bluuebunny/crossref_metadata_2025_split huggingface dataset:
GB_SynP: a modular dCas9-regulated synthetic promoter collection for fine-tuned recombinant gene expression in plants
Authors: Moreno-Giménez Elena , Selma Sara , Calvache Camilo , Orzáez Diego 
Published: May 2022

ABSTRACTProgramable transcriptional factors based on the CRISPR architecture are becoming commonly used in plants for endogenous gene regulation. In plants, a potent CRISPR tool for gene induction is the so-called dCasEV2.1 activation system, which has shown remarkable genome-wide specificity combined with a strong activation capacity. To explore the ability of dCasEV2.1 to act as a transactivator for orthogonal synthetic promoters, a collection of DNA parts was created (GB_SynP) for combinatorial synthetic promoter building. The collection includes (i) minimal promoter parts with the TATA box and 5’UTR regions, (ii) proximal parts containing single or multiple copies of the target sequence for the gRNA, thus functioning as regulatory cis boxes, and (iii) sequence-randomized distal parts that ensure the adequate length of the resulting promoter. A total of 35 promoters were assembled using the GB_SynP collection, showing in all cases minimal background and predictable activation levels depending on the proximal parts used. GB_SynP was also employed in a combinatorial expression analysis of an auto-luminescence pathway in Nicotiana benthamiana, showing the value of this tool in extracting important biological information such as the determination of the limiting steps in an enzymatic pathway.
Molecular Determinants of Tissue Specificity of Flavivirus Nonstructural Protein 1 Interaction with Endothelial Cells
Authors: Lo Nicholas T.N. , Roodsari Susan , Tin Nicole R. , Biering Scott B. , Harris Eva 
Published: May 2022

AbstractMembers of the mosquito-borne flavivirus genus such as dengue (DENV), West Nile (WNV), and Zika (ZIKV) viruses cause distinct diseases and affect different tissues. We previously found that the secreted flaviviral nonstructural protein 1 (NS1) interacts with endothelial cells and disrupts endothelial barrier function in a tissue-specific manner consistent with the disease tropism of the respective viruses. However, the underlying molecular mechanism of this tissue-specific NS1-endothelial cell interaction is not well understood. To elucidate the distinct role(s) that the domains of NS1 (β-roll, wing, and β-ladder) play in NS1 interactions with endothelial cells, we constructed flavivirus NS1 chimeras that exchanged the wing and β-ladder domains in a pair-wise manner between DENV, WNV, and ZIKV NS1. We found that both the NS1 wing and β-ladder domains conferred NS1 tissue-specific endothelial dysfunction, with the wing conferring cell binding and the β-ladder involved in inducing endothelial hyperpermeability as measured by trans-endothelial electrical resistance assay. To narrow down the amino acids dictating cell binding specificity, we utilized the DENV-WNV NS1 chimera and identified residues 91 to 93 (GDI) of DENV NS1 as a molecular motif determining binding specificity. Further, using an in vivo mouse model of localized leak, we found that the GDI motif of the wing domain was essential for triggering DENV NS1-induced vascular leak in mouse dermis. Taken together, we identify molecular determinants of flavivirus NS1 that confer NS1 binding and vascular leak and highlight the importance of the NS1 wing domain for flavivirus pathogenesis.ImportanceFlavivirus NS1 is secreted into the bloodstream from infected cells during a viral infection. Dengue virus NS1 contributes to severe dengue pathology such as endothelial dysfunction and vascular leak independently of the virus. We have shown that multiple flavivirus NS1 proteins result in endothelial dysfunction in a tissue-specific manner consistent with their respective viral tropism. Here, we aimed to identify the molecular determinants that make some, but not other, flavivirus NS1 proteins bind to select endothelial cells in vitro and cause vascular leak in a mouse model. We identified the wing domain of NS1 as a primary determinant conferring differential endothelial dysfunction and vascular leak and narrowed the contributing amino acid residues to a three-residue motif within the wing domain. The insights from this study pave the way for future studies on the effects of flavivirus NS1 on viral dissemination and pathogenesis and offer potential new avenues for antiviral therapies.
Few amino acid mutations in H6 influenza A virus from South American lineage increase pathogenicity in poultry
Authors: Rimondi Agustina , Olivera Valeria S. , Soria Ivana , Parisi Gustavo D. , Rumbo Martin , Perez Daniel R. 
Published: May 2022

