title: PMRT1, a Plasmodium specific parasite plasma membrane transporter is essential for asexual and sexual blood stage development

abstract:
Membrane transport proteins perform crucial roles in cell physiology. The obligate intracellular parasite Plasmodium falciparum, an agent of human malaria, relies on membrane transport proteins for the uptake of nutrients from the host, disposal of metabolic waste, exchange of metabolites between organelles and generation and maintenance of transmembrane electrochemical gradients for its growth and replication within human erythrocytes. Despite their importance for Plasmodium cellular physiology, the functional roles of a number of membrane transport proteins remain unclear, which is particularly true for orphan membrane transporters that have no or limited sequence homology to transporter proteins in other evolutionary lineages. Therefore, in the current study, we applied endogenous tagging, targeted gene disruption, conditional knockdown and knockout approaches to investigate the subcellular localization and essentiality of six membrane transporters during intraerythrocytic development of P. falciparum parasites. They are localized at different subcellular structures - the food vacuole, the apicoplast, and the parasite plasma membrane - and four out of the six membrane transporters are essential during asexual development. Additionally, the plasma membrane resident transporter 1 (PMRT1, PF3D7_1135300), a unique Plasmodium-specific plasma membrane transporter, was shown to be essential for gametocytogenesis and functionally conserved within the genus Plasmodium. Overall, we reveal the importance of four orphan transporters to blood stage P. falciparum development, which have diverse intracellular localizations and putative functions.

ImportancePlasmodium falciparum-infected erythrocytes possess multiple compartments with designated membranes. Transporter proteins embedded in these membranes do not only facilitate movement of nutrients, metabolites and other molecules between these compartments, but are common therapeutic targets and can also confer antimalarial drug resistance. Orphan membrane transporter in P. falciparum without sequence homology to transporters in other evolutionary lineages and divergent to host transporters may constitute attractive targets for novel intervention approaches. Here, we localized six of these putative transporters at different subcellular compartments and probed into their importance during asexual parasite growth using reverse genetic approaches. In total, only two candidates turned out to be dispensable for the parasite, highlighting four candidates as putative targets for therapeutic interventions. This study reveals the importance of several orphan transporters to blood stage P. falciparum development.


category: cell biology


title: Epistatic selection on a selfish Segregation Distorter supergene: drive, recombination, and genetic load

abstract: Meiotic drive supergenes are complexes of alleles at linked loci that together subvert Mendelian segregation to gain preferential transmission. In males, the most common mechanism of drive involves the disruption of sperm bearing alternative alleles. While at least two loci are important for male drive-- the driver and the target-- linked modifiers can enhance drive, creating selection pressure to suppress recombination. In this work, we investigate the evolution and genomic consequences of an autosomal multilocus, male meiotic drive system, Segregation Distorter (SD) in the fruit fly, Drosophila melanogaster. In African populations, the predominant SD chromosome variant, SD-Mal, is characterized by two overlapping, paracentric inversion on chromosome arm 2R and nearly perfect (~100%) transmission. We study the SD-Mal system in detail, exploring its components, chromosomal structure, and evolutionary history. Our findings reveal a recent chromosome-scale selective sweep mediated by strong epistatic selection for haplotypes carrying Sd, the main driving allele, and one or more factors within the double inversion. While most SD-Mal chromosomes are homozygous lethal, SD-Mal haplotypes can recombine with other, complementing haplotypes via crossing over and with wildtype chromosomes only via gene conversion. SD-Mal chromosomes have nevertheless accumulated lethal mutations, excess non-synonymous mutations, and excess transposable element insertions. Therefore, SD-Mal haplotypes evolve as a small, semi-isolated subpopulation with a history of strong selection. These results may explain the evolutionary turnover of SD haplotypes in different populations around the world and have implications for supergene evolution broadly.

category: evolutionary biology


title: An empirical evaluation of genotype imputation of ancient DNA

abstract: With capabilities of sequencing ancient DNA to high coverage often limited by sample quality or cost, imputation of missing genotypes presents a possibility to increase power of inference as well as cost-effectiveness for the analysis of ancient data. However, the high degree of uncertainty often associated with ancient DNA poses several methodological challenges, and performance of imputation methods in this context has not been fully explored. To gain further insights, we performed a systematic evaluation of imputation of ancient data using Beagle 4.0 and reference data from phase 3 of the 1000 Genomes project, investigating the effects of coverage, phased reference and study sample size. Making use of five ancient samples with high-coverage data available, we evaluated imputed data with respect to accuracy, reference bias and genetic affinities as captured by PCA. We obtained genotype concordance levels of over 99% for data with 1x coverage, and similar levels of accuracy and reference bias at levels as low as 0.75x. Our findings suggest that using imputed data can be a realistic option for various population genetic analyses even for data in coverage ranges below 1x. We also show that a large and varied phased reference set as well as the inclusion of low- to moderate-coverage ancient samples can increase imputation performance, particularly for rare alleles. In-depth analysis of imputed data with respect to genetic variants and allele frequencies gave further insight into the nature of errors arising during imputation, and can provide practical guidelines for post-processing and validation prior to downstream analysis.

category: genomics


title: TIAToolbox: An End-to-End Toolbox for Advanced Tissue Image Analytics

abstract: Computational Pathology (CPath) has seen rapid growth in recent years, driven by advanced deep learning (DL) algorithms. These algorithms typically share the same sequence of steps. However, due to the sheer size and complexity of handling large multi-gigapixel whole-slide images, there is no open-source software library that provides a generic end-to-end API for pathology image analysis using best practices for CPath. Most researchers have designed custom pipelines from the bottom-up, restricting the development of advanced CPath algorithms to specialist users. To help overcome this bottleneck, we present TIAToolbox, a Python toolbox designed to make CPath more accessible to new and advanced CPath scientists and pathologists alike. We provide a usable and adaptable library with efficient, cutting-edge and unit-tested tools for data loading, pre-processing, model inference, post-processing and visualization. This enables all kinds of users to easily build upon recent DL developments in the CPath literature. TIAToolbox provides a user-friendly modular API to enable seamless integration of advanced DL algorithms. We show with the help of examples how state-of-the-art DL algorithms can be streamlined using TIAToolbox.

category: bioengineering


title: fingeRNAt - a novel tool for high-throughput analysis of nucleic acid-ligand interactions

abstract:
Computational methods play a pivotal role in drug discovery and are widely applied in virtual screening, structure optimization, and compound activity profiling. Over the last decades, almost all the attention in medicinal chemistry has been directed to protein-ligand binding, and computational tools have been created with this target in mind. With novel discoveries of functional RNAs and their possible applications, RNAs have gained considerable attention as potential drug targets. However, the availability of bioinformatics tools for nucleic acids is limited. Here, we introduce fingeRNAt - a software tool for detecting non-covalent interactions formed in complexes of nucleic acids with ligands. The program detects nine types of interactions: (i) hydrogen and (ii) halogen bonds, (iii) cation-anion, (iv) pi-cation, (v) pi-anion, (vi) pi-stacking, (vii) inorganic ion-mediated, (viii) water-mediated, and (ix) lipophilic interactions. However, the scope of detected interactions can be easily expanded using a simple plugin system. In addition, detected interactions can be visualized using the associated PyMOL plugin, which facilitates the analysis of medium-throughput molecular complexes. Interactions are also encoded and stored as a bioinformatics-friendly Structural Interaction Fingerprint (SIFt) - a binary string where the respective bit in the fingerprint is set to 1 if a particular interaction is present and to 0 otherwise. This output format, in turn, enables high-throughput analysis of interaction data using data analysis techniques. We present applications of fingeRNAt-generated interaction fingerprints for visual and computational analysis of RNA-ligand complexes, including analysis of interactions formed in experimentally determined RNA-small molecule ligand complexes deposited in the Protein Data Bank. We propose interaction-based similarity based on fingerprints as an alternative measure to RMSD to recapitulate complexes with similar interactions but different folding. We present an application of molecular fingerprints for the clustering of molecular complexes. This approach can be used to group ligands that form similar binding networks and thus have similar biological properties.

AUTHOR SUMMARYWe present a novel bioinformatic tool, fingeRNAt, aiming to support scientists in the analysis of complexes of nucleic acids with various types of ligands. The software automatically detects non-covalent interactions and presents them in a form that is understandable to both humans and computers. Such data can help decipher the nature of interactions between nucleic acids and ligands and determine the main factors responsible for forming such complexes in nature. fingeRNAt finds application in multiple studies, both structure- and drug discovery-oriented. Here, we analyzed the experimentally solved structures of RNA complexes with small molecules to determine which binding features are most prevalent, i.e., most common interactions or their hot spots. The results of this analysis may help elucidate the mechanisms of binding and design new active molecules. Moreover, we propose to use the data generated by our software as a new metric for the quantitative comparison of two molecule complexes. We have shown that it is more reliable than the currently used methods in certain "difficult" cases. We have shown that the results of our program can be used for high-throughput analysis of molecular complexes and the search for active molecules. We are confident that fingeRNAt will be a valuable tool for exploring the complex world of interactions of nucleic acids with ligands.

category: bioinformatics


title: Blood-brain Barrier Crossing using Magnetic Stimulated Nanoparticles

abstract: Due to the low permeability and high selectivity of the blood-brain barrier (BBB), existing brain therapeutic technologies are limited by the inefficient BBB crossing of conventional drugs. Magnetic nanoparticles (MNPs) have shown great potential as nano-carriers for efficient BBB crossing under the external static magnetic field (SMF). To quantify the impact of SMF on MNPs in vivo dynamics towards BBB crossing, we developed a physiologically based pharmacokinetic (PBPK) model for intraperitoneal (IP) injected superparamagnetic iron oxide nanoparticles coated by gold and conjugated with poly(ethylene glycol) (PEG) (SPIO-Au-PEG NPs) in mice. Unlike most reported PBPK models that ignore brain permeability, we first obtained the brain permeabilities with and without SMF by determining the concentration of SPIO-Au-PEG NPs in the cerebral blood and brain tissue. This concentration in the brain was simulated by the advection-diffusion equations and was numerically solved in COMSOL Multiphysics. The results from the PBPK model after incorporating the brain permeability showed a good agreement (regression coefficient R2 = 0.825) with the in vivo results, verifying the capability of using the proposed PBPK model to predict the in vivo biodistribution of SPIO-Au-PEG NPs under the exposure to SMF. Furthermore, the in vivo results revealed that the brain bioavailability under the exposure to SMF (4.01%) is slightly better than the control group (3.68%). In addition, the modification of SPIO-Au-PEG NPs with insulin (SPIO-Au-PEG-insulin) showed an improvement of the brain bioavailability by 24.47 % in comparison to the non-insulin group. With the SMF stimulation, the brain bioavailability of SPIO-Au-PEG-insulin was further improved by 3.91 % compared to the group without SMF.

