bluuebunny/crossref_metadata_2025 huggingface dataset:
Staying in touch: how highly specialised moth pollinators track host plant phenology in unpredictable climates
Authors: Finch Jonathan T. D. , Power Sally A. , Welbergen Justin A. , Cook James M.
Published: April 2021

For pollinating insects that visit just a single flowering species, the co-occurrence of flowers and insects in time is likely to have critical implications for both plant and pollinator. Insects often utilise diapause to persist through periods in which resources are unavailable, timing their re-emergence by responding to the same environmental cues as their host plants. The obligate pollination mutualisms (OPMs) betweenEpicephalamoths (Gracillariidae) and their leaf flower host plants are some of the most specialised interactions between plants and insects. However, to date there have been very few studies ofEpicephalamoth lifecycles and none of how they synchronise their activity with the flowering of their host plants.Breynia oblongifolia(Phyllanthaceae) is known to be exclusively pollinated by two highly specific species ofEpicephalamoth (Gracillariidae). We surveyed populations of both the host plant and it’s pollinators over multiple years to determine their annual phenology and then modelled the climatic factors that drive their activity. Using our newly gained knowledge of moth and host plant phenology, we then looked for evidence of diapause at both the egg and pre-pupal stages. Our phenology surveys showed that although female flowers were present throughout the entire year, the abundance of flowers and fruits was highly variable between sites and strongly associated with local rainfall and photoperiod. Fruit abundance, but not flower abundance, was a significant predictor of adultEpicephalaactivity, suggesting that eggs or early instar larvae diapause within dormant flowers and emerge as fruits mature. Searches of overwintering flowers confirmed this, with many containing evidence of pollen and diapausing pollinators. We also observed the behaviour of adultEpicephalaprior to pupation and found that ~10% of the Autumn emergingEpicephalaenter diapause, eclosing to adulthood after 38-56 weeks. The remaining 90% of autumn emerging adults pupate directly with no diapause, suggesting a bet hedging strategy for adult emergence. As such,Epicephalamoths appear to utilise diapause at multiple stages in their lifecycle, and possibly bet hedging, in order to deal with variable flowering phenology and climatic unpredictability.
Simulations of fluorescence imaging in the oral cavity
Authors: Lyu Zheng , Jiang Haomiao , Xiao Feng , Rong Jian , Zhang Tingcheng , Wandell Brian , Farrell Joyce
Published: April 2021

We describe an end-to-end image systems simulation that models a device capable of measuring fluorescence in the oral cavity. Our software includes a 3D model of the oral cavity and excitation-emission matrices of endogenous fluorophores that predict the spectral radiance of oral mucosal tissue. The predicted radiance is transformed by a model of the optics and image sensor to generate expected sensor image values. We compare simulated and real camera data from tongues in healthy individuals and show that the camera sensor chromaticity values can be used to quantify the fluorescence from porphyrins relative to the bulk fluorescence from multiple fluorophores (elastin, NADH, FAD, and collagen). Validation of the simulations supports the use of soft-prototyping in guiding system design for fluorescence imaging.

© 2021 Optical Society of America under the terms of the OSA Open Access Publishing Agreement
Alcohol Cues Elicit Different Abnormalities in Brain Networks of Abstinent Men and Women with Alcohol Use Disorders
Authors: Sawyer Kayle S. , Oscar-Berman Marlene , Ruiz Susan Mosher , Marinkovic Ksenija , Valmas Mary M. , Harris Gordon J.
Published: April 2021

We employed fMRI in 84 men and women with and without a history of alcohol use disorders (ALC and NC, respectively), to explore how gender interacts with alcoholism as reflected in brain activity elicited by alcohol cues. Brain activation was measured in a working memory task (delayed matching-to-sample) with emotional faces as the sample and match cues. During the delay period, intervening distractors were either reward-salient cues (alcoholic beverages) or neutral cues (nonalcoholic beverages or scrambled pictures). ALC women (ALCw) had higher accuracy than ALC men (ALCm). Analyses of scans during the viewing of distractor images revealed significant group-by-gender interactions. Compared to NC men, ALCm evidenced lower activation contrast between reward-salient cues and neutral cues in default mode network regions (including superior prefrontal and precuneus areas), while ALCw had more activation than NC women. Similar interactions were observed for task-regions (including superior parietal, lateral occipital, and prefrontal areas). Region of interest analyses showed that the ALC group had significantly higher levels of activation throughout reward-related circuitry during alcohol distractor interference than during scrambled picture interference. These results suggest that abstinent ALCm and ALCw differ in processing reward-salient cues, which can impact treatment and recovery.

Highlights

Brain reward regions activate highly when individuals with a history of alcohol use disorders view alcoholic beverages.

The brain regions identified subserve vision, memory, and judgement.

