title: Resting-state connectivity underlying cognitive controls association with perspective taking in callous-unemotional traits

abstract: Callous-Unemotional (CU) traits are often associated with impairments in perspective taking and cognitive control; and adolescents with CU traits demonstrate aberrant brain patterns in areas underlying perspective taking and cognitive control. Together cognitive control and perspective taking are thought to link mechanistically to explain CU traits. Because increased cognitive control demands modulate perspective taking ability among both typically developing samples and individuals with elevated CU traits, understanding the neurophysiological substrates of these constructs could inform both identification and interventional efforts to alleviate the societal costs of antisocial behavior. The present study examines the heterogenous functional brain properties underlying cognitive controls influence on perspective taking among adolescents on a continuum of CU traits. Results reveal that cognitive control had a negative indirect association with CU traits via perspective taking; and that greater network connection density within the social and conflict brain networks indirectly associate with lower CU traits via perspective taking and cognitive control (respectively). Additionally, less negative network connection density between the social and conflict networks directly associated with higher CU traits. Our results support the growing literature on cognitive controls influence on socio-cognitive functioning in CU traits and extends that work by identifying underlying functional brain properties.


title: The role of optogenetic stimulations of parvalbumin-positive interneurons in the prefrontal cortex and the ventral hippocampus on an acute MK801 model of schizophrenia-like cognitive inflexibility

abstract:
Background and HypothesisSchizophrenia research arose in the last decades, focusing more on its neural basis. Executive functions such as decision making and cognitive flexibility are the main cognitive areas that are impaired and are considered schizophrenia endophenotypes. Recently, cognitive impairment has been connected with the ablation of glutamatergic NMDARs resulting in increased cortical activity. Selective NMDARs antagonists such as dizocilpine have been used to model cognitive inflexibility in schizophrenia. Moreover, a decreased GABAergic inhibitory activity has been shown elsewhere along with the enhanced cortical activity. This NMDARs/GABA unbalanced ratio may reduce the entrainment of prefrontal gamma and hippocampal theta rhythm, resulting in a prefrontal-hippocampal gamma/theta band desynchronization.

Study DesignThe study addressed the role of acute administrations of dizocilpine to model schizophrenia-like cognitive inflexibility in rats. We used a new version of the attentional set-shifting task, where rats learned switching/reversing the relevant rule. Moreover, we used the new ASST after dizocilpine systemic injections to test cognitive flexibility. Finally, we used in vivo optogenetic stimulations at specific light pulses of parvalbumin-positive interneurons in the prefrontal cortex and ventral hippocampus.

ResultsThe first experiments showed that acute dizocilpine in rats reproduced schizophrenia-like cognitive inflexibility. The second set of experiments demonstrated that appropriate optogenetic light pulses frequencies could rescue the cognitive flexibility previously altered by acute dizocilpine.

ConclusionsThese findings advance our knowledge on the pivotal role of parvalbumin interneurons in schizophrenia-like cognitive impairment and may serve as a standpoint for further research of this severe psychiatric disorder.

category: neuroscience


title: TNFR2 costimulation differentially impacts regulatory and conventional CD4+ T-cell metabolism

abstract: CD4+ conventional T cells (Tconvs) mediate adaptive immune responses, whereas regulatory T cells (Tregs) suppress those responses to safeguard the body from autoimmunity and inflammatory diseases. The opposing activities of Tconvs and Tregs depend on the stage of the immune response and their environment, with an orchestrating role for cytokine- and costimulatory receptors. Nutrient availability also impacts T-cell functionality via metabolic and biosynthetic processes that are largely unexplored. Many data argue that costimulation by Tumor Necrosis Factor Receptor 2 (TNFR2) favors support of Treg over Tconv responses and therefore TNFR2 is a key clinical target. Here, we review the pertinent literature on this topic and highlight the newly identified role of TNFR2 as a metabolic regulator for thymus-derived (t)Tregs. We present novel transcriptomic and metabolomic data that show the differential impact of TNFR2 on Tconv and tTreg gene expression and reveal distinct metabolic impact on both cell types.

category: immunology


title: Rapid differentiation of regulatory CD4+ T cells in the infarcted myocardium blunts in situ inflammation

abstract:
BackgroundMyocardial infarction (MI) is a sterile inflammatory condition associated with tissue injury that results in the activation of T helper cell targeting cardiac antigens. However, the differentiation trajectories and in situ activity of heart-specific CD4+T cells activated in the MI context remain poorly understood.

MethodsHerein, we combined T-cell receptor transgenic models targeting myocardial protein, single-cell transcriptomics, and functional phenotyping to elucidate how the myosin-specific CD4+ T cells (TCR-M) differentiate in the murine infarcted myocardium and ultimately influence tissue repair. Furthermore, we adoptively transferred heart-specific T-cells that were pre-differentiated in vitro towards pro-inflammatory versus regulatory phenotypic states to dissect how they differentially regulate post-myocardial infarction (MI) inflammation.

ResultsFlow cytometry and single-cell transcriptomics findings reveled that transferred TCR-M cells rapidly acquired an induced regulatory phenotype (iTreg) in the infarcted myocardium and blunt local inflammation. Myocardial TCR-M cells differentiated into two main lineages enriched with cell activation and pro-fibrotic transcripts (e.g. Tgfb1) or with suppressor immune checkpoints (e.g. Pdcd1), which we also found in human myocardial tissue. These cells produced high levels of latency-associated peptide (LAP) and inhibited interleukine-17 (IL-17) responses. Notably, TCR-M cells that were pre-differentiated in vitro towards a regulatory phenotype maintained a stable in vivo FOXP3 expression and anti-inflammatory activity when adoptively transferred prior to MI induction. In contrast, TCR-M cells that were pre-differentiated in vitro towards a pro-inflammatory TH17 phenotype were partially converted towards a regulatory phenotype in the injured myocardium and blunted myocardial inflammation.

ConclusionsThese findings reveal that the myocardial milieu provides a suitable environment for iTreg differentiation and reveals novels mechanisms by which the healing myocardium shapes local immunological processes.


title: Multiple alr genes exhibit allorecognition-associated variation in the colonial cnidarian Hydractinia

abstract: The genetics of allorecognition has been determined from inbred lines of Hydractinia symbiolongicarpus, in which genetic control is attributed mainly to the highly polymorphic loci allorecognition 1 (alr1) and allorecognition 2 (alr2) located within the Allorecognition Complex (ARC). While allelic variation at alr1 and alr2 can predict the phenotypes in inbred lines, these two loci do not entirely predict the allorecognition phenotypes in wild-type colonies and their progeny, suggesting the presence of additional uncharacterized genes that improve the prediction of these phenotypes. Comparative genomics analyses were used to identify coding sequence differences in assembled chromosomal intervals of the ARC and genomic scaffold sequences between two incompatible H. symbiolongicarpus siblings from a backcross population. New IgSF-like genes are reported for the ARC, five of these genes are closely related to the alr1 and alr2 genes, suggesting the presence of multiple alr-type genes within this complex. Cloning evidence revealed that the allelic polymorphism of eight Ig-SF-like genes is associated with allorecognition phenotypes in a backcross population of H. symbiolongicarpus. Remarkably, alternative splicing was found as a mechanism that contributes to the functional variability of these genes by changing putative activating receptors to inhibitory receptors, or generating secreted isoforms of allorecognition proteins. Our findings demonstrate that allorecognition in H. symbiolongicarpus is a multigenic phenomenon controlled by genetic variation in at least eight genes in the ARC complex, as well as putative uncharacterized variation outside of this region.


title: A gut commensal protozoa remotely shapes a lung niche for asthma-promoting eosinophils.

abstract:
The gut microbiome influences chronic inflammation of the airways via the gut-lung axis. However, causal connections between microbes and their host, including the underlying mechanisms for this phenomenon remain largely unknown. Here, we show that colonization with the gut commensal protozoa, Tritrichomonas musculis (T.mu), remotely shapes the lung immune landscape and exacerbates allergic airway inflammation. We demonstrate that colonization with T.mu mediates the T and B cell-dependent accumulation and activation of inflammatory group 2 innate lymphoid cells in the lungs to constitute a tripartite immune network that serves as a niche for lung eosinophils. Animals colonized with T.mu show severely exacerbated allergic inflammation in the airways and reveal a new protozoan-driven gut-lung axis that remotely shapes the lung immune network to potentiate chronic pulmonary inflammation.

One-Sentence SummaryA gut microbe exacerbates asthma severity by promoting lung eosinophilia through a tripartite lymphocyte immune network.


title: Phase Separation of a Nodulin Sec14-like protein Maintains Auxin Efflux Carrier Polarity at Arabidopsis Plasma Membranes

abstract:
Robust development can be modulated by local protein accumulation which is sometimes manifested as polarity patterns; yet, the mechanisms that lead to these patterns are largely unknown. Using the model plant Arabidopsis, we report that auxin-induced polar patterns can be reinforced by the phase transition of a SEC14-like lipid-binding protein at the plasma membrane. Using imaging, genetics, and in vitro reconstitutions, we show that the SEC14-like phase transition is promoted outside the stem cell root niche through its association with the caspase-like protease separase and conserved microtubule motors in a plasma membrane interface. Separase cleaves SEC14-like to promote its transition from liquid-like clusters to solid-like filaments. Filaments self-amplify and potentiate the robustness and maintenance of plasma membrane domains through associations with polar proteins. This work uncovers that robust cell patterns involve proteolysis-mediated phase transitions at unpreceded plasma membrane interfaces.

SynopsisThis study provides an example of a protein that undergoes liquid-like phase separation at the plasma membrane in plants. Importantly, this protein known as SFH8 is a lipid-like transferase and can undergo proteolytic-induced solidification, which is an important event in the maintenance of polarized delivery of PINFORMED proteins.

