bluuebunny/crossref_metadata_2025_split huggingface dataset:
Whole human genome 5’-mC methylation analysis using long read nanopore sequencing
Authors: Silva Catarina , Machado Miguel , Ferrão José , Rodrigues Sebastião , Vieira Luís 
Published: May 2021

AbstractDNA methylation is a type of epigenetic modification that affects gene expression regulation and is associated with several human diseases. Microarray and short read sequencing technologies are often used to study 5’-methylcytosine (5’-mC) modification of CpG dinucleotides in the human genome. Although both technologies produce trustable results, the evaluation of the methylation status of CpG sites suffers from the potential side effects of DNA modification by bisulfite and the ambiguity of mapping short reads in repetitive and highly homologous genomic regions, respectively. Nanopore sequencing is an attractive alternative for the study of 5’-mC since the long reads produced by this technology allow to resolve those genomic regions more easily. Moreover, it allows direct sequencing of native DNA molecules using a fast library preparation procedure. In this work we show that 10X coverage depth nanopore sequencing, using DNA from a human cell line, produces 5’-mC methylation frequencies consistent with those obtained by methylation microarray and digital restriction enzyme analysis of methylation. In particular, the correlation of methylation values ranged from 0.73 to 0.90 using an average genome sequencing coverage depth <2X or a minimum read support of 17X for each CpG site, respectively. We also showed that a minimum of 5 reads per CpG yields strong correlations (>0.89) between sequencing runs and an almost uniform variation in methylation frequencies of CpGs across the entire value range. Furthermore, nanopore sequencing was able to correctly display methylation frequency patterns according to genomic annotations, including a majority of unmethylated and methylated sites in the CpG islands and inter-CpG island regions, respectively. These results demonstrate that low coverage depth nanopore sequencing is a fast, reliable and unbiased approach to the study of 5’-mC in the human genome.
Anaerobic derivates of mitochondria and peroxisomes in the free-living amoeba Pelomyxa schiedti revealed by single-cell genomics
Authors: Záhonová Kristína , Treitli Sebastian Cristian , Le Tien , Škodová-Sveráková Ingrid , Hanousková Pavla , Čepička Ivan , Tachezy Jan , Hampl Vladimír 
Published: May 2021

AbstractPelomyxa schiedti is a free-living amoeba belonging to the group Archamoebae, which encompasses anaerobes bearing mitochondrion-related organelles (MROs) – hydrogenosomes in free-living Mastigamoeba balamuthi and mitosomes in the human pathogen Entamoeba histolytica. Anaerobic peroxisomes, another adaptation to anaerobic lifestyle, were identified only recently in M. balamuthi. We found evidence for both these organelles in the single-cell-derived genome and transcriptome of P. schiedti, and corresponding vesicles were tentatively revealed in electron micrographs. In silico reconstructed MRO metabolism seems similar to that of M. balamuthi harboring respiratory complex II, electron-transferring flavoprotein, partial TCA cycle running presumably in reductive direction, pyruvate:ferredoxin oxidoreductase, [FeFe]-hydrogenases, glycine cleavage system, and sulfate activation pathway. The cell disposes with an expanded set of NIF enzymes for iron sulfur cluster assembly, but their localization remains unclear. Quite contrary, out of 67 predicted peroxisomal enzymes, only four were reported also in M. balamuthi, namely peroxisomal processing peptidase, nudix hydrolase, inositol 2-dehydrogenase, and D-lactate dehydrogenase. Other putative functions of peroxisomes could be pyridoxal 5I⍰-phosphate biosynthesis, amino acid and carbohydrate metabolism, and hydrolase activities. Future experimental evidence is necessary to define functions of this surprisingly enzyme-rich anaerobic peroxisome.Author summaryA major part of the microbial diversity cannot be cultured in isolation, and so it escapes from traditional ways of investigation. In this paper, we demonstrate the successful approach for generating good-quality genome and transcriptome drafts from a peculiar amoeba Pelomyxa schiedti using single-cell methods. P. schiedti is a member of Archamoebae clade harboring microaerobic protists including a free-living Mastigamoeba balamuthi and a human parasite Entamoeba histolytica. Mitochondria and peroxisomes represent two organelles that are most affected during adaptation to microoxic or anoxic environments. Mitochondria are known to transform to anaerobic mitochondria, hydrogenosomes, mitosomes, and various transition stages in between, all of which encompass different enzymatic capacity. Anaerobic peroxisomes have been first noticed in M. balamuthi, but their function remained unclear for now. Data obtained in this study were used for revealing the presence and for the detailed functional annotations of anaerobic derivates of mitochondria and peroxisomes in P. schiedti, which were corroborated by transmission electron microscopy.
Sylites: Multipurpose markers for the visualization of inhibitory synapses
Authors: Khayenko Vladimir , Schulte Clemens , Reis Sara L. , Avraham Orly , Schietroma Cataldo , Worschech Rafael , Nordblom Noah F. , Kachler Sonja , Villmann Carmen , Heinze Katrin G. , Schlosser Andreas , Schueler-Furman Ora , Tovote Philip , Specht Christian G. , Maric Hans Michael 
Published: May 2021

