bluuebunny/crossref_metadata_2025_split huggingface dataset:
CODEC enables ‘single duplex’ sequencing
Authors: Bae Jin H. , Liu Ruolin , Nguyen Erica , Rhoades Justin , Blewett Timothy , Xiong Kan , Shea Douglas , Gydush Gregory , Tabrizi Shervin , An Zhenyi , Patel Sahil , Makrigiorgos G. Mike , Golub Todd R. , Adalsteinsson Viktor A. 
Published: June 2021

AbstractDetecting mutations as rare as a single molecule is crucial in many fields such as cancer diagnostics and aging research but remains challenging. Third generation sequencers can read a double-stranded DNA molecule (a ‘single duplex’) in whole to identify true mutations on both strands apart from false mutations on either strand but with limited accuracy and throughput. Although next generation sequencing (NGS) can track dissociated strands with Duplex Sequencing, the need to sequence each strand independently severely diminishes its throughput. Here, we developed a hybrid method called Concatenating Original Duplex for Error Correction (CODEC) that combines the massively parallel nature of NGS with the single-molecule capability of third generation sequencing. CODEC physically links both strands to enable NGS to sequence a single duplex with a single read pair. By comparing CODEC and Duplex Sequencing, we showed that CODEC achieved a similar error rate (10−6) with 100 times fewer reads and conferred ‘single duplex’ resolution to most major NGS workflows.
A secondary metabolite drives intraspecies antagonism in a gut symbiont that is inhibited by cell wall acetylation
Authors: Özçam Mustafa , Oh Jee-Hwan , Tocmo Restituto , Acharya Deepa , Zhang Shenwei , Astmann Theresa , Heggen Mark , Ruiz-Ramírez Silvette , Li Fuyong , Cheng Christopher C. , Vivas Eugenio , Rey Federico E. , Claesen Jan , Bugni Tim , Walter Jens , van Pijkeren Jan-Peter 
Published: June 2021

SUMMARYThe mammalian microbiome encodes numerous secondary metabolite biosynthetic gene clusters, yet their role in microbe-microbe interactions is unclear. Here, we characterized two polyketide synthase gene clusters (funandpks) in the gut symbiontLimosilactobacillus reuteri.Thepks, but not thefuncluster, encodes antimicrobial activity. Forty-one out of 51L. reuteristrains tested are sensitive to Pks products, which was independent of strains’ host origin. The sensitivity to Pks was also established in intraspecies competition experiments in gnotobiotic mice. Comparative genome analyses between Pks-resistant and sensitive strains identified an acyltransferase gene (act) that is unique to Pks-resistant strains. Subsequent cell wall analysis of the wild-type and theactmutant strains showed that Act acetylates cell wall components. Thepksmutants lost their competitive advantage andactmutants lost their Pks resistancein vivo. Thus, our findings provide insight into how closely related gut symbionts can compete and co- exist in the gastrointestinal tract.
CRISPRa screen on a genetic risk locus shared by multiple autoimmune diseases identifies a dysfunctional enhancer that affects IRF8 expression through cooperative lncRNA and DNA methylation machinery
Authors: Zhou Tian , Zhu Xinyi , Ye Zhizhong , Wang Yongfei , Yao Chao , Xu Ning , Zhou Mi , Ma Jianyang , Qin Yuting , Shen Yiwei , Tang Yuanjia , Yin Zhihua , Xu Hong , Zhang Yutong , Zang Xiaoli , Ding Huihua , Yang Wanling , Guo Ya , Harley John B. , Namjou Bahram , Kaufman Kenneth M. , Kottyan Leah C. , Weirauch Matthew T. , Hou Guojun , Shen Nan 
Published: June 2021