O_LIThe complex comprising of the conserved caspase-like protease separase and the kinesin Kin7.3 (known as "KISC") associates with a SEC14-like homolog, the SFH8 protein at apicobasal polar domains
C_LIO_LISeparase cleaves SFH8 to yield a carboxy-terminal proteoform that undergoes a liquid-to-solid transition producing filamentous assemblies at the plasma membrane
C_LIO_LIThe KISC/SFH8 constitutes a feedback loop with auxin that potentiates the interaction, fusion, and maintenance of proteins with polar domains
C_LI

category: cell biology


title: Exosomally targeting microRNA23a ameliorates microvascular endothelial barrier dysfunction following rickettsial infection

abstract: Spotted fever group rickettsioses caused by Rickettsia (R) are devastating human infections, which mainly target microvascular endothelial cells (EC) and can induce lethal EC barrier dysfunction in the brain and lungs. Our previous evidence reveals that exosomes (Exos) derived from rickettsial-infected ECs, namely R-ECExos, can induce disruption of the tight junctional (TJ) protein ZO-1 and barrier dysfunction of human normal recipient brain microvascular endothelial cells (BMECs). However, the underlying mechanism remains elusive. Given that we have observed that microRNA23a (miR23a), a negative regulator of endothelial ZO-1 mRNA, is selectively sorted into R-ECExos, the aim of the present study was to characterize the potential functional role of exosomal miR23a delivered by R-ECExos in normal recipient BMECs. We demonstrated that EC-derived Exos (ECExos) have the capacity to deliver oligonucleotide RNAs to normal recipient BMECs in an RNase-abundant environment. miR23a in ECExos impairs normal recipient BMEC barrier function, directly targeting TJ protein ZO-1 mRNAs. In separate studies using a traditional in vitro model and a novel single living-cell biomechanical assay, our group demonstrated that miR23a anti-sense oligonucleotide-enriched ECExos ameliorate R-ECExo-provoked recipient BMEC dysfunction in association with stabilization of ZO-1 in a dose-dependent manner. These results suggest that Exo-based therapy could potentially prove to be a promising strategy to improve vascular barrier function during bacterial infection and concomitant inflammation.


title: Development of a Novel Microfluidic Co-culture model to study Organoid Vascularization

abstract: Organoids hold great potential for regenerative medicine and biomedical research. While they are able to recapitulate many structural and functional aspects of their in vivo counterparts, they lack a functional vasculature. In this study, we aimed to coculture human small intestinal organoids (hSIO) with a vasculature in a triple co-culture system with endothelial cells (ECs) and fibroblasts in a microfluidic device. Organoids and micro-vessels favour distinct matrix and medium conditions, which were optimized to sustain growth of all cell types. In addition, we found that organoids exhibit poor survival in monoculture in the microfluidic devices. Interestingly, co-culturing hSIO together with ECs and fibroblasts enhances stemness and survival of hSIO. This effect can be further enhanced by adding the GSK-3{beta} inhibitor CHIR99021 to the medium to enhance Wnt signalling. Direct contact of fibroblasts is not required for either vessel formation or for the effect on stem cell maintenance of organoids, as culturing fibroblasts in side-channels yields larger micro-vessels and better organoid survival. Additionally, we observed that the response of cells to different culture media is strongly dependent on the culture set-up as the second setup allows organoids to be cultured in angiogenic medium. Our results suggest that angiocrine signalling by the vasculature affects stem cell maintenance, which should be further investigated. Our co-culture system offers a platform to study angiocrine signalling in the small intestine, which could in the future be used to unravel different mechanisms behind endothelial mediated intestinal homeostasis and (patho) physiology.


title: Lis1-dynein drives corona compaction and error-correction at kinetochores

abstract: Mitotic cell division requires that kinetochores form microtubule attachments that can segregate chromosomes and control mitotic progression via the spindle assembly checkpoint. During prometaphase, kinetochores shed a distal domain called the fibrous corona as microtubule attachments form and mature. This shedding is mediated, in part, by the minus-end directed motor dynein, which  strips kinetochore cargoes along K-fibre microtubules towards the pole. While the main molecular players are well understood, relatively little is known about how dynein stripping is regulated and how it responds to increasing microtubule occupancy. Lis1 is a conserved dynein regulator that associates with kinetochores and is mutated in the severe neurodevelopmental disease lissencephaly. Here, we have combined loss-of-function studies, high-resolution imaging and engineered separation-of-function mutants to define how Lis1 contributes to dynein-mediated corona stripping. We show that cells depleted of Lis1 fail to fully dissemble the corona and delay in metaphase as a result of persistent checkpoint activation. Furthermore, we find that while kinetochore-tethered Lis1-dynein is required for attachment error-correction, the contribution of Lis1 to corona disassembly can be mediated by a rapidly cycling cytosolic pool. These findings support the idea that Lis1 contextualises dynein function at kinetochores to maintain corona disassembly into metaphase and prevent chromosome mis-segregation.


title: Paf1C denoises transcription and growth patterns to achieve organ shape reproducibility

abstract:
In multicellular systems, all cells exhibit transcriptional noise. However, its exact contribution to morphogenesis often remains unclear, especially in animals where cells can also move. Here we take advantage of walled plant cells, where transcriptional noise happens in tissues with a fixed topology. Using synchronously growing guard cells in stomata, we demonstrate that mutation in VIP3, a subunit of the conserved polymerase-associated factor 1 complex (Paf1C), increases transcriptional noise in Arabidopsis. This conclusion could be generalized to other group of cells at the shoot apex. Such noise translates into growth and shape defects. Indeed, in vip3 sepals, we measured higher growth heterogeneity between adjacent cells, with molecular evidence of increased local mechanical conflicts. This even culminated with the presence of negatively growing cells in specific growth conditions. Interestingly, such increased local noise makes the regional pattern of growth less clear-cut. Reduced regional conflicts lead to delay in organ growth arrest, ultimately making final organ shapes and sizes more variable. We propose that transcriptional noise is managed by Paf1C to ensure organ robustness by building up mechanical conflicts at the regional scale, instead of the local scale.

HIGHLIGHTSO_LIPaf1C controls transcriptional noise in Arabidopsis
C_LIO_LIPaf1C reduces growth heterogeneity and dampens local mechanical conflicts
C_LIO_LIImpairing both Paf1C and microtubules can trigger negative growth in Arabidopsis
C_LIO_LIScaling up local conflicts to regional ones with sharp boundaries contributes to organ shape robustness
C_LI

category: developmental biology


title: High-resolution Ribosome Profiling Reveals Translational Selectivity for Transcripts in Bovine Preimplantation Embryo Development

abstract:
High resolution ribosome fractionation and low-input ribosome profiling of bovine oocytes and preimplantation embryos has enabled us to define the translational landscapes of early embryo development at an unprecedented level. We analyzed the transcriptome, polysome- and non-polysome-bound RNA profiles of bovine oocytes (GV and MII stage) and early embryos at 2-, 8-cell, morula, and blastocyst stage, and revealed four modes of translational selectivity: i. selective translation of non-abundant mRNAs, ii. active, but modest translation of a selection of highly expressed mRNAs, iii. translationally suppressed abundant to moderately abundant mRNAs, and iv. mRNAs associated specifically with monosomes. A strong translational selection of low abundance mRNAs encoding protein components involved in metabolic pathways and lysosome was found throughout bovine oocyte and preimplantation development. In particular, genes encoding components involved in mitochondrial function were prioritized for translation. Notably, transcripts encoding proteins regulating chromatin modifications selectively translated in oocytes. We found that the translational dynamics largely reflects transcriptional profiles in oocytes and 2-cell embryos, but observed marked shift in translational control in 8-cell embryos associated with the main phase of embryonic genome activation. Subsequently, transcription and translation become better synchronized in morulae and blastocysts. Together, these data reveal a unique spatiotemporal translational regulation that accompanies bovine preimplantation development.

Significance StatementTranslational control during preimplantation embryo development is poorly understood, mostly due to the scarcity of samples and the corresponding inability to analyze low quantities of these materials. By developing a low-input method, we have been able to explore the transcriptome, polysome- and non-polysome-bound RNA profiles of bovine oocytes (GV and MII stage) and preimplantation embryos at 2-, 8-cell, morula, and blastocyst stages. We reveal four different modes of translational selectivity, plus novel temporal regulatory mechanisms during early embryo development. The spatiotemporal translation dynamics of bovine oocytes and preimplantation embryos offer an entirely new insight into mammalian embryo development research and new possibilities for improving efficiency of assisted reproduction technologies (ARTs).


title: Utilisation of Oxford Nanopore sequencing to generate six complete gastropod mitochondrial genomes as part of a biodiversity curriculum

abstract: High-throughput sequencing has enabled genome skimming approaches to produce complete mitochondrial genomes (mitogenomes) for species identification and phylogenomics purposes. In particular, the portable sequencing device from Oxford Nanopore Technologies (ONT) has the potential to facilitate hands-on training from sampling to sequencing and interpretation of mitogenomes. In this study, we present the results from sampling and sequencing six gastropod mitogenomes (Aplysia argus, Cellana orientalis, Cellana toreuma, Conus ebraeus, Conus miles and Tylothais aculeata) from a graduate level biodiversity course. The students were able to produce mitochondrial genomes from sampling to annotation using existing protocols and programs. Approximately 4Gb of sequence was produced from 15 Flongle and two R9.4 Flowcells, averaging 235Mb and N50=4.4kb per Flowcell. Five of the six 14.1-18kb mitogenomes were circlised containing all 13 core protein coding genes. Additional Illumina sequencing reveal that the ONT assemblies were able to span over highly AT rich sequence in the control region that was otherwise missing in Illumina-assembled mitogenomes, but still contained a base error of one every 70.8-346.7bp with the majority occurring at homopolymer regions. Our findings suggest that ONT are portable and can be used to rapidly produce mitogenomes at low cost and tailored to genomics-based training in biodiversity research.

