title: An Ethogram Identifies Behavioural Markers of Attention to Humans in European Herring Gulls (Larus argentatus)

abstract: Herring gulls are one of the few species that thrive in anthropogenic landscapes. Their longstanding history of urbanisation and familiarity with people makes them an excellent target for studies of the effects of urban living and cognition. Previous studies highlight a connection between food-stealing behaviour, success in anthropogenic areas and an increase in attention towards humans, opening up questions about the exact extent of a gulls knowledge of human food cues and how this manifests itself in behavioural changes. To investigate these questions further, behavioural responses to human cues in a food-related context are investigated and presented in a systematic ethogram, which identifies three distinct markers of attention. Those behaviours - head turns, approaches, and the angular body position relative to the experimenter - differ significantly between control and food conditions, showing that attention towards humans in a food-related context is upregulated and this is reflected in the gulls behaviour. In food condition trials, head turns occur more often and gulls face towards the experimenter ([≤]90{degrees}) with occasional approaches that are never seen in control conditions. Acoustic and behavioural human food-like cues alone were insufficient to elicit a response similar to that seen in presence of a real food item, indicating that gulls specifically pay attention to humans in foraging contexts. The results show situation-dependent attentional modulation in gulls, highlight knowledge gaps about how Herring gulls might identify food objects and provide an ethogram description of attentive behaviours that can be used in further study of attentional modulation in gulls.

category: animal behavior and cognition


title: High-resolution lung adenocarcinoma expression subtypes identify tumors with dependencies on MET, CDK4, CDK6, and PD-L1

abstract:
Lung adenocarcinoma is one of the most common cancer types with various treatment modalities. However, better biomarkers to predict therapeutic response are still needed to improve precision medicine. We utilized a consensus hierarchical clustering approach on 509 LUAD cases from TCGA to identify five robust LUAD expression subtypes. We then integrated genomic (patient and cell line) and proteomic data to help define biomarkers of response to targeted therapies and immunotherapies. This approach defined subtypes with unique proteogenomic and dependency profiles. S4-associated cell lines exhibited specific vulnerability to CDK6 and CDK6-cyclin D3 complex gene, CCND3. S3 was characterized by dependency on CDK4, immune-related expression patterns, and altered MET signaling; experimental validation showed that S3-associated cell lines responded to MET inhibitors, leading to increased PD-L1 expression. We further identified genomic features in S3 and S4 as biomarkers for enabling clinical diagnosis of these subtypes. Overall, our consensus hierarchical clustering approach identified robust tumor expression subtypes, and our subsequent integrative analysis of genomics, proteomics, and CRISPR screening data revealed subtype-specific biology and vulnerabilities. Our lung adenocarcinoma expression subtypes and their biomarkers could help identify patients likely to respond to CDK4/6, MET, or PD-L1 inhibitors, potentially improving patient outcome.

SignificanceThrough integrative analysis of genomic, proteomic, and drug dependency data, we identified robust lung adenocarcinoma expression subtypes and found subtype-specific biomarkers of response, including CDK4/6, MET, and PD-L1 inhibitors.

category: cancer biology


title: Distribution of the National Science Foundation's Advancing Informal STEM Learning Awards (AISL) between 2006-21

abstract: The COVID-19 pandemic tested many fundamental connections between science and society. A growing field working to strengthen those connections exists within the informal STEM learning (ISL) community which provides diverse learning and engagement environments outside the formal classroom. One of the largest funders of ISL initiatives is the National Science Foundation (NSF) which runs the Advanced Informal STEM Learning (AISL) program in the United States. The AISL program supports initiatives through six categories that include pilots and feasibility studies, research in service to practice, innovations in development, broad implementation, literature reviews, syntheses, or meta-analyses, and conferences. However, a number of questions remain unanswered with respect to the distribution and ultimately the broad impact of the awards. In this study, we analyzed publicly available awardee information across a 15-year period (2006-2021) to provide a preliminary analysis of how the AISL grants are distributed across individual states, organizations, and the principal investigators. Several states, organizations, and principal investigators stood out as prolific awardees. Massachusetts and California represented the largest share of awards at 14% and 13% respectively during that time. WGBH Educational Foundation located in Massachusetts and the Exploratorium located in California, received the largest number of awards during the 15 year period. Notably, 67% of the AISL awards list at least one co-principal investigator. Our report brings to light a number of new questions and charts new paths of exploration for future studies.

category: scientific communication and education


title: A glimpse into the DNA virome of the unique "living fossil" Welwitschia mirabilis

abstract: Here, we report the identification and characterization of four novel DNA viruses associated with Welwitschia mirabilis. Complete circular virus-like sequences with affinity to Caulimoviridae members and geminiviruses were detected and characterized from Welwitschia mirabilis genomic data. The two newly Caulimoviridae-like viruses have been tentatively named as Welwitschia mirabilis virus 1 and 2 (WMV1-WMV2); whereas the two identified geminiviruses were named as Welwitschia mirabilis associated geminivirus A and B (WMaGVA-WMaGVB). Phylogenetic analysis suggests that WMV1-2 belong to a proposed genus of Caulimoviridae-infecting gymnosperms. WMaGVA-B are phylogenetically related with both mastreviruses and capulaviruses and likely represent a distinct evolutionary lineage within geminiviruses.

category: microbiology


title: Cell cycle progression requires repression by Groucho during S-phase and its relief at G2-phase

abstract: The cell cycle depends on a sequence of steps that are triggered and terminated via the synthesis and degradation of phase-specific transcripts and proteins. While much is known about how stage-specific transcription is activated, less is understood about how inappropriate gene expression is suppressed. In this paper we demonstrate that Groucho, the Drosophila orthologue of TLE1 and other related human transcriptional corepressors, regulates cell cycle progression in vivo. We show that although Groucho is expressed throughout the cell cycle, its activity is selectively inactivated by phosphorylation, except during S-phase when it represses e2f1 expression. Misregulated Groucho activity causes cell cycle arrest; in particular, both constitutive Groucho activity and failure to repress e2f1 cause cell cycle arrest phenotypes. We also show that the Cdk1 kinase is responsible for stage-specific phosphorylation of Groucho in vivo. We propose that Groucho and its orthologues play key roles in the metazoan cell cycle that may explain the links between TLE corepressors and several types of human cancer.

category: developmental biology


title: A highly sensitive LC-MS/MS method for quantitative determination of 7 vitamin D metabolites in mouse brain tissue

abstract: Despite its critical role in neurodevelopment and brain function, vitamin-D (vit-D) homeostasis, metabolism and kinetics within the central nervous system remain largely undetermined. Thus, it is of critical importance to establish an accurate, highly sensitive and reproducible method to quantitate vit-D in brain tissue. Here, we present a novel liquid chromatography tandem mass spectrometry (LC-MS/MS) method and for the first time, demonstrate detection of seven major vit-D metabolites in brain tissues of C57BL/6J wild-type mice, namely: 1,25(OH)2D3, 3-epi-1,25(OH)2D3, 1,25(OH)2D2, 25(OH)D3, 25(OH)D2, 24,25(OH)2D3, and 24,25(OH)2D2. Chromatographic separation was achieved on a pentaflurophenyol column 3 mM ammonium formate with water/methanol [A] and methanol/isopropanol [B] phases. Detection was by positive-ion electrospray tandem mass spectrometry. We used calibration standards of each metabolite prepared in brain matrices to validate the detection range, precision, accuracy and recovery. Isotopically labelled analogues, 1,25(OH)2D3-d3, 25(OH)D3-C5 and 24,25(OH)2D3-d6, served as the internal standards for the closest molecular related metabolite in all measurements. The calibration range was between 1 fg/mL to 10 ng/mL with an LLOD and LLOQ of 10 fg/mL and 3 fg/mL, respectively. The intra-/inter-day precision and accuracy for measuring brain vit-D metabolites ranged between 0.12-11.53% and 0.28-9.11%, respectively. Recovery ranged between 99.06% and 106.9% for all metabolites. Collectively, the sensitivity and efficiency of our method supersedes previously reported protocols used to measure vit-D and to our knowledge, the first protocol to reveal the abundance of 25(OH)D2, 1,25(OH)D2 and 24,25(OH)2D2, in brain tissue of any species. This technique may be important in supporting the future advancement of pre-clinical research into the function of vit-D in neurophysiological, neuropsychiatric, and neurodegeneration.

category: physiology


title: Neural Kernels for Recursive Support Vector Regression as a Model for Episodic Memory

abstract: Retrieval of episodic memories requires intrinsic reactivation of neuronal activity patterns. The content of the memories are thereby assumed to be stored in synaptic connections. This paper proposes a theory in which these are the synaptic connections that specifically convey the temporal order information contained in the sequences of a neuronal reservoir to the sensory-motor cortical areas that give rise to the subjective impression of retrieval of sensory motor events. The theory is based on a novel recursive version of support vector regression that allows for efficient continuous learning that is only limited by the representational capacity of the reservoir. The paper argues that hippocampal theta sequences are a potential neural substrate underlying this reservoir.The theory is consistent with confabulations and post-hoc alterations of existing memories.

category: neuroscience


title: Wide-field Calcium Imaging Reveals Widespread Changes in Cortical Connectivity Following Repetitive, Mild Traumatic Brain Injury in the Mouse

abstract:
The physiologic basis underlying the long-term consequences of repetitive, mild traumatic brain injury (mTBI) remains poorly understood. Mild traumatic brain injury often results in brief loss of consciousness, impaired attention and concentration, memory problems, impulsivity, and headache, without objective findings on clinical imaging or examination. The effects of mTBI can persist and become cumulative with repetitive injury, suggesting global alterations in cortical networks. Using transparent polymer skulls, we performed mesoscopic Ca2+ imaging in mice to evaluate how repetitive mTBI alters patterns of neuronal interactions across the dorsal cerebral cortex. Spatial Independent Component Analysis (sICA) and Localized semi-Nonnegative Matrix Factorization (LocaNMF) were used to quantify changes in cerebral functional connectivity (FC). Repetitive, mild, controlled cortical impacts induce temporary neuroinflammatory responses, characterized by increased density of microglia exhibiting de-ramified morphology. These temporary neuro-inflammatory changes were not associated with compromised cognitive performance in the Barnes maze or motor function as assessed by rotarod. However, long-term alterations in functional connectivity were observed. Widespread, bilateral changes in FC occurred immediately following impact and persisted for up to 7 weeks, the duration of the experiment. Network alterations include decreases in global efficiency, clustering coefficient, and nodal strength, thereby disrupting functional interactions and information flow throughout the dorsal cerebral cortex. A subnetwork analysis shows the largest disruptions in FC were concentrated near the impact site. Therefore, repetitive mTBI induces a transient neuroinflammation, without alterations in cognitive or motor behavior, and a reorganized cortical network evidenced by the widespread, chronic alterations in cortical FC.