category: pharmacology and toxicology


title: Quantitative T1-relaxation corrected metabolite mapping of 12 metabolites in the human brain at 9.4 T

abstract:
Magnetic resonance spectroscopic imaging (MRSI) is a non-invasive imaging modality that enables observation of metabolites. Applications of MRSI for neuroimaging applications has shown promise for monitoring and detecting various diseases. This study builds off previously developed techniques of short TR, 1H FID MRSI by correcting for T1-weighting of the metabolites and utilizing an internal water reference to produce quantitative (mmol kg-1) metabolite maps. This work reports and shows quantitative metabolite maps for 12 metabolites for a single slice. Voxel-specific T1-corrections for water are common in MRSI studies; however, most studies use either averaged T1-relaxation times to correct for T1-weighting of metabolites or omit this correction step entirely. This work employs the use of voxel-specific T1-corrections for metabolites in addition to water. Utilizing averaged T1-relaxation times for metabolites can bias metabolite maps for metabolites that have strong differences between T1-relaxation for GM and WM (i.e. Glu). This work systematically compares quantitative metabolite maps to single voxel quantitative results and qualitatively compares metabolite maps to previous works.

HighlightsO_LIQuantitative metabolite maps of 12 metabolites human brains acquired at 9.4 T.
C_LIO_LIVoxel-specific T1-weighting corrections for water and metabolites.
C_LIO_LIComparisons of T1-weighted and T1-corrected metabolite maps.
C_LI

category: neuroscience


title: Trans-segmental imaging in the spinal cord of behaving mice

abstract: Spinal cord circuits play crucial roles in transmitting and gating cutaneous somatosensory modalities, such as pain, but the underlying activity patterns within and across spinal segments in behaving mice have remained elusive. To enable such measurements, we developed a wearable widefield macroscope with a 7.9 mm2 field of view, subcellular lateral resolution, 2.7 mm working distance, and <10 g overall weight. We show that highly localized painful mechanical stimuli evoke widespread, coordinated astrocyte excitation across multiple spinal segments.


title: Profiling the Landscape of Drug Resistance Mutations in Neosubstrates to Molecular Glue Degraders

abstract: Targeted protein degradation (TPD) holds immense promise for drug discovery but mechanisms of acquired resistance to degraders remain to be fully identified. Here we used CRISPR-suppressor scanning to identify mechanistic classes of drug resistance mutations to molecular glue degraders in GSPT1 and RBM39, neosubstrates targeted by E3 ligase substrate receptors cereblon and DCAF15, respectively. While many mutations directly alter the ternary complex heterodimerization surface, distal resistance sites were also identified. Several distal mutations in RBM39 led to modest decreases in degradation yet can enable cell survival, underscoring how small differences in degradation can lead to resistance. Integrative analysis of resistance sites across GSPT1 and RBM39 revealed varying levels of sequence conservation and mutational constraint that control the emergence of different resistance mechanisms, highlighting that many regions co-opted by TPD are inessential. Altogether, our study identifies common resistance mechanisms for molecular glue degraders and outlines a general approach to survey neosubstrate requirements necessary for effective degradation.

category: molecular biology


title: Interactions between xylem traits linked to hydraulics during xylem development optimize growth performance in conifer seedlings

abstract:
O_LIClimate change has threatened forests globally, challenging tree species ability to track the rapidly changing environment (e.g., drought and temperature rise). Conifer species face strong environmental filters due to climatic seasonality. Investigating how conifers change their hydraulic architecture during xylem development across the growing season may shed light on possible mechanisms underlying hydraulic adaptation in conifers.
C_LIO_LILaser microscopy was used to assess the three-dimensional hydraulic architecture of balsam fir (Abies balsamea (Linnaeus) Miller), jack pine (Pinus banksiana Lambert), white spruce (Picea glauca (Moench) Voss), and black spruce (Picea mariana (Miller) Britton, Sterns & Poggenburgh) seedlings. We measured hydraulic-related xylem traits in different regions (from early to latewood), during four years of plant growth.
C_LIO_LIThe xylem development of jack pine seedlings contrasts with the other species by maintaining torus overlap (a hydraulic safety-associated xylem trait), relatively constant across the season (from early to latewood), and over the years studied. The expansion of tracheids and torus was positively associated with plant growth.
C_LIO_LIPit aperture-torus covariance is central to the seasonal dynamics of jack pine xylem development, which jointly with a rapid tracheid and pit expansion is consistent with strong growth performance. Linking xylem structural changes during xylem development with hydraulics is a major issue for future research to assess conifers vulnerability to climate change.
C_LI

category: ecology


title: Accumulation and maintenance of information in evolution

abstract: Selection accumulates information in the genome -- it guides stochastically evolving populations towards states (genotype frequencies) that would be unlikely under neutrality. This can be quantified as the Kullback-Leibler (KL) divergence between the actual distribution of genotype frequencies and the corresponding neutral distribution. First, we show that this population-level information sets an upper bound on the information at the level of genotype and phenotype, limiting how precisely they can be specified by selection. Next, we study how the accumulation and maintenance of information is limited by the cost of selection, measured as the genetic load or the relative fitness variance, both of which we connect to the control-theoretic KL cost of control. The information accumulation rate is upper bounded by the population size times the cost of selection. This bound is very general, and applies across models (Wright-Fisher, Moran, diffusion) and to arbitrary forms of selection, mutation and recombination. Finally, the cost of maintaining information depends on how it is encoded: specifying a single allele out of two is expensive, but one bit encoded among many weakly specified loci (as in a polygenic trait) is cheap.


title: PERIOD phosphoclusters control temperature compensation of the Drosophila circadian clock

abstract:
Temperature compensation is a critical feature of circadian rhythms, but how it is achieved remains elusive. Here, we uncovered the important role played by the Drosophila PERIOD (PER) phosphodegron in temperature compensation. Using CRISPR-Cas9, we introduced a series of mutations that altered three Serines (S44, 45 and 47) belonging to the PER phosphodegron, the functional homolog of mammalian PER2s S487 phosphodegron, which impacts temperature compensation. While all three Serine to Alanine substitutions lengthened period at all temperatures tested, temperature compensation was differentially affected. S44A and S45A substitutions caused decreased temperature compensation, while S47A resulted in overcompensation. These results thus reveal unexpected functional heterogeneity of phosphodegron residues in thermal compensation. Furthermore, mutations impairing phosphorylation of the pers phosphocluster decreased thermal compensation, consistent with its inhibitory role on S47 phosphorylation. Interestingly, the S47A substitution caused increased accumulation of hyper-phosphorylated PER at warmer temperatures. This finding was corroborated by cell culture assays in which S47A resulted in excessive temperature compensation of phosphorylation-dependent PER degradation. Thus, we show a novel role of the PER phosphodegron in temperature compensation through temperature-dependent modulation of the abundance of hyper-phosphorylated PER. Our work also reveals interesting mechanistic convergences and differences between mammalian and Drosophila temperature compensation of the circadian clock.

Author summaryCircadian rhythms are critical adaptive mechanisms that enable most organisms to adjust their physiology and behavior to the changes that occur in their environment every day. Ambient temperature varies constantly, but interestingly molecular circadian pacemakers do not accelerate with increasing temperature, while most biochemical reactions are sensitive to temperature. This phenomenon of circadian temperature compensation is poorly understood. Using genome editing and transgenic approaches, we found that two phosphorylated motifs in the Drosophila PERIOD protein, which regulate stability, impact temperature compensation. Moreover, we observed that mutation of a key Serine residue controlling PER degradation, S47, affects the accumulation of phosphorylated PER in a temperature-dependent manner, and causes PER degradation kinetics to become overly protected from increased temperature. As a result, the circadian clock of S47 mutant flies is excessively temperature-compensated. Our work thus reveals an interesting mechanism that controls temperature compensation in Drosophila. Moreover, comparison with mammals reveal interesting similarities, but also important differences in how temperature compensation of the circadian clock is achieved.

category: genetics


title: Identifying mutations in sd1, Pi54 and Pi-ta, and positively selected genes of TN1, the first semidwarf rice in Green Revolution

abstract:
BackgroundTaichung Native 1 (TN1) is the first semidwarf rice cultivar that initiated the Green Revolution. As TN1 is a direct descendant of the Dee-geo-woo-gen cultivar, the source of the sd1 semidwarf gene, the sd1 gene can be defined through TN1. Also, TN1 is susceptible to the blast disease and is described as being drought-tolerant. However, genes related to these characteristics of TN1 are unknown. Our aim was to identify and characterize TN1 genes related to these traits.

ResultsAligning the sd1 of TN1 to Nipponbare sd1, we found a 382-bp deletion including a frameshift mutation. Sanger sequencing validated this deleted region in sd1, and we proposed a model of the sd1 gene that corrects errors in the literature. We also predicted the blast disease resistant (R) genes of TN1. Orthologues of the R genes in Tetep, a well-known resistant cultivar that is commonly used as a donor for breeding new blast resistant cultivars, were then sought in TN1, and if they were present, we looked for mutations. The absence of Pi54, a well-known R gene, in TN1 partially explains why TN1 is more susceptible to blast than Tetep. We also scanned the TN1 genome using the PosiGene software and identified 11 genes deemed to have undergone positive selection. Some of them are associated with drought-resistance and stress response.