category: ecology


title: qad: An R-package to detect asymmetric and directed dependence in bivariate samples

abstract: Correlations belong to the standard repertoire of ecologists for quantifying the strength of dependence between two random variables. Classical dependence measures are usually not capable of detecting non-monotonic or non-functional dependencies. Furthermore, they completely fail to detect asymmetry and direction in dependence, which exist in many situations and should not be ignored. In this paper, we present qad (short for quantification of asymmetric dependence), a non-parametric statistical method to quantify directed and asymmetric dependence of bivariate samples. qad is applicable in general situations, is sensitive to noise in data, exhibits a good small sample performance, detects asymmetry in dependence, shows high power in testing for independence, requires no assumptions regarding the underlying distribution of the data, and reliably quantifies the information gain/predictability of quantity Y given knowledge of quantity X, and vice versa (i.e. q(X,Y) = q(Y, X)). Here, we briefly recall the methodology underlying qad, introduce the functions of the R-package qad, which returns estimates for the measures q(X, Y) denoting the directed dependence of Y on X (or, equivalently, the influence of X on Y), q(Y, X) the directed dependence of X on Y, a(X, Y) [colone] q(X, Y) - q(Y, X) the asymmetry in dependence. Furthermore, qad can be used to predict Y given knowledge of X, and vice versa. Additionally, we compare empirical performance of qad with that of seven other well established measures and demonstrate the applicability of qad on ecological datasets. We illustrate that direction and asymmetry in dependence are universal properties of bivariate associations. qad thus provides additional information gain and the avoidance of model bias and will therefore advance and facilitate the understanding of ecological systems.


title: Novel Methods for Multi-ancestry Polygenic Prediction and their Evaluations in 3.7 Million Individuals of Diverse Ancestry

abstract: Polygenic risk scores are becoming increasingly predictive of complex traits, but subpar performance in non-European populations raises concerns about their potential clinical applications. We develop a powerful and scalable method to calculate PRS using GWAS summary statistics from multi-ancestry training samples by integrating multiple techniques, including clumping and thresholding, empirical Bayes and super learning. We evaluate the performance of the proposed method and a variety of alternatives using large-scale simulated GWAS on ~19 million common variants and large 23andMe Inc. datasets, including up to 800K individuals from four non-European populations, across seven complex traits. Results show that the proposed method can substantially improve the performance of PRS in non-European populations relative to simple alternatives and has comparable or superior performance relative to a recent method that requires a higher order of computational time. Further, our simulation studies provide novel insights to sample size requirements and the effect of SNP density on multi-ancestry risk prediction.

category: genetics


title: SpyChIP identifies cell type-specific transcription factor occupancy from complex tissues

abstract: Chromatin immunoprecipitation (ChIP) is an important technique for characterizing protein-DNA binding in vivo. One drawback of ChIP based techniques is the lack of cell type-specificity when profiling complex tissues. To overcome this limitation, we developed SpyChIP to identify cell type-specific transcription factor (TF) binding sites in native physiological contexts without tissue dissociation or nuclei sorting. SpyChIP takes advantage of a specific covalent isopeptide bond that rapidly forms between the 15 amino acid SpyTag and the 17 kD protein SpyCatcher. In SpyChIP, the target TF is fused with SpyTag by genome engineering, and an epitope tagged SpyCatcher is expressed in cell populations of interest, where it covalently binds to SpyTag-TF. Cell type-specific ChIP is obtained by immunoprecipitating chromatin prepared from whole tissues using antibodies directed against the epitope-tagged SpyCatcher. Using SpyChIP, we identified the genome-wide binding profiles of the Hox protein Ubx in two distinct cell types of the Drosophila haltere disc. Our results revealed extensive region-specific Ubx-DNA binding events, highlighting the significance of cell type-specific ChIP and the limitations of whole tissue ChIP approaches. Analysis of Ubx::SpyChIP results provided novel insights into the relationship between chromatin accessibility and Ubx-DNA binding, as well as different mechanisms Ubx employs to regulate its downstream cis-regulatory modules (CRMs). In addition to SpyChIP, we suggest that SpyTag-SpyCatcher technology, as well as other covalent interaction peptide pairs, has many potential in vivo applications that were previously unachievable.


title: Comparative assessment of computed tomography and magnetic resonance imaging of the spider morph of Python regius and wild type Python regius to evaluate the morphological correlate of the wobble syndrome.

abstract: There is general awareness of artificial selection and its potential implications on health and welfare of animals. Despite growing popularity and increasing numbers of breeds of atypical colour and pattern variants in reptiles, only few studies have investigated the appearance and cause of various diseases associated with colour morphs. Ball pythons (Python regius) are among the most frequently bred reptiles and breeders selected for a multitude of different colour and pattern morphs. Among those colour variants, the spider morph of the ball python is frequently associated with the wobble syndrome. The aim of this study was to determine, whether a morphological variant can be found and brought in association with the clinical occurrence of the wobble syndrome in spider ball pythons, using MRI and CT-imaging as intra-vitam diagnostic methods. Data from eight ball pythons including five spider ball pythons and three wild type ball pythons was assessed and evaluated comparatively. We were able to identify distinctive structural differences in inner ear morphology in spider ball pythons highly probable to relate to the wobble syndrome. To our knowledge, these anomalies are described for the first time and represent a basis for further anatomical and genetic studies and discussions regarding animal welfare in reptile breeding.


title: Limitations of principal components in quantitative genetic association models for human studies

abstract: Principal Component Analysis (PCA) and the Linear Mixed-effects Model (LMM), sometimes in combination, are the most common modern models for genetic association. Previous PCA-LMM comparisons give mixed results and unclear guidance, and have several limitations, including not varying the number of principal components (PCs), simulating overly simple population structures, and inconsistent use of real data and power evaluations. In this work, we thoroughly evaluate PCA and LMM both with varying number of PCs in new realistic genotype and complex trait simulations including admixed families, trees, and large real multiethnic human genotype datasets (1000 Genomes Project, the Human Genome Diversity Panel, and Human Origins) with simulated traits. We find that LMM without PCs performs best in all cases, with the largest effects in the family simulation and all real human datasets. We determined that the large gaps in PCA to LMM performance on the real human datasets is due to the high-dimensional family structure stemming from large numbers of distant relatives, and not from the smaller number of highly related pairs. While it was known that PCA fails on family data, here we report a strong effect on association of cryptic family relatedness in several genetically diverse human datasets, a problem that is not avoided with the common practice of pruning high-relatedness individual pairs. Overall, this work better characterizes the severe limitations of PCA compared to LMM in modeling the complex relatedness structures present in real multiethnic human data and its impact in association studies.


title: Genome-wide signatures of geographic expansion and breeding process in soybean

abstract:
The clarification of genomic signatures left during evolutionary histories of crops is crucial for breeding varieties adapting to changing climate. Soybean, a leguminous crop, provides both plant oil and protein. Here, we analyzed genome sequences of 2,214 soybeans and proposed its evolutionary route, which includes four geographic paths, expansion of annual wild soybean (Glycine soja Sieb. & Zucc.) from Southern China, domestication in Central China, expansion of landrace (G. max (L.) Merr.), and local breeding. We observed that local adaptation of the wild and cultivated soybeans was largely independent, and that genetic introgression was mostly derived from sympatric rather than allopatric wild populations during the range expansion of soybean landraces. Range expansion and breeding processes were accompanied with positive selection of flowering-time genes including GmSPA3c as validated by knock-out mutants. Our study shed lights on the evolutionary history of soybean and provides valuable genetic resources for future breeding.

TeaserThe expansion and selection history of soybean

category: genomics


title: PeerPub: A device for concurrent operant oral self-administration by multiple rats

abstract: The social environment has long been recognized to play an important role in substance abuse disorders (SUD). Operant conditioning is the most widely used rodent model of SUD. However, most operant chambers do not accommodate more than one rat at a time. Here, we introduce PeerPub - a novel social operant chamber. PeerPub uses a touch sensor to record the number of licks on drinking spouts. It then delivers a drop of solution with a fixed volume as the reward to the tip of the spout when the number of licks meets the requirement of a reinforcement schedule. A radio-frequency identification (RFID) chip implanted on top of each rats skull tracks the identity of the rat. The system is controlled by a Raspberry Pi computer. We tested PeerPub using male Wistar Kyoto rats in daily one-hour sessions where supersac, a solution containing glucose and saccharin, was delivered under a fixed ratio 5 schedule. We found that male rats consumed more supersac in group housing rather than in isolated conditions. These data demonstrated the utility of PeerPub in modeling the interaction between motivated behavior and social context. We anticipate devices like PeerPub will help demonstrate the role of the social environment in SUD phenotypes. The design of PeerPub is available at http://github.com/nijie321/PeerPub

category: animal behavior and cognition


title: Structure of the connexin-43 gap junction channel reveals a closed sieve-like molecular gate

abstract: Gap junction channels (GJCs) mediate intercellular communication by connecting two neighboring cells and enabling direct exchange of ions and small molecules. Cell coupling via connexin-43 (Cx43) GJCs is important in a wide range of cellular processes in health and disease 1-3, yet the structural basis of Cx43 function and regulation has not been determined until now. Here we describe the structure of a human Cx43 GJC solved by cryo-EM and single particle analysis at 2.26 [A] resolution. The pore region of Cx43 GJC features several lipid-like densities per Cx43 monomer, located close to a putative lateral access site at the monomer boundary. We found a previously undescribed closed sieve-like gate on the cytosolic side of the pore, formed by the N-terminal domain and the transmembrane helix 2 of Cx43 and stabilized by a small molecule. Structures of the Cx43 GJC and hemichannels in nanodiscs confirm the sieve-like gate arrangement. The features of the Cx43 GJC and hemichannel cryo-EM maps and the channel properties revealed by molecular dynamics simulations suggest that the captured states of Cx43 are closed.

category: biochemistry


title: Pre-equilibrium biosensors: A new approach towards rapid and continuous molecular measurements

abstract: Almost all biosensors that use ligand-receptor binding operate under equilibrium conditions. However, at low ligand concentrations, the equilibration with the receptor (e.g., antibodies and aptamers) become slow and thus equilibrium-based biosensors are inherently limited in making measurements that are both rapid and sensitive. In this work, we provide a theoretical foundation for a novel method through which biosensors can quantitatively measure ligand concentration before reaching equilibrium. Rather than only measuring receptor binding at a single time-point, the pre-equilibrium approach leverages the receptors kinetic response to instantaneously quantify the changing ligand concentration. Importantly, analyzing the biosensor output in frequency domain, rather than in the time domain, we show the degree to which noise in the biosensor affects the accuracy of the pre-equilibrium approach. Through this analysis, we provide the conditions under which the signal-to-noise ratio of the biosensor can be maximized for a given target concentration range and rate of change. As a model, we apply our theoretical analysis to continuous insulin measurement and show that with a properly selected antibody, the pre-equilibrium approach could make the continuous tracking of physiological insulin fluctuations possible.

category: bioengineering


title: Multiplexed Label-Free Biomarker Detection by Targeted Disassembly of Variable-Length DNA Payload Chains

abstract:
Simultaneously studying different types of biomarkers (DNA, RNA, proteins, metabolites) has the potential to significantly improve understanding and diagnosis for many complex diseases. However, extracting biomarkers of different types involves using several technically complex or expensive methodologies, often requiring specialized laboratories and personnel. Streamlining detection through the use of a single multiplexed assay would greatly facilitate the process of accessing and interpreting patient biomarker data. In this work, we present a method for multiplexed biomarker detection based on variable-length DNA payload chains, which are systematically disassembled in the presence of specific biomolecular targets, leading to fragments of different sizes that yield characteristic band patterns in gel electrophoresis. This strategy has enabled us to detect with high sensitivity and specificity DNA sequences including BRCA1, an RNA sequence (miR-141) and the steroid aldosterone. We show that our assay can be multiplexed, enabling simultaneous detection of different types of biomarker. Furthermore, we show that our method suffers no loss of sensitivity when conducted in fetal bovine serum and can be applied using capillary electrophoresis, which may be more amenable to automation and integration in healthcare settings.

