bluuebunny/crossref_metadata_2025 huggingface dataset:
FGF21 defines a potential cardio-hepatic signaling circuit in human heart failure
Authors: Sommakia Salah , Almaw Naredos H. , Lee Sandra H. , Ramadurai Dinesh K. A. , Taleb Iosef , Kyriakopoulos Christos P. , Stubben Chris J. , Ling Jing , Campbell Robert A. , Alharethi Rami A. , Caine William T. , Navankasattusas Sutip , Hoareau Guillaume L. , Abraham Anu E. , Fang James C. , Selzman Craig H. , Drakos Stavros G. , Chaudhuri Dipayan
Published: July 2021

Background

Extrinsic control of cardiac contractility and ultrastructure via neurohormonal signaling is well established, but how other organs regulate cardiomyocyte metabolism is less well understood. Fibroblast growth factor-21 (FGF21) a hormonal regulator of metabolism mainly produced in the liver and adipose tissue, is a prime candidate for such signaling.

Methods

To investigate this further, we examined blood and tissue obtained from human subjects with heart failure with reduced ejection fraction (HFrEF) at the time of left ventricular assist device (LVAD) implantation, and correlated serum FGF21 levels with cardiac gene expression, immunohistochemistry, and clinical parameters.

Results

Circulating FGF21 levels were substantially elevated in HFrEF, compared to healthy subjects (HFrEF: 834.4 ± 101.8 pg/mL, n = 40; controls: 145.9 ± 28.6 pg/mL, n = 20, p = 5.5 × 10−8). There was clear FGF21 staining in diseased cardiomyocytes, and circulating FGF21 levels negatively correlated with the expression of cardiac genes involved in ketone metabolism, consistent with cardiac FGF21 signaling. FGF21 gene expression was low in failing and non-failing hearts, suggesting at least partial extracardiac production of the circulating hormone. Circulating FGF21 levels were correlated with BNP and total bilirubin, markers of chronic cardiac and hepatic congestion.

Conclusions

Circulating FGF21 levels are elevated in HFrEF. The liver is likely the main extracardiac source, and congestive hepatopathy, common in HFrEF, was likely the proximate signal leading to FGF21 elevations. This supports a model of venous congestion from cardiomyopathy driving hepatic FGF21 communication to diseased cardiomyocytes, defining a potential cardio-hepatic signaling circuit in human heart failure.
Clonally expanded virus-specific CD8 T cells acquire diverse transcriptional phenotypes during acute, chronic, and latent infections
Authors: Kuhn Raphael , Sandu Ioana , Agrafiotis Andreas , Hong Kai-Lin , Neumeier Daniel , Merkler Doron , Oxenius Annette , Reddy Sai T. , Yermanos Alexander
Published: July 2021

CD8+ T cells play a crucial role in the control and resolution of viral infections and can adopt a wide range of phenotypes and effector functions depending on the inflammatory context and the duration and extent of antigen exposure. Similarly, viral infections can exert diverse selective pressures on populations of clonally related T cells. Technical limitations have nevertheless made it challenging to investigate the relationship between clonal selection and transcriptional phenotypes of virus-specific T cells. We therefore performed single-cell T cell receptor (TCR) repertoire and transcriptome sequencing of virus-specific CD8 T cells in murine models of acute, chronic and latent infection. We observed clear infection-specific populations corresponding to memory, effector, exhausted, and inflationary phenotypes. We further uncovered a mouse-specific and polyclonal T cell response, despite all T cells sharing specificity to a single viral epitope, which was accompanied by stereotypic TCR germline gene usage in all three infection types. Persistent antigen exposure during chronic and latent viral infections resulted in a higher proportion of clonally expanded T cells relative to acute infection. We furthermore observed a relationship between transcriptional heterogeneity and clonal expansion for all three infections, with highly expanded clones having distinct transcriptional phenotypes relative to lowly expanded clones. Finally, we developed and utilized a bioinformatic pipeline integrating pseudotime and clonality, termed Clonotyme, to further support a model in which expanded virus-specific CD8+ T cells adopt heterogenic, yet preferentially, effector-like phenotypes. Together our work relates clonal selection to gene expression in the context of viral infection and further provides a dataset and accompanying software for the immunological community.
Adaptation in cone photoreceptors contributes to an unexpected insensitivity of On parasol retinal ganglion cells to spatial structure in natural images
Authors: Yu Zhou , Turner Maxwell H. , Baudin Jacob , Rieke Fred
Published: July 2021

Neural circuits are constructed from nonlinear building blocks, and not surprisingly overall circuit behavior is often strongly nonlinear. But neural circuits can also behave near linearly, and some circuits shift from linear to nonlinear behavior depending on stimulus conditions. Such control of the linearity or nonlinearity of circuit behavior is fundamental to neural computation. Here we study a surprising stimulus dependence of the responses of On (but not Off) parasol retinal ganglion cells: these cells respond nonlinearly to spatial structure in temporally-modulated grating stimuli but linearly to spatial structure in flashed gratings and natural visual inputs. We show that this unexpected response linearity can be explained by a shift in the balance of excitatory and inhibitory synaptic inputs that originates at least in part from adaptation in the cone photoreceptors. More generally, this highlights how subtle asymmetries in signaling - here in the cone signals - can qualitatively alter circuit computation.
Classifying COVID-19 variants based on genetic sequences using deep learning models
Authors: Basu Sayantani , Campbell Roy H.
Published: July 2021

