bluuebunny/crossref_metadata_2025 huggingface dataset:
An engineered culture vessel and flow system to improve thein vitroanalysis of volatile organic compounds
Authors: Eshima Jarrett , Pennington Taylor R. , Abdellatif Youssef , Olea Angela Ponce , Lusk Joel F. , Ambrose Benjamin D. , Marschall Ethan , Miranda Christopher , Phan Paula , Aridi Christina , Smith Barbara S.
Published: August 2023

Volatile organic compounds (VOCs) are a biologically important subset of an organism’s metabolome, yetin vitrotechniques for the analysis of these small molecules vary substantially in practice, restricting the interpretation and reproducibility of study findings. Here, we present an engineered culture tool, termed the “Biodome”, designed to enhance analyte sensitivity by integrating dynamic headspace sampling methodology for the recovery of VOCs from viable biological cultures. We validate the functionality of the device forin vitrovolatile metabolomics utilizing computational modeling and fluorescent imaging of mammalian cell culture. We then leverage comprehensive two-dimensional gas chromatography coupled with a time-of-flight mass spectrometer and the enhanced sampling capabilities afforded by our tool to identify seven VOCs not found in the media or exogenously derived from the sampling method (typical pitfalls within vitrovolatilome analysis). We further work to validate the endogenous production of these VOCs using two independent approaches: (i) glycolysis-mediated stable isotopic labeling techniques using13C6–D-glucose and (ii) RNA interference (RNAi) to selectively knockdown β-oxidation via silencing ofCPT2. Isotope labeling reveals 2-Decen-1-ol as endogenously derived with glucose as a carbon source and, through RNAi, we find evidence supporting endogenous production of 2-ethyl-1-hexene, dodecyl acrylate, tridecanoic acid methyl ester and a low abundance alkene (C17) with molecular backbones likely derived from fatty acid degradation. To demonstrate applicability beyond mammalian cell culture, we assess the production of VOCs throughout the log and stationary phases of growth in ampicillin-resistant DH5αEscherichia coli. We identified nine compounds with results supporting endogenous production, six of which were not previously associated withE. coli. Our findings emphasize the improved capabilities of the Biodome forin vitrovolatile metabolomics and provide a platform for the standardization of methodology.
Copy number losses of oncogenes and gains of tumor suppressor genes generate common driver events of human cancer
Authors: Besedina Elizaveta , Supek Fran
Published: August 2023

Cancer driver genes can be under positive selection for various types of genetic alterations, including gain-of-function or loss-of-function point mutations (single-nucleotide variants, SNV), small indels, copy number alterations (CNA) and other structural variants. We studied the landscape of interactions between these different types of alterations affecting the same gene by a statistical method, MutMatch, which can test for significant differences in selection, while accounting for various causes of mutation risk heterogeneity. Analyzing ∼18,000 cancer exomes and genomes, we found that known oncogenes simultaneously exhibit signatures of positive selection and also negative selection, where the latter can mask the former. Consistently, focussing on known positively selected regions identifies additional tumor types where an oncogene is relevant. Next, we characterized the landscape of CNA-dependent selection effects, revealing a general trend of increased positive selection on oncogene mutations not only upon CNA gains but also upon CNA deletions. Conversely, we observe a positive interaction between mutations and CNA gains in tumor suppressor genes. Thus, two-hit events involving point mutations and CNA are universally observed on driver genes regardless of the type of CNA, and may signal new therapeutic opportunities that have been overlooked. An explicit focus on the somatic CNA two-hit events can identify additional driver genes relevant to a tumor type. By a global analysis of CNA-selection effects across many driver genes and tissues, we identified at least four independently varying signatures, and thus generated a comprehensive, data-driven classification of cancer genes by mechanisms of (in)activation by genetic alterations.
Appelmans Protocol forin vitro Klebsiella pneumoniaephage host range expansion leads to induction of a novel temperate linear plasmid prophagevB_KpnS-KpLi5
Authors: Jakob Nadine , Hammerl Jens A , Swierczewski Brett E , Würstle Silvia , Bugert Joachim J
Published: August 2023

Adjuvant therapy with bacteriophage (phage) cocktails in combination with antibiotics is a therapeutic approach currently considered for treatment of infections with encapsulated, biofilm forming, and multidrug-resistantKlebsiella pneumoniae(Kp).Klebsiellaphage are highly selective in targeting a bacterial capsule type. Considering the numerous Kp capsule types and other Kp host restriction factors, phage treatment could be facilitated when generating phages with a broad host range A modified ‘Appelmans protocol’ was used to create phages with an extended host range viain vitroforced DNA recombination. Three T7-like Kp phages with highly colinear genomes were subjected to successive propagation on their susceptible host strains representing the capsule types K64, K27, and K23, and five Kp isolates of the same capsule types initially unsusceptible for phage lysis. After 30 propagation cycles, five phages were isolated via plaque assay. Four output phages represented the original input phages, while the fifth lysed a previously non-permissible Kp isolate, which was not lysed by any of the input phages. Surprisingly, sequence analysis revealed a novel N15/phiKO2-like phage genome (vB_KpnS_KpLi5) lacking substantial homologies to any of the used T7-like phages. This temperate phage was only induced in the presence of all input phages (cocktail), but not by any of them individually. Induction of temperate phages may be a stress response caused by using multiple phages simultaneously. Successive use of different phages for therapeutic purposes may be preferable over simultaneous application in cocktail formulations to avoid undesired induction of temperate phages. (243)
The Effects of Improved Housing on Malaria Transmission in Different Endemic Zones: A Systematic Review and Meta-Analysis
Authors: Nawa Mukumbuta , Adetokunboh Olatunji
Published: August 2023

Introduction

Improved housing has been shown to reduce the risk of malaria infections compared to traditional houses; however, it is unclear if the effects differ in different malaria transmission settings. This study evaluated the effects of improved housing on malaria transmission among different endemic areas.

