title: Evidence of increased Cathepsin B/L and decreased TMPRSS2 usage for cell entry by the SARS-CoV-2 Omicron variant

abstract: The SARS-CoV-2 Omicron variant harbors a large number of mutations in its spike protein compared to the original SARS-CoV-2 strain, which may alter its ability to enter cells. To quantify this effect, we analysed recent datasets on SARS-CoV-2 pseudovirus entry into cells in vitro using a mathematical model. SARS-CoV-2 can enter cells via two pathways, one using the host proteases Cathepsin B/L and the other using the host protease TMPRSS2. We found enhanced entry efficiency of the Omicron variant in cells where the original strain preferentially used Cathepsin B/L and reduced efficiency where it used TMPRSS2, indicating that the Omicron variant had evolved to use the Cathepsin B/L pathway better but at the expense of its ability to use the TMPRSS2 pathway for entry. Applying our model, we estimated >4-fold enhanced efficiency of the Omicron variant in entry via the Cathepsin B/L pathway and >3-fold reduced efficiency in entry via the TMPRSS2 pathway compared to the original or other strains. This differential entry pathway usage may explain the altered cell tropism of the Omicron variant compared to other strains and have implications for its infectivity, transmissibility, and interventions.


title: Trypanosoma brucei: A novel system for metacyclogenesis and complete cyclical development in vitro

abstract: We describe an in vitro system that allows metacyclogenesis of T. brucei ssp. and is not restricted to certain parasite lines. Following the work of Hirumi and co-workers we developed a modified in vitro system which allows metacyclogenesis within 7 to 20 days. The most striking changes were the addition of citrate and cis-aconitate, which triggers transformation from bloodstream to procyclic forms, and the elimination of the remaining bloodstream forms by salicylhydroxamic acid and glycerol. A positive feature of this system is that T. b. gambiense samples isolated from patients, and transformed to procyclic forms for transport, could give rise to infective forms that could be passaged in rodents.

category: microbiology


title: Mucociliary Transport Deficiency and Disease Progression in Syrian Hamsters with SARS-CoV-2 Infection

abstract: Substantial clinical evidence supports the notion that ciliary function in the airways plays an important role in COVID-19 pathogenesis. Although ciliary damage has been observed in both in vitro and in vivo models, consequent impaired mucociliary transport (MCT) remains unknown for the intact MCT apparatus from an in vivo model of disease. Using golden Syrian hamsters, a common animal model that recapitulates human COVID-19, we quantitatively followed the time course of physiological, virological, and pathological changes upon SARS-CoV-2 infection, as well as the deficiency of the MCT apparatus using micro-optical coherence tomography, a novel method to visualize and simultaneously quantitate multiple aspects of the functional microanatomy of intact airways. Corresponding to progressive weight loss up to 7 days post-infection (dpi), viral detection and histopathological analysis in both the trachea and lung revealed steadily descending infection from the upper airways, as the main target of viral invasion, to lower airways and parenchymal lung, which are likely injured through indirect mechanisms. SARS-CoV-2 infection caused a 67% decrease in MCT rate as early as 2 dpi, largely due to diminished motile ciliation coverage, but not airway surface liquid depth, periciliary liquid depth, or cilia beat frequency of residual motile cilia. Further analysis indicated that the fewer motile cilia combined with abnormal ciliary motion of residual cilia contributed to the delayed MCT. The time course of physiological, virological, and pathological progression suggest that functional deficits of the MCT apparatus predispose to COVID-19 pathogenesis by extending viral retention and may be a risk factor for secondary infection. As a consequence, therapies directed towards the MCT apparatus deserve further investigation as a treatment modality.


title: A genome-wide screen identifies SCAI as a modulator of the UV-induced replicative stress response in human cells

abstract: Helix-destabilizing DNA lesions induced by environmental mutagens such as UV light cause genomic instability by strongly blocking the progression of DNA replication forks (RF). At blocked RF, single-stranded DNA (ssDNA) accumulates and is rapidly bound by Replication Protein A (RPA) complexes. Such stretches of RPA-ssDNA constitute platforms for recruitment/activation of critical factors that promote DNA synthesis restart. However, during periods of severe replicative stress, RPA availability may become limiting due to inordinate sequestration of this multifunctional complex on ssDNA, thereby negatively impacting multiple vital RPA-dependent processes. Here, we performed a genome-wide screen to identify factors which restrict the accumulation of RPA-ssDNA during UV-induced replicative stress. While this approach revealed some expected "hits" acting in pathways such as nucleotide excision repair, translesion DNA synthesis, and the intra-S phase checkpoint, it also identifed SCAI, whose role in the replicative stress response was previously unappreciated. Upon UV exposure, SCAI knock-down caused elevated accumulation of RPA-ssDNA during S phase, accompanied by reduced cell survival and compromised RF progression. These effects were independent of the previously reported role of SCAI in 53BP1-dependent DNA double-strand break repair. We also found that SCAI colocalized with stalled RF, and that its depletion promoted nascent DNA degradation. Finally, we (i) provide evidence that EXO1 is the major nuclease underlying ssDNA formation and consequent DNA replication defects in SCAI knockout cells and, consistent with this, (ii) demonstrate that SCAI inhibits EXO1 activity on a ssDNA gap in vitro. Taken together, our data establish SCAI as a novel regulator of the replicative stress response in human cells.

category: molecular biology


title: xia2.multiplex: a multi-crystal data analysis pipeline

abstract:
In macromolecular crystallography radiation damage limits the amount of data that can be collected from a single crystal. It is often necessary to merge data sets from multiple crystals, for example small-wedge data collections on micro-crystals, in situ room-temperature data collections, and collection from membrane proteins in lipidic mesophase. Whilst indexing and integration of individual data sets may be relatively straightforward with existing software, merging multiple data sets from small wedges presents new challenges. Identification of a consensus symmetry can be problematic, particularly in the presence of a potential indexing ambiguity. Furthermore, the presence of non-isomorphous or poor-quality data sets may reduce the overall quality of the final merged data set.

To facilitate and help optimise the scaling and merging of multiple data sets, we developed a new program, xia2.multiplex, which takes data sets individually integrated with DIALS and performs symmetry analysis, scaling and merging of multicrystal data sets. xia2.multiplex also performs analysis of various pathologies that typically affect multi-crystal data sets, including non-isomorphism, radiation damage and preferential orientation. After describing a number of use cases, we demonstrate the benefit of xia2.multiplex within a wider autoprocessing framework in facilitating a multi-crystal experiment collected as part of in situ room-temperature fragment screening experiments on the SARS-CoV-2 main protease.


title: Parallel evolution of amphioxus and vertebrate small-scale gene duplications

abstract:
BackgroundAmphioxus are non-vertebrate chordates characterized by a slow morphological and molecular evolution. They share the basic chordate body-plan and genome organization with vertebrates but lack their "2R" whole-genome duplications and their developmental complexity. For these reasons, amphioxus are frequently used as an outgroup to study vertebrate genome evolution and Evo-Devo. Aside from whole-genome duplications, genes continuously duplicate on a smaller scale. Small-scale duplicated genes can be found in both amphioxus and vertebrate genomes, while only the vertebrate genome has duplicated genes product of their 2R whole-genome duplications. Here, we explore the history of small-scale gene duplications in the amphioxus lineage and compare it to small- and large-scale gene duplication history in vertebrates.

ResultsWe present a study of the European amphioxus (Branchiostoma lanceolatum) gene duplications thanks to a new, high-quality genome reference. We find that, despite its overall slow molecular evolution, the amphioxus lineage has had a history of small-scale duplications similar to the one observed in vertebrates. We find parallel gene duplication profiles between amphioxus and vertebrates, and conserved functional constraints in gene duplication. Moreover, amphioxus gene duplicates show levels of expression and patterns of functional specialization similar to the ones observed in vertebrate duplicated genes. We also find strong conservation of gene synteny between two distant amphioxus species, B. lanceolatum and B. floridae, with two major chromosomal rearrangements.

ConclusionsIn contrast to their slower molecular and morphological evolution, amphioxus small-scale gene duplication history resembles that of the vertebrate lineage both in quantitative and in functional terms.

category: evolutionary biology


title: Super-silencers regulated by chromatin interactions control apoptotic genes

abstract: Human silencers have been shown to exist and regulate developmental gene expression. However, the functional importance of human silencers needs to be elucidated such as the working mechanism and whether they can form "super-silencers". Here, through interrogating two putative silencer components of FGF18 gene, we found that two silencers can cooperate via compensated chromatin interactions to form a "super-silencer". Furthermore, double knock-out of two silencers exhibited synergistic upregulation of FGF18 expression and changes of cell identity. To disturb the "super-silencers", we applied combinational treatment of an EZH2 inhibitor GSK343, and a REST inhibitor, X5050 ("GR"). We found that GR led to severe loss of TADs and loops, while the use of just one inhibitor by itself only showed mild changes. Such changes of TADs and loops may due to reduced CTCF protein level observed upon GR treatment. Moreover, GSK343 and X5050 worked together synergistically to upregulate the apoptotic genes controlled by super-silencers, and thus gave rise to antitumor effects including apoptosis, cell cycle arrest and tumor growth inhibition. Overall, our data demonstrated the first example of a "super-silencer" and showed that combinational usage of GSK343 and X5050 could potentially lead to cancer ablation through disruption of "super-silencers".

category: genomics


title: How Glutamate Promotes Liquid-liquid Phase Separation and DNA Binding Cooperativity of E. coli SSB Protein

abstract: E. coli single-stranded-DNA binding protein (EcSSB) displays nearest-neighbor (NN) and non-nearest-neighbor (NNN)) cooperativity in binding ssDNA during genome maintenance. NNN cooperativity requires the intrinsically-disordered linkers (IDL) of the C-terminal tails. Potassium glutamate (KGlu), the primary E. coli salt, promotes NNN-cooperativity, while KCl inhibits it. We find that KGlu promotes compaction of a single polymeric SSB-coated ssDNA beyond what occurs in KCl, indicating a link of compaction to NNN-cooperativity. EcSSB also undergoes liquid-liquid phase separation (LLPS), inhibited by ssDNA binding. We find that LLPS, like NNN-cooperativity, is promoted by increasing [KGlu] in the physiological range, while increasing [KCl] and/or deletion of the IDL eliminate LLPS, indicating similar interactions in both processes. From quantitative determinations of interactions of KGlu and KCl with protein model compounds, we deduce that the opposing effects of KGlu and KCl on SSB LLPS and cooperativity arise from their opposite interactions with amide groups. KGlu interacts unfavorably with the backbone (especially Gly) and side chain amide groups of the IDL and therefore promotes amide-amide interactions in LLPS and NNN-cooperativity. By contrast, KCl interacts favorably with these amide groups and therefore inhibits LLPS and NNN-cooperativity. These results highlight the importance of salt interactions in regulating the propensity of proteins to undergo LLPS.

