Regulation of ammonium acquisition and use in Oryza longistaminata ramets under nitrogen source heterogeneity Oryza longistaminata, a wild rice, vegetatively reproduces and forms a networked clonal colony consisting of ramets connected by rhizomes. Although water, nutrients, and other molecules can be transferred between ramets via the rhizomes, inter-ramet communication in response to spatially heterogeneous nitrogen availability is not well understood. We studied the response of ramet pairs to heterogeneous nitrogen availability by using a split hydroponic system that allowed each ramet root to be exposed to different conditions. Ammonium uptake was compensatively enhanced in the sufficient-side root when roots of the ramet pairs were exposed to ammonium-sufficient and deficient conditions. Comparative transcriptome analysis revealed that a gene regulatory network for effective ammonium assimilation and amino acid biosynthesis was activated in the sufficient-side roots. Allocation of absorbed nitrogen from the nitrogen-sufficient to the deficient ramets was rather limited. Nitrogen was preferentially used for newly growing axillary buds on the sufficient-side ramets. Biosynthesis of trans-zeatin, a cytokinin, was up-regulated in response to the nitrogen supply, but trans-zeatin appears not to target the compensatory regulation. Our results also implied that the O. longistaminata ortholog of OsCEP1 plays a role as a nitrogen-deficient signal in inter-ramet communication, providing compensatory up-regulation of nitrogen assimilatory genes. These results provide insights into the molecular basis for efficient growth strategies of asexually proliferating plants growing in areas where the distribution of ammonium ions is spatially heterogeneous.

One sentence summaryOryza longistaminata, a rhizomatous wild rice, systemically regulates ammonium acquisition and use in response to spatially heterogeneous nitrogen availability via inter-ramet communication.
Applications of the indole-alkaloid gramine modulate the assembly of individual members of the barley rhizosphere microbiota Microbial communities proliferating at the root-soil interface, collectively referred to as the rhizosphere microbiota, represent an untapped beneficial resource for plant growth, development and health. Integral to a rational manipulation of the microbiota for sustainable agriculture is the identification of the molecular determinants of these communities. In plants, biosynthesis of allelochemicals is centre stage in defining inter-organismal relationships in the environment. Intriguingly, this process has been moulded by domestication and breeding selection. The indole-alkaloid gramine, whose occurrence in barley (Hordeum vulgare L.) is widespread among wild genotypes but has been counter selected in several modern varieties, is a paradigmatic example of this phenomenon. This prompted us to investigate how exogenous applications of gramine impacted on the rhizosphere microbiota of two, gramine-free, elite barley varieties grown in a reference agricultural soil. High throughput 16S rRNA gene amplicon sequencing revealed that applications of gramine interfere with the proliferation of a subset of soil microbes with a relatively broad phylogenetic assignment. Strikingly, growth of these bacteria appeared to be rescued by barley plants in a genotype- and dosage-independent manner. In parallel, we discovered that host recruitment cues can interfere with the impact of gramine application in a host genotype-dependent manner. Interestingly, this latter effect displayed a bias for members of the phyla Proteobacteria. These initial observations indicate that gramine can act as a determinant of the prokaryotic communities inhabiting the root-soil interface.
scTenifoldKnk: an efficient virtual knockout tool for gene function predictions via single-cell gene regulatory network perturbation Bigger PictureO_LIGene knockout (KO) experiments, using genetically altered animals, are a proven powerful approach to elucidate the role of a gene in a biological process. However, systematic KO experiments targeting many genes are usually prohibitive due to limited experimental and animal resources. Here, we present scTenifoldKnk, an efficient virtual KO tool that allows the systematic deletion of many genes individually. scTenifoldKnk uses single-cell RNA sequencing (scRNAseq) data from wild-type (WT) samples to predict gene function in a cell type-specific manner. We show that predictions made by scTenifoldKnk recapitulate findings from real-animal KO experiments. scTenifoldKnk has proven to be a powerful and effective approach for elucidating gene function, prioritizing KO targets, predicting experimental outcomes before real-animal KO experiments are conducted.
C_LI

HighlightsO_LIscTenifoldKnk performs virtual KO experiments using scRNAseq data.
C_LIO_LIscTenifoldKnk only requires data from WT samples; no data is needed from KO samples.
C_LIO_LIPredictions made by scTenifoldKnk recapitulate findings from real-animal KO experiments.
C_LI

Data Science Maturity Level

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=14 SRC="FIGDIR/small/436484v2_ufig1.gif" ALT="Figure 1">
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org.highwire.dtl.DTLVardef@157126aorg.highwire.dtl.DTLVardef@17a2e76org.highwire.dtl.DTLVardef@805c4dorg.highwire.dtl.DTLVardef@12bdc1b_HPS_FORMAT_FIGEXP  M_FIG C_FIG eTOC blurbscTenifoldKnk is a machine learning workflow performing virtual KO experiments to predict gene function. It constructs gene regulatory networks using single-cell RNA sequencing data from wild-type samples and then computationally deletes target genes. Real-data applications demonstrate that scTenifoldKnk recapitulates findings of real-animal KO experiments and accurately predicts gene function in analyzed cells.

