title: Age-related differences in GABA: Impact of analysis technique

abstract: Previous research using magnetic resonance spectroscopy (MRS) has indicated that GABA levels decline in multiple brain regions over the course of healthy aging. However, brain atrophy also occurs during healthy aging, and as a result the tissue composition of MRS voxels (i.e., the percentage of grey matter, white matter, and cerebrospinal fluid in the voxel) may also differ between age groups. Many authors therefore argue for applying a correction to GABA estimates in order to control for differences in tissue composition. Here, we use data from a large healthy aging study to investigate the influence of three tissue correction strategies on age-group differences in GABA. We also evaluate the use of different analysis packages and reference metabolites on group differences. A 3T MEGA-PRESS sequence was used to obtain spectra from seven voxels placed in the visual, auditory, and sensorimotor cortex of 58 young adults (aged 18-29 years) and 85 older adults. We obtained several different estimates of GABA concentrations from the spectra using two analysis software packages (Gannet 3.1 and LCModel), three reference metabolites (water, creatine and N-acetylaspartate) and four tissue correction strategies. Young adults consistently demonstrated significantly higher GABA concentrations in the visual, auditory, and sensorimotor cortex when we used an uncorrected GABA estimate referenced either to water or creatine. When uncorrected GABA was referenced to N-acetylaspartate, age-related differences were observed only in the right ventral visual cortex. Similarly, when any of the four tissue corrections were applied to the data, only age-related differences in the left and right ventral visual cortex voxels remained. Correlations between GABA concentration estimates obtained from different software packages were moderate, as were correlations between uncorrected GABA estimates when different baseline metabolites were used. Correlations between all tissue corrections we explored were extremely high. These results confirm that reports of age-related differences in GABA concentrations are driven, at least in part, by changes in tissue composition.


title: Stability And Shelf-Life Of Plasma Bubbling Treated Cow Milk

abstract: The demand of consumers for naturality of food with minimal processing was forced to the scientists for the discovery of non-thermal plasma which is now an emerging technology for the preservation and decontamination of highly perishable food such as milk. In this study, the microbial and physicochemical characteristics of plasma bubbling of raw cow milk were analysed and a comparison of boiled raw cow milk, commercially available pasteurised and UHT milk was observed. Again, shelf-life study was experimented for the plasma bubbling of milk with respect to raw cow milk sample (control). The plasma bubbling was generated at a voltage (200V), the flow rate of air 10 litres/hour (L/h) and applied to fresh cow milk for 5, 10, and 15 minutes (min) of time interval with 100 mL of the sample volume at room temperature. A significant reduction was observed in coliform and yeast at 200V, 10L/h, 15 min time interval of treatment. The pH of the milk was increased significantly to 6.85 after exposure to plasma bubbling. Whereas, a decreasing value was noticed in total soluble solids (TSS) and titratable acid (TA) with respect to time given by the plasma bubbling. Further a nondetrimental effect was observed for the nutrient content of plasma bubbling of milk. The result of the study indicates that plasma bubbling at (200V, 10L/h,100mL,15 min) treatment improves the quality of milk. This study shows that indirect dielectric barrier discharge (DBD) (plasma bubbling) may offer an effective microbial reduction without affecting the quality attributes. The plasma bubbling processing is an initiative for cow raw milk pasteurisation, which could have a future perspective on industrial food applications.

category: microbiology


title: Sinking the way: a dual role for CCR7 in collective leukocyte migration

abstract: Immune responses crucially rely on the rapid and coordinated locomotion of leukocytes. While it is well established that single-cell migration is often guided by gradients of chemokines and other chemoattractants, it remains poorly understood how such gradients are generated, maintained and modulated. Combining experiment and theory on leukocyte chemotaxis guided by the G protein-coupled receptor (GPCR) CCR7, we demonstrate that in addition to its role as the sensory receptor that steers migration, CCR7 also acts as a generator and modulator of chemotactic gradients. Upon exposure to the CCR7 ligand CCL19, dendritic cells (DCs) effectively internalize the receptor and ligand as part of the canonical GPCR-desensitization response. We show that CCR7 internalization also acts as an effective sink for the chemoattractant, thereby dynamically shaping the spatio-temporal distribution of the chemokine. This mechanism drives complex collective migration patterns, enabling DCs to create or sharpen chemotactic gradients. We further show that these self-generated gradients can sustain the long-range guidance of DCs, adapt collective migration patterns to the size and geometry of the environment, as well as provide a guidance cue for other co-migrating cells. Such dual role of CCR7 as a GPCR that both senses and consumes its ligand can thus provide a novel mode of cellular self-organization.

category: cell biology


title: Liver lipophagy ameliorates nonalcoholic steatohepatitis through lysosomal lipid exocytosis

abstract: Nonalcoholic steatohepatitis (NASH) is a progressive disorder with aberrant lipid accumulation and subsequent inflammatory and profibrotic response. Therapeutic efforts at lipid reduction via increasing cytoplasmic lipolysis unfortunately worsens hepatitis due to toxicity of liberated fatty acid. An alternative approach could be lipid reduction through autophagic disposal, i.e., lipophagy. We engineered a synthetic adaptor protein to induce lipophagy, combining a lipid droplet-targeting signal with optimized LC3-interacting domain. Activating hepatocyte lipophagy in vivo strongly mitigated both steatosis and hepatitis in a diet-induced mouse NASH model. Mechanistically, activated lipophagy promoted the excretion of lipid from hepatocytes via lysosomal exocytosis, thereby suppressing harmful intracellular accumulation of nonesterified fatty acid. A high-content compound screen identified alpelisib and digoxin, clinically-approved compounds, as effective activators of lipophagy. Administration of alpelisib or digoxin in vivo strongly inhibited the transition to steatohepatitis. These data thus identify lipophagy as a promising therapeutic approach to prevent NASH progression.


title: An averaging model for analysis and interpretations of high-order genetic interactions

abstract: While combinatorial genetic data collection from biological systems in which quantitative phenotypes are controlled by functional and non-functional alleles in each of multiple genes (multi-gene systems) is becoming common, a standard analysis method for such data has not been established. A common additive model of the non-functional allele effects contrasted against the functional alleles, based on ANOVA with interaction, has three issues. First, although it is a long tradition of genetics, modeling the effect of the non-functional allele (a null mutant allele) contrasted against that of the functional allele (the wild-type allele) is not suitable for mechanistic understanding of multi-gene systems. Second, an additive model is highly problematic when the system has more than two genes and a limited phenotypic range: errors propagate toward higher order interactions. Third, interpretations of higher-order interactions defined by an additive model are not intuitive. I propose an averaging model, which is suitable for mechanistic understanding of multi-gene systems. The effect of the functional allele is contrasted against the effect of the non-functional allele for easier mechanistic interpretations. Errors in interactions across the orders consistently stay low, which makes the model highly scalable to systems with many genes. The interactions defined by the averaging model are highly intuitive regardless of the orders. Yet, it is still a general linear model, so model fitting is easy and accurate using common statistical tools.

category: genetics


title: Deciphering the tissue-specific regulatory role of intronless genes across cancers

abstract: Intronless genes (IGs) or single-exon genes lacking an intron are found across most Eukaryotes. Notably, IGs display a higher transcriptional fidelity as they are not regulated through alternative splicing, suggesting better predictability biomarkers and easier regulation as targets for therapy. Cancer is a complex disease that relies on progressive uncontrolled cell division linked with multiple dysfunctional biological processes. Tumor heterogeneity remains the most challenging feature in cancer diagnosis and treatment. Given the clinical relevance of IGs, we aim to identify their unique expression profiles and interactome, that may act as functional signatures across eight different cancers. We identified 940 protein-coding IGs in the human genome, of which about 35% were differentially expressed across the analyzed cancer datasets. Specifically, [~]78% of differentially expressed IGs were undergoing transcriptional reprogramming with elevated expression in tumor cells. Remarkably, in all the studied tumors, a highly conserved induction of a group of deacetylase-histones located in a region of chromosome 6 enriched in nucleosome and chromatin condensation processes. This study highlights that differentially expressed human intronless genes across cancer types are prevalent in epigenetic regulatory roles participating in specific PPI networks for ESCA, GBM, and LUAD tumors. We determine that IGs play a key role in the tumor phenotype at transcriptional and post-transcriptional levels, with important mechanisms such as interactomics rewiring.

category: genomics


title: Novel feature selection methods for construction of accurate epigenetic clocks

abstract: Epigenetic clocks allow the accurate prediction of age based on the methylation status of specific CpG sites in a variety of tissues. These predictive models can be used to distinguish the biological age of an organism from its chronological age, and are a powerful tool to measure the effectiveness of aging interventions. There is a growing need for methods to efficiently construct epigenetic clocks. The most common approach is to create clocks using elastic net regression modelling of all measured CpG sites, without first identifying specific features or CpGs of interest. The addition of feature selection approaches provides the opportunity to reduce the cost and time of clock development by decreasing the number of CpG sites included in clocks. Here, we apply both classic feature selection methods and novel combinatorial methods to the development of epigenetic clocks. We perform feature selection on the human whole blood methylation dataset of [~]470,000 CpG features published by Hannum and colleagues (2015). We develop clocks to predict age, using a variety of feature selection approaches, and all clocks have R2 correlation scores of greater than 0.73. The most predictive clock uses 35 CpG sites for a R2 correlation score of 0.87. The five most frequent sites across all clocks are also modelled to build a clock with a R2 correlation score of 0.83. These two clocks are validated on two external datasets where they maintain excellent predictive accuracy and outperform Hannum et als model in accuracy of age prediction despite using significantly less CpGs. We also identify the associated gene regulatory regions of these CpG sites, which may be possible targets for future aging studies. These novel feature selection algorithms will lower the number of sites needed to be sequenced to build clocks and allow conventionally expensive aging epigenetic studies to cost a fraction of what it would normally.

category: bioinformatics


title: Estimation of the firing behaviour of a complete motoneuron pool by combining EMG signal decomposition and realistic motoneuron modelling

abstract:
Our understanding of the firing behaviour of motoneuron (MN) pools during human voluntary muscle contractions is currently limited to electrophysiological findings from animal experiments extrapolated to humans, mathematical models of MN pools not validated for human data, and experimental results obtained from EMG decomposition. These approaches are limited in accuracy or provide information on only small partitions of the MN population. Here, we propose a method based on the combination of high-density EMG (HDEMG) data and realistic modelling for predicting the behaviour of entire pools of motoneurons in humans. The method builds on a physiologically realistic model of a MN pool which predicts, from the experimental spike trains of a smaller number of individual MNs identified from decomposed HDEMG signals, the unknown recruitment and firing activity of the remaining unidentified MNs in the complete MN pool. The MN pool model is described as a cohort of leaky fire- and-integrate (LIF) models of MNs scaled by a physiologically realistic distribution of MN electrophysiological properties and driven by a spinal synaptic input, both derived from decomposed HDEMG data. The MN spike trains and effective neural drive to muscle, predicted with this method, have been successfully validated experimentally. Representative applications of the method are also presented for the prediction of activity-dependant changes in MN intrinsic properties and in MN-driven neuromuscular modelling. The proposed approach provides a validated tool for neuroscientists, experimentalists, and modelers to infer the firing activity of MNs that cannot be observed experimentally, investigate the neurophysiology of human MN pools, support future experimental investigations, and advance neuromuscular modelling for investigating the neural strategies controlling human voluntary contractions.

