bluuebunny/crossref_metadata_2025 huggingface dataset:
A Strategy to Treat COVID-19 Disease with Targeted Delivery of Inhalable Liposomal Hydroxychloroquine: A Non-clinical Pharmacokinetic Study
Authors: Tai Tien-Tzu , Wu Tzung-Ju , Wu Huey-Dong , Tsai Yi-Chen , Wang Hui-Ting , Wang An-Min , Shih Sheue-Fang , Chen Yee-Chun
Published: July 2020

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a newly identified pathogen causing coronavirus disease 2019 (COVID-19) pandemic. Hydroxychloroquine (HCQ), an antimalarial and anti-inflammatory drug, has been shown to inhibit SARS-CoV-2 infectionin vitroand tested in clinical studies. However, lung concentration (6.7 µg/mL) to predict thein vivoantiviral efficacy might not be achievable with the currently proposed oral dosing regimen. Further, a high cumulative doses of HCQ may raise concerns of systemic toxicity, including cardiotoxicity. Here, we described a non-clinical study to investigate the pharmacokinetics of a novel formulation of liposomal HCQ administrated by intratracheal (IT) instillation in Sprague-Dawley (SD) rats which achieved 129.4 µg/g (Cmax) in the lung. Compared to unformulated HCQ administered intravenous (IV), liposomal HCQ with normalized dose showed higher (∼30-fold) lung exposure, longer (∼2.5-fold) half-life in lung, but lower blood exposure with ∼20% of Cmaxand 74% of AUC and lower heart exposure with 24% of Cmaxand 58% of AUC. In conclusion, the pharmacokinetics results in an animal model demonstrate the proof of concept that inhalable liposomal HCQ may provide clinical benefit and serve as a potential treatment for COVID-19.
Zika virus-like particles bearing covalent dimer of envelope protein protect mice from lethal challenge
Authors: De Lorenzo Giuditta , Tandavanitj Rapeepat , Doig Jennifer , Setthapramote Chayanee , Poggianella Monica , Velazquez Ricardo Sanchez , Scales Hannah E. , Edgar Julia M. , Kohl Alain , Brewer James , Burrone Oscar R. , Patel Arvind H.
Published: July 2020

Zika virus (ZIKV) envelope (E) protein is the major target of neutralizing antibodies in infected host, and thus represents a candidate of interest for vaccine design. However, a major concern in the development of vaccines against ZIKV and the related dengue virus is the induction of cross-reactive poorly neutralizing antibodies that can cause antibody-dependent enhancement (ADE) of infection. This risk necessitates particular care in vaccine design. Specifically, the engineered immunogens should have their cross-reactive epitopes masked, and they should be optimized for eliciting virus-specific strongly neutralizing antibodies upon vaccination. Here, we developed ZIKV subunit- and virus-like particle (VLP)-based vaccines displaying E in its wild type form, or E locked in a covalently linked dimeric (cvD) conformation to enhance the exposure of E dimers to the immune system. Compared with their wild-type derivatives, cvD immunogens elicited antibody with higher capacity of neutralizing virus infection of cultured cells. More importantly, these immunogens protected animals from lethal challenge with both the African and Asian lineages of ZIKV, impairing virus dissemination to brain and sexual organs. Moreover, the locked conformation of E reduced the exposure of epitopes recognized by cross-reactive antibodies and therefore showed a lower potential to induce ADEin vitro. Our data demonstrated a higher efficacy of the VLPs in comparison with the soluble dimer and support VLP-cvD as a promising ZIKV vaccine.

Author Summary

Infection with Zika virus (ZIKV) leads to the production by host of antibodies that target the viral surface envelope (E) protein. A subset of these antibodies can inhibit virus infection, thus making E as a suitable candidate for the development of vaccine against the virus. However, the anti-ZIKV E antibodies can cross-react with the E protein of the related dengue virus on account of the high level of similarity exhibited by the two viral proteins. Such a scenario may lead to severe dengue disease. Therefore, the design of a ZIKV vaccine requires particular care. Here, we tested two candidate vaccines containing a recombinant form of the ZIKV E protein that is forced in a covalently stable dimeric conformation (cvD). They were generated with an explicit aim to reduce the exposure of the cross-reactive epitopes. One vaccine is composed of a soluble form of the E protein (sE-cvD), the other is a more complex virus-like particle (VLP-cvD). We used the two candidate vaccines to immunize mice and later infected with ZIKV. The animals produced high level of inhibitory antibodies and were protected from the infection. The VLP-cvD was the most effective and we believe it represents a promising ZIKV vaccine candidate.
Transmission dynamics and control measures of COVID-19 outbreak in China: a modelling study
Authors: Zhang Xu-Sheng , Vynnycky Emilia , Charlett Andre , de Angelis Daniela , Chen Zhengji ,   PHE COVID-19 modelling group, Liu Wei
Published: July 2020