SummaryGene knockout (KO) experiments are a proven, powerful approach for studying gene function. However, systematic KO experiments targeting a large number of genes are usually prohibitive due to the limit of experimental and animal resources. Here, we present scTenifoldKnk, an efficient virtual KO tool that enables systematic KO investigation of gene function using data from single-cell RNA sequencing (scRNAseq). In scTenifoldKnk analysis, a gene regulatory network (GRN) is first constructed from scRNAseq data of wild-type samples, and a target gene is then virtually deleted from the constructed GRN. Manifold alignment is used to align the resulting reduced GRN to the original GRN to identify differentially regulated genes, which are used to infer target gene functions in analyzed cells. We demonstrate that the scTenifoldKnk-based virtual KO analysis recapitulates the main findings of real-animal KO experiments and recovers the expected functions of genes in relevant cell types. Cellular feedback to organotelluranes displays modulation of antioxidant proteins gene expression Organotelluranes RT3 and RT4 are thiol reagents that induce mitochondrial transition pore (MTP) opening in a sensitive and insensitive manner to cyclosporin A. Although RT3 and RT4 promote glutathione depletion, paradoxically, they are also an efficient antioxidant for membrane lipids. These compounds antagonistic effects elicited the challenging question of how the gene expression of antioxidant enzymes would respond to treatment with these compounds. The influence of RT3 and RT4 on antioxidant enzyme expression was investigated in cultured aortic smooth muscle cells (ASMC). RT3 and RT4 promoted disruption of ionic calcium homeostasis, mitochondrial transmembrane potential ({Delta}{Psi}), and cell death in a dose-dependent manner. The cell death mechanisms responded qualitatively to the increase of the organotellurane concentration and changed from apoptosis to necrosis. RT3 and RT4 increased the expression of thioredoxin significantly. RT3 also increased the expression of glutaredoxin and glutathione peroxidase, slightly the catalase expression without significant effects on SOD expression. The results are consistent with GSH and protein thiol depletion and discussed based on the cell toxicity mechanism exhibited by these compounds.



