title: Broadly neutralizing antibodies to SARS-related viruses can be readily induced in rhesus macaques

abstract:
To prepare for future coronavirus (CoV) pandemics, it is desirable to generate vaccines capable of eliciting neutralizing antibody responses against multiple CoVs. Because of the phylogenetic similarity to humans, rhesus macaques are an animal model of choice for many virus-challenge and vaccine-evaluation studies, including SARS-CoV-2. Here, we show that immunization of macaques with SARS-CoV-2 spike (S) protein generates potent receptor binding domain cross- neutralizing antibody (nAb) responses to both SARS-CoV-2 and SARS-CoV-1, in contrast to human infection or vaccination where responses are typically SARS-CoV-2-specific. Furthermore, the macaque nAbs are equally effective against SARS-CoV-2 variants of concern. Structural studies show that different immunodominant sites are targeted by the two primate species. Human antibodies generally target epitopes strongly overlapping the ACE2 receptor binding site (RBS), whereas the macaque antibodies recognize a relatively conserved region proximal to the RBS that represents another potential pan-SARS-related virus site rarely targeted by human antibodies. B cell repertoire differences between the two primates appear to significantly influence the vaccine response and suggest care in the use of rhesus macaques in evaluation of vaccines to SARS-related viruses intended for human use.

ONE SENTENCE SUMMARYBroadly neutralizing antibodies to an unappreciated site of conservation in the RBD in SARS- related viruses can be readily induced in rhesus macaques because of distinct properties of the naive macaque B cell repertoire that suggest prudence in the use of the macaque model in SARS vaccine evaluation and design.


title: Antisense-mediated repression of SAGA-dependent genes involves the HIR histone chaperone

abstract: Eukaryotic genomes are pervasively transcribed by RNA polymerase II (RNAPII), and transcription of long non-coding RNAs often overlaps with coding gene promoters. This might lead to coding gene repression in a process named Transcription Interference (TI). In Saccharomyces cerevisiae (S. cerevisiae), TI is mainly driven by antisense non-coding transcription and occurs through re-shaping of promoter Nucleosome-Depleted Regions (NDRs). In this study, we developed a genetic screen to identify new players involved in Antisense-Mediated Transcription Interference (AMTI). Among the candidates, we found the HIR histone chaperone complex known to be involved in de novo histone deposition. Using genome-wide approaches, we reveal that HIR-dependent histone deposition represses the promoters of SAGA-dependent genes via antisense non-coding transcription. However, while antisense transcription is enriched at promoters of SAGA-dependent genes, this feature is not sufficient to define the mode of gene regulation. We further show that the balance between HIR-dependent nucleosome incorporation and transcription factor binding at promoters directs transcription into a SAGA- or TFIID-dependent regulation. This study sheds light on a new connection between antisense non-coding transcription and the nature of coding transcription initiation.

category: molecular biology


title: A local insulin reservoir ensures developmental progression in condition of nutrient shortage in Drosophila

abstract:
Insulin/IGF signalling (IIS) controls many aspects of development and physiology. In Drosophila, a conserved family of insulin-like peptides (Ilp) is produced by brain neurosecretory cells and exerts systemic functions. Here, we describe the local uptake and storage of Ilps in the Corpora Cardiaca (CC), a group of alpha cell homolog that produces the glucagon-like hormone AKH. Dilp uptake relies on the expression of Impl2, an IGF-BP that accumulates in the CCs. During nutrient shortage, this specific reserve of Ilps is released and activates IIS in a paracrine manner in the prothoracic gland, securing accelerated entry into pupal development through the production of the steroid hormone ecdysone. We therefore uncover a sparing mechanism whereby local Ilp storage and release activates the production of steroids and ensures early developmental progression in adverse food conditions.

Highlights- Dilps are uptaken by CC cells through the IGF-BP Imp-L2
- the CC-Dilp store is released upon nutrient shortage and activates IIS through CC projections on the PG
- upon nutrient shortage, IIS activation in the PG ensures an accelerated transition from larval feeding stage to metamorphosis.

category: developmental biology


title: Variation in mechanical and structural properties of enamel in primate molars

abstract: Mechanical properties of enamel are known to vary across molar crowns in some primates, but the association of this variation with phylogeny, structural properties and tribological behaviour is not well understood. In this study, 20 molars from a range of primate taxa (n=15) were studied using nanoindentation, micro-CT scanning, and SEM imaging. After micro-CT scanning, teeth were sectioned in the lingual-buccal plane through the mesial cusps. Five positions (buccal lateral, buccal cuspal, occlusal middle, lingual cuspal, lingual lateral) were studied in three locations (inner, middle, outer enamel regions). The results show middle enamel had the highest hardness and elastic modulus values in all positions.  Non-functional molar sides (lingual in lower molars and buccal in upper molars) had higher hardness values than their  functional counterparts. Increase in prism size was associated with a decrease in hardness in some tooth positions, and mineral density showed a significant relationship with elastic modulus values. Variation in enamel structure variation (e.g., enamel Schmelzmuster, Hunter-Schreger band thickness), may also be crucial in explaining variation in mechanical properties, with decussation zones associated with higher mechanical properties values. Primate enamel is not a homogeneous material, with variation in mechanical and structural properties across the crown likely associated with functional differences and variation in force distribution. Overall structural and mechanical patterns were similar in the primate species studied despite substantial differences in diet, suggesting these properties are potentially evolutionary conserved.

category: evolutionary biology


title: All-trans retinoic acid induces synaptopodin-dependent metaplasticity in mouse dentate granule cells

abstract: The vitamin A derivative all-trans retinoic acid (atRA) is a key mediator of synaptic plasticity. Depending on the brain region studied, distinct effects of atRA on excitatory and inhibitory neurotransmission have been reported. However, it remains unclear how atRA mediates brain region-specific effects on synaptic transmission and plasticity. Here, we used intraperitoneal injections of atRA (10 mg/kg) in adult male C57BL/6J mice to study the effects of atRA on excitatory and inhibitory neurotransmission in the mouse fascia dentata. In contrast to what has been reported in other brain regions, no major changes in synaptic transmission were observed in the ventral and dorsal hippocampus 6 hours after atRA administration. Likewise, no evidence for changes in the intrinsic properties of hippocampal dentate granule cells was obtained in the atRA-treated group. Moreover, hippocampal transcriptome analysis revealed no essential changes upon atRA treatment. In light of these findings, we tested for the metaplastic effects of atRA, i.e., for its ability to modulate synaptic plasticity expression in the absence of major changes in baseline synaptic transmission. Indeed, in vivo long-term potentiation (LTP) experiments demonstrated that systemic atRA treatment improves the ability of dentate granule cells to express LTP. The plasticity-promoting effects of atRA were not observed in synaptopodin-deficient mice, thus extending our previous results on the relevance of synaptopodin in atRA-mediated synaptic strengthening in the mouse prefrontal cortex. Taken together, our data show that atRA mediates synaptopodin-dependent metaplasticity in mouse dentate granule cells.

category: neuroscience


title: Pupil size dynamics predict dLGN firing mode over a wide range of timescales

abstract: The way in which sensory systems encode information, even at early processing stages, is modulated according to the internal state of the animal. Internal states, such as arousal, are often characterized by relating neural variables to a single "level" of arousal, defined using a behavioral variable such as locomotion or pupil size. Here, we extend the notion of arousal-related modulation by showing that multifaceted aspects of pupil size dynamics can predict changes in neural activity. Specifically, we show that the phases of pupil size fluctuations occurring over multiple timescales modulate firing mode in the dorsal lateral geniculate nucleus (dLGN) of mice. Increased firing rates, driven by a preponderance of tonic spiking, occurred during pupil dilations, while burst spikes preferentially occurred during contractions. We further find that these activity modulations could not be explained solely by pupil size per se, or by transitions between overt locomotion and quiescence, and extend to periods of patterned stimulus viewing. We conclude that dLGN spiking activity is modulated by pupil-indexed arousal processes on various timescales, with implications for the determination of arousal-related cortical rhythms, and for the transfer of sensory information to the cortex.


title: Modulation of dLGN firing mode across multiple timescales is predicted by pupil size dynamics


title: Adenylate Kinase 2 deficiency causes NAD+ depletion and impaired purine metabolism during myelopoiesis

abstract:
Reticular Dysgenesis is a particularly grave form of severe combined immunodeficiency that affects the adaptive and innate immune system. Patients suffer from congenital neutropenia, lymphopenia, and deafness. The disease is caused by biallelic loss of function in mitochondrial Adenylate Kinase 2 (AK2). AK2 mediates the phosphorylation of AMP to ADP, as substrate for ATP synthesis. Accordingly, declining oxidative phosphorylation has been postulated as the driver of disease pathology. The mechanistic basis, however, remains incompletely understood. Single cell RNA-sequencing of patient bone marrow cells implicated altered RNA catabolism and ribonucleoprotein synthesis in the pathogenesis of Reticular Dysgenesis. To investigate these findings, we developed a disease model based on CRISPR-mediated disruption of the AK2 gene in primary human hematopoietic stem cells. We found that AK2-deficient myeloid progenitor cells not only have compromised mitochondrial energy metabolism and increased AMP levels, but also NAD+ and aspartate depletion, metabolites that rely on TCA-cycle activity for regeneration and synthesis. Furthermore, AK2-deficient cells exhibited strikingly increased levels of the purine nucleotide precursor IMP, decreased cellular RNA content, ribosome subunit expression, protein synthesis and a profoundly hypo-proliferative phenotype. The rise in IMP levels stemmed from increased AMP deamination. Pharmacologic inhibition of AMP deaminase normalized IMP levels in AK2-deficient cells, but further aggravated the disease phenotype, pointing to AMP catabolism as a possible metabolic adaptation to mitigate AMP-mediated toxicity. Inducing an adenosine disequilibrium in control cells produced a similar myeloid maturation defect.

