title: Rapid transgenerational adaptation in response to intercropping increases facilitation and reduces competition

abstract: By capitalising on positive biodiversity-productivity relationships, intercropping provides opportunities to improve agricultural sustainability1. However, intercropping is generally implemented using commercial seeds that were bred for maximal productivity in monocultures, which might limit the benefits of crop diversity on yield2,3. Plants can adapt over generations to the level of surrounding plant diversity, notably through increases in niche differentiation4. However, this adaptation potential and the corresponding yield benefit potential have not been explored in annual crop systems. Here we show that plant-plant interactions among annual crops evolved towards increased facilitation and reduced competition when the plants coexistence history matched their current diversity setting, which led to an increase in overyielding of up to 58%. These higher yield benefits were linked to character convergence between species sharing the same coexistence history for two generations. Notably, the six crop species tested converged towards taller phenotypes with lower leaf dry matter content when grown in mixtures. This study provides the first empirical evidence for the importance of parental diversity affecting plant-plant interactions and ecosystem functioning of the following generations in annual cropping systems. These results have important implications for diversified agriculture as they demonstrate the yield potential of targeted cultivars for intercropping, which can be achieved through specific breeding for mixtures.


title: Road encroachment mediates species occupancy, trait filtering and dissimilarity of songbird communities

abstract: Assessing the road effects on biodiversity is challenging because impacts may depend on both wildlife responses to roads and on the spatial arrangement of roads. We questioned whether an increase in road encroachment i.e., the advancement of roads into non-urban areas, leads to significant changes (positive and negative) in species occurrence, and if so, whether those changes are linked to specific traits related to perturbation sensitiveness, therefore acting as filters that increase the community compositional dissimilarity. Using a large citizen-science dataset of point-counts performed throughout Iberian Peninsula (4,459 unique survey sites), we modelled the effect of road density on the occurrence of common songbirds (n=58), while accounting for potential confounding effects of environment and survey effort. We then tested if species occurrence patterns would be linked to specific traits related to the ability to cope with human presence. Finally, we assessed how road density affect the community compositional dissimilarity. We estimated 24 (41%) and 12 (21%) species to be negatively and positively affected by roads, respectively. Increased road encroachment was positively related with a higher prevalence of urban dwellers and negatively related with the occurrence of species nesting and foraging on the ground. Furthermore, increasing road density translated into an increasing community compositional dissimilarity, mostly due to species turnover. Our study support previous research showing that roads have different effects on the occurrence of different species, but we revealed that at least three species' traits have a clear relation with such road responses. Such trait filtering is probably causing a high species turnover between songbird communities occurring in roaded and nearby roadless areas. Overall, we found that different species-specific responses to roads translates into changes at the community level. Landscape and road-network management should be conceived acknowledging that roads are contributing to biodiversity changes. As so, building upon the concepts of Land Sharing / Land Sparing, conservation actions should be tailored according to the different species responses e.g., road verge management targeting species having a positive relation with road density; and compensation actions targeting species showing a negative response toward roads.

category: ecology


title: Functional variability in adhesion and flocculation of yeast megasatellite genes

abstract: Megasatellites are large tandem repeats found in all fungal genomes but especially abundant in the opportunistic pathogen Candida glabrata. They are encoded in genes involved in cell-cell interactions, either between yeasts or between yeast and human cells. In the present work, we have been using an iterative genetic system to delete several C. glabrata megasatellite-containing genes and found that two of them were positively involved in adhesion to epithelial cells, whereas three genes controlled negatively adhesion. Two of the latter, CAGL0B05061g or CAGL0A04851g, are also negative regulators of yeast-to-yeast adhesion, making them central players in controlling C. glabrata adherence properties. Using a series of synthetic Saccharomyces cerevisiae strains in which the FLO1 megasatellite was replaced by other tandem repeats of similar length but different sequences, we showed that the capacity of a strain to flocculate in liquid culture was unrelated to its capacity to adhere to epithelial cells or to invade agar. Finally, in order to understand how megasatellites were initially created and subsequently expanded, an experimental evolution system was set up, in which modified yeast strains containing different megasatellite seeds were grown in bioreactors for more than 200 generations and selected for their ability to sediment at the bottom of the culture tube. Several flocculation-positive mutants were isolated. Functionally relevant mutations included general transcription factors as well as a 230 kb segmental duplication.

category: genetics


title: In vivo Biomechanical Assessment of Iridial Deformations and Muscle Contractions in Human Eyes

abstract: The iris is a muscular organ whose deformations can cause primary angle-closure glaucoma (PACG), a leading cause of blindness. PACG risk assessment does not consider iridial biomechanical factors, despite their expected influence on iris deformations. Here we exploited an existing biometric data set consisting of near-infrared movies acquired during the pupillary light reflex (PLR) as a unique resource to study iris biomechanics. The PLR caused significant (>100%) and essentially spatially uniform radial strains in the iris in vivo, consistent with previous findings. Inverse finite element modeling showed that sphincter muscle tractions were c. 5-fold greater than iridial stroma stiffness (range 4- to 13-fold, depending on sphincter muscle size). This muscle traction is greater than has been previously estimated, which may be due to methodological differences and/or to different patient populations in our study (European descent) vs. previous studies (Asian); the latter possibility is of particular interest due to differential incidence rates of PACG in these populations. Our methodology is fast and inexpensive and may be a useful tool in understanding biomechanical factors contributing to PACG.

category: bioengineering


title: M-TUBE: a fabrication-free microfluidic device for large-volume bacterial electroporation requiring minimal assembly

abstract: Conventional cuvette-based and microfluidics-based bacterial electroporation approaches have distinct advantages, but they are typically limited to relatively small sample volumes, reducing their utility for applications requiring high throughput such as the generation of mutant libraries. Here, we present a disposable, user-friendly microfluidic electroporation device capable of processing large volume bacterial samples yet requiring minimal device fabrication and straightforward operation. We demonstrate that the proposed device can outperform conventional cuvettes in a range of situations, including across Escherichia coli strains with a range of electroporation efficiencies, and we use its large-volume bacterial electroporation capability to generate a library of transposon mutants in the anaerobic gut commensal Bifidobacterium longum.


title: Automated atlas-based multi-label fetal cardiac vesselsegmentation in Congenital Heart Disease

abstract:
Congenital heart disease (CHD) is the most commonly diagnosed birth defect. T2w black blood MRI provides optimal vessel visualisation, aiding prenatal CHD diagnosis. Common clinical practice involves manual segmentation of fetal heart and vessels for visualisation and reporting purposes.

We propose an automated multi-label fetal cardiac vessels deep learning segmentation approach for T2w black blood MRI. Our network is trained using single-label manual segmentations obtained through current clinical practice, combined with a multi-label anatomical atlas with desired multi-label segmentation protocol. Our framework combines deep learning label propagation with 3D residual U-Net segmentation to produce high-quality multi-label output well adapted to the individual subject anatomy.

We train and evaluate the network using forty fetal subjects with suspected coarctation of the aorta, achieving a dice score of 0.79 {+/-} 0.02 for the fetal cardiac vessels region. The proposed network outperforms the label propagation and achieves a statistically equivalent performance to a 3D residual U-Net trained exclusively on manual single-label data (p-value>0.05). This multi-label framework therefore represents an advancement over the single-label approach, providing label-specific anatomical information, particularly useful for assessing specific anomaly areas in CHD.


title: conST: an interpretable multi-modal contrastive learning framework for spatial transcriptomics

abstract:
MotivationSpatially resolved transcriptomics (SRT) shows its impressive power in yielding biological insights into neuroscience, disease study, and even plant biology. However, current methods do not sufficiently explore the expressiveness of the multi-modal SRT data, leaving a large room for improvement of performance. Moreover, the current deep learning based methods lack interpretability due to the "black box" nature, impeding its further applications in the areas that require explanation.

ResultsWe propose conST, a powerful and flexible SRT data analysis framework utilizing contrastive learning techniques. conST can learn low-dimensional embeddings by effectively integrating multi-modal SRT data, i.e. gene expression, spatial information, and morphology (if applicable). The learned embeddings can be then used for various downstream tasks, including clustering, trajectory and pseudotime inference, cell-to-cell interaction, etc. Extensive experiments in various datasets have been conducted to demonstrate the effectiveness and robustness of the proposed conST, achieving up to 10% improvement in clustering ARI in the commonly used benchmark dataset. We also show that the learned embedding can be used in complicated scenarios, such as predicting cancer progression by analyzing the tumour microenvironment and cell-to-cell interaction (CCI) of breast cancer. Our framework is interpretable in that it is able to find the correlated spots that support the clustering, which matches the CCI interaction pairs as well, providing more confidence to clinicians when making clinical decisions.

category: bioinformatics


title: SELINA: Single-cell Assignment using Multiple-Adversarial Domain Adaptation Network with Large-scale References

abstract: The rapid accumulation of single-cell RNA-seq data has provided rich resources to characterize various human cell types. Cell type annotation is the critical step in analyzing single-cell RNA-seq data. However, accurate cell type annotation based on public references is challenging due to the inconsistent annotations, batch effects, and poor characterization of rare cell types. Here, we introduce SELINA (single cELl identity NAvigator), an integrative annotation transferring framework for automatic cell type annotation. SELINA optimizes the annotation for minority cell types by synthetic minority over-sampling, removes batch effects among reference datasets using a multiple-adversarial domain adaptation network (MADA), and fits the query data with reference data using an autoencoder. Finally, SELINA affords a comprehensive and uniform reference atlas with 1.7 million cells covering 230 major human cell types. We demonstrated the robustness and superiority of SELINA in most human tissues compared to existing methods. SELINA provided a one-stop solution for human single-cell RNA-seq data annotation with the potential to extend for other species.


title: The minimizer Jaccard estimator is biased and inconsistent

abstract:
MotivationSketching is now widely used in bioinformatics to reduce data size and increase data processing speed. Sketching approaches entice with improved scalability but also carry the danger of decreased accuracy and added bias. In this paper, we investigate the minimizer sketch and its use to estimate the Jaccard similarity between two sequences.