Significance StatementMore than 2.5 million individuals in the United States suffer minor traumatic brain injuries annually. Because these injuries are typically not associated with visible anatomic injuries or objective clinical findings, they were thought benign and fully recoverable. However, there is increasing awareness of the long-term deleterious consequences, particularly in patients who suffer repeated mTBI. Using long-term, mesoscopic neuronal Ca2+ imaging to characterize the dorsal cerebral cortical connectome following repetitive mTBI, we show extensive, persistent changes in functional connectivity, not only at the site of injury but throughout the cortex. These findings provide new insights into the pathophysiology of mTBI.


title: Chronic Glucocorticoid Stress Reveals Increased Energy Expenditure and Accelerated Aging as Cellular Features of Allostatic Load

abstract: Stress triggers energy-dependent, anticipatory responses that promote survival, a phenomenon termed allostasis. However, chronic activation of allostatic responses results in allostatic load, a dysregulated state that predicts functional decline, accelerates aging, and increases mortality in humans. The cellular basis for the damaging effects of allostatic load remains unclear. By longitudinally profiling primary human fibroblasts across their lifespan, we show that chronic glucocorticoid stress induces a ~60% increase in cellular energy expenditure and a greater reliance on mitochondrial oxidative phosphorylation (OxPhos) for energy production. Chronic stress also causes mtDNA instability, affects age-related cytokines secretion, and accelerates cellular aging, based on DNA methylation clocks, telomere shortening rate, and reduced lifespan. Finally, pharmacologically normalizing OxPhos activity while further increasing energy expenditure induces an exacerbation of the aging phenotype. Altogether, our findings define multi-omic recalibrations of stress adaptation, underscoring increased energy expenditure and accelerated cellular aging as interrelated features of cellular allostatic load.

category: cell biology


title: PERK Signaling Promotes Mitochondrial Elongation By Remodeling Membrane Phosphatidic Acid

abstract: Endoplasmic reticulum (ER) stress and mitochondrial dysfunction are linked in the onset and pathogenesis of numerous diseases. This has led to considerable interest in defining the mechanisms responsible for regulating mitochondria during ER stress. The PERK signaling arm of the unfolded protein response (UPR) has emerged as a prominent ER stress-responsive signaling pathway that regulates diverse aspects of mitochondrial biology. Here, we show that PERK activity also promotes adaptive remodeling of mitochondrial membrane phosphatidic acid (PA) to induce protective mitochondrial elongation during ER stress. We find that PERK activity is required for ER stress-dependent increases in both cellular PA and YME1L-dependent degradation of the intramitochondrial PA transporter PRELID1. Together, these processes lead to the accumulation of PA on the outer mitochondrial membrane where it induces mitochondrial elongation by inhibiting mitochondrial fission. Our results establish a new role for PERK in the adaptive remodeling of mitochondrial phospholipids and demonstrate that PERK-dependent PA regulation functions to adapt organellar shape in response to ER stress.


title: Live-Cell Imaging in Human Colonic Monolayers Reveals Erk Waves Limit the Stem Cell Compartment to Maintain Epithelial Homeostasis

abstract: The establishment and maintenance of different cellular compartments in tissues is a universal requirement across all metazoans. Maintaining the correct ratio of cell types in time and space allows tissues to form patterned compartments and perform complex functions. Patterning is especially evident in the human colon, where tissue homeostasis is maintained by stem cells in crypt structures that balance proliferation and differentiation. Here we developed a 2D patient derived organoid (PDO) screening platform to study tissue patterning and kinase pathway dynamics in single cells across hundreds of conditions. Using this system, we discovered that waves of Erk signaling induced by apoptotic cells play a critical role in maintaining tissue patterning and homeostasis. If Erk is activated acutely across all cells instead of in wavelike patterns, then tissue patterning, and stem cell maintenance are lost. Conversely, if Erk activity is inhibited, then stem cells become unrestricted and expand dramatically. This work demonstrates that the colonic epithelium requires coordinated Erk signaling dynamics to maintain patterning and tissue homeostasis. Our work reveals how Erk can antagonize stem cells yet support cell replacement and the function of the gut.


title: Intergenerational transfer and sex differences of DNA methylation patterns in the Pacific oyster (Crassostrea gigas)

abstract:
Apart from DNA-sequence-based inheritance, inheritance of epigenetic marks such as DNA methylation is controversial across the tree of life. In mammals, post-fertilization and primordial germ cell reprogramming processes erased most parental DNA methylation information. In nonmammalian vertebrates and insects, it has been proposed that DNA methylation is an essential hereditary carrier. However, how and to what extent general DNA methylation reprogramming affects intergenerational inheritance in molluscs remains unclear. Here, we investigated genome-wide DNA methylation in a mollusc model, the Pacific oyster (Crassostrea gigas), to test how epigenetic information transfers from parents to offspring. Analysis of global methylome revealed that the DNA methylation patterns are highly conserved within families. Almost half of the differentially methylated CpG dinucleotides (DMCs) between families in parents could transfer to offspring. These results provided the direct evidence for the hypothesis that the Pacific oyster DNA methylation patterns are inherited in generations. Moreover, distinct DNA methylation differences between male and female somatic tissues in C. gigas are revealed in this study. These sex-differential methylated genes significantly enriched in the regulation of Rho protein signal transduction process, which indicated that DNA methylation might have an essential role in the sexual differentiation of somatic tissues in C. gigas.

Author SummaryTransgenerational inheritance of DNA methylation marks varies across the tree of life. In mammals, post-fertilization and primordial germ cell reprogramming processes obstructed the DNA methylation transmission from parents to child, and only some CpG dinucleotides retain gamete-inherited methylation. However, the DNA methylation inheritance seems apparent in nonmammalian vertebrates and insects. As one of the essential mollusc models, the Pacific oyster Crassostrea gigas have received the most substantial epigenetic studies, mainly focusing on the DNA methylation profiles. While a previous study suggested the existence of paternal inheritance of DNA methylation patterns in C. gigas, more data are needed to confirm this hypothesis. In this study, genome-wide DNA methylation analysis was performed to investigate the epigenetic inheritance in C. gigas. Almost half of the DNA methylation differences between families in parents were found to be transferred to children, indicating the absence of global DNA methylation reprogramming in C. gigas. Besides, extensive hypomethylation in C. gigas females compared with males were also unveiled. These hypomethylated genes were significantly enriched in the regulation of Rho protein signal transduction process. For example, guanine nucleotide exchange factors, including KALRN, FGD1, and FGD6, were hypomethylated in C. gigas females, and the corresponding transcriptions were significantly upregulated. Our findings provided insights into the evolution of DNA methylation patterns, transgenerational epigenetic inheritance, and sexual differentiation in molluscs.

category: evolutionary biology


title: Functional involvement of multiple genes as members of the supergene unit in the female-limited Batesian mimicry of Papilio polytes

abstract: In recent years, many examples have been reported in which complex adaptive traits are regulated by supergene units consisting of multiple genes adjacent to each other on the chromosome. However, few supergene units have been functionally characterized, and it has not been demonstrated that multiple genes in a supergene unit are involved in the adaptive trait. The H locus on chromosome 25 has been shown to be responsible for the female-limited Batesian mimicry in Papilio polytes. The mimetic H locus, about 130 kb, is inverted at both ends from the non-mimetic h locus, and the sequences between H and h locus are diversified (Highly Diversified Region: HDR). The HDR of the H locus contains three genes, doublesex (dsx), U3X, and UXT, and RNAi analyses have shown that dsx is involved in the mimetic pattern formation. Here, we showed that knockdown by RNAi and knockout by Crspr/Cas9 of the transcriptional regulator UXT altered the mimetic pattern to the non-mimetic like pattern in hindwings. In addition, it was also shown by RNAi that long non-coding RNA U3X was functionally involved in the mimetic pattern formation, and by RNA-seq analysis that U3X induced the dsx expression and repressed the UXT expression. In addition, the knockdown of prospero and sir2 genes adjacent to the HDR also affected the hindwing pattern formation. These results indicate that not only genes within the HDR but also adjacent genes outside the HDR may be involved in the mimicry supergene function. This suggests that cis-elements regulating multiple gene expression exists within the HDR of the H-allele, and the need to consider a more extended functional unit of the supergene. This paper shows that multiple genes inside and outside the HDR are functionally involved in the formation of complex adaptive traits, and provides a new perspective on the evolution of the structure and functional units of the supergene.

category: genetics


title: Large haplotypes highlight a complex age structure within the maize pan-genome

abstract: The genomes of maize and other eukaryotes contain stable haplotypes in regions of low recombination. These regions, including centromeres, long heterochromatic blocks and rDNA arrays have been difficult to analyze with respect to their diversity and origin. Greatly improved genome assemblies are now available that enable comparative genomics over these and other non-genic spaces. Using 26 complete maize genomes, we developed methods to align intergenic sequences while excluding genes and regulatory regions. The centromere haplotypes (cenhaps) extend for megabases on either side of the functional centromere regions and appear as evolutionary strata, with haplotype divergence/coalescence times dating as far back as 450 thousand years ago (kya). Application of the same methods to other low recombination regions (heterochromatic knobs and rDNA) and all intergenic spaces revealed that deep coalescence times are ubiquitous across the maize pan-genome. Divergence estimates vary over a broad time scale with peaks at [~]300 kya and 16 kya, reflecting a complex history of gene flow among diverging populations and changes in population size associated with domestication. Cenhaps and other long haplotypes provide vivid displays of this ancient diversity.

category: genomics


title: Human Group Size Puzzle: Why It Is Odd That We Live in Large Societies

abstract: Human groups tend to be much larger than those of non-human primates. This is a puzzle. When ecological factors do not limit primate group size, the problem of coordination creates an upper threshold even when cooperation is guaranteed. This paper offers a simple model of group coordination towards behavioural synchrony to spell out the mechanics of group size limits, and thus show why it is odd that humans live in large societies. The findings suggest that many of our species evolved social behaviours and culturally-maintained social technologies emerged as a solution to this problem.


