Correlation between speed and turning naturally arises for sparsely sampled cell movements Mechanisms regulating cell movement are not fully understood. One feature of cell movement that determines how far cells displace from an initial position is persistence, the ability to perform movements in a direction similar to the previous movement direction. Persistence is thus determined by turning angles between two sequential displacements and be characterized by an average turning angle or persistence time. Recent studies found that a cells average speed and turning are negatively correlated, suggesting a fundamental cell-intrinsic program whereby cells with a lower turning ability (i.e., larger persistence time) are intrinsically faster (or faster cells turn less). By simulating correlated or persistent random walks (PRWs) using two different frameworks (one based on von Mises-Fisher (vMF) distribution and another based on Ornstein-Uhlenbeck (OU) process) we show that the negative correlation between speed and turning naturally arises when cell trajectories are sub-sampled, i.e., when the frequency of sampling is lower than frequency at which cells make movements. This effect is strongest when the sampling frequency is on the order of magnitude with the typical cell persistence time and when cells vary in persistence time. Both conditions are observed for datasets of T cell movements in vivo that we have analyzed. In simulations the correlation arises due to randomness of cell movements resulting in highly variable persistence times for individual cells that with sub-sampling leads to large variability of average cell speeds. Interestingly, previously suggested methodology of calculating displacement of cell cohorts with different speeds resulted in similar results whether or not there is a cell-intrinsic correlation between cell speed and persistence. For both vMF- and OU-based simulations of PRWs we could find parameter values (distribution of persistence times, speeds, and sampling frequency) that matched experimentally measured correlations between speed and turning for two datasets of T cell movement in vivo suggesting that such simple correlations are not fully informative on the intrinsic link between speed and persistence. Our results thus suggest that sub-sampling may contribute to (and perhaps fully explains) the observed correlation between speed and turning at least for some cell trajectory data and emphasize the role of sampling frequency in inference of critical cellular parameters of cell motility such as speeds.

Secondary AbstractMeasurement of cell movements often results in a negative correlation between average speed and average turning angle suggesting an existence of a universal, cell-intrinsic movement program. We show that such a negative correlation may arise if cells in the population differ in their ability for persistent movement when the movement data are sub-sampled. We show that sub-sampling of cell trajectories generated using two different frameworks of persistent random walk can match the experimentally observed correlation between speed and turning for T cell movements in vivo.
Correlation between speed and turning naturally arises for sparsely sampled cell movements Mechanisms regulating cell movement are not fully understood. One feature of cell movement that determines how far cells displace from an initial position is persistence, the ability to perform movements in a direction similar to the previous movement direction. Persistence is thus determined by turning angles between two sequential displacements and be characterized by an average turning angle or persistence time. Recent studies found that a cells average speed and turning are negatively correlated, suggesting a fundamental cell-intrinsic program whereby cells with a lower turning ability (i.e., larger persistence time) are intrinsically faster (or faster cells turn less). By simulating correlated or persistent random walks (PRWs) using two different frameworks (one based on von Mises-Fisher (vMF) distribution and another based on Ornstein-Uhlenbeck (OU) process) we show that the negative correlation between speed and turning naturally arises when cell trajectories are sub-sampled, i.e., when the frequency of sampling is lower than frequency at which cells make movements. This effect is strongest when the sampling frequency is on the order of magnitude with the typical cell persistence time and when cells vary in persistence time. Both conditions are observed for datasets of T cell movements in vivo that we have analyzed. In simulations the correlation arises due to randomness of cell movements resulting in highly variable persistence times for individual cells that with sub-sampling leads to large variability of average cell speeds. Interestingly, previously suggested methodology of calculating displacement of cell cohorts with different speeds resulted in similar results whether or not there is a cell-intrinsic correlation between cell speed and persistence. For both vMF- and OU-based simulations of PRWs we could find parameter values (distribution of persistence times, speeds, and sampling frequency) that matched experimentally measured correlations between speed and turning for two datasets of T cell movement in vivo suggesting that such simple correlations are not fully informative on the intrinsic link between speed and persistence. Our results thus suggest that sub-sampling may contribute to (and perhaps fully explains) the observed correlation between speed and turning at least for some cell trajectory data and emphasize the role of sampling frequency in inference of critical cellular parameters of cell motility such as speeds.

Secondary AbstractMeasurement of cell movements often results in a negative correlation between average speed and average turning angle suggesting an existence of a universal, cell-intrinsic movement program. We show that such a negative correlation may arise if cells in the population differ in their ability for persistent movement when the movement data are sub-sampled. We show that sub-sampling of cell trajectories generated using two different frameworks of persistent random walk can match the experimentally observed correlation between speed and turning for T cell movements in vivo.
