Biology subjects

Watson, M.

Publications and source records attributed to Watson, M..

At least 37 records · Page 2Linked to original sources

bioRxiv2022

Inferring a cell's capabilities from omics data with ImmCellFie

ImmCellFie is a user-friendly, web-based platform for comprehensive analysis of metabolic functions inferred from transcriptomic or proteomic data. It enables researchers to leverage the powerful mechanistic insight provided by complex genome-scale metabolic models with little to no bioinformatics training required. The platform has been integrated with a series of useful tools and richly annotated scientific visualizations for interactive exploration by the user. ImmCellFie pushes beyond simple statistical enrichment and incorporates complex biological mechanisms to quantify cell activity. Graphical abstract

bioinformatics↗

bioRxiv2022

Genome-Wide Epigenomic Profiling of Primary Non-Small Cell Lung Cancer Reveals Specific and Recurrent DNA Methylation Alterations in Smoker Versus Never-Smoker Patients

Epigenetic alterations are widespread in cancer and can complement genetic alterations to influence cancer progression and treatment. To better understand the potential contribution of DNA methylation alterations to tumor phenotype in non-small cell lung cancer (NSCLC) in both smoker and never-smoker patients, we performed a comprehensive, genome-wide profiling of DNA methylation in 17 primary non-small cell lung cancer and 10 matched normal lung samples using the complementary methylation assays MeDIP-seq and MRE-seq. Compared to patient-matched non-malignant lung tissue, we report recurrent methylation changes of several gene promoters, many previously implicated in cancer, including FAM83A and SEPT9 (hypomethylation), and PCDH7, NKX2-1, and SOX17 (hypermethylation). Although smoker and never-smoker patients shared many methylation changes, several were specific and recurrent within a particular smoking status. In particular, never-smokers displayed a greater proportion of hypoDMRs and exhibited a greater number of recurrently hypomethylated promoters, including the promoter of the oncogene ASPSCR1, and others previously linked to cancer, including TOP2A, DPP9, and USP39. Methylation changes outside of promoters were also widespread and often recurrent, particularly the loss of methylation over repetitive elements, highly enriched for ERV1 subfamilies. Recurrent hypoDMRs were also enriched for several transcription factor (TF) binding motifs, often for genes involved in signaling and cell proliferation, including 71% encoding a binding site of NKX2-1, which was found to be significantly upregulated in TCGA LUAD samples. Furthermore, the overwhelming majority of DMRs identified in this study were found to reside in an active chromatin state in at least one tissue profiled using the Roadmap Epigenome data, suggesting that methylation changes may contribute to altered regulatory programs through the adaptation of cell type-specific expression programs.

cancer biology↗

bioRxiv2022

Investigating the impact of database choice on the accuracy of metagenomic read classification for the rumen microbiome

Microbiome analysis is quickly moving towards high-throughput methods such as metagenomic sequencing. Accurate taxonomic classification of metagenomic data relies on reference sequence databases, and their associated taxonomy. However, for understudied environments such as the rumen microbiome many sequences will be derived from novel or uncultured microbes that are not present in reference databases. As a result, taxonomic classification of metagenomic data from understudied environments may be inaccurate. To assess the accuracy of taxonomic read classification, this study classified metagenomic data that had been simulated from cultured rumen microbial genomes from the Hungate collection. To assess the impact of reference databases on the accuracy of taxonomic classification, the data was classified with Kraken 2 using several reference databases. We found that the choice and composition of reference database significantly impacted on taxonomic classification results, and accuracy. In particular, NCBI RefSeq proved to be a poor choice of database. Our results indicate that inaccurate read classification is likely to be a significant problem, affecting all studies that use insufficient reference databases. We observe that adding cultured reference genomes from the rumen to the reference database greatly improves classification rate and accuracy. We also demonstrate that metagenome-assembled genomes (MAGs) have the potential to further enhance classification accuracy by representing uncultivated microbes, sequences of which would otherwise be unclassified or incorrectly classified. However, classification accuracy was strongly dependent on the taxonomic labels assigned to these MAGs. We therefore highlight the importance of accurate reference taxonomic information and suggest that, with formal taxonomic lineages, MAGs have the potential to improve classification rate and accuracy, particularly in environments such as the rumen that are understudied or contain many novel genomes.