ToC Graphic

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title: ChromDMM: A Dirichlet-Multinomial Mixture Model For Clustering Heterogeneous Epigenetic Data

abstract:
MotivationResearch on epigenetic modifications and other chromatin features at genomic regulatory elements elucidates essential biological mechanisms including the regulation of gene expression. Despite the growing number of epigenetic datasets, new tools are still needed to discover novel distinctive patterns of heterogeneous epigenetic signals at regulatory elements.

ResultsWe introduce ChromDMM, a product Dirichlet-multinomial mixture model for clustering genomic regions that are characterised by multiple chromatin features. ChromDMM extends the mixture model framework by profile shifting and flipping that can probabilistically account for inaccuracies in the position and strand-orientation of the genomic regions. Owing to hyper-parameter optimisation, ChromDMM can also regularise the smoothness of the epigenetic profiles across the consecutive genomic regions. With simulated data, we demonstrate that ChromDMM clusters, shifts, and strand-orients the profiles more accurately than previous methods. With ENCODE data, we show that the clustering of enhancer regions in the human genome reveals distinct patterns in several chromatin features. We further validate the enhancer clusters by their enrichment for transcriptional regulatory factor binding sites.

AvailabilityThe software is available at https://github.com/MariaOsmala/ChromDMM

category: bioinformatics


title: ViPal: A Framework for Virulence Prediction of Influenza Viruses with Prior Viral Knowledge Using Genomic Sequences

abstract: Influenza viruses pose significant threats to public health and cause enormous economic loss every year. Previous work has revealed the viral factors that influence the virulence of influenza viruses. However, taking prior viral knowledge represented by heterogeneous categorical and discrete information into account is scarce in the existing work. How to make full use of the preceding domain knowledge into virulence study is challenging but beneficial. This paper proposes a general framework named ViPal for virulence prediction that incorporates discrete prior viral mutation and reassortment information based on all eight influenza segments. The posterior regularization technique is leveraged to transform prior viral knowledge to constraint features and integrated into the machine learning models. Experimental results on influenza genomic datasets validate that our proposed framework can improve virulence prediction performance over baselines. The comparison between ViPal and other existing methods shows the computational efficiency of our framework with superior performance. Moreover, the interpretable analysis through SHAP identifies the scores of constraint features contributing to the prediction. We hope this framework could provide assistance for the accurate detection of influenza virulence and facilitate flu surveillance.


title: Exploring intra-species diversity through non-redundant pangenome assemblies

abstract:
At the genome level, microorganisms are highly adaptable both in terms of allele and gene composition. Such heritable traits emerge in response to different environmental niches and can have a profound influence on microbial community dynamics. As a consequence of this, any individual genome or clonal population will contain merely a fraction of the total genetic diversity of any operationally defined "species", with the collective from that group presenting the broader genomic diversity known as the pangenome. Pangenomes are valuable concepts for studying evolution and adaptation in microorganisms, as they partition genomes into core regions (present in all the genomes, and responsible for housekeeping and species-level niche adaptation) and accessory regions (present only in some genomes, and responsible for ecotype divergence).

Here we present SuperPang, an algorithm capable of producing pangenome assemblies from a set of input genomes of varying quality, including metagenome-assembled genomes or MAGs. SuperPang runs in linear time and its results are complete, non-redundant, preserve gene ordering and contain both coding and non-coding regions. Our approach provides a modular view of the pangenome, identifying operons and genomic islands, and allowing to track their prevalence in different populations. We illustrate our approach by analyzing the intra-species diversity of Polynucleobacter, a clade of ubiquitous freshwater microorganisms characterized by their streamlined genomes and their ecological versatility. We show how SuperPang facilitates the simultaneous analysis of allelic and gene content variation under different environmental pressures, allowing us to study the drivers of microbial diversification at unprecedented resolution.


title: Solution structure of NPSL2, a regulatory element in the oncomiR-1 RNA

abstract: The miR-17~92a polycistron, also known as oncomiR-1, is commonly overexpressed in multiple cancers and has several oncogenic properties. OncomiR-1 encodes six constituent microRNAs (miRs), each enzymatically processed with different efficiencies. However, the structural mechanism that regulates this differential processing remains unclear. Chemical probing of oncomiR-1 revealed that the Drosha cleavage sites of pri-miR-92a are sequestered in a four-way junction. NPSL2, an independent stem loop element, is positioned just upstream of pri-miR-92a and sequesters a crucial part of the sequence that constitutes the basal helix of pri-miR-92a. Disruption of the NPSL2 hairpin structure could promote the formation of a pri-miR-92a structure that is primed for processing by Drosha. Thus, NPSL2 is predicted to function as a structural switch, regulating pri-miR-92a processing. Here, we determined the solution structure of NPSL2 using solution NMR spectroscopy. This is the first high-solution structure of an oncomiR-1 element. NPSL2 adopts a hairpin structure with a large, but highly structured, apical and internal loops. The 10-bp apical loop contains a pH-sensitive A+{middle dot}C mismatch. Additionally, several adenosines within the apical and internal loops have elevated pKa values. The protonation of these adenosines can stabilize the NPSL2 structure through electrostatic interactions. Our study provides fundamental insights into the secondary and tertiary structure of an important RNA hairpin proposed to regulate miR biogenesis.

category: biophysics


title: Targeting tumor stemness switch phenotype by activating pathogen induced stem cell niche defense

abstract: Cancer stem cells (CSCs) reside in their tumor microenvironment (TME) niches, which are often hypoxic. Previously, we found that hypoxia and oxidative stress prevalent in TME may reprogram CSCs to a highly aggressive and inflammatory phenotype, the tumor stemness switch (TSS) phenotype. We previously reported a "stem cell niche defense" mechanism in bone marrow and lung mesenchymal stem cell niche against pathogen. Pathogen induced bystander apoptosis (PIBA) of stem cells harboring intracellular pathogen may be part of this defense mechanism. We speculate that the TSS phenotype may also activate this niche defense mechanism to defend their TME niche against pathogen and therefore could be exploited to target CSCs. Here we report that CSCs of TSS phenotype enriched in post-hypoxia ABCG2+ CSCs of several cell lines of diverse tumors including oral squamous cell carcinoma cell line SCC-25 exhibited bystander apoptosis when infected with either Bacillus Calmette Guerin (BCG) or mutant Mycobacterium tuberculosis (Mtb) strain 18b. The conditioned media (CM) of the infected cells not only exhibited marked anti-tumor activity in vivo, but also showed significant anti-microbial activity. A detailed mechanisms study revealed that some of the infected ABCG2+ CSCs underwent pyroptosis and released a high mobility group box protein 1 (HMGB1)/p53 death signal that can induce a toll like receptor (TLR) 2/4 mediated bystander apoptosis. Thus, our findings suggest that PIBA can be utilized to activate the "niche defense" mechanism in TSS phenotype, which not only target the TSS, but also exhibit marked anti-tumor activity in vivo.

category: cancer biology


title: Adversary of DNA integrity: a long non-coding RNA stimulates driver oncogenic chromosomal rearrangement in human thyroid cells

abstract:
The flurry of publications devoted to the functions of long non-coding RNAs (lncRNAs) published in the last decade leaves no doubt about the exceptional importance of lncRNAs in various areas including tumor biology. Contribution of lncRNAs to the early stages of oncogenesis remains poorly understood. In this study we explored a new role for lncRNAs: stimulation of driver oncogenic mutations that result from specific chromosomal rearrangements. We demonstrated that lncRNA CASTL1 (ENSG00000269945) stimulates the formation of the CCDC6-RET inversion (RET/PTC1) in human thyroid cells subjected to radiation or chemical DNA damage. Facilitation of chromosomal rearrangement requires lncRNA to contain regions complementary to the introns of both CCDC6 and RET genes as deletion of these regions deprives CASTL1 of the ability to stimulate the gene fusion. We found that CASTL1 expression is elevated in tumors with CCDC6-RET fusion which is the most frequent rearrangement in papillary thyroid carcinoma. Our results open a new venue for the studies of early oncogenesis in various tumor types, especially those associated with physical or chemical DNA damage.

