title: Identification and comparative analysis of long non-coding RNAs in the brain of fire ant queens in two different reproductive states

abstract:
BackgroundMany long non-coding RNAs (lncRNAs) have been extensively identified in many higher eukaryotic species. The function of lncRNAs has been reported to play important roles in diverse biological processes, including developmental regulation and behavioral plasticity. However, there are no reports of systematic characterization of long non-coding RNAs in the fire ant Solenopsis invicta.

ResultsIn this study, we performed a genome-wide analysis of lncRNAs in the brains of S. invicta from RNA-seq. In total, 1,393 novel lncRNA transcripts were identified in the fire ant. In contrast to the annotated lncRNA transcripts having at least two exons, novel lncRNAs are monoexonic transcripts with a shorter length. Besides, the transcriptome from virgin alate and dealate mated queens were analyzed and compared. The results showed 295 differentially expressed mRNA genes (DEGs) and 65 differentially expressed lncRNA genes (DELs) between virgin and mated queens, of which 17 lncRNAs were highly expressed in the virgin alates and 47 lncRNAs were highly expressed in the mated dealates. By identifying the DEL:DEG pairs with high association in their expression (Spearmans |rho| > 0.8 and p-value < 0.01), many DELs were co-regulated with DEGs after mating. Furthermore, several remarkable lncRNAs (MSTRG.6523, MSTRG.588, and nc909) that were found to associate with particular coding genes may play important roles in the regulation of brain gene expression in reproductive transition in fire ants.

ConclusionThis study provides the first genome-wide identification of S. invicta lncRNAs in the brains in different reproductive states and will contribute to a fuller understanding of the transcriptional regulation underpinning reproductive changes.


title: In vivo hemostatic capability of a novel Tetra-PEG hydrogel

abstract: TetraStat is a novel synthetic sealant created with a tetra-armed polyethylene glycol (PEG) hydrogel. It has no risk of infection from biological pathogens and has a hemostatic mechanism independent of the blood coagulation pathway and controllable gelation. We evaluated the hemostatic effect of TetraStat in ex vivo and in vivo experiments for future clinical application. In ex vivo experiments using a circulatory system filled with phosphate-buffered saline under high pressure, needle punctures were astricted with TetraStat and two commercially available hemostatic agents (SURGICEL and TachoSil). For in vivo experiments, rat vena cavae were punctured with 14, 18, and 20 gauge needles, and hemorrhage occurred for several seconds. A porous PEG sponge soaked with TetraStat was applied as a hemostatic system for the massive hemorrhage. In the ex vivo experiment, punctures were sealed completely after 1 min astriction with TetraStat gel; in contrast, SURGICEL and TachoSil failed to seal the hole. In vivo experiments demonstrated that TetraStat successfully caused hemostasis in the punctured vena cava within 1 min of application in a dose-dependent manner. For SURGICEL and TachoSil, successful hemostasis occurred after 5 min astriction but was less frequent after 1 min astriction. Ex vivo and in vivo experiments revealed TetraStats high hemostatic ability under high pressure and in rat vena cava injuries under massive hemorrhage. A porous PEG sponge soaked with TetraStat is a promising advancement in hemostatic systems.

category: bioengineering


title: A Robust, Low-Cost Instrument for Real-time Colorimetric Isothermal Nucleic Acid Amplification

abstract: The COVID-19 pandemic has highlighted the need for broader access to molecular diagnostics. Colorimetric isothermal nucleic acid amplification assays enable simplified instrumentation over more conventional PCR diagnostic assays and, as such, represent a promising approach for addressing this need. In particular, colorimetric LAMP (loop-mediated isothermal amplification) has received a great deal of interest recently. However, there do not currently exist robust instruments for performing these kinds of assays in high throughput with real-time readout of amplification signals. To address this need, we developed LARI, the LAMP Assay Reader Instrument. We have deployed over 50 LARIs for routine use in R&D and production environments, with over 12,000 assays run to date. In this paper, we present the design and construction of LARI along with thermal, optical, and assay performance characteristics. LARI can be produced for under $1500 and has broad applications in R&D, point-of-care diagnostics, and global health.


title: Physicochemical and Pharmacokinetic Properties Determining Drug Detection in Skin

abstract: Chemicals, including some systemically administered xenobiotics and their biotransformations, can be detected noninvasively using skin swabs and untargeted metabolomics analysis. We sought to understand the principal drivers that determine whether a drug taken orally or systemically is likely to be observed on the epidermis by using a random forest classifier to predict which drugs would be detected on the skin. A variety of molecular descriptors describing calculated properties of drugs, such as measures of volume, electronegativity, bond energy, and electrotopology, were used to train the classifier. The mean area under the ROC curve was 0.71 for predicting drug detection on the epidermis, and the SHapley Additive exPlanations model interpretation technique was used to determine the most relevant molecular descriptors. Based on the analysis of 2,561 FDA approved drugs, we predict that therapeutic drug classes such as nervous system drugs are more likely to be detected on the skin. Detecting drugs and other chemicals noninvasively on the skin using untargeted metabolomics could be a useful clinical advancement in therapeutic drug monitoring, adherence, and health status.

category: bioinformatics


title: SOX2-phosphorylation toggles a bistable differentiation-switch in squamous cell carcinoma

abstract: The fate choice between stem cell self-renewal and differentiation is regulated by bistable transcriptional networks, which are balanced in homeostasis and imbalanced in tumors. Yet, how stem cells switch from self-renewal to differentiation remains a conundrum. Here, we discover a molecular mechanism that allows stem cell-like tumor propagating cells (TPCs) in squamous cell carcinomas (SCCs) to switch from a mutually exclusive SOX2-PITX1-TP63 self-renewal circuit to a KLF4 driven differentiation program, dependent on the relative occupancy of a novel Klf4-regulatory enhancer cluster (Klf4EC944) by SOX2 or KLF4, respectively. We find SOX2 occupies this site in TPCs to inhibit Klf4 transcription, but upon phosphorylation SOX2 becomes evicted from Klf4EC944, allowing residual KLF4 to occupy this site instead, boost the expression of KLF4 and its downstream targets, and differentiate self-renewing TPCs into post-mitotic SCC cells. This mechanism allows SOX2 to promote self-renewal and tumor formation, while preserving the differentiation potential in SCC cells. Our data suggest that stochastic cell fate decisions depend on the effective concentration of enzymatically regulated transcription factors. The surprising specificity by which SOX2-phosphorylation governs the bistable Klf4EC944 network-switch in SCCs reveals a conceptual framework for the identification of similar switches in other stem cell and cancer types and their potential development into cell type specific differentiation therapies for diseases in which tissue homeostasis has gone awry.

category: cancer biology


title: Effects of the exercise-inducible myokine irisin on proliferation and malignant properties of ovarian cancer cells through the HIF-1 α signaling pathway

abstract:
BackgroundExercise has been shown to be associated with reduced risk and improving outcomes of several types of cancers. Irisin -a novel exercise-related myokine- has been proposed to exert beneficial effects in metabolic disorders including cancer. No previous studies have investigated whether irisin may regulate malignant characteristics of ovarian cell lines.

MethodsIn the present study, we aimed to explore the effect of irisin on viability and proliferation of ovarian cancer cells which was examined by MTT assay. Then, we evaluated migratory and invasive ability of the cells via transwell assays. Moreover, the percentage of apoptosis induction was determined by flowcytometery. Furthermore, the mRNA expression level of genes related to the aerobic respiration (HIF-1, c-Myc, LDHA, PDK1 and VEGF) were detected by real-time PCR.

ResultsOur data revealed that irisin treatment significantly attenuated the proliferation, migration and invasion of ovarian cancer cells. Besides, irisin induced apoptosis in ovarian cancer cells. We also observed that irisin regulated the expression of genes involved in aerobic respiration of ovarian cancer cells.

ConclusionOur results indicate that irisin may play a crucial role in inhibition of cell growth and malignant characteristics of ovarian cancer. This findings may open up avenues for future studies to identify the further therapeutic use of irisin in ovarian cancer management.


title: Utilizing Digital Pathology to Quantify Stromal Caveolin-1 Expression in Malignant and Benign Ovarian Tumors: Associations with Clinicopathological Parameters and Clinical Outcomes

abstract: Loss of stromal caveolin-1 (Cav-1) is a biomarker of a cancer-associated fibroblast (CAF) phenotype and is related to progression, metastasis, and poor outcomes in several cancers. The objective of this study was to evaluate the clinical significance of Cav-1 expression in invasive epithelial ovarian cancer (OvCa). Epithelial and stromal Cav-1 expression were quantified in serous OvCa and benign ovarian tissue in two, independent cohorts - one quantified expression using immunohistochemistry (IHC) and the other using multiplex immunofluorescence (IF) with digital image analysis designed to target CAF-specific expression. Cav-1 expression was significantly downregulated in OvCa stroma compared to non-neoplastic stroma using both the IHC (p=0.002) and IF (p=1.8x10-13) assays. OvCa stroma showed Cav-1 downregulation compared to tumor epithelium with IHC (p=1.2x10-24). Conversely, Cav-1 expression was higher in OvCa stroma compared to tumor epithelium with IF (p=0.002). There was moderate correlation between IHC and IF methods for stromal Cav-1 expression (r2= 0.69, p=0.006) whereas there was no correlation for epithelial expression (r2=0.006, p=0.98). Irrespective of the staining assay, neither response to therapy or overall survival correlated with the expression level of Cav-1 in the stroma or tumor epithelium. Our findings demonstrate a loss of stromal Cav-1 expression in ovarian serous carcinomas. Studies are needed to replicate these findings and explore therapeutic implications, particularly for immunotherapy response.


