title: The social cerebellum: A large-scale investigation of functional and structural specificity and connectivity

abstract: The cerebellum has been traditionally disregarded in relation to non-motor functions, but recent findings indicate it may be involved in language, affective processing, and social functions. Mentalizing is the ability to infer mental states of others and this skill relies on a distributed network of brain regions. Here, we leveraged large-scale multimodal neuroimaging data to elucidate the structural and functional role of the cerebellum in mentalizing. We used functional activations to determine whether the cerebellum has a domain-general or domain-specific functional role, and effective connectivity and probabilistic tractography to map the cerebello-cerebral mentalizing network. We found that the cerebellum is organized in a domain-specific way and that there is a left cerebellar effective and structural lateralization, with more and stronger effective connections from the left cerebellar hemisphere to the right cerebral mentalizing areas, and greater cerebello-thalamo-cortical (CTC) and cortico-ponto-cerebellar (CPC) streamline counts from and to the left cerebellum. Our study provides novel insights to the network organization of the cerebellum, an overlooked brain structure, and mentalizing, one of humans most essential abilities to navigate the social world.


title: Optimal Reinforcement Learning with Asymmetric Updating in Volatile Environments: a Simulation Study

abstract: AO_SCPLOWBSTRACTC_SCPLOWThe ability to predict the future is essential for decision-making and interaction with the environment to avoid punishment and gain reward. Reinforcement learning algorithms provide a normative way for interactive learning, especially in volatile environments. The optimal strategy for the classic reinforcement learning model is to increase the learning rate as volatility increases. Inspired by optimistic bias in humans, an alternative reinforcement learning model has been developed by adding a punishment learning rate to the classic reinforcement learning model. In this study, we aim to 1) compare the performance of these two models in interaction with different environments, and 2) find optimal parameters for the models. Our simulations indicate that having two different learning rates for rewards and punishments increases performance in a volatile environment. Investigation of the optimal parameters shows that in almost all environments, having a higher reward learning rate compared to the punishment learning rate is beneficial for achieving higher performance which in this case is the accumulation of more rewards. Our results suggest that to achieve high performance, we need a shorter memory window for recent rewards and a longer memory window for punishments. This is consistent with optimistic bias in human behavior.

category: neuroscience


title: Microstructural determinants of lens stiffness in rat versus guinea pig lenses

abstract: Proper ocular lens function requires lens biomechanical flexibility which is lost in presbyopia during aging. As increasing lens size has been shown previously to correlate with lens biomechanical stiffness in aging, we tested the hypothesis that whole lens size determines gross biomechanical stiffness. We used an allometric approach to evaluate this hypothesis by comparing lenses from three rodent species (mouse, rats and guinea pigs) of varying size. While rat lenses are larger and stiffer than mouse lenses, guinea pig lenses are even larger than rat lenses but are softer than the rat lens. This indicates that lens size is not a sole determinant of lens stiffness and disproves our hypothesis. Therefore, we investigated the scaling of lens microstructural features that could potentially explain the differences in biomechanical stiffness between rat and guinea pig lenses, including lens capsule thickness, epithelial cell area, fiber cell widths, suture organization, and nuclear size. Capsule thickness, epithelial cell area, and fiber cell widths scaled with lens size (i.e., greater in guinea pig lenses than rats), indicating that sizes of these features do not correlate with the stiffness of rat lenses, while suture organization was similar between rats and guinea pigs. However, we found that the hard rat lens nucleus occupies a greater fraction of the lens than the guinea pig lens nucleus, suggesting a role for nuclear size in determining whole lens stiffness. Therefore, while many features contribute to lens biomechanical properties, the size of the lens nucleus with respect to the size of the lens could be a major determinant of lens stiffness in rats versus guinea pigs.

category: cell biology


title: The Role Of Senescent Cells Removal For Cell Culture Rejuvenation

abstract: It is well known that aging reduces the number of healthy, vital cells, increases the number of dead cells, some of which are replaced by connective tissue. Similar processes occur in cell culture. If endlessly dividing transformed, or cancer cells grow in glass flasks and after their death for various reasons detach from the growth surface, lyzed and thus free up space for the growth of "young" cells, the culture lives arbitrarily long. But if in culture the "senescent" cells do not detach from the growth surface and do not make room for "young" cells, the "young" cells do not have the ability to devide and die. Because of this, the entire population of cells dies. Also in the body, "senescent" cells and intercellular tissues occupy a certain place in the body and do not allow "young", "healthy" cells to divide.


title: A quantitative study of the Golgi retention of glycosyltransferases

abstract: How Golgi glycosyltransferases and glycosidases (hereafter glycosyltransferases) localize to the Golgi is still unclear. Here, we first investigated the post-Golgi trafficking of glycosyltransferases. We found that glycosyltransferases can escape the Golgi to the plasma membrane, where they are subsequently endocytosed to the endolysosome. Post-Golgi glycosyltransferases are probably degraded by the ecto-domain shedding. We discovered that most glycosyltransferases are not retrieved from post-Golgi sites, indicating that retention but not retrieval should be the main mechanism for their Golgi localization. We proposed to use the Golgi residence time to study the Golgi retention of glycosyltransferases quantitatively and systematically. Various chimeras between ST6GAL1 and either transferrin receptor or tumor necrosis factor  quantitatively revealed the contributions of three regions of ST6GAL1, namely the N-terminal cytosolic tail, transmembrane domain and ecto-domain, to the Golgi retention. We found that each of the three regions is sufficient to produce a retention in an additive manner. The N-terminal cytosolic tail length negatively affects the Golgi retention of ST6GAL1, similar to what is known of the transmembrane domain. Therefore, long N-terminal cytosolic tail and transmembrane domain can be a Golgi export signal for transmembrane secretory cargos.


title: Anterior eye development in the brown anole, Anolis sagrei

abstract:
BackgroundAnterior eye development has been explored in different vertebrate species ranging from fish to mammals. However, missing from this diverse group is a representative of reptiles. A promising candidate to fill this void is the brown anole, Anolis sagrei, which is easily raised in the laboratory and for which genome editing techniques exist. Here we provide a detailed histological analysis of the development of the anterior structures of the eye in A. sagrei, which include the cornea, iris, ciliary body, lens, trabecular meshwork, and sclera ossicles.

ResultsDevelopment of the anterior segment in Anoles proceeds as for other vertebrates with the lens forming first followed by the cornea, then the iris, ciliary body, trabecular meshwork, and sclera ossicles. The onset of these latter structures occurs first temporally than nasally. Unlike the eyes of mammals and birds, anoles possess a remarkably thin cornea, flat ciliary body, and a trabecular meshwork that lacks an obvious Schlemms canal.

ConclusionsThis study highlights several features present in anoles and represents an important step towards understanding reptile eye development.

Key FindingsO_LIThe anole cornea epithelium is thin, composed mainly of a single basal cell layer.
C_LIO_LIThe ciliary body lacks a ciliary process.
C_LIO_LIIris and ciliary body formation occur in a spatiotemporal fashion, developing first temporally then nasally.
C_LIO_LIThe anole trabecular meshwork is composed of a spongiform tissue and lacks a Schlemms canal.
C_LI

category: developmental biology


title: On the phenology of protists: Recurrent patterns reveal seasonal variation of protistan (Rhizaria: Cercozoa, Endomyxa) communities in tree canopies

abstract: Tree canopies are colonized by billions of highly specialized microorganisms that are well adapted to the extreme microclimatic conditions, caused by diurnal fluctuations and seasonal changes. In this study we investigated seasonality patterns of protists in tree canopies of a temperate floodplain forest via high-throughput sequencing with group-specific primers for the phyla Cercozoa and Endomyxa. We observed consistent seasonality and identified divergent spring and autumn taxa. Tree crowns were characterized by a dominance of bacterivores and omnivores, while eukaryvores gained a distinctly larger share in litter and soil communities on the ground. Seasonality was largest among communities detected on the foliar surface. Higher variance within alpha diversity of foliar communities in spring indicated greater heterogeneity during community assembly. However, communities underwent distinct changes during the aging of leaves in autumn, reflecting recurring phenological changes during microbial colonization of leaves. Surprisingly, endomyxan root pathogens appeared to be exceptionally abundant across tree canopies during autumn season, demonstrating a potential role of the canopy surface as an important reservoir for wind-dispersed propagules. Overall, about 80% of detected OTUs could not be assigned to known species - representing only a fraction of dozens of microeukaryotic taxa whose canopy inhabitants are waiting to be discovered.

category: ecology


title: Identification of salt tolerant genotypes in wheat using stress tolerance indices

abstract: Salinity is one of the most important causes of yield loss in agricultural products, especially wheat. Wheat cultivation, on the other hand, is carried out on a wide range of agricultural land in saline lands. Therefore, wheat breeding for tolerance to salinity can be an effective way to improve yield and yield stability under these conditions. In this study, twenty wheat genotypes were studied in a randomized complete block design with three replications in two normal and salinity stress conditions in order to identify suitable indices of wheat stress tolerance and also to identify genotypes tolerant to salinity stress. Genotypes were evaluated based on tolerance index (TOL), stress tolerance index (STI), mean productivity (MP), susceptibility to stress (SSI), geometric mean productivity (GMP), harmonic mean productivity (HM), yield stability (YS) and yield index (YS). The results showed that MP, GMP, HM and STI indices are suitable indices because of their positive and highly significant correlation with yield in both normal and salinity stress conditions and it was found that these indices were suitable tools to achieve high yield genotypes in both environments. The results also showed that genotypes 10, 4, 9, 3 and 8 are resistant to salt stress with acceptable yield. Genotypes 5, 11, 12, 14, 16, 17, 18 and 20 were also identified as the most susceptible genotypes.

category: genetics


title: Topoisomerase VI is a chirally-selective, preferential DNA decatenase

abstract: DNA topoisomerase VI (topo VI) is a type IIB DNA topoisomerase found predominantly in archaea and some bacteria, but also in plants and algae. Since its discovery, topo VI has been proposed to be a DNA decatenase, however robust evidence and a mechanism for its preferential decatenation activity was lacking. Using single-molecule magnetic tweezers measurements and supporting ensemble biochemistry, we demonstrate that Methanosarcina mazei topo VI preferentially unlinks, or decatenates, DNA crossings, in comparison to relaxing supercoils, through a preference for certain DNA crossing geometries. In addition, topo VI demonstrates a dramatic increase in ATPase activity, DNA binding and rate of strand passage, with increasing DNA writhe, providing further evidence that topo VI is a DNA crossing sensor. Our study strongly suggests that topo VI has evolved an intrinsic preference for the unknotting and decatenation of interlinked chromosomes by sensing and preferentially unlinking DNA crossings with geometries close to 90{degrees}.

category: biochemistry


title: Quantitative susceptibility mapping of carotid arterial tissue ex vivo: assessing sensitivity to vessel microstructural composition

abstract:
PurposeTo characterise microstructural contributions to the magnetic susceptibility of carotid arteries.

