Emergent selectivity for scenes, object properties, and contour statistics in feedforward models of scene-preferring cortex The scene-preferring portion of the human ventral visual stream, known as the parahippocampal place area (PPA), responds to scenes and landmark objects, which tend to be large in real-world size, fixed in location, and inanimate. However, the PPA also exhibits preferences for low-level contour statistics, including rectilinearity and cardinal orientations, that are not directly predicted by theories of scene- and landmark-selectivity. It is unknown whether these divergent findings of both low- and high-level selectivity in the PPA can be explained by a unified computational theory. To address this issue, we fit feedforward computational models of visual feature coding to the image-evoked fMRI responses of the PPA, and we performed a series of high-throughput experiments on these models. Our findings show that feedforward models of the PPA exhibit emergent selectivity across multiple levels of complexity, giving rise to seemingly high-level preferences for scenes and for objects that are large, spatially fixed, and inanimate/manmade while simultaneously yielding low-level preferences for rectilinear shapes and cardinal orientations. These results reconcile disparate theories of PPA function in a unified model of feedforward feature coding, and they demonstrate how multifaceted selectivity profiles naturally emerge from the feedforward computations of visual cortex and the natural statistics of images.

SIGNIFICANCE STATEMENTVisual neuroscientists characterize cortical selectivity by identifying stimuli that drive regional responses. A perplexing finding is that many higher-order visual regions exhibit selectivity profiles spanning multiple levels of complexity: they respond to highly complex categories, such as scenes and landmarks, but also to surprisingly simplistic features, such as specific contour orientations. Using large-scale computational analyses and human brain imaging, we show how multifaceted selectivity in scene-preferring cortex can emerge from the feedforward, hierarchical coding of visual features. Our work reconciles seemingly divergent findings of selectivity in scene-preferring cortex and suggests that surprisingly simple feedforward feature representations may be central to the category-selective organization of the human visual system.
Lack of Cdk5 activity is involved on Dopamine Transporter expression and function: Evidences from an animal model of Attention-Deficit Hyperactivity Disorder Attention deficit/Hyperactivity disorder (ADHD) is one of the most diagnosed psychiatric disorders nowadays. The core symptoms of the condition include hyperactivity, impulsiveness and inattention. The main pharmacological treatment consists of psychostimulant drugs affecting Dopamine Transporter (DAT) function. We have previously shown that genetically modified mice lacking p35 protein (p35KO), which have reduced Cdk5 activity, present key hallmarks resembling those described in animal models useful for studying ADHD. The p35KO mouse displays spontaneous hyperactivity and shows a calming effect of methylphenidate or amphetamine treatment. Interestingly, dopaminergic neurotransmission is altered in these mice as they have an increased Dopamine (DA) content together with a low DA turnover. This led us to hypothesize that the lack of Cdk5 activity affects DAT expression and/or function in this animal model. In this study, we performed biochemical assays, cell-based approaches, quantitative fluorescence analysis and functional studies that allowed us to demonstrate that p35KO mice exhibit decreased DA uptake and reduced cell surface DAT expression levels in the striatum (STR). These findings are supported by in vitro observations in which the inhibition of Cdk5 activity in N2a cells induced a significant increase in constitutive DAT endocytosis with a concomitant increase in DAT localization to recycling endosomes. Taken together, these data provide evidences regarding the role of Cdk5/p35 in DAT expression and function, thus contributing to the knowledge of DA neurotransmission physiology and also providing therapeutic options for the treatment of DA pathologies such as ADHD.
Comparative immunogenicity of bacterially expressed soluble trimers and nanoparticle displayed influenza hemagglutinin stem immunogens Current influenza vaccines need to be updated annually due to mutations in the globular head of the viral surface protein, hemagglutinin (HA). To address this, vaccine candidates have been designed based on the relatively conserved HA stem domain and have shown protective efficacy in animal models. Oligomerization of the antigens either by fusion to oligomerization motifs or display on self-assembling nanoparticle scaffolds, can induce more potent immune responses compared to the corresponding monomeric antigen due to multivalent engagement of B-cells. Since nanoparticle display can increase manufacturing complexity, and often involves one or more mammalian cell expressed components, it is important to characterize and compare various display and oligomerization scaffolds. Using a structure guided approach, we successfully displayed multiple copies of a previously designed soluble, trimeric influenza stem domain immunogen, pH1HA10, on the ferritin like protein, MsDps2 (12 copies), Ferritin (24 copies) and Encapsulin (180 copies). All proteins were expressed in Escherichia coli. The nanoparticle fusion immunogens were found to be well folded and bound to the influenza stem directed broadly neutralizing antibodies with high affinity. An 8.5 [A] Cryo-EM map of Msdps2-pH1HA10 confirmed the successful design of the nanoparticle fusion immunogen. Mice immunization studies with the soluble trimeric stem and nanoparticle fusion constructs revealed that all of them were immunogenic, and protected mice against homologous (A/Belgium/145-MA/2009) and heterologous (A/Puerto Rico/8/1934) challenge with 10MLD50 mouse adapted virus. Although nanoparticle display conferred a small but statistically significant improvement in protection relative to the soluble trimer in a homologous challenge, heterologous protection was similar in both nanoparticle-stem immunized and trimeric stem immunized groups. Such rapidly producible, bacterially expressed antigens and nanoparticle scaffolds are useful modalities to tackle future influenza pandemics. Glucose inhibits haemostasis and accelerates diet-induced hyperlipidaemia in zebrafish larvae Hyperglycaemia damages the microvasculature in part through the reduced recruitment of immune cells and interference with platelet signalling, leading to poor wound healing and accelerated lipid deposition in mammals. We investigated the utility of zebrafish larvae to model the effect of glucose on neutrophil and macrophage recruitment to a tail wound, wound-induced haemostasis, and chicken egg yolk feed challenge-induced hyperlipidaemia by supplementing larvae with exogenous glucose by immersion or injection. Neither method of glucose supplementation affected the recruitment of neutrophils and macrophages following tail transection. Glucose injection reduced thrombocyte retention and fibrin plug formation while only thrombocyte retention was reduced by glucose immersion following tail transection. We observed accelerated lipid accumulation in glucose-injected larvae challenged with high fat chicken egg yolk feeding. Our study identifies conserved and divergent effects of high glucose on inflammation, haemostasis, and hyperlipidaemia in zebrafish larvae compared to mammals. Trypanosomal variant surface glycoprotein expression in human African trypanosomiasis patients Trypanosoma brucei gambiense, an extracellular protozoan parasite, is the primary causative agent of human African trypanosomiasis. T. b. gambiense is endemic to West and Central Africa, where it is transmitted by the bite of infected tsetse flies. In the bloodstream of an infected host, the parasite evades antibody recognition by altering the Variant Surface Glycoprotein (VSG) that forms a dense coat on its cell surface through a process known as antigenic variation. Each VSG has a variable N-terminal domain exposed to the host and a less variable C-terminal domain that is partially hidden from host antibodies. Our lab developed VSG-seq, a targeted RNA-seq method, to study VSG gene expression in T. brucei. Studies using VSG-seq to characterize antigenic variation in a mouse model have revealed marked diversity in VSG expression within parasite populations, but this finding has not yet been validated in a natural human infection. Here, we used VSG-seq to analyze VSGs expressed in the blood of 12 patients infected with T. b. gambiense. The number of VSGs identified per patient ranged from one to 14; notably, two VSGs were shared by more than one patient. Analysis of expressed VSG N-terminal domain types revealed that 82% of expressed VSGs encoded a type B N-terminus, a bias not seen in datasets from other T. brucei subspecies. C-terminal types in T. b. gambiense infection were also restricted. These results demonstrate a bias either in the underlying VSG repertoire of T. b. gambiense or in this subspecies expression of VSGs during infection. This work demonstrates the feasibility of using VSG-seq to study antigenic variation in human infections and highlights the importance of understanding VSG repertoires in the field.

