bluuebunny/crossref_metadata_2025_split huggingface dataset:
FACT regulates pluripotency through distal regulation of gene expression in murine embryonic stem cells
Authors: Klein David C. , Lardo Santana M. , McCannell Kurtis N. , Hainer Sarah J. 
Published: August 2021

AbstractThe FACT complex is a conserved histone chaperone with essential roles in transcription and histone deposition. FACT is essential in pluripotent and cancer cells, but otherwise dispensable for most mammalian cell types. FACT deletion or inhibition can block induction of pluripotent stem cells, yet the mechanism through which FACT regulates cell fate decisions remains unclear. To determine this mechanism, we used inducible depletion of FACT subunit SPT16 in murine embryonic stem cells paired with genomic factor localization, nascent transcription, and chromatin accessibility analyses. Over a timecourse of SPT16 depletion, nucleosomes invade loci bound by master pluripotency factors and gene-distal DNaseI hypersensitive sites. Simultaneously, transcription of Pou5f1 (OCT4), Sox2, Nanog, and enhancer RNAs produced at the genes’ associated enhancers are downregulated, suggesting that FACT regulates expression of the pluripotency factors themselves. We find that FACT maintains cellular pluripotency through a precise nucleosome-based regulatory mechanism for appropriate expression of both coding and non-coding transcripts associated with pluripotency.
Length Biases in Single-Cell RNA Sequencing of pre-mRNA
Authors: Gorin Gennady , Pachter Lior 
Published: August 2021

AbstractSingle-molecule pre-mRNA and mRNA sequencing data can be modeled and analyzed using the Markov chain formalism to yield genome-wide insights into transcription. However, quantitative inference with such data requires careful assessment and understanding of noise sources. We find that long pre-mRNA transcripts are over-represented in sequencing data, and explore the mechanistic implications. A biological explanation for this phenomenon within our modeling framework requires unrealistic transcriptional parameters, leading us to posit a length-based model of capture bias. We provide solutions for this model, and use them to find concordant and mechanistically plausible parameter trends across data from multiple single-cell RNA-seq experiments in several species.
Modular Fluorescent Nanoparticle DNA Probes for Detection of Peptides and Proteins
Authors: Stawicki Cassandra M. , Rinker Torri E. , Burns Markus , Tonapi Sonal S. , Galimidi Rachel P. , Anumala Deepthi , Robinson Julia K. , Klein Joshua S. , Mallick Parag 
Published: August 2021

AbstractFluorescently labeled antibody and aptamer probes are used in biological studies to characterize binding interactions, measure concentrations of analytes, and sort cells. Fluorescent nanoparticle labels offer an excellent alternative to standard fluorescent labeling strategies due to their enhanced brightness, stability and multivalency; however, challenges in functionalization and characterization have impeded their use. This work introduces a straightforward approach for preparation of fluorescent nanoparticle probes using commercially available reagents and common laboratory equipment. Fluorescent polystyrene nanoparticles, Thermo Fisher Scientific FluoSpheres™, were used in proof-of-principle studies. Particle passivation was achieved by covalent attachment of amine-PEG-azide to carboxylated particles, neutralizing the surface charge from -43 to -15 mV. A conjugation-annealing handle and DNA aptamer probe was attached to the azide-PEG nanoparticle surface either through reaction of pre-annealed handle and probe or through a stepwise reaction of the nanoparticles with the handle followed by aptamer annealing. Nanoparticles functionalized with DNA aptamers targeting histidine tags and VEGF protein had high affinity (EC50s ranging from 3-12 nM) and specificity, and were more stable than conventional labels. This protocol for preparation of nanoparticle probes relies solely on commercially available reagents and common equipment, breaking down the barriers to use nanoparticles in biological experiments.
Emergence of distinct and heterogeneous strains of amyloid beta as Alzheimer’s disease progresses in Down syndrome
Authors: Maxwell Alison M. , Yuan Peng , Rivera Brianna M. , Schaaf Wilder , Mladinov Mihovil , Prasher Vee P. , Robinson Andrew C. , DeGrado William F. , Condello Carlo 
Published: August 2021

