Epigenetic modulation of selected immune response genes and altered functions of T lymphocytes and macrophages collectively contribute to autoimmune diabetes protection. We have previously demonstrated that treatment of female NOD mice with the histone deacetylase inhibitor Trichostatin A (TSA) bestowed irreversible protection against autoimmune diabetes. Herein we show that drug treatment diminished the infiltration of the pancreas with CD4+ and CD8+ T cells and Ly-6C+ monocytes. Significantly, TSA administration selectively repressed the expression of a set of genes exaggerated during diabetes and constitutively expressed primarily in the spleen and rarely in the pancreas. These genes encode lymphokines, macrophage-associated determinants, and transcription factors. Although the copy numbers of many histone deacetylases increased during diabetes in the spleen and pancreas, only those upregulated in the spleen were rendered sensitive to repression by TSA treatment. The T lymphocytes derived from drug-treated donors displayed diminished diabetogenic potential following transfer into immunodeficient NOD.scid mice. In the immunocompromised recipients, diabetes caused by the transfer of activated T lymphocytes from untreated diabetic mice was hampered by the co-transfer of highly purified splenic Ly-6C+ macrophages from drug-treated mice. However, the transfer of Ly-6C+ macrophages from drug-treated mice failed to block ongoing diabetes in wild-type NOD mice. These data demonstrate that the modified gene expression and functional alteration of T lymphocytes and macrophages collectively contribute to diabetes protection afforded by the histone modifier in female NOD mice.
A protein microarray-based in vitro transglutaminase assay platform for epitope mapping and peptide discovery: implication for transglutaminase-mediated immunodominant determination Transglutaminases (TGs) are a family of crosslinking enzymes catalyzing the formation of intra- and inter-molecular glutamine-lysine isopeptide bonds in a calcium dependent manner. Activation of transglutaminases is pathogenically associated with severe human diseases including neurodegenerations, cardiovascular diseases, and autoimmune diseases. Although continuous efforts determining the enzymes substrate preference have been witnessed, a high-throughput assay platform with the "omic" efficiency is still missing for the global identification of substrate-specific TG modification sites. In this study we report a protein microarray-based in vitro TG assay platform for rapid and large-scale (up to 30000 reactions per chip) determination of the glutamine (Q)-bearing TG modification motifs. With this platform we identified the Q16 in superoxide dismutase 1 and Q109 in alpha-synuclein as the primary modification sites for tissue transglutaminase (TG2), the most ubiquitous member of the enzyme family. Of particular interest, we found a close match between the modification motifs and published vaccine epitope sequences in alpha-synuclein, implying an essential and intrinsic role transglutaminase might play in the determination of immunodominant epitopes. Our data collectively suggest the glutamine and its follow-up five residues on the C terminal of a protein compose a minimal determinant motif for TG2 modification and the TG2 modification motifs determined by our platform could finally correspond to the substrates epitope sequences in antigen processing. To screen for site-specific interfering peptides and assist gene editing-based protein engineering, we optimized through onchip amino-acid scanning the TG2 modification motif QQIV in the extracellular matrix protein fibronectin and obtained 14 variants with significantly higher TG2 reactivity that might serve as the competitive inhibitor peptides and 1 with lower reactivity. We further confirmed this optimization approach with 12-mer peptides, the longest ones that could be synthesized on the chip. Taken together, our synthetic transglutaminase assay platform might be able to deliver a precise epitope blueprint for immunotherapeutic targeting and provide proof-of-concept and directional studies for TG-based peptide discovery and protein design.
Crystal structures of alphavirus nonstructural protein 4 (nsP4) reveal an intrinsically dynamic RNA-dependent RNA polymerase fold. Alphaviruses such as Ross River virus (RRV), chikungunya virus, Sindbis virus (SINV), and Venezuelan equine encephalitis virus are mosquito-borne pathogens that can cause arthritis or encephalitis diseases. Nonstructural protein 4 (nsP4) of alphaviruses possesses RNA-dependent RNA polymerase (RdRp) activity essential for viral RNA replication. No 3D structure has been available for nsP4 of any alphaviruses despite its importance for understanding alphaviral RNA replication and for the design of antiviral drugs. Here, we report crystal structures of the RdRp domain of nsP4 from both RRV and SINV determined at resolutions of 2.6 and 1.9 [A]. The structure of the alphavirus RdRp domain appears most closely related to RdRps from pestiviruses, noroviruses, and picornaviruses. Hydrogendeuterium exchange mass spectrometry (HDX-MS) and nuclear magnetic resonance (NMR) methods, showed that in solution, nsP4 is highly dynamic with an intrinsically disordered N-terminal domain. Both full-length nsP4 and the RdRp domain were capable to catalyze RNA polymerization. Structure-guided mutagenesis using a trans-replicase system identified nsP4 regions critical for viral RNA replication.