Opposite abnormalities in activation patterns appeared for alcoholic men and women.
Envelope analysis of the human alpha rhythm reveals EEG Gaussianity
Authors: Hidalgo Víctor Manuel , Díaz Javier , Mpodozis Jorge , Letelier Juan-Carlos
Published: April 2021

The origin of the human alpha rhythm has been a matter of debate since Lord Adrian attributed it to synchronous neural populations in the occipital cortex. Although some authors have pointed out the Gaussian characteristics of the alpha rhythm, their results have been repeatedly disregarded in favor of Adrian’s interpretation; even though the first EEG Gaussianity reports can be traced back to the origins of the field. Here we revisit this problem using the envelope analysis — a method that relies on the fact that the coefficient of variation of the envelope (CVE) for continuous-time zero-mean Gaussian noise (as well as for any filtered sub-band) is equal to, thus making the CVE a fingerprint for Gaussianity. As a consequence, any significant deviation fromis linked to synchronous neural dynamics. We analyzed occipital EEG and iEEG data from massive public databases. Our results showed the human alpha rhythm can be characterized either as a synchronous or as a Gaussian signal based on the value of its CVE. Furthermore, Fourier analysis showed the canonical spectral peak at ≈ 10[Hz] is present in both the synchronous and Gaussian cases, thus demonstrating this same peak can be produced by different underlying neural dynamics. This study confirms the original interpretation of Adrian regarding the origin of the alpha rhythm but also opens the door for the study of Gaussianity in brain dynamics. These results suggest a broader interpretation for event-related synchronization/desynchronization (ERS/ERD) may be needed. Envelope analysis constitutes a novel complement to Fourier-based methods for neural signal analysis relating amplitude modulation patterns (CVE) to signal energy.
CyLaKS: the Cytoskeleton Lattice-based Kinetic Simulator
Authors: Fiorenza Shane A. , Steckhahn Daniel G. , Betterton Meredith D.
Published: April 2021

Interaction of cytoskeletal filaments, motor proteins, and crosslinkers drives important cellular processes including cell division and cell movement. Cytoskeletal networks also undergo nonequilibrium self-organization in reconstituted systems. An emerging problem in cytoskeletal modeling and simulation is spatiotemporal alteration of the dynamics of filaments, motors, and associated proteins. This can occur due to motor crowding and obstacles along filaments, motor interactions and direction switching, and changes, defects, and heterogeneity in the filament lattice. How such spatiotemporally varying cytoskeletal filaments and motor interactions affect their collective properties is not fully understood. We developed the Cytoskeleton Lattice-based Kinetic Simulator (CyLaKS) for problems with significant spatiotemporal variation of motor or filament properties. The simulation builds on previous work modeling motor mechanochemistry into a simulation with many interacting motors and/or associated proteins. CyLaKS also includes detailed-balance in binding kinetics and movement and lattice heterogeneity. The simulation framework is flexible and extensible for future modeling work. Here we illustrate use of CyLaKS to study long-range motor interactions, filament heterogeneity, motion of a heterodimeric motor, and how changing crosslinker number affects filament separation.
Respiratory coupling between prefrontal cortex and hippocampus of rats anesthetized with urethane in theta and non-theta states
Authors: Mofleh Rola , Kocsis Bernat
Published: April 2021

Respiratory modulation of forebrain activity, long considered hard to reliably separate from breathing artifacts, has been firmly established in recent years using a variety of advanced techniques. Respiratory related oscillation (RRO) is derived from rhythmic nasal airflow in the olfactory bulb (OB) and is conveyed to higher order brain networks, including the prefrontal cortex (PFC) and hippocampus (HC), where it may potentially contribute to communication between these structures by synchronizing their activities at the respiratory rate. RRO was shown to change with sleep-wake states, it is strongest in quiet waking, somewhat less in active waking, characterized with theta activity in the HC, and absent in sleep. The goal of this study was to test RRO synchronization between PFC and HC under urethane anesthesia where theta and non-theta states spontaneously alternate. We found that in theta states, PFC-HC coherences significantly correlated with OB-HC but not with OB-PFC, even though RRO was stronger in PFC than in HC. In non-theta states, PFC-HC synchrony correlated with coherences connecting OB to either PFC or HC. Thus, similar to freely behaving rats, PFC-HC synchrony at RRO was primarily dependent on the response of HC to the common rhythmic drive, but only in theta state. The findings help outlining the value and the limits of applications in which urethane-anesthetized rats can be used for modeling the neural mechanisms of RRO in behaving animals.
A mechanism with severing near barbed ends and annealing explains structure and dynamics of dendritic actin networks
Authors: Holz Danielle , Hall Aaron R. , Usukura Eiji , Yamashiro Sawako , Watanabe Naoki , Vavylonis Dimitrios
Published: April 2021

Single molecule imaging has shown that part of actin disassembles within a few seconds after incorporation into the dendritic filament network in lamellipodia, suggestive of frequent destabilization near barbed ends. To investigate the mechanisms behind network remodeling, we created a stochastic model with polymerization, depolymerization, branching, capping, uncapping, severing, oligomer diffusion, annealing, and debranching. We find that filament severing, enhanced near barbed ends, can explain the single molecule actin lifetime distribution, if oligomer fragments reanneal to free ends with rate constants comparable to in vitro measurements. The same mechanism leads to actin networks consistent with measured filament, end, and branch concentrations. These networks undergo structural remodeling, leading to longer filaments away from the leading edge, at the +/- 35° orientation pattern. Imaging of actin speckle lifetimes at sub-second resolution verifies frequent disassembly of newly-assembled actin. We thus propose a unified mechanism that fits a diverse set of basic lamellipodia phenomenology.
ThePseudomonas aeruginosaphosphodiesterase genenbdAis transcriptionally regulated by RpoS and AmrZ
Authors: Gerbracht Katrin , Zehner Susanne , Frankenberg-Dinkel Nicole
Published: April 2021