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. A rationalized definition of tumor suppressor microRNAsexcludes miR-34a While several microRNAs (miRNAs) have been proposed to act as tumor suppressors, a consensual definition of tumor suppressing miRNAs is still missing. Similarly to coding genes, we propose that tumor suppressor miRNAs must show evidence of genetic or epigenetic inactivation in cancers, and exhibit an anti-tumorigenic (e.g., anti-proliferative) activity under endogenous expression levels. Here we observe that this definition excludes the most extensively studied tumor suppressor candidate miRNA, miR-34a. In analyzable cancer types, miR-34a does not appear to be down-regulated in primary tumors relatively to normal adjacent tissues. Deletion of miR-34a is occasionally found in human cancers, but it does not seem to be driven by an anti-tumorigenic activity of the miRNA, since it is not observed upon smaller, miR-34a-specific alterations. Its anti-proliferative action was observed upon large, supra-physiological transfection of synthetic miR-34a in cultured cells, and our data indicates that endogenous miR-34a levels do not have such an effect. Our results therefore argue against a general tumor suppressive function for miR-34a, providing an explanation to the lack of efficiency of synthetic miR-34a administration against solid tumors. Vapor exposure to Δ9-tetrahydrocannabinol (THC) slows locomotion of the Maine Lobster (Homarus americanus) RationaleDespite a long history of use in synaptic physiology, the lobster has been a neglected model for behavioral pharmacology. A restaurateur proposed that exposing lobster to cannabis smoke reduces anxiety and pain during the cooking process. It is unknown if lobster gill respiration in air would result in significant {Delta}9-tetrahydrocannabinol (THC) uptake and whether this would have any detectable behavioral effects.

ObjectiveThe primary goal was to determine tissue THC levels in the lobster after exposure to THC vapor. Secondary goals were to determine if THC vapor altered locomotor behavior or nociception.

MethodsTissue samples were collected (including muscle, brain and hemolymph) from Homarus americanus (N=3 per group) following 30 or 60 minutes of exposure to vapor generated by an e-cigarette device using THC (100 mg/mL in a propylene glycol vehicle). Separate experiments assessed locomotor behavior and hot water nociceptive responses following THC vapor exposure.

ResultsTHC vapor produced duration-related THC levels in all tissues examined. Locomotor activity was decreased (distance, speed, time-mobile) by 30 min inhalation of THC. Lobsters exhibit a temperature-dependent withdrawal response to immersion of tail, antennae or claws in warm water; this is novel evidence of thermal nociception for this species. THC exposure for 60 minutes had only marginal effect on nociception under the conditions assessed.

ConclusionsVapor exposure of lobsters, using an e-cigarette based model, produces dose-dependent THC levels in all tissues and reduces locomotor activity. Hot water nociception was temperature dependent, but only minimal anti-nociceptive effect of THC exposure was confirmed. MTG16 (CBFA2T3) regulates colonic epithelial differentiation, colitis, and tumorigenesis by repressing E protein transcription factors Aberrant epithelial differentiation and regeneration contribute to colon pathologies including inflammatory bowel disease (IBD) and colitis-associated cancer (CAC). MTG16 (CBFA2T3) is a transcriptional corepressor expressed in the colonic epithelium. MTG16 deficiency in mice exacerbates colitis and increases tumor burden in CAC, though the underlying mechanisms remain unclear. Here, we identified MTG16 as a central mediator of epithelial differentiation, promoting goblet and restraining enteroendocrine cell development in homeostasis and enabling regeneration following dextran sulfate sodium (DSS)-induced colitis. Transcriptomic analyses implicated increased E box-binding transcription factor (E protein) activity in MTG16-deficient colon crypts. Using a novel mouse model with a point mutation that disrupts MTG16:E protein complex formation (Mtg16P209T), we established that MTG16 exerts control over colonic epithelial differentiation and regeneration by repressing E protein-mediated transcription. Mimicking murine colitis, MTG16 expression was increased in biopsies from patients with active IBD compared to unaffected controls. Finally, uncoupling MTG16:E protein interactions only partially phenocopied the enhanced tumorigenicity of Mtg16-/- colon in the azoxymethane(AOM)/DSS-induced model of CAC, indicating that MTG16 protects from tumorigenesis through additional mechanisms. Collectively, our results demonstrate that MTG16, via its repression of E protein targets, is a key regulator of cell fate decisions during colon homeostasis, colitis, and cancer.

