title: An innovative optical context to make honeybees crash repeatedly

abstract: To investigate altitude control in honeybees, an optical context was designed to make honeybees crash. It has been widely accepted that honeybees rely on the optic flow generated by the ground to control their altitude. However, identifying an optical context capable of uncorrelating forward speed from altitude in honeybees flight was the first step towards enhancing the optical context to better understand altitude control in honeybees. This optical context aims to put honeybees in the same flight conditions as an open sky flight above mirror-smooth water. An optical manipulation, based on a pair of opposed horizontal mirrors, was designed to remove any visual information coming from the floor and ceiling. Such an optical manipulation reproduced quantitatively the seminal experiment of Heran & Lindauer (1963), and revealed that honeybees control their altitude by detecting the optic flow with a visual field that extends to approximately 165{degrees}.


title: Personalizing the control law of an upper-limb exoskeleton using EMG signal

abstract: Implementing an intuitive control law for an upper-limb exoskeleton dedicated to force augmentation is a challenging issue in the field of human-robot collaboration. The goal of this study is to adapt an EMG-based control system to a user based on individual caracteristics. To this aim, a method has been designed to tune the parameters of control using objective criteria, improving users feedback. The users response time is used as an objective value to adapt the gain of the controller. The proposed approach was tested on 10 participants during a lifting task. Two different conditions have been used to control the exoskeleton: with a generic gain and with a personalized gain. EMG signals was captured on five muscles to evaluate the efficiency of the conditions and the users adaptation. Results showed a statistically significant reduction of mean muscle activity of the deltoid between the beginning and the end of each situation (28.6 {+/-} 13.5% to 17.2 {+/-} 7.3% of Relative Maximal Contraction for the generic gain and from 24.9 {+/-} 8.5% to 18.0 {+/-} 6.8% of Relative Maximal Contraction for the personalized gain). When focusing on the first assisted movements, the personalized gain induced a mean activity of the deltoid significantly lower (29.0 {+/-} 8.0% of Relative Maximal Contraction and 37.4 {+/-} 9.5% of Relative Maximal Contraction, respectively). Subjective evaluation showed that the system with a personalized gain was perceived as more intuitive, and required less concentration when compared to the system with a generic gain.

category: bioengineering


title: Energy efficient convolutional neural networks for arrhythmia detection

abstract: Electrocardiograms (ECG) record the heart activity and are the most common and reliable method to detect cardiac arrhythmias, such as atrial fibrillation (AFib). Lately, many commercially available devices such as smartwatches are offering ECG monitoring. Therefore, there is increasing demand for designing deep learning models with the perspective to be physically implemented on these small portable devices with limited energy supply. In this paper, a workflow for the design of small, energy-efficient recurrent convolutional neural network (RCNN) architecture for AFib detection is proposed. However, the approach can be well generalized to every type of long time series. In contrast to previous studies, that demand thousands of additional network neurons and millions of extra model parameters, the logical steps for the generation of a CNN with only 114 trainable parameters are described. The model consists of a small segmented CNN in combination with an optimal energy classifier. The architectural decisions are made by using the energy consumption as a metric in an equally important way as the accuracy. The optimisation steps are focused on the software which can be embedded afterwards on a physical chip. Finally, a comparison with some previous relevant studies suggests that the widely used huge CNNs for similar tasks are mostly redundant and unessentially computationally expensive.

category: bioinformatics


title: Identification of active modules in interaction networks using node2vec network embedding

abstract:
The identification of condition-specific gene sets from transcriptomic experiments is important to reveal regulatory and signaling mechanisms associated with a given cellular response. Statistical approaches using only expression data allow the identification of genes whose expression is most altered between different conditions. However, a phenotype is rarely a direct consequence of the activity of a single gene, but rather reflects the interplay of several genes to carry out certain molecular processes. Many methods have been proposed to analyze the activity of genes in light of our knowledge of their molecular interactions. However, existing methods have many limitations that make them of limited use to biologists: they detect modules that are too large, too small, or they require the users to specify a priori the size of the modules they are looking for.

We propose AMINE (Active Module Identification through Network Embedding), an efficient method for the identification of active modules. Experiments carried out on artificial data sets show that the results obtained are more reliable than many available methods. Moreover, the size of the modules to be identified is not a fixed parameter of the method and does not need to be specified; rather, it adjusts according to the size of the modules to be found. The applications carried out on real datasets show that the method enables to find important genes already highlighted by approaches solely based on gene variations, but also to identify new groups of genes of high interest. In addition, AMINE method can be used as a web service on your own data (http://amine.i3s.unice.fr).


title: Rescuing Biologically Relevant Consensus Regions Across Replicated Samples

abstract:
MotivationProtein-DNA binding sites of ChIP-seq experiments are identified where the binding affinity is significant based on a given threshold. The choice of the threshold is a trade-off between conservative region identification and discarding weak, but true binding sites.

ResultsWe argue the biological relevance of weak binding sites and the information they add when rescued. The sites are rescued using MSPC, which exploits replicates to lower the threshold required to identify a binding site while keeping a low false-positive rate. We extend MSPC to call consensus regions across any number of replicated samples, accounting for differences between biological and technical replicates. We observed several master transcription regulators (e.g., SP1 and GATA3) and HDAC2-GATA1 regulatory networks on rescued regions.

Availability and implementationAn implementation of the proposed method and the scripts to reproduce the performed analysis are freely available at https://genometric.github.io/MSPC/, MSPC is distributed as a command-line application, an R package available from Bioconductor (https://doi.org/doi:10.18129/B9.bioc.rmspc), and a C# library.


title: Dr. Sim: Similarity Learning for Transcriptional Phenotypic Drug discovery

abstract: Transcriptional phenotypic drug discovery has achieved great success, and various compound perturbation-based data resources, such as Connectivity Map (CMap) and Library of Integrated Network-Based Cellular Signatures (LINCS), have been presented. Computational strategies fully mining these resources for phenotypic drug discovery have been proposed, and among them, a fundamental issue is to define the proper similarity between the transcriptional profiles to elucidate the drug mechanism of actions and identify new drug indications. Traditionally, this similarity has been defined in an unsupervised way, and due to the high dimensionality and the existence of high noise in those high-throughput data, it lacks robustness with limited performance. In our study, we present Dr. Sim, which is a general learning-based framework that automatically infers similarity measurement rather than being manually designed and can be used to characterize transcriptional phenotypic profiles for drug discovery with generalized good performance. We evaluated Dr. Sim on comprehensively publicly available in vitro and in vivo datasets in drug annotation and repositioning using high-throughput transcriptional perturbation data and indicated that Dr. Sim significantly outperforms the existing methods and is proved to be a conceptual improvement by learning transcriptional similarity to facilitate the broad utility of high-throughput transcriptional perturbation data for phenotypic drug discovery. The source code and usage of Dr. Sim is available at https://github.com/bm2-lab/DrSim/.