Methods and Analysis

Electronic databases, clinical trial registries and grey literature were searched for randomised controlled trials, cohort studies, case-control studies, and cross-sectional surveys on housing done between 1987 and 2022. Forest plots were done, and the quality of evidence was assessed using the Grading of Recommendations, Assessments, Development and Evaluation Framework.

Findings

Twenty-two studies were included; twelve were cross-sectional, four were case-control, four were cohort studies and two were RCTs. RCTs indicated that modern houses did not protect against malaria compared to traditional houses. Cohort studies showed an adjusted risk ratio of 0.68 (95% CI 0.48 – 0.96) and Cross-sectional studies indicated an adjusted odds ratio (aOR) of 0.47 (95% CI 0.31 – 0.72). By endemic transmission regions, the adjusted odds ratio in the high endemic was 0.43 (95% CI 0.29 – 0.63); in the moderate transmission regions, aOR = 0.91 (95% CI 0.91 – 1.07) and in the low transmission settings, aOR = 0.42 (95% CI 0.26 – 0.66).

Conclusion

The evidence from observational studies suggests that the risk reduction associated with modern housing is comparable or higher in low malaria transmission settings compared to high transmission settings. Evidence from RCTs in high-transmission settings shows that house improvements may induce risky behaviours such as staying outside for longer hours.

Key Messages

It is known that improved housing reduces the risk of malaria compared to traditional housing; however, the effects of improved housing in different endemic settings are unclear.

To the best of our knowledge, this is the first time a systematic review and meta-analysis has stratified the effect measures of improved housing on malaria transmission in different transmission settings.

Our study found no literature from high-quality research designs such as RCTs and Cohort studies on improved housing in low and moderate transmission settings. We call on researchers and funders to conduct and support high-quality research designs in low and moderate-transmission areas, especially in Africa, as more countries are reducing their malaria burdens due to increased interventions. This will help to achieve and sustain malaria elimination.

Piecemeal improvements, such as closing eaves, screening and iron roofing, are not necessarily associated with a reduced risk of malaria. They may induce risky behaviours due to poor ventilation and higher indoor temperatures resulting in residents staying longer outside thus exposing themselves to infective mosquito bites.
Comparability and Stability of Holotranscobalamin levels in Capillary and Venous Blood
Authors: Woolley Timothy , Rutter Emma , Staudenmaier Macarena
Published: August 2023

Introduction

Self-collected capillary blood has numerous advantages over venous samples taken via phlebotomy, as such the use of this finger-prick testing has increased over recent years, driven in some part by Covid antibody testing. However, there is limited evidence that venous and capillary serum is comparable for many routine analytes. In this study we aimed to determine whether capillary sampling could offer an alternative sampling method to venous for the assessment of vitamin B12 in the form of holotranscobalamin, and if this analyte was stable in unspun capillary blood for three days, thus allowing for samples to be returned by the postal system.

Methods

Matched pairs of capillary and venous blood samples were collected from 25 patients for the determination of holotranscobalamin, one set being processed on day zero, the other set being stored at ambient temperature and then processed on day three to mimic postal samples. Self-collected capillary blood was compared with phlebotomist-collected venous samples using correlation, Passing-Bablock regression, and Altman-Bland analysis.

Results

Passing-Bablock and Bland-Altman analysis showed holotranscobalamin levels in capillary and venous serum to be comparable and that the analyte was stable in unspun blood for at least three days at ambient temperatures.

Conclusions

We believe this is the first published study to determine if capillary blood sampling is an acceptable alternative to venous sampling for determining holotranscobalamin concentration; our data indicates that there is no significant difference in results from unspun venous and capillary blood stored at room temperature for at least 3 days compared to venous blood tested on the same day of collection.
Matching or genetic engineering of HLA Class I and II facilitates successful allogeneic ‘off-the-shelf’ regulatory T cell therapy
Authors: McCallion Oliver , Du Weijie , Glaser Viktor , Milward Kate , Franke Clemens , Kath Jonas , Valkov Mikhail , Yang Mingxing , Künkele Annette , Polansky Julia K. , Schmueck-Henneresse Michael , Volk Hans-Dieter , Reinke Petra , Wagner Dimitrios L. , Hester Joanna , Issa Fadi
Published: August 2023

The potential to harness regulatory T cells (Tregs) for the treatment of autoimmune diseases and transplant rejection has been restricted by several barriers: donor variability, manufacturing complications, and time-consuming expansion processes. These issues further complicate the use of autologous Tregs during acute disease phases or when Tregs are low in number or dysfunctional. Here we explore the potential of ‘off-the-shelf’ allogeneic Tregs, from healthy donors or universal sources, to provide a more practical solution. We discover that the efficacy of these cells is undermined by the recipient’s immune response, and that that rigorous matching of HLA classes I and II overcomes this barrier. Importantly, genetically manipulating HLA expression enables the use of unmatched allogeneic Tregs within vivoefficacy. Our findings underscore the transformative potential of HLA-engineered Tregs, offering a novel, ready-to-use therapeutic avenue for treating a wide array of inflammatory diseases.