This study shows that AK2 deficiency globally curtailed mitochondrial metabolism resulting in NAD+ and aspartate deficiency and disordered purine metabolism. AMP accumulation and its detrimental effects on ribonucleotide synthesis capacity may contribute to the failure of myelopoiesis in Reticular Dysgenesis.

category: cell biology


title: Reticular Dysgenesis-associated Adenylate Kinase 2 deficiency causes failure of myelopoiesis through disordered purine metabolism


title: Genome sequences of the five Sitopsis species of Aegilops and the origin of polyploid wheat B-subgenome

abstract:
Bread wheat (Triticum aestivum L., BBAADD) is a major staple food crop worldwide. The diploid progenitors of the A- and D-subgenomes have been unequivocally identified, that of B however remains ambiguous and controversial but is suspected to be related to species of Aegilops, section Sitopsis. Here, we report the assembly of chromosome-level genome sequences of all five Sitopsis species, namely Ae. bicornis, Ae. longissima, Ae. searsii, Ae. sharonensis, and Ae. speltoides, as well as partial assembly of Ae. mutica genome for phylogenetic analysis. Our results support that the donor of bread wheat B-subgenome is a distinct, probably extinct, diploid species that diverged from an ancestral progenitor of the B-lineage similar to Ae. mutica and Ae. speltoides. The five Sitopsis species have variable genome sizes (4.11-5.89 Gb) with high proportions of repetitive sequences (85.99-89.81%); nonetheless, they retain high collinearity with other wheat genomes. Differences in genome size are primarily due to independent post-speciation amplification of transposons rather than to inter-specific genetic introgression. We also identified a set of Sitopsis genes pertinent to important agronomic traits that can be harnessed for wheat breeding. These resources provide a new roadmap for evolutionary and genetic studies of the wheat group.

SignificanceThe origin of the B-subgenome of hexaploid bread wheat remains unknown. Here we report the assembly of chromosome-level genome sequences of all five Sitopsis species of the genus Aegilops, which are previously considered as possible direct progenitors or contributors to the B-subgenome. Our comparative genomic analyses reveal that the B-subgenome originated from an unknown, most likely extinct species phylogenetically distinct from Ae. speltoides, its extant closest relative. We also provide evidence that Ae. speltoides is neither the direct progenitor of the G-subgenome of tetraploid wheat Triticum timopheevii. The high-quality Sitopsis genomes provide novel avenues to identify new important genes for wheat breeding.


title: A Distributed Network for Multimodal Experiential Representation of Concepts

abstract:
The architecture of the cortical system underlying concept representation is a topic of intense debate. Much evidence supports the claim that concept retrieval selectively engages sensory, motor, and other neural systems involved in the acquisition of the retrieved concept, yet there is also strong evidence for involvement of high-level, supramodal cortical regions. A fundamental question about the organization of this system is whether modality-specific information originating from sensory and motor areas is integrated across multiple "convergence zones" or in a single centralized "hub". We used representational similarity analysis (RSA) of fMRI data to map brain regions where the similarity structure of neural patterns elicited by large sets of concepts matched the similarity structure predicted by a high-dimensional model of concept representation based on sensory, motor, affective, and other modal aspects of experience. Across two studies involving different sets of concepts, different participants, and different tasks, searchlight RSA revealed a distributed, bihemispheric network engaged in multimodal experiential representation, composed of high-level association cortex in anterior, lateral, and ventral temporal lobe; inferior parietal lobule; posterior cingulate gyrus and precuneus; and medial, dorsal, ventrolateral, and orbital prefrontal cortex. These regions closely resemble networks previously implicated in general semantic and "default mode" processing and are known to be high-level hubs for convergence of multimodal processing streams. Supplemented by an exploratory cluster analysis, these results indicate that the concept representation system consists of multiple, hierarchically organized convergence zones supporting multimodal integration of experiential information.

Significance StatementIt has long been known that information about visual, auditory, motor, affective, and other features of our phenomenal experience originate in distinct brain regions. However, it is still unclear how these processing streams converge to form multimodal concept representations. Using fMRI together with a multimodal experiential model of conceptual content, we show in two large studies that concept knowledge is represented across a distributed, bihemispheric network including temporal, parietal, limbic, and prefrontal association cortices. These results argue against the idea of a single centralized "hub" for concept representation, suggesting instead that multiple high-level convergence zones encode conceptual information in terms of multimodal experiential content.


title: Individualized prediction of future cognition using baseline developmental changes in cortical anatomy

abstract: Predictive modeling studies have started to reveal brain measures underlying cognition; however, most studies are based on cross-sectional data (static  final brain measures acquired at one time point). Since brain development comprises of continuously ongoing events leading to cognitive development, predictive modeling studies need to consider  dynamic as opposed to static  final brain measures. Using longitudinal neuroimaging and cognitive data (global executive composite score, an index of executive function) from 82 individuals (aged 5-14 years, scanned 3 times), we built highly accurate prediction models (r=0.61, p=1.6e-09) of future cognition (assessed at visit 3) based on baseline developmental changes in cortical anatomy (from visit 1 to 2). More importantly, dynamic brain measures (change in cortical anatomy from visit 1 to 2) and not static brain measures (cortical anatomy at visit 1 and 2) were critical for predicting future cognition, suggesting the need for considering dynamic brain measures in predicting cognitive outcomes.


title: Inhibitory control of synaptic signals preceding motor action in mouse frontal cortex

abstract:
Preparatory activity in the frontal cortex preceding movement onset is thought to represent a neuronal signature of motor planning. However, how excitatory and inhibitory synaptic inputs to frontal neurons are integrated during movement preparation remains unclear. Here we address this question by performing in vivo whole-cell patch-clamp recordings in the secondary motor cortex (MOs) of head-fixed mice moving on a treadmill. We find that both superficial and deep principal neurons show slowly increasing (~10 s) membrane potential and spike rate ramps preceding the onset of spontaneous, self-paced running periods. By contrast, in animals trained to perform a goal-directed task, both membrane potential and spike ramps are characterized by larger amplitudes and accelerated kinetics during preparation of goal-driven movement. To determine the role of local inhibitory neurons in shaping these task-dependent preparatory signals, we chemogenetically suppressed the activity of specific interneuron subpopulations in untrained animals. Inactivation of parvalbumin-positive (PV+) interneurons leads to depolarized membrane potential ramps with increased amplitudes during preparation of movement, while inactivation of somatostatin-positive (SOM+) interneurons abolishes membrane potential ramps. A computational model of the local MOs circuit shows that SOM+-mediated inhibition of PV+ interneurons in conjunction with recurrent connectivity among the principal neurons can reproduce slow ramping signals, while plasticity of excitatory synapses on SOM+ interneurons can explain the acceleration of these signals in trained animals. Together, our data reveal that local inhibitory neurons play distinct roles in controlling task-dependent preparatory ramping signals when MOs neurons integrate external inputs during motor planning.

HighlightsO_LIPrincipal neurons in MOs show slow preparatory membrane potential and firing rate ramps preceding the onset of spontaneous, self-paced running periods.
C_LIO_LIIn animals trained to perform a goal-directed task, both membrane potential and spike ramps are faster and larger in amplitude.
C_LIO_LIInactivation of PV+ interneurons disinhibits MOs principal neurons and increases the amplitude of membrane potential ramps, while inactivation of SOM+ interneurons abolishes membrane potential ramps.
C_LIO_LIOur modeling results suggest that the concerted action of external inputs and local inactivation shapes task-dependent preparatory motor signals in MOs neurons.
C_LI


title: Discrete Roles of the Ir76b Ionotropic Co-Receptor Impact Olfaction, Blood Feeding, and Mating in the Malaria Vector Mosquito Anopheles coluzzii

abstract:
Anopheline mosquitoes rely on their highly sensitive chemosensory apparatus to detect diverse chemical stimuli that drive the host-seeking and blood-feeding behaviors required to vector pathogens for malaria and other diseases. This process incorporates a variety of chemosensory receptors and transduction pathways. We have used advanced in vivo gene-editing and -labelling approaches to localize and functionally characterize the ionotropic co-receptor AcIr76b in the malaria mosquito Anopheles coluzzii, where it impacts both olfactory and gustatory systems. AcIr76b has a broad expression pattern in female adult antennal grooved pegs, T1 and T2 sensilla on the labellum, stylets, and tarsi, as well as the larval sensory peg. AcIr76b is co-localized with the Orco odorant receptor (OR) co-receptor in a subset of cells across the female antennae and labella. In contrast to Orco and Ir8a, chemosensory co-receptors that appear essential for the activity of their respective sets of chemosensory neurons in mosquitoes, AcIr76b-/- mutants maintain wild-type peripheral responses to volatile amines on the adult palps, labellum, and the larval sensory cone. Interestingly, AcIr76b-/- mutants display significantly increased responses to amines in antennal grooved peg sensilla while coeloconic sensilla reveal significant deficits in responses to several acids and amines. Behaviorally, AcIr76b mutants manifest significantly female-specific insemination deficits and, although AcIr76b-/- mutant females are able to locate, alight, and probe artificial blood hosts, they are incapable of blood feeding successfully. Taken together, our study reveals a multi-dimensional functionality of Ir76b in Anopheline olfactory and gustatory pathways that directly impacts the vectorial capacity of these mosquitoes.