title: Structural basis for RNA-duplex unwinding by the DEAD-box helicase DbpA

abstract: DEAD-box RNA helicases are implicated in most aspects of RNA biology, where these enzymes unwind short RNA duplexes in an ATP-dependent manner. During the central step of the unwinding cycle, the two domains of the helicase core form a distinct closed conformation that destabilizes the RNA duplex, which ultimately leads to duplex melting. Despite the importance of this step for the unwinding process no high resolution structures of this state are available. Here, we employ nuclear magnetic resonance spectroscopy and X-ray crystallography to determine structures of the DEAD- box helicase DbpA in the closed conformation, complexed with substrate duplexes and single- stranded unwinding product. These structures reveal that DbpA initiates duplex unwinding by interacting with up to three base-paired nucleotides and a 5 single-stranded RNA duplex overhang. These high-resolution snapshots, together with biochemical assays, rationalize the destabilization of the RNA duplex and are integrated into a conclusive model of the unwinding process.

category: biochemistry


title: Biochemical characterization of RAD52-mediated D-loop formation using fluorophore-labeled DNA substrates

abstract: The human RAD52 protein is thought to have multiple roles in the mechanisms of repairing DNA double-strand breaks that are caused by replication errors and reactive oxygen species. One such role is to mediate the formation of a displacement loop (D-loop), which is a critical reaction intermediate in homologous recombinational repair. RAD52 is suggested to promote the formation of D-loops when facilitating DNA synthesis at stalled or collapsed replication forks during mitosis. However, RAD52-mediated D-loop formation remains poorly characterized, and the detailed molecular mechanism of the D-loop formation reaction catalyzed by RAD52 is still unclear. In the present study, we developed a gel-based assay that enables rapid detection of RAD52-mediated D-loop formation. This assay utilizes a fluorophore-labeled, single-stranded DNA substrate. In addition to the rapid detection of D-loops, D-loop extension was observed when DNA polymerase was added to the reaction. This assay can also be used for screening large numbers of compounds that either stimulate or inhibit RAD52-mediated D-loop formation. The D-loop formation assay developed in this study is potentially useful for mechanistic studies of DSB repair involving RAD52-mediated D-loop formation, as well as for screening compounds with potential therapeutic effects.


title: Combining SLA 3D printing and soft lithography for fast, versatile, and accessible high-resolution fabrication of customised multiscale cell culture devices with complex designs

abstract:
Commercially available cell culture devices are designed to increase the complexity of simple cell culture models to provide better experimental platforms for biological systems. From microtopography, microwells, plating devices and microfluidic systems to larger constructs for specific applications like live imaging chamber slides, a wide variety of culture devices with different geometries have become indispensable in biology labs. However, the techniques used for their fabrication can be out of reach for most wet labs due to cost and availability of specialised equipment or the need for engineering expertise. Moreover, these techniques also have technical limitations to the volumes, shapes and dimensions they can generate. For these reasons, creating customisable devices tailored to lab-specific biological questions remains difficult to apply.

Taking advantage of low-cost, high-resolution desktop resin 3D printers combined with PDMS soft-lithography we have developed an optimised microfabrication pipeline capable of generating a wide variety of customisable devices for cell culture and tissue engineering in an easy, fast reproducible way for a fraction of the cost of conventional microfabrication or commercial alternatives. This technique enables the manufacture of complex devices across scales bridging the gap between microfabrication and fused deposition moulding (FDM) printing. The method we describe allows for the efficient treatment of resin-based 3D printed constructs for PDMS curing, using a combination of curing steps, washes and surface treatments. Together with the extensive characterisation of the fabrication pipeline, we provide several proof-of-principle applications ranging from simple 2D culture devices to large tissue engineering constructs and organoid formation systems.

We believe this methodology will be applicable in any wet lab, irrespective of prior expertise or resource availability and will therefore enable a wide adoption of tailored microfabricated devices across many fields of biology.

category: bioengineering


title: An in-silico study of the mutation-associated effects on the spike protein of SARS-CoV-2, Omicron variant

abstract: The emergence of Omicron (B.1.1.529), a new Variant of Concern in the COVID-19 outbreak, while accompanied by the ongoing Delta variant infection, has once again fueled fears of a new infection wave and global health concern. In the Omicron variant, the receptor-binding domain (RBD) of its spike glycoprotein is heavily mutated, a feature critical for the transmission rate of the virus by interacting with hACE2. In this study, we used a combination of conventional and advanced neural network-based in silico approaches to predict how these mutations would affect the spike protein. The results demonstrated a decrease in the electrostatic potentials of residues corresponding to receptor recognition sites, an increase in the alkalinity of the protein, a change in hydrophobicity, variations in functional residues, and an increase in the percentage of alpha-helix structure. Our next step was to predict the structural changes of the spike protein using the AI-based tool Alphafold2 and dock it with hACE2. The results revealed that the RBD of the Omicron variant had a higher affinity than the reference. Moreover, all-atom molecular dynamics simulations concluded that the RBD of the Omicron variant exhibits a more dispersed interaction network since mutations resulted in an increased number of hydrophobic interactions and hydrogen bonds with hACE2 compared to the reference RBD. In summary, our current study highlighted the potential structural basis for the enhanced transmissibility and pathogenicity of the Omicron variant, although further research is needed to investigate its epidemiological and biological implications.

category: bioinformatics


title: MetaboListem and TABoLiSTM: Two Deep Learning Algorithms for Metabolite Named Entity Recognition

abstract:
Reviewing the metabolomics literature is becoming increasingly difficult because of the rapid expansion of relevant journal literature. Text-mining technologies are therefore needed to facilitate more efficient literature review. Here we contribute a standardised corpus of full-text publications from metabolomics studies and describe the development of two new metabolite named entity recognition (NER) methods. We introduce two deep learning methods for metabolite NER based on Bidirectional Long Short-Term Memory (BiLSTM) networks incorporating different transfer learning techniques. Our first model (MetaboListem) follows prior methodology using GloVe word embeddings. Our second model exploits BERT and BioBERT for embedding and is named TABoLiSTM (Transformer-Affixed BiLSTM). The methods are trained on a novel corpus annotated using rule-based methods, and evaluated on manually annotated metabolomics articles. MetaboListem (F1 score 0.890, precision 0.892, recall 0.888) and TABoLiSTM (BioBERT version: F1 score 0.909, precision 0.926, recall 0.893) have achieved state-of-the-art performance on metabolite NER. A corpus with >1,200 full-text Open Access metabolomics publications and >116,000 annotated metabolites was created. This work demonstrates that deep learning algorithms are capable of identifying metabolite names accurately and efficiently in text. The proposed corpus and NER algorithms can be used for metabolomics text-mining tasks such as information retrieval, document classification and literature-based discovery.

AvailabilityThe corpus and NER algorithms are freely available with detailed instructions from Github at https://github.com/omicsNLP/MetaboliteNER.


title: Multiscale Modeling of Hepatitis B Virus Capsid Assembly and its Dimorphism

abstract: Hepatitis B Virus (HBV) is an endemic, chronic virus that leads to 800,000 deaths per year. Central to the HBV lifecycle, the viral core has a protein capsid assembled from many copies of a single protein. The capsid protein adopts different (quasi-equivalent) conformations to form icosahedral capsids containing 180 or 240 proteins, T=3 or T=4 respectively in Caspar-Klug nomenclature. HBV capsid assembly has become an important target for new antivirals; nonetheless the assembly pathways and mechanisms that control HBV dimorphism remain unclear. We describe computer simulations of HBV assembly, using a coarse-grained model that has parameters learned from all-atom molecular dynamics simulations of a complete HBV capsid, and yet is computationally tractable. Dynamical simulations with the resulting model reproduce experimental observations of HBV assembly pathways and products. By constructing Markov state models and employing transition path theory, we identify pathways leading to T=3, T=4, and other experimentally observed capsid morphologies. The analysis identifies factors that control this polymorphism, in particular, the conformational free energy landscape of the capsid proteins and their interactions.

category: biophysics


title: The structure of insulin granule core determines secretory capacity being reduced in type-2 diabetes

abstract: Exocytosis in excitable cells is essential for their physiological functions. Although the exocytotic machinery controlling cellular secretion has been well investigated, the function of the vesicular cargo, i.e. secretory granular content remains obscure. Here we combine dSTORM imaging and single-domain insulin antibody, to dissect the in situ structure of insulin granule cores (IGCs) at nano level. We demonstrate that the size and shape of the IGCs can be regulated by the juxta-granular molecules Nucleobindin-2 and Enolase-1, that further contribute to the stimulated insulin secretion. IGCs located at the plasma membrane are larger than those in the cytosol. The IGCs size is decreased by [~]20% after glucose stimulation due to the release of the peripheral part of IGCs through incomplete granule fusion. Importantly, the reduction of the IGCs size is also observed in non-stimulatory pancreatic {beta}-cells from diabetic db/db mice, Akita (Ins2+/-) mice and human Type-2 diabetic donors, in accordance with impaired secretion. These findings overall highlight the structure of exocytotic insulin cores as a novel modality amenable to targeting in the stimulated exocytosis in {beta}-cells with impaired insulin secretion.


title: A model of colour appearance based on efficient coding of natural images

abstract: An objects colour, brightness and pattern are all influenced by its surroundings, and a number of visual phenomena and "illusions" have been discovered that highlight these often dramatic effects. Explanations for these phenomena range from low-level neural mechanisms to high-level processes that incorporate contextual information or prior knowledge. Importantly, few of these phenomena can currently be accounted for when measuring an objects perceived colour. Here we ask to what extent colour appearance is predicted by a model based on the principle of coding efficiency. The model assumes that the image is encoded by noisy spatio-chromatic filters at one octave separations, which are either circularly symmetrical or oriented. Each spatial bands lower threshold is set by the contrast sensitivity function, and the dynamic range of the band is a fixed multiple of this threshold, above which the response saturates. Filter outputs are then reweighted to give equal power in each channel for natural images. We demonstrate that the model fits human behavioural performance in psychophysics experiments, and also primate retinal ganglion responses. Next we systematically test the models ability to qualitatively predict over 35 brightness and colour phenomena, with almost complete success. This implies that contrary to high-level processing explanations, much of colour appearance is potentially attributable to simple mechanisms evolved for efficient coding of natural images, and is a basis for modelling the vision of humans and other animals.


title: Patterns of connectome variability in autism across five functional activation tasks. Fndings from the LEAP project

abstract:
BackgroundAutism spectrum disorder (autism) is a complex neurodevelopmental condition with pronounced behavioural, cognitive, and neural heterogeneities across individuals. Here, our goal was to characterise heterogeneity in autism by identifying patterns of neural diversity as reflected in BOLD fMRI in the way individuals with autism engage with a varied array of cognitive tasks.