Key PointsO_LICrystal structures of alphavirus nsP4 RNA polymerase domain from RRV and SINV.
C_LIO_LInsP4 protein is highly dynamic with an intrinsically disordered N-terminal domain.
C_LIO_LIOptimized RNA elongation activity assay to facilitate antiviral discovery.
C_LI Embracing curiosity eliminates the exploration-exploitation dilemma Balancing exploration with exploitation is seen as a mathematically intractable dilemma that all animals face. In this paper, we provide an alternative view of this classic problem that does not depend on exploring to optimize for reward. We argue that the goal of exploration should be pure curiosity, or learning for learnings sake. Through theory and simulations we prove that explore-exploit problems based on this can be solved by a simple rule that yields optimal solutions: when information is more valuable than rewards, be curious, otherwise seek rewards. We show that this rule performs well and robustly under naturalistic constraints. We suggest three criteria can be used to distinguish our approach from other theories. The spatial configuration of biotic interactions shapes coexistence-area relation-ships in an annual plant community The increase of species richness with area is a universal phenomenon on Earth. How-ever, this observation contrasts with our poor understanding of how these species-area relationships (SARs) emerge from the collective effects of area, spatial heterogeneity, and local interactions. By combining a structuralist approach with five years of empirical observations in a highly-diverse Mediterranean grassland, we show that spatial hetero-geneity plays a little role in the accumulation of species richness with area in our system. Instead, as we increase the sampled area more species combinations are realized, and they coexist mainly due to direct pairwise interactions rather than by changes in single-species dominance or by indirect interactions. We also identify a small set of transient species with small population sizes that are consistently found across spatial scales. These findings empirically support the importance of the architecture of species interactions together with stochastic events for driving coexistence- and species-area relationships. Creatine transporter deficient rat model show motor dysfunction, cerebellar alterations and muscle creatine deficiency without muscle atrophy Creatine (Cr) is a nitrogenous organic acid and plays roles as fast phosphate energy buffer to replenish ATP, osmolyte, antioxidant, neuromodulator, and as a compound with anabolic and ergogenic properties in muscle. Cr is taken from the diet or endogenously synthetized by the enzymes AGAT and GAMT, and specifically taken up by the transporter SLC6A8. Loss-of-function mutations in the genes encoding for the enzymes or the transporter cause Cerebral Creatine Deficiency Syndromes (CCDS). CCDS are characterized by brain Cr deficiency, intellectual disability with severe speech delay, behavioral troubles, epilepsy and motor dysfunction. Among CCDS, the X-linked Cr transporter deficiency (CTD) is the most prevalent with no efficient treatment so far. Different animal models of CTD show reduced brain Cr levels, cognitive deficiencies and together they cover other traits similar to those of patients. However, motor function was poorly explored in CTD models and some controversies in the phenotype exist in comparison with CTD patients. Our recently described Slc6a8Y389C knock-in (KI) rat model of CTD showed mild impaired motor function, morphological alterations in cerebellum, reduced muscular mass, Cr deficiency and increased guanidinoacetate content in muscle, although no consistent signs of muscle atrophy. Our results indicate that such motor dysfunction co-occurred with both nervous and muscle dysfunction, suggesting that muscle strength and performance as well as neuronal connectivity might be affected by this Cr deficiency in muscle and brain. A Hierarchy of Biomolecular Proportional-Integral-Derivative Feedback Controllers for Robust Perfect Adaptation and Dynamic Performance Proportional-Integral-Derivative (PID) feedback controllers have been the most widely used controllers in industry for almost a century due to their good performance, simplicity, and ease of tuning. Motivated by their success in various engineering disciplines, PID controllers recently found their way into synthetic biology, where the design of feedback molecular control systems has been identified as an important goal. In this paper, we consider the mathematical realization of PID controllers via biomolecular interactions. We propose an array of topologies that offer a compromise between simplicity and high performance. We first demonstrate that different Proportional-Integral (PI) controllers exhibit different capabilities for enhancing the dynamics and reducing variance (cell-to-cell variability). Next, we introduce several derivative controllers that are realized based on incoherent feedforward loops acting in a feedback configuration. Alternatively, we show that differentiators can be realized by placing molecular integrators in a negative feedback loop--an arrangement that can then be augmented by PI components to yield PID feedback controllers. We demonstrate that the derivative component can be exploited for enhancing system