SummaryChemosensory receptors play crucial roles across mosquito lifecycles where they often form functional complexes that require cognate co-receptors. To better understand mosquito chemosensory pathways in the malaria vector mosquito An. coluzzii we have utilized advanced gene editing approaches to localize and functionally characterize the ionotropic receptor co-receptor AcIr76b. Expression of AcIr76b was observed in antennal grooved pegs and other accessory olfactory appendages. Mutagenesis of AcIr76b uncovers both reduced and elevated neuronal responses to amines, which suggests a role in response modulation. In addition to olfactory phenotypes, AcIr76b mutants display significantly impaired mating and blood feeding capabilities. Our data reveals discrete roles of AcIr76b across olfactory and gustatory pathways and shed lights on the potential molecular target for vector control strategies.


title: Transcriptional heterogeneity of ventricular zone cells throughout the embryonic mouse forebrain

abstract: The ventricular zone (VZ) of the nervous system contains radial glia cells that were originally considered relatively homogenous in their gene expression. However, a detailed characterization of transcriptional diversity in these VZ cells has not been reported. Here, we performed single-cell RNA sequencing to characterize transcriptional heterogeneity of neural progenitors within the VZ and subventricular zone (SVZ) of the mouse embryonic cortex and ganglionic eminences (GEs). By using a transgenic mouse line to enrich for VZ cells, we detect significant transcriptional heterogeneity within VZ and SVZ progenitors, both between forebrain regions and within spatial subdomains of specific GEs. Additionally, we observe differential gene expression between E12.5 and E14.5 VZ cells, which could provide insights into temporal changes in cell fate. Together, our results reveal a previously unknown spatial and temporal genetic diversity of telencephalic VZ cells that will aid our understanding of initial fate decisions in the forebrain.


title: Skill and value in recruitment forecasting

abstract: Forecasting variation in the recruitment to fish stocks is one of the most challenging and long-running problems in fisheries science and essentially remains unsolved today. Traditionally recruitment forecasts are developed and evaluated based on explanatory and goodness-of-fit approaches that do not reflect their ability to predict beyond the data on which they were developed. Here we propose a new generic framework that allows the skill and value of recruitment forecasts to be assessed in a manner that is relevant to their potential use in an operational setting. We assess forecast skill based on predictive power using a retrospective forecasting approach inspired by meterology, and emphasise the importance of assessing these forecasts relative to a baseline. We quantify the value of these forecasts using an economic cost-loss decision model that is directly relevant to many forecast users. We demonstrate this framework using four stocks of lesser sandeel (Ammodytes marinus) in the North Sea, showing for the first time in an operationally realistic setting that skilful and valuable forecasts are feasible in two of these areas. This result shows the ability to produce valuable short-term recruitment forecasts, and highlights the need to revisit our approach to and understanding of recruitment forecasting.

category: ecology


title: A framework for assessing the skill and value of operational recruitment forecasts


title: Find me if you can: Pre- and Post-hurricane Densities of the Red-bellied Racer (Alsophis rufiventris) on St. Eustatius, and a review of the genus in the Caribbean

abstract: We estimated population densities of the red-bellied racer (Alsophis rufiventris) on the Caribbean island of St. Eustatius in 2011, 2018 and 2019 to determine the likely influence of hurricanes Irma and Maria (September 2017), in addition to evaluating abiotic parameters which may be correlated with its presence. Surveys were conducted at seven sites in 2011 prior to the hurricanes, and at 81 and 108 sites in 2018 and 2019 respectively posterior to the hurricanes. A total of 8.2 ha was surveyed in 2011, and 11.42 ha in 2018/2019. The pre-hurricane (2011) racer density estimate was 9.2/ha (min 7.3 - max 11.6); post-hurricane estimates were 4.6/ha (min 3.4 - max 6.0) in 2018 and 5.0/ha (min 3.8 - max 6.5) in 2019. The pre-hurricane encounter rate of individual racers was 16.0 snakes/hour compared to 0.34 snakes/hour in 2018 and 0.41 snakes/hour in 2019 (post-hurricane). The decrease in encounter rates between 2011 and 2019 implies a negative impact of the hurricanes on racer abundance. Based on calculations of detection probability (0.02 in 2018 and 0.03 in 2019), post-hurricane lambda estimates were 1.82 (95% CI 0.66 - 5.01) in 2018 and 1.60 (95% CI 0.39 - 6.65) snakes/ha in 2019. Given the current small size of the remaining population and the presence of invasive species across the snakes range, this species could be at risk of local extirpation. We suggest conservation actions such as invasive species management and habitat restoration to enable further recovery.


title: Message in a Bottle, Metabarcoding Enables Biodiversity Comparisons Across Ecoregions

abstract:
BackgroundTraditional biomonitoring approaches have delivered a basic understanding of biodiversity, but they cannot support the large-scale assessments required to manage and protect entire ecosystems. This study employed DNA metabarcoding to assess spatial and temporal variation in species richness and diversity in arthropod communities from 52 protected areas spanning three Canadian ecoregions.

ResultsThis study revealed the presence of 26,263 arthropod species in the three ecoregions and indicated that at least another 3,000-5,000 await detection. Results further demonstrate that communities are more similar within than between ecoregions, even after controlling for geographical distance. Overall -diversity declined from east to west, reflecting a gradient in habitat disturbance. Shifts in species composition were high at every site with turnover greater than nestedness, suggesting the presence of many transient species.

ConclusionsDifferences in species composition among their arthropod communities confirm that ecoregions are a useful synoptic for biogeographic patterns and for structuring conservation efforts. The present results also demonstrate that metabarcoding enables large-scale monitoring of shifts in species composition, making it possible to move beyond the biomass measurements that have been the key metric employed in prior efforts to track change in arthropod communities.


title: The crucial role of genome-wide genetic variation in conservation

abstract: The unprecedented rate of extinction calls for efficient use of genetics to help conserve biodiversity. Several recent genomic and simulation-based studies have argued that the field of conservation biology has placed too much focus on the conservation of genome-wide genetic variation, and that this approach should be replaced with another that focuses instead on managing the subset of functional genetic variation that is thought to affect fitness. Here, we critically evaluate the feasibility and likely benefits of this approach in conservation. We find that population genetics theory and empirical results show that the conserving genome-wide genetic variation is generally the best approach to prevent inbreeding depression and loss of adaptive potential from driving populations towards extinction. Focusing conservation efforts on presumably functional genetic variation will only be feasible occasionally, often misleading, and counterproductive when prioritized over genome-wide genetic variation. Given the increasing rate of habitat loss and other environmental changes, failure to recognize the detrimental effects of lost genome-wide variation on long-term population viability will only worsen the biodiversity crisis.


title: Mapping the glycosyltransferase fold landscape using deep learning

abstract: Glycosyltransferases (GTs) play fundamental roles in nearly all cellular processes through the biosynthesis of complex carbohydrates and glycosylation of diverse protein and small molecule substrates. The extensive structural and functional diversification of GTs presents a major challenge in mapping the relationships connecting sequence, structure, fold and function using traditional bioinformatics approaches. Here, we present a convolutional neural network with attention (CNN-attention) based deep learning model that leverages simple secondary structure representations generated from primary sequences to provide GT fold prediction with high accuracy. The model learned distinguishing features free of primary sequence alignment constraints and, unlike other models, is highly interpretable and helped identify common secondary structural features shared by divergent families. The model delineated sequence and structural features characteristic of individual fold types, while classifying them into distinct clusters that group evolutionarily divergent families based on shared secondary structural features. We further extend our model to classify GT families of unknown folds and variants of known folds. By identifying families that are likely to adopt novel folds such as GT91, GT96 and GT97, our studies identify targets for future structural studies and expand the GT fold landscape.

category: bioinformatics


title: CRISPR-VAE: A Method for Explaining CRISPR/Cas12a Predictions, and an Efficiency-aware gRNA Sequence Generator

abstract: Deep learning has shown great promise in the prediction of the gRNA efficiency, which helps optimize the engineered gRNAs, and thus has greatly improved the usage of CRISPR-Cas systems in genome editing. However, the black box prediction of deep learning methods does not provide adequate explanation to the factors that increase efficiency; rectifying this issue promotes the usage of CRISPR-Cas systems in numerous domains. We put forward a framework for interpreting gRNA efficiency prediction, dubbed CRISPR-VAE, that improves understanding the factors that increase gRNA efficiency, and apply it to CRISPR/Cas12a (formally known as CRISPR/Cpf1). We further lay out a semantic articulation of such factors into position-wise k-mer rules. The paradigm consists of building an efficiency-aware gRNA sequence generator trained on available real data, and using it to generate a large amount of synthetic sequences with favorable traits, upon which the explanation of the gRNA prediction is based. CRISPR-VAE can further be used as a standalone sequence generator, where the user has low-level control ability. The framework can be readily integrated with different CRISPR-Cas tools and datasets, and its efficacy is confirmed. The complete implementation of the methods can be found at github.com/AhmadObeid/CRISPR-VAE.


title: Relaxation time asymmetry in stator dynamics of the bacterial flagellar motor

abstract: The bacterial flagellar motor (BFM) is the membrane-embedded rotary molecular motor which turns the flagellum that provides thrust to many bacterial species. This large multimeric complex, composed of a few dozen constituent proteins, has emerged as a hallmark of dynamic subunit exchange. The stator units are inner-membrane ion channels which dynamically bind and unbind to the peptidoglycan at the rotor periphery, consuming the ion motive force (IMF) and applying torque to the rotor when bound. The dynamic exchange is known to be a function of the viscous load on the flagellum, allowing the bacterium to dynamically adapt to its local viscous environment, but the molecular mechanisms of exchange and mechanosensitivity remain to be revealed. Here, by actively perturbing the steady-state stator stoichiometry of individual motors, we reveal a stoichiometry-dependent asymmetry in stator remodeling kinetics. We interrogate the potential effect of next-neighbor interactions and local stator unit depletion and find that neither can explain the observed asymmetry. We then simulate and fit two mechanistically diverse models which recapitulate the asymmetry, finding stator assembly dynamics to be particularly well described by a two-state catch-bond mechanism.

category: biophysics


title: Exploring the Inner Workings of Neuron Circuits That Exhibit Persistent Activity To Explain How Working Memory and Executive Function Are Implemented in The Brain

abstract:
In this research we explore in detail how a phenomenon called "sustained persistent activity" is achieved by circuits of interconnected neurons. Persistent activity is a phenomenon that has been extensively studied (Papoutsi et al. 2013; Kaminski et. al. 2017; McCormick et al. 2003; Rahman, and Berger, 2011). Persistent activity consists of neuron circuits whose spiking activity remains even after the initial stimuli are removed. Persistent activity has been found in the prefrontal cortex (PFC) and has been correlated to working memory and decision making (Clayton E. Curtis and Daeyeol Lee, 2010).

We go beyond the explanation of how persistent activity happens and show how arrangements of those basic circuits encode and store data and are used to perform more elaborated tasks and computations.

