title: Cryptic speciation in freshwaters: are speciation in lentic species shaped by paleohydrological events?

abstract: The Indian subcontinent is extremely diverse in terms of its flora and fauna. However, there is a severe underestimation of the biotic diversity in invertebrate groups as well as a lack of understanding of the processes generating diversity in these groups. Here we aimed to jointly estimate the cryptic diversity of two freshwater snail species complexes - Pila globosa and Pila virens, and uncover the processes behind the speciation in these groups. We employed phylogenetic, morphometric, population genetic and niche modelling technique to address these questions. We found out that both species complexes consist of several genetically and ecologically distinct putative species. The speciation was primarily driven by allopatric isolation into different river basins. A combination of paleoclimatic and paleohydrological changes during the Miocene have shaped the speciation events. Local climatic adaptation also could have contributed towards some of the speciation events. The study sheds light on the complex interaction between the biology of the species and the environment that shape the diversification patterns in a group.


title: The genetic and epigenetic landscape of the Arabidopsis centromeres

abstract:
Centromeres attach chromosomes to spindle microtubules during cell division and, despite this conserved role, show paradoxically rapid evolution and are typified by complex repeats. We used ultra-long-read sequencing to generate the Col-CEN Arabidopsis thaliana genome assembly that resolves all five centromeres. The centromeres consist of megabase-scale tandemly repeated satellite arrays, which support high CENH3 occupancy and are densely DNA methylated, with satellite variants private to each chromosome. CENH3 preferentially occupies satellites with least divergence and greatest higher-order repetition. The centromeres are invaded by ATHILA retrotransposons, which disrupt genetic and epigenetic organization of the centromeres. Crossover recombination is suppressed within the centromeres, yet low levels of meiotic DSBs occur that are regulated by DNA methylation. We propose that Arabidopsis centromeres are evolving via cycles of satellite homogenization and retrotransposon-driven diversification.

One-sentence summaryLong read sequencing and assembly of the Arabidopsis centromeres reveals their genetic and epigenetic topography.

category: genomics


title: Obesity-instructed TREM2high macrophages identified by comparative analysis of diabetic mouse and human kidney at single cell resolution

abstract: Mouse models are a tool for studying the mechanisms underlying complex diseases; however, differences between species pose a significant challenge for translating findings to patients. Here, we used single-cell transcriptomics and orthogonal validation approaches to provide cross-species taxonomies, identifying shared broad cell classes and unique granular cellular states, between mouse and human kidney. We generated cell atlases of the diabetic and obese kidney using two different mouse models, a high-fat diet (HFD) model and a genetic model (BTBR ob/ob), at multiple time points along disease progression. Importantly, we identified a previously unrecognized, expanding Trem2high macrophage population in kidneys of HFD mice that matched human TREM2high macrophages in obese patients. Taken together, our cross-species comparison highlights shared immune and metabolic cell-state changes.


title: The role of visual and olfactory cues in social decisions of guppies and zebrafish

abstract: Vision and olfaction are expensive to maintain, and in many taxa there appears to be a trade-off in investment between the two sensory systems. Previous work has suggested that guppies (Poecilia reticulata) and zebrafish (Danio rerio) may differ in the relative importance they place on these two senses in social interactions. In this study, we directly examined this issue by experimentally contrasting olfactory and visual information in social situations. In the first experiment, we found that guppies spent more time where conspecifics were visible than where they could smell them. On the contrary, zebrafish spent significantly more time in an empty compartment containing smell of conspecifics than in a compartment in which they only saw them. The difference was not large, suggesting that both species integrate various types of information to locate a nearby shoal. In two subsequent experiments, we studied the role of vision and smell in the discrimination of the quality of the social group, namely the number and the familiarity of its members. Zebrafish and guppies were confirmed to rely on different senses to estimate the size of a social group, whereas they did not differ in the discrimination of familiar and non-familiar conspecifics which appears to be based equally on the two senses. Similarly to what happens in other vertebrate clades, we suggest that, among teleosts, there are large differences in the relative importance of the different senses in the perception of the external world.

category: animal behavior and cognition


title: Parameter exploration improves the accuracy of long-read genome assembly

abstract: Long-molecule sequencing is now routinely applied to generate high-quality reference genome assemblies. However, datasets differ in repeat composition, heterozygosity, read lengths and error profiles. The assembly parameters that provide the best results could thus differ across datasets. By integrating four complementary and biologically meaningful metrics, we show that simple fine-tuning of assembly parameters can substantially improve the quality of long-read genome assemblies. In particular, modifying estimates of sequencing error rates improves some metrics more than two-fold. We provide a flexible software, CompareGenomeQualities, that automates comparisons of assembly qualities for researchers wanting a straightforward mechanism for choosing among multiple assemblies.

category: bioinformatics


title: Reconstructing unobserved cellular states from paired single-cell lineage tracing and transcriptomics data

abstract: AO_SCPLOWBSTRACTC_SCPLOWNovel experimental assays now simultaneously measure lineage relationships and transcriptomic states from single cells, thanks to CRISPR/Cas9-based genome engineering. These multimodal measurements allow researchers not only to build comprehensive phylogenetic models relating all cells but also infer transcriptomic determinants of consequential subclonal behavior. The gene expression data, however, is limited to cells that are currently present ("leaves" of the phylogeny). As a consequence, researchers cannot form hypotheses about unobserved, or "ancestral", states that gave rise to the observed population. To address this, we introduce TreeVAE: a probabilistic framework for estimating ancestral transcriptional states. TreeVAE uses a variational autoencoder (VAE) to model the observed transcriptomic data while accounting for the phylogenetic relationships between cells. Using simulations, we demonstrate that TreeVAE outperforms benchmarks in reconstructing ancestral states on several metrics. TreeVAE also provides a measure of uncertainty, which we demonstrate to correlate well with its prediction accuracy. This estimate therefore potentially provides a data-driven way to estimate how far back in the ancestor chain predictions could be made. Finally, using real data from lung cancer metastasis, we show that accounting for phylogenetic relationship between cells improves goodness of fit. Together, TreeVAE provides a principled framework for reconstructing unobserved cellular states from single cell lineage tracing data.


title: Ankyrin repeats in context with human population variation

abstract:
Ankyrin protein repeats bind to a wide range of substrates and are one of the most common protein motifs in nature. Here, we collate a high-quality alignment of 7,407 ankyrin repeats and examine for the first time, the distribution of human population variants from large-scale sequencing of healthy individuals across this family. Population variants are not randomly distributed across the genome but are constrained by gene essentiality and function. Accordingly, we interpret the population variants in context with evolutionary constraint and structural features including secondary structure, accessibility and protein-protein interactions across 383 three-dimensional structures of ankyrin repeats. We find five positions that are highly conserved across homologs and also depleted in missense variants within the human population. These positions are significantly enriched in intra-domain contacts and so likely to be key for repeat packing. In contrast, a group of evolutionarily divergent positions are found to be depleted in missense variants in human but significantly enriched in protein-protein interactions. Our analysis also suggests the domain has three, not two surfaces, each with different patterns of enrichment in protein-substrate interactions and missense variants. Our findings will be of interest to those studying or engineering ankyrin-repeat containing proteins as well as those interpreting the significance of disease variants.

Author SummaryComparison of variation at each position of the amino acid sequence for a protein across different species is a powerful way to identify parts of the protein that are important for its structure and function. Large-scale DNA sequencing of healthy people has recently made it possible to study normal genetic variation within just one species. Our work combines information on genetic differences between over 100,000 people with in-depth analysis of all available three-dimensional structures for Ankyrin repeats which are a widespread family of binding proteins formed by units with similar amino acid sequence that are found in tandem. Our combined analysis identifies sites critical for ankyrin stability as well as the positions most important for substrate interactions and hence function. Although focused only on the Ankyrins, the principles developed in our work are general and can be applied to any protein family.


title: Loss of Rnf43 accelerates Kras-mediated neoplasia and remodels the tumor immune microenvironment in pancreatic adenocarcinoma

abstract: RNF43 is an E3 ubiquitin ligase that is recurrently mutated in pancreatic ductal adenocarcinoma (PDAC) and precursor cystic neoplasms of the pancreas. The impact of RNF43 mutations on PDAC is poorly understood and autochthonous models have not been sufficiently characterized. In this study we describe a genetically engineered mouse model (GEMM) of PDAC with conditional expression of oncogenic Kras and deletion of the catalytic domain of Rnf43 (KRC) in exocrine cells. We demonstrate that Rnf43 loss results in an increased incidence of high-grade cystic lesions of the pancreas and PDAC. Importantly, KRC mice have a significantly decreased survival compared to mice containing only an oncogenic Kras mutation. By use of single cell RNA sequencing we demonstrated that KRC tumor progression is accompanied by a decrease in macrophages, as well as an increase in T and B lymphocytes with evidence of increased immune checkpoint molecule expression and affinity maturation, respectively. This was in stark contrast to the tumor immune microenvironment observed in the Kras/Tp53 driven PDAC GEMM. Furthermore, expression of the chemokine, CXCL5, was found to be specifically decreased in KRC cancer cells by means of epigenetic regulation and emerged as a putative candidate for mediating the unique KRC immune landscape. This GEMM establishes RNF43 as a bona fide tumor suppressor gene in PDAC and puts forth a rationale for an immunotherapy approach in this subset of PDAC cases.

category: cancer biology


title: A connectivity-constrained computational account of topographic organization in high-level visual cortex

abstract:
Inferotemporal cortex (IT) in humans and other primates is topo-graphically organized, containing multiple hierarchically-organized areas selective for particular domains, such as faces and scenes. This organization is commonly viewed in terms of evolved domain-specific visual mechanisms. Here, we develop an alternative, domain-general and developmental account of IT cortical organization. The account is instantiated as an Interactive Topographic Network (ITN), a form of computational model in which a hierarchy of model IT areas, subject to connectivity-based constraints, learns high-level visual representations optimized for multiple domains. We find that minimizing a wiring cost on spatially organized feedforward and lateral connections within IT, combined with constraining the feedforward processing to be strictly excitatory, results in a hierarchical, topographic organization. This organization replicates a number of key properties of primate IT cortex, including the presence of domain-selective spatial clusters preferentially involved in the representation of faces, objects, and scenes, columnar responses across separate excitatory and inhibitory units, and generic spatial organization whereby the response correlation of pairs of units falls off with their distance. We thus argue that domain-selectivity is an emergent property of a visual system optimized to maximize behavioral performance while minimizing wiring costs.