O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/425411v4_ufig1.gif" ALT="Figure 1">
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org.highwire.dtl.DTLVardef@52bb82org.highwire.dtl.DTLVardef@19476d9org.highwire.dtl.DTLVardef@6aa99org.highwire.dtl.DTLVardef@1aa605e_HPS_FORMAT_FIGEXP  M_FIG C_FIG Heat-Dependent Hairpin Melting Drives Thermo-gated TRPV1 Opening The capsaicin receptor TRPV1 is a bio-thermometer in a primary afferent sensory neuron for noxious heat detection. However, the underlying molecular mechanism is unclear. Here, a hairpin thermodynamic model, together with graph theory, was to examine a role of temperature-dependent hairpin sizes in controlling the temperature threshold and sensitivity and gating states of TRPV1. The results showed that heat-dependent melting of the largest hairpin initiates a temperature threshold for channel opening while the smaller hairpins stabilize the open state and increase the heat efficacy or sensitivity. Therefore, this model may provide a broad structural basis for the thermo-gated TRP channels. From GWAS Variant to Function: a Study of ~148,000 Variants for Blood Cell Traits. Genome-wide association studies (GWAS) have identified hundreds of thousands of genetic variants associated with complex diseases and traits. However, most variants are noncoding and not clearly linked to genes, making it challenging to interpret these GWAS signals. We present a systematic variant-to-function study, prioritizing the most likely functional elements of the genome for experimental follow-up, for >148,000 variants identified for hematological traits. Specifically, we developed VAMPIRE: Variant Annotation Method Pointing to Interesting Regulatory Effects, an interactive web application implemented in R Shiny (http://shiny.bios.unc.edu/vampire/). This tool efficiently integrates and displays information from multiple complementary sources, including epigenomic signatures from blood cell relevant tissues or cells, functional and conservation summary scores, variant impact on protein and gene expression, chromatin conformation information, as well as publicly available GWAS and phenome-wide association study (PheWAS) results. Leveraging data generated from independently performed functional validation experiments, we demonstrate that our prioritized variants, genes, or variant-gene links are significantly more likely to be experimentally validated. This study not only has important implications for systematic and efficient revelation of functional mechanisms underlying GWAS variants for hematological traits, but also provides a prototype that can be adapted to many other complex traits, paving the path for efficient variant to function (V2F) analyses. Bulk-assembly of monodisperse coacervates and giant unilamellar vesicles with programmable hierarchical complexity Compartmentalization is an essential step for the emergence of protocellular life on the early earth and the endeavor to bottom-up construct synthetic cells in the lab. Among manifold natural strategies for compartmentalization, assembly of lipid vesicles or coacervate droplets with resemblance to cell envelope or cytoplasm have emerged as two dominant paradigms. However, the spontaneous assembly of lipid vesicles or coacervates generally results in polydisperse microcompartments and lacks the ability to efficiently integrate functional building blocks towards defined complexity, which give rise to the difficulty to achieve the order and complexity towards the scope of that of life via spontaneous assembly. Herein, we show that the interplay of coacervates and colloidal particles provides a bulk-assembly approach to form monodisperse coacervates and giant unilamellar vesicles (GUVs) with programmable hierarchical complexity. The hierarchical complexity of the synthetic cell microcompartments can be delicately engineered from the control of the spatial or temporal organization of components in entity unit to manifold forms of interconnected synthetic cell consortia, achieving a compartment hierarchy analogous to the organelle-cell-tissue structure. The increase in complexity gives rise to emergent properties, for instance, collective morphology evolution of coacervate assemblages, gated permeability of GUVs without sophisticated protein machinery, and remarkable structural and functional stability of GUVs at extreme conditions. This work paves an unprecedented step to form monodisperse, hierarchical coacervates and GUVs via bulk-assembly, which provides a prebiotically plausible approach for the proper integration of inanimate matter towards the order and complexity of life and monodisperse, hierarchical microdroplets for follow-up scientific and technological applications. An exon-intron split framework to prioritize miRNA-driven post-transcriptional regulatory signals and its application to study energy homeostasis in pigs The contribution of microRNAs (miRNAs) to mRNA regulation has often been explored by post hoc selection of downregulated genes and determining whether they harbor binding sites for miRNAs of interest. This approach, however, does not discriminate whether these mRNAs are also downregulated at the transcriptional level. Here, we have characterized the transcriptional and post-transcriptional changes of mRNA expression in two porcine tissues: gluteus medius muscle of fasted and fed Duroc gilts and adipose tissue of lean and obese Duroc-Gottingen minipigs. Exon-intron split analysis (EISA) of RNA-seq data allowed us to identify downregulated mRNAs with high post-transcriptional signals in fed or obese states, and we assessed whether they harbor binding sites for upregulated miRNAs in any of these two physiological states. We found 26 downregulated mRNAs with high post-transcriptional signals in the muscle of fed gilts and 21 of these were predicted targets of upregulated miRNAs also in the fed state. For adipose tissue, 44 downregulated mRNAs in obese minipigs displayed high post-transcriptional signals, and 25 of these were predicted targets of miRNAs upregulated in the obese state. These results suggest that the contribution of miRNAs to mRNA repression is more prominent in the skeletal muscle system. Finally, we identified several genes that may play relevant roles in the energy homeostasis of the pig skeletal muscle (DKK2 and PDK4) and adipose (SESN3 and ESRRG) tissues. By differentiating transcriptional from post-transcriptional changes in mRNA expression, EISA provides a valuable view about the regulation of gene expression, complementary to canonical differential expression analyses. Microbiota-instructed regulatory T cell differentiation is mediated by a distinct RORγt+ antigen presenting cell subset The mutualistic relationship of gut-resident microbiota and cells of the host immune system promotes homeostasis that ensures maintenance of the microbial community and of a poised, but largely non-aggressive, immune cell compartment1, 2. Consequences of disturbing this balance, by environmental or genetic factors, include proximal inflammatory conditions, like Crohns disease, and systemic illnesses, both metabolic and autoimmune. One of the means by which this equilibrium is achieved is through induction of both effector and suppressor or regulatory arms of the adaptive immune system. In mice, Helicobacter species induce regulatory (iTreg) and follicular helper (Tfh) T cells in the colon-draining mesenteric lymph nodes under homeostatic conditions, but can instead induce inflammatory Th17 cells when iTreg cells are compromised3, 4. How Helicobacter hepaticus and other gut bacteria direct T cells to adopt distinct functions remains poorly understood. Here, we investigated which cells and molecular components are required to convey the microbial instruction for the iTreg differentiation program. We found that antigen presentation by cells expressing ROR{gamma}t, rather than by classical dendritic cells, was both required and sufficient for iTreg induction. These ROR{gamma}t+ cells, likely to be type 3 innate lymphoid cells (ILC3) and/or a recently-described population of Aire+ cells termed Janus cells5, require the MHC class II antigen presentation machinery, the chemokine receptor CCR7, and v integrin, which activates TGF-{beta}, for iTreg cell differentiation. In the absence of any of these, instead of iTreg cells there was expansion of microbiota-specific pathogenic Th17 cells, which were induced by other antigen presenting cells (APCs) that did not require CCR7. Thus, intestinal commensal microbes and their products target multiple APCs with pre-determined features suited to directing appropriate T cell differentiation programs, rather than a common APC that they endow with appropriate functions. Our results illustrate the ability of microbiota to exploit specialized functions of distinct innate immune system cells, targeting them to achieve the desired composition of equipoised T cells, thus maintaining tolerance. Nuclear alpha-synuclein is present in the human brain and is modified in dementia with Lewy bodies Dementia with Lewy bodies is pathologically defined by the cytoplasmic accumulation of alpha-synuclein within neuronal cells in the brain. Alpha-synuclein is predominately pre-synaptic, but has been reported present in various subcellular compartments in cell and animal models. In particular, nuclear alpha-synuclein is evident in-vitro and in disease models and has been associated with altered DNA integrity, gene transcription, nuclear homeostasis. However, owing to various factors, the presence of alpha-synuclein in the nuclei of human brain cells remains controversial, as does its role in synucleinopathies.