Graphical Abstract

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org.highwire.dtl.DTLVardef@21948borg.highwire.dtl.DTLVardef@9f60eeorg.highwire.dtl.DTLVardef@194b9fdorg.highwire.dtl.DTLVardef@13eb1a7_HPS_FORMAT_FIGEXP  M_FIG C_FIG Chromatin-contact atlas reveals disorder-mediated protein interactions and moonlighting chromatin-associated RBPs RNA-binding proteins (RBPs) play diverse roles in regulating co-transcriptional RNA-processing and chromatin functions, but our knowledge of the repertoire of chromatin-associated RBPs (caRBPs) and their interactions with chromatin remains limited. Here, we developed SPACE (Silica Particle Assisted Chromatin Enrichment) to isolate global and regional chromatin components with high specificity and sensitivity, and SPACEmap to identify the chromatin-contact regions in proteins. Applied to mouse embryonic stem cells, SPACE identified 1,459 chromatin-associated proteins, [~]48% of which are annotated as RBPs, indicating their dual roles in chromatin and RNA-binding. Additionally, SPACEmap stringently verified chromatin-binding of 404 RBPs and identified their chromatin-contact regions. Notably, SPACEmap showed that about half of the caRBPs bind to chromatin by intrinsically disordered regions (IDRs). Studying SPACE and total proteome dynamics from mES cells grown in 2iL and serum medium indicates significant correlation (R = 0.62). One of the most dynamic caRBPs is Dazl, which we find co-localized with PRC2 at transcription start sites of genes that are distinct from Dazl mRNA binding. Dazl and other PRC2-colocalised caRBPs are rich in intrinsically disordered regions (IDRs), which could contribute to the formation and regulation of phase-separated PRC condensates. Together, our approach provides an unprecedented insight into IDR-mediated interactions and caRBPs with moonlighting functions in native chromatin. 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. Worldwide analysis of reef surveys sorts coral taxa by (adaptive) response to heat stress Coral reefs around the world are under threat due to widespread decline of hard corals caused by anomalous heat waves. Different coral taxa are known to have different sensitivities to heat. However, little is known about how long-term effects of heat exposure differ between coral taxa, nor in particular how such effects might drive adaptive processes. Here, we combined worldwide reef survey data with remotely sensed thermal anomalies to evaluate how local rates of taxa-specific coral cover were associated with heat stress. We found a negative association between coral cover and heat stress: weaker for massive and encrusting Poritidae, for meandroid Favidae and Mussidae; and stronger for branching, tabular and corymbose Acroporidae. Additionally, we found that coral cover decline driven by recent heat stress (measured as the year before a survey) was mitigated by long-term past heat (measured since 1985) in branching, tabular and corymbose Acroporidae and Pocilloporidae, but not in Poritidae, Favidae and Mussidae. This mitigation of coral decline was not observed under the effect of short-term past heat (measured during the two previous years), suggesting that these presumptive adaptive processes are the result of evolutionary adaptation, rather than of physiological acclimatization. We finally queried the Coral Trait Database to characterize morphological, physiological and reproductive traits of different taxa showing similar heat-responses. Further genomics studies should confirm the evolutionary origin of such divergent adaptive responses to heat stress. Reconstitution of minimal motility system based on Spiroplasma swimming by expressing two bacterial actins in synthetic minimal bacterium Motility is one of the most important features of life, but its evolutionary origin remains unknown. In this study, we focused on Spiroplasma, commensal, or parasitic bacteria. They swim by switching the helicity of a ribbon-like cytoskeleton that comprises six proteins, each of which evolved from a nucleosidase and bacterial actin called MreB. We expressed these proteins in a synthetic, non-motile minimal bacterium, JCVI-syn3.0B, whose reduced genome was computer-designed and chemically synthesized. The synthetic bacterium exhibited swimming motility with features characteristic of Spiroplasma swimming. Moreover, some combinations of the two proteins produced a helical cell shape and swimming, suggesting that the swimming originated from the differentiation and coupling of bacterial actins, and we also obtained a minimal system for motility of the synthetic bacterium.