ConclusionsWe have redefined the deletion of the sd1 gene in TN1, a direct descendant of the Dee-geo-woo-gen cultivar, and have corrected some literature errors. Moreover, we have identified blast resistant genes and positively selected genes, including genes that characterize TN1s blast susceptibility and abiotic stress response. These new findings increase the potential of using TN1 to breed new rice cultivars.


title: Temporal clusters of age-related behavioral alterations captured in smartphone touchscreen interactions

abstract: Cognitive and behavioral abilities alter across the adult life span. Smartphones engage various cognitive functions and the corresponding touchscreen interactions may help resolve if and how the behavior is systematically structured by aging. Here, in a sample spanning the adult lifespan (16 to 86 years, N = 598, accumulating 355 million interactions) we analyzed a range of interaction intervals - from a few milliseconds to a minute. We used probability distributions to cluster the interactions according to their next inter-touch interval dynamics to discover systematic age-related changes at the distinct temporal clusters. There were age-related behavioral losses at the clusters occupying short intervals (~ 100 ms, R2 ~ 0.8) but gains at the long intervals (~ 4 s, R2 ~ 0.4). These correlates were independent of the years of experience on the phone or the choice of fingers used on the screen. We found further evidence for a compartmentalized influence of aging, as individuals simultaneously demonstrated both accelerated and decelerated aging at distant temporal clusters. In contrast to these strong correlations, cognitive tests probing sensorimotor, working memory, and executive processes revealed rather weak age-related decline. Contrary to the common notion of a simple behavioral decline with age based on conventional cognitive tests, we show that real-world behavior does not simply decline and the nature of aging systematically varies according to the underlying temporal dynamics. Of all the imaginable factors determining smartphone interactions in the real world, age-sensitive cognitive and behavioral processes can dominatingly dictate smartphone temporal dynamics.

category: animal behavior and cognition


title: Theoretical investigation of active listening behavior based on the echolocation of CF-FM bats

abstract:
Bats perceive the three-dimensional (3D) environment by emitting ultrasound pulses from their nose or mouth and receiving echoes through both ears. To detect the position of a target object, it is necessary to know the distance and direction of the target. Certain bat species synchronize the movement of their pinnae with pulse emission, and it is this behavior that enables 3D direction detection. However, the significance of bats ear motions remains unclear. In this study, we construct a model of an active listening system including the motion of the ears, and conduct mathematical investigations to clarify the importance of ear motion in 3D direction detection. The theory suggests that only certain ear motions, namely three-axis rotation, accomplish accurate and robust 3D direction detection. Our theoretical analysis also strongly supports the behavior whereby bats move their pinnae in the antiphase mode. In addition, we provide the conditions for ear motions to ensure accurate and robust direction detection, suggesting that simple shaped hearing directionality and well-selected uncomplicated ear motions are sufficient to achieve precise and robust 3D direction detection. Our findings and mathematical approach have the potential to be used in the design of active sensing systems in various engineering fields.

Author SummaryMany mammals use visual sensing for primary perception of their surroundings, whereas bats accomplish spatial perception by active acoustic sensing. In particular, by emitting ultrasound pulses and listening to the echoes, bats localize reflective objects, a process known as echolocation. Certain bat species move both of their ears while receiving the echoes, but the essential theory behind this ear movement remains unclear.

This paper describes a simple mathematical model for investigating the active listening strategy employed by bats. The theory suggests that the ear motions employed by bats enables highly accurate direction detection that is robust to observation errors. In addition, we determine what kind of ear motions are optimal for 3D direction detection. This study not only reveals the significance of pinnae motions in bats, but also opens up the possibility of engineering applications for active listening systems.


title: Homogeneity of antibody-drug conjugates critically impacts the therapeutic efficacy in brain tumors

abstract: Glioblastoma multiforme (GBM) is characterized by aggressive growth and the poorest prognosis of all brain tumor types. Most therapies rarely provide clinically meaningful improvements in outcomes of patients with GBM. Antibody-drug conjugates (ADCs) are emerging chemotherapeutics with stunning success in cancer management. Although promising, clinical studies of three ADCs for treating GBM, including Depatux-M, have been discontinued because of safety concerns and limited therapeutic benefits. Here, we report that ADC homogeneity is a critical parameter to maximize the therapeutic potential in GBM therapy. We demonstrate that homogeneous conjugates generated using our linker show enhanced drug delivery to intracranial brain tumors. Notably, compared to heterogeneous ADCs, including a Depatux-M analog, our ADCs provide greatly improved antitumor effects and survival benefits in orthotopic brain tumor models, including a patient-derived xenograft model of GBM. Our findings warrant the future development of homogeneous ADCs as promising molecular entities toward cures for intractable brain tumors.


title: SpaceX: Gene Co-expression Network Estimation for Spatial Transcriptomics

abstract:
MotivationThe analysis of spatially-resolved transcriptome enables the understanding of the spatial interactions between the cellular environment and transcriptional regulation. In particular, the characterization of the gene-gene co-expression at distinct spatial locations or cell types in the tissue enables delineation of spatial co-regulatory patterns as opposed to standard differential single gene analyses. To enhance the ability and potential of spatial transcriptomics technologies to drive biological discovery, we develop a statistical framework to detect gene co-expression patterns in a spatially structured tissue consisting of different clusters in the form of cell classes or tissue domains.

ResultsWe develop SpaceX (spatially dependent gene co-expression network), a Bayesian methodology to identify both shared and cluster-specific co-expression network across genes. SpaceX uses an over-dispersed spatial Poisson model coupled with a high-dimensional factor model which is based on a dimension reduction technique for computational efficiency. We show via simulations, accuracy gains in co-expression network estimation and structure by accounting for (increasing) spatial correlation and appropriate noise distributions. In-depth analysis of two spatial transcriptomics datasets in mouse hypothalamus and human breast cancer using SpaceX, detected multiple hub genes which are related to cognitive abilities for the hypothalamus data and multiple cancer genes (e.g. collagen family) from the tumor region for the breast cancer data.

Availability and implementationThe SpaceX R-package is available at github.com/bayesrx/SpaceX.

Contactxzhousph@umich.edu, veerab@umich.edu

Supplementary informationSupplementary data are available at bookdown.org/satwik91/SpaceX_supplementary/.


title: How much do model organism phenotypes contribute to the computational identification of human disease genes?

abstract: Computing phenotypic similarity has been shown to be useful in identification of new disease genes and for rare disease diagnostic support. Genotype-phenotype data from orthologous genes in model organisms can compensate for lack of human data to greatly increase genome coverage. Work over the past decade has demonstrated the power of cross-species phenotype comparisons, and several cross-species phenotype ontologies have been developed for this purpose. The relative contribution of different model organisms to identifying diseaseassociated genes using computational approaches is not yet fully explored. We use methods based on phenotype ontologies to semantically relate phenotypes resulting from loss-of-function mutations in different model organisms to disease-associated phenotypes in humans. Semantic machine learning methods are used to measure how much different model organisms contribute to the identification of known human gene-disease associations. We find that only mouse phenotypes can accurately predict human gene-disease associations. Our work has implications for the future development of integrated phenotype ontologies, as well as for the use of model organism phenotypes in human genetic variant interpretation.


title: GenOtoScope: Towards automating ACMG classification of variants associated with congenital hearing loss

abstract:
Since next-generation sequencing (NGS) has become widely available, large gene panels containing up to several hundred genes can be sequenced cost-efficiently. However, the interpretation of the often large numbers of sequence variants detected when using NGS is laborious, prone to errors and often not comparable across laboratories. To overcome this challenge, the American College of Medical Genetics and Genomics and the Association for Molecular Pathology (ACMG/AMP) introduced standards and guidelines for the interpretation of sequencing variants. Further gene- and disease-specific refinements regarding hereditary hearing loss have been developed since then. With more than 200 genes associated with hearing disorders, the manual inspection of possible causative variants is especially difficult and time consuming. We developed an open-source bioinformatics tool GenOtoScope, which automates all ACMG/AMP criteria that can be assessed without further individual patient information or human curator investigation, including the refined loss of function criterion ("PVS1"). Two types of interfaces are provided: (i) a command line application to classify sequence variants in batches for a set of patients and (ii) a user-friendly website to classify single variants. We compared the performance of our tool with two other variant classification tools using two hearing loss data sets, which were manually annotated either by the ClinGen Hearing Loss Gene Curation Expert Panel or the diagnostics unit of our human genetics department. GenOtoScope achieved the best average accuracy and precision for both data sets. Compared to the second-best tool, GenOtoScope improved accuracy metric by 25.75% and 4.57% and precision metric by 52.11% and 12.13% on the two data sets respectively. The web interface is freely accessible. The command line application along with all source code, documentation and example outputs can be found via the project GitHub page.

Author summaryNew high-throughput sequencing technologies can produce massive amounts of information and are utilized by laboratories to explain the often complex genetic aetiology of hereditary diseases. The most common sensory disease, hearing loss, is often hereditary and has a high impact on a patients every-day life. To use these sequencing technologies effectively, software tools were developed that can aid researchers interpreting genetic data by semi-automatically classifying the biologic (and thus potentially medical) impact of detected variants (the alterations of the patients genome compared to the human reference genome). The available genetic variant classification tools are either not designed specifically for the interpretation of variants detected in subjects with hearing loss or they do not allow researchers to use them for batch classification of all variants detected, e. g. in a study group. To address this drawback, we developed GenOtoScope, an open-source tool that automates the pathogenicity classification of variants potentially associated with congenital hearing loss. GenOtoScope can be applied for the automatic classification of all variants detected in a set of probands.


title: Small Molecules Targeting the Disordered Transactivation Domain of the Androgen Receptor Induce the Formation of Collapsed Helical States

abstract:
Castration-resistant prostate cancer (CRPC) is a lethal condition suffered by [~]35% of prostate cancer patients who become resistant to existing FDA-approved drugs. Small molecules that target the intrinsically disordered N-terminal domain of the androgen receptor (AR-NTD) have shown promise in circumventing CPRC drug-resistance. A prodrug of one such compound, EPI-002, entered human trials in 2015 but was discontinued after phase I due to poor potency. The compound EPI-7170 was subsequently found to have improved potency, and a related compound entered human trials in 2020. NMR measurements have localized the strongest effects of these compounds to a transiently helical region of the disordered AR-NTD but no detailed structural or mechanistic rationale exists to explain their affinity to this region or the comparative potency of EPI-7170. Here, we utilize all-atom molecular dynamics simulations to elucidate the binding mechanisms of the small molecules EPI-002 and EPI-7170 to the disordered AR-NTD. We observe that both compounds induce the formation of collapsed helical states in the Tau-5 transactivation domain and that these bound states consist of heterogenous ensembles of interconverting binding modes. We find that EPI-7170 has a higher affinity to Tau-5 than EPI-002 and that the EPI-7170 bound ensemble contains a substantially higher population of collapsed helical states than the bound ensemble of EPI-002. We identify a network of interactions in the EPI-7170 bound ensemble that stabilize collapsed helical conformations. Our results provide atomically detailed binding mechanisms for EPI compounds consistent with NMR experiments that will prove useful for drug discovery for CRPC.

