title: A comprehensive comparison on cell type composition inference for spatial transcriptomics data

abstract:
Spatial transcriptomic (ST) technologies allow researchers to examine high-quality RNA-sequencing data along with maintained two-dimensional positional information as well as a co-registered histology image. A popular use of ST omics data is to provide insights about tissue structure and spatially unique features. However, due to the technical nature unique to most ST data, the resolution varies from a diameter of 2-10m to 50-100m instead of single-cell resolution, which brings uncertainty into cell number and cell mixture within each ST spot. Motivated by the important role for spatial arrangement of cell types within a tissue in physiology and disease pathogenesis, several ST deconvolution methods have been developed and are being used to explore gene expression variation and identification of spatial domains. The aim of this work is to review state-of-the-art methods for ST deconvolution, while comparing their strengths and weaknesses. Specifically, we use four real datasets to examine the performance of eight methods across different tissues and technological platforms.

Key PointsO_LICell mixture inference is a critical step in the analysis of spatial transcriptomics (ST) data to prevent downstream analysis suffering from confounding factors at the spot level.
C_LIO_LIExisting ST deconvolution methods can be classified into three groups: probabilistic-based, non-negative matrix factorization and non-negative least squares based, and other deep learning framework-based methods.
C_LIO_LIWe compared eight ST deconvolution methods by using two single cell level resolution datasets and two spot level resolution ST datasets. We provided practical guidelines for the choice of method under different scenarios as well as the optimal subsets of genes to use for each method.
C_LI


title: Effects of Experimental Design, Genetic Architecture and Threshold on Power and False Positive Rate of Genome-Wide Association Studies

abstract: Genome-Wide Association Studies are an important tool for identifying genetic markers associated with a trait, but it has been plagued by the multiple testing problem, which necessitates a multiple testing correction method. While many multiple testing methods have been suggested, e.g. Bonferroni and Benjamini-Hochbergs False Discovery Rate, the quality of the adjusted threshold based on these methods is not as well investigated. The aim of this study was to evaluate the balance between power and false positive rate of a Genome-Wide Association Studies experiment with Bonferroni and Benjamini-Hochbergs False Discovery Rate multiple testing correction methods and to test the effects of various experimental design and genetic architecture parameters on this balance. Our results suggest that when the markers are independent the threshold from Benjamini-Hochbergs False Discovery Rate provides a better balance between power and false positive rate in an experiment. However, with correlations between markers the threshold of Benjamini-Hochbergs False Discovery Rate becomes too lenient with an excessive number of false positives. Experimental design parameters such as sample size and number of markers used, as well as genetic architecture of a trait affect the balance between power and false positive rate. This experiment provided guidance in selecting an appropriate experimental design and multiple testing correction method when conducting an experiment.

category: genetics


title: Steering clear of the confusion effect: aerial predators target fixed points in dense prey aggregations

abstract: Collective behaviours are widely assumed to confuse predators, but empirical support for a confusion effect is often lacking, and its importance must depend on the predators targeting mechanism. Here we show that raptors steer attacks on swarming bats by turning towards a fixed point in the swarm, rather than by using closed-loop pursuit of an individual. Any prey that the predator is on a collision course with will appear on a constant bearing, so target selection emerges naturally from the geometry of a collision. Our results show how predators can simplify the demands on their sensory system by decoupling steering from targeting when grabbing prey from a dense swarm. We anticipate that the same tactic will be used against flocks and schools across a wide range of taxa, in which case a confusion effect is paradoxically more likely to occur in attacks on sparse groups, for which steering and targeting cannot be decoupled.

category: animal behavior and cognition


title: Assessing evidence accumulation and rule learning in humans with an online game

abstract:
Evidence accumulation, how the brain integrates sensory information over time, is an essential component of perception and decision making. In humans, evidence accumulation is commonly modeled as a diffusion process in which noise accumulates linearly with the incoming evidence. However, recent studies in rodents have shown that during perceptual decision making, noise scales non-linearly with the strength of accumulated evidence. The question of whether nonlinear noise scaling also holds for humans has been clouded by differences in the methodologies typically used to collect and analyze human and rodent data. For example, whereas humans are typically given explicit instructions in these tasks, rodents are trained using feedback. Therefore, to evaluate how perceptual noise scales with accumulated evidence, we developed an online evidence accumulation game and nonverbal training pipeline for humans inspired by pulse-based evidence accumulation tasks for rodents. Using this game, we collected and analyzed behavioral data from hundreds of participants trained either with an explicit description of the relevant decision rule or merely with experiential feedback. Across all participants, performance was well described by an accumulation process, in which stimuli were integrated equally across time. Participants trained using feedback alone learned the game rules rapidly and used similar strategies to those who received explicit instructions. Decisions in both groups were influenced in similar ways by biases and perceptual noise, suggesting that explicit instructions did not reduce bias or noise in pulse-based accumulation tasks. Finally, by leveraging data across all participants, we show that perceptual noise during evidence accumulation was best described by a non-linear model of noise scaling, consistent with previous animal studies, but inconsistent with diffusion models widely used in human studies. These results challenge the conventional description of humans accumulation process and suggest that online games inspired by evidence accumulation tasks provide a valuable large-scale behavioral assessment platform to examine perceptual decision making and learning in humans. In addition, the feedback-based training pipeline developed for this game may be useful for evaluating perceptual decision making in human populations with difficulty following verbal instructions.

HighlightsO_LIDevelopment and validation of an online video game to measure perceptual decision making.
C_LIO_LIHumans trained using a feedback-based pipeline exhibit similar strategies and performance compared with those receiving instructions.
C_LIO_LIPerceptual noise increases superlinearly with sensory evidence.
C_LI


title: An Improved Linear Mixed Model for Multivariate Genome-Wide Association Studies

abstract: Current methods of multivariate analysis require complete multivariate phenotypes from each individual and have a computational time complexity of O(n2) per SNP, where n is the sample size. We develop an efficient genomic multivariate analysis tool (GMAT) for genome-wide association studies of multiple correlated traits. The new method can handle incomplete multivariate data with missing records and reduce the time complexity to O(n) per SNP. Simulation studies based on known genotypes and phenotypes of actual populations show that GMAT has increased the statistical power with a proper control of false positivity for association studies compared to the conventional linear mixed model (LMM) that removes individuals with incomplete records. Applications to a balanced donkey data and an unbalanced yeast data show that the computational efficiency of the new method has been increased about tens of times faster than the conventional LMM analysis. The GMAT package can be downloaded at https://github.com/chaoning/GMAT.

category: bioinformatics


title: Single-cell Gene Regulation Network Interference by Large-scale Data Integration

abstract: Single-cell ATAC-seq (scATAC-seq) has proven to be a state-of-art approach to investigating gene regulation at the single-cell level. However, existing methods cannot precisely uncover cell-type-specific binding of transcription regulators (TRs) and construct gene regulation networks (GRNs) in single-cell. ChIP-seq has been widely used to profile TR binding sites in various species, tissues, and samples in the past decades. Here, we developed SCRIP, an integrative method to infer single-cell TR activities and targets based on the integration of scATAC-seq and public bulk ChIP-seq datasets. Our method showed improved performance in evaluating TR binding activities with similar motif information and higher consistency with matched TR expressions. Besides, our method helps in identifying TR-regulated target genes as well as building gene regulation networks (GRN) at single-cell resolution. We demonstrate SCRIPs utility in accurate cell-type clustering, lineage tracing, and inferring cell-type-specific GRNs in multiple biological systems.


title: Natural language signatures of psilocybin microdosing

abstract: Serotonergic psychedelics are being studied as novel treatments for mental health disorders and as facilitators of improved well-being, mental function and creativity. Recent studies have found mixed results concerning the effects of low doses of psychedelics ("microdosing") on these domains. However, microdosing is generally investigated using instruments designed to assess larger doses of psychedelics, which might lack sensitivity and specificity for this purpose. Following a double-blind and placebo-controlled experimental design, we explored natural language as a resource to identify speech produced under the acute effects of psilocybin microdoses, focusing on variables known to be affected by higher doses: verbosity, semantic variability and sentiment scores. Except for semantic variability, these metrics presented significant differences between a typical active microdose of 0.5 g of psilocybin mushrooms and an inactive placebo condition. Moreover, machine learning classifiers trained using these metrics were capable of distinguishing between conditions with high accuracy (AUC{approx}0.8). Our results constitute first proof that low doses of serotonergic psychedelics can be identified from unconstrained natural speech, with potential for widely applicable, affordable, and ecologically valid monitoring of microdosing schedules.

category: pharmacology and toxicology


title: GCNA is a histone binding protein required for spermatogonial stem cell maintenance

abstract: Recycling and de-novo deposition of histones during DNA replication is a critical challenge faced by eukaryotic cells and is coordinated by histone chaperones. However, little is known about how tissue-specific histone chaperones function to maintain tissue homeostasis. Here we show that Germ Cell Nuclear Acidic protein (GCNA), a germ cell specific protein in adult mice, can bind histones and purified GCNA exhibits histone chaperone activity. GCNA associates with the DNA replication machinery and supports progression through S-phase in murine spermatogonial stem cells (SSCs). Whilst GCNA is dispensable for embryonic germ cell development, it is required for the maintenance of the SSC pool and for long-term production of sperm. Our work describes the role of a germ cell specific histone chaperone in SSCs maintenance in mice. These findings provide a mechanistic basis for the male infertility observed in patients carrying mutations in the GCNA locus.

