bluuebunny/crossref_metadata_2025 huggingface dataset:
Cytogenotoxic effects of 3-epicaryoptin inAllium cepaL. root apical meristem cells
Authors: Barman Manabendu , Ray Sanjib
Published: March 2021

Diterpenoid 3-epicaryoptin (C26H36O9) is abundant in the leaves ofClerodendrum inerme, a traditionally used medicinal plant, having insect antifeedant activities. Here, we aim to explore the cytogenotoxic effects of compound 3-epicaryoptin inAllium ceparoot apical meristem cells.A. ceparoots were treated with 3-epicaryoptin (100, 150 & 200 μg mL-1concentration) and the standard compound colchicine (200 μg mL-1concentration) for 2, 4, 4+16 h (4 h treatment followed by 16 h recovery). Cytogenotoxicity was analysed by studying the root growth retardation (RGR), mitotic index (MI), and cellular aberrations. The result showed statistically significant (p<0.01), concentration-dependent RGR effects of 3-epicaryoptin treatment compared with the negative control. Study of cell frequency in different phases of cell division observed a significant (p<0.001) increase in the metaphase cell percentage (66.2±0.58 %, 150 μg mL-1) and which subsequently caused an increase in the frequency of MI (12.29±0.34 %, 150 μg mL-1) at 4h of 3-epicaryoptin treatment and that was comparable with the colchicine action. The cytological study revealed that the 3-epicaryoptin treatment could induce different types of chromosomal abnormalities such as colchicine like metaphase, vagrant chromosomes, sticky chromosomes, anaphase-bridge, and an increased frequency of micronuclei and polyploid cells. These findings indicate that 3-epicaryoptin is cytogenotoxic, and thusC. inermeshould be used with caution in traditional medicine.
Detection of SARS-CoV-2 N501Y mutation by RT-PCR to identify the UK and the South African strains in the population of South Indian state of Telangana
Authors: Ahmed Safaa Muneer , Juvvadi Smita Rao , Kalapala Rakesh , Sreemanthula Jagadeesh Babu
Published: March 2021

Objective

To detect N501Y mutation of the SARS-CoV-2 spike protein by RT-PCR to distinguish (B.1.1.7) UK and (501Y.V2) South African strains from others in the population of Telangana and to determine its clinical implications.

Methods

A primer-probe mix that specifically detects the mutated N501Y strain by real time RT-PCR was designed. 93 samples that were reported positive for COVID-19 by our laboratory in the month of February 2021 were tested using our own primer-probe mix for the presence of N501Y by RT-PCR. The results of RT-PCR were validated by Sanger sequencing in representative samples. Sanger sequencing of other defining spike mutations of B.1.1.7 (del 69-70, del 144, N501Y, A570D, D614G, P681H, T716I, S982A and D1118H) and 501Y.V2 (K417N, E484K, N501Y and D614G) was also investigated.

Findings

Out of 93 COVID-19 positive samples, 12 samples are detected positive for N501Y by RT-PCR. Sanger sequencing of these 12 samples further confirmed the presence of N501Y and other mutations that are characteristic of UK strain (B.1.1.7). The South African strain (501Y.V2) is not detected in any of our samples in this study. But, the E484K mutation that is characteristic of 501Y.V2 is detected in one N501Y negative sample.

Conclusion

Strain-specific RT-PCR for N501Y was developed and validated with Sanger sequencing. Such strategy facilitates quick surveillance for more transmissible and more vaccine resistant strains.
Structural variant selection for high-altitude adaptation using single-molecule long-read sequencing
Authors: Shi Jinlong , Jia Zhilong , Zhao Xiaojing , Sun Jinxiu , Liang Fan , Park Minsung , Zhao Chenghui , Wang Xiaoreng , Chen Qi , Song Xinyu , Yu Kang , Jia Qian , Wang Depeng , Xiao Yuhui , Liu Yinzhe , Wu Shijing , Zhong Qin , Wu Jue , Cui Saijia , Bo Xiaochen , Wu Zhenzhou , Kellis Manolis , He Kunlun
Published: March 2021

Structural variants (SVs) can be important drivers of human adaptation with strong effects, but previous studies have focused primarily on common variants with weak effects. Here, we used large-scale single-molecule long-read sequencing of 320 Tibetan and Han samples, to show that SVs are key drivers of selection under high-altitude adaptation. We expand the landscape of global SVs, apply robust models of selection and population differentiation combining SVs, SNPs and InDels, and use epigenomic analyses to predict driver enhancers, target genes, upstream regulators, and biological functions, which we validate using enhancer reporter and DNA pull-down assays. We reveal diverse Tibetan-specific SVs affecting the cis- and trans-regulatory circuitry of diverse biological functions, including hypoxia response, energy metabolism, lung function, etc. Our study greatly expands the global SV landscape, reveals the central role of gene-regulatory circuitry rewiring in human adaptation, and illustrates the diverse functional roles that SVs can play in human biology.
Single Cell, Single Nucleus and Spatial RNA Sequencing of the Human Liver Identifies Hepatic Stellate Cell and Cholangiocyte Heterogeneity
Authors: Andrews Tallulah S. , Atif Jawairia , Liu Jeff C. , Perciani Catia T. , Ma Xue-Zhong , Thoeni Cornelia , Slyper Michal , Eraslan Gökcen , Segerstolpe Asa , Manuel Justin , Chung Sai , Winter Erin , Cirlan Iulia , Khuu Nicholas , Fischer Sandra , Rozenblatt-Rosen Orit , Regev Aviv , McGilvray Ian D. , Bader Gary D. , MacParland Sonya A.
Published: March 2021