Significance StatementWe introduce the Interactive Topographic Network, a framework for modeling high-level vision, to demonstrate in computational simulations that the spatial clustering of domains in late stages of the primate visual system may arise from the demands of visual recognition under the constraints of minimal wiring costs and excitatory between-area neuronal communication. The learned organization of the model is highly specialized but not fully modular, capturing many of the properties of organization in primates. Our work is significant for cognitive neuroscience, by providing a domain-general developmental account of topo-graphic functional specialization, and for computational neuroscience, by demonstrating how well-known biological details can be successfully incorporated into neural network models in order to account for critical empirical findings.

category: neuroscience


title: A connectivity-constrained computational account of topographic organization in primate high-level visual cortex


title: Competition between distinct ApoE alleles and mCRP for the endothelial receptor CD31 differentially regulates neurovascular inflammation and Alzheimer's disease pathology

abstract:
BACKGROUNDC-reactive protein (CRP) in peripheral inflammation is associated with increased Alzheimers disease (AD) risk in Apolipoprotein E4 (ApoE4), but not ApoE3 or E2, humans. It remains unknown whether peripheral monomeric CRP (mCRP) induces AD pathogenesis through some receptor of blood-facing endothelia in the brain in an ApoE genotype dependent fashion.

METHODSWe used human samples, ApoE knock-in and deficient mouse models, and primary brain endothelia. Different ApoE mice were intraperitoneally (i.p.) injected with mCRP. The characterizations by immunostaining, proximity ligation assay (PLA) and siRNA were conducted to identify the receptor for mCRP. Brain microvessel and endothelia were isolated for RNA sequencing to explore the molecular pathway.

RESULTSWe demonstrate that CD31 (PECAM-1), a blood-facing endothelial receptor in brain, is a competitive target of both mCRP and ApoE protein. ApoE2 competes more strongly with mCRP for CD31 than ApoE4 does, and expressing ApoE4 or knocking out ApoE gene results in higher levels of mCRP-CD31 binding, leading to a decrease of CD31 expression but an increase in CD31 phosphorylation, along with greater cerebrovascular damage and AD pathology. This competitive binding mediates differential endothelial molecular responses depending on ApoE genotype, increasing cerebrovascular inflammation and mitochondria impairment in ApoE4 mice, while inducing vasculogenesis and protective changes in the presence of ApoE2.

CONCLUSIONSOur study reveals a novel and dynamic endothelial ApoE-mCRP-CD31 pathway for AD pathogenesis during chronic inflammation and provides some insight into the opposing ApoE4-neurodegenerative and ApoE2-neuroprotective effects in AD.

Clinical PerspectiveO_ST_ABSWHAT IS NEW?C_ST_ABSO_LICD31 is a competitive target of both mCRP and ApoE in brain endothelia in an ApoE-allele dependent pattern
C_LIO_LImCRP increases CD31 phosphorylation in the brain endothelia and damages cerebrovasculature in ApoE4 carriers and AD brains
C_LIO_LImCRP expression results in neuroprotective or neurodegenerative pathway activation in an ApoE-dependent manner
C_LI

WHAT ARE THE CLINICAL IMPLICATIONS?O_LIAlthough ApoE4 is a major genetic risk factor of AD, some ApoE4 carriers do not develop AD by the age of 90.
C_LIO_LIElderly people often experience peripheral inflammatory attacks and develop chronic low-grade inflammation, which results in the formation and release of mCRP. Because CRP is routine clinical laboratory test, clinicians can use blood CRP level to predict AD risk in ApoE4 carriers.
C_LIO_LIEvidence of Apoe4 genotype and chronic low-grade inflammation stages marked by elevated CRP levels should be targeted in personalized treatment and clinical trials for AD.
C_LI


title: Gene surfing of underdominant alleles promotes formation of hybrid zones

abstract: The distribution of genetic diversity over geographic space has long been investigated in population genetics and serves as a useful tool to understand evolution and history of populations. Within some species or across regions of contact between two species, there are instances where there is no apparent ecological determinant of sharp changes in allele frequencies or divergence. To further understand these patterns of spatial genetic structure and potential species divergence, we model the establishment of clines that occur due to the surfing of underdominant alleles during range expansions. We provide analytical approximations for the fixation probability of underdominant alleles at expansion fronts and demonstrate that gene surfing can lead to clines in 1D range expansions. We extend these results to multiple loci via a mixture of analytical theory and individual-based simulations. We study the interaction between the strength of selection against heterozygotes, migration rates, and local recombination rates on the formation of stable hybrid zones. A key result of our study is that clines created by surfing at different loci can attract each other and align after expansion, if they are sufficiently close in space and in terms of recombination distance. Our findings suggest that range expansions can set the stage for parapatric speciation due to the alignment of multiple selective clines, even in the absence of ecologically divergent selection.

category: evolutionary biology


title: Multi-omics consensus ensemble refines the classification of muscle-invasive bladder cancer with stratified prognosis, tumour microenvironment and distinct sensitivity to frontline therapies

abstract: The molecular classification of muscle-invasive bladder cancer (MIBC) based on transcriptomic signatures has been extensively studied. The complementary nature of information provided by different molecular profiles motivated us to refine MIBC classification by aggregating multi-omics data. We generated a consensus ensemble through ten multi-omics integrative clustering approaches on 396 MIBCs from TCGA. A total of 701 MIBCs from different sequencing technologies were used for external validation. Associations between subtypes and prognosis, molecular profiles, the tumour microenvironment, and potential response to frontline therapies were further analyzed. Nearest template prediction and random forest classification were used to develop a predictive signature/classifier for MIBC refinement. We identified four integrative consensus subtypes of MIBC, which were further designated basal-inflamed, basal-noninflamed, luminal-excluded and luminal-desert by immune profiling. Of note, the refinement of basal-like MIBC classification adds to the literature by identifying a basal-noninflamed MIBC subtype presenting with a significantly poor outcome and a global immune-cold phenotype, which might be triggered by Chr4 deletion and high activation of the oncogenic NRF2 pathway. In contrast, basal-inflamed MIBC showed high immunocyte infiltration and high expression of potential targets for immunotherapy. Using an external metastatic MIBC cohort in which patients received anti-PD-L1 treatment, we suggested that basal-inflamed MIBC had a higher likelihood of responding to immunotherapy than other MIBCs. The R package "refineMIBC" was offered as a research tool to refine MIBC from a single-sample perspective (https://github.com/xlucpu/refineMIBC). This consensus ensemble refines the intrinsic MIBC subtypes, which provides a blueprint for the clinical development of rational targeted and immunotherapeutic strategies.


title: EP400NL is required for cMyc-mediated PD-L1 gene activation by forming a transcriptional coactivator complex

abstract: EP400 is an ATP-dependent chromatin remodeling enzyme that has been implicated in DNA double-strand break repair and transcription regulation including Myc-dependent gene expression. We previously showed that the N-terminal domain of EP400 increases the efficacy of chemotherapeutic drugs against cancer cells. As the EP400 N-terminal-Like (EP400NL) gene resides next to the EP400 gene locus prompted us to investigate whether EP400NL also plays a similar role in epigenetic transcriptional regulation to the full-length EP400 protein. We found that EP400NL forms a human NuA4-like chromatin remodelling complex that lacks both the TIP60 histone acetyltransferase and EP400 ATPase. However, this EP400NL complex displays H2A.Z deposition activity on a chromatin template comparable to the human NuA4 complex, suggesting another associated ATPase such as BRG1 or RuvBL1/RuvBL2 catalyses the reaction. We also demonstrated that the transcriptional coactivator function of EP400NL is required for cMyc and IFN{gamma}-mediated PD-L1 gene activation. Collectively, our studies show that EP400NL plays a role as a transcription coactivator for cMyc-mediated gene expression and provides a potential target to modulate PD-L1 expression in cancer immunotherapy.


title: Non-coding mutations reveal cancer driver cistromes in luminal breast cancer

abstract: Whole genome sequencing of primary breast tumors enabled the identification of cancer driver genes 1,2 and non-coding cancer driver plexuses from somatic mutations 3-6. However, differentiating driver and passenger events among non-coding genetic variants remains a challenge to understand the etiology of cancer and inform delivery of personalized cancer medicine. Herein, we reveal an enrichment of non-coding mutations in cis-regulatory elements that cover a subset of transcription factors linked to tumor progression in luminal breast cancers. Using a cohort of 26 primary luminal ER+PR+ breast tumors, we compiled a catalogue of [~]100,000 unique cis-regulatory elements from ATAC-seq data. Integrating this catalogue with somatic mutations from 350 publicly available breast tumor whole genomes, we identified four recurrently mutated individual cis-regulatory elements. By then partitioning the non-coding genome into cistromes, defined as the sum of binding sites for a transcription factor, we uncovered cancer driver cistromes for ten transcription factors in luminal breast cancer, namely CTCF, ELF1, ESR1, FOSL2, FOXA1, FOXM1 GATA3, JUND, TFAP2A, and TFAP2C in luminal breast cancer. Nine of these ten transcription factors were shown to be essential for growth in breast cancer, with four exclusive to the luminal subtype. Collectively, we present a strategy to find cancer driver cistromes relying on quantifying the enrichment of non-coding mutations over cis-regulatory elements concatenated into a functional unit drawn from an accessible chromatin catalogue derived from primary cancer tissues.


title: MICROMORPHY OFFERS EFFECTIVE DEFENCE AGAINST PREDATION: INSIGHTS FROM THE COST-BENEFIT ANALYSES OF MICRO GASTROPOD PREDATION RECORD

abstract: Predation, an important driver of natural selection, is studied in the fossil record using quantifiable traces like drill holes produced by gastropods and repair scars produced after durophagous attacks. Despite the abundance of such records in molluscan prey, predation records of micromolluscs (<5mm) remained unexplored. Using a Miocene assemblage of microgastropods from the Quilon Limestone, India, we established the predatory-prey dynamics with the help of cost-benefit analyses. The overall predation intensity is low (DF = 0.06, RF= 0.04) and does not depend on the relative abundance of prey groups suggesting a non-random prey selection regardless of the encounter frequency. The predation is selective in terms of taxonomy, ornamentation, and size of the prey. The smallest size class has the lowest DF and RF supporting a negative size refugia. Higher IDF in larger size class and ornamented groups implies morphological defense resulting in higher failure. Microgastropods show a lower predation intensity than their regular-sized counterparts in a global comparison of coeval records. Results of the cost-benefit analyses explain this difference; the net energy gain from predatory drilling is found to increase monotonically with increasing prey size making the small prey less beneficial. Because the predators try to maximize net energy gain from a predatory attack, the microgastropod prey characterized by relatively low net energy yield is not preferred in the presence of larger prey. Micromorphy, therefore, appears a viable strategy for the prey group to be adopted as an evolutionary response against predation, especially in resource-limited conditions that fail to support large body size.

