title: The cross-domain functional organisation of posterior lateral temporal cortex: Insights from ALE meta-analyses of seven cognitive domains spanning 9515 participants

abstract: The posterior lateral temporal cortex is implicated in many verbal, nonverbal and social cognitive domains and processes. Yet without directly comparing these disparate domains, the regions organisation remains unclear; do distinct processes engage discrete subregions, or could different domains engage shared neural correlates and processes? Here, using activation likelihood estimation meta-analyses, the bilateral posterior lateral temporal cortex subregions engaged in seven domains were directly compared. These domains comprised semantics, semantic control, phonology, biological motion, face processing, theory of mind, and representation of tools. Whilst phonology and biological motion were predominantly associated with distinct regions, other domains implicated overlapping areas, perhaps due to shared underlying processes. Theory of mind recruited regions implicated in semantic representation in the left hemisphere, tools engaged semantic control areas, and faces engaged subregions for biological motion and theory of mind. This cross-domain approach provides insight into how posterior lateral temporal cortex is organised and why.


title: Sensory system-specific associations between brain structure and balance

abstract:
Nearly 75% of older adults in the United States report balance problems. Balance difficulties are more pronounced during sensory feedback perturbation (e.g., standing with the eyes closed or on foam). Although it is known that aging results in widespread brain atrophy, less is known about how brain structure relates to balance performance under varied sensory conditions in older age. We measured postural sway of 36 young (18-34 years) and 22 older (66-84 years) adults during four conditions: eyes open, eyes closed, eyes open on foam, and eyes closed on foam. We calculated three summary measures indicating visual, proprioceptive, and vestibular contributions to balance. We also collected T 1-weighted and diffusion-weighted anatomical MRI scans. We aimed to: 1) test for age group differences in brain structure-balance relationships across a range of structural brain measures (i.e., volumetric, surface, and white matter microstructure); and 2) assess how brain structure measures relate to balance, regardless of age. Across both age groups, thinner cortex in multisensory integration regions was associated with greater reliance on visual inputs for balance. Greater gyrification within sensorimotor and parietal cortices was associated with greater reliance on proprioceptive inputs for balance. Poorer vestibular function was correlated with thinner vestibular cortex, greater gyrification within sensorimotor, parietal, and frontal cortices, and lower free water-corrected axial diffusivity in the superior-posterior corona radiata and across the corpus callosum. These results contribute to our scientific understanding of how individual differences in brain structure relate to balance. This has implications for developing brain stimulation interventions to improve balance.

Significance StatementOlder age is associated with greater postural sway, particularly when sensory information is perturbed (e.g., by closing ones eyes). Our work contributes to the field by identifying how individual differences in regional brain structure relate to balance under varying sensory conditions in young and older adults. Across both age groups, lower cortical thickness in sensory integration and vestibular regions, greater gyrification within sensorimotor, parietal, and temporal regions, and lower free water-corrected axial diffusivity in the corpus callosum and corona radiata were related to individual differences in balance scores. We identified brain structures that are associated with specific sensory balance scores; therefore, these results have implications for which brain regions to target in future interventions for different populations.

category: neuroscience


title: pyNeurode: a real-time neural signal processing framework

abstract: Accurate decoding of neural signals often requires assigning extracellular waveforms acquired on the same electrode to their originating neurons, a process known as spike sorting. While many offline sorters are available, accurate online sorting of spikes with many channels is still a challenging problem. Existing online sorters either use simple algorithms with low accuracy, can only process a handful of channels, or depend on a complex runtime environment that is difficult to set up. We have developed a state-of-the-art online spike sorting platform in Python that enables large-scale, fully automatic real-time spike sorting and decoding on hundreds of channels. Our system is cross-platform and works seamlessly with the Open Ephys suite of open-source hardware and software widely used in many neuroscience laboratories worldwide. It also comes with a user-friendly graphical user interface to monitor the cluster quality, spike waveforms and neuronal firing rate. Our platform has comparable accuracy to offline sorters and can achieve an end-to-end sorting latency of around 160 ms for 128-channel signals. It will be useful for research in fundamental neuroscience, closed-loop feedback neuromodulation and brain-computer interfaces.


title: Exploiting Color Space Geometry for Visual Stimulus Design across Animals

abstract: Color vision represents a vital aspect of perception that ultimately enables a wide variety of species to thrive in the natural world. However, unified methods for constructing chromatic visual stimuli in a laboratory setting are lacking. Here, we present stimulus design methods and an accompanying programming package to efficiently probe the color space of any species in which the photoreceptor spectral sensitivities are known. Our hardware-agnostic approach incorporates photoreceptor models within the framework of the principle of univariance. This enables experimenters to identify the most effective way to combine multiple light sources to create desired distributions of light, and thus easily construct relevant stimuli for mapping the color space of an organism. We include methodology to handle uncertainty of photoreceptor spectral sensitivity as well as to optimally reconstruct hyperspectral images given recent hardware advances. Our methods support broad applications in color vision science and provide a framework for uniform stimulus designs across experimental systems.


title: A bivalent EBV vaccine induces neutralizing antibodies that block B and epithelial cell infection and confer immunity in humanized mice

abstract:
Epstein Barr virus (EBV) is the major cause of infectious mononucleosis and is associated with several human cancers. Despite its prevalence and major impact on human health, there are currently no specific vaccines or treatments. Four viral glycoproteins, gp 350 and gH/gL/gp42 mediate entry into the major sites of viral replication, B cells and epithelial cells. Here, we designed a nanoparticle vaccine displaying these proteins and show that it elicits potent neutralizing antibodies that protect against infection in vivo. Based on structural analyses, we designed single chain gH/gL and gH/gL/gp42 proteins that were each fused to bacterial ferritin to form a self-assembling nanoparticles. X-ray crystallographic analysis revealed that single chain gH/gL and gH/gL/gp42 adopted a similar conformation to the wild type proteins, and the protein spikes were observed by electron microscopy. Single chain gH/gL or gH/gL/gp42 nanoparticle vaccines were constructed to ensure product homogeneity needed for clinical development. These vaccines elicited neutralizing antibodies in mice, ferrets, and non-human primates that inhibited EBV entry into both B cells and epithelial cells. When mixed with a previously reported gp350 nanoparticle vaccine, gp350D123, no immune competition was observed. To confirm its efficacy in vivo, humanized mice were challenged with EBV after passive transfer of IgG from mice vaccinated with control, gH/gL/gp42+gp350D123 or gH/gL+gp350D123 nanoparticles. While all control animals (6/6) were infected, only one mouse in each vaccine group that received immune IgG had transient low level viremia (1/6). Furthermore, no EBV lymphomas were detected in immune animals in contrast to non-immune controls. This bivalent EBV nanoparticle vaccine represents a promising candidate to prevent EBV infection and EBV-related malignancies in humans.

One sentence summaryA bivalent gp350 and gH/gL/gp42 nanoparticle vaccine elicits neutralizing antibodies that protect against EBV infection and EBV lymphoma in vivo.

category: immunology


title: CDKN1A is a target for phagocytosis-mediated cellular immunotherapy in acute leukemia

abstract: Most tumor-associated macrophages (TAMs), which are abundant in the tumor microenvironment, demonstrate an immunosuppressive phenotype and contribute to tumor progression, treatment resistance and poor clinical outcomes1,2. Due to their functional plasticity, these cells could be reprogrammed to acquire a pro-inflammatory phenotype and promote tumor clearance1. Several therapeutic approaches targeting TAMs to alleviate their immunosuppressive properties or to harness their tumoricidal capacities have been developed3,4. Inhibition of interactions between phagocytic inhibitor receptors on macrophages and "dont eat me signals" on cancer cells, which promotes cancer cell engulfment, showed therapeutic benefits for several tumor types4-6. Investigating mechanisms involved in macrophage-mediated phagocytosis of tumor cells, we demonstrate here a key role for the cyclin-dependent kinase inhibitor CDKN1A (p21). Through transcriptional repression of SIRP (Signal-Regularity Protein ), which encodes a phagocytic inhibitor, CDKN1A promotes the ability of monocyte-derived macrophages (MDMs) to engulf leukemic cells. In turn, these MDMs acquire a pro-inflammatory phenotype that extends to surrounding MDMs in an Interferon {gamma} (IFN{gamma})-dependent manner. Human monocytes genetically engineered to overexpress p21 (p21TD-Mo) differentiate into anti-inflammatory MDMs that are primed for leukemic cell phagocytosis when transferred into mice xenografted with patient-derived T-cell acute lymphoblastic leukemia (T-ALL) cells. After leukemic cell engulfment, engineered macrophages undergo a pro-inflammatory activation, reducing leukemic burden and substantially prolonging survival of mice. These results reveal p21 as a trigger of phagocytosis-guided pro-inflammatory reprogramming of TAMs and demonstrate the potential for p21TD-Mo-based cell therapy in cancer immunotherapy.


title: A local translation program regulates centriole amplification in the airway epithelium

abstract:
Biogenesis of organelles requires targeting of subset of proteins to specific subcellular domains by signal peptides or mechanisms controlling mRNA localization and local translation. How local distribution and translation of specific mRNAs for organelle biogenesis is achieved remains elusive and likely to be dependent on the cellular context. Here we identify Trinucleotide repeat containing-6a (Tnrc6a), a component of the miRNA pathway, distinctively localized to apical granules of differentiating airway multiciliated cells (MCCs) adjacent to centrioles. In spite of being enriched in TNRC6A and the miRNA-binding protein AGO2, they lack enzymes for mRNA degradation. Instead, we found these apical granules enriched in components of the mRNA translation machinery and newly synthesized proteins suggesting that they are specific hubs for target mRNA localization and local translation in MCCs. Consistent with this, Tnrc6a loss of function prevented formation of these granules and led to a broad reduction, rather than stabilization of miRNA targets. These included downregulation of key genes involved in ciliogenesis and was associated with defective cilia formation both in vivo and in primary airway epithelial cultures. Similar analysis of Tnr6a disruption in yolk sac showed stabilization of miRNA targets, highlighting the potential diversity of these mechanisms across organs.