The COrona VIrus Disease (COVID-19) pandemic led to the occurrence of several variants with time. This has led to an increased importance of understanding sequence data related to COVID-19. In this chapter, we propose an alignment-free k-mer based LSTM (Long Short-Term Memory) deep learning model that can classify 20 different variants of COVID-19. We handle the class imbalance problem by sampling a fixed number of sequences for each class label. We handle the vanishing gradient problem in LSTMs arising from long sequences by dividing the sequence into fixed lengths and obtaining results on individual runs. Our results show that one-vs-all classifiers have test accuracies as high as 92.5% with tuned hyperparameters compared to the multi-class classifier model. Our experiments show higher overall accuracies for B.1.1.214, B.1.177.21, B.1.1.7, B.1.526, and P.1 on the one-vs-all classifiers, suggesting the presence of distinct mutations in these variants. Our results show that embedding vector size and batch sizes have insignificant improvement in accuracies, but changing from 2-mers to 3-mers mostly improves accuracies. We also studied individual runs which show that most accuracies improved after the 20th run, indicating that these sequence positions may have more contributions to distinguishing among different COVID-19 variants.
A quantitative trait variant inGabra2underlies increased methamphetamine stimulant sensitivity
Authors: Goldberg Lisa R. , Yao Emily J. , Kelliher Julia C. , Reed Eric R. , Cox Jiayi Wu , Parks Cory , Kirkpatrick Stacey L. , Beierle Jacob A. , Chen Melanie M. , Johnson William E. , Homanics Gregg E. , Williams Robert W. , Bryant Camron D. , Mulligan Megan K.
Published: July 2021

Psychostimulant (methamphetamine, cocaine) use disorders have a genetic component that remains mostly unknown. Here, we conducted genome-wide quantitative trait locus (QTL) analysis of methamphetamine stimulant sensitivity. To facilitate gene identification, we employed a Reduced Complexity Cross between closely related C57BL/6 mouse substrains and examined maximum speed and distance traveled over 30 min following methamphetamine (2 mg/kg, i.p.). For maximum methamphetamine-induced speed following the second and third administration, we identified a single genome-wide significant QTL on chromosome 11 that peaked near theCyfip2locus [LOD = 3.5, 4.2; peak = 21 cM (36 Mb)]. For methamphetamine-induced distance traveled, we identified a single genome-wide significant QTL on chromosome 5 that peaked near a functional intronic indel inGabra2that codes for the alpha-2 subunit of the GABA-A receptor [LOD = 5.2; peak = 35 cM (67 Mb)]. Striatalcis-expression QTL mapping corroboratedGabra2as a functional candidate gene underlying methamphetamine-induced distance traveled. CRISPR/Cas9-mediated correction of the mutant intronic deletion on the C57BL/6J background to the wild-type C57BL/6NJ allele was sufficient to reduce methamphetamine-induced locomotor activity toward the wild-type C57BL/6NJ-like level, thus validating the quantitative trait variant (QTV). These studies demonstrate the power and efficiency of Reduced Complexity Crosses in identifying causal genes and variants underlying complex traits. Functionally restoringGabra2expression decreased methamphetamine stimulant sensitivity and supports preclinical and human genetic studies implicating the GABA-A receptor in psychostimulant addiction-relevant traits. Importantly, our findings have major implications for investigators studying psychostimulants in the C57BL/6J strain - the gold standard strain in biomedical research.
Temperature Evolution Following Joint Loading Promotes Chondrogenesis by Synergistic Cues via Calcium Signaling
Authors: Nasrollahzadeh Naser , Karami Peyman , Wang Jian , Bagheri Lida , Guo Yanheng , Abdel-Sayed Philippe , Applegate Lee Ann , Pioletti Dominique P.
Published: July 2021

During loading of viscoelastic tissues, part of the mechanical energy is transformed into heat that can locally increase the tissue temperature, a phenomenon known as self-heating. In the framework of mechanobiology, it has been accepted that cells react and adapt to mechanical stimuli. However, the cellular effect of temperature increase as a by-product of loading has been widely neglected. In this work, we focused on cartilage self-heating to present a “thermo-mechanobiological” paradigm, and demonstrate how the synergy of a biomimetic temperature evolution and mechanical loading could influence cell behavior. We thereby developed a customized in vitro system allowing to recapitulate pertinent in vivo physical cues and determined the cells chondrogenic response to thermal and/or mechanical stimuli. Cellular mechanisms of action and potential signaling pathways of thermo-mechanotransduction process were also investigated. We found that co-existence of thermo-mechanical cues had a superior effect on chondrogenic gene expression compared to either signal alone. Specifically, a synergetic effect was observed for upregulation of Sox9 by application of the physiological thermo-mechanical stimulus. Multimodal TRPV4 channels were identified as key mediators of thermo-mechanotransduction process, which becomes ineffective without external calcium sources. We also observed that the isolated temperature evolution, as a by-product of loading, is a contributing factor to the cells response and this could be considered as important as the conventional mechanical loading. Providing an optimal thermo-mechanical environment by synergy of heat and loading portrays new opportunity for development of novel treatments for cartilage regeneration and can furthermore signal key elements for emerging cell-based therapies.
Efficient linear dsDNA tagging using deoxyuridine excision
Authors: Strobel Eric J.
Published: July 2021