SummaryIntrinsically disordered proteins (IDPs), which do not fold into a well-defined three-dimensional structure under physiological conditions, are implicated in many human diseases. Such proteins are difficult to characterize at an atomic level and are extremely challenging drug targets. Small molecules that target a disordered domain of the androgen receptor have entered human trials for the treatment of castration-resistant prostate cancer, but no structural or mechanistic rationale exists to explain their inhibition mechanisms or relative potencies. Here, we utilize molecular dynamics computer simulations to elucidate atomically detailed binding mechanisms of these compounds and understand their inhibition mechanisms. Our results suggest strategies for developing more potent androgen receptor inhibitors and general strategies for IDP drug design.

category: biophysics


title: Fusion protein strategies for cryo-EM study of G protein-coupled receptors

abstract: Single particle cryogenic-electron microscopy (cryo-EM) is used extensively to determine structures of activated G protein-coupled receptors (GPCRs) in complex with G proteins or arrestins. However, applying it to GPCRs without signaling proteins remains challenging because most receptors lack structural features in their soluble domains to facilitate image alignment. In GPCR crystallography, inserting a fusion protein between transmembrane helices 5 and 6 is a highly successful strategy for crystallization. Although the similar strategy has the potential to broadly facilitate cryo-EM structure determination of GPCRs alone without signaling protein, the critical determinants that make this approach successful are not yet clear. Here, we address this shortcoming by exploring different fusion protein designs, which led to structures of antagonist bound A2A adenosine receptor at 3.4[A] resolution and unliganded Smoothened at 3.7[A] resolution. The fusion strategies explored here are likely applicable to cryo-EM interrogation of other GPCRs and small integral membrane proteins.


title: Anticancer immunotherapies transition postcapillary venules into high-endothelial venules that generate TCF1+ T lymphocyte niches through a feed-forward loop

abstract:
The lack of T-cell infiltrates is a major obstacle to effective immunotherapy in cancer. Conversely, the formation of tumor-associated tertiary-lymphoid-like structures (TA-TLS), which are the local site of humoral and cellular immune responses against cancers, are associated with good prognosis and have recently been detected in Immune Checkpoint Blockade (ICB)-responding patients. However, how these lymphoid aggregates develop remains poorly understood. By employing scRNA sequencing, endothelial fate mapping, and functional multiplex immune profiling, we demonstrate that antiangiogenic immune-modulating therapies evoke the transition of postcapillary venules into inflamed high endothelial venules (HEVs), which generate permissive TA-TLS-like lymphocyte niches with PD1neg and PD1+TCF1+CD8 T cell progenitors that differentiate into GrzB+TCF1neg TIM3+ PD1+ CD8 T effector cells. Tumor-HEVs require continuous CD8 and NK cell-derived lymphotoxin signals revealing that tumor-HEV maintenance is actively sculpted by the adaptive immune system through a feed-forward loop.

In BriefHua & Vella et al. reveal that effective antiangiogenic immunotherapy transitions postcapillary venules into inflamed high-endothelial venules (HEV), sustained by CD8 T and NK cell-derived signals through a feed-forward loop. Thereby, tumoral HEVs establish perivascular niches in which TCF1+ PD1+ lymphocytes expand and produce cytolytic PD1+ TIM3+ CD8 T cells that facilitate anti-tumoral immunity.

HighlightsO_LIHigh endothelial venule induction by anticancer immunotherapies generates perivascular immune niches permissive for TCF1+ PD1+ CD8 progenitor T cell expansion and production of TCF1neg PD1+ TIM3+ CD8 effector T cells
C_LIO_LITumoral high-endothelial venules exhibit characteristics of inflamed lymph node HEVs and postcapillary venules
C_LIO_LIPostcapillary venules dynamically transdifferentiate into high-endothelial venules in tumors, which requires continuous signals from surrounding immune cells
C_LIO_LICD8 and NK cells drive tumoral high-endothelial venule formation during antiangiogenic immunotherapies in a feed-forward loop via lymphotoxin beta receptor signaling
C_LI

category: cancer biology


title: Conserved angio-immune subtypes of the cancer microenvironment predict response to immune checkpoint blockade therapy

abstract: Tumor microenvironment (TME) shapes the tumor progression and therapy outcome. Particularly, tumor angiogenesis and immunity impact the effect of immune checkpoint blockade (ICB) therapy. Here, we analyzed the transcriptome from 11,069 patients from The Cancer Genome Atlas (TCGA) to assess 91 functional gene sets corresponding to endothelial and T-cell activity. Intriguingly, TME across 30 non-hematological tumors can be classified into three distinct conserved angio-immune subtypes: high angiogenesis with low immune activity, low angiogenesis with high immune activity, and the one in-between. Remarkably, patients displaying TME with poor angiogenic activity with corresponding high immune activity show the most significant responses to ICB therapy in many cancer types. Notably, re-evaluation of the Javelin Renal 101, renal cell carcinoma clinical trial, provided compelling evidence that the baseline angiogenic state is critical in determining responses to checkpoint blockade. This study offers a clear rationale for incorporating baseline angiogenic state for ICB treatment decision-making.


title: Cell geometry distinguishes migration-associated heterogeneity in two-dimensional systems

abstract:
BackgroundIn vitro migration assays are a cornerstone of cell biology and have found extensive utility in research. Over the past decade, several variations of the two-dimensional (2D) migration assay have improved our understanding of this fundamental process. However, the ability of these approaches to capture the functional heterogeneity during migration and their accessibility to inexperienced users has been limited.

MethodsWe downloaded published time-lapse 2D cell migration datasets and subjected them to feature extraction with the Fiji software. We used the  Analyze Particles tool to extract ten cell geometry features (CGFs), which were grouped into  shape,  size and  position descriptors. Next, we defined the migratory status of cells using the  MTrack2 plugin. All data obtained from Fiji were further subjected to rigorous statistical analysis with R version 4.0.2.

ResultsWe observed consistent associative trends between size and shape descriptors and validated the robustness of our observations across four independent datasets. We used these descriptors to resolve the functional heterogeneity during migration by identifying and characterizing  non-migrators (NM) and  migrators (M). Statistical analysis allowed us to identify considerable heterogeneity in the NM subset, that has not been previously reported. Interestingly, differences in 2D-packing appeared to affect CGF trends and heterogeneity of the migratory subsets for the datasets under investigation.

ConclusionWe developed an analytical pipeline using open source tools, to identify and morphologically characterize functional migratory subsets from label-free, time-lapse migration data. Our quantitative approach identified a previously unappreciated heterogeneity of non-migratory cells and predicted the influence of 2D-packing on migration.


title: Evaluating mono and combination therapy of meropenem and amikacin against Pseudomonas aeruginosa bacteremia in the Hollow-Fiber Infection Model

abstract: Debate continues as to the role of combination antibiotic therapy for the management of Pseudomonas aeruginosa infections. We studied extent of bacterial killing and resistance emergence of meropenem and amikacin as monotherapy and as a combination therapy against susceptible and resistant P. aeruginosa isolates from bacteremic patients using the dynamic in vitro hollow-fiber infection model. Three P. aeruginosa isolates (meropenem MICs 0.125, 0.25 & 64 mg/L) were used simulating bacteremia with an initial inoculum ~1x105 CFU/mL and the expected pharmacokinetics of meropenem and amikacin in critically ill patients. For isolates susceptible to amikacin and meropenem (isolates 1 and 2), the rate of bacterial killing was increased with the combination regimen when compared with monotherapy of either antibiotic. Both the combination and meropenem monotherapy were able to sustain bacterial killing throughout the seven-day treatment course, whereas regrowth of bacteria occurred with amikacin monotherapy after 12 hours. For the meropenem-resistant P. aeruginosa isolate (isolate 3), only the combination regimen demonstrated bacterial killing. Given that tailored antibiotic regimens can maximize potential synergy against some isolates, future studies should explore the benefit of combination therapy against resistant P. aeruginosa.


title: A quadratic analytical solution of root pressure generation provides insights about bamboo and other species

abstract:
We derived a steady-state model of whole root pressure generation through the combined action of all parallel segments of fine roots. This may be the first complete analytical solution for root pressure, which can be applied to complex roots/shoots.

The osmotic volume of a single root is equal to that of the vessel lumen in fine roots and adjacent apoplastic spaces. Water uptake occurs via passive osmosis and active solute uptake ([Formula], osmol s-1), resulting in the osmolal concentration Cr (mol{middle dot}kg-1 of water) at a fixed osmotic volume. Solute loss occurs via two passive processes: radial diffusion of solute Km (Cr-Csoil), where Km is the diffusional constant and Csoil is the soil-solute concentration) from fine roots to soil and mass flow of solute and water into the whole plant from the end of the fine roots.

The proposed model predicts the quadratic function of root pressure [Formula], where b and c are the functions of plant hydraulic resistance, soil water potential, solute flux, and gravitational potential.