Pseudomonas aeruginosais an opportunistic pathogen causing serious infections in immune compromised persons. These infections are difficult to erase with antibiotics, due to the formation of biofilms. The biofilm lifecycle is regulated by the second messenger molecule c-di-GMP (bis-3,5-cyclic di-guanosine monophosphate).P. aeruginosaencodes 40 genes for enzymes presumably involved in the biosynthesis and degradation of c-di-GMP. A tight regulation of expression, subcellular localized function and protein interactions control the activity of these enzymes. In this work we elucidated the transcriptional regulation of the gene encoding the membrane-bound phosphodiesterase NbdA. We previously reported a transcriptional and posttranslational role of nitric oxide (NO) onnbdAand its involvement in biofilm dispersal. NO is released from macrophages during infections but can also be produced byP. aeruginosaitself during anaerobic denitrification. Recently however, contradictory results about the role of NbdA within NO-induced biofilm dispersal were published. Therefore, the transcriptional regulation ofnbdAwas reevaluated to obtain insights into this discrepancy. Determination of the transcriptional start site ofnbdAby 5’-RACE and subsequent identification of the promoter region revealed a shortened open reading frame (ORF) in contrast to the annotated one. In addition, putative binding sites for RpoS and AmrZ were discovered in the newly defined promoter region. Employing chromosomally integrated transcriptionallacZreporter gene fusions demonstrated a RpoS-dependent activation and AmrZ repression ofnbdAtranscription. In order to investigate the impact of NO onnbdAtranscription, conditions mimicking exogenous and endogenous NO were applied. While neither exogenous nor endogenous NO had an influence onnbdApromoter activity, deletion of the nitrite reductase genenirSstrongly increasednbdAtranscription independently of its enzymatic activity during denitrification. The latter supports a role of NirS inP. aeruginosaapart from its enzymatic function.

IMPORTANCE

The opportunistic pathogenPseudomonas aeruginosapossesses a network of genes encoding proteins for the turnover of the second messenger c-di-GMP involved in regulating-among others-the lifestyle switch between planktonic, motile cells and sessile biofilms. Insight into the transcriptional regulation of these genes is important for the understanding of the protein function within the cell. Determination of the transcriptional start site of the phosphodiesterase genenbdArevealed a new promoter region and consequently a shortened open reading frame for the corresponding protein. Binding sites for RpoS and AmrZ were identifiedin silicoand confirmed experimentally. Previously reported regulation by nitric oxide was reevaluated and a strong influence of the moonlighting protein NirS identified.
Oxygen production by an ammonia-oxidizing archaeon
Authors: Kraft Beate , Jehmlich Nico , Larsen Morten , Bristow Laura , Könneke Martin , Thamdrup Bo , Canfield Donald E.
Published: April 2021

Ammonia-oxidizing archaea (AOA) are one of the most abundant groups of microbes in the world’s oceans and are key players in the nitrogen cycle. Their energy metabolism, the oxidation of ammonia to nitrite, requires oxygen. Nevertheless, AOA are abundant in environments where oxygen is undetectable. In incubation experiments, where oxygen concentrations were resolved to the nanomolar range, we show that Nitrosopumilus maritimus produces oxygen (O2) and dinitrogen (N2). The pathway is not completely resolved, but it has nitric oxide as a key intermediate. Part of the oxygen produced is directly used for ammonia oxidation, while some accumulates in the surrounding environment. N. maritimus joins a small handful of organisms known to produce oxygen in the dark, and based on this ability, we re-evaluate their role in oxygen-depleted marine environments.
Both transient and sustained MPK3/6 activities positively control expression of NLR genes in PTI and ETI
Authors: Lang Julien , Genot Baptiste , Bigeard Jean , Colcombet Jean
Published: April 2021

Arabidopsis thalianaMitogen Activated Protein Kinases 3 and 6 (MPK3/6) are known to be activated transiently in PAMP-Triggered Immunity (PTI) and durably in Effector-Triggered Immunity (ETI). However the functional differences between these two kinds of activation kinetics and how they allow coordination of the two layers of plant immunity remain poorly understood. Here, by analysing suppressors of the phenotype caused by a constitutively active form of MPK3, we demonstrate that ETI-mediating nucleotide-binding domain leucine-rich repeat receptors (NLRs) and NLR signaling can act downstream of MPK3 activities. Moreover we provide evidence that both sustained and transient MPK3/6 activities positively control the expression of at least two NLR genes, AT3G04220 and AT4G1110. We further show that the ETI regulators NDR1 and EDS1 also contribute to the upregulations of these two NLRs not only in an ETI context but also in a PTI context. Remarkably, while in ETI, MPK3/6 activities are dependent on NDR1 and EDS1, they are not in PTI, suggesting that if the same actors are involved in the two layers of immunity, the way they are interconnected is different. Finally we demonstrate that expression of the NLR AT3G04220 is sufficient to induce expression of defense genes from the SA branch. Overall this study enlarges our knowledge of MPK3/6 functions during immunity and gives a new insight into the intrication of PTI and ETI.
A SIMPLI (Single-cell Identification from MultiPLexed Images) approach for spatially resolved tissue phenotyping at single-cell resolution
Authors: Bortolomeazzi Michele , Montorsi Lucia , Temelkovski Damjan , Keddar Mohamed Reda , Acha-Sagredo Amelia , Pitcher Michael J. , Basso Gianluca , Laghi Luigi , Rodriguez-Justo Manuel , Spencer Jo , Ciccarelli Francesca D.
Published: April 2021

Multiplexed imaging technologies enable the study of biological tissues at single-cell resolution while preserving spatial information. Currently, high-dimension imaging data analysis is technology-specific and requires multiple tools, restricting analytical scalability and result reproducibility. Here we present SIMPLI (Single-cell Identification from MultiPlexed Images), a novel, flexible and technology-agnostic software that unifies all steps of multiplexed imaging data analysis. After raw image processing, SIMPLI performs a spatially resolved, single-cell analysis of the tissue slide as wells as cell-independent quantifications of marker expression to investigate features undetectable at the cell level. SIMPLI is highly customisable and can run on desktop computers as well as high-performance computing environments, enabling workflow parallelisation for large datasets. SIMPLI produces multiple tabular and graphical outputs at each step of the analysis. Its containerised implementation and minimum configuration requirements make SIMPLI a portable and reproducible solution for multiplexed imaging data analysis.

