Molecular dynamics simulation study reveals effects of key mutations on spike protein structure in SARS-CoV-2 SARS-CoV-2 has been spreading rapidly since 2019 and has produced large-scale mutations in the genomes. The mutation in genes may lead to changes in protein structure, which would have a great impact on the epidemiological characteristics. In this study, we selected the key mutations of SARS-CoV-2 from a real-time monitoring tool, including D614G, A222V, N501Y, T716I, S982A, D1118H of spike (S) protein, and performed molecular dynamics (MD) simulations on single-site mutant D614G, double-site mutant D614G&A222V and penta-site mutant N501Y&D614G&T716I&S982A&D1118H to investigate their effects on protein structure and stability using molecular dynamics (MD) simulations. The results suggested that D614G improved the stability of S protein, while D614G&A222V and N501Y&D614G&T716I&S982A&D1118H showed an increased solvent accessible surface area and they might enhance the ability of protein to react with the outside environment. Our findings could complement the mechanistic link between genotype--phenotype--epidemiological characteristics in the study of SARS-CoV-2. We also found no significant difference between the antigenicity of S protein and the mutants through Ellipro, which may reference for vaccine development and application.
Network Analysis Reveals Different Cellulose Degradation Strategies across Trichoderma harzianum Strains Associated with XYR1 and CRE1 Trichoderma harzianum, whose gene expression is tightly controlled by the transcription factors (TFs) XYR1 and CRE1, is a potential candidate for hydrolytic enzyme production. Here, we performed a network analysis of T. harzianum IOC-3844 and T. harzianum CBMAI-0179 to explore how the regulation of these TFs varies between these strains. In addition, we explored the evolutionary relationships of XYR1 and CRE1 protein sequences among Trichoderma spp. The results of the T. harzianum strains were compared with those of Trichoderma atroviride CBMAI-0020, a mycoparasitic species. Although transcripts encoding carbohydrate-active enzymes (CAZymes), TFs, transporters, and proteins with unknown functions were coexpressed with cre1 or xyr1, other proteins indirectly related to cellulose degradation were identified. The enriched GO terms describing the transcripts of these groups differed across all strains, and several metabolic pathways with high similarity between both regulators but strain-specific differences were identified. In addition, the CRE1 and XYR1 subnetworks presented different topology profiles in each strain, likely indicating differences in the influences of these regulators according to the fungi. The hubs of the cre1 and xyr1 groups included transcripts not yet characterized or described as being related to cellulose degradation. The first-neighbor analyses confirmed the results of the profile of the coexpressed transcripts in cre1 and xyr1. The analyses of the shortest paths revealed that CAZymes upregulated under cellulose degradation conditions are most closely related to both regulators, and new targets between such signaling pathways were discovered. Although the evaluated T. harzianum strains are phylogenetically close and their amino acid sequences related to XYR1 and CRE1 are very similar, the set of transcripts related to xyr1 and cre1 differed, suggesting that each T. harzianum strain used a specific regulation strategy for cellulose degradation. More interestingly, our findings may suggest that XYR1 and CRE1 indirectly regulate genes encoding proteins related to cellulose degradation in the evaluated T. harzianum strains. An improved understanding of the basic biology of fungi during the cellulose degradation process can contribute to the use of their enzymes in several biotechnological applications and pave the way for further studies on the differences across strains of the same species.
Propelling and perturbing appendages together facilitate strenuous ground self-righting Terrestrial animals must self-right when overturned on the ground, but this locomotor task is strenuous. To do so, the discoid cockroach often pushes its wings against the ground to begin a somersault which rarely succeeds. As it repeatedly attempts this, the animal probabilistically rolls to the side to self- right. During winged self-righting, the animal flails its legs vigorously. Here, we studied whether wing opening and leg flailing together facilitate strenuous ground self-righting. Adding mass to increase hind leg flailing kinetic energy increased the animals self-righting probability. We then developed a robot with similar strenuous self-righting behavior and used it as a physical model for systematic experiments. The robots self-righting probability increased with wing opening and leg flailing amplitudes. A potential energy landscape model revealed that, although wing opening did not generate sufficient kinetic energy to overcome the high pitch potential energy barrier to somersault, it reduced the barrier for rolling, facilitating the small kinetic energy from leg flailing to probabilistically overcome it to self-right. The model also revealed that the stereotyped body motion during self-righting emerged from physical interaction of the body and appendages with the ground. Our work demonstrated the usefulness of potential energy landscape for modeling self-righting transitions.