The purpose of the model we propose here is to describe the minimum number of neurons and their interconnections required to explain persistent activity and how this phenomenon is actually a fast storage mechanism required for implementing working memory, task processing and decision making.


title: An advanced automated patch clamp protocol design to investigate drug - ion channel binding dynamics.

abstract: Standard high throughput screening projects using automated patch-clamp instruments often fail to grasp essential details of the mechanism of action, such as binding/unbinding dynamics and modulation of gating. In this study, we aim to demonstrate that depth of analysis can be combined with acceptable throughput on such instruments. Using the microfluidics-based automated patch clamp, IonFlux Mercury, we developed a method for a rapid assessment of the mechanism of action of sodium channel inhibitors, including their state-dependent association and dissociation kinetics. The method is based on a complex voltage protocol, which is repeated at 1 Hz. Using this time resolution we could monitor the onset and offset of both channel block and modulation of gating upon drug perfusion and washout. Our results show that the onset and the offset of drug effects are complex processes, involving several steps, which may occur on different time scales. We could identify distinct sub-processes on the millisecond time scale, as well as on the second time scale. Automated analysis of the results allows collection of detailed information regarding the mechanism of action of individual compounds, which may help the assessment of therapeutic potential for hyperexcitability-related disorders, such as epilepsies, pain syndromes, neuromuscular disorders, or neurodegenerative diseases.

category: pharmacology and toxicology


title: Brain-like functional specialization emerges spontaneously in deep neural networks

abstract: The last quarter century of cognitive neuroscience has revealed numerous cortical regions in humans with distinct, often highly specialized functions, from recognizing faces to understanding language to thinking about what other people are thinking. But it remains unclear why the cortex exhibits this high degree of functional specialization in the first place. Here, we consider the case of face perception, using artificial neural networks to test the hypothesis that functional segregation of face recognition in the brain reflects the computational requirements of the task. We find that networks trained on generic object recognition perform poorly on face recognition and vice versa, and further that networks optimized for both tasks spontaneously segregate themselves into separate systems for faces and objects. Thus, generic visual features that suffice for object recognition are apparently suboptimal for face recognition and vice versa. We then show functional segregation to varying degrees for other visual categories, revealing a widespread tendency for optimization (without built-in task-specific inductive biases) to lead to functional specialization in machines and, we conjecture, also brains.


title: Differential impact of Kv8.2 loss on rod and cone signaling and degeneration.

abstract: Heteromeric Kv2.1/Kv8.2 channels are voltage-gated potassium channels localized to the photoreceptor inner segment. They carry IKx, which is largely responsible for setting the photoreceptor resting membrane potential. Mutations in Kv8.2 result in childhood-onset Cone Dystrophy with Supernormal Rod Response (CDSRR). We generated a Kv8.2 knockout (KO) mouse and examined retinal signaling and photoreceptor degeneration to gain deeper insight into the complex phenotypes of this disease. Using electroretinograms we show that there were delayed or reduced signaling from rods depending on the intensity of the light stimulus, consistent with reduced capacity for light-evoked changes in membrane potential. The delayed response was not seen ex vivo where extracellular potassium levels were controlled by the perfusion buffer, so we propose the in vivo alteration is influenced by genotype-associated ionic imbalance. We observed mild retinal degeneration. Signaling from cones was reduced but there was no loss of cone density. Loss of Kv8.2 altered responses to flickering light with responses attenuated at high frequencies and altered in shape at low frequencies. The Kv8.2 KO line on an all-cone retina background had reduced cone-driven ERG b wave amplitudes and underwent degeneration. Altogether, we provide insight into how a deficit in the dark current affects the health and function of photoreceptors.


title: Disentangling niche theory and beta diversity change

abstract: Beta diversity describes the differences in species composition among communities. Changes in beta diversity over time are thought to be due to selection based on species niche characteristics. For example, theory predicts that selection that favours habitat specialists will increase beta diversity. In practice, ecologists struggle to predict how beta diversity changes. To remedy this problem, we propose a novel solution that formally measures selections effects on beta diversity. Using the Price equation, we show how change in beta diversity over time can be partitioned into fundamental mechanisms including selection among species, variable selection among communities, drift, and immigration. A key finding of our approach is that a species short-term impact on beta diversity cannot be predicted using information on its long-term environmental requirements (i.e. its niche). We illustrate how our approach can be used to partition causes of diversity change in a montane tropical forest before and after an intense hurricane. Previous work in this system highlighted the resistance of habitat specialists and the recruitment of light-demanding species but was unable to quantify the importance of these effects on beta diversity. Using our approach, we show that changes in beta diversity were consistent with ecological drift. We use these results to highlight the opportunities presented by a synthesis of beta diversity and formal models of selection.


title: Population-level inference for home-range areas

abstract:
O_LIHome-range estimates are a common product of animal tracking data, as each range informs on the area needed by a given individual. Population-level inference on home-range areas--where multiple individual home-ranges are considered to be sampled from a population--is also important to evaluate changes over time, space, or covariates, such as habitat quality or fragmentation, and for comparative analyses of species averages. Population-level home-range parameters have traditionally been estimated by first assuming that the input tracking data were sampled independently when calculating home ranges via conventional kernel density estimation (KDE) or minimal convex polygon (MCP) methods, and then assuming that those individual home ranges were measured exactly when calculating the population-level estimates. This conventional approach does not account for the temporal autocorrelation that is inherent in modern tracking data, nor for the uncertainties of each individual home-range estimate, which are often large and heterogeneous.
C_LIO_LIHere, we introduce a statistically and computationally efficient framework for the population-level analysis of home-range areas, based on autocorrelated kernel density estimation (AKDE), that can account for variable temporal autocorrelation and estimation uncertainty.
C_LIO_LIWe apply our method to empirical examples on lowland tapir (Tapirus terrestris), kinkajou (Potos flavus), white-nosed coati (Nasua narica), white-faced capuchin monkey (Cebus capucinus), and spider monkey (Ateles geoffroyi), and quantify differences between species, environments, and sexes.
C_LIO_LIOur approach allows researchers to more accurately compare different populations with different movement behaviors or sampling schedules, while retaining statistical precision and power when individual home-range uncertainties vary. Finally, we emphasize the estimation of effect sizes when comparing populations, rather than mere significance tests.
C_LI


title: Network-level containment of single-species bioengineering

abstract: Ecological systems are facing major diversity losses in this century due to Anthropogenic effects. Habitat loss, overexploitation of resources, invasion and pollution are rapidly jeopardising the survival of whole communities, as revealed by pronounced population losses. Moreover, the potential of future tipping points further complicate their survival and change our perspective of risk. It has been recently suggested that a potential approach to flatten the curve of species extinction and prevent catastrophic shifts would involve the engineering of one selected species within one of these communities, aiming at helping the maintenance of key conditions compatible with high diversity. Such possibility has started to become part of potential intervention scenarios to preserve coral reefs, kelp forests or soil microbiomes in drylands. Despite its potential, very little is known about the actual dynamic responses of complex ecological networks to the introduction of a synthetic strains derived from a resident species. In this paper we address this problem by modelling the response of a competitive community to the addition of a synthetic strain derived from a member of a stable ecosystem. We show that the community interaction matrix largely limits the spread of the engineered strain, thus suggesting that species diversity acts as an ecological firewall. Implications for future restoration and terraformation strategies are discussed.

category: bioengineering


title: Projecting genetic associations through gene expression patterns highlights disease etiology and drug mechanisms

abstract: Understanding how dysregulated transcriptional processes result in tissue-specific pathology requires a mechanistic interpretation of expression regulation across different cell types. It has been shown that this insight is key for the development of new therapies. These mechanisms can be identified with transcriptome-wide association studies (TWAS), which have represented a significant step forward to test the mediating role of gene expression in GWAS associations. However, it is hard to disentangle causal cell types using eQTL data alone, and other methods generally do not use the large amounts of publicly available RNA-seq data. Here we introduce PhenoPLIER, a polygenic approach that maps both gene-trait associations and pharmacological perturbation data into a common latent representation for a joint analysis. This representation is based on modules of genes with similar expression patterns across the same tissues. We observed that diseases were significantly associated with gene modules expressed in relevant cell types, and our approach was accurate in predicting known drug-disease pairs and inferring mechanisms of action. Furthermore, using a CRISPR screen to analyze lipid regulation, we found that functionally important players lacked TWAS associations but were prioritized in phenotype-associated modules by PhenoPLIER. By incorporating groups of co-expressed genes, PhenoPLIER can contextualize genetic associations and reveal potential targets within associated processes that are missed by single-gene strategies.


title: Compound-specific, concentration-independent biophysical properties of sodium channel inhibitor mechanism of action

abstract: We have developed an automated patch-clamp protocol that allows high information content screening of sodium channel inhibitor compounds. We have observed that individual compounds had their specific signature patterns of inhibition, which were manifested irrespective of the concentration. Our aim in this study was to quantify these properties. Primary biophysical data, such as onset rate, the shift of the half inactivation voltage, or the delay of recovery from inactivation, are concentration-dependent. We wanted to derive compound-specific properties, therefore, we had to neutralize the effect of concentration. This study describes how this is done, and shows how compound-specific properties reflect the mechanism of action, including binding dynamics, cooperativity, and interaction with the membrane phase. We illustrate the method using four well-known sodium channel inhibitor compounds, riluzole, lidocaine, benzocaine, and bupivacaine. Compound-specific biophysical properties may also serve as a basis for deriving parameters for kinetic modeling of drug action. We discuss how knowledge about the mechanism of action may help to predict the frequency-dependence of individual compounds, as well as their potential persistent current component selectivity. The analysis method described in this study, together with the experimental protocol described in the accompanying paper, allows screening for inhibitor compounds with specific kinetic properties, or with specific mechanisms of inhibition.


title: Discovery of putative Golgi S-Adenosyl methionine transporters reveals the importance of plant cell wall polysaccharide methylation

abstract: Polysaccharide methylation, especially that of pectin, is a common and important feature of land plant cell walls. Polysaccharide methylation takes place in the Golgi apparatus and therefore relies on the import of S-adenosyl methionine (SAM) from the cytosol into the Golgi. However, to date, no Golgi SAM transporter has been identified in plants. In this work, we studied major facilitator superfamily members in Arabidopsis that we identified as putative Golgi SAM transporters (GoSAMTs). Knock-out of the two most highly expressed GoSAMTs led to a strong reduction in Golgi-synthesised polysaccharide methylation. Furthermore, solid-state NMR experiments revealed that reduced methylation changed cell wall polysaccharide conformations, interactions and mobilities. Notably, the NMR revealed the existence of pectin  egg-box structures in intact cell walls, and showed that their formation is enhanced by reduced methyl-esterification. These changes in wall architecture were linked to substantial growth and developmental phenotypes. In particular, anisotropic growth was strongly impaired in the double mutant. The identification of putative transporters that import SAM into the Golgi lumen in plants provides new insights into the paramount importance of polysaccharide methylation for plant cell wall structure and function.