Author SummaryOur experimental understanding of the firing behaviour of motoneuron (MN) pools during human voluntary muscle contractions is currently limited to the observation of small samples of active MNs obtained from EMG decomposition. EMG decomposition therefore provides an important but incomplete description of the role of individual MNs in the firing activity of the complete MN pool, which limits our understanding of the neural strategies of the whole MN pool and of how the firing activity of each MN contributes to the neural drive to muscle. Here, we combine decomposed high-density EMG (HDEMG) data and a physiologically realistic model of MN population to predict the unknown recruitment and firing activity of the remaining unidentified MNs in the complete MN pool. In brief, an experimental estimation of the synaptic current is input to a cohort of MN models, which are calibrated using the available decomposed HDEMG data, and predict the MN spike trains fired by the entire MN population. This novel approach is experimentally validated and applied to muscle force prediction from neuromuscular modelling, and to investigate neurophysiological properties of the human MN population during voluntary contractions.

category: neuroscience


title: SexFindR: A computational workflow to identify young and old sex chromosomes

abstract: Sex chromosomes have evolved frequently across the tree of life, and have been a source of fascination for decades due to their unique evolutionary trajectories. They are hypothesised to be important drivers in a broad spectrum of biological processes and are the focus of a rich body of evolutionary theory. Whole-genome sequencing provides exciting opportunities to test these theories through contrasts between independently evolved sex chromosomes across the full spectrum of their evolutionary lifecycles. However, identifying sex chromosomes, particularly nascent ones, is challenging, often requiring specific combinations of methodologies. This is a major barrier to progress in the field and can result in discrepancies between studies that apply different approaches. Currently, no single pipeline exists to integrate data across these methods in a statistical framework to identify sex chromosomes at all ages and levels of sequence divergence. To address this, we present SexFindR, a comprehensive workflow to improve robustness and transparency in identifying sex-linked sequences. We validate our approach using publicly available data from five species that span the continuum of sex chromosome divergence, from homomorphic sex chromosomes with only a single SNP that determines sex, to heteromorphic sex chromosomes with extensive degeneration. Next, we apply SexFindR to our large-scale population genomics dataset for sea lamprey, a jawless vertebrate whose sex determination system remains a mystery despite decades of research. We decisively show that sea lamprey do not harbour sex-linked sequences in their somatic genome, leaving open the possibility that sex is determined environmentally or within the germline genome.

category: evolutionary biology


title: period Translation as a Core Mechanism Controlling Temperature Compensation in an Animal Circadian Clock

abstract: The most enigmatic of the canonical properties of circadian clocks is temperature compensation where circadian period length is stable across a wide temperature range despite the temperature dependence of most biochemical reactions. While the core mechanisms of circadian clocks have been well described, the molecular mechanisms of temperature compensation are poorly understood especially in animals. A major gap is the lack of temperature compensation mutants that do not themselves unambiguously affect the temperature dependence of the encoded protein. Here we show that null alleles of two genes encoding components of a complex important for translation of the core clock component period in circadian pacemaker neurons robustly alter the temperature dependence of circadian behavioral period length. These changes are accompanied by parallel temperature dependent changes in oscillations of the PER protein and are consistent with the model that these translation factors mediate the temperature-dependence of PER translation. Consistent with findings from modeling studies, we find that translation of the key negative feedback factor PER plays an instrumental role in temperature compensation.


title: Nucleosome conformation dictates the histone code

abstract: Histone post-translational modifications (PTMs) play a critical role in chromatin regulation. It has been proposed that these PTMs form localized  codes that are read by specialized regions (reader domains) in chromatin associated proteins (CAPs) to regulate downstream function. Substantial effort has been made to define [CAP-histone PTM] specificity, and thus decipher the histone code / guide epigenetic therapies. However, this has largely been done using a reductive approach of isolated reader domains and histone peptides, with the assumption that PTM readout is unaffected by any higher order factors. Here we show that CAP-histone PTM interaction is in fact dependent on nucleosome context. Our results indicate this is due to histone tail accessibility and the associated impact on binding potential of reader domains. We further demonstrate that the in vitro specificity of a tandem reader for PTM-defined nucleosomes is recapitulated in a cellular context. This necessitates we refine the  histone code concept and interrogate it at the nucleosome level.


title: 3D bioprinting of an implantable xeno-free vascularized human skin graft

abstract: Bioengineered tissues or organs produced using matrix proteins or components derived from xenogeneic sources pose risks of allergic responses, immune rejection, or even autoimmunity. Here, we report successful xeno-free isolation, expansion, and cryopreservation of human endothelial cells, fibroblasts, pericytes and keratinocytes from a single donor. We further demonstrate the bioprinting of a human skin substitute with a dermal layer containing xeno-free cultured human endothelial cells (EC), fibroblasts, and pericytes in a xeno-free bioink containing human collagen type I and fibronectin layered in a biocompatible polyglycolic acid (PGA) mesh and subsequently seeded with xeno-free human keratinocytes to form an epidermal layer. Following implantation of such bilayered skin grafts on the dorsum of immunodeficient mice, keratinocytes form a mature stratified epidermis with rete ridge-like structures. The ECs and pericytes form human EC-lined perfused microvessels within 2 weeks after implantation, preventing graft necrosis, and eliciting further perfusion of the graft by angiogenic host microvessels. In summary, we describe the fabrication of a bioprinted vascularized bilayered skin substitute under completely xeno-free culture conditions demonstrating feasibility of a xeno-free approach to complex tissue engineering.

category: bioengineering


title: Multimodal Thrombectomy Device for Treatment of Acute Deep Venous Thrombosis

abstract: Deep vein thrombosis (DVT) is a potentially deadly medical condition that is costly to treat and impacts thousands of Americans every year. DVT is characterized by the formation of blood clots within the deep venous system of the body. If a DVT dislodges it can lead to venous thromboembolism (VTE) and pulmonary embolism (PE), both of which can lead to significant morbidity or death. Current treatment options for DVT are limited in both effectiveness and safety, in part because the treatment of the DVT cannot be confined to a defined sequestered treatment zone. We therefore developed and tested a thrombectomy device that enables the sequesteration of a DVT to a defined treatment zone during fragmentation and evacuation. We observed that, compared to a predicate thrombectomy device, the sequestered approach reduced distal DVT embolization during ex vivo thrombectomy. The sequestered approach also facilitated isovolumetric infusion and suction that enabled clearance of the sequestered treatment zone without significantly impacting vein wall diameter. Results suggest that our novel device using sequestered therapy holds promise for the treatment of high risk large-volume DVTs.


title: Supervised Rank aggregation (SRA): A novel rank aggregation approach for ensemble-based feature selection

abstract:
BackgroundFeature selection (FS) is critical for high dimensional data analysis. Ensemble based feature selection (EFS) is a commonly used approach to develop FS techniques. Rank aggregation (RA) is an essential step of EFS where results from multiple models are pooled to estimate feature importance. However, the literature primarily relies on rule-based methods to perform this step which may not always provide an optimal feature set.

Method and ResultsThis study proposes a novel Supervised Rank Aggregation (SRA) approach to allow RA step to dynamically learn and adapt the model aggregation rules to obtain feature importance. The approach creates a performance matrix containing feature and model performance value from all models and prepares a supervised learning model to get the feature importance. Then, unsupervised learning is performed to select the features using their importance. We evaluate the performance of the algorithm using simulation studies and implement it into real research studies, and compare its performance with various existing RA methods. The proposed SRA method provides better or at par performance in terms of feature selection and predictive performance of the model compared to existing methods.

ConclusionSRA method provides an alternative to the existing approaches of RA for EFS. While the current study is limited to the continuous cross-sectional outcome, other endpoints such as longitudinal, categorical, and time-to-event medical data could also be used.


title: Functional networks of the human bromodomain-containing proteins

abstract:
BackgroundBromodomains are a structurally conserved epigenetic reader domain that bind to acetylated lysine residues in both histone and non-histone proteins. Bromodomain-containing proteins (BRD proteins) function are established scaffolds in the assembly of multi-protein complexes to regulate diverse biological processes. BRD proteins have been classified based on biological and functional similarity, however the functions of many BRD proteins remains unknown. PPI network analysis is useful for revealing organizational roles, identifying functional clusters, and predicting function for BRD proteins.

ResultsWe used available data to construct protein-protein interaction networks (PPINs) to study the properties of the human bromodomain protein family. The network properties of the BRD PPIN establishes that the BRD proteins serve as hub proteins that are enriched near the global center to form an inter-connected PPIN. We identified dense subgraphs formed by BRD proteins and find that different BRD proteins share topological similarity and functional associations. We explored the functional relationships through clustering and Hallmark pathway gene set enrichment analysis and identify potential biological roles for different BRD proteins.