One-Sentence Summary

Matching or engineering of HLA-I and HLA-II facilitates allogeneic ‘off-the-shelf’ regulatory T cells for immunoregulation.
Reversal of Biological Age in Multiple Rat Organs by Young Porcine Plasma Fraction
Authors: Horvath Steve , Singh Kavita , Raj Ken , Khairnar Shraddha , Sanghavi Akshay , Shrivastava Agnivesh , Zoller Joseph A. , Li Caesar Z. , Herenu Claudia B. , Canatelli-Mallat Martina , Lehmann Marianne , Habazin Siniša , Novokmet Mislav , Vučković Frano , Solberg Woods Leah C. , Martinez Angel Garcia , Wang Tengfei , Chiavellini Priscila , Levine Andrew J. , Chen Hao , Brooke Robert T , Gordevicius Juozas , Lauc Gordan , Goya Rodolfo G. , Katcher Harold L.
Published: August 2023

Young blood plasma is known to confer beneficial effects on various organs in mice and rats. However, it was not known whether plasma from young pigs rejuvenates old rat tissues at the epigenetic level; whether it alters the epigenetic clock, which is a highly accurate molecular biomarker of aging. To address this question, we developed and validated six different epigenetic clocks for rat tissues that are based on DNA methylation values derived from n=613 tissue samples. As indicated by their respective names, the rat pan-tissue clock can be applied to DNA methylation profiles from all rat tissues, while the rat brain-, liver-, and blood clocks apply to the corresponding tissue types. We also developed two epigenetic clocks that apply to both human and rat tissues by adding n=1366 human tissue samples to the training data. We employed these six rat clocks to investigate the rejuvenation effects of a porcine plasma fraction treatment in different rat tissues. The treatment more than halved the epigenetic ages of blood, heart, and liver tissue. A less pronounced, but statistically significant, rejuvenation effect could be observed in the hypothalamus. The treatment was accompanied by progressive improvement in the function of these organs as ascertained through numerous biochemical/physiological biomarkers and behavioral responses to assess cognitive functions. An immunoglobulin G (IgG) N-glycosylation pattern shift from pro-to anti-inflammatory also indicated reversal of glycan aging. Overall, this study demonstrates that a young porcine plasma-derived treatment markedly reverses aging in rats according to epigenetic clocks, IgG glycans, and other biomarkers of aging.
HEARTSVG: a fast and accurate method for spatially variable gene identification in large-scale spatial transcriptomic data
Authors: Yuan Xin , Ma Yanran , Gao Ruitian , Cui Shuya , Wang Yifan , Fa Botao , Ma Shiyang , Wei Ting , Ma Shuangge , Yu Zhangsheng
Published: August 2023

Identifying spatially variable genes (SVGs) is crucial for understanding the spatiotemporal characteristics of diseases and tissue structures, posing a distinctive challenge in spatial transcriptomics research. We propose HEARTSVG, a distribution-free, test-based method for fast and accurately identifying spatially variable genes in large-scale spatial transcriptomic data. Extensive simulations demonstrate that HEARTSVG outperforms state-of-the-art methods with higherF1scores (averageF1score=0.903), improved computational efficiency, scalability, and reduced false positives (FPs). Through analysis of twelve real datasets from various spatial transcriptomic technologies, HEARTSVG identifies a greater number of biologically significant SVGs (average recall=0.985, average AUC=0.788) than other comparative methods without prespecifing spatial patterns. Furthermore, by clustering SVGs, we uncover two distinct tumor spatial domains characterized by unique spatial expression patterns, spatial-temporal locations, and biological functions in human colorectal cancer data, unraveling the complexity of tumors.
An Aurora kinase A-BOD1L1-PP2A B56 Axis promotes chromosome segregation fidelity
Authors: Kucharski Thomas J. , Vlasac Irma M. , Higgs Martin R. , Christensen Brock C. , Bechstedt Susanne , Compton Duane A.
Published: August 2023

Cancer cells are often aneuploid and frequently display elevated rates of chromosome mis-segregation in a phenomenon called chromosomal instability (CIN). CIN is commonly caused by hyperstable kinetochore-microtubule (K-MT) attachments that reduces the efficiency of correction of erroneous K-MT attachments. We recently showed that UMK57, a chemical agonist of MCAK (alias KIF2C) improves chromosome segregation fidelity in CIN cancer cells although cells rapidly develop adaptive resistance. To determine the mechanism of resistance we performed unbiased proteomic screens which revealed increased phosphorylation in cells adapted to UMK57 at two Aurora kinase A phosphoacceptor sites on BOD1L1 (alias FAM44A). BOD1L1 depletion or Aurora kinase A inhibition eliminated resistance to UMK57 in CIN cancer cells. BOD1L1 localizes to spindles/kinetochores during mitosis, interacts with the PP2A phosphatase, and regulates phosphorylation levels of kinetochore proteins, chromosome alignment, mitotic progression and fidelity. Moreover, theBOD1L1gene is mutated in a subset of human cancers, andBOD1L1depletion reduces cell growth in combination with clinically relevant doses of taxol or Aurora kinase A inhibitor. Thus, an Aurora kinase A-BOD1L1-PP2A axis promotes faithful chromosome segregation during mitosis. (175 words)
The impact of ZIKV infection on gene expression in neural cells over time
Authors: Rodrigues Moreno Magalhães de Souza , Júnior Antonio Marques Pereira , Fukutani Eduardo Rocha , Araújo-Pereira Mariana , Salgado Vanessa Riesz , de Queiroz Artur Trancoso Lopo
Published: August 2023