0.ABSTRACTAmyloid beta (Aβ) is thought to play a critical role in the pathogenesis of Alzheimer’s disease (AD). Prion-like Aβ polymorphs, or “strains”, can have varying pathogenicity and may underlie the phenotypic heterogeneity of the disease. In order to develop effective AD therapies, it is critical to identify the strains of Aβ that might arise prior to the onset of clinical symptoms and understand how they may change with progressing disease. Down syndrome (DS), as the most common genetic cause of AD, presents promising opportunities to compare such features between early and advanced AD. In this work, we evaluate the neuropathology and Aβ strain profile in the post-mortem brain tissues of 210 DS, AD, and control individuals. We assayed the levels of various Aβ and tau species and used conformation-sensitive fluorescent probes to detect differences in Aβ strains among individuals and populations. We found that these cohorts have some common but also some distinct strains from one another, with the most heterogeneous populations of Aβ emerging in subjects with high levels of AD pathology. The emergence of distinct strains in DS at these later stages of disease suggests that the confluence of aging, pathology, and other DS-linked factors may favor conditions that generate strains that are unique from sAD.
Chronic ethanol drinking in non-human primates induces inflammatory cathepsin gene expression in alveolar macrophages accompanied by functional defects
Authors: Lewis Sloan A. , Doratt Brianna , Sureshchandra Suhas , Jankeel Allen , Newman Natali , Grant Kathleen A. , Messaoudi Ilhem 
Published: August 2021

ABSTRACTChronic alcohol drinking is associated with increased susceptibility to viral and bacterial respiratory pathogens. Investigating the effects of alcohol on the lung is challenging in humans because of the complexity of human drinking behavior and the challenge of obtaining samples. In this study, we utilize a rhesus macaque model of voluntary ethanol self-administration to study the effects of alcohol on the lung in a physiologically and genetically relevant model. We report a heightened activation and inflammatory state in alveolar macrophages (AM) obtained from ethanol drinking animals that is accompanied by increased chromatin accessibility in intergenic regions that regulate inflammatory genes and contain binding motifs for transcription factors AP-1, IRF8, and NFKB p-65. In line with these transcriptional and epigenetic changes at basal state, AM from ethanol drinking animals generate elevated inflammatory mediator responses to LPS and respiratory syncytial virus (RSV). Analysis using scRNA-Seq revealed heterogeneity in lung-resident macrophage and monocyte populations, including increased abundance of activated and cathepsin-expressing clusters and accelerated differentiation with ethanol. Finally, functional assays show increased mitochondrial content in AM from ethanol drinking animals, which is associated with observed increased ROS and decreased phagocytosis capacity. This comprehensive epigenomic, transcriptional and functional profiling of lung macrophages after ethanol drinking in macaques provides previously unidentified mechanisms of ethanol induced infection susceptibility in patients with alcohol use disorders.
Close agreement between deterministic vs. stochastic modeling of first-passage time to vesicle fusion
Authors: Matveev Victor 
Published: August 2021

AbstractCa2+-dependent cell processes such as neurotransmitter or endocrine vesicle fusion are inherently stochastic due to large fluctuations in Ca2+ channel gating, Ca2+ diffusion and Ca2+ binding to buffers and target sensors. However, prior studies revealed closer-than-expected agreement between deterministic and stochastic simulations of Ca2+ diffusion, buffering and sensing, as long as Ca2+ channel gating is not Ca2+-dependent. To understand this result more fully, we present a comparative study complementing prior work, focusing on Ca2+ dynamics downstream of Ca2+ channel gating. Specifically, we compare deterministic (mean-field / mass-action) and stochastic simulations of vesicle exocytosis latency, quantified by the probability density of the first-passage time (FPT) to the Ca2+-bound state of a vesicle fusion sensor, following a brief Ca2+ current pulse. We show that under physiological constraints, the discrepancy between FPT densities obtained using the two approaches remains small even if as few as ∼50 Ca2+ ions enter per single channel-vesicle release unit. Using a reduced two-compartment model for ease of analysis, we illustrate how this close agreement arises from the smallness of correlations between fluctuations of the reactant molecule numbers, despite the large magnitude of the fluctuation amplitudes. This holds if all relevant reactions are heteroreaction between molecules of different species, as is the case for the bimolecular Ca2+ binding to buffers and downstream sensor targets. In this case diffusion and buffering effectively decorrelate the state of the Ca2+ sensor from local Ca2+ fluctuations. Thus, fluctuations in the Ca2+ sensor’s state underlying the FPT distribution are only weakly affected by the fluctuations in the local Ca2+ concentration around its average, deterministically computable value.Statement of SignificanceMany fundamental Ca2+-dependent cell processes are triggered by local Ca2+ elevations involving only a few hundred Ca2+ ions. Therefore, one expects large Ca2+ concentration fluctuations, which are ignored by deterministic reaction-diffusion modeling approaches. However, more accurate stochastic approaches require tracking trajectories of individual Ca2+ ions and its binding targets, which is very computationally expensive. This study reveals conditions under which Ca2+-dependent processes like secretory vesicle fusion can be modeled using efficient deterministic approaches, without sacrificing significant accuracy. We find that deterministic methods can accurately predict the delay to the fusion of a neurotransmitter-containing vesicle, as long as the number of Ca2+ ions is above about 50. We reveal factors that explain the limited impact of stochastic fluctuations in this case.
Uncovering perturbations in human hematopoiesis associated with healthy aging and myeloid malignancies at single cell resolution
Authors: Ainciburu Marina , Ezponda Teresa , Berastegui Nerea , Alfonso-Pierola Ana , Vilas-Zornoza Amaia , Martin-Uriz Patxi San , Alignani Diego , de Espinosa Jose Lamo , Julian Mikel San , Jimenez Tamara , López Félix , Muntion Sandra , Sanchez-Guijo Fermin , Molero Antonieta , Montoro Julia , Serrano Guillermo , Diaz-Mazkiaran Aintzane , Lasaga Miren , Gomez-Cabrero David , Díez-Campelo María , Valcarcel David , Hernaez Mikel , Romero Juan P. , Prosper Felipe 
Published: August 2021