MethodArterial vessels were scanned using high resolution quantitative susceptibility mapping (QSM) at 7T. Models of vessel degradation were generated using ex vivo porcine carotid arteries that were subjected to several different enzymatic digestion treatments that selectively removed microstructural components (smooth muscle cells, collagen and elastin). Magnetic susceptibilities measured in these tissue models were compared to those in untreated (native) porcine arteries. Magnetic susceptibility measured in native porcine carotid arteries was further compared to the susceptibility of cadaveric human carotid arteries to investigate their similarity.

ResultsThe magnetic susceptibility of native porcine vessels was diamagnetic ([X]native = -0.1820ppm), with higher susceptibilities in all models of vessel degradation ([X]elastin degraded = -0.0163ppm; [X]collagen degraded = -0.1158ppm; [X]decellularised = -0.1379ppm; [X]fixed native = -0.2199ppm). Magnetic susceptibility was significantly higher in collagen degraded compared to native porcine vessels (Tukey-Kramer, p<0.01) and between elastin degraded and all other models (including native, Tukey-Kramer, p<0.001). The susceptibility of fixed healthy human arterial tissue was diamagnetic and no significant difference was found between fixed human and fixed porcine arterial tissue susceptibilities (ANOVA, p>0.05).

ConclusionsMagnetic susceptibility measured using QSM is sensitive to the microstructural composition of arterial vessels - most notably to collagen. The similarity of human and porcine arterial tissue susceptibility values provides a solid basis for translational studies. As vessel microstructure becomes disrupted during the onset and progression of carotid atherosclerosis, QSM has the potential to provide a sensitive and specific marker of vessel disease.

category: bioengineering


title: Variability in scRNA-Seq analysis is mitigated by regularized or supervised approaches

abstract: Transcriptional dynamics of evolutionary processes through time are highly complex and require single-cell resolution datasets. This is especially important in cancer during the evolution of resistance, where stochasticity can lead to selection for divergent transcriptional mechanisms. Statistical methods developed to address various questions in single-cell datasets are prone to variability and require careful adjustments of multiple parameter space. To assess the impact of this variation, we utilized commonly used single-cell RNA-Seq analysis tools in a combinatorial fashion to evaluate how repeatable the results are when different methods are combined. In the context of clustering and trajectory estimation, we benchmark the combinatorial space and highlight ares and methods that are sensitive to parameter changes. We have observed that utilizing temporal information in a supervised framework or regularization in latent modeling reduces variability leading to improved overlap when different parameters/methods are used. We hope that future studies can benefit from the results presented here as use of scRNA-Seq analysis tools as out of the box is becoming a standard approach in cancer research.

category: bioinformatics


title: A new field instrument for leaf volatiles reveals an unexpected vertical profile of isoprenoid emission capacities in a tropical forest

abstract:
Both plant physiology and atmospheric chemistry are substantially altered by the emission of volatile isoprenoids (VI), such as isoprene and monoterpenes, from plant leaves. Yet, since gaining scientific attention in the 1950s, empirical research on leaf VI has been largely confined to laboratory experiments and atmospheric observations. Here, we introduce a new field instrument designed to bridge the scales from leaf to atmosphere, by enabling precision VI detection in real time from plants in their natural ecological setting. With a field campaign in the Brazilian Amazon, we reveal an unexpected distribution of leaf emission capacities (EC) across the vertical axis of the forest canopy, with EC peaking in the mid-canopy instead of the sun-exposed canopy surface, and high emissions occurring in understory specialist species. Compared to the simple interpretation that VI protect leaves from heat stress at the hot canopy surface, our results encourage a more nuanced view of the adaptive role of VI in plants. We infer that forest emissions to the atmosphere depend on the dynamic microenvironments imposed by canopy structure, and not simply on canopy surface conditions. We provide a new emissions inventory from 51 tropical tree species, revealing moderate consistency in EC within taxonomic groups. Our self-contained, portable instrument provides real-time detection and live measurement feedback with precision and detection limits better than 0.5 nmol m-2leaf s-1. We call the instrument  PORCO based on the gas detection method: photoionization of organic compounds. We provide a thorough validation of PORCO and demonstrate its capacity to detect ecologically driven variation in leaf emission rates and thus accelerate a nascent field of science: the ecology and ecophysiology of plant volatiles.

Type of paperMethod

category: plant biology


title: The hippocampus shows an own-age bias during unfamiliar face viewing

abstract: The present study investigated the neural correlates of the own-age bias for face recognition in a repetition suppression paradigm. Healthy young and older adults viewed upright and inverted unfamiliar faces. Some of the upright faces were repeated following one of two delays (lag 0 or lag 11). Repetition suppression effects were observed in bilateral fusiform cortex. However, there were no significant effects indicating an own-age bias in repetition suppression. The absence of these effects is arguably inconsistent with perceptual expertise accounts of own-age biases in face processing. By contrast, the right anterior hippocampus showed an own-age bias (greater activity for own-than other-age faces) when viewing an unfamiliar face for the first time. Given the importance of the hippocampus for episodic memory encoding, we conjecture that the increased hippocampal activity for own-age relative to other-age faces reflects differential engagement of neural processes supporting the episodic encoding of faces and might provide insight into the neural underpinnings of own-age biases in face recognition memory.


title: Towards deciphering the Nt17 code: How the sequence and conformation of the first 17 amino acids in Huntingtin regulate the aggregation, cellular properties, and neurotoxicity of mutant Httex1

abstract:
Converging evidence points to the N-terminal domain comprising the first 17 amino acids of the Huntingtin protein (Nt17) as a key regulator of its aggregation, cellular properties and toxicity. In this study, we further investigated the interplay between Nt17 and the polyQ domain repeat length in regulating the aggregation and inclusion formation of exon 1 of the Huntingtin protein (Httex1). In addition, we investigated the effect of removing Nt17 or modulating its local structure on the membrane interactions, neuronal uptake, and toxicity of monomeric or fibrillar Httex1. Our results show that the polyQ and Nt17 domains synergistically modulate the aggregation propensity of Httex1 and that the Nt17 domain plays an important role in shaping the surface properties of mutant Httex1 fibrils and regulating their poly-Q-dependent growth, lateral association and neuronal uptake. Removal of Nt17 or disruption of its transient helical conformations slowed the aggregation of monomeric Httex1 in vitro, reduced inclusion formation in cells, enhanced the neuronal uptake and nuclear accumulation of monomeric Httex1 proteins, and was sufficient to prevent cell death induced by Httex1 72Q overexpression. Finally, we demonstrate that the uptake of Httex1 fibrils into primary neurons and the resulting toxicity are strongly influenced by mutations and phosphorylation events that influence the local helical propensity of Nt17. Altogether, our results demonstrate that the Nt17 domain serves as one of the key master regulators of Htt aggregation, internalization, and toxicity and represents an attractive target for inhibiting Htt aggregate formation, inclusion formation, and neuronal toxicity.

HighlightsO_LIThe Nt17 and polyQ domains synergistically promote Httex1 aggregation.
C_LIO_LIThe Nt17 domain is a key determinant of the lateral association and morphology of fibrils.
C_LIO_LIThe Nt17 domain and conformation regulate the nuclear/cytoplasmic distribution and toxicity of Httex1.
C_LIO_LINt17 conformation is a key determinant of Httex1 fibril membrane interaction and cellular uptake.
C_LIO_LINt17 serves as one of the master regulators of Httex1 aggregation, cellular uptake and toxicity.
C_LI

Graphical abstract

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=147 SRC="FIGDIR/small/431207v2_ufig1.gif" ALT="Figure 1">
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org.highwire.dtl.DTLVardef@7afd68org.highwire.dtl.DTLVardef@92c67aorg.highwire.dtl.DTLVardef@24823dorg.highwire.dtl.DTLVardef@e2a835_HPS_FORMAT_FIGEXP  M_FIG The Nt17 domain: A master switch of Httex1 aggregation, uptake, subcellular localization and neurotoxicity.

In this paper, we showed that 1) the Nt17 and polyQ domains synergistically promote Httex1 aggregation; 2) the Nt17 domain is a key determinant of the lateral association and morphology of fibrils in vitro, 3) Nt17 conformation is a key determinant of Httex1 fibril membrane interaction and cellular uptake in primary neurons; 4) the Nt17 domain and conformation regulate the nuclear/cytoplasmic distribution and toxicity of Httex1 in primary neurons.

The figure was created with Biorender and https://www.vectorstock.com/royalty-free-vector/icon-on-and-off-toggle-switch-button-white-design-vector-30148026

C_FIG


title: The Nt17 domain and its helical conformation regulate the aggregation, cellular properties and neurotoxicity of mutant huntingtin exon 1


title: Deletion of Small Ubiquitin-like Modifier-1 Attenuates Behavioral and Anatomical Deficits by Enhancing Functional Autophagic Activities in Huntington Disease

abstract: Mutant HTT (mHTT) associated with Huntington disease (HD) affects the central nervous system by prominent atrophy in the striatum and promotes psychiatric, cognitive, and choreiform movements, although the exact mechanism remains obscure. Previous studies have shown that SUMO1 (Small Ubiquitin-like Modifier-1) modification of mHTT promotes cellular toxicity, but the in vivo role and functions of SUMO1 in HD pathogenesis are unclear. Here, we report that SUMO1 deletion in Q175DN HD-het knock-in mice (HD mice) prevented age-dependent HD-like motor and neurological impairments and suppressed the striatal atrophy and inflammatory response. SUMO1 deletion caused a drastic reduction in soluble mHtt levels and nuclear and extracellular mHtt inclusions, while increasing cytoplasmic inclusions in the striatum of HD mice. SUMO1 deletion also enhanced autophagic activity, characterized by augmented interactions between mHTT inclusions and a lysosomal marker (LAMP1), increased LC3B/LAMP1 interaction, and decreased sequestosome-1 (p62) and mHTT and diminished p62/LAMP1 interactions in DARPP-32-positive medium spiny neurons (MSNs) in HD mice. Depletion of SUMO1 in an HD cell model also diminished the mHtt levels and enhanced autophagy flux. In addition, the SUMOylation inhibitor ginkgolic acid strongly enhanced autophagy and diminished mHTT levels in human HD fibroblasts. These results indicate that SUMO is a critical therapeutic target in HD and that blocking SUMO may ameliorate HD pathogenesis by improving autophagy activities.