HighlightsO_LITnrc6a is selectively and abundantly present in multiciliated cells (MCC) of the lung.
C_LIO_LILoss of Tnrc6a in the lung leads to reduction, not stabilization of miRNA targets.
C_LIO_LITRNC6A concentrates in apically-localized granules adjacent to centrioles in MCCs.
C_LIO_LIApically-localized granules contain AGO2, but lack mRNA degradation enzymes.
C_LIO_LINewly-synthesized proteins and translation machinery components are concentrated or associated with apically localized granules.
C_LIO_LITnrc6a is required for efficient protein production of miRNA targets and proper centriole amplification during multicilia formation.
C_LI

category: cell biology


title: Biochemistry and nanomechanical properties of human colon cells upon simvastatin, lovastatin and mevastatin supplementations - Raman imaging and AFM studies

abstract: One of the most important areas of medical science is oncology, which is responsible for both the diagnostics and treatment of cancer diseases. Over the years, there has been an intensive development of cancer diagnostics and treatment. This paper shows the comparison of normal (CCD-18Co) and cancerous (CaCo-2) cell lines of the human gastrointestinal tract on the basis of nanomechanical and biochemical properties in order to obtain information on the cancer biomarkers useful in oncological diagnostics. The research techniques used were: Atomic Force Microscopy and Raman spectroscopy and imaging. In addition, the studies included also the effect of the statins compounds: mevastatin, lovastatin and simvastatin and their influence on nanomechanical and biochemical changes of properties of cells tracking using AFM and Raman imaging techniques. The cytotoxicity of mevastatin and lovastatin was determined by using XTT tests.


title: Mitigation of the age-dependent decline in mitochondrial genome integrity

abstract:
Unknown processes promote accumulation of mitochondrial DNA mutations during aging. Accumulation of defective mitochondrial genomes is thought to promote progression of heteroplasmic mitochondrial diseases and degenerative changes with natural aging. We used a heteroplasmic Drosophila model to test 1) whether purifying selection acts to limit the abundance of deleterious mutations during development and aging, 2) whether quality control pathways contribute to purifying selection, 3) whether activation of quality control can mitigate accumulation of deleterious mutations, and 4) whether improved quality control improves healthspan. We show that purifying selection operates during development and growth, but is ineffective during aging. Genetic manipulations suggest that a quality control process known to enforce purifying selection during oogenesis also suppresses accumulation of a deleterious mutation during growth and development. Flies with nuclear genotypes that enhanced purifying selection sustained higher genome quality, retained more vigorous climbing activity and lost fewer dopaminergic neurons. Pharmacological enhancement of quality control produced similar benefits. Importantly, similar pharmacological treatment of aged mice reversed age-associated accumulation of a deleterious mtDNA mutation. Our findings reveal dynamic maintenance of mitochondrial genome fitness, and reduction in the effectiveness of purifying selection during life. Importantly, we describe interventions that mitigate and even reverse age-associated genome degeneration in flies and in mice. Furthermore, mitigation of genome degeneration improved wellbeing in a Drosophila model of heteroplasmic mitochondrial disease.

Significance StatementIn contrast to the orderly segregation of nuclear DNA, mitochondrial genomes compete for replication and segregation. The abundance of mutants that emerge in a cell depends on their success in this competition. We show that quality control mechanisms put deleterious mutations at a disadvantage, but these mechanisms become ineffective during aging. The resulting rise in mutant genomes, which compromises vigor, can be suppressed using genetic backgrounds that enhance quality control. Feeding kinetin to adult Drosophila or mice also reduced the load of mutant mitochondrial genomes. This pharmacological reversal of the age associated deterioration of mtDNA quality suggests possible therapies to alleviate mitochondrial diseases and normal aging.


title: STAT3 regulates cytokine production downstream of TNFR1 in part by inducing expression of TNFAIP3/A20

abstract:
BackgroundPrevious work demonstrated that the Signal Transducer and Activator of Transcription 3 (STAT3) is activated downstream of the Type 1 TNF Receptor. However, whether and how STAT3 regulates gene expression downstream of TNFR1 has not been elucidated.

MethodsGlobal transcriptome analysis by RNA sequencing was performed in wild type and STAT3 knockout mouse embryonic fibroblasts (MEFs) stimulated with TNF. The fold changes in gene expression were assessed bioinformatically. Results of the RNA sequencing were validated at the protein level by using multiplex cytokine assays and immunoblotting.

ResultsStimulation of MEFs with TNF or an agonist antibody to TNFR1 activated STAT3, and this was inhibited by pharmacological inhibition of Jak2 and cSrc. At 4 hours after TNF stimulation, STAT3 knockout MEFs had a greater level than WT MEFs of induction of the chemokines Ccl2, Cxcl1 and Cxcl10 at the RNA and protein levels. Mechanistically, this was due STAT3 promoting the expression of Tnfaip3/A20, a ubiquitin modifying enzyme that inhibits inflammation, in wild-type MEFs at early timepoints after TNFR1 stimulation. In STAT3 knockout MEFs TNF failed to induce the expression of Tnfaip3/A20 or GM-CSF when acting through TNFR1. Expression of A20 into STAT3 knockout MEFs suppressed cytokine expression.

ConclusionSTAT3 limits the induction of Ccl2, Cxcl1 and Cxcl10 in response to TNFR1 activation by promoting the expression of Tnfaip3/A20. On the other hand, STAT3 promotes the expression of GM-CSF in response to TNFR1 stimulation. These results show that STAT3 modulates inflammatory signaling by TNF in normal cells.


title: Honey bee flights under ethanol-exposure show changes in body and wing kinematics

abstract: Flying social insects can provide models of the interactions needed for aerial robot swarms having limited processing resources. The ability to simultaneously make precise measurements of insect wing and body motions on such interacting insects is a recent capability, and chemical exposure may be used to modulate the interactions between insects. These interaction effects must be distinguished from the chemicals effect on non-interacting insects. In this experiment, four high speed cameras (9000 fps) were used to track the wing and body motions of insects (Apis mellifera). Digitization, consisting of data association, hull reconstruction, and segmentation, achieved the first quantitative high speed measurements of ethanol exposed honey bees wing and body motions. Kinematic analysis considered the trial wide mean and maximum values and gross wingstroke parameters, and tested deviations for statistical significance using Welchs t-test and Cohens d test. The results indicate a decrease in maximal heading and pitch rates of the body, and that roll rate is affected at high concentrations (5%). The wingstroke effects include a stroke frequency decrease, stroke amplitude increase, stroke inclination angle increase, and a more planar wingstroke. These effects due to ethanol exposure are valuable tools to separate from interaction effects.

category: animal behavior and cognition


title: Using the UniFrac metric on Whole Genome Shotgun data

abstract: The UniFrac metric has proven useful in revealing diversity across metagenomic communities. Due to the phylogeny-based nature of this measurement, UniFrac has historically only been applied to 16S rRNA data. Simultaneously, Whole Genome Shotgun (WGS) metagenomics has been increasingly widely employed and proven to provide more information than 16S data, but a UniFrac-like diversity metric suitable for WGS data has not previously been developed. The main obstacle for UniFrac to be applied directly to WGS data is the absence of phylogenetic distances in the taxonomic relationship derived from WGS data. In this study, we demonstrate a method to overcome this intrinsic difference and compute the UniFrac metric on WGS data by assigning branch lengths to the taxonomic tree obtained from input taxonomic profiles. We conduct a series of experiments to demonstrate that this WGSUniFrac method is comparably robust to traditional 16S UniFrac and is not highly sensitive to branch lengths assignments, be they data-derived or model-prescribed. Code implementing a prototype of WGSUniFrac along with paper reproducible are available at https://github.com/KoslickiLab/WGSUniFrac.

category: bioinformatics


title: Expansion and Adaptive Evolution of the mTERF Gene Family in Plants

abstract: Mitochondrial transcription termination factor (mTERF) genes are encoded in the nucleus and bind to nucleic acids to regulate the replication, transcription and translation of mitochondrial genomes. Plants possess a large family of mTERF genes that play important roles in regulating organellar gene expression and stress response. However, their origin and expansion in land plants has not been examined. Here, we conducted a comprehensive molecular evolution analysis of 611 mTERF genes identified in 18 plant species, including algae, moss, fern, gymnosperm and flowering plants. Higher plants have more mTERF genes compared to lower plants, forming a huge higher plant-specific clade (M-class mTERF genes). M-class mTERF genes occur in clusters, suggesting that tandem duplication contributed to their expansion. Compared to other mTERF genes, M-class mTERF genes have undergone rapid evolution, and several significant positively selected sites were located in nucleic acid-binding sites. The strong correlation between the number of M-class mTERF genes and corresponding mitochondrial genome variation suggests that the rapid evolution of M-class mTERF genes might account for the changes in the complex machinery for expression regulation of plant mitochondrial genomes, providing molecular evidence for the host-parasite interaction hypothesis between the nucleus and mitochondria.

category: plant biology


title: Denoising diffusion weighted imaging data using convolutional neural networks

abstract: Diffusion weighted imaging (DWI) with multiple, high b-values is critical for extracting tissue microstructure measurements; however, high b-value DWI images contain high noise levels that can overwhelm the signal of interest and bias microstructural measurements. Here, we propose a simple denoising method that can be applied to any dataset, provided a low-noise, single-subject dataset is acquired using the same DWI sequence. The denoising method uses a one-dimensional convolutional neural network (1D-CNN) and deep learning to learn from a low-noise dataset, voxel-by-voxel. The trained model can then be applied to high-noise datasets from other subjects. We validated the 1D-CNN denoising method by first demonstrating that 1D-CNN denoising resulted in DWI images that were more similar to the noise-free ground truth than comparable denoising methods, e.g., MP-PCA, using simulated DWI data. Using the same DWI acquisition but reconstructed with two common reconstruction methods, i.e. SENSE1 and sum-of-square, to generate a pair of low-noise and high-noise datasets, we then demonstrated that 1D-CNN denoising of high-noise DWI data collected from human subjects showed promising results in three domains: DWI images, diffusion metrics, and tractography. In particular, the denoised images were very similar to a low-noise reference image of that subject, more than the similarity between repeated low-noise images (i.e. computational reproducibility). Finally, we demonstrated the use of the 1D-CNN method in two practical examples to reduce noise from parallel imaging and simultaneous multi-slice acquisition. We conclude that the 1D-CNN denoising method is a simple, effective denoising method for DWI images that overcomes some of the limitations of current state-of-the-art denoising methods, such as the need for a large number of subjects for training and accounting for the rectified noise floor.


title: Cell atlas of the human ocular anterior segment: Tissue-specific and shared cell types

abstract:
The anterior segment of the eye consists of the cornea, iris, ciliary body, crystalline lens and aqueous humor outflow pathways. Together, these tissues are essential for the proper functioning of the eye. Disorders of vision have been ascribed to defects in all of them; some, including glaucoma and cataract, are among the most prevalent causes of blindness in the world. To characterize the cell types that comprise these tissues, we generated an anterior segment cell atlas of the human eye using high throughput single-nucleus RNA sequencing (snRNAseq). We profiled 191,992 nuclei from non-diseased anterior segment tissues from 6 human donors, identifying >60 cell types. Many of these cell types were discrete, whereas others, especially in lens and cornea, formed continua corresponding to known developmental transitions that persist in adulthood. Having profiled each tissue separately, we performed an integrated analysis of the entire anterior segment revealing that some cell types are unique to single structure whereas others are shared across tissues. The integrated cell atlas was then used to investigate cell type-specific expression patterns of more than 900 human ocular disease genes identified either through Mendelian inheritance patterns or genome-wide association studies (GWAS).