Zika virus (ZIKV) outbreak caused one of the most significant medical emergencies in the Americas due to associated microcephaly in newborns. To evaluate the impact of ZIKV infection on neuronal cells over time, we retrieved gene expression data from several ZIKV-infected samples obtained at different time point post-infection (pi). Differential gene expression analysis was applied at each time point, with more differentially expressed genes (DEG) identified at 72h pi. There were 5 DEGs (PLA2G2F, TMEM71, PKD1L2, UBD, and TNFAIP3 genes) across all timepoints, which clearly distinguished between infected and healthy samples. The highest expression levels of all five genes were identified at 72h pi. Taken together, our results indicate that ZIKV infection greatly impacts human neural cells at early times of infection, with peak perturbation observed at 72h pi. Our analysis revealed that all five DEGs, in samples of ZIKV-infected human neural stem cells, remained highly upregulated across the timepoints evaluated. Moreover, despite the pronounced inflammatory host response observed throughout infection, the impact of ZIKV is variable over time. Finally, the five DEGs identified herein play prominent roles in infection, and could serve to guide future investigations into virus-host interaction, as well as constitute targets for therapeutic drug development.
Directional uncertainty in chase and escape dynamics
Authors: Treviño Mario , Medina-Coss y León Ricardo , Támez Sergio , Beltrán-Navarro Beatriz , Verdugo Jahir
Published: August 2023

Intercepting and avoiding collisions with moving targets are crucial skills for survival. However, little is known about how these behaviors are implemented when the trajectory of the moving target introduces variability and ambiguity into the perceptual-motor system. We developed a simple visuomotor task in which participants used a joystick to interact with a computer-controlled dot that moved along two-dimensional trajectories. This virtual system allowed us to define the role of the moving object (predator or prey) and adjust its speed and directional uncertainty (i.e.,magnitude and frequency of random directional changes) during chase and escape trials. These factors had a significant impact on participants’ performance in both chasing and escaping trials. We developed a simple geometrical model of potential chaser/escaper interactions to distinguish pursuit from interception chasing trajectories. We found that participants initially pursued the target but switched to a late interception strategy. The amount of late interception strategy followed an inverted U-shaped curve with the highest values at intermediate speeds. We tested the applicability of our task and methods in children who showed a robust developmental improvement in task performance and late interception strategy. Our task constitutes a flexible system in a virtual space for studying chasing and escaping behavior in adults and children. Our analytical methods allow detecting subtle changes in interception strategies, a valuable tool for studying the maturation of predictive and prospective systems, with a high potential to contribute to cognitive and developmental research.
Neural crest cells give rise to non-myogenic mesenchymal tissue in the adult murid ear pinna
Authors: Allen Robyn S. , Biswas Shishir K. , Seifert Ashley W.
Published: August 2023

Despite being a major target of reconstructive surgery, development of the external ear pinna remains poorly studied. As a craniofacial organ highly accessible to manipulation and highly conserved among mammals, the ear pinna represents a valuable model for the study of appendage development and wound healing in the craniofacial complex. Here we provide a cellular characterization of late gestational and postnatal ear pinna development inMus musculusandAcomys cahirinusand demonstrate that ear pinna development is largely conserved between these species. UsingWnt1-cre;ROSAmT/mGmice we find that connective tissue fibroblasts, elastic cartilage, dermal papilla cells, dermal sheath cells, vasculature, and adipocytes in the adult pinna are derived from cranial crest. In contrast, we find that skeletal muscle and hair follicles are not derived from neural crest cells. Cellular analysis using the naturally occurringshort earmouse mutant shows that elastic cartilage does not develop properly in distal pinna due to impaired chondroprogenitor proliferation. Interestingly, while chondroprogenitors develop in a mostly continuous sheet, the boundaries of cartilage loss in theshort earmutant strongly correlate with locations of vasculature-conveying foramen. Concomitant with loss of elastic cartilage we report increased numbers of adipocytes, but this seems to be a state acquired in adulthood rather than a developmental abnormality. In addition, chondrogenesis remains impaired in the adult mid-distal ear pinna of these mutants. Together these data establish a developmental basis for the study of the ear pinna with intriguing insights into the development of elastic cartilage.
RNA-mediated ribonucleoprotein assembly controls TDP-43 nuclear retention
Authors: dos Passos Patricia M. , Hemamali Erandika H. , Mamede Lohany D. , Hayes Lindsey R. , Ayala Yuna M.
Published: August 2023