title: Improving range shift predictions: enhancing the power of traits

abstract: Accurately predicting species range shifts in response to environmental change is a central ecological objective and applied imperative. In synthetic analyses, traits emerge as significant but weak predictors of species range shifts across recent climate change. These studies assume linearity in the relationship between a trait and its function, while detailed empirical work often reveals unimodal relationships, thresholds, and other nonlinearities in many trait-function relationships. We hypothesize that the use of linear modeling approaches fails to capture these nonlinearities and therefore may be under-powering traits to predict range shifts. We evaluate the predictive performance of four different machine learning approaches that can capture nonlinear relationships (ridge-regularized linear regression, ridge-regularized kernel regression, support vector regression, and random forests). We validate our models using four multi-decadal range shift datasets in montane plants, montane small mammals, and marine fish. We show that nonlinear approaches perform substantially better than least-squares linear modeling in reproducing historical range shifts. In addition, using novel model observation and interrogation techniques, the trait classes (e.g. dispersal-or diet-related traits) that we identify as primary drivers of model predictions are consistent with expectations. However, disagreements among models in the directionality of trait predictors suggests limits to trait-based statistical predictive frameworks.


title: RCGAToolbox: A real-coded genetic algorithm software for parameter estimation of kinetic models

abstract:
SummaryKinetic modeling is essential in understanding the dynamic behavior of biochemical networks, such as metabolic and signal transduction pathways. However, parameter estimation remains a major bottleneck in the development of kinetic models. We present RCGAToolbox, software for real-coded genetic algorithms (RCGAs), which accelerates the parameter estimation of kinetic models. RCGAToolbox provides two RCGAs: the unimodal normal distribution crossover with minimal generation gap (UNDX/MGG) and real-coded ensemble crossover star with just generation gap (REXstar/JGG), using the stochastic ranking method. The RCGAToolbox also provides user-friendly graphical user interfaces.

Availability and implementationRCGAToolbox is available from https://github.com/kmaeda16/RCGAToolbox under GNU GPLv3, with application examples. The user guide is provided in the Supplementary Material. RCGAToolbox runs on MATLAB in Windows, Linux, and macOS.

Contactkmaeda@bio.kyutech.ac.jp

Supplementary informationSupplementary Material is available at Bioinformatics online.

category: systems biology


title: Dorsal CA1 Hippocampal Place Cells Form a Multi-Scale Representation of Megaspace

abstract: Spatially firing "place cells" within the hippocampal CA1 region form internal maps of the environment necessary for navigation and memory. In rodents, these neurons have been almost exclusively studied in small environments (<4 m2). It remains unclear how place cells encode a very large open 2D environment, which is more analogous to the natural environments experienced by rodents and other mammals. Such an ethologically realistic environment would require a more complex spatial representation, capable of simultaneously representing space at overlapping multiple fine to coarse informational scales. Here we show that in a  megaspace (18.6 m2), the majority of dorsal CA1 place cells exhibited multiple place subfields of different sizes, akin to those observed along the septo-temporal axis. Furthermore, the total area covered by the subfields of each cell was not correlated with the number of subfields, and this total area increased with the scale of the environment. The multiple different-sized subfields exhibited by place cells in the megaspace suggest that the ensemble population of subfields form a multi-scale representation of space within the dorsal hippocampus. Our findings point to a new dorsal hippocampus ensemble coding scheme that simultaneously supports navigational processes at both fine- and coarse-grained resolutions.


title: FRET-based screening in HEK293T identifies p38 MAPK and PKC inhibition as therapeutic targets for α-synuclein aggregation

abstract: Aggregation of -synuclein is associated with neurodegeneration and a hallmark pathology in synucleinopathies. These aggregates are thought to function as prion-like particles where the conformation of misfolded -synuclein determines the induced pathologys traits similar to prion diseases. Still, little is known about the molecular targets facilitating the conformation-specific biological effects, but their identification could form the basis for new therapeutic intervention. High-throughput screening (HTS) of annotated compound libraries could facilitate mechanistic investigation by identifying targets with impact on -synuclein aggregation. To this end, we developed a FRET-based cellular reporter in HEK293T cells, with sensitivity down to 6.5 nM -synuclein seeds. Using this model system, we identified GF109203X, SB202190, and SB203580 as inhibitors capable of preventing induction of - synuclein aggregation via inhibition of p38 MAPK and PKC, respectively. Our findings highlight the value HTS brings to the mechanistic investigation of -synuclein aggregation while simultaneously identifying novel therapeutic compounds.

category: molecular biology


title: Genome-wide CRISPRi screening identifies OCIAD1 as a prohibitin client and regulatory determinant of mitochondrial Complex III assembly in human cells

abstract: Dysfunction of the mitochondrial electron transport chain (mETC) is a major cause of human mitochondrial diseases. To identify determinants of mETC function, we screened a genome-wide human CRISPRi library under oxidative metabolic conditions with selective inhibition of mitochondrial Complex III and identified OCIA domain-containing protein 1 (OCIAD1) as a Complex III assembly factor. We find that OCIAD1 is an inner mitochondrial membrane protein that forms a complex with supramolecular prohibitin assemblies. Our data indicate that OCIAD1 is required for maintenance of normal steady state levels of Complex III and the proteolytic processing of the catalytic subunit cytochrome c1 (CYC1). In OCIAD1 depleted mitochondria, unprocessed CYC1 is hemylated and incorporated into Complex III. We propose that OCIAD1 acts as an adaptor within prohibitin assemblies to stabilize and/or chaperone CYC1 and to facilitate its proteolytic processing by the IMMP2L protease.


title: Pharmacological inhibition of LSD1 triggers myeloid differentiation by targeting GSE1, a novel oncogene in AML

abstract: The histone de-methylase LSD1 is over-expressed in haematological tumours and has emerged as a promising target for anti-cancer treatment, so that several LSD1 inhibitors are under development and testing, in pre-clinical and clinical settings. However, the complete understanding of their complex mechanism of action is still unreached. Here, we unravelled a novel mode of action of the LSD1 inhibitors MC2580 and DDP-38003, showing that they can induce differentiation of AML cells through the down-regulation of the chromatin protein GSE1. Analysis of the phenotypic effects of GSE1 depletion in NB4 cells showed a strong decrease of cell viability in vitro and of tumour growth in vivo. Mechanistically, we found that a set of genes associated with immune response and cytokine signalling pathways are up-regulated by LSD1 inhibitors through GSE1 protein reduction and that LSD1 and GSE1 co-localise at promoters of a subset of these genes at the basal state, enforcing their transcriptional silencing. Moreover, we show that LSD1 inhibitors lead to the reduced binding of GSE1 to these promoters, activating transcriptional programs that trigger myeloid differentiation. Our study offers new insights on GSE1 as a novel therapeutic target for AML.

category: cancer biology


title: Oxidative stress induced by tBHP in human normal colon cells by label free Raman spectroscopy and imaging.The protective role of natural antioxidants in the form of β-carotene

abstract: The present study aimed to investigate the protective effect of {beta}-carotene on the oxidative stress injury of human normal colon cell line CCD-18Co triggered by tert-Butyl hydroperoxide (tBHP). XTT examination was used to determine cell viability after {beta}-carotene supplementation and to determine the optimal concentration of antioxidant in spectroscopic studies. Cell biochemistry for CCD-18Co control group, after tBHP adding and for cells in {beta}-carotene - tBHP model was studied by using label-free Raman microspectroscopy. Results for stress treated CCD-18Co human colon normal cells and human colon cancer cells Caco-2 based on vibration features were also compared. Pretreatment with {beta}-carotene alleviated damages in CCD-18Co human normal colon cells induced by tBHP and showed the preventative effect on cells apoptosis. Treatment with {beta}-carotene altered the level of ROS investigated based on intensities of Raman peaks typical for lipids, proteins and nucleic acids. Presented study confirmed the antioxidant, protective role of {beta}-carotene against ROS by using spectroscopic label-free Raman techniques.


title: Transfer learning via multi-scale convolutional neural layers for human-virus protein-protein interaction prediction

abstract: To predict interactions between human and viral proteins, we combine evolutionary sequence profile features with a Siamese convolutional neural network (CNN) architecture and a multi-layer perceptron (MLP). Our architecture outperforms various feature encodings-based machine learning and state-of-the-art prediction methods. As our main contribution, we introduce two types of transfer learning methods (i.e.,  frozen type and  fine-tuning type) that reliably predict interactions in a target human-virus domain based on training in a source human-virus domain, by retraining CNN layers. Our transfer learning strategies can effectively apply prior knowledge transfer from large source dataset/task to small target dataset/task to improve prediction performance. Finally, we utilize the  frozen type of transfer learning to predict human-SARS-CoV-2 PPIs, indicating that our predictions are topologically and functionally similar to experimentally known interactions. Source code and datasets are available at https://github.com/XiaodiYangCAU/TransPPI/.


title: A glycan gate controls opening of the SARS-CoV-2 spike protein

abstract: SARS-CoV-2 infection is controlled by the opening of the spike protein receptor binding domain (RBD), which transitions from a glycan-shielded "down" to an exposed "up" state in order to bind the human ACE2 receptor and infect cells. While snapshots of the "up" and "down" states have been obtained by cryoEM and cryoET, details of the RBD opening transition evade experimental characterization. Here, over 130 s of weighted ensemble (WE) simulations of the fully glycosylated spike ectodomain allow us to characterize more than 300 continuous, kinetically unbiased RBD opening pathways. Together with ManifoldEM analysis of cryo-EM data and biolayer interferometry experiments, we reveal a gating role for the N-glycan at position N343, which facilitates RBD opening. Residues D405, R408, and D427 also participate. The atomic-level characterization of the glycosylated spike activation mechanism provided herein achieves a new high-water mark for ensemble pathway simulations and offers a foundation for understanding the fundamental mechanisms of SARS-CoV-2 viral entry and infection.

