Methylation pattern of polymorphically imprinted nc886 is not conserved across mammalia BackgroundIn humans, the nc886 locus is a polymorphically imprinted metastable epiallele. Periconceptional conditions have an effect on the methylation status of nc886, and further, this methylation status is associated with health outcomes in later life, in line with the Developmental Origins of Health and Disease (DOHaD) hypothesis. Animal models would offer opportunities to study the associations between periconceptional conditions, nc886 methylation status and metabolic phenotypes further. Thus, we set out to investigate the methylation pattern of the nc886 locus in non-human mammals.

DataWe obtained DNA methylation data from the data repository GEO for mammals, whose nc886 gene included all three major parts of nc886 and had sequency similarity of over 80% with the human nc886. Our final sample set consisted of DNA methylation data from humans, chimpanzees, bonobos, gorillas, orangutangs, baboons, macaques, vervets, marmosets and guinea pigs.

ResultsIn human data sets the methylation pattern of nc886 locus followed the expected bimodal distribution, indicative of polymorphic imprinting. In great apes, we identified a unimodal DNA methylation pattern with 50% methylation level in all individuals and in all subspecies. In Old World monkeys, the between individual variation was greater and methylation on average was close to 60%. In guinea pigs the region around the nc886 homologue was non-methylated. Results obtained from the sequence comparison of the CTCF binding sites flanking the nc886 gene support the results on the DNA methylation data.

ConclusionsOur results indicate that unlike in humans, nc886 is not a polymorphically imprinted metastable epiallele in non-human primates or in guinea pigs, thus implying that animal models are not applicable for nc886 research. The obtained data suggests that the nc886 region may be classically imprinted in great apes, and potentially also in Old World monkeys, but not in guinea pigs.
Engineering of a fluorescent chemogenetic reporter with tunable color for advanced live-cell imaging Biocompatible fluorescent reporters with spectral properties spanning the entire visible spectrum are indispensable tools for imaging the biochemistry of living cells and organisms in real time. Here, we present the engineering of a fluorescent chemogenetic reporter with tunable optical and spectral properties. A collection of live-cell compatible fluorogenic chromophores with various electronic properties enables to generate bimolecular fluorescent assemblies that cover the visible spectrum from blue to red using a single protein tag engineered and optimized by directed evolution and rational design. We showed that the ability to tune the fluorescence color and properties through simple molecular modulation provides an unprecedent experimental versatility for imaging proteins in live cells, including delicate cultured hippocampal neurons, and in multicellular organisms. The ability to tune the spectral properties and fluorescence performance enables to match the spectral specifications and requirements of the most advanced imaging techniques, and allowed us to achieve efficient stimulated emission depletion (STED) nanoscopy of fusion proteins in live cells and live primary cultured neurons.
Mendelian Randomization for causal inference accounting for pleiotropy and sample structure using genome-wide summary statistics Mendelian Randomization (MR) is a valuable tool for inferring causal relationships among a wide range of traits using summary statistics from genome-wide association studies (GWASs). Existing summary-level MR methods often rely on strong assumptions, resulting in many false positive findings. To relax MR assumptions, ongoing research has been primarily focused on accounting for confounding due to pleiotropy. Here we show that sample structure is another major confounding factor, including population stratification, cryptic relatedness, and sample overlap. We propose a unified MR approach, MR-APSS, which (i) accounts for pleiotropy and sample structure simultaneously by leveraging genome-wide information; and (ii) allows to include more genetic variants with moderate effects as instrument variables (IVs) to improve statistical power without inflating type I errors. We first evaluated MR-APSS using comprehensive simulations and negative controls, and then applied MR-APSS to study the causal relationships among a collection of diverse complex traits. The results suggest that MR-APSS can better identify plausible causal relationships with high reliability. In particular, MR-APSS can perform well for highly polygenic traits, where the IV strengths tend to be relatively weak and existing summary-level MR methods for causal inference are vulnerable to confounding effects. A pilot study on the home range and movement patterns of the Andean Fox Lycalopex culpaeus (Molina 1782) in Cotopaxi National Park, Ecuador This