bluuebunny/crossref_metadata_2025_split huggingface dataset:
Design, 3D-printing, and characterisation of a low-cost, open-source centrifuge adaptor for separating large volume clinical blood samples
Authors: Haque Md Ehtashamul , Marriott Linda , Naeem Noman , Henry Taygan , Conde Alvaro J. , Kersaudy-Kerhoas Maïwenn 
Published: October 2021

AbstractBlood plasma separation is a prerequisite in numerous biomedical assays involving low abundance plasma-borne biomarkers and thus is the fundamental step before many bioanalytical steps. Conventionally, plasma separation is performed using high-capacity refrigerated centrifuges which have the advantage of handling large volume blood samples. These centrifuges are bulky, and prohibitively expensive for low-resource settings, with prices starting from $1,500. Although commercial and existing open-source micro-centrifuges are relatively low-cost, they cannot handle large volume blood samples. Microfluidic blood plasma separation also has been adopted by many researchers to enable low-cost plasma separation, however, these systems still present yield and purity issues for extremely low abundance biomarker detection such as the detection of various fractions of circulating cell-free DNA. To overcome this, we customised the rotor of a commercially available micro-centrifuge ∼$125) using fused filament fabrication to enable centrifugation of large clinical blood samples in resource poor-settings. Our designed adaptor ($15) can hold two 9 mL S-Monovette tubes and maintain the same separation performance (yield, cell count, hemolysis, albumin levels) as the control benchtop refrigerated centrifuge. This low-cost open-source centrifugation system capable of processing clinical blood tubes could be valuable to low-funded laboratories or low-resource settings where centrifugation is required immediately after blood withdrawal for further testing.
Chromosome-encoded IpaH ubiquitin ligases indicate non-human pathogenic Escherichia
Authors: Dranenko Natalia O. , Tutukina Maria , Gelfand Mikhail S. , Kondrashov Fyodor A. , Bochkareva Olga O. 
Published: October 2021

AbstractUntil recently, Shigella and enteroinvasive Escherichia coli were thought to be primate-restricted pathogens. The base of their pathogenicity is the type 3 secretion system (T3SS) encoded by the pINV virulence plasmid, which facilitates host cell invasion and subsequent proliferation. A large family of T3SS effectors, E3 ubiquitin-ligases encoded by the ipaH genes, have a key role in the Shigella pathogenicity through the modulation of cellular ubiquitination that degrades host proteins. However, recent genomic studies identified ipaH genes in the genomes of Escherichia marmotae, a potential marmot pathogen, and an E. coli extracted from fecal samples of bovine calves, suggesting that non-human hosts may also be infected by these strains, potentially pathogenic to humans.We performed a comparative genomic study of the functional repertoires in the ipaH gene family in Shigella and enteroinvasive Escherichia from human and predicted non-human hosts. We found that fewer than half of Shigella genomes had a complete set of ipaH genes, with frequent gene losses and duplications that were not consistent with the species tree and nomenclature. Non-human host IpaH proteins had a diverse set of substrate-binding domains and, in contrast to the Shigella proteins, two different types of the NEL C-terminal domain. Only the ipaH9.8 gene was found in Escherichia derived from both human and non-human hosts. These results provide a framework for understanding of ipaH-mediated host-pathogens interactions and suggest a need for a genomic study of fecal samples from diseased animals.
A kinesin-1 variant reveals motor-induced microtubule damage in cells
Authors: Budaitis Breane G. , Badieyan Somayesadat , Yue Yang , Blasius T. Lynne , Reinemann Dana N. , Lang Matthew J. , Cianfrocco Michael A. , Verhey Kristen J. 
Published: October 2021

