bluuebunny/crossref_metadata_2025 huggingface dataset:
Yolk sac erythromyeloid progenitors sustain erythropoiesis throughout embryonic life
Authors: Soares-da-Silva Francisca , Burlen-Defranoux Odile , Elsaid Ramy , Iturri Lorea , Freyer Laina , Sismeiro Odile , Pinto-do-Ó Perpétua , Gomez-Perdiguero Elisa , Cumano Ana
Published: March 2020

The first hematopoietic cells are produced in the yolk sac and are thought to be rapidly replaced by the progeny of hematopoietic stem cells. Here we document that hematopoietic stem cells do not contribute significantly to erythrocyte production up until birth. Lineage tracing of yolk sac-derived erythromyeloid progenitors, that also contribute to tissue resident macrophages, shows a progeny of highly proliferative erythroblasts, that after intra embryonic injection, rapidly differentiate. These progenitors, similar to hematopoietic stem cells, arec-Mybdependent and are developmentally restricted as they are not found in the bone marrow. We show that erythrocyte progenitors of yolk sac origin require lower concentrations of erythropoietin than their hematopoietic stem cell-derived counterparts for efficient erythrocyte production. Consequently, fetal liver hematopoietic stem cells fail to generate megakaryocyte and erythrocyte progenitors. We propose that large numbers of yolk sac-derived erythrocyte progenitors have a selective advantage and efficiently outcompete hematopoietic stem cell progeny in an environment with limited availability of erythropoietin.
Manganese complex [Mn(CO)3(tpa-κ3N)]Br increases antibiotic sensitivity in multidrug resistantStreptococcus pneumoniae
Authors: Liakopoulos Apostolos , La Ragione Roberto M. , Nagel Christoph , Schatzschneider Ulrich , Rozen Daniel E. , Betts Jonathan W.
Published: March 2020

The emergence of multidrug-resistance (MDR) inStreptococcus pneumoniaeclones and non-vaccine serotypes is of increasing concern, necessitating the development of novel treatment strategies. Here, we determined the efficacy of the Mn complex [Mn(CO)3(tpa-κ3N)]Br against MDRS. pneumoniaestrains. Our data showed that [Mn(CO)3(tpa-κ3N)]Br hasin vitroandin vivoantibacterial activity and has the potential to be used in combination with currently available antibiotics to increase their effectiveness against MDRS. pneumoniae.
Spatially-Resolved Live Cell Tagging and Isolation Using Protected Photoactivatable Cell Dyes
Authors: Genshaft Alex S , Ziegler Carly G. K. , Tzouanas Constantine N. , Mead Benjamin E. , Jaeger Alex M. , Navia Andrew W. , King Ryan P. , Jacks Tyler , Van Humbeck Jeffrey F. , Shalek Alex K.
Published: March 2020

Whether culturedin vitroor part of a complex tissuein vivo, a cell’s phenotype and function are significantly influenced by dynamic interactions with its microenvironment. To explicitly examine how a cell’s spatiotemporal activity impacts its behavior, we developed and validated a strategy termed SPACECAT—Spatially PhotoActivatable Color Encoded Cell Address Tags—to annotate, track, and isolate specific cells in a non-destructive, viability-preserving manner. In SPACECAT, a biological sample is immersed in a photocaged fluorescent molecule, and cells within a location of interest are labeled for further study by uncaging that molecule with user-patterned near-UV light. SPACECAT offers high spatial precision and temporal stability across diverse cell and tissue types, and is compatible with common downstream assays, including flow cytometry and single-cell RNA-Seq. Illustratively, we leveraged this approach in patient-derived intestinal organoids, a spatially complex system less amenable to genetic manipulations, to select for crypt-like regions enriched in stem-like and actively mitotic cells. Moreover, we demonstrate its applicability and utility onex vivotissue sections from four healthy organs and an autochthonous lung tumor model, uncovering spatially-biased gene expression patterns among immune cell subsets and identifying rare myeloid phenotypes enriched around tumor/healthy border regions. In sum, our method provides a minimally invasive and broadly applicable approach to link cellular spatiotemporal features and/or behavioral phenotypes with diverse downstream assays, enabling fundamental insights into the connections between tissue microenvironments and biological (dys)function.
Population Specific Adaptations in Venom Production to Abiotic Stressors in a Widely Distributed Cnidarian
Authors: Sachkova Maria Y. , Macrander Jason , Surm Joachim M. , Aharoni Reuven , Menard-Harvey Shelcie S. , Klock Amy , Leach Whitney B. , Reitzel Adam M. , Moran Yehu
Published: March 2020

Nematostella vectensisis a sea anemone (Actiniaria, Cnidaria) inhabiting estuaries over a broad geographic range where environmental conditions such as temperatures and salinity vary widely. In cnidarians, antagonistic interactions with predators and prey are mediated by their venom, which may be metabolically expensive. In this study, we challengedNematostellapolyps with heat, salinity, UV light stressors and a combination of all three to determine how abiotic stressors impact toxin expression for individuals collected across this species’ range. Transcriptomics and proteomics revealed that the highly abundant toxin Nv1 was the most downregulated gene under heat stress conditions in multiple populations. Physiological measurements demonstrated that venom is metabolically costly to produce suggesting that downregulating venom expression under stressful conditions may be advantageous. Strikingly, under a range of abiotic stressors, individuals from different geographic locations along this latitudinal cline modulate venom production levels differently in a pattern reflecting local adaptation.
Thecis-regulatory codes of response to combined heat and drought stress inArabidopsis thaliana
Authors: Azodi Christina B. , Lloyd John P. , Shiu Shin-Han
Published: March 2020