Impact statementAnimals can, and robots should, use different types of appendages together to propel and perturb themselves to self-right when overturned, a strenuous yet crucial locomotor task. The Temporal Statistics of Musical Rhythm across Western Genres: An Amplitude Modulation Phase Hierarchy Model Statistical learning by the human brain plays a core role in the development of cognitive systems like language and music. Both music and speech have structured inherent rhythms, however the acoustic sources of these rhythms are debated. Theoretically, rhythm structures in both systems may be related to a novel set of acoustic statistics embedded in the amplitude envelope, statistics originally revealed by modelling childrens nursery rhymes. Here we apply similar modelling to explore whether the amplitude modulation (AM) timescales underlying rhythm in music match those in child-directed speech (CDS). Utilising AM-driven phase hierarchy modelling previously applied to infant-directed speech (IDS), adult-directed speech (ADS) and CDS, we test whether the physical stimulus characteristics that yield speech rhythm in IDS and CDS describe rhythm in music. Two models were applied. One utilized a low-dimensional representation of the auditory signal adjusted for known mechanisms of the human cochlear, and the second utilized probabilistic amplitude demodulation, estimating the modulator (envelope) and carriers using Bayesian inference. Both models revealed a similar hierarchically-nested temporal modulation structure across Western musical genres and instruments. Core bands of AM and spectral patterning matched prior analyses of IDS and CDS, and music showed strong phase dependence between slower bands of AMs, again matching IDS and CDS. This phase dependence is critical to the perception of rhythm. Control analyses modelling other natural sounds (wind, rain, storms, rivers) did not show similar temporal modulation structures and phase dependencies. We conclude that acoustic rhythm in language and music has a shared statistical basis. Neuronal Activation of the Gastrointestinal Tract Shapes the Gut Environment in Mice The enteric nervous system (ENS) coordinates responses that shape the intestinal milieu, and participates in bi-directional communication with various organs including the brain. Tools to study, and knowledge of, ENS development and function lag behind brain research. Herein, we deploy recombinant adeno-associated viral (rAAV) vectors with enhanced tropism for the gut to map and activate gut associated neurons in mice with spatial and temporal resolution. rAAV-mediated fluorescent labelling coupled with whole-tissue clearing methods in the small intestine and colon reveal novel neuronal architectures in the gut of mice. Viral delivery of genetically-encoded calcium indicators enable in vivo measurement of ENS activity. Further, we employed chemogentics to specifically activate gut neurons that express choline acetyltransferase (ChAT+) or tyrosine hydroxylase (TH+). Targeted activation of ChAT+ or TH+ neuronal populations associated with the gastrointestinal (GI) tract alters the intestinal transcriptome, mouse and microbial proteomes, metagenome, and fecal metabolome. We reveal previously unknown and broad roles for enteric neurons in modulating GI physiology, mucosal immunity, and microbiome structure, and provide evidence for novel functions by the ENS such as regulating fungal colonization and shaping of bile acid profiles in the gut. These advanced experimental tools and rich datasets with multi-parameter characterization of the gut ecosystem may facilitate expanded understanding and study of the ENS. A protease-activatable luminescent biosensor andreporter cell line for authentic SARS-CoV-2 infection Efforts to define serological correlates of protection against COVID-19 have been hampered by the lack of a simple, scalable, standardised assay for SARS-CoV-2 infection and antibody neutralisation. Plaque assays remain the gold standard, but are impractical for high-throughput screening. In this study, we show that expression of viral proteases may be used to quantitate infected cells. Our assays exploit the cleavage of specific oligopeptide linkers, leading to the activation of cell-based optical biosensors. First, we characterise these biosensors using recombinant SARS-CoV-2 proteases. Next, we confirm their ability to detect viral protease expression during replication of authentic virus. Finally, we generate reporter cells stably expressing an optimised luciferase-based biosensor, enabling viral infection to be measured within 24 h in a 96- or 384-well plate format, including variants of concern. We have therefore developed a luminescent SARS-CoV-2 reporter cell line, and demonstrated its utility for the relative quantitation of infectious virus and titration of neutralising antibodies.