stability, dramatically increasing the molecular control systems dynamic performance, and for reducing the noise effect on the output of interest. The PID controller features are established through various deterministic and stochastic analyses as well as numerical simulations. Finally, we provide an experimental demonstration using a recently developed hybrid setup, the cyberloop, where the controller is implemented in silico to control a biological genetic circuit in vivo. The large array of novel biomolecular PID controllers introduced here forms a basis for the design and construction of advanced high-performance biomolecular control systems that robustly regulate the dynamics of living systems. The neurogenic fate of the hindbrain boundaries relies on Notch-dependent asymmetric cell divisions The generation of cell diversity in the central nervous system occurs during embryogenesis and requires a precise balance between cell proliferation, commitment to specific fates, and further neuronal differentiation. The cellular and molecular mechanisms regulating this balance in the embryonic brain are still poorly understood. Here we study how the neurogenic capacity in the embryonic hindbrain is spatiotemporally allocated, and how the fate and the growth of the hindbrain boundary cells are regulated. By generating a CRISPR-based knock-in zebrafish transgenic line to specifically label the hindbrain boundaries, we unveiled that boundary cells undergo a functional transition to become neurogenic during hindbrain segmentation concurrently as they maintain the progenitor cell pool. Boundary cells engaged in neurogenesis coinciding with the onset of Notch signaling, which triggered their asymmetrical cell division. Our findings reveal that distinct neurogenic phases take place during hindbrain growth and suggest that boundary cells contribute to refine the final number, identity, and proportion of neurons in the brain.

SUMMARYHindbrain boundary cells undergo a functional transition to become neurogenic concurrently as they are maintained as a progenitor cell pool. This involves a behavioral switch from symmetrically dividing progenitor cells to asymmetrically dividing progenitors, which depends on Notch-activity. 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. Stochastic effects during the establishment of β-lactam resistant E. colimutants indicate conditions for collective resistance For antibiotic resistance to arise, new resistant mutants must establish in a bacterial population before they can spread via natural selection. Comprehending the stochastic factors that influence mutant establishment is crucial for a quantitative understanding of antibiotic resistance emergence. Here, we quantify the single-cell establishment probability of four Escherichia coli strains expressing {beta}-lactamase alleles with different activity against the antibiotic cefotaxime, as a function of antibiotic concentration in both unstructured (liquid) and structured (agar) environments. We show that concentrations well below the minimum inhibitory concentration (MIC) can substantially hamper establishment, particularly for highly resistant mutants. While the pattern of establishment suppression is comparable in both tested environments, we find greater variability in establishment probability on agar. Using a simple branching model, we investigate possible sources of this stochasticity, including environment-dependent lineage variability. Lastly, we use the single-cell establishment probability to predict each strains MIC in the absence of social interactions. We observe substantially higher measured than predicted MIC values, particularly for highly resistant strains, which indicates cooperative effects among resistant cells at large cell numbers, such as in standard MIC assays. Citation needed? Wikipedia and the COVID-19 pandemic BackgroundWith the COVID-19 pandemics outbreak, millions flocked to Wikipedia for updated information. Amid growing concerns regarding an "infodemic", ensuring the quality of information is a crucial vector of public health. Investigating if and how Wikipedia remained up to date and in line with science is key to formulating strategies to counter misinformation. Using citation analyses, we asked: which sources informed Wikipedias COVID-19-related articles before and during the pandemics first wave (January-May 2020).

ResultsWe found that coronavirus-related articles referenced trusted media sources and high-quality academic research. Moreover, despite a surge in COVID-19 preprints, Wikipedia had a clear preference for open-access studies published in respected journals and made little use of preprints. Building a timeline of English COVID-19 articles from 2001-2020 revealed a nuanced trade-off between quality and timeliness. It further showed how preexisting articles on key topics related to the virus created a framework for integrating new knowledge. Supported by a rigid sourcing policy, this "scientific infrastructure" facilitated contextualization and regulated the influx of new information. Lastly, we constructed a network of DOI-Wikipedia articles, which showed the shifting landscape of pandemic-related knowledge on Wikipedia and how academic citations create a web of shared knowledge supporting topics like COVID-19 vaccine development.