COVID-19 is reported to have been effectively brought under control in China at its initial start place. To understand the COVID-19 outbreak in China and provide potential lessons for other parts of the world, in this study we combine a mathematical modelling with multiple datasets to estimate its transmissibility and severity and how it was affected by the unprecedented control measures. Our analyses show that before 29thJanuary 2020, the ascertainment rate is 6.9%(95%CI: 3.5 – 14.6%); then it increased to 41.5%(95%CI: 30.6 – 65.1%). The basic reproduction number (R0) was 2.23(95%CI: 1.86 – 3.22) before 8thFebruary 2020; then it dropped to 0.04(95%CI: 0.01 – 0.10). This estimation also indicates that the effect on transmissibility of control measures taken since 23rdJanuary 2020 emerged about two weeks late. The confirmed case fatality rate is estimated at 4.41%(95%CI: 3.65 – 5.30%). This shows that SARS-CoV-2 virus is highly transmissible but less severe than SARS-CoV-1 and MERS-CoV. We found that at the early stage, the majority ofR0comes from the undetected infected people. This implies that the successful control in China was achieved through decreasing the contact rates among people in general populations and increasing the rate of detection and quarantine of the infected cases.
What can the ideal gas say about global pandemics? Reinterpreting the basic reproduction number
Authors: Dickson Bradley M.
Published: July 2020

Through analysis of the ideal gas, we construct a random walk that on average matches the standard susceptible-infective-removed (SIR) model. We show that the most widely referenced parameter, the “basic reproduction number” (ℛ0), is fundamentally connected to the relative odds of increasing or decreasing the infectives population. As a consequence, for ℛ0> 1 the probability that no outbreak occurs is 1/ℛ0. In stark contrast to a deterministic SIR, when ℛ0= 1.5 the random walk has a 67% chance of avoiding outbreak. Thus, an alternative, probabilistic, interpretation of ℛ0arises, which provides a novel estimate of the critical population densityγ/rwithout fitting SIR models. We demonstrate that SARS-CoV2 in the United States is consistent with our model and attempt an estiamte ofγ/r. In doing so, we uncover a significant source of bias in public data reporting. Data are aggregated on political boundaries, which bear no concern for dispersion of population density. We show that this introduces bias in fits and parameter estimates, a concern for understanding fundamental virus parameters and for policy making. Anonymized data at the resolution required for contact tracing would afford access toγ/rwithout fitting. The random walk SIR developed here highlights the intuition that any epidemic is stochastic and recovers all the key parameter values noted by Kermack and McKendrick in 1927.
“An exploratory Integrated Moving Average Time Series Model of the initial outbreak of COVID-19 in six (6) significantly impacted Countries”
Authors: Pascarella Joseph E. , Pascarella Elaina
Published: July 2020

The 2019 Novel Coronavirus SARS-CoV-2 (COVID-19) is a single-stranded RNA virus that has threatened the lives of humans all over the globe. Government officials, policy makers and public health officials have been scrambling and struggling to “flatten the curve” to decelerate the prevalence and spread of COVID-19 given the significant economic destruction of the spread of the virus. Most “flatten the curve” models are based on Compartmental Models. This preliminary research is based on six (6) selected countries significantly impacted by COVID-19 and endeavors to build a new model based on moving averages lagged at different time periods to better hone in on the time the COVID-19 begins to decelerate using the date of first reported case and date of first reported death. This new model, the Consistent Deceleration Model (CDM) is based on each individual country’s date of Peak Increase in Mortality Rate (PINC MR) and the Moving Average since the peak increase in mortality rate (MA POSTINC). The CDM can be utilized of one of many quantitative tools to determine the strength of the deceleration of an infectious outbreak.
A minimal model for household effects in epidemics
Authors: Huber Greg , Kamb Mason , Kawagoe Kyle , Li Lucy M. , Veytsman Boris , Yllanes David , Zigmond Dan
Published: July 2020

Shelter-in-place and other confinement strategies implemented in the current COVID-19 pandemic have created stratified patterns of contacts between people: close contacts within households and more distant contacts between the households. The epidemic transmission dynamics is significantly modified as a consequence.

We introduce a minimal model that incorporates these household effects in the framework of mean-field theory and numerical simulations. We show that the reproduction number R0 depends on the household size in a surprising way: linearly for relatively small households, and as a square root of size for larger households. We discuss the implications of the findings for the lockdown, test, tracing, and isolation policies.
Analysis of exome-sequenced UK Biobank subjects implicates genes affecting risk of hyperlipidaemia
Authors: Curtis David
Published: July 2020

Rare genetic variants in LDLR, APOB and PCSK9 are known causes of familial hypercholesterolaemia and it is expected that rare variants in other genes will also have effects on hyperlipidaemia risk although such genes remain to be identified. The UK Biobank consists of a sample of 500,000 volunteers and exome sequence data is available for 50,000 of them. 11,490 of these were classified as hyperlipidaemia cases on the basis of having a relevant diagnosis recorded and/or taking lipid-lowering medication while the remaining 38,463 were treated as controls. Variants in each gene were assigned weights according to rarity and predicted impact and overall weighted burden scores were compared between cases and controls, including population principal components as covariates. One biologically plausible gene, HUWE1, produced statistically significant evidence for association after correction for testing 22,028 genes with a signed log10 p value (SLP) of −6.15, suggesting a protective effect of variants in this gene. Other genes with uncorrected p<0.001 are arguably also of interest, including LDLR (SLP=3.67), RBP2 (SLP=3.14), NPFFR1 (SLP=3.02) and ACOT9 (SLP=-3.19). Gene set analysis indicated that rare variants in genes involved in metabolism and energy can influence hyperlipidaemia risk. Overall, the results provide some leads which might be followed up with functional studies and which could be tested in additional data sets as these become available. This research has been conducted using the UK Biobank Resource.
Intelligent Pneumonia Identification from Chest X-Rays: A Systematic Literature Review
Authors: Khan Wasif , Zaki Nazar , Ali Luqman
Published: July 2020