Author summaryTechniques for measuring infection with SARS-CoV-2 in the laboratory are laborious and time-consuming, and different laboratories use different approaches. There is therefore no generally agreed way to quantitate neutralising antibodies against SARS-CoV-2, which block infection with the virus and protect people from COVID-19. In this study, we describe a new way to measure SARS-CoV-2 infection, which is much simpler and faster than existing methods. It relies on the production of a specific protease enzyme by the virus, which is able to cleave and activate an engineered protein biosensor in infected cells. This biosensor emits light in the presence of viral infection, and the amount of light released is used as a readout for the amount of infectious SARS-CoV-2 present. The signal is very sensitive, so the number of infected cells required is very small, and the method can be scaled-up to test many samples at once. In particular, we demonstrate how it can be used to detect different variants of SARS-CoV-2, and quantitate neutralising antibodies against these viruses. Cast away on Mindoro island: could density dependence threaten tamaraw (Bubalus mindorensis) on the long term? Endangered species, despite often living at low population densities, may undergo unexpected density-dependent feedbacks in the case of successful recovery or marked reduction in range. Because density-dependence dynamics can increase risk of extinction, these effects can hamper conservation efforts. In this study, we analyse the dynamics of the largest population of the tamaraw (Bubalus mindorensis), a critically endangered species endemic to the Mindoro island, Philippines. The population is located within a < 3,000 ha area in Mounts Iglit-Baco Natural Park, with limited expansion possibilities. We took advantage of a 21 year time series of tamaraw counts to estimate annual population growth rate and possible density-dependence, accounting for sampling errors in the counts. The tamaraw population has been increasing in size at an average rate of +5% per year, as would be expected given its protected status by law. Population growth showed strong spatial structuring, with a population growth close to +10% in the core area of protection, and a reduction of abundance of - 5% at the periphery of its range, inside the protected area. This range constriction is concerning because our best population dynamic model suggests significant negative density-dependence (Bayes factor = 0.9). The contraction of tamaraw range is likely caused by anthropogenic pressures forcing herbivores to live at relatively high densities in the core zone where protection is most effective. Our study highlights the fact that, despite the continuous population growth over the last two decades, the long-term viability of the Mounts Iglit-Baco Natural Park tamaraw population remains uncertain. Genetic integration of behavioural and endocrine components of the stress response The vertebrate stress response comprises a suite of behavioural and physiological traits that must be functionally integrated to ensure organisms cope adaptively with acute stressors. Natural selection should favour functional integration, leading to a prediction of genetic integration of these traits. Despite the implications of such genetic integration for our understanding of human and animal health, as well as evolutionary responses to natural and anthropogenic stressors, formal quantitative genetic tests of this prediction are lacking. Here we demonstrate that acute stress response components in Trinidadian guppies are both heritable and integrated on the major axis of genetic covariation. This integration could either facilitate or constrain evolutionary responses to selection, depending upon the alignment of selection with this axis. Such integration also suggests artificial selection on the genetically correlated behavioural responses to stress could offer a viable non-invasive route to the improvement of health and welfare in captive animal populations. Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. Non-invasive imaging of gene expression and protein secretion dynamics in living mice: identification of ectopic prothrombin expression as driver of thrombosis in cancer The topology of gene expression and protein localization is a crucial characteristic of life, where the spatiotemporal dynamic of secretory proteins instruct higher order organization, including the orchestration of developmental and adaptive programs. However tools to non-invasively interrogate the fate of secretory proteins in vivo are scarce. Here we introduce a genetic tagging strategy for in vivo imaging of the secretion and expression dynamics of secretory proteins in living animals. Applying this to a prototypical liver-derived secretory protein, we demonstrate that this approach, combined with optical in-vivo imaging, uncovers extrahepatic prothrombin expression in multiple novel anatomical sites (including testes, placenta, brain, kidney, heart and lymphatic system) and in emerging tumors, resulting in significant amounts of tumor-derived prothrombin in the blood with procoagulant properties. Syngeneic cell lines from this mouse model enable unravelling regulatory mechanisms in high resolution, and