category: physiology


title: Profiling skeletal muscle-derived secretome with differentiation and acute contractile activity

abstract: Extracellular vesicles (EVs) released from all cells, are essential to cellular communication, and contain biomolecular cargo that can affect recipient cell function. Studies on the effects of contractile activity (exercise) on EVs usually rely on plasma/serum-based assessments, which contain EVs from many different cells. To specifically characterize skeletal muscle-derived vesicles and the effect of acute contractile activity, we used an in vitro model where C2C12 mouse myoblasts were differentiated to form myotubes. EVs were isolated from conditioned media from muscle cells, pre-differentiation (myoblasts) and post-differentiation (myotubes), as well as from acutely stimulated myotubes (1hr @ 14V, C-Pace EM, IonOptix) using total exosome isolation reagent (TEI, ThermoFisher, referred to as extracellular particles [EPs]) and differential ultracentrifugation (dUC; EVs). Myotube-EPs (~98 nm) were 41% smaller than myoblast-EPs (~167 nm, p<0.001, N=8-10). Two-way ANOVA showed a significant main effect for size distribution of myotube vs. myoblast-EPs (p<0.01, N=10-13). Myoblast-EPs displayed a bimodal size distribution profile with peaks at <200 nm and 400-600 nm, compared to myotube-EPs that were largely 50-300 nm in size. Total protein yield from myotube-EPs was nearly 15-fold higher than myoblast-EPs, (p<0.001 N=6-9). Similar biophysical characteristics were observed when EVs were isolated using dUC: myotube-EVs (~195 nm) remained 41% smaller in average size than myoblast-EVs (~330 nm, p=0.07, N=4-6) and had comparable size distribution profiles as EPs isolated via TEI. Myotube-EVs also had 4.7-fold higher protein yield vs. myoblast EVs (p<0.05, N=4-6). Myotube-EPs had significantly decreased expression of exosomal marker proteins TSG101, CD63, ALIX and CD81 compared to myoblast-EPs (p<0.05, N=7-12). Conversely, microvesicle marker ARF6, and lipoprotein marker APO-A1was only found in the myotube-EPs (p<0.05, N=4-12). There was no effect of acute stimulation on myotube-EP biophysical characteristics (N=7), nor on expression of TSG101, ARF6 or CD81 (N=5-6). Myoblasts treated with control or acute stimulation-derived EPs (13 g/well) for 48hrs and 72hrs showed no changes in mitochondrial mass (MitoTracker Red), cell viability or cell count (N=3-4). Myoblasts treated with EP-depleted media (72hrs) had ~90% lower cell counts (p<0.01, N=3). Our data show that EVs differ in size, distribution, protein yield and expression of subtype markers pre- vs. post-skeletal muscle differentiation. There was no effect of acute stimulation on biophysical profile or protein markers in EPs. Acute stimulation-derived EPs did not alter mitochondrial mass nor cell count/viability. Further investigation into the effects of chronic contractile activity on the biophysical characteristics and cargo of skeletal muscle-specific EVs are warranted.


title: Acute thermal stress elicits interactions between gene expression and alternative splicing in a fish of conservation concern

abstract:
Transcriptomics provides a mechanistic understanding of an organisms response to environmental challenges such as increasing temperatures, which can provide key insights into the threats posed by thermal challenges associated with urbanization and climate change. Differential gene expression and alternative splicing are two elements of the transcriptomic stress response that may work in tandem, but relatively few studies have investigated these interactions in fishes of conservation concern. We studied the imperilled redside dace (Clinostomus elongatus) as thermal stress is hypothesised to be an important cause of population declines. We tested the hypothesis that gene expression-splicing interactions contribute to the thermal stress response. Wild fish exposed to acute thermal stress were compared with both handling controls and fish sampled directly from a river. Liver tissue was sampled to study the transcriptomic stress response. Thermally stressed fish showed a prominent transcriptional response (estimated with mRNA transcript abundance) related to transcription regulation and responses to unfolded proteins, and prominent alternatively spliced genes related to gene expression regulation and metabolism. One splicing factor, prpf38b, was upregulated in the thermally stressed group compared to the other treatments. This splicing factor may have a role in the Jun/AP-1 cellular stress response, a pathway with wide-ranging and context-dependent effects. Given large gene interaction networks and the context-dependent nature of transcriptional responses, our results highlight the importance of understanding interactions between gene expression and splicing for understanding transcriptomic responses to thermal stress. Our results also reveal transcriptional pathways that can inform conservation breeding, translocation, and reintroduction programs for redside dace and other imperilled species by identifying appropriate source populations.

SUMMARY STATEMENTGene expression and alternative splicing interact in response to thermal stress in an imperilled fish, with implications for conservation and mechanisms of thermal tolerance in vertebrate ectotherms.


title: Data-Driven Flow-Map Models for Data-Efficient Discovery of Dynamics and Fast Uncertainty Quantification of Biological and Biochemical Systems

abstract: Computational models are increasingly used to investigate and predict the complex dynamics of biological and biochemical systems. Nevertheless, governing equations of a biochemical system may not be (fully) known, which would necessitate learning the system dynamics directly from, often limited and noisy, observed data. On the other hand, when expensive models are available, systematic and efficient quantification of the effects of model uncertainties on quantities of interest can be an arduous task. This paper leverages the notion of flow-map (de)compositions to present a framework that can address both of these challenges via learning data-driven models useful for capturing the dynamical behavior of biochemical systems. Data-driven flow-map models seek to directly learn the integration operators of the governing differential equations in a black-box manner, irrespective of structure of the underlying equations. As such, they can serve as a flexible approach for deriving fast-toevaluate surrogates for expensive computational models of system dynamics, or, alternatively, for reconstructing the long-term system dynamics via experimental observations. We present a data-efficient approach to data-driven flow-map modeling based on polynomial chaos Kriging. The approach is demonstrated for discovery of the dynamics of various benchmark systems and a co-culture bioreactor subject to external forcing, as well as for uncertainty quantification of a microbial electrosynthesis reactor. Such data-driven models and analyses of dynamical systems can be paramount in the design and optimization of bioprocesses and integrated biomanufacturing systems.

category: systems biology


title: Pattern reinstatement and attentional control: overlapping processes during episodic long-term memory retrieval

abstract: Episodic long-term memory (eLTM) retrieval involves the reinstatement of neural patterns from the encoding phase. However, recent evidence suggests that comparable effects on cortical activity patterns can also be linked to attentional control processes on the level of memory representations. The current investigation assesses these two processes independently based on alpha-beta-band activity in the electroencephalogram (EEG). During encoding, subjects were presented with an object on a certain position on the screen and had to imagine it on a new position. In each trial, either the task-irrelevant presentation position or the task- relevant imagination position was lateralized. In the retrieval phase, subjects first made an old/new judgement based on centrally presented objects and then reported the imagination position. Pattern reinstatement should be reflected in similar lateralized alpha-beta activity during encoding and retrieval. Conversely, the influence of attentional control processes during retrieval would be associated with the suppression of alpha-beta power contralateral to the to-be-reported imagination position and with the increase of activity contralateral to the irrelevant presentation position. Our results support this latter pattern. This shows that an experimental differentiation between selective attention and pattern reinstatement processes is necessary when studying the neural basis of eLTM retrieval on cortical level.

category: neuroscience


title: Novel Mathematical Model Based on Cellular Automata for Study of Alzheimer's Disease Progress

abstract: In recent years, extensive research has been done for the prediction, treatment, and recognition of Alzheimers disease (AD). Among these scientific works, mathematical modeling of AD is an efficient way to study the influence of various parameters such as drugs on AD progression. This paper proposes a novel model based on Cellular Automata (CA), a powerful collection of colored cells, for the investigation of AD progress. In our model, the synapses of each neuron have been considered as square cells located around the central cell. The key parameter for the progression of AD in our model is the amount of amyloid-{beta} (A{beta}), which is calculated by differential rate equations of the Puri-Li model. Based on the proposed model in this article, we introduce a new definition of AD Rate for a M x L-neuron network, which can be expanded for the whole space of the hippocampus. To better illustrate the mechanism of this model, we simulate a 3x3 neuron network and discuss the obtained results. Our numerical results show that the variations of some parameters have a great effect on AD progress. For instance, it is obtained that AD Rate is more sensitive to astroglia variations, in comparison to microglia variations. The presented model can improve the scientist's insight into the progress of AD, which will assist them to effectively consider the influence of various parameters on AD.


title: The cellular coding of temperature in the mammalian cortex

abstract: Temperature is a fundamental sensory modality separate from touch, with dedicated receptor channels and primary afferent neurons for cool and warm (Blix, 1882; Filingeri, 2016; Vriens et al., 2014). Unlike other modalities, however, the cortical encoding of temperature remains mysterious, with very few cortical neurons reported that respond to non-painful temperature and the presence of a  thermal cortex is debated (Bokiniec et al., 2018; Craig et al., 2000; Hellon et al., 1973; Milenkovic et al., 2014; Tsuboi et al., 1993). Using widefield and two-photon calcium imaging in the mouse forepaw system, here we identify cortical neurons that respond to cooling and/or warming with distinct spatial and temporal response properties. Surprisingly, we observed a representation of cool, but not warm, in the primary somatosensory cortex, but cool and warm in the posterior insular cortex (pIC). The representation of thermal information in pIC is robust, somatotopicallyarranged and reversible manipulations show a profound impact on thermal perception. Intriguingly, despite being positioned along the same one-dimensional sensory axis, the encoding of cool and warm is distinct, both in highly- and broadly- tuned neurons. Together, our results show that pIC contains the primary cortical representation of skin temperature and may help explain how the thermal system generates sensations of cool and warm.


title: Microglial TNFα controls GABAAR plasticity, slow waves and memory consolidation during sleep

abstract: Microglia sense the changes in their environment. How microglia actively translate these changes into suitable cues to adapt brain physiology is unknown. We reveal an activity-dependent regulation of cortical inhibitory synapses plasticity by microglia, driven by purinergic signaling acting on P2RX7 and mediated by microglia-derived TNF. We demonstrate that sleep induces this microglia-dependent inhibitory plasticity by promoting synaptic enrichment of GABAARs. We further show that in turn, microglia-specific depletion of TNF alters slow waves during NREM sleep and blunts sleep-dependent memory consolidation. Together, our results reveal that microglia orchestrate sleep-intrinsic plasticity of inhibitory synapses, ultimately sculpting sleep slow waves and memory.


title: Distributed network processes account for the majority of variance in localized visual category selectivity

abstract:
In seeking to understand processes fundamental to brain function, the debate between localized versus distributed processing has persisted for over a century. High category selectivity of some visual regions has provided evidence for localized (within-region) processing. However, given recent evidence that distributed connectivity patterns are predictive of localized task-evoked activations, we hypothesized that even highly localized category-selective brain activations are primarily determined by the connectivity-specified convergence of distributed brain processes. Consistent with this, we utilized fMRI data from N=352 human participants to find that distributed activity flow processes (specified by each regions unique "connectivity fingerprint") locally converge on visual cortex regions to confer the majority contribution to visual category selective responses to bodies, faces, places, and tools. This highlights a prominent role for functional network organization in the generation of localized visual category selectivity and provides evidence for a distributed convergence account of localized functionality in the human brain.