category: plant biology


title: Developmental increase of inhibition drives decorrelation of neural activity

abstract: Throughout development, the brain transits from early highly synchronous activity patterns to a mature state with sparse and decorrelated neural activity, yet the mechanisms underlying this process are unknown. The developmental transition has important functional consequences, as the latter state allows for more efficient storage, retrieval and processing of information. Here, we show that, in the mouse medial prefrontal cortex (mPFC), neural activity during the first two postnatal weeks decorrelates following specific spatial patterns. This process is accompanied by a concomitant tilting of excitation/inhibition (E-I) ratio towards inhibition. Using optogenetic manipulations and neural network modeling, we show that the two phenomena are mechanistically linked, and that a relative increase of inhibition drives the decorrelation of neural activity. Accordingly, in two mouse models of neurodevelopmental disorders, subtle alterations in E-I ratio are associated with specific impairments in the correlational structure of spike trains. Finally, capitalizing on EEG data from newborn babies, we show that an analogous developmental transition takes place also in the human brain. Thus, changes in E-I ratio control the (de)correlation of neural activity and, by these means, its developmental imbalance might contribute to the pathogenesis of neurodevelopmental disorders.


title: Target-specific control of olfactory bulb periglomerular cells by GABAergic and cholinergic basal forebrain inputs

abstract: The olfactory bulb (OB), the first relay for odor processing, receives dense GABAergic and cholinergic long-range projections from basal forebrain (BF) nuclei that provide information about the internal state and behavioral context of the animal. However, the targets, impact and dynamics of these afferents are still unclear. I studied how BF synaptic inputs modulate activity in diverse subtypes of periglomerular (PG) interneurons using optogenetic stimulation and loose cell-attached or whole-cell patch-clamp recording in OB slices from adult mice. GABAergic BF inputs potently blocked PG cells firing except in a minority of calretinin-expressing cells in which GABA release elicited spiking. Parallel cholinergic projections excited a previously overlooked PG cell subtype via synaptic activation of M1 muscarinic receptors. Low frequency stimulation of the cholinergic axons drove persistent firing in these PG cells thereby increasing tonic inhibition in principal neurons. Taken together, these findings suggest that modality-specific BF inputs can orchestrate inhibition in OB glomeruli using multiple, potentially independent, inhibitory or excitatory target-specific pathways.


title: Adolescent dopamine neurons represent reward differently during action and state guided learning

abstract: The neuronal underpinning of learning cause-and-effect associations in the adolescent brain remains poorly understood. Two fundamental forms of associative learning are Pavlovian (classical) conditioning, where a stimulus is followed by an outcome, and operant (instrumental) conditioning, where outcome is contingent on action execution. Both forms of learning, when associated with a rewarding outcome, rely on midbrain dopamine neurons in the ventral tegmental area (VTA) and substantia nigra (SN). We find that in adolescent male rats, reward-guided associative learning is encoded differently by midbrain dopamine neurons in each conditioning paradigm. Whereas simultaneously recorded VTA and SN adult neurons have a similar phasic response to reward delivery during both forms of conditioning, adolescent neurons display a muted reward response during operant but a profoundly larger reward response during Pavlovian conditioning suggesting that adolescent neurons assign a different value to reward when it is not gated by action. The learning rate of adolescents and adults during both forms of conditioning was similar further supporting the notion that differences in reward response in each paradigm are due to differences in motivation and independent of state versus action value learning. Static characteristics of dopamine neurons such as dopamine cell number and size were similar in the VTA and SN but there were age differences in baseline firing rate, stimulated release and correlated spike activity suggesting that differences in reward responsiveness by adolescent dopamine neurons are not due to differences in intrinsic properties of these neurons but engagement of different networks.


title: Daily oscillation of the excitation/inhibition ratio is disrupted in two mouse models of autism

abstract: Autism spectrum disorder (ASD) is a prevalent neurodevelopmental disorder involving sensory processing abnormalities. Alterations to the balance between excitation and inhibition (E/I ratio) are postulated to underlie behavioral phenotypes in ASD patients and mouse models. However, in primary visual cortex (V1) of wild type mice, the E/I ratio is not a fixed value, but rather oscillates across the 24h day. Therefore, we hypothesized that the E/I oscillation, rather than the overall E/I ratio, may be disrupted in ASD mouse models. To this end, we measured the E/I ratio in Fmr1 KO and BTBR mice, models of syndromic and idiopathic ASD, respectively. We found that the E/I ratio is dysregulated in both models, but in different ways: the oscillation is flattened in Fmr1 KO and phase-shifted in BTBR mice. These phenotypes cannot be explained by altered sleep timing, which was largely normal in both lines. Furthermore, we found that E/I dysregulation occurs due to alterations in both excitatory and inhibitory synaptic transmission in both models. These findings provide a crucial perspective on the E/I ratio in ASD, suggesting that ASD phenotypes may be produced by a mismatch of E/I to the appropriate behavioral state, rather than alterations to overall E/I levels per se.


title: UVC inactivation of pathogenic samples suitable for cryoEM analysis

abstract: Cryo-electron microscopy has become an essential tool to understand structure and function of biological samples, from individual proteins to whole cells. Especially for pathogens, such as disease-causing bacteria and viruses, insights gained by cryo-EM can aid in developing cures. However, due to the biosafety restrictions of human pathogens, samples are often treated by chemical fixation to render the pathogen inert, affecting the delicate ultrastructure of the sample. Alternatively, researchers use in vitro or ex vivo models, which are non-pathogenic but lack the complexity of the pathogen of interest. Here we show that ultraviolet-C (UVC) radiation at cryogenic temperatures can be used to eliminate or dramatically reduce the infectivity of two model organisms, a pathogenic bacterium (Vibrio cholerae) and a virus-like particle (the ICP1 bacteriophage). We show no discernable structural impact of this treatment of either sample using two cryo-EM methods: cryo-electron tomography (cryo-ET) followed by sub-tomogram averaging (STA), and single particle analysis (SPA). Additionally, we applied the UVC irradiation to the protein apoferritin (ApoF), which is a widely used test sample for high resolution SPA studies. The UVC-treated ApoF sample resulted in a 2.1 [A] structure that did not reveal any discernable structural damage. Together, these results show that the UVC irradiation dose that effectively inactivates cryo-EM samples does not negatively impact their structure. This research demonstrates that UVC treatment is an effective and inexpensive addition to the cryo-EM sample preparation toolbox.


title: Irisin stimulates the release of CXCL1 from differentiating human subcutaneous and deep-neck derived adipocytes via upregulation of NFκB pathway

abstract: Thermogenic brown and beige adipocytes play an important role in combating obesity. Recent studies in rodents and humans have indicated that these adipocytes release cytokines, termed "batokines". Irisin was discovered as a polypeptide regulator of beige adipocytes released by myocytes, primarily during exercise. We performed global RNA sequencing on adipocytes derived from human subcutaneous and deep-neck precursors, which were differentiated in the presence or absence of irisin. Irisin did not exert an effect on the expression of characteristic thermogenic genes, while upregulated genes belonging to various cytokine signaling pathways. Out of the several upregulated cytokines, CXCL1, the highest upregulated, was released throughout the entire differentiation period, and predominantly by differentiated adipocytes. Deep-neck area tissue biopsies also showed a significant release of CXCL1 during 24 hours irisin treatment. Gene expression data indicated upregulation of the NF[kcy]B pathway upon irisin treatment, which was validated by an increase of p50 and decrease of I[kcy]B protein level, respectively. Continuous blocking of the NF{kappa}B pathway, using a cell permeable inhibitor of NF{kappa}B nuclear translocation, significantly reduced CXCL1 release. The released CXCL1 exerted a positive effect on the adhesion of endothelial cells. Together, our findings demonstrate that irisin stimulates the release of a novel "batokine", CXCL1, via upregulation of NF{kappa}B pathway in neck area derived adipocytes, which might play an important role in improving tissue vascularization.


title: PLK1 controls centriole distal appendage formation and centrobin removal via independent pathways

abstract: Centrioles are central structural elements of centrosomes and cilia. They originate as daughter centrioles from existing centrioles in S-phase and reach their full functionality with the formation of distal and subdistal appendages two mitoses later. Current models postulate that the centriolar protein centrobin acts as placeholder for distal appendage proteins that must be removed to complete distal appendage formation. Here, we investigated in non-transformed human epithelial cells the mechanisms controlling centrobin removal and its effect on distal appendage formation. We demonstrate that centrobin is removed from older centrioles due to a higher affinity for the newly born daughter centrioles, under the control of the centrosomal kinase Plk1. Centrobin removal also depends on the presence of subdistal appendage proteins on the oldest centriole. It is, however, not required for distal appendage formation even though this process is equally dependent on Plk1. We conclude that during centriole maturation, Plk1 kinase regulates centrobin removal and distal appendage formation via separate pathways.


title: Phosphorylation of STIM1 at ERK/CDK sites is dispensable for cell migration and ER partitioning in mitosis

abstract: Store-operated Ca2+ entry (SOCE) is a ubiquitous Ca2+ influx required for multiple physiological functions including cell motility. SOCE is activated in response to depletion of intracellular Ca2+ stores following the activation of endoplasmic reticulum (ER) Ca2+ sensor STIM1 which recruits the plasma membrane (PM) Ca2+ channel Orai1 at ER-PM junctions to induce Ca2+ influx. STIM1 is phosphorylated dynamically and this phosphorylation has been implicated in several processes including SOCE inactivation during M-phase, maximal SOCE activation, ER segregation during mitosis, and cell migration. Human STIM1 has 10 Ser/Thr residues in its cytosolic domain that match the ERK/CDK consensus phosphorylation. We recently generated a mouse knock-in line where wild-type STIM1 was replaced by a non-phosphorylatable STIM1 with all 10 S/T mutated to Ala (STIM1-10A). Here, we generate mouse embryonic fibroblasts (MEF) the STIM1-10A mouse line and a control MEF line (WT) that express wild-type STIM1 from a congenic mouse strain. These lines offer a unique model to address the role of STIM1 phosphorylation at endogenous expression and modulation levels in contrast to previous studies that relied mostly on overexpression. We show that STIM1 phosphorylation at ERK/CDK sites is not required for SOCE activation, cell migration, or ER partitioning during mitosis. These results rule out STIM1 phosphorylation as a regulator of SOCE, migration and ER distribution in mitosis.