category: biochemistry


title: MicroRNA Detection in Biological Media Using a Split Aptamer Platform

abstract: Intercellular microRNA (miRNA)-based communication has been implicated in a wide array of functional and dysfunctional biological processes. This has raised attention to the potential use of miRNAs as biomarkers for disease diagnosis and prognosis and produced interest in their detection. Though the list of clinically significant miRNA biomarkers is rapidly expanding, it remains challenging to adapt current tools to investigate new targets in biological environments. Systematic approaches for the rapid development of miRNA biosensors are valuable to reduce this disparity. We describe here a methodology for developing aptamer-based fluorescent biosensors that can specifically detect miRNAs in biological environments, including culture medium from HeLa cells, human serum, and human plasma. This methodology includes the semi-rational design of the hybridization between a pair of split DNA aptamer oligonucleotides and the miRNA target to build a pool of potential sensor designs, and the screening of this pool for designs with high signal-to-background ratio and sequence selectivity. The method uses natural oligonucleotides without chemical modification, and is effective in buffer, 10%, and 30% (v/v) biological media. Following this approach, we developed sensors that detect three miRNA targets (miR-19b, miR-21, and miR-92a) at concentrations as low as 5 nM without amplification and are selective against single-nucleotide mutants. This work expands upon the current design principles of nucleic acid-based biosensors and provides a method to rapidly develop diagnostic tools for novel and niche miRNA targets of interest.


title: De-tensioning of collagen fibers optimizes endometrial receptivity and improves the rate of embryo implantation

abstract:
Successful embryo implantation is associated with intensive endometrium matrix remodeling, in which fibrillar collagens and matrix metalloproteinases play a key role. Here we report that topical treatment with a single dose of human collagenase-1 improves the rate of embryo implantation in murine models regardless of genetic background. Collagenase-1 treatment reduces endometrial collagen-fiber tension affecting their aligned configuration while preserving the overall integrity of the uterine tissue. The collagenase treatment causes enhanced local vascular permeability and creates an inflammatory-like environment, all of which are necessary for successful embryo implantation. We show that this specific treatment rescued heat stress and embryo transfer induced embryo failure in mice. Our findings highlight the clinical feasibility to improve endometrial receptivity for embryo implantation. This treatment approach may be used to improve livestock breeding and for assisted human reproduction medical treatments.

TeaserMild specific collagen remodeling improves endometrial receptivity for natural and embryo- transfer-derived embryo implantation.


title: Ultra high diversity factorizable libraries for efficient therapeutic discovery

abstract: The successful discovery of novel biological therapeutics by selection requires highly diverse libraries of candidate sequences that contain a high proportion of desirable candidates. Here we propose the use of computationally designed factorizable libraries made of concatenated segment libraries as a method of creating large libraries that meet an objective function at low cost. We show that factorizable libraries can be designed efficiently by representing objective functions that describe sequence optimality as an inner product of feature vectors, which we use to design an optimization method we call Stochastically Annealed Product Spaces (SAPS). We then use this approach to design diverse and efficient libraries of antibody CDR-H3 sequences with various optimized characteristics.

category: bioengineering


title: Capturing non-local through-bond effects in molecular mechanics force fields: II. Using fractional bond orders to fit torsion parameters

abstract: Accurate small molecule force fields are crucial for predicting thermodynamic and kinetic properties of drug-like molecules in biomolecular systems. Torsion parameters, in particular, are essential for determining conformational distribution of molecules. However, they are usually fit to computationally expensive quantum chemical torsion scans and generalize poorly to different chemical environments. Torsion parameters should ideally capture local through-space non-bonded interactions such as 1-4 steric and electrostatics and non-local through-bond effects such as conjugation and hyperconjugation. Non-local through-bond effects are sensitive to remote substituents and are a contributing factor to torsion parameters poor transferability. Here we show that fractional bond orders such as the Wiberg Bond Order (WBO) are sensitive to remote substituents and correctly captures extent of conjugation and hyperconjugation. We show that the relationship between WBO and torsion barrier heights are linear and can therefore serve as a surrogate to QC torsion barriers, and to interpolate torsion force constants. Using this approach we can reduce the number of computationally expensive QC torsion scans needed while maintaining accurate torsion parameters. We demonstrate this approach to a set of substituted benzene rings.

category: biophysics


title: AXL-TBK1 driven nuclear AKT3 promotes metastasis

abstract:
Epithelial-to-mesenchymal transition (EMT) contributes to tumor cell survival, immune evasion, migration, invasion, and therapy resistance. Across human cancer, tumors that are high grade, poorly differentiated, and have undergone EMT carry a worse prognosis with a higher likelihood of metastasis. AXL, a receptor tyrosine kinase, drives EMT and is implicated in tumor progression, metastasis, and therapy resistance in multiple cancer types including pancreatic cancer and breast cancer. TANK-binding kinase 1 (TBK1) is central to AXL-driven EMT yet, the mechanism of how TBK1 induces EMT remains unclear. Here, we report that AXL activation stimulates TBK1 binding and phosphorylation of AKT3. TBK1 activation of AKT3 drives binding and phosphorylation of slug/snail resulting in protection from proteasomal degradation and translocation of the complex into the nucleus. We show that nuclear translocation of AKT3 is required for AXL-driven EMT and metastasis. Congruently, nuclear AKT3 expression correlates with worse outcome in aggressive breast cancer. To advance AKT3 as a therapeutic target, an AKT3-isoform selective allosteric small molecule inhibitor, BGB214, was developed. BGB214 inhibits AKT3 nuclear translocation, EMT-TF stability, AKT3-mediated invasion of breast cancer cells and reduces tumor initiation in vivo. Our results suggest that AKT3 nuclear activity is an important feature of AXL-driven epithelial plasticity and that selective AKT3 inhibition represents a novel therapeutic avenue for treating aggressive cancer.

SignificanceNuclear AKT3 activity is an important feature of AXL-TBK1 driven EMT and metastasis, thus selective AKT3 targeting represents a novel approach to treat aggressive cancer.

category: cancer biology


title: Dual Targeting of FLT3 and LSD1 Disrupts the MYC Super-Enhancer Complex in Acute Myeloid Leukemia

abstract: Clinical responses to kinase inhibitor therapy in acute myeloid leukemia (AML) are limited by the inevitable development of resistance. A major contributor to resistance is early epigenetic adaptation, leading to persistence of a small number of leukemia cells. Here we show that inhibition of the epigenetic regulator lysine-specific demethylase 1 (LSD1) augments the response to inhibitors of the FLT3 kinase in AML. We demonstrate that combined FLT3 and LSD1 inhibition results in synergistic cell death of FLT3-mutant AML cells. The drug combination activates a pro-differentiative epigenetic and transcriptional program while simultaneously suppressing the activity of MYC target genes. High-resolution, multi-modal epigenetic analyses revealed that combined FLT3 and LSD1 inhibition results in the suppression of MYC-bound promoters and the activation of PU.1-bound enhancers. Regulon enrichment analysis in primary AML samples nominated STAT5 as a putative regulator of MYC gene expression. Inhibition of FLT3 results in a loss of STAT5 binding at the MYC blood super-enhancer and a loss of super-enhancer activation. In contrast, LSD1 inhibition prevents the removal of repressive H3K9me1 marks at MYC target genes, resulting in suppression of MYC expression. We validated these findings in 66 primary AML samples including 17 FLT3-ITD-positive AML samples, with the majority demonstrating improved responses with the drug combination. High MYC regulon activity was a predictor of response to the drug combination and RNA-seq on drug-treated AML samples revealed suppression of MYC target genes. Finally, single cell ATAC-seq on primary AML blasts treated ex vivo with the FLT3 and LSD1 inhibitor combination results in a shift from MYC super-enhancer-high to a MYC super-enhancer-low cell state. Collectively, these studies provide preclinical rationale for the investigation of dual FLT3 and LSD1 inhibition in a clinical trial.


title: Activation of the Integrated Stress Response overcomes multidrug resistance in FBXW7-deficient cells

abstract: FBXW7 is one of the most frequently mutated tumor suppressors, the deficiency of which has been associated with resistance to some anticancer therapies. Through bioinformatic analyses and genome-wide CRISPR screens, we here reveal that FBXW7 deficiency leads to multidrug resistance (MDR), to a bigger extent than well-established MDR-drivers such as overexpression of the drug-efflux pump ABCB1. Proteomic data from FBXW7-deficient cancer cells identify the upregulation of mitochondrial function as a hallmark of FBXW7 deficiency, which has been previously linked to an increased resistance to chemotherapy. Accordingly, genetic or chemical targeting of mitochondria is preferentially toxic for FBXW7-deficient cells in vitro and in vivo. Mechanistically, we show that the toxicity associated with therapies that target mitochondrial translation such as the antibiotic tigecycline relates to the activation of the Integrated Stress Response (ISR). Furthermore, while searching for additional drugs that could overcome the MDR of FBXW7-deficient cells, we found that all of them unexpectedly also activated the ISR regardless of their currently accepted mechanism of action. Together, our study reveals that one of the most frequent mutations in cancer reduces the sensitivity to the vast majority of available therapies, and identifies a general principle to overcome such resistance.