SIMPLI is available at:https://github.com/ciccalab/SIMPLI.
A biophotoelectrochemical approach to unravelling the role of cyanobacterial cell structures in exoelectrogenesis
Authors: Wey Laura T. , Lawrence Joshua M. , Chen Xiaolong , Clark Robert , Lea-Smith David J. , Zhang Jenny Z. , Howe Christopher J.
Published: April 2021

Photosynthetic microorganisms can export electrons outside their cells, a phenomenon called exoelectrogenesis, which can be harnessed for solar energy conversion. However, the route electrons take from thylakoid membranes to the cell exterior is not understood. Electrochemistry is a powerful analytical technique for studying electron transfer pathways. Here, we show how photoelectrochemistry can be used to compare electron flux from cyanobacterial cells of different growth stages, species and with the outer layers systematically removed. We show that the periplasmic space contributes significantly to the photocurrent profile complexity of whole cells, indicating that it gates electron transfer in exoelectrogenesis. We found that although components of the type IV pili machinery do not have a role in exoelectrogenesis, they contribute significantly to cell-electrode adherence. This study establishes that analytical photoelectrochemistry and molecular microbiology provide a powerful combination to study exoelectrogenesis, enabling future studies to answer biological questions and advance solar energy conversion applications.
Establishing a Role of the Semantic Control Network in Social Cognitive Processing: A Meta-analysis of Functional Neuroimaging Studies
Authors: Diveica Veronica , Koldewyn Kami , Binney Richard J.
Published: April 2021

Most leading models of socio-cognitive processing devote little discussion to the nature and neuroanatomical correlates of cognitive control mechanisms. Recently, it has been proposed that the regulation of social behaviours could rely on brain regions specialised in the controlled retrieval of semantic information, namely the anterior inferior frontal gyrus (IFG) and posterior middle temporal gyrus. Accordingly, we set out to investigate whether the neural activation commonly found in social functional neuroimaging studies extends to these ‘semantic control’ regions. We conducted five coordinate-based meta-analyses to combine results of over 500 fMRI/PET experiments and identified the brain regions consistently involved in semantic control, as well as four social abilities: theory of mind, trait inference, empathy and moral reasoning. This allowed an unprecedented parallel review of the neural networks associated with each of these cognitive domains. The results confirmed that the anterior left IFG region involved in semantic control is reliably engaged in all four social domains. This suggests that social cognition could be partly regulated by the neurocognitive system underpinning semantic control.
Regimes and mechanisms of transient amplification in abstract and biological networks
Authors: Christodoulou Georgia , Vogels Tim P. , Agnes Everton J.
Published: April 2021

We use upper triangular matrices as abstract representations of neuronal networks and directly manipulate their eigenspectra and non-normality to explore different regimes of transient amplification. Counter–intuitively, manipulating the imaginary distribution can lead to highly amplifying regimes. This is noteworthy, because biological networks are constrained by Dale’s law and the non-existence of neuronal self-loops, limiting the range of manipulations in the real dimension. Within these constraints we can further manipulate transient amplification by controlling global inhibition.
Identification of Biochemical and Molecular Markers of Early Aging in Childhood Cancer Survivors
Authors: Ravera Silvia , Vigliarolo Tiziana , Bruno Silvia , Morandi Fabio , Marimpietri Danilo , Sabatini Federica , Dagnino Monica , Petretto Andrea , Bartolucci Martina , Muraca Monica , Biasin Eleonora , Haupt Riccardo , Zecca Marco , Fagioli Franca , Cilloni Daniela , Podestà Marina , Frassoni Francesco
Published: April 2021

Purpose

Survival rates of Childhood Cancer Patients have improved tremendously over the past four decades. However, cancer treatments are associated with an increased risk of developing an anticipated onset of chronic diseases typical of aging. Thus, we aimed to identify molecular/metabolic cellular alterations responsible for early aging in Childhood Cancer Survivors (CCS).

Patients and Methods

Biochemical, proteomic and molecular biology analyses were conducted on mononuclear cells (MNCs) isolated from peripheral blood of 196 CCS, comparing the results with those obtained on MNCs of 154 healthy subjects.

Results

Data demonstrate that CCS-MNCs show: i) inefficient oxidative phosphorylation associated with low energy status and a metabolic switch to lactate fermentation compared with age-matched normal controls; ii) increment of lipid peroxidation due to an unbalance among the oxidative stress production and the activation of the antioxidant defenses; (iii) significantly lower expression of genes and proteins involved in mitochondrial biogenesis and metabolism regulation, such as CLUH, PGC1-α, and SIRT6 in CCS, not observed in the age-matched healthy or elderly subjects. The application of a mathematical model based on biochemical parameters predicts that CCS have a biological age significantly increased by decades compared to the chronological age. Overall, the results show that the impact of chemo/chemoradiotherapy on mitochondria efficiency in 196 CCS was rather homogeneous, irrespective of cancer type, treatment protocols, and time elapsed from the end of the curative period.