Significance StatementA fundamental debate in neuroscience has persisted for over a century with strong evidence on both sides: to what extent do localized versus distributed processes drive brain function? The existence of category-selective regions within the visual cortex provides some of the clearest evidence supporting localized processing, wherein specialized functions (e.g., category selectivity to faces) are primarily generated by within-region processes. This localized account has been countered by the identification of distributed visual cortex category selectivity in the absence of category-selective regions. Here we provide groundwork evidence for a theory of distributed convergence, demonstrating that localized visual-category-selective responses can be accurately generated via distributed activity flow processes. These predictions were generated via each category-selective regions unique connectivity fingerprint, demonstrating that the human brains distributed network architecture is sufficient to generate category selectivity in visual cortex regions that were historically seen as evidence for a primarily local account.


title: Retinoic acid-induced protein 14 controls dendritic spine dynamics associated with depressive-like behaviors

abstract: Dendritic spines are the central postsynaptic machinery that determines synaptic function. The F-actin within dendritic spines regulates their dynamic formation and elimination. Rai14 is an F-actin-regulating protein with a membrane-shaping function. Here, we identified the roles of Rai14 for the regulation of dendritic spine dynamics associated with stress-induced depressive-like behaviors. Rai14-deficient neurons failed to maintain a proper dendritic spine density in the Rai14+/- mouse brain, resulting in impaired functional synaptic activity. Rai14 was protected from degradation by complex formation with Tara, and accumulated in the dendritic spine neck, thereby enhancing spine maintenance. Concurrently, Rai14 deficiency in mice altered gene expression profile relevant to depressive conditions and increased depressive-like behaviors. Moreover, Rai14 expression was reduced in the prefrontal cortex of the mouse stress model, which was blocked by antidepressant treatment. Thus, we propose that Rai14-dependent regulation of dendritic spines may underlie the plastic changes of neuronal connections relevant to depressive-like behaviors.


title: Vigilance, arousal, and acetylcholine: Optimal control of attention in a simple detection task

abstract: Paying attention to particular aspects of the world or being more vigilant in general can be interpreted as forms of  internal action. Such arousal-related choices come with the benefit of increasing the quality and situational appropriateness of information acquisition and processing, but incur potentially expensive energetic and opportunity costs. The choices are likely implemented by widespread ascending neuromodulatory systems, including acetylcholine (ACh) and norepinephrine (NE). The key computational question that elective attention poses for sensory processing is when it is worthwhile paying these costs, and this includes consideration of whether sufficient information has yet been collected to justify the higher signal-to-noise ratio afforded by greater attention and, particularly if a change in attentional state is more expensive than its maintenance, when states of heightened attention ought to persist. We offer a partially observable Markov decision-process treatment of optional attention in a signal detection task, and use it to provide a qualitative model of the results of studies using modern techniques to measure and manipulate ACh in rodents performing a similar task.


title: A connectome-based model of conscious access in monkey cortex

abstract: A growing body of evidence suggests that conscious perception of a sensory stimulus triggers an all-or-none activity across multiple cortical areas, a phenomenon called  ignition. In contrast, the same stimulus, when undetected, induces only transient activity. In this work, we report a large-scale model of the macaque cortex based on recently quantified structural connectome data. We use this model to simulate a detection task, and demonstrate how a dynamical bifurcation mechanism produces ignition-like events in the model network. Within this framework, the model predicts that feedforward excitatory transmission is primarily mediated by the fast AMPA receptors to ensure rapid signal propagation from sensory to associative areas. In contrast, a large fraction of the inter-areal feedback projections and local recurrent excitation depend on the slow NMDA receptors, to ensure ignition of distributed frontoparietal activity. Our model predicts, counterintuitively, that fast-responding sensory areas contain a higher ratio of NMDA to AMPA receptors compared to association cortical areas that show slow, sustained activity. We validate this prediction using in-vitro receptor autoradiography data. Finally, we show how this model can account for various behavioral and physiological effects linked to consciousness. Together, these findings clarify the neurophysiological mechanisms of conscious access in the primate cortex and support the concept that gradients of receptor densities along the cortical hierarchy contribute to distributed cognitive functions.


title: Public volume electron microscopy data: An essential resource to study the brain microvasculature

abstract: Electron microscopy is the primary approach to study ultrastructural features of the cerebrovasculature. However, 2D snapshots of a vascular bed capture only a small fraction of its complexity. Recent efforts to synaptically map neuronal circuitry using volume electron microscopy have also sampled the brain microvasculature in 3D. Here, we perform a meta-analysis of 6 data sets spanning different species and brain regions, including 2 data sets from the MICrONS consortium that have made efforts to segment vasculature in addition to all parenchymal cell types in mouse visual cortex. Exploration of these data have revealed rich information for detailed investigation of the cerebrovasculature. Neurovascular unit cell types (including, but not limited to, endothelial cells, mural cells, perivascular fibroblasts, microglia, and astrocytes) could be discerned across broad microvascular zones. Image contrast was sufficient to identify subcellular details, including endothelial junctions, caveolae, peg-and-socket interactions, mitochondria, Golgi cisternae, microvilli and other cellular protrusions of potential significance to vascular signaling. Additionally, noncellular structures including the basement membrane and perivascular spaces were visible and could be traced between arterio-venous zones along the vascular wall. These explorations revealed structural features that may be important for vascular functions, such as blood-brain barrier integrity, blood flow control, brain clearance, and bioenergetics. They also identified limitations where accuracy and consistency of segmentation could be further honed by future efforts. The purpose of this article is to introduce these valuable community resources within the framework of cerebrovascular research by providing an assessment of their vascular contents, identifying features of significance for further study, and discussing next step ideas for refining vascular segmentation and analysis.


title: Multi-pulse transcranial magnetic stimulation of human motor cortex produces short-latency corticomotor facilitation via two distinct mechanisms

abstract:
BackgroundSingle-pulse transcranial magnetic stimulation of the precentral hand representation (M1HAND) can elicit indirect waves in the corticospinal tract at a periodicity of ~660 Hz, called indirect or I-waves. These synchronized descending volleys are produced by transsynaptic excitation of fastconducting monosynaptic corticospinal axons in M1-HAND. Paired-pulse TMS can induce short-interval intracortical facilitation (SICF) of motor evoked potentials (MEPs) at inter-pulse intervals that match I-wave periodicity.

ObjectiveTo examine whether short-latency corticospinal facilitation engages additional mechanisms independently of I-wave periodicity.

MethodsIn 19 volunteers, one to four biphasic TMS pulses were applied to left M1-HAND with interpulse interval was adjusted to the first peak or first trough of the individual SICF curve. TMS was applied at different intensities to probe the intensity-response relationship.

ResultsPairs, triplets, or quadruplets at individual peak-latency facilitated MEP amplitudes across a wide range of TMS intensities compared to single pulses. Multi-pulse TMSHAND at individual troughlatency also produced a consistent facilitation of MEP amplitude. Short-latency facilitation at trough-latency was less pronounced than short-latency facilitation at peak-latency, but the relative difference in facilitation decreased with increasing stimulus intensity. Increasing the number of pulses from two to four pulses had only a modest effect on MEP facilitation.

ConclusionTwo mechanisms underly short-latency corticomotor facilitation caused by biphasic multi-pulse TMS. An intracortical mechanism is related to I-wave periodicity and engages fast-conducting direct projections to spinal motoneurons. A second corticospinal mechanism does not rely on I-wave rhythmicity and may be mediated by slower conducting indirect pyramidal tract projections from M1-HAND to spinal interneurons. The latter mechanism deserves more attention in TMS studies of the corticomotor system.


title: Seasonal dynamics and polyphenism of butterfly communities in the coastal plains of central Western Ghats, India

abstract: Long-term socioeconomic progress requires a healthy environment/ecosystem, but anthropogenic activities cause environmental degradation and biodiversity loss. Constant ecological monitoring is therefore necessary to assess the state of biodiversity and ecological health. However, baseline data is lacking even for ecologically sensitive regions such as the Western Ghats. We looked at the seasonality and polyphenism of butterflies of the central Western Ghats to get baseline population patterns on these charismatic taxa. We recorded 43118 individuals (175 species) using fortnightly time-constrained counts for two consecutive years, and found the peak abundance (49% of the total individuals) in post-monsoon period (Oct to Jan). The seasonal abundance was correlated with the overall increase in species richness. Habitat differences were stronger than seasonality as samples clustered based on sites. Several species also displayed polyphenism with distinct distributions of wet and dry season forms. Seasonal equitability and indicator species analysis showed distinct inter-species differences in seasonality patterns. This work provides key baseline data on the seasonal dynamics of butterflies of the Western Ghats in the context of climate change and conservation, and will help in future monitoring of this ecologically sensitive region using butterflies.

category: ecology


title: Estimating the distribution of livestock in space and time: a time-series contribution to livestock density mapping

abstract: More than one billion people rely on livestock for income, nutrition, and social cohesion, however livestock keeping can facilitate disease transmission and contribute to climate change. While data on the distribution of livestock thus have broad utility across a range of applications, efforts to map the distribution of livestock on a large scale are limited to the Gridded Livestock of the World (GLW) project. We present a complimentary effort to map the distribution of cattle and pigs in Malawi, Uganda, Democratic Republic of Congo (DRC), and South Sudan. In contrast to GLW, which uses dasymmetric modeling applied to census data to produce time-stratified estimates of livestock counts and spatial density, our work uses complex survey data and distinct modeling methods to generate a time-series of livestock distribution, defining livestock density as the ratio of animals to humans. In addition to favorable cross-validation results and general agreement with national density estimates derived from external data on national human and livestock populations, our results demonstrate extremely good agreement with GLW-3 estimates, supporting the validity of both efforts. Our results furthermore offer a high-resolution time series result and employ a definition of density which is particularly well-suited to the study of livestock-origin zoonoses.


