title: The hyperpolarization-activated current shifts the dynamic range of a voltage-dependent electrical synapse

abstract:
Like their chemical counterparts, electrical synapses show complex dynamics such as rectification and voltage dependence that interact with other electrical processes in neurons. The consequences arising from these interactions for the electrical behavior of the synapse, and the dynamics they create, remain largely unexplored. Using a voltage-dependent electrical synapse between a descending modulatory projection neuron (MCN1) and a motor neuron (LG) in the crustacean stomatogastric ganglion, we find that the influence of the hyperpolarization-activated inward current (Ih) is critical to the function of the electrical synapse. When we blocked Ih with CsCl, the apparent voltage dependence of the electrical synapse shifted by 18.7 mV to more hyperpolarized voltages, placing the dynamic range of the electrical synapse outside of the range of voltages used by the LG motor neuron (-60.2 mV - -44.9 mV). With dual electrode current- and voltage-clamp recordings, we demonstrate that this voltage shift is not due to a change in the properties of the gap junction itself, but is a result of a sustained effect of Ih on the presynaptic MCN1 axon terminal membrane potential. Ih-induced depolarization of the axon terminal membrane potential increased the electrical postsynaptic potentials and currents. With Ih present, the axon terminal resting membrane potential depolarized, shifting the dynamic range of the electrical synapse towards the functional range of the motor neuron. We thus demonstrate that the function of an electrical synapse is critically influenced by a voltage-dependent ionic current (Ih).

New & NoteworthyElectrical synapses and voltage-gated ionic currents are often studied independently from one another, despite mounting evidence that their interactions can alter synaptic behavior. We show that the hyperpolarization-activated inward ionic current shifts the voltage dependence of an electrical synaptic transmission through its depolarizing effect on the membrane potential, enabling it to lie within the functional membrane potential range of a motor neuron. Thus, the electrical synapses function critically depends on the voltage-gated ionic current.


title: The dynamic range of voltage-dependent gap junction signaling is maintained by Ih-induced membrane potential depolarization

category: neuroscience


title: Divergent clonal differentiation trajectories of T cell exhaustion

abstract:
T cells activated by chronic antigen exposure in the setting of viral infections or cancer can adopt an exhausted T cell (Tex) state, characterized by reduced effector function and proliferative capacity, and the upregulation of inhibitory receptors. However, whether all antigen-specific T cell clones follow the same molecular and cellular Tex differentiation trajectory remains unclear. Here, we generate a single-cell multi-omic atlas of T cell exhaustion that redefines the phenotypic diversity and molecular regulation of Tex phenotypes. Longitudinal analysis during chronic viral infection identifies an early effector phenotype that is epigenetically primed for Tex differentiation and two late-stage Tex cell states with either a terminal exhaustion or a killer cell lectin-like receptor (KLR)-expressing cytotoxic gene signature. We define clonal trajectories of antigen-specific T cells using paired single-cell RNA and T cell receptor sequencing and reveal distinct differentiation trajectories resulting in terminal Tex-biased, KLR Tex-biased, or divergent clones that differentiate into both phenotypes. Comparison of Tex phenotypes among shared T cell clones that traffic to multiple organs reveals that clonal differentiation trajectories are maintained across tissues. Finally, we show that differences in clonal differentiation trajectory are driven by TCR signal strength, whereby high-affinity T cell clones preferentially adopt a terminal Tex fate, while low-affinity clones adopt an effector-like KLR Tex fate that is detectable long-term but depleted in high antigen settings. These findings reveal clonal heterogeneity in the T cell response to chronic antigen and genomic programs that underlie Tex fates and persistence.

HighlightsO_LIA single-cell atlas of T cell exhaustion identifies novel early effector and KLR Tex states.
C_LIO_LIClonal T cell analysis defines divergent differentiation trajectories during chronic viral infection leading to terminal and KLR Tex fates.
C_LIO_LIThe heterogeneity of the Tex pool arises from three primary differentiation patterns and are differentially persistent in the setting of high antigen.
C_LIO_LIClonal Tex differentiation patterns are conserved across organ sites and driven by TCR signal strength.
C_LI

category: immunology


title: QTQTN motif upstream of the furin-cleavage site plays key role in SARS-CoV-2 infection and pathogenesis.

abstract: The furin cleavage site (FCS), an unusual feature in the SARS-CoV-2 spike protein, has been spotlighted as a factor key to facilitating infection and pathogenesis by increasing spike processing 1,2. Similarly, the QTQTN motif directly upstream of the FCS is also an unusual feature for group 2B coronaviruses (CoVs). The QTQTN deletion has consistently been observed in in vitro cultured virus stocks and some clinical isolates 3. To determine whether the QTQTN motif is critical to SARS-CoV-2 replication and pathogenesis, we generated a mutant deleting the QTQTN motif ({Delta}QTQTN). Here we report that the QTQTN deletion attenuates viral replication in respiratory cells in vitro and attenuates disease in vivo. The deletion results in a shortened, more rigid peptide loop that contains the FCS, and is less accessible to host proteases, such as TMPRSS2. Thus, the deletion reduced the efficiency of spike processing and attenuates SARS-CoV-2 infection. Importantly, the QTQTN motif also contains residues that are glycosylated4, and disruption its glycosylation also attenuates virus replication in a TMPRSS2-dependent manner. Together, our results reveal that three aspects of the S1/S2 cleavage site - the FCS, loop length, and glycosylation - are required for efficient SARS-CoV-2 replication and pathogenesis.

category: microbiology


title: 4D reconstruction of developmental trajectories using spherical harmonics

abstract: Although the full embryonic development of species such as Drosophila and zebrafish can be 3D imaged in real time, this is not true for mammalian organs, as normal organogenesis cannot be recapitulated in vitro. Currently available 3D data is therefore ex vivo images which provide only a snap shot of development at discrete moments in time. Here we propose a computer-based approach to recreate the continuous evolution in time and space of developmental stages from 3D volumetric images. Our method uses the mathematical approach of spherical harmonics to re-map discrete shape data into a space in which facilitates a smooth interpolation over time. We tested our approach on mouse limb buds (from E10 to E12.5) and embryonic hearts (from 10 to 29 somites). A key advantage of the method is that the resulting 4D trajectory takes advantage of all the available data (i.e. it is not dominated by the choice of a few "ideal" images), while also being able to interpolate well through time intervals for which there is little or no data. This method not only provides a quantitative basis for validating predictive models, but it also increases our understanding of morphogenetic processes. We believe this is the first data-driven quantitative 4D description of limb morphogenesis.

category: developmental biology


title: Social evolution of shared biofilm matrix components

abstract: Biofilm formation is an important and ubiquitous mode of growth among bacteria. Central to the evolutionary advantage of biofilm formation is cell-cell and cell-surface adhesion achieved by a variety of factors, some of which are diffusible compounds that may operate as classical public goods - factors that are costly to produce but may benefit other cells. An outstanding question is how diffusible matrix production, in general, can be stable over evolutionary timescales. In this work, using Vibrio cholerae as a model, we show that shared diffusible biofilm matrix proteins are indeed susceptible to cheater exploitation, and that the evolutionary stability of producing these matrix components fundamentally depends on biofilm spatial structure, intrinsic sharing mechanisms of these components, and flow conditions in the environment. We further show that exploitation of diffusible adhesion proteins is localized within a well-defined spatial range around cell clusters that produce them. Based on this exploitation range and the spatial distribution of cell clusters, we construct a model of costly diffusible matrix production and relate these length scales to the relatedness coefficient in social evolution theory. Our results show that production of diffusible biofilm matrix components is evolutionarily stable under conditions consistent with natural biofilm habitats and host environments. We expect the mechanisms revealed in this study to be relevant to other secreted factors that operate as cooperative public goods in bacterial communities, and the concept of exploitation range and the associated analysis tools to be generally applicable.

category: ecology


title: imageseg: an R package for deep learning-based image segmentation

abstract:
O_LIConvolutional neural networks (CNNs) and deep learning are powerful and robust tools for ecological applications. CNNs can perform very well in various tasks, especially for visual tasks and image data. Image segmentation (the classification of all pixels in images) is one such task and can for example be used to assess forest vertical and horizontal structure. While such methods have been suggested, widespread adoption in ecological research has been slow, likely due to technical difficulties in implementation of CNNs and lack of toolboxes for ecologists.
C_LIO_LIHere, we present R package imageseg which implements a workflow for general-purpose image segmentation using CNNs and the U-Net architecture in R. The workflow covers data (pre)processing, model training, and predictions. We illustrate the utility of the package with two models for forest structural metrics: tree canopy density and understory vegetation density. We trained the models using large and diverse training data sets from a variety of forest types and biomes, consisting of 3288 canopy images (both canopy cover and hemispherical canopy closure photographs) and 1468 understory vegetation images.
C_LIO_LIOverall classification accuracy of the models was high with a Dice score of 0.91 for the canopy model and 0.89 for the understory vegetation model (assessed with 821 and 367 images, respectively), indicating robustness to variation in input images and good generalization strength across forest types and biomes.
C_LIO_LIThe package and its workflow allow simple yet powerful assessments of forest structural metrics using pre-trained models. Furthermore, the package facilitates custom image segmentation with multiple classes and based on color or grayscale images, e.g. in cell biology or for medical images. Our package is free, open source, and available from CRAN. It will enable easier and faster implementation of deep learning-based image segmentation within R for ecological applications and beyond.
C_LI


title: A seasonal pulse of ungulate neonates influences space use by carnivores in a multi-predator, multi-prey system