Conclusions

Our study identifies some biochemical and molecular alterations possibly contributing to the pathophysiology of anticipated aging and metabolic deficiency described in CCS. These results may be useful in identifying approaches to restore the mitochondrial function, slowing down the aging and the associated pathological conditions in CCS.
Comparative anatomy of leaf petioles in temperate trees and shrubs
Authors: Filartiga Arinawa Liz , Klimeš Adam , Altman Jan , Nobis Michael Peter , Crivellaro Alan , Schweingruber Fritz , Doležal Jiří
Published: April 2021

Petioles are important plant organs connecting stems with leaf blades and affecting light-harvesting leaf ability as well as transport of water, nutrient and biochemical signals. Despite petiole’s high diversity in size, shape and anatomical settings, little information is available about their structural adaptations across evolutionary lineages and environmental conditions. To fill our knowledge gap, we investigated the variation of petiole morphology and anatomy in 95 European woody plant species using phylogenetic comparative models. Two major axes of variation were related to leaf area (from large and soft to small and tough leaves), and plant size (from cold-adapted shrubs to warm-adapted tall trees). Larger and softer leaves are found in taller trees of more productive habitats. Their petioles are longer, with a circular outline, thin cuticles without trichomes, and are anatomically characterised by the predominance of sclerenchyma, larger vessels, interfascicular areas with fibers, indistinct phloem rays, and the occurrence of prismatic crystals and druses. In contrast, smaller and tougher leaves are found in shorter trees and shrubs of colder or drier habitats. Their petioles are characterized by teret outline, thick cuticle, simple and non-glandular trichomes, epidermal cells smaller than cortex cells, phloem composed of small cells and radially arranged vessels, fiberless xylem, lamellar collenchyma, acicular crystals and secretory elements. Individual anatomical traits were linked to different internal and external drivers. The petiole length and vessel conduit size increase, while cuticle thickness decreases, with increasing leaf blade area. Epidermis cell walls are thicker in leaves with higher specific leaf area. Collenchyma becomes absent with increasing temperature, epidermis cell size increases with plant height and temperature, and petiole outline becomes polygonal with increasing precipitation. We conclude that species temperature and precipitation optima, plant height, leaf area and thickness exerted a significant control on petiole anatomical and morphological structures not confounded by phylogenetic inertia. Unrelated species with different evolutionary histories but similar thermal and hydrological requirements have converged to similar petiole anatomical structures. Our findings contribute to improving current knowledge about the functional morphoanatomy of the petiole as the key organ that plays a crucial role in the hydraulic pathways in plants.
Fine-tuning the Spike: Role of the nature and topology of the glycan shield in the structure and dynamics of the SARS-CoV-2 S
Authors: Harbison Aoife M. , Fogarty Carl A. , Phung Toan K. , Satheesan Akash , Schulz Benjamin L. , Fadda Elisa
Published: April 2021

The dense glycan shield is an essential feature of the SARS-CoV-2 spike (S) architecture, key to immune evasion and to the activation of the prefusion conformation. Recent studies indicate that the occupancy and structures of the SARS-CoV-2 S glycans depend not only on the nature of the host cell, but also on the structural stability of the trimer; a point that raises important questions about the relative competence of different glycoforms. Moreover, the functional role of the glycan shield in the SARS-CoV-2 pathogenesis suggests that the evolution of the sites of glycosylation is potentially intertwined with the evolution of the protein sequence to affect optimal activity. Our results from multi-microsecond molecular dynamics simulations indicate that the type of glycosylation at N234, N165 and N343 greatly affects the stability of the receptor binding domain (RBD) open conformation, and thus its exposure and accessibility. Furthermore, our results suggest that the loss of glycosylation at N370, a newly acquired modification in the SARS-CoV-2 S glycan shield’s topology, may have contributed to increase the SARS-CoV-2 infectivity as we find that N-glycosylation at N370 stabilizes the closed RBD conformation by binding a specific cleft on the RBD surface. We discuss how the absence of the N370 glycan in the SARS-CoV-2 S frees the RBD glycan binding cleft, which becomes available to bind cell-surface glycans, potentially increases host cell surface localization.
Increasing role of pyrethroid-resistant Anopheles funestus in malaria transmission in the Lake Zone, Tanzania: implications for the evaluation of novel vector control products
Authors: Matowo Nancy S. , Martin Jackline , Kulkarni Manisha A. , Mosha Jacklin F. , Lukole Eliud , Isaya Gladness , Shirima Boniface , Kaaya Robert , Moyes Catherine , Hancock Penelope A. , Rowland Mark , Manjurano Alphaxard , Mosha Franklin W , Protopopoff Natacha , Messenger Louisa A.
Published: April 2021