title: Defensive shimmering responses in the giant honeybee Apis dorsata are triggered by dark stimuli moving against a bright background

abstract: Due to the absence of physical barriers, the open-nesting giant honeybee Apis dorsata has evolved a spectacular collective defence behaviour - known as "shimmering" - against predators, which is characterised by travelling waves generated by individual bees flipping their abdomens in a coordinated and sequential manner across the bee curtain. We examined if shimmering is visually-mediated by presenting moving stimuli of varying sizes and contrasts to the background (dark or light) in bright and dim ambient light conditions. Shimmering was strongest under bright ambient light, and its strength declined in dim-light. A. dorsata shimmered only when presented with the darkest stimulus against a light background, but not when this condition was reversed (light stimulus against dark background). We suggest that this is an effective anti-predatory strategy in opennesting A. dorsata colonies, exposed to high ambient light, as flying predators are more easily detected when they appear as dark moving objects against a bright sky. Moreover, the stimulus detection threshold (smallest visual angular size) is much smaller in this anti-predatory context (1.6{degrees} - 3.4{degrees}) than in the context of foraging (5.7{degrees}), indicating that ecological context affects visual detection threshold.


title: TrieDedup: A fast trie-based deduplication algorithm to handle ambiguous bases in high-throughput sequencing

abstract:
MotivationHigh-throughput sequencing is a powerful tool and is extensively applied in biological studies. However sequencers may report bases with low qualities and lead to ambiguous bases,  Ns. PCR duplicates introduced in library preparation need to be removed in genomics studies, and several deduplication tools have been developed for this purpose. However, the existing tools cannot deal with  Ns correctly or efficiently.

ResultsHere we proposed and implemented TrieDedup, which uses trie (prefix tree) structure to compare and store sequences. TrieDedup can handle ambiguous base  Ns, and efficiently deduplicate at the level of raw sequences. We also reduced its memory usage by approximately 20% by implementing restrictedListDict. We benchmarked the performance of the algorithm and showed that TrieDedup can deduplicate reads up to 160-fold faster than pairwise comparison at a cost of 36-fold higher memory usage. TrieDedup algorithm may facilitate PCR deduplication, barcode or UMI assignment and repertoire diversity analysis of large scale high-throughput sequencing datasets with its ultra-fast algorithm that can account for ambiguous bases due to sequencing errors.

AvailabilityTrieDedup is available at https://github.com/lolrenceH/TrieDedup.


title: Network profiling of hepatocellular carcinoma targets for evidence based pharmacological approach to improve clinical efficacy

abstract:
IntroductionHepatocellular carcinoma (HCC) is the most prevalent malignancy of the liver with limited clinical efficacy of currently used drugs such as sorafenib. Hence in this study we assessed the network proteins of HCC targets to identify the target/s which can achieve optimal clinical efficacy.

Materials and MethodsThe reported HCC targets and their network proteins were identified in the string database. The interactions of the network proteins based on the number of hydrogen bonds formed were evaluated using the chimera software and used to merit the network protein interactions. The merit of network protein interactions in clinical efficacy was assessed based on the expression pattern of the network proteins and corelating their targeting by sorafenib.

Results22 potential HCC targets were identified along with their 152 unique network proteins. The following HCC targets; PDGFRB, IFNA2, VEGFR2, PD1, C-MET, RAR and IGF1R were observed to be among the top networks with the most number of hydrogen bond interactions between them. Among these, C-MET, RAR and IGF1R were significantly expressed in hepatocytes, making them relevant HCC targets. PD-1 and PD-L1, which are immune checkpoint regulators and hence used as part of immune therapy, were observed to form higher numbers of hydrogen bonds with HCC network proteins.

ConclusionOur analysis suggest that selectively targeting IGF1R, C-MET and RAR in hepatocytes together with immunotherapy will result in optimal clinical efficacy in the management of HCC.

category: cancer biology


title: A novel dimeric FAP-targeting small molecule-radio conjugate with high and prolonged tumour uptake

abstract:
Imaging procedures based on small molecule-radio conjugates (SMRCs) targeting fibroblast activation protein (FAP) have recently emerged as a powerful tool for the diagnosis of a wide variety of tumours. However, the therapeutic potential of radiolabeled FAP-targeting agents is limited by their short residence time in neoplastic lesions. In this work, we present the development and in vivo characterization of BiOncoFAP, a new dimeric FAP-binding motif with extended tumour residence time and favorable tumour-to-organ ratio.

MethodsThe binding properties of BiOncoFAP and its monovalent OncoFAP analogue were assayed against recombinant hFAP. Preclinical experiments with [177Lu]Lu-OncoFAP-DOTAGA (177Lu-OncoFAP) and [177Lu]Lu-BiOncoFAP-DOTAGA (177Lu-BiOncoFAP) were performed in mice bearing FAP-positive HT-1080 tumours.

ResultsOncoFAP and BiOncoFAP displayed comparable sub-nanomolar dissociation constants towards hFAP in solution, but the bivalent BiOncoFAP bound more avidly to the target immobilized on solid supports. In a comparative biodistribution study, 177Lu-BiOncoFAP exhibited a more stable and prolonged tumour uptake than 177Lu-OncoFAP ([~]20% ID/g vs [~]4% ID/g, at 24h p.i., respectively). Notably, 177Lu-BiOncoFAP showed favorable tumour-to-organ ratios with low kidney uptake. Both 177Lu-OncoFAP and 177Lu-BiOncoFAP displayed potent anti-tumour efficacy when administered at therapeutic doses in tumour bearing mice.

Conclusions177Lu-BiOncoFAP is a promising candidate for radioligand therapy of cancer, with favorable in vivo tumour-to-organ ratio, long tumour residence time and potent anti-cancer efficacy.


title: Transcriptional response of a target plant to benzoxazinoid and diterpene allelochemicals highlights commonalities in detoxification

abstract: Plants growing in proximity to other plants are exposed to a variety of metabolites that these neighbors release into the environment. Some species produce allelochemicals to inhibit growth of neighboring plants, which in turn have evolved ways to detoxify these compounds. In order to understand how the allelochemical-receiving target plants respond to chemically diverse compounds, we performed whole-genome transcriptome analysis of Arabidopsis thaliana exposed to either the benzoxazinoid derivative 2-amino-3H-phenoxazin-3-one (APO) or momilactone B. These two allelochemicals belong to two very different compound classes, benzoxazinoids and diterpenes, respectively, produced by different cereal crop species. Despite their distinct chemical nature, we observed similar molecular responses of A. thaliana to these allelochemicals. In particular, many of the same or closely related genes belonging to the three-phase detoxification pathway were upregulated in both treatments. Further, we observed an overlap between genes upregulated by allelochemicals and those involved in herbicide detoxification. Our findings highlight the overlap in the transcriptional response of a target plant to natural and synthetic phytotoxic compounds and illustrate how herbicide resistance could arise via pathways involved in plant-plant interaction.

category: plant biology


title: The psychotomimetic ketamine disrupts the transfer of late sensory information in the corticothalamic network

abstract: In prodromal and early schizophrenia, disorders of attention and perception are associated with structural and chemical brain abnormalities, and with dysfunctional, highly distributed corticothalamic networks exhibiting disturbed brain rhythms. The underlying mechanisms are elusive. The non-competitive NMDA receptor antagonist ketamine simulates the symptoms of prodromal and early schizophrenia, including the disturbances in ongoing and task/sensory-related broadband gamma frequency (30-80 Hz) oscillations in corticothalamic networks. In normal healthy subjects and rodents, complex integration processes, like sensory perception, induce transient, large-scale synchronized gamma oscillations in a time window of a few hundreds of ms (200-700 ms) after the presentation of the object of attention (e.g., sensory stimulation). Our goal was to use an electrophysiological multisite network approach to investigate, in lightly anesthetized rats, the effects of a single psychotomimetic low-dose (2.5 mg/kg, subcutaneous) of ketamine on sensory stimulus-induced gamma oscillations. Ketamine transiently increased the power of baseline gamma oscillations and decreased sensory-induced gamma oscillations. In addition, it disrupted information transferability in both the somatosensory thalamus and the related cortex and decreased the stimulus-induced gamma-frequency band thalamocortical connectivity. In conclusion, the present findings support the hypothesis that NMDA receptor antagonism disrupts the transfer of perceptual information in the somatosensory cortico-thalamo-cortical system.


title: A prefrontal to lateral entorhinal pathway disrupts memory.

abstract: While the neural circuits underlying memory encoding, storage, and retrieval are well characterized, the circuits that act to disrupt memory are enigmatic. Here we find that silencing a projection from the prefrontal cortex to the lateral entorhinal cortex surprisingly improves spatial working memory and contextual memory. We then found that the same cell type shows increased activity during errors in a spatial working memory test. Finally we found that optogenetic activation of the same activity patterns can disrupt working memory performance. By using a combination of intersectional genetic and in vivo imaging techniques we advance evidence that a novel prefrontal-entorhinal pathway critically participates in memory disruption.


title: Cochlear theta activity oscillates in phase opposition during interaural attention

abstract: It is widely established that sensory perception is a rhythmic process as opposed to a continuous one. In the context of auditory perception this effect is only established on a cortical and behavioral level. Yet, the unique architecture of the auditory sensory system allows its primary sensory cortex to modulate the processes of its sensory receptors at the cochlear level. Previously, we could demonstrate the existence of a genuine cochlear theta (~6 Hz) rhythm that is modulated in amplitude by intermodal selective attention. As the studys paradigm was not suited to assess attentional effects on the oscillatory phase of cochlear activity the question whether attention can also affect the temporal organization of the cochleas ongoing activity remained open. The present study utilizes an interaural attention paradigm to investigate ongoing otoacoustic activity during a stimulus-free cue-target interval and an omission period of the auditory target in humans. We were able to replicate the existence of the cochlear theta rhythm. Importantly, we found significant phase opposition between the two ears and attention conditions of anticipatory as well as cochlear oscillatory activity during target presentation. Yet, the amplitude was unaffected by interaural attention. These results are the first to demonstrate that intermodal and interaural attention deploy different aspects of excitation and inhibition at the first level of auditory processing. While intermodal attention modulates the level of cochlear activity, interaural attention modulates the timing.