AbstractDysregulated transcription factors represent a major class of drug targets that mediate the abnormal expression of many critical genes involved in SLE and other autoimmune diseases. Although strong evidence suggests that natural human genetic variation affects basal and inducible gene expression, it is still a considerable challenge to establish a biological link between GWAS-identified non-coding genetic risk variants and their regulated gene targets. Here, we combine genetic data, epigenomic data, and CRISPR activation (CRISPRa) assays to screen for functional variants regulating IRF8 expression. Using CRISPR-mediated deletion and 3D chromatin structure analysis, we demonstrate that the locus containing rs2280381 is a cell-type-specific distal enhancer for IRF8 that spatially interacts with the IRF8 promoter. Further, rs2280381 mediates IRF8 expression through enhancer RNA AC092723.1, which recruits TET1 to the IRF8 promoter to modulate IRF8 expression by affecting methylation levels. The alleles of rs2280381 modulate PU.1 binding and chromatin state to differentially regulate AC092723.1 and IRF8 expression. Our work illustrates a strategy to define the functional genetic variants modulating transcription factor gene expression levels and identifies the biologic mechanism by which autoimmune diseases risk genetic variants contribute to the pathogenesis of disease.
Sustained replication of synthetic canine distemper virus defective genomesin vitroandin vivo
Authors: Tilston-Lunel Natasha L. , Welch Stephen R. , Nambulli Sham , de Vries Rory D. , Ho Gregory W , Wentworth David , Shabman Reed , Nichol Stuart T. , Spiropoulou Christina F. , de Swart Rik L. , Rennick Linda J. , Duprex W. Paul 
Published: June 2021

AbstractDefective interfering (DI) genomes restrict viral replication and induce type-I interferon. Since DI genomes have been proposed as vaccine adjuvants or therapeutic antiviral agents, it is important to understand their generation, delineate their mechanism of action, develop robust production capacities, assess their safety andin vivolongevity and determine their long-term effects. To address this, we generated a recombinant (r) canine distemper virus (CDV) from an entirely synthetic molecular clone designed using the genomic sequence from a clinical isolate obtained from a free-ranging raccoon with distemper. rCDV was serially passagedin vitroto identify DI genomes that naturally arise during rCDV replication. Defective genomes were identified by Sanger and next-generation sequencing techniques and predominant genomes were synthetically generated and cloned into T7-driven plasmids. Fully encapsidated DI particles (DIPs) were then generated using a rationally attenuated rCDV as a producer virus to drive DI genome replication. We demonstrate these DIPs interfere with rCDV replication in a dose-dependent mannerin vitro. Finally, we show sustained replication of a fluorescent DIP in experimentally infected ferrets over a period of 14 days. Most importantly, DIPs were isolated from the lymphoid tissues which are a major site of CDV replication. Our established pipeline for detection, generation and assaying DIPs is transferable to highly pathogenic paramyxoviruses and will allow qualitative and quantitative assessment of the therapeutic effects of DIP administration on disease outcome.ImportanceDefective interfering (DI) genomes have long been considered inconvenient artifacts that suppressed viral replicationin vitro. However, advances in sequencing technologies have led to DI genomes being identified in clinical samples, implicating them in disease progression and outcome. It has been suggested that DI genomes could be harnessed therapeutically. Negative strand RNA virus research has provided a rich pool of natural DI genomes over many years and they are probably the best understoodin vitro. Here, we demonstrate identification, synthesis, production and experimental inoculation of novel CDV DI genomes in highly susceptible ferrets. These results provide important evidence that rationally designed and packaged DI genomes can survive the course of a wild-type virus infection.
TRAF2, an innate immune sensor, reciprocally regulates mitophagy and inflammation to maintain cardiac myocyte homeostasis
Authors: Ma Xiucui , Rawnsley David R. , Kovacs Attila , Islam Moydul , Murphy John T. , Zhao Chen , Kumari Minu , Foroughi Layla , Liu Haiyan , Hyrc Krzysztof , Evans Sarah , French Brent A. , Margulies Kenneth B. , Javaheri Ali , Razani Babak , Mann Douglas L. , Mani Kartik , Diwan Abhinav 
Published: June 2021

ABSTRACTMitochondrial damage triggers cell death signaling with catastrophic consequences in long-lived and irreplaceable cells, such as cardiac myocytes. Sensing of leaked mitochondrial DNA upon mitochondrial damage is also a potent trigger of inflammation. Whether the innate immune response pathways monitor mitochondrial damage in mitochondria-rich cardiac myocytes to prevent inflammation and cell death, remains unknown. TRAF2, an adaptor protein downstream of innate immune receptors, localizes to the mitochondria in the unstressed heart, with increased mitochondrial targeting in cardiomyopathic human hearts and after cardiac ischemia-reperfusion injury in mice. Inducible cardiomyocyte-specific deletion of TRAF2 in young adult mice impairs mitophagy with rapid decline in mitochondrial quality, upregulates TLR9 expression in cardiac myocytes, and results in inflammation and cell death manifesting as a fulminant cardiomyopathy. Preventing TLR9-mediated mitochondrial DNA sensing and resultant inflammation provides a short-term reprieve from cardiomyopathy, but persistence of damaged mitochondria results in long-term recrudescence. Restoration of wild-type TRAF2, but not the E3 ubiquitin ligase deficient mutant, improves mitochondrial quality and rescues cardiomyopathy to restore homeostasis. Thus, the innate immune response acts via TRAF2 as the first line of defense against mitochondrial damage by orchestrating homeostatic mitophagy to dampen myocardial inflammation and prevent cell death.
A comprehensive temporal patterning gene network in Drosophila medulla neuroblasts revealed by single-cell RNA sequencing
Authors: Zhu Hailun , Zhao Sihai Dave , Ray Alokananda , Zhang Yu , Li Xin 
Published: June 2021