Author SummaryHuman African Trypanosomiasis is a neglected tropical disease primarily caused by the extracellular parasite Trypanosoma brucei gambiense. To avoid elimination by the host, these parasites repeatedly replace their dense surface coat of Variant Surface Glycoprotein (VSG). Despite the important role of VSGs in prolonging infection, VSG expression during natural human infections is poorly understood. A better understanding of natural VSG gene expression dynamics can clarify the mechanisms that T. brucei uses to alter its VSG coat and improve how trypanosomiasis is diagnosed in humans. We analyzed the expressed VSGs detected in the blood of patients with trypanosomiasis. Our findings indicate that parasites in both natural and experimental infections express diverse sets of VSGs. Modulations in motor unit discharge are related to changes in fascicle length during isometric contractions The integration of electromyography (EMG) and ultrasound imaging has provided important information about the mechanisms of muscle activation and contraction. Unfortunately, EMG does not allow an accurate assessment of the interplay between the neural drive received by muscles, changes in fascicle length (FL) and the force/torque produced. We aimed to assess the relationship between modulations in tibialis anterior (TA) motor unit (MU) firing rate, FL and dorsiflexion torque (DT) using ultrasound-transparent high-density EMG electrodes. EMG and ultrasound images were recorded simultaneously from TA, using a 32-electrode silicon matrix, while performing isometric dorsiflexion, at diverse ankle joint positions (0{degrees} and 30{degrees} plantar flexion) and torques (20% and 40% of maximum). EMG signals were decomposed into individual MUs and changes in FL were assessed with a fascicle-tracking algorithm. MU firings were converted into a cumulative spike train (CST) that was cross-correlated with DT (CST-DT) and FL (CST-FL). High cross-correlations were found for CST-FL, 0.60 (range: 0.31-0.85) and CST-DT 0.71 (range: 0.31-0.88). Cross-correlation lags revealed that the delay between CST-FL (~75ms) was significantly smaller than CST-DT (~150ms, p<0.001). These delays affected the interpretation of MU recruitment/de-recruitment thresholds, with FL showing similar lengths for both recruitment and de-recruitment. This study is the first to demonstrate that changes in TA FL are closely related to both modulations in MU firing frequency and DT. These relationships allow assessment of the interplay between neural drive, muscle contraction and resultant torque, thereby providing a better understanding of the mechanisms responsible for the generation of muscle force.