ConclusionsIn our network analysis we confirmed that BRD proteins are conserved central nodes in the human PPI network and function as scaffolds to form distinctive functional clusters. Overall, this study provides detailed insight into the predictive functions of BRD proteins in the context of functional complexes and biological pathways.


title: Nucleome Browser: An integrative and multimodal data navigation platform for 4D Nucleome

abstract: We introduce Nucleome Browser (http://www.nucleome.org), an interactive, multimodal data visualization and exploration platform for 4D Nucleome research. Our tool effectively integrates heterogeneous datasets (e.g., genomics, imaging, 3D genome structure models, and single-cell data) and external data portals by a new adaptive communication mechanism. Nucleome Browser provides a scalable solution for integrating massive amounts of 4D Nucleome data to navigate multiscale nuclear structure and function in a wide range of biological contexts, enabling hypothesis generation and data sharing with the broad community.


title: Euchromatin activity enhances segregation and compaction of heterochromatin in the cell nucleus

abstract: The large-scale organization of the genome inside the cell nucleus is critical for the cells function. Chromatin - the functional form of DNA in cells - serves as a substrate for active nuclear processes such as transcription, replication and DNA repair. Chromatins spatial organization directly affects its accessibility by ATP-powered enzymes, e.g., RNA polymerase II in the case of transcription. In differentiated cells, chromatin is spatially segregated into compartments - euchromatin and heterochromatin - the former being largely transcriptionally active and loosely packed, the latter containing mostly silent genes and densely compacted. The euchromatin/heterochromatin segregation is crucial for proper genomic function, yet the physical principles behind it are far from understood. Here, we model the nucleus as filled with hydrodynamically interacting active Zimm chains - chromosomes - and investigate how large heterochromatic regions form and segregate from euchromatin through their complex interactions. Each chromosome presents a block copolymer composed of heterochromatic blocks, capable of crosslinking that increases chromatins local compaction, and euchromatic blocks, subjected to stochastic force dipoles that capture the microscopic stresses exerted by nuclear ATPases. These active stresses lead to a dynamic self-organization of the genome, with its coherent motions driving the mixing of chromosome territories as well as large-scale heterochromatic segregation through crosslinking of distant genomic regions. We study the stresses and flows that arise in the nucleus during the heterochromatic segregation, and identify their signatures in Hi-C proximity maps. Our results reveal the fundamental role of active mechanical processes and hydrodynamic interactions in the kinetics of chromatin compartmentalization and in the emergent large-scale organization of the nucleus.

category: biophysics


title: A Novel Chronic in vivo Oral Cadmium Exposure-Washout Mouse Model for Studying Cadmium Toxicity and Complex Diabetogenic Effects

abstract: Type II diabetes mellitus (T2DM) is characterized by insulin resistance, {beta}-cell dysfunction and hyperglycemia. In addition to well known risk factors such as lifestyle and genetic risk score, accumulation of environmental toxicants in organs relevant to glucose metabolism is increasingly recognized as additional risk factors for T2DM. Here, we describe the development of an in vivo oral cadmium (Cd) exposure model. It was shown that oral Cd exposure in drinking water followed by washout and high fat diet (HFD) in C57BL/6N mice results in islet Cd bioaccumulation comparable to that found in native human islets while mitigating the anorexic effects of Cd to achieve the same weight gain required to induce insulin resistance as in Cd naive control mice. Inter individual variation in plasma glucose and insulin levels as well as islet Cd bioaccumulation was observed in both female and male mice. Regression analysis showed an inverse correlation between islet Cd level and plasma insulin following a glucose challenge in males but not in females. This finding highlights the need to account for inter individual target tissue Cd concentrations when interpreting results from in vivo Cd exposure models. No effect of Cd on insulin secretion was observed in islets ex vivo, highlighting differences between in vivo and ex vivo cadmium exposure models. In summary, our oral in vivo Cd exposure-washout with HFD model resulted in islet Cd bioaccumulation that is relevant in the context of environmental cadmium exposure in humans. Here, we showed that islet Cd bioaccumulation is associated with complex cadmium-mediated changes in glucose clearance and {beta}-cell function. The model described here will serve as a useful tool to further examine the relationship between Cd exposure, islet Cd bioaccumulation, dysglycemia and their underlying mechanisms.

category: pharmacology and toxicology


title: Long-term ketogenic diet increases expression of SOX2-dependent oligodendrocyte- and myelination-associated genes in the aged mouse brain.

abstract: Aging is associated with multiple neurodegenerative conditions that severely limit quality of life and shorten lifespan. Studies in rodents indicate that in addition to extending lifespan, the ketogenic diet improves cognitive function in aged animals, thus improving healthspan. To broadly investigate what mechanisms might be activated in the brain in response to ketogenic diet, we conducted transcriptome wide analysis on whole brain samples from 13-month-old mice, 26-month-old mice, and 26-month-old mice fed a ketogenic diet. We observed clear activation of inflammation and complement system pathways in the 26-month-old mice relative to the younger animals. Interestingly, ketogenic diet caused a modest but significant increase in the expression of SOX2-dependent oligodendrocyte/myelination markers.

category: physiology


title: Sonic hedgehog-dependent recruitment of GABAergic interneurons into the developing visual thalamus

abstract: Axons of retinal ganglion cells (RGCs) play critical roles in the development of inhibitory circuits in visual thalamus. We previously reported that RGC axons signal astrocytes to induce the expression of fibroblast growth factor 15 (FGF15), a motogen required for GABAergic interneuron migration into visual thalamus. However, how retinal axons induce thalamic astrocytes to generate Fgf15 and influence interneuron migration remains unknown. Here, we demonstrate that impairing RGC activity had no impact on interneuron recruitment into mouse visual thalamus. Instead, our data show that retinal-derived sonic hedgehog (SHH) is essential for interneuron recruitment. Specifically, we show that thalamus-projecting RGCs express SHH and thalamic astrocytes generate downstream components of SHH signaling. Deletion of RGC-derived SHH leads to a significant decrease in Fgf15 expression, as well as in the percentage of interneurons recruited into visual thalamus. Thus, our findings identify a morphogen-dependent neuron-astrocyte signaling mechanism essential for the migration of thalamic interneurons.


title: Prepronociceptin-expressing neurons in the extended amygdala signal darting away from an aversive odor

abstract: Dysregulation in the neural circuitry that encodes physiological arousal responses is thought to contribute to the manifestation of the maladaptive behaviors observed in neuropsychiatric disorders. We previously found that prepronociceptin-expressing neurons in the bed nucleus of the stria terminalis (PnocBNST neurons) modulate rapid changes in physiological arousal upon presentation of motivationally salient stimuli (Rodriguez-Romaguera et al., 2020). However, whether PnocBNST neurons are necessary to regulate behavioral actions to motivationally salient stimuli is still unknown. Here, we investigated the role of PnocBNST neurons in encoding behavioral responses to motivationally salient stimuli using in vivo calcium imaging and optogenetic approaches in freely behaving mice. We find that the bulk activity of PnocBNST neurons increases when mice are near an aversive odor in comparison to a rewarding odor. However, optogenetic inhibition of PnocBNST neurons does not affect the amount of time mice spend near an aversive odor. Further analysis revealed that a subgroup of PnocBNST neurons that correlate with proximity to the aversive odor also correlate to darting away from the same aversive odor. Since these two behaviors are opposite to each other and since we previously found PnocBNST neurons correlate with arousal responses, we believe these results may be due in part to the encoding of arousal responses that occur when mice approach and dart away from aversive stimuli.


title: Activity of the manganese efflux transporter SLC30A10 in dopaminergic but not GABAergic neurons protects against neurotoxicity

abstract: Metals such as copper, iron, and manganese (Mn) are essential for life, but induce neurotoxicity at elevated levels. Yet, the neuronal mechanisms of metal-induced neurological disease are largely unclear. A primary limitation has been an inability to selectively alter metal levels in specific neurons, so that the role of the targeted neurons in disease biology can be isolated. Here, we show that neuron-specific depletion of metal efflux transporters provides the feasibility to overcome this limitation by focusing on Mn, which accumulates in the basal ganglia and induces motor disease, and the Mn-specific efflux transporter SLC30A10. Pan-neuronal/glial Slc30a10 knockout mice exhibited increased basal ganglia Mn levels and hypolocomotor deficits in early-life (pre-adulthood), which were exacerbated by Mn exposure. The locomotor deficits of the pan-neuronal/glial strain was associated with a reduction in evoked striatal dopamine release without dopaminergic (DAergic) neurodegeneration or changes in striatal tissue dopamine levels. Furthermore, DAergic-specific, but not GABAergic-specific, Slc30a10 knockout mice recapitulated the hypolocomotor phenotype of the pan-neuronal/glial knockouts in early-life although Mn levels were elevated in the targeted basal ganglia regions of both the neuron-specific strains. Put together, our results imply that (1) activity of SLC30A10 in DAergic neurons is necessary to protect against early-life Mn neurotoxicity; (2) increasing Mn levels in DAergic neurons is sufficient to induce early-life motor disease, suggesting that Mn targets DAergic neurons in the early-life period to induce motor deficits; and (3) neuron-specific knockout of metal efflux transporters may be a widely applicable strategy to elucidate mechanisms of metal-induced neurotoxicity.


title: A novel intranasal administration adenoviral vector-based platform for rapid COVID-19 vaccine development

abstract: The coronavirus SARS-CoV-2 has a severe impact on global public health, and the emerging variants threaten the efficacy of the circulating vaccines. Here, we report that a single vaccination with a non-replicated Chimpanzee adenovirus-based vaccine against the SARS-CoV-2 B.1.617.2 variant (JS1-delta) elicits potent humoral, cellular and mucosal immunity in mice. Additionally, a single intranasal administration of JS1-delta provides sufficient protection against B.1.617.2 challenge in mice. This study indicates that Chimpanzee adenovirus type 3 (ChAd3) derived vector represents a promising platform for antiviral vaccine development against respiratory infections, and that JS1-delta is worth further investigation in human clinical trials.

category: immunology


title: Edible formulations of chicken egg derived IgY antibodies neutralize SarsCoV2 Omicron RBD binding to human ACE2

abstract:
SarsCoV2 virus driven pandemic continues to surge propelled by new mutations such as seen in Omicron strain. Omicron is now rapidly becoming the dominant strain globally with more than 30 mutations in the spike protein. The mutations have resulted in Omicron strain escaping most of the neutralizing antibodies generated by the current set of approved vaccines and diluting the protection offered by the vaccines and therapeutic monoclonal antibodies.