Plants respond to their environment by dynamically modulating gene expression. A powerful approach for understanding how these responses are regulated is to integrate information aboutcis-regulatory elements (CREs) into models calledcis-regulatory codes. Transcriptional response to combined stress is typically not the sum of the responses to the individual stresses. However,cis-regulatory codes underlying combined stress response have not been established. Here we modeled transcriptional response to single and combined heat and drought stress inArabidopsis thaliana. We grouped genes by their pattern of response (independent, antagonistic, synergistic) and trained machine learning models to predict their response using putative CREs (pCREs) as features (median F-measure = 0.64). We then developed a deep learning approach to integrate additional omics information (sequence conservation, chromatin accessibility, histone modification) into our models, improving performance by 6.2%. While pCREs important for predicting independent and antagonistic responses tended to resemble binding motifs of transcription factors associated with heat and/or drought stress, important synergistic pCREs resembled binding motifs of transcription factors not known to be associated with stress. These findings demonstrate howin silicoapproaches can improve our understanding of the complex codes regulating response to combined stress and help us identify prime targets for future characterization.
Models for Eco-evolutionary Extinction Vortices and their Detection
Authors: Nabutanyi Peter , Wittmann Meike J.
Published: March 2020

The smaller a population is, the faster it looses genetic variation due to genetic drift. Loss of genetic variation can reduce population growth rate, making populations even smaller and more vulnerable to loss of genetic variation, and so on. Ultimately, the population can be driven to extinction by this “eco-evolutionary extinction vortex”. So far, extinction vortices due to loss of genetic variation have been mainly described verbally. However, quantitative models are needed to better understand when such vortices arise and to develop methods for detecting them. Here we propose quantitative eco-evolutionary models, both individual-based simulations and analytic approximations, that link loss of genetic variation and population decline. Our models assume stochastic population dynamics and multi-locus genetics with different forms of balancing selection. Using mathematical analysis and simulations, we identify parameter combinations that exhibit strong interactions between population size and genetic variation as populations decline to extinction and match our definition of an eco-evolutionary vortex, i.e. the per-capita population decline rates and per-locus fixation rates increase with decreasing population size and number of polymorphic loci. We further highlight cues and early warning signals that may be useful in identifying populations undergoing an eco-evolutionary extinction vortex.
The pd-l1/pd-1 axis blocks neutrophil cytotoxicity in cancer
Authors: Gershkovitz Maya , Yajuk Olga , Fainsod-Levi Tanya , Granot Zvi
Published: March 2020

The PD-L1/PD-1 axis was shown to promote tumor growth and progression via the inhibition of anti-tumor immunity. Blocking this axis was shown to be beneficial in maintaining the anti-tumor functions of the adaptive immune system. Still, the consequences of blocking the PD-L1/PD-1 axis on innate immune responses remain largely unexplored. In this context, neutrophils were shown to consist of different subpopulations, which possess either pro- and anti-tumor properties. PD-L1-expressing neutrophils are considered pro-tumor as they can suppress cytotoxic T-cells. That said, we found that PD-L1 expression is not limited to tumor promoting neutrophils but is also evident in anti-tumor neutrophils. We show that neutrophil cytotoxicity is effectively and efficiently blocked by tumor cell expressed PD-1. Furthermore blocking of either neutrophil PD-L1 or tumor cell PD-1 maintains neutrophil cytotoxicity resulting in enhanced tumor cell apoptosis. Importantly, we show that tumor cell PD-1 blocks neutrophil cytotoxicity and promotes tumor growth via a mechanism independent of adaptive immunity. Taken together, these findings highlight the therapeutic potential of enhancing anti-tumor innate immune responses via blocking of the PD-L1/PD-1 axis.
Emergence ofoptrA-mediated linezolid resistance in multiple lineages and plasmids ofEnterococcus faecalisrevealed by long read sequencing
Authors: McHugh Martin P , Parcell Benjamin J , Pettigrew Kerry A , Toner Geoff , Khatamzas Elham , Karcher Anne Marie , Walker Joanna , Weir Robert , Meunier Danièle , Hopkins Katie L , Woodford Neil , Templeton Kate E , Gillespie Stephen H , Holden Matthew TG
Published: March 2020

Objectives

To characterise the genetic environment ofoptrAin linezolid-resistantEnterococcus faecalisisolates from Scotland.

Methods

Linezolid-resistantE. faecaliswere identified in three Scottish Health Boards and confirmed to carry theoptrAgene at the national reference laboratory. WGS was performed with short read (Illumina MiSeq) and long read (Oxford Nanopore MinION) technologies to generate complete genome assemblies. Illumina reads for 94E. faecalisbloodstream isolates were used to place theoptrA-positive isolates in a larger UK phylogeny.

Results

SixoptrA-positive linezolid-resistantE. faecaliswere isolated from urogenital samples in three Scottish Health Boards (2014-2017). No epidemiological links were identified between the patients, four were community-based, and only one had recent linezolid exposure. Reference-based mapping confirmed the isolates were genetically distinct (>13,900 core SNPs).optrAwas located on a plasmid in each isolate and these plasmids showed limited nucleotide similarity. There was variable presence of transposable elements surroundingoptrA, (including IS1216, IS3, and Tn3) and not always as a recognisable gene cassette. OptrA amino acid sequences were also divergent, resulting in four protein variants differing in 1-20 residues. One isolate belonged to ST16 and clustered with three other isolates in the UK collection (76-182 SNPs), otherwise theoptrA-positive isolates were genetically distinct from the bloodstream isolates (>6,000 SNPs).