The critical functions of the human liver are coordinated through the interactions of hepatic parenchymal and non-parenchymal cells. Recent advances in single cell transcriptional approaches have enabled an examination of the human liver with unprecedented resolution. However, dissociation related cell perturbation can limit the ability to fully capture the human liver’s parenchymal cell fraction, which limits the ability to comprehensively profile this organ. Here, we report the transcriptional landscape of 73,295 cells from the human liver using matched single-cell RNA sequencing (scRNA-seq) and single-nucleus RNA sequencing (snRNA-seq). The addition of snRNA-seq enabled the characterization of interzonal hepatocytes at single-cell resolution, revealed the presence of rare subtypes of hepatic stellate cells previously only seen in disease, and detection of cholangiocyte progenitors that had only been observed duringin vitrodifferentiation experiments. However, T and B lymphocytes and NK cells were only distinguishable using scRNA-seq, highlighting the importance of applying both technologies to obtain a complete map of tissue-resident cell-types. We validated the distinct spatial distribution of the hepatocyte, cholangiocyte and stellate cell populations by an independent spatial transcriptomics dataset and immunohistochemistry. Our study provides a systematic comparison of the transcriptomes captured by scRNA-seq and snRNA-seq and delivers a high-resolution map of the parenchymal cell populations in the healthy human liver.
The nucleotide addition cycle of the SARS-CoV-2 polymerase
Authors: Bera Subhas Chandra , Seifert Mona , Kirchdoerfer Robert N. , van Nies Pauline , Wubulikasimu Yibulayin , Quack Salina , Papini Flávia S. , Arnold Jamie J. , Canard Bruno , Cameron Craig E. , Depken Martin , Dulin David
Published: March 2021

Coronaviruses have evolved elaborate multisubunit machines to replicate and transcribe their genomes. Central to these machines are the RNA-dependent RNA polymerase subunit (nsp12) and its intimately associated cofactors (nsp7 and nsp8). We have used a high-throughput magnetic-tweezers approach to develop a mechanochemical description of this core polymerase. The core polymerase exists in at least three catalytically distinct conformations, one being kinetically consistent with incorporation of incorrect nucleotides. We provide the first evidence that an RdRp uses a thermal ratchet instead of a power stroke to transition from the pre- to post-translocated state. Ultra-stable magnetic tweezers enables the direct observation of coronavirus polymerase deep and long-lived backtrack that are strongly stimulated by secondary structure in the template. The framework presented here elucidates one of the most important structure-dynamics-function relationships in human health today, and will form the grounds for understanding the regulation of this complex.
Rbf/E2F1 control growth and endoreplication via steroid-independent Ecdysone Receptor signalling inDrosophilaprostate-like secondary cells
Authors: Sekar Aashika , Leiblich Aaron , Hellberg Josephine E.E.U. , Sarma Dhruv , Mendes Cláudia C. , Wainwright S. Mark , Gandy Carina , Goberdhan Deborah C.I. , Hamdy Freddie C. , Wilson Clive
Published: March 2021

Dysregulation of cell cycle components results in the development and progression of several cancer types. Unusually, loss of the tumour suppressor gene,Retinoblastoma(Rb), and consequent activation of transcription factor E2F1 have been linked to late-stage tumour progression in prostate cancer, rather than early-stage events. This change is associated with an androgen-independent form of cancer, castration-resistant prostate cancer (CRPC), which frequently still requires androgen receptor (AR) signalling. We have previously shown that binucleate secondary cells (SCs) of theDrosophila melanogastermale accessory gland (AG) share several functional and signalling similarities with human prostate epithelial cells. Upon mating, SC growth regulation switches from a steroid-dependent to a steroid-independent form of Ecdysone Receptor (EcR) control that induces genome endoreplication. Here, we demonstrate that theDrosophilaRb homologue, Rbf, and E2F1, as well as cell cycle regulators, Cyclin D (CycD) and Cyclin E (CycE), are key mediators of SC growth and endoreplication both in virgin and mated males. Importantly, we show that the CycD/Rbf/E2F1 axis requires the EcR, but not ecdysone, to trigger CycE-dependent endoreplication and associated growth in SCs after mating, mirroring changes in CRPC. We also demonstrate that excess Rbf activity reversibly suppresses binucleation in adult SCs. Overall, our work reveals mechanistic parallels between the physiological switch to hormone-independent EcR signalling in SCs, and the pathological switch seen in CRPC, and suggests that the latter may represent the dysregulation of a currently unidentified physiological process, which permits AR signalling when androgen levels are low.
Signal requirement for cortical potential of transplantable human neuroepithelial stem cells
Authors: Varga Balazs V. , Faiz Maryam , Yang Huijuan , Pivonkova Helena , Gao Shangbang , Khelifi Gabriel , Linderoth Emma , Zhen Mei , Hussein Samer M. , Nagy Andras
Published: March 2021