One-Sentence SummaryThe minimal system comprised two bacterial actins that provided cell helicity and swimming to the synthetic minimal bacterium. Morphology of immatures of the thelytokous ant, Monomorium triviale Wheeler (Formicidae: Myrmicinae: Solenopsidini) with descriptions of the extraordinary last-instar queen larvae The ant genus Monomorium is one of the most species-rich but taxonomically problematic groups in the hyperdiverse subfamily Myrmicinae. An East Asian species, M. triviale Wheeler, produces both reproductive queens and sterile workers via obligate thelytokous parthenogenesis. Here, we describe the immature forms of M. triviale based on light and scanning electron microscopy observations, with a note on the striking caste dimorphism in the last larval instar. The last-instar queen larvae were easily recognized by their large size, "aphaenogastroid" body shape, and rows of doorknob-like tubercles on the lateral and dorsal body surface. This type of queen-specific structure has not been found in ants in general, let alone congeneric species found in Japan. In stark contrast to the queen larvae, worker larvae showed a "pheidoloid" body shape and a body surface similar to other ants. The worker larvae were estimated to have three instars, consistent with previously described congeners. The pupae of both castes had no cocoon, a characteristic commonly described in other Myrmicinae species. In total, the developmental period from egg to adult worker averaged 59 days under 25{degrees}C. We discuss possible functions of the tubercles of queen larvae based on previous studies. FGF8-mediated signaling regulates tooth developmental pace during odontogenesis The developing human and mouse teeth constitute an ideal model system to study the regulatory mechanism underlying organ growth control due to the fact that their teeth share highly conserved and well-characterized developmental processes and their developmental tempo vary notably. In the current study, we manipulated heterogenous recombination between human and mouse dental tissues and demonstrate that the dental mesenchyme dominates the tooth developmental tempo and FGF8 could be a critical player during this developmental process. Forced activation of FGF8 signaling in the dental mesenchyme of mice promoted cell proliferation, prevented cell apoptosis via p38 and perhaps PI3K-Akt intracellular signaling, and impelled the transition of the cell cycle from G1- to S-phase in the tooth germ, resulting in the slowdown of the tooth developmental pace. Our results provide compelling evidence that extrinsic signals can profoundly affect tooth developmental tempo and the dental mesenchymal FGF8 could be a pivotal factor in controlling developmental pace in a non-cell-autonomous manner during mammalian odontogenesis. Long term analysis of social structure: evidence of age-based consistent associations in male Alpine ibex Despite its recognized importance for understanding the evolution of animal sociality as well as for conservation, long term analysis of social networks of animal populations is still relatively uncommon. We investigated social network dynamics in males of a gregarious mountain ungulate (Alpine ibex, Capra ibex) over ten years focusing on groups, sub-groups and individuals, exploring the dynamics of sociality over different scales. Despite the social structure changing between seasons, the Alpine ibex population was highly cohesive: fission-fusion dynamics lead almost every male in the population to associate with each other male at least once. Nevertheless, we found that male Alpine ibex showed preferential associations that were maintained across seasons and years. Age seemed to be the most important factor driving preferential associations while other characteristics, such as social status, appeared less crucial. We also found that centrality measures were influenced by age and were also related to individual physical condition. The multi-scale and long-term frame of our study helped us show that ecological constrains, such as resource availability, may play a role in shaping associations in a gregarious species, but they cannot solely explain sociality and preferential association that are likely also to be driven by life-history linked physiological and social needs. Our results highlight the importance of long-term studies based on individually recognizable subjects to help us build on our understanding of the evolution of animal sociality. Enrichment analyses identify shared associations for 25 quantitative traits in over 600,000 individuals from seven diverse ancestries Since 2005, genome-wide association (GWA) datasets have been largely biased toward sampling European ancestry individuals, and recent studies have shown that GWA results estimated from self-identified European individuals are not transferable to non-European individuals due to various confounding challenges. Here, we demonstrate that enrichment analyses which aggregate SNP-level association statistics at multiple genomic scales--from genes to genomic regions and pathways--have been underutilized in the GWA era and can generate biologically interpretable hypotheses regarding the genetic basis of complex trait architecture. We illustrate examples of the robust associations generated by enrichment analyses while studying 25 continuous traits assayed in 566,786 individuals from seven diverse self-identified human ancestries in the UK Biobank and the Biobank Japan, as well as 44,348 admixed individuals from the PAGE consortium including cohorts of African-American, Hispanic and Latin American, Native Hawaiian, and American Indian/Alaska Native individuals. We identify 1,000 gene-level associations that are genome-wide significant in at least two ancestry cohorts across these 25 traits, as well as highly conserved pathway associations with triglyceride levels in