Chest radiography is an important diagnostic tool for chest-related diseases. Medical imaging research is currently embracing the automatic detection techniques used in computer vision. Over the past decade, Deep Learning techniques have shown an enormous breakthrough in the field of medical diagnostics. Various automated systems have been proposed for the rapid detection of pneumonia on chest x-rays images Although such detection algorithms are many and varied, they have not been summarized into a review that would assist practitioners in selecting the best methods from a real-time perspective, perceiving the available datasets, and understanding the currently achieved results in this domain. This paper overviews the current literature on pneumonia identification from chest x-ray images. After summarizing the topic, the review analyzes the usability, goodness factors, and computational complexities of the algorithms that implement these techniques. It also discusses the quality, usability, and size of the available datasets, and ways of coping with unbalanced datasets.
Identification of bioactive metabolites in human iPSC-derived dopaminergic neurons with PARK2 mutation: altered mitochondrial and energy metabolism
Authors: Okarmus Justyna , Havelund Jesper F. , Ryding Matias , Schmidt Sissel I. , Bogetofte Helle , Færgeman Nils J. , Hyttel Poul , Meyer Morten
Published: July 2020

PARK2(parkin) mutations cause early onset of autosomal recessively inherited Parkinson’s disease (PD). Parkin is an ubiquitin E3 ligase and has been reported to participate in several cellular functions, including mitochondrial homeostasis. However, the specific metabolomic changes caused by parkin depletion remain largely unknown. Human induced pluripotent stem cells (iPSCs) withPARK2knockout (KO) provide a valuable model for studying parkin dysfunction in dopaminergic neurons. In the current study, we used isogenic iPSCs to investigate the effect of parkin loss-of-function by comparative metabolomics analysis. The metabolomic profile of thePARK2KO neurons differed substantially from that of healthy controls. We found increased tricarboxylic acid (TCA) cycle activity, perturbed mitochondrial ultrastructure connected with ATP depletion, glycolysis dysregulation with lactate accumulation, and elevated levels of short- and long-chain carnitines. These mitochondrial and energy perturbations in thePARK2KO neurons were combined with increased levels of oxidative stress and a decreased anti-oxidative response. In conclusion, our data describe a unique metabolomic profile associated with parkin dysfunction, demonstrating several PD-related cellular defects. Our findings support and expand previously described PD phenotypic features and show that combining metabolomic analysis with an iPSC-derived dopaminergic neuronal model of PD is a valuable approach to obtain novel insight into the disease pathogenesis.
Panton Valentine leucocidin enhances community-acquired methicillin-resistantStaphylococcus aureuscolonisation of the gut
Authors: Couzon Florence , Bourgeois-Nicolaos Nadège , Benito Yvonne , Larroude Macarena , Tristan Anne , Rasigade Jean-Philippe , Diep Binh An , Doucet-Populaire Florence , Vandenesch François
Published: July 2020

Objective

Community-acquired methicillin resistantStaphylococcus aureus(CA-MRSA) independently emerged and became epidemic at the end of the 20thcentury. Since gut carriage was reported for CA-MRSA and since the common feature of historical CA-MRSA is to harbour Panton Valentine leucocidin (PVL), the question of the possible involvement of this toxin in gut carriage was investigated in mice and cellular models.

Methods

CA-MRSA of three lineages (USA300, USA1100, and ST80) and their isogenic Δpvlderivatives, were tested in competition for gut colonisation in mice and in a model of bacterial adhesion to mucus-producing intestinal epithelial cells.

Results

Mice inoculated with CA-MRSA and their Δpvlderivatives had their gut successfully colonised by the three lineages regardless of the presence of PVL; however, the wild type (WT) CA-MRSA outcompeted the Δpvlderivatives by at least 3 log after 40 days for all lineages tested.In vitrocompetition of CA-MRSA with their Δpvlderivatives showed no fitness disequilibrium after 6 weeks, ensuring that the results obtained in mice did not result from direct bacterial interference. Direct fluorescence assay of mice intestine showedS. aureuslocalised at the mucosal surface of the intestine and within the intestinal crypts, but not within epithelial cells, suggesting a bacterial tropism for the mucus layer. Significant difference in adhesion to intestinal epithelial cells between WT andpvlknockout was only observed on mucus-producing cells, and not on non-producing ones.

Conclusion

PVL enhances CA-MRSA gut colonisation in mice by a mechanism involving adhesion-colonisation of the mucus layer.
Optimization of On-Bead Emulsion Polymerase Chain Reaction Based on Single Particle Analysis
Authors: Siu Ryan H.P. , Liu Yang A. , Chan Kaitlin H. Y. , Ridzewski Clara , Slaughter Liane Siu , Wu Angela R.
Published: July 2020