Anopheles funestus is playing an increasing role in malaria transmission in parts of sub-Saharan Africa, where An. gambiae s.s. has been effectively controlled by long-lasting insecticidal nets. We investigated vector population bionomics, insecticide resistance and malaria transmission dynamics in 86 study clusters in North-West Tanzania. An. funestus s.l. represented 94.5% (4740/5016) of all vectors and was responsible for the majority of malaria transmission (96.5%), with a sporozoite rate of 3.4% and average monthly entomological inoculation rate (EIR) of 4.57 per house. Micro-geographical heterogeneity in species composition, abundance and transmission was observed across the study district in relation to key ecological differences between northern and southern clusters, with significantly higher densities, proportions and EIR of An. funestus s.l. collected from the south. An. gambiae s.l. (5.5%) density, principally An. arabiensis (81.1%) and An. gambiae s.s. (18.9%), was much lower and closely correlated with seasonal rainfall. Both An. funestus s.l. and An. gambiae s.l. were similarly resistant to alpha-cypermethrin and permethrin. Overexpression of CYP9K1, CYP6P3, CYP6P4 and CYP6M2 and high L1014F-kdr mutation frequency were detected in An. gambiae s.s. populations. Study findings highlight the urgent need for novel vector control tools to tackle persistent malaria transmission in the Lake Region of Tanzania.
Growth-rate dependency of ribosome abundance and translation elongation rate inCorynebacterium glutamicumdiffers fromEscherichia coli
Authors: Matamouros Susana , Gensch Thomas , Cerff Martin , Sachs Christian C. , Abdollahzadeh Iman , Hendriks Johnny , Horst Lucas , Tenhaef Niklas , Noack Stephan , Graf Michaela , Takors Ralf , Nöh Katharina , Bott Michael
Published: April 2021

The growth rate µ of bacteria depends on the protein synthesis capacity of the cell and thus on the number of active ribosomes and their translation elongation rate. The relationship between these fundamental growth parameters have only been described for a few bacterial species, in particularEscherichia coli, but are missing for most bacterial phyla. In this study, we systematically analysed the growth-rate dependency of ribosome abundance and translation elongation rate forCorynebacterium glutamicum, a gram-positive model species differing fromE. coliby a lower growth temperature optimum and a lower µmax. Ribosomes were quantified via single-molecule localization microscopy (SMLM) using fluorescently tagged ribosomal proteins and via RNA/protein ratio. Both methods revealed a non-linear relationship with little change in ribosome abundance below µ = 0.4 h-1and a steep increase at higher µ. UnlikeE. coli,C. glutamicumkeeps a large pool of active ribosomes at low µ, but the translation elongation rate declines from ∼9 amino acids s-1at µmaxto <2 aa s-1at µ < 0.1 h-1. A model-based approach shows that depletion of translation precursors at low growth rates can explain the observed decrease in translation elongation rate. Nutrient up-shift experiments support the hypothesis that maintenance of excess ribosomes during poor nutrient conditions enablesC. glutamicumto quickly restart growth when conditions improve.
Impact of sulfamethoxazole on a riverine microbiome
Authors: Borsetto C. , Raguideau S. , Travis E. , Kim D.W. , Lee D.H. , Bottrill A. , Stark R. , Song L. , Cha J.C. , Pearson J. , Quince C. , Singer A.C. , Wellington E.M.H.
Published: April 2021

The continued emergence of bacterial pathogens presenting antimicrobial resistance is widely recognised as a global health threat and recent attention focused on potential environmental reservoirs of antibiotic resistance genes (ARGs). Freshwater environments such as rivers represent a potential hotspot for ARGs and antibiotic resistant bacteria as they are receiving systems for effluent discharges from wastewater treatment plants (WWTPs). Effluent also contains low levels of different antimicrobials including antibiotics and biocides. Sulfonamides are antibacterial chemicals widely used in clinical, veterinary and agricultural settings and are frequently detected in sewage sludge and manure in addition to riverine ecosystems. The impact of such exposure on ARG prevalence and diversity is unknown, so the aim of this study was to investigate the release of a sub-lethal concentration of the sulfonamide compound sulfamethoxazole (SMX) on the river bacterial microbiome using a flume system. This system was a semi-naturalin vitroflume using river water (30 L) and sediment with circulation to mimic river flow. A combination of ‘omics’ approaches were conducted to study the impact of SMX exposure on the microbiomes within the flumes. Metagenomic analysis showed that the addition of low concentrations of SMX (<4 μg L−1) had a limited effect on the bacterial resistome in the water fraction only, with no impact observed in the sediment. Metaproteomics did not show differences in ARGs expression with SMX exposure in water. Overall, the river bacterial community was resilient to short term exposure to sub-lethal concentrations of SMX which mimics the exposure such communities experience downstream of WWTPs throughout the year.
Identification of lectin receptors for conserved SARS-CoV-2 glycosylation sites
Authors: Hoffmann David , Mereiter Stefan , Oh Yoo Jin , Monteil Vanessa , Zhu Rong , Canena Daniel , Hain Lisa , Laurent Elisabeth , Grünwald-Gruber Clemens , Novatchkova Maria , Ticevic Melita , Chabloz Antoine , Wirnsberger Gerald , Hagelkruys Astrid , Altmann Friedrich , Mach Lukas , Stadlmann Johannes , Oostenbrink Chris , Mirazimi Ali , Hinterdorfer Peter , Penninger Josef M.
Published: April 2021