AbstractDuring development, neural stem cells are temporally patterned to sequentially generate a variety of neural types before exiting the cell cycle. Temporal patterning is well-studied in Drosophila, where neural stem cells called neuroblasts sequentially express cascades of Temporal Transcription Factors (TTFs) to control the birth-order dependent neural specification. However, currently known TTFs were mostly identified through candidate approaches and may not be complete. In addition, many fundamental questions remain concerning the TTF cascade initiation, progression, and termination. It is also not known why temporal progression only happens in neuroblasts but not in their differentiated progeny. In this work, we performed single-cell RNA sequencing of Drosophila medulla neuroblasts of all ages to study the temporal patterning process with single-cell resolution. Our scRNA-seq data revealed that sets of genes involved in different biological processes show high to low or low to high gradients as neuroblasts age. We also identified a list of novel TTFs, and experimentally characterized their roles in the temporal progression and neural fate specification. Our study revealed a comprehensive temporal gene network that patterns medulla neuroblasts from start to end. Furthermore, we found that the progression and termination of this temporal cascade also require transcription factors differentially expressed along the differentiation axis (neuroblasts -> -> neurons). Lola proteins function as a speed modulator of temporal progression in neuroblasts; while Nerfin-1, a factor required to suppress de-differentiation in post-mitotic neurons, acts at the final temporal stage together with the last TTF of the cascade, to promote the switch to gliogenesis and the cell cycle exit. Our comprehensive study of the medulla neuroblast temporal cascade illustrated mechanisms that might be conserved in the temporal patterning of neural stem cells.
Silkworm model forBacillus anthracisinfection and virulence determination
Authors: Paudel Atmika , Furuta Yoshikazu , Higashi Hideaki 
Published: June 2021

AbstractBacillus anthracisis an obligate pathogen and a causative agent of anthrax. Its major virulence factors are plasmid-coded; however, recent studies have revealed chromosome-encoded virulence factors, indicating that the current understanding of its virulence mechanism is elusive and needs further investigation. In this study, we established a silkworm (Bombyx mori) infection model ofB. anthracisSterne. We showed that silkworms were killed byB. anthracisand cured of the infection when administered with antibiotics. We quantitatively determined the lethal dose of the bacteria that kills 50% larvae and effective doses of antibiotics that cure 50% infected larvae. Furthermore, we demonstrated thatB. anthracismutants with disruption in virulence genes such aspagA, lef, andatxAhad attenuated silkworm-killing ability and reduced colonization in silkworm hemolymph. The silkworm infection model established in this study can be utilized in large-scale infection experiments to identify novel virulence determinants and develop novel therapeutic options againstB. anthracisinfections.
Hyphal compartmentalization and sporulation in Streptomyces require the conserved cell division protein SepX
Authors: Bush Matthew J. , Gallagher Kelley A. , Chandra Govind , Findlay Kim C. , Schlimpert Susan 
Published: June 2021

AbstractFilamentous actinobacteria such as Streptomyces undergo two distinct modes of cell division, leading to partitioning of growing hyphae into multicellular compartments via cross-walls, and to septation and release of unicellular spores. Specific determinants for cross-wall formation and the importance of hyphal compartmentalization for Streptomyces development are largely unknown. Here we show that SepX, an actinobacterial-specific protein, is crucial for both cell division modes in Streptomyces venezuelae. Importantly, we find that sepX-deficient mutants grow without cross-walls and that this substantially impairs the fitness of colonies and the coordinated progression through the developmental life cycle. Protein interaction studies and live-cell imaging suggest that SepX contributes to the stabilization of the divisome, a mechanism that also requires the dynamin-like protein DynB. Thus, our work identifies an important determinant for cell division in Streptomyces that is required for cellular development and sporulation.
Quantifying live bacterial densities using non-invasive optical measurements of E. coli
Authors: van der Helm Eric , Redl Stephanie M. A. 
Published: June 2021

