Biology subjects

Smith, M.

Publications and source records attributed to Smith, M..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

An improved Solanum verrucosum genome provides insight into potato centromeres and epigenetic regulation

Solanum verrucosum is a wild diploid potato species notable for carrying the uncharacterised Rpi-ver1 gene conferring resistance to Phytophthora infestans, possessing unusually structured centromeres, and exhibiting the rare trait of self-compatibility, the genetic basis of which remains unknown. Here, we present a high-quality genome assembly and annotation of S. verrucosum accession CPC54. A comprehensive inventory of resistance genes is provided, and candidate genes for Rpi-ver1 are identified through remapping approaches. The sequence and epigenetic landscape of centromeres are resolved, revealing a mosaic of repeatless and repetitive domains. These centromeres consist of tandem arrays of kilobase-sized repeats, some of which show signatures of transposable element origin. Specific subfamilies of CRM and Tekay elements appear to have adapted to centromeric regions. The basis of self-compatibility in S. verrucosum is investigated with respect to the S-RNase gene, which is found to be present and intact, but expressed at very low levels. Notably, upstream insertions of transposable elements are present that may interfere with its expression.

plant biology↗

bioRxiv2025

A comprehensive characterization of the phospholipid and cholesterol composition of the uncinate fasciculus in the human brain: evidence of age-related alterations

The uncinate fasciculus (UF) is a long-range association fiber tract that serves to connect the anterior temporal lobe with the orbitofrontal cortex. The UF has been implicated via neuroimaging studies in the neurobiological vulnerability to psychiatric disorders posed by a history of childhood abuse (CA), as well as in the psychopathology underlying depressive disorders. Since the myelin sheath is highly enriched in lipids, white matter (WM) dysfunction may reflect alterations in the myelin lipid profile. In fact, our previous work showed that in the anterior cingulate cortex WM, there was a specific effect of CA in the choline glycerophospholipid fatty acids (FA) involved in the synthesis of arachidonic acid. Given that the UF does not exist in rodents, its molecular properties are highly understudied and its lipid composition is virtually unknown. As such, we sought to quantify the phospholipid FA and cholesterol quantities of the human postmortem UF and measure whether we could detect lipid-related or myelin-constituent gene/protein changes associated with CA and/or depression. Fresh-frozen left hemisphere UF samples were analyzed from individuals with depression who died by suicide with a history of severe CA (DS-CA), individuals with depression who died by suicide without a history of CA (DS), and non-psychiatric control subjects who died naturally or accidentally (CTRL). Phospholipids were separated by thin-layer chromatography. FA and non-derivatized cholesterol were quantified using gas chromatography-flame ionization detection. Relative expression of myelin-constituent genes (PLP1, MAG, CNP, MOG, PLLP, MBP, and MOBP) was measured by RT-qPCR, and levels of myelin-constituent proteins (MAG, MOG, MBP, and PLP) were measured by immunoblotting. We found no robust relationships between depression or CA and any lipid measures, nor in myelin-constituent gene and protein levels. However, in the phospholipids, we observed striking age relationships that varied across fractions, with an overall pattern of increases in monounsaturates and decreases in long chain omega-6 polyunsaturates with age. In tandem, we observed that most myelin-constituent genes and proteins showed decreasing trends with age, with PLP1 and MAG showing significantly decreasing relationships. We hypothesize that the changes in lipid composition and lipid-protein interactions contribute to age-related myelin deficits and declines in cognition. The absence of group differences highlights the importance of regional specificity in molecular studies assessing neurobiological correlates of psychiatric disorders.

neuroscience↗

bioRxiv2025

The genomics of convergent adaptation to intertidal gravel beaches in Mediterranean clingfishes