study reports movement patterns and home range estimates of an Andean fox (Lycalopex culpaeus) in Cotopaxi National Park in Ecuador, representing the first GPS-tagging of the species. The GPS functioned well during the 197-day tracking period. Home range sizes ranged between 4.9 - 8.1 km2, depending on the estimation method. Movement speeds averaged 0.17 km/hr at day vs. 0.87 km/hr. at night, and distance traveled averaged 0.23 km at day vs. 0.89 km at night. These preliminary results highlight the importance of collecting unbiased, high-quality data which enables an enhanced understanding on mammal behavior and human/animal interaction. Fractal polymer with loops recapitulates key features of chromosome organization Chromosomes are exceedingly long polymers compacted in a cell nucleus. While it has long been suggested that mammalian chromosomes during interphase are folded into loops, experimental detection of such loops has remained a daunting task. The most comprehensive experimental information about chromosome spatial organization is provided by Hi-C experiments that measure the frequency of contacts between all chromosomal positions. The lack of a tractable physical model of a polymer folded into loops limits our ability to interpret experimental data. It thus remains unknown how to obtain accurate and quantitative information about the nature of chromosomal looping from Hi-C. Here, we introduce a model of a polymer with random loops, solve it analytically and extend it by simulations for real chains. Remarkably, our model faithfully reproduces complex shapes of experimental contact probability curves universal among mammalian cells. Furthermore, our model allows one to estimate sizes of randomly positioned chromosomal loops from experimental data. We also show that excluded volume in real chains can induce osmotic and topological repulsion between loops, stiffening interphase chromosomes. Thus, our new framework allows interpretation of experimental data and suggests that interphase chromosomes are crumpled polymers further folded into a sequence of randomly positioned loops. Loss of SUMO-specific protease 2 causes isolated glucocorticoid deficiency by blocking adrenal cortex zonal transdifferentiation SUMOylation is a dynamic posttranslational modification, that provides fine-tuning of protein function involved in the cellular response to stress, differentiation, and tissue development. In the adrenal cortex, an emblematic endocrine organ that mediates adaptation to physiological demands, the SUMOylation gradient is inversely correlated with the gradient of cellular differentiation raising important questions about its role in functional zonation and the response to stress. Considering that SUMO-specific protease 2 (SENP2), a deSUMOylating enzyme, is upregulated by ACTH/PKA signalling within the zona Fasciculata (zF), we generated mice with adrenal-specific Senp2 loss to address these questions. Disruption of SENP2 activity in steroidogenic cells leads to specific hypoplasia of the zF, a blunted reponses to ACTH and isolated glucocorticoid deficiency. Mechanistically, overSUMOylation resulting from SENP2 loss shifts the balance between ACTH/PKA and WNT/{beta}-catenin signalling leading to repression of PKA activity and ectopic activation of {beta}-catenin. At the cellular level, this blocks transdifferentiation of {beta}-catenin-positive zona Glomerulosa cells into zF cells and sensitises them to premature apoptosis. Our findings indicate that the SUMO pathway is critical for adrenal homeostasis and stress responsiveness. Balancing sensitivity and specificity in preclinical research The success of scientific discovery in preclinical drug development is based on the different roles of exploration and confirmation. Exploration requires sensitivity while confirmation needs to be specific. Based on published empirical data, we simulated two preclinical research trajectories to examine their effectiveness to detect true effects while keeping animal numbers low. We compare a conventional p-value based approach with a method based on an a priori determined smallest effect size of interest (SESOI). Using a SESOI increases transition rates from exploration to confirmation and results in higher detection rates across the trajectory. Specificity in the SESOI trajectory increases if the prior probability of true effects is low. We conclude that employing a SESOI is superior to a p-value based approach. Based on our findings, we propose a reconsideration of planning and conducting preclinical experiments, especially when the prior probability of true hypotheses is low. The impact of obesity; key features of raised intracranial pressure and clinical sequalae in rodents Elevated intracranial pressure (ICP) is observed in many brain disorders. Obesity has been linked to ICP pathogenesis in disorders such as idiopathic intracranial pressure (IIH). We investigated the effect of diet induced obesity (DIO) on ICP and clinically relevant sequelae.