title: Do we parse the background into separate streams in the cocktail party?

abstract: AO_SCPLOWBSTRACTC_SCPLOWIn the cocktail party situation, people with normal hearing usually follow a single speaker among multiple concurrent ones. However, there is no agreement in the literature as to whether the background is segregated into multiple streams/speakers. The current study varied the number of concurrent speech streams and investigated target detection and memory for the contents of a target stream as well as the processing of distractors. A male-spoken target stream was either presented alone (single-speech), together with one male-spoken (one-distractor), or a male- and a female-spoken distractor (two-distractor). Behavioral measures of target detection and content tracking performance as well as target- and distractor detection related ERPs were assessed. We found that the detection sensitivity and the target N2b amplitude decreased whereas the P3b amplitude increased from the single-speech to the concurrent speech streams conditions. Importantly, the behavioral distractor effect differed between the conditions with one- vs. two-distractor (distraction by the female speaker was lower than that of the male speaker in either condition) and the target N2b elicited in the presence of two distractors was significantly smaller than that elicited in the presence of one distractor. Further, the voltage in the N2b time window significantly differed between the one- and two-distractor conditions for the same (M2) speaker. These results show that speech processing was different in the presence of one vs. two distractors, and thus, the current data suggest that the two background speech streams were segregated from each other.


title: Visual and auditory cortices represent acoustic speech-related information during silent lip reading

abstract:
Speech is an intrinsically multisensory signal and seeing the speakers lips forms a cornerstone of communication in acoustically impoverished environments. Still, it remains unclear how the brain exploits visual speech for comprehension and previous work debated whether lip signals are mainly processed along the auditory pathways or whether the visual system directly implements speech-related processes. To probe this question, we systematically characterized dynamic representations of multiple acoustic and visual speech-derived features in source localized MEG recordings that were obtained while participants listened to speech or viewed silent speech. Using a mutual-information framework we provide a comprehensive assessment of how well temporal and occipital cortices reflect the physically presented signals and speech-related features that were physically absent but may still be critical for comprehension. Our results demonstrate that both cortices are capable of a functionally specific form of multisensory restoration: during lip reading both reflect unheard acoustic features, with occipital regions emphasizing spectral information and temporal regions emphasizing the speech envelope. Importantly, the degree of envelope restoration was predictive of lip reading performance. These findings suggest that when seeing the speakers lips the brain engages both visual and auditory pathways to support comprehension by exploiting multisensory correspondences between lip movements and spectro-temporal acoustic cues.

HighlightsO_LIVisual and auditory cortex represent unheard acoustic information during lip reading
C_LIO_LIAuditory cortex emphasizes the acoustic envelope
C_LIO_LIVisual cortex emphasizes a pitch signature
C_LIO_LITracking of unheard features in auditory cortex is associated with behavior
C_LI


title: Transcriptional profiling of transport mechanisms and regulatory pathways in rat choroid plexus

abstract:
BackgroundDysregulation of brain fluid homeostasis associates with brain pathologies in which fluid accumulation leads to elevated intracranial pressure. Surgical intervention remains standard care, since specific and efficient pharmacological treatment options are limited for pathologies with disturbed brain fluid homeostasis. Such lack of therapeutic targets originates, in part, from the incomplete map of the molecular mechanisms underlying cerebrospinal fluid (CSF) secretion by the choroid plexus.

MethodsThe transcriptomic profile of rat choroid plexus was generated by RNA Sequencing (RNAseq) of whole tissue and epithelial cells captured by fluorescence-activated cell sorting (FACS), and compared to proximal tubules. The bioinformatic analysis comprised mapping to reference genome followed by filtering for type, location, and association with alias and protein function. The transporters and associated regulatory modules were arranged in discovery tables according to their transcriptional abundance and tied together in association network analysis.

ResultsThe transcriptomic profile of choroid plexus displays high similarity between sex and species (human, rat, and mouse) and lesser similarity to another secretory epithelium, the proximal tubules. The discovery tables provide lists of transport mechanisms that could participate in CSF secretion and suggest regulatory candidates.

ConclusionsWith quantification of the transport protein abundance in choroid plexus and their potentially linked regulatory modules, we envision a molecular tool to devise rational hypotheses regarding future delineation of choroidal transport proteins involved in CSF secretion and their regulation. Our vision is to obtain pharmaceutical targets towards modulation of CSF production in pathologies involving disturbed brain water dynamics.


title: Zika virus replicates in the vagina of mice with intact interferon signaling

abstract:
Zika virus (ZIKV) is unusual among flaviviruses in its ability to spread between humans through sexual contact, as well as by mosquitoes. Sexual transmission has the potential to change the epidemiology and geographic range of ZIKV compared to mosquito-borne transmission and potentially could produce distinct clinical manifestations, so it is important to understand the host mechanisms that control susceptibility to sexually transmitted ZIKV. ZIKV replicates poorly in wild-type mice following subcutaneous inoculation, so most ZIKV pathogenesis studies use mice lacking IFN-{beta} signaling (e.g. Ifnar1-/-). However, we found that wild-type mice support ZIKV replication following intravaginal infection, although the infection remained localized to the lower female reproductive tract. Vaginal replication was not a unique property of ZIKV, as other flaviviruses that generally are restricted in wild-type mice also were able to replicate in the vagina. Vaginal ZIKV infection required a high-progesterone state (pregnancy or pre-treatment with depot medroxyprogesterone acetate (DMPA)), identifying a key role for hormonal status in susceptibility to vaginal infection. Progesterone-mediated susceptibility did not appear to result from a compromised epithelial barrier, blunted antiviral gene induction, or changes in vaginal leukocyte populations, leaving open the mechanism by which progesterone confers susceptibility to vaginal ZIKV infection. Progesterone treatment is a key component of mouse vaginal infection models for herpes simplex virus and Chlamydia, but the mechanisms by which DMPA increases susceptibility to those pathogens also remain poorly defined. Understanding how progesterone mediates susceptibility to ZIKV vaginal infection may provide insights into host mechanisms influencing susceptibility to diverse sexually transmitted pathogens.

IMPORTANCEZika virus (ZIKV) is transmitted by mosquitoes, similarly to other flaviviruses. However, ZIKV is unusual in its ability also to spread through sexual transmission. We found that ZIKV was able to replicate in the vaginas of wild-type mice, even though these mice do not support ZIKV replication by other routes, suggesting that the vagina is particularly susceptible to ZIKV infection. Vaginal susceptibility was dependent on a high progesterone state, which is a common feature of mouse vaginal infection models for other pathogens, through mechanisms that have remained poorly defined. Understanding how progesterone mediates susceptibility to ZIKV vaginal infection may provide insights into host mechanisms that influence susceptibility to diverse sexually transmitted pathogens.

category: microbiology


title: Reactive microglia and cytokine release entails microtubule remodeling from acentrosomal to centrosomal arrays

abstract: Microglia reactivity entails a large-scale remodeling of cellular geometry, but the role of the microtubule cytoskeleton during these changes remains unexplored. Here we show that reactive proinflammatory microglia provide a heretofore unique example of microtubule reorganization from a non-centrosomal array of parallel and stable microtubules to a radial array of more dynamic microtubules. While in the homeostatic state microglia nucleate microtubules at Golgi outposts, proinflammatory signaling induces recruitment of nucleating material nearby the centrosome and inhibition of centrosomal maturation enhances NLRP3 inflammasome activation and secretion of IL-1{beta}. Our results demonstrate that a hallmark of microglia reactivity is a striking remodeling of the microtubule cytoskeleton and suggest that pericentrosomal microtubule nucleation may serve as a distinct marker of microglia activation as well as a novel target to modulate cytokine-mediated inflammatory responses in chronic disease and tissue injury.

category: cell biology


title: Microglia reactivity entails microtubule remodeling from acentrosomal to centrosomal arrays


title: Inositol depletion regulates phospholipid metabolism and activates stress signaling in HEK293T cells

abstract: Inositol plays a significant role in cellular function and signaling. Studies in yeast have demonstrated an "inositol-less death" phenotype, suggesting that inositol is an essential metabolite. In yeast, inositol synthesis is highly regulated, and inositol levels have been shown to be a major metabolic regulator, with its abundance affecting the expression of hundreds of genes. Abnormalities in inositol metabolism have been associated with several human disorders. Despite its importance, very little is known about the regulation of inositol synthesis and the pathways regulated by inositol in human cells. The current study aimed to address this knowledge gap. Knockout of ISYNA1 (encoding myo-inositol-3-P synthase 1) in HEK293T cells generated a human cell line that is deficient in de novo inositol synthesis. ISYNA1-KO cells exhibited inositol-less death when deprived of inositol. Lipidomic analysis identified inositol depletion as a global regulator of phospholipid levels in human cells, including downregulation of phosphatidylinositol (PI) and upregulation of the phosphatidylglycerol (PG)/cardiolipin (CL) branch of phospholipid metabolism. RNA-Seq analysis revealed that inositol depletion induced substantial changes in the expression of genes involved in cell signaling, including extracellular signal-regulated kinase (ERK), and genes controlling amino acid transport and protein processing in the endoplasmic reticulum (ER). This study provides the first in-depth characterization of the effects of inositol depletion on phospholipid metabolism and gene expression in human cells, establishing an essential role for inositol in maintaining cell viability and regulating cell signaling and metabolism.