Here, we close this gap and provide a unique demonstration confirming the presence of nuclear alpha-synuclein in post-mortem brain tissue obtained from cases of dementia with Lewy bodies as well as from controls via immunohistochemistry, immunoblot, and label-free mass-spectrometry.

Discrete intra-nuclear alpha-synuclein puncta reactive against phosphorylated serine 129-alpha-synuclein and pan-alpha-synuclein antibodies were observed in cortical neurons and non-neuronal cells in fixed brain sections and in isolated nuclear preparations from Dementia with Lewy bodies cases and matched controls. Subsequent biochemical analysis of subcellular fractionated tissue confirmed alpha-synuclein as present in a nuclear fraction at levels ~ 10-fold lower than in the cytoplasm. Critically, however, an increase in monomeric nuclear alpha-synuclein phosphorylated as serine 129 was observed in cases of dementia with Lewy bodies alongside higher molecular weight pan- and phosphorylation reactive alpha-synuclein species, consistent with the formation of intranuclear phosphorylated alpha-synuclein oligomers. Furthermore, the presence of nuclear alpha-synuclein was confirmed via label free mass spectrometry, as 6 unique alpha-synuclein derived peptide sequences were identified in nuclear fractions (71.4% sequence coverage).

Collectively, our data confirm the presence of nuclear alpha-synuclein in human brain tissue and describe nuclear pathology associated with dementia with Lewy bodies. These findings address a major controversy in the synucleinopathy field by confirming the presence of nuclear alpha-synuclein in autoptic human brain tissue and, for the first time, identify that alpha-synuclein is aggregated into novel and potentially pathological assemblies in the nucleus as part of the disease process associated with dementia with Lewy bodies and thus may contribute to the disease phenotype. Effects of the social environment on movement-integrated habitat selection O_LIMovement links the distribution of habitats with the social environment of animals using those habitats; yet integrating movement, habitat selection, and socioecology remains an opportunity for further study.
C_LIO_LIHere, our objective was to disentangle the roles of habitat selection and social association as drivers of collective movement in a gregarious ungulate. To accomplish this objective, we (1) assessed whether socially familiar individuals form discrete social communities and whether social communities have high spatial, but not necessarily temporal, overlap; and (2) we modelled the relationship between collective movement and selection of foraging habitats using socially informed integrated step selection analysis.
C_LIO_LIWe used social network analysis to assign individuals to social communities and determine short and long-term social preference among individuals. Using integrated step selection functions (iSSF), we then modelled the effect of social processes, i.e., nearest neighbour distance and social preference, and movement behaviour on patterns of habitat selection.
C_LIO_LIBased on assignment of individuals to social communities and home range overlap analyses, individuals assorted into discrete social communities, and these communities had high spatial overlap. By unifying social network analysis with iSSF, we identified movement-dependent social association, where individuals foraged with more familiar individuals, but moved collectively with any between foraging patches.
C_LIO_LIOur study demonstrates that social behaviour and space use are inter-related based on spatial overlap of social communities and movement-dependent habitat selection. Movement, habitat selection, and social behaviour are linked in theory. Here, we put these concepts into practice to demonstrate that movement is the glue connecting individual habitat selection to the social environment.
C_LI The integrity and assay performance of tissue mitochondrial DNA is considerably affected by choice of isolation method The integrity of mitochondrial DNA (mtDNA) isolated from solid tissues is critical for analyses such as long-range PCR, but is typically assessed under conditions that fail to provide information on the individual mtDNA strands. Using denaturing gel electrophoresis, we show that commonly-used isolation procedures generate mtDNA containing several single-strand breaks per strand. Through systematic comparison of DNA isolation methods, we identify a procedure yielding the highest integrity of mtDNA that we demonstrate displays improved performance in downstream assays. Our results highlight the importance of isolation method choice, and serve as a resource to researchers requiring high-quality mtDNA from solid tissues. 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. Modulation of RecFORQ- and RecA-mediated homologous recombination in Escherichia coli by isoforms of translation initiation factor IF2 Homologous recombination (HR) is critically important for chromosomal replication as well as DNA damage repair in all life forms. In Escherichia coli, the process of HR is comprised of (i) two parallel pre-synaptic pathways that are mediated, respectively, by proteins RecB/C/D and RecF/O/R/Q; (ii) a synaptic step mediated by RecA that leads to generation of Holliday junctions (HJs); and (iii) post-synaptic steps mediated sequentially by HJ-acting proteins RuvA/B/C followed by proteins PriA/B/C of replication restart. Combined loss of RuvA/B/C and a DNA helicase UvrD is synthetically lethal, which is attributed to toxicity caused by accumulated HJs since viability in these double mutant strains is restored by removal of the pre-synaptic or synaptic proteins RecF/O/R/Q or RecA, respectively. Here we show that, as in {Delta}uvrD strains, ruv mutations confer synthetic lethality in cells deficient for transcription termination factor Rho, and that loss of RecFORQ pre-synaptic pathway proteins or of RecA suppresses this lethality. Furthermore, loss of IF2-1 (which is one of three isoforms [IF2-1, IF2-2, and IF2-3] of the essential translation initiation factor IF2 that are synthesized from three in-frame initiation codons in infB) also suppressed uvrD-ruv and rho-ruv lethalities, whereas deficiency of IF2-2 and IF2-3 exacerbated the synthetic defects. Our results suggest that Rho deficiency is associated with an increased frequency of HR that is mediated by the RecFORQ pathway along with RecA. They also lend support to earlier reports that IF2 isoforms participate in DNA transactions, and we propose that they do so by modulation of HR functions.