title: Sharks, Lies, and Videotape: A content analysis of 32 years of Shark Week documentaries

abstract: Despite evidence of their importance to marine ecosystems, at least 25% of all chondrichthyan species are estimated or assessed as threatened with extinction. In addition to the logistical difficulties of effectively conserving wide-ranging marine species, shark conservation is believed to have been hindered in the past by public perceptions of sharks as dangerous to humans. Shark Week is a high-profile, international programming event that has potentially enormous influence on public perceptions of sharks, shark research, shark researchers, and shark conservation. However, Shark Week has received regular criticism for poor factual accuracy, fearmongering, bias, and inaccurate representations of science and scientists. This research analyzes the content and titles of Shark Week episodes across its entire 32 years of programming to determine if there are trends in species covered, research techniques featured, expert identity, conservation messaging, type of programming, and portrayal of sharks. We analyzed titles from 272 episodes (100%) of Shark Week programming and the content of all available (201; 73.9%) episodes. Our data demonstrate that the majority of episodes are not focused on shark bites, although such shows are common and many Shark Week programs frame sharks around fear, risk, and adrenaline. While anecdotal descriptions of disproportionate attention to particular charismatic species (e.g. great whites, bull sharks, and tiger sharks) are accurate and supported by data, 79 shark species have been featured briefly at least once. Shark Weeks depictions of research and of scientists are biased towards a small set of (typically visual and expensive) research methodologies and (mostly white, mostly male) scientists, including presentation of many white male non-scientists as experts. While sharks are more often portrayed negatively than positively, limited conservation messaging does appear in 53% of episodes analyzed. Results suggest that as a whole, while Shark Week is likely contributing to the collective perception of sharks as monsters, even relatively small alterations to programming decisions could substantially improve the presentation of sharks and shark science and conservation issues.

category: scientific communication and education


title: DARQ: Deep learning of quality control for stereotaxic registration of human brain MRI

abstract:
Linear registration to stereotaxic space is a common first step in many automated image-processing tools for analysis of human brain MRI scans. This step is crucial for the success of the subsequent image-processing steps. Several well-established algorithms are commonly used in the field of neuroimaging for this task, but none have a 100% success rate. Manual assessment of the registration is commonly used as part of quality control. To reduce the burden of this time-consuming step, we propose Deep Automated Registration Qc (DARQ), a fully automatic quality control method based on deep learning that can replace the human rater and accurately perform quality control assessment for stereotaxic registration of T1w brain scans.

In a recently published study from our group comparing linear registration methods, we used a database of 9325 MRI scans and 64476 registrations from several publicly available datasets and applied seven linear registration tools to them. In this study, the resulting images that were assessed and labeled by a human rater are used to train a deep neural network to detect cases when registration failed. We further validated the results on an independent dataset of patients with multiple sclerosis, with manual QC labels available (n=1200).

In terms of agreement with a manual rater, our automated QC method was able to achieve 89%accuracy and 85% true negative rate (equivalently 15% false positive rate) in detecting scans that should pass quality control in a balanced cross-validation experiments, and 96.1% accuracy and 95.5% true negative rate (or 4.5% FPR) when evaluated in a balanced independent sample, similar to manual QC rater (test-retest accuracy of 93%).

The results show that DARQ is robust, fast, accurate, and generalizable in detecting failure in linear stereotaxic registrations and can substantially reduce QC time (by a factor of 20 or more) when processing large datasets.

category: neuroscience


title: Preservation of eye movements in Parkinson's disease is stimulus and task specific

abstract:
Parkinsons disease (PD) is a neurodegenerative disease that includes motor impairments such as tremor, bradykinesia, and postural instability. Although eye movement deficits are commonly found in saccade and pursuit tasks, preservation of oculomotor function has also been reported. Here we investigate specific task and stimulus conditions under which oculomotor function in PD is preserved. Sixteen PD patients and eighteen healthy, age-matched controls completed a battery of movement tasks that included stationary or moving targets eliciting reactive or deliberate eye movements: pro-saccades, anti-saccades, visually-guided pursuit, and rapid go/no-go manual interception. Compared to controls, patients demonstrated systematic impairments in tasks with stationary targets: pro-saccades were hypometric and anti-saccades were incorrectly initiated toward the cued target in about 35% of trials compared to 14% errors in controls. In patients, task errors were linked to short latency saccades, indicating abnormalities in inhibitory control. However, patients eye movements in response to dynamic targets were relatively preserved. PD patients were able to track and predict a disappearing moving target and make quick go/no-go decisions as accurately as controls. Patients interceptive hand movements were slower on average but initiated earlier, indicating adaptive processes to compensate for motor slowing. We conclude that PD patients demonstrate stimulus- and task-dependency of oculomotor impairments and propose that preservation of eye and hand movement function in PD is linked to a separate functional pathway through the SC-brainstem loop that bypasses the fronto-basal ganglia network.

Significance StatementEye movements are a promising clinical tool to aid in the diagnosis of movement disorders and to monitor disease progression. Although Parkinsons disease (PD) patients show some oculomotor abnormalities, it is not clear whether previously-described eye movement impairments are task specific. We assessed eye movements in PD under different visual (stationary vs. moving targets) and movement (reactive vs. deliberate) conditions. We demonstrate that PD patients are able to accurately track moving objects but make inaccurate eye movements towards stationary targets. The preservation of eye movements towards dynamic stimuli might enable patients to accurately act upon the predicted motion path of the moving target. These results can inform the development of tools for the rehabilitation or maintenance of functional performance.


title: Spatial and Temporal Consistency of Brain Networks for different Multi-Echo fMRI Combination Methods

abstract: The application of multi-echo functional magnetic resonance imaging (fMRI) studies has considerably increased in the last decade due to its superior BOLD sensitivity compared to single-echo fMRI. Various methods have been developed that combine the fMRI time-series derived at different echo times to improve the data quality. Here we evaluated three multi-echo combination schemes, i.e.  optimal combination (T2*-weighted), temporal Signal-to-Noise Ratio (tSNR) weighted, and temporal Contrast-to-Noise Ratio (tCNR) weighted combination. For the first time, the effect of these multi-echo combinations on functional resting-state networks was assessed in the temporal and spatial domain, and compared to networks derived from the second echo (35 ms) functional images. Sixteen healthy volunteers were scanned during a 5 minutes resting-state fMRI session. After obtaining the networks, several temporal and spatial metrics were calculated for their time-series and spatial maps. Our results showed that, compared to the second echo network time-series, the Pearson correlation and root mean square error were the most consistent for the optimal combination time-series and the least with those derived from tSNR-weighted combination. The frequency analysis further suggested that the time-series from the tSNR-weighted combination method reduced hardware- and physiological-related artifacts as reflected by the reduced power for the associated frequencies in almost all networks. Moreover, the spatial stability and extent of the networks significantly increased after multi-echo combination, primarily for the optimal combination, followed by the tSNR-weighted combination. The performance of the tCNR-weighted combination lacked robustness and instead varied remarkedly between resting-state networks in both the temporal and spatial domain. The results highlight the benefits of multi-echo sequences on resting-state networks as well as the importance of adjusting the choice of multi-echo combination method to the research question and domain of interest.


title: The IgE production is initially induced in subcutaneous fat and depends on extrafollicular B cells

abstract:
BackgroundGrowing body of evidence indicates that IgE production can be developed by mechanisms that differ from those responsible for IgG and IgA production. One potential possibility is generation of IgE producing cells from tissue-associated B-cells and/or through extrafollicular pathway. But the role of subcutaneous fat-associated B-cells in this process is poorly investigated. The aim of the present study was to investigate the role of different B- and T- cell subpopulations after long-term antigen administration in IgE response.

MethodsBALB/c mice were immunized 3 times a eeks for 4 weeks in withers region enriched with subcutaneous fat with high and low antigen doses as well as by intraperitoneal route in region enriched with visceral fat for comparison.

ResultsAfter long-term antigen administration that promotes the type of immune response which is more similar to one observed in young allergic children, subcutaneous fat tissue B-cells generates more rapid and active IgE class switched and IgE-produced cells. Although IgE production at later time points was initiated also in regional lymph nodes, the early IgE production was exclusively linked with subcutaneous fat. We observed that low-dose induced strong IgE production accompanied by minimal IgG1 production was linked with extrafollicular B-2 derived plasmablasts as well as extrafollicular T- helpers accumulation. Delayed IgE class switching in regional lymph nodes and visceral fat tissue was characterized by the absence of both stable plasmablasts and T-extrafollicular helpers accumulation.

ConclusionExtrafollicular B- and T-cell responses in subcutaneous fat are necessary for early IgE class switching and sensitization process in the case of allergen penetration through skin.

category: immunology


title: Integration of RT-LAMP and Microfluidic Technology for Detection of SARS-CoV-2 in Wastewater as an Advanced Point-of-care Platform

abstract: Development of lab-on-a-chip (LOC) system based on integration of reverse transcription loop-mediated isothermal amplification (RT-LAMP) and microfluidic technology is expected to speed up SARS-CoV-2 diagnostics allowing early intervention. In the current work, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and RT-LAMP assays were performed on extracted RNA of 7 wastewater samples from COVID-19 hotspots. RT{square}LAMP assay was also performed on wastewater samples without RNA extraction. Current detection of SARS-CoV-2 is mainly by RT-qPCR of ORF (ORF1ab) and N genes so we targeted both to find the best surrogate marker for SARS-CoV-2 detection. We also performed RT-LAMP with/without RNA extraction inside microfluidic device to target both genes. Positivity rates of RT-qPCR and RT-LAMP performed on extracted RNA were 100.0% (7/7) and 85.7% (6/7), respectively. RT-qPCR results revealed that all 7 wastewater samples were positive for N gene (Ct range 37-39), and negative for ORF1ab, suggesting that N gene could be used as a surrogate marker for detection of SARS-CoV-2. RT-LAMP of N and ORF (ORF1a) genes performed on wastewater samples without RNA extraction indicated that all 7 samples remains pink (negative). The color remains pink in all microchannels except microchannels which subjected to RT-LAMP for targeting N region after RNA extraction (yellow color) in 6 out of 7 samples. This study shows that SARS-CoV-2 was successfully detected from wastewater samples using RT-LAMP in microfluidic chips.

category: microbiology


title: Flexible pri-miRNA structures enable tunable production of 5' isomiRs

abstract:
Drosha cleavage of a pri-miRNA defines mature microRNA sequence. Drosha cleavage at alternative positions generates 5 isoforms (isomiRs) which have distinctive functions. To understand how pri-miRNA structures influence Drosha cleavage, we performed a systematic analysis of the maturation of endogenous pri-miRNAs and their variants both in vitro and in vivo. We show that, in addition to previously known features, the overall structural flexibility of pri-miRNA impacts Drosha cleavage fidelity. Internal loops and nearby G{middle dot}U wobble pairs on the pri-miRNA stem induce the use of non-canonical cleavage sites by Drosha, resulting in 5 isomiR production. By analyzing patient data deposited in The Cancer Genome Atlas, we provide evidence that alternative Drosha cleavage of pri-miRNAs is a tunable process that responds to the level of pri-miRNA-associated RNA-binding proteins. Together, our findings reveal that Drosha cleavage fidelity can be modulated by altering pri-miRNA structure, a potential mechanism underlying 5 isomiR biogenesis in tumors.