Rats were fed either a control or high fat diet. Following weight gain long term ICP, headache behavior, body composition and retinal outcome were examined. Post-hoc analysis of retinal histology and molecular analysis of choroid plexus and trigeminal ganglion (TG) were performed.

DIO rats demonstrated raised ICP by 55% which correlated with the abdominal fat percentage and increased non-respiratory slow waves, suggestive of altered cerebral compliance. Concurrently, DIO rats demonstrated a specific cephalic cutaneous allodynia which negatively correlated with the abdominal fat percentage. This sensitivity was associated with increased expression of headache markers in TG. Additionally, DIO rats had an in vivo increased retinal nerve fiber layer thickness associated with raised ICP with a subsequent post-hoc demonstration of neuroretinal degeneration.

This study demonstrates for the first time that DIO leads to raised ICP and subsequent clinically relevant symptom development. This novel model of non-traumatic raised ICP could expand the knowledge regarding disorders with elevated ICP such as IIH. Modular biosynthesis of plant hemicellulose and its impact on yeast cells BackgroundThe carbohydrate polymers that encapsulate plants cells have benefited humans for centuries and have valuable biotechnological uses. In the past five years, exciting possibilities have emerged in the engineering of polysaccharide-based biomaterials. Despite impressive advances on bacterial cellulose-based hydrogels, comparatively little is known about how plant hemicelluloses can be reconstituted and modulated in cells suitable for biotechnological purposes.

ResultsHere, we assembled cellulose synthase-like A (CSLA) enzymes using an optimized Pichia pastoris platform to produce tunable heteromannan (HM) polysaccharides in yeast. By swapping the domains of plant mannan and glucomannan synthases, we engineered chimeric CSLA proteins that made {beta}-1,4-linked mannan in quantities surpassing those of the native enzymes while minimizing the burden on yeast growth. Prolonged expression of a glucomannan synthase from Amorphophallus konjac was toxic to yeast cells: reducing biomass accumulation and ultimately leading to compromised cell viability. However, an engineered glucomannan synthase as well as CSLA pure mannan synthases and a CSLC glucan synthase did not inhibit growth. Interestingly, Pichia cell size could be increased or decreased depending on the composition of the CSLA protein sequence. HM yield and glucose incorporation could be further increased by co-expressing chimeric CSLA proteins with a MANNAN-SYNTHESIS-RELATED (MSR) co-factor from Arabidopsis thaliana.

ConclusionThe results provide novel routes for the engineering of polysaccharide-based biomaterials that are needed for a sustainable bioeconomy. The characterization of chimeric cellulose synthase-like enzymes in yeast offers an exciting avenue to produce plant polysaccharides in a tunable manner. Furthermore, cells modified with non-toxic plant polysaccharides such as {beta}-mannan offer a modular chassis to produce and encapsulate sensitive cargo such as therapeutic proteins. CRSP: Comparative RNA-seq pipeline for species lacking both a reference genome and annotated transcriptome RNA sequencing (RNA-seq) has been a widely used high-throughput method to characterize transcriptomic dynamics spatiotemporally. However, typical RNA-seq data analysis pipelines depend on either a sequenced genome or annotated reference transcripts. This restriction makes the use of RNA-seq for species lacking both of sequenced genomes and reference transcripts challenging. To solve this problem, we developed CRSP, an RNA-seq pipeline integrating a comparative species strategy but not depending on a specific sequenced genome or reference transcripts. Benchmarking suggests the CRSP tool can achieve high accuracy to quantify gene expression levels.