ImportanceThe process of homologous recombination (HR) is important for maintenance of genome integrity in all cells. In Escherichia coli, the RecA protein is a critical participant in HR, which acts at a step common to and downstream of two HR pathways mediated by the RecBCD and RecFOR proteins, respectively. In this study, an isoform (IF2-1) of the translation initiation factor IF2 has been identified as a novel facilitator of RecAs function in vivo during HR. Pseudomonas aeruginosa C-terminal processing protease CtpA assembles into a hexameric structure that requires activation by a spiral-shaped lipoprotein binding partner Pseudomonas aeruginosa CtpA is a carboxyl terminal-processing protease that partners with the outer membrane lipoprotein LbcA to degrade at least five cell wall-associated proteins, four of which are cell wall hydrolases. This activity plays an important role in supporting P. aeruginosa virulence in a mouse model of acute pneumonia. However, almost nothing is known about the molecular mechanisms underlying CtpA and LbcA function. Here, we used structural analysis to show that CtpA alone assembles into an inactive hexamer comprising a trimer of dimers, which limits its substrate access and prevents nonspecific degradation. The adaptor protein LbcA is a right-handed open spiral with 11 tetratricopeptide repeats, which might wrap around a substrate to deliver it to CtpA for degradation. By structure-guided mutagenesis and functional assays, we also showed that the interfaces of the CtpA trimer-of-dimers, and an N-terminal helix of LbcA, are important for LbcA-mediated substrate degradation by CtpA both in vitro and in vivo. This work improves our understanding of the molecular mechanism of a CTP within the C-terminal processing peptidase-3 group.

IMPORTANCECarboxyl-terminal processing proteases (CTPs) are found in all three domains of life. In bacteria, some CTPs have been associated with virulence, raising the possibility that they could be theraputic targets. However, relatively little is known about their molecular mechanisms of action. In Pseudomonas aeruginosa, CtpA supports virulence by working in complex with the outer membrane lipoprotein LbcA to degrade cell wall hydrolases. Here, we report structure-function analyses of CtpA and LbcA, which reveals that CtpA assembles into an inactive hexamer comprising a trimer of dimers. LbcA is monomeric, with an N-terminal region important for binding to and activating CtpA, followed by a spiral structure composed of 11 tetratricopetide repeats, which could wrap around a substrate for delivery to CtpA. This work provides the first structure of a CTP-3 group member, revealing a unique mutimeric arrangement and insight into how this important proteolytic system functions. Changes in body shape implicate cuticle stretch in C. elegans growth control Growth control establishes organism size, requiring mechanisms to sense and adjust growth during development. Studies of single cells revealed that size homeostasis uses distinct control methods. In multicellular organisms, mechanisms that regulate single cell growth must integrate control across organs and tissues during development to generate adult size and shape. We leveraged the roundworm Caenorhabditis elegans as a scalable and tractable model to collect precise growth measurements of thousands of individuals, measure feeding behavior, and quantify changes in animal size and shape during a densely sampled developmental time course. As animals transitioned from one developmental stage to the next, we observed changes in body aspect ratio while body volume remained constant. Then, we modeled a physical mechanism by which constraints on cuticle stretch could cause changes in C. elegans body shape. The model-predicted shape changes are consistent with those observed in the data. Theoretically, cuticle stretch could be sensed by the animal to initiate larval-stage transitions, providing a means for physical constraints to influence developmental timing and growth rate in C. elegans.