HIGHLIGHTSO_LIFlexible pri-miRNA structures lead to 5 isomiR production
C_LIO_LIInternal loops and G{middle dot}U pairs of pri-miRNA contribute to alternative Drosha cleavages
C_LIO_LIAlternative Drosha cleavage results in 5 isomiRs from both strands of pre-miRNAs
C_LIO_LI5 isomiR production is upregulated by pri-miRNA-associated RBPs in cancers
C_LI

GRAPHICAL ABSTRACT

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category: molecular biology


title: Using expert-elicitation to deliver biodiversity monitoring priorities on a Mediterranean island

abstract: Biodiversity monitoring plays an essential role in tracking changes in ecosystems, species distributions and abundances across the globe. Data collected through both structured and unstructured biodiversity recording can inform conservation measures designed to reduce, prevent, and reverse declines in valued biodiversity of many types. However, resources for biodiversity monitoring are limited, it is therefore important that funding bodies prioritise actions relative to the requirements in any given region. We addressed this prioritisation requirement through a three-stage process of expert-elicitation, resulting in a prioritised list of twenty biodiversity monitoring needs for Cyprus. Equal priority was assigned to the twenty monitoring needs within three categories: a top nine, a middle five, and a bottom six. The most highly prioritised biodiversity monitoring needs were those related to the development of robust methodologies, and those ensuring a geographic spread of sufficiently skilled and informed contributors. We suggest ways that the results of our expert-elicitation process could be used to support current and future biodiversity monitoring in Cyprus.

category: ecology


title: Interior of sand fly (Diptera: Psychodidae) abdomen reveals novel structures involved in pheromone release: discovering the Manifold.

abstract: The males of many species of New World Phlebotomines produce volatile terpenoid chemicals which have been shown in Lutzomyia longipalpis s.l. and L. cruciata to be sex/aggregation pheromones which attract female and male conspecifics. Pheromone is produced in secretory cells surrounding a cuticular reservoir which collects the pheromone and passes it through a cuticular duct to the surface of the insect. On the surface the pheromone passes through a specialised structure prior to evaporation. The shape and distribution of the structures are highly diverse and differ according to species. They range in appearance from slightly raised domes (papules) to almost spherical apple shaped structures to slight depressions with central spikes and all with a central pore. They can occur either singly or in many hundreds distributed on most abdominal tergites or grouped on one. The pheromone secreting apparatus in sand flies and other insects have historically been examined from the exterior using scanning electron microscopy (SEM) and from the interior using transmission electron microscopy. In this study we used SEM to examine the interior cuticular structure of 3 members of the Lutzomyia longipalpis s.l. species complex and Migonemyia migonei and found a new structure associated with pheromone release which we have called the Manifold. The Manifold is a substantial structure siting in-line between the cuticular duct and the underside of the tergite. Differences in the size and shape of the Manifold may be related to the chemical structure of the pheromone. In addition to the importance of this hitherto unknown structure in the production, dissemination and ecology of the pheromone, as well as its potential taxonomic value, examination of the interior cuticle by SEM may help locate the secretory apparatus in important vector species where pheromonal activity has been inferred from behavioural studies but the external secretory structures or potential pheromones have not been found.


title: Benchmarking of SpCas9 variants enables deeper base editor screens of BRCA1 and BCL2

abstract: Numerous rationally-designed and directed-evolution variants of SpCas9 have been reported to expand the utility of CRISPR technology. Here, we benchmark PAM preferences, on-target activity, and off-target susceptibility of 11 variants of SpCas9 in cell culture assays with thousands of guides targeting endogenous genes. To enhance the coverage and thus utility of base editing screens, we demonstrate that the SpCas9-NG and SpG variants are compatible with both A>G and C>T base editors, more than tripling the number of guides and assayable residues. We demonstrate the performance of these technologies by screening for loss-of-function mutations in BRCA1 and Venetoclax-resistant mutations in BCL2, identifying both known and new insights into these clinically-relevant genes. We anticipate that the tools and methodologies described here will facilitate the investigation of genetic variants at a finer and deeper resolution for any locus of interest.

category: genomics


title: Breeding displacement in gray wolves (Canis lupus): Three males usurp breeding position and pup rearing from a neighboring pack in Yellowstone National Park

abstract: Gray Wolves (Canis lupus) are territorial, group living carnivores that live in packs typically consisting of a dominant breeding pair and their offspring. Breeding tenures are relatively short and competitive, with vacancies usually occurring following a breeders death, and are often filled by unrelated immigrants or by relatives of the previous breeder. The frequency and conditions of active breeder displacements are poorly understood. Position changes in the dominance hierarchy are common yet rarely documented in detail. We describe a turnover in male breeding position in a wolf pack by males from a neighboring pack in mid-summer 2016 in Yellowstone National Park. Over the course of two months, three males from the Mollies pack displaced the breeding male of the neighboring Wapiti Lake pack, joined the packs two adult females, and subsequently raised the previous males four approximately three-month old pups. In the four years following the displacement (2017 to 2020), at least one of the intruding males has successfully bred with the dominant female and most years with a subordinate female (who was one of the pups at the time of displacement). The pack reared pups to adulthood each year. Male breeding displacements are likely influenced by male-male competition and female mate choice. These changes are the result of individuals competing to improve breeding position and may lead to increased pack stability and greater reproductive success. We report in detail on the behavior of a closely observed breeding displacement and we discuss the adaptive benefits of the change.

category: animal behavior and cognition


title: Subpopulations of soluble, misfolded proteins commonly bypass chaperones: How it happens at the molecular level

abstract:
Subpopulations of soluble, misfolded proteins can bypass chaperones within cells. The scope of this phenomenon and the lifetimes of these states have not been experimentally quantified, and how such misfolding happens at the molecular level is poorly understood. We address the first issue through a meta-analysis of the experimental literature. We find that in all quantitative protein refolding-function studies, there is always a subpopulation of soluble but misfolded and less-functional protein that does not fold in the presence of one or more chaperones. This subpopulation ranges from 8% to 50% of the soluble protein molecules in solution. Fitting the experimental time traces to a kinetic model, we find these chaperone-bypassing misfolded states take months or longer to fold and function in the presence of different chaperones. We next addressed how, at the molecular level, some misfolded proteins can evade chaperones by simulating six different proteins interacting with E. colis GroEL and HtpG chaperones when those proteins are in folded, unfolded, or long-lived, soluble, misfolded states. We observe that both chaperones strongly bind the unfolded state and weakly bind the folded and misfolded states to a similar degree. Thus, these chaperones cannot distinguish between the folded and long-lived misfolded states of these proteins. A structural analysis reveals the misfolded states are highly similar to the native state - having a similar size, amount of exposed hydrophobic surface area, and level of tertiary structure formation. These results demonstrate that in vitro it is common for appreciable subpopulations of proteins to remain misfolded, soluble, and evade the refolding action of chaperones for very long times. Further, these results suggest that this happens because these misfolded subpopulations are near-native and therefore interact with chaperones to a similar extent as properly folded proteins. More broadly, these results indicate a mechanism in which long-time scale changes in protein structure and function can persist in cells because some proteins non-native states can bypass components of the proteostasis machinery.

TEASERNear-native, misfolded protein conformations explain why some soluble proteins fail to refold in the presence of chaperones.

category: biophysics


title: Mapping Vegetation Species Succession in a Mountainous Grassland ecosystem using Landsat and Sentinel-2 data

abstract: Vegetation species succession and composition are significant factors determining the rate of ecosystem biodiversity recovery after being disturbed and subsequently vital for sustainable and effective natural resource management and biodiversity. The succession and composition of grasslands ecosystems worldwide have significantly been affected by the accelerated changes in the environment due to natural and anthropogenic activities. Therefore, understanding spatial data on the succession of grassland vegetation species and communities through mapping and monitoring is essential to gain knowledge on the ecosystem and other ecosystem services. This study used a random forest machine learning classifier on the Google Earth Engine platform to classify grass vegetation species with Landsat 7 ETM+ and ASTER multispectral imager (MI) data resampled with the current Sentinel-2 MSI data to map and estimate the changes in vegetation species succession. The results indicate that ASTER IM has the least accuracy of 72%, Landsat 7 ETM+ 84%, and Sentinel-2 had the highest of 87%. The result also show that other species had replaced four dominant grass species totaling an area of about 49 km2 throughout the study.


title: Sarcoptic mange outbreak decimates South American camelid populations in San Guillermo National Park, Argentina

abstract: Sarcoptic mange epidemics can devastate wildlife populations. In 2014, mange was first detected in vicuna (Vicugna vicugna) and guanaco (Lama guanicoe) in San Guillermo National Park (SGNP), Argentina. This study characterized the potential source and the impacts of the outbreak in 2017-2019. Transect surveys indicated a sharp decrease in the density of live vicuna and guanaco by 68% and 77%, respectively, from May 2017 to June 2018. By April 2019 no vicuna or guanaco were recorded on transect surveys, suggesting a near-extinction at the local level. Clinical signs consistent with mange (e.g. intense pruritus, hyperkeratosis, alopecia) were observed in 24% of live vicuna (n = 478) and 33% of live guanaco (n = 12) during surveys, as well as in 94% of vicuna carcasses (n = 124) and 85% of guanaco carcasses (n = 20) opportunistically examined during the study period. Histological examination (n = 15) confirmed sarcoptic mange as the cause of the cutaneous lesions. Genetic characterization revealed that Sarcoptes scabiei recovered from seven vicuna (n = 13) and three guanaco (n = 11) shared the same genotype, which is consistent with a single source and recent origin of the epidemic. A governmental livestock incentive program introduced llama (Lama glama) in areas adjacent to SGNP in 2009, some of which reportedly had alopecic scaling consistent with sarcoptic mange. We hypothesize that the introduction of mange-infected llama may have triggered the outbreak in wild camelids which has now put them at a high risk of local extinction. This unprecedented event highlights that the accidental introduction of disease may be underestimated at the onset yet can have devastating effects on native ungulate populations with potentially profound effects at the community and ecosystem levels.