This has necessitated the need for newer strategies to prevent this strain from spreading. Towards this unmet need we have developed chicken egg derived anti-RBD IgY antibodies that neutralize the binding of Omicron RBD to human ACE2. Furthermore, we have formulated the edible IgY as flavored beverages to allow for use as oral rinse and prevent the entry of Omicron in the oropharyngeal passage, a major access and accumulation point for this strain in humans.


title: Cryptic Genes for Interbacterial Antagonism distinguish Rickettsia species Infecting Blacklegged Tick from other Rickettsia pathogens

abstract:
BackgroundGenus Rickettsia (Alphaproteobacteria: Rickettsiales) encompasses numerous obligate intracellular species with predominantly ciliate and arthropod hosts. Notable species are pathogens transmitted to mammals by blood-feeding arthropods. Mammalian pathogenicity evolved from basal, non-pathogenic host-associations; however, some non-pathogens are closely related to pathogens. One such species, Rickettsia buchneri, is prevalent in the blacklegged tick, Ixodes scapularis. While I. scapularis transmits several pathogens to humans, it does not transmit Rickettsia pathogens. We hypothesize that R. buchneri established a mutualism with I. scapularis, blocking tick superinfection with Rickettsia pathogens.

MethodsTo improve estimates for assessing R. buchneri infection frequency in blacklegged tick populations, we used comparative genomics to identify an R. buchneri gene (REIS_1424) not present in other Rickettsia species present throughout the I. scapularis geographic range. Bioinformatic and phylogenomics approaches were employed to propose a function for the hypothetical protein (263 aa) encoded by REIS_1424.

ResultsREIS_1424 has few analogs in other Rickettsiales genomes and greatest similarity to non-Proteobacteria proteins. This cohort of proteins varies greatly in size and domain composition, possessing characteristics of Recombination hotspot (Rhs) and contact dependent growth inhibition (CDI) toxins, with similarity limited to proximal C-termini ([~]145 aa). This domain was named CDI-like/Rhs-like CT toxin (CRCT). As such proteins are often found as toxin-antidote (TA) modules, we interrogated REIS_1423 (151 aa) as a putative antidote. Indeed, REIS_1423 is similar to proteins encoded upstream of CRCT domain-containing proteins. Accordingly, we named these proteins CDI-like/Rhs-like C-terminal toxin antidotes (CRCA). R. buchneri expressed both REIS_1423 and REIS_1424 in tick cell culture, and PCR assays showed specificity for R. buchneri over other rickettsiae and utility for positive detection in three tick populations. Finally, phylogenomics analyses uncovered divergent CRCT/CRCA modules in varying states of conservation; however, only R. buchneri and related Tamurae/Ixodes Group rickettsiae carry complete TA modules.

ConclusionWe hypothesize that Rickettsia CRCT/CRCA modules circulate in the Rickettsia mobile gene pool, arming rickettsiae for battle over arthropod colonization. While its functional significance remains to be tested, R. buchneri CRCT/CRCA serves as a marker to positively identify infection and begin deciphering the role this endosymbiont plays in the biology of the blacklegged tick.


title: Antibacterial effects of propolis and brood comb extracts on the causative agent of European Foulbrood (Melissococcus plutonius) in honey bees (Apis mellifera)

abstract: Among a long list of parasites and pathogens that threaten the European honey bee (Apis mellifera), European Foulbrood (EFB) has become an urgent apiary disease, as epidemic outbreaks are becoming increasingly common. EFB is a bacterial disease of larval honey bees, caused by a gram-positive, anaerobic bacterium, Melissococcus plutonius. The most effective current treatment for EFB, oxytetracycline hydrochloride, can disrupt the bee microbiome, cause bee mortality and residues may persist in honey harvested for human consumption. In this study, we explore the efficacy of more sustainable bee-derived solutions, including propolis, honey comb and brood comb ethanol extracts. Using a series of dilutions of these extracts, we determined the minimum inhibitory concertation (MIC) of each bee-derived product on M. plutonius, as well as two model bacterial species, Staphylococcus saprophyticus (gram-positive) and Escherichia coli (gram-negative). Overall, we found that propolis extract was most effective at inhibiting growth of gram-positive bacteria, and that M. plutonious was also susceptible to honey comb (MIC = 16.00 mg/mL) and brood comb (MIC = 45.33 mg/mL) extracts, but at much higher concentrations than that of propolis (MIC = 1.14 mg/mL).


title: Inactivation of B. cereus Spores in Whole Milk and Almond Milk by Novel Serpentine Path Coiled Tube UV-C System

abstract: A novel continuous thin-film (1.59 mm) serpentine path coiled tube (SPCT) UV system operating at 254 nm wavelength was designed and compared with flow field distribution of whole milk with helical path coiled tube (HPCT) UV system using computational fluid dynamics. The results revealed efficient velocity magnitude distribution at serpentine bend geometric locations of the SPCT UV system. Further in this study, we evaluated B. cereus Spores inactivation in whole milk (WM) and almond milk (AM) using the developed SPCT UV system. Experimental data showed that > 4 log reduction of spores was achieved after six and ten passes of WM and AM at a flow rate of 70 and 162 mL/min, respectively. The biodosimetry method was used to verify the delivered reduction equivalent fluence (REF) and reported as 33 {+/-} 0.73 and 36.5 {+/-} 1.9 mJ/cm2. We noticed no significant effect on lipid oxidation and volatiles profile (p > 0.05) up to delivered REF of 60 mJ/cm2. This study demonstrated that high levels of inactivation of B. cereus spores could be feasible with minimal impact on product quality by UV-C processing of dairy and non-dairy opaque scattering fluids.


title: Lysine acetylation regulates antibiotic resistance in Escherichia coli

abstract:
Antibiotic resistance is increasingly becoming a serious challenge to public health. The regulation of metabolism by post-translational modifications (PTMs) has been widely studied; however, the comprehensive mechanism underlying the regulation of acetylation in bacterial resistance against antibiotics is unknown. Herein, with Escherichia coli as the model, we performed quantitative analysis of the acetylated proteome of wild-type sensitive strain (WT) and ampicillin- (Re-Amp), kanamycin- (Re-Kan), and polymyxin B-resistant (Re-Pol) strains. Based on bioinformatics analysis combined with biochemical validations, we found that a common regulatory mechanism exists between the different resistant strains. Acetylation negatively regulates bacterial metabolism to maintain antibiotic resistance, but positively regulates bacterial motility. Further analyses revealed that key enzymes in various metabolic pathways were differentially acetylated. Particularly, pyruvate kinase (PykF), a key glycolytic enzyme regulating bacterial metabolism, and its acetylated form were highly expressed in the three resistant types and were identified as reversibly acetylated by the deacetylase CobB and the acetyl-transferase PatZ, and also could be acetylated by non-enzyme AcP in vitro. Further, the deacetylation of Lys413 of PykF increased the enzyme activity by changing the conformation of ATP binding site of PykF, resulting in an increase in energy production, which in turn increased the sensitivity of drug-resistant strains to antibiotics. This study provides novel insights for understanding bacterial resistance and lays the foundation for future research on regulation of acetylation in antibiotic-resistant strains.

ImportanceThe misuse of antibiotics has resulted in an emergence of a large number of antibiotic-resistant strains, which seriously threaten human health. Bacterial metabolism is tightly controlled by protein post-translational modifications, especially acetylation. However, the comprehensive mechanism underlying regulation of acetylation in bacterial resistance remains unexplored. Here, acetylation was found to positively regulate bacterial motility and negatively regulate energy metabolism, which was common in all the different antibiotic-resistant strains. Moreover, the acetylation and deacetylation process of PykF was uncovered, and deacetylation of the Lys 413 of PykF was found to contribute to bacterial sensitivity to antibiotics. This study provides a new direction for research on development of bacterial resistance through post-translational modifications and provides a theoretical basis for the development of antibacterial drugs.


title: Identification of a CCG-enriched expanded allele in DM1 patients using Amplification-free long-read sequencing

abstract: Myotonic dystrophy type 1 (DM1) exhibits highly heterogeneous clinical manifestations caused by an unstable CTG repeat expansion reaching up to 4,000 CTG. The clinical variability depends on CTG repeat number, CNG repeat interruptions and somatic mosaicism. Currently, none of these factors are simultaneously and accurately determined due to the limitations of gold standard methods used in clinical and research laboratories. An amplicon method for targeting DM1 locus using Single-Molecule Real-Time sequencing was recently developed to accurately analyze expanded alleles. However, amplicon-based sequencing still depends on PCR and the inherent bias towards preferential amplification of smaller repeats can be problematic in DM1. Thus, an amplification-free long-read sequencing method was developed using the CRISPR/Cas9 technology in DM1. This method was used to sequence the DM1 locus in patients with CTG repeat expansion ranging from 130 to > 1000 CTG. We showed that elimination of PCR amplification improves the accuracy of measurement of inherited repeat number and somatic repeat variations, two important key factors in the DM1 severity and age at onset. For the first time, an expansion composed of over 85% CCG repeats was identified using this innovative method in a DM1 family with an atypical clinical profile. No-Amplification targeted sequencing represents a promising method that can overcome research and diagnosis shortcomings, with translational implications for clinical and genetic counseling in DM1.

category: molecular biology


title: Stress granules promote chemoresistance by triggering cellular quiescence

abstract: Cells respond to cellular stress by forming stress granules, molecular condensates containing non-translating messenger ribonucleoproteins. Stress granules form during chemotherapy and promote cell survival and chemoresistance, although the mechanism of this effect is not understood. We provide several lines of evidence that stress granules enhance cell survival by promoting cellular quiescence. First, we see a correlation between spontaneous stress granule formation and cell-cycle exit under non-stress conditions. Second, cells deficient in proteins required for stress granule formation (G3BP1/2) are less likely to exit the cell cycle under non-stress, stress, and chemotherapeutic conditions. Third, rescuing stress granule formation in G3BP1/2 knockout cells restores the fraction of cycling cells to wild-type levels. Finally, cells with enhanced stress granule formation (ddx6 knockout cells) show an increased propensity to exit the cell cycle. These results suggest that stress granules are important regulators of cellular quiescence, which could enable the identification of new anti-chemoresistance therapies that target stress granules.