in a scalable format ex vivo. Beyond discovering new functions of proteins in a targeted manner, this model allows identifying rheostats in the cross-talk between gene expression and availability of a secretory protein. It is also a valuable resource for uncovering novel (tissue-specific) therapeutic vulnerabilities. A high throughput lipidomics method using scheduled multiple reaction monitoring Lipid compositions of cells, tissues and bio-fluids are complex, with varying concentrations and structural diversity, which makes their identification challenging. Newer methods for comprehensive analysis of lipids are thus necessary. Herein, we propose a targeted-mass spectrometry based method for large-scale lipidomics using a combination of variable retention time window and relative dwell time weightage. Using this, we detected more than 1000 lipid species, including structural isomers. The limit of detection varied from femtomolar to nanomolar range and the coefficient of variance <30% for 849 lipid species. We used this method to identify lipids altered due to Vitamin B12 deficiency and found that the levels of lipids with {omega}-3 fatty acid chains decreased while those with {omega}-6 increased. This method enables identification of by far the largest number of lipid species with structural isomers in a single experiment and would significantly advance our understanding of the role of lipids in biological processes. Chromosome breaks in breast cancers occur near herpes tumor virus sequences and explain why the cancer comes back One explanation for why breast cancer has a relentless tendency to come back after treatment is that breast cancer phenomes are fluid, so it is difficult for therapy to target new phenotypes that arise. In breast cancer, viral interactions cause chromosome breakage and fusion, producing anaphase bridges containing two centromeres. Cell division then leads to more chromosome breaks, more mutations, and new cancer phenotypes. Analyses of public data from about 2100 breast cancers implicate Epstein-Barr virus (EBV or Human Herpes Virus 4) in the cycles that break chromosomes and produce anaphase bridges. EBV generates breakage cycles with highly fragmented chromosomes (chromothripsis) as an inherent feature. These episodes continually produce new cancer driver mutations. DNA at chromothripsis boundaries in breast cancers consistently matches EBV tumor variants. Breakpoints are often near known EBV binding sites, and breast cancer shares an EBV methylation signature with other known EBV cancers. Breast cancer breakpoints on all chromosomes cluster around the same positions as in cancers definitively associated with EBV infection (nasopharyngeal cancer and Burkitts lymphoma). The status of EBV in the microbiome and the presence of anaphase bridges are phenotypic markers of breast cancer that are not explicitly targeted by chemotherapy. The breast cancer phenome continually supplies new cancer driver genes that foster breast cancer recurrence.

SummaryBreaks in breast cancer chromosomes and in fragments of anaphase bridges occur mainly near regions that resemble Epstein-Barr virus (EBV) tumor variants

Extensive breaks are an inherent outcome of anaphase bridges and the reassembly of highly fragmented chromosomes continually generates new cancer driver mutations and new cancer phenotypes.

Treatment does not explicitly target the underlying breakage and reassembly cycles so breast cancer often comes back.

The breast cancer phenome is not fixed and can continuously generate new phenotypes

Graphical Abstract

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org.highwire.dtl.DTLVardef@6537cforg.highwire.dtl.DTLVardef@132fa50org.highwire.dtl.DTLVardef@1cfcb04org.highwire.dtl.DTLVardef@11979aa_HPS_FORMAT_FIGEXP  M_FIG C_FIG Anti- SARS-CoV-2 Receptor Binding Domain Antibody Evolution after mRNA Vaccination Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection produces B-cell responses that continue to evolve for at least one year. During that time, memory B cells express increasingly broad and potent antibodies that are resistant to mutations found in variants of concern1. As a result, vaccination of coronavirus disease 2019 (COVID-19) convalescent individuals with currently available mRNA vaccines produces high levels of plasma neutralizing activity against all variants tested1, 2. Here, we examine memory B cell evolution 5 months after vaccination with either Moderna (mRNA-1273) or Pfizer- BioNTech (BNT162b2) mRNA vaccines in a cohort of SARS-CoV-2 naive individuals. Between prime and boost, memory B cells produce antibodies that evolve increased neutralizing activity, but there is no further increase in potency or breadth thereafter. Instead, memory B cells that emerge 5 months after vaccination of naive individuals express antibodies that are similar to those that dominate the initial response. While individual memory antibodies selected over time by natural infection have greater potency and breadth than antibodies elicited by vaccination, the overall neutralizing potency of plasma is greater following vaccination. These results suggest that boosting vaccinated individuals with currently available mRNA vaccines will increase plasma neutralizing activity but may not