genomics↗

bioRxiv2022

mRNA 5' terminal sequences drive 200-fold differences in expression through effects on synthesis, translation and decay

mRNA regulatory sequences control gene expression at multiple levels including translation initiation and mRNA decay. The 5' terminal sequences of mRNAs have unique regulatory potential because of their proximity to key post-transcriptional regulators. Here we have systematically probed the function of 5' terminal sequences in gene expression in human cells. Using a library of reporter mRNAs initiating with all possible 7-mer sequences at their 5' ends, we find an unexpected impact on transcription that underlies 200-fold differences in mRNA expression. Library sequences that promote high levels of transcription mirrored those found in native mRNAs and define two basic classes with similarities to classic Initiator (Inr) and TCT core promoter motifs. By comparing transcription, translation and decay rates, we identify sequences that are optimized for both efficient transcription and growth-regulated translation and stability, including variants of terminal oligopyrimidine (TOP) motifs. We further show that 5' sequences of endogenous mRNAs are enriched for multi-functional TCT/TOP hybrid sequences. Together, our results reveal how 5' sequences define two general classes of mRNAs with distinct growth-responsive profiles of expression across synthesis, translation and decay.

genetics↗

bioRxiv2021

Jaagsiekte sheep retrovirus infection induces changes in microRNA expression in the ovine lung

Ovine pulmonary adenocarcinoma (OPA) is an infectious neoplastic lung disease of sheep caused by jaagsiekte sheep retrovirus. OPA is an important veterinary problem and is also a valuable large animal model for human lung adenocarcinoma. JSRV infects type 2 alveolar epithelial cells in the lung and induces the growth of tumors, but little is known about the molecular events that lead to the activation of oncogenic pathways in infected cells. MicroRNAs (miRNAs) are small RNA molecules of approximately 22 nucleotides with important roles in regulating gene expression in eukaryotes and with well-established roles in cancer. Here we used small-RNA sequencing to investigate the changes in miRNA expression that occur in JSRV-infected ovine lung. After filtering out low abundance miRNAs, we identified expression of 405 miRNAs, 32 of which were differentially expressed in JSRV-infected lung compared to mock-inoculated control lung. Highly upregulated miRNAs included miR-182, miR-183, miR-96 and miR-135b, which have also been associated with oncogenic changes in human lung cancer. Network analysis of genes potentially targeted by the deregulated miRNAs identified their involvement in pathways known to be dysregulated in OPA. We found no evidence to support the existence of miRNAs encoded by JSRV. This study provides the first information on miRNA expression in OPA and identifies a number of targets for future studies into the role of these molecules in the pathogenesis of this unique veterinary model for human lung adenocarcinoma. IMPORTANCEOvine pulmonary adenocarcinoma is a neoplastic lung disease of sheep caused by jaagsiekte sheep retrovirus (JSRV). OPA is a significant welfare and economic concern for sheep producers and is a valuable large animal model for human lung adenocarcinoma. MicroRNAs are small RNA molecules of approximately 22 nucleotides with important functions in regulating gene expression in eukaryotes and with well-established roles in cancer. In this study, we examined the changes in microRNA expression that occur in the lung in response to JSRV infection. We identified differential expression of a number of host-encoded microRNAs in infected tissue, including microRNAs with roles in human cancer. We found no evidence that JSRV encodes a microRNA. This study provides new insights on the cellular response to JSRV infection in the ovine lung, which will inform future studies into the pathogenesis of OPA in sheep and its use as a model for human lung adenocarcinoma.

microbiology↗

bioRxiv2021

Dug: A Semantic Search Engine Leveraging Peer-Reviewed Literature to Span Biomedical Data Repositories

MotivationAs the number of public data resources continues to proliferate, identifying relevant datasets across heterogenous repositories is becoming critical to answering scientific questions. To help researchers navigate this data landscape, we developed Dug: a semantic search tool for biomedical datasets utilizing evidence-based relationships from curated knowledge graphs to find relevant datasets and explain why those results are returned. ResultsDeveloped through the National Heart, Lung, and Blood Institutes (NHLBI) BioData Catalyst ecosystem, Dug has indexed more than 15,911 study variables from public datasets. On a manually curated search dataset, Dugs total recall (total relevant results/total results) of 0.79 outperformed default Elasticsearchs total recall of 0.76. When using synonyms or related concepts as search queries, Dug (0.36) far outperformed Elasticsearch (0.14) in terms of total recall with no significant loss in the precision of its top results. Availability and ImplementationDug is freely available at https://github.com/helxplatform/dug. An example Dug deployment is also available for use at https://search.biodatacatalyst.renci.org/. Contactawaldrop@rti.org or scox@renci.org

bioinformatics↗

bioRxiv2021

Anchored and labile pools of Calcineurin are enabled by a disordered AKAP scaffold