title: The type 1 diabetes gene TYK2 regulates β-cell development and its responses to interferon-α

abstract: Type 1 diabetes (T1D) is an autoimmune disease that results in the destruction of insulin producing pancreatic {beta}-cells. One of the genes associated with T1D is TYK2, which encodes a Janus kinase with critical roles in type-I interferon (IFN) mediated intracellular signaling. To study the role of TYK2 in human pancreatic {beta}-cell development and response to IFN, we generated TYK2 knockout human iPSCs and directed them into the pancreatic endocrine lineage. Here we show that loss of TYK2 compromised the emergence of endocrine precursors by regulating KRAS expression while mature stem cell-islets (SC-islets) function was not affected. In the maturing SC-islets, the loss or inhibition of TYK2 prevented IFN-induced antigen processing and presentation, including MHC Class I expression in pancreatic endocrine and progenitor cells. Furthermore, in a CD8+ cytotoxic T-cell co-culture model, the survival of {beta}-cells was enhanced by a selective TYK2 inhibitor. These results identify an unsuspected role for TYK2 on {beta}-cell development and support TYK2 inhibition in adult {beta}-cells as a potent therapeutic target to halt T1D progression.


title: Redifferentiated cardiomyocytes retain residual dedifferentiation signatures and are protected against ischaemic injury

abstract: Cardiomyocyte renewal by dedifferentiation and proliferation has fueled the field of regenerative cardiology in recent years, while the reverse process of redifferentiation remains largely unexplored. Redifferentiation is characterised by the restoration of function that is lost during dedifferentiation and is key to the healing process following injury. Previously, we showed that ERBB2-mediated heart regeneration has these two distinct phases: dedifferentiation, followed by redifferentiation. Here, using temporal RNAseq and proteomics, we survey the landscape of the dedifferentiation-redifferentiation process in the adult mouse heart. We find well characterised dedifferentiation pathways, such as reduced oxphos, increased proliferation and increased EMT-like features, largely return to normal, though elements of residual dedifferentiation remain, even after contractile function is restored. These hearts appeared rejuvenated and showed robust resistance to ischaemic injury. We find that redifferentiation is driven by negative feedback signalling, notably through LATS1/2 Hippo pathway activity. Disabling LATS1/2 in dedifferentiated cardiomyocytes augments dedifferentiation in vitro and prevents redifferentiation in vivo. Taken together, our data reveal the non-trivial nature of redifferentiation, whereby elements of dedifferentiation linger in a surprisingly beneficial manner. This cycle of dedifferentiation-redifferentiation protects against future insult, in what could become a novel prophylactic treatment against ischemic heart disease for at-risk patients.


title: Breeding site microbiota and oviposition: do female Aedes aegypti shape bacterial communities?

abstract: Aedes aegypti, the main vector of multiple arboviruses, is highly associated with human dwellings. Females exhibit an opportunistic oviposition behavior, seldomly laying eggs on natural containers, but rather distributing them among human-generated breeding sites. Bacterial communities associated with such sites, as well as the compositional shifts they undergo through the development of larval stages, have been described. Some bacteria can play a direct role in supporting the success of mosquito development. Additionally, exposure to different bacteria during larval phases can have an impact on life-history traits. Whether the larvae acquire symbionts from aquatic niches, or just require bacteria as food, is still debated. Based on these facts, we hypothesized that female Ae. aegypti shape the bacterial communities of breeding sites during oviposition as a form of niche construction to favor offspring fitness. Our study presents a series of experiments to address whether gravid females modify bacterial consortia present in larval habitats. For this, we first verified if females can mechanically transfer bacteria into culture media. As evidence of mechanical transmission was obtained, we then elaborated an experimental scheme to dissect effects from factors related to the act of oviposition and mosquito-egg-water interactions. The DNA samples obtained from breeding site water aliquots pertaining to five treatments were subjected to amplicon-oriented sequencing to infer their bacterial community structure. Microbial ecology analyses revealed significant differences between treatments in terms of diversity. Particularly, between-treatment shifts in abundance profiles were detected (pairwise PERMANOVA), also showing that females induce a significant decrease in alpha diversity (1-Simpsons index) through oviposition. In addition, indicator species analysis pinpointed bacterial taxa with significant predicting values and fidelity coefficients for the samples in which single females laid eggs. Furthermore, we provide evidence regarding how one of these indicator taxa, Elizabethkingia, exerts a positive effect upon the development and fitness of mosquito larvae, thus suggesting that the developmental niche construction hypothesis may hold true in this model.

category: ecology


title: Evolutionary changes in the chromatin landscape reshape a developmental gene regulatory network during rapid life history divergence in sea urchins

abstract: Chromatin configuration is highly dynamic during embryonic development in animals, exerting an important point of control in transcriptional regulation. Yet there exists remarkably little information about the contribution of evolutionary changes in chromatin configuration to the evolution of gene expression and organismal traits. Genome-wide assays of chromatin configuration, coupled with whole-genome alignments, can help address this gap in knowledge in several ways. In this study we present a comparative analysis of regulatory element sequences and accessibility throughout embryogenesis in three sea urchin species with divergent life histories: a lecithotroph Heliocidaris erythrogramma, a closely related planktotroph H. tuberculata, and a distantly related planktotroph Lytechinus variegatus. We identified distinct epigenetic and mutational signatures of evolutionary modifications to cis-regulatory element function in H. erythrogramma that have accumulated non-uniformly throughout the genome, suggesting selection, rather than drift, underlies many modifications associated with the derived life history. Specifically, regulatory elements composing the sea urchin developmental gene regulatory network are enriched for signatures of positive selection and accessibility changes which may function to alter binding affinity and access of developmental transcription factors to these sites. Furthermore, regulatory element changes often correlate with divergent expression patterns of genes involved in cell type specification, morphogenesis, and development of other derived traits, suggesting these evolutionary modifications have been consequential for phenotypic evolution in H. erythrogramma. Collectively, our results demonstrate that selective pressures imposed by changes in developmental life history rapidly reshape the cis-regulatory landscape of core developmental genes to generate novel traits and embryonic programs.


title: Characterization of Arabian Peninsula whole exomes: exploring high inbreeding features

abstract: The exome (WES) capture enriched for UTRs on 90 Arabian Peninsula (AP) populations contributed nearly 20,000 new variants from a total over 145,000 total variants. Almost half of these variants were in UTR3, reflecting the low effort we have dedicated to cataloguing these regions, which can bear an important proportion of functional variants, as being discovered in genome-wide association studies. By applying several pathogenic predicting tools, we have demonstrated the high burden in potentially deleterious variants (especially in nonsynonymous and UTR variants located in genes that have been associated mainly with neurologic disease and congenital malformations) contained in AP WES, and the burden was as high as the consanguinity level (inferred as sum of runs of homozygosity, SROH) increased. Arabians had twice SROH values in relation to Europeans and East Asians, and within AP, Saudi Arabia had the highest values and Oman the lowest. We must pursuit cataloguing diversity in populations with high consanguinity, as the potentially pathogenic variants are not eliminated by genetic drift as much as in less consanguineous populations.


title: Different transcriptional responses by the CRISPRa system in distinct types of heterochromatin in Drosophila melanogaster

abstract:
Transcription factors (TFs) activate gene expression by binding to elements close to promoters or enhancers. Some TFs can bind to heterochromatic regions to initiate gene activation, suggesting that if a TF is able to bind to any type of heterochromatin, it can activate transcription. To investigate this possibility, we used the CRISPRa system based on dCas9-VPR as an artificial TF in Drosophila. dCas9-VPR was targeted to the TAHRE subtelomeric element, an example of constitutive heterochromatin, and to promoters and enhancers of the HOX Ultrabithorax (Ubx) and Sex Combs Reduced (Scr) genes in the context of facultative heterochromatin. dCas9-VPR robustly activated TAHRE transcription, showing that although this element is heterochromatic, dCas9-VPR was sufficient to activate its expression. In the case of HOX gene promoters, although these genes are epigenetically silenced by Polycomb complexes, both were ectopically activated. When the artificial TF was directed to enhancers, we found that the expression pattern was different compared to the effect on the promoters. In the case of the Scr upstream enhancer, dCas9-VPR activated the gene ectopically but with less expressivity; however, ectopic activation also occurred in different cells. In the case of the bxI enhancer located in the third intron of Ubx, the presence of dCas9-VPR is capable of increasing transcription initiation while simultaneously blocking transcription elongation, generating a lack of functional phenotype. Our results show that transcription can be activated in any type of heterochromatin by CRISPRa; nevertheless, its effect on transcription is subject to the intrinsic characteristics of each gene or regulatory element.