produce antibodies with breadth equivalent to those obtained by vaccinating convalescent individuals. Loss of the DYRK1A protein kinase results in reduction of ribosomal protein genes expression, ribosome mass and translation defects Ribosomal proteins (RPs) are evolutionary conserved proteins that are essential for protein translation. RP expression must be tightly regulated to ensure the appropriate assembly of ribosomes and to respond to the growth demands of cells. The elements regulating the transcription of RP genes (RPGs) have been characterized in yeast and Drosophila, yet how cells regulate the production of RPs in mammals is less well understood. Here, we show that a subset of RPG promoters is characterized by the presence of the palindromic TCTCGCGAGA motif and marked by the recruitment of the protein kinase DYRK1A. The presence of DYRK1A at these promoters is associated with enhanced binding of the TATA-binding protein, TBP, and it is negatively correlated with the binding of the GABP transcription factor, establishing at least two clusters of RPGs that could be coordinately regulated. However, DYRK1A silencing leads to a global reduction of RPGs, mRNA pointing at DYRK1A activities beyond those dependent on its chromatin association. Significantly, cells in which DYRK1A is depleted have reduced RP levels, fewer ribosomes, reduced global protein synthesis and smaller size. We therefore propose a novel role for DYRK1A in coordinating the expression of genes encoding RPs, thereby controlling cell growth in mammals. The compound of baicalein, wogonin, and oroxylin-A inhibiting EMT in A549 cell line via PI3K/AKT-TWIST1-glycolysis pathway Non-small cell lung cancer (NSCLC) is a worldwide disease with high morbidity and mortality, which is most derived from its metastasis. Some researches show that the epithelial-mesenchymal transition (EMT) process promotes lung cancer cells migration and invasion, leading to NSCLC metastasis. Total Flavonoid Aglycones Extract (TFAE) isolated from Scutellaria baicalensis was reported to inhibit tumor growth and induce apoptosis. In this study, we found that baicalein, wogonin, and oroxylin-A were the active compounds of TFAE. After reconstructing with these three compounds (baicalein (65.8%), wogonin (21.2%), and oroxylin-A (13.0%)), the reconstructed TFAE (reTFAE) present inhibitory effect on the EMT process of A549 cells. Then, bioinformatic technology was employed to elucidate the potential pharmacodynamic mechanism network of reTFAE. We identified the relationship between reTFAE and PI3K/Akt signaling pathways, with TWIST1 as the key protein. LY294002, the inhibitor of PI3K/Akt signaling pathway, and knock-down TWIST1 could significantly enhance the efficacy of reTFAE, with increasing expression of epithelial markers and decreasing expression of mesenchymal markers in A549 cells at the same time. Furthermore, stable isotope dimethyl-labeled proteomics technology was conducted to complement the follow-up mechanism that the EMT-inhibition process may be realized through glycolysis pathway. In conclusion, we claim that TWIST1-targeted flavonoids could provide a new strategy to inhibit EMT progress for the treatment of NSCLC. Cortical ripples in humans Hippocampal ripples index the reconstruction of spatiotemporal neuronal firing patterns essential for the consolidation of memories in the cortex during non-rapid eye movement sleep (NREM). Recently, cortical ripples in humans have been shown to enfold the replay of neuron firing patterns during cued recall. Here, we show that cortical ripples also occur during NREM in humans, with similar density, oscillation frequency (~90Hz), and amplitude to waking. Ripples occurred in all cortical regions with similar characteristics, unrelated to putative hippocampal connectivity, and were less dense and robust in higher association areas. Putative pyramidal and interneuron spiking phase-locked to cortical ripples during NREM, with phase delays consistent with ripple generation through pyramidal-interneuron feedback. Cortical ripples were smaller in amplitude than hippocampal ripples, but were similar in density, frequency, and duration. Cortical ripples during NREM typically occurred just prior to the upstate peak, often during spindles. Upstates and spindles have previously been associated with memory consolidation, and we found that cortical ripples grouped co-firing within the window of spike-timing-dependent plasticity. Thus, human NREM cortical ripples are: ubiquitous and stereotyped with a tightly focused oscillation frequency; similar to hippocampal ripples; associated with upstates and spindles; and associated with unit co-firing. These properties are consistent with cortical ripples possibly contributing to memory consolidation and other functions during NREM in humans. Parallel Characterization of cis-Regulatory Elements for Multiple Genes Using CRISPRpath Current pooled CRISPR screens for cis-regulatory elements (CREs) can only accommodate one gene based on its expression level. Here, we describe CRISPRpath, a scalable screening strategy for parallelly characterizing CREs of genes linked to the same biological pathway and converging phenotypes. We demonstrate the ability of CRISPRpath for simultaneously identifying functional enhancers of six genes in the 6-thioguanine-induced DNA mismatch repair pathway using both CRISPR interference (CRISPRi) and CRISPR nuclease (CRISPRn) approaches. 