The cerebral cortex develops from dorsal forebrain neuroepithelial progenitor cells. Initial expansion of the progenitor cell pool is followed by the generation of neurons of all the cortical layers and later, astrocytes and oligodendrocytes. However, the regulatory pathways that control the expansion and maintenance of the neuroepithelial progenitor cell pool are currently unknown. Here we define six basic pathway components that regulate proliferation of cortically specified human neuroepithelial stem cells (cNESCs)in vitrowithout the loss of developmental potential. We show that activation of FGF and inhibition of BMP and ACTIVIN A signalling are required for long-term cNESC proliferation. We also demonstrate that cNESCs preserve dorsal telencephalon-specific potential when GSK3, AKT and nuclear CATENIN-β1 activity are low. Remarkably, regulation of these six pathway components supports the clonal expansion of cNESCs. Moreover, cNESCs differentiate to lower and upper layer cortical neurons bothin vitroandin vivo. Identifying the mechanisms that drive the self-renewal and fate of cNESCs decision of neuroepithelial stem cells is key to developing new stem cell-based therapeutic approaches to treat neurological conditions.
Single-cell motion of magnetotactic bacteria in microfluidic confinement: interplay between surface interaction and magnetic torque
Authors: Codutti Agnese , Charsooghi Mohammad A. , Cerdá-Doñate Elisa , Taïeb Hubert M. , Robinson Tom , Faivre Damien , Klumpp Stefan
Published: March 2021

Swimming microorganisms often experience complex environments in their natural habitat. The same is true for microswimmers in envisioned biomedical applications. The simple aqueous conditions typically studied in the lab differ strongly from those found in these environments and often exclude the effects of small volume confinement or the influence that external fields have on their motion. In this work, we investigate magnetically steerable microswimmers, specifically magnetotactic bacteria, in strong spatial confinement and under the influence of an external magnetic field. We trap single cells in micrometer-sized microfluidic chambers and track and analyze their motion, which shows a variety of different trajectories, depending on the chamber size and the strength of the magnetic field. Combining these experimental observations with simulations using a variant of an active Brownian particle model, we explain the variety of trajectories by the interplay between the wall interactions and the magnetic torque. We also analyze the pronounced cell-to-cell heterogeneity, which makes single-cell tracking essential for an understanding of the motility patterns. In this way, our work establishes a basis for the analysis and prediction of microswimmer motility in more complex environments.
High Throughput Virtual Screening and Validation of a SARS-CoV-2 Main Protease Non-Covalent Inhibitor
Authors: Clyde Austin , Galanie Stephanie , Kneller Daniel W. , Ma Heng , Babuji Yadu , Blaiszik Ben , Brace Alexander , Brettin Thomas , Chard Kyle , Chard Ryan , Coates Leighton , Foster Ian , Hauner Darin , Kertesz Vilmos , Kumar Neeraj , Lee Hyungro , Li Zhuozhao , Merzky Andre , Schmidt Jurgen G. , Tan Li , Titov Mikhail , Trifan Anda , Turilli Matteo , Van Dam Hubertus , Chennubhotla Srinivas C. , Jha Shantenu , Kovalevsky Andrey , Ramanathan Arvind , Head Martha S. , Stevens Rick
Published: March 2021

Despite the recent availability of vaccines against the acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the search for inhibitory therapeutic agents has assumed importance especially in the context of emerging new viral variants. In this paper, we describe the discovery of a novel non-covalent small-molecule inhibitor, MCULE-5948770040, that binds to and inhibits the SARS-Cov-2 main protease (Mpro) by employing a scalable high throughput virtual screening (HTVS) framework and a targeted compound library of over 6.5 million molecules that could be readily ordered and purchased. Our HTVS framework leverages the U.S. supercomputing infrastructure achieving nearly 91% resource utilization and nearly 126 million docking calculations per hour. Downstream biochemical assays validate this Mproinhibitor with an inhibition constant (Ki) of 2.9µM [95% CI 2.2, 4.0]. Further, using room-temperature X-ray crystallography, we show that MCULE-5948770040 binds to a cleft in the primary binding site of Mproforming stable hydrogen bond and hydrophobic interactions. We then used multipleµs-timescale molecular dynamics (MD) simulations, and machine learning (ML) techniques to elucidate how the bound ligand alters the conformational states accessed by Mpro, involving motions both proximal and distal to the binding site. Together, our results demonstrate how MCULE-5948770040 inhibits Mproand offers a springboard for further therapeutic design.

Significance Statement

The ongoing novel coronavirus pandemic (COVID-19) has prompted a global race towards finding effective therapeutics that can target the various viral proteins. Despite many virtual screening campaigns in development, the discovery of validated inhibitors for SARS-CoV-2 protein targets has been limited. We discover a novel inhibitor against the SARS-CoV-2 main protease. Our integrated platform applies downstream biochemical assays, X-ray crystallography, and atomistic simulations to obtain a comprehensive characterization of its inhibitory mechanism. Inhibiting Mprocan lead to significant biomedical advances in targeting SARS-CoV-2 treatment, as it plays a crucial role in viral replication.
Stress-driven tissue fluidization physically segments vertebrate somites
Authors: Shelton Elijah R. , Kim Sangwoo , Gross Ben J. , Wu Ray , Pochitaloff Marie , Lim Irene , Sletten Ellen M. , Campàs Otger
Published: March 2021