MethodsAll analyses were based on the EU-AIMS/AIMS-2-TRIALS multisite Longitudinal European Autism Project (LEAP) with participants with autism and typically developing controls (TD) between 6 and 30 years of age. We employed a novel task-potency approach which combines the unique aspects of both resting-state fMRI and task-fMRI to quantify task-induced variations in the functional connectome. Normative modelling was used to map atypicality of features on an individual basis with respect to their distribution in neurotypical control participants. We applied robust out-of-sample canonical correlation analysis (CCA) to relate connectome data to behavioural data.

ResultsDeviation from the normative ranges of global functional connectivity was greater for individuals with autism compared to TD in each fMRI task paradigm (all tasks p<0.001). The similarity across individuals of the deviation pattern was significantly increased in autistic relative to TD individuals (p<0.002). The CCA identified significant and robust brainbehavior covariation between functional connectivity atypicality and autism-related behavioral features.

ConclusionsIndividuals with autism engage with tasks in a globally atypical way, but the particular spatial pattern of this atypicality is nevertheless similar across tasks. Atypicalities in the tasks originate mostly from prefrontal cortex and default mode network regions, but also speech and auditory networks. We show, moving forward, sophisticated modeling methods such as task-potency and normative modeling will prove key to unravelling complex heterogeneous conditions like autism.


title: Patterns of connectome variability in autism across five functional activation tasks. Findings from the LEAP project


title: Efficacy and sex differences in the effects on rat brain microglia of the colony-stimulating factor 1 receptor inhibitor-PLX5622

abstract: Microglia play pivotal roles in central nervous system development, homeostasis, responses to trauma, neurodegenerative and neuropsychiatric disorders with significant sex-bias in their symptoms and prevalence. The discovery that the survival of microglia in adult brains depends on the expression of the colony-stimulating factor 1 receptor (CSF1R), along with the development of the effective brain permeant CSF1R inhibitors PLX5622, has boosted the investigation of the role of microglia in health, disease and in relations to sex-bias. The effectiveness of PLX5622 in examining the role of microglia has mainly been demonstrated in mice. Surprisingly, despite the critical importance of rat models in brain research, there are only 4 publications in which PLX5622 was used to investigate the roles of microglia in adult rats. This has been attributed to the "impression" that PLX5622 is "ineffective in rats". In view of the importance and interest in the role of microglia, the indispensability of rats for in vivo electrophysiological brain studies and behavioral research and the high efficacy of PLX5622-chow in eliminating microglia from adult mice brains, we examined the effects of PLX5622-chow on the elimination of the microglia in adult female and male rats. We found significant differences in microglia elimination by ad libitum PLX5622 feeding in male and female rats in different brain regions with significantly greater effectivity in female brains. Our pragmatic study provides practical information on the use and design of PLX5622 in gender-related and rat brain preclinical microglia research.


title: A Neuropsin-based Optogenetic Tool for Precise Control of Gq signaling

abstract: Gq-coupled receptors regulate numerous physiological processes by activating enzymes and inducing intracellular Ca2+ signals. There is a strong need for an optogenetic tool that enables powerful experimental control over Gq signaling. Here, we present chicken opsin 5 (cOpn5) as the long sought-after, single-component optogenetic tool that mediates ultra-sensitive optical control of intracellular Gq signaling with high temporal and spatial resolution. Expressing cOpn5 in mammalian cells enables blue light-triggered, Gq-dependent Ca2+ release from intracellular stores and protein kinase C activation. Strong Ca2+ transients were evoked by brief light pulses of merely 10 ms duration and at 3 orders lower light intensity of that for common optogenetic tools. Photostimulation of cOpn5-expressing cells at the subcellular and single-cell levels generated intracellular and intercellular Ca2+ wave propagation, respectively, thus demonstrating the high spatial precision of cOpn5 optogenetics. The cOpn5-mediated optogenetics could also be applied to activate neurons and control animal behavior in a circuit-dependent manner. We further revealed that optogenetic activation of cOpn5-expressing astrocytes induced massive ATP release and modulation neuronal activation in the brain of awake, behaving mice. cOpn5 optogenetics may find broad applications in studying the mechanisms and functional relevance of Gq signaling in both non-excitable cells and excitable cells in all major organ systems.


title: Chronic neurotransmission increases the susceptibility of lateral-line hair cells to ototoxic insults

abstract: Sensory hair cells receive near constant stimulation by omnipresent auditory and vestibular stimuli. To detect and encode these stimuli, hair cells require steady ATP production, which can be accompanied by a buildup of mitochondrial byproducts called reactive oxygen species (ROS). ROS buildup is thought to sensitize hair cells to ototoxic insults, including the antibiotic neomycin. Work in neurons has shown that neurotransmission is a major driver of ATP production and ROS buildup. Therefore, we tested whether neurotransmission is a significant contributor to ROS buildup in hair cells. Using genetics and pharmacology, we disrupted two key aspects of neurotransmission in zebrafish hair cells: presynaptic calcium influx and the fusion of synaptic vesicles. We find that chronic block of neurotransmission enhances hair-cell survival when challenged with the ototoxin neomycin. This reduction in ototoxin susceptibility is accompanied by reduced mitochondrial activity, likely due to a reduced ATP demand. In addition, we show that mitochondrial oxidation and ROS buildup is reduced when neurotransmission is blocked. Mechanistically, we find that it is the synaptic vesicle cycle rather than presynaptic- or mitochondrial-calcium influx that contributes most significantly to this metabolic stress. Our results comprehensively indicate that, over time, neurotransmission causes ROS buildup that increases the susceptibility of hair cells to ototoxins.


title: Sensory-motor computations in a cortico-pontine pathway

abstract: Subcortical circuits decode the output from multiple cortical areas to create commands for movement. By recording during monkeys smooth pursuit eye movements, we reveal how the dorsolateral pontine nucleus (DLPN) and nucleus reticularis tegmenti pontis (NRTP) decode cortical output. Preparatory activity is inherited from the smooth eye movement region of the frontal eye fields (FEFSEM). Consistent with its effects on eye movement responses to visual motion, preparatory activity modulates the gain of pontine responses to visual motion input from visual area MT. Furthermore, the "place" code for target speed in area MT becomes the motor commands "rate" code in the pons, through linear weighting of MTs output. The division into decoding that happens in the pons and residual computations that occur downstream reveals that the pons is neither a relay nor the source of a final motor command. It is part of a complex, distributed cortical decoder for motor control.


title: RNA-based vaccine demonstrates prophylactic efficacy against Mycobacterium tuberculosis challenge in a mouse model

abstract: Mycobacterium tuberculosis (Mtb) is an opportunistic bacterial pathogen that causes tuberculosis disease (TB) and exerts an extensive burden on global health. The complex intra- and extracellular nature of this bacterium, coupled with different disease stages have made mechanistic studies evaluating the contributions of innate and adaptive host immunity challenging. In this work we leveraged two delivery platforms as prophylactic vaccines to assess immunity and subsequent efficacy against low dose and ultra-low dose aerosol challenge with Mtb H37Rv in C57BL/6 mice. Our proof-of-concept TB vaccine candidate ID91 was produced as a fusion protein formulated with a synthetic TLR4 agonist (glucopyranosyl lipid adjuvant in a stable emulsion) or as a replicating-RNA (repRNA) formulated in a nanostructured lipid carrier (NLC). Results from this work demonstrate that protein subunit- and RNA-based vaccines preferentially elicit cellular immune responses to different ID91 epitopes. In a single prophylactic immunization screen, both platforms reduced pulmonary bacterial burden compared to controls. Excitingly, in prime-boost strategies, groups that received heterologous RNA-prime, protein-boost or combination (simultaneous in different sites) immunizations demonstrated the greatest reduction in bacterial burden and a unique humoral and cellular immune response profile. These data are the first to report that repRNA platforms are a viable system for TB vaccines and should be pursued with high priority Mtb antigens containing CD4+ and CD8+ T cell epitopes.

category: immunology


title: SARS-CoV-2 Variant Delta Potently Suppresses Innate Immune Response and Evades Interferon-Activated Antiviral Responses

abstract: Delta variant of SARS-CoV-2 has caused more severe infections than its previous variants. We studied the host innate immune response to Delta, Alpha and two earlier variants to map the evolution of the recent ones. Our biochemical and transcriptomic studies reveal that Alpha and Delta have progressively evolved over the ancestral variants by silencing innate immune response, thereby limiting cytokine and chemokine production. Though Alpha silenced RLR pathway just as Delta, it failed to persistently silence the innate immune response unlike Delta. Both Alpha and Delta have evolved to resist IFN treatment while they are still susceptible to RLR activation, further highlighting the importance of RLR-mediated, IFN-independent mechanisms in restricting SARS-CoV-2. Our studies reveal that SARS-CoV-2 Delta has integrated multiple mechanisms to silence host innate immune response and evade IFN response. Deltas silent replication and sustained suppression of host innate immune response, possibly resulting in delayed or reduced intervention by the adaptive immune response, could potentially contribute to the severe symptoms and poor recovery index associated with it.