ResultsWe show that the minimizer Jaccard estimator is biased and inconsistent, which means that the expected difference (i.e., the bias) between the estimator and the true value is not zero, even in the limit as the lengths of the sequences grow. We derive an analytical formula for the bias as a function of how the shared k-mers are laid out along the sequences. We show both theoretically and empirically that there are families of sequences where the bias can be substantial (e.g. the true Jaccard can be more than double the estimate). Finally, we demonstrate that this bias affects the accuracy of the widely used mashmap read mapping tool.

AvailabilityScripts to reproduce our experiments are available on GitHub [26].

Contactpzm11@psu.edu


title: Activity of Estrogen Receptor β Agonists in Therapy-Resistant Estrogen Receptor-Positive Breast Cancer

abstract:
BackgroundAmong women, breast cancer is the leading cause of cancer-related death worldwide. Estrogen receptor  positive (ER+) breast cancer accounts for 70% of all breast cancer subtypes. Although ER+ breast cancer initially responds to estrogen deprivation or blockade, resistance emergence compelling the use of more aggressive therapies. While ER is a driver in ER+ breast cancer, ER{beta} plays an inhibitory role in several different cancer types. To date, the lack of highly selective ER{beta} agonists without ER activity has limited the exploration of ER{beta} activation as a strategy for ER+ breast cancer.

MethodsWe measured expression levels of ESR1 and ESR2 genes in immortalized mammary epithelial cells and different breast cancer cell lines. The viability of ER+ breast cancer cell lines upon treatments with specific ER{beta} agonists, including OSU-ERb-12 and LY500307 was assessed. The specificity of the ER{beta} agonists, OSU-ERb-12 and LY500307, was confirmed by reporter assays. The effects of the agonists on cell proliferation, cell cycle, apoptosis, colony formation, cell migration, and expression of tumor suppressor proteins were analyzed. The expression of ESR2 and genes containing ERE-AP1 composite response elements was examined in ER+ human breast cancer samples to determine the correlation between ESR2 expression and overall survival and that of putative ESR2 regulated genes.

ResultsIn this study, we demonstrate the efficacy of highly selective ER{beta} agonists in ER+ breast cancer cell lines and drug-resistant derivatives. ER{beta} agonists blocked cell proliferation, migration and colony formation; and induced apoptosis and S and/or G2/M cell cycle arrest of ER+ breast cancer cell lines. Also, increases in the expression of the key tumor suppressors FOXO1 and FOXO3a were noted. Importantly, the strong synergy between ER{beta} agonists and ER antagonists suggested that the efficacy of ER{beta} agonists is maximized by combination with ER blockade. Lastly, ESR2 (ER{beta} gene) expression was negatively correlated with ESR1 (ER gene) and CCND1 RNA expression in human metastatic ER+/HER2-breast cancer samples.

ConclusionOur results demonstrate that highly selective ER{beta} agonists attenuate the viability of ER+ breast cancer cell lines in vitro and suggest that this therapeutic strategy merits further evaluation for ER+ breast cancer.

category: cancer biology


title: Intracortical microstimulation pulse waveform and frequency recruits distinct spatiotemporal patterns of cortical neuron and neuropil activation.

abstract:
BackgroundNeural prosthetics often use intracortical microstimulation (ICMS) for sensory restoration. To restore natural and functional feedback, we must first understand how stimulation parameters influence the recruitment of neural populations. ICMS waveform asymmetry modulates the spatial activation of neurons around an electrode at 10 Hz; however, it is unclear how asymmetry may differentially modulate population activity at frequencies typically employed in the clinic (e.g. 100 Hz).

ObjectiveWe hypothesized that stimulation waveform asymmetry would differentially modulate preferential activation of certain neural populations, and the differential population activity would be frequency-dependent.

MethodsWe quantified how asymmetric stimulation waveforms delivered at 10 Hz or 100 Hz for 30s modulated spatiotemporal activity of cortical layer II/III pyramidal neurons using in vivo two-photon and mesoscale calcium imaging in anesthetized mice. Asymmetry is defined in terms of the ratio of the leading phase to the return phase of charge-balanced cathodal- and anodal-first waveforms.

ResultsNeurons within 40-60m of the electrode display stable stimulation-induced activity indicative of direct activation, which was independent of waveform asymmetry. The stability of 72% of activated neurons and the preferential activation of 20-90 % of neurons depended on waveform asymmetry. Additionally, this asymmetry-dependent activation of different neural populations was associated with differential progression of population activity. Specifically, neural activity increased over time for some waveforms at 10 Hz, but decreased more at 100 Hz than other waveforms.

ConclusionThese data demonstrate that at frequencies commonly used for sensory restoration, stimulation waveform alters the pattern of activation of different but overlapping populations of excitatory neurons. The impact of these waveform specific responses on the activation of different subtypes of neurons as well as sensory perception merits further investigation.

category: neuroscience


title: Somatodendritic release of cholecystokinin potentiates GABAergic synapses onto ventral tegmental area dopamine cells

abstract: Neuropeptides are contained in nearly every neuron in the central nervous system and can be released from somatodendritic sites as well as from nerve terminals. Cholecystokinin (CCK), among the most abundant neuropeptides in the brain, is expressed in the majority of midbrain dopamine neurons. Here we report that ventral tegmental area (VTA) dopamine neurons release CCK from somatodendritic regions, where it triggers long-term potentiation of GABAergic synapses. The somatodendritic release occurs with trains of action potentials or prolonged but modest depolarization and is dependent on synaptotagmin 7 and T-type Ca2+ channels. Depolarization-induced LTP is blocked by the CCK2R antagonist, LY225910, and mimicked by exogenously added CCK. To test the behavioral role of CCK, we infused it into the mouse VTA. Ca2+ imaging in vivo demonstrated that infused CCK reduced dopamine cell signals during fasted food consumption. Moreover, local infusion of CCK also inhibited food consumption and decreased distance traveled in an open field test. Together our experiments introduce somatodendritic neuropeptide release as a previously unknown feedback regulator of VTA dopamine cell excitability and dopamine-related behaviors.


title: 3D multi-color far-red single-molecule localization microscopy with probability-based fluorophore classification

abstract: Single-Molecule Localization Microscopy remains limited in its ability for robust and simple multi-color imaging. Whereas the fluorophore Alexa647 is widely used due to its brightness and excellent blinking dynamics, other excellent blinking fluorophores, such as CF660 and CF680, spectrally overlap. Here we present Probability-based Fluorophore Classification, a method to perform multi-color SMLM with Alexa647, CF660 and CF680 that uses statistical decision theory for optimal classification. The emission is split in a short and long wavelength channel to enable classification and localization without a major loss in localization precision. Each emitter is classified using a Generalized Maximum Likelihood Ratio Test using the photon statistics of both channels. This easy-to-adopt approach does not require nanometer channel registration, is able to classify fluorophores with tunable low false positive rates (<0.5%) and optimal true positive rates and outperforms traditional ratiometric spectral de-mixing and Salvaged Fluorescence. We demonstrate its applicability on a variety of samples and targets.

category: cell biology


title: Temporal control of late replication and coordination of origin firing by self-stabilizing Rif1-PP1 hubs in Drosophila

abstract:
In the metazoan S phase, coordinated firing of clusters of origins replicates different parts of the genome in a temporal program. Despite advances, neither the mechanism controlling timing nor that coordinating firing of multiple origins is fully understood. Rif1, an evolutionarily conserved inhibitor of DNA replication, recruits protein phosphatase 1 (PP1) and counteracts firing of origins by S-phase kinases. During the mid-blastula transition (MBT) in Drosophila embryos, Rif1 forms subnuclear hubs at each of the large blocks of satellite sequences and delays their replication. Each Rif1 hub disperses abruptly just prior to the replication of the associated satellite sequences. Here, we show that the level of activity of the S-phase kinase, DDK, accelerated this dispersal program, and that the level of Rif1-recruited PP1 retarded it. Further, Rif1-recruited PP1 supported chromatin association of nearby Rif1. This influence of nearby Rif1 can create a "community effect" counteracting kinase-induced dissociation such that an entire hub of Rif1 undergoes switch-like dispersal at characteristic times that shift in response to the balance of Rif1-PP1 and DDK activities. We propose a model in which the spatiotemporal program of late replication in the MBT embryo is controlled by self-stabilizing Rif1-PP1 hubs, whose abrupt dispersal synchronizes firing of associated late origins.

SIGNIFICANCESeventy years ago, it was discovered that large domains of the eukaryotic genome replicate at different times. Detailed descriptions left significant questions unresolved. How are the many origins in the large domains coordinated to fire in unison? What distinguishes different domains and gives rise to a temporal program? When Drosophila embryos first establish late replication, an inhibitor of DNA replication, Rif1, forms hubs over domains of late replicating DNA. Rif1 recruits protein phosphatase 1 (PP1), which prevents kinases from dispersing Rif1 hubs or activating associated origins. When kinase activity eventually exceeds a hub-specific threshold, the self-stabilization of Rif1-PP1 breaks down, hubs disperse abruptly, and all associated origins are free to initiate.


title: Use of object detection in camera trap image identification: assessing a method to rapidly and accurately classify human and animal detections for research and application in recreation ecology

abstract:
Camera traps are increasingly used to answer complex ecological questions. However, the rapidly growing number of images collected presents technical challenges. Each image must be classified to extract data, requiring significant labour, and potentially creating an information bottleneck. We applied an object-detection model (MegaDetector) to camera trap data from a study of recreation ecology in British Columbia, Canada. We tested its performance in detecting humans and animals relative to manual image classifications, and assessed efficiency by comparing the time required for manual classification versus a modified workflow integrating object-detection with manual classification. We also evaluated the reliability of using MegaDetector to create an index of human activity for application to the study of recreation impacts to wildlife. In our application, MegaDetector detected human and animal images with 99% and 82% precision, and 95% and 92% recall respectively, at a confidence threshold of 90%. The overall time required to process the dataset was reduced by over 500%, and the manual processing component was reduced by 840%. The index of human detection events from MegaDetector matched the output from manual classification, with a mean 0.45% difference in estimated human detections across site-weeks. Our test of an open-source object-detection model showed it performed well in partially classifying a camera trap dataset, significantly increasing processing efficiency. We suggest that this tool could be integrated into existing camera trap workflows to accelerate research and application by alleviating data bottlenecks, particularly for surveys processing large volumes of human images. We also show how the model and workflow can be used to anonymize human images prior to classification, protecting individual privacy.