AvailabilityCRSP and documents are available at: https://github.com/pjiang1105/CRSP. Discovery of highly potent pancoronavirus fusion inhibitors that also effectively inhibit COVID-19 variants from the UK (Alpha), South Africa (Beta), and India (Delta) We report here the discovery of several highly potent small molecules that showed low nM potency against SARS-CoV (IC50: as low as 13 nM), SARS-CoV-2 (IC50: as low as 23 nM), and MERS-CoV (IC50: as low as 76 nM) in pseudovirus based assays with excellent selectivity indices (SI: as high as > 5000) demonstrating their pancoronavirus inhibition. Some compounds also show 100% inhibition of CPE (IC100) at 1.25 {micro}M against an authentic SARS-CoV-2 (US_WA-1/2020). Furthermore, the most active inhibitors also potently inhibited variants of concerns (VOCs), such as the UK (B.1.1.7), South Africa (B.1.351), and Delta variant (B.1.617.2), originated in India. We confirmed that one of the potent inhibitors binds to the prefusion spike protein trimer of SARS-CoV-2 by SPR. Besides, we showed that they inhibit virus-mediated cell-cell fusion. The ADME data of one of the most active inhibitors, NBCoV1, show drug-like properties. In vivo PK of NBCoV1 in rats demonstrated excellent half-life (t1/2) of 11.3 h, mean resident time (MRT) of 14.2 h, and oral bioavailability. We expect the lead inhibitors to pave the way for further development to preclinical and clinical candidates. Host phenology can drive the evolution of intermediate virulence strategies in some parasites AO_SCPLOWBSTRACTC_SCPLOWThe traditional mechanistic trade-offs resulting in a negative correlation between transmission and virulence are the foundation of nearly all current theory on the evolution of parasite virulence. Several ecological factors have been shown to modulate the optimal virulence strategies predicted from mechanistic trade-off models, but these ecological factors have not yet been shown to be sufficient to explain the intermediate virulence strategies observed in any natural system. The timing of seasonal activity, or phenology, is a common factor that influences the types and impact of many ecological interactions but is difficult to incorporate into virulence evolution studies. We develop a mathematical model of a disease system with seasonal host activity to study the evolutionary consequences of host phenology on the virulence of obligate-killer parasite. Results from this model demonstrated that seasonal host activity is sufficient to drive the evolution of intermediate parasite virulence in some types of natural disease systems, even when a traditional mechanistic trade-off between transmission and virulence is not assumed in the modeling framework. The optimal virulence strategy in these systems can be determined by both the duration of the host activity period as well as the variation in the host emergence timing. Parasites with low virulence strategies are favored in environments with long host activity periods and in environments in which hosts emerge synchronously. The results demonstrate that host phenology can be sufficient to select for intermediate optimal virulence strategies, providing an alternative mechanism to account for virulence evolution in some natural systems. 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. 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. Map7D2 and Map7D1 facilitate microtubule stabilization through distinct mechanisms in neuronal cells Microtubule (MT) dynamics are modulated through the coordinated action of various MT-associated proteins (MAPs). However, the regulatory mechanisms underlying MT dynamics remain unclear. We show that the MAP7 family protein Map7D2 stabilizes MTs to control cell motility and neurite outgrowth. Map7D2 directly bound to MTs through its N-terminal half and stabilized MTs in vitro. Map7D2 localized prominently to the centrosome and partially on MTs in mouse N1-E115 neuronal cells, which expresses two of the four MAP7 family members, Map7D2 and Map7D1. Map7D2 loss decreased the resistance to the MT-destabilizing agent, nocodazole without affecting acetylated/detyrosinated stable MTs, suggesting that Map7D2 stabilizes MTs via direct binding. In addition, Map7D2 loss increased the rate of random cell migration and neurite outgrowth, presumably by disturbing the balance between MT stabilization and destabilization. Map7D1 exhibited similar subcellular localization and gene knock-down phenotypes to Map7D2. However, in contrast to Map7D2, Map7D1 was required for the maintenance of acetylated stable MTs. Taken together, our data suggest that Map7D2 and Map7D1 facilitate MT stabilization through distinct mechanisms in cell motility and neurite outgrowth. Amino acids targeted based metabolomics study in non-segmental Vitiligo: a pilot study IntroductionVitiligo is an asymptomatic disorder that results from the loss of pigments (melanin), causing skin or mucosal depigmentation and impairs beauty.