title: Mucosal Immunization of Cynomolgus Macaques with Adenoviral Vector Vaccine Elicits Neutralizing Nasal and Serum Antibody to Several SARS-CoV-2 Variants

abstract: The emergence of SARS-CoV-2 variants continues to be a major obstacle for controlling the global pandemic. Despite the currently authorized SARS-CoV-2 vaccines ability to reduce severe disease and hospitalization, new immunization strategies are needed that enhance mucosal immune responses, inhibit community transmission, and provide protection against emerging variants. We have developed a mucosally delivered, non-replicating recombinant adenovirus vector (rAd5) vaccine, that has proven efficacy in the clinic against other respiratory viruses [1]. Here we evaluated the immunogenicity of three candidate SARS-CoV-2 vaccines in cynomolgus macaques that contained spike (S) and/or nucleocapsid (N) from either the Wuhan or the beta variant to select a candidate for future clinical development. Mucosal immunization with the Wuhan specific S vaccine (ED90) induced significant cross-reactive serum IgG responses against to Wuhan, beta, gamma and delta lineages, and generated substantial serum neutralizing activity. In nasal samples, ED90 immunization induced 1000-fold increases in IgA to all variants of concern tested and had neutralizing activity against Wuhan and delta. While immunization with the beta specific vaccine (ED94) enhanced IgG and IgA responses to homologous beta variant S and RBD, this approach resulted in less cross-reactive responses to other variants in the serum and nasal passages compared to ED90. As ED90 immunization induced the most robust cross-reactive systemic and mucosal antibody responses, this candidate was chosen for future clinical development.


title: A single-dose of the deactivated rabies virus vectored COVID-19 vaccine, CORAVAX, is highly efficacious and alleviates lung inflammation in the hamster model.

abstract: Without sufficient herd immunity through either vaccination or natural infection, the Coronavirus Disease 2019 pandemic is unlikely to be controlled. Waning immunity with the currently approved vaccines suggests the need to evaluate vaccines causing the induction of long-term responses. Here we report the immunogenicity and efficacy of our adjuvanted single-dose Rabies vectored SARS CoV-2 S1 vaccine, CORAVAX, in hamsters. CORAVAX induces high SARS CoV-2 S1 specific and virus-neutralizing antibodies (VNA) that prevent weight loss, viral loads, disease, lung inflammation, and the cytokine storm in hamsters. We also observed high Rabies VNA titers. In summary, CORAVAX is a promising dual antigen vaccine candidate for clinical evaluation against SARS CoV-2 and Rabies virus.


title: Novel and Reported Compensatory Mutations in rpoABC Associate Specifically with Predominant Mycobacterium tuberculosis Rifampicin Resistance Marker rpoB:S450L

abstract:
BackgroundRifampicin (RIF) is a key first-line drug used to treat tuberculosis, a pulmonary disease caused by Mycobacterium tuberculosis. However antibiotic resistance to RIF is prevalent despite an apparent fitness cost. RIF resistance is primarily caused by mutations in the RIF resistance determining region in the rpoB gene, at the cost of slower growth in rich media. Compensatory mutations in the genes rpoA and rpoC have been shown to alleviate this fitness cost. These compensatory mutations may explain how RIF resistant strains have spread so rapidly. However, the effect of compensation on transmission is still unclear, partly because of uncertainty over which rpoABC mutations compensate for which RIF resistance markers.

ObjectivesWe performed an association study on a globally representative set of 4309 whole genome sequenced clinical M. tuberculosis isolates to identify novel putative compensatory mutations, determine the prevalence of known and previously reported putative compensatory mutations, and determine which RIF resistance markers associate with these compensatory mutations.

Results and ConclusionsOnly 20.0% (216/1079) of RIF resistant isolates carried previously reported high-probability compensatory mutations, suggesting existence of other compensatory mutations. Using a strict phylogenetic approach, we identified 18 novel putative compensatory mutations in rpoC, rpoB, and rpoA. Novel and previously reported compensatory mutations were strongly associated with the RIFR marker rpoB:S450L, suggesting compensation may be specific to particular RIFR markers. These findings will aid identification of RIF-resistant M. tuberculosis strains with restored fitness. Such strains pose a greater risk of causing resistant outbreaks.


title: Characterization of various remdesivir-resistant mutations of SARS-CoV-2 by mathematical modeling and molecular dynamics simulation.

abstract:
Mutations continue to accumulate within the SARS-CoV-2 genome, and the ongoing epidemic has shown no signs of ending. It is critical to predict problematic mutations that may arise in clinical environments and assess their properties in advance to quickly implement countermeasures against future variant infections. In this study, we identified mutations resistant to remdesivir, which is widely administered to SARS-CoV-2-infected patients, and discuss the cause of resistance. First, we simultaneously constructed eight recombinant viruses carrying the mutations detected in in vitro serial passages of SARS-CoV-2 in the presence of remdesivir. Time course analyses of cellular virus infections showed significantly higher infectious titers and infection rates in mutant viruses than wild type virus under treatment with remdesivir. Next, we developed a mathematical model in consideration of the changing dynamic of cells infected with mutant viruses with distinct propagation properties and defined that mutations detected in in vitro passages canceled the antiviral activities of remdesivir without raising virus production capacity. Finally, molecular dynamics simulations of the NSP12 protein of SARS-CoV-2 revealed that the molecular vibration around the RNA-binding site was increased by the introduction of mutations on NSP12. Taken together, we identified multiple mutations that affected the flexibility of the RNA binding site and decreased the antiviral activity of remdesivir. Our new insights will contribute to developing further antiviral measures against SARS-CoV-2 infection.

Significance StatementConsidering the emerging Omicron strain, quick characterization of SARS-CoV-2 mutations is important. However, owing to the difficulties in genetically modifying SARS-CoV-2, limited groups have produced multiple mutant viruses. Our cutting-edge reverse genetics technique enabled construction of eight reporter-carrying mutant SARS-CoV-2 in this study. We developed a mathematical model taking into account sequential changes and identified antiviral effects against mutant viruses with differing propagation capacities and lethal effects on cells. In addition to identifying the positions of mutations, we analyzed the structural changes in SARS-CoV-2 NSP12 by computer simulation to understand the mechanism of resistance. This multidisciplinary approach promotes the evaluation of future resistance mutations.


title: Prevalence Of Multi Drug Resistant Bacteria On Environmental And Medical Device Surfaces Of Korea Nepal Friendship Hospital

abstract:
The microbial monitoring of environmental and medical-devices surface is used to evaluate efficacy of routine cleaning and disinfection practises and to detect the presence of specific Nosocomial Pathogens. The prevalence of Multidrug Resistance organisms in hospital premises projects serious problems in transmitting to susceptible hosts which is difficult to treat. A cross sectional descriptive research was conducted from December 2016 to June 2017 at the pathology laboratory of Korea Nepal Friendship Hospital (KNFH). A total 140 samples were considered, encompassing the medical devices of the hospital (100), housekeeping surfaces (15) and air (25). Susceptibility test for bacterial isolates was done by disk diffusion assay. Of the total 140 samples taken and analysed, 100% showed growth positivity. In most of the swabs taken, Coagulase Negative Staphylococci was dominant, followed by Staphylococcus aureus, Streptococcus spp. Micrococcus spp., E coli, Pseudomonas spp., Bacillus spp., Acinetobacter spp., Klebsiella spp., Fungi, and least Proteus spp. The dry surfaces were dominantly contaminated by gram positive bacteria whereas moistened surfaces like wash basin were contaminated by gram negative as well as gram positive bacteria. Total 277 strains were exposed to various class of antibiotics, among the gram positive environmental isolates, Coagulase Negative Staphylococci 16 (34.78%) had highest MDR prevalence followed by Staphylococcus aureus 8 (29.62%), Streptococcus spp. 4 (12.90%), Micrococcus spp. 4 (9.30%) and no MDR was shown by any Bacillus spp isolates. Whereas, in case of gram negative, Klebsiella spp. 6 (35.29%) had highest MDR prevalence

followed by Acinetobacter spp. 6 (31.57%), E. coli 8 (27.58%), Pseudomonas spp. 4 (18.18%), and lastly Proteus spp. with no MDR at all. The thick dirt covering the cotton swabs and heavy microbial load on them has displayed not only disinfecting practice but also cleaning practice is missing. Heavy contamination shows possible NIs breakout, its important to have routine microbial assessment with standard protocol and find ways to decrease its load.


title: Tracking influenza a virus infection in the lung from hematological data with machine learning

abstract: The tracking of pathogen burden and host responses with minimal-invasive methods during respiratory infections is central for monitoring disease development and guiding treatment decisions. Utilizing a standardized murine model of respiratory Influenza A virus (IAV) infection, we developed and tested different supervised machine learning models to predict viral burden and immune response markers, i.e. cytokines and leukocytes in the lung, from hematological data. We performed independently in vivo infection experiments to acquire extensive data for training and testing purposes of the models. We show here that lung viral load, neutrophil counts, cytokines like IFN-{gamma} and IL-6, and other lung infection markers can be predicted from hematological data. Furthermore, feature analysis of the models shows that blood granulocytes and platelets play a crucial role in prediction and are highly involved in the immune response against IAV. The proposed in silico tools pave the path towards improved tracking and monitoring of influenza infections and possibly other respiratory infections based on minimal-invasively obtained hematological parameters.


title: Serological screening in wild ruminants in Germany, 2021/22: No evidence of SARS-CoV-2, bluetongue virus or pestivirus spread but high seroprevalences against Schmallenberg virus

abstract: Wildlife animals may be susceptible for multiple infectious agents of public health or veterinary relevance, thereby potentially forming a reservoir that bears the constant risk of re-introduction into the human or livestock population. Here, we serologically investigated 493 wild ruminant samples collected in the 2021/22 hunting season in Germany for the presence of antibodies against the severe acute respiratory coronavirus 2 (SARS-CoV-2) and four viruses pathogenic for domestic ruminants, namely the orthobunyavirus Schmallenberg virus (SBV), the reovirus bluetongue virus (BTV) and ruminant pestiviruses like bovine viral diarrhoea virus or border disease virus. The animal species comprised fallow deer, red deer, roe deer, mouflon and wisent. For coronavirus serology, additional 307 fallow, roe and red deer samples collected between 2017 and 2020 at three military training areas were included. While antibodies against SBV could be detected in about 13.6% of the samples collected in 2021/22, only one fallow deer of unknown age tested positive for anti-BTV antibodies and all samples reacted negative for antibodies against ruminant pestiviruses. In an ELISA based on the receptor-binding domain (RBD) of SARS-CoV-2, 25 out of 493 (5.1%) samples collected in autumn and winter 2021/22 scored positive. This sero-reactivity could not be confirmed by the highly specific virus neutralization test, occurred also in 2017, 2018 and 2019, i.e. prior to the human SARS-CoV-2 pandemic, and was likewise observed against the RBD of the related SARS-CoV-1. Therefore, the SARS-CoV-2-seroreactivity was most likely induced by another, hitherto unknown deer virus belonging to the subgenus Sarbecovirus of betacoronaviruses.