The present study investigates the theoretical dependencies of Pr on the factors detailed above and demonstrates the root pressure-mediated distribution of water through the hydraulic architecture of a 6.8-m-tall bamboo shoot.

One sentence SummaryAn analytical solution for root pressure is derived and illustrated by applying it to the measured hydraulic architecture of a 7-meter bamboo shoot.

category: physiology


title: HSFA1 proteins mediate heat-induced accumulation of CPT7-derived polyprenols affecting thylakoid organization

abstract:
Polyprenols are ubiquitous isoprenoid compounds that accumulate in large quantities in plant photosynthetic tissues. While our knowledge of polyprenol biochemistry is constantly expanding, the regulation of their biosynthesis as well as the molecular basis of their cellular action are still poorly understood. In Arabidopsis, the polyprenols Pren-9, -10 and -11, synthesized by cis-prenyltransferase 7 (CPT7), are localized in plastidial membranes and affect the photosynthetic performance of chloroplasts.

In this report we present evidence that plastidial polyprenols are among the major constituents of thylakoid membranes. Disturbances in polyprenol level, caused by alterations in CPT7 expression, change chloroplast ultrastructure, affect aggregation of LHCII complexes and modulate non-photochemical quenching (NPQ). Moreover, we show that Arabidopsis responds to high temperature by upregulating expression of CPT7 and increasing the accumulation of CPT7-derived polyprenols. These heat-induced changes in polyprenol biosynthesis are mediated by Heat Shock Transcription Factors of the HSFA1 family, the master regulators of heat stress response. Collectively, results presented in this report bring us closer to understanding the mechanisms by which polyprenols affect plant physiology and provide an additional link between chloroplast biology and plant responses to changing environmental conditions.

One sentence summaryHeat Shock Transcription Factors induce biosynthesis of polyprenols - isoprenoid compounds that affect the organization and function of chloroplasts.

category: plant biology


title: COVID-19: Salient Aspects of Coronavirus Infection, Vaccines and Vaccination Testing and their Implications

abstract:
In the present study, three basic aspects related to COVID-19 are presented.

O_LIThe occurrence of coronavirus infection is analyzed statistically as number of coronaviruses infected alveolar cells compared to normal alveolar cells in human lungs. The mole concept is used to estimate the number of normal alveolar cells per human lung. The number of coronavirus infections in infected alveolar cells is estimated from the published Lower Respiratory Tract (LRT) load data. The Poisson probability distribution is aptly applied to imply the incubation period of the coronavirus infection to be within day-3 to day-7, with the cumulative probability of 75%. The incubation period within day-0 to day-10 has a cumulative probability of 98%. It implies a 10-day quarantine to isolate an uninfected individual as a precautionary measure.
C_LIO_LIThree vaccines to combat COVID-19, which adopt distinct paradigms while preparing them, are analyzed. These are Modernas mRNA-1273, Oxford-AstraZenecas ChAdOx1 nCoV-19 and Bharat BioTechs COVAXIN. The mole concept is used to estimate the antigen mass density per dose of each of these vaccines as 10 g cm-3, 0.1 g cm-3 and 1 g cm-3, respectively. The vaccines are deemed to be compatible to neutralize the infection.
C_LIO_LIA statistical analysis is performed of the Modernas mRNA-1273 vaccine efficacy of 94.1% and Oxfords ChAdOx1 nCoV-19 vaccine efficacy of 62.1% in terms of groups of volunteers testing negative to vaccine by chance. In the Moderna vaccination testing scenario, since the probability of negative response of vaccine is small, the Poisson probability distribution for 95% cumulative probability is used to describe the vaccination testing in 300 samples of 47 volunteers each. Thus, 87% of samples have average group of 3 volunteers testing negative to vaccine. About 6% of samples have all volunteers testing positive to vaccine. In the Oxford vaccination testing scenario, since the probability of negative response of vaccine is finite, the Gaussian probability distribution for 95% probability is used to describe the vaccination testing in 75 samples of 120 volunteers each. Thus, 68% of samples have average group of 45 volunteers testing negative to vaccine. No sample has all volunteers testing positive to vaccine. A vaccine, irrespective of its efficacy being high or low, is necessary for mass immunization.
C_LI

category: scientific communication and education


title: The development of bi-directionally coupled self-organizing neurovascular networks captures orientation-selective neural and hemodynamic cortical responses

abstract: The network of neurons in the brain is considered the primary substrate of information processing. Despite growing evidence on the possible role of cerebral blood flow in information processing, the cerebrovascular network is generally viewed as an irrigation system that ensures a timely supply of oxygen, glucose, and nutrients to the neural tissue. However, a recent study has shown that cerebral microvessels, like neurons, also exhibit tuned responses to sensory stimuli. Tuned neural responses to sensory stimuli are certainly enhanced with experience-dependent Hebbian plasticity and other forms of learning. Hence it is possible that the densely interconnected microvascular network might also be subject to some form of plasticity or competitive learning rules during early postnatal development such that its fine-scale structure becomes optimized for metabolic delivery to a given neural micro-architecture. To explore the possibility of adaptive lateral interactions and tuned responses in cerebral microvessels, we modeled the cortical neurovascular network by interconnecting two laterally connected self-organizing networks (Laterally Interconnected Synergetically Self-Organizing Map - LISSOM). The afferent and lateral connections of the LISSOM were defined by trainable weights. By varying the topology of lateral connectivity in the vascular network layer, we observed that the partial correspondence of feature selectivity between neural and hemodynamic responses could be explained by lateral coupling across local blood vessels such that the central domain receives an excitatory drive of more blood flow and a more distal surrounding region where blood flow is reduced. Critically, our simulations suggest a new role for feedback from the vascular to the neural network because the radius of vascular perfusion seems to determine whether the cortical neural map develops into a clustered and columnar vs. salt-and-pepper organization.


title: Excess interictal activity marks seizure prone cortical areas and mice in a genetic epilepsy model

abstract:
Genetic epilepsies are often caused by variants in widely expressed genes, potentially impacting numerous brain regions and functions. For instance, gain-of-function (GOF) variants in the widely expressed Na+-activated K+ channel gene KCNT1 alter basic neurophysiological and synaptic properties of cortical neurons, leading to developmental epileptic encephalopathy. Yet, aside from causing seizures, little is known about how such variants reshape interictal brain activity, and how this relates to epileptic activity and other disease symptoms.

To address this knowledge gap, we monitored neural activity across the dorsal cortex in a mouse model of human KCNT1-related epilepsy using in vivo, awake widefield Ca2+ imaging. We observed 52 spontaneous seizures and 1700 interictal epileptiform discharges (IEDs) in homozygous mutant (Kcnt1m/m) mice, allowing us to map their appearance and spread at high spatial resolution. Outside of seizures and IEDs, we detected ~46,000 events, representing interictal cortical activity, in both Kcnt1m/m and wild-type (WT) mice, and we classified them according to their spatial profiles.

Spontaneous seizures and IEDs emerged within a consistent set of susceptible cortical areas, and seizures propagated both contiguously and non-contiguously within these areas in a manner influenced, but not fully determined, by underlying synaptic connectivity. Seizure emergence was predicted by a progressive concentration of total cortical activity within the impending seizure emergence zone. Outside of seizures and IEDs, similar events were detected in WT and Kcnt1m/m mice, suggesting that the spatial structure of interictal activity was largely preserved. Several features of these events, however, were altered in Kcnt1m/m mice. Most event types were briefer, and their intensity more variable, across Kcnt1m/m mice; mice showing more intense activity spent more time in seizure. Furthermore, the rate of events whose spatial profile overlapped with where seizures and IEDs emerged was increased in Kcnt1m/m mice.

Taken together, these results demonstrate that an epilepsy-causing K+ channel variant broadly alters physiology. Yet, outside of seizures and IEDs, it acts not to produce novel types of cortical activity, but rather to modulate its amount. The areas where seizures and IEDs emerge show excessively frequent and intense interictal activity and the mean intensity of an individuals cortical activity predicts its seizure burden. These findings provide critical guidance for targeting future research and therapy development.


title: Multiplex, translaminar imaging in the spinal cord of behaving mice

abstract: While the spinal cord is known to play critical roles in sensorimotor processing, including pain-related signaling, corresponding activity patterns in genetically defined cell types across spinal laminae have remained elusive. Calcium imaging has enabled cellular activity measurements in behaving rodents but is currently limited to superficial regions. Using chronically implanted microprisms, we imaged sensory and motor evoked activity in regions and at speeds inaccessible by other high-resolution imaging techniques. To enable translaminar imaging in freely behaving animals through implanted microprisms, we additionally developed wearable microscopes with custom-compound microlenses. This new integrated system addresses multiple challenges of previous wearable microscopes, including their limited working distance, resolution, contrast, and achromatic range. The combination of these innovations allowed us to uncover that dorsal horn astrocytes in behaving mice show somatosensory program-dependent and lamina-specific calcium excitation. Additionally, we show that tachykinin precursor 1 (Tac1)-expressing neurons exhibit upper laminae-restricted activity to acute mechanical pain but not locomotion.


title: Pattern and component responses of primate MT neurons recorded with multi-contact electrodes, stimulated with 1D and 2D noise patterns.

abstract:
Direction selective neurons in primary visual cortex (area V1) are affected by the aperture problem, i.e., they are only sensitive to motion orthogonal to their preferred orientation. A solution to this problem first emerges in the middle temporal (MT) area, where a subset of neurons (called pattern cells) combine motion information across multiple orientations and directions, becoming sensitive to pattern motion direction. These cells are expected to play a prominent role in subsequent neural processing, but they are intermixed with cells that behave like V1 cells (component cells), and others that do not clearly fall in either group. The picture is further complicated by the finding that cells that behave like pattern cells with one type of pattern, might behave like component cells for another.

We recorded from macaque MT neurons using multi-contact electrodes while presenting both type I and unikinetic plaids, in which the components were 1D noise patterns. We found that the indices that have been used in the past to classify neurons as pattern or component cells work poorly when the properties of the stimulus are not optimized for the cell being recorded, as is always the case with multi-contact arrays. We thus propose alternative measures, which considerably ameliorate the problem, and allow us to gain insights in the signals carried by individual MT neurons.