HighlightsO_LIBody size measurements of thousands of animals in a dense developmental time course
C_LIO_LIGrowth rate exhibits nonlinear dynamics in both length and width
C_LIO_LIChanges in body shape but not volume occur during periods of increased quiescence
C_LIO_LIDynamics of animal shape consistent with a length-based threshold in cuticle stretch
C_LIO_LIModeling of cuticle stretch dynamics suggests a novel mode for growth control
C_LI Naturally occurring combinations of receptors from single cell transcriptomics in endothelial cells VEGF inhibitor drugs have been successful, especially in ophthalmology, but not all patients respond to them. Combinations of drugs are likely to be needed for more effective therapies of angiogenesis-related diseases. In this paper we describe naturally occurring combinations of receptors in endothelial cells that might help to understand how cells communicate and to identify targets for drug combinations. We also develop and share a new software tool called DECNEO to identify them.

Single-cell gene expression data are used to identify a set of co-expressed endothelial cell receptors, conserved among species (mice and humans) and enriched, within a network, of connections to up-regulated genes. This set includes several receptors previously shown to play a role in angiogenesis. Multiple statistical tests from large datasets, including an independent validation set, support the reproducibility, evolutionary conservation and role in angiogenesis of these naturally occurring combinations of receptors. We also show tissue-specific combinations and, in the case of choroid endothelial cells, consistency with both well-established and recent experimental findings, presented in a separate paper.

The results and methods presented here advance the understanding of signaling to endothelial cells. The methods are generally applicable to the decoding of intercellular combinations of signals. Force profile of the two-handed hardstyle kettlebell swing in novice older adults: an exploratory profile BackgroundUnderstanding the force profile of an exercise increases clinical confidence when assessing the benefits and potential risks of a prescribed exercise. This exploratory study presents the force profile of the hardstyle kettlebell swing in novice older adults and compares peak force with kettlebell deadlifts. These data will help inform healthcare providers and coaches who are considering prescribing kettlebell exercises for older adults.

MethodsThirty-five community-dwelling males and females (59-79 years) were recruited, from applicants to participate in the BELL trial. Two-handed hardstyle swings were performed with 8-16 kg. Deadlifts were performed with 8-24 kg and 8-32 kg for females and males, respectively. Ground reaction force was obtained from a floor-mounted force platform. Pairwise comparisons of peak force, forward force, rate of force development, swing cadence, sex, and kettlebell mass, were investigated for the kettlebell swing, with representative force-time curves described. Pairwise comparisons of peak force, sex and kettlebell mass were investigated for the deadlift, with comparisons of peak force, kettlebell mass, and sex, between swings and deadlifts.

ResultsFor kettlebells up to 16 kg, paired samples T-tests show a large exercise effect ({delta} > 1.4) with peak force higher for swings than deadlifts. Data shows: (i) higher peak force during swings than deadlifts ({delta} = 1.77), reaching 4.5 (1.0) N.kg-1, (ii) peak force during an 8 kg swing was greater than a 32kg deadlift, (iii) negligible difference in normalised peak force between males and females performing kettlebell swings, but a moderately large effect size during deadlifts (males > females, {delta} = 0.69), (iv) mean rate of force development of 19.9 (4.7) N.s-1.kg-1 with a very weak, positive correlation with kettlebell mass (y = 14.4 + 0.32x), and trivial effect of sex, (v) mean forward force equal to 5.5% of vertical force during swings, increasing from 3.8 (1.6) % with 8 kg to 7.1 (2.6) % with 16 kg.