category: paleontology


title: Distinct Brain Mechanisms for Conflict Adaptation Within and Across Conflict Types

abstract: Cognitive conflict, like other cognitive processes, shows the characteristic of adaptation, i.e., conflict effects are attenuated when immediately following a conflicting event, a phenomenon known as the conflict adaptation effect (CAE). One important aspect of CAE is its sensitivity to the intertrial coherence of conflict type, i.e., behavioral CAE occurs only if consecutive trials are of the same conflict type. Although reliably observed behaviorally, the neural mechanisms underlying such a phenomenon remains elusive. With a paradigm combining the classic Simon task and Stroop task, this fMRI study examined neural correlates of conflict adaptation both within and across conflict types. The results revealed that when the conflict type repeated (but not when it alternated), the CAE-like neural activations were observed in dorsal anterior cingulate cortex, inferior frontal gyrus, superior parietal lobe, etc. (i.e., regions within typical task-positive networks). In contrast, when the conflict type alternated (but not when it repeated), we found CAE-like neural deactivations in a range of regions including bilateral superior and medial frontal gyri, bilateral angular cortex, bilateral temporal cortices, etc. (i.e., regions within the typical task-negative network). Moreover, this CAE-like neural deactivation predicts behavior performance. Network analyses suggested that these regions (for CAE-like neural activities within and across conflict type[s] respectively) can be clustered into two antagonistic networks. This evidence suggests that our adaptation to cognitive conflicts within a conflict type and across different types may rely on these two distinct neural mechanisms.


title: Epigenetic regulation of innate immune memory in microglia

abstract: Microglia are the tissue-resident macrophages of the CNS. They originate in the yolk sac, colonize the CNS during embryonic development and form a self-sustaining population with limited turnover. A consequence of their relative slow turnover is that microglia can serve as a long-term memory for inflammatory or neurodegenerative events. We characterized the epigenomes and transcriptomes of microglia exposed to different stimuli; an endotoxin challenge (LPS) and genotoxic stress (DNA repair deficiency-induced accelerated aging). Whereas the enrichment of permissive epigenetic marks at enhancer regions explains training (hyperresponsiveness) of primed microglia to LPS challenge, the tolerized response of microglia seems to be regulated by loss of permissive epigenetic marks. Here, we identify that inflammatory stimuli and accelerated aging because of genotoxic stress activate distinct gene networks. These gene networks and associated biological processes are partially overlapping, which is likely driven by specific transcription factor networks, resulting in altered epigenetic signatures and distinct functional (desensitized vs. primed) microglia phenotypes.


title: Identification of Leucinostatins from Ophiocordyceps sp. as Antiparasitic Agents Against Trypanosoma cruzi

abstract: Safe and effective treatments for Chagas disease, a potentially fatal parasitic infection associated with cardiac and gastrointestinal pathology and caused by the kinetoplastid parasite Trypanosoma cruzi, have yet to be developed. Benznidazole and nifurtimox, which are currently the only available drugs against T. cruzi, are associated with severe adverse effects and questionable efficacy in the late stage of the disease. Natural products have proven to be a rich source of new chemotypes for other infectious agents. We utilized a microscopy-based high-throughput phenotypic screen to identify inhibitors of T. cruzi from a library of natural product samples obtained from fungi procured through a Citizen Science Soil Collection Program (https://whatsinyourbackyard.org/), and the Great Lakes (USA) benthic environment. We identified five leucinostatins (A, B, F, NPDG C and NPDG D) as potent inhibitors of the intracellular amastigote form of T. cruzi. Leucinostatin B also showed in vivo efficacy in a mouse model of Chagas disease. Given prior reports that leucinostatins A and B have antiparasitic activity against the related kinetoplastid T. brucei, our findings suggest a potential cross-trypanocidal compound class and provide a platform for further chemical derivatization of a potent chemical scaffold against T. cruzi.

category: microbiology


title: Single cell analysis of spondyloarthritis regulatory T cells identifies distinct synovial gene expression patterns and clonal fates

abstract: Regulatory T cells (Tregs) play an important role in controlling inflammation and limiting autoimmunity, but their phenotypes at inflammatory sites in human disease are poorly understood. We here analyze the single-cell transcriptome of >16,000 Tregs obtained from peripheral blood and synovial fluid of two patients with HLA-B27+ ankylosing spondylitis and three patients with psoriatic arthritis, closely related forms of inflammatory spondyloarthritis. We identify multiple Treg clusters with distinct transcriptomic profiles, including, among others, a regulatory CD8+ subset expressing cytotoxic markers/genes, and a Th17-like RORC+ Treg subset characterized by IL-10 and LAG-3 expression. Synovial Tregs show upregulation of interferon signature and TNF receptor superfamily genes, and marked clonal expansion, consistent with tissue adaptation and antigen contact respectively. Individual synovial Treg clones map to different clusters indicating cell fate divergence. Finally, we demonstrate that LAG-3 directly inhibits IL-12/23 and TNF secretion by patient-derived monocytes, a mechanism with translational potential in SpA. Our detailed characterization of Tregs at an important inflammatory site illustrates the marked specialization of Treg subpopulations.

category: immunology


title: Nuclear and cytoplasmic RNA exosomes and PELOTA1 prevent miRNA-induced secondary siRNA production in Arabidopsis

abstract: Amplification of short interfering RNA (siRNAs) via RNA dependent RNA Polymerases (RdRPs) is of fundamental importance in RNA silencing. In plants, silencing by microRNAs (miRNAs) generally does not lead to engagement of RdRPs, in part thanks to an as yet poorly understood activity of the cytoplasmic exosome adaptor SKI2. Here, we show that mutation of the cytoplasmic exosome subunit RRP45B results in siRNA production very similar to what is observed in ski2 mutants. Furthermore, loss of the nuclear exosome adaptor HEN2 leads to secondary siRNA production from miRNA targets largely distinct from those producing siRNAs in ski2. Importantly, mutation of the Release Factor paralogue PELOTA1 required for subunit dissociation of stalled ribosomes causes siRNA production from miRNA targets overlapping with, but distinct from, those affected in ski2 and rrp45b mutants. We also show that miRNA-induced illicit secondary siRNA production correlates with miRNA levels rather than accumulation of stable 5-cleavage fragments. We propose that stalled RNA-induced Silencing Complex (RISC) and ribosomes, but not stable target mRNA cleavage fragments released from RISC, trigger secondary siRNA production, and that the exosome limits siRNA amplification by reducing RISC dwell time on miRNA target mRNAs while PELOTA1 does so by reducing ribosome stalling.

category: molecular biology


title: Evolution of brilliant iridescent feather nanostructures

abstract: The brilliant iridescent plumage of birds creates some of the most stunning color displays known in the natural world. Iridescent plumage colors are produced by nanostructures in feathers and have evolved in a wide variety of birds. The building blocks of these structures--melanosomes (melanin-filled organelles)--come in a variety of forms, yet how these different forms contribute to color production across birds remains unclear. Here, we leverage evolutionary analyses, optical simulations and reflectance spectrophotometry to uncover general principles that govern the production of brilliant iridescence. We find that a key feature that unites all melanosome forms in brilliant iridescent structures is thin melanin layers. Birds have achieved this in multiple ways: by decreasing the size of the melanosome directly, by hollowing out the interior, or by flattening the melanosome into a platelet. The evolution of thin melanin layers unlocks color-producing possibilities, more than doubling the range of colors that can be produced with a thick melanin layer and simultaneously increasing brightness. We discuss the implications of these findings for the evolution of iridescent structures in birds and propose two evolutionary paths to brilliant iridescence.


title: Early Disruption of Photoreceptor Cell Architecture and Loss of Vision in a Humanized Pig Model of Usher Syndrome

abstract: Usher syndrome (USH) is the most common form of monogenic deaf-blindness. Loss of vision is untreatable and, so far, there are no suitable animal models for testing therapeutic strategies. By introducing a human mutation into the harmonin-encoding USH1C gene in pigs, we generated the first translational animal model for USH type 1 with characteristic hearing defect, vestibular dysfunction and visual impairment. Changes in photoreceptor architecture, quantitative motion analysis and electroretinography were characteristics of the reduced retinal virtue in USH1C pigs. Primary cells from those animals and USH1C patients showed significantly elongated primary cilia, compared to wild-type, confirming the nature of USH as a true and general ciliopathy and proving the therapeutic capacity of gene supplementation and gene repair approaches.

category: genetics


title: Cryo-EM Structure of the Photosynthetic LH1-RC Complex from Rhodospirillum rubrum

abstract: We present a cryo-EM structure of the light-harvesting-reaction center (LH1-RC) core complex from purple phototrophic bacterium Rhodospirillum (Rsp.) rubrum at 2.76 [A] resolution. The LH1 complex forms a closed, slightly elliptical ring structure with 16 {beta}-polypeptides surrounding the RC. Our biochemical analysis detected rhodoquinone (RQ) molecules in the purified LH1-RC, and the cryo-EM density map specifically positions RQ at the QA site in the RC. The geranylgeraniol sidechains of bacteriochlorophyll (BChl) aG coordinated by LH1 {beta}-polypeptides exhibit a highly homologous tail-up conformation that allows for interactions with the bacteriochlorin rings of nearby LH1 -associated BChls aG. The structure also revealed key protein-protein interactions in both N- and C-terminal regions of the LH1 {beta}-polypeptides, mainly within a face-to-face structural subunit. Our findings enable to evaluate past experimental and computational results obtained with this widely used organism and provide crucial information for more detailed exploration of light-energy conversion, quinone transport, and structure-function relationships in pigment-protein complexes.