ConclusionsUnderstanding how scientific research interacts with the digital knowledge-sphere during the pandemic provides insight into how Wikipedia can facilitate access to science. It also reveals how, aided by what we term its "citizen encyclopedists", it successfully fended off COVID-19 disinformation and how this unique model may be deployed in other contexts. Population differences in the length and early-life dynamics of telomeres among European pied flycatchers Telomere length and shortening rate are increasingly used as biomarkers for long-term costs in ecological and evolutionary studies because of their relationships with survival and fitness. Both early-life conditions and growth, and later-life stressors can create variation in telomere shortening rate. Studies on between- population telomere length and dynamics are scarce, despite the expectation that populations exposed to varying environmental constraints would present divergent telomere length patterns. The pied flycatcher (Ficedula hypoleuca) is a passerine bird breeding across Eurasia (from Spain to western Siberia) and migrating through the Iberian Peninsula to spend the non-breeding period in sub-Saharan Africa. Thus, different populations show marked differences in migration distance. We studied the large-scale variation of telomere length and early-life dynamics in the pied flycatcher by comparing six European populations across a north- south gradient (Finland, Estonia, England, and Spain) predicting negative effect of migration distance on adult telomere length, and of nestling growth on nestling telomere dynamics. There were clear population differences in telomere length, with English birds from mid-latitudes having the longest telomeres. Telomere length did not thus show consistent latitudinal variation and was not linearly linked to differences in migration distance. Early-life telomere shortening rate tended to vary between populations. Fast growth was associated with shorter telomeres in the early life, but faster nestling growth affected telomeres more negatively in northern than southern populations. While the sources of between-population differences in telomere-related biology remain to be more intensively studied, our study illustrates the need to expand telomere studies at the between-population level. Comprehensive analysis of pyroptosis-associated in molecular classification, immunity and prognostic of glioma Integrative analysis was performed in the Chinese Glioma Genome Atlas and The Cancer Genome Atlas to describe the pyroptosis-associated molecular classification and prognostic signature in glioma. Pyroptosis-related genes were used for consensus clustering and to develop a prognostic signature. The immune statuses, molecular alterations and clinical features of differentially expressed genes were analyzed among different subclasses and risk groups. A lncRNA-miRNA-mRNA network was built, and drug sensitivity analysis was used to identify small molecular drugs for the identified genes. Glioma can be divided into two subclasses using 30 pyroptosis-related genes. Cluster 1 displayed high immune signatures and poor prognosis as well as high immune-related function scores. A prognostic signature based on 15 pyroptosis-related genes of the CGGA cohort can predict the overall survival of glioma and was well validated in the TCGA cohort. Cluster 1 had higher risk scores. The high-risk group had high immune cell and function scores and low DNA methylation of pyroptosis-related genes. The differences in pyroptosis-related gene mutations and somatic copy numbers were significant between the high-risk and low-risk groups. The ceRNA regulatory network uncovered the regulatory patterns of different risk groups in glioma. Nine pairs of target genes and drugs were identified. In vitro, CASP8 promotes the progression of glioma cells. Pyroptosis-related genes can reflect the molecular biological and clinical features of glioma subclasses. The established prognostic signature can predict prognosis and distinguish molecular alterations in glioma patients. Our comprehensive analyses provide valuable guidelines for improving glioma patient management and individualized therapy. Multi-echo Quantitative Susceptibility Mapping: How to Combine Echoes for Accuracy and Precision at 3 T PurposeTo compare different multi-echo combination methods for MRI quantitative susceptibility mapping (QSM). Given the current lack of consensus, we aimed to elucidate how to optimally combine multi-echo gradient-recalled echo (GRE) signal phase information, either before or after applying Laplacian-base methods (LBMs) for phase unwrapping or background field removal.