Site-specific strategies for exchanging segments of dsDNA are important for DNA library construction and molecular tagging. Deoxyuridine (dU) excision is an approach for generating 3’ ssDNA overhangs in gene assembly and molecular cloning procedures. Unlike approaches that use a multi-base pair motif to specify a DNA cut site, dU excision requires only a dT→dU substitution. Consequently, excision sites can be embedded in biologically active DNA sequences by placing dU substitutions at non-perturbative positions. In this work, I describe a molecular tagging method that uses dU excision to exchange a segment of a dsDNA strand with a long synthetic oligonucleotide. The core workflow of this method, called deoxyUridine eXcision-tagging (dUX-tagging), is an efficient one-pot reaction: strategically positioned dU nucleotides are excised from dsDNA to generate a 3’ overhang so that additional sequence can be appended by annealing and ligating a tagging oligonucleotide. The tagged DNA is then processed by one of two procedures to fill the 5’ overhang and remove excess tagging oligo. To facilitate its widespread use, all dUX-tagging procedures exclusively use commercially available reagents. As a result, dUX-tagging is a concise and easily implemented approach for high-efficiency linear dsDNA tagging.
A genetic toolkit for studying transposon control in the Drosophila melanogaster ovary
Authors: ElMaghraby Mostafa F. , Tirian Laszlo , Senti Kirsten-André , Meixner Katharina , Brennecke Julius
Published: July 2021

Argonaute proteins of the PIWI class complexed with PIWI-interacting RNAs (piRNAs) protect the animal germline genome by silencing transposable elements. One of the leading experimental systems for studying piRNA biology is the Drosophila melanogaster ovary. In addition to classical mutagenesis, transgenic RNA interference (RNAi), which enables tissue-specific silencing of gene expression, plays a central role in piRNA research. Here, we establish a versatile toolkit focused on piRNA biology that combines germline transgenic RNAi, GFP marker lines for key proteins of the piRNA pathway, and reporter transgenes to establish genetic hierarchies. We compare constitutive, pan-germline RNAi with an equally potent transgenic RNAi system that is activated only after germ cell cyst formation. Stage-specific RNAi allows us to investigate the role of genes essential for germline cell survival, for example nuclear RNA export or the SUMOylation pathway, in piRNA-dependent and independent transposon silencing. Our work forms the basis for an expandable genetic toolkit provided by the Vienna Drosophila Resource Center.
Varenicline Prevents SARS-CoV-2 Infection In Vitro and in Rhesus Macaques
Authors: Nau Jeffrey , Luthra Priya , Lanzer Kathleen , Szaba Frank , Cookenham Tres , Carlson Eric
Published: July 2021

Background

SARS-CoV-2 infections have resulted in a global pandemic, but an antiviral therapy for this novel strain of coronavirus does not currently exist. The objective of our study was to investigate the antiviral potential of the nicotinic acetylcholine receptor (nACHR) agonist varenicline tartrate against SARS-CoV-2.

Methods

We assessed antiviral activity usingin vitrohuman cell assays and we assessedin vivoefficacy in a rhesus macaque model.

Results

In vitrostudies found that varenicline tartrate, over a range of concentrations, reduced the infectivity of SARS-CoV-2 wildtype, alpha, and beta variants in Calu-3 cells and Caco-2 cells, with maintenance of cell viability.In vivostudies found that varenicline tartrate, administered as a nasal spray to rhesus macaques, reduced SARS-CoV-2 wildtype viral load and inhibited viral replication in the nasal mucosa and upper airway.

Conclusion

Although the study reported here was exploratory, we have confirmed that the nAChR agonist varenicline has the potential to interact with and inhibit SARS-CoV-2 infection and replication.
Predicting trait phenotypes from knowledge of the topology of gene networks
Authors: Beatty Andy , Winkler Christopher R. , Hagen Thomas , Cooper Mark
Published: July 2021

In many fields there is interest in manipulating genes and gene networks to realize improved trait phenotypes. The practicality of doing so, however, requires accepted theory on the properties of gene networks that is well-tested by empirical results. The extension of quantitative genetics to include models that incorporate properties of gene networks expands the long tradition of studying epistasis resulting from gene-gene interactions. Here we consider NK models of gene networks by applying concepts from graph theory and Boolean logic theory, motivated by a desire to model the parameters that influence predictive skill for trait phenotypes under the control of gene networks; N defines the number of graph nodes, the number of genes in the network, and K defines the number of edges per node in the graph, representing the gene-gene interactions. We define and consider the attractor period of an NK network as an emergent trait phenotype for our purposes. A long-standing theoretical treatment of the dynamical properties of random Boolean networks suggests a transition from long to short attractor periods as a function of the average node degree K and the bias probability P in the applied Boolean rules. In this paper we investigate the appropriateness of this theory for predicting trait phenotypes on random and real microorganism networks through numerical simulation. We show that: (i) the transition zone between long and short attractor periods depends on the number of network nodes for random networks; (ii) networks derived from metabolic reaction data on microorganisms also show a transition from long to short attractor periods, but at higher values of the bias probability than in random networks with similar numbers of network nodes and average node degree; (iii) the distribution of phenotypes measured on microorganism networks shows more variation than random networks when the bias probability in the Boolean rules is above 0.75; and (iv) the topological structure of networks built from metabolic reaction data is not random, being best approximated, in a statistical sense, by a lognormal distribution. The implications of these results for predicting trait phenotypes where the genetic architecture of a trait is a gene network are discussed.
Toxicological profile of Umbilical Cord Blood-derived small Extracellular Vesicles
Authors: Rodrigues S.C. , Cardoso R.M.S. , Gomes C.F. , Duarte F.V. , Freire P. C. , Neves R. , Simões-Correia J.
Published: July 2021