New SARS-CoV-2 variants are continuously emerging with critical implications for therapies or vaccinations. All 22 N-glycan sites of SARS-CoV-2 Spike remain highly conserved among the variants B.1.1.7, 501Y.V2 and P.1, opening an avenue for robust therapeutic intervention. Here we used a comprehensive library of mammalian carbohydrate-binding proteins (lectins) to probe critical sugar residues on the full-length trimeric Spike and the receptor binding domain (RBD) of SARS-CoV-2. Two lectins, Clec4g and CD209c, were identified to strongly bind to Spike. Clec4g and CD209c binding to Spike was dissected and visualized in real time and at single molecule resolution using atomic force microscopy. 3D modelling showed that both lectins can bind to a glycan within the RBD-ACE2 interface and thus interferes with Spike binding to cell surfaces. Importantly, Clec4g and CD209c significantly reduced SARS-CoV-2 infections. These data report the first extensive map and 3D structural modelling of lectin-Spike interactions and uncovers candidate receptors involved in Spike binding and SARS-CoV-2 infections. The capacity of CLEC4G and mCD209c lectins to block SARS-CoV-2 viral entry holds promise for pan-variant therapeutic interventions.
Predicted Utility Modulates Working Memory Fidelity in the Brain
Authors: Levin Emily J. , Brissenden James A. , Fengler Alexander , Badre David
Published: April 2021

The predicted utility of information stored in working memory (WM) is hypothesized to influence the strategic allocation of WM resources. Prior work has shown that when information is prioritized, it is remembered with greater precision relative to other remembered items. However, these paradigms often complicate interpretation of the effects of predicted utility on item fidelity due to a concurrent memory load. Likewise, no fMRI studies have examined whether the predicted utility of an item modulates fidelity in the neural representation of items during the memory delay without a concurrent load. In the current study, we used fMRI to investigate whether predicted utility influences fidelity of WM representations in the brain. Using a generative model multivoxel analysis approach to estimate the quality of remembered representations across predicted utility conditions, we observed that items with greater predicted utility are maintained in memory with greater fidelity, even when they are the only item being maintained. Further, we found that this pattern follows a parametric relationship where more predicted utility corresponded to greater fidelity. These precision differences could not be accounted for based on a redistribution of resources among already-remembered items. Rather, we interpret these results in terms of a gating mechanism that allows for pre-allocation of resources based on predicted value alone. This evidence supports a theoretical distinction between resource allocation that occurs as a result of load and resource pre-allocation that occurs as a result of predicted utility.
Interactome analysis ofC. eleganssynapses by TurboID-based proximity labeling
Authors: Artan Murat , Barratt Stephen , Flynn Sean M. , Begum Farida , Skehel Mark , Nicolas Armel , de Bono Mario
Published: April 2021

Proximity labelling provides a powerfulin vivotool to characterize the proteome of sub-cellular structures and the interactome of specific proteins. Using the highly active biotin ligase TurboID, we optimize a proximity labelling protocol forC. elegans. We use this protocol to characterise the proteomes of the worm’s gut, muscle, skin, and nervous system. We express TurboID exclusively in the pair of AFD neurons and show we can identify known and previously unknown proteins expressed selectively in AFD. We knock TurboID into the endogenouselks-1gene, which encodes a presynaptic active zone protein. We identify many known ELKS-1 interacting proteins as well as previously uncharacterised synaptic proteins. Versatile vectors, and the inherent advantage ofC. elegansfor biochemistry, make proximity labelling a valuable addition to the nematode’s armory.

Teaser

We optimize a TurboID proximity labeling protocol for C. elegans and use it to characterize tissue and synaptic proteomes
SARS-CoV-2 binding to ACE2 triggers pericyte-mediated angiotensin-evoked cerebral capillary constriction
Authors: Hirunpattarasilp Chanawee , James Gregory , Freitas Felipe , Sethi Huma , Kittler Josef T. , Huo Jiandong , Owens Raymond J. , Attwell David
Published: April 2021

The SARS-CoV-2 receptor, ACE2, is found on pericytes, contractile cells enwrapping capillaries that regulate brain, heart and kidney blood flow. ACE2 converts vasoconstricting angiotensin II into vasodilating angiotensin-(1-7). In brain slices from hamster, which has an ACE2 sequence similar to human ACE2, angiotensin II alone evoked only a small capillary constriction, but evoked a large pericyte-mediated capillary constriction generated by AT1 receptors in the presence of the SARS-CoV-2 receptor binding domain (RBD). The effect of the RBD was mimicked by blocking ACE2. A mutated non-binding RBD did not potentiate constriction. A similar RBD-potentiated capillary constriction occurred in human cortical slices. This constriction reflects an RBD-induced decrease in the conversion of angiotensin II to angiotensin-(1-7). The clinically-used drug losartan inhibited the RBD-potentiated constriction. Thus AT1 receptor blockers could be protective in SARS-CoV-2 infection by reducing pericyte-mediated blood flow reductions in the brain, and perhaps the heart and kidney.
A high-throughput fluorescence polarization assay to discover inhibitors of arenavirus and coronavirus exoribonucleases
Authors: Hernández Sergio , Feracci Mikael , De Jesus Carolina Trajano , El-Kazzi Priscila , Kaci Rafik , Garlatti Laura , Decroly Etienne , Canard Bruno , Ferron François , Alvarez Karine
Published: April 2021

Viral exoribonucleases are uncommon in the world of RNA viruses. To date, this activity has been identified only in theArenaviridaeand theCoronaviridaefamilies. These exoribonucleases play important but different roles in both families: for mammarenaviruses the exoribonuclease is involved in the suppression of the host immune response whereas for coronaviruses, exoribonuclease is both involved in a proofreading mechanism ensuring the genetic stability of viral genomes and participating to evasion of the host innate immunity. Because of their key roles, they constitute attractive targets for drug development. Here we present a high-throughput assay using fluorescence polarization to assess the viral exoribonuclease activity and its inhibition. We validate the assay using three different viral enzymes from SARS-CoV-2, lymphocytic choriomeningitis and Machupo viruses. The method is sensitive, robust, amenable to miniaturization (384 well plates) and allowed us to validate the proof-of-concept of the assay by screening a small focused compounds library (23 metal chelators). We also determined the IC50 of one inhibitor common to the three viruses.