title: Identification of the cell-type-specific ER membrane protein Tanmp expressed in hypothalamic tanycytes and subsets of neurons

abstract: Genomes of higher eukaryotes encode many uncharacterized proteins, and the functions of these proteins cannot be predicted from the primary sequences due to a lack of conserved functional domains. During a screening of novel noncoding RNAs abundantly expressed in mouse brains, we incidentally identified a gene termed Tanmp, which encoded an endoplasmic reticulum (ER) protein without known functional domains. Tanmp is specifically expressed in the nervous system, and the highest expression was observed in a specialized cell type called tanycyte that aligns the ventral wall of the third ventricle in the hypothalamus. Immunostaining of Tanmp revealed the fine morphology of tanycytes with highly branched apical ER membranes. Immunoprecipitation revealed that Tanmp associates with mitochondrial ATPase at least in vitro, and ER and mitochondrial signals occasionally overlapped in tanycytes. Mutant mice lacking Tanmp did not exhibit overt phenotypes, suggesting that Tanmp is not essential in mice reared under normal laboratory conditions. We also found that RNA probes that are predicted to uniquely detect Tanmp mRNA cross-reacted with uncharacterized RNAs, highlighting the importance of experimental validation of the specificity of probes during the hybridization-based study of RNA localization.


title: Tumour Suppressor Parafibromin/Hyrax Governs Cell Polarity and Centrosome Assembly in Neural Stem Cells

abstract: Neural stem cells (NSCs) divide asymmetrically to balance their self-renewal and differentiation, an imbalance in which can lead to NSC overgrowth and tumour formation. The functions of Parafibromin, a conserved tumour suppressor, in the nervous system are not established. Here, we demonstrate that Drosophila Parafibromin/Hyrax (Hyx) inhibits NSC overgrowth by governing cell polarity. Hyx is essential for the apicobasal polarization of NSCs, through its role in the asymmetric distribution of polarity proteins. hyx depletion results in the symmetric division of NSCs, leading to the formation of supernumerary NSCs in the larval brain. Importantly, we show that human Parafibromin can fully rescue NSC overgrowth and cell polarity defects in Drosophila hyx mutant brains. We have also discovered a novel role for Hyx in regulating the formation of interphase microtubule-organizing center and mitotic spindles in NSCs. Moreover, Hyx is required for the proper localization of a key centrosomal protein, Polo, and the microtubule-binding proteins Msps and D-TACC in dividing NSCs. Furthermore, Hyx directly regulates the polo expression in vitro. Altogether, our study provides the first evidence that the brain tumour suppressor-like role and polarity establishing functions of Hyx are mediated by its role in regulating microtubule growth and centrosomal assembly. The new paradigm that Parafibromin orchestrates cell polarization by regulating centrosomal assembly may be relevant to future studies on Parafibromin/HRPT2-associated cancers.


title: Predicting RNA splicing from DNA sequence using Pangolin

abstract: Recent progress in deep learning approaches have greatly improved the prediction of RNA splicing from DNA sequence. Here, we present Pangolin, a deep learning model to predict splice site strength in multiple tissues that has been trained on RNA splicing and sequence data from four species. Pangolin outperforms state of the art methods for predicting RNA splicing on a variety of prediction tasks. We use Pangolin to study the impact of genetic variants on RNA splicing, including lineage-specific variants and rare variants of uncertain significance. Pangolin predicts loss-of-function mutations with high accuracy and recall, particularly for mutations that are not missense or nonsense (AUPRC = 0.93), demonstrating remarkable potential for identifying pathogenic variants.

category: genomics


title: Molecular mechanism of target site selection and remodeling by type V CRISPR-associated transposons

abstract: Although the canonical function of CRISPR-Cas systems is to provide adaptive immunity against mobile genetic elements1, type I-F, I-B and V-K systems have been adopted by Tn7-like transposons to direct RNA-guided transposon insertion2-7. Type V-K CRISPR-associated transposons rely on the activities of the pseudonuclease Cas12k, the transposase TnsB, the AAA+ ATPase TnsC and the zinc-finger protein TniQ7. However, the molecular and structural details of RNA-directed DNA transposition have remained elusive. Here we report cryo-electron microscopic structures of a Cas12k-guide RNA-target DNA complex and a DNA-bound, polymeric TnsC filament. The Cas12k complex structure reveals an intricate guide RNA architecture and critical interactions mediating RNA-guided target DNA recognition. The assembly of the TnsC helical filament is ATP-dependent and accompanied by structural remodeling of the bound DNA duplex. In vivo transposition assays corroborate key features of the structures, and biochemical experiments further show that TniQ restricts TnsC polymerization, while the TnsB transposase interacts directly with TnsC filaments to trigger their disassembly upon ATP hydrolysis. Together, these results suggest a mechanistic model whereby RNA-directed target selection by Cas12k primes TnsC polymerization and DNA remodeling, generating a recruitment platform for TnsB to catalyze site-specific transposon insertion. The present work advances our mechanistic understanding of the cross-talk between CRISPR effectors and the transposition machinery and will inform design efforts to harness CRISPR-associated transposons as programmable site-specific gene insertion tools for genome engineering applications.

category: biochemistry


title: The molecular mechanism of cytoadherence to placenta or tumor cells through VAR2CSA from Plasmodium falciparum

abstract: Pregnancy Associated Malaria (PAM) threatens more than one million pregnant women and their infants in endemic regions due to poor outcomes. VAR2CSA plays a vital role in the cytoadherence of infected erythrocytes (IEs) to placenta against immuno-clearance via binding to Chondroitin sulfate A (CSA), which is displayed mostly on the surface of placental or tumor cells. In this study, we determined the cryo-EM structures of VAR2CSA ectodomain and its complex with CSA at a resolution of 3.6 [A] and 3.4 [A], respectively and revealed that CSA binding induces significant conformational change for ligand accommodation. Beyond structural studies, we generated VAR2CSA fragments and mutation and evaluated their binding activity to either isolated CSA or placental and tumor cell line using multi-disciplinary techniques. The results showed that 9-site mutation in DBL2X abolished the CSA binding activity and also disrupted its binding to both placental and tumor cells. Overall, our work clearly elucidated the molecular mechanism of cytoadherence to placental or tumor cells through VAR2CSA. Which may facilitate new PAM vaccine development and improve the drug delivery systems targeting both placenta and tumors.


title: Algorithms for discovery of cis-eQTL signals in woody species: the vine (Vitis vinifera L.) as a study model

abstract: Expression quantitative trait loci (eQTLs), are associations between genetic variants, such as Single Nucleotide Polymorphisms (SNPs), and gene expression. eQTLs are an important tool to understand the genetics of gene expression of complex phenotypes. eQTLs analysis are common in human studies and in model species such as mice, rats and yeast but are very scarce in wood crop species such as fruit trees or grapevines. In this study a comprehensive bioinformatic pipeline has been carried out using genomics and expression data from 10 genotypes of grape, which has been used as model species. As a result of this study a total of 10,618 genetic variants that regulate gene expression levels of 525 genes were detected. A 78% of them, 411, received a functional annotation from UniProtKB or DAVID, between the annotated protein-coding genes are Germin-like proteins (GLPs), auxin-regulatory factors, GRFS, ANK_REP_REGION domain-containing protein, Kinesin motor domain-containing protein or RPP13like protein 2(LOC100852873). This new inventory of cis eQTLs influencing gene expression during the ripening of fruits of Vitis vinifera L. will be an important resource for future research to understand the mechanistic basis for variation in gene regulation in this species. In the future, this methodology may be applied to other woody species, once the necessary databases are generated.


title: The Phytochemical Diversity of Commercial Cannabis in the United States

abstract: The legal status of Cannabis is changing, fueling an increased diversity of Cannabis-derived products. Because Cannabis contains dozens of chemical compounds with potential psychoactive or medicinal effects, understanding its phytochemical diversity is crucial. The legal Cannabis industry heavily markets products to consumers based on widely used labelling systems purported to predict the effects of different Cannabis "strains." We analyzed the cannabinoid and terpene content of tens of thousands of commercial Cannabis samples across six US states, finding distinct chemical phenotypes (chemotypes) which are reliably present. After careful descriptive analysis of the phytochemical diversity and comparison to the commercial labels commonly attached to Cannabis samples, we show that commercial labels do not consistently align with the observed chemical diversity. However, certain labels are statistically overrepresented for specific chemotypes. These results have important implications for the classification of commercial Cannabis, the design of animal and human research, and the regulation of legal Cannabis marketing.


title: Glial place cells: complementary encoding of spatial information in hippocampal astrocytes

abstract: Calcium dynamics into astrocytes influence the activity of nearby neuronal structures. However, because previous reports show that astrocytic calcium signals largely mirror neighboring neuronal activity, current information coding models neglect astrocytes. Using simultaneous two-photon calcium imaging of astrocytes and neurons in the hippocampus of mice navigating a virtual environment, we demonstrate that astrocytic calcium signals actively encode spatial information. Calcium events carrying spatial information occurred in topographically organized astrocytic subregions. Importantly, astrocytes encoded spatial information that was complementary and synergistic to that carried by neurons, improving spatial position decoding when astrocytic signals were considered alongside neuronal ones. These results suggest that the complementary place-dependence of localized astrocytic calcium signals regulates clusters of nearby synapses, enabling dynamic, context-dependent, variations in population coding within brain circuits.