category: biochemistry


title: The molecular basis of ubiquitin-specific protease 8 autoinhibition by the WW-like domain

abstract: Ubiquitin-specific protease 8 (USP8) is a deubiquitinating enzyme involved in multiple membrane trafficking pathways. The enzyme activity is inhibited by binding to 14-3-3 proteins, and mutations of the 14-3-3 binding motif in USP8 are related to Cushings disease. However, the molecular basis of USP8 enzyme activity regulation remains unclear. Here, we identified amino acids 645-684 of USP8 as an autoinhibitory region, which our pull-down and single-molecule FRET assay results suggested interacts with the catalytic USP domain. In silico modelling indicated that the region forms a WW-like domain structure, plugs the catalytic cleft, and narrows the entrance to the ubiquitin-binding pocket. Furthermore, 14-3-3 was found to inhibit USP8 enzyme activity partly by enhancing the interaction between the WW-like and USP domains. These findings provide the molecular basis of USP8 autoinhibition via the WW-like domain. Moreover, they suggest that the release of autoinhibition may underlie Cushings disease caused by USP8 mutations.


title: Deep-learning on-chip DSLM enabling video-rate volumetric imaging of neural activities in moving biological specimens

abstract: Volumetric imaging of dynamic signals in a large, moving, and light-scattering specimen is extremely challenging, owing to the requirement on high spatiotemporal resolution and difficulty in obtaining high-contrast signals. Here we report that through combing a microfluidic chip-enabled digital scanning light-sheet illumination strategy with deep-learning based image restoration, we can realize isotropic 3D imaging of crawling whole Drosophila larva on an ordinary inverted microscope at single-cell resolution and high volumetric imaging rate up to 20 Hz. Enabled with high performances even unmet by current standard light-sheet fluorescence microscopes, we intoto record the neural activities during the forward and backward crawling of 1st instar larva, and successfully correlate the calcium spiking of motor neurons with the locomotion patterns.

category: bioengineering


title: Micro-Meta App: an interactive software tool to facilitate the collection of microscopy metadata based on community-driven specifications

abstract: For the information content of microscopy images to be appropriately interpreted, reproduced, and meet FAIR (Findable Accessible Interoperable and Reusable) principles, they should be accompanied by detailed descriptions of microscope hardware, image acquisition settings, image pixel and dimensional structure, and instrument performance. Nonetheless, the thorough documentation of imaging experiments is significantly impaired by the lack of community-sanctioned easy-to-use software tools to facilitate the extraction and collection of relevant microscopy metadata. Here we present Micro-Meta App, an intuitive open-source software designed to tackle these issues that was developed in the context of nascent global bioimaging community organizations, including BioImaging North America (BINA) and QUAlity Assessment and REProducibility in Light Microscopy (QUAREP-LiMi), whose goal is to improve reproducibility, data quality and sharing value for imaging experiments. The App provides a user-friendly interface for building comprehensive descriptions of the conditions utilized to produce individual microscopy datasets as specified by the recently proposed 4DN-BINA-OME tiered-system of Microscopy Metadata model. To achieve this goal the App provides a visual guide for a microscope-user to: 1) interactively build diagrammatic representations of hardware configurations of given microscopes that can be easily reused and shared with colleagues needing to document similar instruments. 2) Automatically extracts relevant metadata from image files and facilitates the collection of missing image acquisition settings and calibration metrics associated with a given experiment. 3) Output all collected Microscopy Metadata to interoperable files that can be used for documenting imaging experiments and shared with the community. In addition to significantly lowering the burden of quality assurance, the visual nature of Micro-Meta App makes it particularly suited for training users that have limited knowledge of the intricacies of light microscopy experiments. To ensure wide-adoption by microscope-users with different needs Micro-Meta App closely interoperates with MethodsJ2 and OMERO.mde, two complementary tools described in parallel manuscripts.


title: Mechanistic insights into the Japanese Encephalitis Virus RNA dependent RNA polymerase protein inhibition by bioflavonoids from Azadirachta indica

abstract: Japanese encephalitis (JE) virus, belongs to the genus flavivirus, is a major cause of viral encephalitis, which results in neurological damage. RNA-dependent RNA polymerase (RdRp) protein is the sole enzyme responsible for viral genome replication in RNA viruses and serves as an excellent target for anti-viral therapeutic development, as its homolog is not present in humans. In this study, the crystal structure of JE RNA-dependent RNA polymerase (jRdRp) protein was obtained from protein data bank while 43 by bioflavonoids reported in Azadirachta indica were retrieved from the PubChem database. Following, structure based virtual screening was employed using MTiOpenScreen server and top four compounds, viz. Gedunin, Nimbolide, Ohchinin acetate, and Kulactone, with the most negative docking scores (> -10 kcal/mol) conformations were redocked using AutoDock Vina; these complexes showed mechanistic interactions with Arg474, Gly605, Asp668, and Trp800 residues in the active site of jRdRp protein against reference ligand, i.e., Guanosine-5-Triphosphate. Furthermore, 100 ns classical molecular dynamics simulation and binding free energy calculation showed considerable stability (via hydrogen bonding and hydrophobic interactions) of docked bioflavonoids in the active pocket of jRdRp and significant contribution of van der Waals interactions for docked complex stability during simulation, respectively. Therefore, the outcome of this study predicted the substantial anti-viral activity of Gedunin, Nimbolide, Ohchinin acetate, and Kulactone against jRdRp protein and can be considered for further antiviral drug development.


title: findere: fast and precise approximatemembership query

abstract:
MotivationApproximate membership query (AMQ) structures as Cuckoo filters or Bloom filters are widely used for representing large sets of elements. Their lightweight space usage explains their success, mainly as they are the only way to scale hundreds of billions or trillions of elements. However, they suffer by nature from non-avoidable false-positive calls that bias downstream analyses of methods using these data structures.

ResultsIn this work we propose a simple strategy and its implementation for reducing the false-positive rate of any AMQ data structure indexing k-mers (words of length k). The method we propose, called findere, enables to speed-up the queries by a factor two and to decrease the false-positive rate by two order of magnitudes. This achievement is done one the fly at query time, without modifying the original indexing data-structure, without generating false-negative calls and with no memory overhead. With no drawback, this method, as simple as it is effective, reduces either the false-positive rate or the space required to represent a set given a user-defined false-positive rate.

Availabilityhttps://github.com/lrobidou/findere


title: Structural and biochemical analysis of ATPase activity and EsxAB substrate binding of M. tuberculosis EccCb1 enzyme

abstract: The EccC enzyme of M. tuberculosis ESX-1 system is a promising target for antivirulence drug development. The EccC enzyme comprises two polypeptides (i) EccCa1, a membrane bound enzyme having two ATPase domains D2 & D3 (ii) cytosolic EccCb, which contains two ATPase domains. In current study, we have analyzed the low-resolution structure of EccCb1, performed ATPase activity and EsxAB substrate binding analysis. The EccCb1 enzyme eluted as oligomer from size exclusion column and small angle X-ray scattering analysis revealed the double hexameric structure in solution. The EccCb1 enzyme showed catalytic efficiency (kcat/KM)[~] 0.020{+/-}0.005 M-1 min-1, however [~] 3.7 fold lower than its D2 and [~]1.7 fold lower than D3 domains respectively. The D2 and D3 domains exhibited the ATPase activity and mutation of residues involved in ATP+Mg2+ binding have yielded 56-94% reduction in catalytic efficiency for both D2 and D3 domains. The EccCb1 binds the EsxAB substrate with KD [~] 11.4{+/-}3.4 nM via specific groove located at C-terminal region of D3 domain. ATP binding to EccCb1 enhanced the EsxAB substrate binding by [~] 3 fold, indicating ATPase energy involvement in EsxAB substrate translocation. We modeled the dodecameric EccCb1+EsxAB+ ATP+Mg2+ complex, which showed the binding pockets involved in ATP+Mg2+ and EsxAB substrate binding. The enzyme dynamics involved in ATP+Mg2+ and EsxAB substrate recognition were identified and showed the enhanced stability of EccCb1 enzyme as a result of ligand binding. Overall, our structural and biochemical analysis showed the low-resolution structure and mechanism involved in ATPase activity and EsxAB substrate binding and dynamics involved in EsxAB substrate and ATP+Mg2+ recognition. Overall, our structural and biochemical data on EccCb1 will contribute significantly in development of antivirulence inhibitors, which will prevent virulence factor secretion by M. tuberculosis ESX-1 system.

category: biophysics


title: Deep learning models for poorly differentiated colorectal adenocarcinoma classification in whole slide images using transfer learning

abstract: Colorectal poorly differentiated adenocarcinoma (ADC) is known to have a poor prognosis as compared with well to moderately differentiated ADC. The frequency of poorly differentiated ADC is relatively low (usually less than 5% among colorectal carcinomas). Histopathological diagnosis based on endoscopic biopsy specimens is currently the most cost effective method to perform as part of colonoscopic screening in average risk patients, and it is an area that could benefit from AI-based tools to aid pathologists in their clinical workflows. In this study, we trained deep learning models to classify poorly differentiated colorectal ADC from Whole Slide Images (WSIs) using a simply transfer learning method. We evaluated the models on a combination of test sets obtained from five distinct sources, achieving receiver operator curve (ROC) area under the curves (AUCs) in the range of 0.94-0.98.

category: pathology


title: PIF7 controls leaf cell proliferation through an AN3 substitution-repression mechanism

abstract: Plants are agile, plastic organisms, able to adapt to ever-changing circumstances. Responding to far-red (FR) wavelengths from nearby vegetation, shade-intolerant species elicit the adaptive Shade Avoidance Syndrome (SAS), characterised by elongated petioles, leaf hyponasty and smaller leaves. We utilised end-of-day FR (EODFR) treatments to interrogate molecular processes that underlie the SAS leaf response. Genetic analysis establishes PHYTOCHROME INTERACTING FACTOR 7 (PIF7) is required for EODFR-mediated constraint of leaf blade cell division, while EODFR mRNAseq data identified ANGUSTIFOLIA3 (AN3) as a potential PIF7 target. We show PIF7 can suppress AN3 transcription through a sequestering mechanism that prevents AN3 activation of its own expression. We also establish PIF7 and AN3 impose antagonistic control of gene expression via common cis-acting promoter motifs in several cell cycle regulator genes. EODFR triggers the molecular substitution of AN3 to PIF7 at G-box/PBE-box promoter regions, and a switch from promotion to repression of gene expression.

