Biology subjects

Allen, B.

Publications and source records attributed to Allen, B..

16 recordsLinked to original sources

bioRxiv2025

Intermediate tissue oxygen level is required to observe murine FLASH skin sparing

ObjectiveThis study evaluated the hypothesis that baseline tissue oxygen (pO2) would modulate FLASH toxicity sparing in murine skin, using a wide range of pO2 values, with ultra-high dose rate (UHDR) versus conventional dose rate (CDR) irradiation. ApproachMurine leg tissue pO2 was systematically varied and measured during irradiation from a FLASH Mobetron 9 MeV linac at 25 Gy, comparing UHDR ({approx}240 Gy/s) to CDR ({approx}0.16 Gy/s), for radiation induced skin toxicity outcomes. Baseline tissue pO2 was systematically modulated in 5 different treatment cohorts, using different ranges of inhaled gas (room air, 100% oxygen, or carbogen) and through varying limb vascular compression (partial or full). Radiolytic oxygen consumption, gO2 (mmHg/Gy), was quantified in vivo, and induced macroscopic skin toxicity was scored daily post treatment. Main ResultsFLASH skin sparing was observed at a fixed dose of 25 Gy in groups with partial leg clamping (pO2{approx}7{+/-}4mmHg), inhaled air (pO2{approx}12{+/-}6mmHg) and 100% oxygen (pO2{approx}16{+/-}4mmHg), while reduction in ulceration progression was significant only in the air inhalation group. No FLASH effect was observed under anoxic conditions, via complete blood flow occlusion (pO2{approx}0{+/-}1mmHg), or when modulated by inhaled carbogen (pO2{approx}21{+/-}7mmHg). In vivo measurements of radiolytic oxygen consumption, gO2, correlated to initial pO2 under UHDR conditions (pO2{approx}4-16mmHg), with ulceration predominantly occurring at pO2 values above 16mmHg. Inspired carbogen induced the highest pO2 at which point there was no FLASH sparing, for any dose groups between 25 to 15 Gy, despite having large changes in damage with dose. At the specific dose level of 25 Gy studied, the toxicity scores under anoxia for both UHDR and CDR were low (toxicity scores < 1) with no differences observed. SignificanceThese findings point to the fact that moderate and low tissue pO2 is associated with diminished oxygen-mediated damage at UHDR but not CDR, seen with inspired room air or 100% oxygen. Anoxic and hyperoxic murine skin are associated with minimal and maximal radiation damage respectively, but also exhibit no apparent FLASH toxicity sparing effect, with further investigation warranted into if the FLASH toxicity sparing effect persists at higher doses under anoxia.

cancer biology↗

bioRxiv2025

High-Throughput Screening of FRET-Based Protein Rulers Using a Hyperspectral Microcapillary Array

FRET (Fluorescence Resonance Energy Transfer) is a versatile technique used in biology, chemistry, and materials science that involves distance-dependent energy transfer between light-sensitive molecules. Current FRET screening methods are limited when handling large libraries. Challenges include a low dynamic range, overlapping spectra, multiple fluorophores, and high background signals. A high-throughput FRET approach enables rapid, multiplexed screening of interactions, conformations, and dynamics with accurate, real-time measurements. We developed a hyperspectral high-throughput microcapillary array (HyCAP), which is ideal for sorting large clone libraries based on fluorescence assays. Our platform provides sensitive, high-resolution hyperspectral imaging with a throughput of approximately 105 per array. FRET protocols can be used to study variations in protein structure and assemblies. HyCAP demonstrates potential in fluorophore engineering, including emission changes related to environment (such as pH, enzymatic activity, and temperature), stoichiometry differences, fluorophore orientation, and photobleaching through time-resolved experiments.

synthetic biology↗

bioRxiv2025

Adaptive dynamics of sexual signaling and discrimination under different mating conditions

A wide variety of animals engage in same-sex sexual behavior (SSB). Some instances of SSB appear to arise from individuals mating indiscriminately without regard to the sex of their partners. Theory suggests that indiscriminate mating strategies can be selected if sexes are not completely distinguishable--that is, without strong signals of sexual identity. We model the coevolution of sexual signaling and discrimination using the adaptive dynamics approach to long-term trait evolution. Our model relaxes two major assumptions from past phenomenological theory. First, we consider a mechanistic mating process with tunable encounter rates and mating costs. Second, we vary reproductive investment roles to allow for better correspondence with empirical examples of SSB. We confirm that coevolution of sexual signaling and discrimination can lead to two distinct equilibria: one with no sexual signals and indiscriminate mating; the other with perfect signaling and exclusively different-sex sexual behavior. Selection for indiscriminate mating is strongest at very low or very high encounter rates. Mating costs lead to selection for reduced overall mating rate, but this can lead to either more or less SSB, depending on how this reduction is achieved. Overall, our model highlights the importance of ecologically relevant parameters for the evolution of SSB.