SIGNIFICANCE STATEMENTSeveral of the most prevalent blinding ocular conditions worldwide, including glaucoma, cataract and uncorrected refractive error, involve structures of the anterior segment of the human eye, which consists of the cornea, iris, ciliary body, crystalline lens and aqueous humor outflow pathways. In addition to providing transcriptomic profiles of the cell types within individual tissues, this work contributes to our understanding of the relatedness and diversity of these cell types across contiguous tissues by generating an integrated anterior segment cell atlas and documenting the expression of over 900 disease-associated genes in each cell type. By allowing simultaneous interrogation of cell-type specific expression of genes across multiple tissues, the atlas may yield broad insight into normal and disease-associated anterior segment functions.


title: SAIBR: A simple, platform-independent method for spectral autofluorescence correction

abstract:
Biological systems are increasingly viewed through the lens of mathematics, physics, and systems approaches that demand accurate quantification of gene expression and local protein concentrations. Such approaches have benefited greatly from the revolution in genetic engineering sparked by CRISPR/Cas9. By facilitating the tagging of genes at their genomic loci, CRISPR/Cas9 allows us to use fluorescence to monitor proteins that are expressed at or near endogenous levels under native regulatory control. However, due to their typically lower expression levels, quantitative experiments using endogenously-tagged genes can run into limits imposed by autofluorescence (AF). AF is often a particular challenge in the illumination bands occupied by the most efficient fluorescent proteins (GFP, mNeonGreen). Stimulated by our work in C. elegans, we describe and validate Spectral Autofluorescence Image correction By Regression (SAIBR), a simple, platform-independent protocol, and associated GUI-based FIJI plugin to correct for autofluorescence using standard filter sets and illumination conditions. Fully validated for use in C. elegans embryos and tested in diverse systems, including starfish oocytes and fission yeast, SAIBR achieves accurate quantitation of fluorophore signal and enables reliable detection and quantification of even weakly expressed proteins. Thus, SAIBR provides a highly accessible, low barrier way to incorporate AF correction as standard for researchers working on a broad variety of cell and developmental systems.

Summary StatementImplemented as an easy-to-use Fiji Plugin, SAIBR provides effective autofluorescence correction for cells and tissues using standard imaging conditions.


title: Identification of replication fork-associated proteins in Drosophila embryos and cultured cells using iPOND coupled to quantitative mass spectrometry

abstract: Replication of the eukaryotic genome requires the formation of thousands of replication forks that must work in concert to accurately replicate the genetic and epigenetic information. Defining replication fork-associated proteins is a key step in understanding how genomes are replicated and repaired in the context of chromatin to maintain genome stability. To identify replication fork-associated proteins, we performed iPOND (Isolation of Proteins on Nascent DNA) coupled to quantitative mass spectrometry in Drosophila embryos and cultured cells. We identified 76 and 278 fork-associated proteins in post-MZT embryos and Drosophila cultured S2 cells, respectively. By performing a targeted screen of a subset of these proteins, we demonstrate that BRWD3, a targeting specificity factor for the DDB1/Cul4 ubiquitin ligase complex (CRL4), functions at replication forks to promote fork progression and maintain genome stability. Altogether, our work provides a valuable resource for those interested in the DNA replication, repair and chromatin assembly during development.

category: biochemistry


title: SERM: a self-consistent deep learning solution forrapid and accurate gene expression recovery

abstract: Single cell RNA sequencing (scRNA-seq) is a promising technique to determine the states of individual cells and classify novel cell subtypes. Computationally, the processing of scRNA-seq data presents a daunting challenge because of the noisy nature and humongous size and dimensionality of the data. Compromised solution by omitting the genes with low expression is commonly taken in current scRNA-seq analysis, which leads to inaccurate gene counts. In this paper, we introduce a broadly applicable data-driven gene expression recovery framework, referred to as the self-consistent expression recovery machine (SERM), to impute the missing gene expression. Using deep learning, SERM first learns from a subset of the noisy gene expression data to estimate the underlying data distribution. SERM then recovers the overall gene expression data by imposing a self-consistency on the gene expression matrix, thus ensuring that the expression levels are similarly distributed in different parts of the matrix. We show that SERM significantly improves the accuracy of gene imputation with at least 100-fold increase in computational efficiency in comparison to the state-of-the-art techniques. Thus SERM promises to provide an urgently needed computational solution for rapid and accurate recovery of big genomic expression data. SERM is available as a web-based computational tool (https://www.analyxus.com/compute/serm) and its source codes can be found in https://github.com/xinglab-ai/self-consistent-expression-recovery-machine.


title: Redefining the architecture of ferlin proteins: insights into multi-domain protein structure and function

abstract: Ferlins are complex, multidomain proteins whose functions involve membrane fusion, membrane repair, and exocytosis. Progress in studying the domain composition of ferlins has been problematic due to the large size of the ferlin proteins and the difficulty in computing accurate residue ranges for each respective ferlin protein domain. However, recent advances in in silico protein folding methods have significantly enhanced the understanding of the domain structure of these complex proteins. To compute unbiased domain boundaries, we used RoseTTAFold to assemble full-length models for each of the six human ferlin proteins (dysferlin, myoferlin, otoferlin, Fer1L4, Fer1L5, and Fer1L6). Despite the differences in amino acid sequence between the ferlin proteins, the domain ranges and the distinct subdomains in some of the ferlin domains were remarkably consistent. Further, the RoseTTAFold/AlphaFold2 in silico boundary prediction methods allowed us to describe and characterize a previously unknown C2 domain, ubiquitous in all human ferlins, which we refer to as C2-FerA. At present, the ferlin domain-domain interactions implied by the full-length in silico models are predicted to have a low accuracy; however, the use of RoseTTAFold and AlphaFold2 as a domain finder has proven to be a powerful research tool in understanding ferlin structure.

category: biophysics


title: Curcumin and NCLX Inhibitors Share Anti-Tumoral Mechanisms in Microsatellite-Instability-Driven Colorectal Cancer

abstract:
Colorectal cancer (CRC) is associated with high mortality worldwide and new targets are needed to overcome treatment resistance. Recent evidences highlight a role of the mitochondria calcium homeostasis in the development of CRC. In this context, we aimed to evaluate the role of the mitochondrial sodium-calcium-lithium exchanger (NCLX) and its targeting in CRC. We also identified curcumin as a new potential inhibitor of NCLX.

In vitro, curcumin exerted strong anti-tumoral activity through its action on NCLX with mtCa2+ and reactive oxygen species overload associated with a mitochondrial membrane depolarization, leading to reduced ATP production and apoptosis through mitochondrial permeability transition pore opening concomitant with G2/M cell cycle arrest. NCLX inhibition with either CGP37157 (a benzodiazepine derivative), small interfering RNA-mediated knock-down or knockout approaches reproduced the effects of curcumin. Altered mitochondrial respiration, cellular aerobic glycolysis and endoplasmic reticulum-mitochondria membrane perturbations participated in these mechanisms. In a xenograft mouse model, NCLX inhibitors decreased CRC tumor growth. Both transcriptomic analysis of The Cancer Genome Atlas dataset and immunohistochemical analysis of tissue microarrays from 381 patients with microsatellite instability (MSI)-driven CRC demonstrated that higher NCLX expression was associated with MSI status and for the first time NCLX expression was significantly associated with recurrence-free survival in MSI CRC patients.

Our findings provide strong evidence that blocking NCLX inhibits CRC in vitro and in vivo. We highlight a novel anti-tumoral mechanism of curcumin through its action on NCLX and mitochondria calcium overload that could benefit for therapeutic treatment of patients with MSI CRC.

category: cancer biology


title: In Vitro and In Vivo Pathogenicity of A. alternata to Sugar beet and Assessment of Sensitivity to Fungicides

abstract: A. alternata is a weak and opportunistic pathogen, but recently this pathogen has been reported in major sugar beet growing regions in the US. Therefore, we were interested to investigate the in-vitro and in-vivo pathogenesis of A. alternata to sugar beet seed, seedlings, tissue slices, stecklings, and roots. The in-vitro study demonstrated that A. alternata reduces seed germination, causes damping-off of seedlings, and decomposes beet tissue. The in-vivo study showed the same inoculum responsible for Alternaria leaf spots, and black circular spots on beetroot. Furthermore, we did a preliminary study to determine in vitro sensitivities of some fungicides to determine the effective concentration that inhibited 50% of radial mycelial growth (EC50 values) of A. alternata. The results indicated that EC50 values ranged from 0.0023 to 0.0058 for PROLINE, from 0.0004 to 0.0021 for OMEGA, and from 0.0027 to 0.0053 for Super TIN. Among the potential fungicides, the efficiency of quinone outside inhibitors (QoI- PRIAXOR) was relatively higher to inhibit the radial growth of A. alternata, irrespective of concentrations.


title: Physiological responses of plants to in vivo XRF radiation damage: insights from elemental, histochemical, anatomical and ultrastructural analyses

abstract: X-ray fluorescence spectroscopy (XRF) is a powerful technique for the in vivo assessment of plant tissues. However, the potential X-ray exposure damages might affect the structure and elemental composition of living plant tissues leading to artefacts on the recorded data. Herein, we exposed soybean (Glycine max (L.) Merrill) leaves to several X-ray doses through a polychromatic benchtop microprobe X-ray fluorescence spectrometer, modulating the photon flux by adjusting either the beam size, focus, or exposure time. The structure and physiological responses of the irradiated plant tissues were investigated through light and transmission electron microscopy (TEM). Histochemical tests were also conducted. Depending on the dose, the X-ray exposure induced decreased K and X-ray scattering intensities, and increased Ca, P, and Mn signals on soybean leaves. Anatomical analysis indicated necrosis of the epidermal and mesophyll cells on the irradiated spots, where TEM images revealed the collapse of cytoplasm and cell-wall breaking. Furthermore, the histochemical analysis detected the production of reactive oxygen species, as well as inhibition of chlorophyll autofluorescence in these areas. Under certain X-ray exposure conditions, e.g., high photon flux and exposure time, XRF measurements may affect the soybean leaves structures and ultrastructure, elemental composition, cellular ultrastructure, and induce programmed cell death. These results shed light on the characterization of the radiation damage, and thus, help to assess the X-ray radiation limits and strategies for in vivo for XRF analysis.