Conclusions

We report multiple variants of the linezolid resistance geneoptrAin diverseE. faecalisstrain and plasmid backgrounds, suggesting multiple introductions of the gene into theE. faecalispopulation and selection driving recent emergence.
Adding carbon fiber to shoe soles does not improve running economy: a muscle-level explanation
Authors: Beck Owen N. , Golyski Pawel R. , Sawicki Gregory S.
Published: March 2020

In an attempt to improve their distance-running performance, many athletes race with carbon fiber plates embedded in their shoe soles. Accordingly, we sought to establish whether, and if so how, adding carbon fiber plates to shoes soles reduces athlete aerobic energy expenditure during running (improves running economy). We tested 15 athletes during running at 3.5 m/s in four footwear conditions that varied in shoe sole carbon fiber plate bending stiffness. For each condition, we quantified athlete aerobic energy expenditure and performed biomechanics analyses, which included the use of ultrasound imaging to examine soleus muscle dynamicsin vivo. Overall, increased footwear bending stiffness lengthened ground contact time (p=0.048), but did not affect ankle (p≥0.060), knee (p≥0.128), or hip (p≥0.076) joint angles or moments. Additionally, increased footwear bending stiffness did not affect muscle activity (all seven measured leg muscles (p≥0.146)), stride averaged active soleus volume, (p=0.068) or aerobic power (p=0.458) during running. Hence, footwear bending stiffness does not appear to alter the volume of aerobic energy consuming muscle in the soleus, or any other leg muscle, during running. Therefore, adding carbon fiber plates to shoe soles slightly alters whole-body and calf muscle biomechanics but does not improve running economy.
Mitochondrial DNA Sequence Diversity in Mammals: a Correlation Between the Effective and Census Population Sizes
Authors: James Jennifer , Eyre-Walker Adam
Published: March 2020

What determines the level of genetic diversity of a species remains one of the enduring problems of population genetics. Since, neutral diversity depends upon the product of the effective population size and mutation rate there is an expectation that diversity should be correlated to measures of census population size. This correlation is often observed for nuclear but not for mitochondrial DNA. Here we revisit the question of whether mitochondrial DNA sequence diversity is correlated to census population size by compiling the largest dataset to date from 639 mammalian species. In a multiple regression we find that nucleotide diversity is significantly correlated to both range size and mass-specific metabolic rate, but not a variety of other factors. We also find that a measure of the effective population size, the ratio of non-synonymous to synonymous diversity, is also significantly negatively correlated to both range and mass-specific metabolic rate. These results together suggest that species with larger ranges have larger effective population sizes. The slope of the relationship between diversity and range is such that doubling the range increases diversity by 12 to 20%, providing one of the first quantifications of the relationship between effective and census population sizes.
Ruler elements in chromatin remodelers set nucleosome array spacing and phasing
Authors: Oberbeckmann Elisa , Niebauer Vanessa , Watanabe Shinya , Farnung Lucas , Moldt Manuela , Schmid Andrea , Cramer Patrick , Peterson Craig L. , Eustermann Sebastian , Hopfner Karl-Peter , Korber Philipp
Published: March 2020

Arrays of regularly spaced nucleosomes dominate chromatin and are often phased by alignment to reference sites like active promoters. How the distances between nucleosomes (spacing), and between phasing sites and nucleosomes are determined remains unclear, and specifically, how ATP dependent chromatin remodelers impact these features. Here, we used genome-wide reconstitution to probe howSaccharomyces cerevisiaeATP dependent remodelers generate phased arrays of regularly spaced nucleosomes. We find that remodelers bear a functional element named the ‘ruler’ that determines spacing and phasing in a remodeler-specific way. We use structure-based mutagenesis to identify and tune the ruler element residing in the Nhp10 and Arp8 modules of the INO80 remodeler complex. Generally, we propose that a remodeler ruler regulates nucleosome sliding direction bias in response to (epi)genetic information. This finally conceptualizes how remodeler-mediated nucleosome dynamics determine stable steady-state nucleosome positioning relative to other nucleosomes, DNA bound factors, DNA ends and DNA sequence elements.
Large-Scale Analysis of Cell Death Phenotypic Heterogeneity
Authors: Inde Zintis , Forcina Giovanni C. , Denton Kyle , Dixon Scott J.
Published: March 2020

Individual cancer cells within a population can exhibit substantial phenotypic heterogeneity such that exposure to a lethal agent will kill only a fraction of cells at a given time. Whether common molecular mechanisms govern this fractional killing in response to different lethal stimuli is poorly understood. In part, this is because it is difficult to compare fractional killing between conditions using existing approaches. Here, we show that fractional killing can be quantified and compared for hundreds of populations in parallel using high-throughput time-lapse imaging. We find that fractional killing is highly variable between lethal agents and between cell lines. At the molecular level, we find that the antiapoptotic protein MCL1 is an important determinant of fractional killing in response to mitogen activated protein kinase (MAPK) pathway inhibitors but not other lethal stimuli. These studies lay the foundation for the large-scale, quantitative analysis of cell death phenotypic heterogeneity.
Moving Beyond the Mean: Subgroups and dimensions of brain activity and cognitive performance across domains
Authors: Hawco Colin , Dickie Erin W. , Jacobs Grace , Daskalakis Zafiris J. , Voineskos Aristotle N.
Published: March 2020