ABSTRACTIn chickens, infections with influenza A virus (IAV) can range from mild to severe and lethal. The study of IAV infections in poultry has been mostly limited to strains from the North American and Eurasian lineages, whereas limited information exists on similar studies with strains from the South American lineage (SAm). To better evaluate the risk of introduction of a prototypical SAm IAV strain into poultry, chickens were infected with a wild-type SAm origin strain (WT557/H6N2). The resulting virus progeny was serially passage in chickens 20 times and the immunopathological effects in chickens of the last passage virus, 20Ch557/H6N2, was compared to those of the parental strain. Comparison of complete viral genome sequences indicated that the 20Ch557/H6N2 strain contained 13 amino acid differences compared to the wild type strain. Five of these mutations are in functionally relevant regions of the viral surface glycoproteins hemagglutinin (HA) and neuraminidase (NA). However, despite higher and more prolonged virus shedding in chickens inoculated with the 20Ch557/H6N2 strain compared to those that received the WT557/H6N2 strain, transmission to naïve chickens was not observed for either group. Analyses by flow cytometry of mononuclear cells and lymphocyte subpopulations from the lamina propria and intraepithelial lymphocytic cells (IELs) from ileum revealed significant increase in the percentages of CD3+TCRγδ+ IELs in chickens inoculated with the 20Ch557/H6N2 strain compared to those inoculated with the WT557/H6N2 strain.
Signals Generated by Neutrophil Receptors for Danger Molecules Transactivate Allosterically Modulated FFA2R: Distinct response patterns are mediated by modulators recognized by different allosteric receptor sites
Authors: Lind Simon , Granberg Kenneth L. , Forsman Huamei , Dahlgren Claes 
Published: May 2022

AbstractPositive allosteric modulators for free fatty acid receptor 2 (FFA2R/GPR43), that affect receptor function through binding to two distinct allosteric binding sites, were used to determine the correlation between the responses induced in neutrophils by two distinct activation modes; FFA2R was activated either by the orthosteric agonist propionate or by a receptor transactivation mechanism that activated FFA2R from the cytosolic side of the neutrophil plasma membrane by signals generated by the neutrophil PAFR (receptor for platelet activating factor), P2Y2R (receptor for ATP), FPR1 (receptor for fMLF) and FPR2 (receptor for WKYMVM). We show that the transactivation signals that activate FFA2R in the absence of any orthosteric agonist were generated downstream of the signaling G protein that couple to PAFR and P2Y2R. This transactivation of allosterically modulated FFA2Rs, by signals generated by PAFR/P2Y2R, represents a novel mechanism by which a G protein coupled receptor can be activated. Weak correlations were obtained when the FFA2R activity was induced by the transactivation signals generated by PAFRs and P2Y2Rs were compared with the FFA2R activity induced by the orthosteric agonist propionate. Comparison of the responses for each allosteric modulator revealed that the ratio values, calculated from the peak values of the ATP and propionate responses, varied from 0.2 to 1. Depending on the allosteric modulator, the response induced by the two different mechanisms (orthosteric activation and receptor transactivation, respectively), was equal or the propionate response was more pronounced. Importantly, we conclude that FFA2R activation from outside (orthosteric activation) and inside (receptor cross-talk/transactivation) can be selectively affected by an allosteric FFA2R modulator.The allosterically modulated FFA2R is transactivated by signals generated by other GPCRs.The PAF and ATP receptors transactivate FFA2R from the cytosolic side of the membrane.The mechanisms that regulates activation of FFA2R from outside and inside differ.
CpG dinucleotide enrichment in the influenza A virus genome as a live attenuated vaccine development strategy
Authors: Sharp Colin P , Thompson Beth H , Nash Tessa J , Diebold Ola , Pinto Rute M , Thorley Luke , Lin Yao-Tang , Sives Samantha , Wise Helen , Hendry Sara Clohisey , Grey Finn , Vervelde Lonneke , Simmonds Peter , Digard Paul , Gaunt Eleanor R 
Published: May 2022