title: Single cell profiling of γδ hepatosplenic T-cell lymphoma unravels tumor cell heterogeneity associated with disease progression

abstract: Hepatosplenic T-cell lymphoma (HSTCL), mostly derived from {gamma}{delta} T cells, is a rare but very aggressive lymphoma with poor outcomes. The molecular pathogenesis driving HSTCL is largely unknown while only limited treatment options are available. In this study, by performing paired single cell RNA-seq and T cell receptor (TCR) sequencing on biopsies from a HSTCL patient pre- and post-chemotherapy treatments, we characterized unique gene expressing signatures of malignant {gamma}{delta} T cells, with a set of marker genes were newly identified in HSTCL (AREG, PLEKHA5, VCAM1 etc.). Although the malignant {gamma}{delta} T cells were expanded from a single TCR clonotype according to their TCR identities, they evolved into two transcriptional distinct tumor subtypes during the disease progression. The Tumor_1 subtype was dominant in pre-treatment samples with highly aggressive phenotypes. While the Tumor_2 had relative mild cancer hallmark signatures but expressed genes associated with tumor survival signal and drug resistance (IL32, TOX2, AIF1, AKAP12 etc.), and finally became the main tumor subtype post-treatment. We further dissected the tumor microenvironment of the HSTCL and noticed that CD8 memory T cells were clonal expanded post-treatment. In addition, we discovered dynamically rewiring cell-cell interaction networks during the treatment. The tumor cells had reduced communications with the microenvironment post-treatment. Our study reveals heterogenous and dynamic tumor and microenvironment underlying pathogenesis of HSTCL and may contribute to identify novel targets for diagnosis and cure of HSTCL in the future.


title: SUMO/deSUMOylation of the BRI1 brassinosteroid receptor modulates plant growth responses to temperature

abstract:
Brassinosteroids (BRs) are a class of steroid molecules perceived at the cell surface that act as plant hormones. The BR receptor BRI1 offers a model to understand receptor-mediated signaling in plants and the role of post-translational modifications. Here we identify SUMOylation as a new modification, targeting BRI1 to regulate its activity. BRI1 is SUMOylated in planta on two lysine residues and the levels of BRI1-SUMO conjugates are controlled by the Desi3a SUMO protease. We demonstrate that BRI1 is deSUMOylated at elevated temperature by Desi3a, leading to increased BRI1 interaction with the negative regulator of BR signaling BIK1 and enhancing BRI1 endocytosis. Loss of Desi3a or BIK1 results in increased response to temperature elevation, indicating that BRI1 deSUMOylation acts as a safety mechanism necessary to keep temperature responses in check. Altogether, our work establishes BRI1 deSUMOylation as a molecular crosstalk mechanism between temperature and BR signaling, allowing plants to translate environmental inputs into growth response.

SIGNIFICANCE STATEMENTThe brassinosteroid (BR) receptor BRI1 provides a paradigm for understanding receptor-mediated signaling in plants and contribution of post-translational modifications. Here, we show that BRI carries SUMO modifications in planta on two intracellular lysine residues and that temperature elevation triggers BRI1 deSUMOylation mediated by the Desi3a SUMO protease. Importantly, BRI1 deSUMOylation leads to downregulation of BR signaling via increased BRI1 interaction with the BIK1 negative regulator and increased BRI1 endocytosis. Loss of BRI1 deSUMOylation in desi3a mutants boosts plant responses to heat, indicating that BRI1 deSUMOylation acts as a brake to keep temperature responses in check. Our study uncovers a new post-translational modification targeting BRI1 and sheds light on its functional outcome for environmentally-controlled plant growth.

category: plant biology


title: Systematic histone H4 replacement in Arabidopsis thaliana reveals a role for H4R17 in regulating flowering time

abstract: Despite the broad array of roles for epigenetic mechanisms on regulating diverse processes in eukaryotes, no experimental system for the direct assessment of histone function is currently available in plants. In this work, we present the development of a genetic strategy in Arabidopsis thaliana in which modified H4 transgenes can completely replace the expression of endogenous histone H4. Using this strategy, we established a collection of plants expressing different H4 point mutants targeting residues that may be post-translationally modified in vivo. To demonstrate the utility of this new H4 mutant collection, we screened it to uncover substitutions in H4 that alter flowering time. We identified different mutations in the tail (H4R17A) and the globular domain (H4R36A, H4R39K, H4R39A, and H4K44A) of H4 that strongly accelerate the floral transition. Furthermore, we found a conserved regulatory relationship between H4R17 and the ISWI chromatin remodeling complex in plants. Similar to other biological systems, H4R17 regulates nucleosome spacing via ISWI. Overall, this work provides a large set of H4 mutants to the plant epigenetics community that can be used to systematically assess histone H4 function in A. thaliana and a roadmap to replicate this strategy for studying other histone proteins in plants.


title: Fam49b dampens TCR signal strength to regulate survival of positively selected thymocytes

abstract: The fate of developing T cells is determined by the strength of T cell receptor (TCR) signal they receive in the thymus. This process is finely regulated through tuning of positive and negative regulators in thymocytes. The Family with sequence similarity 49 member B (Fam49b) protein is a newly discovered negative regulator of TCR signaling that has been shown to suppress Rac-1 activity in vitro in cultured T cell lines. However, the contribution of Fam49b to thymic development of T cells is unknown. To investigate this important issue, we generated a novel mouse line deficient in Fam49b (Fam49b-KO). We observed that Fam49b-KO double positive (DP) thymocytes underwent excessive negative selection, whereas the positive selection stage was unaffected. This altered development process resulted in significant reductions in CD4 and CD8 single positive thymocytes as well as peripheral T cells. Interestingly, a large proportion of the TCR{gamma}{delta}+ and CD8+TCR{beta}+ gut intraepithelial T lymphocytes were absent in Fam49b-KO mice. Our results demonstrate that Fam49b dampens thymocytes TCR signaling in order to escape negative selection during development, uncovering the function of Fam49b as a critical regulator of selection process to ensure normal thymocyte development.

category: immunology


title: Arresting microbiome development limits immune system maturation and resistance to infection.

abstract:
Disruptions to the intestinal microbiome during weaning lead to long-term negative effects on host immune function. However, the critical host-microbe interactions occurring during weaning required for healthy immune system development remain poorly understood. We find that restricting microbiome maturation during weaning leads to stunted immune system development and increased susceptibility to enteric infection. We developed a gnotobiotic mouse model of the early-life microbiome designated as Pediatric Community (PedsCom). This nine-member consortium of microbes derived from intestinal microbiomes of preweaning mice stably colonized germfree adult mice and was efficiently transmitted to offspring for multiple generations. Unexpectedly, the relative abundance of PedsCom microbes were largely unaffected by the transition from a milk-based to a fiber rich solid food diet. PedsCom mice developed less peripheral regulatory T cells and Immunoglobulin A, hallmarks of microbiota-driven immune system development. Consistent with defects in maturation, adult PedsCom mice retain high susceptibility to salmonella infection characteristic of young mice and humans. Altogether, our work illustrates how the post-weaning transition in intestinal microbiome composition contributes to normal immune maturation and protection from enteric infection. Accurate modelling of the pre-weaning microbiome provides a window into the microbial requirements of healthy immune development and suggests an opportunity to design microbial interventions at weaning to improve immune system development in human infants.

One Sentence SummaryArresting microbiome development stunts immune ontogeny


title: Polyfunctional IL-17A+ MAIT cells are expanded in the peripheral blood of patients with HLA-B27+ axial spondyloarthritis

abstract:
ObjectivesStudies in axial spondyloarthritis (axSpA) have yielded conflicting results regarding the identity of the major IL-17A-producing lymphocyte populations. The goal of this study was to comprehensively assess the production of IL-17A and related cytokines by peripheral blood lymphocytes in axSpA.

MethodsPeripheral blood mononuclear cells were isolated from patients with axSpA and healthy controls matched for age, sex and HLA-B27 status. Unstimulated cells and cells activated with PMA/Ionomycin were analyzed by 25-parameter fluorescent flow cytometry. Data were analyzed by hierarchical gating, UMAP and SPICE.

ResultsExcept for a reduced frequency of mucosal-associated invariant T (MAIT) cells and natural killer (NK) cells, there were no other significant differences in abundance of major lymphocyte populations in axSpA patients compared with controls. Increased IL-17A production in axSpA was observed in total non-B lymphocytes and in MAIT cells. The fraction of MAIT cells expressing the tissue residency markers CD69 and CD103 was increased in axSpA. CD103 positive MAIT cells were enriched for IL-17A producers. axSpA patients demonstrated an expansion of MAIT cell subsets producing IL-17A, IL-17F, GM-CSF and TNF. This expansion was only observed in HLA-B27+ patients.

ConclusionsWe document an expansion of polyfunctional IL-17A+ MAIT cells in the peripheral blood of HLA-B27+ patients with axSpA. These results are consistent with the implied role of intestinal dysbiosis or inflammation in axSpA pathogenesis.

Key messagesWhat is already known about this subject? O_LIVarious IL-17A-producing lymphocyte populations have been implicated in the pathogenesis of axSpA.
C_LI

What does this study add? O_LIPolyfunctional MAIT cells capable of producing IL-17A, IL-17F, GM-CSF and TNF are expanded in the peripheral blood of HLA-B27+ patients with axSpA.
C_LI

How might this impact on clinical practice or future developments? O_LIOverproduction of IL-17A by MAIT cells is the most consistent finding of peripheral blood lymphocyte studies in axSpA.
C_LIO_LIOur data support the pathogenetic link between intestinal and axial inflammation in axSpA.
C_LI


title: Development of an HIV reporter virus that identifies latently infected CD4+ T cells

abstract: There is no cure for HIV infection as the virus establishes a latent reservoir, which escapes highly active antiretroviral treatments. One major obstacle is the difficulty to identify cells that harbor latent proviruses. We devised a novel viral vector that carries a series of versatile reporter molecules that are expressed in an LTR-dependent or LTR-independent manner and allows to accurately distinguish productive from latent infection. Using primary human CD4+ T cells, we show that transcriptionally silent proviruses make up over 50% of all infected cells. These latently infected cells harbor proviruses, but lack evidence for viral transcription. LTR silent integrations occurred to variable degrees in all CD4+ T-subsets examined with CD4+ TEM and CD4+ Treg displaying the highest frequency. Viral vectors such as the one described here, permit interrogation HIV latency at a single-cell resolution, revealing mechanisms of latency establishment and allowing for the characterization of effective latency reversing agents.


title: Genomic surveillance of Bacillus cereus sensu lato strains isolated from meat and poultry products in South Africa enables inter- and intra-national surveillance and source tracking

abstract:
Members of the Bacillus cereus sensu lato (s.l.) species complex, also known as the B. cereus group, vary in their ability to cause illness, but are frequently isolated from foods, including meat products; however, food safety surveillance efforts that employ whole-genome sequencing (WGS) often neglect these potential pathogens. Here, WGS was used to characterize B. cereus s.l. strains (n = 25) isolated during surveillance of meat products in South Africa. Strains were collected from beef, poultry, and mixed meat products obtained from (i) retail outlets, processing plants, and butcheries across six South African provinces (n = 15, 7, and 1, respectively), and (ii) imports in cold stores (n = 2). Strains were assigned to panC Groups IV, III, II, and V (n = 18, 5, 1, and 1, respectively) and spanned multiple genomospecies, regardless of the taxonomy used. All strains possessed diarrheal toxin-encoding genes, while one sequence type 26 (ST26) strain possessed cereulide (emetic toxin) synthetase-encoding genes. No strains harbored anthrax toxin- or capsule-encoding genes. The 25 strains were partitioned into 15 lineages via in silico seven-gene multi-locus sequence typing (MLST), six of which contained multiple strains sequenced in this study, which were identical or nearly identical at the whole-genome scale. Five MLST lineages contained (nearly) identical genomes collected from two or three South African provinces; one MLST lineage contained nearly identical genomes from two countries (South Africa and the Netherlands), indicating that B. cereus s.l. can spread intra- and inter-nationally via foodstuffs.