Human neuroimaging during cognitive tasks has provided unique and important insights into the neurobiology of cognition. However, the vast majority of research relies upon group aggregate or average statistical maps of activity, which do not fully capture the rich variability which exists across individuals. To better characterize individual variability, hierarchical clustering was performed separately on six fMRI tasks in 822 participants from the Human Connectome Project. Across all tasks, clusters ranged from a predominantly ‘deactivating’ pattern towards a more ‘activating’ pattern of brain activity, with differences in out-of-scanner cognitive test scores between clusters. Cluster stability was assessed via bootstrapping approach. Cluster probability did not indicate distinct/clear clustering. However, when participants were plotted in a dimensionally reduced ‘similarity space’ derived from bootstrapping, variability in brain activity among participants was best represented multidimensionally. A ‘positive to negative’ axis of activity was the strongest driver of individual differences.
ARHGEF3 regulates skeletal muscle regeneration and strength through autophagy
Authors: You Jae-Sung , Singh Nilmani , Reyes-Ordonez Adriana , Khanna Nidhi , Bao Zehua , Zhao Huimin , Chen Jie
Published: March 2020

Skeletal muscle regeneration is essential for restoring muscle function upon injury and for the maintenance of muscle health with aging. ARHGEF3, a Rho-specific GEF, negatively regulates myoblast differentiation via mammalian target of rapamycin complex 2 (mTORC2)-Akt signaling in a GEF-independent mannerin vitro. Here, we investigated ARHGEF3’s role in skeletal muscle regeneration by creating ARHGEF3 KO mice. These mice exhibited no discernible phenotype under normal conditions. Upon injury, however, ARHGEF3 deficiency enhanced the mass, fiber size and function of regenerating muscles in both young and aged mice. Surprisingly, these effects were not mediated by mTORC2-Akt signaling, but by the GEF activity of ARHGEF3. Furthermore, ARHGEF3 KO promoted muscle regeneration through activation of autophagy, a process that is also critical for maintaining muscle strength. Accordingly, in old mice, ARHGEF3 depletion prevented muscle weakness by restoring autophagy flux. Collectively, our findings identify an unexpected link between ARHGEF3 and autophagy-related muscle pathophysiology.
Notch1 regulates breast cancer stem cell function via a non-canonical cleavage-independent pathway
Authors: Sui Lufei , Wang Suming , Rodriguez Roberto K. , Sim Danielle , Bhattacharya Nandita , Blois Anna L. , Chen Siqi , Aziz Sura , Schlaeger Thorsten , Rogers Michael S. , Bielenberg Diane , Akslen Lars A. , Watnick Randolph S.
Published: March 2020

Current treatment of triple negative breast cancer patients is hindered by a high incidence of chemoresistance (30-50%). The prevailing theory is that resistance and subsequent recurrence is driven by cancer stem cells. Unfortunately, the functional characterization of cancer stem cells at the molecular level is still incomplete. We show here, that within the canonical breast cancer stem cell population, a subset of cells characterized by high Notch1 expression possesses the tumor-initiating property associated with cancer stem cells. Moreover, the tumor initiating property of these high Notch1-expressing breast cancer stem cells is mediated by a cleavage independent Notch signaling pathway culminating in the repression of SIRT1. Of note, the Notch1-mediated repression of SIRT1 is required not only for tumor initiation, but also for chemoresistance in breast cancer stem cells. Strikingly, inhibition of SIRT1 obviates the requirement for Notch1, marking the first example of conferring cancer stem cell function by inhibiting the activity of a single protein. We also demonstrate that progenitor-like mammary epithelial cells, which possess both luminal and basal properties, are also characterized by high Notch1 expression and repression of SIRT1 via the non-canonical pathway. These findings provide the first functional mechanistic requirements for tumor initiation by breast cancer stem cells and suggest that activation of the non-canonical Notch1 pathway is hardwired into tumor-initiating progenitor cells and thus a prerequisite for tumor initiation.

Statement of Significance

We demonstrate that chemoresistant and tumor-initiating properties of breast cancer stem cells are driven by repression of SIRT1 via non-canonical Notch signaling, suggesting a novel therapeutic strategy for triple negative breast cancer.
Association of Cardiovascular Manifestations with In-hospital Outcomes in Patients with COVID-19: A Hospital Staff Data
Authors: Liu Ru , Ming Xiaoyan , Xu Ou , Zhou Jianli , Peng Hui , Xiang Ning , Zhang Jiaming , Zhu Hong
Published: March 2020

Background

The outbreaks of coronavirus disease 2019 (COVID-19) caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remain a huge threat to the public health worldwide. Clinical data is limited up to now regarding the risk factors in favor of severe conversion of non-severe case with COVID-19.

Aims

This study analyzed a hospital staff data to figure out general clinical features of COVID-19 in terms of the association of cardiovascular manifestations (CVMs) with in-hospital outcomes of COVID-19 cases.

Methods

Retrospective, single-center case series of 41 consecutive hospitalized health staff with confirmed COVID-19 were collected at the Central Hospital of Wuhan in Wuhan, China, from January 15 to January 24, 2020. Epidemiological, demographic, clinical, laboratory, radiological, treatment data and in-hospital adverse events were collected and analyzed. Final date of follow-up was March 3, 2020. A comparative study was applied between cases with CVMs and those without CVMs.