Shaping functional structures during embryonic development requires both genetic and physical control. During somitogenesis, cell-cell coordination sets up genetic traveling waves in the presomitic mesoderm (PSM) that orchestrate somite formation. While key molecular and genetic aspects of this process are known, the mechanical events required to physically segment somites from the PSM remain unclear. Combining direct mechanical measurements during somite formation, live imaging of cell and tissue structure, and computer simulations, here we show that somites are mechanically sectioned off from the PSM by a large, actomyosin-driven increase in anisotropic stress at the nascent somite-somite boundary. Our results show that this localized increase in stress drives the regional fluidization of the tissue adjacent to the forming somite border, enabling local tissue remodeling and the shaping of the somite. Moreover, we find that active tension fluctuations in the tissue are optimized to mechanically define sharp somite boundaries while minimizing somite morphological defects. Altogether, these results indicate that mechanical changes at the somite-somite border and optimal tension fluctuations in the tissue are essential physical aspects of somite formation.
A Synthesis of Game Theory and Quantitative Genetic Models of Social Evolution
Authors: McGlothlin Joel W. , Akçay Erol , Brodie Edmund D. , Moore Allen J. , Van Cleve Jeremy
Published: March 2021

Two popular approaches for modeling social evolution, evolutionary game theory and quantitative genetics, ask complementary questions but are rarely integrated. Game theory focuses on evolutionary outcomes, with models solving for evolutionarily stable equilibria, whereas quantitative genetics provides insight into evolutionary processes, with models predicting short-term responses to selection. Here we draw parallels between evolutionary game theory and interacting phenotypes theory, which is a quantitative genetic framework for understanding social evolution. First, we show how any evolutionary game may be translated into two quantitative genetic selection gradients, nonsocial and social selection, which may be used to predict evolutionary change from a single round of the game. We show that synergistic fitness effects may alter predicted selection gradients, causing changes in magnitude and sign as the population mean evolves. Second, we show how evolutionary games involving plastic behavioral responses to partners can be modeled using indirect genetic effects, which describe how trait expression changes in response to genes in the social environment. We demonstrate that repeated social interactions in models of reciprocity generate indirect effects and conversely, that estimates of parameters from indirect genetic effect models may be used to predict the evolution of reciprocity. We argue that a pluralistic view incorporating both theoretical approaches will benefit empiricists and theorists studying social evolution. We advocate the measurement of social selection and indirect genetic effects in natural populations to test the predictions from game theory, and in turn, the use of game theory models to aid in the interpretation of quantitative genetic estimates.
A Chemical-Enhanced System for CRISPR-Based Nucleic Acid Detection
Authors: Li Zihan , Zhao Wenchang , Ma Shixin , Li Zexu , Yao Yingjia , Fei Teng
Published: March 2021

The CRISPR-based nucleic acid detection systems such as SHERLOCK, DETECTR and HOLMES have shown great potential for point-of-care testing of viral pathogens, especially in the context of COVID-19 pandemic. Here we optimize several key parameters of reaction chemistry and develop a Chemical Enhanced CRISPR Detection system for nucleic acid (termed CECRID). For the Cas12a/Cas13a-based signal detection phase, we determine buffer conditions and substrate range for optimal detection performance. By comparing several chemical additives, we find that addition of L-proline can secure or enhance Cas12a/Cas13a detection capability. For isothermal amplification phase with typical LAMP and RPA methods, inclusion of L-proline can also enhance specific target amplification as determined by CRISPR detection. Using SARS-CoV-2 pseudovirus, we demonstrate CECRID has enhanced detection sensitivity over chemical additive-null method with either fluorescence or lateral flow strip readout. Thus, CECRID provides an improved detection power and system robustness towards practical application of CRISPR-based diagnostics.
A neural marker of eye contact highly impaired in autism spectrum disorder
Authors: Lio Guillaume , Corazzol Martina , Fadda Roberta , Doneddu Giuseppe , Demily Caroline , Sirigu Angela
Published: March 2021

Attention to faces and eye contact are key behaviors for establishing social bonds in humans. In Autism Spectrum Disorders (ASD) a neurodevelopmental disturbance characterized by poor communication skills, impaired face processing and gaze avoidance are critical clinical features for its diagnosis. The biological alterations underlying these impairments are not clear yet. Using high-density electroencephalography coupled with multi-variate pattern classification and group blind source separation methods we searched for face- and face components-related neural signals that could best discriminate neurotypical and ASD visual processing. First, we isolated a face-specific neural signal in the superior temporal sulcus peaking at 240ms after stimulus onset. A machine learning algorithm applied on the extracted neural component reached 74% decoding accuracy at the same latencies, dissociating the neurotypical population from ASD subjects in whom this signal was weak. Further, by manipulating attention to face parts we found that the signal-evoked power in neurotypical subjects varied as a function of the distance of the eyes in the face stimulus with respect to the viewers’ fovea, i.e. it was strongest when the eyes were projected on the fovea and weakest when projected in the retinal periphery. Such selective face and face-components neural modulations were not found in ASD individuals although they showed typical early face related P100 and the N170 signals. These findings show that dedicated cortical mechanisms related to face perception set neural priority for attention to eyes and that these mechanisms are altered in individuals with ASD.
Multi-omic meta-analysis of transcriptomes and the bibliome uncovers novel hypoxia-inducible genes
Authors: Ono Yoko , Bono Hidemasa
Published: March 2021