Emulsion polymerase chain reaction (ePCR) enables parallel amplification of millions of different DNA molecules while avoiding bias and chimeric byproducts, essential criteria for applications including next generation sequencing, aptamer selection, and protein-DNA interaction studies. Despite these advantages, ePCR remains underused due to the lack of optimal starting conditions, straightforward methods to evaluate success, and guidelines for tuning the reaction. This knowledge has been elusive for bulk emulsion generation methods, such as stirring, the only methods that can emulsify libraries of ≥108sequences within minutes, because these emulsions have not been characterized in ways that preserve the heterogeneity that defines successful ePCR. Our study systematically quantifies the influence of tuning individual ePCR parameters by single particle analysis, which preserves this heterogeneity. We combine ePCR with magnetic microbeads and quantify the amplification yield via qPCR and the proportion of clonal and saturated beads via flow cytometry. Our single particle level analysis of thousands of beads resolves two key criteria that define the success of ePCR: 1) whether the target fraction of 20% clonal beads predicted by the Poisson distribution is achieved, and 2) whether those beads are partially or maximally covered by amplified DNA. We found that increasing the polymerase concentration 20-fold in ePCR over recommended concentrations for conventional PCR was necessary to obtain sufficient PCR products. Surprisingly, dramatical increases in the concentrations of reverse primer and nucleotides recommended in literature gave no measurable change in outcome. We thus provide evidence-based starting conditions for effective and economical ePCR for real DNA libraries and a straightforward workflow for evaluating the success of tuning ePCR prior to downstream applications.
Analysis of neuronal Ca2+handling properties by combining perforated patch clamp recordings and the added buffer approach
Authors: Hess Simon , Pouzat Christophe , Paeger Lars , Pippow Andreas , Kloppenburg Peter
Published: July 2020

Ca2+functions as an important intracellular signal for a wide range of cellular processes. These processes are selectively activated by controlled spatiotemporal dynamics of the free cytosolic Ca2+. Intracellular Ca2+dynamics are regulated by numerous cellular parameters. Here, we established a new way to determine neuronal Ca2+handling properties by combining the ‘added buffer’ approach (Neher and Augustine, 1992) with perforated patch-clamp recordings (Horn and Marty, 1988). Since the added buffer approach typically employs the standard whole-cell configuration for concentration-controlled Ca2+indicator loading, it only allows for the reliable estimation of the immobile fraction of intracellular Ca2+buffers. Furthermore, crucial components of intracellular signaling pathways are being washed out during prolonged whole-cell recordings, leading to cellular deterioration. By combining the added buffer approach with perforated patch-clamp recordings, these issues are circumvented, allowing the precise quantification of the cellular Ca2+handling properties, including immobile as well as mobile Ca2+buffers.
Widespread retention of ohnologs in key developmental gene families following whole genome duplication in arachnopulmonates
Authors: Harper Amber , Gonzalez Luis Baudouin , Schönauer Anna , Janssen Ralf , Seiter Michael , Holzem Michaela , Arif Saad , McGregor Alistair P. , Sumner-Rooney Lauren
Published: July 2020

Whole genome duplications have occurred multiple times during animal evolution, including in lineages leading to vertebrates, teleosts, horseshoe crabs and arachnopulmonates. These dramatic events initially produce a wealth of new genetic material, generally followed by extensive gene loss. It appears, however, that developmental genes such as homeobox genes, signalling pathway components and microRNAs are frequently retained as duplicates (so called ohnologs) following whole-genome duplication. These not only provide the best evidence for whole-genome duplication, but an opportunity to study its evolutionary consequences. Although these genes are well studied in the context of vertebrate whole-genome duplication, similar comparisons across the extant arachnopulmonate orders are patchy. We sequenced embryonic transcriptomes from two spider species and two amblypygid species and surveyed three important gene families, Hox, Wnt and frizzled, across these and twelve existing transcriptomic and genomic resources for chelicerates. We report extensive retention of putative ohnologs, including amblypygids, further supporting the ancestral arachnopulmonate whole-genome duplication. We also find evidence of consistent evolutionary trajectories in Hox and Wnt gene repertoires across three of the five arachnopulmonate orders, with inter-order variation in the retention of specific paralogs. We identify variation between major clades in spiders and are better able to reconstruct the chronology of gene duplications and losses in spiders, amblypygids, and scorpions. These insights shed light on the evolution of the developmental toolkit in arachnopulmonates, highlight the importance of the comparative approach within lineages, and provide substantial new transcriptomic data for future study.
Mechanical transmission of African swine fever virus byStomoxys calcitrans: insights from a mechanistic model
Authors: Timothée Vergne , Mathieu Andraud , Sarah Bonnet , Nick De Regge , Marc Desquesnes , Johanna Fite , Florence Etore , Mutien-Marie Garigliany , Ferran Jori , Laetitia Lempereur , Marie-Frédérique Le Potier , Elsa Quillery , Claude Saegerman , Laurence Vial , Emilie Bouhsira
Published: July 2020

African swine fever (ASF) represents a global threat with huge economic consequences for the swine industry. Even though direct contact is likely to be the main transmission route from infected to susceptible hosts, recent epidemiological investigations have raised questions regarding the role of hematophagous arthropods, in particular the stable fly (Stomoxys calcitrans). In this study, we developed a mechanistic vector-borne transmission model for ASF virus (ASFV) within an outdoor domestic pig farm in order to assess the relative contribution of stable flies to the spread of the virus. The model was fitted to the ecology of the vector, its blood-feeding behaviour and pig-to-pig transmission dynamic. Model outputs suggested that in a context of low abundance (<5 flies per pig), stable flies would play a minor role in the spread of ASFV, as they are expected to be responsible for around 10% of transmission events. However, with abundances of 20 and 50 stable flies per pig, the vector-borne transmission would likely be responsible for almost 30% and 50% of transmission events, respectively. In these situations, time to reach a pig mortality of 10% would be reduced by around 26% and 40%, respectively. The sensitivity analysis emphasised that the expected relative contribution of stable flies was strongly dependent on the volume of blood they regurgitated and the infectious dose for pigs. This study identified crucial knowledge gaps that need to be filled in order to assess more precisely the potential contribution of stable flies to the spread of ASFV, including a quantitative description of the populations of hematophagous arthropods that could be found in pig farms, a better understanding of blood-feeding behaviours of stable flies, and the quantification of the probability that stable flies partially fed with infectious blood transmits the virus to a susceptible pig during a subsequent blood-feeding attempt.
A Model of the Transition to Behavioral and Cognitive Modernity using Reflexively Autocatalytic Networks
Authors: Gabora Liane , Steel Mike
Published: July 2020