title: Multidimensional MRI for characterization of subtle axonal injury accelerated using an adaptive nonlocal multispectral filter

abstract: Multidimensional MRI is an emerging approach that simultaneously encodes water relaxation (T1 and T2) and mobility (diffusion) and replaces voxel-averaged values with subvoxel distributions of those MR properties. While conventional (i.e., voxel-averaged) MRI methods cannot adequately quantify the microscopic heterogeneity of biological tissue, using subvoxel information allows to selectively map a specific T1-T2-diffusion spectral range that corresponds to a group of tissue elements. The major obstacle to the adoption of rich, multidimensional MRI protocols for diagnostic or monitoring purposes is the prolonged scan time. Our main goal in the present study is to evaluate the performance of a nonlocal estimation of multispectral magnitudes (NESMA) filter on reduced datasets to limit the total acquisition time required for reliable multidimensional MRI characterization of the brain. Here we focused and reprocessed results from a recent study that identified potential imaging biomarkers of axonal injury pathology from the joint analysis of multidimensional MRI, in particular voxelwise T1-T2 and diffusion-T2 spectra in human Corpus Callosum, and histopathological data. We tested the performance of NESMA and its effect on the accuracy of the injury biomarker maps, relative to the co-registered histological reference. Noise reduction improved the accuracy of the resulting injury biomarker maps, while permitting data reduction of 35.7% and 59.6% from the full dataset for T1-T2 and diffusion-T2 cases, respectively. As successful clinical proof-of-concept applications of multidimensional MRI are continuously being introduced, reliable and robust noise removal and consequent acquisition acceleration would advance the field towards clinically-feasible diagnostic multidimensional MRI protocols.


title: Small molecules enhance the potency of natural antimicrobial peptides

abstract: The skin-associated microbiome plays an important role in general well-being and in a variety of treatable conditions. In this regard, endogenous antimicrobial peptides have a role in controlling the microbial population. We demonstrate here that certain small molecular species can amplify the potency of naturally-occurring antimicrobial peptides. For example, Niacinamide is a vitamin B3 analogue naturally found in foods and widely used in topical skin care products, and here we have investigated its cooperativity with the human antimicrobial peptide LL37 on the bacterium Staphylococcus aureus. We have also studied two other structurally related B3 analogs. We observed a clear synergistic effect of niacinamide and, to some extent, methyl niacinamide, whereas isonicotinamide showed no significant cooperativity with LL37. Adaptively-biased molecular dynamics simulations revealed that the analogs partition into the head group region of an anionic bilayer used to mimic the bacterial membrane. The observed effects on the physical properties of the membrane are well correlated with experimental activity. In contrast, the analogs have little effect on zwitterionic bilayers which mimic a mammalian membrane. We conclude that these vitamin B3 analogues can potentiate the activity of host peptides by modulating the physical properties of the bacterial membrane, and to a lesser extent through direct interactions with the peptide. The level of cooperativity is strongly dependent on the detailed chemistry of the additive, suggesting an opportunity to fine-tune the behaviour of host peptides.


title: Managing human mediated range shifts: understanding spatial, temporal and genetic variation in marine non- native species

abstract: The use of molecular methods to manage natural resources is increasingly common. However, DNA-based methods are seldom used to understand the spatial and temporal dynamics of species range shifts. This is important when managing range-shifting species such as non-native species (NNS), which can have negative impacts on biotic communities. Here we investigated the range-shifting NNS Ciona robusta, Clavelina lepadiformis, Microcosmus squamiger and Styela plicata using a combined methodological approach. We first conducted non-molecular biodiversity surveys for these NSS along the South African coastline, and compared the results with historical surveys. We detected no consistent change in range size across species, with some displaying range stability and others showing range shifts. We then sequenced a section of cytochrome c oxidase subunit I (COI) from tissue samples and found genetic differences along the coastline but no change over recent times. Finally, we found that environmental DNA metabarcoding data showed broad congruence with both the non-molecular biodiversity and the COI datasets, but failed to capture complete incidence of all NSS. Overall, we demonstrated how a combined methodological approach can effectively detect spatial and temporal variation in genetic composition and range size, which is key for managing biodiversity changes of both threatened and NSS.


title: A polygenic architecture with conditionally neutral effects underlies ecological differentiation in Silene

abstract:
O_LIEcological differentiation can drive speciation but it is unclear how the genetic architecture of habitat-dependent fitness contributes to lineage divergence. We investigated the genetic architecture of cumulative flowering, a fitness component, in second-generation hybrids between Silene dioica and S. latifolia transplanted into the natural habitat of each species.
C_LIO_LIWe used reduced-representation sequencing and Bayesian Sparse Linear Mixed Models (BSLMMs) to analyze the genetic control of cumulative flowering in each habitat.
C_LIO_LIOur results point to a polygenic architecture of cumulative flowering. Allelic effects were mostly beneficial or deleterious in one habitat and neutral in the other. The direction of allelic effects was associated with allele frequency differences between the species: positive-effect alleles were often derived from the native species, whereas negative-effect alleles, at other loci, tended to originate from the non-native species.
C_LIO_LIWe conclude that ecological differentiation is governed and maintained by many loci with small, conditionally neutral effects. Conditional neutrality may result from differences in selection targets in the two habitats and provides hidden variation upon which selection can act. Polygenic architectures of adaptive differentiation are expected to be transient during lineage divergence and may therefore be unrelated to high genetic differentiation at the underlying loci.
C_LI


title: CCAR-1 has a novel role in regulating the Caenorhabditis elegans germline

abstract: 1.The Cell Division Cycle and Apoptosis Regulator (CCAR) protein family members are putative transcription regulators that have been characterized for modulating the cell cycle, apoptosis, metabolism, and the heat shock response. Mammals have two CCAR family members, CCAR1 and CCAR2/DBC1, that evolved from the founding family member CCAR-1 that is expressed in Caenorhabditis elegans. Mammalian CCAR2, the most well-studied family member, has been shown to regulate genes involved in metabolism in cultured cells. However, the regulation of gene expression by CCAR family members at an organismal level is unknown. Here, we use whole transcriptome RNA sequencing to examine the effects of CCAR-1 on gene expression in Caenorhabditis elegans. We show that CCAR-1 regulates germline transcription, reproduction, lifespan, and DNA-damage induced apoptosis. This study shows the role of CCAR-1 in vital physiological functions in the C. elegans germline that have not been investigated before.


title: MiDAS 4: A global catalogue of full-length 16S rRNA gene sequences and taxonomy for studies of bacterial communities in wastewater treatment plants

abstract: Biological wastewater treatment and an increased focus on resource recovery is fundamental for environmental protection, human health, and sustainable development. Microbial communities are responsible for these processes, but our knowledge of their diversity and function is still poor, partly due to the lack of good reference databases and comprehensive global studies. Here, we sequenced more than 5 million high-quality, full-length 16S rRNA gene sequences from 740 wastewater treatment plants (WWTPs) across the world and used the sequences to construct MiDAS 4, a full-length amplicon sequence variant resolved 16S rRNA gene reference database with a comprehensive taxonomy from the domain to species-level for all references. Using a study-independent amplicon dataset from the Global Water Microbiome Consortium project (269 WWTPs), we showed that the MiDAS 4 database provides much better coverage for bacteria in WWTPs worldwide compared to commonly applied universal references databases, and greatly improved the rate of genus and species-level classification. Hence, MiDAS 4 provides a unifying taxonomy for the majority of prokaryotic diversity in WWTPs globally, which can be used for linking microbial identities with their functions across studies. Taking advantage of MiDAS 4, we carried out an amplicon-based, global-scale microbial community profiling of activated sludge plants using two common sets of primers targeting the V1-V3 and V4 region of the 16S rRNA gene. We found that the V1-V3 primers were generally best suited for this ecosystem, and revealed how environmental conditions and biogeography shape the activated sludge microbiota. We also identified process-critical taxa (core and conditionally rare or abundant taxa), encompassing 966 genera and 1530 species. These represented approximately 80% and 50% of the accumulated read abundance, respectively, and represent targets for further investigations. Finally, we showed that for well-studied functional guilds, such as nitrifiers or polyphosphate accumulating organisms, the same genera were prevalent worldwide, with only a few abundant species in each genus.

category: microbiology


title: Impairment of a cyanobacterial glycosyltransferase that modifies a pilin results in biofilm development

abstract:
A biofilm inhibiting mechanism operates in the cyanobacterium Synechococcus elongatus. Here, we demonstrate that the glycosyltransferase homolog, Ogt, participates in the inhibitory process - inactivation of ogt results in robust biofilm formation. Furthermore, a mutational approach shows requirement of the glycosyltransferase activity for biofilm inhibition. This enzyme is necessary for glycosylation of the pilus subunit and for adequate pilus formation. In contrast to wild-type culture in which most cells exhibit several pili, only 25% of the mutant cells are piliated, half of which possess a single pilus. In spite of this poor piliation, natural DNA competence was similar to that of wild-type, therefore, we propose that the unglycosylated pili facilitate DNA transformation. Additionally, conditioned medium from wild-type culture, which contains a biofilm inhibiting substance(s), only partially blocks biofilm development by the ogt-mutant. Thus, we suggest that inactivation of ogt affects multiple processes including production or secretion of the inhibitor as well as the ability to sense or respond to it.

Originality-Significance StatementThe molecular mechanisms that underlie biofilm development in cyanobacteria are just emerging. Using the cyanobacterium S. elongatus as a model, we demonstrate that glycosylation of the pilus subunit is crucial for the biofilm self-suppression mechanism, however, it is dispensable for DNA competence.


title: Chasing the metabolism of novel syntrophic acetate-oxidizing bacteria in thermophilic methanogenic chemostats

abstract:
BackgroundAcetate is the major intermediate of anaerobic digestion of organic waste to CH4. In anaerobic methanogenic systems, acetate degradation is carried out by either acetoclastic methanogenesis or a syntrophic degradation by a syntrophy of acetate oxidizers and hydrogenotrophic methanogens. Due to challenges in isolation of syntrophic acetate-oxidizing bacteria (SAOB), the diversity and metabolism of SAOB, as well as the mechanisms of their interactions with methanogenic partners remain poorly understood.