ABSTRACTEarly hematopoiesis is a continuous process in which hematopoietic stem and progenitor cells (HSPCs) gradually differentiate toward specific lineages. Aging and myeloid malignant transformation are characterized by changes in the composition and regulation of HSPCs. In this study, we used single cell RNA sequencing (scRNAseq) to characterize an enriched population of human hematopoietic stem and progenitor cells (HSPC) obtained from young and elderly healthy individuals.Based on their transcriptional profile, we identified changes in the proportions of progenitor compartments during aging, and differences in their functionality, as evidenced by gene set enrichment analysis. Trajectory inference revealed that altered gene expression dynamics accompanied cell differentiation, which could explain age-associated changes in hematopoiesis. Next, we focused on key regulators of transcription by constructing gene regulatory networks and detected regulons that were specifically active in elderly individuals. Using previous findings in healthy cells as a reference, we analyzed scRNA-seq data obtained from patients with myelodysplastic syndrome (MDS) and detected specific alterations of the expression dynamics of genes involved in erythroid differentiation in all patients with MDS such as TRIB2. In addition, the comparison between transcriptional programs and GRN regulating normal HSPCs and MDS HSPCs allowed identification of regulons that were specifically active in MDS cases such as SMAD1, HOXA6, POU2F2 and RUNX1 suggesting a role of these TF in the pathogenesis of the disease.In summary, we demonstrate that the combination of single cell technologies with computational analysis tools enable the study of a variety of cellular mechanisms involved in early hematopoiesis and can be used to dissect perturbed differentiation trajectories associated with aging and malignant transformation. Furthermore, the identification of abnormal regulatory mechanisms associated with myeloid malignancies could be exploited for personalized therapeutic approaches in individual patients.
Circadian Proteins Cry and Rev-erb Converge to Deepen Cellular Quiescence by Downregulating Cyclin D and Cdk4,6
Authors: Wang Xia , Liu Bi , Pan Qiong , Kwon Jungeun Sarah , Miller Matthew A. , Croce Kimiko Della , Yao Guang 
Published: August 2021