evolutionary biology↗

bioRxiv2024

The probability and duration of immigration in microbial communities

Immigration is a fundamental feature of microbial communities. We propose a method to determine the probability (Pi) that a number of immigrants (i) can attain an abundance N using the ratio of the probabilities of death q and division or "birth" p and the gamblers ruin equation: Pi= (1-(q/p)i/(1-(q/p)N). We estimate the probability of successful bioaugmentation or transplantation, the fate of a mutation infection and extinction. For example, an inoculum of 108 bacteria with a q/p of 1.00000001 has a 10-43 chance of attaining an abundance of 1010. The immigration parameter used in neutral models, m is 1/(1-q/p). We calculated the long-term average value of m and q/p in a wastewater treatment plant. The value of m varies by >5 orders of magnitude, with a curious bimodal distribution. However, all the values of q/p are very close to, but greater than, 1. We expect the long-term average value of q/p to be [~]1 in all stable microbial communities. In the absence of migration, bacterial populations with a q/p [≥]1 will go extinct with probability 1. The link between q/p and infectious dose is known and we demonstrate that, in principle, the gamblers ruin equation can estimate the infectious dose in naturally occurring infections, using Vibrio cholerae "carriers" to illustrate the point. We use the ratio q/p to estimate the time (measured in events or solar time) for a given change in abundance to happen. When q/p=1, extinction in even a small microbial population will take thousands of years.

microbiology↗

bioRxiv2024

Density-dependent network structuring within and across wild animal systems

High population density should drive individuals to more frequently share space and interact, producing better-connected spatial and social networks [1-4]. Although this theory is fundamental to our understanding of disease dynamics [2,5-8], it remains unconfirmed how local density generally drives individuals positions within their networks, which reduces our ability to understand and predict density-dependent processes [4,9,10]. Here we provide the first general evidence that density drives greater network connectedness at fine spatiotemporal scales, at the scale of individuals within wild animal populations. We analysed 36 datasets of simultaneous spatial and social behaviour in >58,000 individual animals, spanning 30 species of fish, reptiles, birds, mammals, and insects. 80% of systems exhibited strong positive relationships between local density and network centrality. However, >80% of relationships were nonlinear and 75% became shallower at higher values, signifying that demographic and behavioural processes counteract densitys effects, thereby producing saturating trends [11-15]. Densitys effect was much stronger and less saturating for spatial than social networks, such that individuals become disproportionately spatially connected rather than socially at higher densities. Consequently, ecological processes that depend on spatial connections (e.g. indirect pathogen transmission, resource competition, and territory formation) are likely more density-dependent than those involving social interactions (e.g. direct pathogen transmission, aggression, and social learning). These findings reveal fundamental ecological rules governing societal structuring, with widespread implications. Identifying scaling rules based on processes that generalise across systems, such as these patterns of density dependence, might provide the ability to predict network structures in novel systems.

ecology↗

bioRxiv2023

Microbiota-dependent indole production is required for the development of collagen-induced arthritis

Altered tryptophan catabolism has been identified in inflammatory diseases like rheumatoid arthritis (RA) and spondyloarthritis (SpA), but the causal mechanisms linking tryptophan metabolites to disease are unknown. Using the collagen-induced arthritis (CIA) model we identify alterations in tryptophan metabolism, and specifically indole, that correlate with disease. We demonstrate that both bacteria and dietary tryptophan are required for disease, and indole supplementation is sufficient to induce disease in their absence. When mice with CIA on a low-tryptophan diet were supplemented with indole, we observed significant increases in serum IL-6, TNF, and IL-1{beta}; splenic ROR{gamma}t+CD4+ T cells and ex vivo collagen-stimulated IL-17 production; and a pattern of anti-collagen antibody isotype switching and glycosylation that corresponded with increased complement fixation. IL-23 neutralization reduced disease severity in indole-induced CIA. Finally, exposure of human colon lymphocytes to indole increased expression of genes involved in IL-17 signaling and plasma cell activation. Altogether, we propose a mechanism by which intestinal dysbiosis during inflammatory arthritis results in altered tryptophan catabolism, leading to indole stimulation of arthritis development. Blockade of indole generation may present a novel therapeutic pathway for RA and SpA.

immunology↗

bioRxiv2022

Glia-neuron coupling via a bipartite sialylation pathway promotes neural transmission and stress tolerance in Drosophila