MethodsMulti-echo GRE data were simulated in a numerical head phantom, and multiecho GRE images were acquired at 3 T in ten healthy volunteers. To enable image-based estimation of GRE signal noise, five volunteers were scanned twice in the same session without repositioning. Five QSM processing pipelines were designed: one applied nonlinear phase fitting over echo times (TEs) before LBMs; two applied LBMs to the TE-dependent phase and then combined multiple TEs via either TE-weighted or signal-to-noise ratio (SNR)-weighted averaging; two calculated TE-dependent susceptibility maps via either multi-step or single-step QSM and then combined multiple TEs via magnitude-weighted averaging. Results from different pipelines were compared using visual inspection; summary statistics of susceptibility in deep gray matter, white matter, and venous regions; phase noise maps (error propagation theory); and, in the healthy volunteers, regional fixed bias analysis (Bland-Altman) and regional differences between the means (nonparametric tests).

ResultsNonlinearly fitting the multi-echo phase over TEs before applying LBMs provided the highest regional accuracy of{chi} and the lowest phase noise propagation compared to averaging the LBM-processed TE-dependent phase. This result was especially pertinent in high-susceptibility venous regions.

ConclusionFor multi-echo QSM, we recommend combining the signal phase by nonlinear fitting before applying LBMs. Intron-mediated induction of phenotypic heterogeneity Introns are universally present in the nuclear genomes of eukaryotes1. The budding yeast, an otherwise intron-poor species, preserves two sets of ribosomal protein (RP) genes differing primarily in their introns2-4. Despite recent findings on the role of RP introns under stress and starvation5-7, understanding the contribution of introns to ribosome regulation remains challenging. Here, combining isogrowth profiling8 with single-cell protein measurements9, we found that introns can mediate inducible phenotypic heterogeneity conferring a clear fitness advantage. Osmotic stress leads to bimodal expression of the small ribosomal subunit protein Rps22B, mediated by an intron in the 5 untranslated region of its transcript. The two resulting yeast subpopulations differ in their ability to cope with starvation. Low Rps22B protein levels resulted in prolonged survival under sustained starvation, while high Rps22B levels enabled cells to grow faster after transient starvation. Further, yeast growing at high sugar concentrations - similar to those in ripe grapes - exhibit bimodal Rps22B expression when approaching stationary phase. Differential intron-mediated regulation of RP genes thus provides a way to diversify the population when starvation looms in natural environments. Our findings reveal a new role for introns in inducing phenotypic heterogeneity in changing environments and suggest that duplicated RP genes in yeast contribute to resolving the evolutionary conflict between precise expression control and environmental responsiveness10. The genome of the cereal pest Sitophilus oryzae: a transposable element haven BackgroundThe rice weevil Sitophilus oryzae is one of the most important agricultural pests, causing extensive damage to cereal in fields and to stored grains. S. oryzae has an intracellular symbiotic relationship (endosymbiosis) with the Gram-negative bacterium Sodalis pierantonius and is a valuable model to decipher host-symbiont molecular interactions.

ResultsWe sequenced the Sitophilus oryzae genome using a combination of short and long reads to produce the best assembly for a Curculionidae species to date. We show that S. oryzae has undergone successive bursts of transposable element (TE) amplification, representing 72% of the genome. In addition, we show that many TE families are transcriptionally active, and changes in their expression are associated with insect endosymbiotic state. S. oryzae has undergone a high gene expansion rate, when compared to other beetles. Reconstruction of host-symbiont metabolic networks revealed that, despite its recent association with cereal weevils (30 Kyear), S. pierantonius relies on the host for several amino acids and nucleotides to survive and to produce vitamins and essential amino-acids required for insect development and cuticle biosynthesis.

ConclusionsHere we present the genome of an agricultural pest beetle, which may act as a foundation for pest control. In addition, S. oryzae may be a useful model for endosymbiosis, and studying TE evolution and regulation, along with the impact of TEs on eukaryotic genomes. Comparative analysis of cattle breeds as satellite cell donors for cultured beef BackgroundCultured meat is a promising new field with the potential for considerable environmental and animal welfare benefits. One technological approach to cultured meat production utilises the proliferative and differentiative capacity of muscle-derived satellite cells (SCs) to produce large volumes of cultured muscle tissue from small biopsies of donor animals. Differing genotypes between cattle breeds lead to predictable phenotypic traits, resulting in breeds being favoured for their respective meat or milk production characteristics in the livestock industry. However, whilst these breeds show significant differences in muscle growth, it is unclear whether the physiological differences observed between them in vivo are reflected in differences in SC behaviour in vitro, particularly with respect to proliferation, differentiation and cellular longevity, and hence whether particular breeds might represent preferred SC donors for a cultured beef bioprocess.