abstract:
O_LIThe behavioral mechanisms by which predators encounter prey are poorly resolved. In particular, the extent to which predators engage in active search for prey versus incidentally encountering them is unknown. The distinction between search and incidental encounter influences prey population dynamics with active search exerting a stabilizing force on prey populations by alleviating predation pressure on low-density prey and increasing it for high-density prey.
C_LIO_LIParturition of many large herbivores occurs during a short and predictable temporal window in which young are highly vulnerable to predation. Our study aims to determine how a suite of carnivores responds to the seasonal pulse of newborn ungulates using contemporaneous GPS locations of four species of predators and two species of prey.
C_LIO_LIWe used step-selection functions to assess whether coyotes, cougars, black bears, and bobcats actively searched for parturient females in a low-density population of mule deer and a high-density population of elk. We then assessed whether searching carnivores shifted their habitat use toward areas exhibiting a high probability of encountering neonates.
C_LIO_LINone of the four carnivore species encountered parturient mule deer more often than expected by chance suggesting that predation of young resulted from incidental encounters. By contrast, we determined that cougar and male bear movements positioned them in proximity of parturient elk more often than expected by chance which is evidence of searching behavior. Although both male bears and cougars searched for neonates, only male bears used elk parturition habitat in a way that dynamically tracked the phenology of the elk birth pulse suggesting that maximizing encounters with juvenile elk was a motivation when selecting resources.
C_LIO_LIOur results support the existence of a stabilizing mechanism to prey populations through active search behavior by predators because carnivores in our study searched for the high-density prey species (elk) but ignored the low-density species (mule deer). We conclude that prey density must be high enough to warrant active search, and that there is high interspecific and intersexual variability in foraging strategies among large mammalian predators and their prey.
C_LI


title: Genomic epidemiological models describe pathogen evolution across fitness valleys

abstract: Genomics is fundamentally changing epidemiological research. However, exploring hypotheses about pathogen evolution in different epidemiological contexts poses new challenges. Models intertwining pathogen epidemiology and genomic evolution can help understand processes such as the emergence of novel pathogen genotypes with higher transmission or resistance to treatment. In this work, we present Opqua, a computational framework for flexible simulation of pathogen epidemiology and evolution. We use Opqua to study determinants of evolution across fitness valleys. We confirm that competition can limit evolution in high transmission environments and find that low transmission, host mobility, and complex pathogen life cycles facilitate reaching new adaptive peaks through population bottlenecks and decoupling selective pressures. The results show the potential of genomic epidemiological modeling as a tool in infectious disease research.

category: evolutionary biology


title: Dissecting supergene: evidence for recombination suppression among multiple functional loci within inversions.

abstract: Supergenes are genetic architectures associated with discrete and concerted variation in multiple traits. It has long been suggested that supergenes control these complex polymorphisms by suppressing recombination between set of coadapted genes. However, because recombination suppression hinders the dissociation of the individual effects of genes within supergenes, there is still little evidence that supergenes evolve by tightening linkage between coadapted genes. Here, combining an landmark-free phenotyping algorithm with multivariate genome wide association studies, we dissected the genetic basis of wing pattern variation in the butterfly Heliconius numata. We showed that the supergene controlling the striking wing-pattern polymorphism displayed by this species contains many independent loci associated with different features of wing patterns. The three chromosomal inversions of this supergene suppress recombination between these loci, supporting the hypothesis that they may have evolved because they captured beneficial combinations of alleles. Some of these loci are associated with colour variations only in morphs controlled by inversions, indicating that they were recruited after the formation of these inversions. Our study shows that supergenes and clusters of adaptive loci in general may form via the evolution of chromosomal rearrangements suppressing recombination between co-adapted loci but also via the subsequent recruitment of linked adaptive mutations.


title: Supergene formation: evidence for recombination suppression among multiple functional loci within inversions.


title: The genome sequence of the lesser marbled fritillary, Brenthis ino, and evidence for a segregating neo-Z chromosome

abstract: The lesser marbled fritillary, Brenthis ino (Rottemburg, 1775), is a species of Palearctic butterfly. Male B. ino individuals have been reported to have between 12 and 14 pairs of chromosomes, a much reduced chromosome number than is typical in butterflies. Here we present a chromosome-level genome assembly for B. ino, as well as gene and transposable element annotations. The assembly is 411.8 Mb in span with contig and scaffold N50s of 9.6 and 29.5 Mb respectively. We also show evidence that the male individual from which we generated HiC data was heterozygous for a neo-Z chromosome, consistent with inheriting 14 chromosomes from one parent and 13 from the other. This genome assembly will be a valuable resource for studying chromosome evolution in Lepidoptera, as well as for comparative and population genomics more generally.


title: A practical guide for generating unsupervised, spectrogram-based latent space representations of animal vocalizations

abstract: The manual detection, analysis, and classification of animal vocalizations in acoustic recordings is laborious and requires expert knowledge. Hence, there is a need for objective, generalizable methods that detect underlying patterns in these data, categorize sounds into distinct groups, and quantify similarities between them. Among all computational methods that have been proposed to accomplish this, neighborhood-based dimensionality reduction of spectrograms to produce a latent-space representation of calls stands out for its conceptual simplicity and effectiveness. Using a dataset of manually annotated meerkat (Suricata suricatta) vocalizations, we demonstrate how this method can be used to obtain meaningful latent space representations that reflect the established taxonomy of call types. We analyze strengths and weaknesses of the proposed approach, give recommendations for its usage and show application examples, such as the classification of ambiguous calls and the detection of mislabeled calls. All analyses are accompanied by example code to help researchers realize the potential of this method for the study of animal vocalizations.

category: animal behavior and cognition


title: Isolation disrupts social interactions and destabilizes brain development in bumblebees

abstract: Social isolation, particularly in early life, leads to deleterious physiological and behavioral outcomes. Few studies, if any, have been able to capture the behavioral and neurogenomic consequences of early life social isolation together in a single social animal system. Here, we leverage new high-throughput tools to comprehensively investigate the impact of isolation in the bumblebee (Bombus impatiens) from behavioral, molecular, and neuroanatomical perspectives. We reared newly emerged bumblebees either in complete isolation, small groups, or in their natal colony, and then analyzed their behaviors while alone or paired with another bee. We find that when alone, individuals of each rearing condition show distinct behavioral signatures. When paired with a conspecific, bees reared in small groups or in the natal colony express similar behavioral profiles. Isolated bees, however, showed increased social interactions. To identify the neurobiological correlates of these differences, we quantified brain gene expression and measured the volumes of key brain regions for a subset of individuals from each rearing condition. Overall, we find that isolation increases social interactions and disrupts gene expression and brain development. Limited social experience in small groups is sufficient to preserve typical patterns of brain development and social behavior.


title: Cancer metabolic subtypes and their association with molecular and clinical features

abstract:
Alterations of metabolic pathways in cancer have been investigated for many years, beginning way before the discovery of the role of oncogenes and tumor suppressors, and the last few years have witnessed a renewed interest in this topic. Large-scale molecular and clinical data on tens of thousands of samples allow us today to tackle the problem from a general point of view.

Here we show that transcriptomic profiles of tumors can be exploited to define metabolic cancer subtypes, which can be systematically investigated for association with other molecular and clinical data. We find thousands of significant associations between metabolic subtypes and molecular features such as somatic mutations, structural variants, epigenetic modifications, protein abundance and activation; and with clinical/phenotypic data including survival probability, tumor grade, and histological types.

Our work provides a methodological framework and a rich database of statistical associations, accessible from metaminer.unito.it, that will contribute to the understanding of the role of metabolic alterations in cancer and to the development of precision therapeutic strategies.

category: cancer biology


title: Fat Body Phospholipid State Dictates Hunger Driven Feeding Behavior

abstract:
Diet-induced obesity (DIO) leads to dysfunctional feeding behavior. But the precise molecular nodes that are dysregulated by DIO that alter satiety sensing and feeding motivation are not fully disentangled. The fruit fly is a simple genetic model system yet displays significant evolutionary conservation to mammalian nutrient sensing and energy balance. Using a longitudinal high sugar regime in Drosophila, we sought to address how lipid alteration in fat cells alters feeding motivation. We find that long-term exposure to an HSD increases baseline feeding in flies. However, prolonged exposure to HSD degrades the hunger-driven feeding (HDF) response. Lipidomics analysis reveals that longitudinal exposure to HSD significantly alters whole body phospholipid profiles. Then, performing a systematic screen for phospholipid enzymes, we identify a specific enzyme, PECT, a rate-limiting enzyme in the phosphatidylethanolamine (PE) biosynthesis pathway and the fly ortholog of human PCYT2, was critical to maintaining hunger-driven feeding motivation. We show that disrupting PECT only in the fat body causes insulin-resistant phenotypes and a loss of hunger-driven feeding. Excitingly, we find that overexpression of PECT restores HSD induced loss of hunger-driven feeding response. Strikingly human studies have noted a correlation between PCYT2/PECT levels and clinical obesity. Now, our unbiased studies in Drosophila provide specific genetic evidence for PECT in maintaining nutrient sensing during DIO. Our study provides novel insights on the role of phospholipids in interorgan communication of nutrient status.

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC="FIGDIR/small/472854v3_ufig1.gif" ALT="Figure 1">
View larger version (41K):
org.highwire.dtl.DTLVardef@1486dfaorg.highwire.dtl.DTLVardef@2c38beorg.highwire.dtl.DTLVardef@1415fc8org.highwire.dtl.DTLVardef@f75aae_HPS_FORMAT_FIGEXP  M_FIG C_FIG

category: physiology


category: genetics


title: Seed-coat thickness explains contrasting germination responses to smoke and heat by Leucadendron species

abstract:
Fire stimulates germination of most seeds in fire-prone vegetation. Fruits of Leucadendron (Proteaceae) are winged achenes or nutlets that correlate with their requirements for smoke and/or heat in promoting germination. We describe five possible smoke-heat dormancy-release/germination syndromes among plants, of which Leucadendron displays three (no response, smoke only, smoke and heat). As seed-coat thickness varies with seed-storage location (plant or soil) and morphology (winged or wingless), we tested its possible role in water uptake and germination. Species with winged seeds achieved 100% germination in the absence of smoke/heat, seed coats were an order of magnitude thinner, and their permeability greatly exceeded that of nutlets. As seed-coat thickness increased a) imbibitional water uptake declined at a decreasing rate, and b) the response to smoke, and to a lesser extent heat, increased linearly to reach levels of germination approaching those of winged seeds. For species responsive to smoke and heat, there was no additive effect when applied together, suggesting that they may have promoted the same physiological process. By what mechanisms a) the smoke response is greater the thicker the seed coat, and b) smoke chemicals could increase water permeability to explain the non-additive effect of smoke and heat, warrant further investigation.