category: plant biology


title: Longevity interventions temporally scale healthspan in Caenorhabditis elegans

abstract: Human centenarians and longevity mutants of model organisms show lower incidence rates of late-life morbidities than the average population. However, whether longevity is caused by a compression of the portion of life spent in a state of morbidity, i.e., "sickspan," is highly debated even in isogenic C. elegans. Here, we developed a microfluidic device that employs acoustophoretic force fields to quantify the maximum muscle strength and dynamic power in aging C. elegans. Together with different biomarkers for healthspan, we found a stochastic onset of morbidity, starting with a decline in dynamic muscle power and structural integrity, culminating in frailty. Surprisingly, we did not observe a compression of sickspan in longevity mutants but instead observed a temporal scaling of healthspan. Given the conservation of these longevity interventions, this raises the question of whether the healthspan of mammalian longevity interventions is also temporally scaled.

category: systems biology


title: Single-Cell Profiling Resolved Transcriptional Alterations and Lineage Dynamics of Subventricular Zone after Mild Traumatic Brain Injury

abstract: Mild traumatic brain injury (mTBI) is the most common form of brain trauma caused by physical impact. The subventricular zone (SVZ) is a neurogenetic niche that contributes to homeostasis and repair after brain injury. It is particularly challenging to fully elucidate the molecular alterations in the SVZ occurring in response to injury due to its cell diversity and the complex network. In this study, we aimed to address this issue using a novel transcriptomic technique- unbiased single-cell RNA sequencing. We resolved previous unknown cell subpopulations harbored in the niche, and uncovered cell type-specific alterations in gene expression, enriched pathways, and cell-cell crosstalk following mTBI. Notably, we also report novel lineage trajectories and molecular hallmarks that govern neurogenesis. This study dissects the delicate transcriptome changes of individual cell types as well as the reprogramming process of cells in the SVZ niche after mTBI, and our findings are expected to facilitate the development of therapeutic interventions or diagnostic tests for mTBI.


title: Co-expression patterns of microglia markers Iba1, TMEM119 and P2RY12 in Alzheimer's disease

abstract: Microglia have been identified as key players in Alzheimers disease pathogenesis, and other neurodegenerative diseases. Iba1, and more specifically TMEM119 and P2RY12 are gaining ground as presumedly more specific microglia markers, but comprehensive characterization of the expression of these three markers individually as well as combined is currently missing. Here we used a multispectral immunofluorescence dataset, in which over seventy thousand microglia from both aged controls and Alzheimer patients have been analysed for expression of Iba1, TMEM119 and P2RY12 on a single-cell level. For all markers, we studied the overlap and differences in expression patterns and the effect of proximity to {beta}-amyloid plaques. We found no difference in absolute microglia numbers between control and Alzheimer subjects, but the prevalence of specific combinations of markers (phenotypes) differed greatly. In controls, the majority of microglia expressed all three markers. In Alzheimer patients, a significant loss of TMEM119+-phenotypes was observed, independent of the presence of {beta}-amyloid plaques in its proximity. Contrary, phenotypes showing loss of P2RY12, but consistent Iba1 expression were increasingly prevalent around {beta}-amyloid plaques. No morphological features were conclusively associated with loss or gain of any of the markers or any of the identified phenotypes. All in all, none of the three markers were expressed by all microglia, nor can be wholly regarded as a pan- or homeostatic marker, and preferential phenotypes were observed depending on the surrounding pathological or homeostatic environment. This work could help select and interpret microglia markers in previous and future studies.


title: Structural and functional connectivity reconstruction with CATO - A Connectivity Analysis TOolbox

abstract: We describe a Connectivity Analysis TOolbox (CATO) for the reconstruction of structural and functional brain connectivity based on diffusion weighted imaging and resting-state functional MRI data. CATO is an integrative and modular software package that enables researchers to run end-to-end reconstructions from MRI data to structural and functional connectome maps, customize their analysis and utilize different software packages during the data preprocessing. The structural and functional connectome maps can be reconstructed with respect to user-defined (sub)cortical atlases providing aligned connectivity matrices for integrative multimodal analyses. We outline the structural and functional processing pipelines in CATO, the implementation in MATLAB and associated stand-alone application, and the calibration of performance with respect to simulated diffusion weighted imaging data and resting-state functional MRI data from the ICT2015 challenge and the Human Connectome Project. CATO is free open-source software and available at www.dutchconnectomelab.nl/CATO.


title: An ancient divide in outer membrane tethering systems in Bacteria

abstract: Recent data support the hypothesis that Gram-positive bacteria (monoderms) arose from Gram-negatives (diderms) through loss of the outer membrane (OM). However how this happened remains unknown. Considering that tethering of the OM is essential for cell envelope stability in diderm bacteria we hypothesize that its destabilization may have been involved in OM loss. Here, we present an in-depth analysis of the four main OM tethering systems across all Bacteria. We show that their distribution strikingly follows the bacterial phylogeny with a bimodal distribution matching the deepest phylogenetic cleavage between Terrabacteria (a clade encompassing Cyanobacteria, Deinococcus/Thermus, Firmicutes, etc.) and Gracilicutes (a clade encompassing Proteobacteria, Bacteroidetes, Spirochaetes, etc.). Diderm Terrabacteria display as the main system OmpM, a porin that attaches non-covalently to modified peptidoglycan or to secondary cell wall polymers. In contrast, the lipoprotein Pal is restricted to the Gracilicutes along with a more sporadic occurrence of OmpA. While Brauns lipoprotein Lpp is largely considered as the textbook example of OM attachment, it is actually present only in a subclade of Gammaproteobacteria. We propose an evolutionary scenario whereby the last common bacterial ancestor used a system based on OmpM, which was later replaced by one based on the lipoprotein Pal concomitantly to the emergence of the Lol machinery to address lipoproteins to the OM, with OmpA as a possible transition state. We speculate that the existence of only one main OM tethering system in the Terrabacteria would have allowed the multiple emergences of the monoderm phenotype specifically observed in this clade through OmpM perturbation. We test this hypothesis by inactivating all four ompM gene copies in the genetically tractable diderm Firmicute Veillonella parvula. The resulting mutant is severely affected in growth and displays high sensitivity to OM stress. High resolution imaging and tomogram reconstructions reveal a dramatic - yet non-lethal - phenotype, in which vast portions of the OM detach, producing large vesicles surrounding multiple monoderm-like cells sharing a common periplasm. Complementation by a single OmpM rescues the phenotype to a normal cell envelope. Together, our results highlight an ancient shift in bacterial evolution involving OM tethering systems. They suggest a possible mechanism for OM loss and a high flexibility of the cell envelope in diderm Firmicutes, making them ideal models to further refine our understanding of the mechanisms involved in bacterial OM stability, and opening the way to recapitulate the monoderm/diderm transition in the laboratory.


title: Breaching the cell-envelope barriers of gram-positive and fungal microbes by a type VI secretion system in Acidovorax citrulli

abstract: The type VI secretion system (T6SS) is a double-tubular toxin-injection nanomachine widely found in gram-negative human and plant pathogens. The current model depicts that the T6SS spear-like Hcp tube is powered by the contraction of an outer sheath to drill through the envelope of a neighboring cell, achieving cytosol to cytosol delivery. However, gram-positive bacteria seem to be impenetrable to such T6SS action. Here we report that a plant pathogen Acidovorax citrulli (AC) deploys a highly potent T6SS to kill a range of bacteria including Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis, and Mycobacterium smegmatis as well as fungal species including Candida albicans and Pichia pastoris. Using bioinformatic and biochemical assays, we identified a group of T6SS effectors and characterized one effector RhsB that is critical for interspecies interaction. We report that RhsB contains a conserved YD-repeat domain and a C-terminal nuclease domain. Toxicity of RhsB was neutralized by its downstream immunity proteins through direct interaction. RhsB was cleaved at the C-terminal end and a catalytic mutation within the internal aspartic protease abolished such cleavage. Collectively, the T6SS of AC displays potent activities to penetrate the cell envelope barriers of gram-positive and fungal species, highlighting the greatly expanded capabilities of T6SS in modulating microbiome compositions in complex environments.


title: The neurogenic fate of the hindbrain boundaries: a Notch-dependent behavioral switch triggers asymmetric division of boundary stem cells

abstract:
The generation of cell diversity in the central nervous system occurs during embryogenesis and requires a precise balance between cell proliferation, commitment to specific fates, and further neuronal differentiation. The cellular and molecular mechanisms regulating this balance in the embryonic brain are still poorly understood. Here we study how the neurogenic capacity in the embryonic hindbrain is spatiotemporally allocated, and how the fate and the growth of the hindbrain boundary cells are regulated. By generating a CRISPR-based knock-in zebrafish transgenic line to specifically label the hindbrain boundaries, we unveiled that boundary cells undergo a functional transition to become neurogenic during hindbrain segmentation concurrently as they maintain the progenitor cell pool. Boundary cells engaged in neurogenesis coinciding with the onset of Notch signaling, which triggered their asymmetrical cell division. Our findings reveal that distinct neurogenic phases take place during hindbrain growth and suggest that boundary cells contribute to refine the final number, identity, and proportion of neurons in the brain.