Abstract

This paper proposes a model of the cognitive mechanisms underlying the transition to behavioral and cognitive modernity in the Upper Paleolithic using autocatalytic networks. These networks have been used to model life’s origins. More recently, they have been applied to the emergence ofcognitivestructure capable of undergoingculturalevolution. Mental representations of knowledge and experiences play the role of catalytic molecules, the interactions among them (e.g., the forging of new associations or affordances) play the role of reactions, and thought processes are modeled as chains of these interactions. We posit that one or more genetic mutations may have allowed thought to be spontaneously tailored to the situation by modulating the degree of (1) divergence (versus convergence), (2) abstractness (versus concreteness), and (3) context-specificity. This culminated in persistent, unified autocatalytic semantic networks that bridged previously compartmentalized knowledge and experience. We explain the model using one of the oldest-known uncontested examples of figurative art: the carving of the Hohlenstein-Stadel Löwenmensch, or lion-man. The approach keeps track of where in a cultural lineage each innovation appears, and models cumulative change step by step. It paves the way for a broad scientific framework for the origins of both biological and cultural evolutionary processes.
Functional and structural characterization of allosteric activation of Phospholipase Cε by Rap1A
Authors: Sieng Monita , Selvia Arielle F. , Garland-Kuntz Elisabeth E. , Hopkins Jesse B. , Fisher Isaac J. , Marti Andrea T. , Lyon Angeline M.
Published: July 2020

Phospholipase Cε (PLCε) is activated downstream of G protein-coupled receptors (GPCRs) and receptor tyrosine kinases (RTKs) through direct interactions with small GTPases, including Rap1A and Ras. While Ras has been reported to allosterically activate the lipase, it is not known whether Rap1A has the same ability, or what its molecular mechanism might be. Rap1A activates PLCε in response to the stimulation of β-adrenergic receptors (β-ARs), translocating the complex to the perinuclear membrane. Because the C-terminal Ras association (RA2) domain of PLCε was proposed to the primary binding site for Rap1A, we first confirmed using purified proteins that the RA2 domain is indeed essential for activation by Rap1A. However, we also showed that the PLCε pleckstrin homology (PH) domain and first two EF hands (EF1/2) are required for Rap1A activation, and identified hydrophobic residues on the surface of the RA2 domain that are also necessary for activation by the GTPase. Finally, small angle X-ray scattering (SAXS) showed that Rap1A binding induces and stabilizes discrete conformational states in PLCε variants that can be activated by the GTPase. This data, together with the recent structure of a catalytically active fragment of PLCε, provide the first evidence that Rap1A, and by extension Ras, allosterically activate the lipase by promoting and stabilizing interactions between the RA2 domain and the PLC core.
Selective cross-linking of coinciding protein assemblies by in-gel cross-linking mass-spectrometry
Authors: Hevler Johannes F. , Lukassen Marie V. , Cabrera-Orefice Alfredo , Arnold Susanne , Pronker Matti F. , Franc Vojtech , Heck Albert J.R.
Published: July 2020

Cross-linking mass spectrometry has developed into an important method to study protein structures and interactions. The in-solution cross-linking workflows involve time and sample consuming steps and do not provide sensible solutions for differentiating cross-links obtained from co-occurring protein oligomers, complexes, or conformers. Here we developed a cross-linking workflow combining blue native PAGE with in-gel cross-linking mass spectrometry (IGX-MS). This workflow circumvents steps, such as buffer exchange and cross-linker concentration optimization. Additionally, IGX-MS enables the parallel analysis of co-occurring protein complexes using only small amounts of sample. Another benefit of IGX-MS observed by experiments on GroEL and purified bovine heart mitochondria, is the substantial reduction of artificial over-length cross-links when compared to in-solution cross-linking. We next used IGX-MS to investigate the complement components C5, C6, and their hetero-dimeric C5b6 complex. The obtained cross-links were used to generate a refined structural model of the complement component C6, resembling C6 in its inactivated state. This finding shows that IGX-MS can be used to provide new insights into the initial stages of the terminal complement pathway.
Evaluation ofTH-Cre knock-in cell lines for detection and specific targeting of stem cell-derived dopaminergic neurons
Authors: Fiorenzano A , Nelander Wahlestedt J , Parmar M
Published: July 2020