Impact StatementWe developed and tested a workflow for classifying camera trap images that integrated an existing object-detection model with manual image classification. Our workflow demonstrates an increase in efficiency of 500% over manual labelling, and additionally includes a method to anonymize human images prior to archiving and classification. We provide an example of the application of these tools to ease data processing, particularly for studies focused on recreation ecology which record high volumes of human images. Data lags due to processing delays have the potential to result in sub-optimal conservation decisions, which may be alleviated by accelerated processing. To our knowledge, this is the first in-depth assessment of the practical application of such technology to real world workflows focused on human detections.


title: Assessing the global impact of targeted conservation actions on species abundance

abstract: In recent years, vertebrate population abundance has declined at unprecedented rates (WWF 2020). In response, targeted conservation measures - such as breeding programs or species-specific habitat management - have been applied to halt population declines, aid population recovery, and reduce and reverse the loss of biodiversity (Salafsky et al. 2008; Bolam et al. 2020). Until now, assessments of conservation actions have focused on the extent to which they reduce extinction risk, impact populations within protected areas, or increase the global area of land under protection (Hoffmann et al. 2010, 2015; Barnes et al. 2016; Maxwell et al. 2020; Bolam et al. 2020; Grace et al. 2021a). Here, we record and analyze conservation actions for 26,904 vertebrate populations from 4,629 species, to measure the impact of targeted conservation on vertebrate abundance. Using a counterfactual approach to represent population trends in the absence of conservation, we demonstrate that targeted actions have delivered substantial positive effects on the abundance of recipient vertebrate populations worldwide. We show that, in the absence of conservation, a global indicator of vertebrate abundance would have declined even more. Positive population trends were associated with vertebrate populations subject to species or habitat management. We demonstrate that targeted conservation actions can help to reverse global biodiversity loss and show the value of counterfactual analysis for impact evaluation - an important step towards reversing biodiversity declines.


title: Experimental characterisation of de novoproteins and their unevolved random-sequencecounterparts

abstract: De novo gene emergence provides a route for new proteins to be formed from previously non-coding DNA. Proteins born in this way are considered random sequences, and typically assumed to lack defined structure. While it remains unclear how likely a de novo protein is to assume a soluble and stable tertiary structure, intersecting evidence from random-sequence and de novo-designed proteins suggests that native-like biophysical properties are abundant in sequence space. Taking putative de novo proteins identified in human and fly, we experimentally characterise a library of these sequences to assess their solubility and structure propensity. We compare this library to a set of synthetic random proteins with no evolutionary history. Bioin-formatic prediction suggests that de novo proteins may have remarkably similar distributions of biophysical properties to unevolved random sequences of a given length and amino acid composition. However, upon expression in vitro, de novo proteins exhibit higher solubility which is further induced by the DnaK chaperone system. We suggest that while synthetic ran-dom sequences are a useful proxy for de novo proteins in terms of structure propensity, de novo proteins may be better integrated in the cellular system given their higher solubility.

category: evolutionary biology


title: SLiMAn: an integrative web server for exploring short linear motif-mediated interactions in interactomes.

abstract: Cells express thousands of macromolecules, and their functioning relies on multiple networks of intermolecular interactions. These interactions can be experimentally determine at different spatial and temporal resolutions. But, physical interfaces are not often delineated directly especially in high-throughput experiments. However, numerous three-dimensional structures of complexes have been already solved and sequence conservation allows comparative modeling of additional complexes. A large fraction of protein-protein interactions involves domain and so-called SLiMs (for Short Linear Motifs). Often, SLiMs lie in disordered regions or loops. Their small size and loosely folded nature prevent straightforward detection. SLiMAn (Short Linear Motif Analysis), a new web server is provided to help thorough analysis of interactomics data. Starting from a list of putative interactants such as the output of an interactomics study, SLiMs (from ELM) and SLiM-recognition domains (from Pfam) are extracted and potential pairing are displayed. Additionally, filters are available to dig into the predicted results such as the motif E-value, IUpred2 scoring functions for disorder or BioGRID interaction matches. When structural templates are available, a given SLiM and its recognition domain can be modeled using SCWRL. We illustrate, here, the use of SLiMAn on three distinct examples including one real-case study. We oversee wide-range applications for SLiMAn in the context of massive analysis of protein-protein interactions at proteome-wide scale. This new web server is made freely available at http://sliman.cbs.cnrs.fr.


title: Automatic grading of cervical biopsies by combining full and self-supervision

abstract:
In computational pathology, the application of Deep Learning to the analysis of Whole Slide Images (WSI) has provided results of unprecedented quality. Due to their enormous size, WSIs have to be split into small images (tiles) which are first encoded and whose representations are then agglomerated in order to solve prediction tasks, such as prognosis or treatment response. The choice of the encoding strategy plays a key role in such algorithms. Current approaches include the use of encodings trained on unrelated data sources, full supervision or self-supervision. In particular, self-supervised learning (SSL) offers a great opportunity to exploit all the unlabelled data available. However, it often requires large computational resources and can be challenging to train. On the other end of the spectrum, fully-supervised methods make use of valuable prior knowledge about the data but involve a costly amount of expert time.

This paper proposes a framework to reconcile SSL and full supervision and measures the trade-off between long SSL training and annotation effort, showing that a combination of both has the potential to substantially increase performance. On a recently organized challenge on grading Cervical Biopsies, we show that our mixed supervision scheme reaches high performance (weighted accuracy (WA): 0.945), outperforming both SSL (WA: 0.927) and transfer learning from ImageNet (WA: 0.877). We further provide insights and guidelines to train a clinically impactful classifier with a limited expert and/or computational workload budget. We expect that the combination of full and self-supervision is an interesting strategy for many tasks in computational pathology and will be widely adopted by the field.


title: A biphasic Deep Semi-supervised framework for Suptype Classification and biomarker discovery

abstract: To take full advantage of the unprecedented development of -omics technologies and generate further biological insights into human disease, it is a pressing need to develop novel computational methods for integrative analysis of multi-omics data. Here we proposed a biphasic Deep Semi-supervised multi-omics integration framework for Subtype Classification and biomarker discovery, DeepSSC. In phase 1, each denoising autoencoder was used to extract a compact representation for each -omics data, and then they were concatenated and put into a feed-forward neural network for subtype classification. In phase 2, our Biomarker Gene Identification procedure leveraged that neural network classifier to render subtype-specific important biomarkers. We also validated our given results on independent dataset. We demonstrated that DeepSSC exhibited better performance over other state-of-the-art techniques concerning classification tasks. As a result, DeepSSC successfully detected well-known biomarkers and hinted at novel candidates from different -omics data types related to the investigated biomedical problems.


title: Generic principles of space compartmentalization in protocell patterns

abstract: Self-organization of cells into higher-order structures is key for multicellular organisms, e.g. during embryonic epithelium formation via repetitive replication of template-like founder cells. Yet, very similar spatial arrangements of cell-like compartments (protocells) are also seen in cell extracts in the absence of template structures and genetic material. Here we show that protocell patterns are highly organized, featuring a spatial arrangement and coarsening like two-dimensional foams but without signatures of disordered hyperuniformity. These features even remain unaffected when enforcing smaller protocells by stabilizing microtubule filaments. Comparing our data to generic models, we conclude that protocell patterns emerge by simultanous formation of randomly placed seeds that grow at a uniform rate until fusion of adjacent protocells drives coarsening. The strong similarity of our observations to the recently reported organization of epithelial monolayers suggests common generic principles for space allocation in living matter.

category: biophysics


title: Glycation renders alpha-synuclein oligomeric strain and modulates microglia activation

abstract: -Synuclein is known to involve in the pathogenesis of Parkinsons diseases (PD) and related disorders. However, it is unclear how its aggregation causes neuronal degeneration and neuroinflammation. Due to intrinsic disorder nature, -synuclein produces a large number of structural ensembles and diverse aggregation intermediates. The post-translational modifications add a new layer of complexity to the aggregation mechanism. Recently, it has been demonstrated that glycation of -synuclein restricts into oligomeric intermediates and causes neuronal toxicity. However, the understanding of aggregation mechanism, dopaminergic neuronal death, and neuroinflammation by the glycated -synuclein is yet to be elucidated. The present study aims to address how glycated synuclein differs in oligomerization and neuroinflammation. The glycation of -synuclein perturbs the aggregation kinetics and prevents the fibrilization through the alteration of surface charges of N-terminal domain residues which prevents membrane binding and seed amplification mechanism. Mass spectrometry-based proteomics analysis of BV2 cells treated with glycated oligomers provides evidence of alteration of endocytic mechanism, mitochondrial dysfunction, and inflammatory cascade. Here, we show that -synuclein oligomers strongly bind to TLR2 and activate the TLR2 mediated signaling. However, glycated -synuclein oligomers impair the TLR2 binding and compromise TLR2 signaling. Interestingly, we also find that the glycated -synuclein oligomers favor NLRP3 inflammasome mediated neuroinflammation compared to non- glycated -synuclein oligomers. In conclusion, our findings suggest that microglia response towards -synuclein is conformation-specific and glycated oligomers can contribute to neurodegeneration differently.


title: Absence of the fragile X mental retardation protein alters response patterns to sounds in the auditory midbrain

abstract: Among the different autism spectrum disorders, Fragile X Syndrome (FXS) is the most common inherited cause of mental retardation. Sensory and especially auditory hypersensitivity is a key symptom in patients, which is well mimicked in the Fmr1 -/- mouse model. However, the physiological mechanisms underlying FXSs acoustic hypersensitivity in particular remain poorly understood. Here, we categorized spike response patterns to pure tones of different frequencies and intensities from neurons in the Inferior Colliculus (IC), a central integrator in the ascending auditory pathway. Based on this categorization we analyzed differences in response patterns between IC neurons of WT and Fmr1 -/- mice. Our results report broadening of frequency tuning, an increased firing in response to monaural as well as binaural stimuli, an altered balance of excitation-inhibition, and reduced response latencies, all expected features of acoustic hypersensitivity. Furthermore, we were surprised to notice that all neuron response types in Fmr1 -/- mice displayed enhanced offset-rebound activity outside their excitatory frequency response area. These results provide evidence that the loss of Fmr1 not only increases spike responses in IC neurons similar to auditory brainstem neurons, but also changes response patterns such as offset spiking. One can speculate this to be an underlying aspect of the receptive language problems associated with Fragile X syndrome.