We conclude that arranging cells along a component-to-pattern continuum is an oversimplification, and that the signals carried by individual cells only make sense when embodied in larger populations.


title: A personalizable autonomous neural mass model of epileptic seizures

abstract: Work in the last two decades has shown that neural mass models (NMM) can realistically reproduce and explain epileptic seizure transitions as recorded by electrophysiological methods (EEG, SEEG). In previous work, advances were achieved by increasing excitation and heuristically varying network inhibitory coupling parameters in the models. Based on these early studies, we provide a laminar NMM capable of realistically reproducing the electrical activity recorded by SEEG in the epileptogenic zone during interictal to ictal states. With the exception of the external noise input into the pyramidal cell population, the model dynamics are autonomous. By setting the system at a point close to bifurcation, seizure-like transitions are generated, including pre-ictal spikes, low voltage fast activity, and ictal rhythmic activity. A novel element in the model is a physiologically motivated algorithm for chloride dynamics: the gain of GABAergic post-synaptic potentials is modulated by the pathological accumulation of chloride in pyramidal cells due to high inhibitory input and/or dysfunctional chloride transport. In addition, in order to simulate SEEG signals for comparison with real seizure recordings, the NMM is embedded first in a layered model of the neocortex and then in a realistic physical model. We compare modelling results with data from four epilepsy patient cases. By including key pathophysiological mechanisms, the proposed framework captures succinctly the electrophysiological phenomenology observed in ictal states, paving the way for robust personalization methods based on NMMs.


title: Enhanced learning and sensory salience in a cerebellar mouse autism model

abstract: Among the impairments manifested by autism spectrum disorder (ASD) are sometimes islands of enhanced function1,2. Although neuronal mechanisms for enhanced functions in ASD are unknown, the cerebellum is a major site of developmental alteration, and early-life perturbation to it leads to ASD with higher likelihood than any other brain region3,4. Here we report that a cerebellum-specific transgenic mouse model of ASD shows faster learning on a sensory evidence-accumulation task5. In addition, transgenic mice showed enhanced sensitivity to touch and auditory cues, and prolonged electrophysiological responses in Purkinje-cell complex spikes and associative neocortical regions. These findings were replicated by pairing cues with optogenetic stimulation of Purkinje cells. Computational latent-state analysis of behavior6-8 revealed that both groups of mice with cerebellar perturbations exhibited enhanced focus on current rather than past information, consistent with a role for the cerebellum in retaining information in memory9. We conclude that cerebellar perturbation can activate neocortex via complex spike activity and reduce reliance on prior experience, consistent with a weak-central-coherence account in which ASD traits arise from enhanced detail-oriented processing1. This recasts ASD not so much as a disorder but as a variation that, in particular niches, can be adaptive.


title: Pharmacological blockade of muscle afferents and perception of effort: a systematic review with meta-analysis

abstract:
BackgroundThe perception of effort (PE) provides information on task difficulty and influences physical exercise regulation and human behavior. This perception differs from other-exercise related perceptions such as pain. There is no consensus on the role of group III-IV muscle afferents as a signal processed by the brain to generate PE.

ObjectiveThe aim of this meta-analysis was to investigate the effect of pharmacologically blocking muscle afferents on the PE.

MethodsSix databases were searched to identify studies measuring the ratings of perceived effort (RPE) during physical exercise, with and without pharmacological blockade of muscle afferents. Articles were coded based on the operational measurement used to distinguish studies in which PE was assessed specifically (effort dissociated) or as a composite experience including other exercise-related perceptions (effort not dissociated). Articles that did not provide enough information for coding were assigned to the unclear group.

ResultsThe effort dissociated group (n=6) demonstrated a slight RPE increase with reduced muscle afferents feedback (standard mean change raw (SMCR), 0.39; 95%CI, 0.13 to 0.64). The group effort not dissociated (n=2) did not reveal conclusive results (SMCR, -0.29; 95%CI, -2.39 to 1.8). The group unclear (n=8) revealed a slight RPE decrease with reduced muscle afferents feedback (SMCR, -0.27; 95%CI, -0.50 to -0.04).

ConclusionsThe heterogeneity in results between groups reveals that the inclusion of perceptions other than effort in its rating influences the RPE scores reported by the participants. The absence of decreased RPE in the effort dissociated group suggests that muscle afferents feedback is not a sensory signal of PE.

Key pointsO_LITo date, there is no consensus on the neurophysiological signal processed by the brain to generate the perception of effort.
C_LIO_LIFollowing a systematic search in six databases, this meta-analysis suggests that reducing afferent feedback from the working muscles via epidural anesthesia does not reduce perception of effort.
C_LIO_LIThis systematic review suggests that afferent feedback from the working muscles is not the neurophysiological signal processed by the brain to generate the perception of effort.
C_LI


title: Toll signaling enhances mosquito antiplasmodial immunity by promoting differentiation of hemocytes to the Megacyte lineage

abstract: Activation of Toll signaling in Anopheles gambiae, by silencing Cactus, eliminates Plasmodium ookinetes by enhancing local release of hemocytes-derived microvesicles that promote activation of the mosquito complement-like system. A new effector hemocyte subpopulation of large granulocytes, the megacytes, was recently identified. We report that Cactus silencing dramatically increases the proportion of megacytes, from 5 to 79% of circulating granulocytes. Transcriptomic and morphological analysis, as well as in situ hybridization and expression of cell-specific markers, indicate that Cactus silencing triggers granulocyte differentiation into megacytes. Megacytes are very plastic cells that can extend long filopodia and tend to form clusters in vivo. Moreover, megacytes are massively recruited to the basal midgut surface in response to bacterial feeding and Plasmodium infection. We propose that Toll signaling promotes differentiation of granulocytes to the megacyte lineage, a major cellular effector of antibacterial and antiplasmodial immunity.

category: microbiology


title: PCBP1 Deficient Pigs Hold the Potential to Inhibit CSFV Infection

abstract:
Classical swine fever virus (CSFV), pathogen of classic swine fever, has caused severe economic losses worldwide. Poly (rC)-binding protein 1 (PCBP1), interacting with Npro of CSFV, plays a vital role in CSFV growth. Here, our research is the first report to generate PCBP1 knockout pigs via gene editing technology. The PCBP1 knockout pigs exhibited normal birth weight, reproductive-performance traits, and developed normally. Viral challenge results indicated that primary cells isolated from F0 and F1 generation pigs could significantly reduce CSFV infection. Additional mechanism exploration further confirmed that PCBP1 KO mediated antiviral effect is related with the activation of type I interferon. Beyond showing that gene editing strategy can be used to generate PCBP1 KO pigs, our study introduces a valuable animal model for further investigating infection mechanisms of CSFV that help to develop better antiviral solution.

ImportanceAs a negative regulator in immune modulation, the effects of PCBP1 on viral replication have been found to be valuable. Here, this study was the first report to generate PCBP1 knockout pigs with normal pregnancy rate and viability. Primary cells isolated from F0 and F1 generation PCBP1 knockout pigs could significantly reduce CSFV infection. The PCBP1 knockout pigs could be used as a natural host models for investigating the effects of PCBP1-mediating critical interactions on viral replication and helping to develop better antiviral solution.


title: An expanded toolkit for Drosophila gene tagging using synthesized homology donor constructs for CRISPR mediated homologous recombination

abstract: Previously, we described a large collection of Drosophila strains that each carry an artificial exon containing a T2AGAL4 cassette inserted in an intron of a target gene based on CRISPR-mediated homologous recombination (Lee et al., 2018). These alleles permit numerous applications and have proven to be very useful. Initially, the homologous recombination-based donor constructs had long homology arms (>500 bps) to promote precise integration of large constructs (>5kb). Recently, we showed that in vivo linearization of the donor constructs enables insertion of large artificial exons in introns using short homology arms (100-200 bps) (Kanca et al., 2019a). Shorter homology arms make it feasible to commercially synthesize homology donors and minimize the cloning steps for donor construct generation. Unfortunately, about 50% of Drosophila genes lack suitable coding introns for integration of artificial exons. Here, we report the development of new set of constructs that allow the replacement of the coding region of genes that lack suitable introns with a KozakGAL4 cassette, generating a knock-out/knock-in allele that expresses GAL4 similarly as the targeted gene. We also developed custom vector backbones to further facilitate and improve transgenesis. Synthesis of homology donor constructs in custom plasmid backbones that contain the target gene sgRNA obviates the need to inject a separate sgRNA plasmid and significantly increases the transgenesis efficiency. These upgrades will enable the targeting of nearly every fly gene, regardless of exon-intron structure, with a 70-80% success rate.


title: Structure, dynamics, and inhibition of Staphylococcus aureus m1A22-tRNA methyltransferase

abstract: The enzyme m1A22-tRNA methyltransferase (TrmK) catalyses the transfer of a methyl group from SAM to the N1 of adenine 22 in tRNAs. TrmK is essential for Staphylococcus aureus survival during infection, but has no homologue in mammals, making it a promising target for antibiotic development. Here we describe the structural and functional characterisation of S. aureus TrmK. Crystal structures are reported for S. aureus TrmK apoenzyme and in complexes with SAM and SAH. Isothermal titration calorimetry showed that SAM binds to the enzyme with favourable but modest enthalpic and entropic contributions, whereas SAH binding leads to an entropic penalty compensated by a large favourable enthalpic contribution. Molecular dynamics simulations point to specific motions of the C-terminal domain being altered by SAM binding, which might have implications for tRNA recruitment. Activity assays for S. aureus TrmK-catalysed methylation of WT and position 22 mutants of tRNALeu demonstrate that the enzyme requires an adenine at position 22 of the tRNA. Intriguingly, a small RNA hairpin of 18 nucleotides is methylated by TrmK depending on the position of the adenine. In-silico screening of compounds suggested plumbagin as a potential inhibitor of TrmK, which was confirmed by activity measurements. Furthermore, LC-MS indicated the protein was covalently modified by one equivalent of the inhibitor, and proteolytic digestion coupled with LC-MS identified Cys92, in the vicinity of the SAM-binding site, as the sole residue modified. These results these results identify a cryptic binding pocket of S. aureus TrmK and lay the foundation for future structure-based drug discovery.