Graphical abstract

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title: CHA1: A New Combinatorial Therapy That Reciprocally Regulates Wnt and JAK/STAT/Interferon Signaling to Re-program Breast Tumors and the Tumor-Resident Landscape.

abstract: Triple negative breast cancers (TNBC) pose exceptional challenges with fatal brain metastases as a clear and unmet need. Immune checkpoint inhibitors (ICIs) are promising therapeutic strategies, but most TNBC are resistant, or "cold" tumors, due to lack of tumor-resident immune engagement. No FDA-approved therapies exist which promote a "cold-to-hot" transition or induce the important biomarker PD-L1, often used for ICI clinical decision-making. Maximal ICI susceptibility, or a full "cold-to-hot" transition, requires reciprocal Wnt signaling inhibition and Jak/STAT/interferon signaling activation. We report a new compound combination (CHA1) that fits the above criteria. CHA1 is comprised of EGCG (epigallocatechin-3-gallate; green-tea compound) and decitabine (DNA-methyltransferase (DNMT1) inhibitor; 5deaza-cytidine; FDA-approved for hematologic malignancies). We used immune-compromised and syngeneic TNBC pre-clinical models to investigate tumor-intrinsic and tumor-resident T-cell effects, respectively. All results required CHA1 (but not EGCG or decitabine alone) and utilized attainable human dose equivalences with manageable safety profiles. CHA1 triggered efficient Wnt signaling inhibition by elevating Wnt pathway inhibitors (HBP1 and SFRP1) and traversed the blood-brain barrier to reduce both tumor and brain metastatic growth. Transcriptomic and expression analyses revealed that CHA1 treatment effectuated a robust tumor-intrinsic JAK/STAT/IFN response 1) to induce PDL1 and 2) to induce antigen presentation and processing genes, including MHC-1, MHC-2 and numerous genes attributed to professional antigen-presenting cells; 3) to induce CD8+-T-cell infiltration and activation. Additionally, CHA1 pre-treatment improved anti-PDL1 efficacy in a syngeneic setting. Lastly, we derived a composite gene signature emblematic of CHA1 treatment and of a favorable clinical prognosis in-silico. Together, our work supports a model in which CHA1 influences epigenetics, Wnt and Jak/STAT/IFN signaling mechanisms--all to reprogram an epithelial-mesenchymal TNBC tumor to express antigen-presenting properties and to recruit and activate tumor-resident CD8+-T-cells. We discuss our findings in the context of cancer biology and immunity with implications for improving ICI susceptibility for TNBC.


title: A cancer stem cell population underlies a multi-lineage phenotype and drug resistance in prostate cancer

abstract: To resist lineage-dependent therapies, cancer cells adopt a plastic stem-like state, leading to phenotypic heterogeneity. Here we dissect the cellular origins of such heterogeneity in a metastatic castration-resistant prostate cancer (CRPC) patient-derived adenocarcinoma organoid model displaying a range of luminal and neuroendocrine phenotypes and driven by mutations in cell cycle (CDKN1B) and epigenetic (ARID1A, and ARID1B) regulators. As shown by lineage tracing, metastatic tumor heterogeneity originated from distinct subclones of infrequent stem/progenitor cells that each produced a full distribution of differentiated lineage markers, suggesting multiclonal evolution to a relatively stable bipotential state. Single cell ATAC-seq analyses revealed the co-occurrence of transcription factor activities associated with multiple disparate lineages in the stem/progenitors: WNT and RXR stem factors, AR and FOXA1 luminal epithelial drivers, and NR2F1 and ASCL1 neural factors. Inhibition of AR in combination with AURKA but not EZH2 blocked tumor growth. These data provide insight into the origins and dynamics of cancer cell plasticity and stem targeted therapy.


title: Cuticle and cuticular sensilla in Agnostina

abstract: The microstructure of the cuticle of several Agnostina species is described. For the first time, it has been shown that their cuticle consists of three layers: thick outer and transitional layers and a thin inner layer. The layers are made up of stacks of laminae: the laminae in the outer layer are thicker than in the transitional and inner ones. The evolution of the agnostoid cuticle proceeded, apparently, in the direction of decreasing the thickness of the inner layer which in contrast to all other arthropods, is thinner than the outer layer. Apparently, the inner layer was mineralized to a lesser extent than the outer one. In the structure of the cuticle, Agnostina are close to horseshoe crabs and trilobites. On the outer and inner layers, a set of sensilla was revealed; they can be attributed to trichoid, campaniform, and digitiform sensilla (with mechanoreceptor function) and celoconic sensilla (with chemoreceptor function). Also, unique sensorial ensembles were found on cephalons and pygidia; the function of the former is probably mechanosensorial, and the purpose of the latter cannot be assumed on this material.

category: paleontology


title: Physiological and molecular mechanisms regulated mesophyll conductance under severe drought in water-saving drought-resistant rice

abstract: Water-saving and drought-resistant rice (WDR) is a new type of rice varieties. It plays an important role in responding to drought with high yield and has been widely planted in central China at present. High photosynthetic production potential caused by high mesophyll conductance (gm) is the main factor promoted high yield formation in drought for WDR. But little is known about physiological and molecular mechanisms regulated gm in drought for WDR. Therefore, WDR cultivar HY73 and drought-sensitive cultivar HLY898 were used for comparative studies with three irrigation regimes before applying severe drought treatment at heading to create different differential individuals of photosynthetic potential and gm. The results showed that cultivar HY73 had lower up-regulation different expression genes (DEGs) than cultivar HLY898 in drought at transcriptional level. Conversely, DEGs of down-regulation was higher in cultivar HY73 than cultivar HLY898. In addition, 3071 DEGs were clustered in 3 modules named Midnightblue (734 DEGs), Blue (921 DEGs), and Turquoise (1416 DEGs) in severe drought merged three irrigation regimes and both cultivars, which the modules had significant correlational relationship with gm based on weighted gene co-expression network analysis (P<0.05). Only DEGs in midnightblue module were enriched in photosynthesis process and positively regulated gm (P<0.05). The main biological process were photosynthesis (GO:0015979), light harvesting in photosystem I (GO:0009768), reductive pentose-phosphate cycle (GO:0019253), protein-chromophore linkage (GO:0018298), photosynthetic electron transport in photosystem I (GO:0009773), and photosystem II repair (GO:0010206). These results indicate that gm and energy distribution in PSI and PSII systems could synergistic effect photosynthetic production potential in severe drought for rice plants. In the modules, the 18 most highly connected hub genes were screened using co-expression networks method. RT-PCR analysis indicated that CSP41B, PGLP1A, LHCA5, and GSTU6 genes had a similar variation trend with gm among treatments for both cultivar. LHCA5 and CSP41B genes were significantly up-regulated in HY73 compared with HLY898 in drought (P<0.05). And the both genes locates in thylakoid membrane in photosystems. Therefore, LHCA5 and CSP41B genes could be key genes to synergistically manage gm and energy distribution in photosystems. Our results provide some new physiological and molecular mechanisms regulated gm in severe drought for WDR.

category: physiology


title: SegVeg: Segmenting RGB images into green and senescent vegetation by combining deep and shallow methods

abstract:
The pixels segmentation of high resolution RGB images into background, green vegetation and senescent vegetation classes is a first step often required before estimating key traits of interest including the vegetation fraction, the green area index, or to characterize the sanitary state of the crop. We developed the SegVeg model for semantic segmentation of RGB images into the three classes of interest. It is based on a U-net model that separates the vegetation from the background. It was trained over a very large and diverse dataset. The vegetation pixels are then classified using a SVM shallow machine learning technique trained over pixels extracted from grids applied to images. The performances of the SegVeg model are then compared to a three classes U-net model trained using weak supervision over RGB images with predicted pixels by SegVeg as groundtruth masks.

Results show that the SegVeg model allows to segment accurately the three classes, with however some confusion mainly between the background and the senescent vegetation, particularly over the dark and bright parts of the images. The U-net model achieves similar performances, with some slight degradation observed for the green vegetation: the SVM pixel-based approach provides more precise delineation of the green and senescent patches as compared to the convolutional nature of U-net. The use of the components of several color spaces allows to better classify the vegetation pixels into green and senescent ones. Finally, the models are used to predict the fraction of the three classes over the grids pixels or the whole images. Results show that the green fraction is very well estimated (R2=0.94) by the SegVeg model, while the senescent and background fractions show slightly degraded performances (R2=0.70 and 0.73, respectively).

We made SegVeg publicly available as a ready-to-use script, as well as the entire dataset, rendering segmentation accessible to a broad audience by requiring neither manual annotation nor knowledge, or at least, offering a pre-trained model to more specific use.

category: plant biology


title: SWEET11 and SWEET12 transporters function in tandem to modulate sugar flux in Arabidopsis: An account of the underlying unique structure-function relationship

abstract: Sugar will eventually be exported transporters (SWEETs) have been identified as a unique class of sugar efflux transporters in all biological kingdoms. AtSWEET11 and AtSWEET12 in Arabidopsis act synergistically to perform distinct physiological roles, particularly in apoplasmic phloem loading, seed filling, and sugar level alteration at the site of pathogen infection. Plasma membrane-localized AtSWEET11 and AtSWEET12 transporters exclusively facilitate sucrose transport along the concentration gradient. This article examines the sucrose binding pocket of AtSWEET11 and AtSWEET12 using docking studies, and how they act synergistically in various functions throughout plant development and during abiotic and biotic stresses. Further, we highlight the phylogenetic and the in-silico analyses of AtSWEET11 and AtSWEET12 orthologs from 39 economically important plant species that could provide new platforms for future studies on sugar allocation mechanisms across the different plant families. In-depth understanding of these transporters and their molecular regulatory mechanisms could be harnessed for crop improvement and crop protection.


title: Digital twins and hybrid modelling for simulation of physiological variables and stroke risk

abstract: One of the more interesting ideas for achieving personalized, preventive, and participatory medicine is the concept of a digital twin. A digital twin is a personalized computer model of a patient. So far, digital twins have been constructed using either mechanistic models, which can simulate the trajectory of physiological and biochemical processes in a person, or using machine learning models, which for example can be used to estimate the risk of having a stroke given a cross-section profile at a given timepoint. These two modelling approaches have complementary strengths which can be combined into a hybrid model. However, even though hybrid modelling combining mechanistic modelling and machine learning have been proposed, there are few, if any, real examples of hybrid digital twins available. We now present such a hybrid model for the simulation of ischemic stroke. On the mechanistic side, we develop a new model for blood pressure and integrate this with an existing multi-level and multi-timescale model for the development of type 2 diabetes. This mechanistic model can simulate the evolution of known physiological risk factors (such as weight, diabetes development, and blood pressure) through time, under different intervention scenarios, involving a change in diet, exercise, and certain medications. These forecast trajectories of the physiological risk factors are then used by a machine learning model to calculate the 5-year risk of stroke, which thus also can be calculated for each timepoint in the simulated scenarios. We discuss and illustrate practical issues with clinical implementation, such as data gathering and harmonization. By improving patients understanding of their body and health, the digital twin can serve as a valuable tool for patient education and as a conversation aid during the clinical encounter. As such, it can facilitate shared decision-making, promote behavior change towards a healthy lifestyle, and improve adherence to prescribed medications.

category: systems biology


title: An anchored experimental design and meta-analysis approach to address batch effects in large-scale metabolomics

abstract:
Large-scale untargeted metabolomics studies suffer from individual variation, batch effects and instrument variability, making comparisons of common spectral features across studies difficult. One solution is to compare studies after compound identification. However, compound identification is expensive and time consuming. We successfully identify common spectral features across multiple studies, with a generalizable experimental design approach. First, we included an anchor strain, PD1074, during sample and data collection. Second, we collected data in blocks with multiple controls. These anchors enabled us to successfully integrate three studies of Caenorhabditis elegans for nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS) data from five different assays. We found 34% and 14% of features to be significant in LC-MS and NMR, respectively. Between 20-50% of spectral features differ in a mutant and among a set of genetically diverse natural strains, suggesting this reduced set of spectral features are excellent targets for compound identification.