The development and adoption of cell therapies has been largely limited by difficulties associated with their safety, handling and storage. Extracellular vesicles (EV) have recently emerged as a likely mediator for the therapeutic effect of cells, offering several advantages over cell therapies. Due to their small size and inability to expand and metastasize, EV are generally considered safer than cell transplantation. Nevertheless, few studies have scrutinized the toxicity profile of EV, particularly after repeated high dose administration. The present study aimed to evaluate a preparation of small EV obtained from umbilical cord blood mononuclear cells (UCB-MNC-sEV) for its cytotoxicity in different cell lines, as well as its differential accumulation, distribution and toxicity following repeated intravenous (IV) administrations in a rodent model. In vitro, repeated sEV exposure in concentrations up to 1×1011 particles/ml had no deleterious impact on the viability or metabolic activity of peripheral blood mononuclear cells, THP-1 monocytes, THP-1-derived macrophages, normal dermal human fibroblasts or human umbilical vein endothelial cells. DiR-labeled sEV, injected IV for four weeks in healthy rats, were detected in clearance organs, particularly kidneys, spleen and liver, similarly to control dye. Moreover, repeated administrations during six and twelve weeks of up to 1×1010 total particles of sEV-dye were well tolerated, with no changes in general hematological cell counts, or kidney and liver toxicity markers. Importantly, unlabeled sEV likewise did not induce significant alterations in cellular and biochemical blood parameters, nor any morphological changes in heart, kidney, lung, spleen, or liver tissue. In sum, our data shows that UCB-MNC-sEV have no significant toxicity in vitro or in vivo, even when administered repeatedly at high concentrations, therefore confirming their safety profile and potential suitability for future clinical use.
Early neuroadaptations to an obesogenic diet identify the schizophrenia-related ErbB4 receptor in obesity-induced hippocampal abnormalities
Authors: Vega-Torres Julio David , Ontiveros-Angel Perla , Terrones Esmeralda , Stuffle Erwin C. , Solak Sara , Tyner Emma , Oropeza Marie , Peña Ike Dela , Obenaus Andre , Ford Byron D. , Figueroa Johnny D.
Published: July 2021

Childhood obesity leads to hippocampal atrophy and altered cognition. However, the molecular mechanisms underlying these impairments are poorly understood. The neurotrophic factor neuregulin-1 (NRG1) and its cognate ErbB4 receptor play critical roles in hippocampal maturation and function. This study aimed to determine whether altered NRG1-ErbB4 activities may partly explain hippocampal abnormalities in rats exposed to an obesogenic Western-like diet (WD). Lewis rats were randomly divided into four groups (12 rats/group): 1) control diet+vehicle (CDV); 2) CD+NRG1 (CDN) (daily intraperitoneal injections: 5 μg/kg/day; between postnatal day, PND 21-PND 41); 3) WD+VEH (WDV); 4) WD+NRG1 (WDN). Neurobehavioral assessments were performed at PND 43-49. Brains were harvested for MRI and molecular analyses at PND 49. We found that NRG1 administration reduced hippocampal volume (7%) and attenuated hippocampal-dependent cued fear conditioning in CD rats (56%). NRG1 administration reduced PSD-95 protein expression (30%) and selectively reduced hippocampal cytokine levels (IL-33, GM-CSF, CCL-2, IFN-γ) while significantly impacting microglia morphology (increased span ratio and reduced circularity). WD rats exhibited reduced right hippocampal volume (7%), altered microglia morphology (reduced density and increased lacunarity), and increased levels of cytokines implicated in neuroinflammation (IL-1α, TNF-α, IL-6). Notably, NRG1 synergized with the WD to increase hippocampal ErbB4 phosphorylation and the tumor necrosis alpha converting enzyme (TACE/ADAM17) protein levels. Together, these data suggest a novel interaction between obesogenic diet exposure and TACE/ADAM17-NRG1-ErbB4 signaling during hippocampal maturation. Our results indicate that supraoptimal ErbB4 activities may contribute to the abnormal hippocampal structure and cognitive vulnerabilities observed in obese individuals.

Highlights

Obesogenic diet consumption during adolescence induces anxiety-like behaviors before the onset of obesity-related changes in metabolism.

Obesogenic diet-driven abnormal behaviors co-occurred with alterations in hippocampal pro-inflammatory cytokine profiles.