ResultsComparing SCs isolated from five breeds (Belgian Blue, Holstein Friesian, Galloway, Limousin and Simmental), we found that the proliferation rates were largely unaffected by the donor breed. In contrast, potentially meaningful differences were observed in the kinetics and extent of myogenic differentiation. Furthermore, whilst differentiation dropped for all breeds with increasing population doublings (PDs), SCs from Belgian Blue and Limousin cattle showed significantly longer retention of differentiation capacity over long-term passaging.

ConclusionSCs from all breeds were able to proliferate and differentiate, although Limousin and (particularly) Belgian Blue cattle, both breeds commonly used for traditional meat production, may represent preferred donors for cultured beef production. Global diversity and distribution of prophages are lineage-specific within the Ralstonia solanacearum plant pathogenic bacterium species complex Ralstonia solanacearum is a destructive plant pathogenic bacterium and the causative agent of bacterial wilt disease, infecting over 200 plant species worldwide. In addition to chromosomal genes, its virulence is mediated by mobile genetic elements including integrated DNA of bacteriophages, i.e., prophages, which may carry fitness-associated auxiliary genes or modulate host gene expression. Although experimental studies have characterised several prophages that shape R. solanacearum virulence, the global diversity, distribution, and wider functional gene content of R. solanacearum prophages is unknown. In this study, prophages were identified in a diverse collection of 192 R. solanacearum draft genome assemblies originating from six continents. Prophages were identified bioinformatically and their diversity investigated using genetic distance measures, gene content, GC, and total length. Prophage distribution was characterised using metadata on R. solanacearum geographic origin and lineage classification (phylotypes), and their functional gene content was assessed by identifying putative prophage-encoded auxiliary genes. In total, 343 intact prophages were identified, forming ten genetically distinct clusters. These included five prophage clusters belonging to the Inoviridae, Myoviridae, and Siphoviridae phage families, and five uncharacterised clusters, possibly representing novel, previously undescribed phages. The prophages had broad geographical distribution being present across multiple continents. However, they were generally host phylogenetic lineage-specific, and overall, prophage diversity was proportional to the genetic diversity of their hosts. The prophages contained a myriad of auxiliary genes involved in metabolism and virulence of both phage and bacteria. Our results show that while R. solanacearum prophages are highly diverse globally, they make lineage-specific contributions to the R. solanacearum accessory genome, which could have resulted from shared coevolutionary history. Robust and Remote Measurement of Heart Rate Based on A Surveillance Camera Arrhythmia is a marked symptom of many cardiovascular diseases (CVDs). The accurate and timely detection of heart rate can greatly reduce the harm of arrhythmia to people. However, it is still a challenge to robustly and remotely measure heart rate in daily life due to the changing environmental conditions during measurement, such as the varying light intensity, the movement of people, and the uncertain distance between the sensor to people. In this study, we propose a method to accurately measure human heart rate within a distance of 4.5 meters under different light intensities by simply using a surveillance camera. After a 20-second color video of a persons hand is captured by the camera, a method based on the Fast Fourier Transform (FFT) algorithm is designed to extract the blood volume pulse wave to calculate the heart rate. According to the comparison between the real heart rate and results measured by electrocardiography (ECG), the proposed method achieves an accuracy of 95.8% when the measurement is performed within a distance of 4.5 meters and 90% when within 5.0 meters. Our experiments show that when the illuminance varies between 100-1000 lux (lighting level indoor), we still get the correct results. Our experiments also demonstrate that the proposed method accurately obtain heart rate even when the light intensity is below 32 lux (300-500 lux in a workplace environment). The methods strong adaptability to changing environmental conditions makes it applicable to many scenarios, such as homes of the elderly, classrooms, and other public spaces. LEA motifs promote desiccation tolerance in vivo BackgroundCells and organisms typically cannot survive in the absence of water. However, there are some notable exceptions, including animals such as nematodes, tardigrades, rotifers, and some arthropods. One class of proteins known to play a role in desiccation resistance is the late embryogenesis abundant (LEA) proteins. These largely disordered proteins protect plants and animals from desiccation. A multitude of studies have characterized stress-protective capabilities of LEA proteins in vitro and in heterologous systems. However, the extent to which LEA proteins exhibit such functions in vivo, in their native contexts in animals, is unclear.