Modification by sialylated glycans can affect protein functions, underlying mechanisms that control animal development and physiology. Sialylation relies on a dedicated pathway involving evolutionarily conserved enzymes, including CMP-sialic acid synthetase (CSAS) and sialyltransferase (SiaT) that mediate the activation of sialic acid and its transfer onto glycan termini, respectively. In Drosophila, CSAS and DSiaT genes function in the nervous system, affecting neural transmission and excitability. We found that these genes function in different cells: the function of CSAS is restricted to glia, while DSiaT functions in neurons. This partition of the sialylation pathway allows for regulation of neural functions via a glia-mediated control of neural sialylation. The sialylation genes were shown to be required for tolerance to heat and oxidative stress and for maintenance of the normal level of voltage-gated sodium channels. Our results uncovered a unique bipartite sialylation pathway that mediates glia-neuron coupling and regulates neural excitability and stress tolerance. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=152 SRC="FIGDIR/small/486211v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@15b8de3org.highwire.dtl.DTLVardef@1c037e8org.highwire.dtl.DTLVardef@9674bborg.highwire.dtl.DTLVardef@1c9c010_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2021

Looking for lipases and lipolytic organisms in low-temperature anaerobic reactors treating domestic wastewater

Poor lipid degradation limits low-temperature anaerobic treatment of domestic wastewater even when psychrophiles are used. We combined metagenomics and metaproteomics to find lipolytic bacteria and their potential, and actual, cold-adapted extracellular lipases in anaerobic membrane bioreactors treating domestic wastewater at 4{degrees}C and 15{degrees}C. Of the 40 recovered putative lipolytic metagenome-assembled genomes (MAGs), only three (Chlorobium, Desulfobacter, and Mycolicibacterium) were common and abundant (relative abundance [≥] 1%) in all reactors. Notably, some MAGs that represented aerobic autotrophs contained lipases. Therefore, we hypothesised that the lipases we found are not always associated with exogenous lipid degradation and can have other roles such as polyhydroxyalkanoates (PHA) accumulation/degradation and interference with the outer membranes of other bacteria. Metaproteomics did not provide sufficient proteome coverage for relatively lower abundant proteins such as lipases though the expression of fadL genes, long-chain fatty acid transporters, was confirmed for four genera (Dechloromonas, Azoarcus, Aeromonas and Sulfurimonas), none of which were recovered as putative lipolytic MAGs. Metaproteomics also confirmed the presence of 15 relatively abundant ([≥]1%) genera in all reactors, of which at least 6 can potentially accumulate lipid/polyhydroxyalkanoates. For most putative lipolytic MAGs, there was no statistically significant correlation between the read abundance and reactor conditions such as temperature, phase (biofilm and bulk liquid), and feed type (treated by ultraviolet light or not). Results obtained by metagenomics and metaproteomics did not confirm each other and further work is required to identify the true lipid degraders in these systems.

molecular biology↗

bioRxiv2021

Genome-wide association identifies candidate genes for drought tolerance in coast redwood and giant sequoia

Drought is a major limitation for survival and growth in plants. With more frequent and severe drought episodes occurring due to climate change, it is imperative to understand the genomic and physiological basis of drought tolerance to be able to predict how species will respond in the future. In this study, univariate and multitrait multivariate GWAS methods were used to identify candidate genes in two iconic and ecosystem-dominating species of the western US - coast redwood and giant sequoia - using ten drought-related physiological and anatomical traits and genome-wide sequence-capture SNPs. Population level phenotypic variation was found in carbon isotope discrimination, osmotic pressure at full turgor, xylem hydraulic diameter and total area of transporting fibers in both species. Our study identified new 78 new marker x trait associations in coast redwood and six in giant sequoia, with genes involved in a range of metabolic, stress and signaling pathways, among other functions. This study contributes to a better understanding of the genomic basis of drought tolerance in long-generation conifers and helps guide current and future conservation efforts in the species. Significance StatementClimate change brings more frequent and severe drought events that challenge the survival of natural populations of plants. While most of our knowledge about drought tolerance comes from annual and domesticated plants, the genomic basis of drought tolerance in long-generation trees is poorly understood. Here, we aim to fill this gap by identifying candidate genes in two conifer species, coast redwood and giant sequoia.