HighlightWe show Leucadendron seeds are either plant-stored with thin, highly permeable seed-coats that germinate readily; or soil-stored and the thicker their seed-coat, the lower their permeability and greater their need for smoke/heat to promote germination.

category: plant biology


title: A recurring YYDRxG pattern in broadly neutralizing antibodies to a conserved site on SARS-CoV-2, variants of concern, and related viruses

abstract:
Studying the antibody response to SARS-CoV-2 informs on how the human immune system can respond to antigenic variants as well as other SARS-related viruses. Here, we structurally and functionally characterized a potent human antibody ADI-62113 that also neutralizes SARS-CoV- 2 variants of concern and cross-reacts with many other sarbecoviruses. A YYDRxG motif encoded by IGHD3-22 in CDR H3 facilitates targeting to a highly conserved epitope on the SARS-CoV-2 receptor binding domain. A computational search for a YYDRxG pattern in publicly available sequences identified many antibodies with broad neutralization activity against SARS-CoV-2 variants and SARS-CoV. Thus, the YYDRxG motif represents a common convergent solution for the human humoral immune system to counteract sarbecoviruses. These findings also suggest an epitope targeting strategy to identify potent and broadly neutralizing antibodies that can aid in the design of pan-sarbecovirus vaccines and antibody therapeutics.

Short SummaryDecryption of a recurrent sequence feature in anti-SARS-CoV-2 antibodies identifies how potent pan-sarbecovirus antibodies target a conserved epitope on the receptor binding domain.


title: Distinguishing COVID-19 infection and vaccination history by T cell reactivity

abstract: SARS-CoV-2 infection and COVID-19 vaccines elicit memory T cell responses. Here, we report the development of two new pools of Experimentally-defined T cell epitopes derived from the non-spike Remainder of the SARS-CoV-2 proteome (CD4RE and CD8RE). The combination of T cell responses to these new pools and Spike (S) were used to discriminate four groups of subjects with different SARS-CoV-2 infection and COVID-19 vaccine status: non-infected, non-vaccinated (I-V-); infected and non-vaccinated (I+V-); infected and then vaccinated (I+V+); and non-infected and vaccinated (I-V+). The overall classification accuracy based on 30 subjects/group was 89.2% in the original cohort and 88.5% in a validation cohort of 96 subjects. The T cell classification scheme was applicable to different mRNA vaccines, and different lengths of time post-infection/post-vaccination. T cell responses from breakthrough infections (infected vaccinees, V+I+) were also effectively segregated from the responses of vaccinated subjects using the same classification tool system. When all five groups where combined, for a total of 239 different subjects, the classification scheme performance was 86.6%. We anticipate that a T cell-based immunodiagnostic scheme able to classify subjects based on their vaccination and natural infection history will be an important tool for longitudinal monitoring of vaccination and aid in establishing SARS-CoV-2 correlates of protection.


title: An infectious SARS-CoV-2 B.1.1.529 Omicron virus escapes neutralization by several therapeutic monoclonal antibodies

abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the global COVID-19 pandemic resulting in millions of deaths worldwide. Despite the development and deployment of highly effective antibody and vaccine countermeasures, rapidly-spreading SARS-CoV-2 variants with mutations at key antigenic sites in the spike protein jeopardize their efficacy. Indeed, the recent emergence of the highly-transmissible B.1.1.529 Omicron variant is especially concerning because of the number of mutations, deletions, and insertions in the spike protein. Here, using a panel of anti-receptor binding domain (RBD) monoclonal antibodies (mAbs) corresponding to those with emergency use authorization (EUA) or in advanced clinical development by Vir Biotechnology (S309, the parent mAbs of VIR-7381), AstraZeneca (COV2-2196 and COV2-2130, the parent mAbs of AZD8895 and AZD1061), Regeneron (REGN10933 and REGN10987), Lilly (LY-CoV555 and LY-CoV016), and Celltrion (CT-P59), we report the impact on neutralization of a prevailing, infectious B.1.1.529 Omicron isolate compared to a historical WA1/2020 D614G strain. Several highly neutralizing mAbs (LY-CoV555, LY-CoV016, REGN10933, REGN10987, and CT-P59) completely lost inhibitory activity against B.1.1.529 virus in both Vero-TMPRSS2 and Vero-hACE2-TMPRSS2 cells, whereas others were reduced ([~]12-fold decrease, COV2-2196 and COV2-2130 combination) or minimally affected (S309). Our results suggest that several, but not all, of the antibody products in clinical use will lose efficacy against the B.1.1.529 Omicron variant and related strains.


title: Genome-wide, integrative analysis of microRNA-mediated gene regulation in human wounds reveals cooperating microRNAs as therapeutic targets

abstract: MicroRNAs (miR) are important posttranscriptional regulators and exhibit a high potential to be utilized in diagnosis and therapy. However, our insufficient knowledge of the miR-mediated gene regulation in human skin wound healing severely hinders the identification of clinically relevant miRs. Here, we performed paired small RNA and long RNA sequencing in human tissue samples, including matched skin and acute wounds collected at each healing stage and chronic non-healing venous ulcers (VU). With integrative small and long RNA-omics analysis, we developed a compendium (https://www.xulandenlab.com/humanwounds-mirna-mrna), which will be an open, comprehensive resource to broadly aid wound healing research. With this first clinical, wound-centric resource of miRs and mRNAs, we identified 17 pathologically relevant miRs that exhibited abnormal VU expression and displayed their targets enriched explicitly in the VU gene signature. Intermeshing regulatory networks controlled by these miRs revealed their high cooperativity in contributing to chronic wound pathology characterized by persistent inflammation and proliferative phase initiation failure. Furthermore, we demonstrated that miR-34a, miR-424, and miR-516, upregulated in VU, cooperatively suppressed keratinocyte growth while promoting inflammatory response. Collectively, our study opens the possibility of developing innovative wound treatment that targets pathologically relevant cooperating miRs to attain higher therapeutic efficacy and specificity.

category: systems biology


title: Integrative small and long RNA-omics analysis of human healing and non-healing wounds discovers cooperating microRNAs as therapeutic targets


title: A dual reporter system identifies an intermediate state and sequential regulators of 2-cell-like-to-pluripotent state transition

abstract: Mouse embryonic stem cells (ESCs) cycle in and out of 2-cell-like (2C-like) state in culture. The molecular mechanism governing the exit of 2C-like state remains obscure, partly due to the lack of a reporter system that can genetically mark intermediate states during exiting process. Here, we identify an intermediate state that is marked by the co-expression of MERVL::tdTomato and OCT4-GFP (MERLOT) during 2C-like-to-pluripotent state transition (2CLPT). Transcriptome and epigenome analyses demonstrate that MERLOT cells cluster closely with 8-16 cell stage mouse embryos, suggesting that 2CLPT partly mimics early preimplantation development. Through a CRISPRa screen, we identify an ARRDC3-NEDD4-OCT4 regulatory axis that plays an essential role in controlling 2CLPT. Furthermore, re-evaluating previously reported 2C-like state regulators reveals dual function of Chaf1a in regulating the entry and exit of 2C-like state. Finally, ATAC-Seq footprinting analysis uncovers Klf3 as an essential transcription factor required for efficient 2CLPT. Together, our study identifies a genetically traceable intermediate state during 2CLPT and provides a valuable tool to study molecular mechanisms regulating this process.

category: cell biology


title: Drosophila hemocytes recognize lymph gland tumors of mxc mutants and activate the Toll pathway in reactive oxygen species-dependent manner

abstract:
Mechanisms of cancer cell recognition and elimination by the innate immune system remains unclear. Circulating hemocytes are associated with the hematopoietic tumors in Drosophila mxcmbn1 mutant larvae. The innate immune signalling pathways are activated in the fat body to suppress the tumor growth by inducing antimicrobial peptides (AMP). Here, we investigated the regulatory mechanism underlying the activation in the mutant. Reactive oxygen species accumulated in the hemocytes due to induction of dual oxidase and its activator. The hemocytes were also localized on the fat body. These were essential for transmitting the information on tumors toward the fat body to induce AMP expression. Regarding to the tumor recognition, we found that matrix metalloproteinase 1 (MMP1) and MMP2 were highly expressed in the tumors. Ectopic expression of MMP2 was associated with AMP induction in the mutants. Furthermore, the basement membrane components in the tumors were reduced and ultimately lost. The hemocytes may recognize the disassembly in the tumors. Our findings highlight the underlying mechanism via which macrophage-like hemocytes recognize tumor cells and relay the information toward the fat body to induce AMPs. and contribute to uncover the immune systems roles against cancer.