NEW AND NOTEWORTHYBy employing ultrasound-transparent high-density surface EMG electrodes, we show that modulations in tibialis anterior motor unit discharge rate were closely related to both changes in its fascicle length and resultant torque. These relationships allowed quantifying delays between neural drive and muscle shortening as well as muscle shortening and torque during submaximal isometric contractions, providing an accurate estimation of the time required to generate muscle force and subsequent production of torque via the tendon. Classification of daily crop phenology in PhenoCams using deep learning and hidden markov models Near surface cameras, such as those in the PhenoCam network, are a common source of ground truth data in modelling and remote sensing studies. Despite having locations across numerous agricultural sites, few studies have used near surface cameras to track the unique phenology of croplands. Due to management activities, crops do not have a natural vegetation cycle which many phenological extraction methods are based on. For example, a field may experience abrupt changes due to harvesting and tillage throughout the year. A single camera can also record several different plants due to crop rotations, fallow fields, and cover crops. Current methods to estimate phenology metrics from image time series compress all image information into a relative greenness metric, which discards a large amount of contextual information. This can include the type of crop present, whether snow or water is present on the field, the crop phenology, or whether a field lacking green plants consists of bare soil, fully senesced plants, or plant residue. Here we developed a modelling workflow to create a daily time series of crop type and phenology, while also accounting for other factors such as obstructed images and snow covered fields. We used a mainstream deep learning image classification model, VGG16. Deep learning classification models do not have a temporal component, so to account for temporal correlation among images our workflow incorporates a hidden markov model in the post-processing. The initial image classification model had out of sample F1 scores of 0.83-0.85, which improved to 0.86-0.91 after all post-processing steps. The resulting time series show the progression of crops from emergence to harvest, and can serve as a daily, local scale dataset of field states and phenological stages for agricultural research. pkgndep: a tool for heaviness analysis on R package dependencies SummaryNumerous R packages have been developed for bioinformatics analysis in the last decade and dependencies among packages have become critical issues to consider. In this work, we proposed a new metric named dependency heaviness that measures the number of unique dependencies that a parent brings to a package, and we proposed possible solutions for reducing the complexity of dependencies by optimizing the use of heavy parents. We implemented the metric in a new R package pkgndep which provides an intuitive way for dependency heaviness analysis. Based on pkgndep, we additionally performed a global analysis of dependency heaviness on CRAN and Bioconductor ecosystems and we revealed top packages that have significant contributions of high dependency heaviness to their child packages.