AbstractKinesins drive the transport of cellular cargoes as they walk along microtubule tracks, however, recent work has suggested that the physical act of kinesins walking along microtubules can stress the microtubule lattice. Here, we describe a kinesin-1 KIF5C mutant with an increased ability to generate defects in the microtubule lattice as compared to the wild-type motor. Expression of the mutant motor in cultured cells resulted in microtubule breakage and fragmentation, suggesting that kinesin-1 variants with increased damage activity would have been selected against during evolution. The increased ability to damage microtubules is not due to the altered motility properties of the mutant motor as expression of the kinesin-3 motor KIF1A, which has similar single-motor motility properties, also caused increased microtubule pausing, bending, and buckling but not breakage. In cells, motor-induced microtubule breakage could not be prevented by increased a-tubulin K40 acetylation, a post-translational modification known to increase microtubule flexibility. In vitro, lattice damage induced by wild-type KIF5C was repaired by soluble tubulin and resulted in increased rescues and microtubule growth whereas lattice damage induced by the KIF5C mutant resulted in larger repair sites that made the microtubule vulnerable to breakage and fragmentation when under mechanical stress. These results demonstrate that kinesin-1 motility causes defects in and damage to the microtubule lattice in cells. While cells have the capacity to repair lattice damage, conditions that exceed this capacity result in microtubule breakage and fragmentation and may contribute to human disease.
In vitro experiments and kinetic models of pollen hydration show that MSL8 is not a simple tension-gated osmoregulator
Authors: Miller Kari , Strychalski Wanda , Nickaeen Masoud , Carlsson Anders , Haswell Elizabeth S. 
Published: October 2021

SummaryPollen, a neighbor-less cell that contains the male gametes, undergoes multiple mechanical challenges during plant sexual reproduction, including desiccation and rehydration. It was previously showed that the pollen-specific mechanosensitive ion channel MscS-Like (MSL)8 is essential for pollen survival during hydration and proposed that it functions as a tension-gated osmoregulator. Here we test this hypothesis with a combination of mathematical modeling and laboratory experiments. Time-lapse imaging revealed that wild-type pollen grains swell and then stabilize in volume rapidly during hydration.msl8mutant pollen grains, however, continue to expand and eventually burst. We found that a mathematical model wherein MSL8 acts as a simple tension-gated osmoregulator does not replicate this behavior. A better fit was obtained from variations of the model wherein MSL8 inactivation is independent of its membrane tension gating threshold or MSL8 strengthens the cell wall without osmotic regulation. Experimental and computational testing of several perturbations, including hydration in an osmolyte-rich solution, hyper-desiccation of the grains, and MSL8-YFP overexpression, indicated that the Cell Wall Strengthening Model best simulated experimental responses. Finally, expression of a non-conducting MSL8 variant did not complement themsl8overexpansion phenotype. These data indicate that, contrary to our hypothesis and to known MS ion channel function in single-cell systems, MSL8 does not act as a simple membrane tension-gated osmoregulator. Instead, they support a model wherein ion flux through MSL8 is required to alter pollen cell wall properties. These results demonstrate the utility of pollen as a cellular-scale model system and illustrate how mathematical models can correct intuitive hypotheses.
T cell receptor and IL-2 signaling strength control memory CD8+ T cell functional fitness via chromatin remodeling
Authors: Chin Shu Shien , Guillen Erik , Chorro Laurent , Achar Sooraj , Ng Karina , Oberle Susanne , Alfei Francesca , Zehn Dietmar , Altan-Bonnet Grégoire , Delahaye Fabien , Lauvau Grégoire 
Published: October 2021

AbstractCognate antigen signal controls CD8+ T cell priming, expansion size and effector versus memory cell fates, but it is not known if and how it modulates the functional features of memory CD8+ T cells. Here we show that the strength of T cell receptor (TCR) signaling determines the requirement for interleukin-2 (IL-2) signals to form a pool of memory CD8+ T cells that competitively re-expand upon secondary antigen encounter. Combining strong TCR and intact IL-2 signaling synergistically induces genome-wide chromatin accessibility in regions targeting a wide breadth of biological processes, consistent with their greater functional fitness. Chromatin accessibility in promoters of genes encoding for stem cell, cell cycle and calcium-related proteins correlated with faster intracellular calcium accumulation, initiation of cell cycle and more robust expansion. High-dimensional flow-cytometry analysis also highlights higher subset diversity and phenotypes. These results formally establish that epitope selection in vaccine design strongly impacts memory CD8+ T cell epigenetic programming and functions.One Sentence SummaryThe strength of antigenic and interleukin 2 signals received by CD8+ T cells during vaccination epigenetically programs their ability to form functional memory.
A within-host infection model to explore tolerance and resistance
Authors: Duneau David , Lafont Pierre D. M. , Lauzeral Christine , Parthuisot Nathalie , Faucher Christian , Jin Xuerong , Buchon Nicolas , Ferdy Jean-Baptiste 
Published: October 2021