ObjectiveDue to the complexity of the pathogenesis of this disease and various theories including self-safety theory, oxidative stress, neurological theory and internal defects of melanocytes behind it, and finally, the vast role of amino acids in body metabolism and various activities of the body, amino acids targeted based metabolomics was set up to follow any fluctuation inside this disease.

MethodologyThe study of amino acid profiles in plasma of people with non-segmental vitiligo using a liquid chromatography equipped with fluorescent detector was performed to find remarkable biomarkers for the diagnosis and evaluation of disease severity of patients with vitiligo. Twenty-two amino acids derivatized with o-phthalaldehyde (OPA) and fluorylmethyloxycarbonyl chloride (FMOC), were precisely determined. Next, the concentrations of these twenty-two amino acids and their corresponding molar ratios were calculated in 37 patients (including 18 females and 19 males) and corresponding 34 healthy individuals (18 females and 16 males). Using R programing, the data were completely analyzed between the two groups of patients and healthy to find suitable and reliable biomarkers.

ResultsInterestingly, comparing the two groups, in the patient group, tyrosine, cysteine, the ratio of tyrosine to lysine and the ratio of cysteine to ornithine were increased while, arginine, lysine, ornithine and glycine ratios to cysteine have been decreased. These amino acids were selected for identification of patients with accuracy of detection of approximately 0.95 using the assessment of logistic regression.

ConclusionThese results indicate a disruption of the production of melanin, increased immune activity and oxidative stress, which are also involved in the effects of vitiligo. Therefore, these amino acids can be used as biomarker for the evaluation of risk, prevention of complications in individuals at risk and monitoring of treatment process. Co-option of immune effectors by the hormonal signalling system triggering metamorphosis in Drosophila melanogaster Insect metamorphosis is regulated by the production, secretion and degradation of two peripheral hormones: 20-hydroxyecdysone (ecdysone) and juvenile hormone (JH). In addition to their roles in developmental regulation, increasing evidence suggests that these hormones are involved in innate immunity processes, such as phagocytosis and the induction of antimicrobial peptide (AMP) production. AMP regulation includes systemic responses as well as local responses at surface epithelia that contact with the external environment. At pupariation, Drosophila melanogaster increases dramatically the expression of three AMP genes, drosomycin (drs), drosomycin-like 2 (drsl2) and drosomycin-like 5 (drsl5). We show that the systemic action of drs at pupariation is dependent on ecdysone signalling in the fat body and operates via the ecdysone downstream target, Broad-Z4. In parallel, ecdysone also regulates local responses, specifically through the activation of drsl2 expression in the gut. Finally, we confirm the relevance of this ecdysone dependent AMP expression for the control of bacterial persistence by showing that flies lacking drs expression in the fat body have higher bacterial persistence over metamorphosis. Together, our data emphasize the importance of the association between ecdysone signalling and immunity using in vivo studies and establish a new role for ecdysone at pupariation, which impacts developmental success by regulating the immune system in a stage-dependent manner. We speculate that this co-option of immune effectors by the hormonal system may constitute a pre-emptive mechanism to control bacterial numbers in the pupa at the core of metamorphosis evolution. Glucose inhibits haemostasis and accelerates diet-induced hyperlipidaemia in zebrafish larvae Hyperglycaemia damages the microvasculature in part through the reduced recruitment of immune cells and interference with platelet signalling, leading to poor wound healing and accelerated lipid deposition in mammals. We investigated the utility of zebrafish larvae to model the effect of glucose on neutrophil and macrophage recruitment to a tail wound, wound-induced haemostasis, and chicken egg yolk feed challenge-induced hyperlipidaemia by supplementing larvae with exogenous glucose by immersion or injection. Neither method of glucose supplementation affected the recruitment of neutrophils and macrophages following tail transection. Glucose injection reduced thrombocyte retention and fibrin plug formation while only thrombocyte retention