category: biochemistry


title: MAFFIN: Metabolomics Sample Normalization Using Maximal Density Fold Change with High-Quality Metabolic Features and Corrected Signal Intensities

abstract:
Sample normalization is a critical step in metabolomics to remove differences in total sample amount or concentration of metabolites between biological samples. Here, we present MAFFIN, an accurate and robust post-acquisition sample normalization workflow that works universally for metabolomics data collected by mass spectrometry (MS)-based platforms. The most important design of MAFFIN is the calculation of normalization factor using maximal density fold change (MDFC) value computed by a kernel density-based approach. MDFC is more accurate than traditional median FC-based normalization, especially when the numbers of up- and down-regulated metabolic features are different. In addition, we showcase two essential steps that are overlooked by conventional normalization methods, and incorporated them into MAFFIN. First, instead of using all detected metabolic features, MAFFIN automatically extracts and uses only the high-quality features to calculate FCs and determine the normalization factor. In particular, multiple orthogonal criteria are proposed to pick up the high-quality features. Second, to guarantee the accuracy of the FCs, the MS signal intensities of the high-quality features are corrected using serial quality control (QC) samples. Using simulated data and urine metabolomics datasets, we demonstrated the critical need of high-quality feature selection, MS signal correction, and MDFC. We also show the superior performance of MAFFIN over other commonly used post-acquisition sample normalization methods. Finally, a biological application on a human saliva metabolomics study shows that MAFFIN provides robust sample normalization, leading to better data separation in principal component analysis (PCA) and the identification of more significantly altered metabolic features.

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title: MEMO: Mass Spectrometry-based Sample Vectorization to Explore Chemodiverse Datasets

abstract:
In natural products research, chemodiverse extracts coming from multiple organisms are explored for novel bioactive molecules, sometimes over extended periods. Samples are usually analyzed by liquid chromatography coupled with fragmentation mass spectrometry to acquire informative mass spectral ensembles. Such data is then exploited to establish relationships among analytes or samples (e.g. via molecular networking) and annotate metabolites. However, the comparison of samples profiled in different batches is challenging with current metabolomics methods. Indeed, the experimental variation - changes in chromatographical or mass spectrometric conditions - often hinders the direct comparison of the profiled samples. Here we introduce MEMO - MS2 BasEd SaMple VectOrization - a method allowing to cluster large amounts of chemodiverse samples based on their LC-MS/MS profiles in a retention time agnostic manner. This method is particularly suited for heterogeneous and chemodiverse sample sets. MEMO demonstrated similar clustering performance as state-of-the-art metrics taking into account fragmentation spectra. More importantly, such performance was achieved without the requirement of a prior feature alignment step and in a significantly shorter computational time. MEMO thus allows the comparison of vast ensembles of samples, even when analyzed over long periods of time, and on different chromatographic or mass spectrometry platforms. This new addition to the computational metabolomics toolbox should drastically expand the scope of large-scale comparative analysis.

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title: Single cell RNA-seq identifies developing corneal cell fates in the human cornea organoid

abstract:
The cornea is a protective and refractive barrier in the eye crucial for vision. Understanding the human cornea in health, disease and cell-based treatments can be greatly advanced with cornea organoids developed in culture from induced pluripotent stem cells. While a limited number of studies have investigated the single-cell transcriptomic composition of the human cornea, its organoids have not been examined similarly. Here we elucidated the transcriptomic cell fate map of 4 month-old human cornea organoids and the central cornea from three donors. The organoids harbor cell clusters representing corneal epithelium, stroma and endothelium with sub populations that capture signatures of early developmental states. Unlike the adult cornea where the largest cell population is stromal, the organoids develop almost equal proportion of the three major cell types. These corneal organoids offer a three-dimensional platform to model corneal diseases and integrated responses of the different cell types to treatments.

TeaserTranscriptomic Map of Cornea Organoid and Human Cornea

category: developmental biology


title: Monospecific and bispecific monoclonal SARS-CoV-2 neutralizing antibodies that maintain potency against B.1.617

abstract: COVID-19 pathogen SARS-CoV-2 has infected hundreds of millions and caused over 5 million deaths to date. Although multiple vaccines are available, breakthrough infections occur especially by emerging variants. Effective therapeutic options such as monoclonal antibodies (mAbs) are still critical. Here, we report the development, cryo-EM structures, and functional analyses of mAbs that potently neutralize SARS-CoV-2 variants of concern. By high-throughput single cell sequencing of B cells from spike receptor binding domain (RBD) immunized animals, we identified two highly potent SARS-CoV-2 neutralizing mAb clones that have single-digit nanomolar affinity and low-picomolar avidity, and generated a bispecific antibody. Lead antibodies showed strong inhibitory activity against historical SARS-CoV-2 and several emerging variants of concern. We solved several cryo-EM structures at [~]3 [A] resolution of these neutralizing antibodies in complex with prefusion spike trimer ectodomain, and revealed distinct epitopes, binding patterns, and conformations. The lead clones also showed potent efficacy in vivo against authentic SARS-CoV-2 in both prophylactic and therapeutic settings. We also generated and characterized a humanized antibody to facilitate translation and drug development. The humanized clone also has strong potency against both the original virus and the B.1.617.2 Delta variant. These mAbs expand the repertoire of therapeutics against SARS-CoV-2 and emerging variants.

category: immunology


title: Chaperoning of the histone octamer by the acidic domain of DNA repair factor APLF

abstract: Nucleosome assembly requires the coordinated deposition of histone complexes H3-H4 and H2A-H2B to form a histone octamer on DNA. In the current paradigm, specific histone chaperones guide the deposition of first H3-H4 and then H2A-H2B(1-5). Here, we show that the acidic domain of DNA repair factor APLF (APLFAD) can assemble the histone octamer in a single step, and deposit it on DNA to form nucleosomes. The crystal structure of the APLFAD-histone octamer complex shows that APLFAD tethers the histones in their nucleosomal conformation. Mutations of key aromatic anchor residues in APLFAD affect chaperone activity in vitro and in cells. Together, we propose that chaperoning of the histone octamer is a mechanism for histone chaperone function at sites where chromatin is temporarily disrupted.


title: Time-resolved analysis of transcription kinetics in single live mammalian cells

abstract: In eukaryotic cells, RNA polymerase II synthesizes mRNA in three stages, initiation, elongation, and termination, and numerous factors determine how quickly a gene is transcribed to produce mRNA molecules through these steps. However, there are few techniques available to measure the rate of each step in living cells, which prevents a better understanding of transcriptional regulation. Here, we present a quantitative analysis method to extract kinetic rates of transcription from time-lapse imaging data of fluorescently labeled mRNA in live cells. Using embryonic fibroblasts cultured from two knock-in mouse models, we monitored transcription of {beta}-actin and Arc mRNA labeled with MS2 and PP7 stem-loop systems, respectively. After inhibiting transcription initiation, we measured the elongation rate and the termination time by fitting the time trace of transcription intensity with a mathematical model function. We validated our results by comparing them with steady-state fluctuation analysis and stochastic simulations. This live-cell transcription analysis method will be useful for studying the regulation of elongation and termination steps and may provide insight into the diverse mechanisms of transcriptional processes.


title: An early Cambrian polyp reveals an anemone-like ancestor for medusozoan cnidarians

abstract: Extant cnidarians are a disparate phylum of non-bilaterians and their diploblastic body plan represents a key step in animal evolution. Anthozoans (anemones, corals) are benthic polyps, while adult medusozoans (jellyfishes) are dominantly pelagic medusae. A sessile polyp is present in both groups and is widely conceived as the ancestral form of their last common ancestor. However, the nature and anatomy of this ancestral polyp, particularly of medusozoans, are controversial, owing to the divergent body plans of both groups in the extant lineages and the rarity of medusozoan soft tissues in the fossil record. Here we redescribe the enigmatic Conicula striata Luo et Hu from the early Cambrian Chengjiang biota, south China, which has previously been interpreted as a polyp, lophophorate or deuterostome. We show that C. striata possessed features of both anthozoans and medusozoans. Its stalked polyp and fully encasing conical, annulated organic skeleton (periderm) are features of medusozoans. However, the gut is partitioned by [~]28 mesenteries, and has a tubular pharynx, resembling anthozoans. Our phylogenetic analysis recovers C. striata as a stem medusozoan, indicating that the enormously diverse medusozoans were derived from an anemone-like ancestor, with the pharynx lost and number of mesenteries reduced prior to the origin of crown group Medusozoa.

category: paleontology


title: Hypodopaminergic state of the nigrostriatal pathway drives compulsive alcohol use

abstract: The neurobiological mechanisms underlying compulsive alcohol use, a cardinal feature of alcohol use disorder, remain elusive. The key modulator of motivational processes dopamine (DA) is suspected to play an important role in this pathology, but its exact implication remains to be determined. Here, we found that rats expressing compulsive alcohol-related behavior, operationalized as punishment-resistant self-administration, showed a decrease in DA levels restricted to the dorsolateral territories of the striatum, the main output structure of the nigrostriatal DA pathway. We then causally demonstrated that a chemogenetic-induced selective hypodopaminergia of this pathway results in compulsive alcohol self-administration in rats otherwise resilient, accompanied by the emergence of alcohol withdrawal-like motivational impairments. These results demonstrate a major implication of tonic nigrostriatal hypodopaminergic state in alcohol addiction and provide new insights into our understanding of the neurobiological mechanisms underlying compulsive alcohol use.