Hypoxia is a condition in which cells, tissues, or organisms are deprived of sufficient oxygen supply. Aerobic organisms have a hypoxic response system, represented by hypoxia-inducible factor 1-α (HIF1A), to adapt to this condition. Due to publication bias, there has been little focus on genes other than well-known signature hypoxia-inducible genes. Therefore, in this study, we performed a meta-analysis to identify novel hypoxia-inducible genes. We searched publicly available transcriptome databases to obtain hypoxia-related experimental data, retrieved the metadata, and manually curated it. We selected the genes that are differentially expressed by hypoxic stimulation, and evaluated their relevance in hypoxia by performing enrichment analyses. Next, we performed a bibliometric analysis using gene2pubmed data to examine genes that have not been well studied in relation to hypoxia. Gene2pubmed data that provides information about the relationship between gene and publication. We calculated and evaluated the number of reports and similarity coefficients of each gene to HIF1A, which is a representative gene in hypoxia studies. In this data-driven study, we report that several genes that were not known to be associated with hypoxia, including the G protein-coupled receptor 146 gene, are upregulated by hypoxic stimulation.
GCN5 Maintains Muscle Integrity by Acetylating YY1 to Promote Dystrophin Expression
Authors: Addicks Gregory C. , Zhang Hongbo , Ryu Dongryeol , Vasam Goutham , Marshall Philip L. , Green Alex E. , Patel Sonia , Kang Baeki E. , Kim Doyoun , Katsyuba Elena , Williams Evan G. , Renaud Jean-Marc , Auwerx Johan , Menzies Keir J.
Published: March 2021

This work identifies a novel role for the acetyltransferase GCN5 in regulating muscle integrity through inhibition of DNA binding activity of the transcriptional repressor YY1. Here we report that in mice a muscle-specific knockout of GCN5 (Gcn5skm-/-) reduces the expression of key structural muscle proteins, including dystrophin, resulting in myopathy. Supporting our observation, a meta-analysis between the differential transcriptome ofGcn5skm-/-muscle and all available open-access data sets identified top correlations with musculoskeletal diseases in humans. GCN5 was found to acetylate YY1 at two residues (K392 and K393), which disrupts the interaction between the YY1 zinc-finger region and DNA. De/acetylation mimics for these YY1 post-translational modifications modulated muscle structural gene expression and DNA binding. Analysis of human GTEx data also found positive and negative correlations between fiber diameter and GCN5 and YY1 respectively. Collectively, our results demonstrate that GCN5 acetyltransferase activity regulates YY1 DNA binding and expression of dystrophin to modulate muscle integrity.
Bacterial retrons enable precise gene editing in human cells
Authors: Zhao Bin , Chen Shi-An A. , Lee Jiwoo , Fraser Hunter B.
Published: March 2021

Retrons are bacterial genetic elements involved in anti-phage defense. They have the unique ability to reverse transcribe RNA into multicopy single-stranded DNA (msDNA) that remains covalently linked to their template RNA. Retrons coupled with CRISPR-Cas9 in yeast have been shown to improve editing efficiency of precise genome editing via homology-directed repair (HDR). HDR editing efficiency has been limited by challenges associated with delivering extracellular donor DNA encoding the desired mutation. In this study, we tested the ability of retrons to produce msDNA as donor DNA and facilitate HDR by tethering msDNA to guide RNA in HEK293T and K562 cells. Through heterologous reconstitution of retrons from multiple bacterial species with the CRISPR-Cas9 system, we demonstrated HDR rates of up to 11.3%. Overall, our findings represent the first step in extending retron-based precise gene editing to human cells.
Biasing RNA coarse-grained folding simulations with Small–Angle X–ray Scattering (SAXS) data
Authors: Mazzanti L. , Alferkh L. , Frezza E. , Pasquali S.
Published: March 2021

RNA molecules can easily adopt alternative structures in response to different environmental conditions. As a result, a molecule’s energy landscape is rough and can exhibits a multitude of deep basins. In the absence of a high-resolution structure, Small Angle X-ray Scattering data (SAXS) can narrow down the conformational space available to the molecule and be used in conjunction with physical modeling to obtain high-resolution putative structures to be further tested by experiments. Because of the low-resolution of this data, it is natural to implement the integration of SAXS data into simulations using a coarse-grained representation of the molecule, allowing for much wider searches and faster evaluation of SAXS theoretical intensity curves than with atomistic models. We present here the theoretical framework and the implementation of a simulation approach based on our coarse-grained model HiRE-RNA combined with SAXS evaluations “on-the-fly” leading the simulation toward conformations agreeing with the scattering data, starting from partially folded structures as the ones that can easily be obtained from secondary structures predictions based tools. We show on three benchmark systems how our approach can successfully achieve high-resolution structures with remarkable similarity with the native structure recovering not only the overall shape, as imposed by SAXS data, but also the details of initially missing base pairs.
Attentional dynamics of efficient codes
Authors: Młynarski Wiktor , Tkačik Gašper
Published: March 2021