title: Exploring the phycosphere of Emiliania huxleyi: from bloom dynamics to microbiome assembly experiments

abstract: Coccolithophores have global ecological and biogeochemical significance as the most important calcifying marine phytoplankton group. The structure and selection of prokaryotic communities associated with the most abundant coccolithophore and bloom-forming species, Emiliania huxleyi, are still poorly known. In this study, we assessed the diversity of bacterial communities associated with an E. huxleyi bloom in the Celtic Sea, exposed axenic E. huxleyi cultures to prokaryotic communities derived from bloom and non-bloom conditions and followed the dynamics of their microbiome composition over one year. Bloom-associated prokaryotic communities were dominated by SAR11, Marine group II Euryarchaeota, Rhodobacterales and contained substantial proportions of known indicators of phytoplankton bloom demises such as Flavobacteriaceae and Pseudoalteromonadaceae. Taxonomic richness of replicated co-cultures resulting from natural communities with axenic E. huxleyi rapidly shifted and then stabilized over time, presumably by ecological selection favoring more beneficial populations. Recruited microbiomes from the environment were consistently dependent on the composition of the initial bacterioplankton community. Phycosphere-associated communities derived from the E. huxleyi bloom depth were highly similar to one another, suggesting deterministic processes, whereas cultures from non-bloom conditions show an effect of both deterministic processes and stochasticity. Overall, this work sheds new light on the importance of the initial inoculum composition in microbiome recruitment and elucidates the temporal dynamics of its composition and long-term stability.


title: Hormone levels increase in response to video playback in the Siamese fighting fish

abstract: The mechanisms of action of hormones regulating aggression in fish remain poorly understood. One possibly confounding variable is the lack of standardization in the type of stimuli used to elicit aggression. The presentation of controlled stimuli in videos, a.k.a. video playback, can provide better control of the fight components. However, this technique has produced conflicting results in animal behaviour studies and needs to be carefully validated. Here, the response of male Betta splendens to a matched for size conspecific fighting behind a one-way mirror presented live or through video playback was compared. Despite their non-interactive nature, both stimuli elicited a strong aggressive response by focal fish, which started with stereotypical threat displays and progressed to overt attacks. Overall, the frequency and duration of aggressive behaviours and swimming activity were similar towards live and video stimuli. Post-fight plasma levels of the androgen 11-ketotestosterone were elevated as compared to controls, regardless of the type of stimuli. Cortisol also increased in response to the video images, as previously described for interactive live fights in this species. The study shows for the first time in a fish a robust endocrine response to video stimuli and supports the use of this technique for researching aggressive behaviour in B. splendens.


title: Integrating phylogenetic and functional data in microbiome studies

abstract: Microbiome functional data are frequently analyzed to identify associations between microbial gene families and sample groups of interest. This is most often performed with approaches focused on the metagenome-wide relative abundance of microbial functions. Although such approaches can provide valuable insights, it is challenging to distinguish between different possible explanations for variation in community-wide functional profiles by considering functions alone. To help address this problem, we have developed a novel, phylogeny-aware framework to expand taxonomic balance tree-based approaches to identify enriched functions more robustly. The key focus of our approach, termed POMS, is on identifying functions that are consistently enriched in sample groups across independent taxonomic lineages. Based on simulated data we demonstrate that POMS can more accurately identify gene families that confer a selective advantage compared with commonly used differential abundance approaches. We also show that POMS can identify enriched functions in real-world metagenomics datasets that are potential targets of strong selection on multiple members of the microbiome. While this framework may not be able to identify all potential functional enrichments, the enrichments it does identify are more interpretable and conservative compared with those identified by existing differential abundance approaches. More generally, POMS is a novel approach for exploring microbiome functional data, which could be used to complement standard analyses. POMS is freely available as an R package at: https://github.com/gavinmdouglas/POMS.


title: Discovering genomic regions associated with the phenotypic differentiation of European local pig breeds

abstract:
BackgroundIntensive selection of modern pig breeds resulted in genetic improvement of productive traits while local pig breeds remained less performant. As they have been bred in extensive systems, they have adapted to specifical environmental conditions resulting in a rich genotypic and phenotypic diversity. In this study, European local pig breeds were genotypically and phenotypically characterised using DNA-pool sequencing data and breed level phenotypes related to stature, fatness, growth and reproductive performance traits. These data were analysed using a dedicated approach to detect selection signatures linked to phenotypic traits in order to uncover potential candidate genes that may be under adaptation to specific environments.

ResultsGenetic data analysis of European pig breeds revealed four main axes of genetic variation represented by Iberian and modern breeds (i.e. Large White, Landrace, and Duroc). In addition, breeds clustered according to their geographical origin, for example French Gascon and Basque breeds, Italian Apulo Calabrese and Casertana breeds, Spanish Iberian and Portuguese Alentejano breeds. Principal component analysis of phenotypic data distinguished between larger and leaner breeds with better growth potential and reproductive performance on one hand and breeds that were smaller, fatter, and had low growth and reproductive efficiency on the other hand. Linking selection signatures with phenotype identified 561 significant genomic regions. Among them, several regions contained candidate genes with possible biological effect on stature, fatness, growth and reproduction performance traits. For example, strong associations were found for stature in two regions containing ANXA4 and ANTXR1 genes, for growth performance containing TLL1 gene, for fatness containing DNMT3A and POMC genes and for reproductive performance containing HSD17B7 gene.

ConclusionsThe present study on European local pig breeds used a newly developed approach for searching selection signatures supported by phenotypic data at the breed level to identify potential candidate genes that may have adapted to different living environments and production systems. Results can be useful to define conservation programs of local pig breeds.


title: Modular capsid decoration boosts adenovirus vaccine-induced humoral and cellular immunity against SARS-CoV-2

abstract:
Adenovirus vector vaccines have been widely and successfully deployed in response to COVID-19. However, despite inducing potent T cell immunity, improvement of vaccine-specific antibody responses upon homologous boosting is modest compared to other technologies. Here, we describe a system to enable modular decoration of adenovirus capsid surfaces with protein antigens and demonstrate induction of potent humoral immunity against these displayed antigens. Ligand attachment via a covalent isopeptide bond was achieved in a rapid and spontaneous reaction, requiring simple co-incubation of ligand and vector components. We used a recently described protein superglue, DogTag/DogCatcher, which is similar to the widely used SpyTag/SpyCatcher ligation system but performs better in loop structures. DogTag was inserted into surface-exposed loops in the adenovirus hexon protein to allow attachment of DogCatcher-fused ligands on virus particles. Efficient coverage of the capsid surface was achieved using a variety of ligands and vector infectivity was retained in each case. Capsid decoration shielded particles from anti-adenovirus neutralizing antibodies. In prime-boost regimens, proof-of-concept COVID-19 adenovirus vaccines decorated with the receptor-binding domain (RBD) of SARS-CoV-2 spike induced >10-fold higher SARS-CoV-2 neutralization titers compared to an undecorated adenovirus vector encoding spike. Importantly, decorated vectors retained robust T cell immunogenicity to encoded antigens, a key hallmark of adenovirus vector vaccines. We propose capsid decoration via protein superglue-mediated covalent ligation as a novel strategy to improve the efficacy and boostability of adenovirus-based vaccines and therapeutics.

One Sentence SummaryDecorating the capsid surface of adenovirus vaccine vectors using a spontaneous protein superglue induces potent pathogen-specific immunity

category: bioengineering


title: Response dynamics of spiking network models to incoming seizure-like perturbation.

abstract:
Epilepsies are characterized by electrophysiological crises in the brain, which were first observed thanks to electroencephalograms. However, it is known that seizures originating from one or more specific regions may or may not spread to the rest of the brain, while the exact mechanisms are unclear. We propose three computational models at the neural network scale to study the underlying dynamics of seizure propagation, understand which specific features play a role, and relate them to clinical or experimental observations. We consider both network features, such as the internal connectivity structure and single neuron model, and input properties in our characterization. We show that a paroxysmal input leads to a dynamical heterogeneity inside the network, non-trivially related with its architecture, which may or may not entrain it into a seizure. Although hard to anticipate because of the intricate nature of the instability involved, the seizure propagation might be circumvented upon acting on the network during a specific time window. As we deal with a complex system, which seems to depend non trivially on various parameters, we propose a probabilistic approach to the propagative/non-propagative scenarios, which may serve as a guide to control the seizure by using appropriate stimuli.

SignificanceOur computational study shows the specific role that the inhibitory population can have and the possible dynamics regarding the propagation of seizure-like behavior in three different neuronal networks. The study conducts in this paper results from the combination of structural aspects and time-continuous measures, which helps us unravel the internal dynamics of the network. We show the existence of a specific time window favorable to the reversal of the seizure propagation.


title: Granulocyte Colony Stimulating Factor causes cerebellar deficits and anxiety in a mouse model of CSF-1 receptor-related leukoencephalopathy

abstract: Colony stimulating factor (CSF) receptor-1 (CSF-1R)-related leukoencephalopathy (CRL) is an adult-onset, demyelinating neurodegenerative disease caused by autosomal dominant mutations in CSF1R, modeled by the Csf1r+/- mouse. The expression of Csf2, encoding granulocyte- macrophage CSF (GM-CSF) and of Csf3, encoding granulocyte CSF (G-CSF), are elevated in both mouse and human CRL brains. While monoallelic targeting of Csf2 has been shown to attenuate many behavioral and histological deficits of mouse CRL, including cognitive dysfunction and demyelination, the contribution of Csf3 has not been explored. In this manuscript, we investigate the behavioral, electrophysiological and histopathological phenotypes of CRL mice following monoallelic targeting of Csf3. We show that Csf3 heterozygosity normalized the Csf3 levels in Csf1r+/- mouse brains and ameliorated anxiety-like behavior, motor coordination and social interaction deficits, but not their cognitive impairment. Consistent with this, Csf3 heterozygosity attenuated microglial activation in the cerebellum and in the ventral but not in the dorsal hippocampus. Csf3 heterozygosity also failed to prevent demyelination. Csf1r+/- mice exhibited altered synaptic activity in the deep cerebellar nuclei (DCN) associated with increased deposition of the complement factor C1q on glutamatergic synapses and with increased engulfment of glutamatergic synapses by DCN microglia. These phenotypes were significantly ameliorated by monoallelic deletion of Csf3. Our findings indicate that G-CSF and GM-CSF play non-overlapping roles in mouse CRL development and suggest that G-CSF could be an additional therapeutic target in CRL.