GRAPHICAL ABSTRACT

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/485859v1_ufig1.gif" ALT="Figure 1">
View larger version (47K):
org.highwire.dtl.DTLVardef@84abe1org.highwire.dtl.DTLVardef@3066fforg.highwire.dtl.DTLVardef@1ba166corg.highwire.dtl.DTLVardef@164cf0_HPS_FORMAT_FIGEXP  M_FIG C_FIG Fourteen C. elegans strains are used in three individual studies. PD1074, the anchor control strain (orange), is grown alongside test strains (green, yellow, purple). Multiple biological replicates of PD1074 captures environmental variation in growth conditions. Non-polar and polar metabolic data across the three studies (i.e., natural strains, central metabolism mutants, and UGT mutants) were collected by nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS). Data acquisition controls in each block included biological reference material and pooled PD1074 samples. Biological replicates of PD1074 (n = 42 for LC-MS, n = 52 for NMR) were included in all batches. Meta-analysis provided comparable inferences to mixed effects models, and the estimated relative effects of each test strain to PD1074 and straightforward comparisons of test strains across experiments.


title: ALS sensitive spinal motor neurons enter a degenerative downward spiral of impaired splicing and proteostasis

abstract: Despite clear therapeutic potential, the mechanisms that confer differential neuronal sensitivity are not well understood. During Amyotrophic Lateral Sclerosis (ALS), sensitive spinal motor neurons (SpMN) die while a subset of rostral cranial motor neurons (CrMN) survive. In this work, we optimized a protocol to differentiate CrMNs and SpMNs from human induced pluripotent stem cells (iPSCs) by direct programming and positional patterning. Human iCrMNs are more resistant than iSpMNs to proteotoxic stress and rely on the proteasome to maintain proteostasis. iCrMNs better prevent mislocalization of TDP43 from the nucleus, a hallmark of ALS progression. iSpMNs contain more splicing defects than iCrMNs in response to ALS-related stress with genes involved in splicing and proteostasis maintenance. Therefore, iCrMNs resist ALS at two levels, preventing protein accumulation and reducing splicing defects in response to TDP43 nuclear depletion. Thus, ALS-sensitive iSpMNs appear to enter a downward spiral compromising their ability to maintain proteostasis and splicing.

category: neuroscience


title: Retinotopic tiling of retinal ganglion cell axons at single-cell precision in the mouse superior colliculus

abstract: Retinotopy, like all long-range projections, can arise from the axons themselves or their targets, but details of the connectivity remain unknown. Using two-photon axonal imaging, we functionally mapped the mouse retinocollicular projection at single-cell resolution. We found a near-perfect match between the spatial patterns of retinal ganglion cell axon terminals in the superior colliculus and their corresponding receptive fields, indicating that retinotopy arises from topographic axonal organizations at single-cell precision.


title: Topographic projection of retinal ganglion cell axons at single-cell precision underlies retinotopy in the mouse superior colliculus


title: Xenon LFP Analysis Platform is a Novel Graphical User Interface for Analysis of Local Field Potential from Large-Scale MEA Recordings

abstract:
High-density multi-electrode array (HD-MEA) has enabled neuronal measurements at high spatial resolution to record local field potentials (LFP), extracellular spikes, and network activity with ease. Whilst we have advanced recording systems with over 4000 electrodes, capable of recording data at over 20 kHz, it still presents computational challenges to handle, process, extract, and view information from these large recordings. It can be challenging for researchers to extract and view even a single channel that has more than a billion data points, let alone process a group of channels.

We have created a computational method, and an open-source toolkit built on Python, rendered on a web browser using Plotlys Dash for extracting and viewing the data, and creating interactive visualization. In addition to extracting and viewing entire or small chunks of data sampled at lower or higher frequencies, respectively, it provides a framework to collect user inputs, analyze channel groups, generate raster plots, view quick summary measures for LFP activity, detect and isolate noise channels, and generate plots and visualization in both time and frequency domain. Incorporated into our Graphical User Interface (GUI), we also have created a novel seizure detection method, which can be used to detect the onset of seizures in all or a selected group of channels and provide the following measures of seizures: distance, duration, and propagation across the region of interest.

We demonstrate the utility of this toolkit, using datasets collected from the 3Brain BioCAM duplex system. For the current analysis, we demonstrate the toolkit and methods with a low sampling frequency dataset (300 Hz) and a group of approximately 400 channels. Using this toolkit, we present novel data demonstrating increased seizure propagation speed from slices of Scn1aHet mice compared to littermate controls.

With advances in HD-MEA recording systems with high spatial and temporal resolution, limited tools are available for researchers to view and process these big datasets. We now provide a user-friendly toolkit to analyze LFP activity obtained from large-scale MEA recordings with translatable applications to EEG recordings, and demonstrate the utility of this new graphic user interface with novel biological findings.


title: Effects of variability in manually contoured spinal cord masks on fMRI co-registration and interpretation

abstract: Functional magnetic resonance imaging (fMRI) of the human spinal cord (SC) is an emerging method for understanding neurovascular responses to stimuli. Group-analysis of SC fMRI data involves co-registration of subject-level data to standard space, which requires manual masking of the cord and may result in bias of group-level SC fMRI results. To test this, we examined variability in SC masks drawn in fMRI data from 21 healthy participants from a completed study mapping responses to sensory stimuli of the C7 dermatome. Masks were drawn on temporal mean functional image by eight raters with varying levels of neuroimaging experience, and the rater from the original study acted as an expert reference. Spatial agreement between rater and expert masks was measured using the Dice Similarity Coefficient, and the influence of rater and dataset was examined using ANOVA. Each raters masks were used to register functional data to the PAM50 template. Gray matter-white matter signal contrast of registered functional data was used to evaluate the spatial normalization accuracy across raters. Subject- and group-level analyses of activation during left- and right-sided sensory stimuli were performed for each raters co-registered data. Agreement with the experts SC mask was associated with both rater experience level (F(7,140) = 32.12, P < 2x10-16,{eta} 2 = 0.29) and dataset (F(20,140) = 20.58, P < 2x10-16,{eta} 2 = 0.53). Dataset variations may reflect image quality metrics: the ratio between the signal intensity of spinal cord voxels and surrounding cerebrospinal fluid was correlated with DSC results (p<0.001). As predicted, variability in the manually-drawn masks influenced spatial normalization, and GM:WM contrast in the registered data showed significant effects of rater and dataset (rater: F(8,160) = 23.57, P < 2x10-16,{eta} 2 = 0.24; dataset: F(20,160) = 22.00, P < 2x10-16,{eta} 2 = 0.56). Registration differences propagated into subject-level activation maps which showed rater-dependent agreement with the expert. Although group-level activation maps differed between raters, no systematic bias was identified. Increasing consistency in manual contouring of spinal cord fMRI data improved co-registration and inter-rater agreement in activation mapping, however our results suggest that improvements in image acquisition and post-processing are also critical to address.


title: Females outperform males in spatial learning despite increased amyloid plaques and microgliosis in a TgF344-AD rat model of Alzheimer's disease

abstract: Alzheimers disease (AD) is a progressive neurodegenerative disease and is the sixth leading cause of death in the US. AD is more prevalent in females than males. While estrogen provides neuroprotection in females, sex mediated differences in the development of AD pathology are not fully elucidated. Therefore, a comparison of the events that develop between sexes in the early-stage of AD pathology may reveal new potential targets for more effective therapeutic intervention. To address sex differences, we analyzed early stage 9-month male and female TgF344-AD (Tg-AD) rats, an AD model carrying the APPswe and Presenilin 1 (PS1{Delta}E9) mutations that develops progressive age-dependent AD pathology similar to humans. Using active place avoidance (aPAT) tests that assess hippocampal-dependent spatial learning and memory, we found significant deficits in Tg-AD females compared to wild type females, but no significant difference between the two male genotypes. Moreover, significant sex differences were observed in that Tg-AD females outperformed Tg-AD males in several measures of the aPAT test. Unexpectedly, Tg-AD females displayed higher levels of hippocampal amyloid plaques and amoeboid microglia than their Tg-AD male littermates. Furthermore, Tg-AD females experienced less hippocampal neuronal loss and had higher GluA2 subunit levels than Tg-AD males. Based on our findings, we propose that estrogen may protect females against cognitive impairment at early stages of AD by regulating GluA2 levels independently of amyloid plaque deposition and gliosis. Elucidating this potential protective mechanism of action of estrogen in AD could lead to new targets for early intervention.


