bluuebunny/crossref_metadata_2025_split huggingface dataset:
A high-throughput genome-integrated assay reveals spatial dependencies governing Tcf7l2 binding
Authors: Szczesnik Tomasz , Chu Lendy , Ho Joshua W. K. , Sherwood Richard 
Published: March 2020

2 SummaryPredicting where transcription factors bind in the genome from theirin-vitroDNA binding affinity is confounded by the large number of possible interactions with nearby transcription factors. To characterise the binding logic for the Wnt effector transcription factor Tcf7l2, we have developed a high-throughput screening platform in which thousands of 99-bp synthesised DNA sequences are inserted into a specific genomic locus through CRISPR/Cas9-based homology-directed repair, followed by measurement of Tcf7l2 binding by DamID. Using this platform at two genomic loci in mouse embryonic stem cells, we show that while the binding of Tcf7l2 closely follows thein-vitromotif binding strength and is influenced by local chromatin accessibility, it is also strongly affected by the surrounding 99-bp of sequence. The presence of nearby Oct4 and Klf4 motifs promote Tcf7l2 binding, particularly in the adjacent ~20 to 50-bp nearby and oscillating with a 10.8-bp phasing relative to these cofactor motifs, which matches the turn of a DNA helix. This novel high-throughput DamID assay provides a powerful platform to determine local DNA sequence grammars that causally influence transcription factor binding in controlled genomic contexts.
Prospects and limitations of expansion microscopy in chromatin ultrastructure determination
Authors: Kubalová Ivona , Černohorská Markéta Schmidt , Huranová Martina , Weisshart Klaus , Houben Andreas , Schubert Veit 
Published: March 2020

AbstractExpansion Microscopy (ExM) is a method to magnify physically a specimen with preserved ultrastructure. It has the potential to explore structural features beyond the diffraction limit of light. The procedure has been successfully used for different animal species, from isolated macromolecular complexes through cells to tissue slices. Expansion of plant-derived samples is still at the beginning, and little is known whether the chromatin ultrastructure becomes altered by physical expansion.In this study, we expanded isolated barley nuclei and compared whether ExM can provide a structural view of chromatin comparable with super-resolution microscopy. Different fixation and denaturation/digestion conditions were tested to maintain the chromatin ultrastructure. We achieved up to ∼4.2-times physically expanded nuclei corresponding to a maximal resolution of ∼50-60 nm when imaged by wild-field (WF) microscopy. By applying structured illumination microscopy (SIM, super-resolution) doubling the WF resolution the chromatin structures were observed at a resolution of ∼25-35 nm.WF microscopy showed a preserved nucleus shape and nucleoli. Moreover, we were able to detect chromatin domains, invisible in unexpanded nuclei. However, by applying SIM we observed that the preservation of the chromatin ultrastructure after expansion was not complete and that the majority of the tested conditions failed to keep the ultrastructure.Nevertheless, using expanded nuclei we detected successfully centromere repeats by fluorescencein situhybridization (FISH) and the centromere-specific histone H3 variant CENH3 by indirect immunostaining. However, although these repeats and proteins were localized at the correct position within the nuclei (indicating a Rabl orientation) their ultrastructural arrangement was impaired.
Minor spliceosome disruption causes limb growth defects without altering patterning
Authors: Drake Kyle D , Lemoine Christopher , Aquino Gabriela S , Vaeth Anna M , Kanadia Rahul N 
Published: March 2020

AbstractDisruption of the minor spliceosome causes primordial dwarfism in microcephalic osteodysplastic primordial dwarfism type 1. Similarly, primordial dwarfism in domesticated animals is linked to positive selection in minor spliceosome components. Despite the importance of minor intron splicing in limb size regulation, its role in limb development remains unexplored. Here we show that loss of U11 small nuclear RNA, an essential minor spliceosome component, results in stunted limbs that maintain patterning. Notably, earlier loss of U11 corresponded to increased severity. We find that limb size is reduced due to elevated minor intron retention in minor intron-containing genes that regulate cell cycle. Limb progenitor cells experience delayed prometaphase to metaphase transition and prolonged S-phase, resulting in death of rapidly dividing, distally located progenitors. Consequently, crucial limb patterning genes are upregulated and their expression is maintained spatially to achieve basic patterning. Overall, these findings reveal a potential mechanism shared in disease and domestication.
TPX2 activation by GM130 controls astral microtubule formation and spindle orientation
Authors: Guo Haijing , Wei Jen-Hsuan , Seemann Joachim 
Published: March 2020