title: Neural correlates of individual odor preference in Drosophila

abstract: Behavior varies even among genetically identical animals raised in the same environment1-10. However, little is known about the circuit or anatomical underpinnings of this individuality11,12. Drosophila olfaction is an ideal system for discovering the origins of behavioral individuality among genetically identical individuals. The fly olfactory circuit is well-characterized13-16,17-19 and stereotyped15, yet stable idiosyncrasies in odor preference4, neural coding4, and neural wiring20,21 are present and may be relevant to behavior. Using paired behavior and two-photon imaging measurements, we show that individual odor preferences in odor-vs-air and odor-vs-odor assays are predicted by idiosyncratic calcium dynamics in Olfactory Receptor Neurons (ORNs) and Projection Neurons (PNs), respectively. This suggests that circuit variation at the sensory periphery determines individual odor preferences. Furthermore, paired behavior and immunohistochemistry measurements reveal that variation in ORN presynaptic density also predicts odor-vs-odor preference. This point in the olfactory circuit appears to be a locus of individuality where microscale variation gives rise to idiosyncratic behavior. To unify these results, we constructed a leaky-integrate-and-fire model of 3,062 neurons in the antennal lobe. In these simulations, stochastic fluctuations at the glomerular level, like those observed in our ORN immunohistochemistry, produce variation in PN calcium responses with the same structure as we observed experimentally, the very structure that predicts idiosyncratic behavior. Thus, our results demonstrate how minute physiological and structural variations in a neural circuit may produce individual behavior, even when genetics and environment are held constant.


title: Selfee: Self-supervised Features Extraction of animal behaviors

abstract: Fast and accurately characterizing animal behaviors is crucial for neuroscience research. Deep learning models are efficiently used in laboratories for behavior analysis. However, it has not been achieved to use a fully unsupervised method to extract comprehensive and discriminative features directly from raw behavior video frames for annotation and analysis purposes. Here, we report a self-supervised feature extraction (Selfee) convolutional neural network with multiple downstream applications to process video frames of animal behavior in an end-to-end way. Visualization and classification of the extracted features (Meta-representations) validate that Selfee processes animal behaviors in a comparable way of human understanding. We demonstrate that Meta-representations can be efficiently used to detect anomalous behaviors that are indiscernible to human observation and hint in-depth analysis. Furthermore, time-series analyses of Meta-representations reveal the temporal dynamics of animal behaviors. In conclusion, we present a self-supervised learning approach to extract comprehensive and discriminative features directly from raw video recordings of animal behaviors and demonstrate its potential usage for various downstream applications.


title: New Insights into Molecular Basis Identification of Three Novel Strains of the Bacillus Subtilis Group Produce Cry Proteins Isolated from Soil Samples in Adana, Turkey

abstract:
AimsThis study aimed to analyze the evolutionary relationship between Bacillus species isolated from agricultural soil using in-silico tools.

Methods and ResultsAcross-sectional study was conducted in Adana province, in Turkey. A total of 120 Bacillus species were isolated from 80 soil samples. However, the phylogenetic tree diverged into two lineages; one belongs to B. subtilis group while the other belongs to B. cereus group. Interestingly, three native strains (SY27.1A, SY35.3A, and SY58.5A), which produce Cry proteins, shared high similarity with B. subtilis group (over 99%) and less than 95% similarity with known B. thuringiensis and other species of B. cereus group. Furthermore, 11 canonical SNPs (canSNPs) were identified in strains that belong to B. pumilus group when compared with B. subtilis reference sequences.

ConclusionsPhylogenetic analysis of 16S rRNA sequences was found valuable for differentiation between Bacillus species isolated from soil samples. In addition, SNPs analysis provided more intra-specific information in the cases of B. subtilis group.

Significance and Impact of StudyA detailed analysis was provided for the SNPs present in a conserved region of 16S rRNA gene of Bacillus species. Also, we proposed three novel Bacillus strains that produce Cry proteins and belong to B. subtilis group.


title: The Kishony Mega-Plate Experiment, a Markov Process

abstract: A correct understanding of the DNA evolution of drug resistance is critical in developing strategies for suppressing and preventing this process. The Kishony Mega-Plate Experiment demonstrates this important phenomenon that occurs in the practice of medicine, that of the evolution of drug-resistance. The evolutionary process which the bacteria in this experiment are doing is called a Markov Process or Markov Chain. Understanding this process enables clinicians and researchers to predict the evolution of drug-resistance and develop strategies to prevent this process. This paper will show how to apply the Markov Chain model of DNA evolution to the Kishony Mega-Plate Experiment and why the experiment behaves the way it does by contrasting the Jukes-Cantor model of DNA evolution (a stationary model) with a modification of the Jukes-Cantor model that makes it a non-stationary, non-equilibrium Markov Chain. The numerical behaviors of the stationary and non-stationary models are compared. What this analysis shows is that DNA evolution is a non-stationary, non-equilibrium process and that by using the correct non-stationary, non-equilibrium model that one can simulate and predict the behavior of real evolutionary examples and that these analytical tools can give the clinician guidance on how to use antimicrobial selection pressures for treating infectious diseases. This in turn can help reduce the numbers and costs of hospitalization for sepsis, pneumonia and other infectious diseases.

category: evolutionary biology


title: Genomic bases of insularity and ecological divergence in barn owls (Tyto alba) of the Canary Islands

abstract: Islands, and the particular organisms that populate them, have long fascinated biologists. Due to their isolation, islands offer unique opportunities to study the effect of neutral and adaptive mechanisms in determining genomic and phenotypical divergence. In the Canary Islands, an archipelago rich in endemics, the barn owl (Tyto alba) is thought to have diverged into a subspecies (T. a. gracilirostris) on the eastern islands, Fuerteventura and Lanzarote. Taking advantage of 40 whole-genomes and modern population genomics tools, we provide the first look at the origin and genetic makeup of barn owls of this archipelago. We show that the Canaries hold diverse, long-standing and monophyletic populations with a neat distinction of gene pools from the different islands. Using new method, less sensitive to structure than classical FST, to detect regions involved in local adaptation to the insular environment, we identified a haplotype-like region likely under positive selection in all Canaries individuals. Genes in this region suggest morphological adaptations to insularity. In the eastern islands, where the subspecies T. a. gracilirostris is present, genomic traces of selection pinpoint signs of locally adapted body proportions and blood pressure, consistent with the smaller size of this population living in a hot arid climate. In turn, genomic regions under selection in the western barn owls from Tenerife showed an enrichment in genes linked to hypoxia, a potential response to inhabiting a small island with a marked altitudinal gradient. Our results illustrate the interplay of neutral and adaptive forces in shaping divergence and early onset speciation.


title: Graphene Oxide reinforced Agarose-Hydroxyapatite Bioprinted 3-D Scaffolds for Bone Regeneration

abstract:
Three-dimensional (3D) bioprinting is an emerging technology for fabricating cells, biomaterials and extracellular matrix (ECM) into customized shapes and patterns. Here, we report additive manufacturing to create a customized 3D bioactive constructs for regenerative medicine. We have attempted to emphasize the use of agarose and graphene oxide as a promising material for the conceptualization of bioink unpaid to its unique physicochemical properties. The 3D printed structure is able to regenerating bone tissues and regulates the cellular differentiation without any significant morphological changes. The presence of graphene oxide enhances the osteoinductive behavior of the developed scaffolds, which is further supplemented by encapsulating human mesenchymal stem cells (hMSCs) on the 3D printed scaffolds.

A significant enhanced expression of early osteogenic markers like morphogenetic protein (BMP), Runx-2, collagen-1, osteopontin, osteocalcin as well as mineralized ECM are observed on agarose-hydroxyapatite and graphene oxide 3D printed scaffolds compared to agarose-hydroxyapatite 3D printed scaffolds. Thus, the outcomes of the developed 3D bioprinted scaffolds provide a promising strategy for development of personalized bone grafts for tissue regeneration.

category: bioengineering


title: Phenotyping single-cell motility in microfluidic confinement

abstract: At all scales, the movement patterns of organisms serve as dynamic read-outs of their behaviour and physiology. We devised a novel droplet microfluidics assay to encapsulate single algal microswimmers inside closed arenas, and comprehensively studied their roaming behaviour subject to a large number of environmental stimuli. We compared two model species, Chlamydomonas reinhardtii (freshwater alga, 2 cilia), and Pyramimonas octopus (marine alga, 8 cilia), and detailed their highly-stereotyped behaviours and the emergence of a trio of macroscopic swimming states (smooth-forward, quiescent, tumbling or excitable backward). Harnessing ultralong timeseries statistics, we reconstructed the species-dependent reaction network that underlies the choice of locomotor behaviour in these aneural organisms, and discovered the presence of macroscopic non-equilibrium probability fluxes in these active systems. We also revealed for the first time how microswimmer motility changes instantaneously when a chemical is added to their microhabitat, by inducing deterministic fusion between paired droplets - one containing a trapped cell, and the other, a pharmacological agent that perturbs cellular excitability. By coupling single-cell entrapment with unprecedented tracking resolution, speed and duration, our approach offers unique and potent opportunities for diagnostics, drug-screening, and for querying the genetic basis of micro-organismal behaviour.


title: Systemic influences of mammary cancer on monocytes in mice

abstract: There is a growing body of evidence that cancer causes systemic changes. These influences are most evident in the bone marrow and blood, particularly the myeloid compartment. Here we show using mouse models of breast cancer caused by the mammary epithelial expression of the Polyoma middle T antigen that there is an increase in the number of circulating and splenic monocytes. In the circulation, cancer does not affect ratios of classical to non-classical populations monocytes nor their halflives. Single cell RNA sequencing also indicates that cancer does not induce any new monocyte populations. In the bone marrow cancer does not change monocytic progenitor number is unaffected but the proliferation rate of monocytes is higher thus providing an explanation for expansion in the circulating number. Deep RNA sequencing of these monocytic populations reveals cancer causes changes in the classical monocyte compartment with changes evident in bone marrow monocytes but more in the blood suggesting influences in both compartments. Down regulation of interferon type 1 signalling and antigen presentation were the most prominent. Consistent with this analysis down regulated genes are enriched with STAT1/STAT2 binding sites in their promoter, transcription factors required for type 1 interferon signalling. However, these transcriptome changes in mice did not replicate those found in patients with breast cancer. Consequently, mouse models of cancer may be insufficient to study the systemic influences of human cancer.