Obesogenic diet consumption attenuates exogenous NRG1 effects on hippocampal-related behaviors and structure.

Exogenous NRG1 administration during adolescence resulted in reduced hippocampal volumes and domain-specific cognitive impairments.

Exogenous NRG1 administration has potent immunomodulatory actions and alters hippocampal microglia morphology.
Rewiring the Three-Carbon Metabolism Abrogates Multiple MAPK-Induced Cellular Dysfunctions During Metabolic Disorder
Authors: Dubé Alexandre K. , Malenfant Nicolas , Ladonne Florence , Piano Amanda , Mohammad Karamat , Bélanger Marc , Bégin Frédéric , LeBel-Guay Florence , Titorenko Vladimir I. , Bourbonnais Yves
Published: July 2021

Withdrawal notice

The authors have withdrawn their manuscript. The lipidomic data presented in the manuscript were based on an excel summary sheet provided by VIT, whose group (AP and KM) conducted the analysis using an Orbitrap mass spectrometer. However, after submitting to BioRxiv, the VIT group could not recover the RAW (primary source) files from the lipidomic platform because these files were deleted due to the maintenance protocol used for the Orbitrap mass spectrometer. It was agreed to reconduct the lipidomic analysis. As the Orbitrap mass spectrometer was out of service at that time, the lipidomic analysis was conducted with the help of a QToF mass spectrometer. Important differences were noted for the relative abundance and species of many lipids across the strains compared to the previously reported data hence casting some doubt on their interpretation. Therefore, until further analysis can be carried out the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.
spiralize: an R Package for Visualizing Data on Spirals
Authors: Gu Zuguang , Hübschmann Daniel
Published: July 2021

Spiral layout has two major advantages for data visualization. First, it is able to visualize data with long axes, which greatly improves the resolution of visualization. Second, it is efficient for time series data to reveal periodic patterns. Here we present the R package spiralize that provides a general solution for visualizing data on spirals. spiralize implements numerous graphics functions so that self-defined high-level graphics can be easily implemented by users. The flexibility and power of spiralize are demonstrated by five examples from real-world datasets.

Availability and implementation

The spiralize package and documentations are freely available at the Comprehensive R Archive Network (CRAN) https://CRAN.R-project.org/package=spiralize.

Contact

z.gu@dkfz.de or d.huebschmann@dkfz.de

Supplementary information

Supplementary data are available at Bioinformatics online.
Shortening of 3’ UTRs in most cell types composing tumor tissues implicates alternative polyadenylation in protein metabolism
Authors: Burri Dominik , Zavolan Mihaela
Published: July 2021

During pre-mRNA maturation 3’ end processing can occur at different polyadenylation sites in the 3’ untranslated region (3’ UTR) to give rise to transcript isoforms that differ in the length of their 3’UTRs. Longer 3’ UTRs contain additional cis-regulatory elements that impact the fate of the transcript and/or of the resulting protein.

Extensive alternative polyadenylation (APA) has been observed in cancers, but the mechanisms and roles remain elusive. In particular, it is unclear whether the APA occurs in the malignant cells or in other cell types that infiltrate the tumor. To resolve this, we developed a computational method, called SCUREL, that quantifies changes in 3’UTR length between groups of cells, including cells of the same type originating from tumor and control tissue. We used this method to study APA in human lung adenocarcinoma (LUAD).

SCUREL relies solely on annotated 3’UTRs and on control systems, such as T cell activation and spermatogenesis gives qualitatively similar results at much greater sensitivity compared to the previously published scAPA method.

In the LUAD samples, we find a general trend towards 3’UTR shortening not only in cancer cells compared to the cell type of origin, but also when comparing other cell types from the tumor vs. the control tissue environment. However, we also find high variability in the individual targets between patients. The findings help to understand the extent and impact of APA in LUAD, which may support improvements in diagnosis and treatment.
Accurate Detection of Convergent Mutations in Large Protein Alignments with ConDor
Authors: Morel Marie , Lemoine Frédéric , Zhukova Anna , Gascuel Olivier
Published: July 2021

Evolutionary convergences are observed at all levels, from phenotype to DNA and protein sequences, and changes at these different levels tend to be highly correlated. Notably, convergent and parallel mutations can lead to convergent changes in phenotype, such as changes in metabolism, drug resistance, and other adaptations to changing environments.

We propose a two-step approach to detect mutations under convergent evolution in protein alignments. We first select mutations that emerge more often than expected under neutral evolution and then test whether their emergences correlate with the convergent phenotype under study. The first step can be used alone when no phenotype is available, as is often the case with microorganisms. In the first step, a phylogeny is inferred from the data and used to simulate the evolution of each alignment position. These simulations are used to estimate the expected number of mutations under neutral conditions, which is compared to what is observed in the data. Next, using a comparative phylogenetic approach, we measure whether the presence of mutations occurring more often than expected correlates with the convergent phenotype.