AbstractWe introduce Sylites – small and versatile fluorogenic affinity probes for high-contrast visualization of inhibitory synapses. Having stoichiometric labeling and exceptional selectivity for neuronal gephyrin, a hallmark protein of the inhibitory post-synapse, Sylites enable superior synapse staining compared with antibodies. Combined with super-resolution microscopy, Sylites allow precise nanoscopic measurements of the synapse. In brain tissue, Sylites reveal the three-dimensional distribution of inhibitory synapses within just an hour.
A Dynamic Affective Core to bind the Contents, Context, and Value of Conscious Experience
Authors: Kishida Kenneth T. , Sands L. Paul 
Published: May 2021

AbstractThe private and dynamic nature of conscious subjective experience poses an empirical challenge that has led neuroscience-based theories about consciousness to note the importance of ‘the hard problem’ of explaining how subjective phenomenal experience can arise from neural activity but set it aside and focus on the ‘easier’ problems associated with information representation and behavior. This approach leaves a major gap in our understanding of the neural mechanisms underlying conscious subjective experience and its dynamic nature. However, computational methods integrated with a variety of tools for measuring human brain activity are beginning to link dynamic changes in subjective affect with reproducible neurobehavioral signals in humans. In particular, research applying computational reinforcement learning theory has shown tremendous utility in investigating human choice behavior and the role the dopaminergic system plays in dynamic behavioral control. This research is beginning to reveal an explicit connection between the dynamics of dopaminergic signals and dynamic changes in subjective affect. However, it should be obvious that the dopaminergic system alone is not sufficient to explain all of the complexities of affective dynamics. We review foundational work, highlight current problems and open questions, and propose a Dynamic Affective Core Hypothesis that integrates advances in our understanding of the representation of the content and context of conscious experiences with our nascent understanding about how these representations acquire and retain affective subjective value.
Crosstalk between Fat Mass and Obesity-related (FTO) and multiple WNT signaling pathways
Authors: Kim Hyunjoon , Jang Soohyun , Lee Young-suk 
Published: May 2021

ABSTRACTFat Mass and Obesity-related (FTO) gene is associated with a diverse set of human diseases. Yet, the functional landscape of FTO remains largely unknown, most likely owing to its wide range of mechanistic roles and cell-type-specific targets. Here, we discover the intricate role of FTO in multiple WNT signaling pathways. Re-analyses of public data identified the bifurcation of canonical and noncanonical WNT pathways as the major role of FTO. In FTO-depleted cells, we find that the canonical WNT/β-Catenin signaling is inhibited in a non-cell autonomous manner via the upregulation of DKK1. Simultaneously, this upregulation of DKK1 promotes cell migration via activating the noncanonical WNT/PCP pathway. Unexpectedly, we also find that the canonical WNT/STOP signaling induces the accumulation of cytoplasmic FTO proteins. This subsequently leads to the stabilization of mRNAs via RNA demethylation, revealing a previously uncharacterized mode of WNT action in RNA regulation. Altogether, this study places the functional context of FTO at the branching point of multiple WNT signaling pathways which may explain the wide spectrum of FTO functions.
Multiomic Characterization of Stage I Lung Adenocarcinoma Reveals Distinct Genetic and Immunologic Features of Recurrent Disease
Authors: Kratz Johannes R. , Li Jack Z. , Tsui Jessica , Lee Jen C. , Ding Vivianne W. , Rao Arjun A. , Mann Michael J. , Chan Vincent , Combes Alexis J. , Krummel Matthew F. , Jablons David M. 
Published: May 2021