category: biophysics


title: Multiple time scales of adaptation allow cones to encode the inputs created by visual exploration of natural scenes

abstract: Primates explore their visual environment by making frequent saccades, discrete and ballistic eye movements that direct the fovea to specific regions of interest. Saccades produce large and rapid changes in input. The magnitude of these changes and the limited signaling range of visual neurons means that effective encoding requires rapid adaptation. Here, we explore how cone photoreceptors maintain sensitivity under these conditions. Adaptation makes cone responses to naturalistic stimuli highly nonlinear and dependent on stimulus history. Such responses cannot be explained by linear or linear-nonlinear models but are well explained by a biophysical model of phototransduction with fast and slow adaptational mechanisms. The resulting model can predict cone responses to a broad range of stimuli and enables the design of stimuli that elicit specific (e.g. linear) cone photocurrents. These advances will provide a foundation for investigating the contributions of cones and post-cone processing to visual function.


title: A chicken IgY can efficiently inhibit the entry and replication of SARS-CoV-2 by targeting the ACE2 binding domain in vitro

abstract:
COVID-19 pneumonia has now spread widely in the world. Currently, no specific antiviral drugs are available. The vaccine is the most effective way to control the epidemic. Passive immune antibodies are also an effective method to prevent and cure COVID-19 pneumonia. We used the SARS-CoV-2 S receptor-binding domain (RBD) as an antigen to immunize layers in order to extract, separate, and purify SARS-CoV-2-IgY from egg yolk. SARS-CoV-2-IgY (S-IgY)can block the entry of SARS-CoV-2 into the Cells and reduce the viral load in cells. The Half effective concentration (EC50) of W3-IgY (S-IgY in the third week after immunization) is 1.35 {+/-} 0.15nM. The EC50 of W9-IgY (S-IgY in the ninth week after immunization) is 2.76 {+/-} 1.54 nM. When the dose of S-IgY is 55 nM, the fluorescence representing intracellular viral protein is obviously weakened in Immunofluorescence microscopy.

Results of Sars-CoV-2 /Vero E6 cell experiment confirmed that S-IgY has a strong antiviral effect on SARS-CoV-2, and its (EC50) is 27.78 {+/-}1.54 nMvs 3,259 {+/-} 159.62 nM of Redesivir (differ > 106 times P<0.001).

S-IgY can inhibit the entry and replication of SARS-CoV-2, which is related to its targeting the ACE2 binding domain.

S-IgY is safe, efficient, stable, and easy to obtain. This antibody can be an effective tool for preventing and treating COVID-19 pneumonia.

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The figure briefly illustrates that the preparation and extraction of S-IgY and its anti-S-CoV-2 mechanism is to inhibit the entry and replication of SARS-CoV-2 by targeting the ACE2.

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category: immunology


title: Characterization of humoral and SARS-CoV-2 specific T cell responses in people living with HIV

abstract:
There is an urgent need to understand the nature of immune responses generated against SARS-CoV-2, to better inform risk-mitigation strategies for people living with HIV (PLWH). Although not all PLWH are considered immunosuppressed, residual cellular immune deficiency and ongoing inflammation could influence COVID-19 disease severity, the evolution and durability of protective memory responses. Here, we performed an integrated analysis, characterizing the nature, breadth and magnitude of SARS-CoV-2-specific immune responses in PLWH, controlled on ART, and HIV negative subjects. Both groups were in the convalescent phase of predominately mild COVID-19 disease. The majority of PLWH mounted SARS-CoV-2 Spike- and Nucleoprotein-specific antibodies with neutralizing activity and SARS-CoV-2-specific T cell responses, as measured by ELISpot, at levels comparable to HIV negative subjects. T cell responses against Spike, Membrane and Nucleocapsid were the most prominent, with SARS-CoV-2-specific CD4 T cells outnumbering CD8 T cells. Notably, the overall magnitude of SARS-CoV-2-specific T cell responses related to the size of the naive CD4 T cell pool and the CD4:CD8 ratio in PLWH, in whom disparate antibody and T cell responses were observed. Both humoral and cellular responses to SARS-CoV-2 were detected at 5-7 months post-infection, providing evidence of medium-term durability of responses irrespective of HIV serostatus. Incomplete immune reconstitution on ART and a low CD4:CD8 ratio could, however, hamper the development of immunity to SARS-CoV-2 and serve as a useful tool for risk stratification of PLWH. These findings have implications for the individual management and potential effectiveness of vaccination against SARS-CoV-2 in PLWH.

One Sentence SummaryAdaptive immune responses to SARS-CoV-2 in the setting of HIV infection


title: Favourable antibody responses to human coronaviruses in children and adolescents with autoimmune rheumatic diseases

abstract: Differences in humoral immunity to coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), between children and adults remain unexplained and the impact of underlying immune dysfunction or suppression unknown. Here, we examined the antibody immune competence of children and adolescents with prevalent inflammatory rheumatic diseases, juvenile idiopathic arthritis (JIA), juvenile dermatomyositis (JDM) and juvenile systemic lupus erythematosus (JSLE), against the seasonal human coronavirus (HCoV)-OC43 that frequently infects this age group. Despite immune dysfunction and immunosuppressive treatment, JIA, JDM and JSLE patients mounted comparable or stronger responses than healthier peers, dominated by IgG antibodies to HCoV-OC43 spike, and harboured IgG antibodies that cross-reacted with SARS-CoV-2 spike. In contrast, responses to HCoV-OC43 and SARS-CoV-2 nucleoproteins exhibited delayed age-dependent class-switching and were not elevated in JIA, JDM and JSLE patients, arguing against increased exposure. Consequently, autoimmune rheumatic diseases and their treatment were associated with a favourable ratio of spike to nucleoprotein antibodies.


title: Metabolic constraints on nitrogen fixation by rhizobia in legume nodules

abstract: Rhizobia induce nodule formation on legume roots and differentiate into bacteroids, which use plant-derived dicarboxylates as energy and electron sources for reduction of atmospheric N2 into ammonia for secretion to plants. Using heterogeneous genome-scale datasets, we reconstructed a model of bacteroid metabolism to investigate the effects of varying dicarboxylate and oxygen supply on carbon and nitrogen allocation. Modelling and 13C metabolic flux analysis in bacteroids indicate that microaerobiosis restricts the decarboxylating arm of the TCA cycle and limits ammonia assimilation into glutamate. Catabolism of dicarboxylates induces a higher oxygen demand but also a higher NADH/NAD+ ratio compared to sugars. Carbon polymer synthesis and alanine secretion by bacteroids facilitate redox balance in microaerobic nodules with alanine secretion increasing as oxygen tension decreases. Our results provide a framework for understanding fundamental constraints on rhizobial metabolism during symbiotic nitrogen fixation.

category: microbiology


title: Seasonal Dynamics in the Number and Composition of Coliform Bacteria in Drinking Water Reservoirs

abstract:
Worldwide, surface waters like lakes and reservoirs are one of the major sources for drinking water production, especially in regions with water scarcity. In the last decades, they have undergone significant changes due to climate change. This includes not only an increase of the water temperature but also microbiological changes. In recent years, increased numbers of coliform bacteria have been observed in these surface waters. In our monitoring study we analyzed two drinking water reservoirs (Klingenberg and Kleine Kinzig Reservoir) over a two-year period in 2018 and 2019. We detected high numbers of coliform bacteria up to 2.4 x 104 bacteria per 100 ml during summer months, representing an increase of four orders of magnitude compared to winter. Diversity decreased to one or two species that dominated the entire water body, namely Enterobacter asburiae and Lelliottia spp., depending on the reservoir. Interestingly, the same, very closely related strains have been found in several reservoirs from different regions. Fecal indicator bacteria Escherichia coli and enterococci could only be detected in low concentrations. Furthermore, fecal marker genes were not detected in the reservoir, indicating that high concentrations of coliform bacteria were not due to fecal contamination. Microbial community revealed Frankiales and Burkholderiales as dominant orders. Enterobacterales, however, only had a frequency of 0.04% within the microbial community, which is not significantly affected by the extreme change in coliform bacteria number. Redundancy analysis revealed water temperature, oxygen as well as nutrients and metals (phosphate, manganese) as factors affecting the dominant species. We conclude that this sudden increase of coliform bacteria is an autochthonic process that can be considered as a mass proliferation or "coliform bloom" within the reservoir. It is correlated to higher water temperatures in summer and is therefore expected to occur more frequently in the near future, challenging drinking water production.

HighlightsO_LIColiform bacteria proliferate in drinking water reservoirs to values above 104 per 100 ml
C_LIO_LIThe genera Lelliottia and Enterobacter can form these "coliform blooms"
C_LIO_LIMass proliferation is an autochthonic process, not related to fecal contaminations
C_LIO_LIIt is related to water temperature and appears mainly in summer
C_LIO_LIIt is expected to occur more often in future due to climate change
C_LI

Graphical abstract

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=75 SRC="FIGDIR/small/428560v1_ufig1.gif" ALT="Figure 1">
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title: Organic electron donors and terminal electron acceptors structure anaerobic microbial communities and interactions in a permanently stratified sulfidic lake

abstract: The extent to which nutrients structure microbial communities in permanently stratified lakes is not well understood. This study characterized microbial communities from the anoxic layers of the meromictic and sulfidic Fayetteville Green Lake (FGL), NY, and investigated the roles of organic electron donors and terminal electron acceptors in shaping microbial community structure and interactions. Bacterial communities from the permanently stratified layer below the chemocline (monimolimnion) and from enrichment cultures inoculated by lake sediments were analyzed using 16S rRNA gene sequencing. Results showed that anoxygenic phototrophs dominated microbial communities in the upper monimolimnion (21 m), which harbored little diversity, whereas the most diverse communities resided at the bottom of the lake (~52 m). Organic electron donors explained 54% of the variation in the microbial community structure in aphotic cultures enriched on an array of organic electron donors and different inorganic electron acceptors. Electron acceptors only explained 10% of the variation, but were stronger drivers of community assembly in enrichment cultures supplemented with acetate or butyrate compared to the cultures amended by chitin, lignin or cellulose. We identified a range of habitat generalists and habitat specialists in both the water column and enrichment samples using Levins index. Network analyses of interactions among microbial groups revealed Chlorobi and sulfate reducers as central to microbial interactions in the upper monimolimnion, while Syntrophaceae and other fermenting organisms were more important in the lower monimolimnion. The presence of photosynthetic microbes and communities that degrade chitin and cellulose much below the chemocline supported the downward transport of microbes, organic matter and oxidants from the surface and the chemocline. Collectively, our data suggest niche partitioning of bacterial communities by interactions that depend on the availability of different organic electron donors and terminal electron acceptors. Thus, light, as well as the diversity and availability of chemical resources drive community structure and function in FGL, and likely in other stratified, meromictic lakes.


title: Live attenuated SARS-CoV-2 vaccine candidate: Protective immunity without serious lung lesions in Syrian hamsters

abstract:
Various COVID-19 vaccine candidates are currently under clinical trial. However, no live attenuated vaccine has been developed yet, despite their generally high efficacy. Here, we established temperature-sensitive mutant strains of SARS-CoV-2, whose growth was significantly slower than that of the parent strain at 37{degrees}C. One of the strains, A50-18, which presented mutations in nonstructural protein 14, did not replicate at all at 37{degrees}C in vitro. In vivo experiments demonstrated that this strain replicated inefficiently in the lungs of Syrian hamsters, and intra-nasal inoculation induced sufficient anti-SARS-CoV-2-neutralizing antibodies to protect against wild type virus infection. These results suggest that the A50-18 strain could be a promising live attenuated vaccine candidate against SARS-CoV-2.