IntroductionUnderstanding the genetic basis of widespread phenotypic convergence, particularly for complex morphological traits, remains a major challenge in evolutionary biology. The Mediterranean gravel beach clingfishes of the genus Gouania provide an excellent system to study this phenomenon. Within this genus, two distinct morphotypes, "slender" and "stout", have repeatedly evolved, adapting to different microhabitats. These morphotypes differ in multiple complex traits, including body elongation, head compression, vertebral number, eye size, and the structure of the adhesive disc. Materials and MethodsIn a first step, to scrutinize phylogenetic convergence, we combined 3D morphometrics of the pelvic girdle and skull, with molecular species delimitation based on >660 DNA barcodes, and a phylogenomic framework based on more than 3,400 single-copy orthologs. Secondly, by employing whole-genome resequencing and a novel "convergence score" statistic, we examined genomic convergence across multiple levels: nucleotides, sequences, genes, and functional pathways. Results and DiscussionWhile we found no evidence of large-scale genomic or protein-level convergence, we identified promising candidate regions at the level of single variants, genes, and biological pathways. Notably, a longer shared (but interrupted) haplotype around the morphogene adam12 was associated with convergent traits. The lack of simple genomic patterns may reflect the radiations age and the complex genetic basis of the underlying morphological traits (e.g., eye-size, neurocranium shape). Altogether, our findings highlight the importance of assessing genomic convergence at multiple molecular levels to uncover diagnostic signals across varying evolutionary processes and timescales. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=149 SRC="FIGDIR/small/675783v1_ufig1.gif" ALT="Figure 1"> View larger version (61K): org.highwire.dtl.DTLVardef@58265eorg.highwire.dtl.DTLVardef@13f1deaorg.highwire.dtl.DTLVardef@c51d79org.highwire.dtl.DTLVardef@112217d_HPS_FORMAT_FIGEXP M_FIG C_FIG

evolutionary biology↗

bioRxiv2025

Phased Potato Genome Assembly and Association Genetics Enable Characterisation of the Elusive H1 Resistance Locus Against Potato Cyst Nematodes

The complexity of potato genetics, characterised by tetrasomic inheritance, has contributed to slower genetic gain in potato compared to other major crops. Disease resistance genes, often found in large clusters of highly similar paralogs and alleles, further complicate genetic studies. The H1 resistance locus, introgressed into potato cultivars from Solanum tuberosum spp. andigena, has been successfully used for over 60 years to control Globodera rostochiensis in Europe. Although previous genetic studies mapped this resistance to chromosome 5, the complete structure of the locus remained elusive. To reduce genomic complexity, we generated a dihaploid of the cultivar Athlete, DH4_Athlete, carrying the H1 resistance locus, and produced a phased haplotype representation of the H1 interval using Oxford Nanopore sequencing. Combined with RenSeq-based association genetics, this approach allowed us to reconstruct the entire H1 locus, including recombination points at both the 5' and 3' ends of the interval.

plant biology↗

bioRxiv2025

Geographic distribution of genetic diversity of Heterocephalus glaber analyzed using whole genome sequencing and a chromosome-scale genome assembly

Naked mole-rats (Heterocephalus glaber) are a species of rodent endemic to the Horn of Africa, notable among mammals for their long lifespans, resistances to a variety of stresses, and eusocial mating behavior. Though their natural range extends across large portions of Kenya, Ethiopia, Somalia, and Djibouti, the large majority of genetic and genomic analyses focus on Kenyan specimens. Here, we constructed a chromosome-scale reference genome assembly for H.glaber, along with new reference assemblies of both the Damaraland mole-rat (Fukomys damarensis) and guinea pig (Cavia porcellus) genomes to aid annotation. We leveraged our H.glaber assembly, along with modern whole-genome sequencing, to characterize the genetic diversity of specimens deriving from Kenya, southern Ethiopia, and eastern Ethiopia. We found the Kenyan and southern Ethiopian specimens to be closely related to each other and highly diverged from eastern Ethiopian specimens. We also found specimens collected from nearby locations in southern Ethiopia to be more closely related to Kenyan specimens than to each other. This unexpected distribution of shared genetic diversity highlights the importance of local migration barriers to gene flow in wild H.glaber populations.

ecology↗

bioRxiv2025

A Portable RPA-CRISPR/Cas12a-Based Biosensing Platform for On-Site Detection of the Microcystin Synthetase E Gene in Lake Water