AbstractBackgroundRecurrence after surgery for early-stage lung cancer is common, occurring between 30-50% of the time. Despite the popularization of prognostic gene signatures in early-stage lung cancer that allow us to better predict which patients may recur, why patients recur after surgery remains unclear.MethodsUsing a large cohort of lung adenocarcinoma patients with complete genetic, genomic, epigenetic and clinical profiling, a recurrence classifier was developed which identifies patients at highest risk of recurrence. The genetic, genomic, and epigenetic profiles of stage I patients with low-vs. high-risk of recurrence were compared. To characterize the tumor immune microenvironment of recurrent stage I tumors, single cell RNA-seq was performed on fresh tissue samples undergoing lung adenocarcinoma resection at UCSF to identify unique immune population markers and applied to the large stage I lung adenocarcinoma cohort using digital cytometry.ResultsRecurrence high-risk stage I lung adenocarcinomas demonstrated a higher mutation burden than low-risk tumors, however, none of the known canonical lung cancer driver mutations were more prevalent in high-risk tumors. Transcriptomic analysis revealed widespread activation of known cancer and cell cycle pathways with simultaneous downregulation of immune response pathways including antigen presentation and Th1/Th2 activation. Tumors at high-risk of recurrence displayed depleted adaptive immune populations, and depletion of adaptive immune populations was independently prognostic of recurrence in stage I lung adenocarcinomas.ConclusionRecurrent stage I lung adenocarcinomas display distinct features of genomic and genetic instability including increased tumor mutation burden, neoantigen load, activation of numerous mitotic and cell cycle genes, and decreased genome-wide methylation burden. Relative depletion of infiltrating adaptive immune populations may allow these tumors to escape immunosurveillance and recur after surgery.
GPI anchor remodeling by the plant PGAP1 ortholog HLD1 is essential for Papaver self-incompatibility
Authors: Lin Zongcheng , Xie Fei , Triviño Marina , Zhao Tao , Coppens Frederik , Sterck Lieven , Bosch Maurice , Franklin-Tong Vernonica E. , Nowack Moritz K. 
Published: May 2021

AbstractIn eukaryotes, glycosylphosphatidylinositol anchored proteins (GPI-APs) are tethered to the outer leaflet of the plasma membrane where they function as key regulators of a plethora of biological processes. Self-incompatibility (SI) plays a pivotal role regulating fertilization in higher plants through recognition and rejection of ‘self’ pollen. Here we used Arabidopsis thaliana lines engineered to be self-incompatible by expression of Papaver SI determinants for an SI suppressor screen. We identify HLD1, an ortholog of human GPI-inositol deacylase PGAP1, whose mutation completely abolishes the SI response. We show that HLD1 functions as a GPI-inositol deacylase and that this GPI-remodeling activity is essential for SI. Using GFP-SKU5 as a representative GPI-AP, we show that HLD1 mutation does not affect GPI-AP production and targeting, but alters the configuration of mature GPI-APs. This prevents GPI-AP release from the plasma membrane, suggesting that this process plays a critical role in the regulation of SI. Our data not only identify GPI anchoring as a new pathway of SI providing new directions to investigate SI mechanisms, but identifies for the first time a function for GPI-AP remodeling by inositol deacylation in plants.One sentence summaryThe Papaver self-incompatibility response requires GPI-anchor modification by HLD1, an ortholog of the mammalian PGAP1.
Analysis of viromes and microbiomes from pig fecal samples reveals that phages and prophages are not vectors of antibiotic resistance genes
Authors: Billaud Maud , Lamy-Besnier Quentin , Lossouarn Julien , Moncaut Elisabeth , Dion Moira B. , Moineau Sylvain , Traoré Fatoumata , Le Chatelier Emmanuelle , Denis Catherine , Estelle Jordi , Achard Caroline , Zemb Olivier , Petit Marie-Agnès 
Published: May 2021