ImportanceNation-wide foodborne pathogen surveillance programs that employ high-resolution genomic methods have been shown to provide vast public health and economic benefits. However, B. cereus s.l. are often overlooked during large-scale, routine WGS efforts. Thus, to our knowledge, no studies to date have evaluated the potential utility of WGS for B. cereus s.l. surveillance and source tracking in foodstuffs. In this proof-of-concept study, we applied WGS to B. cereus s.l. strains collected via South Africas national surveillance program of domestic and imported meat products, and we provide strong evidence that B. cereus s.l. can be disseminated intra- and inter-nationally via the agro-food supply chain. Our results showcase that WGS can be used for source tracking of B. cereus s.l. in foods, although future WGS and isolate metadata collection efforts are needed to ensure that B. cereus s.l. surveillance initiatives are on par with those of other foodborne pathogens.


title: Elevation of Clavibacter michiganensis subsp. californiensis to species level as Clavibacter californiensis sp. nov., merging and re-classification of Clavibacter michiganensis subsp. chilensis and Clavibacter michiganensis subsp. phaseoli as Clavibacter chilensis sp. nov. based on complete genome in-silico analyses

abstract: The Gram-positive Clavibacter genus is currently divided into seven species (C. michiganensis, C. nebraskensis, C. capsici, C. sepedonicus, C. tessellarius, C. insidiosus and C. zhangzhiyongii) and three subspecies (C. michiganensis subsp. californiensis, C. michiganensis subsp. chilensis and C. michiganensis subsp. phaseoli). Recent studies have indicated that the taxonomic rank of the subspecies must be re-evaluated. In this research, we assessed the taxonomy position of the three C. michiganensis subspecies and clarified the taxonomic nomenclature of other 75 Clavibacter strains. The complete genomes of the type strains of the three Clavibacter subspecies, type strain of C. tessellarius and C. nebraskensis A6096 were sequenced using PacBio RSII technology. Application of whole-genome-based computational approaches such as average nucleotide identity (ANI), digital DNA-DNA hybridization (dDDH), multi-locus sequence analysis (MLSA) of seven housekeeping genes (acnA, atpD, bipA, icdA, mtlD, recA and rpoB), phylogenomic tree reconstructed from 1,028 core genes, and ANI-based phylogeny pinpointed conclusive evidence to raise C. michiganensis subsp. californiensis to the species status. These results led us to propose the establishment of C. californiensis sp. nov. as a species with its type strain C55 (=CFBP 8216=ATCC BAA-2691). Moreover, the orthologous and in-silico dot plot analyses, along with the aforementioned bioinformatic strategies, revealed a high degree of homology between C. michiganensis subsp. chilensis and C. michiganensis subsp. phaseoli. Based on these outcomes, we proposed to combine both subspecies into a single taxon and elevate its rank to the species level as C. chilensis sp. nov., with ZUM3936 (= ATCC BAA-2690 = CFBP 8217) as the type strain.


title: A family of C. elegans VASA homologs control Argonaute pathway specificity and promote transgenerational silencing

abstract: Germline Argonautes direct transcriptome surveillance within peri-nuclear membraneless organelles called nuage. In C. elegans, a family of Vasa-related Germ Line Helicase (GLH) proteins localize in, and promote the formation of nuage called P granules. Previous studies have implicated GLH proteins in inherited silencing but direct roles in amplification of small RNAs, or in target mRNA or Argonatue binding have not been identified. Here we show that GLH proteins compete with each other to control Argonaute pathway specificity, bind directly to Argonaute-target mRNAs and act to promote the amplification of small RNAs required for transgenerational inheritance. We show that the ATPase cycle of GLH-1 regulates its direct binding to the Argonaute WAGO-1 which engages amplified small RNAs. Our findings support a dynamic and direct role for GLH proteins in inherited silencing beyond their role as structural components of nuage.


title: Distinct roles for SOX2 and SOX21 in differentiation, distribution and maturation of pulmonary neuroendocrine cells

abstract: Pulmonary neuroendocrine (NE) cells represent a small population in the airway epithelium, but despite this, hyperplasia of NE cells is associated with several lung diseases, such as congenital diaphragmatic hernia and bronchopulmonary dysplasia. The molecular mechanisms causing the development of NE cell hyperplasia remains poorly understood. Previously, we showed that the SOX21 modulates the SOX2-initiated differentiation of epithelial cells in the airways (Eenjes, 2021). Here, we show that precursor NE cells start to develop in the SOX2+SOX21+ airway region and that SOX21 suppresses the differentiation of airway progenitors to precursor NE cells. During development, clusters of NE cells start to form and NE cells mature by expressing neuropeptide proteins, such as CGRP. Deficiency in SOX2 resulted in decreased clustering, while deficiency in SOX21 increased both the numbers of NE ASCL1+ precursor cells early in development, and the number of cell clusters at E18.5. In addition, at the end of gestation (E18.5), a number of NE cells in Sox2 heterozygous mice, did not yet express CGRP suggesting a delay in maturation. In conclusion, SOX2 and SOX21 function in the initiation, migration and maturation of NE cells.

category: developmental biology


title: Assessing Myf5 and Lbx1 contribution to carapace development by reproducing their turtle-specific signatures in mouse embryos

abstract:
BackgroundThe turtle carapace is an evolutionary novelty resulting from changes in the processes that build ribs and their associated muscles in most tetrapod species. Turtle embryos have several unique features that might play a role in this process, including the carapacial ridge, a Myf5 gene with shorter coding region that generates an alternative splice variant lacking exon 2, and unusual expression patterns of Lbx1 and HGF.

ResultsWe generated Myf5 alleles reproducing the Myf5 turtle expression features. At mid gestation, mouse embryos expressing Myf5 lacking exon 2 reproduced some early properties of turtle somites, but still developed into viable and fertile mice. Extending Lbx1 expression into the hypaxial dermomyotomal lip of trunk somites to mimic the turtle Lbx1 expression pattern, produced fusions in the distal part of the ribs.

ConclusionsTurtle-like Myf5 activity might generate a plastic state in developing trunk somites under which they can either enter carapace morphogenetic routes, possibly triggered by signals from the carapacial ridge, or still engage in the development of a standard tetrapod ribcage in the absence of those signals. In addition, trunk Lbx1 expression might play a later role in the formation of the lateral border of the carapace.


title: Selection analysis identifies unusual clustered mutational changes in Omicron lineage BA.1 that likely impact Spike function

abstract: Among the 30 non-synonymous nucleotide substitutions in the Omicron S-gene are 13 that have only rarely been seen in other SARS-CoV-2 sequences. These mutations cluster within three functionally important regions of the S-gene at sites that will likely impact (i) interactions between subunits of the Spike trimer and the predisposition of subunits to shift from down to up configurations, (ii) interactions of Spike with ACE2 receptors, and (iii) the priming of Spike for membrane fusion. We show here that, based on both the rarity of these 13 mutations in intrapatient sequencing reads and patterns of selection at the codon sites where the mutations occur in SARS-CoV-2 and related sarbecoviruses, prior to the emergence of Omicron the mutations would have been predicted to decrease the fitness of any genomes within which they occurred. We further propose that the mutations in each of the three clusters therefore cooperatively interact to both mitigate their individual fitness costs, and adaptively alter the function of Spike. Given the evident epidemic growth advantages of Omicron over all previously known SARS-CoV-2 lineages, it is crucial to determine both how such complex and highly adaptive mutation constellations were assembled within the Omicron S-gene, and why, despite unprecedented global genomic surveillance efforts, the early stages of this assembly process went completely undetected.


title: In vitro demonstration of broad host range mobile genetic elements transferring antibiotic resistance from the human microbiome

abstract: Mobile genetic elements (MGEs) carrying antibiotic resistance genes (ARGs) disseminate ARGs when they mobilise into new bacterial hosts. To investigate the scale of such horizontal gene transfer (HGT) events between human gut commensals and enteropathogens for the first time, we compared 1354 cultured commensal strains (540 species) to 45,403 enteropathogen strains (12 species) and found 64,188 MGE-mediated ARG transfer events between the two groups using established methods. Among the 5931 MGEs involved, we found 15 broad host range elements predicted to have crossed different bacterial phyla while also occurring in animal and environmental microbiomes. We experimentally demonstrated that predicted broad host range MGEs can mobilise from commensals Dorea longicatena and Hungatella hathewayi to enteropathogens Klebsiella oxytoca, crossing phyla simultaneously. Our work establishes the MGE-mediated ARG dissemination network between human gut commensals and enteropathogens and highlights novel broad host range MGEs as targets for future ARG dissemination management.


title: Video based phenotyping platform for the laboratory mouse

abstract: Automated detection of complex animal behavior remains a challenge in behavioral neurogenetics. Developments in computer-vision have greatly advanced automated behavior detection and allow high-throughput pre-clinical studies. An integrated hardware and software solution is necessary to facilitate the adoption of these advances in the field of behavioral neurogenetics, particularly for non-computational labs. We have published a series of papers using an open field arena to annotate complex behaviors such as grooming, posture, and gait as well as higher level constructs such as frailty. Here, we describe in detail our integrated hardware and software solution for robust, high quality video monitoring of mice. We provide safety data for mice in long-term monitoring by recording environmental conditions in the arenas as well as health markers. The details provided here offer one potential solution for standardized high quality video data for computer vision tasks, and enables replication of our work.

category: animal behavior and cognition


title: Metabolic competition between lipid metabolism and histone methylation regulates sexual differentiation in human malaria parasites.

abstract:
Like most pathogens, malaria parasites balance persistence in the current host with transmission to the next. For Plasmodium falciparum, the most widespread and virulent human malaria parasite, persistence depends on continuous asexual replication in red blood cells while transmission requires differentiation into non-replicating gametocytes, the male and female cells able to infect the mosquito vector. This developmental decision is controlled via stochastic derepression of a heterochromatin-silenced locus encoding the transcription factor PfAP2-G, which acts as the master switch for sexual differentiation. Recent work showed that the frequency of pfap2-g derepression is responsive to extracellular levels of phospholipid precursors. However, the regulatory mechanisms linking these metabolites to epigenetic regulation of pfap2-g was hitherto unknown. Using chemical, metabolomic, and genetic approaches, we show that this response is mediated by metabolic competition for the methyl donor S-adenosylmethionine between histone methyltransferases and phosphoethanolamine methyltransferase, a critical enzyme in the parasites pathway for de novo phosphatidyl-choline synthesis. When phosphatidyl-choline precursors are scarce, increased consumption of S-adenosylmethionine for de novo synthesis of phosphatidyl-choline results in a decrease in histone methylation marks that mediate silencing of pfap2-g, thereby up-regulating its transcription and increasing the frequency of sexual differentiation. Our findings reveal a key connection between metabolite utilization and gene expression in malaria parasites that forms the mechanistic link between phosphatidyl-choline metabolism and the frequency of sexual commitment.