AbstractSpindle orientation is important in multiple developmental processes as it determines cell fate and function. The correct orientation of the mitotic spindle depends on the assembly of a proper astral microtubule network. Here, we report that the spindle assembly factor TPX2 regulates the astral microtubule network. TPX2 in the spindle pole area is activated by GM130 on mitotic Golgi membranes to promote astral microtubule growth. GM130 relieves TPX2 inhibition by competing for importin α binding. During mitosis, phosphorylation of importin α at Serine 62 by Cdk1 switches its substrate preference from TPX2 to GM130, thereby enabling competition-based activation. Importin α S62A impedes local TPX2 activation and compromises astral microtubule formation, ultimately resulting in misoriented spindles. Blocking the GM130-importin α-TPX2 pathway impairs astral microtubule growth. Our results reveal a novel role for TPX2 in the organization of astral microtubules. Furthermore, we show that the substrate preference of the important mitotic modulator importin α is regulated by Cdk1 phosphorylation.
Alterations in the human plasma lipidome in response to Tularemia vaccination
Authors: Maner-Smith Kristal M. , Ford David A. , Goll Johannes B. , Jensen Travis L. , Khadka Manoj , Colucci Jennifer K , Gelber Casey E. , Albert Carolyn J. , Bosinger Steve , Franke Jacob D. , Natrajan Muktha , Rouphael Nadine , Johnson Robert , Sanz Patrick , Anderson Evan J. , Hoft Daniel F. , Mulligan Mark , Ortlund Eric A. 
Published: March 2020

AbstractTularemia is a rare but highly contagious and potentially fatal disease caused by bacteriaFrancisella tularensiswhere as few as ten inhaled organisms can lead to an infection, making it one of the most infectious microorganisms known and a potential bioweapon. To better understand the response to a live, attenuated tularemia vaccine and the biological pathways altered post-vaccination, healthy adults were vaccinated by scarification and plasma was collected pre- and post-vaccination for longitudinal lipidomics studies. Using tandem mass spectrometry, we identified and quantified individual lipid molecular species within representative lipid classes in plasma to characterize alterations in the plasma lipidome during the vaccine response. Separately, we targeted oxylipins, a subset of lipid mediators involved in inflammatory pathways. We identified 14 differentially abundant lipid species from eight lipid classes. These included 5-Hydroxyeicosatetraenoic acid (5-HETE), an eicosanoid produced following arachidonic acid liberation and epoxygenation, which is indicative of lipoxygenase activity and, subsequently, inflammation. Results suggest that 5-HETE was metabolized to a dihydroxyeicosatrienoic acid (DHET) by Day 7 post-vaccination, shedding light on the kinetics of the 5-HETE-mediated inflammatory response. In addition to 5-HETE and DHET, we observed pronounced changes in 34:1 phosphatidylinositol, anandamide, oleamide, ceramides, 16:1 cholesteryl ester, and several glycerophospholipids, several of these changes in abundance were correlated with serum cytokines and T cell activation. These data provide new insights into alterations in plasma lipidome post tularemia vaccination, potentially identifying key mediators and pathways involved in vaccine response and efficacy.
Rottlerin ameliorates DSS-induced colitis by improving intestinal barrier function via activation of the Epac-2/Rap-1 signaling pathway
Authors: Song Xue , Zuo Lugen , Wang Luyao , Zhu Zihan , Tao Jing , Jiang Yifan , Wu Xiaopei , Wang Zhikun , Nian Jing , Xiang Ping , Zhang Xiaofeng , Zhao Hao , Yu Liang , Li Jing , Hu Jianguo 
Published: March 2020