category: zoology


title: Prediction Under Uncertainty: Dissociating Sensory from Cognitive Expectations in Highly Uncertain Musical Contexts

abstract: Predictive models in the brain rely on the continuous extraction of regularities from the environment. These models are thought to be updated by novel information, as reflected in prediction error responses such as the mismatch negativity (MMN). However, although in real life individuals often face situations in which uncertainty prevails, it remains unclear whether and how predictive models emerge in high-uncertainty contexts. Recent research suggests that uncertainty affects the magnitude of MMN responses in the context of music listening. However, musical predictions are typically studied with MMN stimulation paradigms based on Western tonal music, which are characterized by relatively high predictability. Hence, we developed an MMN paradigm to investigate how the high uncertainty of atonal music modulates predictive processes as indexed by the MMN and behavior. Using MEG in a group of 20 subjects without musical training, we demonstrate that the magnetic MMN in response to pitch, intensity, timbre, and location deviants is evoked in both tonal and atonal melodies, with no significant differences between conditions. In contrast, in a separate behavioral experiment involving 39 non-musicians, participants detected pitch deviants more accurately and rated confidence higher in the tonal than in the atonal musical context. These results indicate that contextual tonal uncertainty modulates processing stages in which conscious awareness is involved, although deviants robustly elicit low-level pre-attentive responses such as the MMN. The achievement of robust MMN responses, despite high tonal uncertainty, is relevant for future studies comparing groups of listeners MMN responses to increasingly ecological music stimuli.


title: Goal-oriented predictive representations in the human hippocampus

abstract: Recent work in cognitive and systems neuroscience has suggested that the hippocampus might support planning, imagination, and navigation by forming "cognitive maps" that capture the structure of physical spaces, tasks, and situations. Critically, navigation involves planning within a context and disambiguating similar contexts to reach a goal. We examined hippocampal activity patterns in humans during a goal-directed navigation task to examine how contextual and goal information are incorporated in the construction and execution of navigational plans. Results demonstrate that, during planning, the hippocampus carries a context-specific representation of a future goal. Importantly, this effect could not be explained by stimulus or spatial information alone. During navigation, we observed reinstatement of activity patterns in the hippocampus ahead of participants required actions, which was strongest for behaviorally relevant points in the sequence. These results suggest that, rather than simply representing overlapping associations, hippocampal activity patterns are powerfully shaped by context and goals.


title: Goal-centered representations in the human hippocampus


title: A comparison of the IgM and IgT repertoires reveals the structure of B cell communities in rainbow trout is a consequence of mechanistic and evolutionary forces

abstract: In rainbow trout (Oncorhynchus mykiss), three classes of antibodies have been identified: IgM, IgD, and IgT, which differ in their abundance and effector functions. Though the process of VDJ recombination that creates these antibodies is often described as a stochastic process, recent findings in mammals suggest that there are biases in antibody construction. Because class switching is absent in teleosts, studying the IgM and IgT communities of rainbow trout provides the opportunity to see how evolution has differentially shaped the IgM and IgT repertoires. Even though it has not yet been demonstrated that biases exist in variable region construction for trout immunoglobulins, it seems reasonable that both natural and artificial selection have driven preferential gene-segment usage and pairing biases in rainbow trout. In this study, we sequenced the heavy chain variable regions of membrane IgM and IgT from multiple fish and predicted that given the more generalist role of the abundant IgM there would be less pressure to bias V gene segment usage and DJ pairings; rather, natural selection would have favored diversity and the ability to respond to a plethora of pathogens. Conversely, as IgT is substantially less abundant in the serum than IgM and specialized in its function, there would be selective pressure to make the most out of a little, thus favoring biased V segment usage and preferential DJ pairings. In support of our hypotheses, for IgT, over 70% of DJ pairs were biased and over 60% of antibodies were constructed with just two V gene segments. These biases were not prevalent in the IgM repertoire, where only 4% of DJ pairs were biased and no single V gene segment was utilized in more than 10% of antibodies. We found that these biases have profound influences in the richness and evenness of the repertoires, with the IgM repertoire investing more equitably in nearly double the number of VDJ combinations compared with IgT.


title: Does plant diversity determine the diversity of the rhizosphere microbial community?

abstract:
The rhizosphere community represents an "ecological interface" between plant and soil, providing the plant with a number of advantages. Close connection and mutual influence in this communication allow to talk about the self-adjusting "plant-rhizosphere community" system, which should be be studied in connection. Diversity estimation is one of the ways of describing both bacterial and plant communities. Based on the literature, there are two assumptions of how the diversity of plant communities related to the diversity of bacterial communities: 1) an increase in the species richness of plants leads to an increase in the number of available micro-niches, and increasing of microbial diversity, 2) an increase in the species richness of plants is accompanied by the predominant development of bacteria from highly productive specific taxa and decreasing in the diversity of microorganisms. Experimental studies show controversial results.

We analyzed field sites (rye crop field and two fallow sites), using DNA isolation of both the plant root mass (followed by sequencing of the ITS1 region) and rhizosphere microorganisms (followed by sequencing of the 16s rDNA V4 region). This allowed us to 1) accurately determine the abundance and taxonomic position of plant communities; 2) extract information about both plant and microbial communities from the same sample.

There was no correlation between alpha-diversity indices of plants and rhizosphere communities. Alpha-diversity connection should be explored in similar plant communities, such as synusia. We hypothesize, that the significant differences in plant abundances lead to significant changes in exudation profiles, and the loss of diversity connection. The beta-diversity between rhizosphere communities and plant communities is highly correlated, in particular in terms of the abundance of taxa. This can be explained by a potential correlation (as reported in the literature) or by the presence of statistical artifacts.


title: Self-organization of human dorsal-ventral forebrain structures by light induced SHH

abstract: Organizing centers secrete morphogens that specify the emergence of germ layers and the establishment of the bodys axes during embryogenesis. While traditional experimental embryology tools have been instrumental in dissecting the molecular aspects of organizers in model systems, they are impractical in human in-vitro model systems to quantitatively dissect the relationships between signaling and fate along embryonic coordinates. To systematically study human embryonic organizer centers, we devised a collection of optogenetic ePiggyBac vectors to express a photoactivatable Cre-loxP recombinase, that allows the systematic induction of organizer structures by shining blue-light on hESCs. We used a light stimulus to geometrically confine SHH expression in neuralizing hESCs. This led to the self-organization of mediolateral neural patterns from the organizer. scRNA-seq analysis established that these structures represent the dorsal-ventral forebrain, at the end of the first month of development. Here, we show that morphogen light-stimulation is a scalable tool that induces self-organizing centers.

category: developmental biology


title: Cascading effects of elephant-human interactions in a savanna ecosystem and the implications for ecology and conservation

abstract:
Our study monitored the changes in elephant numbers, distribution and ecological impact over a fifty-year period as the free-ranging intermingled movements of wildlife and traditional subsistence pastoralists across the Amboseli ecosystem were disrupted by a national park, livestock ranches, farms, settlements and changing lifestyles and economies.

Elephants compressed into the national park by poaching and settlement turned woodlands to grassland and shrublands and swamps into short grazing lawns, causing a reduction of plant and herbivore diversity and resilience to extreme events. The results echo the ecological findings of high-density elephant populations in protected areas across eastern and southern Africa. The impact has led to the view of elephants in parks being incompatible with biodiversity and to population control measures.

In contrast to Amboseli National Park, we found woody vegetation grew and plant diversity fell in areas abandoned by elephants. We therefore used naturalist and exclosure experiments to determine the density-dependent response of vegetation to elephants. We found plant richness to peak at the park boundary where elephants and livestock jostled spatially, setting up a creative browsing-grazing tension and a patchwork of habitats explaining the plant richness.

A review of prehistorical and historical literature lends support to the Amboseli findings that elephants and people, the two dominant keystone species in the savannas, have been intimately entangled prior to the global ivory trade and colonialism. The findings point to the inadequacy of parks for conserving mega herbivores and as ecological baselines.

The Amboseli study underlines the significance of space and mobility in expressing the keystone role of elephants, and to community-based conservation as a way to win space and mobility for elephants, alleviate the ecological disruption of compressed populations and minimize population management.


title: The relationship between benthic nutrient fluxes and bacterial community in Aquaculture tail-water treatment systems

abstract: Constructed-wetlands, Biofilms, and sedimentation are potential aquaculture tail-water treatments however their roles on the distribution of benthic microbial community and the way they affect the interaction between microbial community and inorganic nutrient fluxes have not been fully explored. This study applied 16S rRNA high-throughput sequencing technology to investigate the microbial community distribution and their link with nutrient fluxes in an aquaculture tail-water bioremediation system. Results showed that bacterial community compositions were significantly different in constructed-wetland and biofilm treatments (p<0.05) relative to sedimentation. The composition of the 16S rRNA genes among all the treatments was enriched with Proteobacteria, Bacteroidetes, Firmicutes, and Flavobacteria. NMDS analysis showed that the bacterial composition in constructed-wetland and biofilm samples clustered separately compared to those in sedimentation. The Functional-Annotation-of-Prokaryotic-Taxa analysis indicated that the proportions of sediment-microbial-functional groups (aerobic-chemoheterophy, chemoheterotrophy, and nitrate-ammonification combined) in the constructed-wetland treatment were 47%, 32% in biofilm and 13% in sedimentation system. Benthic-nutrient fluxes for phosphate, ammonium, nitrite, nitrate and sediment oxygen consumption differed markedly among the treatments (p<0.05). Canonical correspondence analysis indicated constructed-wetland had the strongest association between biogeochemical contents and the bacterial community relative to other treatments. This study suggests that the microbial community distributions and their interactions nutrient fluxes were most improved in the constructed-wetland followed by the area under biofilm and sedimentation treatment.