AbstractUnderstanding the transmission of antibiotic resistance genes (ARGs) is critical for human health. For this, it is necessary to identify which type of mobile genetic elements is able to spread them from animal reservoirs into human pathogens. Previous research suggests that in pig feces, ARGs may be encoded by bacteriophages. However, convincing proof for phage-encoded ARGs in pig viromes is still lacking, because of bacterial DNA contaminating issues. We collected 14 pig fecal samples and performed deep sequencing on both highly purified viral fractions and total microbiota, in order to investigate phage and prophage-encoded ARGs. We show that ARGs are absent from the genomes of active, virion-forming phages (below 0.02% of viral contigs from viromes), but present in three prophages, representing 0.02% of the viral contigs identified in the microbial dataset. However, the corresponding phages were not detected in the viromes, and their genetic maps suggest they might be defective. Furthermore, our dataset allows for the first time a comprehensive view of the interplay between prophages and viral particles.
High vitamin B2 production by the Lactiplantibacillus plantarum B2 strain carrying a mutation in the aptamer P1 helix of the FMN-riboswitch that regulates expression of the bacterial rib operon
Authors: Ripa Inés , Ruiz-Masó José Ángel , De Simone Nicola , Russo Pasquale , Spano Giuseppe , del Solar Gloria 
Published: May 2021

SummaryManufacturing of probiotics and functional foods using lactic acid bacteria that overproduce vitamin B2 has gained growing interest due to ariboflavinosis problems affecting populations of both developing and affluent countries. Two isogenic Lactiplantibacillus plantarum strains, namely a riboflavin-producing parental strain (UFG9) and a roseoflavin-resistant strain (B2) that carries a mutation in the FMN-aptamer of the potential rib operon riboswitch, were analyzed for production and intra- and extracellular accumulation of flavins, as well as for regulation of the rib operon expression. Strain B2 accumulated in the medium one of the highest levels of riboflavin+FMN ever reported for lactic acid bacteria, exceeding by ~25 times those accumulated by UFG9. Inside the cells, concentration of FAD was similar in both strains, while that of riboflavin+FMN was ~6-fold higher in B2. Mutation B2 could decrease the stability of the aptamer’s regulatory P1 helix even in the presence of the effector, thus promoting the antiterminator structure of the riboswitch ON state. Although the B2-mutant riboswitch showed an impaired regulatory activity, it retained partial functionality being still sensitive to the effector. The extraordinary capacity of strain B2 to produce riboflavine, together with its metabolic versatility and probiotic properties, can be exploited for manufacturing multifunctional foods.
Structural basis of PETISCO complex assembly during piRNA biogenesis inC. elegans
Authors: Perez-Borrajero Cecilia , Podvalnaya Nadezda , Holleis Kay , Lichtenberger Raffael , Karaulanov Emil , Simon Bernd , Basquin Jérôme , Hennig Janosch , Ketting René F. , Falk Sebastian 
Published: May 2021

AbstractPiwi-interacting RNAs (piRNAs) constitute a class of small RNAs that bind PIWI proteins and are essential to repress transposable elements in the animal germline, thereby promoting genome stability and maintaining fertility.C. eleganspiRNAs (21U RNAs) are transcribed individually from minigenes as precursors that require 5’ and 3’ processing. This process depends on the PETISCO complex, consisting of four proteins: IFE-3, TOFU-6, PID-3, and ERH-2. We employ biochemical and structural biology approaches to characterize the PETISCO architecture and its interaction with RNA, together with its effector proteins TOST-1 and PID-1. These two proteins define different PETISCO functions: PID-1 governs 21U processing whereas TOST-1 links PETISCO to an unknown process essential for early embryogenesis.Here, we show that PETISCO forms an octameric assembly with each subunit present in two copies. Determination of structures of the TOFU-6/PID-3 and PID-3/ERH-2 subcomplexes, supported byin vivostudies of subunit interaction mutants, allows us to propose a model for the formation of the TOFU-6/PID-3/ERH-2 core complex, and its functionality in germ cells and early embryos. Using NMR spectroscopy, we demonstrate that TOST-1 and PID-1 bind to a common surface on ERH-2, located opposite its PID-3 binding site, explaining how PETISCO can mediate different cellular roles.
Biphasic effects of IL-27 during Staphylococcus aureus implant-associated osteomyelitis in mice
Authors: Morita Yugo , Franchini Anthony M. , Owen John R. , Martinez John C. , Daiss John L. , de Mesy Bentley Karen L. , Kates Stephen L. , Schwarz Edward M. , Muthukrishnan Gowrishankar 
Published: May 2021