SignificanceWhether transcription only depends on activating factors binding to chromatin, even though it is found in a silent state as heterochromatin, remains an open question. In this work, we addressed this question using the CRISPRa system via dCas9-VPR as a synthetic transcriptional activator in Drosophila. This activator was directed to a constitutive heterochromatin element and to promoters and enhancers of two HOX genes, which in the tissues where they are not expressed, are present as facultative heterochromatin. In all cases, the CRISPRa system was able to activate transcription, showing that its sole presence is sufficient for this to occur. Although transcription in constitutive heterochromatin was very robust, in the case of promoters and enhancers of HOX genes, the degree of expressivity, penetrance and ectopic effect was different between promoters and enhancers. These results indicate that the presence of a synthetic activator can activate transcription by binding to transcriptional regulatory elements; however, its effect depends on the particular characteristics of each one. These results show how artificial transcription factors can be used to understand transcription regulation at the organismal level.


title: piRNA-like small RNAs target transposable elements in a Clade IV parasitic nematode

abstract: The small RNA (sRNA) pathways identified in the model organism Caenorhabditis elegans are not widely conserved across nematodes. For example, the PIWI pathway and PIWI-interacting RNAs (piRNAs) are involved in regulating and silencing transposable elements (TE) in most animals but have been lost in nematodes outside of the Caenorhabditis elegans group (Clade V), and little is known about how nematodes regulate TEs in the absence of the PIWI pathway. Here, we investigated the role of sRNAs in the Clade IV parasitic nematode Strongyloides ratti by comparing two genetically identical adult stages (the parasitic female and free-living female). We identified putative small-interfering RNAs, microRNAs and tRNA-derived sRNA fragments that are differentially expressed between the two adult stages. Two classes of sRNAs were predicted to regulate TE activity including (i) a parasite-associated class of 21-22 nt long sRNAs with a 5 uracil (21-22Us) and monophosphate modification, and (ii) 27 nt long sRNAs with a 5 guanine/adenine (27GAs) and polyphosphate modification. The 21-22Us show striking resemblance to the 21U PIWI-interacting RNAs found in C. elegans, including an AT rich upstream sequence, overlapping loci and physical clustering in the genome.


title: Introducing an argonaute-facilitated PCR platform

abstract: Argonaute proteins are characterized by highly efficient guide DNA/RNA-directed binding to target nucleic acids with high fidelity. Employing this feature, we designed an argonaute-facilitated PCR platform by making use of an argonaute derived from Mesorhizobium japonicum (MejAgo) which does not carry nuclease activity and exposes the 3 end of guide DNA when binding. Each reaction cycle of the MejAgo-PCR platform consists of a denaturing step and a polymerase-mediated extension step, omitting the annealing step required by the traditional PCR. More importantly, MejAgo-PCR could significantly improve the sensitivity of PCR in template detection due to the argonaute-facilitated pairing between guide DNA/primer and template. Thus, an argonaute-facilitated PCR has the potential to be developed as an advanced PCR platform with higher sensitivity and efficiency.

category: biochemistry


title: Structure-based design and characterization of Parkin activating mutations

abstract:
Human autosomal recessive mutations in the Parkin gene are causal for Parkinsons disease (PD). Parkin encodes a ubiquitin E3 ligase that functions together with the PD associated kinase, PINK1, in a mitochondrial quality control pathway. Structural studies reveal that Parkin exists in an inactive conformation mediated by multiple autoinhibitory domain interfaces. Here we have performed comprehensive mutational analysis of both human and rat Parkin to unbiasedly determine Parkin activating mutations across all major autoinhibitory interfaces. Out of 31 mutations tested, we identify 11 activating mutations clustered near the RING0:RING2 or REP:RING1 interfaces, which reduce the thermal stability of Parkin. Of these, we demonstrate that three mutations, V393D, A401D, and W403A located at the REP:RING1 interface were able to completely rescue a Parkin S65A mutant, defective in mitophagy, in cell-based studies. Overall our data extends previous analysis of Parkin activation mutants and suggests that small molecules that mimic REP:RING1 destabilisation offer therapeutic potential for PD patients harbouring select Parkin mutations.

Summary blurbParkin, an E3 ubiquitin ligase involved in Parkinsons disease, is inactive in the basal state and is activated by PINK1 to mediate mitophagy. Here we characterized 31 mutations and discovered three that activate Parkin and rescue loss of PINK1 phosphorylation.


title: Truvari: Refined Structural Variant Comparison Preserves Allelic Diversity

abstract: For multi-sample structural variant analyses like merging, benchmarking, and annotation, the fundamental operation is to identify when two SVs are the same. Commonly applied approaches for comparing SVs were developed alongside technologies which produce ill-defined boundaries. As SV detection becomes more exact, algorithms to preserve this refined signal are needed. Here we present Truvari - a SV comparison, annotation and analysis toolkit - and demonstrate the effect of SV comparison choices by building population-level VCFs from 36 haplotype-resolved long-read assemblies. We observe over-merging from other SV merging approaches which causes up to a 2.2x inflation of allele frequency relative to Truvari.


title: LazyPair: scalable prediction of protein-protein interactions and interaction types

abstract:
MotivationAlmost all cellular processes require protein-protein interactions. Common interaction types include binding, post-translational modifications, and catalysis. However, existing prediction tools do not take these interaction types into account and do not scale well on proteome-wide prediction.

ResultsHere we show that a random forest classifier trained on per-residue physicochemical and biochemical properties is useful for predicting protein-protein interactions. Counterintuitively, we find that training random forests by individual interaction types improves accuracy. Furthermore, a combination of these specialised classifiers improves generalisability. We call our protein-protein interaction prediction tool LazyPair. More importantly, LazyPair outperforms the state-of-the-art in accuracy, generalisability and scalability.

Availability and implementationLazyPair and the source code and data for reproducing our analysis are freely available at https://github.com/Gardner-BinfLab/PPI_Analysis_2022 and https://doi.org/10.5281/zenodo.6071630. The web server version and the source code are freely available at https://tisigner.com/lazypair/ and https://github.com/Gardner-BinfLab/TISIGNER-ReactJS, respectively.


title: Structural Basis of HIV-1 Maturation Inhibitor Binding and Activity

abstract: HIV-1 maturation inhibitors (MIs) interfere with the final step in the viral lifecycle by disrupting the ordered proteolytic processing of the viral Gag polyprotein into its individual domains. Bevirimat (BVM) and its analogs interfere with the final catalytic cleavage of spacer peptide 1 (SP1) from the capsid protein (CA) C-terminal domain (CACTD), by binding to and stabilizing the CACTD-SP1 region. MIs are under development as alternative drugs to augment current antiretroviral therapies. Although promising, their mechanism of action and associated virus resistance pathways remain poorly understood at the molecular, biochemical, and structural levels. Here, we report atomic-resolution magic angle spinning (MAS) NMR structures of microcrystalline assemblies of CACTD-SP1 complexed with BVM and/or the assembly cofactor inositol hexakisphosphate (IP6). BVM and IP6 can bind simultaneously to SP1, with BVM positioned in the center of its 6-helix bundle in a unique conformation. Importantly, the NMR-observed structural effects of BVM on IP6 binding suggest that the inhibitor stabilizes the 6-helix bundle in multiple ways. In addition, BVM-resistant SP1-A1V and SP1-V7A variants exhibit distinct conformational and binding characteristics. Taken together, our results reveal a novel allosteric mechanism by which BVM disrupts maturation and provide a structural explanation for BVM resistance as well as important guidance for the design of new MIs.


title: SKOR1 mediates FER kinase-dependent invasive growth of breast cancer cells

abstract:
High expression of the tyrosine kinase FER is an independent prognostic factor that correlates with poor survival in breast cancer patients. To investigate whether the kinase activity is essential for FER oncogenic properties, we developed an ATP analogue-sensitive knock-in allele (FERASKI). Specific FER kinase inhibition in MDA-MD-231 cells reduces migration, invasion, and metastasis in a mouse model of breast cancer. Using the FERASKI system, we identify SKI family transcriptional corepressor 1 (SKOR1) as a direct FER kinase substrate. SKOR1 loss phenocopies FER inhibition, leading to impaired proliferation, migration and invasion, and inhibition of breast cancer growth and metastasis formation in mice. We show that the candidate FER phosphorylation residue, SKOR1-Y234, is essential for FER-dependent tumor progression features. Finally, our work suggests that the SKOR1-Y234 residue promotes Smad2/3 signaling through SKOR1 binding to Smad3 attenuation.

Our study thus identifies SKOR1 as a mediator of FER-dependent breast cancer progression, advocating FER kinase inhibition as a candidate strategy to treat high-grade breast cancers.

SummaryThe SKI FAMILY TRANSCRIPTIONAL COREPRESSOR 1 (SKOR1) has been mainly associated with neuronal development. Now, Sluimer et al. identify SKOR1 as a new substrate of the oncogenic tyrosine kinase FER, and a driver of TNBC progression.

category: cancer biology


title: Loss of E-cadherin Induces IGF1R Activation Revealing a Targetable Pathway in Invasive Lobular Breast Carcinoma

abstract: No Special Type breast cancer (NST; commonly known as Invasive Ductal Carcinoma (IDC)) and Invasive Lobular Carcinoma (ILC) are the two major histological subtypes of breast cancer with significant differences in clinicopathological and molecular characteristics. The defining pathognomonic feature of ILC is loss of cellular adhesion protein, E-cadherin (CDH1). We have previously shown that E-cadherin functions as a negative regulator of the Insulin-like Growth Factor 1 Receptor (IGF1R) and propose that E-cadherin loss in ILC sensitizes cells to growth factor signaling which thus alters their sensitivity to growth factor signaling inhibitors and their downstream activators. To investigate this potential therapeutic vulnerability, we generated CRISPR-mediated CDH1 knockout (CDH1 KO) IDC cell lines (MCF7, T47D, ZR75.1) to uncover the mechanism by which loss of E-cadherin results in IGF pathway activation. CDH1 KO cells demonstrated enhanced invasion and migration that was further elevated in response to IGF1, serum and Collagen I. CDH1 KO cells exhibited increased sensitivity to IGF resulting in elevated downstream signaling. Despite minimal differences in membranous IGF1R levels between wildtype (WT) and CDH1 KO cells, significantly higher ligand-receptor interaction was observed in the CDH1 KO cells, potentially conferring enhanced downstream signaling activation. Critically, increased sensitivity to IGF1R, PI3K, Akt and MEK inhibitors was observed in CDH1 KO cells and ILC patient-derived organoids, suggesting that these targets require further exploration in ILC treatment and that CDH1 loss may be exploited as a biomarker of response for patient stratification.


title: Social dominance and reproduction result in increased integration of oxidative state in males of an African cichlid fish

abstract: Oxidative stress is a potential cost of social dominance and reproduction, which could mediate life history trade-offs between current and future reproductive fitness. However, the evidence for an oxidative cost of social dominance and reproduction is mixed, in part because organisms have efficient protective mechanisms that can counteract oxidative insults. Further, previous studies have shown that different aspects of oxidative balance, including oxidative damage and antioxidant function, varies dramatically between tissue types, yet few studies have investigated oxidative cost in terms of interconnectedness and coordination within the system. Here, we tested whether dominant and subordinate males of the cichlid Astatotilapa burtoni differ in integration of different components of oxidative stress. We assessed 7 markers of oxidative stress, which included both oxidative damage and antioxidant function in various tissue types (total of 14 measurements). Across all oxidative stress measurements, we found more co-regulated clusters in dominant males, suggesting that components of oxidative state are more functionally integrated in dominant males than they are in subordinate males. We discuss how a high degree of functional integration reflects increased robustness or efficiency of the system (e.g. increased effectiveness of antioxidant machinery in reducing oxidative damage), but we also highlight potential costs (e.g. activation of cytoprotective mechanisms may have unwanted pleiotropic effects). Overall, our results suggest that quantifying the extent of functional integration across different components of oxidative stress could reveal insights into the oxidative cost of important life history events.