O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

category: systems biology


title: Exposure to 1800 MHz LTE electromagnetic fields under proinflammatory conditions decreases the response strength and increases the acoustic threshold of auditory cortical neurons.

abstract: Increased needs in mobile phone communications have raised successive generations (G) of wireless technologies, which could differentially affect biological systems. We have assessed how a single 2h head-only exposure to a 4G long term evolution (LTE)-1800 MHz electromagnetic field (EMF) impacts on the microglial space coverage and the electrophysiological neuronal activity in the primary auditory cortex (ACx) in rats submitted to an acute neuroinflammation induced by lipopolysaccharide. The mean specific absorption rate in the ACx was 0.5 W/kg. Multiunit recording revealed that LTE-EMF triggered reduction in the response strength to pure tones and to natural vocalizations, together with an increase in acoustic threshold in the low and medium frequencies. Iba1 immunohistochemistry showed no change in the area covered by microglia cell bodies and processes. In healthy rats, the same LTE-exposure induced no change in response strength and acoustic threshold. Our data indicate that an acute neuroinflammation sensitize neuronal responses to LTE-EMF, which leads to an altered processing of acoustic stimuli in the ACx.

category: neuroscience


title: Impact of pre-breeding feeding practices on rabbit mammary gland development at mid-pregnancy.

abstract:
Optimizing rabbit does preparation during early life to improve reproductive performance is a major challenge for breeders. Does selected for reproduction have nutritional needs, which may not be supplied with the common practice of feed restriction during rearing in commercial rabbit production. Nutrition during early life was already known to influence metabolism, reproduction and mammary gland development later in life, in particular during pregnancy. The aim of this study was to analyze four different restriction feeding strategies during post-weaning and over the pubertal periods (high or moderate restriction feeding applied from 5 to 9 weeks of age and/or restricted or ad libitum over the following 3 weeks constituting the pubertal period).

Unlike food intake, which remains regular, mean body weight gain was inversely proportional to the dietary restriction applied over the considered periods. The feeding strategies in place for the four groups have no effect on the reproductive parameters of the females, as opposed to certain metabolic parameters such as cholesterolemia, that vary with dietary intake. Furthermore, restriction programs have impacted mammary tissular structures at mid-pregnancy. The expression of lipid metabolism enzymes (Fatty acid synthase N and Stearoyl co-A desaturase) is also modified in mammary epithelial cells by the dietary strategies implemented. Moreover, milk gene expression, used as differentiation markers, indicates a better mammary epithelial development regarding further lactation, in the case of the less restrictive strategies during early life period, especially the higher feeding allowance.

Our results highlight the importance of investigating feeding conditions of young female rabbits and nutrition in early life rearing, in order to provide specific recommendations for optimizing lactation and thus preventing neonatal mortality of the offspring.

category: physiology


title: Identification of differential hypothalamic DNA methylation and gene expression associated with sexual partner preferences in rams

abstract: The sheep is a valuable model to test whether hormone mechanisms that sexually differentiate the brain underlie the expression of sexual partner preferences because as many as 8% of rams prefer same-sex partners. Epigenetic factors such as DNA methylation act as mediators in the interaction between steroid hormones and the genome. Variations in the epigenome could be important in determining morphological or behavior differences among individuals of the same species. In this study, we explored DNA methylation differences in the hypothalamus of male oriented rams (MORs) and female oriented rams (FORs). We employed reduced representation bisulfite sequencing (RRBS) to generate a genome-wide map of DNA methylation and RNA-Seq to profile the transcriptome. We found substantial DNA methylation and gene expression differences between FORs and MORs. Although none of the differentially methylated genes yielded significant functional terms directly associated with sex development, three differentially expressed genes were identified that have been associated previously with sexual behaviors. We hypothesize that these differences are involved in the phenotypic variation in ram sexual partner preferences, whereas future studies will have to find the specific mechanisms. Our results add an intriguing new dimension to sheep behavior that should be useful for further understanding epigenetic and transcriptomic involvement.


title: Algorithms for Estimating Time-Locked Neural Response Components in Cortical Processing of Continuous Speech

abstract:
ObjectiveThe Temporal Response Function (TRF) is a linear model of neural activity time-locked to continuous stimuli, including continuous speech. TRFs based on speech envelopes typically have distinct components that have provided remarkable insights into the cortical processing of speech. However, current methods may lead to less than reliable estimates of single-subject TRF components. Here, we compare two established methods, in TRF component estimation, and also propose novel estimation algorithms that utilize prior knowledge of these components, bypassing the full TRF estimation.

MethodsWe compared two established algorithms, ridge and boosting, and two novel algorithms based on Subspace Pursuit and Expectation Maximization, which directly estimate TRF components given plausible assumptions regarding component characteristics. Single-channel, multi-channel, and source-localized TRFs were fit on simulations and real magnetoencephalographic data. Performance metrics included model fit and component estimation accuracy.

ResultsBoosting and ridge have comparable performance in component estimation. The novel algorithms outperformed the others in simulations, but not on real data, possibly due to the plausible assumptions not actually being met. Ridge had marginally better model fits on real data, but also more spurious TRF activity.

ConclusionResults indicate that both smooth (ridge) and sparse (boosting) algorithms perform comparably at TRF component estimation. The SP and EM algorithms may be accurate, but rely on assumptions of component characteristics.

SignificanceThis systematic comparison establishes the suitability of widely used and novel algorithms for estimating robust TRF components, which is essential for improved subject-specific investigations into the cortical processing of speech.


title: Rethinking Statistical Learning as a Dynamic Stochastic Process, from The Motor Systems Perspective

abstract: The brain integrates streams of sensory input and builds accurate predictions, while arriving at stable percepts under disparate time scales. This stochastic process bears different dynamics for different people, yet statistical learning (SL) currently averages out, as noise, individual fluctuations in data streams registered from the brain as the person learns. We here adopt the motor systems perspective to reframe SL. Specifically, we rethink this problem using the demands that the persons brain faces to predict, and control variations in biorhythmic activity akin to those present in bodily motions. This new approach harnesses gross data as the important signals, to reassess how individuals learn predictive information in stable and unstable environments. We find two types of learners: narrow-variance learners, who retain explicit knowledge of the regularity embedded in the stimuli -the goal. They seem to use an error-correction strategy steadily present in both stable and unstable cases. In contrast, broad-variance learners emerge only in the unstable environment. They undergo an initial period of memoryless learning characterized by a gamma process that starts out exponentially distributed but converges to Gaussian. We coin this mode exploratory, preceding the more general error-correction mode characterized by skewed-to-symmetric distributions and higher signal content from the start. Our work demonstrates that statistical learning is a highly dynamic and stochastic process, unfolding at different time scales, and evolving distinct learning strategies on demand.


title: Effect of habitual reading direction on saccadic eye movements

abstract: Cognitive processes can influence the characteristics of saccadic eye movements. Reading habits, including habitual reading direction, also affects cognitive and visuospatial processes, favouring attention to the side where reading begins. Few studies have investigated the effect of habitual reading direction on saccade directionality of low-cognitive-demand stimuli (such as dots). The current study examined horizontal prosaccade, antisaccade and self-paced saccade in subjects with two primary habitual reading directions. We hypothesised that saccades responding to the target in subjects habitual reading direction would show a longer prosaccade latency and lower antisaccade error rate (errors being a reflexive glance to a sudden-appearing target, rather than a saccade away from it). Sixteen young Chinese participants with primary habitual reading direction from left to right and sixteen young Arabic and Persian participants with primary habitual reading direction from right to left were recruited. Subjects needed to look towards a 5{degrees}/ 10{degrees}target in the prosaccade task or look towards the mirror image location of the target in the antisaccade task and look between two 10-degree targets in the self-paced saccade task. Only Arabic and Persian participants showed a shorter and directional prosaccade latency towards 5{degrees}target against their habitual reading direction. No significant effect of primary reading direction on antisaccade latency towards the correct directions was found. However, we found that Chinese readers generated significantly shorter prosaccade latencies and higher antisaccade directional errors compared with Arabic and Persian readers. The present study provides an insight into the effect of reading habits on saccadic eye movements in response to low-cognitive-demand stimuli and offers a platform for future studies to investigate the relationship between reading habits and neural mechanisms of eye movement behaviours.


title: Neuron-specific protein network mapping of autism risk genes identifies shared biological mechanisms and disease relevant pathologies

abstract:
Autism spectrum disorder (ASD) is a genetically heterogeneous disorder. Sequencing studies have identified hundreds of risk genes for autism spectrum disorder (ASD), but the signaling networks of genes at the protein level remain largely unexplored, which can provide insight into previously unknown individual and convergent disease pathways in the brain. To address this gap, we used neuron- specific proximity-labeling proteomics (BioID) to identify protein-protein interaction (PPI) networks of 41 ASD-risk genes. Network analysis revealed the combined 41 risk gene PPI network map had more shared connectivity between unrelated ASD-risk genes than represented in existing public databases. We identified common pathways between established and uncharacterized risk genes, including synaptic transmission, mitochondrial/metabolic processes, Wnt signaling pathways, ion channel activity and MAPK signaling. Investigation of the mitochondrial and metabolic network using gene knockouts revealed a functional hub in neurons for multiple risk genes not previously associated with this pathway. Further, we identified that the uncharacterized ASD-risk gene PPP2R5D localizes to the synapse, which is disrupted by patient de novo missense mutations. Investigation of de novo missense variants of additional synaptic ASD-risk genes demonstrated that changes in PPI networks can capture synaptic transmission deficits. The neuronal 41 ASD-risk gene PPI network map also revealed enrichment for an additional 112 ASD-risk genes and human brain cell types implicated in ASD pathology. Interestingly, clustering of ASD-risk genes based on their PPI network connectivity identified multiple gene groups that correlate mutation-type to clinical behavior scores. Together, our data reveal that using PPI networks to map ASD risk genes can identify previously unknown individual and convergent neuronal signaling networks, provide a method to assess the impact of patient variants, and reveal new biological insight into disease mechanisms.

