bluuebunny/crossref_metadata_2025_split huggingface dataset:
Multiplex knockout of trichome-regulating MYB duplicates in hybrid poplar using a single gRNA
Authors: Bewg W. Patrick , Harding Scott A. , Engle Nancy L. , Vaidya Brajesh N. , Zhou Ran , Reeves Jacob , Horn Thomas W. , Joshee Nirmal , Jenkins Jerry W. , Shu Shengqiang , Barry Kerrie W. , Yoshinaga Yuko , Grimwood Jane , Schmitz Robert J. , Schmutz Jeremy , Tschaplinski Timothy J. , Tsai Chung-Jui 
Published: September 2021

ABSTRACTAs the focus for CRISPR edited plants moves from proof-of-concept to real world applications, precise gene manipulation will increasingly require concurrent multiplex editing for polygenic traits. A common approach for editing across multiple sites is to design one gRNA per target; however, this complicates construct assembly and increases the possibility of off-target mutations. In this study, we utilized one gRNA to targetMYB186, a known positive trichome regulator, as well as its paralogsMYB138andMYB38at a consensus site for mutagenesis inPopulus tremula×P. albaINRA 717-1B4. Unexpected duplications ofMYB186andMYB138resulted in a total of eight alleles for the three targeted genes in the hybrid poplar. Deep sequencing and PCR analyses confirmed editing across all eight targets in nearly all of the resultant glabrous mutants, ranging from small indels to large genomic dropouts, with no off-target activity detected at four potential sites. This highlights the effectiveness of a single gRNA targeting conserved exonic regions for multiplex editing. Additionally, cuticular wax and whole leaf analyses showed a complete absence of triterpenes in the trichomeless mutants, hinting at a previously undescribed role for the non-glandular trichomes of poplar.ONE SENTENCE SUMMARYTargeting conserved sequences with a single gRNA allowed efficient mutagenesis of a multigene family and the recovery of trichomeless and triterpene-free poplar mutants.
Pro-apoptotic caspase deficiency reveals a cell-extrinsic mechanism of NK cell regulation
Authors: Olsen Tayla M. , Tan Wei Hong , Knudsen Arne C. , Rongvaux Anthony 
Published: September 2021

AbstractRegulated cell death is essential for the maintenance of cellular and tissue homeostasis. In the hematopoietic system, genetic defects in apoptotic cell death generally produce the accumulation of immune cells, inflammation and autoimmunity. In contrast, we found that genetic deletion of caspases of the mitochondrial apoptosis pathway reduces natural killer (NK) cell numbers and makes NK cells functionally defectivein vivoandin vitro. Caspase deficiency results in constitutive activation of a type I interferon (IFN) response, due to leakage of mitochondrial DNA and activation of the cGAS/STING pathway. The NK cell defect in caspase-deficient mice is independent of the type I IFN response, but the phenotype is partially rescued by cGAS or STING deficiency. Finally, caspase deficiency alters NK cells in a cell-extrinsic manner. Type I IFNs and NK cells are two essential effectors of antiviral immunity, and our results demonstrate that they are both regulated in a caspase-dependent manner. Beyond caspase-deficient animals, our observations may have implications in infections that trigger mitochondrial stress and caspase-dependent cell death.
Rapid, multianalyte detection of opioid metabolites in wastewater
Authors: Kumar Narendra , Rana Muhit , Geiwitz Michael , Khan Niazul Islam , Catalano Matthew , Ortiz-Marquez Juan C. , Kitadai Hikari , Weber Andrew , Dweik Badawi , Ling Xi , van Opijnen Tim , Argun Avni , Burch Kenneth S. 
Published: September 2021

AbstractBy monitoring opioid metabolites, wastewater-based epidemiology (WBE) could be an excellent tool for real-time information on consumption of illicit drugs. A key limitation of WBE is the reliance on costly laboratory-based techniques that require substantial infrastructure and trained personnel, resulting in long turnaround times. Here, we present an aptamer-based graphene field effect transistor (AptG-FET) platform for simultaneous detection of three different opioid metabolites. This platform provides a reliable, rapid, and inexpensive method for quantitative analysis of opioid metabolites in wastewater (WW). The platform delivers a limit of detection (LOD) 2-3 orders of magnitude lower than previous reports, but in line with the concentrations range (pg/ml to ng/ml) of these opioid metabolites present in real samples. To enable multianalyte detection we developed a facile, reproducible, and high yield fabrication process producing twenty G-FETs with integrated side gate platinum (Pt) electrodes on a single chip. Our devices achieved the simultaneous and selective multianalyte detection of three different metabolites: Noroxycodone (NX), 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), and Norfentanyl (NF) in wastewater.
Characterization of aquaporin1b (AQP1b) mRNA in mud loach (Misgurnus mizolepis) in response to heavy metal and immunostimulant stimuli
Authors: Lee Sang Yoon , Nam Yoon Kwon , Kim Yi Kyung 
Published: September 2021