title: Phenotypic Pliancy and the Breakdown of Epigenetic Polycomb Mechanisms

abstract:
Epigenetic regulatory mechanisms allow multicellular organisms to develop distinct specialized cell identities despite having the same total genome. Cell-fate choices are based on gene expression programs and environmental cues that cells experience during embryonic development, and are usually maintained throughout the life of the organism despite new environmental cues. The evolutionarily conserved Polycomb group (PcG) proteins form Polycomb Repressive Complexes (PRCs) that help orchestrate these developmental choices. Post-development, these complexes actively maintain the resulting cell fate, even in the face of environmental perturbations. Given the crucial role of these polycomb mechanisms in providing phenotypic fidelity (i.e. maintenance of cell fate), we hypothesize that their dysregulation after development will lead to decreased phenotypic fidelity allowing dysregulated cells to sustainably switch their phenotype in response to environmental changes. We call this abnormal phenotypic switching phenotypic pliancy. We introduce a computational evolutionary model that allows us to test our systems-level phenotypic pliancy hypothesis in-silico and in a context-independent manner. We find that 1) phenotypic fidelity is an emergent systems-level property of PcG-like mechanisms, and 2) phenotypic pliancy is an emergent systems-level property resulting from this mechanisms dysregulation. Since there is evidence that metastatic cells behave in a phenotypically pliant manner and PcG dysregulation is common in cancer, we hypothesize that progression to metastasis is driven by the emergence of phenotypic pliancy in cancer cells as a result of PcG mechanism dysregulation. We corroborate our hypothesis and the results of our computational model using single-cell RNA-sequencing data from metastatic cancers. We find that metastatic cancer cells are phenotypically pliant in the same manner as predicted by our model.

Significance StatementWe introduce the concept of cellular phenotypic pliancy- sustained abnormal phenotypic switching in response to environmental changes- and demonstrate that such behavior can be caused by dysregulation of Polycomb mechanisms. To overcome the incomplete knowledge about these mechanisms in higher organisms, we develop an abstract computational model to study the emergence of phenotypic pliancy from a general systems-level view by confirming our hypothesis over a wide range of simulated gene-regulatory networks and Polycomb mechanisms patterns. We corroborate our hypothesis and model predictions using single-cell RNA-seq metastatic cancer datasets. Our hypothesis has the potential to shed light on a general phenomenon for complex diseases where abnormal phenotypic switching is relevant.

category: systems biology


title: Interplay between murine sex-biased gene expression and hepatic zonation revealed by single nucleus RNA sequencing

abstract:
The zonation of liver metabolic processes is well-characterized; however, little is known about the cell type-specificity and zonation of sexually dimorphic gene expression or its growth hormone (GH)-dependent transcriptional regulators. We address these issues using single nucleus RNA sequencing of 32,000 nuclei representing nine major liver cell types. Nuclei were extracted from livers from young adult male and female mice, from male mice infused with GH continuously to mimic the female plasma GH pattern, and from mice treated with TCPOBOP, a xenobiotic agonist ligand of the liver nuclear receptor CAR (Nr1i3). Analysis of these rich transcriptomic datasets revealed: 1) expression of sex-biased genes and their key GH-dependent transcriptional regulators is primarily restricted to hepatocytes and is not a feature of liver non-parenchymal cells; 2) many sex-biased transcripts show sex-dependent zonation within the liver lobule; 3) gene expression is substantially feminized in both periportal and pericentral hepatocytes when male mice are infused with GH continuously; 4) sequencing nuclei increases the sensitivity for detecting thousands of nuclear-enriched lncRNAs and enables determination of their liver cell type-specificity, sex bias and hepatocyte zonation profiles; 5) the periportal to pericentral hepatocyte cell ratio is significantly higher in male than female liver; and 6) TCPOBOP exposure disrupts sex-specific gene expression and hepatocyte zonation within the liver lobule. These findings highlight the complex interconnections between hepatic sexual dimorphism and zonation at the single cell level and reveal how endogenous hormones and foreign chemical exposure can alter these interactions across the liver lobule with large effects on both protein-coding genes and lncRNAs.

SynopsisSingle nucleus RNA-seq analysis elucidated the cell type-specificity and zonation of the sex-biased murine liver transcriptome, including thousands of long non-coding RNAs. Xenobiotic exposure induced widespread dysregulation, including both gain and loss of sex-biased gene expression and changes in zonation.


title: BIN1 regulates electrical activity and network synchronization in hiPSC-derived neurons

abstract: Alzheimers disease (AD) is a multifactorial disease with a strong genetic background. Recent genomic wide association studies have identified several loci linked to an increased risk of AD. However, genes regulated by these variants and the pathophysiological mechanisms regulated by those genes remain largely elusive. In this work, we studied the role of Bridging Integrator 1 (BIN1), the second most important AD risk gene after APOE, in neurons generated from human induced pluripotent stem cells (hiPSCs) in bi-dimensional cultures and cerebral organoids. We show that deletion of BIN1 is sufficient to cause neuronal hyperactivation and network desynchronization in a cell-autonomous fashion. These functional changes are correlated with increased Tau phosphorylation and transcriptional changes similar to those observed in the brains of AD patients, particularly in glutamatergic neurons. Together, our results reveal a role for BIN1 in the regulation of electrical activity in human neurons and suggest that its implication in AD pathogenesis could be related to the neuronal and network dysfunctions observed in the AD brain.


title: c-Fos protein shRNA blockade in the central amygdala nucleus interfere with rats emotional reactivity on behavioral and autonomic level

abstract: This report is focusing on a function of the c-Fos protein in an associative, stress-induced memory. The shRNA vector injections were utilised to functionally silence the central amygdala nucleus in adult Wistar rats. Subsequently the operated animals and their control counterparts were screened in a selection of an emotionally-dependent tests and in a few standard behavioral neuroscience tools. Rats from the c-Fos silencing (ShFos) group expressed contra-depressive-like behaviors in Porsolt Swimming Test, spending more time actively searching for escape way then the rats from the control group. ShFos rats also had engaged in a more rapid activity in the Open Field Test, showing a decline in the neo-phobia. Micturition was decreased in shFos animals, indicating a change in the emotionality on an autonomic level. Presented results are showcasing a multi-directional regulation of the behaviors from the central amygdala nucleus by the c-Fos activity.


title: Central role of Prominin-1 in lipid rafts during liver regeneration

abstract: Prominin-1 (PROM1), a lipid raft protein, is required for maintaining cancer stem cell properties in hepatocarcinoma cell lines, but its physiological roles in the liver have not been well studied. Here, we investigated the role of PROM1 in lipid rafts with a precise molecular mechanism during liver regeneration. We found that the expression of PROM1 increased during liver regeneration after 2/3 partial hepatectomy (PHx) or CCl4 injection. Interestingly, hepatocyte proliferation and liver regeneration were attenuated in liver-specific Prom1 knockout (Prom1LKO) mice compared to wild-type (Prom1f/f) mice. Detailed mechanistic studies revealed that PROM1 interacted with the interleukin-6 signal transducer glycoprotein 130 (GP130) and confined GP130 to lipid rafts so that STAT3 signaling by IL-6 was effectively activated. Moreover, the overexpression of the glycosylphosphatidylinsositol (GPI)-anchored first extracellular domain of PROM1 (PROM1GPI-EX1), which is a domain that binds to GP130, rescued the proliferation of hepatocytes and liver regeneration in Prom1LKO mice. PROM1 is upregulated in hepatocytes during liver regeneration, and upregulated PROM1 recruits GP130 into lipid rafts and activates the IL6-GP130-STAT3 axis. Thus, we conclude that PROM1 plays an important role in lipid rafts during liver regeneration and might be a promising target for therapeutic applications of liver transplantation.


title: PP1 phosphatases control PAR-2 localization and polarity establishment in C. elegans embryos

abstract:
Cell polarity relies on the asymmetric distribution of the conserved PAR proteins, which is regulated by phosphorylation/dephosphorylation reactions. While the kinases involved have been well studied, the role of phosphatases remains poorly understood. In C. elegans zygotes, phosphorylation of the posterior PAR-2 protein by the atypical protein kinase PKC-3 inhibits PAR-2 cortical localization. Polarity establishment depends on loading of PAR-2 at the posterior cortex. We show that the PP1 phosphatases GSP-1 and GSP-2 are required for polarity establishment in embryos. We find that co-depletion of GSP-1 and GSP-2 abrogates the cortical localization of PAR-2 and that GSP-1 and GSP-2 interact with PAR-2 via a PP1 docking motif in PAR-2. Mutating this motif in vivo, to prevent binding of PAR-2 to PP1, abolishes cortical localization of PAR-2, while optimizing this motif extends PAR-2 cortical localization. Our data suggest a model in which GSP-1/-2 counteract PKC-3 phosphorylation of PAR-2 allowing its cortical localization at the posterior and polarization of the one-cell embryo.