One Sentence SummaryA live attenuated virus provided immunity against SARS-CoV-2 in an animal model, making it a promising vaccine candidate.


title: Global detection of DNA repair outcomes induced by CRISPR-Cas9

abstract: CRISPR-Cas9 generates double-stranded DNA breaks (DSBs) to activate cellular DNA repair pathways for genome editing. The repair of DSBs leads to small insertions or deletions (indels) and other complex byproducts, including large deletions and chromosomal translocations. Indels are well understood to disrupt target genes, while the other deleterious byproducts remain elusive. We developed a new in silico analysis pipeline for the previously described primer-extension-mediated sequencing assay to comprehensively characterize CRISPR-Cas9-induced DSB repair outcomes in human or mouse cells. We identified tremendous deleterious DSB repair byproducts of CRISPR-Cas9 editing, including large deletions, plasmid integrations, and chromosomal translocations. We further elucidated the important roles of microhomology, chromosomal interaction, recurrent DSBs, and DSB repair pathways in the generation of these byproducts. Our findings provide an extra dimension for genome editing safety besides off-targets. And caution should be exercised to avoid not only off-target damages but also deleterious DSB repair byproducts during genome editing.

category: genomics


title: 501Y.V2 and 501Y.V3 variants of SARS-CoV-2 lose binding to Bamlanivimab in vitro

abstract: We generated several versions of the receptor binding domain (RBD) of the Spike protein with mutations existing within newly emerging variants from South Africa and Brazil. We found that the mutant RBD with K417N, E484K, and N501Y exchanges has higher binding affinity to the human receptor compared to the wildtype RBD. This mutated version of RBD also completely abolishes the binding to a therapeutic antibody, Bamlanivimab, in vitro.


title: Identification of common key genes and pathways between Covid-19 and lung cancer by using protein-protein interaction network analysis

abstract: COVID-19 is indeed an infection that is caused by a recently found coronavirus group, a type of virus proven to cause human respiratory diseases. The high mortality rate was observed in patients who had pre-existing health conditions like cancer. However, the molecular mechanism of SARS-CoV-2 infection in lung cancer patients was not discovered yet at the pathway level. This study was about determining the common key genes of COVID-19 and lung cancer through network analysis. The hub genes associated with COVID-19 and lung cancer were identified through Protein-Protein interaction analysis. The hub genes are ALB, CXCL8, FGF2, IL6, INS, MMP2, MMP9, PTGS2, STAT3 and VEGFA. Through gene enrichment, it is identified both COVID-19 and lung cancer have a common pathway in EGFR tyrosine kinase inhibitor resistance, IL-17 signalling pathway, AGE-RAGE signalling pathway in diabetic complications, HIF-1 signalling pathway and pathways in cancer.


title: Correlation between structure and function in phosphatidylinositol lipid-dependent Kir2.2 gating

abstract: Inward rectifier K+(Kir) channels regulate cell membrane potential. Different Kir channels respond to unique ligands, but all are regulated by phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). Using planar lipid bilayers we show that Kir2.2 exhibits bursts of openings separated by long quiescent inter-burst periods. Increasing PI(4,5)P2 concentration shortens the Kir2.2 inter-burst duration and lengthens the burst duration without affecting dwell times within a burst. From this, we propose that burst and inter-burst durations correspond to the CTD-docked and CTD-undocked conformations observed in the presence and absence of PI(4,5)P2 in atomic structures. We also studied the effect of different phosphatidylinositol lipids on Kir2.2 activation and conclude that the 5 phosphate is essential to Kir2.2 pore opening. Other phosphatidylinositol lipids can compete with PI(4,5)P2 but cannot activate Kir2.2 without the 5 phosphate. PI(4)P, which is directly interconvertible to and from PI(4,5)P2, might thus be a regulator of Kir channels in the plasma membrane.


title: Pharmacologic Therapy for Engraftment Arrhythmia Induced by Transplantation of Human Cardiomyocytes

abstract:
BackgroundEngraftment arrhythmias (EAs) are observed in large animal studies of intramyocardial transplantation of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) for myocardial infarction. Although transient, the risk posed by EA presents a barrier to clinical translation.

ObjectivesWe hypothesized that clinically approved antiarrhythmic drugs can prevent EA-related mortality as well as suppress tachycardia and arrhythmia burden.

MethodshPSC-CM were transplanted into the infarcted porcine heart by surgical or percutaneous delivery to induce EA. Following a screen of antiarrhythmic agents, a prospective study was conducted to determine the effectiveness of amiodarone plus ivabradine in preventing cardiac death and suppressing EA.

ResultsEA was observed in all subjects, and amiodarone-ivabradine treatment was well-tolerated. None of the treated subjects experienced the primary endpoint of cardiac death, unstable EA or heart failure compared to 5/8 (62.5%) in the control cohort (hazard ratio 0.00; 95% confidence interval, 0-0.297; p = 0.002). Overall survival including two deaths in the treated cohort from immunosuppression-related infection showed borderline improvement with treatment (hazard ratio 0.21; 95% confidence interval, 0.03-1.01; p = 0.05). Without treatment, peak heart rate averaged 305 {+/-} 29 beats per min (bpm), whereas in treated subjects peak daily heart rate was significantly restricted to 185{+/-}9 bpm (p = 0.006). Similarly, treatment reduced peak daily EA burden from 96.8 {+/-} 2.9% to 76.5 {+/-} 7.9% (p = 0.003). Antiarrhythmic treatment was safely discontinued after approximately one-month of treatment without recrudescence of arrhythmia.

ConclusionsThe risk of engraftment arrhythmia following hPSC-CM transplantation can be reduced significantly by combined amiodarone and ivabradine drug therapy.

Condensed AbstractEngraftment arrhythmia (EA) is a transient but serious complication of cardiac remuscularization therapy. Using a porcine model of cardiac remuscularization and EA, ivabradine and amiodarone were independently effective in suppressing tachycardia and arrhythmia, respectively. Baseline amiodarone combined with adjunctive ivabradine successfully prevented cardiac death, unstable EA and heart failure (hazard ratio 0.00; 95% confidence interval, 0-0.297; p = 0.002) and significantly suppressed peak daily heart rate and arrhythmia burden (p=0.006 and 0.003, respectively). Antiarrhythmic treatment was successfully discontinued after one-month without recrudescence of arrhythmia. We conclude that EA can be suppressed by combined amiodarone and ivabradine drug therapy.

category: pharmacology and toxicology


title: Gender and geographical disparity in editorial boards of journals in psychology and neuroscience

abstract: While certain metrics of diversity have seen great improvement in recent years in academic psychology and neuroscience, unequal representation remains for many positions of power. Here, we reviewed publicly available information in order to infer the proportion of editors by gender and their country of affiliation in the top 50 journals worldwide in each of the two fields. The sample included a total of 2,864 editors for psychology journals and 3,093 editors for neuroscience journals. There was a statistically significant difference in the proportion of male and female editors in both fields, both across editorial roles, and within various role categories, including editor-in-chief and their deputies at neuroscience journals, associate and section editors in both fields, and editorial and advisory board members in both fields. The only category in which there was not a significant imbalance of male and female scholars was the editors-in-chief of psychology journals and their deputies. Geographically, USA-based academics significantly outnumbered those from other countries as editors in both psychology and neuroscience. Results also indicated that over three quarters of psychology journals (76%) were comprised of more than 50% male editors, while only 20% had a similar proportion of female editors. In neuroscience, 88% of journals were comprised of more than 50% male editors, while only 10% of journals included a similar, proportional majority of female editors. Findings suggest that editorial positions in academic journals, possibly one of the most powerful decision-making roles in academic psychology and neuroscience, are not balanced in gender or geographical representation.

category: scientific communication and education


title: A compressed representation of spatial distance in the rodent hippocampus

abstract:
Principal cells in the rodent hippocampus often fire in response to traversal through a specific spatial location (place cells), as well as elapsed time during an imposed temporal delay or after stimulus offset (time cells). Sequences of time cells unfold rapidly at first, with many time cells with narrow time fields. As the triggering event recedes into the past, time cells are fewer and have broader fields. This means that the representation of time in the hippocampus is compressed with greater resolution for time points near the present. Using tetrode recordings we measured individual CA1 units while rats traveled along a track that could be changed in length. Consistent with previous results, most place cells coded for distance from the starting point of the trajectory. Critically, place cells became less numerous and showed gradually widening fields with distance from the starting location. These results suggest that as the animal leaves a landmark, the hippocampal place code forms a compressed representation of distance from the starting location. The representation of time and space in the hippocampus have similar properties suggesting that they arise from similar computational mechanisms.

Significance StatementThe hippocampus represents relationships between events in time and space. It has been hypothesized that temporal and spatial relationships are the result of a common computational mechanism. Previous work has shown that the representation of time in the hippocampus is compressed, with less neural resolution for more temporally remote events, consistent with the observation that temporal memory is worse for events further in the past. This paper shows an analogous result for spatial relationships. Place cells coded for distance from the start of a journey. As distance increased, place fields became broader and less numerous, showing a decrease in spatial resolution. This result suggests a unified coding scheme for the dimensions of time and space in the rodent hippocampus.

category: neuroscience


title: Nicotine-mediated recruitment of GABAergic neurons to a dopaminergic phenotype attenuates motor deficits in alpha-synuclein Parkinson's model.

abstract:
BACKGROUNDPrevious work revealed an inverse correlation between smoking and Parkinsons disease (PD) that is associated with nicotine-induced neuroprotection of dopaminergic (DA) neurons against nigrostriatal damage in PD primates and rodent models. Nicotine, a neuroactive component of tobacco, can directly alter the activity of midbrain DA neurons and induce non-DA neurons in the substantia nigra (SN) to acquire a DA phenotype. We investigated the recruitment mechanism of nigrostriatal GABAergic neurons to express DA phenotypes, such as transcription factor Nurr1 and DA-synthesizing enzyme tyrosine hydroxylase (TH), and the concomitant effects on motor function.