AbstractAquaporins (AQPs) facilitate the transport of water or other small solutes into cells in the presence of osmotic gradients. However, the current understanding of piscine AQP gene with cellular stress responses has been still limitedly exemplified. In present study, we characterized the mud loach AQP1b gene at the nucleotide and amino acid levels. We identified three AQP 1b transcript variants (mmAQP1b_tv1, mmAQP1b_tv2, and mmAQP1b_tv3). Then, we examined the AQP1b promoter region and observed several transcription factor binding sites (TFBS) for nuclear factor of activated T-cells (NFAT), SRY-box, c-AMP responsive element binding protein (CREB), GATA binding factor, and hepatic nuclear factor-1. Interestingly, mmAQP1b transcription was differentially modulated by heavy metal or immunostimulant challenge. Further studies to deepen the knowledge of fish AQP-mediated adaptation response potentially relevant to molecular pathogenesis are warranted.Summary statementWe identified mud loach AQP1b transcript variants and consensus sequences involved in stress or innate immunity in promotor region. AQP1b transcription was differentially modulated by heavy metal or immunostimulant challenge.
Tumor Suppressor p53 Restrains Cancer Cell Dissemination by Modulating Mitochondrial Dynamics
Authors: Phan Thi Tuyet Trinh , Lin Yu-Chun , Chou Yu-Ting , Wu Chien-Wei , Lin Lih-Yuan 
Published: September 2021

AbstractTumor suppressor p53 plays a central role in preventing tumorigenesis. Here, we unravel how p53 modulates mitochondrial dynamics to restrain the metastatic properties of cancer cells. p53 inhibits the mammalian target of rapamycin complex 1 (mTORC1) signaling to attenuate the protein level of mitochondrial fission process 1 (MTFP1), which fosters the pro-fission dynamin-related protein 1 (Drp1) phosphorylation. This regulatory mechanism allows p53 to restrict cell migration and invasion governed by Drp1-mediated mitochondrial fission. Downregulating p53 expression or elevating the molecular signature of mitochondrial fission correlates with aggressive tumor phenotypes and poor prognosis in cancer patients. Upon p53 loss, exaggerated mitochondrial fragmentation stimulates the activation of the extracellular signal-regulated kinase 1/2 (ERK1/2) signaling resulting in epithelial-to-mesenchymal transition (EMT)-like changes in cell morphology, accompanied by accelerated matrix metalloproteinase-9 (MMP9) expression and invasive cell migration. Notably, blocking the activation of mTORC1/MTFP1/Drp1/ERK1/2 axis completely abolishes the p53 deficiency-driven cellular morphological switch, MMP9 expression, and cancer cell dissemination. Our findings unveil a hitherto unrecognized mitochondria-dependent molecular mechanism underlying the metastatic phenotypes of p53-compromised cancers.
Effect of dietary zinc supplementation on the gastrointestinal microbiota and host gene expression in theShank3B−/−mouse model of autism spectrum disorder
Authors: Wong Giselle C. , Jung Yewon , Lee Kevin , Fourie Chantelle , Handley Kim M. , Montgomery Johanna M. , Taylor Michael W. 
Published: September 2021