SUMMARY STATEMENTDrosophila blood cells transmit information regarding the existence of tumor cells to the fat body and induce the production of anti-tumor peptides via the generation of reactive oxygen species.


title: Inferring community assembly processes from functional seed trait variation along temperature gradient

abstract:
O_LIAssembly of plant communities has long been scrutinized through the lens of trait-based ecology. Studies generally analyze functional traits related to the vegetative growth, survival and resource acquisition and thus ignore how ecological processes may affect plants at other stages of their lifecycle, particularly when seeds disperse, persist in soil and germinate.
C_LIO_LIHere, we analyzed an extensive data set of 16 traits for 167 species measured in-situ in 36 grasslands located along an elevational gradient and compared the impact of abiotic filtering, biotic interactions and dispersal on traits reflecting different trait categories: plant vegetative growth, germination, dispersal, and seed morphology. For each community, we quantified community weighted mean (CWM) and functional diversity (FD) for all traits and established their relationships to mean annual temperature.
C_LIO_LIThe seed traits were weakly correlated to vegetative traits and thus constituted independent axes of plant phenotypical variation that were affected differently by the ecological processes considered. Abiotic filtering impacted mostly the vegetative traits and to a lesser extent on seed germination and morphological traits. Increasing low-temperature stress towards colder sites selected for short-stature, slow-growing and frost-tolerant species that produce small quantity of smaller seeds with higher degree of dormancy, high temperature requirements for germination and comparatively low germination speed.
C_LIO_LIBiotic interactions, specifically competition in the lowlands and facilitation in uplands, also filtered certain functional traits in the study communities. The benign climate in lowlands promoted plant with competitive strategies including fast growth and resource acquisition (vegetative growth traits) and early and fast germination (germination traits), whereas the effects of facilitation on the vegetative and germination traits were cancelled out by the strong abiotic filtering.
C_LIO_LIThe changes in the main dispersal vector from zoochory to anemochory along the gradient strongly affected the dispersal and the seed morphological trait structure of the communities. Specifically, stronger vertical turbulence and moderate warm-upwinds combined with low grazing intensity selected for light and non-round shaped seeds with lower terminal velocity and endozoochorous potential.
C_LIO_LISynthesis. We clearly demonstrate that, in addition to vegetation traits, seed traits can substantially contribute to functional structuring of plant communities along environmental gradients. Thus, the, hard seed traits related to germination and dispersal are critical to detect multiple, complex community assembly rules. Consequently, such traits should be included in core lists of plant traits and, when applicable, be incorporated into analysis of community assembly.
C_LI


title: Genome Assemblies of the Warthog and Kenyan Domestic Pig Provide Insights into Suidae Evolution and Candidate Genes for African Swine Fever Tolerance

abstract: As warthog (Phacochoerus africanus) has innate immunity against African swine fever (ASF), it is critical to understand the evolutionary novelty of warthog to explain its specific ASF resistance. Here, we present two completed new genomes of one warthog and one Kenyan domestic pig, as the fundamental genomic references to decode the genetic mechanism on ASF tolerance. Our results indicated, multiple genomic variations, including gene losses, independent contraction and expansion of specific gene families, likely moulded warthogs genome to adapt the environment. Importantly, the analysis of the presence and absence of genomic sequences revealed that, the warthog genome had a DNA sequence absence of the lactate dehydrogenase B (LDHB) gene on chromosome 2 compared to the reference genome. The overexpression and siRNA of LDHB indicated that its inhibition on the replication of ASFV. Combining with large-scale sequencing data of 123 pigs from all over the world, contraction and expansion of TRIM genes families revealed that TRIM family genes in the warthog genome were potentially responsible for its tolerance to ASF. Our results will help further improve the understanding of genetic resistance ASF in pigs.


title: CONGA: Copy number variation genotyping in ancient genomes and low-coverage sequencing data

abstract: AO_SCPLOWBSTRACTC_SCPLOWTo date, ancient genome analyses have been largely confined to the study of single nucleotide polymorphisms (SNPs). Copy number variants (CNVs) are a major contributor of disease and of evolutionary adaptation, but identifying CNVs in ancient shotgun-sequenced genomes is hampered by (i) most published genomes being <1 x coverage, (ii) ancient DNA fragments being typically <80 bps. These characteristics preclude state-of-the-art CNV detection software to be effectively applied to ancient genomes. Here we present CONGA, an algorithm tailored for genotyping deletion and duplication events in genomes with low depths of coverage. Simulations and down-sampling experiments show that CONGA can genotype deletions >1 kbps with F-scores >0.75 at [≥]1 x, and distinguish between heterozygous and homozygous states. Using CONGA, we analyse deletion events at 10,018 loci in 56 ancient human genomes spanning the last 50,000 years, with coverages 0.4 x -26 x. We show that inter-individual genetic diversity measured using deletions and SNPs are highly correlated, as in modern-day genomes, confirming that deletion frequencies broadly reflect demographic history. We also identify signatures of strong purifying selection on deletions in ancientgenomes, such as an excess of singletons compared to those in SNPs. CONGA paves the way for systematic studies of drift, mutation load, and adaptation in ancient and modern-day gene pools through the lens of CNVs.

category: genomics


title: The grape MYB24 mediates the coordination of light-induced terpene and flavonol accumulation in response to berry anthocyanin sunscreen depletion

abstract: The presence of naturally-occurring color mutants in plants has permitted the identification of many regulatory genes implicated in the synthesis of discrete metabolic compounds, mostly anthocyanins and carotenoids. Conversely, transcription factors that coordinate more than one specialized metabolic pathway seem challenging to screen from a forward genetics perspective. We explored the relationship between different branches of the phenylpropanoid and isoprenoid pathways while examining an infrequent berry skin color variegation in grapevine. Red and white berry skin sections were compared at the genetic, transcriptomic and metabolomic levels showing that, as in most cultivated white grape varieties, the uncolored skin section convened the non-functional alleles of the anthocyanin regulators MYBA1 and MYBA2, explaining the lack of pigments. In contrast, light-responsive flavonols and monoterpenes increased in anthocyanin-depleted areas. We disclosed an enrichment of the flavonol, terpene and carotenoid pathways among up-regulated genes from white-skin sections, accompanied by increased expressions of flavonol regulators and the still uncharacterized MYB24 gene. We used DAP-seq to examine the in vitro binding of affinity-purified MYB24 protein to genomic DNA and demonstrated its binding in the promoter regions of terpene (22) and carotenoid (6) genes, in addition to more than 30 photosynthesis and light-response genes, including the flavonol-regulator HY5 homologue (HYH). We confirmed the activation of TPS35 and HYH promoter:luciferase reporters in the presence of MYB24 and the grape bHLH MYC2, all of which correlate in their higher expression in white skin variegated sections. The integration of several datasets allowed to define a list of high confidence targets, suggesting MYB24 as a modulator of light responses including the synthesis of flavonoids (flavonols) and isoprenoids (terpenes, and putatively carotenoids). The correspondence between MYB24 and monoterpenes in all conditions surveyed implies that this regulatory network is broadly triggered towards berry ripening, and that the absence of anthocyanin sunscreens accelerates its activation most likely in a dose-dependent manner due to increased radiation exposure.


title: Mirtazapine treatment in a young female mouse model of Rett syndrome identifies time windows for the rescue of early phenotypes during development

abstract: Rett Syndrome (RTT) is a rare X-linked neurodevelopmental disorder, mainly caused by mutations in the MECP2 gene. Reduction in monoamine levels in RTT patients and mouse models suggested the possibility to rescue clinical phenotypes through antidepressants. Accordingly, we tested mirtazapine (MTZ), a noradrenergic and specific-serotonergic tetracyclic antidepressant (NaSSA). In previous studies, we showed high tolerability and significant positive effects of MTZ in male Mecp21m1.1Bird-knock-out mice, adult female Mecp2tm1.1Bird-heterozygous (Mecp2+/-) mice, and adult female RTT patients. However, it remained to explore MTZ efficacy in female Mecp2+/- mice at young ages. As RTT-like phenotypes in young Mecp2+/- mice have been less investigated, we carried out a behavioural characterization to analyze Mecp2+/- mice in "early adolescence" (6 weeks) and "late adolescence/young adulthood" (11 weeks) and identified several progressive phenotypes. Then, we evaluated the effects of either a 15- or a 30-day MTZ treatment on body weight and impaired motor behaviours in 11-week-old Mecp2+/- mice. Finally, since defective cortical development is a hallmark of RTT, we performed a histological study on the maturation of perineuronal nets (PNNs) and parvalbuminergic (PV) neurons in the primary motor cortex. The 30-day MTZ treatment was more effective than the shorter 15-day treatment, leading to the significant rescue of body weight, hindlimb clasping and motor learning in the accelerating rotarod test. Behavioral improvement was associated with normalized PV immunoreactivity levels and PNN thickness. These results support the use of MTZ as a new potential treatment for adolescent girls affected by RTT and suggest a possible mechanism of action.


title: Low Frequency Oscillations drive EEG's complexity changes during wakefulness and sleep

abstract: Recently, the sleep-wake states have been analysed using novel complexity measures, complementing the classical analysis of EEGs by frequency bands. This new approach consistently shows a decrease in EEGs complexity during slow-wave sleep, yet it is unclear how cortical oscillations shape these complexity variations. In this work, we analyse how the frequency content of brain signals affects the complexity estimates in freely moving rats. We find that the low-frequency spectrum - including the Delta, Theta, and Sigma frequency bands - drives the complexity changes during the sleep-wake states. This happens because low-frequency oscillations emerge from neuronal population patterns, as we show by recovering the complexity variations during the sleep-wake cycle from micro, meso, and macroscopic recordings. Moreover, we find that the lower frequencies reveal synchronisation patterns across the neocortex, such as a sensory-motor decoupling that happens during REM sleep. Overall, our works shows that EEGs low frequencies are critical in shaping the sleep-wake states complexity across cortical scales.


title: Nanoscale patterning of in vitro neuronal circuits

abstract: Methods for patterning neurons in vitro have gradually improved and are used to investigate questions difficult to address in or ex vivo. Though these techniques guide axons between groups of neurons, multiscale control of neuronal connectivity, from circuits to synapses, is yet to be achieved in vitro. As studying neuronal circuits with synaptic resolution in vivo poses significant challenges, an in vitro alternative could serve as a testbed for in vivo experiments or as a platform for validating biophysical and computational models. In this work we use a combination of electron beam and photolithography to create polydimethylsiloxane (PDMS) structures with features ranging from 150 nanometers to a few millimeters. Leveraging the difference between average axon and dendritic spine diameters, we restrict axon growth while allowing spines to pass through nanochannels to guide synapse formation between small groups of neurons (i.e. nodes). We show this technique can be used to generate large numbers of isolated feed-forward circuits where connections between nodes are restricted to regions connected by nanochannels. Using a genetically encoded calcium indicator in combination with fluorescently tagged post synaptic protein, PSD-95, we demonstrate functional synapses can form in this region. Although more work needs to be done to control connectivity in vitro, we believe this is a significant step in that direction.