genomics↗

bioRxiv2021

Synchronous seasonality in the gut microbiota of wild wood mouse populations

O_LIThe gut microbiome performs many important functions in mammalian hosts, with community composition shaping its functional role. However, what factors drive individual microbiota variation in wild animals and to what extent these are predictable or idiosyncratic across populations remains poorly understood. C_LIO_LIHere, we use a multi-population dataset from a common rodent species (the wood mouse, Apodemus sylvaticus), to test whether a consistent set of core gut microbes is identifiable in this species, and to what extent the predictors of microbiota variation are consistent across populations. C_LIO_LIBetween 2014 and 2018 we used capture-mark-recapture and 16S rRNA profiling to intensively monitor two wild UK mouse populations and their gut microbiota, as well as characterising the microbiota from a laboratory-housed colony of the same species. C_LIO_LIAlthough broadly similar at high taxonomic levels and despite being only 50km apart, the two wild populations did not share a single bacterial amplicon sequence variant (ASV). Meanwhile, the laboratory-housed colony shared many ASVs with one of the wild populations from which it is thought to have been founded decades ago. Despite strong taxonomic divergence in the microbiota, the factors predicting compositional variation in each wild population were remarkably similar. We identified a strong and consistent pattern of seasonal microbiota restructuring that occurred at both sites, in all years, and within individual mice. While the microbiota was highly individualised, seasonal convergence in the gut microbiota among individuals occurred in late winter/early spring. C_LIO_LIThese findings reveal highly repeatable seasonal gut microbiota dynamics across distinct populations of this species, despite divergent taxa being involved. Providing a platform for future work to understand the drivers and functional implications of such predictable seasonal microbiome restructuring, including whether it might provide the host with adaptive seasonal phenotypic plasticity. C_LI

ecology↗

bioRxiv2020

Social networks strongly predict the gut microbiota of wild mice

The mammalian gut teems with beneficial microbes, yet how hosts acquire these symbionts remains poorly understood. Research in primates suggests that microbes can be picked up via social contact, but the role of social interactions in non-group-living species remains unexplored. Here, we use a passive tracking system to collect high resolution spatiotemporal activity data from wild mice (Apodemus sylvaticus). Social network analysis revealed social association strength to be the strongest predictor of microbiota similarity among individuals, controlling for factors including spatial proximity and kinship, which had far smaller or nonsignificant effects. This social effect was limited to interactions involving males (male-male and male-female), implicating sex-dependent behaviours as driving processes. Social network position also predicted microbiota richness, with well-connected hub individuals having the most diverse microbiotas. Overall, these findings suggest social contact provides a key transmission pathway for gut symbionts even in relatively asocial mammals, that strongly shapes the adult gut microbiota. This work underlines the potential for individuals to pick up beneficial symbionts as well as pathogens from social interactions.

ecology↗

bioRxiv2020

The Hedgehog Co-Receptor BOC Differentially Regulates SHH Signaling During Craniofacial Development

The Hedgehog (HH) pathway controls multiple aspects of craniofacial development. HH ligands signal through the canonical receptor PTCH1, and three co-receptors- GAS1, CDON and BOC. Together, these co-receptors are required during embryogenesis to mediate proper HH signaling. Here we investigated the individual and combined contributions of GAS1, CDON and BOC to HH-dependent mammalian craniofacial development. Individual deletion of either Gas1 or Cdon results in variable holoprosencephaly phenotypes, characterized by the failure to divide and form the telencephalon and midfacial structures. In contrast, we find that Boc deletion results in facial widening consistent with increased HH pathway activity. Additionally, the deletion of Boc in a Gas1 null background partially rescues the craniofacial defects observed in Gas1 single mutants; a phenotype that persists over developmental time. This contrasts with HH-dependent phenotypes in other tissues that significantly worsen following combined deletion of Gas1 and Boc. Mechanistically, BOC selectively restricts neural crest-derived mesenchymal proliferation. Together, these data indicate that BOC acts as a multi-functional regulator of HH signaling during craniofacial development, alternately promoting or restraining HH pathway activity in a tissue-specific fashion. Summary statementHere we identify dual, tissue-specific roles for the Hedgehog co-receptor BOC in both the promotion and antagonism of Hedgehog signaling during craniofacial development.

developmental biology↗

bioRxiv2019

High-quality chromosome-scale assembly of the walnut (Juglans regia L) reference genome