AbstractShank genes are implicated in ~1% of people with autism and mice with Shank3 knock out mutations exhibit autism-like behaviours. Zinc deficiency and gastrointestinal problems can be common among people with autism, and zinc is a key element required for SHANK protein function and gut development. InShank3B−/−mice, a supplementary zinc diet reverses autism behaviours. We hypothesise that dietary zinc may alter the gut microbiome, potentially affecting the gut-microbiome-brain axis, which may contribute to changes in autism-like behaviours. To test this, four types of gastrointestinal samples (ileum, caecum, colon, faecal) were collected from wild-type and knock-outShank3B−/−mice on either control or supplemented-zinc diets. Cage, genotype and zinc diet each contributed significantly to bacterial community variation (accounting for 12.8%, 3.9% and 2.3% of the variation, respectively). Fungal diversity differed significantly between wild-type and knock-outShank3B−/−mice on the control zinc diet, and the fungal biota differed among gut locations. RNA-seq analysis of host (mouse) transcripts revealed differential expression of genes involved in host metabolism that may be regulated by the gut microbiota and genes involved in anti-microbial interactions. By utilising theShank3B−/−knock-out mouse model we were able to examine the influence of – and interactions between – dietary zinc and ASD-linked host genotype. These data broaden understanding of the gut microbiome in autism and pave the way towards potential microbial therapeutics for gastrointestinal problems in people with autism.ImportancePreviously, supplemental dietary zinc in the Shank3B−/−mouse model of autism spectrum disorder resulted in observations of ASD behaviours reversal; in this study we also used theShank3B−/−mouse model to examine the influence of – and interaction between – dietary zinc and ASD-linked host genotype. Sample location along the gastrointestinal tract, genotype and zinc diet explained some of the variation in the microbiota data, with notable bacterial differences between treatment groups. Differential expression of host genes between treatment groups, including antimicrobial interaction genes and gut microbiota-regulated host metabolism genes, suggests that the interplay between gut microbes, the gastrointestinal tract and the brain may play a major role towards the observed amelioration of ASD behaviours seen previously with supplemented dietary zinc. These results widen the scope towards manipulating both dietary zinc and the microbiota itself to ameliorate ASD-related behaviours and associated gastrointestinal issues.
Advanced understanding of prokaryotic biofilm formation using a cost-effective and versatile multi-panel adhesion (mPAD) mount
Authors: Schulze Stefan , Schiller Heather , Solomonic Jordan , Telhan Orkan , Costa Kyle , Pohlschroder Mechthild 
Published: September 2021

AbstractMost microorganisms exist in biofilms, which comprise aggregates of cells surrounded by an extracellular matrix that provides protection from external stresses. Based on the conditions under which they form, biofilm structures vary in significant ways. For instance, biofilms that develop when microbes are incubated under static conditions differ from those formed when microbes encounter the shear forces of a flowing liquid. Moreover, biofilms develop dynamically over time. Here, we describe a cost-effective, 3D-printed coverslip holder that facilitates surface adhesion assays under a broad range of standing and shaking culture conditions. This multi-panel adhesion (mPAD) mount further allows cultures to be sampled at multiple time points, ensuring consistency and comparability between samples and enabling analyses of the dynamics of biofilm formation. As a proof of principle, using the mPAD mount for shaking, oxic cultures, we confirm previous flow chamber experiments showing that Pseudomonas aeruginosa wild type and a phenazine deletion mutant (Δphz) form similar biofilms. Extending this analysis to anoxic conditions, we reveal that microcolony and biofilm formation can only be observed under shaking conditions and are decreased in the Δphz mutant compared to wild-type cultures, indicating that phenazines are crucial for the formation of biofilms if oxygen as an electron acceptor is not available. Furthermore, while the model archaeon Haloferax volcanii does not require archaella for attachment to surfaces under static conditions, we demonstrate that H. volcanii mutants that lack archaella are negatively affected in their early stages of biofilm formation under shaking conditions.ImportanceDue to the versatility of the mPAD mount, we anticipate that it will aid the analysis of biofilm formation in a broad range of bacteria and archaea. Thereby, it contributes to answering critical biological questions about the regulatory and structural components of biofilm formation and understanding this process in a wide array of environmental, biotechnological, and medical contexts.
An open label, adaptive, phase 1 trial of high-dose oral nitazoxanide in healthy volunteers: an antiviral candidate for SARS-CoV-2
Authors: Walker Lauren E , FitzGerald Richard , Saunders Geoffrey , Lyon Rebecca , Fisher Michael , Martin Karen , Eberhart Izabela , Woods Christie , Ewings Sean , Hale Colin , Rajoli Rajith KR , Else Laura , Dilly-Penchala Sujan , Amara Alieu , Lalloo David G , Jacobs Michael , Pertinez Henry , Hatchard Parys , Waugh Robert , Lawrence Megan , Johnson Lucy , Fines Keira , Reynolds Helen , Rowland Timothy , Crook Rebecca , Byrne Kelly , Mozgunov Pavel , Jaki Thomas , Khoo Saye , Owen Andrew , Griffiths Gareth , Fletcher Thomas E 
Published: September 2021