title: Population coding and oscillatory subspace synchronization integrate context into actions

abstract: Contextual cues and prior evidence guide human goal-directed behavior. To date, the neurophysiological mechanisms that implement contextual priors to guide subsequent actions remain unclear. Here we demonstrate that increasing behavioral uncertainty introduces a shift from an oscillatory to a continuous processing mode in human prefrontal cortex. At the population level, we found that oscillatory and continuous dynamics reflect dissociable signatures that support distinct aspects of encoding, transmission and execution of context-dependent action plans. We show that prefrontal population activity encodes predictive context and action plans in serially unfolding orthogonal subspaces, while prefrontal-motor theta oscillations synchronize action-encoding population subspaces to mediate the hand-off of action plans. Collectively, our results reveal how two key features of large-scale population activity, namely continuous population trajectories and oscillatory synchrony, operate in concert to guide context-dependent human behavior.


title: Tryptophol acetate and tyrosol acetate, metabolites secreted by a probiotic yeast, halt cytokine storm

abstract: Probiotic fermented foods are perceived as contributing to human health and capable of protecting against inflammation, however solid mechanistic evidence for the presumptive therapeutic benefits is lacking. Here we report that tryptophol acetate and tyrosol acetate, small molecule metabolites secreted by the probiotic milk-fermented yeast Kluyveromyces marxianus exhibit remarkable anti-inflammatory properties. Comprehensive in vivo, ex vivo and in vitro experiments, employing LPS-induced  cytokine storm models, reveal dramatic effects of the two molecules, added in tandem, on mice morbidity, laboratory parameters and mortality. In parallel, significant attenuation of pro-inflammatory cytokines including IL-6, IL-1, IL-1{beta} and TNF-, and reduction of reactive oxygen species were recorded. Importantly, tryptophol acetate and tyrosol acetate did not completely suppress cytokine generation, but rather brought their concentrations back to baseline levels, further maintaining core immune functions, including phagocytosis. The anti-inflammatory effects of tryptophol acetate and tyrosol acetate were mediated through downregulation of TLR4, IL-1R, and TNFR signaling pathways and increased A20 expression, attenuating NF-{kappa}B level. In addition, the two molecules had a significant impact on mice microbiome, increasing the abundance of the genus Bactericides, known to exhibit anti-inflammatory properties. Overall, this work illuminates pronounced and broad-based immune modulation properties of probiotic yeast-secreted metabolites, uncovering their mechanism of action and underscoring potential new therapeutic avenues for severe inflammation.


title: LONG-TERM MONITORING OF THE EUROPEAN ROLLER (CORACIAS GARRULUS) IN UKRAINE: IS CLIMATE BEHIND THE CHANGES?

abstract: Since the 1980s there has been a long-term decline in numbers and contraction of range in Europe, including Ukraine. Our specific goals were to reconstruct the climatically suitable range of the species in Ukraine before the 1980s, gain better knowledge on its requirements, compare the past and current suitable areas, infer the regional and environmental variables that best explain its occurrence, and quantify the overall range change in the country. For these purposes we created a database consisting of 347 records of the roller made ever in Ukraine. We employed a species distribution modeling (SDM) approach to hindcast changes in the suitable range of the roller during historical times across Ukraine and to derive spatially explicit predictions of climatic suitability for the species under current climate. SDMs were created for three time intervals (before 1980, 1985-2009, 2010-2021) using corresponding climate data extracted from the TerraClim database. SDMs show a decline of suitable for rollers areas in the country from 85 to 46%. Several factors, including land cover and use, human population density and climate, that could have contributed to the decline of the species in Ukraine were considered. We suggest climate change and its speed (velocity) have been responsible for shaping the contemporary home range of the European roller.


title: Classification and Regression Models for Genomic Selection of Skewed Phenotypes: A Case for Disease Resistance in Winter Wheat (Triticum aestivum L.)

abstract: Most genomic prediction models are linear regression models that assume continuous and normally distributed phenotypes, but responses to diseases such as stripe rust (caused by Puccinia striiformis f. sp. tritici) are commonly recorded in ordinal scales and percentages. Disease severity (SEV) and infection type (IT) data in germplasm screening nurseries generally do not follow these assumptions. On this regard, researchers may ignore the lack of normality, transform the phenotypes, use generalized linear models, or use supervised learning algorithms and classification models with no restriction on the distribution of response variables, which are less sensitive when modeling ordinal scores. The goal of this research was to compare classification and regression genomic selection models for skewed phenotypes using stripe rust SEV and IT in winter wheat. We extensively compared both regression and classification prediction models using two training populations composed of breeding lines phenotyped in four years (2016-2018, and 2020) and a diversity panel phenotyped in four years (2013-2016). The prediction models used 19,861 genotyping-by-sequencing single-nucleotide polymorphism markers. Overall, square root transformed phenotypes using rrBLUP and support vector machine regression models displayed the highest combination of accuracy and relative efficiency across the regression and classification models. Further, a classification system based on support vector machine and ordinal Bayesian models with a 2-Class scale for SEV reached the highest class accuracy of 0.99. This study showed that breeders can use linear and non-parametric regression models within their own breeding lines over combined years to accurately predict skewed phenotypes.


title: Resistance to miltefosine results from amplification of the RTA3 floppase or inactivation of flippases in Candida parapsilosis

abstract:
Flippases and floppases are two classes of proteins that have opposing functions in the maintenance of lipid asymmetry of the plasma membrane. Flippases translocate lipids from the exoplasmic leaflet to the cytosolic leaflet, and floppases act in the opposite direction. Phosphatidylcholine (PC) is a major component of the eukaryotic plasma membrane and is asymmetrically distributed, being more abundant in the exoplasmic leaflet. Here we show that gene amplification of a putative PC floppase or double disruption of two PC flippases in the pathogenic yeast Candida parapsilosis results in resistance to miltefosine, an alkylphosphocholine drug that affects PC metabolism that has recently been granted orphan drug designation approval by the US FDA for treatment of invasive candidiasis. We analysed the genomes of 170 C. parapsilosis isolates and found that 107 of them have copy number variations (CNVs) at the RTA3 gene. RTA3 encodes a putative PC floppase whose deletion is known to increase the inward translocation of PC in Candida albicans. RTA3 copy number ranges from 2 to >40 across the C. parapsilosis isolates. Interestingly, 16 distinct CNVs with unique endpoints were identified, and phylogenetic analysis shows that almost all of them have originated only once. We found that increased copy number of RTA3 correlates with miltefosine resistance. Additionally, we conducted an adaptive laboratory evolution experiment in which two C. parapsilosis isolates were cultured in increasing concentrations of miltefosine over 26 days. Two genes, CPAR2_303950 and CPAR2_102700, gained homozygous protein-disrupting mutations in the evolved strains and code for putative PC flippases homologous to S. cerevisiae DNF1. Our results indicate that alteration of lipid asymmetry across the plasma membrane is a key mechanism of miltefosine resistance. We also find that C. parapsilosis is likely to gain resistance to miltefosine rapidly, because many isolates carry loss-of-function alleles in one of the flippase genes.

Author summaryMiltefosine was developed as an anticancer drug but is commonly used to treat infections with the protozoan parasites Leishmania and Trypanosoma cruzi. More recently, it has been used to treat fungal infections, and in 2021 it was designated as an orphan drug by the US Food and Drug Administration for treatment of invasive candidiasis. Miltefosine is a derivative of phosphatidylcholine (PC), a major constituent of the cell membrane. PC and other phospholipids are asymmetrically distributed across the cell membrane. The mechanism of action of miltefosine is unknown. Here, we show that either increasing the activity of a putative floppase, which controls outward "flop" movement of phospholipids, or decreasing the activity of flippases, which control inward "flip" movement, results in increased resistance of the fungal pathogen Candida parapsilosis to miltefosine. This result suggests that miltefosine acts by controlling the localisation of PC or other phospholipids in the membrane. Importantly, we find that many C. parapsilosis isolates carry mutations in one flippase gene, which renders them partially resistant to miltefosine, and prone to easily acquiring increased resistance.


title: Amino acid substitutions in human growth hormone affect coiled-coil content and receptor binding

abstract: The interaction between human Growth Hormone (hGH) and hGH Receptor (hGHR) has great relevance to human diseases such as acromegaly and cancer. HGH has been extensively engineered by other workers to improve binding and other properties. We used a computational screen to select substitutions at single hGH positions within the hGHR-binding site. We find that, while many successfully slow down dissociation of the hGH-hGHR complex once bound, they also slow down the association of hGH to hGHR. We are particularly interested in E174 which belongs to the hGH zinc-binding triad, and which spans coiled-coil helices and obeys the coiled-coil heptad pattern. Surprisingly, substituting E174 with A leads to substantial increase in an experimental measure of coiled-coil content. E174A is known to increase affinity of hGH against hGHR; here we show that this is simply because the off-rate is slowed down more than the on-rate, in line with what has been found for other affinity-improving mutations. For E174Y (and mutations at other sites) the slowdown in on-rate was greater, leading to decreased affinity. The results point to a link between coiled-coiling, zinc binding, and hGHR-binding affinity in hGH, and also suggest rules for choosing affinity-increasing substitutions.

category: bioengineering


title: 3D-Strudel - a novel model-dependent map-feature validation method for high-resolution cryo-EM structures

abstract: Recent technological advances in electron cryo-microscopy (cryo-EM) have led to significant improvements in the resolution of many single-particle reconstructions and a sharp increase in the number of entries released in the Electron Microscopy Data Bank (EMDB) every year, which in turn has opened new possibilities for data mining. Here we present a resolution-dependent library of rotamer-specific amino-acid map motifs mined from entries in the EMDB archive with reported resolution between 2.0 and 4.0[A]. We further describe 3D-Strudel, a method for map/model validation based on these libraries. 3D-Strudel calculates linear correlation coefficients between the map values of a map-motif from the library and the experimental map values around a target residue. We also present "Strudel Score", a plug-in for ChimeraX, as a user-friendly tool for visualisation of 3D-Strudel validation results.

category: bioinformatics


title: Structural, functional and computational studies of membrane recognition by Plasmodium Perforin-Like Proteins 1 and 2.