TDP-43 is an essential RNA-binding protein strongly implicated in the pathogenesis of neurodegenerative disorders characterized by cytoplasmic aggregates and loss of nuclear TDP-43. The protein shuttles between nucleus and cytoplasm, yet maintaining predominantly nuclear TDP-43 localization is important for TDP-43 function and for inhibiting cytoplasmic aggregation. We previously demonstrated that specific RNA binding mediates TDP-43 self-assembly and biomolecular condensation, requiring multivalent interactions via N- and C-terminal domains. Here, we show that these complexes play a key role in TDP-43 nuclear retention. TDP-43 forms macromolecular complexes with a wide range of size distribution in cells and we find that defects in RNA binding or inter-domain interactions, including phase separation, impair the assembly of the largest species. Our findings suggest that recruitment into these macromolecular complexes prevents cytoplasmic egress of TDP-43 in a size-dependent manner. Our observations uncover fundamental mechanisms controlling TDP-43 cellular homeostasis, whereby regulation of RNA-mediated self-assembly modulates TDP-43 nucleocytoplasmic distribution. Moreover, these findings highlight pathways that may be implicated in TDP-43 proteinopathies and identify potential therapeutic targets.
Dietaryindex: A User-Friendly and Versatile R Package for Standardizing Dietary Pattern Analysis in Epidemiological and Clinical Studies
Authors: Zhan Jiada James , Hodge Rebecca A , Dunlop Anne L , Lee Matthew M. , Bui Linh , Liang Donghai , Ferranti Erin P
Published: August 2023

Background

Few standardized and open-source tools exist for calculating dietary pattern indexes from dietary intake data in epidemiological and clinical studies. Miscalculations of dietary indexes, with suspected erroneous findings, are occasionally noted in the literature.

Objective

The primary aim is to develop and validate dietaryindex, a user-friendly and versatile R package that standardizes the calculation of dietary indexes.

Methods

Dietaryindex utilizes a two-step process: an initial calculation of serving size for each food and nutrient category, followed by the calculation of individual dietary indexes. It includes generic functions that accept any preprocessed serving sizes of food groups and nutrients, with the standard serving sizes defined according to the methodologies used in well-known prospective cohort studies. For ease of use, dietaryindex also offers one-step functions that directly reference common datasets and tools, including the National Health and Nutrition Examination Survey (NHANES) and Block Food Frequency Questionnaire, eliminating the need for data preprocessing. At least two independent researchers validated the serving size definitions and scoring algorithms of dietaryindex.

Results

Dietaryindex can calculate multiple dietary indexes of high interest in research, including Healthy Eating Index (HEI) - 2020, Alternative Healthy Eating Index 2010, Dietary Approaches to Stop Hypertension Index, Alternate Mediterranean Diet Score, Dietary Inflammatory Index, American Cancer Society 2020 dietary index, and Planetary Health Diet Index from the EAT-Lancet Commission. In our validation process, dietaryindex demonstrated full accuracy (100%) in all generic functions with two-decimal rounding precision in comparison to hand-calculated results. Similarly, using NHANES 2017-2018 data and ASA24 and DHQ3 example data, the HEI2015 outputs from dietaryindex aligned (99.95% - 100%) with results using the SAS codes from the National Cancer Institute.

Conclusions

Dietaryindex is a user-friendly, versatile, and validated informatics tool for standardized dietary index calculations. We have open-sourced all the validation files and codes with detailed tutorials on GitHub (https://github.com/jamesjiadazhan/dietaryindex).
Cell-free chromatin particles from dying cells promote the induction of an immune response in human lymphocytes
Authors: Shabrish Snehal , Raghuram Gorantla V. , Raphael Nimisha , Narayanan Ranjani , Lopes Relestina , Kondhalkar Dipali , Khare Naveen Kumar , Mittra Indraneel
Published: August 2023

It is long established that cell death and immune response are closely related, although the nature of this relationship has remained unclear. We earlier reported that cell-free chromatin particles (cfChPs) released from the billions of cells that die in the body every day to enter into the blood circulation, or those that are released locally from dying cells, can readily enter into healthy cells to induce DNA damage and activate inflammatory cytokines. In this study we investigated whether cfChPs from dying cells might be the missing link between cell death and immune response. We treated human lymphocytes with cfChPs isolated from sera of healthy individuals or with cfChPs released from hypoxia-induced dying lymphocytes. We observed that cfChPs from both sources readily entered into lymphocytes to accumulate in their nuclei within 2 h. This was associated with marked activation of CD69 and release of inflammatory cytokines, as well as of p-STING and cGAS expression. The addition of the STING protein inhibitor H151 to the cfChPs treated cells abolished the release of inflammatory cytokines. These findings lead us to suggest that cfChPs from dying cells are the critical triggers of immune response which act via cGAS-STING pathway.
Visualizing and quantifying structural diversity around mobile resistance genes
Authors: Shaw Liam P. , Neher Richard A.
Published: August 2023

Understanding the evolution of mobile genes is important for understanding the spread of antimicrobial resistance (AMR). Many clinically important AMR genes have been mobilized by mobile genetic elements (MGEs) on the kilobase scale, such as integrons and transposons, which can integrate into both chromosomes and plasmids and lead to rapid spread of the gene through bacterial populations. Looking at the flanking regions of these mobile genes in diverse genomes can highlight common structures and reveal patterns of MGE spread. However, historically this has been a largely descriptive process, relying on gene annotation and expert knowledge. Here we describe a general method to visualize and quantify the structural diversity around genes usingpangraphto find blocks of homologous sequence. We apply this method to a set of twelve clinically important beta-lactamase genes and provide interactive visualizations of their flanking regions athttps://liampshaw.github.io/flanking-regions. We show that nucleotide-level variation in the mobile gene itself generally correlates with increased structural diversity in its flanking regions, demonstrating a relationship between rates of mutational evolution and rates of structural evolution, and find a bias for greater structural diversity upstream. Our framework is a starting point to investigate general rules that apply to the horizontal spread of new genes through bacterial populations.