Highlights

ArenaviridaeandCoronaviridaeviral families share an exoribonuclease activity of common evolutionary origin

ArenaviridaeandCoronaviridaeexoribonuclease is an attractive target for drug development

We present a high-throughput assay in 384 well-plates for the screening of inhibitors using fluorescence polarization

We validated the assay by screening of a focused library of 23 metal chelators against SARS-CoV-2, Lymphocytic Choriomeningitis virus and Machupo virus exoribonucleases

We determined the IC50by fluorescence polarization of one inhibitor common to the three viruses.
Choosing between cocaine and sucrose under the influence: testing the effect of cocaine tolerance
Authors: Vandaele Y. , Ahmed S.H
Published: April 2021

Cocaine use not only depends on the reinforcing properties of the drug, but also on its intoxicating effects on alternative nondrug activities. In animal models investigating choice between cocaine and alternative sweet rewards, the latter influence can have a dramatic impact on choice outcomes. When cocaine intoxication at the moment of choice is prevented by imposing sufficiently long intervals between choice trials, animals typically prefer the sweet reward. However, when choosing under the drug influence is permitted, animals shift their preference in favor of cocaine. We previously hypothesized that this preference shift is mainly due to a direct suppression of responding for sweet reward by cocaine intoxication. Here we tested this hypothesis by making rats tolerant to this drug-induced behavioral suppression. Contrary to our expectation, tolerance did not prevent rats from shifting their preference to cocaine when choosing under the influence. Thus, other mechanisms must be invoked to explain the influence of cocaine intoxication on choice outcomes.
Native or non-native protein-protein docking models? Molecular dynamics to the rescue
Authors: Jandova Zuzana , Vargiu Attilio Vittorio , Bonvin Alexandre M. J. J.
Published: April 2021

Molecular docking excels at creating a plethora of potential models of protein-protein complexes. To correctly distinguish the favourable, native-like models from the remaining ones remains, however, a challenge. We assessed here if a protocol based on molecular dynamics (MD) simulations would allow to distinguish native from non-native models to complement scoring functions used in docking. To this end, first models for 25 protein-protein complexes were generated using HADDOCK. Next, MD simulations complemented with machine learning were used to discriminate between native and non-native complexes based on a combination of metrics reporting on the stability of the initial models. Native models showed higher stability in almost all measured properties, including the key ones used for scoring in the CAPRI competition, namely the positional root mean square deviations and fraction of native contacts from the initial docked model. A Random Forest classifier was trained, reaching 0.85 accuracy in correctly distinguishing native from non-native complexes. Reasonably modest simulation lengths in the order of 50 to 100 ns are already sufficient to reach this accuracy, which makes this approach applicable in practice.
Structural heterogeneity in biliverdin modulates spectral properties of Sandercyanin fluorescent protein
Authors: Ghosh Swagatha , Mondal Sayan , Yadav Keerti , Aggarwal Shantanu , Schaefer Wayne F. , Narayana Chandrabhas , Ramaswamy S.
Published: April 2021

Sandercyanin, a blue homo-tetrameric lipocalin protein purified from Canadian walleye (Stizostedion vitreus), is the first far-red fluorescent protein reported in vertebrates(1–3). Sandercyanin binds non-covalently to biliverdin IXα (BLA) and fluoresces at 675nm on excitation at 375nm and 635nm(1). Sandercyanin fluorescence can be harnessed for manyin vivoapplications when engineered into a stable monomeric form. Here, we report the spectral properties and crystal structures of engineered monomeric Sandercyanin-BLA complexes. Compared to wild-type protein, monomeric Sandercyanin (∼18kDa) binds BLA with similar affinities and show a broad red-shifted absorbance spectra but possess reduced quantum efficiency. Crystal structures reveal D-ring pyrrole of BLA rotated around the C14-C15 bond, which is stabilized by neighboring aromatic residues and increased water-mediated polar contacts in the BLA-binding pocket. A tetrameric Sandercyanin variant (Tyr-142-Ala) co-displaying red- and far-red absorbing states, and reduced fluorescence shows similar conformational changes in BLA binding pocket. Our results suggest that D-ring flexibility of BLA and its rearrangement reduces the fluorescence quantum-yield of monomeric Sandercyanin. Structures of monomeric Sandercyanin could be utilized as prototypes to generate bright BLA-inducible fluorescent proteins. Further, our study postulates a mechanism for modulating photo-states in BLA-bound lipocalins, known only in phytochromes till date.

Significance Statement

Sandercyanin is a tetrameric red fluorescent protein from a blue variant of walleye (Stizostedion vitreum)that binds to biliverdin IXα (BLA). Its biophysical properties and structures have been published earlier(1). A bright and stable monomeric Sandercyanin could be utilized as a fusion protein for fluorescence-based applications. Here we report the first structures and spectral properties of fluorescent monomeric Sandercyanin-BLA complexes and describe the molecular basis of modulated spectral properties due to rotated D-ring pyrrole around C14-C15 bond and re-shuffling of BLA-binding pocket. BLA-bound monomeric Sandercyanin could be engineered into brighter variants forin-vivoapplications. Our study also reveals an unfamiliar mechanism in BLA-binding lipocalins that regulates red- and far-red absorbance states.