Availability and implementationThe package pkgndep and documentations are freely available from the Comprehensive R Archive Network (CRAN) https://cran.r-project.org/package=pkgndep. The dependency heaviness analysis for all 21,741 CRAN and Bioconductor packages retrieved on 2021-10-28 are available at https://pkgndep.github.io/. The Rab GTPase activating protein TBC-2 regulates endosomal localization of DAF-16 FOXO and lifespan FOXO transcription factors have been shown to regulate longevity in model organisms and are associated with longevity in humans. To gain insight into how FOXO functions to increase lifespan, we examined the subcellular localization of DAF-16 in C. elegans. We show that DAF-16 is localized to endosomes and that this endosomal localization is increased by the insulin-IGF signaling (IIS) pathway. Endosomal localization of DAF-16 is modulated by endosomal trafficking proteins. Disruption of the Rab GTPase activating protein TBC-2 increases endosomal localization of DAF-16, while inhibition of TBC-2 targets, RAB-5 or RAB-7 GTPases, decreases endosomal localization of DAF-16. Importantly, the amount of DAF-16 that is localized to endosomes has functional consequences as increasing endosomal localization through mutations in tbc-2 reduced the lifespan of long-lived daf-2 IGFR mutants, depleted their fat stores, and DAF-16 target gene expression. Overall, this work identifies endosomal localization as a mechanism regulating DAF-16 FOXO, which is important for its functions in metabolism and aging. 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. Data-Driven Network Models for Genetic Circuits From Time-Series Data with Incomplete Measurements Synthetic gene networks are frequently conceptualized and visualized as static graphs. This view of biological programming stands in stark contrast to the transient nature of biomolecular interaction, which is frequently enacted by labile molecules that are often unmeasured. Thus, the network topology and dynamics of synthetic gene networks can be difficult to verify in vivo or in vitro, due to the presence of unmeasured biological states. Here we introduce the dynamical structure function as a new mesoscopic, data-driven class of models to describe gene networks with incomplete measurements. We introduce a network reconstruction algorithm and a code base for reconstructing the dynamical structure function from data, to enable discovery and visualization of graphical relationships in a genetic circuit diagram as time-dependent functions rather than static, unknown weights. We prove a theorem, showing that dynamical structure functions can provide a data-driven estimate of the size of crosstalk fluctuations from an idealized model. We illustrate this idea with numerical examples. Finally, we show how data-driven estimation of dynamical structure functions can explain failure modes in two experimentally implemented genetic circuits, a historical genetic circuit and a new E. coli based transcriptional event detector. Microbial succession on degrading micropollutants under recurrent exposure The increasing-volume release of micropollutants into natural surface waters has raised great concern due to their environmental accumulation. Persisting micropollutants can impact multiple generations of organisms, but their microbially-mediated degradation and their influence on community assembly remain understudied. Here, freshwater microbes were treated with several common micropollutants, alone or in combination, and then transferred every 5 days to fresh medium containing the same micropollutants to mimic the repeated exposure of microbes. Metabarcoding of 16S rRNA gene makers was chosen to study the succession of bacterial assemblages following micropollutant exposure. The removal rates of micropollutants were then measured to assess degradation capacity of the associated communities. The degradation of micropollutants did not accelerate over time but altered the microbial community composition. Community assembly was dominated by stochastic processes during early exposure, via random community changes and emergence of seedbanks, and deterministic processes later in the exposure, via advanced community succession. Early exposure stages were characterized by the presence of sensitive microorganisms such as Actinobacteria and Planctomycetes, which were then replaced by more tolerant bacteria such as Bacteroidetes and Gammaproteobacteria. Our findings have important implication for ecological feedback between microbe-micropollutants under anthropogenic climate change scenarios. A genomic assessment of the marine-speciation paradox within the toothed whale superfamily Delphinoidea The importance of post-divergence gene flow in speciation has been documented across a range of taxa in recent years, and may have been especially widespread in highly mobile, wide-ranging marine species, such as cetaceans. Here, we studied individual genomes from nine species across the three families of the toothed whale superfamily Delphinoidea (Delphinidae, Phocoenidae, Monodontidae). To investigate the role of post-divergence gene flow in the speciation process, we used a multifaceted approach, including: (i) phylogenomics, (ii) the distribution of shared derived alleles, and (iii) demographic inference. We found the divergence of lineages within Delphinoidea did not follow a process of pure bifurcation, but was much more complex. Sliding-window phylogenomics reveal a high prevalence of discordant topologies within the superfamily, with further analyses indicating these discordances arose due to both incomplete lineage sorting and gene flow. D-statistics, D-foil, and f-branch analyses supported gene flow between members of Delphinoidea, with the vast majority of gene flow occurring as ancient interfamilial events. Demographic analyses provided evidence that introgressive gene flow has likely ceased between all species pairs tested, despite reports of contemporary interspecific hybrids. Our study provides the first steps towards resolving the large complexity of speciation within Delphinoidea; we reveal the prevalence of ancient interfamilial gene flow events prior to the diversification of each family, and suggests that contemporary hybridisation events may be disadvantageous, as hybrid individuals do not appear to contribute to the parental species gene pools. Naming of human diseases on the wrong side of history BackgroundIn the medical sphere, understanding naming conventions strengthen the integrity of naming human diseases remains nominal rather than substantial yet. Since the current nosology-based standard for human diseases could not offer a one-size-fits-all corrective mechanism, many idiomatic but flawed names frequently appear in scientific literature and news outlets at the cost of sociocultural impacts.

ObjectiveWe attempt to examine the ethical oversights of current naming practices and propose heuristic rationales and approaches to determine a pithy name instead of an inopportune nosology.