Microcystins are potent cyanotoxins produced by toxigenic cyanobacteria during harmful algal blooms (HABs), posing risks to ecosystems and human health. In this study, we developed a portable RPA-CRISPR/Cas12a biosensing platform for the rapid, on-site detection of the microcystin synthetase E (mcyE) gene, a key biomarker for microcystin-producing strains. The developed RPA-CRISPR/Cas12a assays enable detection of the mcyE gene within 50 min, with either fluorescence or lateral flow assay readouts. The fluorescence readouts have an analytical detection limit of 48.4 copies/{micro}L and a dynamic range of 1.2 x 102 to 1.2 x 107 copies/{micro}L. To enable field deployment, a magnetic bead-based DNA extraction method was integrated, achieving extraction within 1 hour without centrifugation. The complete workflow demonstrated a method LOD of 8.4 x 102 cells/mL in spiked lake water. Applicability was validated using non-spiked environmental water samples collected from multiple HAB-affected lakes. Importantly, a systematic matrix effect assessment was conducted for the CRISPR sensing step, evaluating environmental variables such as pH, ions, nutrients, and natural organic matter. This study establishes a practical, sensitive, and selective detection tool for proactive HAB monitoring. The platforms simplicity, portability, and completeness, from sample pretreatment to signal readout, highlight its potential for real-world environmental biosensing applications.

microbiology↗

bioRxiv2025

Morning Elevation in Insulin Enhances Afternoon Postprandial Insulin Action and Glucose Effectiveness

The second-meal phenomenon refers to the improved glycemic response to a subsequent identical meal. We previously showed that morning (AM) hyperinsulinemia is a key mediator, priming the liver for enhanced net hepatic glucose uptake (NHGU) and glycogen storage during an afternoon (PM) hyperinsulinemic-hyperglycemic clamp. Postprandial NHGU is regulated by three primary mechanisms: insulin action (IA), initiated by hyperinsulinemia; glucose effectiveness (GE), driven by hyperglycemia; and the portal glucose signal (PGS), a neurally-mediated signal activated by glucose delivery into the hepatoportal circulation. It remained unclear, however, which of these mechanisms govern the increase in PM NHGU following AM insulin exposure. To address this, dogs underwent an AM clamp with either a 4-hour hyperinsulinemic prime (Prime, n=8) or basal insulin delivery (No Prime, n=8). After a 1.5-hour rest, both groups underwent a PM hyperglycemic clamp with portal glucose delivery under basal insulin conditions to isolate the effects of an AM insulin prime on PM glucose-mediated hepatic signals (GE/the PGS). Mean PM NHGU was significantly greater in the Prime group (2.2 {+/-} 0.3 mg/kg/min) compared to the No Prime group (0.1 {+/-} 0.3 mg/kg/min, p=0.005), accompanied by augmented net glycolytic and glycogen flux. These findings indicate that morning insulin can enhance glucose-mediated PM NHGU independently of a rise in PM insulin. However, maximal second-meal NHGU also requires elevated PM insulin. Together, this suggests that strategically timed early-day insulin or insulinotropic interventions could potentially improve hepatic responsiveness in settings of impaired postprandial glycemic control, such as insulin resistance or diabetes. Article HighlightsO_LIElevated morning insulin primes the liver for enhanced afternoon net hepatic glucose uptake (NHGU), but it was unclear whether augmentation of insulin action (IA), glucose effectiveness (GE), or the portal glucose signal (PGS) mediates this effect. C_LIO_LIDogs underwent a morning euglycemic clamp with either elevated or basal insulin delivery, followed by an afternoon euinsulinemic-hyperglycemic clamp to isolate the effect of morning insulin priming on afternoon GE/PGS. C_LIO_LIMorning insulin priming enhanced afternoon NHGU via increased glucose-mediated mechanisms, though maximal afternoon NHGU also requires elevated afternoon insulin. C_LIO_LIThese findings identify mechanisms underlying insulin-induced hepatic metabolic memory, providing a framework to inform strategies improving postprandial glucose handling in diabetes. C_LI

physiology↗

bioRxiv2025

Decoding PDI diversity: insights into structure, domains, and functionality in sorghum