title: Modeling the tonotopic map using a two-dimensional array of neural oscillators

abstract: We present a model of a tonotopic map known as the Oscillatory Tonotopic Self-Organizing Map (OTSOM). It is a 2-dimensional, self-organizing array of Hopf oscillators, capable of performing a Fourier-like decomposition of the input signal. While the rows in the map encode the input phase, the columns encode frequency. Although Hopf oscillators exhibit resonance to a sinusoidal signal when there is a frequency match, there is no obvious way to also achieve phase tuning. We propose a simple method by which a pair of Hopf oscillators, unilaterally coupled through a coupling scheme termed as modified power coupling, can exhibit tuning to the phase offset of sinusoidal forcing input. The training of OTSOM is performed in 2 stages: while the frequency tuning is adapted in stage 1, phase tuning is adapted in stage 2. Earlier tonotopic map models have modeled frequency as an abstract parameter unconnected to any oscillation. By contrast, in OTSOM, frequency tuning emerges as a natural outcome of an underlying resonant process. The OTSOM model can be regarded as an approximation of the tonotopic map found in the primary auditory cortices of mammals, particularly exemplified in the studies of echolocating bats.


title: Antimicrobial resistance in dairy slurry tanks: a critical point for measurement and control

abstract: Waste from dairy production is one of the worlds largest sources of contamination from antimicrobial resistant bacteria (ARB) and genes (ARGs). However, studies to date do not provide necessary evidence to inform antimicrobial resistance (AMR) countermeasures. We undertook a detailed, interdisciplinary, longitudinal analysis of dairy slurry waste. The slurry contained a population of ARB and ARGs, with resistances to current, historical and never-used on-farm antibiotics; resistances were associated with Gram-negative and Gram-positive bacteria and mobile elements (ISEcp1, Tn916, Tn21-family transposons). Modelling and experimental work suggested that these populations are in dynamic equilibrium, with microbial death balanced by fresh input. Consequently, storing slurry without further waste input for at least 60 days was predicted to reduce ARB spread onto land, with >99% reduction in cephalosporin resistant Escherichia coli. The model also indicated that for farms with low antibiotic use, further reductions are unlikely to reduce AMR further. We conclude that the slurry tank is a critical point for prevalence and control of AMR, and that measures to limit the spread of AMR from dairy waste should combine responsible antibiotic use, including low total quantity, avoidance of human critical antibiotics, and choosing antibiotics with shorter half-lives, coupled with appropriate slurry storage.


title: Systematic analysis of ADP-ribose detection reagents and optimisation of sample preparation to detect ADP-ribosylation in vitro and in cells

abstract: Recent evidence suggests that modification of substrates with a single ADP-ribose (ADPr) is important in for example antiviral immunity and cancer. However, the endogenous substrates and the extent of mono-ADP-ribosylation are still largely unclear. Several reagents were developed to detect ADP-ribosylation but it is unknown whether they recognise only ADPr, amino acid-ADPr linkages or a combination of ADPr with a protein backbone. We screened the affinity of selected reagents for enzymatically, chemically and in cell generated ADP-ribosylation on glutamate, cysteine, serine, arginine, threonine and RNA by blotting, as well as analysed the subcellular sites of ADP-ribosylation using immunofluorescence confocal microscopy. We furthermore observed that the modification is heat-labile and optimised sample preparation procedures. Our comparison of the available reagents, as well as optimisation of sample preparation, will allow future work further dissecting the function of ADP-ribosylation in cells, both on protein and on RNA substrates.


title: MolEvolvR: a web-app for characterizing proteins using molecular evolution and phylogeny

abstract: Studying proteins through the lens of evolution can help identify conserved features and lineage-specific variants, and potentially, their functions. MolEvolvR (http://jravilab.org/molevolvr) is a web-app that enables researchers to run a general-purpose computational workflow for characterizing the molecular evolution and phylogeny of their proteins of interest. The web-app accepts input in multiple formats: protein/domain sequences, homologous proteins, or domain scans. MolEvolvR returns detailed homolog data, along with dynamic graphical summaries, e.g., MSA, phylogenetic trees, domain architectures, domain proximity networks, phyletic spreads, and co-occurrence patterns across lineages. Thus, MolEvolvR provides a powerful, easy-to-use interface for computational protein characterization.


title: High-dimension to high-dimension screening for detecting genome-wide epigenetic regulators of gene expression

abstract:
MotivationThe advancement of high-throughput technology characterizes a wide range of epigenetic modifications across the genome involved in disease pathogenesis via regulating gene expression. The high-dimensionality of both epigenetic and gene expression data make it challenging to identify the important epigenetic regulators of genes. Conducting univariate test for each epigenetic-gene pair is subject to serious multiple comparison burden, and direct application of regularization methods to select epigenetic-gene pairs is computationally infeasible. Applying fast screening to reduce dimension first before regularization is more efficient and stable than applying regularization methods alone.

ResultsWe propose a novel screening method based on robust partial correlation to detect epigenetic regulators of gene expression over the whole genome, a problem that includes both high-dimensional predictors and high-dimensional responses. Compared to existing screening methods, our method is conceptually innovative that it reduces the dimension of both predictor and response, and screens at both node (epigenetic features or genes) and edge (epigenetic-gene pairs) levels. We develop data-driven procedures to determine the conditional sets and the optimal screening threshold, and implement a fast iterative algorithm. Simulations and two applications to long non-coding RNA and DNA methylation regulation in Kidney cancer and Glioblastoma Multiforme illustrate the validity and advantage of our method.

AvailabilityThe R package, related source codes and real data sets used in this paper are provided at https://github.com/kehongjie/rPCor.


title: Tie-lines reveal interactions driving heteromolecular condensate formation

abstract: Phase separation of biomolecules give rise to membraneless organelles that contribute to the spatiotemporal organisation of the cell. In most cases, such biomolecular condensates contain multiple components, but the manner in which interactions between components control the stability of condensates remained challenging to elucidate. Here, we develop an approach to determine tie-line gradients in ternary liquid-liquid phase separation (LLPS) systems, based on measurements of the dilute phase concentration of only one component. We show that the sign of the tie-line gradient is related to the cross-interaction energy between the polymers in the system and discriminates between competitive and cooperative phase separation. Using this approach, we studied the interaction between protein Fused in Sarcoma (FUS) and polyethylene glycol (PEG) polymer chains, and measured positive tie-line gradients. Our results show that PEG drives LLPS through an associative interaction with FUS and is not an inert crowder. We further studied the interaction between PolyA RNA (3.0{+/-}0.5kDa) and the protein G3BP1, and using the tie-line gradient as a reporter for the stoichiometry of polymers in the condensate we determined a G3BP1-to-PolyA RNA molar ratio of 1:4 in the dense phase. Our framework for measuring tie-line gradients opens up a route for the characterisation of interaction types and compositions in ternary LLPS systems.


title: A microfluidic platform for characterizing the structure and rheology of biofilm streamers

abstract: Biofilm formation is the most successful survival strategy for bacterial communities. In the biofilm lifestyle, bacteria embed themselves in a self-secreted matrix of extracellular polymeric substances (EPS), which acts as a shield against mechanical and chemical insults. When ambient flow is present, this viscoelastic scaffold can take a streamlined shape, forming biofilm filaments suspended in flow, called streamers. Streamers significantly disrupt the fluid flow by causing rapid clogging and affect transport in aquatic environments. Despite their relevance, the structural and rheological characterization of biofilm streamers is still at an early stage. In this work, we present a microfluidic platform that allows the reproducible growth of biofilm streamers in controlled physico-chemical conditions and the characterization of their biochemical composition, morphology, and rheology in situ. We employed isolated micropillars as nucleation sites for the growth of single biofilm streamers under the continuous flow of a diluted bacterial suspension. By combining fluorescent staining of the EPS components and epifluorescence microscopy, we were able to characterize the biochemical composition and morphology of the streamers. Additionally, we optimized a protocol to perform hydrodynamic stress tests in situ, by inducing controlled variations of the fluid shear stress exerted on the streamers by the flow. Thus, the reproducibility of the formation process and the testing protocol make it possible to perform several consistent experimental replicates that provide statistically significant information. By allowing the systematic investigation of the role of biochemical composition on the structure and rheology of streamers, this platform will advance our understanding of biofilm formation.


title: The RNA binding protein Nab2 genetically interacts with multiple RNA exosome cofactors to regulate target RNAs

abstract: RNA binding proteins play important roles in the processing and precise regulation of RNAs. Highlighting the biological importance of RNA binding proteins is the increasing number of human diseases that result from mutations in genes that encode these proteins. We recently discovered that mutations in the ZC3H14 gene, which encodes an evolutionarily conserved polyadenosine RNA-binding protein, cause intellectual disability. Studies of the budding yeast orthologue of ZC3H14, Nuclear Poly(A) Binding protein 2 (Nab2), have provided insight into the functions of this protein. The NAB2 gene is essential in S. cerevisiae, and conditional nab2 mutants cause defects in a number of steps in RNA processing. To explore the critical functions of the Nab2/ZC3H14 protein family, we performed a high-copy suppressor screen on nab2 mutant cells. This screen identified genes encoding two core subunits of the RNA exosome, as well as Nrd1 and Ski7, nuclear and cytoplasmic cofactors of the RNA exosome, respectively. Nrd1 is an RNA binding protein that is part of the Nrd1-Nab3-Sen1 (NNS) complex, which plays an important role in transcription termination of non-coding RNAs. Ski7 is a GTP-binding protein that mediates interaction between the RNA exosome and the Ski complex, which targets RNA transcripts to the exosome for processing and degradation in the cytoplasm. To explore the functional interactions between the RNA exosome and Nab2, we employed RNA-seq analysis to identify the transcripts most impacted by overexpression of these exosome cofactors in nab2 mutant cells. This analysis revealed that many transcripts show small changes in steady-state levels, consistent with a global role of Nab2 in modulating transcript stability. This study uncovers functional interactions between the RNA exosome and Nab2 in both the nucleus and the cytoplasm.