MethodsFirst, we examined the compiled global online news volumes and emotional tones on some inopportune nosology like German measles, Middle Eastern Respiratory Syndrome, Spanish flu, Hong Kong flu, and Huntingtons disease in the wake of COVID-19. Second, we prototypically scrutinize the lexical dynamics and pathological differentials of German measles and common synonyms by leveraging the capacity of the Google Books Ngram Corpus. Third, we demonstrated the empirical approaches to curate an exclusive substitute for an anachronistic nosology German measles based on deep learning models and post-hoc explanations.

ResultsThe infodemiological study shows that the public informed the offensive names with extremely negative tones in textual and visual narratives. The findings of the historiographical study indicate that many synonyms of German measles did not survive, while German measles became an anachronistic usage, and rubella has taken the dominant place since 1994. The PubMedBERT model could identify rubella as a potential substitution for German measles with the highest semantic similarity. The results of the semantic drift experiments further indicate that rubella tends to survive during the ebb and flow of semantic drift.

ConclusionsOur findings indicate that the nosological evolution of anachronistic names could result in sociocultural impacts without a corrective mechanism. To mitigate such impacts, we introduce some ethical principles for formulating an improved naming scheme. Based on deep learning models and post-hoc explanations, our illustrated experiments could provide hallmark references to the remedial mechanism of naming practices and pertinent credit allocations. Sgs1BLM independent role of Dna2DNA2 nuclease at DNA double strand break is inhibited by Nej1XLF The two major pathways of DNA double strand break (DSB) repair, non-homologous end-joining (NHEJ) and homologous recombination (HR), are highly conserved from yeast to mammals. The regulation of 5 DNA resection controls repair pathway choice and influences repair outcomes. Nej1 was first identified as a canonical NHEJ factor involved in stimulating the ligation of broken DNA ends and more recently, it was shown to be important for DNA end-bridging and inhibiting 5 resection mediated by Dna2-Sgs1. Nej1 interacts with Sae2 and this impacts DSB repair in three ways. First, Nej1 inhibits MRX-Sae2 interactions and Sae2 localization to a DSB. Second, Nej1 inhibits Sae2-dependent recruitment of Dna2 in the absence of Sgs1. Third, NEJ1 and SAE2 showed an epistatic relationship for DNA end-bridging, an event that restrains the broken ends and reduces the frequency of genomic deletions from developing at the DSB. Deletion of NEJ1 suppressed the synthetic lethality of sae2{Delta} sgs1{Delta} and was dependent on the nuclease activity of Dna2. These Nej1 functions promote end-joining DSB repair, but could also be relevant for controlling resection initiation during HR repair.

HighlightsO_LINej1 physically interacts with Sae2 and inhibits end-resection at a DSB.
C_LIO_LINej1 inhibits Sae2 interactions with the MRX complex.
C_LIO_LINej1 inhibits Sae2-dependent recruitment of Dna2 to a DSB.
C_LIO_LINEJ1 and SAE2 are epistatic for DNA end-bridging.
C_LIO_LIDeletion of NEJ1 suppresses the synthetic lethality of sae2{Delta} sgs1{Delta}, which is dependent on Dna2 nuclease activity.
C_LI 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. Comparative Analysis of non-coding small-RNAs in Pseudomonas aeruginosa Keratitis Strains with Different Antibiotic Susceptibility Pseudomonas aeruginosa, is a gram-negative bacterium causes opportunistic or nosocomial infections in immunocompromised individuals. In recent years, a steady increase in human corneal infections of P. aeruginosa has been reported with increased multi-drug resistance (MDR) or extensively drug resistance (XDR). Several non-coding sRNAs, has been identified to regulate various physiological processes in P. aeruginosa, including biofilm formation, quorum sensing. However, the regulatory mechanism of sRNAs in MDR/XDR pathways of P. aeruginosa keratitis strains is not yet studied. In this study, we identified bacterial sRNAs in publicly available P. aeruginosa keratitis genomes and investigated their regulatory role in MDR/XDR pathways using bioinformatic analysis. Totally, 46 P. aeruginosa keratitis strains from different geographical regions were included. Of 46, Eight (30%) out of Twenty-seven and Nine (52) out of Nineteen P. aeruginosa strains from India and Australia were identified as not-MDR. Whereas, 10 (38%) Indian and 9 (47%) Australian strains were identified as MDR. Eight Indian strains were identified as XDR. Out of 46 strains, 23 (50%) carried ExoU, 21(45%) carried ExoS and two (5%) strains carried both ExoU and ExoS, exotoxins for their virulence. The sRNA, SPA0021 was identified in 18 MDR/XDR and 6 not-MDR strains along with UCBPP-PA14. Interestingly, majority of the imipenem resistant P. aeruginosa keratitis strains from the present study was found to be carried SPA0023 sRNA (18 out of 30 strains). The outer membrane porin protein OprD, identified as binding target of SPA0023. Negative regulation or inactivation of OprD, reported in increased imipenem resistance in P. aeruginosa. Mutation analysis revealed that SPA0023 carrying P. aeruginosa keratitis strains contains a lesser number of amino acid changes in OprD protein than other strains. These findings indicate, imipenem resistance in SPA0023 carried strains might arose from the negative regulation or inhibition of OprD by SPA0023. However, functional studies are warranted with large number of P. aeruginosa keratitis strains to confirm the negative regulation of OprD by SPA0023 and imipenem resistance. Improved radiation expression profiling in blood by sequential application of sensitive and specific gene signatures PurposeCombinations of expressed genes can discriminate radiation-exposed from normal control blood samples by machine learning based signatures (with 8 to 20% misclassification rates). These signatures can quantify therapeutically-relevant as well as accidental radiation exposures. The prodromal symptoms of Acute Radiation Syndrome (ARS) overlap those present in Influenza and Dengue Fever infections. Surprisingly, these human radiation signatures misclassified gene expression profiles of virally infected samples as false positive exposures. The present study investigates these and other confounders, and then mitigates their impact on signature accuracy.