abstract: Perforin-like proteins (PLPs) play key roles in the mechanisms associated with parasitic disease caused by apicomplexans such as Plasmodium (malaria) and Toxoplasma. The T. gondii PLP1 (TgPLP1) mediates tachyzoite egress from cells, while the five Plasmodium PLPs carry out various roles in the life cycle of the parasite and with respect to the molecular basis of disease. Here we focus on Plasmodium vivax PLP1 and PLP2 (PvPLP1 and PvPLP2) compared to TgPLP1; PvPLP1 is important for invasion of mammalian hosts by the parasite and establishment of a chronic infection, PvPLP2 is important during the symptomatic blood stage of the parasite life cycle. Determination of the crystal structure of the membrane-binding APC{beta} domain of PvPLP1 reveals notable differences with that of TgPLP1, which are reflected in its inability to bind lipid bilayers in the way that TgPLP1 and PvPLP2 can be shown to. Molecular dynamics simulations combined with site-directed mutagenesis and functional assays allow a dissection of the binding interactions of TgPLP1 and PvPLP2 on lipid bilayers, and reveal a similar tropism for lipids found enriched in the inner leaflet of the mammalian plasma membrane. In addition to this shared mode of membrane binding PvPLP2 displays a secondary synergistic interaction side-on from its principal bilayer interface. This study underlines the substantial differences between the biophysical properties of the APC{beta} domains of Apicomplexan PLPs, which reflect their significant sequence diversity. Such differences will be important factors in determining the cell targeting and membrane-binding activity of the different proteins, in their different developmental roles within parasite life cycles.

category: biophysics


title: Proteolytic cleavage of the extracellular domain affects signaling of parathyroid hormone receptor 1

abstract: Parathyroid hormone 1 receptor (PTH1R) is a member of the class B family of G protein-coupled receptors, which are characterized by a large extracellular domain required for ligand binding. We have previously shown that the extracellular domain of PTH1R is subject to metalloproteinase cleavage in vivo that is regulated by ligand-induced receptor trafficking and leads to impaired stability of PTH1R. In this work, we localize the cleavage site in the first loop of the extracellular domain using amino-terminal protein sequencing of purified receptor and by mutagenesis studies. We further show, that a receptor mutant not susceptible to proteolytic cleavage exhibits reduced signaling to Gs and increased activation of Gq compared to wild-type PTH1R. These findings indicate that the extracellular domain modulates PTH1R signaling specificity, and that its cleavage affects receptor signaling.

category: pharmacology and toxicology


title: Numerical methods for the detection of phase defect structures in excitable media

abstract: Electrical waves that rotate in the heart organize dangerous cardiac arrhythmias. Finding the region around which such rotation occurs is one of the most important practical questions for arrhythmia management. For many years, the main method for finding such regions was so-called phase mapping, in which a continuous phase was assigned to points in the heart based on their excitation status and defining the rotation region as a point of phase singularity. Recent analysis, however, showed that in many rotation regimes there exist phase discontinuities and the region of rotation must be defined not as a point of phase singularity, but as a phase defect line. In this paper, we use this novel methodology and perform a comparative study of three different phase definitions applied to in silico data and to experimental data obtained from optical voltage mapping experiments on monolayers of human atrial myocytes. We introduce new phase defect detection algorithms and compare them with those that appeared in literature already. We find that the phase definition is more important than the algorithm to identify sudden spatial phase variations. Sharp phase defect lines can be obtained from a phase derived from local activation times observed during one cycle of arrhythmia. Alternatively, similar quality can be obtained from a reparameterization of the classical phase obtained from observation of a single timeframe of transmembrane potential. We found that the phase defect line length was (35.9 {+/-} 6.2) mm in the Fenton-Karma model and (4.01 {+/-} 0.55) mm in cardiac human atrial myocyte monolayers. As local activation times are obtained during standard clinical cardiac mapping, the methods are also suitable to be applied to clinical datasets. All studied methods are publicly available and can be downloaded from an institutional web-server.


title: Dopamine and GPCR-mediated modulation of DN1 clock neurons gates the circadian timing of sleep

abstract: The metronome-like circadian regulation of sleep timing must still adapt to an uncertain environment. Recent studies in Drosophila indicate that neuromodulation not only plays a key role in clock neuron synchronization but also affects interactions between the clock network and brain sleep centers. We show here that the targets of neuromodulators, G-Protein Coupled Receptors (GPCRs), are highly enriched in the fly brain circadian clock network. Single cell sequencing indicates that they are not only differentially expressed but also define clock neuron identity. We generated a comprehensive guide library to mutagenize individual GPCRs in specific neurons and verified the strategy with a targeted sequencing approach. Combined with a behavioral screen, the mutagenesis strategy revealed a novel role of dopamine in sleep regulation by identifying two dopamine receptors and a clock neuron subpopulation that gate the timing of sleep.

category: neuroscience


title: Assembly of recombinant tau into filaments identical to those of Alzheimer's disease and chronic traumatic encephalopathy

abstract: Abundant filamentous inclusions of tau are characteristic of more than 20 neurodegenerative diseases that are collectively termed tauopathies. Electron cryo-microscopy (cryo-EM) structures of tau amyloid filaments from human brain revealed that distinct tau folds characterise many different diseases. A lack of laboratory-based model systems to generate these structures has hampered efforts to uncover the molecular mechanisms that underlie tauopathies. Here, we report in vitro assembly conditions with recombinant tau that replicate the structures of filaments from both Alzheimers disease (AD) and chronic traumatic encephalopathy (CTE), as determined by cryo-EM. Our results suggest that post-translational modifications of tau modulate filament assembly, and that previously observed additional densities in AD and CTE filaments may arise from the presence of inorganic salts, like phosphates and sodium chloride. In vitro assembly of tau into disease-relevant filaments will facilitate studies to determine their roles in different diseases, as well as the development of compounds that specifically bind to these structures or prevent their formation.


title: Disrupting ATXN1 Nuclear Localization in a Knock-in SCA1 Mouse Model Improves a Spectrum of SCA1-Like Phenotypes and their Brain Region Associated Transcriptomic Profiles

abstract:
Spinocerebellar ataxia type 1 (SCA1) is a dominant trinucleotide repeat neurodegenerative disease characterized by motor dysfunction, cognitive impairment, and premature death. Degeneration of cerebellar Purkinje cells is a frequent and prominent pathological feature of SCA1. We previously showed that transport of ATXN1 to Purkinje cell nuclei is required for pathology, where mutant ATXN1 alters transcription. To examine the role of ATXN1 nuclear localization broadly in SCA1-like disease pathogenesis, CRISPR-Cas9 was used to develop a mouse with the amino acid alteration (K772T) in the nuclear localization sequence of the expanded ATXN1 protein.

Characterization of these mice indicates proper nuclear localization of mutant ATXN1 contributes to many disease-like phenotypes including motor dysfunction, cognitive deficits, and premature lethality. RNA sequencing analysis of genes whose expression was corrected to WT levels in Atxn1175QK772T/2Q mice indicates that transcriptomic aspects of SCA1 pathogenesis differ between the cerebellum, brainstem, cerebral cortex, hippocampus, and striatum.


title: Reconstitution of T cell immunity against EBV in the immunocompromised host by adoptive transfer of peptide-stimulated T cells after allogeneic stem cell transplantation

abstract:
Reconstitution of T cell repertoire after allogeneic stem cell transplantation is a long and often incomplete process. As a result, reactivation of Epstein-Barr virus (EBV) is a frequent complication that may be treated by adoptive transfer of donor-derived EBV-specific T cells. We generated donor-derived EBV-specific T cells by peptide stimulation and adoptively transferred them to a patient with angioimmunoblastic T-cell lymphoma (AITL), who had developed persisting high titers of EBV concomitant to relapse after transplantation. T cell receptor beta (TCR{beta}) deep sequencing showed that the T cell repertoire of the patient early after transplantation (day 60) was strongly reduced and only very low numbers of EBV-specific T cells were detectable. Manufacturing and in vitro expansion of donor-derived EBV-specific T cells resulted in enrichment of EBV epitope-specific, HLA-restricted T cells. Monitoring after adoptive transfer revealed that the dominant TCR sequences from peptide-stimulated T cells persisted long-term and established an EBV-specific TCR clonotype repertoire in the host, with many of the EBV-specific TCRs present in the donor. This reconstituted repertoire was associated with immunological control of EBV and with lack of further AITL relapse.

Author summaryA characteristic feature of all herpesviruses is their persistence in the hosts body after primary infection. Hence, the hosts immune system is confronted with the problem to control these viruses life-long. Well-known representative of the herpesvirus group are the classic Herpes-Simplex Virus (HSV-1) and Varicella Zoster Virus (VZV, causing chicken pox); a less known representative is Epstein Barr Virus (EBV, causing mononucleosis). When the immune system is severely compromised, for example after stem cell transplantation from a foreign (allogeneic) donor, these viruses can reappear, as they are already in the hosts body. Especially EBV cause life-threatening complications after stem cell transplantation and only reinforcement of the hosts immune system can reestablish viral control. Here we show that ex vivo manufactured EBV-specific T cells can reestablish long-term control of EBV and that these cells persist in the hosts body over months. These results give us a better understanding of viral immune reconstitution post-transplant and of clinically-relevant T cell populations against EBV.

category: immunology


title: Prediction of prophages and their host ranges in pathogenic and commensal Neisseria species

abstract:
The genus Neisseria includes two pathogenic species, N. gonorrhoeae and N. meningitidis, and numerous commensal species. Neisseria species frequently exchange DNA with one other, primarily via transformation and homologous recombination, and via multiple types of mobile genetic elements (MGEs). Few Neisseria bacteriophages (phages) have been identified and their impact on bacterial physiology is poorly understood. Furthermore, little is known about the range of species that Neisseria phages can infect. In this study, we used three virus prediction tools to scan 248 genomes of 21 different Neisseria species and identified 1302 unique predicted prophages. Using comparative genomics, we found that many predictions are dissimilar from other prophages and MGEs previously described to infect Neisseria species. We also identified similar predicted prophages in genomes of different Neisseria species. Additionally, we examined CRISPR-Cas targeting of each Neisseria genome and predicted prophage. While CRISPR targeting of chromosomal DNA appears to be common among several Neisseria species, we found that 20% of the prophages we predicted are targeted significantly more than the rest of the bacterial genome in which they were identified (i.e., backbone). Furthermore, many predicted prophages are targeted by CRISPR spacers encoded by other species. We then used these results to infer additional host species of known Neisseria prophages and predictions that are highly targeted relative to the backbone. Together, our results suggest that we have identified novel Neisseria prophages, several of which may infect multiple Neisseria species. These findings have important implications for understanding horizontal gene transfer between members of this genus.