title: siRNA-mediated gene knockdown via electroporation in hydrozoan jellyfish embryos

abstract: As the sister group to bilaterians, cnidarians stand in a unique phylogenetic position that provides insights into evolutionary aspects of animal development, physiology, and behavior. While cnidarians are classified into two types, sessile polyps and free-swimming medusae, most studies at the cellular and molecular levels have been conducted on representative polyp-type cnidarians and have established techniques of genetic manipulation. Recently, gene knockdown by delivery of short hairpin RNAs into eggs via electroporation has been introduced to two polyp-type cnidarians, Nematostella vectensis and Hydractinia symbiolongicarpus, enabling systematic loss-of-function experiments. By contrast, current methods of genetic manipulation for medusa-type cnidarians, or jellyfish, are quite limited, and reliable techniques are required to interrogate function of specific genes. Here, we present a method to knockdown target genes by small interfering RNA (siRNA) delivery into fertilized eggs via electroporation, using the hydrozoan jellyfish, Clytia hemisphaerica and Cladonema paciificum. We show that siRNAs targeting endogenous GFP1 and Wnt3 in Clytia efficiently knockdown gene expression and result in known planula phenotypes: loss of green fluorescence and defects in axial patterning, respectively. We also successfully knock down endogenous Wnt3 in Cladonema by siRNAs electroporation, which circumvents the technical difficulty of microinjecting small eggs. Unexpectedly, Wnt3 knockdown in Cladonema does not induce morphological defects in planula, suggesting a distinct mechanism that operates during embryogenesis. Our method for electroporation of siRNAs targeting specific genes is applicable to other animals, including and beyond jellyfish species, and will facilitate the investigation and understanding of myriad aspects of animal development.


title: Myosin waves and a mechanical asymmetry guide the oscillatory migration of Drosophila cardiac progenitors

abstract: Heart development begins with the formation of a tube, as cardiac progenitors migrate from opposite sides of the embryo and meet medially. Defective cardiac progenitor movements cause congenital heart defects. However, the mechanisms of cell migration during early heart development remain poorly understood. We investigated the mechanisms of movement of the Drosophila cardiac progenitors, the cardioblasts. Using quantitative time-lapse microscopy, we found that cardioblasts did not advance monotonically. Instead, cardioblasts took periodic forward and backward steps as they migrated. The forward steps were greater in both amplitude and duration, resulting in net forward movement of the cells. The molecular motor non-muscle myosin II displayed an alternating pattern of localization between the leading and trailing ends of migrating cardioblasts, forming oscillatory waves that traversed the cells. The alternating pattern of myosin polarity was associated with the alternative contraction and relaxation of the leading and trailing edges of the cell. Mathematical modelling predicted that forward migration requires the presence of a stiff boundary at the trailing edge of the cardioblasts. Consistent with this, we found a supracellular actin cable at the trailing edge of the cardioblasts. When we reduced the tension sustained by the actin cable, the amplitude of the backward steps of cardioblasts increased, thus reducing the net forward speed of migration. Together, our results indicate that periodic cell shape changes coupled with a polarized actin cable produce asymmetrical forces that guide cardioblast migration.


title: Strategies for sampling pseudo-absences for species distribution models in complex mountainous terrain

abstract:
O_LIPredictions from species distribution models (SDMs) that rely on presence-only data are strongly influenced by how pseudo-absences are derived. However, which strategies to generate pseudo-absences give rise to faithful SDMs in complex mountainous terrain, and whether species-specific or generic strategies perform better remain open questions.
C_LIO_LIHere, across 500 plant species, we investigated comprehensively how predictions of SDMs at a 93 m spatial resolution are influenced by pseudo-absence strategies, using the complex topography of the Swiss mountains as a model system. We used five generic (random, equal-stratified, proportional-stratified, target, density) and three species-specific (target specific, density specific and geographic specific) approaches to derive pseudo-absence data. We conducted performance tests for each of our eight strategies in combination with (a) spatial bias, generated within our occurrence dataset on sites with highest sampling density, to investigate how this common bias problem influences the performance of pseudo-absence sampling strategies, and (b) a new approach to reduce model extrapolation in environmental space by including background data from all environmental conditions of the study area. SDMs were evaluated against an independent and well-sampled dataset of true presences and absences.
C_LIO_LIThe random, the density (generic), and the geographic specific (species-specific) strategies consistently performed best, even in cases of strong spatial sampling bias in the occurrence data. Including a background of environmentally stratified pseudo-absences improved predictions of species distributions towards environmental extremes, and significantly reduced spatial extrapolations of model predictions in environmental space.
C_LIO_LIOur results indicate that both generic and species-specific pseudo-absence strategies allow estimating robust SDMs and we provide clear recommendations which strategies to choose in complex terrain and when presence data are prone to high sampling bias. In datasets with strong sampling bias, most pseudo-absence strategies produce extrapolation problems and we additionally recommend environmentally stratified pseudo-absences in these cases. Overall, in species rich datasets the use of complex and computationally demanding, species-specific pseudo-absence strategies may not always be justified compared to simpler generic approaches.
C_LI


title: Footprints of domestication in dry-cured meat Penicillium fungi: convergent specific phenotypes and horizontal gene transfers

abstract: Many fungi have been domesticated for food production, with genetic differentiation between populations from food and wild environments, and food populations often acquiring beneficial traits through horizontal gene transfers. We studied the population structures and phenotypes of two distantly related Penicillium species used for dry-cured meat production, P. nalgiovense, the most common species in the dry-cured meat food industry, and P. salamii, used locally by farms. Both species displayed low genetic diversity, with no differentiation between strains isolated from dry-cured meat and those from other environments. Nevertheless, the strains collected from dry-cured meat within each species displayed slower proteolysis and lipolysis than their wild-type conspecifics, and those of P. nalgiovense were whiter. The phenotypes of the non-dry-cured meat strains were more similar to their sister species than to their conspecific dry-cured meat strains, indicating an evolution of specific phenotypes in dry-cured meat strains. A comparison of available Penicillium genomes from various environments revealed evidence of multiple horizontal gene transfers, particularly between P. nalgiovense and P. salamii. Some horizontal gene transfers involving P. biforme, also found in dry-cured meat products, were also detected. We also detected positive and purifying selection based on amino-acid changes. Our genetic and phenotypic findings suggest that human selection has shaped the P. salamii and P. nalgiovense populations used for dry-cured meat production, which constitutes domestication. Several genetic and phenotypic changes were similar in P. salamii, P. nalgiovense, and P. biforme, providing an interesting case of convergent adaptation to the same human-made environment.

category: evolutionary biology


title: Metabolism of glucose activates TORC1 through multiple mechanisms in Saccharomyces cerevisiae

abstract: Target of Rapamycin Complex 1 (TORC1) is a conserved eukaryotic protein complex that links the presence of nutrients with cell growth. In Saccharomyces cerevisiae, TORC1 activity is positively regulated by the presence of amino acids and glucose in the medium. However, mechanisms underlying nutrient-induced TORC1 activation remain poorly understood. By utilizing a TORC1 activation assay, we demonstrate that differential metabolism of glucose activates TORC1 through three distinct pathways in yeast. The first  canonical Rag GTPase-dependent pathway requires conversion of glucose to fructose 1,6-bisphosphate which activates TORC1 via the Rag GTPase heterodimer Gtr1GTP/Gtr2GDP. The second  non-canonical Rag GTPase-dependent pathway requires conversion of glucose to glucose 6-phosphate which activates TORC1 via Gtr1GTP/Gtr2GTP. The third  Rag GTPase-independent pathway requires complete glycolysis and vacuolar ATPase reassembly for TORC1 activation. Glucose-induced TORC1 activation can be uncoupled from glucose-induced AMPK inactivation. We have established a roadmap to deconstruct the link between glucose metabolism and TORC1 activation.


title: Mobile elements in human population-specific genome and phenotype divergence

abstract: Mobile genetic elements (MEs) are heritable mutagens that contribute to divergence between lineages by recursively generating structural variants. ME variants (MEVs) are difficult to genotype, obscuring their impact on recent genome and trait diversification. We developed a tool that uses short-read sequence data to accurately genotype MEVs, enabling us to study them using statistical genetics methods in global human genomes. We observe population-specific differences in the distribution of Alu insertions that distinguish Japanese from other populations. We integrated MEVs with epigenomic and expression quantitative trait loci (eQTL) maps to determine how they impact traits. This reveals coherent patterns by which specific MEs regulate tissue-specific gene expression, including creating or attenuating enhancers and recruiting post-transcriptional regulators. We pinpoint MEVs as genetic causes of disease risk, including a LINE-1 insertion linked to keloid and other diseases of fibroblast inflammation, by introducing MEVs into the genome-wide association study (GWAS) framework. In addition to nominating previously-hidden MEVs as causes of human diseases, this work highlights MEs as accelerators of human population divergence and begins to decipher the semantics of MEs.


title: Multiplexed microfluidic chip for cell co-culture

abstract: Paracrine signaling is challenging to study in vitro, as conventional culture tools dilute soluble factors and offer little to no spatiotemporal control over signaling. Microfluidic chips offer potential to address both of these issues. However, few solutions offer both control over onset and duration of cell-cell communication, and high throughput. We have developed a microfluidic chip designed to culture cells in adjacent chambers, separated by valves to selectively allow or prevent exchange of paracrine signals. The chip features 16 fluidic inputs and 128 individuallyaddressable chambers arranged in 32 sets of 4 chambers. Media can be continuously perfused or delivered by diffusion, which we model under different culture conditions to ensure normal cell viability. Immunocytochemistry assays can be performed in the chip, which we modeled and fine-tuned to reduce total assay time to 1h. Finally, we validate the use of the chip for co-culture studies by showing that HEK293Ta cells respond to signals secreted by RAW 264.7 immune cells in adjacent chambers, only when the valve between the chambers is opened.


title: ScanExitronLR: characterization and quantification of exitron splicing events in long-read RNA-seq data

abstract:
SummaryExitron splicing is a type of alternative splicing where coding sequences are spliced out. Recently, exitron splicing has been shown to increase proteome plasticity and play a role in cancer. Long-read RNA-seq is well suited for quantification and discovery of alternative splicing events; however, there are currently no tools available for detection and annotation of exitrons in long-read RNA-seq data. Here we present ScanExitronLR, an application for the characterization and quantification of exitron splicing events in long-reads. From a BAM alignment file, reference genome and reference gene annotation, ScanExitronLR outputs exitron events at the transcript level. Outputs of ScanExitronLR can be used in downstream analyses of differential exitron splicing. A companion tool, AnnotateExitron, reports exitron annotations such as truncation or frameshift type, nonsense-mediated decay status, and Pfam domain interruptions. We demonstrate that ScanExitronLR performs better on noisy long-reads than currently published exitron detection algorithms designed for short-read data.

Availability and ImplementationScanExitronLR is freely available at https://github.com/ylab-hi/ScanExitronLR and distributed as a pip package on the Python Package Index.