Furthermore, little is known about the distribution of LEA proteins in multicellular organisms or tissue-specific requirements in conferring stress protection.

ResultsTo study the endogenous function of an LEA protein in an animal, we created a true null mutant of C. elegans LEA-1, as well as endogenous fluorescent reporters of the protein. We confirmed that C. elegans lacking LEA-1 are sensitive to desiccation. LEA-1 mutant animals were also sensitive to heat and osmotic stress and were prone to protein aggregation. During desiccation, LEA-1 expression increased and became more widespread throughout the body. LEA-1 was required at high levels in body wall muscle for animals to survive desiccation and osmotic stress. We identified minimal motifs within C. elegans LEA-1 that are sufficient to increase desiccation survival of E. coli. To test whether such motifs are central to LEA-1s in vivo functions, we then replaced the sequence of lea-1 with these minimal motifs and found that C. elegans survived mild desiccation and osmotic stress at the same levels as worms with the full-length protein.

ConclusionsOur results provide insights into the endogenous functions and expression dynamics of an LEA protein in a multicellular animal. The results show that LEA-1 buffers animals from a broad range of stresses. Our identification of LEA motifs that can function in both bacteria and in a multicellular organism suggests the possibility of engineering LEA-1-derived peptides for optimized desiccation protection. Gradient-mixing LEGO robots for purifying DNA origami nanostructures of multiple components by rate-zonal centrifugation DNA origami purification is critical in emerging applications of functionalized DNA nanostructures from basic fundamental biophysics, nanorobots to therapeutics. Advances in DNA origami purification have led to the establishment of rate-zonal centrifugation (RZC) as a scalable, high-yield, and contamination-free approach to purifying DNA origami nanostructures. In RZC purification, a linear density gradient is created using viscous agents, such as glycerol and sucrose, to separate molecules based on their mass and shape during high-rpm centrifugation. However, current methods for creating density gradients are typically time-consuming because of their reliance on slow passive diffusion. Here, we built a LEGO gradient mixer to rapidly create a quasi-continuous density gradient with minimal layering of concentrations using simple rotational motion. We found that rotating two layers of different concentrations at an angle can reduce the diffusion time from a few hours to mere minutes. The instrument needed to perform the movement can be constructed from low-cost components, such as Arduino and LEGO Mindstorms pieces, and has comparable efficacy to commercial gradient mixers currently available. Our results demonstrate that the creation of a linear density gradient can be achieved with minimal labor, time, and cost with this machine. With the recent advances in DNA origami production, we anticipate our findings to further improve the viability of scaling up DNA origami purification in grams quantities. Our simple process enables automated large-scale purification of functionalized DNA origami more feasible in resource-constrained settings. Size dependent regeneration capacity of functionalized Capra ear derived micro-tissue scaffolds for treatment of cartilage defects Cartilage regeneration remains a great challenge in orthopedic treatment owing to their avascular in nature and lack of self-healing ability. Various clinical treatment options are widely used including osteochondral graft transplantation, micro-fracture, blood clot formation and tissue debridement to recover the damaged cartilage. In this circumstance, cartilage defect recovery via tissue engineered functional micro tissue delivery is becoming an emerging trend in musculoskeletal therapeutics. In this study, functional micro-scaffolds (MS) were generated from Capra ear cartilage, and were separated into size-wise groups. The scaffolds were decellularized via NaOH treatment. The cell adhesion study indicated that Capra adipose tissue derived mesenchymal stem cells (ADMSCs) adhesion is more in ~100 m MSs in comparison to 150-300 m MSs. It may be assumed that the cells are compatible to grow on fibrous surface area (100 m) in comparison to dense surface (150-300 m). Further, 100 m MSs were transformed into functional micro tissues (FMTs) in presence of high density ADMSCs in a hanging droplet culture system. The FMTs was transferred to F127 block polymer hydrogel for 3D culture. After 21 d cultures, the FMT clusters were evaluated for quantitative gene expression. To assess the in vivo cartilage defect regeneration potential, FMTs were delivered to rabbit auricular cartilage defect for 15, 30 and 60 d studies. The H&E-stained histological analysis showed that the cartilage defect is