Main HighlightsO_LINeuron-specific protein interaction screening of 41 ASD-risk genes to identify new disease mechanisms at the protein level
C_LIO_LIHigh connectivity between multiple unrelated ASD-risk genes at the protein interaction level
C_LIO_LIPPI networks show disease-relevant pathways including synaptic transmission, metabolic pathways, Wnt signaling, ion channel activity, MAPK signaling
C_LIO_LIMetabolic pathways, such as TCA cycle and pyruvate metabolism, are altered in neurons by multiple ASD-risk genes not previously linked to this pathway
C_LIO_LINovel localization of uncharacterized ASD-risk gene PPP2R5D at the synapse, which is disrupted by de novo mutations identified in patients
C_LIO_LIClustering of ASD-risk genes based on PPI connectivity identifies multiple gene groups that show correlation between mutation-type and clinical behavior scores, revealing the importance of understanding PPI networks in ASD
C_LI


title: Computed tomographic analysis of the dental system of three Jurassic ceratopsians: implications for the evolution of the tooth replacement pattern and diet in early-diverging ceratopsians

abstract: The dental system of ceratopsids is among the most specialized structure in Dinosauria, and includes high angled wear surfaces, split tooth roots, and multiple teeth in each tooth family. However, the early evolution of this unique dental system is generally poorly understood due to a lack of knowledge of the dental morphology and development in early-diverging ceratopsians. Here we study the dental system of three of the earliest-diverging Chinese ceratopsians: Yinlong and Hualianceratops from the early Late Jurassic of Xinjiang, and Chaoyangsaurus from the Late Jurassic of Liaoning. By using micro-computed tomographic analyses, our study has revealed significant new information regarding the dental system of these early ceratopsians, including no more than five replacement teeth in each jaw quadrant; at most one generation of replacement teeth in each alveolus; nearly full resorption of the functional tooth root during tooth replacement; and occlusion with low-angled, concave wear facets that differs significantly from the shearing occlusal system seen in ceratopsids. Yinlong displays an increase in the number of maxillary tooth alveoli and a decrease in the number of replacement teeth during ontogeny as well as the retention of remnants of functional teeth in the largest individual. Early-diverging ceratopsians thus display a relatively slow tooth replacement rate compared to late-diverging ceratopsians. Combined with paleobotany and palaeoenvironment data, Yinlong likely uses gastroliths to triturate foodstuffs, and the difference in diet strategy might have influenced the pattern of tooth replacement in later-diverging ceratopsians.

category: paleontology


title: Re-use of labware reduces CO2 equivalent footprint and running costs in laboratories

abstract: Laboratory-based research is resource intensive in terms of financial costs and its carbon footprint. Research laboratories require immense amounts of energy to power equipment, as well as large volumes of materials, particularly of single-use item consumption. In fact, many laboratories have essentially become reliant on single-use plastics. Understanding the full carbon footprint of consumable usage is increasingly important as many research institutes commit to carbon neutrality. To date, no carbon footprint assessment has been conducted to detail the differences between single-use plastics, and reusable glass in a laboratory setting. Here, we analyse the CO2 equivalent (CO2e) footprint of utilising single-use plastics, and re-use of glass or plastic items within laboratory environments. We focused our assessment on four commonly utilised consumables for mammalian cell and bacterial culture, and found that re-use scenarios resulted in substantial reduction in CO2e footprint up to 11-fold. In addition, we estimated the long-term financial costs of re-use and single-use scenarios, and found that re-use had either similar or much lower running costs even when including technical staff wage. We concluded that research facilities must foster re-use in laboratory consumables, while reserving single-use items for select, defined cases. Our study highlights the need to account for indirect CO2e footprint in designing a carbon-neutral lab.

category: cell biology


title: CaPTrends: A global database of Carnivoran Population Trends

abstract:
MotivationPopulation trend information is an  Essential Biodiversity Variable for monitoring change in biodiversity over time. Here, we present a global dataset of 1122 population trends describing changes in abundance over time in large mammals from the Order Carnivora - some of the worlds most charismatic and functionally important fauna.

Main types of variables includedKey data fields for each record: species, coordinates, trend timeframe, methods of data collection and analysis, and population timeseries or summarised trend value. Population trend values are reported using quantitative metrics in 75% of records that collectively represent more than 6500 population estimates. The remaining records qualitatively describe population change (e.g. increase).

Spatial location and grainRecords represent locations across the globe (latitude: -51.0 to 80.0; longitude: -166.0 to 166.0) with more trends found within the northern temperate zone.

Time period and grainRecords span from 1726 to 2017, with 92% of trends starting after 1950.

Major taxa and level of measurementWe conducted a semi-systematic search for population trend data in 87 species from four families in the order Carnivora: Canidae, Felidae, Hyaenidae and Ursidae. We compiled data for 50 of the 87 species.

Software format.csv

category: ecology


title: A protocol for locating and counting transgenic sequences from laboratory animals using a map-then-capture (MapCap) sequencing workflow: procedure and application of results.

abstract:
Transgenic rodent models for human diseases have been widely used over the past 50 years and are a mainstay of many biomedical research programs. Oftentimes the sequence of the transgenic segment of DNA is carefully designed but incorporation of this DNA into the host genome is less well understood. Structural variation and insertional mutagenesis may occur at transgenic insertion sites. Here, we present a robust workflow including identification of the transgene locus via selective Illumina sequencing followed by Cas9-mediated target DNA enrichment of the locus, which successfully identified beginning and end sites of a large transgenic insertion into a murine model for human amylin-induced type II diabetes. Enriched sequences were mapped via Oxford Nanopore sequencing. Although the insertion was too long for a single mapped genetic sequence to encompass, the method provided multiple insights relevant to the animal model: a minimum number of forward- and reverse-facing transcript copies as well as characterization of an inversion point within the insertion site. The insertion start point containing both murine and human DNA was used to identify and separate animals hemizygous for the transgenic insertion from homozygous animals. This identification could be performed early in the rodent life cycle prior to maturation (i.e. breeding age), thus allowing for management of colony phenotypes and eliminating the need to "genotype by phenotype" later on (onset of amylin-induced type II diabetes does not occur until ~8-10 weeks of age for this model). We further confirmed our homozygous diabetic mice function the same as colonies established in other labs and present full antibody and fluorescent-staining protocols (available in SI). Lastly, we note that, due to our genotyping, a novel animal was identified within our colony: non-diabetic homozygous mice. Indeed, only 37% of homozygous mice bred in our colony became diabetic.

AUTHOR SUMMARY (for broad audience)Transgenic rodent models are important to studying human diseases. When creating a new rodent model, one may insert new DNA into a well-characterized background genome. However, it is oftentimes not known where the new DNA was incorporated, how many times it was incorporated, or if any coding sequences or regulatory elements within native DNA were disrupted. Here, we have developed a method to characterize transgenic animals, and have applied it to a popular model for studying human amylin-induced type II diabetes.

category: genetics


title: Viability, task switching, and fall avoidance of the simplest dynamic walker

abstract: Humans display great versatility when performing goal-directed tasks while walking. However, the extent to which such versatility helps with fall avoidance remains unclear. We recently demonstrated a functional connection between the motor regulation needed to achieve task goals (e.g. maintaining walking speed) and a simple walkers ability to reject large disturbances. Here, for the same model, we identify the viability kernel--the state space region in which the walker can step forever via at least one sequence of push-off inputs per state. We further find that only a few basins of attraction of the speed-regulated walkers steady-state gaits can fully cover the viability kernel. This highlights a potentially important role of task-level motor regulation in fall avoidance. Therefore, we posit an adaptive hierarchical control/regulation strategy that switches between different task-level regulators to avoid falls. Our hierarchical task switching controller only requires a target value of the regulated observable--a  task switch--at each walking step, each chosen from a small, predetermined collection. Because humans have typically already learned to perform such tasks during nominal walking conditions, this suggests that the  information cost of biologically implementing such controllers for the nervous system, including cognitive demands in humans, could be relatively low.


title: NestedBD: Bayesian Inference of Phylogenetic Trees From Single-Cell DNA Copy Number Profile Data Under a Birth-Death Model

abstract:
Copy number aberrations (CNAs) are ubiquitous in many types of cancer. Inferring CNAs from cancer genomic data could help shed light on the initiation, progression, and potential treatment of cancer. While such data have traditionally been available via "bulk sequencing", the more recently introduced techniques for single-cell DNA sequencing (scDNAseq) provide the type of data that makes CNA inference possible at the single-cell resolution.