Proteins play indispensable roles in cellular function, acting as both structural components and catalysts for essential biological processes. Their proper folding into three-dimensional structures is critical for functionality. To ensure correct folding, proteins interact with chaperones and folding catalysts such as Protein Disulfide Isomerases (PDIs), which assist in the formation and rearrangement of disulfide bonds that stabilize proteins by linking cysteine residues. PDIs are part of the thioredoxin (TRX) superfamily and are characterized by a conserved CXXC motif that contributes to their redox potential. They exhibit isomerase and oxidoreductase activities, that enable them to rearrange and form new disulfide bonds. PDI family members in sorghum (SbPDI) present a broad and largely unexplored diversity in domain order, structure, and architecture between or even within species. To shed light on this diversity, we identified and characterized PDI family members in sorghum in silico to explore their domain architecture, three-dimensional structure and functionality. Author summaryIn this work, we explore how genomic and molecular tools can improve our understanding of the function and diversity of the protein disulfide isomerase (PDI) family in plants, using sorghum as a model and humans as a reference. By analysing domain architecture and predicted structures across the PDI family, we lay the groundwork for future studies investigating their roles in plant development and stress responses, including through targeted gene editing approaches. Although PDIs have been widely studied in humans, their structural and functional diversity in plants remains largely unexplored. With this study, I aim to help close this knowledge gap and highlight the structural differences of PDIs in plants.

bioinformatics↗

bioRxiv2025

ezscore-f: A Set of Freely Available, Validated Sleep Stage Classifiers for Forehead EEG

The increasing availability of wearable forehead EEG devices, such as the Hypnodyne ZMax, open-source DCM, CGX PatchEEG, and many others, has significantly expanded opportunities for convenient, at-home sleep monitoring. However, most publicly available sleep classifiers are trained on scalp EEG from traditional polysomnography (PSG) data, and thus generalize poorly to forehead EEG due to differences in electrode placement, referencing montages, and higher susceptibility to artifacts from user movements and electrode displacement. Conventional classifiers typically do not explicitly account for these artifacts, resulting in inaccurate and misleading sleep stage scoring. To address this gap, we developed a suite of artifact-aware sleep stage classifiers trained specifically using forehead EEG data, leveraging two comprehensive datasets--Wearanize+ and Donders2022--that contain concurrent forehead EEG and clinical PSG recordings. We further introduce two classifier variants: one optimized for real-time applications that operates directly on raw EEG amplitudes, and another optimized for offline analysis utilizing normalized EEG signals. Validation results indicate robust and reliable classification performance across standard sleep stages (Wake, N1, N2, N3, REM), along with effective identification of artifact epochs. Importantly, the developed classifiers generalize well to forehead EEG devices beyond the original training platform. These validated classifiers are freely available to the sleep research community through the open-source ezscore-f package, providing versatile and practical tools for forehead EEG-based sleep stage analysis.

neuroscience↗

bioRxiv2025

eDNA sampling reveals no negative effects of beaver recolonisation on the catchment-scale distribution of migratory fish

O_LIReintroduction of keystone species is considered part of the solution to the current biodiversity crisis. The Eurasian beaver (Castor fiber) is one such species, shaping its habitat by felling trees, building dams and creating wetlands. However, while the potential benefits to aquatic biodiversity and ecological functioning have been studied on a local scale, the impacts of beavers on catchment-scale processes such as fish migration remain understudied. C_LIO_LISequencing of environmental DNA (eDNA metabarcoding) from water samples is a cost-effective method to study species distributions across large geographical scales. Here, eDNA samples (n=426) were collected from 142 sites across the UKs oldest and largest established wild beaver population, located on Tayside, East Scotland, and analysed using a vertebrate specific metabarcoding assay. We combined presence/absence data from eDNA results with other environmental and anthropogenic variables to model the effects of beaver presence on the distribution of three migratory fish species. C_LIO_LIUsing generalised linear models, we found no effects of the current beaver presence on the distribution of Atlantic salmon or lamprey, but a positive co-occurrence with European eel at the catchment scale. Model outputs also reinforced previous findings on the impact of barriers to migration and other abiotic and biotic factors on fish species, demonstrating the effectiveness of eDNA sampling in rivers for understanding species distributions at a catchment scale. C_LIO_LIThis case study provides novel insights into the co-distribution of beavers and migratory fish, and highlights how catchment-wide eDNA monitoring can be applied by environmental managers to aid decision-making and impact assessment more generally. C_LI

ecology↗

bioRxiv2025

Profiling metabotropic glutamate receptor 7 expression in Rett syndrome: consequences for pharmacotherapy