IMPORTANCEDrug-resistant Neisseria gonorrhoeae is a major threat to human health. Commensal Neisseria species are thought to serve as reservoirs of antibiotic resistance and virulence genes for the pathogenic species N. gonorrhoeae and N. meningitidis. Therefore, it is important to understand both the diversity of mobile genetic elements (MGEs) that can mediate horizontal gene transfer within this genus, and the breadth of species these MGEs can infect. In particular, few bacteriophages (phages) have been identified and characterized in Neisseria species. In this study, we identified a large number of candidate phages integrated within the genomes of commensal and pathogenic Neisseria species, many of which appear to be novel phages. Importantly, we discovered extensive interspecies targeting of predicted phages by Neisseria CRISPR-Cas systems, which may reflect their movement between different species. Uncovering the diversity and host range of phages is essential for understanding how they influence the evolution of their microbial hosts.

category: microbiology


title: Biology and engineering of integrative and conjugative elements: Construction and analyses of hybrid ICEs reveal element functions that affect species-specific efficiencies

abstract:
Integrative and conjugative elements (ICEs) are mobile genetic elements that reside in a bacterial host chromosome and are prominent drivers of bacterial evolution. They are also powerful tools for genetic analyses and engineering. Transfer of an ICE to a new host involves many steps, including excision from the chromosome, DNA processing and replication, transfer across the envelope of the donor and recipient, processing of the DNA, and eventual integration into the chromosome of the new host (now a stable transconjugant). Interactions between an ICE and its hosts throughout the life cycle likely influence the efficiencies of acquisition by new hosts. Here, we investigated how different functional modules of two ICEs, Tn916 and ICEBs1, affect the transfer efficiencies into different host bacteria. We constructed hybrid elements that utilize the high-efficiency regulatory and excision modules of ICEBs1 and the conjugation genes of Tn916. These elements produced more transconjugants than Tn916, likely due to increased excision frequencies. We also found that several Tn916 and ICEBs1 components can substitute for one other. Using B. subtilis donors and three Enterococcus species as recipients, we found that different hybrid elements were more readily acquired by some species than others, demonstrating species-specific interactions in steps of the ICE life cycle. This work demonstrates that hybrid elements utilizing the efficient regulatory functions of ICEBs1 can be built to enable efficient transfer into and engineering of a variety of other species.

Author summary (non-technical)Horizontal gene transfer helps drive microbial evolution, enabling bacteria to rapidly acquire new genes and traits. Integrative and conjugative elements (ICEs) are mobile genetic elements that reside in a bacterial host chromosome and are prominent drivers of horizontal gene transfer. They are also powerful tools for genetic analyses and engineering. Some ICEs carry genes that confer obvious properties to host bacteria, including antibiotic resistances, symbiosis, and pathogenesis. When activated, an ICE-encoded machine is made that can transfer the element to other cells, where it then integrates into the chromosome of the new host. Specific ICEs transfer more effectively into some bacterial species compared to others, yet little is known about the determinants of the efficiencies and specificity of acquisition by different bacterial species. We made and utilized hybrid ICEs, composed of parts of two different elements, to investigate determinants of transfer efficiencies. Our findings demonstrate that there are species-specific interactions that help determine efficiencies of stable acquisition, and that this explains, in part, the efficiencies of different ICEs. These hybrid elements are also useful in genetic engineering and synthetic biology to move genes and pathways into different bacterial species with greater efficiencies than can be achieved with naturally occurring ICEs.


title: A high-throughput neutralization assay for yellow fever serodiagnostics

abstract:
Quick and accurate detection of neutralizing antibodies (nAbs) against yellow fever is essential in serodiagnosis during outbreaks, for surveillance and to evaluate vaccine efficacy in population-wide studies. All this requires serological assays that can process a large number of samples in a highly standardized format. Albeit being laborious, time-consuming and limited in throughput, classical plaque reduction neutralization test (PRNT) is still considered gold standard for the detection and quantification of nAbs due to its sensitivity and specificity. Here we report the development of an alternative fluorescence-based serological assay (SNTFLUO) with an equally high sensitivity and specificity that is fit for high-throughput testing with the potential for automation. Finally, our novel SNTFLUO was cross-validated in several reference laboratories and against international WHO standards showing its potential to be implemented in clinical use. SNTFLUO assays with similar performance are available for the Japanese encephalitis, Zika and dengue viruses amenable for differential diagnostics.

IMPORTANCEFast and accurate detection of neutralizing antibodies (nAbs) against yellow fever virus (YFV) is key in yellow fever serodiagnosis, outbreak surveillance and monitoring of vaccine efficacy. Although classical PRNT still remains gold standard for measuring YFV nAbs, this methodology suffers from inherent limitations such as a low throughput and an overall high labor intensity. We present a novel fluorescence-based serum neutralization test (SNTFLUO) with equally high sensitivity and specificity that is fit for processing large number of samples in a highly standardized manner and has the potential to be implemented in clinical use. In addition, we present SNTFLUO assays with similar performance for Japanese encephalitis, Zika and dengue viruses opening new avenues for differential diagnostics.


title: The Trypanosoma cruzi kinetoplast DNA minicircle sequences transfer biomarker of the multidrug treatment of Chagas disease

abstract:
BackgroundThe Trypanosoma cruzi infection renders the transfer of the mitochondrion kinetoplast DNA minicircle sequences into the hosts genome. The Aves are refractory to the infection, but chicks hatched from the T. cruzi inoculated eggs integrate the DNA minicircle sequences into the germ line cells. Rabbits, mice and chickens with the minicircle sequences mutations develop the Chagas cardiomyopathy and the DNA transfer underpins the heart disease.

MethodologyThe PCR with the specific primer sets revealed the Protist nuclear DNA and the kinetoplast DNA in the agarose gels bands probed with the radiolabel specific sequences from tissues of the T. cruzi-infected rabbits and of the mice. A target- primer TAIL-PCR amplification employing primer sets from the chickens, rabbits and mice, in combination with primer sets from the the T. cruzi kinetoplast minicircle sequences was used. This approach led us to disclose the integration sites of the kinetoplast DNA biomarker, then, used to monitor the effect of multidrug treatment of the T. cruzi infected mice.

Principal findingsThe Southern hybridization, clone and sequence of the amplification products revealed the DNA minicircle sequences integrations sites in the LINE transposable elements. An array of inhibitors of eukaryote cells division was used to arrest the DNA transfer. It was shown that nine out of 12 inhibitors prevented the kinetoplast DNA integration into the macrophage genome. The multidrug treatment of the acutely T. cruzi-infected mice with Benznidazole, Azidothymidine and Ofloxacin lessened circa 2.5-fold the rate of the minicircle sequences integrations in the mouse genome and inhibited the rejection of the target heart cells.

Conclusion and significanceThe T. cruzi mitochondrion kinetoplast minicircle sequences transfer driven pathogenesis of Chagas disease is an ancient Cross-Kingdom DNA phenomenon of evolution and, therefore, paradigm research with effective purposing inhibitors is needed.

Authors summaryChagas disease is considered the main cause of human heart failure in the Western Hemisphere. The treatment of the clinically manifested Chagas heart disease is considered unsatisfactory. Perhaps the most important problem in the field of Chagas disease is determination of the pathogenesis of the target heart cells lysis. We showed the transfer of the T. cruzi kDNA minicircle sequences into the genome of rabbits and mice, and to Bird refractory to the infections. The inoculation of a few T. cruzi in the fertile chicken eggs renders the kDNA sequences integration in the stem cells. Interestingly, the chicks that hatched retain the kDNA and develop the Chagas-like cardiomyopathy indistinguishable to that in the rabbits and mice. This result prompted the multidrug treatment of the Chagas heart disease with inhibitors of the eukaryotic cells division. We showed that nine out of 12 inhibitors prevented the transfer of the kDNA mutations into the macrophage genome, and that the treatment of the acutely T. cruzi-infected mice with Benznidazole + Ofloxacin + Azidothymidine lowered circa 2.5-fold the rate of the mutations in the chromosomes. These findings translated to the pathology showing inhibition of the heart lesions in the treated T. cruzi-infected mice. We suggest purposing new inhibitors should be tested to overturning the Chagas heart disease.

category: molecular biology


title: p166 links membrane and intramitochondrial modules of the trypanosomal tripartite attachment complex

abstract:
The protist parasite Trypanosoma brucei has a single mitochondrion with a single unit genome termed kinetoplast DNA (kDNA). Faithfull segregation of replicated kDNA is ensured by a complicated structure termed tripartite attachment complex (TAC). The TAC physically links the basal body of the flagellum with the kDNA spanning the two mitochondrial membranes. Here, we characterized p166 as the only TAC subunit that is anchored in the inner membrane. Its C-terminal transmembrane domain separates the protein into a large N-terminal region that interacts with the kDNA-localized TAC102 and a 34 aa C-tail that binds to the intermembrane space-exposed loop of the integral outer membrane protein TAC60. Thus, in contrast to the outer membrane TAC region which requires four essential subunits for proper function a single inner membrane TAC subunit is sufficient to bridge the distance from the OM to the kDNA. Surprisingly, non-functional p166 lacking the C-terminal 34 aa still localizes to the TAC region. This suggests the existence of non-essential TAC-associated proteins in the OM. These proteins can loosely bind to non-functional p166 lacking the C-terminal 34 aa and keep it at the TAC but their binding would not be strong enough to withstand the mechanical force upon kDNA segregation.