title: SARS-CoV-2 Viroporins Activate The NLRP3-Inflammasome Via The Mitochondrial Permeability Transition Pore

abstract: Cytokine storm precipitated by activation of the host innate immune defenses is a major cause of COVID19 death. To elucidate how SARS-CoV-2 initiates this inflammatory process, we studied viroporin proteins E and Orf3a (2-E+2-3a). Expression of 2-E+2-3a in human 293T cells resulted in increased cytosolic Ca++ and then elevated mitochondrial Ca++, taken up through the MUCi11-sensitive mitochondrial calcium uniporter (MCU). Increased mitochondrial Ca++ resulted in stimulation of mitochondrial reactive oxygen species (mROS) production, which was blocked by mitochondrially-targeted catalase or MnTBAP. To determined how mROS activates the inflammasome, we transformed 293T cells with NLRP3, ASC, pro-caspase-1 and pro-IL-1{beta} plus used THP1 derived macrophages to monitor the secretion of mature IL-1{beta}. This revealed that mROS activates a factor that is released via the NIM811-sensitive mitochondrial permeability pore (mtPTP) to activate the inflammasome. Hence, interventions targeting mROS and the mtPTP may mitigate the severity of COVID19 cytokine storms.

category: immunology


title: Alterations in SARS-CoV-2 Omicron and Delta peptides presentation by HLA molecules

abstract: The T-cell immune response is a major determinant of effective SARS-CoV-2 clearance. Here, using the recently developed T-CoV bioinformatics pipeline (https://t-cov.hse.ru) we analyzed the peculiarities of the viral peptide presentation for the Omicron, Delta and Wuhan variants of SARS-CoV-2. First, we showed the absence of significant differences in the presentation of SARS-CoV-2-derived peptides by the most frequent HLA class I/II alleles and the corresponding HLA haplotypes. Then, the analysis was limited to the set of peptides originating from the Spike proteins of the considered SARS-CoV-2 variants. The major finding was the destructive effect of the Omicron mutations on PINLVRDLPQGFSAL peptide, which was the only tight binder from the Spike protein for HLA-DRB1*03:01 allele and some associated haplotypes. Specifically, we predicted a dramatical decline in binding affinity of HLA-DRB1*03:01 and this peptide after N211 deletion, L212I substitution and EPE 212-214 insertion. The computational prediction was experimentally validated by ELISA with the use of corresponding thioredoxin-fused peptides and recombinant HLA-DR molecules. Another finding was the significant reduction in the number of tightly binding Spike peptides for HLA-B*07:02 HLA class I allele (both for Omicron and Delta variants). Overall, the majority of HLA alleles and haplotypes was not significantly affected by the mutations, suggesting the maintenance of effective T-cell immunity against the Omicron and Delta variants. Finally, we introduced the Omicron variant to T-CoV portal and added the functionality of haplotype-level analysis to it.


title: Efficient recall of Omicron-reactive B cell memory after a third dose of SARS-CoV-2 mRNA vaccine

abstract:
Despite a clear role in protective immunity, the durability and quality of antibody and memory B cell responses induced by mRNA vaccination, particularly by a 3rd dose of vaccine, remains unclear. Here, we examined antibody and memory B cell responses in a cohort of individuals sampled longitudinally for [~]9-10 months after the primary 2-dose mRNA vaccine series, as well as for [~]3 months after a 3rd mRNA vaccine dose. Notably, antibody decay slowed significantly between 6- and 9-months post-primary vaccination, essentially stabilizing at the time of the 3rd dose. Antibody quality also continued to improve for at least 9 months after primary 2-dose vaccination. Spike- and RBD-specific memory B cells were stable through 9 months post-vaccination with no evidence of decline over time, and [~]40-50% of RBD-specific memory B cells were capable of simultaneously recognizing the Alpha, Beta, Delta, and Omicron variants. Omicron-binding memory B cells induced by the first 2 doses of mRNA vaccine were boosted significantly by a 3rd dose and the magnitude of this boosting was similar to memory B cells specific for other variants. Pre-3rd dose memory B cell frequencies correlated with the increase in neutralizing antibody titers after the 3rd dose. In contrast, pre-3rd dose antibody titers inversely correlated with the fold-change of antibody boosting, suggesting that high levels of circulating antibodies may limit reactivation of immunological memory and constrain further antibody boosting by mRNA vaccines. These data provide a deeper understanding of how the quantity and quality of antibody and memory B cell responses change over time and number of antigen exposures. These data also provide insight into potential immune dynamics following recall responses to additional vaccine doses or post-vaccination infections.

Graphical Summary

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title: Omicron booster in ancestral strain vaccinated mouse augments protective immunities against both the Delta and Omicron variants

abstract: A booster vaccination is called for constraining the evolving epidemic of SARS-CoV-2. However, the necessity of a new COVID-19 vaccine is currently unclear. To compare the effect of an Omicron-matched S DNA vaccine and an ancestral S DNA vaccine in boosting cross-reactive immunities, we firstly immunized mice with two-dose of a DNA vaccine encoding the spike protein of the ancestral Wuhan strain. Then the mice were boosted with DNA vaccines encoding spike proteins of either the Wuhan strain or the Omicron variant. Specific antibody and T cell responses were measured at 4 weeks post boost. Our data showed that the Omicron-matched vaccine efficiently boosted RBD binding antibody and neutralizing antibody responses against both the Delta and the Omicron variants. Of note, antibody responses against the Omicron variant elicited by the Omicron-matched vaccine were much stronger than those induced by the ancestral S DNA vaccine. Meanwhile, CD8+ T cell responses against both the ancestral Wuhan strain and the Omicron strain also tended to be higher in mice boosted by the Omicron-matched vaccine than those in mice boosted with the ancestral S DNA vaccine, albeit no significant difference was observed. Our findings suggest that an Omicron-matched vaccine is preferred for boosting cross-reactive immunities.


title: Omicron booster in ancestral strain vaccinated mice augments protective immunities against both the Delta and Omicron variants


title: Identification of a SARS-CoV-2 host metalloproteinase-dependent entry pathway differentially used by SARS-CoV-2 and variants of concern Alpha, Delta, and Omicron

abstract: To infect cells, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) binds to angiotensin converting enzyme 2 (ACE2) via its spike glycoprotein (S), delivering its genome upon S-mediated membrane fusion. SARS-CoV-2 uses two distinct entry pathways: 1) a surface, serine protease-dependent or 2) an endosomal, cysteine protease-dependent pathway. In investigating serine protease-independent cell-cell fusion, we found that the matrix metalloproteinases (MMPs), MMP2/9, can activate SARS-CoV-2 S fusion activity, but not that of SARS-CoV-1. Importantly, metalloproteinase activation of SARS-CoV-2 S represents a third entry pathway in cells expressing high MMP levels. This route of entry required cleavage at the S1/S2 junction in viral producer cells and differential processing of variants of concern S dictated its usage. In addition, metalloproteinase inhibitors reduced replicative Alpha infection and abrogated syncytia formation. Finally, we found that the Omicron S exhibit reduced metalloproteinase-dependent fusion and viral entry. Taken together, we identified a MMP2/9-dependent mode of activation of SARS-CoV-2 S. As MMP2/9 are released during inflammation and severe COVID-19, they may play important roles in SARS-CoV-2 S-mediated cytopathic effects, tropism, and disease outcome.


title: Targeted Down Regulation Of Core Mitochondrial Genes During SARS-CoV-2 Infection

abstract:
Defects in mitochondrial oxidative phosphorylation (OXPHOS) have been reported in COVID-19 patients, but the timing and organs affected vary among reports. Here, we reveal the dynamics of COVID-19 through transcription profiles in nasopharyngeal and autopsy samples from patients and infected rodent models. While mitochondrial bioenergetics is repressed in the viral nasopharyngeal portal of entry, it is up regulated in autopsy lung tissues from deceased patients. In most disease stages and organs, discrete OXPHOS functions are blocked by the virus, and this is countered by the host broadly up regulating unblocked OXPHOS functions. No such rebound is seen in autopsy heart, results in severe repression of genes across all OXPHOS modules. Hence, targeted enhancement of mitochondrial gene expression may mitigate the pathogenesis of COVID-19.

One-Sentence SummaryCovid-19 is associated with targeted inhibition of mitochondrial gene transcription.

category: molecular biology


title: Blockage of Lamin-A/C loss diminishes the pro-inflammatory macrophage response

abstract: Mutations and defects in nuclear lamins can cause major pathologies in affected tissues. Recent studies have also established potential links between lamins, inflammation, and inflammatory diseases but the underlying molecular mechanisms are unknown. We now report that pro-inflammatory activation of macrophages reduces levels of Lamin-A/C to augment pro-inflammatory gene expression and cytokine secretion. We show that activation of bone-marrow derived macrophages (BMDMs) degrades Lamin-A/C, as preceded by its phosphorylation, which is mediated by Caspase-6 and CDK1, respectively. Inhibiting Lamin-A/C phosphorylation and degradation severely inhibits pro-inflammatory gene expression and cytokine secretion. Using LPS-activated Lamin-A/C Knock Out BMDMs, we confirmed that the activation of the IFN-{beta}-STAT pathway is amplified due to Lamin-A/C reduction, which ultimately augments the pro-inflammatory response. As Lamin-A/C is a previously unappreciated regulator of the pro-inflammatory macrophage response, our findings could provide novel opportunities to treat inflammatory diseases. In first proof-of-concept studies we show that macrophage pro-inflammation, as induced by Lipopolysaccharide or pathogenic E. coli, could be reduced by inhibiting Lamin-A/C phosphorylation and degradation. The inhibition of macrophage pro-inflammation could also be achieved by inhibiting members of the Lamin-A/C regulated IFN-{beta}-STAT pathway, i.e., phospho-STAT1 and phospho-STAT3. This newly found mechanism to suppress the pro-inflammatory response of macrophages will provoke a re-thinking of how inflammation can be addressed therapeutically.


title: Development of a physiological insulin resistance model in human stem cell-derived adipocytes

abstract:
Adipocytes are key regulatory cells of human metabolism, and their dysfunction in insulin signaling is central to metabolic diseases such as type II diabetes mellitus (T2D). However, the progression of insulin resistance that leads to T2D is still poorly understood. This limited understanding is due, in part, to the dearth of suitable models of insulin signaling in human adipocytes. Traditionally, in vitro adipocyte models fail to recapitulate in vivo insulin signaling, possibly due to exposure to supraphysiological nutrient and hormone conditions. Here, we have developed a sensitization protocol for human pluripotent stem cell-derived adipocytes that uses physiologically relevant nutrient conditions to produce a potent signaling response comparable to in vivo adipocytes. After systematically optimizing conditions, this protocol allows for robust insulin-stimulated glucose uptake and transcriptional insulin response. Furthermore, exposure of these sensitized adipocytes to physiologically relevant hyperinsulinemic conditions dampens insulin-stimulated glucose uptake and dysregulates transcription of insulin-responsive genes. Overall, this sensitization methodology provides a novel platform for the mechanistic study of insulin signaling and resistance using human pluripotent stem cell-derived adipocytes.