title: Endothelial transmigration hotspots limit vascular leakage through heterogeneous expression of ICAM1

abstract: Upon inflammation, leukocytes leave the circulation by crossing the endothelial monolayer at specific transmigration  hotspot regions. Although these regions support leukocyte transmigration, their functionality is not clear. We found that endothelial hotspots function to limit vascular leakage during transmigration events. Using the photo-convertible probe mEos4b, we traced back and identified original endothelial transmigration hotspots. Using this method, we show that the heterogeneous distribution of ICAM-1 determines the location of the transmigration hotspot. Interestingly, loss of ICAM-1 heterogeneity either by CRISPR/Cas9-induced knockout of ICAM-1 or equalizing the distribution of ICAM-1 in all endothelial cells results in loss of TEM hotspots but not necessarily in reduced TEM events. Functionally, loss of endothelial hotspots results in increased vascular leakage during TEM. Mechanistically, we demonstrate that the 3 extracellular Ig-like domains of ICAM-1 are crucial for hotspot recognition. However, the intracellular tail of ICAM-1 and the 4th Ig-like dimerization domain are not involved, indicating that intracellular signalling or ICAM-1 dimerization is not required for hotspot recognition. Together, we discovered that hotspots function to limit vascular leakage during inflammation-induced extravasation.

category: immunology


title: Human Mucosal Associated Invariant T cell proliferation is dependent on a MYC-SLC7A5-Glycolysis metabolic axis

abstract:
Mucosal Associated Invariant T (MAIT) cells are an abundant population of innate T cells which recognise bacterial ligands presented by the MHC class-I like molecule MR1. MAIT cells play a key role in host protection against bacterial and viral pathogens. Upon activation MAIT cells undergo proliferative expansion and increased production of effector molecules such as cytokines. The molecular and metabolic mechanisms controlling MAIT cell effector functions are still emerging. In this study, we found that expression of the key metabolism regulator and transcription factor MYC is upregulated in MAIT cells upon immune stimulation. Using quantitative mass spectrometry, we identified the activation of two MYC controlled metabolic pathways; amino acid transport and glycolysis, both of which are critical for MAIT cell proliferation. Finally, we show that MYC expression in response to immune activation is diminished in MAIT cells isolated from people with obesity, resulting in defective MAIT cell proliferation and functional responses. Collectively our data details for the first time the importance of MYC regulated metabolism for MAIT cell proliferation, and provides additional insight into the molecular defects underpinning functional failings of MAIT cells in obesity.

Graphical Abstract

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title: The role of intraspecific competition during post-glacial recolonization: a Norway spruce case study

abstract:
O_LITo a large extent, the current distribution and population structure of many species were shaped by the cycles of isolation in glacial refugia and subsequent population expansions. Isolation in, and post-glacial expansion through, heterogeneous environments led to either neutral or adaptive divergence. Norway spruce is no exception, and its current distribution is the consequence of a complex interplay between evolutionary forces.
C_LIO_LIWe investigated population differentiation and adaptations of Norway spruce for juvenile growth, diameter of the trunk at breast height and its components: wood density and tracheid traits. Data from 4,461 phenotyped and genotyped Norway spruce from 396 half-sib families in two progeny tests were used to test for divergent selection in the framework of QST vs FST, and to carry out multiple Genome-Wide Association Studies.
C_LIO_LIWe show that (i) the macroscopic resultant trait (trunk diameter), unlike its microscopic components (tracheid dimensions) and juvenile growth, was under divergent selection, and (ii) adaptation predated the Last Glaciation Maximum.
C_LIO_LIAltogether, the current variation in these phenotypic traits in Norway spruce is better explained by local adaptation to ancestral environments than to current ones, where populations were partly pre-adapted mainly through growth-related traits.
C_LI


title: Factors behind intraspecific competition during post-glacial recolonization: a Norway spruce case study


title: MeShClust v3.0: High-quality clustering of DNA sequences using the mean shift algorithm and alignment-free identity scores

abstract:
BackgroundTools for accurately clustering biological sequences are among the most important tools in computational biology. Two pioneering tools for clustering sequences are CD-HIT and UCLUST, both of which are fast and consume reasonable amounts of memory; however, there is a big room for improvement in terms of cluster quality. Motivated by this opportunity for improving cluster quality, we applied the mean shift algorithm in MeShClust v1.0. The mean shift algorithm is an instance of unsupervised learning. Its strong theoretical foundation guarantees the convergence to the true cluster centers. Our implementation of the mean shift algorithm in MeShClust v1.0 was a step forward; however, it was not the original algorithm. In this work, we make progress toward applying the original algorithm while utilizing alignment-free identity scores in a new tool: MeShClust v3.0.

ResultsWe evaluated CD-HIT, MeShClust v1.0, MeShClust v3.0, and UCLUST on 22 synthetic sets and five real sets. These data sets were designed or selected for testing the tools in terms of scalability and different similarity levels among sequences comprising clusters. On the synthetic data sets, MeShClust v3.0 outperformed the related tools on all sets in terms of cluster quality. On two real data sets obtained from human microbiome and maize transposons, MeShClust v3.0 outperformed the related tools by wide margins, achieving 55%--300% improvement in cluster quality. On another set that includes degenerate viral sequences, MeShClust v3.0 came third. On two bacterial sets, MeShClust v3.0 was the only applicable tool because of the long sequences in these sets. MeShClust v3.0 requires more time and memory than the related tools; almost all personal computers at the time of this writing can accommodate such requirements. MeShClust v3.0 can estimate an important parameter that controls cluster membership with high accuracy.

ConclusionsThese results demonstrate the high quality of clusters produced by MeShClust v3.0 and its ability to apply the mean shift algorithm to large data sets and long sequences. Because clustering tools are utilized in many studies, providing high-quality clusters will help with deriving accurate biological knowledge.


title: Novel MBLs inhibitors screened from FDA-approved drug library restore the susceptibility of carbapenems to NDM-1-harbouring bacteria

abstract: The production of metallo-{beta}-lactamases (MBLs) is one of the major mechanisms adopted by bacterial pathogens to resist carbapenems. One promising strategy to overcome MBLs-mediated carbapenems resistance is to develop effective inhibitors. Repurposing approved drugs to restore the efficacy of carbapenems represents an efficient and cost-effective approach to fight infections caused by carbapenem resistant pathogens. Here, twelve FDA-approved compounds were screened to neutralize the ability of NDM-1. Among these compounds, dexrazoxane, embelin, candesartan cilexetil (CAN) and nordihydroguaiaretic acid (NDGA) were further demonstrated to inhibit all tested MBLs, and showed an in vitro synergistic bactericidal effect with meropenem against MBLs-producing bacteria. Mechanistic studies revealed that dexrazoxane, embelin and CAN are metal ion chelating agents, while the inhibition of NDM-1 by NDGA involves its direct binding with the active region of NDM-1. Furthermore, dexrazoxane, embelin and CAN and NDGA dramatically rescued the treatment efficacy of meropenem in three infection models. Our observations indicated that dexrazoxane, embelin, CAN and NDGA are promising carbapenem adjuvants against MBLs-positive carbapenem resistant bacterial pathogens.

category: microbiology


title: Rapid exchange of stably bound protein and DNA cargo on a DNA origami receptor

abstract: Biomolecular complexes can form stable assemblies yet can also rapidly exchange their subunits to adapt to environmental changes. Simultaneously allowing for both stability and rapid exchange expands the functional capacity of biomolecular machines and enables continuous function while navigating a complex molecular world. Inspired by biology, we design and synthesize a DNA origami receptor that exploits multi-valent interactions to form stable complexes that are simultaneously capable of rapid subunit exchange. The system utilizes a mechanism first outlined in the context of the DNA replisome, known as multi-site competitive exchange, and achieves a large separation of time scales between spontaneous subunit dissociation, which requires days, and rapid subunit exchange, which occurs in minutes. In addition, we use the DNA origami receptor to demonstrate stable interactions with rapid exchange of both DNA and protein subunits, thus highlighting the applicability of our approach to arbitrary molecular cargo; an important distinction with canonical toehold exchange between single-stranded DNA. We expect this study to be the first of many that use DNA origami structures to exploit multi-valent interactions for the design and synthesis of a wide range of possible kinetic behaviors.

category: synthetic biology


title: Meta-analysis of the black soldier fly (Hermetia illucens) microbiota based on 16S rRNA gene amplicon sequencing

abstract: Black soldier fly larvae (BSFL) belong to the most widely reared insect species as an alternative protein source at industrial scale. Bacteria in the larval gut can provide benefits for the animal, though some bacteria can also be pathogenic for the insect. Accurate characterization of the BSFL microbiota is important for the production of BSFL in terms of yield and microbiological safety. In this study, 16S ribosomal RNA gene sequence data sets from 11 studies were re-analysed to gain better insights in the BSFL gut microbiota, potential factors that influence their composition, and differences between the gut and the whole larvae microbiota. A core gut microbiota was found consisting of members of Enterococcus, Klebsiella, Morganella, Providencia, and Scrofimicrobium. Further, the factors "Study", "Age" and "Feed" significantly affected the microbiota gut composition. When compared to whole larvae, a significantly lower number of observed zero-radius Operational Taxonomic Units and a lower diversity was found for gut samples, suggesting that the larvae harboured additional microbes on their cuticle or in the insect body. Universal choices in insect sample type, primer selection and bio-informatics can strengthen future meta-analyses and improve our understanding of the BSFL gut microbiota towards the optimization of insect production.


title: Conserved Untranslated Regions of Multipartite Viruses: Natural Markers of Novel Viral Genomic Components and Tags of Viral Evolution

abstract:
Viruses with split genomes are categorized as being either segmented or multipartite according to whether their genomic segments occur in single or different virions. Some complexity will exist, in that inherited "core" vital segments viruses may renew the others once host and environmental alterations keep driving viral evolution. Despite this uncertainty, empirical observations have been made across the split genomes in the untranslated regions (UTRs) on the short or long stretches of conserved or identical sequences. In this study, we describe a methodology that combines RNA and small RNA sequencing, conventional BLASTx, and iterative BLASTn of UTRs to detect viral genomic components even if they encode orphan genes (ORFans). Within the phylum Kitrinoviricota, novel putative multipartite viruses and viral genomic components were annotated using data obtained from our sampling or publicly available sources. The novel viruses, as extensions or intermediate nodes, enriched the information of the evolutionary networks. Furthermore, the diversity of novel genomic components emphasized the evolutionary roles of reassortment and recombination, as well as genetic deletion, strongly supporting the genomic complexity. These data also suggest insufficient knowledge of these genomic components for categorizing some extant viral taxa. The relative conservation of UTRs at the genome level may explain the relationships between monopartite and multipartite viruses and how the multipartite viruses can have a life strategy involving multiple host cells.