ABSTRACTSynonymous recoding of RNA virus genomes is a promising approach for generating attenuated viruses to use as vaccines. Problematically, recoding typically hinders virus growth, but this may be rectified using CpG dinucleotide enrichment. CpGs are recognised by cellular zinc-finger antiviral protein (ZAP), and so in principle, removing ZAP sensing from a virus propagation system will reverse attenuation of a CpG-enriched virus, enabling high titre yield of a vaccine virus. We tested this using a vaccine strain of influenza A virus (IAV) engineered for increased CpG content in genome segment 1. Virus attenuation was mediated by the short isoform of ZAP, correlated with the number of CpGs added, and was enacted via turnover of viral transcripts. The CpG-enriched virus was strongly attenuated in mice, yet conveyed protection from a potentially lethal challenge dose of wildtype virus. Importantly for vaccine development, CpG-enriched viruses were genetically stable during serial passage. Unexpectedly, in both MDCK cells and embryonated hens’ eggs that are used to propagate live attenuated influenza vaccines, the ZAP-sensitive virus was fully replication competent. Thus, ZAP- sensitive CpG enriched viruses that are defective in human systems can yield high titre in vaccine propagation systems, providing a realistic, economically viable platform to augment existing live attenuated vaccines.AUTHOR SUMMARYCpG dinucleotides are under-represented in vertebrate genomes, wherein cytosines in the CpG conformation are methylated to regulate transcription. Methylated cytosines are prone to deamination, resulting in TpG dinucleotides replacing CpGs. The resultant CpG suppression has provided a route by which vertebrate cells can recognise RNA from invading pathogens, using cellular Zinc-finger Antiviral Protein (ZAP) as a CpG sensor. Vertebrate-infecting RNA viruses also genomically suppress CpGs, and it is believed that this is an evolved trait to evade detection by ZAP. Here, we engineered an influenza A virus (IAV) with elevated CpG content and characterised how this impacts viral replication. CpG addition resulted in viral attenuation, mediated by ZAP activity. CpG suppression is conserved in dog and chicken genomes (relevant for live attenuated IAV vaccine propagation), and it is logical to predict that ZAP-mediated CpG sensing would also be conserved in these species. However, when we propagated ZAP-sensitive IAV in cognate culture systems, we saw no replication defect. This unexpected result raises questions about why viruses infecting these species suppress CpG in their genomes, and importantly delivers a new, tractable approach to augment rational live attenuated IAV vaccine design.
Neryl acetate, the major component of Corsican Helichrysum Italicum essential oil, mediates its biological activities on skin barrier
Authors: Lemaire Géraldine , Olivero Malvina , Rouquet Virginie , Moga Alain , Pagnon Aurélie , Cenizo Valérie , Portes Pascal 
Published: May 2022

ABSTRACTCorsican Helichrysum Italicum essential oil is characterized by high concentrations of neryl acetate, and we previously demonstrated that Corsican Helichrysum Italicum essential oil increases the expression of genes that are part of the differentiation complex (involucrin, small proline rich proteins, late cornified envelope, S100 protein family). The biological activities of Helichrysum Italicum essential oil and neryl acetate were compared to identify how neryl acetate contributes to Helichrysum Italicum essential oil activity on human skin. Neryl acetate, as a part component of Helichrysum Italicum essential oil, was tested on skin explant models for 24 hours and 5 days in comparison with Helichrysum Italicum essential oil. We analyzed the biological regulations in the skin explant by transcriptomic analysis, skin barrier protein immunofluorescence, lipid staining and ceramide analysis by liquid chromatography-mass spectrometry. Transcriptomic analysis revealed that 41.5% of Helichrysum Italicum essential oil-modulated genes were also regulated by neryl acetate and a selected panel of genes were confirmed by quantitative reverse transcription PCR analysis. Those genes are involved in epidermal differentiation, skin barrier formation and ceramide synthesis. Involucrin, involved in formation of the cornified envelope, was upregulated at both gene and protein levels after 24 hours and 5 days respectively. After 5 days of treatment, total lipids and ceramides were also increased. Our results demonstrate that neryl acetate mediates a large part of Corsican Helichrysum Italicum essential oil activity on skin barrier formation.
Intracellular injection of brain extracts from Alzheimer’s disease patients trigger unregulated Ca2+ release from intracellular stores that hinders cellular bioenergetics
Authors: Pensalfini Anna , Umar Abdul Rahim , Glabe Charles , Parker Ian , Ullah Ghanim , Demuro Angelo 
Published: May 2022