was reduced by glucose immersion following tail transection. We observed accelerated lipid accumulation in glucose-injected larvae challenged with high fat chicken egg yolk feeding. Our study identifies conserved and divergent effects of high glucose on inflammation, haemostasis, and hyperlipidaemia in zebrafish larvae compared to mammals. A chromosome-level genome assembly of a maize elite breeding line Dan340 The maize cultivar Dan340 is an excellent backbone inbred line of Luda Red Cob Group with several desirable characters, such as disease resistance, lodging resistance, high combining ability, wide adaptability and so on. In this study, we constructed a high-quality chromosome-level reference genome for Dan340 by combining PacBio long HiFi sequencing, Illumina short reads and chromosomal conformational capture (Hi-C) sequencing reads. The final assembly of Dan340 genome was 2,348.72 Mb, including 2,738 contigs and 2,315 scaffolds with a N50 of 41.49 Mb and 215.35 Mb, respectively. The percent of high quality Illumina reads mapped to the reference genome was up to 97.48%. The assembly of this genome will not only facilitate our understanding about intraspecific genome diversity in maize, but also provides a novel resource for maize breeding.

Key findingsO_LIThe final assembly of Dan340 genome was 2,348.72 Mb, including 2,738 contigs and 2,315 scaffolds with a N50 of 41.49 Mb and 215.35 Mb, respectively.
C_LIO_LIThe percent of reads mapped to the reference genome was up to 97.48%.
C_LIO_LIThe results showed that 96.84% of the plant single-copy orthologues were complete. Complete single-copy and multi copy genes accounted for 87.36% and 9.48% of the genes, respectively. Taken together, these results indicated that our Dan340 genome assembly presented high quality and coverage.
C_LI Bacterial and fungal communities experience rapid secondary succession during the first year following a wildfire in a California chaparral. The rise in wildfire frequency and severity across the globe has increased interest in secondary succession. However, despite the role of soil microbial communities in the regeneration of post-fire vegetation and biogeochemical cycling, the lack of measurements immediately post-fire and at high temporal resolution has resulted in a limited understanding of microbial secondary succession. To fill this knowledge gap, we sampled soils at 17, 25, 34, 67, 95, 131, 187, 286, and 376 days after a Southern California wildfire in fire-adapted chaparral shrublands. We assessed bacterial and fungal biomass with qPCR of 16S and 18S and richness and composition with Illumina MiSeq sequencing of 16S and ITS2 amplicons. Fire severely reduced bacterial biomass by 47% and richness by 46%, but the impacts were stronger for fungi, with biomass decreasing by 86% and richness by 68%. While bacterial biomass and richness oscillated in response to precipitation, the declines in biomass and richness for both groups persisted for the entire year. In contrast to the unburned plots, burned fungal and bacterial communities experienced rapid succession, with 5-6 compositional turnover periods. Analogous to plants, "fire-loving" pyrophilous microbes increased post-fire and drove successional dynamics. Interestingly, many of the same pyrophilous taxa found in forests worldwide also dominated post-fire chaparral and traded off markedly in abundance over time. For fungi, an initial peak in the Basidiomycete yeast Geminibasidium traded off with the filamentous Ascomycetes Pyronema, Aspergillus, and Penicillium. For bacteria, the Proteobacteria genus Massilia dominated burned plots throughout the year, but the following most dominant bacteria, the Firmicute Bacillus and Proteobacteria Noviherbaspirillum, traded off in abundance over time. Our unprecedented high-resolution temporal sampling allowed us to capture post-fire microbial secondary successional dynamics and show that putative tradeoffs in thermotolerance, colonization, and competition among dominant pyrophilous microbes control microbial succession with possible implications for ecosystem function. Testing for phylogenetic signal in single-cell RNA-seq data AO_SCPLOWBSTRACTC_SCPLOWPhylogenetic methods are emerging as a useful tool to understand cancer evolutionary dynamics, including tumor structure, heterogeneity, and progression. Most currently used approaches utilize either bulk whole genome sequencing (WGS) or single-cell DNA sequencing (scDNA-seq) and are based on calling copy number alterations and single nucleotide variants (SNVs). Here we explore the potential of single-cell