SUMMARYCalvi et al. identify PP1 protein phosphatases as regulators of cell polarity in C. elegans embryos. Their results show that two redundant phosphatases, GSP-1 and GSP-2, interact with the polarity protein PAR-2 and control its localization and polarity establishment.


title: Overlapping transcriptional programs promote survival and axonal regeneration of injured retinal ganglion cells

abstract: Injured neurons in the adult mammalian central nervous system often die and seldom regenerate axons. To uncover transcriptional pathways that could ameliorate these disappointing responses we analyzed three interventions that increase survival and regeneration of mouse retinal ganglion cells (RGCs) following optic nerve crush (ONC) injury, albeit not to a clinically useful extent. We assessed gene expression in each of 46 RGC types by single cell transcriptomics following ONC and treatment. We also compared RGCs that regenerated to those that survived but did not regenerate. Each intervention enhanced survival of most RGC types, but type-independent axon regeneration required manipulation of multiple pathways. Distinct computational methods converged on separate sets of genes selectively expressed by RGCs likely to be dying, surviving, or regenerating. Overexpression of genes associated with the regeneration program enhanced axon regeneration in vivo, indicating that mechanistic analysis can be used to identify novel methods for promoting regeneration of injured neurons.


title: Discovery of a SARS-CoV-2 Broadly-Acting Neutralizing Antibody with Activity against Omicron and Omicron + R346K Variants

abstract: The continual emergence of SARS-CoV-2 variants of concern, in particular the newly emerged Omicron (B.1.1.529) variant, has rendered ineffective a number of previously EUA approved SARS-CoV-2 neutralizing antibody therapies. Furthermore, even those approved antibodies with neutralizing activity against Omicron are reportedly ineffective against the subset of Omicron variants that contain a R346K substitution, demonstrating the continued need for discovery and characterization of candidate therapeutic antibodies with the breadth and potency of neutralizing activity required to treat newly diagnosed COVID-19 linked to recently emerged variants of concern. Following a campaign of antibody discovery based on the vaccination of Harbour H2L2 mice with defined SARS-CoV-2 spike domains, we have characterized the activity of a large collection of Spike-binding antibodies and identified a lead neutralizing human IgG1 LALA antibody, STI-9167. STI-9167 has potent, broad-spectrum neutralizing activity against the current SARS-COV-2 variants of concern and retained activity against the Omicron and Omicron + R346K variants in both pseudotype and live virus neutralization assays. Furthermore, STI-9167 nAb administered intranasally or intravenously provided protection against weight loss and reduced virus lung titers to levels below the limit of quantitation in Omicron-infected K18-hACE2 transgenic mice. With this established activity profile, a cGMP cell line has been developed and used to produce cGMP drug product intended for use in human clinical trials.

category: microbiology


title: Computation of Antigenicity Predicts SARS-CoV-2 Vaccine Breakthrough Variants

abstract: It has been reported that multiple SARS-CoV-2 variants of concerns (VOCs) including B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma), and B.1.617.2 (Delta) can reduce neutralisation by antibodies, resulting in vaccine breakthrough infections. Virus-antiserum neutralisation assays are typically performed to monitor potential vaccine breakthrough strains. However, such experimental-based methods are slow and cannot instantly validate whether newly emerging variants can break through current vaccines or therapeutic antibodies. To address this, we sought to establish a computational model to predict the antigenicity of SARS-CoV-2 variants by sequence alone and in real time. In this study, we firstly identified the relationship between the antigenic difference transformed from the amino acid sequence and the antigenic distance from the neutralisation titres. Based on this correlation, we obtained a computational model for the receptor binding domain (RBD) of the spike protein to predict the fold decrease in virus-antiserum neutralisation titres with high accuracy (~0.79). Our predicted results were comparable with experimental neutralisation titres of variants, including B.1.1.7 (Alpha), B.1.351 (Beta), B.1.617.2 (Delta), B.1.429 (Epsilon), P.1 (Gamma), B.1.526 (Iota), B.1.617.1 (Kappa), and C.37 (Lambda), as well as SARS-CoV. Here, we firstly predicted the fold of decrease of B.1.1.529 (Omicron) as 17.4-fold less susceptible to neutralisation. We visualised all 1521 SARS-CoV-2 lineages to indicate variants including B.1.621 (Mu), B.1.630, B.1.633, B.1.649, and C.1.2, which can induce vaccine breakthrough infections in addition to reported VOCs B.1.351 (Beta), P.1 (Gamma), B.1.617.2 (Delta), and B.1.1.529 (Omicron). Our study offers a quick approach to predict the antigenicity of SARS-CoV-2 variants as soon as they emerge. Furthermore, this approach can facilitate future vaccine updates to cover all major variants. An online version can be accessed at http://jdlab.online.

category: evolutionary biology


title: MaxQuant and MSstats in Galaxy enable reproducible cloud-based analysis of quantitative proteomics experiments for everyone

abstract: Quantitative mass spectrometry-based proteomics has become a high-throughput technology for the identification and quantification of thousands of proteins in complex biological samples. Two de facto standard tools, MaxQuant and MSstats, allow for the analysis of raw data and finding proteins with differential abundance between conditions of interest. To enable accessible and reproducible quantitative proteomics analyses in a cloud environment, we have integrated MaxQuant (including TMTpro 16/18plex), Proteomics Quality Control (PTXQC), MSstats and MSstatsTMT into the open-source Galaxy framework. This enables the web-based analysis of label-free and isobaric labeling proteomics experiments via Galaxys graphical user interface on public clouds. MaxQuant and MSstats in Galaxy can be applied in conjunction with thousands of existing Galaxy tools and integrated into standardized, sharable workflows. Galaxy tracks all metadata and intermediate results in analysis histories, which can be shared privately for collaborations or publicly, allowing full reproducibility and transparency of published analysis. To further increase accessibility, we provide detailed hands-on training materials. The integration of MaxQuant and MSstats into the Galaxy framework enables their usage in a reproducible way on accessible large computational infrastructures, hence realizing the foundation for high throughput proteomics data science for everyone.


title: The kinase inhibitor Palbociclib is a potent and specific RNA-binding molecule

abstract: The growing awareness of the role of RNA in human disease has motivated significant efforts to discover drug-like small molecules that target RNA. However, high throughput screening campaigns report very low hit rates and generally identify compounds with weak affinity, while most structures reported in Academic studies also lack the pharmacological properties of successful drugs. Even FDA-approved RNA-targeting drugs have only weak (10 M) binding activity. Thus, it is often stated that only complex RNA structures, such as the ribosome or riboswitches, are amenable to small molecule chemistry. We report that the kinase inhibitor Palbociclib/Ibrance is a nM ligand for the HIV-1 TAR. It inhibits recruitment of the positive transcription elongation factor complex at nM concentrations and discriminates >20 fold. We further show that RNA binding can be fully decoupled from kinase inhibition, yielding a new molecule with even higher affinity for RNA. We thus demonstrate that nM affinity, specificity, and potent biochemical activity against  undruggable RNAs can be found in the chemical space of blockbuster drugs.


title: AI-based graptolite identification improves shale gas exploration

abstract: Graptolites are fossils from the mid-Cambrian to lower Carboniferous periods that inform both our understanding of evolution and the exploration of shale gas [1-4]. The identification of graptolite species remains a visual task carried out by experienced taxonomists because their fine-grained morphologies and incomplete preservation in multi-fossil samples have hindered automation. Artificial intelligence (AI) holds great promise for transforming such meticulous tasks, and has already proven useful in applications ranging from animal classification to medical diagnostics [5-15]. Here we demonstrate that graptolites can be identified with taxonomist accuracy using a deep learning model [16-18]. We develop a convolutional neural network to classify macrofossils, and construct a comprehensive dataset of >34,000 images of 113 graptolite species annotated at pixel-level resolution to train the model. We validate the models performance by comparing its ability to identify 100 images of graptolite species that are significant for rock dating and shale gas exploration with 21 experienced taxonomists from research institutes and the shale gas industry. Our model achieves 86% and 81% accuracy when identifying the genus and species of graptolites, respectively; outperforming taxonomists in terms of accuracy, time, and generalization. By investigating the decisions made by the neural network, we further show that it can recognise fine-grained morphological details better than taxonomists. Our AI approach, providing taxonomist-level graptolite identification, can be deployed on web and mobile apps to extend graptolite identification beyond research institutes and improve the efficiency of shale gas exploration.

category: paleontology


title: Longitudinal Assessment of SARS-CoV-2 Specific T Cell Cytokine-Producing Responses for 1 Year Reveals Persistence of Multi-Cytokine Proliferative Responses, with Greater Immunity Associated with Disease Severity

abstract:
Cellular-mediated immunity is critical for long-term protection against most viral infections, including coronaviruses. We studied 23 SARS-CoV-2-infected survivors over a one year post symptom onset (PSO) interval by ex vivo cytokine ELISpot assay. All subjects demonstrated SARS-CoV-2-specific IFN-{gamma}, IL-2, and Granzyme B (GzmB) T cell responses at presentation, with greater frequencies in severe disease. Cytokines, mainly produced by CD4+ T cells, targeted all structural proteins (Nucleocapsid, Membrane, Spike) except Envelope, with GzmB > IL-2 > IFN-{gamma}. Mathematical modeling predicted that: 1) cytokine responses peaked at 6 days for IFN-{gamma}, 36 days for IL-2, and 7 days for GzmB, 2) severe illness was associated with reduced IFN-{gamma} and GzmB, but increased IL-2 production rates, 3) males displayed greater production of IFN-{gamma}, whereas females produced more GzmB. Ex vivo responses declined over time with persistence of IL-2 in 86% and of IFN-{gamma} and GzmB in 70% of subjects at a median of 336 days PSO. The average half-life of SARS-CoV-2-specific cytokine-producing cells was modelled to be 139 days ([~]4.6 months). Potent T cell proliferative responses persisted throughout observation, were CD4 dominant, and were capable of producing all 3 cytokines. Several immunodominant CD4 and CD8 epitopes identified in this study were shared by seasonal coronaviruses or SARS-CoV-1 in the Nucleocapsid and Membrane regions. Both SARS-CoV-2-specific CD4+ and CD8+ T cell clones were able to kill target cells, though CD8 tended to be more potent.