The focal and progressive degeneration of dopaminergic (DA) neurons in ventral midbrain has made Parkinson’s disease (PD) a particularly interesting target of cell-based therapies. However, ethical issues and limited tissue availability have so far hindered the widespread use of human fetal tissue in cell-replacement therapy. DA neurons derived from human pluripotent stem cells (hPSCs) offer unprecedented opportunities to access a renewable source of cells suitable for PD therapeutic applications. To better understand the functional properties of stem-cell derived DA neurons, we generated targeted hPSC lines with the gene coding for Cre recombinase knocked into theTHlocus. When combined with flexed GFP, they serve as reporter cell lines able to identify and isolate TH+ neuronsin vitroand after transplantationin vivo. TheseTH-Cre lines provide a valuable genetic tool to manipulate DA neurons useful for the design of more precise DA differentiation protocols and the study of these cells after transplantation in pre-clinical animal models of PD.
Intermolecular Channels Direct Crystal Orientation in Mineralized Collagen
Authors: Xu YiFei , Nudelman Fabio , Eren E. Deniz , Wirix Maarten J. M. , Cantaert Bram , Nijhuis Wouter H. , Hermida-Merino Daniel , Portale Giuseppe , Bomans Paul H. H. , Ottmann Christian , Friedrich Heiner , Bras Wim , Akiva Anat , Orgel Joseph P. R. O. , Meldrum Fiona C. , Sommerdijk Nico A. J. M.
Published: July 2020

The mineralized collagen fibril is the basic building block of bone, commonly pictured as a parallel array of ultrathin carbonated hydroxyapatite (HAp) platelets distributed throughout the collagen. This orientation is often attributed to an epitaxial relationship between the HAp and collagen molecules inside 2D voids within the fibril. Although recent studies have questioned this model, the structural relationship between the collagen matrix and HAp, and the mechanisms by which collagen directs mineralization remain unclear. Here, we use XRD to reveal that the voids in the collagen are in fact cylindrical pores with diameters of ∼2 nm, while electron microscopy shows that the HAp crystals in bone are only uniaxially oriented with respect to the collagen. Fromin vitromineralization studies with HAp, CaCO3and γ-FeOOH we conclude that confinement within these pores, together with the anisotropic growth of HAp, dictates the orientation of HAp crystals within the collagen fibril.
Discovery of widespread transcription initiation at microsatellites predictable by sequence-based deep neural network
Authors: Grapotte Mathys , Saraswat Manu , Bessière Chloé , Menichelli Christophe , Ramilowski Jordan A. , Severin Jessica , Hayashizaki Yoshihide , Itoh Masayoshi , Tagami Michihira , Murata Mitsuyoshi , Kojima-Ishiyama Miki , Noma Shohei , Noguchi Shuhei , Kasukawa Takeya , Hasegawa Akira , Suzuki Harukazu , Nishiyori-Sueki Hiromi , Frith Martin C. ,   FANTOM consortium, Chatelain Clément , Carninci Piero , de Hoon Michiel J.L. , Wasserman Wyeth W. , Bréhélin Laurent , Lecellier Charles-Henri
Published: July 2020

Using the Cap Analysis of Gene Expression (CAGE) technology, the FANTOM5 consortium provided one of the most comprehensive maps of Transcription Start Sites (TSSs) in several species. Strikingly, ~ 72% of them could not be assigned to a specific gene and initiate at unconventional regions, outside promoters or enhancers. Here, we probed these unassigned TSSs and showed that, in all species studied, a significant fraction of CAGE peaks initiate at microsatellites, also called short tandem repeats (STRs). To confirm this transcription, we developed Cap Trap RNA-seq, a technology which combines cap trapping and long reads MinION sequencing. We trained sequence-based deep learning models able to predict CAGE signal at STRs with high accuracy. These models unveiled the importance of STR surrounding sequences not only to distinguish STR classes, as defined by the repeated DNA motif, one from each other, but also to predict their transcription. Excitingly, our models predicted that genetic variants linked to human diseases affect STR-associated transcription and correspond precisely to the key positions identified by our models to predict transcription. Together, our results extend the repertoire of non-coding transcription associated with DNA tandem repeats and complexify STR polymorphism.
KLF4 Recruits SWI/SNF to Increase Chromatin Accessibility and Reprogram the Endothelial Enhancer Landscape under Laminar Shear Stress
Authors: Moonen Jan-Renier A.J. , Chappell James , Shi Minyi , Shinohara Tsutomu , Li Dan , Mumbach Maxwell R. , Zhang Fan , Nasser Joseph , Mai Daniel H. , Taylor Shalina , Wang Lingli , Metzger Ross J. , Chang Howard Y. , Engreitz Jesse M. , Snyder Michael P. , Rabinovitch Marlene
Published: July 2020

Physiologic laminar shear stress (LSS) induces an endothelial gene expression profile that is vasculo-protective. In this report, we delineate how LSS mediates changes in the epigenetic landscape to promote this beneficial response. We show that under LSS, KLF4 interacts with the SWI/SNF nucleosome remodeling complex to increase accessibility at enhancer sites that promote expression of homeostatic endothelial genes. By combining molecular and computational approaches we discovered enhancers that loop to promoters of known and novel KLF4- and LSS-responsive genes that stabilize endothelial cells and suppress inflammation, such asBMPR2andDUSP5. By linking enhancers to genes that they regulate under physiologic LSS, our work establishes a foundation for interpreting how non-coding DNA variants in these regions might disrupt protective gene expression to influence vascular disease.
Phylogeny and biogeography ofCeibaMill. (Malvaceae, Bombacoideae)
Authors: Pezzini Flávia Fonseca , Dexter Kyle G. , de Carvalho-Sobrinho Jefferson G. , Kidner Catherine A. , Nicholls James A. , de Queiroz Luciano P. , Pennington R. Toby
Published: July 2020