Results

Of all, clinicians and clinical nurses accounted for 80.5%, while 87.8% of all had history of patient contact. The population was presented with a mean age of 39.1 ± 9.2 and less comorbidities than community population. The three most frequent symptoms of COVID-19 cases analyzed were fever (82.9%), myalgia or fatigue (80.5%) and cough (63.4%). While, the three most frequent initial symptoms were myalgia or fatigue (80.5%), fever (73.2%) and cough (41.5%). There were 95.1% cases featured as non-severe course of disease according to the official standard in China. Patients with CVMs and those without CVMs accounted for 58.5% and 41.5%, respectively. Compared with cases without CVMs, patients with CVMs were presented with lower baseline lymphocyte count (0.99 ± 0.43 and 1.55 ± 0.61, P<0.001), more who had at least once positive nucleic acid detection of throat swab during admission (50.0% and 11.8%, P=0.011), and more received oxygen support (79.2% and 23.5%, P<0.001). The rate of in-hospital adverse events was significantly higher in patients with CVMs group (75.0% and 23.5%, P=0.001). Multivariable logistic regression model indicated that, coexisting with CVMs in COVID-19 patients was not independently associated with in-hospital adverse events.

Conclusions

Most of hospital staff with COVID-19 had history of patient contact, featured non-severe course of disease. Cases with CVMs suffered from more in-hospital adverse events than those without CVMs. But concomitant CVMs were not independently associated with in-hospital adverse events in COVID-19 patients.
Light environment drives evolution of color vision genes in butterflies and moths
Authors: Sondhi Yash , Ellis Emily A. , Theobald Jamie C. , Kawahara Akito Y.
Published: March 2020

Opsins are the primary light-sensing molecules in animals. Opsins have peak sensitivities to specific wavelengths which allows for color discrimination. The opsin protein family has undergone duplications and losses, dynamically expanding and contracting the number of opsins, throughout invertebrate evolution, but it is unclear what drives this diversity. Light availability, however, appears to play a significant role. Dim environments are associated with low opsin diversity in deep-sea fishes and cave-dwelling animals. Correlations between high opsin diversity and bright environments, however, are tenuous. Insects are a good system to test whether opsin expansion is associated with greater light availability because they are enormously diverse and consequently display large variation in diel activity. To test this, we used 200 insect transcriptomes and examined the patterns of opsin diversity associated with diel-niche. We focused on the butterflies and moths (Lepidoptera) because this group has significant variation in diel-niche, substantial opsin recovery (n=100), and particularly well-curated transcriptomes. We identified opsin duplications using ancestral state reconstruction and examined rates of opsin evolution, and compared them across diel-niches. We find Lepidoptera species active in high light environments have independently expanded their opsins at least 10 times. Opsins from diurnal taxa also evolve faster; 13 amino acids were identified across different opsins that were under diversifying selection. Structural models reveal that four of these amino acids overlap with opsin color-tuning regions. By parsing nocturnal and diurnal switches, we show that light environment can influence gene diversity, selection, and protein structure of opsins in Lepidoptera.
Correlating predicted epigenetic marks with expression data to find interactions between SNPs and genes
Authors: Despinasse Antoine , Park Yongjin , Lapi Michael , Kellis Manolis
Published: March 2020

Despite all the work done, mapping GWAS SNPs in non-coding regions to their target genes remains a challenge. The SNP can be associated with target genes by eQTL analysis. Here we introduce a method to make these eQTLs more robust. Instead of correlating the gene expression with the SNP value like in eQTLs, we correlate it with epigenomic data. This epigenomic data is very expensive and noisy. We therefore predict the epigenomic data from the DNA sequence using the deep learning framework DeepSEA (Zhou and Troyanskaya, 2015).
Systematic tools for reprogramming plant gene expression in a simple model,Marchantia polymorpha
Authors: Sauret-Güeto Susanna , Frangedakis Eftychios , Silvestri Linda , Rebmann Marius , Tomaselli Marta , Markel Kasey , Delmans Mihails , West Anthony , Patron Nicola J. , Haseloff Jim
Published: March 2020

We present the OpenPlant toolkit, a set of interlinked resources and techniques to develop Marchantia as testbed for bioengineering in plants. Marchantia is a liverwort, a simple plant with an open form of development that allows direct visualization of gene expression and dynamics of cellular growth in living tissues. We describe new techniques for simple and efficient axenic propagation and maintenance of Marchantia lines with no requirement for glasshouse facilities. Marchantia plants spontaneously produce clonal propagules within a few weeks of regeneration, and lines can be amplified million-fold in a single generation by induction of the sexual phase of growth, crossing and harvesting of progeny spores. The plant has a simple morphology and genome with reduced gene redundancy, and the dominant phase of its life cycle is haploid, making genetic analysis easier. We have built robust Loop assembly vector systems for nuclear and chloroplast transformation and genome editing. These have provided the basis for building and testing a modular library of standardized DNA elements with highly desirable properties. We have screened transcriptomic data to identify a range of candidate genes, extracted putative promoter sequences, and tested themin vivoto identify new constitutive promoter elements. The resources have been combined into a toolkit for plant bioengineering that is accessible for laboratories without access to traditional facilities for plant biology research. The toolkit is being made available under the terms of the OpenMTA and will facilitate the establishment of common standards and the use of this simple plant as testbed for synthetic biology.
Transient opening of tricellular vertices controls paracellular transport through the follicle epithelium duringDrosophilaoogenesis
Authors: Isasti-Sanchez Jone , Münz-Zeise Fenja , Luschnig Stefan
Published: March 2020