European, East Asian, and Native Hawaiian cohorts. Alpha oscillations and stimulus-evoked activity dissociate metacognitive reports of attention, visibility and confidence in a rapid visual detection task Variability in the detection and discrimination of weak visual stimuli has been linked to oscillatory neural activity. In particular, the amplitude of activity in the alpha-band (8-12 Hz) has been shown to impact upon the objective likelihood of stimulus detection, as well as measures of subjective visibility, attention, and decision confidence. We aimed to clarify how preparatory alpha influences performance and phenomenology, by recording simultaneous subjective measures of attention and confidence (Experiment 1), or attention and visibility (Experiment 2) on a trial-by-trial basis in a visual detection task. Across both experiments, alpha amplitude was negatively and linearly correlated with the intensity of subjective attention. In contrast to this linear relationship, we observed a quadratic relationship between the strength of alpha oscillations and subjective ratings of confidence and visibility. We find that this same quadratic relationship links alpha amplitude to the strength of stimulus evoked responses. Visibility and confidence judgements corresponded to the strength of evoked responses, but confidence, uniquely, incorporated information about attentional state. As such, our findings reveal distinct psychological and neural correlates of metacognitive judgements of attentional state, stimulus visibility, and decision confidence. Algal photosystem I dimer and high resolution model of PSI:plastocyanin complex Photosystem I (PSI) enables photo-electron transfer and regulates photosynthesis in the bioenergetic membranes of cyanobacteria and chloroplasts. Being a multi-subunit complex, its macromolecular organization affects the dynamics of photosynthetic membranes. Here, we reveal a chloroplast PSI from the green alga Chlamydomonas reinhardtii that is organized as a homodimer, comprising 40 protein subunits with 118 transmembrane helices that provide scaffold for 568 pigments. Our cryo-EM structure identifies that the absence of PsaH and Lhca2 gives rise to a head-to-head relative orientation of the PSI-LHCI monomers in a way that is essentially different from the oligomer formation in cyanobacteria. The light-harvesting protein Lhca9 is the key element for mediating this dimerization. The interface between the monomers is lacking PsaH, and thus partially overlaps with the surface area that would bind one of the LHCII complexes in state transitions. We also define the most accurate available PSI-LHCI model at 2.3 [A] resolution, including a flexibly bound electron donor plastocyanin, and assign correct identities and orientations of all the pigments, as well as 621 water molecules that affect energy transfer pathways. Re-convolving the compositional landscape of primary and recurrent glioblastoma reveals prognostic andtargetable tissue states Glioblastoma is an aggressive diffusely infiltrating neoplasm that spreads beyond surgical resection margins, where it intermingles with non-neoplastic brain cells. This complex microenvironment harbouring infiltrating glioma and non-neoplastic brain cells is the origin of tumor recurrence. Thus, understanding the cellular and molecular features of the glioma microenvironment is therapeutically and prognostically important. Here, we used single-nucleus RNA sequencing (snRNAseq) of primary and recurrent glioma to define three compositional  tissue-states rooted in cohabitation of cell-types and transcriptional states. These comprise a state featuring A) abundant normal brain microenvironment cells, B) reactive/inflamed brain microenvironment, and C) cellular/proliferative tumor. All these states exhibited variable degrees of infiltration by glioma cells. Spatial transcriptomics confirmed that the cell-types and transcriptomics states which compositionally cohabitate tissue states also colocalize in space. Tissue states are clinically significant because they correlate with radiographic, histopathologic, and prognostic features. We found that enrichment of tissue state B signature correlated with shorter survival. Importantly, we found that our compositionally defined tissue states are enriched in distinct metabolic pathways. One such pathway is fatty acid biosynthesis, which was enriched in tissue state B - a state enriched in recurrent glioblastoma and composed of astrocyte-like/mesenchymal glioma cells, reactive astrocytes resembling those seen in neurodegeneration, and monocyte-like myeloid cells. We showed that treating acute slices of GBM with a fatty acid synthesis inhibitor is sufficient to deplete the transcriptional signature of tissue state B. Our findings define a novel compositional approach to the glioma-infiltrated tissue which allows us to discover prognostic and targetable features, paving the way to new mechanistic and therapeutic discoveries. GUIdEStaR (G-quadruplex, uORF, IRES, Epigenetics, Small RNA, Repeats), the integrated matadatabase: transcript-indexed binary information creation for chaining with neural network analysis GUIdEStaR integrates existing databases of various types of G-quadruplex, upstream Open Reading Frame (uORF), Internal Ribosome Entry Site (IRES), methylation to RNA and histone protein, small RNA, and repeats. GUIdEStaR consists of approx. 