Author summaryThe current workflows for virus identification are largely based on high-throughput sequencing and coupled protein sequence homology-dependent analysis methods and tools. However, for viruses with split genomes, the identification of genomic components whose deduced protein sequences are not homologous to known sequences is inadequate. Furthermore, many plant-infecting multipartite viruses contain conserved UTRs across their genomic components. Based on this, we propose a practical method of UTR-backed iterative BLASTn (UTR-iBLASTn) to explore the components with ORFans and study virus evolution using the UTRs as signals. These shed light on viral "dark matter"--unknown/omitted genomic components of segmented/multipartite viruses from different kingdoms and hosts, and the origins of these components.


title: Out of the blue: the independent activity of sulfur-oxidizers and diatoms mediate the sudden color shift of a tropical river

abstract: Rio Celeste ("Sky-Blue River") is a river located in the Tenorio National Park (Costa Rica) that has become an important hotspot for eco-tourism due to its striking sky-blue color. A previous study suggested that this color is not caused by dissolved chemical species, but by precipitation of lightscattering aluminosilicate particles at the mixing point of two colorless streams, the acidic Quebrada Agria and the neutral Rio Buenavista. We now present microbiological information on Rio Celeste and its two tributaries, as well as a more detailed characterization of the particles that occur at the mixing point. Our results overturn the previous belief that the light scattering particles are formed by the aggregation of smaller particles coming from Rio Buenavista, and rather point to chemical formation of hydroxyaluminosilicate colloids with Quebrada Agria as the main contributor to the phenomenon, with Rio Buenavista acting as a secondary source of silica to the reaction. We also show how the sky-blue color of Rio Celeste arises from the tight interaction between chemical and biological processes. Sulfur-oxidizing bacteria generate an acidic environment in Quebrada Agria, which in turn cause dissolution and mobilization of aluminum and other metals, while in Rio Buenavista the growth of diatoms transforms dissolved silicon into colloidal biogenic forms. The local interaction between these two well-known biological activities gives rise to the unique Rio Celeste phenomenon, in what constitutes a textbook example of emergent behavior in environmental microbiology.


title: Using strain-resolved analysis to identify contamination in metagenomics data

abstract: Metagenomics analyses can be negatively impacted by DNA contamination. While external sources of contamination such as DNA extraction kits have been widely reported and investigated, contamination originating within the study itself remains underreported. Here we applied high-resolution strain-resolved analyses to identify contamination in two large-scale clinical metagenomics datasets. By mapping strain sharing to DNA extraction plates, we identified well-to-well contamination in both negative controls and biological samples in one dataset. Such contamination is more likely to occur among samples that are on the same or adjacent columns or rows of the extraction plate than samples that are far apart. Our strain-resolved workflow also reveals the presence of externally derived contamination, primarily in the other dataset. Overall in both datasets, contamination is more significant in samples with lower biomass. Our work demonstrates that genome-resolved strain tracking, with its essentially genome-wide nucleotide-level resolution, can be used to detect contamination in sequencing-based microbiome studies. Our results underscore the value of strain-specific methods to detect contamination and the critical importance of looking for contamination beyond negative and positive controls.


title: Mod(mdg4) variants repress telomeric retrotransposonHeT-A by blocking subtelomeric enhancers

abstract: Telomeres in Drosophila are composed of sequential non-LTR retrotransposons: HeT-A, TART, and TAHRE. Although they are repressed by the piRNA pathway in the germline, how these retrotransposons are regulated in somatic cells is poorly understood. Here, we show that specific splice variants of Mod(mdg4) repress HeT-A by blocking subtelomeric enhancers in ovarian somatic cells. We found that the Mod(mdg4)-N variant represses HeT-A expression most efficiently among the variants. Mod(mdg4)-N mutant flies show elevated HeT-A expression and female sterility. Mod(mdg4)-N-binding subtelomeric sequences exhibit enhancer-blocking activity, and recruitment of RNA polymerase II (Pol II) on subtelomeres by Mod(mdg4)-N is essential for this enhancer-blocking. Moreover, Mod(mdg4)-N functions to form chromatin boundaries of higher-order chromatin conformation but this mechanism is independent of its Pol II recruitment activity at telomeres/subtelomeres. This study provides a link between enhancer-blocking and telomere regulation, and two different molecular mechanisms exhibited by an insulator protein to orchestrate precise gene expression.

category: molecular biology


title: Live-cell fluorescence lifetime multiplexing using synthetic fluorescent probes

abstract: Fluorescence lifetime multiplexing requires fluorescent probes with distinct fluorescence lifetimes but similar spectral properties. Even though synthetic probes for many cellular targets are available for multicolor live-cell fluorescence microscopy, few of them have been characterized for their use in fluorescence lifetime multiplexing. Here we demonstrate that from a panel of 18 synthetic probes, eight pairwise combinations are suitable for fluorescence lifetime multiplexing in living mammalian cell lines. Moreover, combining multiple pairs in different spectral channels enables us to image up to six different biological targets, effectively doubling the number of observable targets. The combination of synthetic probes with fluorescence lifetime multiplexing is thus a powerful approach for live-cell imaging.


title: The central role of the interspecific interactions in the evolution of microbial communities

abstract: The interspecific interactions play an important role in the establishment of a community phenotype. Furthermore, the evolution of a community can not only occur through an evolution of the species composing the community but also of the interactions among them. In this study, we investigated how widespread was the evolution of interspecific interactions in the evolutionary response of eight two-bacterial species communities regarding productivity. We found evidence for an evolution of the interactions in half of the studied communities which gave rise to a mean change of 15% in community productivity as compared to what was expected from the individual responses. Even when the interactions did not evolve themselves, they influenced the evolutionary responses of the bacterial strains within the communities which further affected community response. We found that the evolution within a community often promoted an adaptation of the bacterial strains to the abiotic environment, especially for the dominant strain in a community. Overall, this study suggested that the evolution of the interspecific interactions was frequent and that it could increase community response to evolution. We propose that the existence of an evolution of the interspecific interactions can justify the consideration of the community as a unit of selection.


title: Palatal morphology predicts the paleobiology of early salamanders

abstract: Ecological preferences and life history strategies have enormous impacts on the evolution and phenotypic diversity of salamanders, but the yet established reliable ecological indicators hinder investigations on the paleobiology of early salamanders. Here we statistically demonstrate using geometric morphometric analysis that both the shape of the palate and many non-shape variables particularly associated with vomerine teeth are ecologically informative in early stem- and basal crown-group salamanders. The morphology of the palate is heavily impacted by convergence constrained by feeding mechanisms and also exhibits clear stepwise evolutionary patterns with alternative phenotypic designs to cope with similar functional demand. Paleoecological disparities in early salamanders had took place before the Middle Jurassic and have achieved all ecological preferences in the Early Cretaceous. Metamorphosis is significant in the expansion of ecomorphospace of the palate in early salamanders. The common ancestor of salamanders is metamorphosed and terrestrial, and share unified lifestyles with other modern amphibians.


title: Thermodynamic analysis of Zα domain-nucleic acid interactions

abstract: DNA/RNA molecules adopting the left-handed conformation (Z-form) have been attributed with immunogenic properties. However, their biological role and importance has been a topic of debate for many years. The discovery of Z-DNA/RNA binding domains (Z domains) in varied proteins that are involved in the innate immune response, such as the interferon inducible form of the RNA editing enzyme ADAR1 (p150), Z-DNA binding protein 1 (ZBP1), the fish kinase PKZ and the poxvirus inhibitor of interferon response E3L, indicates important roles of Z-DNA/RNA in immunity and self/non-self-discrimination. Such Z domain-containing proteins recognise left-handed Z-DNA/RNA in a conformation-specific manner. Recent studies have implicated these domains in virus recognition. Given these important emerging roles for the Z domains, it is pivotal to understand the mechanism of recognition of the Z-DNA/Z-RNA by these domains. To this end, we assessed the binding thermodynamics of Z domain from ORF112 and ADAR1 on T(CG)3 and T(CG)6 oligonucleotides which have high propensity to adopt the Z-conformation. Our study highlights important differences in the mode of binding by the two Z domains originating from different proteins. Site-directed mutagenesis was employed together with isothermal titration calorimetry to tease apart finer details of the binding thermodynamics. Our work advances the understanding on binding thermodynamics of Z domains to their cognate nucleic acid substrates and contributes to the efforts to gain a complete appreciation of this process.