Disordered scaffold proteins provide multivalent landing pads that, via a series of embedded Short Linear Motifs (SLiMs), bring together the components of a complex to orchestrate precise spatial and temporal regulation of cellular processes. One such protein is AKAP5, which contains SLiMs that anchor PKA and Calcineurin, and recruit substrate (the TRPV1 receptor). Calcineurin is anchored to AKAP5 by a well-characterised PxIxIT SLiM. Here we show, using a combination of biochemical and biophysical approaches, that the Calcineurin PxIxIT-binding groove also recognises several hitherto unknown lower-affinity SLiMs in addition to the PxIxIT motif. We demonstrate that the assembly is in reality a complex system with conserved SLiMs spanning a wide affinity range. The capture is analogous to that seen for many DNA-binding proteins that have a weak non-specific affinity for DNA outside the canonical binding site, but different in that it involves (i) two proteins, and (ii) hydrophobic rather than electrostatic interactions. It is also compatible with the requirement for both stable anchoring of the enzyme and responsive downstream signalling. We conclude that the AKAP5 C-terminus is enriched in lower-affinity/mini-SLiMs that, together with the canonical SLiM, maintain a structurally disordered but tightly regulated signalosome.

biophysics↗

bioRxiv2021

New insights from 33,813 publicly available metagenome-assembled-genomes (MAGs) assembled from the rumen microbiome

Recent rumen microbiome metagenomics papers1,2 have published hundreds of metagenome-assembled genomes (MAGs), comparing them to 4,941 MAGs published by Stewart et al3 in order to define novelty. However, there are many more publicly available MAGs from ruminants. In this paper, for the first time, all available resources are combined, catalogued and de-replicated to define putative species-level bins. As well as providing new insights into the constitution of the rumen microbiome, including an updated estimate of the number of microbial species in the rumen, this work demonstrates that a lack of community-adopted standards for the release and annotation of MAGs hinders progress in microbial ecology and metagenomics.

microbiology↗

bioRxiv2021

A Kiosk Station for the Assessment of Multiple Cognitive Domains and Enrichment of Monkeys

BackgroundNonhuman primates (NHPs) are self-motivated to perform cognitive tasks on touchscreens in their animal housing setting. To leverage this ability, fully integrated hardware and software solutions are needed, that work within housing and husbandry routines while also spanning cognitive task constructs of the Research Domain Criteria (RDoC). New MethodWe describe a Kiosk Station (KS-1) that provides robust hardware and software solutions for running cognitive tasks in cage-housed NHPs. KS-1 consists of a frame for mounting flexibly on housing cages, a touchscreen animal interface with mounts for receptables, reward pumps and cameras, and a compact computer cabinet with an interface for controlling behavior. Behavioral control is achieved with a unity3D program that is virtual-reality capable, allowing semi-naturalistic visual tasks to assess multiple cognitive domains. ResultsKS-1 is fully integrated into the regular housing routines of monkeys. A single person can operate multiple KS-1s. Monkeys engage with KS-1 at high motivation and cognitive performance levels at high intra-individual consistency. Comparison with Existing MethodsKS-1 is optimized for flexible mounting onto standard apartment cage systems. KS-1 has a robust animal interface with options for gaze/reach monitoring. It has an integrated user interface for controlling multiple cognitive task using a common naturalistic object space designed to enhance task engagement. All custom KS-1 components are open-sourced. ConclusionsKS-1 is a versatile tool for cognitive profiling and enrichment of cage-housed monkeys. It reliably measures multiple cognitive domains which promises to advance our understanding of animal cognition, inter-individual differences and underlying neurobiology in refined, ethologically meaningful behavioral foraging contexts.

animal behavior and cognition↗

bioRxiv2020

Metagenomic analysis of the cow, sheep, reindeer and red deer rumen

The rumen microbiota comprises a community of microorganisms which specialise in the degradation of complex carbohydrates from plant-based feed. These microbes play a highly important role in ruminant nutrition and could also act as sources of industrially useful enzymes. In this study, we performed a metagenomic analysis of samples taken from the ruminal contents of cattle (Bos Taurus), sheep (Ovis aries), reindeer (Rangifer tarandus) and red deer (Cervus elaphus). We constructed 391 metagenome-assembled genomes originating from 16 microbial phyla. We compared our genomes to other publically available microbial genomes and found that they contained 279 novel species. We also found significant differences between the microbiota of different ruminant species in terms of the abundance of microbial taxonomies, carbohydrate-active enzyme genes and KEGG orthologs. However, we found that the vast majority of carbohydrate-active enzymes were present in all of our sample types, which may indicate that there is a core set of these enzymes which are present across ruminants and are independent of diet and environmental conditions. We present a dataset of rumen-derived genomes which in combination with other publicly-available rumen genomes can be used as a reference dataset in future metagenomic studies. Data SummaryThe paired-read fastq files supporting the conclusions of this article are available in the European Nucleotide Archive repository (https://www.ebi.ac.uk/ena/browser/view/PRJEB34458). The RUG fasta files supporting the conclusions of this article are available in the Edinburgh DataShare repository (https://doi.org/10.7488/ds/2640).

microbiology↗

bioRxiv2019

Single-cell RNA-seq reveals CD16- monocytes as key regulators of human monocyte transcriptional response to Toxoplasma