Paracellular permeability is regulated to allow solute transport or migration of cells across epithelial or endothelial barriers. However, how occluding junction dynamics controls paracellular permeability is poorly understood. Here we describe patency, a developmentally regulated process inDrosophilaoogenesis, during which cell vertices in the follicle epithelium open transiently to allow paracellular transport of yolk proteins for uptake by the oocyte. We show that the sequential removal of E-Cadherin, N-Cadherin, NCAM/Fasciclin-2 and Sidekick from vertices precedes their basal-to-apical opening, while the subsequent assembly of tricellular occluding junctions terminates patency and seals the paracellular barrier. E-Cadherin-based adhesion is required to limit paracellular channel size, whereas stabilized adherens junctions, prolonged NCAM/Fasciclin-2 expression, impeded endocytosis, or increased actomyosin contractility prevent patency. Our findings reveal a key role of cell vertices as gateways controlling paracellular transport, and demonstrate that the dynamic regulation of adhesion and actomyosin contractility at vertices governs epithelial barrier properties.
An Improved 4’-Aminomethyltroxsalen-Based DNA Crosslinker for Biotinylation of DNA
Authors: Wielenberg Kevin , Wang Miao , Yang Min , Ozer Abdullah , Lis John T. , Lin Hening
Published: March 2020

Nucleic acid crosslinkers that covalently join commentary strands have applications as both pharmaceuticals and biochemical probes. Psoralen is a popular crosslinker moiety that reacts with double stranded DNA and RNA upon irradiation with long wave UV light. A commercially available compound EZ-Link Psoralen-PEG3-Biotin has been used in many studies to crosslink DNA and double strand RNA for genome-wide investigations. Here we present a novel probe, AP3B, which uses a psoralen derivative, 4’-aminomethyltrioxsalen, to biotinylate nucleic acids. We show that this compound is 8-fold more effective at labeling DNA in cells and several hundred-fold more effective at crosslinking two strands of DNAin vitrothan the commercially available compound EZ-Link Psoralen-PEG3-Biotin.
Trypanosoma bruceiultra-structure cell modifications caused by knockdown of the ribonuclease Rrp44/Dis3
Authors: Cesaro Giovanna , Hiraiwa Priscila M. , Carneiro Flavia R. G. , Rouam Valérie , Legrand Pierre , Conesa José Javier , Réfrégiers Matthieu , Pereiro Eva , Guimarães Beatriz G. , Jamme Frédéric , Zanchin Nilson I. T.
Published: March 2020

Rrp44/Dis3 is an essential protein conserved in all eukaryotes that functions in the maturation of many different RNA precursors and RNA surveillance. TheTrypanosoma bruceiRrp44/Dis3 homologue (TbRRP44) is required for maturation of pre-rRNA, spliced leader, and U3 snoRNA precursors. Its depletion leads to inhibition of cell proliferation and eventually to cell death. In this work, we showed that TbRRP44 depletion causes a massive expansion of acidic and lysosome-derived vacuoles, enlargement of cell and nuclei sizes without changes in DNA content, mitochondrial inactivation, and autophagy induction. Consistently, 3D reconstructions using cryo-soft X-ray tomography revealed extreme vacuolation of the cytoplasm and numerous cellular alterations, including an increase in size and number of calcium-containing vesicles and lipid droplets. These multiple defects indicate that a combination of alterations converge to induce lysosome expansion. With time, the cytoplasm is taken up by lysosome-derived vacuoles, which may be a final stage leading to the cell death triggered by TbRRP44 depletion. These studies provide the first evidence on the ultra-structure cell modifications caused by deficiency of this essential ribonuclease inT. brucei.
Environmental variability and the evolution of bet-hedging strategies
Authors: Slimon Kelley , Germain Rachel M.
Published: March 2020

Bet-hedging strategies, such as dispersal and dormancy, are predicted to evolve in varying and uncertain environments and are critical to ecological models of biodiversity maintenance. Theories of the specific ecological scenarios that favor the evolution of dispersal, dormancy, or their covariance are rarely tested, particularly for naturally-evolved populations that experience complex patterns of spatiotemporal environmental variation. We grew 23 populations ofVulpia microstachys, an annual grass native to California, in a greenhouse, and on the offspring generation measured seed dispersal ability and dormancy rates. We hypothesized that seed dormancy rates and dispersal abilities would be highest in populations from more productive, temporally variable sites, causing them to covary positively. Contrary to our hypothesis, our data suggest that both dispersal and dormancy evolve to combat different axes and scales of spatial heterogeneity, and are underlain by different seed traits, allowing them to evolve independently. Dormancy appears to have evolved as a strategy for overcoming microgeographic heterogeneity rather than temporal climate fluctuations, an outcome that to our knowledge has not been considered by theory. In sum, we provide much needed empirical data on the evolution of bet hedging, as well as a new perspective on the ecological function dormancy provides in heterogeneous landscapes.
ABO blood group, glycosyltransferase activity and risk of Venous Thrombosis
Authors: Kosta Manal Ibrahim , Bailly Pascal , Silvy Monique , Saut Noemie , Suchon Pierre , Morange Pierre-Emmanuel , Chiaroni Jacques , Trégouët David-Alexandre , Goumidi Louisa
Published: March 2020

Introduction

ABO blood group influence the risk of venous thrombosis (VT) by modifying A and B glycosyltransferases (AGT and BGT) activities that further modulates Factor VIII (FVIII) and von Willebrand Factor (VWF) plasma levels. The aim of this work was to evaluate the association of plasma GTs activities with VWF/FVIII plasma levels and VT risk in a case-control study.