Our method is implemented in a standalone workflow and a webserver, called ConDor. We apply ConDor to three datasets: sedges PEPC proteins, HIV reverse transcriptase and fish rhodopsin. The results show that the two components of ConDor complement each other, with an overall accuracy that compares favorably to other available tools, especially on large datasets.
Does Communal Breeding Promote an Increase in Social Immunity in Burying Beetles? A Test Case with Nicrophorus Defodiens
Authors: Suswaram M. , Belk M. , Creighton C.
Published: July 2021

Communal breeding is a reproductive system in which more than a single pair of individuals share parental care duties. Burying beetles (genus Nicrophorus) breed on small vertebrate carcasses, which is used as a food source for their young. On larger carcasses, burying beetles will breed communally, forming multiple male-female associations. A significant and costly component of parental investment by burying beetles is the preservation of the carcass with secretions containing immune molecules. Because this immune investment is for the benefit of the offspring, the behavior is a form of social immunity. We test the hypothesis that communal breeding in burying beetles evolved as a mechanism to increase the social immune investment on larger carcasses, which are more difficult to preserve. We used N. defodiens, a communal breeding burying beetle species to test the hypothesis. There were two experimental treatments wherein, the females either bred communally or non-communally. Our results show that the combined immune activity in the secretions were higher in communally breeding pairs than in the immune contribution of single male-female pairs. However, subordinate females were rarely observed on the carcass, and the level of social immune activity of dominant females was lower than females breeding singly. These data suggest that communal breeding in N. defodiens decreases the level of investment in social immunity. Our results demonstrate that the presence of multiple females, which is common under natural conditions, can greatly complicate patterns of social immunity investment in burying beetles.
Complex Mutation Profiles in Mismatch Repair and Ribonucleotide Reductase Mutants Reveal Novel Repair Substrate Specificity of MutS Homolog (MSH) Complexes
Authors: Lamb Natalie A. , Bard Jonathan , Loll-Krippleber Raphael , Brown Grant W. , Surtees Jennifer A.
Published: July 2021

Determining mutation signatures is standard for understanding the etiology of human tumors and informing cancer treatment. Multiple determinants of DNA replication fidelity prevent mutagenesis that leads to carcinogenesis, including the regulation of free deoxyribonucleoside triphosphate (dNTP) pools by ribonucleotide reductase (RNR) and repair of replication errors by the mismatch repair (MMR) system. We identified genetic interactions between rnr1 alleles that elevate dNTP levels and MMR. We then utilized a targeted deep-sequencing approach to determine mutational signatures associated with MMR pathway defects. By combining rnr1 and msh mutations to increase dNTP levels and alter the mutational load, we uncovered previously unreported specificities of Msh2-Msh3 and Msh2-Msh6. Msh2-Msh3 is uniquely able to direct repair of G/C single base deletions in GC runs, while Msh2-Msh6 specifically directs repair of substitutions at G/C dinucleotides. We also identified broader sequence contexts that influence variant profiles in different genetic backgrounds. Finally, we observed that the mutation profiles in double mutants were not necessarily an additive relationship of mutation profiles in single mutants. Our results have implications for interpreting mutation signatures from human tumors, particularly when MMR is defective.
Malignancy and NF-kB signalling strengthen coordination between the expression of mitochondrial and nuclear-encoded oxidative phosphorylation genes
Authors: Perez Marcos Francisco , Sarkies Peter
Published: July 2021

Mitochondria are ancient endosymbiotic organelles crucial to eukaryotic growth and metabolism. Mammalian mitochondria carry a small genome containing thirteen protein-coding genes with the remaining mitochondrial proteins encoded by the nuclear genome. Little is known about how coordination between the two sets of genes is achieved. Correlation analysis of RNA-seq expression data from large publicly-available datasets is a common method to leverage genetic diversity to infer gene co-expression modules. Here we use this method to investigate nuclear-mitochondrial gene expression coordination. We identify a pitfall in correlation analysis that results from the large variation in the proportion of transcripts from the mitochondrial genome in RNA-seq data. Commonly used normalization techniques based on total read count (such as FPKM or TPM) produce artefactual negative correlations between mitochondrial- and nuclear-encoded transcripts. This also results in artefactual correlations between pairs of nuclear-encoded genes, thus having important consequences for inferring co-expression modules beyond mitochondria. We show that these effects can be overcome by normalizing using the median-ratio normalization (MRN) or trimmed mean of M values (TMM) methods. Using these normalizations, we find only weak and inconsistent correlations between mitochondrial and nuclear-encoded mitochondrial genes in the majority of healthy human tissues from the GTEx database. However, a subset of healthy tissues with high expression of NF-κB show significant coordination supporting a role for NF-κB in retrograde signalling. Contrastingly, most cancer types show robust coordination of nuclear and mitochondrial OXPHOS gene expression, identifying this as a feature of gene regulation in cancer.
Intrinsic Network Activity Reflects the Fluctuating Experience of Tonic Pain
Authors: Deak Bettina , Eggert Thomas , Mayr Astrid , Stankewitz Anne , Filippopulos Filipp , Jahn Pauline , Witkovsky Viktor , Straube Andreas , Schulz Enrico
Published: July 2021

Although we know sensation is continuous, research on long-lasting and continuously changing stimuli is scarce and the dynamic nature of ongoing cortical processing is largely neglected.