category: biochemistry


title: Crystal Structure of Caskin1/CASK complex reveals the molecular basis of the binding specificity of CASK_CAMK domain and its binding partners

abstract: CASK is a unique scaffold protein in the synapse system. It links numerous proteins to the pre- or post-synaptic region and is critical to the homeostasis of synaptic vesicles. The N-terminus of CASK is a calcium/calmodulin-dependent protein kinase (CAMK) domain, which has diverse functions and interacts with downstream proteins to form a scaffold platform. Caskin1 is one of the brain-specific adaptor proteins of CASK. Previous studies showed that CASK_CAMK domain interacts with Caskin1 CID domain with relatively low affinity. In this study, we re-visit this interaction by remapping the interaction boundary and solving their complex structure. Based on the structure, we systematically compared the interactions between CASK_CAMK and other binding partners. Our results showed that CAMK domain occupies the CID peptide by using its C-lobe groove (between the 1 and 2) and there is a highly conserved signature motif ({zeta}-x-{psi}-W-{psi}-x-R) in the CID domain, where {zeta} is acidic side chain containing residues, x is any amino acid residue, {psi} is hydrophobic residues, W is for tryptophan, and R is arginine. These findings allowed us to identify several new potential cytoplasmic binding partners for CASK_CAMK.


title: An efficient simplified method for the generation of corneal epithelial cells from human pluripotent stem cells

abstract: Corneal epithelial cells derived from human pluripotent stem cells (hPSCs) are an important cell source for preclinical models to test ophthalmic drugs. However, current differentiation protocols lack instructions regarding optimal culturing conditions, which hinders the quality of cells and limits scale-up. Here, we introduce a simplified small molecule-based corneal induction method (SSM-CI) to generate corneal epithelial cells from hPSCs. SSM-CI provides the advantage of minimizing cell culturing time using two defined culturing media containing TGF-{beta}, and Wnt/{beta}-catenin pathway inhibitors, and bFGF growth factor over 25 days. Compared to the conventional human corneal epithelial cell line (HCE-T) and human primary corneal epithelial cells (hPCEpCs), corneal epithelial cells generated by SSM-CI are well-differentiated and express relevant maturation markers, including PAX6 and CK12. RNA-seq analysis indicated the faithful differentiation of hPSCs into corneal epithelia, with significant upregulation of corneal progenitor and adult corneal epithelial phenotypes. Furthermore, despite the initial inhibition of TGF-{beta} and Wnt/{beta}-catenin, upregulation of these pathway-related transcripts was observed in the later stages, indicating their necessity in the generation of mature corneal epithelial cells. Moreover, we observed a shift in gene signatures associated with the metabolic characteristics of mature corneal epithelial cells, involving a decrease in glycolysis- and an increase in fatty acid oxidation. This was also attributed to the overexpression of metabolic enzymes and transporter-related transcripts responsible for fatty acid metabolism. Thus, SSM-CI provides a comprehensive method for the generation of functional corneal epithelial cells for use in preclinical models.


title: piggyBac-system-mediated genomic integration of large variant libraries for deep mutational scanning in mammalian cells

abstract: There is a need of introducing large variant libraries into mammalian cells to facilitate deep mutational scanning (DMS). In this study, we develop an approach to integrate in-vitro ligation dsDNA variant library into cultured mammalian cell genome via the piggyBac transposon system. T4 DNA ligase was employed to produce dsDNA ligation products containing variant library sequence flanked by piggyBac inverted terminal repeat sequences (ITRs) at both ends. The ligation products were then cotransfected with a piggyBac transposase expression vector into mammalian cells for library integration. The clone formation assay results showed that about 29,000 resistance clones were generated from a single well of 6-well-plate transfected HEK293T cells. As a proof of principle of using the strategy for DMS, a GFP chromophore random variant library was integrated into the HEK293T cell genome. By FACS selection, we enriched GFP-positive cells. The next-generation sequencing (NGS) results showed that the GFP-chromophore amino acid sequences were restored after selection.


title: RT-qPCR as a screening platform for mutational and small molecule impacts on structural stability of RNA tertiary structures

abstract: The exponential increase in the discovery and characterization of RNA tertiary structures has highlighted their active role in a variety of human disease, yet often their interactome and specific function remain unknown. Small molecules offer opportunities to both decode these cellular roles and develop therapeutics, yet there are few examples of small molecules that target biologically relevant RNA tertiary structures. While RNA triple helices are a particular attractive target, discovery of triple helix modulators has been hindered by the lack of correlation between small molecule affinity and effect on structural modulation, thereby limiting the utility of affinity-based screening as a primary filtering method. To address this challenge, we developed a high-throughput RT-qPCR screening platform that reports on the effect of mutations and additives, such as small molecules, on the structuredness of triple helices. Using the 3-end of the oncogenic non-coding RNA MALAT1 as an example, we demonstrated the applicability of both a two-step and a one-pot method to assess the impact of mutations and small molecules on the stability of the triple helix. Employment of a functional high-throughput assay as a primary screen will significantly expedite the discovery of probes that modulate RNA triple helices structural landscape and, consequently, help gain insight into the roles of these pervasive structures.


title: In vivo inactivation of RAD51-mediated homologous recombination leads to premature aging, but not to tumorigenesis

abstract: Genetic instability is a hallmark of both cancer and aging. Homologous recombination (HR) is a prominent DNA repair pathway maintaining genomic integrity. Mutations in many HR genes lead to cancer predisposition. Paradoxically, the consequences of mutations in the pivotal HR player, RAD51, on cancer development remain puzzling. Moreover, in contrast with other HR genes, RAD51 mouse models are not available to experimentally address the role of RAD51 on aging and carcinogenesis, in vivo. Here, we engineered a mouse model with an inducible dominant negative form of RAD51 (SMRad51) that suppresses RAD51-mediated HR without stimulating alternative non-conservative repair pathways. We found that, in vivo expression of SMRad51 did not trigger tumorigenesis, but instead induced premature aging. We propose that these in vivo phenotypes result from the exhaustion of proliferating progenitors submitted to chronic endogenous replication stress resulting from RAD51-mediated HR suppression. Our data underline the importance of the RAD51 activity for progenitors homeostasis, preventing aging, and more generally for the balance between cancer and aging.


title: Blocking Kir6.2 channels with SpTx1 potentiates glucose-stimulated insulin secretion from murine pancreatic β cells and lowers blood glucose in diabetic mice

abstract: ATP-sensitive K+ (KATP) channels in pancreatic {beta} cells comprise pore-forming subunits (Kir6.2) and modulatory sulfonylurea receptor subunits (SUR1). The ATP sensitivity of these channels enables them to couple metabolic state to insulin secretion in {beta} cells. Antidiabetic sulfonylureas such as glibenclamide target SUR1 and indirectly suppress Kir6.2 activity. Glibenclamide acts as both primary and secondary secretagogues to trigger insulin secretion and potentiate glucose-stimulated insulin secretion, respectively. We tested whether blocking Kir6.2 itself causes the same effects as glibenclamide, and found that the Kir6.2 pore-blocker SpTx1 acts as a strong secondary, but not a primary, secretagogue. SpTx1 triggered a transient rise of plasma insulin and lowered the elevated blood glucose of diabetic mice over-expressing Kir6.2 but did not affect those of non-diabetic mice. This proof-of-concept study suggests that blocking Kir6.2 may serve as an effective treatment for diabetes and other diseases stemming from Kir6.2 hyperactivity that cannot be suppressed with sulfonylureas.

category: physiology


title: Comparative Phenotyping of Two Commonly Used Chlamydomonas reinhardtii Background Strains: CC-1690 (21gr) and CC-5325 (the CLiP Mutant Library Background)

abstract: The unicellular green alga Chlamydomonas reinhardtii is an excellent model organism to investigate many essential cellular processes in photosynthetic eukaryotes. Two commonly used background strains of Chlamydomonas are CC-1690 and CC-5325. CC-1690, also called 21gr, has been used for the Chlamydomonas genome project and several transcriptome analyses. CC-5325 is the background strain for the Chlamydomonas Library Project (CLiP). Photosynthetic performance in CC-5325 has not been evaluated in comparison with CC-1690. Additionally, CC-5325 is often considered to be cell-wall deficient, although detailed analysis is missing. The circadian rhythms in CC-5325 are also unclear. To fill these knowledge gaps and facilitate the use of the CLiP mutant library for various screens, we performed phenotypic comparisons between CC-1690 and CC-5325. Our results showed that CC-5325 grew faster heterotrophically in dark and equally well in mixotrophic liquid medium as compared to CC-1690. CC-5325 had lower photosynthetic efficiency and was more sensitive to heat than CC-1690. Furthermore, CC-5325 had an intact cell wall with comparable integrity to that in CC-1690, though appears to have reduced thickness. Finally, CC-5325 could perform phototaxis, but could not maintain a sustained circadian rhythm of phototaxis as CC160 did. Our results will be useful for researchers in the Chlamydomonas community to choose suitable background strains for mutant analysis and employ the CLiP mutant library for genome-wide mutant screens under appropriate conditions, especially in the areas of photosynthesis, thermotolerance, cell wall, and circadian rhythms.

category: plant biology


title: Stomatal anatomy, leaf structure and nutrients of tropical rainforest tree species respond to altitude in a coordinated manner in accordance with the leaf economics spectrum

abstract: Understanding taxon level responses of key plant traits to environmental variation across tropical rainforests (TRFs) is important to determine their response to climate change. We used an altitudinal gradient (from 150 to 2100 m above sea level, asl) across TRFs in Sri Lanka to answer the following questions: (a) Does the response to altitude by stomatal traits differ among plant taxa in TRFs?; (b) Are the altitudinal responses of key leaf structural traits (e.g. leaf mass per area, LMA) and major leaf nutrient (nitrogen, N, and phosphorus, P) concentrations linked to the corresponding variation in stomatal traits in a coordinated response across taxa?; (c) How strong is the influence of climatic variation on responses of leaf traits to altitude?. Leaf samples were collected in permanent sampling plots within rainforest reserves at Kanneliya (150 m asl), Sinharaja-Enasalwatta (1050 m), Hakgala (1800 m) and Piduruthalagala (2100 m) from 19 species in three plant genera (Calophyllum, Semecarpus and Syzygium). Stomatal density, guard cell length and epidermal density showed variation among taxa, but did not respond to altitude. Potential conductance index (PCI), a proxy for photosynthetic capacity, decreased with increasing altitude, in a common response across taxa. We found evidence that altitudinal responses of LMA, leaf N and P were linked to stomatal responses in a coordinated manner, where key features were the negative correlations between PCI and LMA and between proxy photosynthetic N- and P-use efficiencies ( PNUE and  PPUE) and LMA. We found strong responses to climatic variation across taxa and altitudes, where PCI,  PNUE and  PPUE increased and LMA decreased with increasing temperature, precipitation and solar irradiance. We conclude that stomatal traits of tree species in TRFs form part of a coordinated leaf trait response to environmental change which is in accordance with the leaf economics spectrum.