Top-down attention is hypothesized to dynamically allocate limited neural resources to task-relevant computations. According to this view, sensory neurons are driven not only by stimuli but also by feedback signals from higher brain areas that adapt the sensory code to the goals of the organism and its belief about the state of the environment. Here we formalize this view by optimizing a model of population coding in the visual cortex for maximally accurate perceptual inference at minimal activity cost. The resulting optimality predictions reproduce measured properties of attentional modulation in the visual system and generate novel hypotheses about the functional role of top-down feedback, response variability, and noise correlations. Our results suggest that a range of seemingly disparate attentional phenomena can be derived from a general theory combining probabilistic inference with efficient coding in a dynamic environment.
Fast ontogenetic growth drives steep evolutionary scaling of metabolic rate
Authors: Norin Tommy
Published: March 2021

Metabolic rate (MR) changes with body mass (BM) as MR = aBMb, where a is a normalisation constant (log–log intercept) and b the scaling exponent (log–log slope). This scaling relationship is fundamental to biology and widely applied, yet a century of research has provided little consensus on why and how steeply metabolic rate scales with body mass. I here show that ontogenetic (within-individual) b can be strongly and positively related to growth rates of juvenile fish when food availability is naturally restricted, with fast growing individuals having steep and near-isometric metabolic scaling (b ≈ 1). I suggest that the steep evolutionary (among-species) scaling also found for fishes (b also approaching 1) is a by-product of natural selection for these fast growing individuals early in ontogeny, and that a weaker relationship between metabolic scaling and growth later in life causes variation in b at lower taxonomic levels (within orders or species). I support these ideas by showing that b within fish orders is linked to natural mortality rates of fish larvae.
PD-1 expression on NK cells can be related to cytokine stimulation and tissue residency
Authors: Wagner Arnika K , Kadri Nadir , Tibbitt Chris , van de Ven Koen , Bagawath-Singh Sunitha , Oliinyk Denys , LeGresly Eric , Campbell Nicole , Trittel Stephanie , Riese Peggy , Sandalova Tatyana , Achour Adnane , Kärre Klas , Chambers Benedict J
Published: March 2021

Although PD-1 was shown to be a hallmark of T cells exhaustion, controversial studies have been reported on the role of PD-1 on NK cells. Here, we found by flow cytometry and single cell RNA sequencing analysis that PD-1 can be expressed on MHC class I-deficient tumor-infiltrating NK cellsin vivo. We also demonstrate distinct alterations in the phenotype ofPD-1-deficient NK cells which in part could be attributed to a decrease in tumor-infiltrating NK cells inPD-1-deficient mice. NK cells fromPD-1-deficient mice exhibited a more mature phenotype which might reduce their capacity to migrate and killin vivo. Finally, our results demonstrate that PD-L1 molecules in membranes ofPD-1-deficient NK cells migrate faster than in NK cells from wildtype mice, suggesting that PD-1 and PD-L1 formcisinteractions with each other on NK cells.
Sexual maturation in Atlantic salmon male parr may be triggered both in early spring and late summer under standard farming conditions
Authors: Ciani Elia , von Krogh Kristine , Nourizadeh-Lillabadi Rasoul , Mayer Ian , Fontaine Romain , Weltzien Finn-Arne
Published: March 2021

Male Atlantic salmon (Salmo salar) display different sexual strategies, maturing either as parr during the freshwater phase (as sneaky spawners), or as post smolts following one or several years at sea. First sexual maturation (puberty) occurs at different times depending on environmental and genetic factors. To improve our knowledge on the timing (age and season) of first sexual maturation in Atlantic salmon male parr, we investigated pubertal activation in second generation farmed salmon from the Norwegian river Figgjo, reared under natural conditions of photoperiod and water temperature. Histological analysis, in combination with morphometric measurements, plasma androgen levels and pituitary gonadotropin gene expression analysis revealed that, as previously reported, some male parr initiated early sexual maturation in spring at one year of age. Interestingly, some male parr were observed to initiate sexual maturation already in autumn, six months after hatching (under-yearlings), much earlier than reported in previous studies. One-year old maturing males showed a low induction in gonadotropin levels, while under-yearling maturing males displayed a significant increase infshbtranscripts as compared to immature fish. Plasma testosterone, detectable also in immature males, increased constantly during testes development, while 11-ketotestosterone, undetectable in immature and early maturing males, increased during more advanced stages of maturation. A mild feminization of the testes (ovotestes) was detected in a subset of samples. This study brings new knowledge on the little investigated field of sexually maturing under-yearlings in Atlantic salmon. This is also the first study comparing the physiology of under-yearling vs one-year old maturing male parr, thus bringing new insights to the remarkable plasticity of Atlantic salmon puberty.
Fluorescence lifetime imaging of pH along the secretory pathway
Authors: Linders Peter T.A. , Ioannidis Melina , Beest Martin ter , van den Bogaart Geert
Published: March 2021