40,000 genes and 320,000 transcripts. An mRNA transcript is divided into 5 regions (5UTR, 3UTR, exon, intron, and biological region) where each region contains presence-absence data of 169 different types of elements. Recently, artificial intelligence (AI) based analysis of sequencing data has been gaining popularity in the area of bioinformatics. GUIdEStaR generates datasets that can be used as inputs to AI methods. At the GUIdEStaR homepage, users submit gene symbols by clicking a "Send" button, and shortly result files in CSV format are available for download at the result website. Users have an option to send the result files to their email addresses. Additionally, the entire database and the example Java codes are also freely available for download. Here, we demonstrate the database usage with three neural network classification studies-1) small RNA study for classifying transcription factor (TF) genes into either one of TF mediated by small RNA originated from SARS-CoV-2 or by human microRNA (miRNA), 2) cell membrane receptor study for classifying receptor genes as either with virus interaction or without one, and 3) nonsense mediated mRNA decay (NMD) study for classifying cell membrane and nuclear receptors as either NMD target or non-target. GUIdEStaR is available for access to the easy-to-use web-based database at www.guidestar.kr and for download at https://sourceforge.net/projects/guidestar. Relationship between shear wave velocity and muscle activation is inconsistent across different muscle types There is an increasing use of shear wave ultrasound elastography to quantify mechanical properties of muscles under various conditions such as changes muscle length and levels of activation in healthy and pathological muscle. However, little is known about the variability in shear wave velocity among muscles as most studies investigate one specific muscle. The purpose of this study was to determine if the relationship between SWV and muscle activation is consistent across muscles with different architectures: biceps brachii, tibialis anterior, and medial gastrocnemius, All measures were made at matching levels of activation and approximately at the optimal length for each muscle to control for length-dependent changes in the relationship between activation and force or stiffness. We also conducted a control experiment to determine how the passive force within a muscle alters the relationship between muscle activation and shear wave velocity. The relationship between shear wave velocity-squared and activation above 10% MVC differed across muscles, with biceps brachii and medial gastrocnemius showing a lower slope than tibialis anterior. Shear wave velocity-squared also differed between muscles at the shortest length (p<0.001) and the increase in shear wave velocity-squared with muscle lengthening also differed between muscle types (p = 0.005) Muscle-specific differences could not be explained by the architectural features such as pennation angle, fiber length, and physiological cross-sectional area. Our results demonstrate that there is not a unique relationship between muscle activation and shear wave velocity, highlighting the importance of understanding the many factors contributing to shear wave propagation in muscle before elastography can be used to make quantitative comparisons across muscle types. Testing for phylogenetic signal in single-cell RNA-seq data AO_SCPLOWBSTRACTC_SCPLOWPhylogenetic methods are emerging as a useful tool to understand cancer evolutionary dynamics, including tumor structure, heterogeneity, and progression. Most currently used approaches utilize either bulk whole genome sequencing (WGS) or single-cell DNA sequencing (scDNA-seq) and are based on calling copy number alterations and single nucleotide variants (SNVs). Here we explore the potential of single-cell RNA sequencing (scRNA-seq) to reconstruct cancer evolutionary dynamics. scRNA-seq is commonly applied to explore differential gene expression of cancer cells throughout tumor progression. The method exacerbates the single-cell sequencing problem of low yield per cell with uneven expression levels. This accounts for low and uneven sequencing coverage and makes SNV detection and phylogenetic analysis challenging. In this paper, we demonstrate for the first time that scRNA-seq data contains sufficient evolutionary signal and can be utilized in phylogenetic analyses. We explore and compare results of such analyses based on both expression levels and SNVs called from scRNA-seq data. Both techniques are shown to be useful for reconstructing phylogenetic relationships between cells, reflecting the clonal composition of a tumor. Both standardized expression values and SNVs appear to be equally capable of reconstructing a similar pattern of phylogenetic relationship. This pattern is stable even when phylogenetic uncertainty is taken in account. Our results open up a new direction of somatic phylogenetics based on scRNA-seq data. Further research is required to refine and improve these approaches to capture the full picture of somatic evolutionary dynamics in cancer. Bioinformatic and mechanistic analysis of the palmerolide PKS-NRPS biosynthetic pathway from the microbiome of an Antarctic ascidian Complex interactions exist between microbiomes and their hosts. Increasingly, defensive metabolites that have been attributed to host biosynthetic capability are now being recognized as products of host-associated microbes. These unique metabolites often have bioactivity targets in human disease and can be purposed as pharmaceuticals. Polyketides are a complex family of natural products that often serve as defensive metabolites for competitive or pro-survival purposes for the producing organism, while demonstrating bioactivity in human diseases as cholesterol lowering agents, anti-infectives, and anti-tumor agents. Marine invertebrates and microbes are a rich source of polyketides. Palmerolide A, a polyketide isolated from the Antarctic ascidian Synoicum adareanum, is a vacuolar-ATPase inhibitor with potent bioactivity against melanoma cell lines. The biosynthetic gene clusters (BGCs) responsible for production of secondary metabolites are encoded in the genomes of the producers as discrete genomic elements. A candidate palmerolide BGC was identified from a S. adareanum microbiome-metagenome based