We have reported that levels of metabotropic glutamate receptor 7 (mGlu7) are dramatically decreased in brain samples from Rett syndrome patients carrying truncation mutations in the Methyl-CpG Binding Protein 2 (MECP2) gene. Additionally, we identified decreases in mGlu7 levels in Mecp2+/- female mice and demonstrated that administration of a positive allosteric modulator (PAM) with activity at mGlu7 corrected deficits in cognitive, social, and respiratory domains. Here, we expanded our studies to a larger cohort of RTT samples covering a range of mutations and evaluated expression of the three widely expressed group III mGlu receptors (mGlu4,7 and 8). We found significant decreases in mGlu7, but not mGlu4 or mGlu8, mRNA expression across this larger cohort; additionally, we identified a previously unknown and robust correlation in the expression of mGlu4 and mGlu8 in control individuals. Stratification of RTT patients into individuals with mutations that are clinically correlated with severe versus mild disease revealed statistically significant decreases in mGlu7 expression only in patients with mutations that induce more severe symptoms. We then administered the PAM VU0422288 to mice modeling the mild R306C mutation (Mecp2R306C/+) and found a significant reduction in apneas induced by VU0422888 administration despite no decreases in mGlu7 expression in the brainstem or cortex. These results provide the first evidence of potentially broad utility for mGlu7 PAMs in reducing apneas in RTT patients.

neuroscience↗

bioRxiv2025

ERC 2.0 - evolutionary rate covariation update provides more powerful inference of functional interactions across large phylogenies

Evolutionary Rate Covariation (ERC) is an established comparative genomics method that identifies sets of genes sharing patterns of sequence evolution, which suggests shared function. Whereas many functional predictions of ERC have been empirically validated, its predictive power has hitherto been limited by its inability to tackle the large numbers of species in contemporary comparative genomics datasets. This study introduces ERC2.0, an enhanced methodology for studying ERC across phylogenies with hundreds of species and tens of thousands of genes. ERC2.0 improves upon previous iterations of ERC in algorithm speed, normalizing for heteroskedasticity, and normalizing correlations via Fisher transformations. These improvements have resulted in greater statistical power to predict biological function. In exemplar yeast and mammalian datasets, we demonstrate that the predictive power of ERC2.0 is improved relative to the previous method, ERC1.0, and that further improvements are obtained by using larger yeast and mammalian phylogenies. We attribute the improvements to both the larger datasets and improved rate normalization. We demonstrate that ERC2.0 has high predictive accuracy for known annotations and can predict the functions of genes in non-model systems. Our findings underscore the potential for ERC2.0 to be used as a single-pass computational tool in candidate gene screening and functional predictions.

evolutionary biology↗

bioRxiv2025

Hepatic Metabolic Memory Triggered by AM Exposure to Glucagon Alters Afternoon Glucose Metabolism