AbstractHow are some individuals surviving infections while others die? The answer lies in how infected individuals invest into controlling pathogen proliferation and mitigating damage, two strategies respectively called resistance and disease tolerance. Pathogen within-host dynamics (WHD), influenced by resistance, and its connection to host survival, determined by tolerance, decide the infection outcome. To grasp these intricate effects of resistance and tolerance, we used a deterministic theoretical model where pathogens interact with the immune system of a host. The model describes the positive and negative regulation of the immune response, consider the way damage accumulate during the infection and predicts WHD. When chronic, infections stabilize at a Set-Point Pathogen Load (SPPL). Our model predicts that this situation can be transient, the SPPL being then a predictor of life span which depends on initial condition (e.g. inoculum). When stable, the SPPL is rather diagnostic of non lethal chronic infections. In lethal infections, hosts die at a Pathogen Load Upon Death (PLUD) which is almost independent from the initial conditions. As the SPPL, the PLUD is affected by both resistance and tolerance but we demonstrate that it can be used in conjunction with mortality measurement to distinguish the effect of disease tolerance from that of resistance. We validate empirically this new approach, usingDrosophila melanogasterand the pathogenProvidencia rettgeri. We found that, as predicted by the model, hosts that were wounded or deficient of key antimicrobial peptides had a higher PLUD, while Catalase mutant hosts, likely to have a default in disease tolerance, had a lower PLUD.
Interaction of human CRX and NRL in live HEK293T cells measured using fluorescence resonance energy transfer (FRET)
Authors: Zhuo Xinming , Knox Barry E. 
Published: October 2021

AbstractCRX and NRL are retina-specific transcription factors that control rod photoreceptor differentiation and synergistically activate rod phototransduction gene expression. Previous experiments showed they interact in vitro and in yeast two-hybrid assays. Here, we examined CRX-NRL interaction in live HEK293T cells using two fluorescence resonance energy transfer (FRET) approaches: confocal microscopy and flow cytometry (FC-FRET). FC-FRET can provide measurements from many cells having wide donor-acceptor expression ranges. FRET efficiencies were calibrated with a series of donor (EGFP)-acceptor (mCherry) fusion proteins separated with linkers between 6-45 amino acids. CRX and NRL were fused at either terminus with EGFP or mCherry to create fluorescent proteins, and all combinations were tested in transiently transfected cells. FRET signals between CRX or NRL homo-pairs were highest with both fluorophores fused to the DNA binding domains (DBD), lower with both fused to the activation domains (AD), and not significant when fused on opposite termini. NRL had stronger FRET signals than CRX. A significant FRET signal between CRX and NRL hetero-pairs was detected when donor was fused to the CRX DNA binding domain and the acceptor fused to the NRL activation domain. FRET signals increased with CRX or NRL expression levels at a rate much higher than expected for collisional FRET alone. Together, our results show the formation of CRX-NRL complexes in live HEK293T cells that are close enough for FRET.
BioCompute Objects to communicate a viral detection pipeline with potential for use in a regulatory environment
Authors: Gulzar Naila , Keeney Jonathon , Baker Jack B. , Klempir Ondrej , Hannigan Geoffrey , Bitton Danny A. , Maritz Julia M , King Charles Hadley S. , Patel Janisha A. , Duncan Paul , Mazumder Raja 
Published: October 2021