MethodsThis study investigated recall by previous and novel radiation signatures independently derived from multiple Gene Expression Omnibus datasets on common and rare non-malignant blood disorders and blood-borne infections (thromboembolism, S. aureus bacteremia, malaria, sickle cell disease, polycythemia vera, and aplastic anemia). Normalized expression levels of signature genes are used as input to machine learning-based classifiers to predict radiation exposure in other hematological conditions.

ResultsExcept for aplastic anemia, these blood-borne disorders modify the normal baseline expression values of genes present in radiation signatures, leading to false-positive misclassification of radiation exposures in 8 to 54% of individuals. Shared changes, predominantly in DNA damage response and apoptosis-related gene transcripts in radiation and confounding hematological conditions, compromise the utility of these signatures for radiation assessment. These confounding conditions (sickle cell disease, thromboembolism, S. aureus bacteremia, malaria) induce neutrophil extracellular traps, initiated by chromatin decondensation, DNA damage response and fragmentation followed by programmed cell death. Riboviral infections (for example, Influenza or Dengue fever) have been proposed to bind and deplete host RNA binding proteins, inducing R-loops in chromatin. R-loops that collide with incoming replication forks can result in incompletely repaired DNA damage, inducing apoptosis and releasing mature virus. To mitigate the effects of confounders, we evaluated predicted radiation-positive samples with novel gene expression signatures derived from radiation-responsive transcripts encoding secreted blood plasma proteins whose expression levels are unperturbed by these conditions.

ConclusionsThis approach identifies and eliminates misclassified samples with underlying hematological or infectious conditions, leaving only samples with true radiation exposures. Diagnostic accuracy is significantly improved by selecting genes that maximize both sensitivity and specificity in the appropriate tissue using combinations of the best signatures for each of these classes of signatures. Flexible reprogramming of Pristionchus pacificus motivation for attacking Caenorhabditis elegans in predator-prey competition Animals with diverse diets must adapt their food priorities to a wide variety of environmental conditions. This diet optimization problem is especially complex for predators that compete with prey for food. Although predator-prey competition is widespread and ecologically critical, it remains difficult to disentangle predatory and competitive motivations for attacking competing prey. Here, we dissect the foraging decisions of the omnivorous nematode Pristionchus pacificus to reveal that its seemingly failed predatory attempts against Caenorhabditis elegans are actually motivated acts of efficacious territorial aggression. While P. pacificus easily kills and eats larval C. elegans with a single bite, adult C. elegans typically survives and escapes from bites. However, nonfatal biting can provide competitive benefits by reducing access of adult C. elegans and its progeny to bacterial food that P. pacificus also eats. We show that P. pacificus considers costs and benefits of both predatory and territorial outcomes to decide which food goal, prey or bacteria, should guide its motivation for biting. These predatory and territorial motivations impose different sets of rules for adjusting willingness to bite in response to changes in bacterial abundance. In addition to biting, predatory and territorial motivations also influence which search tactic P. pacificus uses to increase encounters with C. elegans. When treated with an octopamine receptor antagonist, P. pacificus switches from territorial to predatory motivation for both biting and search. Overall, we demonstrate that P. pacificus assesses alternate outcomes of attacking C. elegans and flexibly reprograms its foraging strategy to prioritize either prey or bacterial food. Heterogeneous but not random: Cargo degradation in phagosomes is kinetically coupled to 1 their intracellular mobility Immune cells degrade internalized pathogens in phagosomes through sequential biochemical changes. The degradation must be fast enough for effective infection control. The presumption is that each phagosome degrades cargos autonomously with a distinct but stochastic kinetic rate. Here we report that the degradation kinetics of individual phagosomes is not stochastic but coupled to their intracellular motility. By engineering RotSensors that are optically anisotropic, magnetic responsive, and fluorogenic in response to degradation activities in phagosomes, we monitored cargo degradation kinetics in single phagosomes simultaneously with their translational and rotational dynamics. We show that phagosomes that move faster centripetally are more likely to encounter and fuse with lysosomes, thereby acidifying faster and degrading cargos more efficiently. The degradation rates increase nearly linearly with the translational and rotational velocities of phagosomes. Our results indicate that the centripetal motion of phagosomes functions as a clock for controlling the progression of cargo degradation. Pore dynamics and asymmetric cargo loading in an encapsulin nanocompartment Encapsulins are protein nanocompartments that house various cargo enzymes, including a family of decameric ferritin-like proteins. Here, we study a recombinant Haliangium ochraceum encapsulin:encapsulated ferritin complex using electron cryo-microscopy and hydrogen/deuterium exchange mass spectrometry to gain insight into the structural relationship between the encapsulin shell and its protein cargo. An asymmetric single particle reconstruction reveals four encapsulated ferritin decamers in a tetrahedral arrangement within the encapsulin nanocompartment. This leads to a symmetry mismatch between the protein cargo and the icosahedral encapsulin shell. The encapsulated ferritin decamers are offset from the interior face of the encapsulin shell. Using HDX-MS, we observed dynamic behavior of the major five-fold pore in the encapsulin shell and show the pore opening via the movement of the encapsulin A-domain. These data will accelerate efforts to engineer the encapsulation of heterologous cargo proteins and to alter the permeability of the encapsulin shell via pore modifications.