ABSTRACTOBJECTIVESRottlerin, a pan PDE inhibitor, has a variety of pharmacological activities, including enhancing barrier function and mediating anti-inflammatory activity by changing the distribution of occludin and ZO-1. Nevertheless, the function of rottlerin on Crohn disease (CD) keep unknown. Our aim of the study is to investigate the role of rottlerin on CD-like colitis and its mechanism.METHODSWild-type mice which were 8-10 weeks old were randomly divided into three treatment groups: (i) the normal feeding, no administration (control) group, (ii) the group administered 3% dextran sodium sulfate (DSS) alone, and (iii) the group administered rottlerin (100 mg/kg) and 3% DSS. In this study, the effect of rottlerin on the function and structure of the intestinal barrier was investigated, and the possible mechanism was discussed. We performed signaling pathway analysis and flow cytometry to identify the detailed mechanisms by which rottlerin (10 μg/mL) treatment inhibits cell growth arrest and the attenuation of TJ proteins in LPS-treated FHs 74 int cells.RESULTSRottlerin treatment significantly ameliorated colitis induced by DSS in WT mice, which was manifested by a decrease in inflammation score, the attenuation of inflammatory factors and the inhibition of destruction on intestinal barrier structure. Rottlerin enhanced the levels of occludin and ZO-1, and improved the function of intestinal barrier, which may have been why rottlerin ameliorated colitis in WT mice. The anti-inflammatory effect of rottlerin may be partly due to the activation of Epac-2/Rap-1 signaling.CONCLUSIONSRottlerin may treat CD in humans via enhancing TJ proteins expression and improving the function of intestinal barrier.
Single-shot Mesoporous Silica Rods Scaffold for Induction of Humoral Responses Against Small Antigens
Authors: Dellacherie Maxence O , Li Aileen , Lu Beverly Y , Verbeke Catia S , Gu Luo , Stafford Alexander G. , Doherty Edward J. , Mooney David J. 
Published: March 2020

AbstractVaccines have shown significant promise in eliciting protective and therapeutic responses. However, most effective vaccines require several booster shots, and it is challenging to generate potent responses against small molecules and synthetic peptide antigens often used to increase target specificity and improve vaccine stability. As continuous antigen uptake and processing by APCs and persistent toll-like receptor (TLR) priming have been shown to amplify antigen specific humoral immunity, we explored whether a single injection of a mesoporous silica micro-rod (MSR) vaccine containing synthetic molecules and peptides can effectively generate potent and durable antigen-specific humoral immunity. A single injection of the MSR vaccine against a gonadotropin-releasing hormone (GnRH) decapeptide elicited highly potent anti-GnRH response that lasted for over 12 months. The MSR vaccine generated higher titers than bolus or alhydrogel alum vaccine formulations. Moreover, a MSR vaccine directed against a Her2/neu peptide within the Trastuzumab binding domain showed immunoreactivity to native Her2 protein on tumor cell surface and, when directed against nicotine, generated long-term anti-nicotine antibodies. Mechanistically, we found that the MSR vaccine induced persistent germinal center (GC) B-cell activity for more than 3 weeks after a single injection, generation of memory B cells, and that at least 7 days of immunostimulation by the vaccine was required to generate an effective humoral response. Together, these data suggest that the MSR vaccine represents a promising technology for synthetic antigen vaccines to bypass the need for multiple immunizations and enhance long-term production of antibodies against endogenous antigens in the context of reproductive biology, cancer, and chronic addiction.
Substrate deformation regulates DRM2-mediated DNA methylation in plants
Authors: Fang Jian , Leichter Sarah M. , Jiang Jianjun , Biswal Mahamaya , Lu Jiuwei , Zhang Zhi-Min , Ren Wendan , Zhai Jixian , Cui Qiang , Zhong Xuehua , Song Jikui 
Published: March 2020

AbstractDNA methylation is an important epigenetic mechanism that critically regulates gene expression and genomic stability. In plants, Domains Rearranged Methyltransferase 2 (DRM2) preferentially mediates CHH methylation (H=C, T, A), a substrate specificity distinct from that of mammalian DNA methyltransferases. However, the underlying mechanism is unknown. Here, we report structure-function characterizations of DRM2-mediated methylation. An arginine finger from the catalytic loop intercalates into DNA minor groove, inducing large DNA deformation that impacts the substrate specificity of DRM2. To accommodate the substrate deformation, the target recognition domain of DRM2 embraces the enlarged DNA major groove via shape complementarity, disruption of which via C397R mutation shifts the substrate specificity of DRM2 toward CHG DNA. This study uncovers DNA deformation as a mechanism in regulating the substrate specificity of DRM2, implicative of transposon-specific repression in plants.
Blood vessel occlusion byCryptococcus neoformansis a mechanism for haemorrhagic dissemination of infection
Authors: Gibson Josie F , Bojarczuk Aleksandra , Evans Robert J , Kamuyango Alfred , Hotham Richard , Lagendijk Anne K , Hogan Benjamin M , Ingham Philip W , Renshaw Stephen A , Johnston Simon A 
Published: March 2020