ImportanceOur findings highlight the relative importance of SARS-CoV-2-specific GzmB-producing T cell responses in SARS-CoV-2 control, shared CD4 and CD8 immunodominant epitopes in seasonal coronaviruses or SARS-CoV-1, and indicate robust persistence of T cell memory at least one year after infection. Our findings should inform future strategies to induce T cell vaccines against SARS-CoV-2 and other coronaviruses.


title: Hurricane Harvey Impacts on Water Quality and Microbial Communities in Houston, TX Waterbodies

abstract: Hurricanes, and other extreme weather events, can temporarily alter the structure of coastal systems and generate floodwaters that are contaminated with fecal-associated microbes; however, every coastal system and extreme weather event is unique, so identification of trends and commonalities in these episodic events is challenging. To improve our understanding of the resilience of coastal systems to the disturbance of extreme weather events and the risk exposure to floodwaters poses to the public, we monitored water quality, fecal indicator bacteria (FIB) at three stations within Clear Lake, an estuary between Houston and Galveston, and three stations in bayous that feed into the estuary. Water samples were collected immediately before Hurricane Harvey (pre-HH, August 25th, 2017), immediately after (HH, August 30th) and then throughout the fall of 2017 (post-HH). FIB levels were monitored by culturing E. coli and Enterococci. Microbial community structure was profiled by high throughput sequencing of PCR-amplified 16S rRNA gene fragments. Water quality and FIB data was also compared to historical data for these water body segments. Before Harvey, salinity within Clear Lake ranged from 9 to 11 practical salinity units (PSU). Immediately after the storm, salinity dropped to < 1 PSU and then gradually increased to pre-storm and historical levels over two months. Dissolved inorganic nutrient levels were also relatively low immediately after Harvey and returned, within a couple of months, to pre-HH and historical levels. FIB levels were elevated immediately after the storm; however, after one week, E. coli levels had decreased to acceptable levels for freshwater. Enterococci levels collected several weeks after the storm were within the range of historical levels for these water bodies. Microbial community structure shifted from a system dominated by Cyanobacteria sp. before HH to a system dominated by Proteobacteria and Bacteroidetes immediately after. Further, several sequences observed only in floodwater showed similarity to sequences previously reported for samples collected following Hurricane Irene. These changes in beta diversity corresponded to salinity and nitrate/nitrite concentrations. Differential abundance analysis of metabolic pathways, predicted from 16S sequences, suggested that pathways associated with virulence and antibiotic resistance were elevated in floodwater. Overall, these results suggest that recovery of the Clear Lake system following Hurricane Harvey took at least a month and floodwater generated from these extreme events may have high levels of fecal contamination, antibiotic resistant bacteria and bacteria rarely observed in other systems.


title: Evidence of antibodies against SARS-CoV-2 in wild mustelids from Brittany (France)

abstract: In the French region of Brittany, mainly in the department of the Cotes dArmor, during the first semester of 2021, seropositivity for SARS-CoV-2 was detected in five wild mustelids out of 32 animals tested. Anti-SARS-CoV-2 IgG against at least four out of five recombinant viral proteins (S1 receptor binding domain, nucleocapsid, S1 subunit, S2 subunit and spike) were detected using automated western blot technique in three martens (Martes martes) and two badgers (Meles meles). An ELISA test also objectified seropositivities. Although the 171 qPCRs carried out on samples from the 33 mustelids were all negative, these preliminary results (observational study) nevertheless bear witness to infections of unknown origin. The epidemiological surveillance of Covid-19 in wildlife must continue, in particular with the tools of efficient serology.


title: High abundance of transcription regulators compacts the nucleoid in Escherichia coli

abstract:
In enteric bacteria organization of the circular chromosomal DNA into a highly dynamic and toroidal shaped nucleoid involves various factors such as DNA supercoiling, nucleoid-associated proteins (NAPs), the structural maintenance of chromatin (SMC) complex, and macro-domain organizing proteins. Here we show that ectopic expression of transcription regulators at high levels leads to nucleoid compaction. This serendipitous result was obtained by fluorescence microscopy upon ectopic expression of the transcription regulator and phosphodiesterase PdeL of Escherichia coli of a strain expressing the mCherry-tagged HU- subunit (HupA) for nucleoid staining. Nucleoid compaction by PdeL depends on DNA-binding, but not on its enzymatic phosphodiesterase activity. Nucleoid compaction was also observed upon high-level ectopic expression of the transcription regulators LacI, RutR, RcsB, LeuO and Cra, which range from single target gene regulators to global regulators. In case of LacI its high-level expression in presence of the gratuitous inducer IPTG also led to nucleoid compaction indicating that compaction is caused by unspecific DNA-binding. In all cases nucleoid compaction correlated with misplacement of the FtsZ ring and loss of MukB foci, a subunit of the SMC complex. Thus, high levels of several transcription regulators cause nucleoid compaction with consequences on transcription, replication, and cell division.

ImportanceThe bacterial nucleoid is a highly organized and dynamic structure for simultaneous, transcription, replication and segregation of the bacterial genome. Compaction of the nucleoid and disturbance of DNA segregation and cell division by artificially high levels of transcription regulators, as described here, reveals that an excess of DNA-binding protein disturbs nucleoid structuring. The results suggest that ectopic expression levels of DNA-binding proteins for genetic studies of their function but also for their purification should be carefully controlled and adjusted.


title: Genomic library of Bordetella

abstract:
BackgroundThe re-emergence of whooping cough and geographic disparities in vaccine escape or antimicrobial resistance dynamics, underline the importance of a unified definition of Bordetella pertussis strains. Understanding of the evolutionary adaptations of Bordetella pathogens to humans and animals requires comparative studies with environmental bordetellae.

MethodsWe have set-up a unified library of Bordetella genomes by merging previously existing Oxford and Pasteur databases, importing genomes from public repositories, and developing harmonized genotyping schemes. We developed a genus-wide cgMLST genotyping scheme and incorporated a previous B. pertussis cgMLST scheme. Specific schemes were developed to define antigenic, virulence and macrolide resistance profiles. Genomic sequencing of 83 French B. bronchiseptica isolates and of B. tumulicola, B. muralis and B. tumbae type strains was performed.

ResultsThe public library currently includes 2,581 Bordetella isolates and their provenance data, and 2,084 genomes. The "classical Bordetella" (B. bronchiseptica, B. parapertussis and B. pertussis), which form a single genomic species (B. bronchiseptica genomic species, BbGS), were overrepresented (n=2,382). The phylogenetic analysis of Bordetella genomes associated the three novel species B. tumulicola, B. muralis and B. tumbae in a clade with B. petrii and revealed 18 yet undescribed species. A sister lineage of the classical bordetellae, provisionally named Bbs lineage II, was uncovered and may represent a novel species (average nucleotide identity with BbGS strains: [~]95%). It comprised strain HT200 from India, two strains of  genogroup 6 from the USA and six clinical isolates from France; this lineage lacked ptxP and its fim2 gene was divergent. Within B. pertussis, vaccine antigen sequence types marked important phylogenetic subdivisions, and macrolide resistance markers (23S_rRNA allele 13 and fhaB3) confirmed the current restriction of this phenotype in China with few exceptions.

ConclusionsThe genomic platform provides an expandable resource for unified genotyping of Bordetella strains and will facilitate collective evolutionary and epidemiological understanding of the re-emergence of whooping cough and other Bordetella infections.

Data summaryBordetella genomes list and accession numbers: Supplementary Table S4

Bordetella genus phylogeny dataset (92 isolates): https://bigsdb.pasteur.fr/cgi-bin/bigsdb/bigsdb.pl?db=pubmlst_bordetella_isolates&page=query&project_list=23&submit=1

B. bronchiseptica phylogeny dataset (213 isolates): https://bigsdb.pasteur.fr/cgi-bin/bigsdb/bigsdb.pl?db=pubmlst_bordetella_isolates&page=query&project_list=24&submit=1

B. pertussis phylogeny (124 isolates): https://bigsdb.pasteur.fr/cgi-bin/bigsdb/bigsdb.pl?db=pubmlst_bordetella_isolates&page=query&project_list=25&submit=1

iTOL interactive trees: https://itol.embl.de/shared/1l7Fw0AvKOoCF


title: Omicron variant of SARS-CoV-2 exhibits an increased resilience to the antiviral type I interferon response

abstract: The new variant of concern (VOC) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Omicron (B.1.1.529), is genetically very different from other VOCs. We compared Omicron with the preceding VOC Delta (B.1.617.2) and the wildtype strain (B.1) with respect to their interactions with the antiviral type I interferon (IFN-alpha/beta) response in infected cells. Our data indicate that Omicron has gained an elevated capability to suppress IFN-beta induction upon infection and to better withstand the antiviral state imposed by exogenously added IFN-alpha.


title: FoodMicrobionet v4: a large, integrated, open and transparent database for food bacterial communities

abstract:
With the availability of high-throughput sequencing techniques our knowledge of the structure and dynamics of food microbial communities has made a quantum leap. However, this knowledge is dispersed in a large number of papers and hard data are only partly available through powerful on-line databases and tools such as QIITA, MGnify and the Integrated Microbial Next Generation Sequencing platform, whose annotation is not optimized for foods.

Here, we present the 4th iteration of FoodMicrobionet, a database of the composition of bacterial microbial communities of foods and food environments. With 180 studies and 10,151 samples belonging to 8 major food groups FoodMicrobionet 4.1.2 is arguably the largest and best annotated database on food bacterial communities. This version includes 1,684 environmental samples and 8,467 food samples, belonging to 16 L1 categories and 196 L6 categories of the EFSA FoodEx2 classification and is approximately 4 times larger than previous version (3.1, https://doi.org/10.1016/j.ijfoodmicro.2019.108249).

Using data in FoodMicrobionet we confirm that taxonomic assignment at the genus level can be performed confidently for the majority of amplicon sequence variants using the most commonly used 16S RNA gene target regions (V1-V3, V3-V4, V4), with best results with higher quality sequences and longer fragment lengths, but that care should be exercised in confirming the assignment at species level.

Both FoodMicrobionet and related data and software conform to FAIR (findable, accessible, interoperable, reusable/reproducible) criteria for scientific data and software and are freely available on public repositories (GitHub, Mendeley data).

Even if FoodMicrobionet does not have the sophistication of QIITA, IMNGS and MGnify, we feel that this iteration, due to its size and diversity, provides a valuable asset for both the scientific community and industrial and regulatory stakeholders.


title: Close kin dyads indicate intergenerational dispersal and barriers

abstract: The movement of individuals through continuous space is typically constrained by dispersal ability and dispersal barriers. A range of approaches have been developed to investigate these. KO_SCPLOWINDISPERSEC_SCPLOW is a new approach that infers intergenerational dispersal ({sigma}) from close kin dyads, and appears particularly useful for investigating taxa that are difficult to observe individually. This study, focusing on the mosquito Aedes aegypti, shows how the same close kin data can also be used for barrier detection. We empirically demonstrate this new extension of the method using genome-wide sequence data from 266 Ae. aegypti. First, we use the spatial distribution of full-sib dyads collected within one generation to infer past movements of ovipositing female mosquitoes. These dyads indicated the relative barrier strengths of two roads, and performed favourably against alternative genetic methods for detecting barriers. The difference in variance between the sib and first cousin spatial distributions was used to infer movement over the past two generations, providing estimates of intergenerational dispersal ({sigma} = 81.5-197.1 m.gen-1/2) and density ({rho} = 833-4864 km-2). Dispersal estimates showed general agreement with those from mark-release-recapture studies. Barriers, {sigma}, {rho}, and neighbourhood size (331-526) can inform forthcoming releases of dengue-suppressing Wolbachia bacteria into this mosquito population.