AbstractRepurposing approved drugs may rapidly establish effective interventions during a public health crisis. This has yielded immunomodulatory treatments for severe COVID-19, but repurposed antivirals have not been successful to date because of redundancy of the target in vivo or suboptimal exposures at studied doses. Nitazoxanide is an FDA approved antiparasitic medicine, that physiologically-based pharmacokinetic (PBPK) modelling has indicated may provide antiviral concentrations across the dosing interval, when repurposed at higher than approved doses. Within the AGILE trial platform (NCT04746183) an open label, adaptive, phase 1 trial in healthy adult participants was undertaken with high dose nitazoxanide. Participants received 1500mg nitazoxanide orally twice-daily with food for 7 days. Primary outcomes were safety, tolerability, optimum dose and schedule. Intensive pharmacokinetic sampling was undertaken day 1 and 5 with Cmin sampling on day 3 and 7. Fourteen healthy participants were enrolled between 18th February and 11th May 2021. All 14 doses were completed by 10/14 participants. Nitazoxanide was safe and well tolerated with no significant adverse events. Moderate gastrointestinal disturbance (loose stools) occurred in 8 participants (57.1%), with urine and sclera discolouration in 12 (85.7%) and 9 (64.3%) participants, respectively, without clinically significant bilirubin elevation. This was self-limiting and resolved upon drug discontinuation. PBPK predictions were confirmed on day 1 but with underprediction at day 5. Median Cmin was above the in vitro target concentration on first dose and maintained throughout. Nitazoxanide administered at 1500mg BID with food was safe and well tolerated and a phase 1b/2a study is now being initiated in COVID-19 patients.
Universal inverse square relationship between heart rate variability and heart rate
Authors: Maltsev Anna V. , Monfredi Oliver , Maltsev Victor A. 
Published: September 2021

AbstractIn our previous study, we analyzed heart rate variability and heart rate from a large variety of cardiac preparations (including humans, living animals, Langendorff-perfused isolated hearts, and single sinoatrial nodal cells) in diverse species, combining our data with those of previously published articles. The analysis revealed that regardless of conditions, heart rate variability (for the purposes of the study assessed as standard deviation of beat-to-beat intervals) vs. heart rate follows a universal exponential decay-like relationship. Numerical simulations of diastolic interval variability by adding a randomly fluctuating term (Iper) to net current revealed a similar relationship. In the present study, using a Taylor series, we found that this relationship is, in fact, inverse square, and we derive an explicit formula for the standard deviation (sd) of the cycle length (CL) as a function of heart rate (HR) with biophysically meaningful parameters: sd(CL)=sd(Iper)*(60,000/mean(HR) -APD)^2/(ΔV*C), where CL is in ms, HR in beats per minute, Iper in pA, APD in ms is an average AP duration of pacemaker cells, C in pF is cell membrane capacitance, and ΔV is the magnitude of diastolic depolarization in mV. This relationship gives direct insight into heart rate variability mechanisms at the basic level of individual pacemaker cells, i.e. their intrinsic CL variability linked to stochastic operation of ion channels (both Ca release and cell membrane channels) generating Iper. Our explicit formula may be also used for a more precise biomedical interpretation of heart rate variability after respective corrections for heart rate.
Stem cell-derived macrophages as a new platform for studying host-pathogen interactions in livestock
Authors: Meek Stephen , Watson Tom , Eory Lel , McFarlane Gus , Wynne Felicity J. , McCleary Stephen , Dunn Laura E.M. , Charlton Emily M. , Criag Chloe , Shih Barbara , Regan Tim , Taylor Ryan , Sutherland Linda , Gossner Anton , Chintoan-Uta Cosmin , Fletcher Sarah , Beard Philippa M. , Hassan Musa A. , Grey Finn , Hope Jayne C. , Stevens Mark P. , Nowak-Imialek Monika , Niemann Heiner , Ross Pablo J. , Tait-Burkard Christine , Brown Sarah M. , Lefevre Lucas , Thomson Gerard , McColl Barry W. , Lawrence Alistair B. , Archibald Alan L. , Steinbach Falko , Crooke Helen R. , Gao Xuefei , Liu Pentao , Burdon Tom 
Published: September 2021