Materials and Methods

420 cases were matched with 420 controls for age and ABO blood group. GT activities in plasma were measured using the quantitative transfer of tritiated N-acetylgalactosamine or galactose to the 2’-fucosyl-lactose and expressed in disintegration per minute/30µL of plasma and 2 hours of reaction (dpm/30µL/2H). FVIII and VWF plasma levels were respectively measured using human FVIII-deficient plasma in a 1-stage factor assay and STA LIATEST VWF (Diagnostica Stago).

Results

A and B GT activities were significantly lower in cases than in controls (8119±4027 vs 9682±4177 dpm/30µL/2H, p=2.03 × 10−5, and 4931±2305 vs 5524±2096 dpm/30µL/2H, p=0.043 respectively). This association was observed whatever the ABO blood groups. The ABO A1 blood group was found to explain∼80% of AGT activity. After adjusting for ABO blood groups, AGT activity was not correlated to VWF/FVIII plasma levels. Conversely, there was a moderate correlation (ρ∼0.30) between BGT activity and VWF/ FVIII plasma levels in B blood group carriers.

Conclusion

This work showed, for the first time, that GT activities were decreased in VT patients in comparison to controls with the same ABO blood group. The biological mechanisms responsible for this association remained to be determined.
JNK pathway restricts DENV, ZIKV and CHIKV infection by activating complement and apoptosis in mosquito salivary glands
Authors: Chowdhury Avisha , Modahl Cassandra M. , Tan Siok Thing , Xiang Benjamin Wong Wei , Missé Dorothée , Vial Thomas , Kini R. Manjunatha , Pompon Julien Francis
Published: March 2020

Arbovirus infection ofAedes aegyptisalivary glands (SGs) determines transmission. However, there is a dearth of knowledge on SG immunity. Here, we characterized SG immune response to dengue, Zika and chikungunya viruses using high-throughput transcriptomics. The three viruses regulate components of Toll, IMD and JNK pathways. However, silencing of Toll and IMD components showed variable effects on SG infection by each virus. In contrast, regulation of JNK pathway produced consistent responses. Virus infection increased with depletion of component Kayak and decreased with depletion of negative regulator Puckered. Virus-induced JNK pathway regulates complement and apoptosis in SGs via TEP20 and Dronc, respectively. Individual and co-silencing of these genes demonstrate their antiviral effects and that both may function together. Co-silencing eitherTEP20orDroncwithPuckeredannihilates JNK pathway antiviral effect. We identified and characterized the broad antiviral function of JNK pathway in SGs, expanding the immune arsenal that blocks arbovirus transmission.
Colonization with heterologous bacteria reprograms aCaenorhabditis elegansnutritional phenotype
Authors: Sun Qing , Vega Nicole M. , Cervantes Bernardo , Mancuso Christopher P. , Mao Ning , Taylor Megan , Collins James J. , Khalil Ahmad S. , Gore Jeff , Lu Timothy K.
Published: March 2020

Animals rely on the gut microbiome to process complex food compounds that the host cannot digest and to synthesize nutrients that the host cannot produce. New systems are needed to study how the expanded metabolic capacity provided by the gut microbiome impacts the nutritional status and health of the host. Here we colonized the nematodeCaenorhabditis elegansgut with cellulolytic bacteria that enabledC. elegansto utilize cellulose, an otherwise indigestible substrate, as a carbon source. The nutritional benefits of colonization with cellulolytic bacteria were assayed directly, by incorporation of isotopic biomass, and indirectly, as host larval yield resulting from glucose release in the gut. As a community component in the worm gut, cellulolytic bacteria can also support additional bacterial species with specialized roles, which we demonstrate by usingLactobacillusto protect againstSalmonellainfection. As a model system,C. eleganscolonized with cellulolytic bacteria can be used to study microbiome-host interactions. Engineered microbiome communities may provide host organisms with novel functions, such as the ability to use more complex nutrient sources and to fight against pathogen infections.

One Sentence Summary

Heterologous bacteria colonizing an animal gut help digest complex sugars to provide nutrition for the host in a model system.
Plant and Bacterial Metaxin-like Proteins: Novel Proteins Related to Vertebrate Metaxins Involved in Uptake of Nascent Proteins into Mitochondria
Authors: Adolph Kenneth W.
Published: March 2020