title: Partitioning gene-level contributions to complex-trait heritability by allele frequency identifies disease-relevant genes

abstract: Recent works have shown that SNP-heritability--which is dominated by low-effect common variants--may not be the most relevant quantity for localizing high-effect/critical disease genes. Here, we introduce methods to estimate the proportion of phenotypic variance explained by a given assignment of SNPs to a single gene (genelevel heritability). We partition gene-level heritability across minor allele frequency (MAF) classes to find genes whose gene-level heritability is explained exclusively by "low-frequency/rare" variants (0.5% [≤] MAF < 1%). Applying our method to ~17K protein-coding genes and 25 quantitative traits in the UK Biobank (N=290K), we find that, on average across traits, ~2.5% of nonzero-heritability genes have a rare-variant component, and only ~0.8% (370 gene-trait pairs) have heritability exclusively from rare variants. Of these 370 gene-trait pairs, 37% were not detected by existing gene-level association testing methods, likely because existing methods combine signal from all variants in a region irrespective of MAF class. Many of the additional genes we identify are implicated in phenotypically related Mendelian disorders or congenital developmental disorders, providing further evidence of their trait-relevance. Notably, the rare-variant component of gene-level heritability exhibits trends different from those of common-variant gene-level heritability. For example, while total gene-level heritability increases with gene length, the rare-variant component is significantly larger among shorter genes; the cumulative distributions of gene-level heritability also vary across traits and reveal differences in the relative contributions of rare/common variants to overall gene-level polygenicity. We conclude that the proportion of gene-level heritability attributable to low-frequency/rare variation can yield novel insights into complex-trait genetic architecture.

category: genetics


title: A genealogical estimate of genetic relationships

abstract: The application of genetic relationships among individuals, characterized by a genetic relationship matrix (GRM), has far-reaching effects in human genetics. However, the current standard to calculate the GRM generally does not take advantage of linkage information and does not reflect the underlying genealogical history of the study sample. Here, we propose a coalescent-informed framework to infer the expected relatedness between pairs of individuals given an ancestral recombination graph (ARG) of the sample. Through extensive simulations we show that the eGRM is an unbiased estimate of latent pairwise genome-wide relatedness and is robust when computed using genealogies inferred from incomplete genetic data. As a result, the eGRM better captures the structure of a population than the canonical GRM, even when using the same genetic information. More importantly, our framework allows a principled approach to estimate the eGRM at different time depths of the ARG, thereby revealing the time-varying nature of population structure in a sample. When applied to genotyping data from a population sample from Northern and Eastern Finland, we find that clustering analysis using the eGRM reveals population structure driven by subpopulations that would not be apparent using the canonical GRM, and that temporally the population model is consistent with recent divergence and expansion. Taken together, our proposed eGRM provides a robust tree-centric estimate of relatedness with wide application to genetic studies.


title: How synonymous mutations alter enzyme structure andfunction over long time scales

abstract: The specific activity of enzymes can be altered over long time scales in cells by synonymous mutations, which change an mRNA molecules sequence but not the encoded proteins primary structure. How this happens at the molecular level is unknown. Here, we investigate this issue by applying multiscale modeling to three E. coli enzymes - type III chloramphenicol acetyltransferase, D-alanine-D-alanine ligase B, and dihydrofolate reductase. This modeling involves coarse-grained simulations of protein synthesis and post-translational behavior, all-atom simulations as a test of robustness, and QM/MM calculations to characterize enzymatic function. We first demonstrate that our model predicts experimentally measured changes in specific activity due to synonymous mutations. Then, we show that changes in codon translation rates induced by synonymous mutations cause shifts in co-translational and post-translational folding pathways that kinetically partition molecules into subpopulations that very slowly interconvert to the native, functional state. These long-lived states exhibit reduced catalytic activity, as demonstrated by their increased activation energies for the reactions they carry out. Structurally, these states resemble the native state, with localized misfolding near the active sites of the enzymes. The localized misfolding involves noncovalent lasso entanglements - a topology in which the protein backbone forms a loop closed by noncovalent native contacts which is then threaded by another portion of the protein. Such entanglements are often kinetic traps, as they can require a large proportion of the protein to unfold, which is energetically unfavorable, before they can disentangle and attain the native state. The near-native structures of these misfolded states allow them to bypass the proteostasis machinery and remain soluble, as they exhibit similar hydrophobic surface areas as the native state. These entangled structures persist in all-atom simulations as well, indicating that these conclusions are independent of model resolution. Moreover, the structures of long-lived entangled states are supported by agreement with limited-proteolysis mass spectrometry results. Thus, synonymous mutations cause shifts in the co- and post-translational structural ensemble of proteins, whose altered subpopulations lead to long-term changes in the specific activities of some enzymes. The formation of entangled subpopulations is a plausible mechanism through which changes in translation elongation rate alter ensemble-averaged specific activities, which can ultimately affect the efficiency of biochemical pathways and phenotypic traits.


title: Drug resistance mechanisms create targetable proteostatic vulnerabilities in Her2+ breast cancers

abstract: Oncogenic kinase inhibitors show short-lived responses in the clinic due to high rate of acquired resistance. We previously showed that pharmacologically exploiting oncogene-induced proteotoxic stress can be a viable alternative to oncogene-targeted therapy. Here, we performed extensive analyses of the transcriptomic, metabolomic and proteostatic perturbations during the course of treatment of Her2+ breast cancer cells with a Her2 inhibitor covering the drug response, resistance, relapse and drug withdrawal phases. We found that acute Her2 inhibition, in addition to blocking mitogenic signaling, leads to significant decline in the glucose uptake, and shutdown of glycolysis and of global protein synthesis. During prolonged therapy, compensatory overexpression of Her3 allows for the reactivation of mitogenic signaling pathways, but fails to re-engage the glucose uptake and glycolysis, resulting in proteotoxic ER stress, which maintains the protein synthesis block and growth inhibition. Her3-mediated cell proliferation under ER stress during prolonged Her2 inhibition is enabled due to the overexpression of the eIF2 phosphatase GADD34, which uncouples protein synthesis block from the ER stress response to allow for active cell growth. We show that this imbalance in the mitogenic and proteostatic signaling created during the acquired resistance to anti-Her2 therapy imposes a specific vulnerability to the inhibition of the endoplasmic reticulum quality control machinery. The latter is more pronounced in the drug withdrawal phase, where the de-inhibition of Her2 creates an acute surge in the downstream signaling pathways and exacerbates the proteostatic imbalance. Therefore, the acquired resistance mechanisms to oncogenic kinase inhibitors may create secondary vulnerabilities that could be exploited in the clinic.


title: Divergent transcriptional and transforming properties of PAX3-FOXO1 and PAX7-FOXO1 paralogs

abstract: The hallmarks of the alveolar subclass of rhabdomyosarcoma are chromosomal translocations that generate chimeric PAX3-FOXO1 or PAX7-FOXO1 transcription factors. Both PAX-FOXO1s result in related cell transformation in animal models, but both mutations are associated with distinct pathological manifestations in patients. To assess the mechanisms underlying these differences, we generated isogenic fibroblast lines expressing either PAX-FOXO1 paralog. Mapping of their genomic recruitment using CUT&Tag revealed that the two chimeric proteins have distinct DNA binding preferences. In addition, PAX7-FOXO1 causes stronger de novo transactivation of its bound regions than PAX3-FOXO1, resulting in greater transcriptomic dynamics involving genes regulating cell shape and cycle. Consistently, PAX3-FOXO1 accentuates fibroblast cellular traits associated with contractility and surface adhesion and limits entry into M phase. In contrast, PAX7-FOXO1 drives cells to adopt an amoeboid shape, reduces entry into S phase, and causes more genomic instabilities. Altogether, our results argue that the diversity of rhabdomyosarcoma manifestation arises, in part, from the divergence between the transcriptional activities of PAX3-FOXO1 and PAX7-FOXO1. Furthermore, the identified pronounced deleterious effects of PAX7-FOXO1 provide an explanation for the low frequency of the translocation generating this factor in patients with rhabdomyosarcoma.


title: Cannabinoid signaling modulation through JZL184 restores key phenotypes of a mouse model for Williams-Beuren syndrome

abstract: Williams-Beuren syndrome (WBS) is a rare genetic multisystemic disorder characterized by mild to moderate intellectual disability and hypersocial phenotype, while the most life-threatening features are cardiovascular abnormalities. Nowadays, there are no available treatments to ameliorate the main traits of WBS. The endocannabinoid system (ECS), given its relevance for both cognitive and cardiovascular function, could be a potential druggable target in this syndrome. We analyzed the components of the ECS in the complete deletion (CD) mouse model of WBS and assessed the impact of its pharmacological modulation in key phenotypes relevant for WBS. CD mice showed the characteristic hypersociable phenotype with no preference for social novelty and poor object-recognition performance. Brain cannabinoid type-1 receptor (CB1R) in CD male mice showed alterations in density and coupling with no detectable change in main endocannabinoids. Endocannabinoid signaling modulation with sub-chronic (10 d) JZL184, a selective inhibitor of monoacylglycerol lipase (MAGL), specifically normalized the social and cognitive phenotype of CD mice. Notably, JZL184 treatment improved cardiac function and restored gene expression patterns in cardiac tissue. These results reveal the modulation of the ECS as a promising novel therapeutic approach to improve key phenotypic alterations in WBS.


title: scFlow: A Scalable and Reproducible Analysis Pipeline for Single-Cell RNA Sequencing Data

abstract: Advances in single-cell RNA-sequencing technology over the last decade have enabled exponential increases in throughput: datasets with over a million cells are becoming commonplace. The burgeoning scale of data generation, combined with the proliferation of alternative analysis methods, led us to develop the scFlow toolkit and the nf-core/scflow pipeline for reproducible, efficient, and scalable analyses of single-cell and single-nuclei RNA-sequencing data. The scFlow toolkit provides a higher level of abstraction on top of popular single-cell packages within an R ecosystem, while the nf-core/scflow Nextflow pipeline is built within the nf-core framework to enable compute infrastructure-independent deployment across all institutions and research facilities. Here we present our flexible pipeline, which leverages the advantages of containerization and the potential of Cloud computing for easy orchestration and scaling of the analysis of large case/control datasets by even non-expert users. We demonstrate the functionality of the analysis pipeline from sparse-matrix quality control through to insight discovery with examples of analysis of four recently published public datasets and describe the extensibility of scFlow as a modular, open-source tool for single-cell and single nuclei bioinformatic analyses.