The Neotropics is the most species-rich area in the world and the mechanisms that generated and maintain its biodiversity are still debated. This paper contributes to the debate by investigating the evolutionary and biogeographic history of the genusCeibaMill. (Malvaceae: Bombacoideae).Ceibacomprises 18 mostly neotropical species endemic to two major biomes, seasonally dry tropical forests (SDTFs) and rain forests and its species are one of the most characteristic elements of neotropical SDTF, one of the most threatened biomes in the tropics. Phylogenetic analyses of DNA sequence data from the nuclear ribosomal internal transcribed spacers (ITS) for 30 accessions representing 14 species ofCeibarecovered the genus as monophyletic and showed geographical and ecological structure in three main clades: (i) a rain forest lineage of nine accessions ofC. pentandrasister to the remaining species; (ii) a highly supported clade composed ofC. schottiiandC. aesculifoliafrom Central American and Mexican SDTF plus two accessions ofC. samaumafrom inter Andean valleys from Peru; and (iii) a highly supported South American SDTF clade including 10 species showing little sequence variation. Within this South American clade, no species represented by multiple accessions were resolved as monophyletic. We demonstrate that the patterns of species age, monophyly and geographical structure previously reported for SDTF species within the Leguminosae family are not shared byCeiba, suggesting that further phylogenetic studies of unrelated groups are required.

HIGHLIGHTS

This paper provides a well sampled phylogeny of the iconic genusCeiba, one of the most characteristic tree genera of neotropical Seasonally Dry Tropical Forest (SDTF).

There is a clear phylogenetic signal for biome preference and geographic structure inCeiba.

We estimate a mid Miocene origin forCeiba, with the stem node age of the genus estimated at 21.1 Ma and the crown node age at 12.7 Ma.

Ceibaspecies have young stem ages in the SDTF clade but old stem ages in rain forest species.

Patterns of species age, monophyly, ecological and geographical structure reported for SDTF species are only partially shared byCeiba, an iconic genus of neotropical SDTF.
Metabolic features of mouse and human retinas: rods vs. cones, macula vs. periphery, retina vs. RPE
Authors: Li Bo , Zhang Ting , Liu Wei , Wang Yekai , Xu Rong , Zeng Shaoxue , Zhang Rui , Zhu Siyan , Gillies Mark C , Zhu Ling , Du Jianhai
Published: July 2020

Photoreceptors, especially cones, which are enriched in the human macula, have high energy demands, making them vulnerable to metabolic stress. Metabolic dysfunction of photoreceptors and their supporting retinal pigment epithelium (RPE) is an important underlying cause of degenerative retinal diseases. However, how cones and the macula support their exorbitant metabolic demand and communicate with RPE is unclear. By profiling metabolite uptake and release and analyzing metabolic genes, we have found cone-rich retinas and human macula share specific metabolic features with upregulated pathways in pyruvate metabolism, mitochondrial TCA cycle and lipid synthesis. Human neural retina and RPE have distinct but complementary metabolic features. Retinal metabolism centers on NADH production and neurotransmitter biosynthesis. The retina needs aspartate to sustain its aerobic glycolysis and mitochondrial metabolism. RPE metabolism is directed toward NADPH production and biosynthesis of acetyl-rich metabolites, serine and others. RPE consumes multiple nutrients, including proline, to produce metabolites for the retina.
ThePeroxiredoxin 6gene plays a critical role in the homeostatic regulation of fear response
Authors: Phasuk Sarayut , Pairojana Tanita , Suresh Pavithra , Huang Shun-Ping , Pakaprot Narawut , Chompoopong Supin , Yang Chee-Hing , Chang Hsueh-Kai , Chen Chien-Chang , Liu Ingrid Y.
Published: July 2020

Peroxiredoxin 6 (PRDX6) is a multifunctional enzyme implicated in redox regulation and expressed in many organs including the brain. It is known to participate in many psychiatric functions, but its role in fear memory is unknown. The present study demonstrates thatPRDX6plays a critical role in the regulation of fear response. UsingPrdx6knockout (Prdx6−/−) mice, we identified that PRDX6 acts as a suppressor in fear memory formation. Lack ofPrdx6leads to the faster fear acquisition and enhanced contextual fear response. This phenomenon was confirmed by the fact that injection of lentivirus-carried human PRDX6-V5 into the hippocampus ofPrdx6−/−mice restored the enhanced fear response to the wild-type level. In the hippocampus ofPrdx6−/−mice, calcium-dependent PLA2 level was increased, which may compensate for the lack of aiPLA2 function to maintain normal synaptic membranes. On the other hand, reactive oxygen species (ROS) levels did not change, indicating loss of peroxidase function did not affect the regulation of fear response.
ASIC1a is required for neuronal activation via low-intensity ultrasound stimulation in mouse brain
Authors: Lim Jormay , Chu Ya-Cherng , Hao Chen-Ming , Liao Wei-Hao , Lin Shao-Shien , Hsu Sherry , Tai Hsiao-Hsin , Chien Ya-Chih , Lai Dar-Ming , Chen Wen-Shiang , Chen Chih-Cheng , Wang Jaw-Lin
Published: July 2020

Accumulating evidence has shown transcranial low-intensity ultrasound can be potentially a non-invasive neural modulation tool to treat brain diseases. However, the underlying mechanism remains elusive, because the majority of studies on animal models applying rather high-intensity ultrasound that cannot be safely used in humans. Here we showed low-intensity ultrasound was able to activate neurons in the mouse brain and repeated ultrasound stimulation resulted in adult neurogenesis in specific brain regions.In vitrocalcium imaging studies showed that a specific ultrasound stimulation mode, which combined with both ultrasound-induced pressure and acoustic streaming mechanotransduction, is required to activate cultured cortical neurons. ASIC1a and the tether-mode mechanotransduction were involved in the low-intensity ultrasound-mediated mechanotransduction and cultured neuron activation, which was inhibited by ASIC1a blockade and cytoskeleton-modified agents. In contrast, the inhibition of mechanical sensitive channels involved in bilayer-model mechanotransduction like Piezo or TRP proteins did not affect the ultrasound-mediated neuronal activation.