category: genetics


title: Whole genome analysis sheds light on the genetic origin of Huns, Avars and conquering Hungarians

abstract: Huns, Avars and conquering Hungarians were Migration Period nomadic groups which arrived in three successive waves in the Carpathian Basin between the 5th and 9th centuries. Based on historical data each of these groups are thought to have arrived from Asia, although their exact origin and relation to other ancient and modern populations has been debated. In this study we have sequenced 9 Hun, 143 Avar and 113 Hungarian conquest period samples, and identified three core populations, representing immigrants from each period, with no recent European ancestry. Our results suggest that this "immigrant core" of both Huns and Avars originated in present day Mongolia, and their origin can be traced back to Xiongnus. On the other hand, the "immigrant core" of the conquering Hungarians derived from an earlier admixture of Mansis, early Sarmatians and descendants of late Xiongnus. In addition, we detected shared Hun-related ancestry in numerous Avar and Hungarian conquest period genetic outliers indicating a genetic link between these successive nomadic groups. Aside from the immigrant core groups we identified that the majority of the individuals from each period were local residents, harboring "native European" ancestry.


title: How a Formate Dehydrogenase Responds to Oxygen: Unexpected O2 Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and  Clusters.

abstract: The reversible two-electron interconversion of formate and CO2 is catalyzed by both non-metallo and metallo-formate dehydrogenases (FDHs). The latter group comprises molybdenum-or tungsten-containing enzymes with the metal coordinated by two equivalents of a pyranopterin cofactor, a cysteinyl or selenocysteinyl ligand supplied by the polypeptide, and a catalytically essential terminal sulfido ligand. In addition, these biocatalysts incorporate one or more [4Fe-4S] clusters for facilitating long-distance electron transfer. But an interesting dichotomy arises when attempting to understand how the metallo-FDHs react with O2. Whereas existing scholarship portrays these enzymes as being unable to perform in air due to extreme O2 lability of their metal centers, studies dating as far back as the 1930s emphasize that some of these systems exhibit formate oxidase (FOX) activity, coupling formate oxidation to O2 reduction. Therefore, to reconcile these conflicting views, we explored context-dependent functional linkages between metallo-FDHs and their cognate electron acceptors within the same organism vis-a-vis catalysis under atmospheric conditions. Here, we report the discovery and characterization of an O2-insensitive FDH2 from the sulfate-reducing bacterium Desulfovibiro vulgaris Hildenborough that ligates tungsten, selenocysteine, and four [4Fe-4S] clusters. Notably, we advance a robust expression platform for its recombinant production, eliminating both the requirement of nitrate or azide during purification and reductive activation with thiols and/or formate prior to catalysis. Because the distinctive spectral signatures of formate-reduced DvH-FDH2 remain invariant under anaerobic and aerobic conditions, we benchmarked the enzyme activity in air, identifying CO2 as the bona fide product of catalysis. Full reaction progress curve analysis uncovers a high catalytic efficiency when probed with an artificial electron acceptor pair. Furthermore, we show that DvH-FDH2 enables hydrogen peroxide production sans superoxide release to achieve O2 insensitivity. Direct electron transfer to cytochrome c in air also reveals that electron bifurcation is operational in this system. Taken together, our work unambiguously proves for the first time the coexistence of redox bifurcated FDH and FOX activities within a metallo-FDH scaffold. These findings have important implications for engineering O2-tolerant FDHs and bio-inspired artificial metallocatalysts, as well as for the development of authentic formate/air biofuel cells, modulation of catalytic bias, assessing the limits of reversible catalysis, understanding directional electron transfer, and discerning formate bioenergetics of gut microbiota.


title: Microtubule-Inspired Functionalization of Carbon Nanotubes: A Biomimetic Carrier Design

abstract: We propose a bioinspired, non-covalent carbon nanotubes (CNTs) functionalization strategy to augment their bioavailability and alleviate their biotoxicity. For functionalization, select amphiphilic peptides from a cytoskeletal biopolymer, microtubule (MT), were used. The peptides are involved in the MT polymerization by maintaining the essential lateral interactions among the MTs - and {beta}-tubulin subunits. They also participate in forming the MT-binding sites for hosting several MT-targeting antimitotics. Utilizing in silico methods, this study showed the peptides influenced CNTs diffusivity and aqueous solubility. The hydrodynamic shield formed by the peptides from {beta}-tubulin was more widespread on the CNT than the -tubulin peptides; however, the latter created a broader hydrophobic CNT coating than those from the {beta}-tubulin. In particular, the peptides consisting of the H1-B2, H10, H1-B2, and the M-loop, demonstrated structural features that serve to augment CNTs water solubility and dispersibility. The performance of the peptide-functionalized CNTs as drug carriers was examined by studying seventeen antimitotics. The CNT-peptides structural composition was identified as a suitable carrier for phomopsin A, laulimalide, epothilone A, epothilone D, discodermolide, eribulin, and docetaxel. The peptides played dual roles displaying affinities to the antimitotics and the CNT; in particular, the peptides from the H1-B2 and H2-B3 loops of {beta}-tubulin exhibited exceptional binding properties. Specific mutations on the wildtype peptides, including those from the -tubulin M-loop and H2-B3, or the {beta}-tubulin H1-B2, are proposed to refine their hydrophobicity, eliminate unfavorable inter-peptides electrostatic interactions or the spatial hindrance at certain regions, to enhance their conformational steadiness and exposure to the tube surface. A combination of the select amphiphilic peptides from both tubulin subunits is suggested to improve CNTs bioavailability and efficiency for carrying insoluble hydrophobic cargos.


title: Exo-Enzymatic Cell-Surface Glycan Labeling for Capturing Glycan-Protein Interactions through Photo-Crosslinking

abstract: Tools to interrogate glycoconjugate-protein interactions in the context living cells are highly attractive for the identification of critically important functional binding partners of glycan-binding proteins. These interactions are challenging to interrogate due to low affinity and rapid dissociation rates of glycan-protein binding events. The use of photo-crosslinkers to capture glycan-protein interaction complexes has shown great promise for identifying binding partners involved in these interactions. Current methodologies use metabolic oligosaccharide engineering (MOE) to incorporate photo-crosslinking sugars. However, these MOE strategies are not amenable to all cell types and can result in low incorporation and cell-surface display of the photo-crosslinking probe, limiting their utility for studying many types of interactions. We describe here an exo-enzymatic strategy for selectively introducing photo-crosslinking probes into cell-surface glycoconjugates using the recombinant human sialyltransferase ST6GAL1 and a diazirine-linked CMP-Neu5Ac derivative. Probe introduction is highly efficient, amenable to different cell types and resulted in improved crosslinking when compared to MOE. This exo-enzymatic labeling approach can selectively introduce the photo-crosslinking sugar on to specific glycan epitopes and subclasses by harnessing the specificity of the sialyltransferase employed, underscoring its potential as a tool to interrogate and identify glycoconjugate ligands for diverse glycan-binding proteins.


title: Spatially resolved profiling of protein conformation and interactions by biocompatible chemical cross-linking in living cells

abstract: The protein structures and interactions that maintain and regulate cellular processes in different subcellular organelles are heterogeneous and dynamic. However, it remains challenging to characterize the subcellular specificity and translocation of protein complexes in terms of conformation and interactions. Herein, we developed a spatially resolved protein complex profiling approach by biocompatible chemical cross-linking in living cells (SPACX) to monitor the dynamics of protein conformation, interactions and translocation. The advancement of fast capturing protein complexes in the physiological state, coupled with efficient enrichment of the cross-linked peptides, ensured deep-coverage analysis of the protein interactome in living cells. By ensemble structure refinement with cross-linking restraints, subcellular-specific conformation heterogeneity was identified for PTEN. PTEN displayed a broader range of dynamic conformation changes on the dual specificity domains in the nucleus than in the cytoplasm. Moreover, based on conformational differences, different interacting assemblies involving 25 cytoplasm-exclusively and 18 nucleus-exclusively PTEN-interacting proteins were found to account for diverse biological functions. Upon ubiquitin-proteasome system (UPS) stress, the assembly of PTEN and its interacting partners changed obviously during translocation. We newly identified 36 PTEN-interacting proteins, which were found to be highly enriched in functional pathways closely related to cell apoptosis. Inspiringly, the interactions among PTEN isoforms and their interacting proteins were accessible by the determination of sequence-unique cross-linking interfaces for direct interactions. All these results indicate the promise of SPACX to elucidate the functional heterogeneity of proteins in individual subcellular sociology.


title: Targeting SARS-CoV-2 infection through CAR T cells and bispecific T cell engagers

abstract: Despite advances in antibody treatments and vaccines, COVID-19 caused by SARS-CoV-2 infection remains a major health problem resulting in excessive morbidity and mortality and the emergence of new variants has reduced the effectiveness of current vaccines. Here, as a proof-of-concept we engineered primary CD8 T cells to express SARS-CoV-2 Spike protein-specific CARs, using extracellular region of ACE2, and demonstrated their highly specific and potent cytotoxicity towards Spike-expressing target cells. To improve on this concept as a potential therapeutic, we developed a bispecific T cell engager combining ACE2 with an anti-CD3 scFv (ACE2-Bite) to target infected cells and the virus. Similar to CAR-T cell approach, ACE2-Bite endowed cytotoxic cells to selectively kill Spike-expressing targets. Furthermore, ACE2-Bite neutralized the pseudoviruses of SARS-CoV, SARS-CoV-2 wild-type and variants including Delta and Omicron, as a decoy protein. Remarkably, ACE2-Bite molecule showed a higher binding and neutralization affinity to Delta and Omicron variants compared to SARS-CoV-2 wild-type Spike proteins, suggesting the potential of this approach as a variant-proof, therapeutic strategy for future SARS-CoV-2 variants, employing both humoral and cellular arms of the adaptive immune response.

category: bioengineering


title: Targeting SARS-CoV-2 infection through CAR-T like bispecific T cell engagers incorporating ACE2


title: Controlled adhesion, membrane pinning and vesicle transport by Janus particles

abstract: The interactions between biomembranes and particles are key to many applications, but the lack of controllable model systems to study them limits the progress in their research. Here, we describe how Janus polystyrene microparticles, half coated with iron, can be partially engulfed by artificial cells, namely giant vesicles, with the goals to control and investigate their adhesion and degree of encapsulation. The interaction between the Janus particles and these model cell membrane systems is mediated by electrostatic charge, offering a further mode of modulation in addition to the iron patches. The ferromagnetic particle coatings also enable the ability to manipulate and transport the vesicles by magnetic fields.