title: Working memory capacity of crows and monkeys arises from similar neuronal computations

abstract: Complex cognition relies on flexible working memory, which is severely limited in its capacity. The neuronal computations underlying these capacity limits have been extensively studied in humans and in monkeys, resulting in competing theoretical models. We probed the working memory capacity of crows (Corvus corone) in a change detection task, developed for monkeys (Macaca mulatta), while we performed extracellular recordings of the prefrontal-like area nidopallium caudolaterale. We found that neuronal encoding and maintenance of information were affected by item load, in a way that is virtually identical to results obtained from monkey prefrontal cortex. Contemporary neurophysiological models of working memory employ divisive normalization as an important mechanism that may result in the capacity limitation. As these models are usually conceptualized and tested in an exclusively mammalian context, it remains unclear if they fully capture a general concept of working memory or if they are restricted to the mammalian neocortex. Here we report that carrion crows and macaque monkeys share divisive normalization as a neuronal computation that is in line with mammalian models. This indicates that computational models of working memory developed in the mammalian cortex can also apply to non-cortical associative brain regions of birds.


title: IL-4 and helminth infection downregulate MINCLE-dependent macrophage response to mycobacteria and Th17 adjuvanticity

abstract: The myeloid C-type lectin receptor (CLR) MINCLE senses the mycobacterial cell wall component trehalose-6,6-dimycolate (TDM). Recently, we found that IL-4 down-regulates MINCLE expression in macrophages. IL-4 is a hallmark cytokine in helminth infections, which appear to increase the risk for mycobacterial infection and active tuberculosis. Here, we investigated functional consequences of IL-4 and helminth infection on MINCLE-driven macrophage activation and Th1/Th17 adjuvanticity. IL-4 inhibited MINCLE and cytokine induction after macrophage infection with Mycobacterium bovis Bacille Calmette-Guerin (BCG). Infection of mice with BCG upregulated MINCLE on myeloid cells, which was inhibited by IL-4 plasmid injection and by infection with the nematode Nippostrongylus brasiliensis in monocytes. To determine the impact of helminth infection on MINCLE-dependent immune responses, we vaccinated mice with a recombinant protein together with the MINCLE ligand Trehalose-6,6-dibehenate (TDB) as adjuvant. Concurrent infection with N. brasiliensis or with Schistosoma mansoni promoted T cell-derived IL-4 production and suppressed Th1/Th17 differentiation in the spleen. In contrast, helminth infection did not reduce Th1/Th17 induction by TDB in draining peripheral lymph nodes, where IL-4 levels were unaltered. Upon use of the TLR4-dependent adjuvant G3D6A, N. brasiliensis infection impaired selectively the induction of splenic antigen-specific Th1 but not of Th17 cells. Thus, helminth infection attenuated the Th17 response to MINCLE-dependent immunization in an organ-specific manner. Taken together, our results demonstrate down-regulation of MINCLE expression on monocytes and macrophages by IL-4 as a possible mechanism of thwarted Th17 vaccination responses by underlying helminth infection.


title: Cyclin-G-associated kinase GAK/Aux orchestrates glial autophagy via Atg9 phosphorylation in Parkinson's disease

abstract: Glia serve as double-edged swords to modulate neuropathology in Parkinsons disease (PD), but how they react opposingly to be beneficial or detrimental under pathological conditions, like promote or eliminate -Synuclein inclusions, remain elusive. Here we present evidence that the PD risk factor Cyclin G-associated kinase (GAK)/dAuxilin (Aux) is a new component in glial autophagy. Lack of GAK/Aux promotes autophagic induction, disrupts lysosome acidification, and blocks glial clearance of substrates. Aux regulates Atg9 trafficking to autophagosomes and lysosomes for autophagosome biogenesis and autolysosome maturation, respectively, via Atg9 phosphorylation at newly identified residues T62 and T69. GAK/Aux-mediated glial autophagy is required for -Syn degradation and contributes to a broad spectrum of parkinsonian symptoms in Drosophila and mice. Our findings indicate that glial autophagy is a critical component in PD progression and identify a new autophagy factor functioning in a tissue-specific manner; targeting glial autophagy is potentially a therapeutic solution for PD.


title: Learning to be on time: temporal coordination of neural dynamics by activity-dependent myelination

abstract: Activity-dependent myelination is the mechanism by which myelin changes as a function of neural activity, and plays a fundamental role in brain plasticity. Mediated by structural changes in glia, activity-dependent myelination regulates axonal conduction velocity. It remains unclear how neural activity impacts myelination to orchestrate the timing of neural signaling. We developed a model of spiking neurons enhanced with neuron-glia feedback. Inspired by experimental data and use-dependent synaptic plasticity, we introduced a learning rule, called the Activity-Dependent Myelination (ADM) rule, by which conduction velocity scales with firing rates. We found that the ADM rule implements a homeostatic control mechanism that promotes and preserves synchronization. ADM-mediated plasticity was found to optimize synchrony by compensating for variability in axonal lengths by scaling conduction velocity in an axon-specific way. This property was maintained even when the network structure is altered. We further explored how external stimuli interact with the ADM rule to trigger bidirectional and reversible changes in conduction delays. These results highlight the role played by activity-dependent myelination in synchronous neural communication and brain plasticity.


title: Macrophages in the reticuloendothelial system inhibit the propagation phase of mouse apolipoprotein A-II amyloidosis

abstract: Amyloidosis refers to a group of degenerative diseases that are characterized by the deposition of misfolded protein fibrils in various organs. Deposited amyloid may be removed by a phagocyte-dependent innate immune system; however, the precise mechanisms during disease progression remain unclear. We herein investigated the properties of macrophages that contribute to amyloid degradation and disease progression using transmissible apolipoprotein A-II amyloidosis model mice. Intravenously injected AApoAII amyloid was efficiently engulfed by reticuloendothelial macrophages in the liver and spleen and disappeared by 24 h. While cultured murine macrophages degraded AApoAII via the endosomal-lysosomal pathway, AApoAII fibrils reduced cell viability and phagocytic capacity. Furthermore, the depletion of reticuloendothelial macrophages prior to the induction of AApoAII markedly increased hepatic and splenic AApoAII deposition. These results highlight the physiological role of reticuloendothelial macrophages against inter-individual amyloid propagation and suggest the maintenance of phagocytic integrity as a therapeutic strategy to inhibit disease progression.


title: Epigenetic mechanisms of partial dosage compensation in an avian, female heterogametic system

abstract: The evolution of genetic sex determination is often accompanied by degradation of one of the proto sex chromosomes. Male heterogametic systems have evolved convergent, epigenetic mechanisms restoring the resulting imbalance in gene dosage between diploid autosomes (AA) and the hemizygous sex chromosome (X). Female heterogametic systems (AAf ZWf, AAm ZZm) tend to only show partial dosage compensation (0.5 < Zf:AAf < 1) and dosage balance (0.5<Zf:ZZm<1). The underlying mechanism remains largely elusive. Here, we quantified gene expression for a total of 15 male and female Eurasian crows (Corvus (corone) spp.) raised under common garden conditions. In addition, we characterized aspects of the regulatory landscape quantifying genome-wide ATAC-seq and 5mC methylation profiles. Partial dosage compensation was explained by female upregulation of Z-linked genes accompanied by increased chromatin accessibility on the female Z chromosome. 5mC methylation was strongly reduced in open chromatin-regions and GC islands and showed chromosome-, but no sex-specific variation. With the exception of the pseudo-autosomal region (PAR), female upregulation of gene expression was evenly spread across the Z chromosome without evidence for regional epigenetic regulation, as has for example been suggested for the male hypermethylated region (MHM) in chicken. Our results support the hypothesis that partial dosage compensation in female heterogametic systems is subject to chromosome-wide, epigenetic control mediated by differential chromatin accessibility between the sexes.


title: Noradrenaline and ATP regulate white adipocyte adiponectin exocytosis: disturbed adrenergic and purinergic signalling in obesity-associated diabetes.

abstract:
White adipocyte adiponectin exocytosis is triggered by cAMP and a concomitant increase of cytosolic Ca2+ potentiates its release. White adipose tissue is richly innervated by sympathetic nerves co-releasing noradrenaline (NA) and ATP that may act on receptors in the adipocyte plasma membrane to increase cAMP via adrenergic receptors and Ca2+ via purinergic receptors, respectively. Here we determine the importance of NA and ATP for the regulation of white adipocyte adiponectin exocytosis, at the cellular and molecular level, and we specifically detail the ATP signalling pathway. Immunohistochemical staining demonstrates that tyrosine hydroxylase (enzyme involved in catecholamine synthesis) is dramatically reduced in inguinal white adipose tissue (IWAT) isolated from mice with diet-induced obesity; this is associated with diminished levels of NA in IWAT and with lowered serum adiponectin. Adiponectin exocytosis (measured as increase in plasma membrane capacitance and as secreted product) is triggered by NA or ATP alone in cultured and primary mouse IWAT adipocytes, and enhanced by a combination of the two secretagogues. The ATP-induced adiponectin exocytosis is largely Ca2+-dependent and activated via P2Y2 receptors (P2Y2Rs) and the Gq11/PLC pathway. Adiponectin release induced by the nucleotide is abrogated in adipocytes isolated from obese/diabetic mice and this is associated with [~]70% reduced abundance of P2Y2Rs. The NA-triggered adiponectin exocytosis is likewise abolished in "obese adipocytes", concomitant with a 50% lower gene expression of beta 3 adrenergic receptors ({beta}3ARs). The NA-stimulated adiponectin secretion does not contain Ca2+-dependent components. Collectively, our data suggest that sympathetic innervation is a principal regulator of adiponectin exocytosis and that disruptions of this control are associated with the obesity-associated reduction of circulating levels of HMW adiponectin.