METHODSWild-type and -syn-overexpressing (PD) mice treated with chronic nicotine were assessed by behavioral pattern monitor (BPM) and immunohistochemistry/in-situ hybridization to measure behavior and the translational/transcriptional regulation of neurotransmitter phenotype following selective Nurr1 overexpression or DREADD-mediated chemogenetic activation.

RESULTSNicotine treatment led to a transcriptional TH and translational Nurr1 upregulation within a pool of SN GABAergic neurons in wild-type animals. In PD mice, nicotine increased Nurr1 expression, reduced the number of -syn-expressing neurons, and simultaneously rescued motor deficits. Hyperactivation of GABA neurons alone was sufficient to elicit de novo translational upregulation of Nurr1 in non-DA neurons. Retrograde labeling revealed that a fraction of these GABAergic neurons projects to the dorsal striatum.

CONCLUSIONSNicotine exposure initiates neuroprotective mechanisms counteracting the neurodegenerative effects of -syn accumulation in DA neurons and contributing to Nurr1-mediated therapeutic effects. Revealing the mechanism of nicotine-induced DA plasticity protecting SN neurons against nigrostriatal damage could contribute to developing new strategies for neurotransmitter replacement in PD.


title: Dynamin is primed at endocytic sites for ultrafast endocytosis

abstract: Dynamin mediates fission of vesicles from the plasma membrane during endocytosis. Typically, dynamin is recruited from the cytosol to endocytic sites, requiring seconds to tens of seconds. However, ultrafast endocytosis in neurons internalizes vesicles as quickly as 50 ms during synaptic vesicle recycling. Here we demonstrate that Dynamin 1 is pre-recruited to endocytic sites for ultrafast endocytosis. Specifically, Dynamin 1xA, a splice variant of Dynamin 1, interacts with Syndapin 1 to form molecular condensates on the plasma membrane when the proline-rich domain of this variant is dephosphorylated. When this domain is mutated to include phosphomimetic residues or Syndapin 1s dynamin-interacting domain is mutated, Dynamin 1xA becomes diffuse, and consequently, ultrafast endocytosis slows down by [~]100-fold. Mechanistically, Syndapin 1 acts as an adaptor by binding the plasma membrane and stores Dynamin 1xA at endocytic sites. This cache bypasses the recruitment step and accelerates endocytosis at synapses.


title: Development of chemosynthetic microbial communities in organic falls deployed in the deep Southwest Atlantic Ocean

abstract: The assembly and successional processes of microbial communities inhabiting deep-sea whale and wood falls are highly complex and vastly unknown, as a myriad of factors may affect the development of a chemosynthetic-based ecosystem on these organic islands. The chemoautotrophy supported by organic substrates is the basis of long-lasting ecosystems, considered biodiversity hotspots in the oligotrophic deep sea. Understanding how these microbial communities develop and the factors affecting them could shed light on processes related to the maintenance of biodiversity in this environment. We performed a whale- and wood-fall experiment in the southwest Atlantic on the Brazilian continental margin and investigated biofilm-forming bacterial and archaeal communities colonising these substrates, deployed at 1500 and 3300 m depth. The composition of the prokaryotic communities shared some similarities with previously reported organic falls in the north Pacific and the Mediterranean Sea, mainly regarding sulphur oxidising chemolithotrophic taxa from the phyla Campylobacterota and Proteobacteria. Communities were found to be highly different between the organic substrates, as whale fall associated biofilms presented a higher dominance of sulphur oxidising chemolithotrophs. We also observed a significant difference between the two sites, with the whale associated communities at the 1500 isobath presenting a faster establishment of the chemosynthetic taxa.


title: Characterization of heliorhodopsins detected via functional metagenomics in freshwater Actinobacteria, Chloroflexi and Archaea

abstract:
Rhodopsins are widespread in microbes residing in diverse aquatic environments across the globe. Recently, a new unusual rhodopsin family, the heliorhodopsins (HeRs), was discovered, distributed among diverse bacteria, archaea, eukarya and even viruses. Here, using functional metagenomics on samples from Lake HaHula and Ein Afek reserve, we found and characterized ten HeRs representing divergent members of the family. The expressed HeRs absorb light in the green and yellow wavelengths and originate from Actinobacteria, Chloroflexi and Archaea. The photocycle of the HeR from Chloroflexi revealed a low accumulation of the M-intermediate that we connect to the lack of two conserved histidine residues in transmembrane helices 1 and 2 in this protein. Another of HeR, from Actinobacteria, exhibited an unusually fast photocycle (166 ms, 5 times faster than HeR-48C12). To further explore the still unresolved question of the HeR function, we performed an analysis of protein families among genes neighboring HeRs, in our clones and thousands of other microbes. This analysis revealed a putative connection between HeRs and genes involved in oxidative stress. At the same time, very few protein families were found to distinguish genes surrounding prokaryotic HeRs from those surrounding rhodopsin pumps. The strongest association was found with the DegV family involved in activation of fatty acids and uncharacterized family DUF2177, which allowed us to hypothesize that HeRs are involved in membrane lipid remodeling. This work further establishes functional metagenomics as a simple and fruitful method of screening for new rhodopsins.

SignificanceThe recently discovered divergent rhodopsin family of heliorhodopsins is abundant in freshwater environments. In this study, we sampled a habitat rich in dissolved organic matter to increase our chances of finding spectrally shifted rhodopsins. Using functional metagenomics, diverse heliorhodopsins absorbing green and yellow light were discovered. The metagenomic clones originated from diverse prokaryotic groups: Actinobacteria, Chloroflexi and even Archaea, emphasizing the versatility of the E. coli expression system used. Photocycles of representative heliorhodopsins were measured and exhibited diverse kinetic characteristics. Analysis of genes neighboring heliorhodopsins in diverse prokaryotes revealed their putative connection to membrane lipid re-modeling and oxidative stress. Our findings suggest that functional metagenomics is a productive method for the discovery of new and diverse rhodopsins.


title: Pilot study: Trehalose-induced remodelling of the human microbiota affects Clostridioides difficile infection outcome in an in vitro colonic model

abstract: Within the human intestinal tract, dietary, microbial- and host-derived compounds are used as signals by many pathogenic organisms, including Clostridioides difficile. Trehalose has been reported to enhance virulence of certain C. difficile ribotypes; however, such variants are widespread and not correlated with clinical outcomes for patients suffering from C. difficile infection (CDI). Here, we make preliminary observations to how to trehalose supplementation affects the microbiota in an in vitro model and show that trehalose can reduce the outgrowth of C. difficile, preventing simulated CDI. Three clinically reflective human gut models simulated the effects of sugar (trehalose or glucose) or saline ingestion on the microbiota. Models were instilled with sugar or saline and further exposed to C. difficile spores. The recovery of the microbiota following antibiotic treatment and CDI induction was monitored in each model. The human microbiota remodeled to utilise the bioavailable trehalose. Clindamycin induction caused simulated CDI in models supplemented with either glucose or saline; however, trehalose supplementation did not result in CDI, although limited spore germination did occur. The absence of CDI in trehalose model was associated with enhanced abundances of Finegoldia, Faecalibacterium and Oscillospira, and reduced abundances of Klebsiella and Clostridium spp., compared with the other models. Functional analysis of the microbiota in the trehalose model revealed differences in the metabolic pathways, such as amino acid metabolism, which could be attributed to prevention of CDI. Our data show that trehalose supplementation remodelled the microbiota, which prevented simulated CDI, potentially due to enhanced recovery of nutritionally competitive microbiota against C. difficile.


title: Nanoscale resolution of microbial fiber degradation in action

abstract:
Deconstruction of plant cell walls is imperative to global carbon cycling and sustainability efforts. Selected microbes degrade plant fibers using extremely efficient multi-enzymatic cellulosomes assemblies. Organization of cellulosomes on the bacterial cell surface and their ecological regulation remain elusive. By combining structural methodologies with molecular and biochemical approaches on the canonical Clostridium thermocellum system, we provide an unprecedented view into the in-situ structure and distribution of cellulosomal enzymes while interacting with their cellulosic substrate during fiber degradation. Structural exploration of growing cultures revealed isogenic phenotypic heterogeneity of cellulosome organization on single cells across the bacterial population, suggesting a division-of labor strategy driven by product-dependent dynamics. This study demonstrates how structural biology under near-physiological conditions can be employed to develop ecological hypotheses to understand microbial plant-fiber degradation at the single-cell nanoscale level.

One Sentence SummaryThis study contributes critical insights into the in-situ organization of cellulosomes and their cellulosic substrates and provides evidence for phenotypic heterogeneity, with dynamic, growth phase-dependent organization of the fiber-degrading machinery.


title: Monoclonal antibodies targeting the influenza virus N6 neuraminidase

abstract:
Influenza A viruses are a diverse species that include 16 hemagglutinin (HA) subtypes and 9 neuraminidase (NA) subtypes. While the antigenicity of many HA subtypes is reasonably well studied, less is known about NA antigenicity, especially when it comes to non-human subtypes that only circulate in animal reservoirs. The N6 NA subtypes are mostly found in viruses infecting birds. However, they have also been identified in viruses that infect mammals, such as swine and seals. More recently, highly pathogenic H5N6 subtype viruses have caused rare infections and mortality in humans. Here, we generated murine mAbs to the N6 NA, characterized their breadth and antiviral properties in vitro and in vivo and mapped their epitopes by generating escape mutant viruses. We found that the antibodies had broad reactivity across the American and Eurasian N6 lineages, but relatively little binding and inhibition of the H5N6 NA. Several of the antibodies exhibited strong NA inhibition activity and some also showed activity in the antibody dependent cellular cytotoxicity reporter assay and neutralization assay. In addition, we generated escape mutant viruses for six monoclonal antibodies and found mutations on the lateral ridge of the NA. Lastly, we observed variable protection in H4N6 and H5N6 mouse challenge models when the antibodies were given prophylactically.