Impact statement

Understanding the evolution and spread of mobile resistance genes is challenging because of the high variability in their genomic contexts. Here we outline a fast computational approach that identifies stretches of homologous sequence in the flanking regions of a gene, simultaneously producing interactive visualizations of these regions and quantifying the diversity within them. As an example, we apply the method to twelve clinically important beta-lactamase genes. We find that structural diversity around the resistance gene is correlated with mutations within it and that there is greater structural diversity upstream of many genes. There may be other general patterns about the evolution of mobile resistance genes that can be recovered with this kind of analysis.
Enhancement of High-Density Lipoprotein-Associated Protease Inhibitor Activity Prevents Atherosclerosis Progression
Authors: Mobilia Maura , Karakashian Alexander , Whitus Callie , Neupane Khaga R. , Johnson Lance A. , Graf Gregory A. , Gordon Scott M.
Published: August 2023

Background

Inflammatory cells within atherosclerotic lesions secrete various proteolytic enzymes that contribute to lesion progression and destabilization, increasing the risk for an acute cardiovascular event. The relative contributions of specific proteases to atherogenesis is not well understood. Elastase is a serine protease, secreted by macrophages and neutrophils, that may contribute to the development of unstable plaque. We have previously reported interaction of endogenous protease-inhibitor proteins with high-density lipoprotein (HDL), including alpha-1-antitrypsin, an inhibitor of elastase. These findings support a potential role for HDL as an endogenous modulator of protease activity. In this study, we test the hypothesis that enhancement of HDL-associated elastase inhibitor activity is protective against atherosclerotic lesion progression.

Methods

We designed an HDL-targeting protease inhibitor (HTPI) that binds to HDL and confers elastase inhibitor activity. Lipoprotein binding and the impact of HTPI on atherosclerosis was examined using mouse models.

Results

HTPI is a small (1.6 kDa) peptide with an elastase inhibitor domain, a soluble linker, and an HDL-targeting domain. When incubated with human plasmaex vivo, HTPI predominantly binds to HDL. Intravenous administration of HTPI to mice resulted in its binding to plasma HDL and increased elastase inhibitor activity on isolated HDL. Accumulation of HTPI within plaque was observed after systemic administration toApoe-/-mice. To examine the effect of HTPI treatment on atherosclerosis, prevention and progression studies were performed usingLdlr-/-mice fed Western diet. In both study designs, HTPI-treated mice had reduced lipid deposition in plaque. Histology and immunofluorescence staining of aortic root sections were used to examine the impact of HTPI on lesion morphology and inflammatory features.

Conclusions

These data support the hypothesis that HDL-associated anti-elastase activity can improve the atheroprotective potential of HDL and highlight the potential utility of HDL enrichment with anti-protease activity as an approach for stabilization of atherosclerotic lesions.
Inference of Selective Force on House Mouse Genomes during Secondary Contact in East Asia
Authors: Fujiwara Kazumichi , Kubo Shunpei , Endo Toshinori , Takada Toyoyuki , Shiroishi Toshihiko , Suzuki Hitoshi , Osada Naoki
Published: August 2023

The house mouse (Mus musculus), commensal to humans, has spread globally via human activities, leading to secondary contact between genetically divergent subspecies. This pattern of genetic admixture can provide insights into the selective forces at play in this well-studied model organism. Our analysis of 163 house mouse genomes, with a particular focus on East Asia, revealed substantial admixture between the subspeciescastaneusandmusculus, particularly in Japan and southern China. We revealed, despite the admixture, all Y chromosomes in the East Asian samples belonged to themusculus-type haplogroup, potentially explained by genomic conflict under sex ratio distortion due to varying copy numbers of ampliconic genes on sex chromosomes. We also investigated the influence of selection on the post-hybridization of the subspeciescastaneusandmusculusin Japan. Even though the genetic background of most Japanese samples closely resembles the subspeciesmusculus, certain genomic regions overrepresented thecastaneus-like genetic components, particularly in immune-related genes. Furthermore, a large genomic block containing a vomeronasal/olfactory receptor gene cluster predominantly harboredcastaneus-type haplotypes in the Japanese samples, highlighting the crucial role of olfaction-based recognition in shaping hybrid genomes.
Compartment specific responses to contractility in the small intestinal epithelium
Authors: Hinnant Taylor , Ning Wenxiu , Lechler Terry
Published: August 2023

Tissues are subject to multiple mechanical inputs at the cellular level that influence their overall shape and function. In the small intestine, actomyosin contractility can be induced by many physiological and pathological inputs. However, we have little understanding of how contractility impacts the intestinal epithelium on a cellular and tissue level. In this study, we probed the cell and tissue-level effects of contractility by using mouse models to genetically increase the level of myosin activity in the two distinct morphologic compartments of the intestinal epithelium, the crypts and villi. We found that increased contractility in the villar compartment caused shape changes in the cells that expressed the transgene and their immediate neighbors. While there were no discernable effects on villar architecture, even low levels of transgene induction in the villi caused non-cell autonomous hyperproliferation of the transit amplifying cells in the crypt, driving increased cell flux through the crypt-villar axis. In contrast, induction of increased contractility in the proliferating cells of the crypts resulted in nuclear deformations, DNA damage, and apoptosis. This study reveals the complex and diverse responses of different intestinal epithelial cells to contractility and provides important insight into mechanical regulation of intestinal physiology.