Key point listO_LIWhite adipose tissue is richly innervated by sympathetic nerves that co-release noradrenaline (NA) and ATP.
C_LIO_LIProtein levels of tyrosine hydroxylase and NA are dramatically decreased in white adipose tissue from obese/diabetic mice, concomitant with reduced serum levels of high-molecular weight (HMW) adiponectin.
C_LIO_LINA and ATP stimulate white adipocyte adiponectin exocytosis via beta adrenergic and purinergic receptors respectively. The ATP-induced adiponectin secretion is chiefly Ca2+-dependent and activated via the P2Y2/Gq11/PLC pathway.
C_LIO_LIThe purinergic signalling is abrogated in adipocytes from obese/diabetic mice, due to reduced abundance of P2Y2Rs. The response to NA is likewise abolished in "obese adipocytes", associated with lowered gene expression of beta 3 adrenergic receptors ({beta}3ARs).
C_LIO_LIWe propose that sympathetic innervation is central in regulation of adiponectin exocytosis via co-secretion of NA and ATP and that this control is disrupted in obesity-associated diabetes, leading to lower circulating levels of HMW adiponectin.
C_LI

category: physiology


title: Selection for seed size has indirectly shaped specialized metabolite abundance in oat (Avena sativa L.)

abstract: Plant breeding strategies to optimize metabolite profiles are necessary to develop health-promoting food crops. In oats (Avena sativa L.), seed metabolites are of interest for their antioxidant properties and their agronomic role in mitigating disease severity, yet have not been a direct target of selection in breeding. In a diverse oat germplasm panel spanning a century of breeding, we investigated the degree of variation of these specialized metabolites and how it has been molded by selection for other traits, like yield components. We also ask if these patterns of variation persist in modern breeding pools. Integrating genomic, transcriptomic, metabolomic and phenotypic analyses for three types of seed specialized metabolites - avenanthramides, avenacins, and avenacosides - we found reduced genetic variation in modern germplasm compared to diverse germplasm, in part due to increased seed size associated with more intensive breeding. Specifically, we found that abundance of avenanthramides increases with seed size, but additional variation is attributable to expression of biosynthetic enzymes, but avenacoside abundance decreases with seed size and plant breeding intensity. Overall, we show that increased seed size associated with plant breeding has uneven effects on the seed metabolome, and broadly contributes to understanding how selection shapes plant specialized metabolism.

category: plant biology


title: Deep Bayesian networks for uncertainty estimation and adversarial resistance of white matter hyperintensity segmentation

abstract: White matter hyperintensities (WMH) are frequently observed on structural neuroimaging of elderly populations and are associated with cognitive decline and increased risk of dementia. Many existing WMH segmentation algorithms produce suboptimal results in populations with vascular lesions or brain atrophy, or require parameter tuning and are computationally expensive. Additionally, most algorithms do not generate a confidence estimate of segmentation quality, limiting their interpretation. MRI-based segmentation methods are often sensitive to acquisition protocols, scanners, noise-level, and image contrast, failing to generalize to other populations and out-of-distribution datasets. Given these concerns, we propose a novel Bayesian 3D Convolutional Neural Network (CNN) with a U-Net architecture that automatically segments WMH, provides uncertainty estimates of the segmentation output for quality control and is robust to changes in acquisition protocols. We also provide a second model to differentiate deep and periventricular WMH. 432 subjects were recruited to train the CNNs from four multi-site imaging studies. A separate test set of 158 subjects was used for evaluation, including an unseen multi-site study. We compared our model to two established state-of-the-art techniques (BIANCA and DeepMedic), highlighting its accuracy and efficiency. Our Bayesian 3D U-Net achieved the highest Dice similarity coefficient of 0.89 {+/-} 0.08 and the lowest modified Hausdorff distance of 2.98 {+/-} 4.40 mm. We further validated our models highlighting their robustness on  clinical adversarial cases simulating data with low signal-to-noise ratio, low resolution, and different contrast (stemming from MRI sequences with different parameters). Our pipeline and models are available at: https://hypermapp3r.readthedocs.io


title: Trait empathy modulates music-related functional connectivity.

abstract: It has been well established through behavioural studies that empathy significantly influences music perception. Such individual differences typically manifest as variability in whole brain functional connectivity patterns. To date, nobody has examined the modulatory effect of empathy on functional connectivity patterns during continuous music listening. In the present study, we seek to investigate the global and local connectivity patterns of 36 participants whose fMRI scanning was done by employing the naturalistic paradigm wherein they listened to a continuous piece of music. We used graph-based measures of functional connectivity to identify how cognitive and affective components of empathy modulate functional connectivity. Partial correlation between Eigenvector centrality and measures of empathy showed that cognitive empathy is associated with higher centrality in the sensorimotor regions responsible for motor mimicry while affective empathy showed higher centrality in regions related to auditory affect processing. We furthermore identified a left-hemispheric dominance in the modulatory effect of affective empathy particularly in the Orbitofrontal cortex and the temporal pole. Results are discussed in relation to various theoretical models of empathy and music cognition.


title: Semantic scene-object consistency modulates N300/400 EEG components, but does not automatically facilitate object representations

abstract: During natural vision, objects rarely appear in isolation, but often within a semantically related scene context. Previous studies reported that semantic consistency between objects and scenes facilitates object perception, and that scene-object consistency is reflected in changes in the N300 and N400 components in EEG recordings. Here, we investigate whether these N300/400 differences are indicative of changes in the cortical representation of objects. In two experiments, we recorded EEG signals while participants viewed semantically consistent or inconsistent objects within a scene; in Experiment 1, these objects were task-irrelevant, while in Experiment 2, they were directly relevant for behavior. In both experiments, we found reliable and comparable N300/400 differences between consistent and inconsistent scene-object combinations. To probe the quality of object representations, we performed multivariate classification analyses, in which we decoded the category of the objects contained in the scene. In Experiment 1, in which the objects were not task-relevant, object category could be decoded from around 100 ms after the object presentation, but no difference in decoding performance was found between consistent and inconsistent objects. By contrast, when the objects were task-relevant in Experiment 2, we found enhanced decoding of semantically consistent, compared to semantically inconsistent, objects. These results show that differences in N300/400 components related to scene-object consistency do not index changes in cortical object representations, but rather reflect a generic marker of semantic violations. Further, our findings suggest that facilitatory effects between objects and scenes are task-dependent rather than automatic.


title: The Rules of Pulsatile Neurostimulation

abstract: Electrical stimulation of neural responses is used both scientifically in brain mapping studies and in many clinical applications such as cochlear, vestibular, and retinal implants. Due to safety considerations, stimulation of the nervous system is restricted to short biphasic pulses. Despite decades of research and development, neural implants are far from optimal in their ability to restore function and lead to varying improvements in patients. In this study, we provide an explanation for how pulsatile stimulation affects individual neurons and therefore leads to variability in restoration of neural responses. The explanation is grounded in the physiological response of channels in the axon and represented with mathematical rules that predict firing rate as a function of pulse rate, pulse amplitude, and spontaneous activity. We validate these rules by showing that they predict recorded vestibular afferent responses in macaques and discuss their implications for designing clinical stimulation paradigms and electrical stimulation-based experiments.


title: Perinatal Inflammation Disturbs Secondary Germinal Zone Neurogenesis and Gliogenesis Producing Deficits in Sociability

abstract:
Epidemiologic studies have demonstrated that infections during pregnancy increase the risk of offspring developing Schizophrenia, Autism, Depression and Bipolar Disorder and have implicated interleukin-6 (IL-6) as a causal agent. However, other cytokines have been associated with psychiatric disorders; therefore, it remains to be established whether elevating IL-6 is sufficient to alter the trajectory of neural development. Furthermore, most rodent studies have manipulated the maternal immune system at mid-gestation, which affects the stem cells and progenitors in both the primary and secondary germinal matrices. Therefore, a question that remains to be addressed is whether elevating IL-6 when the secondary germinal matrices are most active will affect brain development. Here, we have increased IL-6 from postnatal days 3-6, when the secondary germinal matrices are rapidly expanding. Using Nestin-CreERT2 fate mapping we show that this transient increase in IL-6 decreased neurogenesis in the dentate gyrus of the dorsal hippocampus, reduced astrogliogenesis in the prefrontal cortex and amygdala and decreased oligodendrogenesis in the body and splenium of the corpus callosum all by [~]50%. Moreover, the IL-6 treatment elicited behavioral changes classically associated with neurodevelopmental disorders. As adults, IL-6 injected male mice lost social preference in the social approach test, spent [~]30% less time socially engaging with sexually receptive females and produced [~]50% fewer ultrasonic vocalizations during mating. They also engaged [~]50% more time in self-grooming behavior and had an increase in inhibitory avoidance. Altogether, these data provide new insights into the biological mechanisms linking perinatal immune activation to complex neurodevelopmental brain disorders.