ABSTRACTThe proper balance and transition between cellular quiescence and proliferation are critical to tissue homeostasis, and their deregulations are commonly found in many human diseases, including cancer and aging. Recent studies showed that the reentry of quiescent cells to the cell cycle is subjected to circadian regulation. However, the underlying mechanisms are largely unknown. Here, we report that two circadian proteins, Cryptochrome (Cry) and Rev-erb, deepen cellular quiescence in rat embryonic fibroblasts, resulting in stronger serum stimulation required for cells to exit quiescence and reenter the cell cycle. This finding was opposite from what we expected from the literature. By modeling a library of possible regulatory topologies linking Cry and Rev-erb to a bistable Rb-E2f gene network switch that controls the quiescence-to-proliferation transition and by experimentally testing model predictions, we found Cry and Rev-erb converge to downregulate Cyclin D/Cdk4,6 activity, leading to an ultrasensitive increase of the serum threshold to activate the Rb-E2f bistable switch. Our findings suggest a mechanistic role of circadian proteins in modulating the depth of cellular quiescence, which may have implications in the varying potentials of tissue repair and regeneration at different times of the day.
A single nucleotide change in thepolCDNA polymerase III inClostridium thermocellumis sufficient to create a hypermutator phenotype
Authors: Lanahan Anthony , Zakowicz Kamila , Tian Liang , Olson Daniel G. , Lynd Lee R. 
Published: August 2021

AbstractClostridium thermocellumis a thermophilic, anaerobic, bacterium that natively ferments cellulose to ethanol, and is a candidate for cellulosic biofuel production. Recently, we identified a hypermutator strain ofC. thermocellumwith a C669Y mutation in thepolCgene, which encodes a DNA polymerase III enzyme. Here we reintroduce this mutation using recently-developed CRISPR tools to demonstrate that this mutation is sufficient to recreate the hypermutator phenotype. The resulting strain shows an approximately 30-fold increase in the mutation rate. This mutation appears to function by interfering with metal ion coordination in the PHP domain responsible for proofreading. The ability to selectively increase the mutation rate inC. thermocellumis a useful tool for future directed evolution experiments.ImportanceCellulosic biofuels are a promising approach to decarbonize the heavy duty transportation sector. A longstanding barrier to cost-effective cellulosic biofuel production is the recalcitrance of the material to solubilization. Native cellulose-consuming organisms, such asClostridium thermocellum, are promising candidates for cellulosic biofuel production, however they often need to be genetically modified to improve product formation. One approach is adaptive laboratory evolution. Our findings demonstrate a way to increase the mutation rate in this industrially-relevant organism, which can reduce the time needed for adaptive evolution experiments.
Telmisartan restricts Chikungunya virus infection in vitro and in vivo through the AT1/ PPAR-γ/MAPKs pathways
Authors: De Saikat , Mamidi Prabhudutta , Ghosh Soumyajit , Keshry Supriya Suman , Mahish Chandan , Pani Sweta Smita , Laha Eshna , Ray Amrita , Datey Ankita , Chatterjee Sanchari , Singh Sharad , Mukherjee Tathagata , Khamaru Somlata , Chattopadhyay Subhasis , Subudhi Bharat Bhusan , Chattopadhyay Soma 
Published: August 2021

ABSTRACTChikungunya virus (CHIKV) has re-emerged as a global public health threat. The inflammatory pathways of RAS and PPAR-γ are usually involved in viral infections. Thus, Telmisartan (TM) with known capacity to block AT1 receptor and activate PPAR-γ, was investigated against CHIKV. The anti-CHIKV effect of TM was investigated in vitro (Vero, RAW 264.7 cells and hPBMCs) and in vivo (C57BL/6 mice). TM was found to abrogate CHIKV infection efficiently (IC50 of 15.34-20.89µM in the Vero and RAW 264.7 cells respectively). Viral RNA and proteins were reduced remarkably with the TM driven modulation of host m-TOR signaling. Additionally, TM interfered in the early and late stages of CHIKV life cycle with efficacy in both pre and post-treatment assay. Moreover, the agonist of AT1 receptor and antagonist of PPAR-γ increased CHIKV infection suggesting TM’s anti-viral potential by modulating host factors. Besides, reduced activation of all major MAPKs, NF-κB (p65) and cytokines by TM through the inflammatory axis supported the fact that the anti-CHIKV efficacy of TM is partly mediated through the AT1/PPAR-γ/MAPKs pathways. Interestingly, at the human equivalent dose, TM abrogated CHIKV infection and inflammation significantly leading to reduced clinical score and complete survival of C57BL/6 mice. Additionally, TM reduced infection in hPBMC derived monocyte-macrophage populations in vitro. Hence, TM was found to reduce CHIKV infection by targeting both viral and host factors. Considering its safety and in vivo efficacy, it can be a suitable candidate in future for repurposing against CHIKV.
External Validation of a Mobile Clinical Decision Support System for Diarrhea Etiology Prediction in Children: A Multicenter Study in Bangladesh and Mali
Authors: Garbern Stephanie C , Nelson Eric J , Nasrin Sabiha , Keita Adama Mamby , Brintz Ben J , Gainey Monique , Badji Henry , Nasrin Dilruba , Howard Joel , Taniuchi Mami , Platts-Mills James A. , Kotloff Karen L , Haque Rashidul , Levine Adam C , Sow Samba O , Alam Nur H , Leung Daniel T 
Published: August 2021