AbstractStrong evidence indicates that amyloid beta (Aβ) inflicts its toxicity in Alzheimer’s disease (AD) by promoting uncontrolled elevation of cytosolic Ca2+ in neurons. We have previously shown that synthetic Aβ42 oligomers stimulate abnormal intracellular Ca2+ release from the endoplasmic reticulum stores, suggesting that a similar mechanism of Ca2+ toxicity may be common to the endogenous Aβs oligomers. To investigate this possibility, we use human postmortem brain extracts from control and AD-affected patients and test their ability to trigger Ca2+ fluxes when injected intracellularly into Xenopus oocytes. Immunological characterization of samples from AD patients revealed elevated content of soluble Aβ oligomers, detected by the conformation-dependent OC-antibody, whereas no immunoreactivity was detected in the normal samples. Intracellular injection of brain extracts from control patients failed to trigger detectable changes in intracellular Ca2+. Conversely, brain extracts from AD patients triggered Ca2+ events consisting of local and global Ca2+ fluorescent transients rising within few seconds after injection and persisting for several seconds. Pre-incubation of brain extracts with the conformation specific OC antibody completely suppressed brain extract ability to trigger cytosolic Ca2+ events. Comparison of the elementary events triggered by brain extracts and synthetic Aβ42 oligomer showed comparable temporal evolution and amplitudes to events triggered by direct injection of IP3. Moreover, bath application of caffeine reversibly inhibited local and global Ca2+ signals in all the samples confirming the involvement of Ca2+ release from the ER. Analysis of the recorded Ca2+ fluorescence signals by computational modeling allowed quantification of the IP3 and Ca2+ generated by each sample. The model further shows that the abnormal increase of Ca2+ and IP3 may affect mitochondrial bioenergetics. These results, supports the hypothesis that endogenous amyloid oligomer contained in neurons of AD-affected brains may represent the toxic agents responsible for neurons malfunctioning and death, associated with the disruption of neuronal Ca2+ homeostasis.
Atomic model of Vesicular Stomatitis Virus and Mechanism of Assembly
Authors: Zhou Kang , Si Zhu , Ge Peng , Tsao Jun , Luo Ming , Zhou Z. Hong 
Published: May 2022

AbstractLike other negative-strand RNA viruses (NSVs) such as influenza and rabies, vesicular stomatitis virus (VSV) has a three-layered organization: a layer of matrix protein (M) resides between the membrane envelope, studded by glycoprotein (G), and the nucleocapsid, composed of the nucleocapsid protein (N) and the encapsidated genomic RNA. Lack of in situ atomic structures of these viral components has limited our understanding of the virion assembly mechanism. Here, by cryoEM and sub-particle reconstruction, we have determined the in situ structures of M and N inside VSV at 3.47 Å resolution. In the virion, N and M have a stoichiometry of 1:2. The in situ structures of both N and M differ from their crystal structures in their N-terminal segments and oligomerization loops. N-RNA, N-N, and N-M-M interactions govern the formation of the capsid. A double layer of M contributes to packaging of the helical nucleocapsid: the inner M (IM) joins neighboring turns of the N helix, while the outer M (OM) contacts G and the membrane envelope. The pseudo-crystalline organization of G is further mapped by cryoET. The mechanism of VSV assembly is delineated by the network interactions of these viral components.
Neurocomputational evidence that conflicting prosocial motives guide distributive justice
Authors: Li Yue , Hu Jie , Ruff Christian C. , Zhou Xiaolin 
Published: May 2022