Significance statementIn these studies, we doubled circulating IL-6 levels in mice from postnatal days 3-6 to test the hypothesis that this would be sufficient to disturb neural development. More specifically, we hypothesized that IL-6 would affect postnatal neural stem cell and progenitor expansion and specification. We show that this transient increase in IL-6 decreases the numbers of granule neurons in the dorsal hippocampus, astrocytes in the prefrontal cortex and amygdala and oligodendrocytes in the corpus callosum. Importantly, this transient increase in IL-6 changes the sociability, communication and repetitive behaviors of the treated mice as adults, which are core symptoms pertinent to several neurodevelopmental psychiatric disorders.


title: Imaging-guided fine tuning of CAR synapse dynamics and T cell triggering

abstract: T cells typically recognize their ligands using microvilli to scan for cognate antigen and scaffold T cell receptors (TCRs) during antigen recognition. Conventional chimeric antigen receptors (CARs) are often built using single-chain variable fragments (scFvs) with far greater affinity than that of natural TCRs. The implications of this for T cell function are not well understood. Using high-resolution microscopy, we studied the membrane dynamics in cells bearing anti-human epidermal growth factor receptor 2 (HER2) CARs and found these hyper-stabilized microvillar contacts relative to TCRs. While these CARs also impaired synapse resolution, a monomerized CAR with a lower affinity scFv rescued synapse dynamics and improved antigen-dose discrimination. The dimeric low-affinity CAR improved synapse dynamics and minimized early indicators of exhaustion, while maintaining or improving effector functions. This work highlights the importance of designing CAR binding dynamics that more closely resemble natural TCR antigen sensing to optimize T cell quality and function.


title: THE TRANSCRIPTIONAL RESPONSE TO OXIDATIVE STRESS IS INDEPENDENT OF STRESS-GRANULE FORMATION

abstract: Cells respond to stress with translational arrest, robust transcriptional changes, and transcription-independent formation of mRNP assemblies termed stress granules (SGs). Despite considerable interest in the role of SGs in oxidative, unfolded-protein, and viral stress responses, whether and how SGs contribute to stress-induced transcription has not been rigorously examined. To address this issue, we characterized transcriptional changes in Drosophila S2 cells induced by acute oxidative-stress and assessed how these were altered under conditions that disrupted SG assembly. Sodium-arsenite stress for 3 hours predominantly resulted in the induction or upregulation of stress-responsive mRNAs whose levels peaked during cell recovery after stress cessation. The stress-transcriptome is enriched in mRNAs coding for protein chaperones, including HSP70 and low molecular-weight heat shock proteins, glutathione transferases, and several non-coding RNAs. Oxidative stress also induced prominent cytoplasmic stress granules that disassembled 3-hours after stress cessation. As expected, RNAi-mediated knockdown of the conserved G3BP1/ Rasputin protein inhibited stress-granule assembly. However, this disruption had no significant effect on the stress-induced transcriptional response or stress-induced translational arrest. Thus, SG assembly and stress-induced effects on gene expression appear to be driven by distinctive signaling processes. We suggest that while SG assembly represents a fast, transient mechanism, the transcriptional response enables a slower, longer-lasting mechanism for adaptation to and recovery from cell stress.

category: cell biology


title: Deep learning-based algorithm for predicting the live birth potential of mouse embryos

abstract: In assisted reproductive technology (ART), embryos produced by in vitro fertilization (IVF) are graded according to their live birth potential, and high-grade embryos are preferentially transplanted. However, the rate of live birth following clinical ART remains low worldwide, suggesting that grading is inaccurate. One explanation is that grading is classically based on the characteristic shape of embryos at a limited number of developmental stages and does not consider the shape of embryos and intracellular structures, e.g., nuclei, at various stages important for normal embryogenesis. Therefore, here we developed a Normalized Multi-View Attention Network (NVAN) that directly predicts live birth potential from nuclear structural features in live-cell fluorescence images taken of mouse embryos across a wide range of stages. The classification accuracy of our method was 83.87%, which greatly exceeded that of existing machine-learning methods and that of visual inspection by embryo culture specialists. By visualizing the features that contributed most to the prediction of live birth potential, we found that the size and shape of the cell nucleus at the morula stage and at the time of cell division were important for live birth prediction. We anticipate that our method will help ART and developmental engineering as a new basic technology for IVF embryo selection.


title: Range-wide habitat use and Key Biodiversity Area coverage for a lowland tropical forest raptor across an increasingly deforested landscape

abstract: Quantifying habitat use is important for understanding how animals meet their requirements for survival and provides useful information for conservation planning. Currently, assessments of range-wide habitat use that delimit species distributions are incomplete for many taxa. The harpy eagle (Harpia harpyja) is a raptor of conservation concern, widely distributed across Neotropical lowland forests, that currently faces threats from increasing habitat loss and fragmentation. Here, we use a logistic regression modelling framework to identify habitat resource selection and predict habitat suitability based on a new method developed from the International Union for the Conservation of Nature Area of Habitat range metric. From the habitat use model, we performed a gap analysis to identify areas of high habitat suitability in regions with limited coverage in the Key Biodiversity Area (KBA) network. Range-wide habitat use indicated that harpy eagles prefer areas of 70-75 % evergreen forest cover, low elevation, and high vegetation heterogeneity. Conversely, harpy eagles avoid areas of >10 % cultivated landcover and mosaic forest, and topographically complex areas. Our habitat use model identified a large continuous area across the pan-Amazonia region, and a habitat corridor from the Choco-Darien ecoregion of Colombia running north along the Caribbean coast of Central America. Little habitat was predicted across the Atlantic Forest biome, which is now severely degraded. The current KBA network covered [~]18 % of medium to high suitability harpy eagle habitat exceeding the target representation (10 %). Four major areas of high suitability habitat lacking coverage in the KBA network were identified in the Choco-Darien ecoregion of Colombia, western Guyana, and north-west Brazil. We recommend these multiple gaps of habitat as new KBAs for strengthening the current KBA network. Modelled area of habitat estimates as described here are a useful tool for large-scale conservation planning and can be readily applied to many taxa.


title: Deep tissue scattering compensation with three-photon F-SHARP

abstract: Optical imaging techniques are widely used in biological research, but their penetration depth is limited by tissue scattering. Wavefront shaping techniques are able to overcome this problem in principle, but are often slow and their performance depends on the sample. This greatly reduces their practicability for biological applications. Here we present a scattering compensation technique based on three-photon (3P) excitation, which converges faster than comparable two-photon (2P) techniques and works reliably even on densely labeled samples, where 2P approaches fail. To demonstrate its usability and advantages for biomedical imaging we apply it to the imaging of dendritic spines on GFP-labeled layer 5 neurons in an anesthetized mouse.

category: bioengineering


title: Fetal Electrocardiogram Extraction from the Mother's Abdominal Signal Using the Ensemble Kalman Filter

abstract: Fetal electrocardiogram (fECG) assessment is essential throughout pregnancy to monitor the wellbeing and development of the fetus, and to possibly diagnose potential congenital heart defects. Due to the high noise incorporated in the abdominal ECG (aECG) signals, the extraction of fECG has been challenging. And it is even a lot more difficult for fECG extraction if only one channel of aECG is provided, i.e. in a compact patch device. In this paper, we propose a novel algorithm based on the Ensemble Kalman Filter (EnKF) for non-invasive fECG extraction from a single-channel aECG signal. To assess the performance of the proposed algorithm, we used our own clinical data, obtained from a pilot study with 10 subjects each of 20 min recording, and data from the PhysioNet 2013 Challenge bank with labeled QRS complex annotations. The proposed methodology shows the average positive predictive value (PPV) of 97.59%, sensitivity (SE) of 96.91%, and F1-score of 97.25% from the PhysioNet 2013 Challenge bank. Our results also indicate that the proposed algorithm is reliable and effective, and it outperforms the recently proposed Extended Kalman Filter (EKF) based algorithm.


title: Guided Axon Outgrowth of Neurons by Real-Time Femtosecond Laser Penetration on a 4 μm Thick Thin-Glass Sheet

abstract: Transplantation of scaffold-embedded guided neurons has been reported to increase neuronal regeneration following brain injury. However, precise axonal integration between host and transplant neurons to form functional synapses remains a major problem. This study aims to develop a real-time femtosecond (fs) laser penetration on a 4 m thick thin-glass sheet to promote guided axon outgrowth influenced by molecular gradients in a microfluidic device. The device enables the introduction of the guidance molecule (i.e., netrin-1), neuronal culture, and manipulation by fs laser. After fabricating multiple micro-holes on the thin-glass sheet using fs laser, netrin-1 gradients with radial concentrations are generated in the chamber, affecting axon outgrowth and guidance. A majority of axons (~92%) experiences guided outgrowth with positive angular changes towards netrin-1 gradients. These results demonstrate the capability of the precise and real-time manipulation system based on a fs laser and a microfluidic device to control the growth of neurons.


title: Fluorogenic EXO-Probe Aptamers for Imaging and Tracking Exosomal RNAs

abstract: Small extracellular vesicles (sEVs), or exosomes, play important roles in physiological and pathological cellular communication. sEVs contain both short and long non-coding RNAs that regulate gene expression and epigenetic processes. Studying the intricacies of sEV function and RNA-based communication requires tools capable of labeling sEV RNA. Here we developed a novel genetically encodable reporter system for tracking sEV RNAs comprising an sEV-loading RNA sequence, termed the EXO-Code, fused to a fluorogenic RNA Mango aptamer for RNA imaging. This fusion construct allowed the visualization and tracking of RNA puncta and colocalization with markers of multivesicular bodies; imaging RNA puncta within sEVs; and quantification of sEVs. This technology represents a useful and versatile tool to interrogate the role of sEVs in cellular communication via RNA trafficking to sEVs, cellular sorting decisions, and sEV RNA cargo transfer to recipient cells.


title: AllerStat: Finding Statistically Significant Allergen-Specific Patterns in Protein Sequences by Machine Learning

abstract:
Cutting-edge technologies such as genome editing and synthetic biology allow us to produce novel foods and functional proteins. However, their toxicity and allergenicity must be accurately evaluated. Allergic reactions are caused by specific amino-acid sequences in proteins (Allergen Specific Patterns, ASPs), of which, many remain undiscovered. In this study, we introduce a data-driven approach and a machine-learning (ML) method to find undiscovered ASPs. The proposed method enables an exhaustive search for amino-acid sub-sequences whose frequencies are statistically significantly higher in allergenic proteins. As a proof-of-concept (PoC), we created a database containing 21,154 proteins of which the presence or absence allergic reactions are already known, and the proposed method was applied to the database. The detected ASPs in the PoC study were consistent with known biological findings, and the allergenicity prediction accuracy using the detected ASPs was higher than extant approaches.

TeaserWe propose a computational method for finding statistically significant allergen-specific amino-acid sequences in proteins.

category: bioinformatics


title: svaRetro and svaNUMT: modular packages for annotation of retrotransposed transcripts and nuclear integration of mitochondrial DNA in genome sequencing data

abstract:
BackgroundThe biological significance of structural variation is now more widely recognized. However, due to the lack of available tools for downstream analysis, including processing and annotating, interpretation of structural variant calls remains a challenge.

FindingsHere we present svaRetro and svaNUMT, R packages that provide functions for annotating novel genomic events such as non-reference retro-copied transcripts and nuclear integration of mitochondrial DNA. We evaluate the performance of these packages to detect events using simulations and public benchmarking datasets, and annotate processed transcripts in a public structural variant database.

ConclusionssvaRetro and svaNUMT provide efficient, modular tools for downstream identification and annotation of structural variant calls.