ConclusionDuring kettlebell swings, there is negligible difference in normalised net peak force between novice males and females using the same absolute loads. Where ground reaction force is a therapeutic target, kettlebell swings with an 8 kg kettlebell could have similar effects to much heavier deadlifts (>24 kg). Kettlebell swings performed with lighter loads, could provider similar therapeutic value to much heavier deadlifts, and may be a more appealing, affordable, and convenient option for older adults. Trade-offs between cost of ingestion and rate of intake drive defensive toxin use Animals that ingest toxins can become unpalatable and even toxic to predators and parasites through toxin sequestration. Because most animals rapidly eliminate toxins to survive their ingestion, it is unclear how populations transition from susceptibility and toxin elimination to tolerance and accumulation as chemical defense emerges. Studies of chemical defense have generally focused on species with active toxin sequestration and target-site insensitivity mutations or toxin-binding proteins that permit survival without necessitating toxin elimination. Here, we investigate whether animals that presumably rely on toxin elimination for survival can utilize ingested toxins for defense. We use the A4 and A3 Drosophila melanogaster fly strains from the Drosophila Synthetic Population Resource (DSPR), which respectively possess elevated and reduced metabolic nicotine resistance amongst DSPR fly lines. We find that ingesting nicotine increased A4 but not A3 fly survival against Leptopilina heterotoma wasp parasitism.Further, we find that despite possessing genetic variants that enhance toxin elimination, A4 flies accrued more nicotine than A3 individuals likely by consuming more media. Our results suggest that enhanced toxin metabolism can allow for greater toxin intake by offsetting the cost of toxin ingestion. Passive toxin accumulation that accompanies increased toxin intake may underlie the early origins of chemical defense. Cell cycle-independent integration of stress signals promotes Non-G1/G0 quiescence entry Cellular quiescence is a non-proliferative state required for cell survival under stress and during development. In most quiescent cells, proliferation is stopped in a reversible state of low Cdk1 kinase activity; in many organisms, however, quiescent states with high Cdk1 activity can also be established through still uncharacterized stress or developmental mechanisms. Here, we used a microfluidics approach coupled to phenotypic classification by machine learning to identify stress pathways associated with starvation-triggered high-Cdk1 quiescent states in Saccharomyces cerevisiae. We found that low- and high-Cdk1 quiescent states shared a core of stress-associated processes, such as autophagy, protein aggregation, and mitochondrial upregulation, but differed in the nuclear accumulation of the stress transcription factors Xbp1, Gln3, and Sfp1. The decision between low- or high-Cdk1 quiescence was controlled by cell cycle-independent accumulation of Xbp1, which acted as a time-delayed integrator of the duration of stress stimuli. Our results show how cell cycle-independent stress-activated factors promote cellular quiescence outside of G1/G0. Charting the human amygdala development across childhood and adolescence: Manual and automatic segmentation The developmental pattern of the amygdala throughout childhood and adolescence has been inconsistently reported in previous neuroimaging studies. Given the relatively small size of the amygdala on full brain MRI scans, discrepancies may be partly due to methodological differences in amygdalar segmentation. To investigate the impact of volume extraction methods on amygdala volume, we compared FreeSurfer, FSL and volBrain segmentation measurements with those obtained by manual tracing. The manual tracing method, which we used as the  gold standard, exhibited almost perfect intra- and inter-rater reliability. We observed systematic differences in amygdala volumes between automatic (FreeSurfer and volBrain) and manual methods. Specifically, compared with the manual tracing, FreeSurfer estimated larger amygdalae, and volBrain produced smaller amygdalae while FSL demonstrated a mixed pattern. The tracing bias was not uniform, but higher for smaller amygdalae. We further modeled amygdalar growth curves using accelerated longitudinal cohort data from the Chinese Color Nest Project. Trajectory modeling and statistical assessments of the manually traced amygdalae revealed linearly increasing and parallel developmental patterns for both girls and boys, although the amygdalae of boys were larger than those of girls. Compared to these trajectories, the shapes of developmental curves were similar when using the volBrain derived volumes. FreeSurfer derived trajectories had more nonlinearities and appeared flatter. FSL derived trajectories demonstrated an inverted U shape and were significantly different from those derived from manual tracing method. The use of amygdala volumes adjusted for total gray-matter volumes, but not intracranial volumes, resolved the shape discrepancies and led to reproducible growth curves curves between manual tracing and the automatic methods (except FSL). Our findings revealed steady growth of the human amygdala, mirroring its functional development across the school age. Methodological improvements are warranted for current automatic tools to achieve more accurate tracing of the amygdala at school age, calling for next generation tools. TeamTree analysis: a new approach to evaluate scientific production Advances in science and technology depend on the work of research teams and the publication of results through peer-reviewed articles representing a growing socio-economic resource. Current methods to mine the scientific literature regarding a field of interest focus on content, but the workforce credited by authorship remains largely unexplored, and appropriate measures of scientific production are debated. Here, a new bibliometric approach named TeamTree analysis is introduced that visualizes the development and composition of the workforce driving a field. A new citation-independent measure that scales with the H index estimates impact based on publication record, genealogical ties and collaborative connections. This author-centered approach complements existing tools to mine the scientific literature and to evaluate research across disciplines. Viral spillover risk in High Arctic increases in a glacierised watershed While many viruses have a single natural host, host restriction can be incomplete, hereby