60% of the identified enhancers are known promoters with distinct epigenomic features compared to other active promoters, including increased chromatin accessibility and interactivity. Furthermore, by imposing different levels of selection pressure, CRISPRpath can distinguish enhancers exerting strong impact on gene expression from those exerting weak impact. Our results offer a nuanced view of cis-regulation and demonstrate that CRISPRpath can be leveraged for understanding the complex gene regulatory program beyond transcriptional output at scale. Serotonin and common antidepressants regulate lipoprotein(a) macropinocytosis via enhanced cell surface binding Multiple receptors have been implicated in lipoprotein(a) (Lp(a)) uptake, yet each receptor only partially accounts for the uptake, and no endocytic pathway has been described for Lp(a) internalisation to date. In this study, we define macropinocytosis as the endocytic pathway responsible for internalising Lp(a). In liver and macrophage cells, Lp(a) uptake was dependent on extracellular calcium and inhibited by amiloride or its derivative EIPA (5-(N-ethyl-N-isopropyl)-Amiloride). Uptake of the unique protein component of Lp(a), apolipoprotein(a) (apo(a)) alone was also dependent on macropinocytosis, indicating that it is the likely mediator of Lp(a) uptake. In macrophages, the tricyclic antidepressant, imipramine, and the selective serotonin reuptake inhibitors (SSRIs) citalopram, fluoxetine and sertraline all inhibited Lp(a) uptake, likely due to macrophage de-differentiation effects and dynamin inhibition (in the case of sertraline). In liver cells, imipramine and citalopram strongly stimulated Lp(a) uptake, while sertraline inhibited uptake. Imipramine and citalopram enhanced cell surface binding of Lp(a) rather than upregulating macropinocytosis per se, indicating that immobilisation on the plasma membrane is an important step in Lp(a) macropinocytosis. Serotonin treatment also increased Lp(a) cell surface binding, promoting subsequent macropinocytotic uptake, consistent with imipramine and citalopram boosting extracellular serotonin levels through serotonin transporter inhibition. Imipramine and citalopram increased Lp(a) delivery to Rab11-positive recycling endosomes which promoted recycling of the apo(a) component of Lp(a) into the culture media. These findings support the utility of imipramine and citalopram as promising Lp(a)-lowering therapeutics in people suffering from depression who often have elevated Lp(a) levels and an increased risk of cardiovascular disease.

SummaryElevated lipoprotein(a) (Lp(a)) levels have been found in individuals suffering from depression posing an increased risk of cardiovascular disease (CVD). Efforts to effectively reduce circulating Lp(a) levels have been limited due to an inability to ascribe a clear cellular uptake pathway, despite multiple receptors being implicated in Lp(a) uptake. We report here that Lp(a) uptake is mediated by the fluid-phase endocytic process, macropinocytosis. Furthermore, serotonin and commonly prescribed antidepressants promote Lp(a) uptake by macropinocytosis in liver cells by increasing cell surface association. These findings represent a major paradigm shift in Lp(a) biology since previous studies have attributed Lp(a) uptake to receptor-mediated endocytosis. They also imply that treatment with antidepressants may improve cardiovascular as well as mental health. A vascularized tumoroid model for human glioblastoma angiogenesis Glioblastoma (GBM) angiogenesis is critical for tumor growth and recurrence, making it a compelling therapeutic target. Here, a disease-relevant, vascularized tumoroid in vitro model with stem-like features and stromal surrounds is reported. The model is used to recapitulate how individual components of the GBMs complex brain microenvironment such as hypoxia, vasculature-related stro-mal cells and growth factors support GBM angiogenesis. It is scalable, tractable, cost-effective and can be used with biologically-derived or biomimetic matrices. Patient-derived primary GBM cells are found to closely participate in blood vessel formation in contrast to a GBM cell line containing differentiated cells. Exogenous growth factors amplify this effect under normoxia but not at hypoxia suggesting that a significant amount of growth factors is already being produced under hypoxic conditions. Under hypoxia, primary GBM cells and umbilical vein endothelial cells are found to strongly co-localize in a mosaic pattern to form sprouting vascular networks, with GBM cells acquiring an endothelial-like behaviour, which has been reported to occur in vivo. These findings demonstrate that our 3D tumoroid in vitro model exhibits biomimetic attributes that may permit its use as a preclinical model in studying microenvironment cues of tumor angiogenesis. Balancing sensitivity and specificity in preclinical research The success of scientific discovery in preclinical drug development is based on the different roles of exploration and confirmation. Exploration requires sensitivity while confirmation needs to