SUMMARYHindbrain boundary cells undergo a functional transition to become neurogenic concurrently as they are maintained as a progenitor cell pool. This involves a behavioral switch from symmetrically dividing progenitor cells to asymmetrically dividing progenitors, which depends on Notch-activity.

category: developmental biology


title: The neurogenic fate of the hindbrain boundaries relies on Notch-dependent asymmetric cell divisions


title: gcaPDA: A Haplotype-resolved Diploid Assembler

abstract: Generating chromosome-scale haplotype resolved assembly is important for functional studies. However, current de novo assemblers are either haploid assemblers that discard allelic information, or diploid assemblers that can only tackle genomes of low complexity. Here, we report a diploid assembler, gcaPDA (gamete cells assisted Phased Diploid Assembler), which exploits haploid gamete cells to assist in resolving haplotypes. We generate chromosome-scale phased diploid assemblies for the highly heterozygous and repetitive genome of a maize F1 hybrid using gcaPDA and evaluate the assembly result thoroughly. With applicability of coping with complex genomes and fewer restrictions on application than other diploid assemblers, gcaPDA is likely to find broad applications in studies of eukaryotic genomes.

category: genomics


title: A model of exercise-induced changes in metabolism, signaling, and gene expression in skeletal muscle

abstract: Skeletal muscle is the principal contributor to exercise-induced changes in human metabolism. Strikingly, although it has been demonstrated that a lot of metabolites accumulating in blood and human skeletal muscle during an exercise activate different signaling pathways and induce expression of many genes in working muscle fibres, the system understanding of signaling-metabolic pathways interrelations with downstream genetic regulation in the skeletal muscle is still elusive. Herein, a physiologically based computational model of skeletal muscle comprising energy metabolism, Ca2+ and AMPK signalling pathways, and expression regulation of genes with early and delayed responses has been developed based on a modular modeling approach. The integrated modular model validated on diverse including original experimental data and different exercise modes provides a comprehensive in silico platform in order to decipher and track cause-effect relationships between metabolic, signaling and gene expression levels in the skeletal muscle.


title: Tetracycline exposure alters key gut microbiota in Africanized honey bees (Apis mellifera scutellata x spp.)

abstract: Honey bees play a critical role in ecosystem health, biodiversity maintenance, and crop yield. Antimicrobials, such as tetracyclines, are used widely used across agriculture, medicine, and in bee keeping, and bees can be directly or indirectly exposed to tetracycline residues in the environment. In European honey bees, tetracycline exposure has been linked with shifts in the gut microbiota that negatively impact bee health. However, the effects of antimicrobials on Africanized honey bee gut microbiota have not been examined. The aim of this study was to investigate the effects of tetracycline exposure on the gut microbial community of Africanized honey bees (Apis mellifera scutellata x spp), which are important pollinators in South, Central, and North America. Bees (n=1,000) were collected from hives in Areia-PB, Northeastern Brazil, placed into plastic chambers and kept under controlled temperature and humidity conditions. The control group (CON) was fed daily with syrup (10g) consisting of a 1:1 solution of demerara sugar and water, plus a solid protein diet (10g) composed of 60% soy extract and 40% sugar syrup. The tetracycline group (TET) was fed identically but with the addition of tetracycline hydrochloride (450 {micro}g/g) to the sugar syrup. Bees were sampled from each group before (day 0), and after tetracycline exposure (days 3, 6 and 9). Abdominal contents dissected out of each bee underwent DNA extraction and 16S rRNA sequencing (V3-V4) on an Illumina MiSeq. Sequences were filtered and processed through QIIME2 and DADA2. Microbial community composition and diversity and differentially abundant taxa were evaluated by treatment and time. Bee gut microbial composition (Jaccard) and diversity (Shannon) differed significantly and increasingly over time and between CON and TET groups. Tetracycline exposure was associated with decreased relative abundances of Bombella and Fructobacillus, along with decreases in key core microbiota such as Snodgrassella, Gilliamella, Rhizobiaceae, and Apibacter. These microbes are critical for nutrient metabolism and pathogen defense, and decreased abundances of these microbes could negatively affect bee health. Considering the global ecological and economic importance of honey bees as pollinators, it is critical to understand the effects of agrochemicals including antimicrobials on honey bees.

category: ecology


title: Successful intergeneric transfer of a major apple scab resistance gene (Rvi6) from apple to pear and precise comparison of the downstream molecular mechanisms of this resistance in both species

abstract:
BackgroundScab is the most important fungal disease of apple and pear. Apple (Malus x domestica Borkh.) and European pear (Pyrus communis L.) are genetically related but they are hosts of two different fungal species: Venturia inaequalis for apple and V. pyrina for European pear. The apple/V. inaequalis pathosystem is quite well known, whereas knowledge about the pear/V. pyrina pathosystem is still limited. The aim of our study was to analyse the mode of action of a major resistance gene of apple (Rvi6) in transgenic apple and pear plants interacting with the two scab species (V. inaequalis and V. pyrina), in order to determine the degree of functional transferability between the two pathosystems.

ResultsTransgenic pear clones constitutively expressing the Rvi6 gene from apple were compared to a scab transgenic apple clone carrying the same construct. After inoculation in greenhouse with V. pyrina, strong defense reactions and very limited sporulation were observed on all transgenic pear clones tested. Microscopic observations revealed frequent aborted conidiophores in the Rvi6 transgenic pear / V. pyrina interaction. The macro- and microscopic observations were very comparable to the Rvi6 apple / V. inaequalis interaction. However, this resistance in pear proved variable according to the strain of V. pyrina, and one of the strains tested overcame the resistance of most of the transgenic pear clones. Comparative transcriptomic analyses of apple and pear resistant interactions with V. inaequalis and V. pyrina, respectively, revealed different cascades of molecular mechanisms downstream of the pathogen recognition by Rvi6 in the two species. Signal transduction was triggered in both species with calcium (and G-proteins in pear) and interconnected hormonal signaling (jasmonic acid in pear, auxins in apple and brassinosteroids in both species), without involvement of salicylic acid. This led to the induction of defense responses such as a remodeling of primary and secondary cell wall, lipids biosynthesis (galactolipids in apple and cutin and cuticular waxes in pear), systemic acquired resistance signal generation (in apple) or perception in distal tissues (in pear), and the biosynthesis of phenylpropanoids (flavonoids in apple but also lignin in pear).

ConclusionThis study is the first example of a successful intergeneric transfer of a resistance gene among Rosaceae, with a resistance gene functioning towards another species of pathogen.


title: SARS-CoV-2-specific memory B cells can persist in the elderly despite loss of neutralising antibodies

abstract:
Memory B cells (MBC) can provide a recall response able to supplement waning antibodies with an affinity-matured response better able to neutralise variant viruses. We studied a cohort of vulnerable elderly care home residents and younger staff, a high proportion of whom had lost neutralising antibodies (nAb), to investigate their reserve immunity from SARS-CoV-2-specific MBC. Class-switched spike and RBD-tetramer-binding MBC with a classical phenotype persisted five months post-mild/asymptomatic SARS-CoV-2 infection, irrespective of age. Spike/RBD-specific MBC remained detectable in the majority who had lost nAb, although at lower frequencies and with a reduced IgG/IgA isotype ratio. Functional spike/S1/RBD-specific recall was also detectable by ELISpot in some who had lost nAb, but was significantly impaired in the elderly, particularly to RBD. Our findings demonstrate persistence of SARS-CoV-2-specific MBC beyond loss of nAb, but highlight the need for careful monitoring of functional defects in RBD-specific B cell immunity in the elderly.

One sentence summaryCirculating class-switched spike and RBD-specific memory B cells can outlast detectable neutralising antibodies but are functionally constrained in the elderly.


title: Functional characterization of SARS-CoV-2 vaccine elicited antibodies in immunologically naive and pre-immune humans

abstract: As the COVID-19 pandemic continues, the authorization of vaccines for emergency use has been crucial in slowing down the rate of infection and transmission of the SARS-CoV-2 virus that causes COVID-19. In order to investigate the longitudinal serological responses to SARS-CoV-2 natural infection and vaccination, a large-scale, multi-year serosurveillance program entitled SPARTA (SARS SeroPrevalence and Respiratory Tract Assessment) was initiated at 4 locations in the U.S. The serological assay presented here measuring IgG binding to the SARS-CoV-2 receptor binding domain (RBD) detected antibodies elicited by SARS-CoV-2 infection or vaccination with a 95.5% sensitivity and a 95.9% specificity. We used this assay to screen more than 3100 participants and selected 20 previously infected pre-immune and 32 immunologically naive participants to analyze their antibody binding to RBD and viral neutralization (VN) responses following vaccination with two doses of either the Pfizer-BioNTech BNT162b2 or the Moderna mRNA-1273 vaccine. Vaccination not only elicited a more robust immune reaction than natural infection, but the level of neutralizing and anti-RBD antibody binding after vaccination is also significantly higher in pre-immune participants compared to immunologically naive participants (p<0.0033). Furthermore, the administration of the second vaccination did not further increase the neutralizing or binding antibody levels in pre-immune participants (p=0.69). However, ~46% of the immunologically naive participants required both vaccinations to seroconvert.


title: Complete protection by a single dose skin patch delivered SARS-CoV-2 spike vaccine

abstract: SARS-CoV-2 has infected over 160 million people and resulted in more than 3.3 million deaths, and we still face many challenges in the rollout of vaccines. Here, we use the high-density microarray patch to deliver a SARS-CoV-2 spike subunit vaccine directly to the skin. We show the vaccine, dry-coated on the patch is thermostable, and delivery of spike via HD-MAP induced greater cellular and antibody immune responses, with serum able to potently neutralize clinically relevant isolates including those from the B.1.1.7 and B.1.351 lineages. Finally, a single dose of HD-MAP-delivered spike provided complete protection from a lethal virus challenge, demonstrating that HD-MAP delivery of a SARS-CoV-2 vaccine is superior to traditional needle-and-syringe vaccination and has the potential to greatly impact the ongoing COVID-19 pandemic.


title: Kite-shaped molecules block SARS-CoV-2 cell entry at a post-attachment step

abstract: Anti-viral small molecules are currently lacking for treating coronavirus infection. The long development timescales for such drugs are a major problem, but could be shortened by repurposing existing drugs. We therefore screened a small library of FDA-approved compounds for potential severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) antivirals using a pseudovirus system that allows a sensitive read-out of infectivity. A group of structurally-related compounds, showing moderate inhibitory activity with IC50 values in the 1-5{micro}M range, were identified. Further studies demonstrated that these  kite-shaped molecules were surprisingly specific for SARS-CoV and SARS-CoV-2 and that they acted early in the entry steps of the viral infectious cycle, but did not affect virus attachment to the cells. Moreover the compounds were able to prevent infection in both kidney- and lung-derived human cell lines. The structural homology of the hits allowed the production of a well-defined pharmacophore that was found to be highly accurate in predicting the anti-viral activity of the compounds in the screen. We discuss the prospects of repurposing these existing drugs for treating current and future coronavirus outbreaks.


title: Identification of Pex34p as a component of the peroxisomal de novo biogenesis machinery in yeast

abstract: Cells can regulate the abundance and composition of peroxisomes to adapt to environmental changes. In the bakers yeast, S. cerevisiae, peroxisomes represent the only site for degradation of fatty acids. Hence, it is not surprising that growth of yeast cells on oleic acid results in a massive proliferation of peroxisomes. New peroxisomes can form either by division of pre-existing peroxisomes or de novo in a Pex25p-dependent process with the involvement of the Endoplasmic Reticulum (ER). In search for further factors involved in de novo formation of peroxisomes, we screened ~6,000 yeast mutants that were depleted of peroxisomes by conditional inhibition of PEX19 expression. Screening the mutants for the reappearance of peroxisomes upon expression of PEX19 identified Pex34p, in addition to the well-known component Pex25p, as crucial determinants for de novo biogenesis. Pex34p interacts with Pex19p and with different Peroxisomal Membrane Proteins (PMPs) in a PEX19-dependent manner. Depletion of Pex34p results in reduced numbers of import-competent peroxisomes formed de novo and Pex3p is partly retained and distributed in ER-like structures. We suggest that Pex25p and Pex34p are both required to maintain peroxisome number in a cell and that they perform non-redundant roles in the de novo formation of peroxisomes.

category: cell biology


title: BrainEXP-NPD: a database of transcriptomic profiles of human brains of six neuropsychiatric disorders

abstract:
BackgroundSpatio-temporal gene expression has been widely used to study gene functions and biological mechanisms in diseases. Numerous microarray and RNA sequencing data focusing on brain transcriptomes in neuropsychiatric disorders have accumulated. However, their consistency, reproducibility has not been properly evaluated. Except for a few psychiatric disorders, like schizophrenia, bipolar disorder and autism, most have not been compared to each other for cross-disorder comparisons.