In this paper, we introduce a new birth-death evolutionary model of CNAs as well as a Bayesian method, NestedBD, for the inference of evolutionary trees (topologies and branch lengths with relative mutation rates) from single-cell data under this model. We assessed the accuracy of our method on both simulated and biological data and compared it to the accuracy of two standard phylogenetic tools, namely neighbor-joining and maximum parsimony (MP). We show through simulations that our method infers more accurate topologies and branch lengths. We also studied the ancestral state reconstruction accuracy with the birth-death evolutionary model and found it outperformed MP. Finally, running all three methods on a colorectal cancer data set, we observed that among all three methods, only the phylogeny inferred by NestedBD clearly separated the primary tumor cells from the metastatic ones, providing a more plausible history of the tumor cells.


title: Conservation and divergence of myelin proteome and oligodendrocyte transcriptome profiles between humans and mice

abstract: Human myelin disorders are commonly studied in mouse models. Since both clades evolutionarily diverged approximately 85 million years ago, it is critical to know to what extent the myelin protein composition has remained similar. Here we use quantitative proteomics to analyze myelin purified from human white matter and find that the relative abundance of the structural myelin proteins PLP, MBP, CNP and SEPTIN8 correlates well with that in C57Bl/6N- mice. Conversely, multiple other proteins were identified exclusively or predominantly in human or mouse myelin. This is exemplified by peripheral myelin protein-2 (PMP2), which was specific to human CNS myelin, while tetraspanin-2 (TSPAN2) and connexin-29 (CX29/GJC3) were confined to mouse myelin. Assessing published scRNA-seq-datasets, human and mouse oligodendrocytes display well-correlating transcriptome profiles but divergent expression of distinct genes including Pmp2, Tspan2 and Gjc3. Species-dependent diversity of oligodendroglial mRNA-expression and myelin protein composition can be informative when translating from mouse models to humans.


title: UbE3-APA: A Bioinformatic Strategy to Elucidate Ubiquitin E3 Ligase Activities in Quantitative Proteomics Study

abstract:
MotivationUbiquitination is widely involved in protein homeostasis and cell signaling. Ubiquitin E3 ligases are critical regulators of ubiquitination that recognize and recruit specific ubiquitination targets for the final rate-limiting step of ubiquitin transfer reactions. Understanding the ubiquitin E3 ligase ac-tivities will provide knowledge in the upstream regulator of the ubiquitination pathway and reveal po-tential mechanisms in biological processes and disease progression. Recent advances in mass spec-trometry-based proteomics have enabled deep profiling of ubiquitylome in a quantitative manner. Yet, functional analysis of ubiquitylome dynamics and pathway activity remains challenging.

ResultsHere, we developed a UbE3-APA, a computational algorithm and stand-alone python-based software for Ub E3 ligase Activity Profiling Analysis. Combining an integrated annotation database with statistical analysis, UbE3-APA identifies significantly activated or suppressed E3 ligases based on quantitative ubiquitylome proteomics datasets. Benchmarking the software with published quantitative ubiquitylome analysis confirms the genetic manipulation of SPOP enzyme activity through overexpres-sion and mutation. Application of the algorithm in the re-analysis of a large cohort of ubiquitination proteomics study revealed the activation of PARKIN and the co-activation of other E3 ligases in mito-chondria depolarization-induced mitophagy process. We further demonstrated the application of the algorithm in the DIA-based quantitative ubiquitylome analysis.

AvailabilitySource code and binaries are freely available for download at URL: https://github.com/Chenlab-UMN/Ub-E3-ligase-Activity-Profiling-Analysis, implemented in python and supported on Linux and MS Windows

Contactyuechen@umn.edu

Supplementary informationSupplementary data are available.

category: bioinformatics


title: Impaired iron recycling from erythrocytes is an early iron-dependent hallmark of aging

abstract: Aging affects iron homeostasis, as evidenced by tissue iron loading and toxicity and common anemia in the elderly. Iron needs in mammals are met primarily by iron-recycling from senescent red blood cells (RBCs), a task chiefly accomplished by splenic red pulp macrophages (RPMs) via erythrophagocytosis. Given that RPMs continuously process iron, their cellular functions might be susceptible to age-dependent decline, a condition that has been largely unexplored to date. Here, we found that 10-11-months-old female mice exhibit iron loading, diminished lysosomal activity, and decreased erythrophagocytosis rate in RPMs. These impairments lead to the retention of senescent hemolytic RBCs in the spleen, and the formation of undegradable iron- and heme-rich extracellular protein aggregates, likely derived from ferroptotic RPMs. We further found that feeding mice an iron-reduced diet alleviates iron accumulation in RPMs, enhances their ability to clear erythrocytes, and limits ferroptosis. Consequently, this diet ameliorates hemolysis of splenic RBCs and the formation of iron-rich aggregates, increasing serum iron availability in aging mice. Using RPM-like cells, we show that the diminished iron-recycling capacity of RPMs is underlain by iron accumulation and reduced expression of heme-catabolizing enzyme heme oxygenase 1 (HO-1). Taken together, we identified RPM collapse as an early hallmark of aging and demonstrated that dietary iron reduction improves iron turnover efficacy.


title: Dogma and belief: the primary lesion in age-related hearing loss is old news

abstract: Presbycusis, or age-related hearing loss, is the most common sensory deficit globally, and the biggest modifiable risk factor for a later dementia diagnosis. Despite its ubiquity, however, the primary pathology contributing to presbycusis is reportedly contentious, particularly the relative role of damage to the sensory outer hair cells compared to the stria vascularis, an important inner ear structure that maintains the ionic concentration of inner ear fluids that surround it. To determine what might be the "dogma" of the field regarding the primary pathology in presbycusis, we conducted an online Google survey (https://forms.gle/GPreoePmRxBBkchc7) asking relevant respondents in the field their opinions on the matter. In Question (Q1), respondents were asked to rate  in your opinion from  least likely to  most likely (on a scale 1 to 4 [being  most likely])  what is the primary pathology contributing to presbycusis? in terms of  damage to: i) the inner hair cells, ii) outer hair cells (OHCs), iii) spiral ganglion, iv) stria vascularis. The term  dogma suggests that the proportion of people in the field who believe that the main cause of presbycusis is damage to the stria vascularis is at least 50%. The results of our survey estimated this proportion to be 19/101 = 0.188 (95% CI [0.124 0.275]) and a two-sided test of the null hypothesis that this proportion is at least 0.5 was rejected (p < 10-10). When it came to opining what  other professionals in the field consider to be the primary pathology contributing to presbycusis (Q2), the tendency to rank  damage to OHCs as being the primary pathology was 45%. Stria vascularis was least likely to be ranked 4 (11%) by professionals in the field opining about the beliefs of others. Even when  noise damage was excluded as a factor (Q3), the  most likely contributing factor to presbycusis was stated to be damage to the OHCs (42%). Our data suggest the dogma of the field is that damage to outer hair cells is the primary pathology in presbycusis.


title: Transcriptomic and functional genetic evidence for distinct ecophysiological responses across complex life cycle stages

abstract:
Organisms with complex life cycles demonstrate a remarkable ability to change their phenotypes across development, presumably as an evolutionary adaptation to developmentally variable environments. Developmental variation in environmentally sensitive performance, and thermal sensitivity in particular, has been well documented in holometabolous insects. For example, thermal performance in adults and juvenile stages exhibit little genetic correlation (genetic decoupling) and can evolve independently, resulting in divergent thermal responses. Yet, we understand very little about how this genetic decoupling occurs. We tested the hypothesis that genetic decoupling of thermal physiology is driven by fundamental differences in physiology between life stages, despite a potentially conserved Cellular Stress Response. We used RNAseq to compare transcript expression in response to a cold stressor in Drosophila melanogaster larvae and adults and used RNAi (RNA interference) to test whether knocking down nine target genes differentially affected larval and adult cold tolerance. Transcriptomic responses of whole larvae and adults during and following exposure to -5{degrees}C were largely unique both in identity of responding transcripts and in temporal dynamics. Further, we found no relationship between stage-specificity and tissue-specificity of transcripts, suggesting that the differences are not simply driven by differences in tissue composition across development. In addition, RNAi of target genes resulted in largely stage-specific and sometimes sex-specific effects on cold tolerance. The combined evidence suggests that thermal physiology is largely stage-specific at the level of gene expression, and thus natural selection may be acting on different loci during the independent thermal adaptation of different life stages.

Summary StatementRNAseq and gene knockout via transgenic RNAi lines suggest that physiological responses to low temperatures are largely distinct across life stages of the fly Drosophila melanogaster.

category: evolutionary biology


title: Pain-related learning signals in the human insula

abstract:
Pain is not only a perceptual phenomenon, but also a preeminent learning signal. In reinforcement learning models, prediction errors (PEs) play a crucial role, i.e. the mismatch between expectation and sensory input. In particular, advanced learning models require the representation of different types of PEs, namely signed PEs (whether more or less pain was expected) to specify the direction of learning, and unsigned PEs (the absolute deviation from an expectation) to adapt the learning rate. The insula has been shown to play an important role in pain intensity coding and in signaling surprise. However, mainly unsigned PEs could be identified in the anterior insula. It remains an open question whether these PEs are specific to pain, and whether signed PEs are also represented in the insula.

To answer these questions, 47 subjects learned associations of two conditioned stimuli (CS) with four unconditioned stimuli (US; painful heat or loud sound, of one low and one high intensity each) while undergoing functional magnetic resonance imaging (fMRI) and skin conductance response (SCR) measurements. CS-US associations reversed multiple times between intensities and between sensory modalities, generating frequent PEs.

SCRs indicated comparable nonspecific characteristics of the two modalities. fMRI analyses focusing on the insular and opercular cortices contralateral to painful stimulation showed that activation in the anterior insula correlated with unsigned intensity PEs. Importantly, this unsigned PE signal was similar for pain and aversive sounds and also modality PEs, indicating an unspecific aversive surprise signal. Conversely, signed pain intensity PE signals were modality-specific and located in the dorsal posterior insula, an area previously implicated in pain intensity processing.