be specific. Based on published empirical data, we simulated two preclinical research trajectories to examine their effectiveness to detect true effects while keeping animal numbers low. We compare a conventional p-value based approach with a method based on an a priori determined smallest effect size of interest (SESOI). Using a SESOI increases transition rates from exploration to confirmation and results in higher detection rates across the trajectory. Specificity in the SESOI trajectory increases if the prior probability of true effects is low. We conclude that employing a SESOI is superior to a p-value based approach. Based on our findings, we propose a reconsideration of planning and conducting preclinical experiments, especially when the prior probability of true hypotheses is low. Presence of a SARS-COV-2 protein enhances Amyloid Formation of Serum Amyloid A A marker for the severeness and disease progress of COVID-19 is overexpression of serum amyloid A (SAA) to levels that in other diseases are associated with a risk for SAA amyloidosis. In order to understand whether SAA amyloidosis could also be a long-term risk of SARS-COV-2 infections we have used long all-atom molecular dynamic simulations to study the effect of a SARS-COV-2 protein segment on SAA amyloid formation. Sampling over 40 {micro}s we find that presence of the nine-residue segment SK9, located at the C-terminus of the Envelope protein, increases the propensity for SAA fibril formation by three mechanisms: it reduces the stability of the lipid-transporting hexamer shifting the equilibrium toward monomers, it increases the frequency of aggregation-prone configurations in the resulting chains, and it raises the stability of SAA fibrils. Our results therefore suggest that SAA amyloidosis and related pathologies may be a long-term risk of SARS-COV-2 infections. Polycystins are upregulated in fibrotic kidneys and promote renal fibrosis Autosomal polycystic kidney disease (ADPKD) is caused by mutations in PKD1or PKD2 gene, which encodes polycystin-1 or polycystin-2 respectively. Paradoxically, polycystins are up-regulated in human ADPKD kidneys and overexpression of polycystin-1 even induced renal cyst formation in mice, suggesting that up-regulation of polycystin is pathogenic. In this study, we determined the role of polycystin in renal fibrosis and explored its underlying mechanisms. We found that the expression of polycystin-1 or polycystin-2 was positively correlated with the expression of collagen-I in the kidney of three different fibrotic mouse models. Pharmaceutical inhibition or genetic knockout of polycystin-2 reduced the expression of pro-fibrotic markers in fibrotic kidneys of two animal models. Moreover, conditional knockout of Pkd1 gene attenuated renal fibrosis in two mouse models. Enhancer of Zeste homolog 2 (EZH2) is a methyltransferase which plays important role in renal fibrosis. We found that EZH2 was positively correlated with the expression of polycystins in fibrotic kidneys. Conditional knockout of the Ezh2 attenuated the anti-fibrotic responses induced by Pkd1 deletion in fibrotic kidneys.

In conclusion, polycystins are up-regulated in injured kidneys and polycystins promote renal fibrosis through up-regulation of EZH2 in fibrotic kidneys, suggesting that renal fibrosis in polycystic kidneys is not the direct effect of PKD gene mutations.

SignificanceMutations of polycystins cause autosomal polycystic kidney disease (ADPKD) which is associated with increased renal fibrosis in cystic kidneys. However, the role of polycystin in renal fibrosis remains unclear. Here, we showed that the expression of polycystins is positively correlated with the development of renal fibrosis in injured kidneys. Pharmaceutical inhibition or genetic deletion of polycystin-2 attenuated renal fibrosis in fibrotic kidneys. Moreover, genetic deletion of polycystin-1 inhibited renal fibrosis in two different animal models. Finally, we showed that ablation of the methyltransferase EZH2 mitigated the anti-fibrotic effect of polycystin-1 deletion in fibrotic kidneys. We conclude that polycystin promotes through EZH2 and renal fibrosis in cystic kidney is not the direct effect of PKD gene mutations. Protective host-dependent antagonism among 	Pseudomonas in the Arabidopsis phyllosphere The plant microbiome is a rich biotic environment, comprising numerous taxa. The community structure of these colonizers is constrained by multiple factors, including host-microbe and microbe-microbe interactions, as well as the interplay between the two. While much can be learned from pairwise relationships between individual hosts and microbes, or individual microbes with themselves, the ensemble of interrelations between the host and microbial consortia may lead to different outcomes that are not easily predicted from the individual interactions. Their study can thus provide new insights into the complex relationship between plants and microbes. Of particular importance is how strain-specific such plant-microbe-microbe interactions are, and how they eventually affect plant health. Here, we test strain-level interactions in the phyllosphere between groups of co-existing commensal and pathogenic Pseudomonas among each other and with A. thaliana, by employing