MethodsWe organized 48 human brain transcriptome datasets from six sources. The original brain donors include patients with schizophrenia (SCZ, N=427), bipolar disorder (BD, N=312), major depressive disorder (MDD, N=219), autism spectrum disorder (ASD, N=53), Alzheimers disease (AD, N=765), Parkinsons disease (PD, N=163) as well as controls as unaffected by such disorders (CTRL, N=6,378), making it a total of 8,317 samples. Raw data included multiple brain regions of both sexes, with ages ranging from embryonic to seniors. After standardization, quality control, filtering and removal of known and unknown covariates, we performed comprehensive meta- and mega-analyses, including gene differential expression and gene co-expression network.

ResultsA total of 6922, 3011, 2703, 4389, 3507, 4279 significantly differentially expressed genes (FDR q < 0.05) were detected in the comparisons of 6 brain regions of SCZ-CTRL, 5 brain regions of BD-CTRL, 6 brain regions of MDD-CTRL, 4 brain regions of ASD-CTRL, 7 brain regions of AD-CTRL, and 6 brain regions of PD-CTRL, respectively. Most differentially expressed genes were brain region-specific and disease-specific. SCZ and BD have a maximal transcriptome similarity in striatum ({rho}=0.42) among the four brain regions, as measured by Spearmans correlation of differential expression log2 FC values. SCZ and MDD have a maximal transcriptome similarity in hippocampus ({rho}=0.30) among the five brain regions. BD and MDD have a maximal transcriptome similarity in frontal cortex ({rho}=0.45) among the five brain regions. Other disease pairs have a less transcriptome similarity ({rho}<0.1) in all brain regions. PD is negatively correlated with SCZ, BD, and MDD in cerebellum and striatum. We also performed coexpression network analyses for different disorders and controls separately. We developed a database named BrainEXP-NPD (http://brainexpnpd.org:8088/BrainEXPNPD/), to provide a userfriendly web interface for accessing the data, and analytical results of meta- and mega-analyses, including gene differential expression and gene co-expression networks between cases and controls on different brain regions, sexes and age groups. Discussion: BrainEXP-NPD compiled the largest collection of brain transcriptomic data of major neuropsychiatric disorders and presented lists of differentially expressed genes and coexpression modules in multiple brain regions of six major disorders.

category: bioinformatics


title: Skeletal Muscle Histidine Containing Dipeptide Contents are Increased in Freshwater Turtles (Chrysemys picta bellii) with Cold-Acclimation

abstract:
Freshwater turtles found in higher latitudes can experience extreme challenges to acid-base homeostasis while overwintering, due to a combination of cold temperatures along with the potential for environmental hypoxia. Histidine containing dipeptides (HCDs; carnosine, anserine and balenine) may facilitate pH regulation in response to these challenges, through their role as pH buffers. We measured the HCDs content of three tissues (liver, cardiac muscle and skeletal muscle) from the anoxia-tolerant painted turtle (Chrysemys picta bellii) acclimated to either 3 or 20{degrees}C. HCDs were detected in all tissues, with the highest content shown in the skeletal muscle. Turtles acclimated to 3{degrees}C had more HCD in their skeletal muscle than those acclimated to 20{degrees}C (carnosine = 20.8{+/-}4.5 vs 12.5{+/-}5.9 mmol{middle dot}kg DM-1; ES = 1.59 (95%CI: 0.16 - 3.00), P = 0.013). The higher HCD content observed in the skeletal muscle of the cold-acclimated turtles suggests a role in acid-base regulation in response to physiological challenges associated with living in the cold, with the increase possibly related to the temperature sensitivity of carnosines dissociation constant and buffering power of the skeletal muscle during anoxic submergence.

HighlightsO_LIpH regulation is a major challenge for overwintering freshwater turtles.
C_LIO_LIHistidine containing dipeptides are important intracellular buffers.
C_LIO_LITurtles acclimated to 3{degrees}C had higher HCD content than those at 20{degrees}C.
C_LIO_LIHCDs may be important pH regulators in cold-acclimated turtles.
C_LI

category: physiology


title: Reporting and Misreporting of Sex Differences in the Biological Sciences

abstract: As part of an initiative to improve rigor and reproducibility in biomedical research, the U. S. National Institutes of Health now requires the consideration of sex as a biological variable in preclinical studies. This new policy has been interpreted by some as a call to compare males and females with each other. Researchers testing for sex differences may not be trained to do so, however, increasing risk for misinterpretation of results. Using a list of recently published articles curated by Woitowich et al. (eLife, 2020; 9:e56344), we examined reports of sex differences and non-differences across nine biological disciplines. Sex differences were claimed in the majority of the 147 articles we analyzed; however, statistical evidence supporting those differences was often missing. For example, when a sex-specific effect of a manipulation was claimed, authors usually had not tested statistically whether females and males responded differently. Thus, sex-specific effects may be over-reported. In contrast, we also encountered practices that could mask sex differences, such as pooling the sexes without first testing for a difference. Our findings support the need for continuing efforts to train researchers how to test for and report sex differences in order to promote rigor and reproducibility in biomedical research.

category: scientific communication and education


title: Serial Dependence in Temporal Perception Reveals the Dynamics of Constructing an Internal Reference Frame

abstract: Temporal perception is crucial to cognitive functions. To better estimate temporal durations, the observers need to construct an internal reference frame based on past experience and apply it to guide future perception. However, how this internal reference frame is constructed remains largely unclear. Here we showed the dynamics of the internal reference construction from the perspective of serial dependence in temporal reproduction tasks. We found the current duration estimation is biased towards both perceived and reproduced durations in previous trials. Moreover, this effect is regulated by the variability of sample durations. The influence of previous trials was stronger when the observers were exposed to context with more variable durations, which is inconsistent with previous theories that the similarity between successive stimuli induces serial dependence. We proposed a Bayesian model with an adaptive reference updated continuously after each observation, which can better explain the serial dependence observed in temporal perception.

category: neuroscience


title: Microfabricated disk technology: rapid scale up in midbrain organoid generation

abstract: By providing a three-dimensional in vitro culture system with key features of the substantia nigra region in the brain, 3D neuronal organoids derived from human induced pluripotent stem cells (iPSCs) provide living neuronal tissue resembling the midbrain region of the brain. However, a major limitation of conventional brain organoid culture is that it is often labor-intensive, requiring highly specialized personnel for moderate throughput. Additionally, the methods published for long-term cultures require time-consuming maintenance to generate brain organoids in large numbers. With the increasing need for human midbrain organoids (hMOs) to better understand and model Parkinsons disease (PD) in a dish, there is a need to implement new workflows and methods to both generate and maintain hMOs, while minimizing batch to batch variation. In this study, we developed a method with microfabricated disks to scale up the generation of hMOs. This opens up the possibility to generate larger numbers of hMOs, in a manner that minimizes the amount of labor required, while decreasing variability and maintaining the viability of these hMOs over time. Taken together, producing hMOs in this manner opens up the potential for these to be used to further PD studies.


title: Spatial regulation of coordinated excitatory and inhibitory synaptic plasticity at dendritic synapses

abstract: The induction of synaptic plasticity at an individual dendritic glutamatergic spine can affect neighboring spines. This local modulation generates dendritic plasticity microdomains believed to expand the neuronal computational capacity. Here, we investigate whether local modulation of plasticity can also occur between glutamatergic synapses and adjacent GABAergic synapses. Using MNI-glutamate and DPNI-GABA double uncaging combined with electrophysiology, live-cell imaging and single-particle tracking, we find that the induction of LTP at an individual glutamatergic spine causes the depression of nearby GABAergic inhibitory synapses (within 3 microns), whereas more distant ones are potentiated. Notably, L-type calcium channels and calpain are required for this plasticity spreading. Overall, our data support a model whereby input-specific glutamatergic postsynaptic potentiation induces a spatially-regulated rearrangement of inhibitory synaptic strength in the surrounding area through short-range heterosynaptic interactions. Such local coordination of excitatory and inhibitory synaptic plasticity is expected to profoundly influence dendritic information processing and integration.


title: An olfactory pattern generator for on-demand combinatorial control of receptor activities

abstract: Olfactory systems employ combinatorial receptor codes for odors. Systematically generating stimuli that address the combinatorial possibilities of an olfactory code poses unique challenges. Here, we present a stimulus method to probe the combinatorial code, demonstrated using the Drosophila larva. This method leverages a set of primary odorants, each of which targets the activity of one olfactory receptor neuron (ORN) type at an optimal concentration. Our setup uses microfluidics to mix any combination of primary odorants on demand to activate any desired combination of ORNs. We use this olfactory pattern generator to demonstrate a spatially distributed olfactory representation in the dendrites of a single interneuron in the antennal lobe, the first olfactory neuropil of the larva. In the larval mushroom body, the next processing layer, we characterize diverse receptive fields of a population of Kenyon cells. The precision and flexibility of the olfactory pattern generator will facilitate systematic studies of processing and transformation of the olfactory code.