ImportanceThe N6 NA has recently gained prominence due to the emergence of highly pathogenic H5N6 viruses. Currently, there is limited characterization of the antigenicity of avian N6 neuraminidase. Our data is an important first step towards a better understanding of the N6 NA antigenicity.


title: Rapid active-reflection display on dorsal stripes of Atherinomorus lacunosus

abstract: A large number of living creatures are able to use environmental light effectively as a biological display. The biological structural colors are very attractive not only within the coloring species but also to humans. However, the detailed function of bio-reflectors, which constitute the structural color with respect to communication, remains unknown. Atherinomorus lacunosus has alignments of iridophore spots on its dorsal part. Here it is found that a spot with a diameter of approximately 0.1 mm causes a rhythmic blinking of light owing to rapid reflection changes in iridophores existing inside the spot. The iridophores contain reflecting particles which show similar rotational responses to magnetic field under a light exposure. The speed of the intensity change of light at a frequency of approximately 1 Hz is proposed to be controlled by the nervous system of A. lacunosus. This kind of passive illumination may contribute to the development of a new optical device with low energy consumption.


title: Differential contributions of M1 and pre-M1 to ion selectivity in ASICs and ENaCs

abstract: The ability to discriminate between different ionic species, termed ion selectivity, is a key feature of ion channels and forms the basis for their physiological function. Members of the degenerin/epithelial sodium channel (DEG/ENaC) superfamily of trimeric ion channels are typically sodium selective, but to a surprisingly variable degree. While acid-sensing ion channels (ASICs) are weakly sodium selective (sodium:potassium around 10:1), ENaCs show a remarkably high preference for sodium over potassium (>500:1). The most obvious explanation for this discrepancy may be expected to originate from differences in the pore-lining second transmembrane segment (M2). However, these show a relatively high degree of sequence conservation between ASICs and ENaCs and previous functional and structural studies could not unequivocally establish that differences in M2 alone can account for the disparate degrees of ion selectivity. By contrast, surprisingly little is known about the contributions of the first transmembrane segment (M1) and the preceding pre-M1 region. In this study, we use conventional and non-canonical amino acid-based mutagenesis in combination with a variety of electrophysiological approaches to show that the pre-M1 and M1 regions of mASIC1a channels are major determinants of ion selectivity. Mutational investigations of the corresponding regions in hENaC show that they contribute less to ion selectivity, despite affecting ion conductance. In conclusion, our work supports the notion that the remarkably different degrees of sodium selectivity in ASICs and ENaCs are achieved through different mechanisms. The results further highlight how M1 and pre-M1 are likely to differentially affect pore structure in these related channels.


title: The M1 and pre-M1 segments contribute differently to ion selectivity in ASICs and ENaCs


title: Detection of a biolistic delivery of fluorescent markers and CRISPR/Cas9 to the pollen tube

abstract: In recent years, genome-editing techniques, such as the CRISPR/Cas9 system, have been highlighted as a new approach to plant breeding. Agrobacterium-mediated transformation has been widely utilized to generate transgenic plants by introducing plasmid DNA containing CRISPR/Cas9 into plant cells. However, this method is generally applicable to a limited range of plants, such as model species. To overcome this limitation, we developed a method to genetically modify male germ cells without the need for Agrobacterium-mediated transfection and tissue culture, by using tobacco as a model. In this study, plasmid DNA containing sequences of Cas9, guide RNA, and fluorescent reporter was introduced into pollen using a biolistic delivery system. Based on the transient expression of fluorescent reporters, the Arabidopsis UBQ10 promoter was found to be the most suitable for driving expression of the delivered gene in pollen tubes. We also evaluated delivery efficiency in male germ cells in the pollen by expression of the introduced fluorescent marker. Mutations were detected in the target gene in the genomic DNA extracted from CRISPR/Cas9 introduced pollen tubes but were not detected in the negative control. Bombarded pollen germinated pollen tubes on the stigma and produced two sperm cells within the pistil. We also observed ovules showing fluorescence derived from bombarded pollen. The findings of this study provide important insights into the editing of pollen tube genomes and the delivery of genome-modified male germ cells for seed production.


title: Hepatitis C Virus E1 Protein Enhances Macrophage iNOS Expression In Vitro

abstract:
ObjectiveHepatitis C virus (HCV) is a single-stranded RNA virus that causes chronic hepatitis, cirrhosis, and liver cancer. Approximately 170 million individuals are infected with HCV worldwide. The pathogenesis of HCV-associated liver injury is thought to be due to the host antiviral immune response, including the T cell response, and excessive production of proinflammatory cytokines, reactive oxygen species, and nitric oxide (NO).

Interferon-{gamma} (IFN-{gamma}) is a key cytokine in the adaptive immune response that is primarily secreted from CD4+ T helper cells to induce cytotoxic T lymphocyte (CTL) cell response against HCV infection. Another important role of IFN-{gamma} is the activation of macrophages in the liver resulting in inhibition of viral replication and increased NO production.

Enhanced inducible nitric oxide synthase (iNOS) expression and NO production observed in the liver of HCV-infected patients is positively correlated with viral load and hepatic inflammation. HCV-infected macrophages are major producers of NO in the liver. It is not completely understood how HCV proteins affect iNOS expression and what the role of IFN-{gamma} is in HCV protein expression in HCV-infected macrophages. In this study, we examined the effect of INF-{gamma} and HCV proteins on iNOS expression in the Raw264.7 cell line.

ResultsConsistent with other studies, HCV core and NS5A proteins induced iNOS expression in macrophages. Moreover, HCV E1 protein-enhanced iNOS expression is highest in the presence and absence of IFN-{gamma} activation.

ConclusionThese results indicate that hepatitis C virus core, NS5A, E1 protein regulates iNOS protein expression in IFN-{gamma}-activated and resting macrophage cell lines. These findings points to a future research direction for understanding the pathogenesis of HCV-related liver inflammation.


title: Single-Domain SARS-CoV-2 S1 and RBD Antibodies Isolated from Immunized Llama Effectively Bind Targets of the Wuhan, UK, and South African Strains in vitro

abstract: The spreading of SARS-CoV-2 variants has become a major challenge of the current fight against the pandemic. Of particular concerns are the strains that have arisen from the United Kingdom (UK) and South Africa. The UK variant spreads rapidly and is projected to overtake the original strain in the US as early as in March 2021, while the South African variant appears to evade some effects of the current vaccines. Potential false-negative diagnosis using currently available antigen kits that may not recognize these variants could cause another wave of community infection. Therefore, it is imperative that antibodies used in the detection kits are validated for binding against these variants. Here we report that the nanoantibodies (nAbs in our terminology, also referred to as VHH fragments, single domain antibodies, nanobodies) that we have developed for rapid antigen detection test bind the receptor binding domain (RBD) of the S1 protein from the original COVID-SARS-2 virus as well as those from the UK and South African variants. This finding validates our antibodies used in our assay for the detection of these major variant strains.


title: Bacteria elicit a phage tolerance response subsequent to infection of their neighbors

abstract: Plaque occurrence on a bacterial lawn manifests successive rounds of bacteriophage infection. Yet, mechanisms evolved by bacteria to limit plaque spread have been hardly explored. Here we investigated the dynamics of plaque development by lytic phages infecting the bacterium Bacillus subtilis. We report that plaque expansion is followed by a constriction phase owing to bacterial growth into the plaque zone. This phenomenon is caused by an adaptive process, herein termed "phage tolerance response", elicited by non-infected bacteria located at the plaque rim upon sensing infection of their neighbors. The temporary phage-tolerance is executed by the stress response RNA polymerase sigma factor {sigma}X, primarily through activation of the dlt operon, encoding enzymes that catalyze D-alanylation of cell wall teichoic acid polymers, the major attachment sites for phages infecting Gram-positive bacteria. D-alanylation impedes phage binding and hence infection, thus enabling the uninfected bacteria to form a protective shield opposing plaque spread.


title: The Host Interactome of Spike Expands the Tropism of SARS-CoV-2

abstract: The SARS-CoV-2 virus causes severe acute respiratory syndrome (COVID-19) and has rapidly created a global pandemic. Patients that survive may face a slow recovery with long lasting side effects that can afflict different organs. SARS-CoV-2 primarily infects epithelial airway cells that express the host entry receptor Angiotensin Converting Enzyme 2 (ACE2) which binds to spike protein trimers on the surface of SARS-CoV-2 virions. However, SARS-CoV-2 can spread to other tissues even though they are negative for ACE2. To gain insight into the molecular constituents that might influence SARS-CoV-2 tropism, we determined which additional host factors engage with the viral spike protein in disease-relevant human bronchial epithelial cells (16HBEo-). We found that spike recruited the extracellular proteins laminin and thrombospondin and was retained in the endoplasmatic reticulum (ER) by the proteins DJB11 and FBX2 which support re-folding or degradation of nascent proteins in the ER. Because emerging mutations of the spike protein potentially impact the virus tropism, we compared the interactome of D614 spike with that of the rapidly spreading G614 mutated spike. More D614 than G614 spike associated with the proteins UGGT1, calnexin, HSP7A and GRP78/BiP which ensure glycosylation and folding of proteins in the ER. In contrast to G614 spike, D614 spike was endoproteolytically cleaved, and the N-terminal S1 domain was degraded in the ER even though C-terminal  S2 only proteoforms remained present. D614 spike also bound more laminin than G614 spike, which suggested that extracellular laminins may function as co-factor for an alternative,  S2 only dependent virus entry. Because the host interactome determines whether an infection is productive, we developed a novel proteome-based cell type set enrichment analysis (pCtSEA). With pCtSEA we determined that the host interactome of the spike protein may extend the tropism of SARS-CoV-2 beyond mucous epithelia to several different cell types, including macrophages and epithelial cells in the nephron. An  S2 only dependent, alternative infection of additional cell types with SARS-CoV-2 may impact vaccination strategies and may provide a molecular explanation for a severe or prolonged progression of disease in select COVID-19 patients.


title: N-Terminal finger stabilizes the reversible feline drug GC376 in SARS-CoV-2 Mpro

abstract: The main protease (Mpro, also known as 3CL protease) of SARS-CoV-2 is a high priority drug target in the development of antivirals to combat COVID-19 infections. A feline coronavirus antiviral drug, GC376, has been shown to be effective in inhibiting the SARS-CoV-2 main protease and live virus growth. As this drug moves into clinical trials, further characterization of GC376 with the main protease of coronaviruses is required to gain insight into the drugs properties, such as reversibility and broad specificity. Reversibility is an important factor for therapeutic proteolytic inhibitors to prevent toxicity due to off-target effects. Here we demonstrate that GC376 has nanomolar Ki values with the Mpro from both SARS-CoV-2 and SARS-CoV strains. Restoring enzymatic activity after inhibition by GC376 demonstrates reversible binding with both proteases. In addition, the stability and thermodynamic parameters of both proteases were studied to shed light on physical chemical properties of these viral enzymes, revealing higher stability for SARS-CoV-2 Mpro. The comparison of a new X-ray crystal structure of Mpro from SARS-CoV complexed with GC376 reveals similar molecular mechanism of inhibition compared to SARS-CoV-2 Mpro, and gives insight into the broad specificity properties of this drug. In both structures, we observe domain swapping of the N-termini in the dimer of the Mpro, which facilitates coordination of the drugs P1 position. These results validate that GC376 is a drug with an off-rate suitable for clinical trials.