TeaserA new protocol to generate hPSC-adipocytes that respond to physiological insulin levels and can model diabetes.


title: Cell cycle-dependent binding between Cyclin B1 and Cdk1 revealed by time-resolved Fluorescence Correlation Spectroscopy

abstract:
Measuring the dynamics with which the regulatory complexes of the cell cycle machinery assemble and disassemble is a crucial barrier to our understanding that until now has been difficult to address. This considerable gap in our understanding is due to the difficulty of reconciling biochemical assays with single cell-based techniques, but recent advances in microscopy and gene editing techniques now enable the measurement of protein-protein interaction kinetics in living cells.

Here, we apply Fluorescence Correlation Spectroscopy (FCS) and Fluorescence Cross-Correlation Spectroscopy (FCCS) to study the dynamics of the cell cycle machinery, beginning with Cyclin B1 and its binding to its partner kinase Cdk1 that together form the major mitotic kinase. Although Cyclin B1 and Cdk1 are known to bind with high affinity, our results reveal that in living cells there is a pool of Cyclin B1 that is not bound to Cdk1. Furthermore, we provide evidence that the affinity of Cyclin B1 for Cdk1 increases during the cell cycle, indicating that the assembly of the complex is a regulated step. Our work lays the groundwork for studying the kinetics of protein complex assembly and disassembly during the cell cycle in living cells.


title: Katnip is required to maintain microtubule function and lysosomal delivery to autophagosomes and phagosomes

abstract:
The efficient delivery of lysosomes is essential for many cell functions, such as the degradation of unwanted intracellular components by autophagy and the killing and digestion of extracellular microbes within phagosomes. Using the amoeba Dictyostelium discoideum we find that cells lacking Katnip (Katanin interacting protein) have a general defect in lysosome delivery and whilst able to make autophagosomes and phagosomes correctly are then unable to digest them.

Katnip is largely unstudied yet highly conserved across evolution. Previously studies found Katnip mutations in animals cause defects in cilia structure. Here we show that Katnip plays a general role in maintaining microtubule function. We find that loss of Katnip has no overall effect on microtubule dynamics or organisation, but is important for the transport and degradation of endocytic cargos. Strikingly, Katnip mutants become highly sensitive to GFP-tubulin expression, which leads to microtubule tangles, defective anaphase extension and slow growth. Our findings establish a conserved role for Katnip in the function of all microtubules, not just cilia as previously reported. We speculate this is via a key function in microtubule repair, required to maintain endosomal trafficking and lysosomal degradation.


title: Impaired Lef1 activation accelerates iPSC-derived keratinocytes differentiation in Progeria

abstract: Hutchinson-Gilford progeria syndrome (HGPS) is a detrimental premature aging disease caused by a point mutation in the human LMNA gene. This mutation results in the abnormal accumulation of a truncated pre-lamin A protein called progerin. Among the drastically accelerated signs of aging in HGPS patients, severe skin phenotypes such as alopecia and sclerotic skins always develop with the disease progression. Here, we study the HGPS molecular mechanisms focusing on early skin development by differentiating patient-derived induced pluripotent stem cells (iPSCs) to a keratinocyte lineage. Interestingly, HGPS iPSCs showed an accelerated commitment to the keratinocyte lineage than the normal control. To study potential signaling pathways that accelerated skin development in HGPS, we investigated the WNT pathway components during HGPS iPSCs-keratinocytes induction. Surprisingly, despite the unaffected {beta}-catenin activity, the expression of a critical WNT transcription factor LEF1 was diminished from an early stage in HGPS iPSCs-keratinocytes differentiation. Chromatin immunoprecipitation (ChIP) experiment further revealed strong bindings of LEF1 to the early-stage epithelial developmental markers K8 and K18 and that the LEF1 silencing by siRNA down-regulates the K8/K18 transcription. During the iPSCs-keratinocytes differentiation, correction of HGPS mutation by Adenine base editing (ABE), while in a partial level, rescued the phenotypes for accelerated keratinocyte lineage-commitment. ABE also reduced the cell death in HGPS iPSCs-derived keratinocytes. These findings brought new insight into the molecular basis and therapeutic application for the skin abnormalities in HGPS.

category: developmental biology


title: Estrogen suppresses DMRT1 expression during ovarian development in the chicken

abstract:
Primary sex determination is the choice between two different developmental fates, a testis or an ovary. This selection is thought to require the action of a master regulator gene that triggers expression of a gene cascade in the bipotential gonad precursor in one sex. The selection of a particular developmental pathway is also thought to suppress the alternative developmental pathway.

In birds, where the male is the homogametic sex (ZZ) and females the heterogametic sex (ZW), the Z-linked transcription factor DMRT1 is considered the master regulator and has been shown to be essential for testis development, and to also inhibit the ovarian pathway. Here, we characterise in detail, DMRT1 transcription and protein levels during gonadal development in the chick. Our analysis suggests that DMRT1 protein levels are equivalent in male and female gonads during the bipotential phase of development, and that DMRT1 protein levels are reduced in the developing ovary during the differentiation phase. The reduction in DMRT1 protein levels in the somatic cells of the female medulla, coincides with the induction of aromatase expression and the initiation of estrogen synthesis. Analysis of sex-reversed gonads and mixed-sex chimeric gonads, suggests that the reduction in DMRT1 protein is due to inhibition of DMRT1 expression by estrogen.

Our data suggests that estrogen signalling is involved in primary sex determination by regulation of DMRT1 protein expression.


title: Zebrafish fin regeneration requires generic and regeneration-specific responses of osteoblasts to trauma

abstract:
Successful regeneration requires the coordinated execution of multiple cellular responses to injury. In amputated zebrafish fins, mature osteoblasts dedifferentiate, migrate towards the injury and form proliferative osteogenic blastema cells. We show that osteoblast migration is preceded by cell elongation and alignment along the proximodistal axis, which require actomyosin, but not microtubule turnover. Surprisingly, osteoblast dedifferentiation and migration can be uncoupled. Using pharmacological and genetic interventions, we found that NF-{kappa}B and retinoic acid signalling regulate dedifferentiation without affecting migration, while the complement system and actomyosin dynamics are required for migration but not dedifferentiation. Furthermore, by removing bone at two locations within a fin ray, we established a trauma model containing two injury sites. We found that osteoblasts dedifferentiate at and migrate towards both sites, while accumulation of osteogenic progenitor cells and regenerative bone formation only occur at the distal-facing injury. Together, these data indicate that osteoblast dedifferentiation and migration represent generic injury responses that are differentially regulated and can occur independently of each other and of regenerative growth. Successful bone regeneration appears to require the coordinated execution of generic and regeneration-specific responses of osteoblast to trauma.

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title: Complete sequence of a 641-kb insertion of mitochondrial DNA in the Arabidopsis thaliana nuclear genome

abstract:
Intracellular transfers of mitochondrial DNA continue to shape nuclear genomes. Chromosome 2 of the model plant Arabidopsis thaliana contains one of the largest known nuclear insertions of mitochondrial DNA (numts). Estimated at over 600 kb in size, this numt is larger than the entire Arabidopsis mitochondrial genome. The primary Arabidopsis nuclear reference genome contains less than half of the numt because of its structural complexity and repetitiveness. Recent datasets generated with improved long-read sequencing technologies (PacBio HiFi) provide an opportunity to finally determine the accurate sequence and structure of this numt. We performed a de novo assembly using sequencing data from recent initiatives to span the Arabidopsis centromeres, producing a gap-free sequence of the Chromosome 2 numt, which is 641-kb in length and has 99.933% nucleotide sequence identity with the actual mitochondrial genome. The numt assembly is consistent with the repetitive structure previously predicted from fiber-based fluorescent in situ hybridization. Nanopore sequencing data indicate that the numt has high levels of cytosine methylation, helping to explain its biased spectrum of nucleotide sequence divergence and supporting previous inferences that it is transcriptionally inactive. The original numt insertion appears to have involved multiple mitochondrial DNA copies with alternative structures that subsequently underwent an additional duplication event within the nuclear genome. This work provides insights into numt evolution, addresses one of the last unresolved regions of the Arabidopsis reference genome, and represents a resource for distinguishing between highly similar numt and mitochondrial sequences in studies of transcription, epigenetic modifications, and de novo mutations.

Significance statementNuclear genomes are riddled with insertions of mitochondrial DNA. The model plant Arabidopsis has one of largest of these insertions ever identified, which at over 600-kb in size represents one of the last unresolved regions in the Arabidopsis genome more than 20 years after the insertion was first identified. This study reports the complete sequence of this region, providing insights into the origins and subsequent evolution of the mitochondrial DNA insertion and a resource for distinguishing between the actual mitochondrial genome and this nuclear copy in functional studies.

category: genomics