TeaserCryo-EM and HDX-MS analysis of an encapsulin nanocompartment shows that the pores at the five-fold icosahedral vertex of the shell are flexible. OrganoID: a versatile deep learning platform for tracking and analysis of single-organoid dynamics Organoids have immense potential as ex vivo disease models for drug discovery and personalized drug screening. Dynamic changes in individual organoid morphology, number, and size can indicate important drug responses, however these metrics are difficult and labor-intensive to obtain for high-throughput image datasets. Here, we present OrganoID, a robust image analysis platform that automatically recognizes, labels, and tracks single organoids, pixel-by-pixel, in brightfield and phase-contrast microscopy experiments. The platform was trained on images of pancreatic cancer organoids and validated on separate images of pancreatic, lung, colon, and adenoid cystic carcinoma organoids, which showed excellent agreement with manual measurements of organoid count (96%) and size (95%) without any parameter adjustments. Single-organoid tracking accuracy remained above 89% over a four-day time-lapse microscopy study. Automated single-organoid morphology analysis of a chemotherapy dose-response experiment identified decreased organoid circularity as an important morphological feature reflecting drug response. OrganoID enables straightforward, detailed, and accurate image analysis to accelerate the use of organoids in high-throughput, data-intensive biomedical applications. N-VEGF, the autoregulatory arm of VEGF-A Vascular endothelial growth factor A (VEGF-A) is a secreted protein that stimulates angiogenesis in response to hypoxia. Under hypoxic conditions, a non-canonical long isoform called L-VEGF is concomitantly expressed with VEGF-A. Once translated, L-VEGF it is proteolytically cleaved to generate N-VEGF and VEGF-A. Interestingly, while VEGF-A is secreted and affects the surrounding cells, N-VEGF is mobilized to the nucleus. This suggests that N-VEGF participates in transcriptional response to hypoxia. In this study, we performed a series of complementary experiments to examine the functional role of N-VEGF. Strikingly, we found that the mere expression of N-VEGF followed by its hypoxia-independent mobilization to the nucleus was sufficient to induce key genes associated with angiogenesis, such as Hif1, VEGF-A isoforms, as well as genes associated with cell survival under hypoxia. Complementarily, when N-VEGF was genetically depleted, key hypoxia-induced genes were downregulated and cells were significantly susceptible to hypoxia-mediated apoptosis. This is the first reports of N-VEGF serving as an autoregulatory arm of VEGF-A. Further experiments will be needed to determine the role of N-VEGF in cancer and embryogenesis. Coupled myovascular expansion directs cardiac growth and regeneration BackgroundHeart regeneration requires multiple cell types to enable cardiomyocyte (CM) proliferation. How these cells interact to create growth niches is unclear. Here we profile proliferation kinetics of cardiac endothelial cells (CECs) and CMs in the neonatal mouse heart and find that they are spatiotemporally coupled.