Author Summary

The small intestine epithelium is comprised of two main compartments: the villi which contain differentiated cells that function in nutrient absorption, and the crypts which are made up of undifferentiated cells which serve to replenish cells of the villi. Because of their physical location within the tissue, villi and crypts are subjected to different types of insults and mechanical forces. We sought to directly test how villi and crypts respond to mechanical changes in the epithelia by genetically inducing actomyosin contraction. Increasing contractility in villar cells resulted in cell shape changes without affecting their overall polarity or organization. However, it led to a non-autonomous increase in proliferation of the undifferentiated cells of the intestine. In contrast, increased contractility in the proliferative cells of the crypt resulted in nuclear shape changes, DNA damage and ultimately a rapid cell death. Thus, our work demonstrates that the crypt and villi epithelia respond differently to mechanical changes and highlights long-range regulation between villi and crypt compartments.
Variant annotation across homologous proteins (“Paralogue Annotation”) identifies disease-causing missense variants with high precision, and is widely applicable across protein families
Authors: Li Nicholas , Mazaika Erica , Theotokis Pantazis , Zhang Xiaolei , Jang Mikyung , Ahmad Mian , Powell George , Heyne Henrike O. , Lal Dennis , Barton Paul JR , Walsh Roddy , Whiffin Nicola , Ware James S
Published: August 2023

Background

Distinguishing pathogenic variants from those that are rare but benign remains a key challenge in clinical genetics, especially for variants not previously observed and characterised in humans.In vitroandin vivofunctional characterisation are typically resource intensive, and model systems may not accurately predict influence on human disease. Manyin silicotools have been developed to predict which variants are disease-causing, but typically lack precision. Here we demonstrate the applicability of a framework, called Paralogue Annotation, that draws on information from previously-characterised variants in homologous proteins to predict whether variants in a gene of interest are likely disease causing.

Methods

We assessed the performance of Paralogue Annotation through three orthogonal approaches: (1) comparison to establishedin silicovariant prediction tools using 47,360 missense variants from ClinVar across 3,524 genes representing a broad range of diverse protein classes, by calculating precision and sensitivity; (2) evaluation against large-scale functional assays of variant effect inTP53andPPARG; and (3) comparing odd ratios calculated from case-control association tests for inherited cardiac arrhythmia syndromes, and neurodevelopmental disorders with epilepsy, stratifying variants by Paralogue Annotation.

Results

Paralogue Annotation correctly annotates 4,328 ClinVar pathogenic variants, with 245 false positives, yielding a precision of 0.95. This increases to 0.99 with more stringent annotation parameters (requiring greater conservation of amino acids between annotated orthologues) at the expense of sensitivity. Compared to established tools, Paralogue Annotation has higher precision for identification of pathogenic variants, albeit with lower sensitivity across diverse test sets. Extending the technique by transferring annotations between homologous protein domains, rather than full-length protein paralogues, increases sensitivity. Rare variants predicted pathogenic by Paralogue Annotation were more strongly disease-associated (increased odds ratio) than unstratified rare variants for six out of eight genes tested with case-control cohort approaches.

Conclusions

Paralogue Annotation has high precision for detection of pathogenic missense variants, outperformingin silicomethods where data are available to make a prediction. As the number of characterised variants increases in reference datasets such as ClinVar, Paralogue Annotation will further increase in sensitivity and applicability.
Extracellular vesicles are the main contributor to the non-viral protected extracellular sequence space
Authors: Lücking Dominik , Mercier Coraline , Alarcón-Schumacher Tomas , Erdmann Susanne
Published: August 2023

Marine environmental virus metagenomes, commonly referred to as ’viromes’, are typically generated by physically separating virus-like particles (VLPs) from the microbial fraction based on their size and mass. However, most methods used to purify VLPs, enrich extracellular vesicles (EVs) and gene transfer agents (GTAs) simultaneously. Consequently, the sequence space traditionally referred to as a ’virome’ contains host-associated sequences, transported via EVs or GTAs. We therefore propose to call the genetic material isolated from size-fractionated (0.22 µm) and DNase-treated samplesprotected environmental DNA (peDNA).This sequence space contains viral genomes, DNA transduced by viruses and DNA transported in EVs and GTAs. Since there is no genetic signature for peDNA transported in EVs, GTAs and virus particles, we rely on the successful removal of contaminating remaining cellular and free DNA when analyzing peDNA. Using marine samples collected from the North Sea, we generated a thoroughly purified peDNA dataset and developed a bioinformatic pipeline to determine the potential origin of the purified DNA. This pipeline was applied to our dataset as well as existing global marine ’viromes’. Through this pipeline, we identified known GTA and EV producers, as well as organisms with actively transducing proviruses as the source of the peDNA, thus confirming the reliability of our approach. Additionally, we identified novel and widespread EV producers, and found quantitative evidence suggesting that EV-mediated gene transfer plays a significant role in driving horizontal gene transfer (HGT) in the world’s oceans.
Vitamin C activates young LINE-1 elements in mouse embryonic stem cells via H3K9me3 demethylation
Authors: Cheng Kevin C.L. , Frost Jennifer M. , Sánchez-Luque Francisco J. , García-Canãdas Marta , Taylor Darren , Yang Wan R. , Irayanar Branavy , Sampath Swetha , Patani Hemalvi , Agger Karl , Helin Kristian , Ficz Gabriella , Burns Kathleen H. , Ewing Adam , García-Pérez José L. , Branco Miguel R.
Published: August 2023