Many cellular processes are dependent on correct pH levels, and this is especially important for the secretory pathway. Defects in pH homeostasis in distinct organelles cause a wide range of diseases, including disorders of glycosylation and lysosomal storage diseases. Ratiometric imaging of the pH-sensitive mutant of green fluorescent protein (GFP), pHLuorin, has allowed for targeted pH measurements in various organelles, but the required sequential image acquisition is intrinsically slow and therefore the temporal resolution is unsuitable to follow the rapid transit of cargo between organelles. We therefore applied fluorescence lifetime imaging microscopy (FLIM) to measure intraorganellar pH with just a single excitation wavelength. We first validated this method by confirming the pH in multiple compartments along the secretory pathway and compared the pH values obtained by the FLIM-based measurements with those obtained by conventional ratiometric imaging. Then, we analyzed the dynamic pH changes within cells treated with Bafilomycin A1, to block the vesicular ATPase, and Brefeldin A, to block ER-Golgi trafficking. Finally, we followed the pH changes of newly-synthesized molecules of the inflammatory cytokine tumor necrosis factor (TNF)-α while they were in transit from the endoplasmic reticulum via the Golgi to the plasma membrane. The toolbox we present here can be applied to measure intracellular pH with high spatial and temporal resolution, and can be used to assess organellar pH in disease models.
Conserved and context-dependent roles for Pdgfrb signaling during zebrafish vascular mural cell development
Authors: Ando Koji , Shih Yu-Huan , Ebarasi Lwaki , Grosse Ann , Portman Daneal , Chiba Ayano , Mattonet Kenny , Gerri Claudia , Stainier Didier Y.R. , Mochizuki Naoki , Fukuhara Shigetomo , Betsholtz Christer , Lawson Nathan D.
Published: March 2021

Platelet derived growth factor beta and its receptor, Pdgfrb, play essential roles in the development of vascular mural cells, including pericytes and vascular smooth muscle. To determine if this role was conserved in zebrafish, we analyzedpdgfbandpdgfrbmutant lines. Similar to mouse,pdgfbandpdgfrbmutant zebrafish lack brain pericytes and exhibit anatomically selective loss of vascular smooth muscle coverage. Despite these defects,pdgfrbmutant zebrafish did not otherwise exhibit circulatory defects at larval stages. However, beginning at juvenile stages, we observed severe cranial hemorrhage and vessel dilation associated with loss of pericytes and vascular smooth muscle cells inpdgfrbmutants. Similar to mouse,pdgfrbmutant zebrafish also displayed structural defects in the glomerulus, but normal development of hepatic stellate cells. We also noted defective mural cell investment on coronary vessels with concomitant defects in their development. Together, our studies support a conserved requirement for Pdgfrb signaling in mural cells. In addition, these mutants provide an important model for definitive investigation of mural cells during early embryonic stages without confounding secondary effects from circulatory defects.

Summary statement

Genetic analysis in zebrafish demonstrates the conserved role of Pdgfb/Pdgfrb signaling in pericyte and vascular smooth muscle cell formation during vascular development in vertebrates.
A MALDI-TOF assay identifies nilotinib as an inhibitor of inflammation in acute myeloid leukaemia
Authors: Marín-Rubio José Luis , Heap Rachel E. , Heunis Tiaan , Dueñas Maria Emilia , Inns Joseph , Scott Jonathan , Simpson A. John , Blair Helen , Heidenreich Olaf , Allan James M. , Saxty Barbara , Trost Matthias
Published: March 2021

Inflammatory responses are important in cancer, particularly in the context of monocyte-rich aggressive myeloid neoplasm. We developed a label-free cellular phenotypic drug discovery assay to identify anti-inflammatory drugs in human monocytes derived from acute myeloid leukaemia (AML), by tracking several biological features ionizing from only 2,500 cells using matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry. A proof-of-concept screen showed that the BCR-ABL inhibitor nilotinib, but not the structurally similar imatinib, blocks inflammatory responses. In order to identify the cellular (off-)targets of nilotinib, we performed thermal proteome profiling (TPP). Unlike imatinib, nilotinib and other later generation BCR-ABL inhibitors inhibit the p38α-MK2/3 signalling axis which suppressed the expression of inflammatory cytokines, cell adhesion and innate immunity markers in activated human monocytes derived from AML. Thus, our study provides a tool for the discovery of new anti-inflammatory drugs, which could contribute to the treatment of inflammation in myeloid neoplasms and other diseases.

Key Points

Label-free cell-based assay identifies new anti-inflammatory drugs using MALDI-TOF MS. Nilotinib reduces inflammation by inhibition of MAPK14-MK2/3 signalling axis in AML.
Long-read cDNA sequencing identifies functional pseudogenes in the human transcriptome
Authors: Troskie Robin-Lee , Jafrani Yohaann , Mercer Tim R. , Ewing Adam D. , Faulkner Geoffrey J. , Cheetham Seth W.
Published: March 2021

Pseudogenes are gene copies presumed to mainly be functionless relics of evolution due to acquired deleterious mutations or transcriptional silencing. When transcribed, pseudogenes may encode proteins or enact RNA-intrinsic regulatory mechanisms. However, the extent, characteristics and functional relevance of the human pseudogene transcriptome are unclear. Short-read sequencing platforms have limited power to resolve and accurately quantify pseudogene transcripts owing to the high sequence similarity of pseudogenes and their parent genes. Using deep full-length PacBio cDNA sequencing of normal human tissues and cancer cell lines, we identify here hundreds of novel transcribed pseudogenes. Pseudogene transcripts are expressed in tissue-specific patterns, exhibit complex splicing patterns and contribute to the coding sequences of known genes. We survey pseudogene transcripts encoding intact open reading frames (ORFs), representing potential unannotated protein-coding genes, and demonstrate their efficient translation in cultured cells. To assess the impact of noncoding pseudogenes on the cellular transcriptome, we delete the nucleus-enriched pseudogene PDCL3P4 transcript from HAP1 cells and observe hundreds of perturbed genes. This study highlights pseudogenes as a complex and dynamic component of the transcriptional landscape underpinning human biology and disease.
The DBL-1/TGF-β signaling pathway regulates pathogen-specific innate immune responses inC. elegans
Authors: Madhu Bhoomi , Gumienny Tina L.
Published: March 2021