RNA sequencing (scRNA-seq) to reconstruct cancer evolutionary dynamics. scRNA-seq is commonly applied to explore differential gene expression of cancer cells throughout tumor progression. The method exacerbates the single-cell sequencing problem of low yield per cell with uneven expression levels. This accounts for low and uneven sequencing coverage and makes SNV detection and phylogenetic analysis challenging. In this paper, we demonstrate for the first time that scRNA-seq data contains sufficient evolutionary signal and can be utilized in phylogenetic analyses. We explore and compare results of such analyses based on both expression levels and SNVs called from scRNA-seq data. Both techniques are shown to be useful for reconstructing phylogenetic relationships between cells, reflecting the clonal composition of a tumor. Both standardized expression values and SNVs appear to be equally capable of reconstructing a similar pattern of phylogenetic relationship. This pattern is stable even when phylogenetic uncertainty is taken in account. Our results open up a new direction of somatic phylogenetics based on scRNA-seq data. Further research is required to refine and improve these approaches to capture the full picture of somatic evolutionary dynamics in cancer. PKR-mediated Stress Response Enhances Dengue and Zika Virus Replication The mechanisms by which Flaviviruses use non-canonical translation to support their replication in host cells are largely unknown. Here we investigated how the integrated stress response (ISR), which promotes translational arrest by eIF2 phosphorylation (p-eIF2), regulates Flavivirus replication. During Dengue virus (DENV) and Zika virus (ZIKV) infection, eIF2 activation peaked at 24 hours post infection and was dependent on PKR but not type I interferon. The ISR is activated downstream of p-eIF2 during infection with either virus, but translation arrest only occurred following DENV infection. Despite this difference, both DENV and ZIKV replication was impaired in cells lacking PKR, independent of IFN-I/NF-kB signaling or cell viability. By using a ZIKV 5UTR reporter system as a model, we found that this region of the genome is sufficient to promote an enhancement of viral mRNA translation in the presence of an active ISR. Together we provide evidence that Flaviviruses escape ISR translational arrest and co-opt this response to increase viral replication. Design, Mutate, Screen: High-throughput creation of genetic clocks with different period-amplitude characteristics Directed evolution has become an invaluable tool in protein engineering and has also greatly influenced the construction of synthetic gene circuits. The ability to generate diversity at precise targets for directed evolution approaches has improved vastly, allowing researchers to create large, specific mutant libraries with relative ease. Screening approaches for large mutant libraries have similarly come a long way, especially when the desired behavior can easily be tested for with static, single time-point assays. For more complex gene circuits with dynamic phenotypes that change over time, directed evolution approaches to controlling and tuning circuit behavior have been hindered by the lack of sufficiently high-throughput screening methods to isolate variants with desired characteristics. Here we utilize directed mutagenesis and multiplexed microfluidics to develop a workflow for creating, screening and tuning dynamic gene circuits that operate at the population level. Specifically, we create a mutant library of an existing oscillator, the synchronized lysis circuit, and tune its dynamics while uncovering principles regarding its behavior. Lastly, we utilize this directed evolution workflow to construct a new synchronized genetic oscillator that exhibits robust dynamics over long time scales. Integrative Modelling of Signalling Network Dynamics Identifies Cell Type-selective Therapeutic Strategies for FGFR4-driven Cancers Oncogenic FGFR4 signalling represents a potential therapeutic target in many cancer types, including triple negative breast cancer (TNBC) and hepatocellular carcinoma (HCC). However, resistance to single-agent therapy directed at FGFR4 remains a major challenge, prompting the need to identify more effective combinatorial therapeutic strategies. Here, we integrated computational network modelling and experimental validation to characterise dynamic reprogramming of the FGFR4 signalling network in TNBC following FGFR4 kinase inhibition. We found that AKT, which signals downstream of FGFR4, displayed a rapid and potent