AUTHOR SUMMARYMitochondria evolved from a single endosymbiotic event and are a hallmark of eukaryotes. The large majority of genes for mitochondrial proteins are nuclear encoded now and only a small number are found in the mitochondrial genome. The protist Trypanosoma brucei is an extreme eukaryote in many aspects. For instance, trypanosomes have a single mitochondrion and its genome - called kinetoplast DNA (kDNA) - locates as a single unit inside the mitochondrion close to the basal body of the flagellum. The tripartite attachment complex (TAC) forms a connection between the basal body and the kDNA ensuring faithful segregation of kDNA among the daughter cells upon cytokinesis. Recently, several TAC subunits of the cytoplasm, the outer mitochondrial membrane (OM) and the mitochondrial matrix have been characterized. Here, we identify p166 as the first TAC subunit of the inner mitochondrial membrane. It is anchored with a single transmembrane domain separating the protein into a N-terminal moiety located in the matrix and a short C-tail. The latter reaches into the intermembrane space and binds the OM subunit TAC60 whereas the N-terminus interacts with the matrix subunit TAC102. Thus, with p166 we identified the missing link required to connect different modules of the TAC.


title: LTBP1 promotes fibrillin incorporation into the extracellular matrix

abstract: LTBP1 is a large extracellular matrix protein and an associated ligand of fibrillinmicrofibrils. Knowledge of LTBP1 functions is largely limited to its role in targeting and sequestering TGF{beta} growth factors within the extracellular matrix, thereby regulating their bioavailability. However, the recent description of a wide spectrum of phenotypes in multiple tissues in patients harboring LTBP1 pathogenic variants suggests a multifaceted role of the protein in the homeostasis of connective tissues. To better understand the human pathology caused by LTBP1 deficiency it is important to investigate its functional role in extracellular matrix formation. In this study, we show that LTBP1 coordinates the incorporation of fibrillin-1 and -2 into the extracellular matrix in vitro. We also demonstrate that this function is differentially exerted by the two isoforms, the short and long forms of LTBP1. Thereby our findings uncover a novel TGF{beta}-independent LTBP1 function potentially contributing to the development of connective tissue disorders.


title: A self-generated Toddler gradient guides mesodermal cell migration

abstract:
The sculpting of germ layers during gastrulation relies on coordinated migration of progenitor cells, yet the cues controlling these long-range directed movements remain largely unknown. While directional migration often relies on a chemokine gradient generated from a localized source, we find that zebrafish ventrolateral mesoderm is guided by the uniformly expressed and secreted protein Toddler/ELABELA/Apela, acting as a self-generated gradient. We show that the Apelin receptor, which is specifically expressed in mesodermal cells, has a dual role during gastrulation, acting as a scavenger receptor to generate a Toddler gradient, and as a chemokine receptor to sense this guidance cue. Thus, we uncover a single receptor-based self-generated gradient as the enigmatic guidance cue that can robustly steer the directional migration of mesoderm through the complex and continuously changing environment of the gastrulating embryo.

One sentence summaryAplnr has a dual role to self-generate and sense a Toddler gradient directing mesodermal cells during zebrafish gastrulation.

category: developmental biology


title: eXNVerify: coverage analysis for long and short-read sequencing data in clinical context

abstract: Accurate identification of genetic variants to a large extent is based on type of experimental technology, quality of the material and coverage of obtained sequencing data. Our motivation was to create a tool that will evaluate genome coverage and accelerate the introduction of long-read sequencing to medical diagnostics and clinical practice. Here we present eXNVerify: a tool for inspection of clinical data in the context of pathogenic variants. The tool calculates Clinical Depth Coverage - a measure of coverage which we introduce to evaluate loci with pathogenic germline and somatic variants reported in ClinVar. The tool additionally provides visualization options for user-defined genes of interest. Finally, we present an examples of BRCA1, TP53, CFTR application and results of a test conducted in the Extensive Sequence Dataset of Gold-Standard Samples for Benchmarking and Development. eXNVerify is available at https://github.com/porebskis/eXNVerify and can be directly pulled from the DockerHub repository: docker pull porebskis/exnverify:1.0.


title: LY6S, a New Interferon-Inducible Human Member of the Ly6a-Subfamily Expressed by Spleen Cells and Associated with Inflammation and Viral Resistance

abstract:
Syntenic genomic loci on human chromosome 8 (hChr8) and mouse chromosome 15 (mChr15) code for LY6/Ly6 (lymphocyte antigen 6) family proteins. The 23 murine Ly6 family genes include eight genes that are flanked by the murine Ly6e and Ly6l genes and form an Ly6 subgroup referred to here as the Ly6a subfamily gene cluster. Ly6a, also known as Sca1 (Stem Cell Antigen-1) and TAP (T-cell activating protein), is a member of the Ly6a subfamily gene cluster. No LY6 genes have been annotated within the syntenic LY6E to LY6L human locus. We report here on LY6S, a solitary human LY6 gene that is syntenic with the murine Ly6a subfamily gene cluster, and with which it shares a common ancestry. LY6S codes for the interferon-inducible GPI-linked LY6S-iso1 protein that contains only 9 of the 10 consensus LY6 cysteine residues and is most highly expressed in a non-classical cell population. Its expression leads to distinct shifts in patterns of gene expression, particularly of genes coding for inflammatory and immune response proteins, and LY6S-iso1 expressing cells show increased resistance to viral infection. Our findings reveal the presence of a previously un-annotated human interferon-stimulated gene, LY6S, which has a one to eight ortholog relationship with the genes of the Ly6a subfamily gene cluster, is most highly expressed in spleen cells of a non-classical cell-lineage and whose expression induces viral resistance and is associated with an inflammatory phenotype and with the activation of genes that regulate immune responses.

One Sentence SummaryLY6S is a newly discovered human interferon-inducible gene associated with inflammation and with resistance to viral replication.


title: Optimal spatial allocation of enzymes as an investment problem

abstract: Given a limited number of molecular components, cells face various allocation problems demanding decisions on how to distribute their resources. For instance, cells decide which enzymes to produce at what quantity, but also where to position them. Here we focus on the spatial allocation problem of how to distribute enzymes such as to maximize the total reaction flux produced by them in a system with given geometry and boundary conditions. So far, such distributions have been studied by computational optimization, but a deeper theoretical understanding was lacking. We derive an optimal allocation principle, which demands that the available enzymes are distributed such that the marginal flux returns at each occupied position are equal. This  homogeneous marginal returns criterion (HMR criterion) corresponds to a portfolio optimization criterion in a scenario where each investment globally feeds back onto all payoffs. The HMR criterion allows us to analytically understand and characterize a localization-delocalization transition in the optimal enzyme distribution that was previously observed numerically. In particular, our analysis reveals the generality of the transition, and produces a practical test for the optimality of enzyme localization by comparing the reaction flux to the influx of substrate. Based on these results, we devise an additive construction algorithm, which builds up optimal enzyme arrangements systematically rather than by trial and error. Taken together, our results reveal a common principle in allocation problems from biology and economics, which can also serve as a design principle for synthetic biomolecular systems.


title: Multi-omics subtyping of hepatocellular carcinoma patients using a Bayesian network mixture model

abstract:
Comprehensive molecular characterization of cancer subtypes is essential for predicting clinical outcomes and searching for personalized treatments. We present bnClustOmics, a statistical model and computational tool for multi-omics unsupervised clustering, which serves a dual purpose: Clustering patient samples based on a Bayesian network mixture model and learning the networks of omics variables representing these clusters. The discovered networks encode interactions among all omics variables and provide a molecular characterization of each patient subgroup. We conducted simulation studies that demonstrated the advantages of our approach compared to other clustering methods in the case where the generative model is a mixture of Bayesian networks. We applied bnClustOmics to a hepatocellular carcinoma (HCC) dataset comprising genome (mutation and copy number), transcriptome, proteome, and phosphoproteome data. We identified three main HCC subtypes together with molecular characteristics, some of which are associated with survival even when adjusting for the clinical stage. Cluster-specific networks shed light on the links between genotypes and molecular phenotypes of samples within their respective clusters and suggest targets for personalized treatments.

Author summaryMulti-omics approaches to cancer subtyping can provide more insights into molecular changes in tumors compared to single-omics approaches. However, most multi-omics clustering methods do not take into account that gene products interact, for example, as parts of protein complexes or signaling networks. Here we present bnClustOmics, a Bayesian network mixture model for unsupervised clustering of multi-omics data, which can represent dependencies among molecular changes of various omics types explicitly. Unlike other approaches that use data from public interaction databases as ground truth, bnClustOmics learns the dependencies between genes from the analyzed multi-omics dataset. At the same time, our approach can also account for prior knowledge from public interaction databases and use it to guide network learning without losing the ability to learn new dependencies. We applied bnClustOmics to a multi-omics HCC dataset and identified three subtypes similar to those identified in other HCC studies. The cluster-specific networks learned by bnClustOmics revealed additional insights into the molecular characterization of the discovered subgroups and highlighted the changes in signaling networks leading to distinct HCC phenotypes.


title: ICoRD: Iterative Correlation-Based ROI Detection Method for the Extraction of Neural Signals in Calcium Imaging

abstract: In vivo calcium imaging is a standard neuroimaging technique that allows selective observation of target neuronal activities. In calcium imaging, neuron activation signals provide key information for the investigation of neural circuits. For efficient extraction of the calcium signals of neurons, selective detection of the region of interest (ROI) pixels corresponding to the active subcellular region of the target neuron is essential. However, current ROI detection methods for calcium imaging data exhibit a relatively low signal extraction performance from neurons with a low signal-to-noise power ratio (SNR). This is problematic because a low SNR is unavoidable in many biological experiments. Therefore, we propose an iterative correlation-based ROI detection (ICoRD) method that robustly extracts the calcium signal of the target neuron from a calcium imaging series with severe noise. ICoRD extracts calcium signals closer to the ground-truth calcium signal than the conventional method from simulated calcium imaging data in all low SNR ranges. Additionally, this study confirmed that ICoRD robustly extracts activation signals against noise, even within in vivo environments. ICoRD showed reliable detection from neurons with a low SNR and sparse activation, which were not detected by conventional methods. ICoRD will facilitate our understanding of neural circuit activity by providing significantly improved ROI detection in noisy images.