Innate immunity in animals is orchestrated by multiple cell signaling pathways, including the TGF-β superfamily pathway. While the role of TGF-β signaling in innate immunity has been clearly identified, the requirement for this pathway in generating specific, robust responses to different bacterial challenges has not been characterized. Here, we address the role of DBL-1/TGF-β in regulating signature host defense responses to a wide range of bacteria inC. elegans. This work reveals a role of DBL-1/TGF-β in animal survival, organismal behaviors, and molecular responses in different environments. Additionally, we identify a novel role for SMA-4/Smad that suggests both DBL-1/TGF-β-dependent and -independent functions in host avoidance responses. RNA-seq analyses and immunity reporter studies indicate DBL-1/TGF-β differentially regulates target gene expression upon exposure to different bacteria. Furthermore, the DBL-1/TGF-β pathway is itself differentially affected by the bacteria exposure. Collectively, these findings demonstrate bacteria-specific host immune responses regulated by the DBL-1/TGF-β signaling pathway.
Vagabond: bond-based parametrization reduces overfitting for refinement of proteins
Authors: Ginn Helen M.
Published: March 2021

Structural biology methods have delivered over 150 000 high-resolution structures of macromolecules, which have fundamentally altered our understanding of biology and our approach to developing new medicines. However, the description of molecular flexibility is instrinsically flawed and in almost all cases, regardless of the experimental method used for structure determination, there remains a strong overfitting bias during molecular model building and refinement. In the worst case this can lead to wholly incorrect structures and thus incorrect biological interpretations. Here, by reparametrizing the description of these complex structures in terms of bonds rather than atomic positions, and by modelling flexibility using a deterministic ensemble of structures, it is demonstrated that structures can be described using fewer parameters than in conventional refinement. The current implementation, applied to X-ray diffraction data, significantly reduces the extent of overfitting, allowing the experimental data to reveal more biological information in electron-density maps.
Theoretical and practical approaches to improve the performance of local correlation algorithms for volume data analysis and shape recognition
Authors: Titarenko Valeriy , Roseman Alan M.
Published: March 2021

In this paper, several approaches to be used to accelerate algorithms for fitting an atomic structure into a given 3D density map determined by cryo-EM are discussed. Rotation and translation of the atomic structure to find similarity scores are used and implemented with discrete Fourier transforms. Several rotations can be combined into groups to accelerate processing. The finite resolution of experimental and simulated maps allows a reduction in the number of rotations and translations needed in order to estimate similarity-score values.
Pre-service teachers' acquisition of scientific knowledge and scientific skills through inquiry-based laboratory activity
Authors: Valls-Bautista Cristina , Solé-LLussà Anna , Casanoves Marina
Published: March 2021

Purpose

Scientific inquiry is a leading methodology that promotes science process skills to acquire scientific knowledge. There is evidence that primary school teachers have difficulties introducing inquiry-based activities in their classrooms. Hence, adequate teacher instruction in inquiry methodology is important to apply inquiry-based activities in school science lessons. This work aims to analyse if pre-service teachers succeeded in developing scientific knowledge and scientific skills through the application of an inquiry laboratory activity.

Design/methodology/approach

This article is presented as a case study developed in a group of 82 pre-service teachers. This research methodology involved qualitative and quantitative data.

Findings

The results demonstrate that pre-service teachers could improve their scientific skills and knowledge through inquiry-based laboratory activity.

Originality/value

The present study assesses not only the scientific knowledge but also if students can acquire scientific skills by doing the inquiry laboratory activity and if these skills are related to low-order cognitive skills or high-order cognitive skills.
Five leading-edge K-12 districts’ decision-making processes for EdTech innovations
Authors: Dexter Sara , Francisco Aubrey , Luke Luna Christina
Published: March 2021

Purpose

The purpose of this study was to better understand K-12 district leaders' reasoning and processes for selecting and deploying EdTech instructional products, including which, if any, types of data are used to support decision-making.

Design/methodology/approach

This multisite case study of educational technology (EdTech) decision-making comprises five purposely selected districts at the leading edge of EdTech innovation. The unit of analysis was a recent purchase they had made of an instructional, classroom-oriented digital product (defined as a product used by teachers and/or students in the classroom for the purposes of student learning). The key leader heading up the purchase was interviewed, as were other leaders and a teacher who were involved in the decision-making process.

Findings

The processes districts used to make their purchasing decisions involved teachers, district leaders and technical specialists who considered usability, usage data and alignment with student learning and interoperability, respectively. While in some cases there were plans to collect data on student learning outcomes, districts did not uniformly emphasize that in their decision-making processes. Instead, the type of educational technology tool that was purchased influenced whether or not districts planned to seek out student-level outcome data as evidence of the product's efficacy. For the purchases associated with access to content, school leaders considered usage or log data generated by the program itself as sufficient indication that the program is “working.” Where the software's functionality encompassed skill development, leaders stated future plans to look at student-level outcomes as a means for judging if the program “worked.”

Originality/value

Few accounts of district decision-making about the adoption of educational technology innovations are present in the literature. These five cases provide insight into the role evidence plays in decisions to adopt educational technology.