almost healed in 60 d study in comparison to 15 and 30 d study. Comparative toxicogenomics of glyphosate and Roundup herbicides by mammalian stem cell-based genotoxicity assays and molecular profiling in Sprague-Dawley rats Whether glyphosate-based herbicides (GBHs) are more potent than glyphosate alone at activating cellular mechanisms, which drive carcinogenesis remains controversial. As GBHs are more cytotoxic that glyphosate, we reasoned they may also be more capable of activating carcinogenic pathways. We tested this hypothesis by comparing the effects of glyphosate with Roundup GBHs both in vitro and in vivo. First, glyphosate was compared with representative GBHs namely MON 52276 (EU), MON 76473 (UK) and MON 76207 (USA) using the mammalian stem cell-based ToxTracker system. Here, MON 52276 and MON 76473, but not glyphosate and MON 76207, activated oxidative stress and unfolded protein responses. Second, molecular profiling of liver was performed in female Sprague-Dawley rats exposed to glyphosate or MON 52276 (both at 0.5, 50, 175 mg/kg bw/day glyphosate) for 90 days. MON 52276 but not glyphosate increased hepatic steatosis and necrosis. MON 52276 and glyphosate altered the expression of genes in liver reflecting TP53 activation by DNA damage and circadian rhythm regulation. Genes most affected in liver were similarly altered in kidneys. Small RNA profiling in liver showed decreased amounts of miR-22 and miR-17 from MON 52276 ingestion. Glyphosate decreased mir-30 while miR-10 levels were increased. DNA methylation profiling of liver revealed 5,727 and 4,496 differentially methylated CpG sites between the control and glyphosate and MON 52276 exposed animals respectively. Apurinic/apyrimidinic DNA damage formation in liver was increased with glyphosate exposure. Altogether, our results show that Roundup formulations cause more biological changes linked with carcinogenesis than glyphosate. DeLUCS: Deep Learning for (Unsupervised) Clustering of DNA Sequences We present a novel Deep Learning method for the Unsupervised Clustering of DNA Sequences (DeLUCS) that does not require sequence alignment, sequence homology, or (taxonomic) identifiers. DeLUCS uses Frequency Chaos Game Representations (FCGR) of primary DNA sequences, and generates "mimic" sequence FCGRs to self-learn data patterns (genomic signatures) through the optimization of multiple neural networks. A majority voting scheme is then used to determine the final cluster assignment for each sequence. The clusters learned by DeLUCS match true taxonomic groups for large and diverse datasets, with accuracies ranging from 77% to 100%: 2,500 complete vertebrate mitochondrial genomes, at taxonomic levels from sub-phylum to genera; 3,200 randomly selected 400 kbp-long bacterial genome segments, into clusters corresponding to bacterial families; three viral genome and gene datasets, averaging 1,300 sequences each, into clusters corresponding to virus subtypes. DeLUCS significantly outperforms two classic clustering methods (K-means++ and Gaussian Mixture Models) for unlabelled data, by as much as 47%. DeLUCS is highly effective, it is able to cluster datasets of unlabelled primary DNA sequences totalling over 1 billion bp of data, and it bypasses common limitations to classification resulting from the lack of sequence homology, variation in sequence length, and the absence or instability of sequence annotations and taxonomic identifiers. Thus, DeLUCS offers fast and accurate DNA sequence clustering for previously intractable datasets. 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. Rapid evolution of seed dormancy during sunflower de-domestication Hybridization between crops and their wild relatives may promote the evolution of de-domesticated (feral) weeds. Wild sunflower (Helianthus annuus L.) is typically found in ruderal environments, but crop-wild hybridization may facilitate the evolution of weedy populations. Using one crop-specific mitochondrial marker (CMS-PET1) and 14 nuclear SSR markers, we studied the origin and genetic diversity of a recently discovered weedy population of sunflower (named BRW). Then, using a resurrection approach, we tested for rapid evolution of weedy traits (seed dormancy, herbicide resistance, and competitive ability) by sampling weedy and wild populations 10 years apart (2007 and 2017). All the weedy plants present the CMS-PET1 cytotype, confirming their feral origin. At the nuclear markers, BRW showed higher genetic diversity than the cultivated lines and low differentiation with one wild population, suggesting that wild hybridization increased their genetic diversity. We found support for rapid evolution towards higher seed dormancy, but not for higher competitive ability or herbicide resistance. Our results highlight the importance of seed dormancy during the earliest stages of adaptation and show that crop-wild hybrids can evolve quickly in agricultural environments.