Contactyang4414@umn.edu

Supplementary InformationSupplementary data are available at Bioinformatics online.


title: BET inhibition sensitizes immunologically-cold Rb-deficient prostate cancer to immune checkpoint blockade

abstract:
PurposeNon-T cell-inflamed immunologically "cold" tumor microenvironments (TME) are associated with poor responsiveness to immune checkpoint blockade (ICB), and can be sculpted by tumor cell genomics. Here we evaluated how Retinoblastoma (Rb) tumor suppressor loss of function (LOF), one of the most frequent alterations in human cancer and associated with lineage plasticity, poor prognosis and therapeutic outcomes, alters the TME, and whether therapeutic strategies targeting the molecular consequences of Rb loss enhance ICB efficacy.

Experimental DesignWe performed bioinformatics analysis to elucidate the impact of endogenous Rb LOF on the immune TME in human primary and metastatic tumors. Next, we utilized isogenic murine models of Rb-deficient prostate cancer (PC) for in vitro and in vivo mechanistic studies to examine how Rb loss and bromodomain and extraterminal (BET) domain inhibition (BETi) reprograms the immune landscape, and evaluated in vivo therapeutic efficacy of BETi, singly and in combination with ICB and androgen deprivation therapy.

ResultsRb loss was enriched in non-T cell-inflamed tumors, and Rb-deficient murine tumors demonstrated decreased immune infiltration in vivo. The BETi JQ1 increased immune infiltration into the TME through enhanced tumor cell STING/NF-{kappa}B activation and type I interferon (IFN) signaling within tumor cells, resulting in differential macrophage and T cell-mediated tumor growth inhibition and sensitization of Rb-deficient PC to ICB.

ConclusionsBETi can reprogram the immunologically cold Rb-deficient TME via STING/NF-{kappa}B/IFN signaling to sensitize Rb-deficient PC to ICB. These data provide the mechanistic rationale to test combinations of BETi and ICB in clinical trials of Rb-deficient PC.

STATEMENT OF TRANSLATIONAL RELEVANCERb LOF is one of the most common genomic alterations in human cancer, occurring in approximately 1/3 of advanced malignancies, Furthermore, loss of Rb correlates with enhanced aggressiveness and poor therapeutic outcomes. In this study, we demonstrate that loss of Rb is also associated with an immunosuppressive tumor microenvironment and lack of responsiveness to immune checkpoint blockade (ICB). As a strategy to overcome Rb LOF induced immunosuppression, we have demonstrated that BETi treatment drives STING/NF-{kappa}B signaling and type I interferon production within tumor cells, resulting in immune-mediated tumor control in Rb-deficient PC, which is accentuated by the combination with ICB and ADT. These findings provide a roadmap for maximizing the clinical translation of BET inhibitors into the clinic to treat aggressive-variant Rb-deficient PC.


title: Single Nucleus RNA Sequencing of Pre-Malignant Liver Reveals Disease-Associated Hepatocyte State with HCC Prognostic Potential

abstract: Current approaches to stage chronic liver diseases have limited utility to directly predict liver cancer risk. Here, we employed single nucleus RNA sequencing (snRNA-seq) to characterize the cellular microenvironment of healthy and chronically injured pre-malignant livers using two distinct mouse models. Analysis of 40,748 hepatic nuclei unraveled a previously uncharacterized disease-associated hepatocyte transcriptional state (daHep). These cells were absent in healthy livers, but were increasingly prevalent as chronic liver disease progressed towards hepatocarcinogenesis. Gene expression deconvolution of 1,439 human liver transcriptomes from publicly available datasets revealed that daHep frequencies highly correlate with current histopathological liver disease staging systems. Importantly, we show that high daHep levels precede carcinogenesis in mice and humans and predict a higher risk of hepatocellular carcinoma (HCC) development. This novel transcriptional signature with diagnostic and, more importantly, prognostic significance has the potential to change the way chronic liver disease patients are staged, surveilled and risk-stratified.


title: Genetic mechanisms and biological processes underlying host response to ophidiomycosis (Snake Fungal Disease) inferred from tissue-specific transcriptome analyses.

abstract:
There is growing concern about infectious diseases in wildlife species caused by pathogenic fungi. Detailed knowledge exists about host pathology and the molecular mechanisms underlying host physiological response to some fungal diseases affecting amphibians and bats but is lacking for others with potentially significant impacts on large groups of animals. One such disease is ophidiomycosis (Snake Fungal Disease; SFD) which is caused by the fungus Ophidiomyces ophidiicola and impacts diverse species of snakes. Despite this potential, the biological mechanisms and molecular changes occurring during infection are unknown for any snake species. To gain this information, we performed a controlled experimental infection of captive Prairie rattlesnakes (Crotalus viridis) with O. ophidiicola at different temperatures. We then generated liver, kidney, and skin transcriptomes from control and infected snakes to assess tissue specific genetic responses to infection. Given previous SFD histopathological studies and the fact that snakes are ectotherms, we expected highest fungal activity on skin and a significant impact of temperature on host response. In contrast, we found that most of the differential gene expression was restricted to internal tissues and fungal-infected snakes showed transcriptome profiles indicative of long-term inflammation of specific tissues. Infected snakes at the lower temperature had the most pronounced overall host functional response whereas, infected snakes at the higher temperature had overall expression profiles similar to control snakes possibly indicating recovery from the disease. Overall, our results suggest SFD is a systemic disease with a chronic host response, unlike acute response shown by amphibians to Batrachochytrium dendrobatidis infections. Our analysis also generated a list of candidate protein coding genes that potentially mediate SFD response in snakes, providing tools for future comparative and evolutionary studies into variable species susceptibility to ophidiomycosis.

Author summaryOphidiomycosis (Snake Fungal Disease; SFD) is an infectious fungal disease in snakes that has been documented in more than 40 species over the past 20 years. Though many snake species seem vulnerable to SFD, little is known about how snake physiology changes in response to infection with the causative fungus, Ophidiomyces ophidiicola. Here we report results from the first experimental transcriptomic study of SFD in a snake host. Our goals were to identify genes with a putative role in host response, use this information to understand what biological changes occur in different tissues in snakes when infected with O. ophidiicola, and determine if temperature has an impact in these ectothermic animals. We conclude that SFD is a systemic disease with a chronic inflammation leading to deterioration of internal organs and that these physiological impacts are more pronounced at low rather than high temperatures. These results contrast with fungal infections in amphibians where hosts show an acute response mostly restricted to skin. Our list of candidate genes carry utility in potentially diagnosing genetic susceptibility to SFD in snake species of conservation concern.

category: pathology


title: HPCA1 is required for systemic ROS and calcium cell-to-cell signaling and plant acclimation to stress

abstract: As multicellular organisms, plants constantly balance and coordinate many metabolic, physiological, and molecular responses between different cell types and tissues. This process is essential for plant development, growth, and response to different environmental cues. Because plants lack a nervous system, they transmit different signals over long distances via cell-to-cell signaling. Recent studies revealed that reactive oxygen species (ROS), produced by respiratory burst oxidase homologs (RBOHs) at the apoplast play a key role in cell-to-cell signaling. A state of enhanced ROS production by one cell is thereby sensed by a neighboring cell, causing it to produce ROS, creating a continuous chain of cell-to-cell ROS accumulation termed the  ROS wave. This process was found to mediate systemic signals throughout the plant and is required for plant acclimation to different stresses. Although RBOHs were found to produce ROS essential for this process, the identity of the receptor(s) perceiving the apoplastic ROS signal is currently unknow. Here we reveal that the leucine-rich-repeat receptor-like kinase HPCA1 (H2O2-induced Ca2+ increases 1) acts as a central ROS receptor required for the propagation of cell-to-cell ROS signals, systemic signaling in response to different biotic and abiotic stresses, and plant acclimation to stress. We further report that HPCA1 is required for systemic calcium signals, but not systemic membrane depolarization responses, and identify key calcium-dependent signal transduction proteins involved in this process. Our findings reveal that HPCA1 plays a key role in mediating and coordinating systemic cell-to-cell ROS and calcium signals that are required for plant acclimation to stress.


title: Petascale pipeline for precise alignment of images from serial section electron microscopy

abstract: The reconstruction of neural circuits from serial section electron microscopy (ssEM) images is being accelerated by automatic image segmentation methods. Segmentation accuracy is often limited by the preceding step of aligning 2D section images to create a 3D image stack. Precise and robust alignment in the presence of image artifacts is challenging, especially as datasets are attaining the petascale. We present a computational pipeline for aligning ssEM images with several key elements. Self-supervised convolutional nets are trained via metric learning to encode and align image pairs, and they are used to initialize iterative fine-tuning of alignment. A procedure called vector voting increases robustness to image artifacts or missing image data. For speedup the series is divided into blocks that are distributed to computational workers for alignment. The blocks are aligned to each other by composing transformations with decay, which achieves a global alignment without resorting to a time-consuming global optimization. We apply our pipeline to a whole fly brain dataset, and show improved accuracy relative to prior state of the art. We also demonstrate that our pipeline scales to a cubic millimeter of mouse visual cortex. Our pipeline is publicly available through two open source Python packages.


title: UBQLN2 restrains the domesticated retrotransposon PEG10 to maintain neuronal health in ALS

abstract: Amyotrophic Lateral Sclerosis (ALS) is a fatal, neurodegenerative disease characterized by progressive motor neuron dysfunction and loss. A portion of ALS is caused by mutation of the proteasome shuttle factor Ubiquilin 2 (UBQLN2), but the molecular pathway leading from UBQLN2 dysfunction to disease remains unclear. Here, we demonstrate a function of UBQLN2 in regulating activity of the domesticated gag-pol retrotransposon  paternally expressed gene 10 (PEG10) in human cells and tissues. In cells, the PEG10 gag-pol protein cleaves itself in a mechanism reminiscent of retrotransposon self-processing to generate a liberated  nucleocapsid fragment, which uniquely localizes to the nucleus and changes expression of genes involved in axon remodeling. In spinal cord tissue from ALS patients, PEG10 gag-pol is elevated compared to healthy controls. These findings implicate the retrotransposon-like activity of PEG10 as a contributing mechanism in ALS through regulation of gene expression, and restraint of PEG10 as a primary function of UBQLN2.

category: cell biology