synthetic communities of genome-barcoded isolates. We found that commensal Pseudomonas prompted a host response leading to a selective inhibition of a specific pathogenic lineage, resulting in plant protection. The extent of plant protection, however, was dependent on plant genotype, indicating that these effects are host-mediated. There were similar genotype-specific effects on the microbe side, as we could pinpoint an individual Pseudomonas isolate as the predominant cause for this differential interaction. Collectively, our work highlights how within-species genetic differences on both the host and microbe side can have profound effects on host-microbe-microbe dynamics. The paradigm that we have established provides a platform for the study of host-dependent microbe-microbe competition and cooperation in the A. thaliana-Pseudomonas system. The Functional Nanopore Screen: A Versatile High-throughput Assay to Study and Engineer Protein Nanopores in Escherichia coli Nanopores comprise a versatile class of membrane proteins that carry out a range of key physiological functions and are increasingly developed for different biotechnological applications. Yet, a capacity to study and engineer protein nanopores by combinatorial means has so far been hampered by a lack of suitable assays that combine sufficient experimental resolution with throughput. Addressing this technological gap, the Functional Nanopore (FuN) screen now provides a quantitative and dynamic read-out of nanopore assembly and function in E. coli. The assay is based on genetically-encoded fluorescent protein sensors that resolve the nanopore-dependent influx of Ca2+ across the inner membrane of E. coli. Illustrating its versatile capacity, the FuN screen is first applied to dissect the molecular features that underlie the assembly and stability of nanopores formed by the S2168 holin. In a subsequent step, nanopores are engineered by recombining the transmembrane module of S2168 with different ring-shaped oligomeric protein structures that feature defined hexa-, hepta- and octameric geometries. Library screening highlights substantial plasticity in the ability of the S2168 transmembrane module to oligomerize in alternative geometries while the functional properties of the resultant nanopores can be fine-tuned through the identity of the connecting linkers. Overall, the FuN screen is anticipated to facilitate both fundamental studies and complex nanopore engineering endeavors with many potential applications in biomedicine, biotechnology and synthetic biology. Multiscale mechanistic modelling of the host defense in invasiveaspergillosis reveals leukocyte activation and iron acquisition as driversof infection outcome Aspergillus species are ubiquitous environmental molds, the spores of which are inhaled daily by most humans. Immunocompromised hosts can develop an invasive infection resulting in high mortality. There is, therefore, a pressing need for host-centric therapeutics for this infection. To address this need, we created a multi-scale computational model of the infection, focused on its interaction with the innate immune system and iron, a critical nutrient for the pathogen. The model, parameterized using published data, was found to recapitulate a wide range of biological features and was experimentally validated in vivo. Conidial swelling was identified as critical in fungal strains with high growth, whereas the siderophore secretion rate is an essential prerequisite for establishment of the infection in low-growth strains. In immunocompetent hosts, high growth, high swelling probability, and impaired leukocyte activation lead to a high conidial germination rate. Similarly, in neutropenic mice, high fungal growth was achieved through synergy between high growth rate, high swelling probability, slow leukocyte activation, and high siderophore secretion. In summary, the model reveals a small set of parameters related to fungal growth, iron acquisition, and leukocyte activation as key determinants of the fate of the infection. Conserved organ-specific microbial assemblages in different populations of a terrestrial crab Brachyuran crabs originated in the oceans and evolved specific morphological and physiological adaptations to live in freshwater, intertidal and even terrestrial habitats but the role of a selection mechanism involving symbiotic microorganisms long these colonization processes are not known. In this work we investigated the associated microbiota of three populations of a terrestrial brachyuran crab, Chiromantes haematocheir, to find evidence of a conserved crab-specific microbiome unrelated to the population of origin and dissimilar from environmental microbial assemblages. Bacterial 16S rRNA gene and fungal ITS sequences were obtained from selected crab organs and environmental matrices to profile microbial communities. In spite of the presence of truly marine larval stages and no gregarious behaviour, favouring microbiota exchanges, we found common, organ-specific microbiota, associated to the gut and the gills of the crabs (with more than 15% of the genera detected specifically enriched only in one organ). Our results suggest an early establishment of a new common, stable microbiota in the transition from water to land.