title: The neural G protein Gαo tagged with GFP at an internal loop is functional in C. elegans

abstract: Go is the alpha subunit of the major heterotrimeric G protein in neurons and mediates signaling by every known neurotransmitter, yet the signaling mechanisms activated by Go remain to be fully elucidated. Genetic analysis in Caenorhabditis elegans has shown that Go signaling inhibits neuronal activity and neurotransmitter release, but studies of the molecular mechanisms underlying these effects have been limited by lack of tools to complement genetic studies with other experimental approaches. Here we demonstrate that inserting the green fluorescent protein (GFP) into an internal loop of the Go protein results in a tagged protein that is functional in vivo and that facilitates cell biological and biochemical studies of Go. Transgenic expression of Go-GFP rescues the defects caused by loss of endogenous Go in assays of egg laying and locomotion behaviors. Defects in body morphology caused by loss of Go are also rescued by Go-GFP. The Go-GFP protein is localized to the plasma membrane of neurons, mimicking localization of endogenous Go. Using GFP as an epitope tag, Go-GFP can be immunoprecipitated from C. elegans lysates to purify Go protein complexes. The Go-GFP transgene reported in this study enables studies involving in vivo localization and biochemical purification of Go to complement the already well-developed genetic analysis of Go signaling.


title: Diversity of excitatory release sites

abstract: The molecular mechanisms underlying the diversity of cortical glutamatergic synapses is still only partially understood. Here, we tested the hypothesis that presynaptic active zones (AZs) are constructed from molecularly uniform, independent release sites (RSs), the number of which scales linearly with the AZ size. Paired recordings between hippocampal CA1 pyramidal cells and fast-spiking interneurons followed by quantal analysis demonstrate large variability in the number of RSs (N) at these connections. High resolution molecular analysis of functionally characterized synapses reveals highly variable Munc13-1 content of AZs that possess the same N. Replica immunolabeling also shows a 3-fold variability in the Munc13-1 content of AZs of identical size. Munc13-1 is clustered within the AZs; cluster size and density are also variable. Our results provide evidence for quantitative molecular heterogeneity of RSs and support a model in which the AZ is built up from variable numbers of molecularly heterogeneous, but independent RSs.


title: Functional basis for calmodulation of the TRPV5 calcium channel

abstract: Within the transient receptor potential (TRP) superfamily of ion channels, TRPV5 is a highly Ca2+-selective channel important for active reabsorption of Ca2+ in the kidney. Its channel activity is controlled by a negative feedback mechanism involving calmodulin (CaM) binding. Combining advanced microscopy techniques and biochemical assays, this study characterized the dynamic bilobal CaM regulation and binding stoichiometry. We demonstrate for the first time that functional (full-length) TRPV5 interacts with CaM in the absence of Ca2+, and this interaction is intensified at increasing Ca2+ concentrations sensed by the CaM C-lobe that achieves channel pore blocking. Channel inactivation occurs without CaM N-lobe calcification. Moreover, we reveal a 1:2 stoichiometry of TRPV5:CaM binding by implementing single molecule photobleaching counting, a technique with great potential for studying TRP channel regulation. In conclusion, our study proposes a new model for CaM- dependent regulation - calmodulation - of the Ca2+-selective TRPV5 that involves apoCaM interaction and lobe-specific actions.

category: cell biology


title: Dynamics of hunchback translation in real time and at single mRNA resolution in the Drosophila embryo

abstract: The Hunchback (Hb) transcription factor is critical for anterior-posterior patterning of the Drosophila embryo. Despite the maternal hb mRNA acting as a paradigm for translational regulation, due to its repression in the posterior of the embryo, little is known about the translatability of zygotically transcribed hb mRNAs. Here we adapt the SunTag system, developed for imaging translation at single mRNA resolution in tissue culture cells, to the Drosophila embryo to study the translation dynamics of zygotic hb mRNAs. Using singlemolecule imaging in fixed and live embryos, we provide evidence for translational repression of zygotic SunTag-hb mRNAs. While the proportion of SunTag-hb mRNAs translated is initially uniform, translation declines from the anterior over time until it becomes restricted to a posterior band in the expression domain. We discuss how regulated hb mRNA translation may help establish the sharp Hb expression boundary, which is a model for precision and noise during developmental patterning. Overall, our data show how use of the SunTag method on fixed and live embryos is a powerful combination for elucidating spatiotemporal regulation of mRNA translation in Drosophila.

category: developmental biology


title: Endothelial cells elicit a pro-inflammatory response to SARS-COV-2 without productive viral infection

abstract:
ObjectivesThrombotic and microvascular complications are frequently seen in deceased COVID-19 patients. However, whether this is caused by direct viral infection of the endothelium or inflammation-induced endothelial activation remains highly contentious.

MethodsHere, we use patient autopsy samples, primary human endothelial cells and an in vitro model of the pulmonary epithelial-endothelial cell barrier to show that primary human endothelial cells express very low levels the SARS-CoV-2 receptor ACE2 and the protease TMPRSS2.

ResultsAccordingly, endothelial cells can only be infected when SARS-CoV-2 is present at very high concentrations. However, this is not a productive infection (i.e. no infectious virus is produced) and viral entry induces an inflammatory response. We also show that SARS-CoV-2 does not infect endothelial cells in 3D vessels under flow conditions. We further demonstrate that in a co-culture model endothelial cells are not infected with SARS-CoV-2. They do however sense and respond to infection in the adjacent epithelial cells, increasing ICAM-1 expression and releasing pro-inflammatory cytokines.

ConclusionsTaken together, these data suggest that in vivo, endothelial cells are unlikely to be infected with SARS-CoV-2 and that infection is only likely to occur if the adjacent pulmonary epithelium is denuded (basolateral infection) or a high viral load is present in the blood (apical infection). In such a scenario, whilst SARS-CoV-2 infection of the endothelium can occur, it does not contribute to viral amplification. However, endothelial cells are still likely to play a key role in SARS-CoV-2 pathogenesis by sensing adjacent infection and mounting a pro-inflammatory response to SARS-CoV-2.


title: High-resolution genetic mapping reveals cis-regulatory and copy number variation in loci associated with cytochrome P450-mediated detoxification in a generalist arthropod pest

abstract:
Chemical control strategies are driving the evolution of pesticide resistance in pest populations. Understanding the genetic mechanisms of these evolutionary processes is of crucial importance to develop sustainable resistance management strategies. The acaricide pyflubumide is one of the most recently developed mitochondrial complex II inhibitors with a new mode of action that specifically targets spider mite pests. In this study, we characterize the molecular basis of pyflubumide resistance in a highly resistant population of the spider mite Tetranychus urticae. Classical genetic crosses indicated that pyflubumide resistance was incompletely recessive and controlled by more than one gene. To identify resistance loci, we crossed the resistant population to a highly susceptible T. urticae inbred strain and propagated resulting populations with and without pyflubumide exposure for multiple generations in an experimental evolution set-up. High-resolution genetic mapping by a bulked segregant analysis approach led to the identification of three quantitative trait loci (QTL) linked to pyflubumide resistance. Two QTLs were found on the first chromosome and centered on the cytochrome P450 CYP392A16 and a cluster of CYP392E6-8 genes. Comparative transcriptomics revealed a consistent overexpression of CYP392A16 and CYP392E8 in the experimental populations that were selected for pyflubumide resistance. We further corroborated the involvement of CYP392A16 in resistance by in vitro functional expression and metabolism studies. Collectively, these experiments uncovered that CYP392A16 N-demethylates the toxic carboxamide form of pyflubumide to a non-toxic compound. A third QTL coincided with cytochrome P450 reductase (CPR), a vital component of cytochrome P450 metabolism. We show here that the resistant population harbors three gene copies of CPR and that this copy number variation is associated with higher mRNA abundance. Together, we provide evidence for detoxification of pyflubumide by cytochrome P450s that is likely synergized by gene amplification of CPR.

Author summaryOur understanding of the causal genetic variants that drive the evolution of quantitative traits, such as polygenic pesticide resistance, remains very limited. Here, we followed a high-resolution genetic mapping approach to localize the genetic variants that cause pyflubumide resistance in the two-spotted spider mite Tetranychus urticae. Three well-supported QTL were uncovered and pointed towards a major role for cytochrome P450-mediated detoxification. Cis-regulatory variation for cytochrome P450s was observed, and in vitro cytochrome P450 experiments showed that pyflubumide was metabolized into a non-toxic derivate. A third QTL centered on cytochrome P450 reductase (CPR), which is required for cytochrome P450 activity, and is amplified in pyflubumide resistant populations. Our results indicate that pyflubumide resistance is mediated by cytochrome P450 detoxification that is enhanced by gene amplification at the CPR locus.

Data availabilitySequence data has been deposited at the Sequence Read Archive (PRJNA596790). Phenotypic quantification data is available at Supplementary Data S1.

category: genetics


title: Supervised biomedical semantic similarity

abstract:
BackgroundSemantic similarity between concepts in knowledge graphs is essential for several bioinformatics applications, including the prediction of protein-protein interactions and the discovery of associations between diseases and genes. Although knowledge graphs describe entities in terms of several perspectives (or semantic aspects), state-of-the-art semantic similarity measures are general-purpose. This can represent a challenge since different use cases for the application of semantic similarity may need different similarity perspectives and ultimately depend on expert knowledge for manual fine-tuning.

ResultsWe present a new approach that uses supervised machine learning to tailor aspect-oriented semantic similarity measures to fit a particular view on biological similarity or relatedness. We implement and evaluate it using different combinations of representative semantic similarity measures and machine learning methods with four biological similarity views: protein-protein interaction, protein function similarity, protein sequence similarity and phenotype-based gene similarity.

ConclusionsThe results demonstrate that our approach outperforms non-supervised methods, producing semantic similarity models that fit different biological perspectives significantly better than the commonly used manual combinations of semantic aspects.