AbstractProfiling the growth of bacterial cultures over time can be a tedious and error-prone process. Here, we present the development and evaluation of the use of the ODity platform to optically measure bacterial cell densities non-invasively. The digital growth data for E. coli MG1655 was calibrated against colony forming units (CFU/mL) obtained by plating on solid media. Diauxic-like shifts of liquid E. coli MG1655 cultures grown at 37°C in LB media were observed at densities as low as 2.9 × 107 ± 1.2 CFU/mL. The shift occurred at a significantly higher cell density (6.0 × 107 ± 1.2 CFU/mL) when the bacteria were cultured at 31°C. These shifts were only short lived, 15.2 ± 1.5 and 20.8 ± 1.8 min at 37°C and 31°C, respectively, with the previous growth rate restored thereafter. We measured minimum doubling times of 17.0 ± 1.1 and 24.8 ± 0.9 min at 37°C and 31°C, respectively. These results demonstrate that the growth and growth rate of bacterial cultures can be accurately determined non-invasively using the ODity device.
Harnessing methylotrophs as a bacterial platform to reduce adverse effects of the use of the heavy lanthanide gadolinium in magnetic resonance imaging
Authors: Good Nathan M. , Lee Harvey , Hawker Emily R. , Gilad Assaf A. , Martinez-Gomez N. Cecilia 
Published: June 2021

ABSTRACTGadolinium is a key component of magnetic resonance imaging contrast agents that are critical tools for enhanced detection and diagnosis of tissue and vascular abnormalities. Untargeted post-injection deposition of gadolinium in vivo, and association with diseases like nephrogenic systemic fibrosis, has alerted regulatory agencies to re-evaluate their widespread use and generated calls for safer gadolinium-based contrast agents (GBCAs). Increasing anthropogenic gadolinium in surface water has also raised concerns of potential bioaccumulation in plants and animals. Methylotrophic bacteria can acquire, transport, store and use light lanthanides as part of a cofactor complex with pyrroloquinoline quinone (PQQ), an essential component of XoxF-type methanol dehydrogenases (MDHs), a critical enzyme for methylotrophic growth with methanol. We report robust gadolinium-dependent methanol growth of a genetic variant of Methylorubrum extorquens AM1, named evo-HLn, for “evolved for heavy lanthanides”. Genetic adaptation of evo-HLn resulted in increased xox1 promoter and XoxF MDH activities, transport and storage of Gd3+, and augmented biosynthesis of PQQ. Gadolinium-grown cells exhibited a shorter T1 relaxation time compared to cells with lanthanum or no lanthanide when analyzed by MRI. In addition, evo-HLn was able to grow on methanol using the GBCA Gd-DTPA as the sole gadolinium source, showing the potential of this strain for the development of novel GBCAs and gadolinium recovery from medical waste and/or wastewater.
Estimation of cancer cell fractions and clone trees from multi-region sequencing of tumors
Authors: Zheng Lily , Wood Laura , Karchin Rachel , Scharpf Robert 
Published: June 2021

AbstractMulti-region sequencing of one or multiple biopsies of solid tumors from a patient can be used to improve our understanding of the diversity of subclones in the patient’s tumor and shed light on the evolutionary history of the disease. Due to the large number of possible evolutionary relationships between clones and the fundamental uncertainty of the mutational composition of subclones, elucidating the most probable evolutionary relationships poses statistical and computational challenges. We developed a Bayesian hierarchical model called PICTograph to model uncertainty in the assignment of mutations to subclones and an approach to reduce the space of possible graphical models that postulate their evolutionary origin. Compared to available methods, our approach provided more consistent and accurate estimates of cancer cell fractions and better tree topology reconstruction over a range of simulated clonal diversity. Application of PICTograph to whole exome sequencing data of individuals with pancreatic cancer precursor lesions confirmed known early occurring mutations and indicated substantial molecular diversity, including multiple distinct subclones (range 6 - 12) and intra-sample mixing of subclones. As the complete evolutionary history for some patients was not identifiable, we used ensemble-based visualizations to distinguish between highly probable evolutionary relationships recovered in multiple models from uncertain relationships occurring in a small subset of models. These analyses indicate that PICTograph provides a useful approximation to evolutionary inference, particularly when the evolutionary course of a patient’s cancer is complex.
Comparative therapeutic strategies for preventing aortic rupture in a mouse model of vascular Ehlers Danlos syndrome
Authors: Legrand Anne , Guery Charline , Faugeroux Julie , Fontaine Erika , Beugnon Carole , Gianfermi Amélie , Loisel-Ferreira Irmine , Verpont Marie-Christine , Adham Salma , Mirault Tristan , Hadchouel Juliette , Jeunemaitre Xavier 
Published: June 2021