AbstractInfectious diseases of farmed and wild animals pose a recurrent threat to food security and human health. The macrophage, a key component of the innate immune system, is the first line of defence against many infectious agents and plays a major role in shaping the adaptive immune response. However, this phagocyte is a target and host for many pathogens. Understanding the molecular basis of interactions between macrophages and pathogens is therefore crucial for the development of effective strategies to combat important infectious diseases. We explored how pluripotent stem cells (PSCs) can provide a limitlessin vitrosupply of genetically and experimentally tractable macrophages from livestock. Porcine and bovine PSC-derived macrophages (PSCdMs) exhibited molecular and functional characteristics ofex vivoprimary macrophages. Pig PSCdMs were productively infected by Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) and African Swine Fever Virus (ASFV), two of the most economically important and devastating viruses in pig farming. Moreover, Pig PSCdMs were readily amenable to genetic modification by CRISPR/Cas9 gene editing applied in parental stem cells, or directly by lentiviral vector transduction. PSCs and differentiated derivatives therefore provide a useful and ethical experimental platform to investigate the genetic and molecular basis of host-pathogen interactions in livestock.
Precision fMRI reveals that the language-selective network supports both phrase-structure building and lexical access during language production
Authors: Hu Jennifer , Small Hannah , Kean Hope , Takahashi Atsushi , Zekelman Leo , Kleinman Daniel , Ryan Elizabeth , Nieto-Castañón Alfonso , Ferreira Victor , Fedorenko Evelina 
Published: September 2021

AbstractA fronto-temporal brain network has long been implicated in language comprehension. However, this network’s role in language production remains debated. In particular, it remains unclear whether all or only some language regions contribute to production, and which aspects of production these regions support. Across three fMRI experiments that rely on robust individual-subject analyses, we characterize the language network’s response to high-level production demands. We report three novel results. First, sentence production, spoken or typed, elicits a strong response throughout the language network. Second, the language network responds to both phrase-structure building and lexical access demands, although the response to phrase-structure building is stronger and more spatially extensive, present in every language region. Finally, contra some proposals, we find no evidence of brain regions—within or outside the language network—that selectively support phrase-structure building in production relative to comprehension. Instead, all language regions respond more strongly during production than comprehension, suggesting that production incurs a greater cost for the language network. Together, these results align with the idea that language comprehension and production draw on the same knowledge representations, which are stored in a distributed manner within the language-selective network and are used to both interpret and generate linguistic utterances.
Genetic variations within human gained enhancer elements affect human brain sulcal morphology
Authors: Lemaitre Herve , Le Guen Yann , Tilot Amanda K. , Stein Jason L. , Philippe Cathy , Mangin Jean-François , Fisher Simon E. , Frouin Vincent 
Published: September 2021

AbstractThe expansion of the cerebral cortex is one of the most distinctive changes in the evolution of the human brain. Cortical expansion and related increases in cortical folding may have contributed to emergence of our capacities for high-order cognitive abilities. Molecular analysis of humans, archaic hominins, and non-human primates has allowed identification of chromosomal regions showing evolutionary changes at different points of our phylogenetic history. In this study, we assessed the contributions of genomic annotations spanning 30 million years to human sulcal morphology measured via MRI in more than 18,000 participants from the UK Biobank. We found that variation within brain-expressed human gained enhancers, regulatory genetic elements that emerged since our last common ancestor with Old World monkeys, explained more trait heritability than expected for the left and right calloso-marginal posterior fissures and the right central sulcus. Intriguingly, these are sulci that have been previously linked to the evolution of locomotion in primates and later on bipedalism in our hominin ancestors.
Mechanical Causation of Biological Structure: Productive Pulls Produce Persistent Filaments in a Human Fibroblast Model of Matrix Development
Authors: Silverman Alexandra A. , Olszewski Jason D. , Siadat Seyed Mohammad , Ruberti Jeffrey W. 
Published: September 2021

ABSTRACTThe principal mechanisms driving the synthesis and organization of durable animal structure have been the subject of intense investigation for decades. Here, we present evidence that mechanical strains can direct the formation of extracellular matrix (ECM) filaments via a mechanochemical cascade. This process is driven by cooperative cell contractions (“pulls”) that organize and precipitate structure via strain-induced polymer assembly. In anin vitromodel of ECM synthesis, we use high-resolution optical microscopy to observe the kinematics of cell motion during their growth to confluency and identified cell-to-cell pulls that result in the production of persistent ECM filaments. Using live-cell confocal imaging, we confirmed that these pulls can directly cause the formation of fibronectin filaments that then bind collagen, producing persistent structures aligned with the direction of the pull. The finding suggests a new model for initial durable structure formation in animals based on local cell contraction, extensional strain and polymer mechanochemistry. The results have important implications for ECM development, growth and life-threatening pathologies of the ECM such as fibrosis.