AbstractThe volume of nucleic acid sequence data has exploded in recent years, and with it, the challenge of finding and transforming relevant data into meaningful information. Processing the abundance of data can require a dynamic ecosystem of customized tools. As analysis pipelines become more complex, there is an increased difficulty in communicating analysis details in a way that is understandable yet of sufficient detail to make informed decisions about results or repeat the analysis. This may be of particular interest to institutions and private companies that need to communicate complex computations in a regulatory environment. To meet this need for standard reporting, the open source BioCompute framework was developed as a standardized mechanism for communicating the details of an analysis in a concise and organized way, and other tools and interfaces were subsequently developed according to the standard. The goal of BioCompute is to streamline the process of communicating computational analyses. Reports that conform to the BioCompute standard are called BioCompute Objects (BCOs). Here, a comprehensive suite of BCOs is presented, representing interconnected elements of a computation that is modeled after those that might be found in a regulatory submission, but which can be shared publicly. Because BCOs are human and machine readable, they can be displayed in customized ways to further improve their utility, and an example of a collapsible format is shown. The work presented here serves as a real world implementation that imitates actual submissions, providing concrete examples. As an example, a pipeline designed to identify viral contaminants in biological manufacturing, such as for vaccines, is developed and rigorously tested to establish a rate of false positive detection, and is described in a BCO report. That pipeline relies on a specially curated database for alignment, and a set of synthetic reads for testing, both of which are also descriptively packaged in their own BCOs. All of the sufficiently complex processes associated with this analysis are therefore represented as BCOs that can be cross-referenced, demonstrating the modularity of BCOs, their ability to organize tremendous complexity, and their use in a lifelike regulatory environment.
KIMGENS: A novel method to estimate kinship in organisms with mixed haploid diploid genetic systems robust to population structure
Authors: Wang Yen-Wen , Ané Cécile 
Published: October 2021

AbstractMotivationKinship estimation is necessary for evaluating violations of assumptions or testing certain hypotheses in many population genomic studies. However, kinship estimators are usually designed for diploid systems and cannot be used in populations with mixed haploid diploid genetic systems. The only estimators for different ploidies require datasets free of population structure, limiting their usage.ResultsWe present KIMGENS, an estimator for kinship estimation among individuals of various ploidies, that is robust to population structure. This estimator is based on the popular KING-robust estimator but uses diploid relatives of the individuals of interest as references of heterozygosity and extends its use to haploid-diploid and haploid pairs of individuals. We demonstrate that KIMGENS estimates kinship more accurately than previously developed estimators in simulated panmictic, structured and admixed populations, but has lower accuracy when the individual of interest is inbred. KIMGENS also outperforms other estimators in a honeybee dataset. Therefore, KIMGENS is a valuable addition to a population geneticist’s toolbox.Availability and ImplementationKIMGENS and its association simulation tool are implemented and available open-source at https://github.com/YenWenWang/HapDipKinship.ContactYen-Wen Wang Email: ywang883@wisc.edu
Novel graph theoretic biological pathway network analytics methods for analyzing and discovering Alzheimer’s disease related genes
Authors: Saha Subrata , Soliman Ahmed , Rajasekaran Sanguthevar 
Published: October 2021

AbstractBackgroundAlzheimer’s disease (AD) is the most common form of dementia among older people. It is a complex disease and the genetics and environmental factors behind it are not conclusive yet. Traditional statistical analyses are inadequate to identify variants, genes, or pathways capable of explaining AD as a unit. In this context, pathway network analysis based on a set of curated AD-specific genes identified in the literature can elucidate biological mechanisms underneath AD. Through the network, we can infer influential pathways that can together explain AD. Consequently, we can target those pathways and corresponding genes for further analysis to develop new drugs, discover novel AD-related genes, combine multiple hypotheses, and so forth.MethodsWe have developed a novel graph theoretic algorithm that can elucidate complex biology from a given set of disease-related genes. It constructs a weighted network of enriched pathways where similarity score between a pair of pathways is defined in a context-specific manner. To make the network robust, we employ topological overlap techniques on top of the raw similarity measure. We then provide the importance of each pathway with respect to the entire network, functional modules and importance of each pathway in a specific module, gene clusters, and so forth. We also provide a method to identify a set of novel genes that can further explain the disease-related genes and the disease itself.ResultsWe have employed our algorithms onto a set of AD-specific genes. It identified three distinct functional modules that are related to metabolism, cancer, and infectious disease related pathways. These findings are matched with three recognized hypotheses in Alzheimer’s disease, e.g. “metabolism hypothesis,” “cell cycle hypothesis,” and “infectious disease hypothesis.” By analyzing the curated genes common among those functional modules, we can attain more understanding about this fateful disease. We have also identified 24 novel AD-related genes of which at least 14 genes are known to be involved in AD.ConclusionsWe developed a computational framework for analyzing biological pathways in a context-specific manner. It can be used in any sets of disease-related genes. We manifest its efficacy, reliability, and accuracy by employing a set of AD-specific genes.
Nanoscale organization of ryanodine receptor distribution and phosphorylation pattern determines the dynamics of calcium sparks
Authors: Hernández Mesa M. , van den Brink J. , Louch W.E. , McCabe K.J. , Rangamani P. 
Published: October 2021