AbstractWe created a knock-in Col3a1+/G182R mouse model with spontaneous mortality caused by thoracic aortic rupture that recapitulates a rare vascular genetic disease of type III collagen, the vascular Ehlers-Danlos syndrome (vEDS). Investigation of this model showed lower survival rate in males caused by aortic rupture, thin non-inflammatory arteries and altered arterial collagen. Transcriptomic analysis of aortas showed upregulation of genes related to inflammation and cell stress response. Compared to water, survival rate of Col3a1+/G182R mice was not affected by beta-blockers (propranolol or celiprolol). Two other vasodilating anti-hypertensive agents (hydralazine, amlodipine) gave opposite results on aortic rupture and mortality rate. There was a spectacular beneficial effect of losartan, reversed by the cessation of its administration, and a marked deleterious effect of exogenous angiotensin II. These results suggest that blockade of the renin angiotensin system should be tested as a first-line medical therapy in patients with vEDS.
Contributions of distinct auditory cortical inhibitory neuron types to the detection of sounds in background noise
Authors: Lakunina Anna A. , Menashe Nadav , Jaramillo Santiago 
Published: June 2021

AbstractThe ability to separate background noise from relevant acoustic signals is essential for appropriate sound-driven behavior in natural environments. Examples of this separation are apparent in the auditory system, where neural responses to behaviorally relevant stimuli become increasingly noise-invariant along the ascending auditory pathway. However, the mechanisms that underlie this reduction in responses to background noise are not well understood. To address this gap in knowledge, we first evaluated the effects of auditory cortical inactivation on mice of both sexes trained to perform a simple auditory signal-in-noise detection task, and found that outputs from the auditory cortex are important for the detection of auditory stimuli in noisy environments. Next, we evaluated the contributions of the two most common cortical inhibitory cell types, parvalbumin-expressing (PV+) and somatostatin-expressing (SOM+) interneurons, to the perception of masked auditory stimuli. We found that inactivation of either PV+ or SOM+ cells resulted in a reduction in the ability of mice to determine the presence of auditory stimuli masked by noise. These results indicate that a disruption of auditory cortical network dynamics by either of these two types of inhibitory cells is sufficient to impair the ability to separate acoustic signals from noise.Significance StatementAppropriate behavior in a natural environment relies on the ability to separate background noise from relevant signals. We found that auditory cortical inhibitory neurons play a causal role in separating environmental noise from behaviorally relevant auditory signals. These results advance our understanding of the computations performed by the auditory system to decompose and analyze acoustic stimuli in the presence of noise.
Segmenting German housing markets using principal component and cluster analyses
Authors: Wiersma Simon , Just Tobias , Heinrich Michael 
Published: June 2021

Purpose
Germany has a polycentric city structure. This paper aims to reduce complexity of this structure and to find a reliable classification scheme of German housing markets at city level based on 17 relevant market parameters.

Design/methodology/approach
This paper uses a two-step clustering algorithm combining k-means with Ward’s method to develop the classification scheme. The clustering process is preceded by a principal component analysis to merely retain the most important dimensions of the market parameters. The robustness of the results is investigated with a bootstrapping method.

Findings
It is found that German residential markets can best be segmented into four groups. Geographic contiguity plays a specific role, but is not a main factor. Our bootstrapping analysis identifies the majority of pairwise city relations (88.5%) to be non-random.

Research limitations/implications
A deeper discussion concerning the most relevant market parameters is required. The stability of the clusters is to be re-investigated in the future, as the bootstrapping analysis indicates that some clusters are more homogeneous than others.

Practical implications
The developed classification scheme provides insights into opportunities and risks associated with specific city groups. The findings of this study can be used in portfolio management to reduce unsystematic investment risks and to formulate investment strategies.

Originality/value
To the best of the authors’ knowledge, this is the first paper to offer insights into the German housing markets which applies principal component, cluster and bootstrapping analyses in a sole integrated approach.