leading to spillovers to other host species. However, such spillover risks are difficult to quantify. As climate change is rapidly transforming environments, it is becoming critical to quantify the potential for spillovers. To address this issue, we resorted to an unbiased metagenomics approach, and focused on two environments, soil and lake sediments from Lake Hazen, the largest High Arctic freshwater lake in the world. We used DNA and RNA sequencing to reconstruct the lakes virosphere and its range of eukaryotic hosts, and estimated the spillover risk by measuring the congruence between the viral and the eukaryotic host phylogenetic trees. We show that spillover risk is higher in lake sediments than in soil and increased with runoff from glacier melt, a proxy for climate change. Should climate change also shift species range of potential viral vectors and reservoirs northwards, the High Arctic could become fertile ground for emerging pandemics. Deep learning: step forward to high-resolution in vivo shortwave infrared imaging Shortwave infrared window (SWIR: 1000-1700 nm) represents a major improvement compared to the NIR-I region (700-900 nm) in terms of temporal and spatial resolutions in depths down to 4 mm. SWIR is a fast and cheap alternative to more precise methods such as X-ray and opto-acoustic imaging. Main obstacles in SWIR imaging are the noise and scattering from tissues and skin that reduce the precision of the method. We demonstrate that the combination of SWIR in vivo imaging in the NIRIIb region (1500-1700 nm) with advanced deep learning image analysis allows to overcome these obstacles and making a large step forward to high resolution imaging: it allows to precisely segment vessels from tissues and noise, provides morphological structure of the vessels network, with learned pseudo-3D shape, their relative position, dynamic information of blood vascularization in depth in small animals and distinguish the vessels types: artieries and veins. For demonstration we use neural network IterNet that exploits structural redundancy of the blood vessels, which provides a useful analysis tool for raw SWIR images. Genomically Mined Acoustic Reporter Genes Enable In Vivo Monitoring of Tumors and Tumor-Homing Bacteria A major outstanding challenge in the fields of biological research, synthetic biology and cell-based medicine is the difficulty of visualizing the function of natural and engineered cells noninvasively inside opaque organisms. Ultrasound imaging has the potential to address this challenge as a widely available technique with a tissue penetration of several centimeters and spatial resolution below 100 {micro}m. Recently, the first genetically encoded acoustic reporters were developed based on bacterial gas vesicles to link ultrasound signals to molecular and cellular function. However, the properties of these first-generation acoustic reporter genes (ARGs) resulted in limited sensitivity and specificity for imaging in the in vivo context. Here, we describe second-generation ARGs with greatly improved acoustic properties and expression characteristics, identified through a phylogenetic screen of candidate gene clusters from diverse bacteria and archaea. The resulting constructs offer major qualitative and quantitative improvements, including much stronger ultrasound contrast, the ability to produce nonlinear signals distinguishable from background tissue, and stable long-term expression. We demonstrate the utility of these next-generation ARGs by imaging in situ gene expression in mouse models of breast cancer and tumor-homing probiotic bacteria, revealing the unique spatial distributions of tumor growth and colonization by therapeutic cells noninvasively in living subjects. Changes in Behavior and Salivary Serum Amyloid A in cows with Subclinical Mastitis The aim of this study was to identify detailed changes in behavior, and in salivary serum amyloid A (SAA), associated with subclinical mastitis. This included standard sickness behaviors (which form part of an adaptive response to conserve energy, minimize heat loss and facilitate recovery following infection and inflammation) and lesser-studied behaviors, that are biologically predicted to change at low-level infection, and therefore particularly relevant for detecting subclinical mastitis (here labelled  luxury behaviors). SAA is a physiological marker of systemic inflammation, with levels in milk and serum already known to increase during subclinical mastitis. We investigated whether the same was true for SAA in cow saliva. Data were collected for n = 34 commercial barn-housed dairy cows, tested in 17 matched-pairs. Each pair comprised a cow with subclinical mastitis (SCM) and a healthy control (CTRL), identified using somatic cell count (SCC) (SCM: SCC >200 x1000 cells/ml; CTRL: SCC <100 x1000 cells/ml). SCM cows were selected for study ad-hoc, at which point they were paired with a CTRL cow, based upon parity and calving date; consequently, the full data set was accrued over several months. Data were collected for each pair over 3 days: SCC (l4:00-l5:00h) Day 1; behavior (24h from 00:00h) Day 2; salivary serum amyloid-A (SAA) Day 3. We report, for the first time, that an increase in salivary SAA occurs during subclinical mastitis; SAA was higher in SCM cows and demonstrated a positive (weak) correlation with SCC. The behavioral comparisons revealed that SCM cows had reductions in activity (behavioral transitions and distance moved), social exploration, social reactivity (here: likelihood to move away/be displaced following receipt of agonism), performance of social grooming and head butts, and the receipt of agonistic non-contact challenges. In addition, SCM cows received more head swipes, and spent a greater proportion of time lying with their head on their flank than CTRL cows. SCM cows also displayed an altered feeding pattern; they spent a greater proportion of feeding time in direct contact with two conspecifics, and a lower proportion of feeding time at self-locking feed barriers, than CTRL cows. Behavioral measures were found to correlate, albeit loosely, with serum SAA in a direction consistent with predictions for sickness behavior. These included positive correlations with both lying duration and the receipt of all agonistic behavior, and negative correlations with feeding, drinking, the performance of all social and all agonistic behavior, and social reactivity. We conclude that changes in salivary SAA, social behavior, and activity offer potential in the detection of subclinical mastitis and recommend further investigation to substantiate and refine our findings.