title: Resistance of Dickeya solani strain IPO 2222 to lytic bacteriophage vB_Dsol_D5 ({Phi}D5) results in fitness tradeoffs for the bacterium during infection

abstract: Resistance to bacteriophage infections protects bacteria in phage-full environments, allowing them to survive and multiply in the presence of their viral predators. However, such resistance may cause direct costs for strains linked with the ecological fitness expressed as reduced competitiveness for resources or reduced virulence or both. Unfortunately, limited knowledge exists about such costs paid by phage-resistant plant pathogenic bacteria in their natural environments. This study analyzed the costs of phage resistance paid by broad host phytopathogenic pectinolytic bacterium Dickeya solani both in vitro and in potato (Solanum tuberosum L.) plants. Thirteen D. solani IPO 2222 Tn5 mutants were identified that exhibited resistance to infection caused by lytic bacteriophage vB_Dsol_D5 ({Phi}D5). The genes disrupted in these 13 mutants encoded proteins involved in the synthesis of the bacterial envelope components (viz. LPS, EPS and capsule). The ability of {Phi}D5-resistant D. solani mutants to colonize and cause symptoms on potato plants as well as other phenotypes that are known to contribute to the ecological fitness of D. solani in-plant environment, including growth rate, production of effectors, swimming and swarming motility, use of various carbon and nitrogen sources and biofilm formation were assessed. Although phage resistance did not affect most of the phenotypes of {Phi}D5-resistant D. solani evaluated in vitro, all phage resistant mutants were significantly compromised in their ability to survive on and colonize and cause disease symptoms in potato plants. This study is, to our knowledge, one of few to show the direct link between phage resistance and the fitness of plant pathogenic bacteria and the first one to assess phage-host associations for D. solani.


title: Potent Neutralizing Activity of Polyclonal Equine Antibodies against Omicron SARS-CoV-2

abstract: Using a polyclonal approach of equine anti-SARS-CoV-2 F(ab)2 antibodies we have achieved a high level of neutralizing potency against all SARS-CoV-2 variants tested. Neutralization titers were in the range of 105-106 IU/mL including Omicron: 111,403 UI/mL, which is 2-3 orders of magnitude what is normally achieved in response to SARS-CoV-2 infection and/or vaccination. The presence of high titers of a repertoire of antibodies targeting conserved epitopes in different regions of the spike protein could plausibly account for this remarkable breadth of neutralization. These results warrant the clinical investigation of anti-SARS-CoV-2 equine polyclonal F(ab)2 antibodies as a novel therapeutic strategy against COVID-19


title: Transertion is used for localized expression and assembly of Vibrio parahaemolyticus T3SS2

abstract:
Using environmental cues, bacteria commit to the assembly of transmembrane complexes such as the type III secretion system 2 (T3SS2), a membrane-bound, syringe-like secretory apparatus used during infection to inject host cells with virulence factors. Here we report Vibrio parahaemolyticus uses transertion, localized transcription, translation, and membrane insertion, to assemble its T3SS2. Upon binding bile acids, the membrane bound receptor and transcription factor VtrA/VtrC captures the T3SS2 pathogenicity island at the inner membrane. Activated VtrA/VtrC induces production of VtrB, the membrane bound master T3SS2 transcriptional regulator. VtrB then induces the membrane-proximal T3SS2 genes to undergo transertion for assembly of the membrane inserted secretion machinery. Transertion is a process that can be used for the efficient assembly of membrane-bound molecular complexes in response to extracellular signals.

One-Sentence SummaryLocalized transcription, translation, and membrane insertion of multi-protein complexes in bacteria in response to host cues.


title: BET-independent MLV integration is retargeted in vivo and selects distinct genomic elements for lymphomagenesis

abstract: Moloney murine leukemia virus (MLV) infects BALB/c mice and induces T-cell lymphoma in mice. Retroviral integration is mediated by the interaction of the MLV integrase (IN) with members of the bromodomain and extra-terminal motif (BET) protein family (BRD2, BRD3 and BRD4). Introduction of the W390A mutation in MLV IN abolishes BET interaction. Here we compared the replication of W390A MLV and WT MLV in adult BALB/c mice to study the role of BET proteins in replication, integration and tumorigenesis in vivo. Comparing WT and W390A MLV infection revealed similar viral loads in blood, thymus and spleen cells. Interestingly, W390A MLV integration was retargeted away from GC-enriched genomic regions. However, both WT MLV and W390A MLV developed T cell lymphoma after a similar latency represented by an enlarged thymus and spleen and multi-organ tumor infiltration. Integration site sequencing from splenic tumor cells revealed clonal expansion in all WT MLV- and W390A MLV-infected mice. However, the integration profile of W390A MLV and WT MLV differed significantly. Integrations were enriched in enhancers and promoters but compared to WT, W390A MLV integrated less frequently into enhancers and more into oncogene bodies, such as Notch1 and Ppp1r16b. We conclude that host factors direct MLV in vivo integration site selection. Although, BET proteins target WT MLV integration preferentially towards enhancers and promoters, insertional lymphomagenesis can occur independently from BET, likely due to the intrinsically strong enhancer/promoter of the MLV LTR.


title: Discovery and functional interrogation of SARS-CoV-2 protein-RNA interactions

abstract: The COVID-19 pandemic is caused by severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2). The betacoronvirus has a positive sense RNA genome which encodes for several RNA binding proteins. Here, we use enhanced crosslinking and immunoprecipitation to investigate SARS-CoV-2 protein interactions with viral and host RNAs in authentic virus-infected cells. SARS-CoV-2 proteins, NSP8, NSP12, and nucleocapsid display distinct preferences to specific regions in the RNA viral genome, providing evidence for their shared and separate roles in replication, transcription, and viral packaging. SARS-CoV-2 proteins expressed in human lung epithelial cells bind to 4773 unique host coding RNAs. Nine SARS-CoV-2 proteins upregulate target gene expression, including NSP12 and ORF9c, whose RNA substrates are associated with pathways in protein N-linked glycosylation ER processing and mitochondrial processes. Furthermore, siRNA knockdown of host genes targeted by viral proteins in human lung organoid cells identify potential antiviral host targets across different SARS-CoV-2 variants. Conversely, NSP9 inhibits host gene expression by blocking mRNA export and dampens cytokine productions, including interleukin-1/{beta}. Our viral protein-RNA interactome provides a catalog of potential therapeutic targets and offers insight into the etiology of COVID-19 as a safeguard against future pandemics.

category: molecular biology


title: Expression of Unfolded Protein Response Genes in Post-transplantation Liver Biopsies

abstract:
Cholestatic liver diseases are a major source of morbidity and mortality that can progress to end-stage liver disease. There are few effective medical therapies for primary biliary cholangitis, primary sclerosing cholangitis and other cholestatic liver diseases, in part, due to our incomplete understanding of the pathogenesis of cholestatic liver injury. The hepatic unfolded protein response (UPR) is an adaptive cellular response to endoplasmic reticulum stress that is important in the pathogenesis of many liver diseases and recent animal studies have demonstrated the importance of the UPR in the pathogenesis of cholestatic liver injury. However, the role of the UPR in human cholestatic liver diseases is largely unknown. In this study, we utilized liver biopsies from patients after liver transplantation as a disease model to determine the transcriptional profile and hepatic UPR gene expression that is associated with liver injury and cholestasis. RNA-seq analysis revealed that patients with hyperbilirubinemia had enhanced expression of hepatic UPR pathways. Alternatively, liver biopsy samples from patients with acute rejection had enhanced gene expression of LAG3 and CDK1. Pearson correlation analysis of serum alanine aminotransferase, aspartate aminotransferase and total bilirubin levels demonstrated significant correlations with the hepatic expression of several UPR genes, as well as genes involved in hepatic bile acid metabolism and inflammation. In contrast, serum alkaline phosphatase levels were correlated with the level of hepatic bile acid metabolism gene expression but not liver UPR gene expression.

ConclusionOverall, these data indicate that hepatic UPR pathways are increased in cholestatic human liver biopsy samples and supports an important role of the UPR in the mechanism of human cholestatic liver injury.


title: Toxoplasma gondii-s basal complex: the other apicomplexan business end is multifunctional

abstract: The Apicomplexa are famously named for their apical complex, a constellation of organelles at their apical end dedicated to invasion of their host cells. In contrast, at the other end of the cell, the basal complex (BC) has been in the limelight since it is much less prominent and specific functions were not immediately obvious. However, in the past decade a staggering array of functions have been associated with the BC, and strides have been made in understanding its structure. Here, these collective insights are supplemented with new data to provide an overview of the understanding of the BC in Toxoplasma gondii. The emerging picture is that the BC is a dynamic and multifunctional complex, with a series of (putative) functions. The BC has multiple roles in cell division: it is the site where building blocks are added to the cytoskeleton scaffold; it exerts a two-step stretch and constriction mechanism as contractile ring; and it is key in organelle division. Furthermore, the BC has numerous putative roles in  import, such as the recycling of mother cell remnants, the acquisition of host-derived vesicles, possibly the uptake of lipids derived from the extracellular medium, and the endocytosis of micronemal proteins. The latter process ties the BC to motility, whereas an additional role in motility is conferred by Myosin C. Furthermore, the BC acts on the assembly and/or function of the intravacuolar network, which may directly or indirectly contribute to the establishment of chronic tissue cysts. Here we provide experimental support for molecules acting in several of these processes, and identify several new BC proteins critical to maintaining the cytoplasmic bridge between divided parasites. However, the dispensable nature of many BC components leaves many questions unanswered regarding its function. In conclusion, the BC in T. gondii is a dynamic and multifunctional structure at the posterior end of the parasite.


title: A light-gated transcriptional recorder for detecting cell-cell contacts

abstract: Technologies for detecting cell-cell contacts are powerful tools for studying a wide range of biological processes, from neuronal signaling to cancer-immune interactions within the tumor microenvironment. Here we report TRACC, a GPCR-based transcriptional recorder of cellular contacts, which converts contact events into stable transgene expression. TRACC is derived from our previous protein-protein interaction recorders, SPARK (Kim et al., 2017) and SPARK2 (Kim et al., 2019). TRACC incorporates light gating via the LOV domain, which provides user-defined temporal control of tool activation and reduces background. We show that TRACC detects cell-cell contacts with high specificity and sensitivity in mammalian cell culture and that it can be used to interrogate interactions between neurons and glioma, a form of brain cancer.