AbstractMeningitis caused by infectious pathogens are associated with vessel damage and infarct formation, however the physiological cause is unknown.Cryptococcus neoformans, is a human fungal pathogen and causative agent of cryptococcal meningitis, where vascular events are observed in up to 30% of cases, predominantly in severe infection. Therefore, we aimed to investigate how infection may lead to vessel damage and associated pathogen dissemination using a zebrafish model forin vivolive imaging. We find that cryptococcal cells become trapped within the vasculature (dependent on there size) and proliferate there resulting in vasodilation. Localised cryptococcal growth, originating from a single or small number of cryptococcal cells in the vasculature was associated with sites of dissemination and simultaneously with loss of blood vessel integrity. Using a cell-cell junction tension reporter we identified dissemination from intact blood vessels and where vessel rupture occurred. Finally, we manipulated blood vessel stifness via cell junctions and found increased stiffness resulted in increased dissemination. Therefore, global vascular vasodilation occurs following infection, resulting in increased vessel tension which subsequently increases dissemination events, representing a positive feedback loop. Thus, we identify a mechanism for blood vessel damage during cryptococcal infection that may represent a cause of vascular damage and cortical infarction more generally in infective meningitis.
Asparagine signals mitochondrial respiration and can be targeted to impair tumour growth
Authors: Krall Abigail S. , Mullen Peter J. , Surjono Felicia , Momcilovic Milica , Schmid Ernst W. , Halbrook Christopher J. , Thambundit Apisadaporn , Mittelman Steven D. , Lyssiotis Costas A. , Shackelford David B. , Knott Simon R.V. , Christofk Heather R. 
Published: March 2020

AbstractMitochondrial respiration is critical for cell proliferation. In addition to producing ATP via the electron transport chain (ETC), respiration is required for the generation of TCA cycle-derived biosynthetic precursors, such as aspartate, an essential substrate for nucleotide synthesis. Because mTORC1 coordinates availability of biosynthetic precursors with anabolic metabolism, including nucleotide synthesis, a link between respiration and mTORC1 is fitting. Here we show that in addition to depleting intracellular aspartate, ETC inhibition depletes aspartate-derived asparagine and impairs mTORC1 activity. Providing exogenous asparagine restores mTORC1 activity, nucleotide synthesis, and proliferation in the context of ETC inhibition without restoring intracellular aspartate in a panel of cancer cell lines. As a therapeutic strategy, the combination of ETC inhibitor metformin, which limits tumour asparagine synthesis, and either asparaginase or dietary asparagine restriction, which limit tumour asparagine consumption, effectively impairs tumour growth in several mouse models of cancer. Because environmental asparagine is sufficient to restore proliferation with respiration impairment, bothin vitroandin vivo, our findings suggest that asparagine synthesis is a fundamental purpose of mitochondrial respiration. Moreover, the results suggest that asparagine signals active respiration to mTORC1 to communicate biosynthetic precursor sufficiency and promote anabolism.
The impact of enterprise unit policy change on the quantity demanded for crop insurance
Authors: Bulut Harun 
Published: March 2020