title: Activation of the dorsal septum increases alcohol consumption in male C57BL/6J mice.

abstract: Binge drinking is a common pattern of excessive alcohol consumption associated with Alcohol Use Disorder (AUD) and unraveling the neurocircuitry that promotes this type of drinking is critical to the development of novel therapeutic interventions. The septal region was once a focal point of alcohol research yet has seen limited study over the last decade in relation to binge drinking. Numerous studies point to involvement of the dorsal septum (dSep) in excessive drinking and withdrawal, but few studies have manipulated this region in the context of binge drinking behavior. The present experiments were primarily designed to determine the effect of chemogenetic manipulation of the dSep on binge-like alcohol drinking in male and female C57BL/6J mice. Mice received bilateral infusion of AAVs harboring hM4Di, hM3Dq, or mCherry into the dSep and subjects were challenged with systemic administration of clozapine-N-oxide (CNO; 3 mg/kg) and vehicle (saline; 0.9%) in the context of binge-like alcohol consumption, locomotor activity, and sucrose drinking. CNO-mediated activation (hM3Dq) of the dSep resulted in a significant increase in binge-like alcohol consumption, locomotor activity, and sucrose intake in male mice. DSep activation promoted sucrose drinking in female mice, but alcohol intake and locomotor activity were unaffected. Conversely, silencing (hM4Di) of the dSep modestly decreased locomotor activity in males and did not influence alcohol or sucrose intake in either sex. Lastly, CNO was without effect in mCherry-expressing control groups. These data support a role for the dSep in promoting excessive, binge-like drinking behavior in a sex-dependent fashion and suggests a broad role for the region in the modulation of locomotor activity as well as general appetitive behavior such as sucrose drinking.


title: Human tRNA methyltransferase ALKBH8 targets wider repertoire of tRNAs and non-coding RNAs

abstract: Transfer RNAs acquire a large plethora of chemical modifications. Among those, modifications of the anticodon loop play important roles in translational fidelity and tRNA stability. Four human wobble U containing tRNAs obtain 5-methoxycarbonylmethyluridine (mcm5U34) or 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U34), which play a role in decoding. This mark involves a cascade of enzymatic activities. The last step is mediated by Alkylation Repair Homolog 8 (ALKBH8). In this study, we performed a transcriptome-wide analysis of the repertoire of ALKBH8 RNA targets. Using a combination of HITS-CLIP-seq and RIP-seq analyses, we uncover ALKBH8-bound RNAs. It targets an additional wobble U-containing tRNA tRNALys(UUU). More interestingly, the spectrum of bound RNAs includes other types of non-coding RNAs, such as C/D box snoRNAs, 7SK RNA or some miRNAs, respectively.


title: The oncogene FOXQ1 is a selective activator of Wnt/β-catenin signalling

abstract: The forkhead box transcription factor FOXQ1 is aberrantly induced in various cancers, and contributes to tumour growth and metastasis. It has been suggested that FOXQ1 exacerbates cancer by activating the oncogenic Wnt/{beta}-catenin signalling pathway. However, the mode of action of FOXQ1 in the Wnt pathway remains to be resolved. Here, we report that FOXQ1 is a bimodal transcriptional activator of Wnt target gene expression in normal and cancer cells. Using co-immunoprecipitation, proximity proteomics, and reporter assays, we show that FOXQ1 engages the Wnt transcriptional complex to promote gene expression via TCF/LEF transcription factors. In parallel, FOXQ1 differentially regulates the expression of Wnt target genes independently of {beta}-catenin and TCF/LEFs, which is facilitated by spatially separated activator and repressor domains. Our results suggest that FOXQ1 is a novel component of the Wnt transcriptional complex that reinforces and specifies Wnt signalling in a context-dependent manner.


title: Exocyst stimulates each step of exocytic SNARE complex assembly and vesicle fusion

abstract: The exocyst is a large multisubunit tethering complex essential for targeting and fusion of secretory vesicles in eukaryotic cells. Although the assembled exocyst complex is proposed to tether vesicles to the plasma membrane and activate the SNARE proteins for membrane fusion, only little is known about the key biochemical steps that exocyst stimulates in the course of SNARE complex assembly, a critical question defining the essential molecular role of the exocyst complex. Here, we use a combination of single molecule and bulk fluorescence assays with purified octameric yeast exocyst complexes to examine the role of exocyst in a reconstituted SNARE assembly and fusion system. We show that the exocyst complex simulates multiple steps spanning from SNARE protein activation to ternary complex assembly, rather than affecting only a specific subset of steps. We also observed that the exocyst has important downstream roles in driving membrane fusion, up to full content mixing of vesicle lumens. Our results suggest that the exocyst complex provides extensive chaperoning functions for the entire process of SNARE complex assembly, presumably using its multi-faceted structure provided by the eight subunits.


title: Divergent age-related methylation patterns in long and short-lived mammals

abstract: Age-related changes to cytosine methylation have been extensively characterized across the mammalian family. Some cytosines that are conserved across mammals exhibit age-related methylation changes that are so consistent that they were used to successfully develop cross-species age predictors. In a similar vein, methylation levels of some conserved cytosines correlate highly with species lifespan, leading to the development of highly accurate lifespan predictors. Surprisingly, little to no commonality is found between these two sets of cytosines even though the relationship between aging and lifespan is by most measures linked. We ventured to address this conundrum by first identifying age-related cytosines whose methylation levels change in opposite directions between short and long-lived species. We hypothesized that age-related CpGs that are also associated with species lifespan would tap into biological processes that simultaneously impact aging and lifespan. To this end, we analyzed age-related cytosine methylation patterns in 82 mammalian species. For each CpG, we correlated the intra-species age correlation with maximum lifespan across mammalian species. We refer to this correlation of correlations as Lifespan Uber Correlation (LUC). This approach is unique in incorporating age and species lifespan in a single analysis. We identified 629 CpGs with opposing methylation aging patterns in long and short-lived species. Many of these are found to be near BCL11B, NPTN, and HOXC4 loci. Methylation and transcription analyses of Bcl11b knockout mice indicate that this gene partially regulates the methylation state of LUC CpGs. We developed DNAm age estimators (epigenetic clocks) based on LUC CpGs. These LUC clocks exhibited the expected behavior in benchmark aging interventions such as caloric restriction, growth hormone receptor knockout and high-fat diet. Furthermore, we found that Bcl11b heterozygous knockout mice exhibited an increased epigenetic age in the striatum. Overall, we present a bioinformatics approach that identified CpGs and their associated genes implicated both in aging and lifespan. These cytosines lend themselves to developing epigenetic clocks that are sensitive to perturbations that impact both age and lifespan.


title: A novel mechanism for left-right asymmetry establishment involving tissue remodeling and MyoID

abstract:
How novel left-right asymmetries evolve in animals is unknown. Drosophila pachea males display a unique left-right asymmetry in the genital organs and mate in a right-sided position onto the female. In flies, male genitalia undergo a clockwise 360{degrees} rotation during development. To test if this tissue remodeling process is linked to asymmetry development in the male genitalia of Drosophila pachea, we developed CRIPSR and transgenesis in this singular species. As in D. melanogaster, mutants of the MyoID gene homolog undergo reverse rotation of the developing genitalia. In addition, they have mirror-image asymmetric genitalia. Although their genital asymmetry is reversed mutants still adopt a right-sided copulation posture. Our results reveal a novel mechanism for establishing left-right asymmetry in animals, involving complex tissue remodeling and MyoID.

One Sentence SummaryD. pachea genital left-right size asymmetry is established through MyoID-mediated tissue remodelling and does not determine the direction of mating posture.


title: Population-specific association of Clock gene polymorphism with annual cycle timing in stonechats

abstract: Timing is essential for survival and reproduction of organisms across the tree of life. The core circadian clock gene Clk has been implicated in annual timing and shows highly conserved sequence homology across vertebrates except for one variable region of poly Glutamine repeats. Clk genotype varies in some species with latitude, seasonal timing and migration. However, findings are inconsistent, difficult to disentangle from environmental responses, and biased towards high latitudes. Here we combine field data with a common-garden set up to study associations of Clk with latitude, migration and annual-cycle timing within the stonechat species complex with trans-equatorial distribution range. Including 950 individuals from nine populations with diverse migratory strategies. Gene diversity was lowest in resident African and Canary Island populations and increased with latitude, independently of migration distance. Repeat length and annual-cycle timing was linked in a population-specific way. Specifically, equatorial African stonechats showed delayed timing with longer repeat length for all annual-cycle stages. Our data suggest that at low latitudes with constant photoperiod, Clk genotype might orchestrate a range of consistent, individual chronotypes. In contrast, the influence of Clk on annual-cycle timing at higher latitudes might be mediated by its interactions with genes involved in (circadian) photoperiodic pathways.


title: Development of efficient RNAi methods in the corn leafhopper Dalbulus maidis, a promising application for pest control

abstract:
BACKGROUNDThe corn leafhopper Dalbulus maidis is the main vector of three important stunting pathogens that affect maize production. The most common control strategy against this species is the use of insecticides that provide minimal, short-term protection. In this context, genomic-based technologies such as RNA interference (RNAi) could be a suitable approach to control this pest in a highly specific manner, avoiding the adverse effects associated with insecticide misuse. Therefore, the objective of the present work was to assess the application of RNAi on D. maidis through different dsRNA delivery methods and known the function of target gene, Bicaudal C (BicC).