AbstractIn the history of humanity, most conflicts within and between societies have originated from perceived inequality in resource distribution. How humans achieve and maintain distributive justice has therefore been an intensely studied issue. However, most research on the corresponding psychological processes has focused on inequality aversion and has been largely agnostic of other motives that may either align or oppose this behavioral tendency. Here we provide behavioral, computational, and neuroimaging evidence that distribution decisions are guided by three distinct motives - inequality aversion, harm aversion, and rank reversal aversion – that interact and in fact can also deter individuals from pursuing equality. At the neural level, we show that these three motives are encoded by separate neural systems, compete for representation in various brain areas processing equality and harm signals, and are integrated in the striatum, which functions as a crucial hub for translating the motives to behavior. Our findings provide a comprehensive framework for understanding the cognitive and biological processes by which multiple prosocial motives are coordinated in the brain to guide redistribution behaviors. This framework enhances our understanding of the brain mechanisms underlying equality-related behavior, suggests possible neural origins of individual differences in social preferences, and provides a new pathway to understand the cognitive and neural basis of clinical disorders with impaired social functions.
Choosing memory retrieval strategies: a critical role for inhibition in the dentate gyrus
Authors: Albrecht Anne , Müller Iris , Weiglein Aliće , Pollali Evangelia , Çalışkan Gürsel , Stork Oliver 
Published: May 2022

AbstractRemembering the location of food is essential for survival. Rodents and humans employ mainly hippocampus-dependent spatial strategies, but when being stressed they shift to striatum-mediated stimulus-based strategies. To investigate underlying brain circuits, we tested mice with a heightened stress susceptibility due to a lack of the GABA-synthetizing enzyme GAD65 (GAD65-/- mice) in a dual solution task. Here, GAD65-/- mice preferred to locate a food reward in an open field via a proximal cue, while their wildtype littermates preferred a spatial strategy. The analysis of cFos co-activation across brain regions and of stress-induced mRNA expression changes of GAD65 pointed towards the hippocampal dorsal dentate gyrus (dDG) as a central structure for mediating stress effects on strategy choices via GAD65. Reducing the GAD65 expression locally in the dDG by a shRNA mediated knock down was sufficient to replicate the phenotype of the global GAD65 knock out and to increase dDG excitability. Using DREADD vectors to specifically interfere with dDG circuit activity during dual solution retrieval but not learning confirmed that the dDG modulates strategy choices and that a balanced excitability of this structure is necessary to establish spatial strategy preference. These data highlight the dDG as a critical hub for choosing between spatial and non-spatial foraging strategies.
Gendered sense of safety and coping strategies in public places: a study in Atatürk Meydanı of Izmir
Authors: Şenol Fatma 
Published: April 2022

PurposeA threatened sense of safety in public spaces is a problem for liveable communities. For better public policies, this study investigates multi-dimensional and multi-scalar aspects of gendered perceived safety and strategies by women and men in daily public spaces.Design/methodology/approachA face-to-face survey with 40 men and 50 women in a public space (Izmir, Turkey) is deployed. Descriptive statistics and regression analysis compare participants' perceptions of and strategies for safety across the city, neighbourhood and the study site.FindingsTheir experienced-based familiarities in public places increase women's perceived safety. As safety strategies, different place-based and gendered-preconditions appear for women and men going “outside” especially “alone” (i.e. unaccompanied). Reaffirming female vulnerability in public places, gendered preconditions include individuals' attributes. Of place-based preconditions, crowd and police are significant mechanisms for safety but emphasized differently by women and men. Housewives' female companionship in the study site develops a class- and gender-based claim for a safe place away from their underserved neighbourhood.Practical implicationsGendered- and place preconditions for women's safety can inform design policies about surveillance and permeability of public spaces. Lack of data about class-based differences about perceived safety is a limitation.Originality/valueAmong a few, it takes perceived safety as performative acts with learned strategies across (rather than momentary perceptions in) socio-spatial spaces and provides a research framework that considers such acts with individual and spatial dimensions across multiple socio-spatial scales.