title: Scaling principles of white matter brain connectivity

abstract: Brains come in many shapes and sizes. Nature has endowed big-brained primate species like humans with a proportionally large cerebral cortex. White matter connectivity - the brains infrastructure for long-range communication - might not always scale at the same pace as the cortex. We investigated the consequences of this allometric scaling for white matter brain network connectivity. Structural T1 and diffusion MRI data were collated across fourteen primate species, describing a comprehensive 350-fold range in brain volume. We report volumetric scaling relationships that point towards a restriction in macroscale connectivity in larger brains. Building on previous findings, we show cortical surface to outpace white matter volume and the corpus callosum, suggesting the emergence of a white matter  bottleneck of lower levels of connectedness through the corpus callosum in larger brains. At the network level, we find a potential consequence of this bottleneck in shaping connectivity patterns, with homologous regions in the left and right hemisphere showing more divergent connectivity in larger brains. Our findings show conserved scaling relationships of major brain components and their consequence for macroscale brain circuitry, providing a comparative framework for expected connectivity architecture in larger brains such as the human brain.


title: Differential microRNA expression analyses across two brain regions in Alzheimer's disease

abstract:
BackgroundDysregulation of microRNAs (miRNAs) is involved in the pathogenesis of neurodegenerative diseases, including Alzheimers disease (AD). Hitherto, sample sizes from differential miRNA expression studies in AD are exceedingly small aggravating any biological inference. To overcome this limitation, we investigated six candidate miRNAs in a large collection of brain samples.

MethodsBrain tissue was derived from superior temporal gyrus (STG) and entorhinal cortex (EC) from 99 AD patients and 91 controls. Expression of six miRNAs was examined by qPCR (STG) or small RNA sequencing (EC). Brain region-dependent differential miRNA expression was investigated in a transgenic AD mouse model using qPCR and FISH. Total RNA sequencing was used to assess differential expression of miRNA target genes.

ResultsMiR-129-5p, miR-132-5p, and miR-138-5p were significantly downregulated in AD vs. controls both in STG and EC, while miR-125b-5p and miR-501-3p showed no evidence for differential expression in this dataset. In addition, miR-195-5p was significantly upregulated in EC but not STG in AD patients. The brain region-specific pattern of miR-195-5p expression was corroborated in vivo in transgenic AD mice. Total RNA sequencing identified several novel and functionally interesting target genes of these miRNAs involved in synaptic transmission (GABRB1), the immune-system response (HCFC2) or AD-associated differential methylation (SLC16A3).

ConclusionsUsing two different methods (qPCR and small RNA-seq) in two separate brain regions in 190 individuals we more than doubled the available sample size for most miRNAs tested. Differential gene expression analyses confirm the likely involvement of miR-129-5p, miR-132-5p, miR-138-5p, and miR-195-5p in AD pathogenesis and highlight several novel potentially relevant target mRNAs.

FundingThis work was supported by the Deutsche Forschungsgemeinschaft (DFG) and the National Science Foundation China (NSFC) as a Joint Sino-German research project ("MiRNet-AD", #391523883). Additional support was provided by the DFG Research Infrastructure NGS_CC (project 407495230) as part of the Next Generation Sequencing Competence Network (#423957469) and the Cure Alzheimer s Fund (CAF) as part of the CIRCUITS consortium project.


title: Transcriptional characterization of the glial response due to chronic neural implantation of flexible microprobes

abstract: Long term implantation of (micro-)probes into neural tissue cause unique and disruptive responses to these foreign bodies. In this study, we present the transcriptional trajectory of glial cells responding to chronic implantation of flexible micro-probes for up to 18 weeks. Transcriptome analysis shows a rapid activation of microglial cells and a strong upregulation of reactive astrocytic genes, which is lost over the full duration of the implant period. Most interestingly, animals that were implanted for 18 weeks show a transcriptional profile similar to non-implanted controls, with increased expression of genes associated with wound healing and angiogenesis, which raises hope of a normalization of the neuropil to the pre-injury state when using flexible probes. Nevertheless, our data show, that a subset of genes upregulated after 18 weeks belong to the family of immediate early genes, which would indicate that structural and functional remodeling has not been completed at this time point. Our results confirm and extend previous work on the molecular changes resulting from the presence of intraneural probes and provide a rational basis for developing intervention strategies to control them.


title: Pre-therapeutic Microglia Activation and Sex Determine Therapy Effects of Chronic Immunomodulation

abstract: Modulation of the innate immune system is emerging as a promising therapeutic strategy against Alzheimers disease (AD). However, determinants of a beneficial therapeutic effect are ill-understood. Thus, we investigated the potential of 18 kDa translocator protein positron-emission-tomography (TSPO-PET) for assessment of microglial activation in mouse brain before and during chronic immunomodulation. Serial TSPO-PET was performed during five months of chronic microglia modulation by stimulation of peroxisome proliferator-activated receptor (PPAR)-{gamma} with pioglitazone in two different mouse models of AD (PS2APP, AppNL-G-F). Using mixed statistical models on longitudinal TSPO-PET data, we tested for effects of therapy and sex on treatment response. We tested correlations of baseline with longitudinal measures of TSPO-PET, and correlations between PET results with spatial learning performance and {beta}-amyloid accumulation of individual mice. Immunohistochemistry was used to determine the molecular source of the TSPO-PET signal. Pioglitazone-treated female PS2APP and AppNL-G-F mice showed attenuation of the longitudinal increases in TSPO-PET signal when compared to vehicle controls, whereas treated male AppNL-G-F mice showed the opposite effect. Baseline TSPO-PET strongly predicted changes in microglial activation in treated mice (R=-0.874, p<0.0001) but not in vehicle controls (R=-0.356, p=0.081). Reduced TSPO-PET signal upon treatment was associated with better spatial learning and higher fibrillar {beta}-amyloid accumulation. Immunohistochemistry confirmed activated microglia to be the source of the TSPO-PET signal (R=0.952, p<0.0001). TSPO-PET represents a sensitive biomarker for monitoring of immunomodulation and closely reflects activated microglia. Pre-therapeutic assessment of baseline microglial activation and sex are strong predictors of individual immunomodulation effects and could serve for responder stratification.


title: Eosinophils are an integral component of the pulmonary granulocyte response in Tuberculosis and promote host resistance in mice

abstract: Host resistance to Mycobacterium tuberculosis infection requires the activities of multiple leukocyte subsets, yet the roles of the different innate effector cells during tuberculosis are incompletely understood. Here we uncover an unexpected association between eosinophils and Mtb infection. In humans, eosinophils are decreased in the blood but enriched in resected human tuberculosis lung lesions and autopsy granulomas. Influx of eosinophils is also evident in infected zebrafish, mice, and nonhuman primate granulomas, where they are functionally activated and degranulate. Importantly, employing complementary genetic models of eosinophil deficiency, we demonstrate that, in mice, eosinophils are required for optimal pulmonary bacterial control and host survival after Mtb infection. Collectively, our findings uncover an unexpected recruitment of eosinophils to the infected lung tissue and a protective role for these cells in the control of Mtb infection in mice.

category: immunology


title: Viruses roam the wheat phyllosphere

abstract: The phyllosphere comprises all the above-ground sections of plants. This niche is colonised by complex microbial communities, including algae, fungi, archaea, and bacteria1-3. They are known to induce plant growth and promote health4,5 or act as causative agents of plant diseases6-8. It is thought that the most abundant organisms are phyllobacteria2, with an estimate between 106 to 107 cells per cm2. Viruses are highly abundant across many environments, often outnumbering bacteria 10 to 19. However, not much is known about their abundance and composition in the phyllosphere, a harsh environment for viruses due to environmental variability and high UV exposure. To investigate this niche in detail, we extracted, sequenced and analysed phyllosphere virome. Using leaf samples from winter wheat (Triticum aestivum), we identified a total of 876 viral populations (vOTUs), mostly belonging to the Caudovirales order. Most of these were predicted to be lytic. Remarkably, 810 of these viral populations correspond to new viral species with no matches to known sequences. We estimate a minimum of 2.0 x106 viral particles per leaf. Overall, these findings suggest that the phyllosphere ecosystem harbours an abundant and active community of novel viruses that play essential roles in shaping this habitat.

category: microbiology


title: The giant Staphylococcus epidermidis surface protein Embp binds selectively to fibrillated fibronectin through a Velcro-like mechanism

abstract:
Staphylococcus epidermidis causes some of the most hard-to-treat clinical infections by forming biofilms: Multicellular communities of bacteria encased in a protective matrix, supporting immune evasion and tolerance against antibiotics. Biofilms occur most commonly on medical implants, and a key event in implant colonization is the robust adherence to the surface, facilitated by interactions between bacterial surface proteins and host matrix components. S. epidermidis is equipped with a giant adhesive protein, Embp, which facilitates bacterial interactions with surface-deposited, but not soluble fibronectin. The structural basis behind this selective binding process has remained obscure. Using a suite of single-cell and single-molecule analysis techniques, we show that S. epidermidis is capable of such distinction because Embp binds specifically to fibrillated fibronectin on surfaces, while ignoring globular fibronectin in solution. S. epidermidis adherence is critically dependent on multi-valent interactions involving 50 fibronectin-binding repeats of Embp. This unusual, Velcro-like interaction proved critical for colonization of surfaces under high flow, making this newly identified attachment mechanism particularly relevant for colonization of intravascular devices, such as prosthetic heart valves or vascular grafts. Other biofilm-forming pathogens, such as Staphylococcus aureus, express homologs of Embp and likely deploy the same mechanism for surface colonization. Our results may open for a novel direction in efforts to combat devastating, biofilm-associated infections, as the development of implant materials that steer the conformation of adsorbed proteins is a much more manageable task than avoiding protein adsorption altogether.

Graphical abstractFibronectin exists in two different conformations in the body. It circulates in the bodily fluids in globular conformation, however, it become fibrillated once adsorbed to an implant surface. S. epidermidis possess a giant 1 MDa receptor known as Embp bind specifically to fibrillated Fn but not to the globular Fn.

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title: The giant staphylococcal protein Embp facilitates colonization of surfaces through Velcro-like attachment to fibrillated fibronectin