Methods and ResultsWe show that coupled myovascular expansion during cardiac growth or regeneration is dependent upon VEGF-VEGFR2 signaling, as genetic deletion of Vegfr2 from CECs or inhibition of VEGFA abrogates both CEC and CM proliferation. Repair of cryoinjury displays poor spatial coupling of CEC and CM proliferation. Boosting CEC density after cryoinjury with virus encoding Vegfa enhances regeneration. Using Mendelian randomization, we demonstrate that circulating VEGFA levels are positively linked with human myocardial mass, suggesting that Vegfa can stimulate human cardiac growth.

ConclusionsOur work demonstrates the importance of coupled CEC and CM expansion and reveals a myovascular niche that may be therapeutically targeted for heart regeneration. A General Biphasic-Bodyweight Model for Scaling Basal Metabolic Rate, Glomerular Filtration Rate and Drug Clearance from Birth to Adulthood Understanding the maturation process of human physiology and metabolism has broad medical and pharmaceutical implications. Age and bodyweight are frequently considered as separate variables in modeling the dynamical changes of human organ functions and of drug clearance from birth to adulthood. The objective of this study is to propose a unified, continuous and bodyweight-only equation to quantify the changes of human basal metabolic rate (BMR), glomerular filtration rate (GFR) and drug clearance (CL) from infancy to adulthood. The BMR datasets were retrieved from a comprehensive historical database of male and female subjects (0.02 to 64 years). The CL datasets for 17 drugs and the GFR dataset were generated from age-incorporated maturation-and-growth models with reported parameter values. The model used in the simulation is independent of the proposed model. A statistical approach was used to simulate the model generated CL and GFR data for a hypothetical population with 26 age groups (ranging from 0 to 20 years). Besides, individual CL data for one drug, and sparse PBPK-modeled CL values for two drugs were also included for analysis. A 4-parameter, mixed-allometry equation with two power-law functions of bodyweight was proposed and evaluated as a general model using nonlinear regression and dimensionless analysis. All datasets universally reveal biphasic curves with two distinct linear segments on log-log plots. Compared with simple allometry, the biphasic model fits satisfactorily to all datasets (based on Akaikes Information Criterion and residual plots). The biphasic equation consists of two reciprocal allometric terms that asymptotically determine the overall curvature. The fitting results show a superlinear scaling phase (slope >1; ca. 1.5 - 3.5) below the characteristic bodyweight at the phase transition; and above which, a sublinear scaling phase (slope <1; ca. 0.5 - 0.7) is evident. The phase-transition bodyweight is ranging from 5 to 20 kg (corresponding to 0.5 - 9 years) and the mean value is around 10 kg ([~]2 years) for all data sets. The dimensionless analysis generalizes, and offers quantitative realization of, the maturation and growth process. In conclusion, the proposed mixed-allometry equation is a generic model that quantitatively describes the phase transition occurring in the human maturation process of BMR, GFR and drug CL. Decreased cell stiffness facilitates detachment and migration of breast cancer cells in 3D collagen matrices: An exploratory study BackgroundTumour-cell detachment is a critical early event in the metastatic cascade. However, the role of the cells mechanical properties in detachment and migration is not well understood. This exploratory study aimed to assess how intracellular stiffness changes these processes.

MethodsMDA-MB-231 cells were embedded as 10,000-cell spheroids in 2 and 4 mg/ml collagen matrices. Intracellular stiffness was assessed with mitochondria tracking microrheology of cells that migrated distances equivalent to four and six times the cell diameter (dC) from the spheroid and compared to cells at the spheroid surface (0dC), representing medium, high and no migration, respectively.

Results and discussionThe mitochondrial mean square displacement and intracellular stiffness decreased during detachment and migration for both collagen concentrations (i.e. rigidities). The mean square displacement of 4dC and 6dC cells was similar, whereas cell stiffness was lower for 4dC than for 6dC cells. With increasing matrix rigidity, the intracellular stiffness decreased for 0dC cells and did not change for 4dC and 6dC cells. It is proposed that decreased cell stiffness drives detachment and migration and increased matrix rigidity physically hinders migration, and cells need to become softer or remodel the environment to migrate. The independence of the stiffness of migrated cells from matrix rigidity suggests that cells remodel their environment through matrix proteins cleavage to migrate.

ConclusionsThe study revealed the collective effects of enhanced migratory conditions and increased matrix rigidity on the mechanical properties of the cells. The expression of matrix metalloproteinases and transforming growth factor {beta} and the role of cell volume on detachment and migration in matrices with varying pore sizes are proposed targets for further studies on metastatic cancer cells.