reactivation following FGFR4 targeting. Through model-based simulation and systematic prediction of the effect of co-targeting specific network nodes, we predicted, and validated experimentally, strong synergism of co-targeting FGFR4 and particular ErbB kinases or AKT, but not the upstream kinase PI3K. Further, incorporation of protein expression data from hundreds of cancer cell types enabled us to adapt our model to other diverse cellular contexts, leading to the prediction that while AKT rebound occurs frequently, it is not a general phenomenon. Instead, ERK is reactivated in a subset of cell types, including the FGFR4-driven HCC cell line Hep3B. This was subsequently corroborated, and moreover, co-targeting FGFR4 and MEK in Hep3B cells markedly enhanced inhibition of cell proliferation. Overall, these findings provide novel insights into the dynamics of drug-induced network remodelling in cancer cells, highlight the impact of protein expression heterogeneity on network response to targeted therapy and identify candidate cell type-selective combination treatments for FGFR4-driven cancer. Plant biomechanics and resilience to environmental changes are controlled by specific lignin chemistries in each vascular cell type and morphotype The biopolymer lignin, deposited in the cell walls of vascular cells, is essential for long-distance water conduction and structural support of plants. Independently of the species, each different vascular cell type contains a conserved lignin chemistry with specific aromatic and aliphatic substitutions. Yet, the biological role of this conserved and specific lignin chemistry for each cell type remained unclear. Herein, we investigate the role of specific lignin chemistries for cellular function by producing single cell analyses on vascular cell morphotypes, all enabling sap conduction but differing in morphology. We found that specific lignin chemistries accumulate in each morphotype. Moreover, lignin accumulates dynamically, increasing in quantity and changing composition, to alter the cell wall biomechanics of each morphotype during their maturation. For similar aromatic substitution, residues with alcohol aliphatic functions increased stiffness whereas aldehydes increased flexibility. Modifying this specific lignin chemistry impairs the cell wall biomechanics of each morphotype and consequently reduces their capacity to optimally conduct water in normal conditions, and to recover from drought. Altogether, lignin chemistry is differently controlled for each sap conducting cell types during their maturation to dynamically adjust their biomechanics and hydraulic properties to adapt to developmental and environmental constraints. Scaling effects of temperature on parasitism from individuals to host-parasite systems Parasitism is expected to change in a warmer future, but whether warming leads to substantial increases in parasitism remains unclear. Understanding how warming effects on parasitism in individual hosts (e.g., parasite load) translate to effects on population-level parasitism (e.g., prevalence, R0) remains a major knowledge gap. We analyzed the temperature dependence of parasitism at both host and population levels in thirteen empirical vector-borne host-parasite systems and found a strong, significant positive correlation between the thermal optima of individual- and population-level parasitism. We also found a significant, positive correlation in eleven environmentally-transmitted parasite systems, though several of these systems exhibited thermal optima >5{o}C apart between individual and population levels. Similarly, parasitism thermal optima were close to host performance thermal optima in vector-borne systems but not in environmentally-transmitted systems. We then adapted and simulated simple models for both transmission modes and found a similar pattern to the empirical systems: thermal optima in vector-borne systems were more strongly correlated across scales compared to environmentally-transmitted systems. Generally, our results suggest that information on the temperature-dependence, and specifically the thermal optimum, at either the individual- or population-level should provide a useful--though not quantitatively exact--baseline for predicting temperature dependence at the other level, especially in vector-borne parasite systems. Environmentally-transmitted parasitism may operate by a different set of rules, in which temperature-dependence is decoupled in some systems, requiring the need for trait-based studies of temperature-dependence at individual and population levels.