In a longitudinal study with 152 fMRI sessions, participants were asked to continuously rate the intensity of applied tonic heat pain for 20 minutes. Using group independent component analysis and dual-regression, we extracted the subjects’ time courses of intrinsic network activity. The relationship between the dynamic fluctuation of network activity with the varying time courses of three pain processing entities was computed: pain intensity, the direction of pain intensity changes and temperature.

We were able to dissociate the spatio-temporal patterns of objective (temperature) and subjective (pain intensity/changes of pain intensity) aspects of pain processing in the human brain. We found two somatosensory networks with distinct functions: one network which encodes the small fluctuations in temperature and consists mainly of bilateral SI. A second right-lateralised network that encodes the intensity of the subjective experience of pain consists of SI, SII, the PCC, and the thalamus.

We revealed the somatosensory dynamics that build up towards a current subjective percept of pain. The timing suggests a cascade of subsequent processing steps towards the current pain percept.
CRISPR/Cas9-mediated gene disruption of endogenous co-receptors confers broad resistance to HIV-1 in human primary cells and humanized mice
Authors: Li Shasha , Holguin Leo , Burnett John C.
Published: July 2021

In this project, we investigated the CRISPR/Cas9 system for creating HIV resistance by targeting the human CCR5 and CXCR4 genes, which encode cellular co-receptors required for HIV-1 infection. Using a clinically scalable system for transient ex vivo delivery of Cas9/gRNA ribonucleoprotein (RNP) complexes, we demonstrated that CRISPR-mediated disruption of CCR5 and CXCR4 in T-lymphocytes cells significantly reduced surface expression of the co-receptors, thereby establishing resistance to HIV-1 infection by CCR5 (R5)-tropic, CXCR4 (X4)-tropic, and dual (R5/X4)-tropic strains. CRISPR-mediated disruption of the CCR5 alleles in human CD34+ hematopoietic stem and progenitor cells (HSPCs) led to the differentiation of HIV-resistant macrophages. In human CD4+ T cells transplanted into a humanized mouse model, disruption of CXCR4 inhibited replication of X4-tropic HIV-1, thus leading to the virus-mediated enrichment CXCR4-disrupted cells in the peripheral blood and spleen. However, in human CD4+ T cells with both CCR5 and CXCR4 disruption, we observed poor engraftment in bone marrow, although significant changes were not observed in the lung, spleen, or peripheral blood. This study establishes a clinically scalable strategy for the dual knockout of HIV-1 co-receptors as a therapeutic strategy, while also raising caution of disrupting CXCR4, which may abate engraftment of CD4+ T cells in bone marrow.
Proximity-dependent biotinylation to elucidate the interactome of TNK2 non-receptor tyrosine kinase
Authors: Tahir Raiha , Madugundu Anil K. , Udainiya Savita , Cutler Jevon A. , Renuse Santosh , Wang Li , Pearson Nicole A. , Mitchell Chris , Mahajan Nupam , Pandey Akhilesh , Wu Xinyan
Published: July 2021

Non-receptor tyrosine kinases represent an important class of signaling molecules which are involved in driving diverse cellular pathways. Although, the large majority have been well-studied in terms of their protein-binding partners, the interactomes of some of the key non-receptor tyrosine kinases such as TNK2 (also known as activated Cdc42-associated kinase 1 or ACK1) have not been systematically investigated. Aberrant expression and hyperphosphorylation of TNK2 has been implicated in a number of cancers. However, the exact proteins and cellular events that mediate phenotypic changes downstream of TNK2 are unclear. Biological systems that employ proximity-dependent biotinylation methods, such as BioID, are being increasingly used to map protein-protein interactions as they provide increased sensitivity in discovering interaction partners. In this study, we employed BioID coupled to the biotinylation site identification technology (BioSITe) method that we recently developed to perform molecular mapping of intracellular proteins associated with TNK2. We also employed stable isotope labeling with amino acids in cell culture (SILAC) to quantitatively explore the interactome of TNK2. By performing a controlled comparative analysis between full-length TNK2 and its truncated counterpart, we were not only able to confidently identify site-level biotinylation of previously well-established TNK2 binders and substrates such as NCK1, NCK2, CTTN, STAT3, but also discover several novel TNK2 interacting partners. We validated TNK2 interaction with one of the novel TNK2 interacting protein, clathrin interactor 1 (CLINT1), using immunoblot analysis. Overall, this work reveals the power of the BioSITe method coupled to BioID and highlights several molecules that warrant further exploration to assess their functional significance in TNK2-mediated signaling.
High-throughput variant detection using a color-mixing strategy
Authors: Xie Nina Guanyi , Zhang Kerou , Song Ping , Li Renqiang , Luo Junfeng , Zhang David Yu
Published: July 2021