PurposeThe article examines the impact of policy change on enterprise unit subsidies that took place in 2009 on the quantity demanded for crop insurance.Design/methodology/approachThe analysis covers corn, soybeans, and wheat that are grown in six economic regions and uses various measures of purchasing such as acres insured, unit structure, coverage levels, as well as crop hail use as proxies for the quantity demanded.The analysis first employs time series econometric tools to analyze whether the time path of the share of enterprise units within buyup acres is influenced by the policy change in enterprise unit subsidies. It then comparatively examines the insurance experience between 2008 (right before the change) and 2015 (well after the change).FindingsFor corn, soybean, and wheat, the analysis establishes that the time path of the share of enterprise units within buyup coverage acres is statistically and economically influenced by the intervention. The analysis further quantifies the intervention's immediate and long-term impacts and finds that farmers' unit choices are highly responsive (elastic) to subsidy rates in those units.Between 2008 and 2015, the insurance experience generally indicates that the share of enterprise units within buyup coverage surged, the share of acres under catastrophic coverage declined, and the share acres in high coverage levels increased. Meanwhile, growers have increasingly utilized crop-hail policies.Originality/valueThis appears to be the first study (1) quantifying the sensitivity of farmers' unit choices with respect to subsidy rates in those units and finding that such choices are actually highly responsive (elastic), and (2) pointing out the interaction between MPCI and crop-hail products and offering insights as to their combined use. The findings should be of considerable value to policymakers, academics, bankers, and producers in regards to the design and use of risk management tools.
Effect of entrepreneurial orientation, market orientation and total quality management on performance
Authors: Ali Gamal Abdualmajed , Hilman Haim , Gorondutse Abdullahi Hassan 
Published: March 2020

PurposeThis study aims to examine the joint effect of entrepreneurial orientation (EO), market orientation (MO), total quality management (TQM) and organisational performance in the Kingdom of Saudi Arabia’s (KSA's) small and medium enterprise (SME) sector.Design/methodology/approachThis paper used a quantitative research design. A total of 393 questionnaires were distributed to and collected from owners/managers of SMEs in KSA to acquire requisite data for examining the hypothesised model of the study. Partial least squares structural equation modelling was used to analyse the research data.FindingsThe results indicate that EO, MO and TQM are positively and significantly related to the organisational performance of SMEs. In addition, the findings reveal that TQM has contributed the most to the enhancement of organisational development, which is followed by EO and finally, MO.Practical implicationsDeveloping economies consider SMEs as an approach to generate new jobs and enhance economic growth. The results provide owners/managers, practitioners and academicians with an enhanced understanding of the relationship and effects of EO, MO and TQM on organisational performance, particularly in the SME sector. Thus, owners/managers are guided to develop improved and further effective decisions for the implementation of TQM practices with strong EO and MO. Consequently, firms realise superior performance and continuously compete within their market.Originality/valueFrom the perspective of developing countries, this study contributes to the existing literature by providing empirical evidence regarding the effects of EO, MO, TQM and SMEs' performance. This study is the first to empirically examine the SME sector within the KSA's economy regarding the new orientation among decision makers in terms of increasing the significance of non-oil activities. This study also confirms the usability of resource-based view theory and strategic orientations variables in the KSA SMEs.
Interest rate liberalization and bank liquidity creation: evidence from China
Authors: Zhang Jiaming , Deng Xiangrong 
Published: March 2020

PurposeThis study aims to empirically analyze how interest rate liberalization affects bank liquidity creation, and investigate whether the relationship between them is linear.Design/methodology/approachBased on panel data on 145 banks in China over the period 1997–2015, this paper regresses the econometric model by conducting feasible generalized least square estimation.FindingsThe regression results show that, first, interest rate liberalization has a nonlinear impact on bank liquidity creation, and the relationship between them is inverted U-shaped. In other words, as interest rate liberalization progresses, bank liquidity creation increases first, and then decreases. Second, through the mediation effect tests, this study found that interest rate liberalization affects bank liquidity creation through bank risk-taking. That is, interest rate liberalization leads to changes in bank risk-taking, thus resulting in changes in bank liquidity creation.Research limitations/implicationsThe effect of interest rate liberalization on bank liquidity creation is nonlinear, so promoting interest rate liberalization faces a trade-off because excessive bank liquidity creation may lead to asset price bubbles, while insufficient bank liquidity creation may inhibit economic growth.Practical implicationsInterest rate liberalization has a significant impact on bank liquidity creation; therefore, bank liquidity creation should be added to the objective function of the regulator that determines interest rate liberalization reform in China.Social implicationsInterest rate liberalization has a direct impact on bank risk-taking, so the consequences of interest rate liberalization should be included in the framework of macro-prudential supervision.Originality/valueInterest rate liberalization is one of the most important financial reforms in China, yet its potential impact on firm-level bank liquidity is little explored. This paper attempts to fill the gap.