The Staub-Traugott effect, or second-meal phenomenon, describes improved glucose disposal after a second identical meal. We previously showed that morning hyperinsulinemia primes the liver to enhance afternoon net hepatic glucose uptake and glycogen storage. However, mixed meals trigger co-secretion of insulin and glucagon, and glucagon is traditionally viewed as opposing insulins hepatic actions. Whether glucagon modifies the persistence of insulins priming effects across sequential metabolic challenges is unknown. Therefore, we investigated whether morning hyperglucagonemia alters the ability of morning hyperinsulinemia to prime subsequent hepatic glucose metabolism. Conscious dogs underwent two pancreatic clamp periods separated by a 1.5h rest period. Endogenous insulin and glucagon were suppressed with somatostatin and replaced intraportally at defined rates. During a 4h morning hyperinsulinemic-euglycemic clamp, dogs received matched insulin prime infusions with either basal glucagon (AM INS; n=8) or elevated glucagon (AM INS+GCG; n=8). After the rest period, both groups underwent a 2.5h afternoon hyperinsulinemic-hyperglycemic clamp under identical hormonal conditions. Afternoon net hepatic glucose uptake, glycogen, glycolytic, and gluconeogenic flux rates were quantified using arteriovenous difference methods and [3-3H]-glucose tracer kinetics. Liver biopsies were collected before and after the afternoon clamp to assess gene transcription and protein regulators of hepatic glucose metabolism. During the afternoon clamp, despite matched insulin, glucagon, and glucose levels, net hepatic glucose uptake was 41% lower in AM INS+GCG (3.6{+/-}0.4 mg/kg/min) than in AM INS (6.1{+/-}0.6 mg/kg/min; p<0.003). This was accompanied by a trend toward incomplete suppression of hepatic glucose production in AM INS+GCG (1.4{+/-}0.4 mg/kg/min), whereas it was fully suppressed in the AM INS group (p=0.06). Direct glycogen synthesis was also 44% lower in AM INS+GCG (1.8{+/-}0.2 vs 3.2{+/-}0.7 mg/kg/min; p<0.015), along with reductions in net glycogen synthesis and glycolytic flux. Morning insulin with basal glucagon increased hepatic glucokinase mRNA and protein before the afternoon clamp, whereas concurrent glucagon prevented this induction. In summary, antecedent morning hyperglucagonemia attenuates insulin-mediated hepatic priming, reducing hepatic glucose flux during a later hyperinsulinemic-hyperglycemic challenge. These findings identify glucagon as a regulator of hepatic metabolic memory alongside insulin and demonstrate that early-day insulin-glucagon dynamics shape the livers response to subsequent challenges, providing mechanistic insight into postprandial glucose regulation and implications for metabolic health and diabetes risk. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=121 SRC="FIGDIR/small/639957v3_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@6d210org.highwire.dtl.DTLVardef@1c93840org.highwire.dtl.DTLVardef@18c0b49org.highwire.dtl.DTLVardef@138ee9f_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2025

A Broad Survey and Functional Analysis of Immunoglobulin Loci Variation in Rhesus Macaques

Rhesus macaques (RMs) are a vital model for studying human disease and invaluable to pre-clinical vaccine research, particularly for the study of broadly neutralizing antibody responses. Such studies require robust genetic resources for antibody-encoding genes within the immunoglobulin (IG) loci. The complexity of the IG loci has historically made them challenging to characterize accurately. To address this, we developed novel experimental and computational methodologies to generate the largest collection to date of integrated antibody repertoire and long-read genomic sequencing data in 106 Indian origin RMs. We created a comprehensive resource of IG heavy and light chain variable (V), diversity (D), and joining (J) alleles, as well as leader, intronic, and recombination signal sequences (RSSs), including the curation of 1474 novel alleles, unveiling tremendous diversity, and expanding existing IG allele sets by 60%. This publicly available, continually updated resource (https://vdjbase.org/reference_book/Rhesus_Macaque) provides the foundation for advancing RM immunogenomics, vaccine discovery, and translational research.

immunology↗

bioRxiv2025

Susceptibility of bovine respiratory and mammary epithelial cells to avian and mammalian derived clade 2.3.4.4b H5N1 highly pathogenic avian influenza viruses

Zoonotic transmission of avian influenza viruses into mammals is relatively rare due to anatomical differences in the respiratory tract between species. Recently, clade 2.3.4.4b highly pathogenic H5N1 avian influenza viruses were detected circulating in North American cattle. Sporadic transmission between cattle, humans, and other animals proximal to cattle or after consuming products from infected cattle has occurred, but thus far there is no evidence of human-to-human transmission. However, the virus has the potential to adapt to the mammalian respiratory tract with every transmission event that occurs, making it crucial to understand cellular and species tropism of the H5N1 2.3.4.4b viruses. We compared viral kinetics of clade 2.3.4.4b viruses isolated from birds and mammals in respiratory epithelial cells derived from cattle, human, swine, and ferret. We found that avian derived viruses could replicate in swine cells only, yet mammalian derived strains could replicate efficiently in all tracheal and nasal epithelial cells tested. Interestingly, only bovine mammary epithelial cells (MEC) and swine respiratory epithelial cells were permissive to both avian and mammalian derived strains, possibly due to increased sialic acid expression on bovine MEC compared to bovine tracheal epithelial cells (TEC). However, sialic acid expression differed between dairy and beef cows: TEC derived from a dairy cow had increased expression of 2,3 sialic acid receptors compared to TEC from a beef-dairy cow cross. This study highlights the ability of clade 2.3.4.4b H5N1 viruses derived from mammals but not wild birds to infect the respiratory epithelium of other mammalian hosts.