on a high degree of congruence with a chemical structure-based retrobiosynthetic prediction. Protein family homology analysis, conserved domain searches, active site and motif identification were used to identify and propose the function of the [~]75 kb trans-acyltransferase (AT) polyketide synthase-non-ribosomal synthase (PKS-NRPS) domains responsible for the stepwise synthesis of palmerolide A. Though PKS systems often act in a predictable co-linear sequence, this BGC includes multiple trans-acting enzymatic domains, a non-canonical condensation termination domain, a bacterial luciferase-like monooxygenase (LLM), and is found in multiple copies within the metagenome-assembled genome (MAG). Detailed inspection of the five highly similar pal BGC copies suggests the potential for biosynthesis of other members of the palmerolide chemical family. This is the first delineation of a biosynthetic gene cluster from an Antarctic microbial species, recently proposed as Candidatus Synoicohabitans palmerolidicus. These findings have relevance for fundamental knowledge of PKS combinatorial biosynthesis and could enhance drug development efforts of palmerolide A through heterologous gene expression. An Ohio State Scenic River Shows Elevated Antibiotic Resistance Genes, Including Acinetobacter Tetracycline and Macrolide Resistance, Downstream of Wastewater Treatment Plant Effluent The entry of antibiotic resistance genes (ARGs) into aquatic systems has been documented for large municipal wastewater treatment plants, but there is less study of the impact of smaller plants that are situated on small rural rivers. We sampled water metagenomes for ARG and taxa composition from the Kokosing River, a small rural river in Knox County, Ohio, which has been designated an Ohio State Scenic River for retention of natural character. Samples were obtained 1.0 km upstream, 120 m downstream, and 6.4 km downstream from the effluent release of the Mount Vernon wastewater treatment plant (WWTP). ARGS were identified in metagenomes using ShortBRED markers from the CARD database screened against UniPROT. Through all seasons, the metagenome just downstream of the WWTP effluent showed a substantial elevation of at least 15 different ARGs, including 6 ARGs commonly associated with Acinetobacter baumannii such as msrE, mphE (macrolide resistance) and tet(39) (tetracycline resistance). The ARGs most prevalent near the effluent pipe persisted 6.4 km downriver. Using MetaPhlAn2 clade-specific marker genes, the taxa distribution near the effluent showed elevation of reads annotated as Acinetobacter species as well as gut-associated taxa, Bacteroides and Firmicutes. The ARG levels and taxa prevalence showed little dependence on seasonal chlorination of the effluent. Nitrogen and phosphorus were elevated near the effluent pipe but had no consistent correlation with ARG levels. We show that in a rural river microbiome, year-round wastewater effluent substantially elevates ARGs including those associated with multidrug-resistant A. baumanii.

IMPORTANCEAntibiotic resistance is a growing problem worldwide, with frequent transmission between pathogens and environmental organisms. Rural rivers can support high levels of recreational use by people unaware of inputs from treated wastewater, while WWTPs can generate a small but significant portion of flow volume into a river surrounded by forest and agriculture. There is little information on the rural impacts of WWTP effluent on the delivery and transport of antibiotic resistance genes. In our study, the river water proximal to wastewater effluent shows evidence for the influx of multidrug-resistant Acinetobacter baumanii, an opportunistic pathogen of concern for hospitals but also widespread in natural environments. Our work highlights the importance of wastewater effluent in management of environmental antibiotic resistance, even in high quality, rural river systems. Python-Microscope: A new open source Python library for the control of microscopes Custom built microscopes often require control of multiple hardware devices and precise hardware coordination. It is also desirable to have a solution that is scalable to complex systems and translatable between components from different manufacturers. Here we report Python-Microscope, a free and open source Python library for high performance control of arbitrarily complex and scalable custom microscope systems. Python-Microscope offers simple to use Python-based tools, abstracting differences between physical devices by providing a defined interface for different device types. Concrete implementations are provided for a range of specific hardware and a framework exists for further expansion. Python-Microscope supports the distribution of devices over multiple computers while maintaining synchronisation via highly precise hardware triggers. We discuss the architecture choices of Python-Microscope that overcome the performance problems often raised against Python and demonstrate the different use cases that drove its design: its integration in user facing projects, namely in the Microscope-Cockpit project; in controlling complex microscopes at high speed while using the Python programming language; and as a microscope simulation tool for software development. Phylogenomic discordance suggests polytomies along the backbone of the large genus Solanum Premise of the studyEvolutionary studies require solid phylogenetic frameworks, but increased volumes of phylogenomic data have revealed incongruent topologies among gene trees in many organisms both between and within genomes. Some of these incongruences indicate polytomies that may remain impossible to resolve. Here we investigate the degree of gene-tree discordance in Solanum, one of the largest flowering plant genera that includes the cultivated potato, tomato, and eggplant, as well as 24 minor crop plants.