Vitamin C (vitC) enhances the activity of 2-oxoglutarate-dependent dioxygenases, including TET enzymes, which catalyse DNA demethylation, and Jumonji-domain histone demethylases. The epigenetic remodelling promoted by vitC improves the efficiency of induced pluripotent stem cell derivation, and is required to attain a ground-state of pluripotency in embryonic stem cells (ESCs) that closely mimics the inner cell mass of the early blastocyst. However, genome-wide DNA and histone demethylation can lead to upregulation of transposable elements (TEs), and it is not known how vitC addition in culture media affects TE expression in pluripotent stem cells. Here we show that vitC increases the expression of evolutionarily young LINE-1 (L1) elements in mouse ESCs. We find that TET activity is dispensable for these effects, and that instead L1 upregulation occurs largely as a result of H3K9me3 loss mediated by KDM4A/C histone demethylases. Despite increased L1 levels, we did not detect increased somatic insertion rates in vitC-treated cells. Notably, treatment of human ESCs with vitC also increases L1 protein levels, which could impact the genetic and epigenetic stability of human pluripotent stem cells.
CurveCurator: A recalibrated F-statistic to assess, classify, and explore significance of dose-response curves
Authors: Bayer Florian P. , Gander Manuel , Kuster Bernhard , The Matthew
Published: August 2023

Dose-response curves are key metrics in pharmacology and biology to assess phenotypic or molecular actions of bioactive compounds in a quantitative fashion. Yet, it is often unclear whether or not a response significantly differs from a curve without regulation, particularly in high-throughput applications or unstable assays. Treating potency and effect size estimates from random and true curves with the same level of confidence can lead to incorrect hypotheses and issues in training machine learning models. Here, we present CurveCurator, an open-source software and interactive dashboard that provides reliable dose-response characteristics by computing p-values and false discovery rates based on a recalibrated F-statistic and a novel thresholding procedure. Application of CurveCurator to large-scale data sets demonstrates its scalable utility across several application areas.
Activity-dependent Mitochondrial ROS Signaling Regulates Recruitment of Glutamate Receptors to Synapses
Authors: Doser Rachel L. , Knight Kaz M. , Deihl Ennis , Hoerndli Frederic J.
Published: August 2023

Our understanding of mitochondrial signaling in the nervous system has been limited by the technical challenge of analyzing mitochondrial functionin vivo. In the transparent genetic modelCaenorhabditis elegans,we were able to manipulate and measure mitochondrial signaling as well as neuronal activityin vivo. Using this approach, we provide evidence supporting a novel role for mitochondrial signaling in dendrites. Specifically, we show that dendritic mitochondria take up calcium (Ca2+) via the mitochondrial Ca2+uniporter MCU-1 causing an upregulation of mitochondrial reactive oxygen species (mitoROS) production. We also observed that mitochondria are positioned in close proximity to synaptic clusters of GLR-1, theC. elegansortholog of the AMPA subtype of glutamate receptors that mediate neuronal excitation. We show that synaptic recruitment of GLR-1 is downregulated by mitoROS signaling resulting from mitochondrial Ca2+uptake via MCU-1. Thus, postsynaptic mitochondria provide a means of negative feedback that regulates excitatory synapse function which may be vital for neuronal homeostasis by preventing excitotoxicity and energy depletion.
Phylonumtomics uncovers diverse evolutionary trajectories of mitogenomic fossils buried in mammalian and avian genomes
Authors: Chen Yu-Chi , Vendrami David L. J. , Huber Maximilian L. , Handel Luisa E. Y. , Cooney Christopher R. , Hoffman Joseph I. , Gossmann Toni I.
Published: August 2023

Sporadically genetic material that originates from an organelle genome integrates into the nuclear genome. However it is unclear what processes maintain such an integration over longer evolutionary time. Recently it was shown that nuclear DNA of mitochondrial origin (NUMTs) may harbour genes with intact mitochondrial reading frames despite the fact that they are highly divergent to the host’s mitochondrial genome. Two major hypotheses have been put forward to explain the existence of such mitocoding nuclear genes: (A) recent introgression from another species and (B) long-term selection. To address whether these intriguing possibilities do play a role we scanned the genomes of more than 1,000 avian and mammalian species for NUMTs. We indeed identified that the subclass of divergent NUMTs harbouring mitogenes with intact reading frames are widespread across mammals and birds. We can show that for these NUMTs signatures of cross-species introgression are widespread in birds, but not mammals with the exception of ungulates. We can also show that a substantial fraction of deeply divergent NUMTs are maintained by selection. For a small number of NUMT genes we identify an evolutionary signature that is consistent with adaptive evolution including one human NUMT that is shared among seven ape species. This highlights the intriguing possibility that NUMT insertions occasionally may be functional.