title: Maternal and/or direct supplementation with a combination of a casein hydrolysate and yeast β-glucan on post-weaning performance and intestinal health in the pig

abstract: A 2 x 2 factorial experiment was conducted to investigate the effect of maternal supplementation from day 83 of gestation and/or direct supplementation from weaning of a bovine casein hydrolysate plus a yeast {beta}-glucan (CH-YBG) on pig performance and intestinal health on day ten post-weaning. Twenty cross bred gilts (Large White x Landrace) were randomly assigned to one of two dietary groups (n = 10 gilts/group): basal diet (basal sows) and basal diet supplemented with CH-YBG (supplemented sows) from day 83 of gestation until weaning (2g/sow/day). At weaning, 120 pigs (6 pigs/sow) were selected. The two dam groups were further divided, resulting in four experimental groups (10 replicates/group; 3 pigs/pen) as follows: 1) BB (basal sows + basal pigs); 2) BS (basal sows + supplemented pigs); 3) SB (supplemented sows + basal pigs); 4) SS (supplemented sows + supplemented pigs). Supplemented pigs were offered 0.5g CH-YBG/kg of feed for 10 days post-weaning. On day 10 post-weaning, 1 pig/pen was humanely sacrificed and samples were taken from the gastrointestinal tract for analysis. Pigs weaned from supplemented sows (SS, SB) had reduced faecal scores and incidence of diarrhoea (P<0.05) compared to pigs weaned from basal sows (BB, BS), with SS pigs not displaying the transient rise in faecal scores seen in the other three groups from day 3 to day 10 post-weaning (P<0.05). Pigs weaned from supplemented sows had reduced feed intake (P<0.05), improved feed efficiency (P<0.05), increased butyrate concentrations (P<0.05), increased abundance of Lactobacillus (P<0.05) and decreased abundance of Enterobacteriaceae and Campylobacteraceae (P<0.05) compared to pigs weaned from basal sows. In conclusion, a combination of both maternal and direct supplementation with CH-YBG is the desirable feeding strategy to ameliorate the post-weaning challenge in pig production systems.


title: Coursing hyenas and stalking lions: the potential for inter- and intraspecific interactions

abstract: Resource partitioning promotes coexistence among guild members, and carnivores reduce interference competition through behavioural mechanisms that promote spatio-temporal separation. We analyzed sympatric lion and spotted hyena movements and activity patterns to ascertain the mechanisms facilitating their coexistence within semi-arid and wetland ecosystems. We identified recurrent high-use (revisitation) and extended stay (duration) areas within home ranges, and correlated environmental variables with movement-derived measures of inter- and intraspecific interactions. Spatial overlaps among lions and hyenas occurred at edges of home ranges, around water-points, along pathways between patches of high-use areas, and expanded during the wet season. Lions shared more of their home ranges with spotted hyenas in arid ecosystems, but shared more of their ranges with conspecifics in mesic environments. Despite shared space use, we found evidence for subtle temporal differences in the nocturnal movement and activity patterns between the two predators, suggesting a fine localized-scale avoidance strategy. Revisitation frequency and duration within home ranges were influenced by interspecific interactions, subsequent to land cover categories and diel cycles. Intraspecific interactions were also important for lions and, for hyenas, moon illumination and ungulates attracted to former anthrax carcass sites in Etosha, with distance to water in Chobe/Linyanti. Recursion and duration according to locales of competitor probabilities were similar among female lions and both sexes of hyenas, but different for male lions. Our results suggest that lions and spotted hyenas mediate the potential for interference competition through subtle differences in temporal activity, fine-scale habitat use differentiation, and localized reactive-avoidance behaviours. These findings enhance our understanding of the potential effects of interspecific interactions among large carnivore space-use patterns within an apex predator system, and show adaptability across heterogeneous and homogeneous environments. Future conservation plans should emphasize the importance of inter- and intraspecific competition within large carnivore communities, particularly moderating such effects within increasingly fragmented landscapes.

category: ecology


title: Anatomy of the heart of the leatherback turtle

abstract: Non-crocodylian reptiles have hearts with a single ventricle, which is partially separated by a muscular ridge that provide some separation of blood flows. An exceptional situation exists in monitor lizards and pythons, where the ventricular left side generates a much higher systolic blood pressure than the right side, thus resembling mammals and birds. This functional division of the ventricle depends on a large muscular ridge and may relate to high metabolic demand. The large leatherback turtle (<1000 kg), with its active ocean-going lifestyle and elevated body temperatures, may have similar adaptations. Here, we report on the anatomy the hearts of two leatherback turtles. One stranded in Ballum, Denmark in 2020, and was examined in detail, supplemented by observations and photos of an additional stranding specimen from Canada. The external morphology of the leatherback heart resembles that of other turtles, but it is large. We made morphometric measurements of the Ballum heart and created an interactive 3D model using high resolution MRI. The volume of the ventricle was 950 ml, from a turtle of 300 kg, which is almost twice as large as in other reptiles. The Ballum heart was compared to MRI scans of the hearts of a tortoise, a python, and a monitor lizard. Internally, the leatherback heart is typical of non-crocodylian reptiles, and did not contain the well-developed septation found in pythons and monitor lizards. We conclude that if leatherback turtles have exceptional circulation needs, they are sustained with a relatively large but otherwise typical non-crocodylian reptile heart.


title: Adaptations for bipedal walking: Musculoskeletal structure and three-dimensional joint mechanics of humans and bipedal chimpanzees (Pan troglodytes)

abstract: Humans are unique among apes and other primates in the musculoskeletal design of their lower back, pelvis and lower limbs. Here, we describe the three-dimensional ground reaction forces and lower/hind limb joint mechanics of human and bipedal chimpanzee walking over a full stride and test whether: 1) the estimated limb joint work and power during stance phase, especially the single-support period, is lower in humans than bipedal chimpanzees, 2) the limb joint work and power required for limb swing is lower in humans than in bipedal chimpanzees, and 3) the estimated total mechanical power during walking, accounting for the storage of passive elastic strain energy in humans, is lower in humans than in bipedal chimpanzees. Humans and bipedal chimpanzees were compared at matched dimensionless and dimensional velocities. Our results indicate that humans walk with significantly less work and power output in the first double-support period and the single-support period of stance, but markedly exceed chimpanzees in the second-double support period (i.e., push-off). Humans generate less work and power in limb swing, although the species difference in limb swing power was not statistically significant. We estimated that total mechanical positive  muscle fiber work and power were 46.9% and 35.8% lower, respectively, in humans than bipedal chimpanzees at matched dimensionless speeds. This is due in part to mechanisms for the storage and release of elastic energy at the ankle and hip in humans. Further, these results indicate distinct heel strike and lateral balance mechanics in humans and bipedal chimpanzees, and suggest a greater dissipation of mechanical energy through soft tissue deformations in humans. Together, our results document important differences between human and bipedal chimpanzee walking mechanics over a full stride, permitting a more comprehensive understanding of the mechanics and energetics of chimpanzee bipedalism and the evolution of hominin walking.


title: Preferred synonymous codons are translated more accurately: Proteomic evidence, among-species variation, and mechanistic basis

abstract: A commonly stated cause of the widespread phenomenon of unequal uses of synonymous codons is their differential translational accuracies. However, this long-standing translational accuracy hypothesis (TAH) of codon usage bias has had no direct evidence beyond anecdotes. Analyzing proteomic data from Escherichia coli, we observe higher translational accuracies of more frequently used synonymous codons, offering direct, global evidence for the TAH. The experimentally measured codon-specific translational accuracies validate a sequence-based proxy; this proxy provides support for the TAH from the vast majority of over 1000 taxa surveyed in all domains of life. We find that the relative translational accuracies of synonymous codons vary substantially among taxa and are strongly correlated with the amounts of cognate tRNAs relative to those of near-cognate tRNAs. These and other observations suggest a model in which selections for translational efficiency and accuracy drive codon usage bias and its coevolution with the tRNA pool.


title: Systemic RNA interference-defective (SID) genes modulate dopaminergic neurodegeneration in C. elegans

abstract:
The fine-tuning of gene expression is critical for all cellular processes; aberrations in this activity can lead to pathology, and conversely, resilience. As their role in coordinating organismal responses to both internal and external factors have increasingly come into focus, small non-coding RNAs have emerged as an essential component to disease etiology. Using Systemic RNA interference Defective (SID) mutants of the nematode Caenorhabditis elegans, deficient in endogenous gene silencing, we examined the potential consequences of dysfunctional epigenomic regulation in the context of Parkinsons disease (PD). Specifically, the loss of either the sid-1 or sid-3 genes, which encode a dsRNA transporter and an endocytic regulatory non-receptor tyrosine kinase, respectively, conferred neuroprotection to dopaminergic neurons in an established transgenic C. elegans strain wherein overexpression of human -synuclein (-syn) from a chromosomally integrated multicopy transgene causes neurodegeneration. We further show that knockout of a specific microRNA, mir-2, attenuates -syn neurotoxicity; suggesting that the native targets of mir-2-dependent gene silencing represent putative neuroprotective modulators. In support of this, we demonstrated that RNAi knockdown of multiple mir-2 targets enhanced -syn-induced dopaminergic neurodegeneration. Moreover, we demonstrate that mir-2 overexpression originating in the intestine can induce neurodegeneration of dopaminergic neurons, an effect that was reversed by pharmacological inhibition of SID-3 activity. Interestingly, sid-1 mutants retained mir-2-induced enhancement of neurodegeneration. Transcriptomic analysis of -syn animals with and without a sid-1 mutation revealed 27 differentially expressed genes with human orthologs related to a variety of diseases, including PD. Among these was pgp-8, encoding a P-glycoprotein-related ABC transporter. Notably, sid-1; pgp-8 double mutants abolished the neurodegeneration resulting from intestinal mir-2 overexpression. This research positions known regulators of small RNA-dependent gene silencing within a framework that facilitates mechanistic evaluation of epigenetic responses to exogenous and endogenous factors influencing dopaminergic neurodegeneration, revealing a path toward new targets for therapeutic intervention of PD.