AbstractSuper-resolution imaging techniques have provided a better understanding of the relationship between the nanoscale organization of function of ryanodine receptors (RyRs) in cardiomy-ocytes. Interestingly recent data have indicated that this relationship is disrupted in heart failure (HF), as RyRs are dispersed into smaller and more numerous clusters. However, RyRs are also hyperphosphorylated in this condition, and this is reported to occur preferentially within the cluster centre. Thus, the combined impact of RyR relocalization and sensitization on Ca2+ spark generation in failing cardiomyocytes is likely complex and these observations suggest that both the nanoscale organization of RyRs and the pattern of phosphorylated RyRs within clusters could be critical determinants of Ca2+ spark dynamics. To test this hypothesis, we used computational modeling to quantify the relationships between RyR cluster geometry, phosphorylation patterns, and sarcoplasmic reticulum (SR) Ca2+ release. We found that RyR cluster disruption results in a decrease in spark fidelity and longer sparks with a lower amplitude. Phosphorylation of some RyRs within the cluster can play a compensatory role, recovering healthy spark dynamics. Interestingly, our model predicts that such compensation is critically dependent on the phosphorylation pattern, as phosphorylation localized within the cluster center resulted in longer Ca2+ sparks and higher spark fidelity compared to a uniformly distributed phosphorylation pattern. Our results strongly suggest that both the phosphorylation pattern and nanoscale RyR reorganization are critical determinants of Ca2+ dynamics in HF.Significance StatementRyRs are ion channels located on the membrane of the sarcoplasmic reticulum that are responsible for an increase in cytosolic Ca2+ during cell excitation. Here, we investigate how the geometry of RyR clusters combined with spatial phosphorylation patterns impacts on Ca2+ spark generation and kinetics. The findings from our study show that both phosphorylation pattern and RyR cluster shape and dispersion have implications on Ca2+ spark activity and provide insights into altered Ca2+ dynamics during HF.
Survival of hatchery grown native oysters is associated with specific gut-associated bacteria at various locations within the Puget Sound
Authors: Kunselman Emily , Minich Jeremiah J , Horwith Micah , Gilbert Jack A , Allen Eric E 
Published: October 2021

AbstractThe Olympia oyster (Ostrea lurida) of the Puget Sound suffered a dramatic population crash, but restoration efforts hope to revive this native species. One overlooked variable in the process of assessing ecosystem health is association of bacteria with marine organisms and the environments they occupy. Oyster microbiomes are known to differ significantly between species, tissue type, and the habitat in which they are found. The goals of this study were to determine the impact of field site and habitat on the oyster microbiome and to identify core oyster-associated bacteria in the Puget Sound. Olympia oysters from one parental family were deployed at four sites in the Puget Sound both inside and outside of eelgrass (Zostera marina) beds. Using 16S rRNA gene amplicon sequencing of the oyster gut, shell, surrounding seawater and sediment, we demonstrate that gut-associated bacteria are distinct from the surrounding environment and vary by field site. Furthermore, regional differences in the gut microbiota are associated with the survival rates of oysters at each site after two months of field exposure. However, habitat type had no influence on microbiome diversity. Further work is needed to identify the specific bacterial dynamics that are associated with oyster physiology and survival rates.ImportanceThe outcomes of this study demonstrate the need to monitor microbial ecology at potential oyster restoration sites, specifically for the native Olympia oyster in the Puget Sound. Some areas of Puget Sound may be less amenable to Olympia oyster restoration than others due to microbiome colonization trends. Furthermore, this study puts the oyster microbiome in the context of its surroundings, providing a holistic perspective on the factors that may influence oyster performance and microbial ecology.