title: Unveiling molecular interactions that stabilize bacterial adhesion pili

abstract: Adhesion pili assembled by the chaperone-usher pathway are superelastic helical filaments on the surface of bacteria, optimized for attachment to target cells. Here, we investigate the biophysical function and structural interactions that stabilize P pili from uropathogenic bacteria. Using optical tweezers we measure P pilus subunit-subunit interaction dynamics and show that pilus compliance is contour-length dependent. Atomic details of subunit-subunit interactions of pili under tension are shown using steered molecular dynamics (sMD) simulations. sMD results also indicate that the N-terminal "staple" region of P pili significantly stabilizes the helical filament structure, consistent with previous structural data, suggesting more layer-to-layer interactions could compensate for the lack of a staple in Type 1 pili. This study informs our understanding of essential structural and dynamic features of adhesion pili, supporting the hypothesis that the biophysical function of pili is niche-adapted rather than a direct consequence of genetic similarity or diversity.


title: Septin-Mediated Mechanobiological Reprogramming of T Cell Transmigration and 3D Motility

abstract:
T cells migrate in nearly every healthy, inflamed or diseased tissue. Such  all-terrain motility is achieved by a dynamic mechanobiological balance between amoeboid and mesenchymal-like migration modes. Here, we report that septin proteins function as a key regulator of migratory balance in T cells. We show that active septins compartmentalize the lymphocytes cortex into a peristaltically treadmilling  tube during avoidance response to mechanically crowding hindrances. T cell cortical peristaltism mechanically channels nucleus and cytoplasm between mechanically crowding 3D collagen fibers. Septins inactivation shifts T cell motility balance towards mesenchymal-like mode, characterized by distinct contact guidance and MAP4-, SEPT9-, HDAC6-mediated enhancement of microtubules and microtubule-associated dynein contractility. The non-stretchable microtubular cables secure structurally coherent cell passage through confining spaces and long-distance transmission of dynein-generated forces, which replace diminished actomyosin contractility. Thus, septins provide T cells with a structural and signaling molecular switch between actomyosin-driven amoeboid and dynein-driven mesenchymal-like migration.

GLOSSARYO_LISteric interactions - objects interactions by the means of their spatial collision, determined by objects shapes.
C_LIO_LISteric guidance - T cell navigation within crowded 3D environments, determined by the available passages around and between steric hindrances.
C_LIO_LIPeristaltic treadmilling - locomotion mode by the means of repeated series of polarized cell cortex extension, stabilization, and retraction, accompanied by translocation of nucleus and cytoplasm via circumferential cortex contractility.
C_LIO_LIDynein contractility - contractile forces within a system of non-stretchable, force-transmitting microtubular cables, generated by dynein motor proteins.
C_LI


title: The CRISPR-Cas System differentially regulates surface-attached and pellicle-biofilm in Salmonella enterica serovar Typhimurium

abstract: CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR associated) system has been studied for its role in biofilm regulation and expression of outer membrane proteins in Salmonella. We investigated the CRISPR-Cas mediated biofilm regulation in Salmonella enterica serovar Typhimurium by deleting CRISPR-Cas components,{Delta} crisprI,{Delta} crisprII, {Delta}{Delta}crisprI crisprII, and{Delta} cas op. We determined that the system positively regulates surface-biofilm while inhibiting pellicle-biofilm. In knockout strains, flagellar (fliC, flgK) and curli (csgA) genes were repressed, causing reduced surface-biofilm. Conversely, they displayed altered pellicle-biofilm architecture possessing bacterial multilayers and a denser ECM with enhanced cellulose and lesser Curli, ergo weaker pellicle. The intracellular cellulose concentration was less in the knockout strains due to upregulation of bcsC, necessary for cellulose export. We hypothesized that exported cellulose integrates into the pellicle. We determined that crp is upregulated in the knockout strains, thereby inhibiting the expression of csgD, hence csgA and bcsA. The conflicting upregulation of bcsC, the last gene of bcsABZC operon, could be independently regulated by the CRISPR-Cas system owing to a partial match between the CRISPR-spacers and bcsC gene. The CRP-mediated regulation of the flagellar genes in the knockout strains was probably circumvented through the regulation of Yddx governing the availability of {sigma}28 factor that further regulates class3 flagellar genes (fliC, flgK). Additionally, the variations in the LPS profile and expression of LPS-related genes (rfaC, rfbG, rfbI) in the knockout strains could also contribute to the altered pellicle architecture. Collectively, we establish that the CRISPR-Cas system differentially regulates the formation of surface-attached and pellicle-biofilm.


title: First detection of SARS-CoV-2 infection in Canadian wildlife identified in free-ranging white-tailed deer (Odocoileus virginianus) from southern Quebec, Canada

abstract: White-tailed deer are susceptible to SARS-CoV-2 and represent a relevant species for surveillance. We investigated SARS-CoV-2 infection in white-tailed deer in Quebec, Canada. In November 2021, 251 nasal swabs and 104 retropharyngeal lymph nodes from 258 deer were analyzed for SARS-CoV-2 RNA, whole genome sequencing and virus isolation and 251 thoracic cavity fluid samples were tested for neutralizing antibodies. We detected SARS-CoV-2 RNA in three nasal swabs from the Estrie region and virus was isolated from two samples; evidence of past exposure was detected among deer from the same region. Viral sequences were assigned to lineage AY.44, a sublineage of B.1.617.2. All deer sequences clustered with human GISAID sequences collected in October 2021 from Vermont USA, which borders the Estrie region. Mutations in the S-gene and a deletion in ORF8 encoding a truncated protein were detected. These findings underscore the importance of ongoing surveillance of key wildlife species for SARS-CoV-2.


title: High-throughput suppressor screens unveil functional convergence of single-gene lysis proteins

abstract: In contrast to canonical dsDNA phages, lysis in ssRNA Fiersviridae and ssDNA Microviridae phages is encoded by a minimal single gene (sgl), to meet the size constraints of some of the smallest genomes on earth. To achieve lysis, Sgl proteins exploit evolutionary "weak spots" in the bacterial cell wall by inhibiting specific steps in cell wall synthesis. Although a handful of these proteins have been characterized, the potentially diverse "weak spots" targeted by most Sgls remains enigmatic. Here, we repurpose Dub-seq for genome-wide assessment of host suppressors of Sgl activity and apply to eight diverse Sgls awaiting molecular target characterization. In addition to known molecular mechanisms, we discover a complex network of suppressors across the spectrum of genes involved in cell wall biogenesis. We highlight the first case of Sgl convergent evolution between unrelated proteins by determining the molecular target of SglPP7 as MurJ, suggesting the existence of universal "weak spots" in the bacterial cell wall.


title: Pathogenicity of SARS-CoV-2 Omicron in Syrian hamsters and its neutralization with different Variants of Concern

abstract: SARS-CoV-2 Omicron variant is rampantly spreading across the globe. Animal models are useful in understanding the disease characteristics as well as properties of emerging SARS-CoV-2 variants. We assessed the pathogenicity and immune response generated by BA.1 sub-lineage of SARS-CoV-2 Omicron variant with R346K mutation in 5 to 6-week old Syrian hamsters. Virus shedding, organ viral load, lung disease and immune response generated were sequentially assessed. The disease characteristics of Omicron were found to be similar to that of other SARS-CoV-2 variants of concerns in hamsters like high viral replication in the respiratory tract and interstitial pneumonia. The infected hamsters demonstrated lesser body weight gain in comparison to the uninfected control hamsters. Viral RNA could be detected in nasal washes and respiratory organs (nasal turbinate, trachea, bronchi and lungs) till 10 and 14 days respectively. The clearance of the virus was observed from nasal washes and lungs by day 7. Neutralizing antibody response against Omicron variant was detected from day 5 with rising antibody titers till 14 days. However, the cross-neutralization titre of the sera against other variants showed severe reduction ie., 7 fold reduction against Alpha and no titers against B.1, Beta and Delta. This preliminary data shows that Omicron variant infection can produce moderate to severe lung disease and the neutralizing antibodies produced in response to Omicron variant infection shows poor neutralizing ability against other co-circulating SARS-CoV-2 variants like Delta which necessitates caution as it may lead to increased cases of reinfection.


title: Intra-granuloma accumulation and inflammatory differentiation of neutrophils underlie mycobacterial ESX-1-mediated immunopathology

abstract:
The conserved ESX-1 type VII secretion system is a major virulence determinant of pathogenic mycobacteria, including Mycobacterium tuberculosis and Mycobacterium marinum. ESX-1 is known to interact with infected macrophages, but its potential roles in regulating other host cells and immunopathology have remained largely unexplored. Using a murine M. marinum infection model we identify neutrophils and Ly6C+MHCII+ monocytes as the main cellular reservoirs for the bacteria. We show that ESX-1 promotes intra-granuloma accumulation of neutrophils and that neutrophils have a previously unrecognized required role in executing ESX-1-mediated pathology. To explore if ESX-1 also regulates the function of recruited neutrophils we performed single cell RNA-seq analysis indicating that ESX-1 drives newly recruited uninfected neutrophils into an inflammatory phenotype via an extrinsic mechanism. In contrast, monocytes restricted the accumulation of neutrophils and immunopathology, demonstrating a major host-protective function for monocytes specifically by suppressing ESX-1-mediated neutrophilic inflammation. iNOS activity was required for the suppressive mechanism and we identified Ly6C+MHCII+ monocytes as the main iNOS-expressing cell type in the infected tissue. These results suggest that ESX-1 mediates immunopathology by driving neutrophil accumulation and phenotypic differentiation in the infected tissue, and they demonstrate an antagonistic interplay between monocytes and neutrophils by which monocytes suppress host-detrimental neutrophilic inflammation.

Author SummaryThe ESX-1 type VII secretion system is required for virulence of pathogenic mycobacteria, including Mycobacterium tuberculosis. ESX-1 interacts with infected macrophages, but its potential roles in regulating other host cells and immunopathology have remained largely unexplored. We demonstrate that ESX-1 promotes immunopathology by driving intra-granuloma accumulation of neutrophils, which upon arrival adopt an inflammatory phenotype in an ESX-1-dependent manner. In contrast, monocytes limited the accumulation of neutrophils and neutrophil-mediated pathology via an iNOS-dependent mechanism, suggesting a major host-protective function for monocytes specifically by restricting ESX-1-mediated neutrophilic inflammation. These findings provide insight into how ESX-1 promotes disease, and they reveal an antagonistic functional relationship between monocytes and neutrophils that may regulate immunopathology not only in mycobacterial infection, but also in other infections as well as in inflammatory conditions and cancer.