Author SummaryThe progressive death of neurons that produce the neurotransmitter dopamine is a clinical hallmark of Parkinsons disease (PD). An integrated response to environmental and genetic factors leads to expression changes in specific genes and non-protein-coding regulatory molecules called dsRNAs that influence the pathology underlying PD. Here we report, for the first time in an animal model of PD, that mutations in genes encoding proteins which function in the cellular import of dsRNA protect dopamine neurons from degeneration. By generating a profile of individual genes affected when dsRNA transport is incapacitated, we established a foundation for the systematic analysis of their distinct contributions to neuroprotection. One subclass of dsRNAs, termed microRNAs, function analogously to a conductor of an orchestra by controlling hundreds of genes, simultaneously, to coordinate cellular processes. We identified a single microRNA that suppresses the activity of numerous genes in dopamine neurons and contributes to neurodegeneration. Genomic knockout of this deleterious microRNA, or prevention of its transport into dopamine neurons, unmasked a set of previously unreported neuroprotective proteins. This study supports a hypothesis whereby mechanisms that "fine tune" dopamine availability intersect with regulators of dsRNA transport to cooperatively maintain an optimal balance between neuronal activity, survival, and neurodegeneration.


title: Genome-wide Association Study Reveals that PvGUX1_1 is Associated with Pod Stringlessness in Snap Bean (Phaseolus vulgaris L.)

abstract: The suture strings is a particularly important pod trait that determines the quality and texture of snap bean (Phaseolus vulgaris L.). The St locus on chromosome 2 has been described as a major locus associated with suture strings. However, the gene and genetic basis underlying this locus remain unknown. Here, we investigated the suture strings of 138 snap bean accessions across two years. A total of 3.66 million single-nucleotide polymorphisms (SNPs) were obtained by deep resequencing. Based on these SNPs, we identified a strong association signal on Chr02 and a promising candidate gene, PvGUX1_1. Further analysis revealed that the 2-bp deletion in exon of PvGUX1_1 was significantly associated with stringlessness. Comparative mapping indicated that PvGUX1_1 was a domesticated locus and diverged from PvGUX1_2 during an early stage. Our study provides important insights into the genetic mechanism of suture string formation and useful information for snap bean improvement.


title: Extracellular RNA signatures of mutant KRAS(G12C) lung adenocarcinoma cells

abstract: Extracellular RNAs (exRNAs) are actively secreted from cells in membrane-bound extracellular vesicles (EVs). Diverse classes of RNAs are secreted as exRNAs, including messenger RNAs (mRNAs), long noncoding RNAs (lncRNAs), and transposable element RNAs (TE RNAs). However, the full composition and clinical utility of exRNAs secreted in response to oncogenic signaling are unknown. Here we use both affinity- and nanofiltration-based EV isolation approaches to show that mutant KRAS(G12C) signaling results in the secretion of specific lncRNAs, TE RNAs, and mRNAs, some of which are prognostic for lung adenocarcinoma (LUAD) patient survival. We found that inhibition of KRAS(G12C) signaling broadly reprograms the noncoding transcriptome, as evidenced by a substantial increase in TE RNA secretion. KRAS(G12C) inhibition also increased the abundance of secreted lncRNAs and retained intron-containing transcripts, while decreasing the mRNA content of EVs. Oncogenic KRAS(G12C) signaling was required for the secretion of mRNAs from a set of 20 genes that are significantly associated with unfavorable clinical outcomes in LUAD. Our study suggests that both coding and noncoding RNAs that are secreted in EVs may serve as KRAS(G12C)-specific signatures for diagnosing lung cancer.


title: Enrichment of somatic mutations in schizophrenia brain targets prenatally active transcription factor bindings sites

abstract: Schizophrenia (SCZ) is a complex neuropsychiatric disorder in which both germline genetic mutations and maternal factors, such as infection and immune activation, have been implicated, but how these two strikingly different mechanisms might converge on the same phenotype is unknown. During development, cells accumulate somatic, mosaic mutations in ways that can be shaped by the cellular environment or endogenous processes, but these early developmental mutational patterns have not been studied in SCZ. Here we analyzed deep (267x) whole-genome sequencing (WGS) of DNA from cerebral cortical neurons isolated from 61 SCZ and 25 control postmortem brains to capture mutations occurring before or during fetal neurogenesis. SCZ cases showed a >15% increase in genome-wide sSNV compared to controls (p < 2e-10). Remarkably, mosaic T>G mutations and CpG transversions (CpG>GpG or CpG>ApG) were 79- and 62-fold enriched, respectively, at transcription factor binding sites (TFBS) in SCZ, but not in controls. The pattern of T>G mutations resembles mutational processes in cancer attributed to oxidative damage that is sterically blocked from DNA repair by transcription factors (TFs) bound to damaged DNA. The CpG transversions similarly suggest unfinished DNA demethylation resulting in abasic sites that can also be blocked from repair by bound TFs. Allele frequency analysis suggests that both localized mutational spikes occur in the first trimester. We call this prenatal mutational process "skiagenesis" (from the Greek skia, meaning shadow), as these mutations occur in the shadow of bound TFs. Skiagenesis reflects as-yet unidentified prenatal factors and is associated with SCZ risk in a subset ([~]13%) of cases. In turn, mutational disruption of key TFBS active in fetal brain is well positioned to create SCZ-specific gene dysregulation in concert with germline risk genes. Skiagenesis provides a fingerprint for exploring how epigenomic regulation and prenatal factors such as maternal infection or immune activation may shape the developmental mutational landscape of human brain.


title: Confirming Multiplex Q-PCR Use in COVID-19 with Next Generation Sequencing: Strategies for Epidemiological Advantage

abstract:
Rapid classification and tracking of emerging SARS-CoV-2 variants are critical for understanding the transmission dynamics and developing strategies for interrupting the transmission chain. Next-Generation Sequencing (NGS) is an exceptional tool for whole-genome analysis and deciphering new mutations. The technique has been instrumental in identifying the Variants of Concern and tracking this pandemic. However, NGS remains expensive and time-consuming for large-scale monitoring of COVID-19. This study analyzed a total of 78 de-identified samples that screened positive for SARS-CoV-2 from two timeframes, August 2020 and July 2021. All 78 samples were classified into WHO lineages by whole genome sequencing then compared with two commercially available Q-PCR assays for spike protein mutation(s). The data showed good concordance with Q-PCR and NGS analysis for specific SARS-COV-2 lineages and characteristic mutations.

Deployment of Q-PCR testing to detect known SARS-COV-2 variants may be extremely beneficial. These assays are quick and cost-effective, thus can be implemented as an alternative to sequencing for screening known mutations of SARS-COV-2 for clinical and epidemiological interest. The findings support the great potential for Q-PCR to be an effective strategy offering several COVID-19 epidemiological advantages.


title: Deep learning-based system for real-time behavior recognition and closed-loop control of behavioral mazes using depth sensing

abstract: Robust quantification of animal behavior is fundamental in experimental neuroscience research. Systems providing automated behavioral assessment are an important alternative to manual measurements avoiding problems such as human bias, low reproducibility and high cost. Integrating these tools with closed-loop control systems creates conditions to correlate environment and behavioral expressions effectively, and ultimately explain the neural foundations of behavior. We present an integrated solution for automated behavioral analysis of rodents using deep learning networks on video streams acquired from a depth-sensing camera. The use of depth sensors has notable advantages: tracking/classification performance is improved and independent of animals coat color, and videos can be recorded in dark conditions without affecting animals natural behavior. Convolutional and recurrent layers were combined in deep network architectures, and both spatial and temporal representations were successfully learned for a 4- classes behavior classification task (standstill, walking, rearing and grooming). Integration with Arduino microcontrollers creates an easy-to-use control platform providing low-latency feedback signals based on the deep learning automatic classification of animal behavior. The complete system, combining depth-sensor camera, computer, and Arduino microcontroller, allows simple mapping of input-output control signals using the animals current behavior and position. For example, a feeder can be controlled not by pressing a lever but by the animal behavior itself. An integrated graphical user interface completes a user-friendly and cost-effective solution for animal tracking and behavior classification. This open-software/open-hardware platform can boost the development of customized protocols for automated behavioral research, and support ever more sophisticated, reliable and reproducible behavioral neuroscience experiments.

category: animal behavior and cognition


title: Rats emit unique distress calls in social inequality conditions

abstract: Humans show aversion toward inequality of social reward, and this aversion plays important roles for the establishment of social cooperation. However, it has remained unknown whether commonly used experimental animals show negative responses to social reward inequality. In this study, we found that rats showed bonding-like behavior to an affiliative human who repeatedly stroked the rats. In addition, these rats emitted distress calls, an index of negative emotion, when an affiliative human stroked another rat in front of them. These distress calls had acoustic characteristics different from those emitted in response to physical stress stimuli such as air-puff. Rats emitted calls with higher frequency (28 kHz) and shorter durations (0.05 sec) in an inequality condition than the frequency and durations of calls emitted when receiving air-puff. Our results suggested that rats exhibited negative emotion with unique distress calls in response to a social inequality condition.


title: Dynamics of allosteric regulation of the phospholipase C-γ isozymes upon recruitment to membranes

abstract: Numerous receptor tyrosine kinases and immune receptors activate phospholipase C-gamma (PLC-{gamma}) isozymes at membranes to control diverse cellular processes including phagocytosis, migration, proliferation, and differentiation. The molecular details of this process are not well understood. Using hydrogen-deuterium exchange mass spectrometry (HDX-MS), we show that PLC-{gamma}1 is relatively inert to lipid vesicles that contain its substrate, PIP2, unless first bound to the kinase domain of the fibroblast growth factor receptor (FGFR1). Exchange occurs throughout PLC-{gamma}1 and is exaggerated in PLC-{gamma}1 containing an oncogenic substitution (D1165H) that allosterically activates the lipase. These data support a model whereby initial complex formation shifts the conformational equilibrium of PLC-{gamma}1 to favor activation. This receptor-induced priming of PLC-{gamma}1 also explains the capacity of a kinase-inactive fragment of FGFR1 to modestly enhance the lipase activity of PLC-{gamma}1 operating on lipid vesicles but not a soluble analog of PIP2 and highlights cooperativity between receptor engagement and membrane proximity. Priming is expected to be greatly enhanced for receptors embedded in membranes and nearly universal for the myriad of receptors and co-receptors that bind the PLC-{gamma} isozymes.