MethodsA densely sampled species-level phylogeny of Solanum is built using unpublished and publicly available Sanger sequences comprising 60% of all accepted species (742 spp.) and nine regions (ITS, waxy, and seven plastid markers). The robustness of this topology is tested by examining a full plastome dataset with 140 species and a nuclear target-capture dataset with 39 species of Solanum (Angiosperms353 probe set).

Key resultsWhile the taxonomic framework of Solanum remained stable, gene tree conflicts and discordance between phylogenetic trees generated from the target-capture and plastome datasets were observed. The latter correspond to regions with short internodal branches, and network analysis and polytomy tests suggest the backbone is composed of three polytomies found at different evolutionary depths. The strongest area of discordance, near the crown node of Solanum, could potentially represent a hard polytomy.

ConclusionsWe argue that incomplete lineage sorting due to rapid diversification is the most likely cause for these polytomies, and that embracing the uncertainty that underlies them is crucial to understand the evolution of large and rapidly radiating lineages. Integrative small and long RNA-omics analysis of human healing and non-healing wounds discovers cooperating microRNAs as therapeutic targets MicroRNAs (miR) are important posttranscriptional regulators and exhibit a high potential to be utilized in diagnosis and therapy. However, our insufficient knowledge of the miR-mediated gene regulation in human skin wound healing severely hinders the identification of clinically relevant miRs. Here, we performed paired small RNA and long RNA sequencing in human tissue samples, including matched skin and acute wounds collected at each healing stage and chronic non-healing venous ulcers (VU). With integrative small and long RNA-omics analysis, we developed a compendium (https://www.xulandenlab.com/humanwounds-mirna-mrna), which will be an open, comprehensive resource to broadly aid wound healing research. With this first clinical, wound-centric resource of miRs and mRNAs, we identified 17 pathologically relevant miRs that exhibited abnormal VU expression and displayed their targets enriched explicitly in the VU gene signature. Intermeshing regulatory networks controlled by these miRs revealed their high cooperativity in contributing to chronic wound pathology characterized by persistent inflammation and proliferative phase initiation failure. Furthermore, we demonstrated that miR-34a, miR-424, and miR-516, upregulated in VU, cooperatively suppressed keratinocyte growth while promoting inflammatory response. Collectively, our study opens the possibility of developing innovative wound treatment that targets pathologically relevant cooperating miRs to attain higher therapeutic efficacy and specificity. Impact of circadian time of dosing on cardiomyocyte-autonomous effects of glucocorticoids Mitochondrial capacity is critical to adapt the high energy demand of the heart to circadian oscillations and diseased states. Glucocorticoids regulate the circadian cycle of energy metabolism, but little is known about how circadian timing of exogenous glucocorticoid dosing directly regulates heart metabolism through cardiomyocyte-autonomous mechanisms. While chronic oncedaily intake of glucocorticoids promotes metabolic stress and heart failure, we recently discovered that intermittent once-weekly dosing of exogenous glucocorticoids promoted muscle metabolism in normal and obese skeletal muscle. However, the effects of glucocorticoid intermittence on heart metabolism and heart failure remain unknown. Here we investigated the extent to which circadian time of dosing regulates the effects of the glucocorticoid prednisone in heart metabolism and function in conditions of single pulse or chronic intermittent dosing. In WT mice, we found that prednisone improved cardiac content of NAD+ and ATP with light-phase dosing (ZT0), while the effects were blocked by dark-phase dosing (ZT12). The drug effects on mitochondrial function were cardiomyocyte-autonomous, as shown by inducible cardiomyocyte-restricted glucocorticoid receptor (GR) ablation, and depended on an intact cardiomyocyte clock, as shown by inducible cardiomyocyte-restricted ablation of Brain and Muscle ARNT-like 1 (BMAL1). Conjugating time-of-dosing with chronic intermittence, we found that once-weekly prednisone improved metabolism and function in heart after myocardial injury dependent on circadian time of intake, i.e. with lightphase but not dark-phase dosing. Our study identifies cardiac-autonomous mechanisms through which circadian-specific intermittent dosing reconverts glucocorticoid drugs to metabolic boosters for the heart.