The metaxins were originally identified as vertebrate proteins of the outer mitochondrial membrane involved in protein import into mitochondria. Metaxin proteins have also been found in diverse invertebrate phyla. The present study is concerned with examining whether metaxin-like proteins occur in plants and bacteria. Metaxin-like proteins were revealed by their homology with human metaxins and the possession of characteristic GST_Metaxin protein domains. The results demonstrate that metaxin-like proteins exist in plants that include a wide variety of angiosperms, both eudicots and monocots, and other plant groups. Metaxin-like proteins can also be detected in bacteria, particularly in the Proteobacteria phylum, but also in different bacterial phyla. Phylogenetic analysis indicates that plant metaxin-like proteins, bacterial metaxin-like proteins, and vertebrate metaxins form distinct phylogenetic groups, but are related. Metaxin-like proteins, however, are only distantly related to GSTs (glutathione S-transferase proteins). A similar degree of homology is found in aligning the amino acid sequences of plant and bacterial metaxin-like proteins with human metaxins 1, 2, and 3 and other vertebrate metaxins. The amino acid identities range from about 22%-28% for each alignment. The presence of two conserved protein domains, GST_N_Metaxin and GST_C_Metaxin, in both plant and bacterial metaxin-like proteins provides evidence that these proteins are related to the vertebrate and invertebrate metaxins. The metaxin-like proteins have predicted secondary structures that are dominated by alpha-helical segments, like the vertebrate and invertebrate metaxins.
Phosphorylation barcode-dependent signal bias of the dopamine D1 receptor
Authors: Kaya Ali I. , Perry Nicole A. , Gurevich Vsevolod V. , Iverson T.M.
Published: March 2020

Agonist-activated G protein-coupled receptors (GPCRs) must correctly select from hundreds of potential downstream signaling cascades and effectors. To accomplish this, GPCRs first bind to an intermediary signaling protein, such as G protein or arrestin. These intermediaries initiate signaling cascades that promote the activity of different effectors, including several protein kinases. The relative roles of G proteins versus arrestins in initiating and directing signaling is hotly debated, and it remains unclear how the correct final signaling pathway is chosen given the ready availability of protein partners. Here, we begin to deconvolute the process of signal bias from the dopamine D1 receptor (D1R) by exploring factors that promote the activation of ERK1/2 or Src, the kinases that lead to cell growth and proliferation. We found that ERK1/2 activation involves both arrestin and Gαs, while Src activation depends solely on arrestin. Interestingly, we found that the phosphorylation pattern influences both arrestin and Gαs coupling, suggesting an additional way the cells regulate G protein signaling. The phosphorylation sites in the D1R intracellular loop 3 are particularly important for directing the binding of G protein versus arrestin and for selecting between the activation of ERK1/2 and Src. Collectively, these studies correlate functional outcomes with a physical basis for signaling bias and provide fundamental information on how GPCR signaling is directed.

Significance Statement

The functional importance of receptor phosphorylation in GPCR regulation has been demonstrated. Over the past decade, the phospho-barcode concept was developed to explain the multi-dimensional nature of the arrestin-dependent signaling network downstream of GPCRs. Here, we used the dopamine-1 receptor (D1R) to explore the effect of receptor phosphorylation on G protein-dependent and arrestin-dependent ERK and Src activation. Our studies suggest that D1R intracellular loop-3 phosphorylation affects both G proteins and arrestins. Differential D1R phosphorylation can direct signaling toward ERK or Src activation. This implies that phosphorylation induces different conformations of receptor and/or bound arrestin to initiate or select different cellular signaling pathways.
Genome-wide targets identification by CRISPRi-Omics for high-titer production of free fatty acids inEscherichia coli
Authors: Fang Lixia , Fan Jie , Wang Congya , Cao Yingxiu , Song Hao
Published: March 2020

To construct a superior microbial cell factory for chemical synthesis, a major challenge is to fully exploit cell potential via identifying and engineering beneficial gene targets in the sophisticated metabolic networks. Here, we develop an approach that integrates CRISPR interference (CRISPRi) to readily modulate genes expression and omics analyses to identify potential targets in multiple cellular processes, enabling systematical discovery of beneficial chromosomal gene targets that can be engineered to optimize free fatty acids (FFAs) production inEscherichia coli. We identify 56 beneficial genes via synergistic CRISPRi-Omics strategy, including 46 novel targets functioning in cell structure and division, and signaling transduction that efficiently facilitate FFAs production. Upon repressingihfAand overexpressingaidBandtesA’inE. coli, the recombinant strain LihfA-OaidB results in a FFAs titer of 21.6 g L-1in fed-batch fermentation, which, to our best knowledge, is the maximum FFAs titer by the recombinantE. colireported to date.
The Recombinational Landscape inDaphnia pulex
Authors: Lynch Michael , Ye Zhiqiang , Maruki Takahiro
Published: March 2020

Through the analysis of linkage disequilibrium from genome-wide sequencing data for multiple individuals from eight populations, the general features of the recombinational landscape are revealed in the microcrustaceanDaphnia pulex.The data suggest an exceptionally uniform pattern of recombination across the genome, while also confirming general patterns that are inconsistent with existing population-genetic models for the relationship between linkage dis-equilibrium and physical distances between genomic sites. Patterns of linkage disequilibrium are highly consistent among populations, and average rates of recombination are quite similar for all chromosomes. There is no evidence of recombination hotspots, and although there does appear to be suppressed recombination in the vicinity of gene bodies, this effect is quite small. Although this species reproduces asexually in ∼ 80% of generations, the mean per-generation recombination rate per nucleotide site is ∼ 37× the per-nucleotide mutation rate. Contrary to expectations for models in which crossing-over is the primary mechanism of recombination, and consistent with data for other species, the gradient of linkage disequilibrium with increasing physical distance between sites is far too high at short distances and far too low at long distances, suggesting an important role for factors such as the nonindependent appearance of pairs of mutations on haplotypes and long-range gene-conversion-like processes. Combined with other observations on patterns of nucleotide variation, these results provide a strong case for the utility ofD. pulexas a model system for the study of mechanisms of evolution in natural populations.