microbiology↗

bioRxiv2024

Extended Quality (eQual): Radial threshold clustering based on n-ary similarity

We are transforming Radial Threshold Clustering (RTC), an O(N 2) algorithm, into Extended Quality Clustering, an O(N) algorithm with several novel features. Daura et als RTC algorithm is a partitioning clustering algorithm that groups similar frames together based on their similarity to the seed configuration. Two current issues with RTC is that it scales as O(N 2) making it inefficient at high frame counts, and the clustering results are dependent on the order of the input frames. To address the first issue, we have increased the speed of the seed selection by using k-means++ to select the seeds of the available frames. To address the second issue and make the results invariant with respect to frame ordering, whenever there is a tie in the most populated cluster, the densest and most compact cluster is chosen using the extended similarity indices. The new algorithm is able to cluster in linear time and produce more compact and separate clusters.

biophysics↗

bioRxiv2024

Large scale capsid-mediated mobilisation of bacterial genomic DNA in the gut microbiome

Transducing bacteriophages and gene transfer agents (GTA) are constrained by their capsids structural properties in the length of host DNA they can package. Nanopore sequencing of intact capsid-packaged DNA molecules with full-sized reads can be used to establish the precise lengths and identity of individual packaged DNA molecules and their association with specific bacterial hosts. This approach was validated using a few well-characterised transducing systems, and then applied to study bacterial DNA encapsidation in the faecal microbiomes from three healthy human donors. Bacterial DNA encapsidation appears to be widespread in the microbiome with up to 5.4% of capsid-packaged DNA in the gut virome being of bacterial (non-prophage) origin. Generalised transduction and GTA activity are especially prevalent in the families Oscillospiraceae and Ruminococcaceae, whereas an example of lateral transduction was observed in genus Bacteroides. In addition to that, induction of prophages in a variety of highly prevalent gut bacteria was observed.

microbiology↗

bioRxiv2024

RNA methylation controls stress granule formation and function during erythropoiesis

Stress granules (SGs) are crucial in RNA regulation, affecting cell fate and function. SGs contain RNAs, some of which can be methylated. We studied m6A RNA modifications during the human CD34+ HSPCs (hCD34+) differentiating into erythroid cells and found that mRNAs encoding many erythroid-specific proteins had decreased methylation during differentiation. Increased levels ALKBH5 demethylase during erythropoiesis controls the levels of the 3UTR methylation of these mRNAs. hCD34+ carrying ALKBH5 mutations demonstrated a block in erythropoiesis, and mass-spectrometry studies of the mutant cells showed decreased levels of SG proteins, including the core granule protein ATXN2. ALKBH5 directly regulates the methylation of the mRNA of ATXN2. ATXN2 overexpression accelerated the erythroid differentiation of HSPCs and rescued the erythroid differentiation of ALKBH5 mutant cells. Very few SGs are found in normal human erythroid progenitors. SGs accumulated substantially in ALKBH5 mutant cells, and surprisingly overexpression of ATNX2 reduced SG numbers to normal. Polysome analysis demonstrated m6A-modified RNAs to be enriched in the pre-polysome fractions that were less translated. This work establishes a mechanism by which during stress, ATXN2 facilitates the release of SG-stored m6A-modified RNAs including erythroid-specific and SG-enriched RNAs that are loaded onto functional ribosomes, allowing better translation and accelerated erythroid differentiation during stress.

cell biology↗