Previous studies have identified abnormal insula function and abnormal learning as potential causes of pain chronification. Our findings link these results and suggest one potential mechanism, namely a misrepresentation of learning relevant prediction errors in the insular cortex.


title: Structural basis for Nirmatrelvir in vitro efficacy against the Omicron variant of SARS-CoV-2

abstract: The COVID-19 pandemic continues to be a public health threat with emerging variants of SARS-CoV-2. Nirmatrelvir (PF-07321332) is a reversible, covalent inhibitor targeting the main protease (Mpro) of SARS-CoV-2 and the active protease inhibitor in PAXLOVID (nirmatrelvir tablets and ritonavir tablets). We evaluated the in vitro catalytic activity and in vitro potency of nirmatrelvir against the main protease (Mpro) of prevalent variants of concern (VOC) or variants of interest (VOI): Alpha (, B.1.1.7), Beta ({beta}, B.1.351), Delta ({delta}, B1.617.2), Gamma ({gamma}, P.1), Lambda ({lambda}, B.1.1.1.37/C37), Omicron (o, B.1.1.529) as well as the original Washington or wildtype strain. These VOC/VOI carry prevalent mutations at varying frequencies in the Mpro specifically for: , {beta},{gamma} (K90R), {lambda} (G15S) and o (P132H). In vitro biochemical enzymatic assay characterization of the enzyme kinetics of the mutant Mpros demonstrate that they are catalytically comparable to wildtype. Nirmatrelvir has similar potency against each mutant Mpro including P132H that is observed in the Omicron variant with a Ki of 0.635 nM as compared to a Ki of 0.933nM for wildtype. The molecular basis for these observations were provided by solution-phase structural dynamics and structural determination of nirmatrelvir bound to the o, {lambda} and {beta} Mpro at 1.63 - 2.09 [A] resolution. These in vitro data suggest that PAXLOVID has the potential to maintain plasma concentrations of nirmatrelvir many-fold times higher than the amount required to stop the SARS-CoV-2 VOC/VOI, including Omicron, from replicating in cells (1).

category: biochemistry


title: Structural basis for Nirmatrelvir in vitro efficacy against SARS-CoV-2 variants


title: Nirmatrelvir, an orally active Mpro inhibitor, is a potent inhibitor of SARS-CoV-2 Variants of Concern

abstract: New variants of SARS-CoV-2 with potential for enhanced transmission, replication, and immune evasion capabilities continue to emerge causing reduced vaccine efficacy and/or treatment failure. As of January 2021, the WHO has defined five  variants of concern (VOC): B.1.1.7 (Alpha, ), B.1.351 (Beta, {beta}), P.1 (Gamma, {gamma}), B.1.617.2 (Delta, {delta}), and B.1.1.529 (Omicron, o). To provide a therapeutic option for the treatment of COVID-19 and variants, Nirmatrelvir, the antiviral component of PAXLOVID, an oral outpatient treatment recently authorized for conditional or emergency use treatment of COVID-19, was developed to inhibit SARS-CoV-2 replication. Nirmatrelvir (PF-07321332) is a specific inhibitor of coronavirus main protease (Mpro, also referred to as 3CLpro), with potent antiviral activity against several human coronaviruses, including SARS-CoV-2, SARS-CoV, and MERS (Owen et al, Science 2021. doi: 10.1126/science.abl4784). Here, we evaluated PF-07321332 against the five SARS-CoV-2 VOC (, {beta}, {gamma}, {delta},, o) and two Variants of Interest or VOI, C.37 ({lambda}) and B.1.621 (), using qRT-PCR in VeroE6 cells lacking the P-glycoprotein (Pgp) multidrug transporter gene (VeroE6 P-gp knockout cells). Nirmatrelvir potently inhibited USA-WA1/2020 strain, and , {beta}, {gamma}, {lambda}, {delta}, , and o variants in VeroE6 P-gp knockout cells with mean EC50 values 38.0 nM, 41.0 nM, 127.2 nM, 24.9 nM, 21.2 nM, 15.9 nM, 25.7 nM and 16.2 nM, respectively. Sequence analysis of the Mpro encoded by the variants showed ~100% identity of active site amino acid sequences, reflecting the essential role of Mpro during viral replication leading to ability of Nirmatrelvir to exhibit potent activity across all the variants.

category: microbiology


title: ccnWheat: A Database for Comparing Co-expression Networks Analysis of Allohexaploid Wheat and Its Progenitors

abstract: Genetic and epigenetic changes after polyploidization events could result in variable gene expression and modified regulatory networks. Here, using large-scale transcriptome data, we constructed co-expression networks for diploid, tetraploid, and hexaploid wheat species, and built a platform for comparing co-expression networks of allohexaploid wheat and its progenitors, named ccnWheat. ccnWheat is a platform for searching and comparing specific functional co-expression networks, as well as identifying the related functions of the genes clustered therein. Functional annotations like pathway, gene family, protein-protein interactions, microRNA (miRNA), and several lines of epigenome data are integrated in this platform, and Gene Ontology (GO) annotation, gene set enrichment analysis (GSEA), motif identification, and other useful tools are also included. Using ccnWheat, we found that the network of WHEAT ABERRANT PANICLE ORGANIZATION 1 (WAPO1) has more co-expressed genes related to spike development in hexaploid wheat than its progenitors. We also found a novel motif of CArG specifically in the promoter region of WAPO-A1, suggesting that neofunctionalization of the WAPO-A1 gene affects spikelet development in hexaploid wheat. ccnWheat is useful for investigating co-expression networks and conducting other associated analyses, and thus facilitates comparative and functional genomic studies in wheat. ccnWheat is freely available at http://bioinformatics.cau.edu.cn/ccnWheat.


title: Particle flow modulates growth dynamics and nanoscale-arrested growth of transcription factor condensates in living cells

abstract: Liquid-liquid phase separation (LLPS) is emerging as key physical principle for biological organization inside living cells, forming condensates that play important roles in the regulation of multiple functions. Inside living nuclei, transcription factor (TF) condensates regulate transcriptional initiation and amplify transcriptional output of expressed genes. Yet, the biophysical parameters controlling TF condensation are still poorly understood. Here we applied a battery of single molecule imaging tools, theory and simulations to investigate the physical properties of TF condensates of the Progesterone Receptor (PR) in vivo. Analysis of individual PR trajectories at different ligand concentrations showed marked signatures of a ligand-tunable and regulated LLPS process. Using a machine learning architecture, we uncovered that diffusion within condensates follows fractional Brownian motion, reflecting viscoelastic interactions between PR and chromatin within condensates. High density single molecule localization maps further revealed that condensate growth dynamics is dominated by Brownian motion coalescence (BMC) at shorter times, but deviate at longer timescales reaching a growth plateau with nanoscale condensate sizes. To understand our observations we developed an extension of the BMC model by including stochastic unbinding of particles within condensates. The model reproduced the BMC behavior together with finite condensate sizes a steady-state, fully recapitulating our experimental data. Our results are thus consistent with droplet growth dynamics being regulated by the escaping probability of TFs molecules from condensates. The interplay between condensation assembly and molecular escaping maintains an optimum physical condensate size. Such phenomena must have implications for the biophysical regulation of other TF condensates and could also operate in multiple biological scenarios.

category: biophysics


title: TGFβ receptor inhibition unleashes interferon-β production by tumor-associated macrophages and enhances radiotherapy efficacy

abstract:
BackgroundTransforming growth factor-beta (TGF{beta}) can limit the efficacy of cancer treatments, including radiotherapy (RT), by inducing an immunosuppressive tumor environment. The association of TGF{beta} with impaired T cell infiltration and antitumor immunity is known, but the mechanisms by which TGF{beta} participates in immune cell exclusion and limits the efficacy of antitumor therapies warrant further investigations.

MethodsWe used the clinically relevant TGF{beta} receptor 2 (TGF{beta}R2)-neutralizing antibody MT1 and the small molecule TGF{beta}R1 inhibitor LY3200882 and evaluated their efficacy in combination with RT against murine orthotopic models of head and neck and lung cancer.

ResultsWe demonstrated that TGF{beta} pathway inhibition strongly increased the efficacy of RT. TGF{beta}R2 antibody upregulated interferon beta (IFN{beta}) expression in tumor-associated macrophages (TAMs) within the irradiated tumors and favored T cell infiltration at the periphery and within the core of the tumor lesions. We highlighted that both the antitumor efficacy and inhibition of immune exclusion observed with the combination of MT1 and RT were dependent on type I interferon signaling.

ConclusionsThese data shed new light on the role of TGF{beta} in limiting the efficacy of RT, identifying a novel mechanism involving the inhibition of macrophage-derived type I interferon production, and fostering the use of TGF{beta}R inhibition in combination with RT in therapeutic strategies for the management of head and neck and lung cancer.

category: cancer biology


title: The Life Span of the Great Black Wasp Sphex pensylvanicus L> (Hymenoptera: Sphecidae) and Other Observations from a Nesting Aggregation in Sioux City, Iowa, USA> 2003 -2017

abstract: In a fifteen year period, the observed life span of both male and female Sphex pensylvanicus was documented in an aggregation of these wasps behind a retaining wall on a residential lot in Sioux City, Iowa, USA. In addition, early season emergence of the males before the females (protandry) and instances of male territorial behavior were also confirmed. Half or more of the wasps disappeared; possible out-migration?

category: zoology


title: Prediction and validation of host cleavage targets of SARS-CoV-2 3C like protease

abstract: How SARS-CoV-2 causes the observed range of clinical manifestations and disease severity remains poorly understood. SARS-CoV-2 encodes for two proteases (3CLPro and PLPro), vital for viral production, but also promiscuous with respect to host protein targets, likely contributing to the range of disease. Pharmacological inhibition of the 3C-like3 protease has revealed remarkable reduction in hospitalization and death in phase 2/3 clinical studies. However, the mechanisms responsible for the pathology mediated by those proteases are still unclear. In this study, we develop a bioinformatic algorithm, leveraging experimental data from SARS-CoV, to predict host cleavage targets of the SARS-CoV-2 3C-like protease, or 3CLPro. We capture targets of the 3CL protease described previously for SARS-CoV, and we identify hundreds of new putative targets. We experimentally validate a number of these predicted targets, including the giant sarcomeric protein Obscurin, and show that expression of 3CL protease alone recapitulates the sarcomeric disorganization seen by SARS-CoV-2 infection of hiPSC-derived cardiomyocytes. Our data provide a resource to identify putative host cleavage targets of 3CL protease that contribute to mechanisms and heterogeneity of disease in COVID-19 and future coronavirus outbreaks.


title: Nirmatrelvir, Molnupiravir, and Remdesivir maintain potent in vitro activity against the SARS-CoV-2 Omicron variant

abstract: Variants of SARS-CoV-2 have become a major public health concern due to increased transmissibility, and escape from natural immunity, vaccine protection, and monoclonal antibody therapeutics. The highly transmissible Omicron variant has up to 32 mutations within the spike protein, many more than previous variants, heightening these concerns of immune escape. There are now multiple antiviral therapeutics that have received approval for emergency use by the FDA and target both the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) and the main protease (Mpro), which have accumulated fewer mutations in known SARS-CoV-2 variants. Here we test nirmatrelvir (PF-07321332), and other clinically relevant SARS-CoV-2 antivirals, against a panel of SARS-CoV-2 variants, including the novel Omicron variant, in live-virus antiviral assays. We confirm that nirmatrelvir and other clinically relevant antivirals all maintain activity against all variants tested, including Omicron.