AbstractInterleukin-27 is a pleiotropic cytokine whose reported functions during bacterial infections are debated as an area of active research. To address this, we investigated the role of IL-27 signaling during Staphylococcus aureus osteomyelitis. Clinically, we observed elevated serum IL-27 levels (20-fold higher, p<0.05) in patients with S. aureus osteomyelitis compared to uninfected patients undergoing elective total joint replacement. Remarkably, IL-27 serum levels immediately following septic death were 60-fold higher vs. uninfected patients (p<0.05), suggesting that IL-27 may be a biomarker of end-stage infection and/or cytokine storm. To test this, we hypothesized that IL-27 mediates bacterial clearance during the acute phase of S. aureus osteomyelitis, and subsequently suppresses inflammation to prevent cytokine storm and osteolysis during chronic infection. In mice, we observed that systemic IL-27 delivery by a recombinant adeno-associated viral vector (rAAV-IL-27) ameliorates surgical site soft tissue infection and peri-implant bone loss during the establishment of implant-associated S. aureus osteomyelitis. This effect was not observed in IL-27 receptor α knock-out mice, suggesting a direct role of IL-27/IL-27R signaling on immune and bone cell functions. Examination of IL-27-mediated immune responses via transcriptome analyses of infected tibiae demonstrated that IL-27 is a biphasic cytokine with IL-27/IL-27R activating immunostimulatory responses including Th17, IL-2, TLR, and iNOS signaling early, and subsequently suppressing these pathways during chronic infection. Ex vivo confirmation using murine macrophages revealed that IL-27 co-stimulates TLR signaling to increase the production of nitric oxide, and immunomodulatory cytokines such as IL-10, IL-21, IL-31, and TNF-β, but is not a chemokine.Author SummaryStaphylococcus aureus is the most common pathogen in orthopaedic infections, and hard-to-treat (MRSA) strains cause >50% of these infections. Thus, there is an urgent need to develop immunotherapies to treat these life-threatening S. aureus infections. Currently, the role of multifunctional IL-27 on S. aureus osteomyelitis is unknown. In a clinical study, we observed that IL-27 is an important biomarker for identifying S. aureus osteomyelitis patients, and that elevated serum IL-27 levels correlated with adverse clinical outcomes, such as septic death. In our efforts to uncover the underlying mechanisms, we reveal that IL-27 is a biphasic cytokine, activating proinflammatory immune pathways, including Th17 responses, early during acute S. aureus osteomyelitis, and subsequently repressing them during the chronic phase to prevent cytokine storm and bone damage. These results indicate that immune modulation of IL-27/IL-27R signaling could be a viable therapeutic strategy in mitigating S. aureus osteomyelitis.
Topic: Hyper immune Bovine Colostrum as a Low-Cost, Large-Scale Source of Antibodies against COVID-19
Authors: Nili Hassan , Bouzari Majid , Attaran Hamid Reza , Golab Nader Ghaleh , Rabani Mohammad 
Published: May 2021

AbstractMany different strategies have been used to fight against COVID-19 pandemic as a therapeutics or prophylaxis approaches. However, none of them so far have used, passive immune transfer using products from immunized farm animals. Hyper immune bovine colostrums (HBC) have been used against many different respiratory and gastrointestinal tracts infections during past decades.Six mixed Holstein X Semental dairy cattle’s in their 6-7 months of gestation period years were chosen for hyper immunization with COVID-19 vaccine. An isolated and very well protected site was selected and equipped according to animal husbandry code of practice, used for animal experimentation. Specific IgG level against SARS-CoV-2 virus was measured before and after vaccination in the sera, and in the colostrum following parturition. Very high specific IgG level was detected one week following second vaccination in the sera and in first colostrums after parturition. Safety of the product was approved following phase 1 of clinical trials in 40 healthy volunteers. Phase 2 of the clinical trials is underway. Early results show effectiveness of the product in reducing sore throat and cough in early stages of SARS-CoV-2 infection.