title: Population structure and genetic connectivity of the scalloped hammerhead shark (Sphyrna lewini) across nursery grounds from the Eastern Tropical Pacific: implications for management and conservation

abstract: Defining demographically independent units and understanding gene flow between them is essential for managing and conserving exploited populations. The scalloped hammerhead shark, Sphyrna lewini, is a coastal semi-oceanic species found worldwide in tropical and subtropical waters. Pregnant females give birth in shallow coastal estuarine habitats that serve as nursery grounds for neonates and small juveniles, and adults move offshore and become highly migratory. We evaluated the population structure and connectivity of S. lewini in coastal areas across the Eastern Tropical Pacific (ETP) using both sequences of the mitochondrial DNA control region (mtCR) and nuclear-encoded microsatellite loci. The mtCR defined two genetically discrete geographic groups: the Mexican Pacific and the central-southern Eastern Tropical Pacific (Guatemala, Costa Rica, Panama, and Colombia). Overall, the mtCR data showed low levels of haplotype diversity ranged from 0.000 to 0.608, while nucleotide diversity ranged from 0.000 to 0.0015. A more fine-grade population structure analysis was detected using microsatellite loci where Guatemala, Costa Rica, and Panama differed significantly. Genetic diversity analysis with nuclear markers revealed an observed heterozygosity ranging from 0.68 to 0.71 and an allelic richness from 5.89 to 7.00. Relatedness analysis revealed that individuals within nursery areas were more closely related than expected by chance, suggesting that S. lewini may exhibit reproductive philopatric behaviour within the ETP. Findings of at least two different management units, and evidence of philopatric behaviour call for intensive conservation actions for this critically endangered species in the ETP.


title: Dynamic Sex Differences in Reactive and Appetitive Aggression

abstract: Aggression is an evolutionarily conserved, adaptive component of social behavior. Studies in male mice illustrate that aggression is influenced by numerous factors including the degree to which an individual finds aggression rewarding and will work for access to attack and subordinate mice. While such studies have expanded our understanding of the molecular and circuit mechanisms of male aggression very little is known about female aggression, owed in part to limited availability of valid mouse models in females. Here we use an ethologically relevant model of male vs. female aggression by pair housing adult male and female outbred CFW mice with opposite sex cage mates. We assess reactive (defensive) aggression in the resident intruder (RI) test and appetitive (rewarding) aggression in the aggression conditioned place preference (CPP) and operant self-administration (SA) tests. Our results show dramatic sex differences in both qualitative and quantitative aspects of reactive vs. appetitive aggression. Males exhibit more wrestling and less investigative behavior during RI, find aggression rewarding and will work for access to a subordinate to attack. Females exhibit more bites, alternate between aggressive behaviors and investigative behaviors more readily during RI, however, they do not find aggression to be rewarding or reinforcing. These results establish sex differences in aggression in mice, providing an important resource for the field to better understand the circuit and molecular mechanisms of aggression in both sexes.


title: Long-term learning transforms prefrontal cortex selectivity during working memory

abstract: The lateral prefrontal cortex (lPFC) is reliably active during working memory (WM) across human and animal models, but the role of lPFC in successful WM is under debate. For instance, non-human primate (NHP) electrophysiology research finds that lPFC circuitry stores WM representations. Human neuroimaging instead suggests that lPFC plays a control function over WM content that is stored in sensory cortices. These seemingly incompatible WM accounts are often confounded by differences in the amount of task training and stimulus exposure across studies (i.e., NHPs tend to be trained extensively). Here, we test the possibility that such long-term training may alter the role of lPFC in WM maintenance. We densely sampled WM-related activity across learning, in three human participants, using a longitudinal functional MRI (fMRI) protocol. Over three months, participants trained on (1) a serial reaction time (SRT) task, wherein complex fractal stimuli were embedded within probabilistic sequences, and (2) a delayed recognition task probing WM for trained or novel stimuli. Participants were scanned frequently throughout training, to track how WM activity patterns change with repeated stimulus exposure and long-term associative learning. WM task performance improved for trained (but not novel) fractals and, neurally, delay activity significantly increased in distributed lPFC voxels across learning. Pattern similarity analyses also found that item-level WM representations emerged within lPFC, but not in sensory cortices, and lPFC delay activity increasingly reflected sequence relationships from the SRT task, even though that information was task-irrelevant for WM. These findings demonstrate that human lPFC develops stimulus-selective WM responses with learning and WM representations are shaped by long-term experience. Influences from training and long-term memory may reconcile competing accounts of lPFC function during WM.


title: Influence of claustrum on cortex varies by area, layer, and cell type

abstract: The claustrum is a small subcortical structure with widespread connections with disparate regions of the cortex. These far-reaching projections have led to many hypotheses concerning its function. However, we know little about how claustrum input affects neural activity in cortex, particularly beyond frontal areas. Here, using optogenetics and multi-regional Neuropixels recordings from over 15,000 neurons in awake mice, we demonstrate that the effect of claustrum input differs depending on brain area, layer, and cell type. Brief claustrum stimulation produces approximately 1 spike per claustrum neuron, which affects many fast-spiking (FS; putative inhibitory) but very few regular-spiking (RS; putative excitatory) cortical neurons. Prolonged claustrum stimulation affects many more cortical FS and RS neurons. More inhibition occurs in frontal regions and deeper layers, while more excitation occurs in posterior regions and superficial layers. These differences imply that the function of claustrum input to cortex depends on the area, supporting the idea that claustro-cortical circuits are organized into functional modules.


title: Associative memory of structured knowledge

abstract:
A long standing challenge in biological and artificial intelligence is to understand how new knowledge can be constructed from known building blocks in a way that is amenable for computation by neuronal circuits. Here we focus on the task of storage and recall of structured knowledge in longterm memory. Specifically, we ask how recurrent neuronal networks can store and retrieve multiple knowledge structures. We model each structure as a set of binary relations between events and attributes (attributes may represent e.g., temporal order, spatial location, role in semantic structure), and map each structure to a distributed neuronal activity pattern using a vector symbolic architecture (VSA) scheme.

We then use associative memory plasticity rules to store the binarized patterns as fixed points in a recurrent network. By a combination of signal-to-noise analysis and numerical simulations, we demonstrate that our model allows for efficient storage of these knowledge structures, such that the memorized structures as well as their individual building blocks (e.g., events and attributes) can be subsequently retrieved from partial retrieving cues. We show that long-term memory of structured knowledge relies on a new principle of computation beyond the memory basins. Finally, we show that our model can be extended to store sequences of memories as single attractors.


title: Systematic Elucidation and Pharmacological Targeting of Tumor-Infiltrating Regulatory T Cell Master Regulators

abstract:
Due to their immunosuppressive role, tumor-infiltrating regulatory T cells (TI-Tregs) represent attractive therapeutic targets. Analysis of TI vs. peripheral Tregs (P-Tregs) from 36 patients, across four malignancies, identified 17 candidate Master Regulators (MRs), predicted to mechanistically regulate TI-Tregs transcriptional state. In vivo, pooled CRISPR-KO screening, using a hematopoietic stem cell transplant model, confirmed essentiality of 7 of 17 MRs in TI-Treg recruitment and/or retention to the TME, without affecting other T cell subtypes, while individual knockout of the most significant MR (TRPS1) significantly reduced tumor allograft growth. TI-Treg drug perturbation profile analysis identified drugs capable of inverting the TI-Treg-specific MR activity signature at low concentration. Low dose treatment with gemcitabine (top prediction) inhibited tumor growth in immunocompetent but not immunocompromised allografts, increased PD-1 inhibitor efficacy, and depleted TI-Tregs in vivo. The study provides key insight into Treg infiltration mechanism and a gene reporter assay to identify additional small molecule inhibitors.

Graphical Abstract

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title: Honey bees change the microbiota of pollen

abstract: Pollen, as all other plant tissues, harbors different microorganisms. As honey bees (Apis mellifera) collect and pack pollen they add regurgitated nectar to moisten and glue the pollen grains, possibly changing the microbial composition. In this study, we compared the microbiota of clean Brassica napus pollen with that of bee-worked corbicular pollen. We found that by working pollen, bees increase the bacterial diversity of pollen, by adding honey bee symbionts such as Bombella, Frischella, Gilliamella and Snodgrassella, bee pathogens as Spiroplasma and nectar dwelling Lactobacillus to the new pollen microbiota. The bee gut is an important source of inoculum of the corbicular pollen microbiota. We discuss the implications of these findings and propose future research avenues.


title: Practical application of indicators for genetic diversity in CBD post-2020 Global Biodiversity Framework implementation

abstract:
Genetic diversity is a key aspect of biological variation for the adaptability and survival of populations of species, which must be monitored to assure maintenance. We used data from the Swedish Red-List 2020 and a recent government report to apply three indicators for genetic diversity proposed for the post-2020 Global Biodiversity Framework of the Convention on Biological Diversity (CBD). We made more detailed indicator assessments for mammals and herptiles.

For indicator 1, the proportion of populations with effective population size Ne>500, 33% of 22557 investigated species had a population estimate. For herptiles and mammals, 70% and 49%, respectively likely had Ne>500.

For indicator 2, the proportion of populations or range remaining within species, 20% of all species evaluated for the Red-List have data. Meanwhile, 32% of the herptile and 84% of the mammal populations are maintaining their populations/range.

For indicator 3, the number of species/populations in which genetic diversity is monitored using DNA-based methods, there are studies on 3% of species, and 0.3% are monitored. In contrast 68% of mammals and 29% of herptiles are studied using DNA, and 8% of mammals and 24% of herptiles are genetically monitored.

We conclude that the Red List provide data suitable for evaluating the genetic indicators, but the data quality can be improved. There is a synergy in estimating the genetic indicators in parallel with the Red-Listing process. We propose that indicator values could be included in national Red-Listing as a new category - "genetically threatened", based on the genetic indicators.