The release of the first reference genome of walnut (Juglans regia L.) enabled many achievements in the characterization of walnut genetic and functional variation. However, it is highly fragmented, preventing the integration of genetic, transcriptomic, and proteomic information to fully elucidate walnut biological processes. Here we report the new chromosome-scale assembly of the walnut reference genome (Chandler v2.0) obtained by combining Oxford Nanopore long-read sequencing with chromosome conformation capture (Hi-C) technology. Relative to the previous reference genome, the new assembly features an 84.4-fold increase in N50 size, and the full sequence of all 16 chromosomal pseudomolecules, nine of which present telomere sequences at both ends. Using full-length transcripts from single-molecule real-time sequencing, we predicted 40,491 gene models, with a mean gene length higher than the previous gene annotations. Most of the new protein-coding genes (90%) are full-length, which represents a significant improvement compared to Chandler v1.0 (only 48%). We then tested the potential impact of the new chromosome-level genome on different areas of walnut research. By studying the proteome changes occurring during catkin development, we observed that the virtual proteome obtained from Chandler v2.0 presents fewer artifacts than the previous reference genome, enabling the identification of a new potential pollen allergen in walnut. Also, the new chromosome-scale genome facilitates in-depth studies of intraspecies genetic diversity by revealing previously undetected autozygous regions in Chandler, likely resulting from inbreeding, and 195 genomic regions highly differentiated between Western and Eastern walnut cultivars. Overall, Chandler v2.0 is a valuable resource to understand and explore walnut biology better.

genomics↗

bioRxiv2018

Depletion of microbiome-derived molecules in the host using Clostridium genetics

The gut microbiota produce hundreds of molecules that are present at high concentrations in circulation and whose levels vary widely among humans. In most cases, molecule production has not been linked to specific bacterial strains or metabolic pathways, and unraveling the contribution of each molecule to host biology remains difficult. A general system to toggle molecules in this pool on/off in the host would enable interrogation of the mechanisms by which they modulate host biology and disease processes. Such a system has been elusive due to limitations in the genetic manipulability of Clostridium and its relatives, the source of many molecules in this pool. Here, we describe a method for reliably constructing clean deletions in a model commensal Clostridium, C. sporogenes (Cs), including multiply mutated strains. We demonstrate the utility of this method by using it to toggle off the production of ten Cs-derived molecules that accumulate in host tissues. By comparing mice colonized by wild-type Cs versus a mutant deficient in the production of branched short-chain fatty acids, we discover a previously unknown IgA-modulatory activity of these abundant microbiome-derived molecules. Our method opens the door to interrogating and sculpting a highly concentrated pool of chemicals from the microbiome.

microbiology↗

bioRxiv2018

Genome-wide chromatin accessibility and transcriptome profiling show minimal epigenome changes and coordinated transcriptional dysregulation of hedgehog signaling in Danforth’s short tail mice

Danforths short tail (Sd) mice provide an excellent model for investigating the underlying etiology of human caudal birth defects, which affect 1 in 10,000 live births. Sd animals exhibit aberrant axial skeleton, urogenital, and gastrointestinal development similar to human caudal malformation syndromes including urorectal septum malformation, caudal regression, VACTERL association, and persistent cloaca. Previous studies have shown that the Sd mutation results from an endogenous retroviral (ERV) insertion upstream of the Ptf1a gene resulting in its ectopic expression at E9.5. Though the genetic lesion has been determined, the resulting epigenomic and transcriptomic changes driving the phenotype have not been investigated. Here, we performed ATAC-seq experiments on isolated E9.5 tailbud tissue, which revealed minimal changes in chromatin accessibility in Sd/Sd mutant embryos. Interestingly, chromatin changes were localized to a small interval adjacent to the Sd ERV insertion overlapping a known Ptf1a enhancer region, which is conserved in mice and humans. Furthermore, mRNA-seq experiments revealed increased transcription of PTF1A target genes and, importantly, downregulation of hedgehog pathway genes. Reduced sonic hedgehog (SHH) signaling was confirmed by in situ hybridization and immunofluorescence suggesting that the Sd phenotype results, in part, from downregulated SHH signaling. Taken together, these data demonstrate substantial transcriptome changes in the Sd mouse, and indicate that the effect of the ERV insertion on Ptf1a expression may be mediated by increased chromatin accessibility at a conserved Ptf1a enhancer. We propose that human caudal dysgenesis disorders may result from dysregulation of hedgehog signaling pathways.

genetics↗

bioRxiv2016

ULTRAFAST LASER-PROBING SPECTROCOPY FOR STUDYING MOLECULAR STRUCTURE OF POLYMERIC PROTEINS

We report the development of a new technique to screen protein crystallinity quantitatively based on laser-probing spectroscopy with sub-picosecond resolution. First, we show theoretically that the temperature dependence of the refractive index of a polymeric protein is correlated to its crystallinity. Then, we performed time-domain thermo-transmission experiments on purified semi-crystalline proteins, both native and recombinant (i.e., silk and squid ring teeth), and also on intact E. coli cells bearing overexpressed recombinant protein. Our results demonstrate, for the first time, quantification of crystallinity in real time for polymeric proteins. Our approach can potentially be used for screening an ultra-large number of polymeric proteins in vivo.

biophysics↗