SIGNIFICANCE

CNS neurons have no sensory function, protected by the skull. For this reason, brain neuromodulation by ultrasound were either done at a high intensity or through auditory nerves. We demonstrate in this study CNS neurons react to ultrasound stimulation at an intensity (5 mW/cm2) far lower than typical therapeutic ultrasound (>30 mW/cm2). Using micropipette ultrasound in calcium imaging, we show the reactions of CNS neurons to ultrasound is through ASIC1a channels, pointing to the molecular basis for direct ultrasound neuromodulation at low intensity. Furthermore, we also show evidence of neurogenesis with the same ultrasound stimulation, suggesting potential therapeutic translation.
Ral GTPases promote metastasis by controlling biogenesis and organ colonization of exosomes
Authors: Ghoroghi S , Mary B , Larnicol A , Klein A , Osmani N , Busnelli I , Delalande F , Paul N , Halary S , Gros F , Fouillen L , Haeberle AM , Royer C , Spiegelhalter C , André-Grégoire G , Murphy K , Timpson P , Carapito R , Blot-Chabaud M , Gavard J , Carapito C , Vitale N , Lefebvre O , Goetz JG , Hyenne V
Published: July 2020

Cancer extracellular vesicles (EVs) mainly exert pro-tumoral functions by changing the phenotypes of stromal cells to the benefit of tumor growth and metastasis. They shuttle to distant organs and fertilize pre-metastatic niches facilitating subsequent seeding by circulating tumor cells. The levels of tumor secreted EVs correlate with tumor aggressiveness, however, the link between EV secretion mechanisms and their capacity to form pre-metastatic niches remains obscure. Here, we show that GTPases of the Ral family control, through the phospholipase D1, multi-vesicular bodies homeostasis and thereby tune the biogenesis and secretion of pro-metastatic EVs. RalA and RalB promote lung metastasis in a syngeneic mouse model. Importantly, EVs from RalA or RalB depleted cells have limited organotropic capacities in vivo and, as a consequence, are less efficient in promoting lung metastasis. RalA or RalB modulate the EV levels of the adhesion molecule MCAM/CD146, which mediates lung colonization. Finally, RalA and RalB, but also MCAM/CD146, are factors of poor prognosis in human breast cancer patients. Altogether, our study identifies Ral GTPases as central molecules linking the mechanisms of EVs secretion, cargo loading to their capacity to disseminate and induce pre-metastatic niches.
Species coexistence and overlapping distributions in a metacommunity are compatible with niche differences and competition at a local scale
Authors: Dubart Maxime , David Patrice , Ben-Ami Frida , Haag Christoph R. , Pajunen V. Ilmari , Ebert Dieter
Published: July 2020

Niche partitioning is the most studied factor structuring communities of competing species. In fragmented landscapes, however, a paradox can exist: different taxa may competitively dominate different types of habitat patches, resulting in a form of spatial niche partitioning, yet differences in long-term distributions among species can appear surprisingly small. This paradox is illustrated by an emblematic metacommunity - that ofDaphniaspp. in rockpools on the Finnish Baltic coast, where three species compete with each other, have distinct ecological preferences, yet largely overlap in long-term distributions. Here we examine how metacommunity models that explicitly estimate species-specific demographic parameters can solve the apparent paradox. Our research confirms previous studies that local extinction rates are influenced by environmental variables in a strong and species-specific way and are considerably increased by interspecific competition. Yet, our simulations show that this situation exists alongside interspecific differences in realized niches that are, overall, small, and identified three main explanations for this compatibility. Our results illustrate how state-space modelling can clarify complex metacommunity dynamics and explain why local competition and niche differentiation do not always scale up to the landscape level.
Acoel single-cell transcriptomics: cell-type analysis of a deep branching bilaterian
Authors: Duruz Jules , Kaltenrieder Cyrielle , Ladurner Peter , Bruggmann Rémy , Martìnez Pedro , Sprecher Simon G.
Published: July 2020

Bilaterian animals display a wide variety of cell types, organized into defined anatomical structures and organ systems, which are mostly absent in pre-bilaterian animals. Xenacoelomorpha are an early-branching bilaterian phylum displaying an apparently relatively simple anatomical organization that have greatly diverged from other bilaterian clades. In this study, we use whole-body single-cell transcriptomics on the acoelIsodiametra pulchrato identify and characterize different cell types. Our analysis identifies the existence of ten major cell-type categories in acoels all contributing to main biological functions of the organism: metabolism, locomotion and movements, behavior, defense and development. Interestingly, while most cell clusters express core fate markers shared with other animal clades, we also describe a surprisingly large number of clade-specific marker genes, suggesting the emergence of clade-specific common molecular machineries functioning in distinct cell types. Together, these results provide novel insight into the evolution of bilaterian cell-types and open the door to a better understanding of the origins of the bilaterian body plan and their constitutive cell types.