RESULTSWe have identified and characterized the core components of the RNAi machinery in silico and established two methods of exogenous double-stranded RNAs (dsRNA) delivery to D. maidis. BicC -an important regulator of insect oogenesis-dsRNA was successfully delivered via injection or ingestion to adult females, causing significant reductions in the transcript levels and ovipositions and observable phenotypes in the ovaries when compared to control females. The small doses of dsRNABicC administered were enough to trigger a strong RNAi response, demonstrating that D. maidis is highly sensitive to RNAi.

CONCLUSIONThis is, to our knowledge, the first report describing RNAi application in D. maidis, a tool that can be used to advance towards a novel, insecticide-free control strategy against this pest.


title: A chromosome-scale genome assembly of the false clownfish, Amphiprion ocellaris

abstract:
BackgroundThe false clownfish Amphiprion ocellaris is a popular fish species and an emerging model organism for studying the ecology, evolution, adaptation, and developmental biology of reef fishes. Despite this, high-quality genomic resources for this species are scarce, hindering advanced genomic analyses. Leveraging the power of PacBio long-read sequencing and Hi-C chromosome conformation capture techniques, we constructed a high-quality chromosome-scale genome assembly for the clownfish A. ocellaris.

ResultsThe initial genome assembly comprised of 1,551 contigs of 861.42 Mb, with an N50 of 863.85 kb. Hi-C scaffolding of the genome resulted in 24 chromosomes containing 856.61 Mb. The genome was annotated with 26,797 protein-coding genes and had 96.62 % completeness of conserved actinopterygian genes, making this genome the most complete and high quality among published anemonefish genomes. Transcriptomic analysis identified tissue-specific gene expression patterns, with the brain and optic lobe having the largest number of expressed genes. Further, comparative genomic analysis revealed 91 genome elements conserved only in A. ocellaris and its sister species Amphiprion percula, and not in other anemonefish species. These elements are close to genes that are involved in various nervous system functions and exhibited distinct expression patterns in brain tissue, potentially highlighting the genetic toolkits involved in lineage-specific divergence and behaviors of the clownfish branch.

ConclusionsOverall, our study provides the highest quality A. ocellaris genome assembly and annotation to date, whilst also providing a valuable resource for understanding the ecology and evolution of reef fishes.

category: genomics


title: ISSAAC-seq enables sensitive and flexible multimodal profiling of chromatin accessibility and gene expression in single cells

abstract: Joint profiling of chromatin accessibility and gene expression from the same single cell/nucleus provides critical information about cell types in a tissue and cell states during a dynamic process. These emerging multi-omics techniques help the investigation of cell-type resolved gene regulatory mechanisms1-7. However, many methods are currently limited by low sensitivity, low throughput or complex workflow. Here, we developed in situ SHERRY after ATAC-seq (ISSAAC-seq), a highly sensitive and flexible single cell multi-omics method to interrogate chromatin accessibility and gene expression from the same single nucleus. We demonstrated that ISSAAC-seq is sensitive and provides high quality data with orders of magnitude more features than existing methods. Using the joint profiles from over 10,000 nuclei from the mouse cerebral cortex, we uncovered major and rare cell types and cell-type specific regulatory elements and identified heterogeneity at the chromatin level within established cell types defined by gene expression. Finally, we revealed distinct dynamics and relationships of gene expression and chromatin accessibility during an oligodendrocyte maturation trajectory.


title: User-guided local and global copy-number segmentation for tumor sequencing data

abstract:
MotivationCopy-number aberrations (CNA) are genetic alterations that amplify or delete the number of copies of large genomic segments. Although they are ubiquitous in cancer and subsequently a critical area of current cancer research, CNA identification from DNA sequencing data is challenging because it requires partitioning of the genome into complex segments that may not be contiguous. Existing segmentation algorithms address these challenges either by leveraging the local information among neighboring genomic regions, or by globally grouping genomic regions that are affected by similar CNAs across the entire genome. However, both approaches have limitations: overclustering in the case of local segmentation, or the omission of clusters corresponding to focal CNAs in the case of global segmentation. Importantly, inaccurate segmentation will lead to inaccurate identification of important CNAs.

ResultsWe introduce CNAViz, a web-based tool that enables the user to simultaneously perform local and global segmentation, thus overcoming the limitations of each approach. Using simulated data, we demonstrate that by several metrics, CNAViz yields more accurate segmentations relative to existing local and global segmentation methods. Moreover, we analyze six bulk DNA sequencing samples from three breast cancer patients. By validating with parallel single-cell DNA sequencing data from the same samples, we show that CNAVizs more accurate segmentation improves accuracy in downstream copy-number calling.

Availability and implementationhttps://github.com/elkebir-group/cnaviz

Contacts.zaccaria@ucl.ac.uk, melkebir@illinois.edu


title: Genome sequence data reveal at least two distinct incursions of the tropical race 4 (TR4) variant of Fusarium wilt into South America

abstract: The global banana industry is threatened by one of the most devastating diseases: Fusarium wilt (FWB). FWB is caused by the soil-borne fungus Fusarium oxysporum f. sp. cubense (Foc), which almost annihilated the banana production in the late 1950s. A new strain of Foc, known as tropical race 4 (TR4), attacks a wide range of banana varieties including Cavendish clones which are the source of 99% of banana exports. In 2019, Foc TR4 was reported in Colombia, and more recently (2021) in Peru. In this study, we sequenced three fungal isolates identified as Foc TR4 from La Guajira (Colombia) and compared them against 19 whole-genome sequences of Foc TR4 publicly available, including four genome sequences recently released from Peru. To understand the genetic relatedness of the Colombian Foc TR4 isolates and those from Peru, we conducted a phylogenetic analysis based on a genome-wide set of single nucleotide polymorphisms (SNPs). Additionally, we compared the genomes of the 22 available Foc TR4 isolates looking for the presence-absence of gene polymorphisms and genomic regions. Our results reveal that (i) the Colombian and Peruvian isolates are genetically distant, which could be better explained by independent incursions of the pathogen to the continent, and (ii) there is a high correspondence between the genetic relatedness and geographic origin of Foc TR4. The profile of present/absent genes and the distribution of missing genomic regions showed a high correspondence to the clades recovered in the phylogenetic analysis, supporting the results obtained by SNP-based phylogeny.


title: The coupling between healthspan and lifespan in Caenorhabditis depends on complex interactions between compound intervention and genetic background

abstract: Aging is characterized by declining health that results in decreased neuromuscular function and cellular resilience. The relationship between lifespan and health, and the influence of genetic background on that relationship, has important implications in the development of anti-aging interventions. Here we combined survival under thermal and oxidative stress with swimming performance, to evaluate health effects across a nematode genetic diversity panel for three compounds previously studied in the Caenorhabditis Intervention Testing Program - NP1, propyl gallate, and resveratrol. We show that oxidative stress resistance and thermotolerance vary with compound intervention, genetic background, and age. The effects of tested compounds on swimming locomotion, in contrast, are largely species-specific. Additionally, thermotolerance, but not oxidative stress or swimming ability, correlates with lifespan. Our results demonstrate the importance of assessing health and lifespan across genetic backgrounds in the effort to identify reproducible aging interventions.


title: Tuftsin: a natural molecule against SARS-CoV-2 infection

abstract: Coronavirus disease 2019 (COVID-19) continuously proceeds despite the application of a variety of vaccines. It is still urgent to find effective ways to treat COVID-19. Recent studies indicate that NRP1, an important receptor of the natural peptide tuftsin, facilitates SARS-CoV-2 infection. Importantly, tuftsin is a natural human molecule released from IgG. Here, we found 91 overlapping genes between tuftsin targets and COVID-19-associated genes. Bioinformatics analyses indicated that tuftsin could also target ACE2 and exert some immune-related functions to treat COVID-19. Using surface plasmon resonance (SPR) analysis, we confirmed that tuftsin can bind ACE2 and NRP1 directly. Moreover, tuftsin effectively impairs the binding of SARS-CoV-2 S1 to ACE2. Thus, tuftsin is an attractive drug against COVID-19. And tuftsin as natural immunostimulating peptide in human, we speculate that tuftsin may has crucial roles in asymptomatic carriers or mild cases of COVID-19.


title: A deep learning pipeline for segmentation of Proteus mirabilis colony patterns

abstract: The motility mechanisms of microorganisms are critical virulence factors, enabling their spread and survival during infection. Motility is frequently characterized by qualitative analysis of macroscopic colonies, yet the standard quantification method has mainly been limited to manual measurement. Recent studies have applied deep learning for classification and segmentation of specific microbial species in microscopic images, but less work has focused on macroscopic colony analysis. Here, we advance computational tools for analyzing colonies of Proteus mirabilis, a bacterium that produces a macroscopic bullseye-like pattern via periodic swarming, a process implicated in its virulence. We present a dual-task pipeline for segmenting (1) the macroscopic colony including faint outer swarm rings, and (2) internal ring boundaries, unique features of oscillatory swarming. Our convolutional neural network for patch-based colony segmentation and U-Net with a VGG-11 encoder for ring boundary segmentation achieved test Dice scores of 93.28% and 83.24%, respectively. The predicted masks at times improved on the ground truths from our automated annotation algorithms. We demonstrate how application of our pipeline to a typical swarming assay enables ease of colony analysis and precise measurements of more complex pattern features than those which have been historically quantified.


title: Basis of narrow-spectrum activity of fidaxomicin on gut pathogen Clostridioides difficile

abstract: Fidaxomicin (Fdx) is widely used to treat Clostridioides difficile (Cdiff) infections (CDIs), but the molecular basis of its narrow-spectrum activity in the human gut microbiome remains enigmatic. CDIs are a leading cause of nosocomial deaths. Fdx, which inhibits RNA polymerase (RNAP), targets Cdiff with minimal effects on gut commensals, reducing CDI recurrence. Here, we present the cryo-electron microscopy structure of Cdiff RNAP in complex with Fdx, allowing us to identify a crucial Fdx-binding determinant of Cdiff RNAP that is absent in most gut microbiota like Proteobacteria and Bacteroidetes. By combining structural, biochemical, and bioinformatic analyses, we establish that a single RNAP residue is a sensitizing element for Fdx narrow-spectrum activity. Our results provide a blueprint for targeted drug design against an important human pathogen.