AbstractBackgroundDiarrheal illness is a leading cause of antibiotic use for children in low- and middle-income countries. Determination of diarrhea etiology at the point-of-care without reliance on laboratory testing has the potential to reduce inappropriate antibiotic use.MethodsThis prospective observational study aimed to develop and externally validate the accuracy of a mobile software application (“App”) for the prediction of viral-only etiology of acute diarrhea in children 0-59 months in Bangladesh and Mali. The App used previously derived and internally validated models using combinations of “patient-intrinsic” information (age, blood in stool, vomiting, breastfeeding status, and mid-upper arm circumference), pre-test odds using location-specific historical prevalence and recent patients, climate, and viral seasonality. Diarrhea etiology was determined with TaqMan Array Card using episode-specific attributable fraction (AFe) >0.5.ResultsOf 302 children with acute diarrhea enrolled, 199 had etiologies above the AFe threshold. Viral-only pathogens were detected in 22% of patients in Mali and 63% in Bangladesh. Rotavirus was the most common pathogen detected (16% Mali; 60% Bangladesh). The viral seasonality model had an AUC of 0.754 (0.665-0.843) for the sites combined, with calibration-in-the-large α=-0.393 (−0.455 – -0.331) and calibration slope β=1.287 (1.207 – 1.367). By site, the pre-test odds model performed best in Mali with an AUC of 0.783 (0.705 - 0.86); the viral seasonality model performed best in Bangladesh with AUC 0.710 (0.595 - 0.825).ConclusionThe App accurately identified children with high likelihood of viral-only diarrhea etiology. Further studies to evaluate the App’s potential use in diagnostic and antimicrobial stewardship are underway.
COVID-19 vaccine response in people with multiple sclerosis
Authors: Tallantyre Emma C , Vickaryous Nicola , Anderson Valerie , Asardag Aliye Nazli , Baker David , Bestwick Jonathan , Bramhall Kath , Chance Randy , Evangelou Nikos , George Katila , Giovannoni Gavin , Grant Leanne , Harding Katharine E , Hibbert Aimee , Ingram Gillian , Jones Meleri , Kang Angray S , Loveless Samantha , Moat Stuart J , Robertson Neil P , Schmierer Klaus , Shah Sita Navin , Simmons Jessica , Upcott Matthew , Willis Mark , Jolles Stephen , Dobson Ruth 
Published: August 2021

AbstractObjectiveTo investigate the effect of disease modifying therapies on serological response to SARS-CoV2 vaccines in people with multiple sclerosisMethods473 people with multiple sclerosis from 5 centres provided one or more dried blood spot samples and questionnaires about COVID-19. Information about disease and drug history was extracted from their medical records. Dried blood spots were eluted and tested for antibodies to SARS-CoV2 receptor binding domain. Seropositivity was expressed according to validated cut-off indices. Antibody titers were partitioned into tertiles using data from people on no disease modifying therapy as a reference. We calculated the odds ratio of seroconversion (Univariate logistic regression) and compared quantitative vaccine response (Kruskal Wallis) following SARS-CoV2 vaccine according to disease modifying therapy. We used regression modelling to explore the effect of factors including vaccine timing, treatment duration, age, vaccine type and lymphocyte count on vaccine response.ResultsCompared to no disease modifying therapy, the use of anti-CD20 monoclonal antibodies (odds ratio 0.03; 95% confidence interval 0.01-0.06, p<0.001) and fingolimod (odds ratio 0.41; 95% confidence interval 0.01-0.12) were associated with lower seroconversion following SARS-CoV2 vaccine. All other drug groups did not differ significantly from the untreated cohort. Time since last anti-CD20 treatment and total time on treatment were significantly related with response to vaccination. Vaccine type significantly predicted seroconversion, but not in those on anti-CD20 medications.InterpretationSome disease modifying therapies carry a risk of attenuated response to SARS-CoV2 vaccination in people with MS. We provide recommendations for the practical management of this patient group.