ResultsIn this study, we successfully enriched previously unknown SAOB by operating continuous thermophilic anaerobic chemostats fed with acetate, propionate, butyrate, or isovalerate as the sole carbon and energy source. They represent novel clades belonging to Clostridia, Thermoanaerobacteraceae, Anaerolineae, and Gemmatimonadetes. In these SAOB, acetate is degraded through reverse Wood-Ljungdahl pathway or an alternative pathway mediated by the glycine cleavage system, while the SAOB possessing the latter pathway dominated the bacterial community. Moreover, H2 is the major product of the acetate degradation by these SAOB, which is mediated by [FeFe]-type electron-confurcating hydrogenases, formate dehydrogenases, and NADPH reoxidation complexes. We also identified the methanogen partner of these SAOB in acetate-fed chemostat, Methanosarcina thermophila, which highly expressed genes for CO2-reducing methanogenesis and hydrogenases to supportively consuming H2 at transcriptional level. Finally, our bioinformatical analyses further suggested that these previously unknown syntrophic lineages were prevalent and might play critical roles in thermophilic methanogenic reactors.

ConclusionThis study expands our understanding on the phylogenetic diversity and in situ biological functions of uncultured syntrophic acetate degraders, and presents novel insights on how they interact with their methanogens partner. These knowledges strengthen our awareness on the important role of SAO in thermophilic methanogenesis and may be applied to manage microbial community to improve the performance and efficiency of anaerobic digestion.


title: Isoform resolved measurements of absolute translational efficiency define interplay of HIF and mTOR dysregulation in kidney cancer

abstract: Hypoxia inducible factor (HIF) and mammalian target of rapamycin (mTOR) pathways orchestrate responses to oxygen and nutrient availability. These pathways are frequently dysregulated in cancer, but their interplay is poorly understood, in part because of difficulties in simultaneous measurement of global and mRNA-specific translation. Here we describe a workflow for measurement of absolute translational efficiency of mRNAs resolved by their transcription start sites (TSSs). Its application to kidney cancer cells revealed a remarkable extent of translational reprogramming by mTOR, strongly affecting many metabolic enzymes and pathways. By contrast, global effects of HIF on translation were limited, and we did not observe reported translational activation by HIF2A. In contrast, HIF-dependent alterations in TSS usage were associated with robust changes in translational efficiency in a subset of genes. Analyses of the interplay of HIF and mTOR revealed that specific classes of HIF1A and HIF2A transcriptional target gene manifest markedly different sensitivity to mTOR, in a manner that supports combined use of HIF2A and mTOR inhibitors in treatment of kidney cancer.


title: TAF8 regions important for TFIID lobe B assembly, or for TAF2 interactions, are required for embryonic stem cell survival

abstract: The human general transcription factor TFIID is composed of the TATA-binding protein (TBP) and 13 TBP-associated factors (TAFs). In eukaryotic cells, TFIID is thought to nucleate RNA polymerase II (Pol II) preinitiation complex formation on all protein coding gene promoters and thus, be crucial for Pol II transcription. TFIID is composed of three lobes, named A, B and C. Structural studies showed that TAF8 forms a histone fold pair with TAF10 in lobe B and participates in connecting lobe B to lobe C. In the present study, we have investigated the requirement of the different regions of TAF8 for in vitro TFIID assembly, and the importance of certain TAF8 regions for mouse embryonic stem cell (ESC) viability. We have identified a TAF8 region, different from the histone fold domain of TAF8, important for assembling with the 5TAF core complex in lobe B, and four regions of TAF8 each individually required for interacting with TAF2 in lobe C. Moreover, we show that the 5TAF coreinteracting TAF8 domain, and the proline rich domain of TAF8 that interacts with TAF2, are both required for mouse embryonic stem cell survival. Thus, our study demonstrates that distinct TAF8 regions involved in connecting lobe B to lobe C are crucial for TFIID function and consequent ESC survival.


title: Selective sorting of microRNAs into exosomes by phase-separated YBX1 condensates

abstract: Exosomes may mediate cell-to-cell communication by transporting various proteins and nucleic acids to neighboring cells. Some protein and RNA cargoes are significantly enriched in exosomes. How cells efficiently and selectively sort them into exosomes remains incompletely explored. Previously we reported that YBX1 is required in sorting of miR-223 into exosomes. Here we show that YBX1 undergoes liquid-liquid phase separation (LLPS) in vitro and in cells. YBX1 condensates selectively recruit miR-223 in vitro and into exosomes secreted by cultured cells. Point mutations that inhibit YBX1 phase separation impair the incorporation of YBX1 protein into biomolecular condensates formed in cells, and perturb miR-233 sorting into exosomes. We propose that phase separation-mediated local enrichment of cytosolic RNA binding proteins and their cognate RNAs enables their targeting and packaging by vesicles that bud into multivesicular bodies. This provides a possible mechanism for efficient and selective engulfment of cytosolic proteins and RNAs into intraluminal vesicles which are then secreted as exosomes from cells.


title: Perfluorononanoic acid impedes mouse oocyte maturation by inducing mitochondrial dysfunction and oxidative stress

abstract: Perfluorononanoic acid (PFNA), a member of PFAS, is frequently detected in human blood and tissues, even in follicular fluid of women. The exposure of PFNA, but not PFOA and PFOS, is positively correlated with miscarriage and increased time to pregnancy. Toxicological studies indicated that PFNA exposure is associated with immunotoxicity, hepatotoxicity, developmental toxicity, and reproductive toxicity in animals. However, there is little information regarding the toxic effects of PFNA on oocyte maturation. In this study, we investigated the toxic effects of PFNA exposure on mouse oocyte maturation in vitro. Our results showed that 600 M PFNA significantly inhibited germinal vesicle breakdown (GVBD) and polar body extrusion (PBE) in mouse oocytes. Our further study revealed that PFNA induced abnormal metaphase I (MI) spindle assembly, evidenced by malformed spindles and mislocalization of p-ERK1/2 in PFNA-treated oocytes. We also found that PFNA induced abnormal mitochondrial distribution and increased mitochondrial membrane potential. Consequently, PFNA increased reactive oxygen species (ROS) levels, leading to oxidative stress, DNA damage, and eventually early-stage apoptosis in oocytes. In addition, after 14 h culture, PFNA disrupted the formation of metaphase II (MII) spindle in most PFNA-treated oocytes with polar bodies. Collectively, our results indicate that PFNA interferes with oocyte maturation in vitro via disrupting spindle assembly, damaging mitochondrial functions, and inducing oxidative stress, DNA damage, and early-stage apoptosis.


title: Characterization of the Drosophila adult hematopoietic system reveals a rare cell population with differentiation and proliferation potential.

abstract: While many studies have described Drosophila embryonic and larval blood cells, the hematopoietic system of the imago remains poorly characterized and conflicting data have been published concerning adult hematopoiesis. Using a combination of blood cell markers, we show that the adult hematopoietic system is essentially composed of a few distinct mature blood cell types. In addition, our transcriptomics results indicate that adult and larval blood cells have both common and specific features and it appears that adult hemocytes reactivate many gene expressed in embryonic blood cells. Interestingly, we identify a small set of blood cells that do not express differentiation markers but maintain the progenitor marker domeMeso. Yet, we show that these cells are derived from the posterior signaling center, a specialized population of cells present in the larval lymph gland, rather than from larval blood cell progenitors, and that their maintenance depends on the EBF transcription factor Collier. Furthermore, while these cells are normally quiescent, we find that some of them can differentiate and proliferate in response to bacterial infection. In sum, our results indicate that adult flies harbor a small population of specialized cells with limited hematopoietic potential and further support the idea that no substantial hematopoiesis takes place during adulthood.


title: Downregulation of WT1 transcription factor gene expression is required to promote myocardial fate

abstract: During cardiac development, cells from the precardiac mesoderm fuse to form the primordial heart tube, which then grows by addition of further progenitors to the venous and arterial poles. In the zebrafish, wilms tumor 1 transcription factor a (wt1a) and b (wt1b) are expressed in the pericardial mesoderm at the venous pole of the forming heart tube. The pericardial mesoderm forms a single layered mesothelial sheet that contributes to further the growth of the myocardium, and forms the proepicardium. Proepicardial cells are subsequently transferred to the myocardial surface and give rise to the epicardium, the outer layer covering the myocardium in the adult heart. wt1a/b expression is downregulated during the transition from pericardium to myocardium, but remains high in proepicardial cells. Here we show that sustained wt1 expression impaired cardiomyocyte maturation including sarcomere assembly, ultimately affecting heart morphology and cardiac function. ATAC-seq data analysis of cardiomyocytes overexpressing wt1 revealed that chromatin regions associated with myocardial differentiation genes remain closed upon wt1b overexpression in cardiomyocytes, suggesting that wt1 represses a myocardial differentiation program. Indeed, a subset of wt1a/b-expressing cardiomyocytes changed their cell adhesion properties, delaminated from the myocardial epithelium, and upregulated the expression of epicardial genes, as confirmed by in vivo imaging. Thus, we conclude that wt1 acts as a break for cardiomyocyte differentiation by repressing chromatin opening at specific genomic loci and that sustained ectopic expression of wt1 in cardiomyocytes can lead to their transformation into epicardial cells.


title: C2CD6 is required for assembly of the CatSper calcium channel complex and fertilization

abstract: The CatSper cation channel is essential for sperm capacitation and male fertility. The multi-subunit CatSper complexes form highly organized calcium signaling nanodomains on flagellar membranes. Here we report identification of an uncharacterized protein C2CD6 as a novel subunit of the CatSper ion channel complex. C2CD6 contains a calcium-dependent membrane targeting C2 domain. C2CD6 interacts with the CatSper calcium-selective core forming subunits. Deficiency of C2CD6 depletes the CatSper nanodomains from the flagellum and results in male sterility. C2CD6-deficient sperm are defective in hyperactivation and fail to fertilize oocytes both in vitro and in vivo. Interestingly, transient treatments with either Ca2+ ionophore, starvation, or a combination of both restore the fertilization capacity of C2CD6-deficient sperm in vitro. C2CD6 interacts with EFCAB9, a pH-dependent calcium sensor in the CatSper complex. We postulate that C2CD6 may regulate CatSper assembly, target the CatSper complex to flagellar plasma membrane, and function as a calcium sensor. The identification of C2CD6 as an essential subunit may facilitate the long-sought reconstitution of the CatSper ion channel complex in a heterologous system for male contraceptive development.