Many diseases are related to multiple genetic alterations along a single gene. Probing for highly multiple (>10) variants in a single qPCR tube is not possible due to a limited number of fluorescence channels and one variant per channel, so many more tubes are needed. Here, we experimentally validate our novel color-mixing strategy that uses fluorescence combinations as digital color codes to probe multiple variants simultaneously. The color-mixing strategy relies on a simple intra-tube assay that can probe for 15 variants as part of an inter-tube assay that can probe for an exponentially increased number of variants. The color-mixing strategy is achieved using multiplex double-stranded toehold probes modified with fluorophores and quenchers; the probes are designed to be quenched or luminous after binding to wildtype or variant templates. We used the color-mixing strategy to probe for 21 pathogenic mutations in thalassemia and to distinguish between heterozygous and homozygous variants in 6 tubes, with a specificity of 99% and a sensitivity of 94%. To support tuberculosis diagnosis, we used the same strategy to simultaneously probe in Mycobacterium tuberculosis for rifampicin-resistance mutations occurring within one 81-bp region and one 48-bp region in rpoB gene, plus five isoniazid-resistance mutations in inhA and katG genes.
A model of rapid homeostatic plasticity accounts for hidden, long-lasting changes in a neuronal circuit after exposure to high potassium
Authors: Rue Mara C.P. , Alonso Leandro , Marder Eve
Published: July 2021

Neural circuits must both function reliably and flexibly adapt to changes in their environment. We studied how both biological neurons and computational models respond to high potassium concentrations. Pyloric neurons of the crab stomatogastric ganglion (STG) initially become quiescent, then recover spiking activity in high potassium saline. The neurons retain this adaptation and recover more rapidly in subsequent high potassium applications, even after hours in control saline. We constructed a novel activity-dependent computational model that qualitatively captures these results. In this model, regulation of conductances is gated on and off depending on how far the neuron is from its target activity. This allows the model neuron to retain a trace of past perturbations even after it returns to its target activity in control conditions. Thus, perturbation, followed by recovery of normal activity, can hide cryptic changes in neuronal properties that are only revealed by subsequent perturbations.
Testing Hybridoma Cells for Mycoplasma Contamination
Authors: Greenfield Edward A.
Published: July 2021

Because mycoplasmas are a diverse group of organisms and are difficult to culture, several different strategies for detecting mycoplasma contamination have been developed. To date, no one test is suitable for detecting all of the possible mycoplasmas that may contaminate hybridoma or myeloma cultures. Therefore, it is sensible to consider using several methods. The most commonly used techniques are described here. Mycoplasma screening by growth on microbial medium can be performed on agar plates or in broth culture. Cultures are grown under both aerobic and anaerobic conditions because some common strains of Mycoplasma prefer the lack of oxygen (Mycoplasma buccale, Mycoplasma faucium, M. orale, and M. salivarium). Mycoplasma colonies form a characteristic “fried egg” appearance on agar plates, and this is the diagnostic feature used to confirm mycoplasma contamination. The colonies are small and are most easily seen with an inverted microscope. A quicker method for testing for mycoplasma takes advantage of the DNA-intercalating dye Hoechst 33258. Fixed cells are stained with the dye, and contaminated cultures are detected by the bright, punctate cytoplasmic staining of the Mycoplasma DNA. Finally, commercial kits for the detection of mycoplasmas using colorimetric assays or reporter cells are also described.
Ridding Hybridoma Cells of Mycoplasma Contamination
Authors: Greenfield Edward A.
Published: July 2021

Resistance to the most common antibiotics is what makes elimination of mycoplasma contamination so difficult, but not impossible. Different species of Mycoplasma have varied sensitivities to each of the major classes of antibiotics. One method presented here entails selection in antibiotic-containing medium combined with single-cell cloning over activated macrophage feeders. Its success rate is as high as 70%. As a method of last resort, growing hybridoma cells as ascites tumors is one of the most effective methods of removing mycoplasma contamination. The mycoplasmas are removed from the hybridoma cell surface by the immune system of the mouse. Mice must be of the same genetic background as the hybridomas. This technique is the same as the method for ridding cells of bacterial or fungal infection, and the success rate is perhaps 80%.
What older adults do in online communities when they co-create and co-destruct value
Authors: Kamalpour Mostafa , Eden Rebekah , Syed Rehan A. , Buys Laurie , Tariq Amina , Watson Jason
Published: July 2021

Purpose

This study aims to explain the value co-creation and co-destruction practices of older adults in an online community (OC).

Design/methodology/approach

Adopting practice theory and service-dominant logic as a theoretical perspective, this paper examined an OC of older adults by conducting an inductive thematic analysis of the interactions of the participants in the community.

Findings

The analysis revealed older adults engage with three value co-creation plus one value co-destruction practices in the OC including, communal coping practices, happiness creation practices, social capital generation practices and disparaging practices for older adults.

Research limitations/implications

Illustrated in a conceptual model, this study extends previous work evidencing OCs serve as a platform for value co-creation and value co-destruction activities in the context of older adults. Further, it suggests OCs facilitate resilience of older adults through value co-creation practices. Recognition of value co-destruction in OCs is critical as it is detrimental to the resilience of older adults. This study provides the needed foundation to advance knowledge on the use of OCs by older adults and suggests future research directions.

Practical implications

Identifying co-creation and co-destruction practices of older adults in OCs enables service providers (e.g. caregivers) to engage better in online value co-creation practices. Further, the findings of this study address one of the main priorities of service science to investigate the impact of value co-creation on well-being.

Originality/value

Despite the significant engagement of older adults in OCs, there is a lack of enough knowledge in the literature regarding value co-creation and co-destruction practices of older adults in OCs. This study addressed this gap by explaining how older adults co-create and co-destruct value in online spaces.