Monocytes are among the major myeloid cells that respond to Toxoplasma, a ubiquitous foodborne that infects [≥]1 billion people worldwide, in human peripheral blood. As such, a molecular understanding of human monocyte-Toxoplasma interactions can expedite the development of novel human toxoplasmosis control strategies. Current molecular studies on monocyte-Toxoplasma interactions are based on average cell or parasite responses across bulk cell populations. Although informative, population-level averages of monocyte responses to Toxoplasma have sometimes produced contradictory results, such as whether CCL2 or IL12 define effective monocyte response to the parasite. Here, we used single-cell dual RNA sequencing (scDual-Seq) to comprehensively define, for the first time, the monocyte and parasite transcriptional responses that underpin human monocyte-Toxoplasma encounters at the single cell level. We report extreme transcriptional variability between individual monocytes. Furthermore, we report that Toxoplasma-exposed and unexposed monocytes are transcriptionally distinguished by a reactive subset of CD14++CD16- monocytes. Functional cytokine assays on sorted monocyte populations show that the infection-distinguishing monocytes secrete high levels of chemokines, such as CCL2 and CXCL5. These findings uncover the Toxoplasma-induced monocyte transcriptional heterogeneity and shed new light on the cell populations that largely define cytokine and chemokine secretion in human monocytes exposed to Toxoplasma.

genomics↗

bioRxiv2019

Microscaled Proteogenomic Methods for Precision Oncology

Cancer proteogenomics integrates genomics, transcriptomics and mass spectrometry (MS)-based proteomics to gain insights into cancer biology and treatment efficacy. A proteogenomics approach was therefore developed for frozen core biopsies using tissue-sparing specimen processing with a \"microscaled\" proteomics workflow. For technical proof-of-principle, biopsies from ERBB2 positive breast cancers before and 48-72 hours after the first dose of neoadjuvant trastuzumab-based chemotherapy were analyzed. ERBB2 protein and phosphosite levels, as well as mTOR target phosphosites, were significantly more suppressed upon treatment in cases associated with pathological complete response, suggesting MS-based pharmacodynamics is achievable. Furthermore, integrated analyses indicated potential causes of treatment resistance including the absence of ERBB2 amplification (false-ERBB2 positive) and insufficient ERBB2 activity for therapeutic sensitivity despite ERBB2 amplification (pseudo-ERBB2 positive). Candidate resistance features in true-ERBB2+ cases, including androgen receptor signaling, mucin expression and an inactive immune microenvironment were observed. Thus, proteogenomic analysis of needle core biopsies is feasible and clinical utility should be investigated.

cancer biology↗

bioRxiv2019

Development of the duodenal, ileal, jejunal and caecal microbiota in chickens

BackgroundThe chicken intestinal microbiota plays a large role in chicken health and productivity and a greater understanding of its development may lead to interventions to improve chicken nutrition, disease resistance and welfare.\n\nResultsIn this study we examine the duodenal, jejunal, ileal and caecal microbiota of chickens from day of hatch to 5 weeks of age (day 1, 3, 7, 14 and week 5). DNA was extracted from intestinal content samples and the V4 region of the 16S rRNA gene was amplified and sequenced. We identify significant differences in microbial community composition, diversity and richness between samples taken from different locations within the chicken intestinal tract. We also characterise the development of the microbiota at each intestinal site over time.\n\nConclusionsOur study builds upon existing literature to further characterise the development of the chicken intestinal microbiota.

microbiology↗

bioRxiv2019

Assembly of hundreds of novel bacterial genomes from the chicken caecum

Chickens are a highly important source of protein for a large proportion of the human population. The caecal microbiota plays a crucial role in chicken nutrition through the production of short chain fatty acids, nitrogen recycling and amino acid production. In this study we sequenced DNA from caecal contents samples taken from 24 chickens belonging to either a fast or slower growing breed consuming either a vegetable-only diet or a diet containing fish meal. We utilised 1.6T of Illumina data to construct 469 draft metagenome-assembled bacterial genomes, including 460 novel strains, 283 novel species and 42 novel genera. We compared our genomes to data from eight EU countries and show that these genomes are abundant within European chicken flocks. We also compared the abundance of our genomes, and the carbohydrate active enzymes they produce, between our chicken groups and demonstrate that there are both breed- and diet-specific microbiomes, as well as an overlapping core microbiome. This data will form the basis for future studies examining the composition and function of the chicken caecal microbiota.

microbiology↗

bioRxiv2019

An improved pig reference genome sequence to enable pig genetics and genomics research

The domestic pig (Sus scrofa) is important both as a food source and as a biomedical model with high anatomical and immunological similarity to humans. The draft reference genome (Sscrofa10.2) of a purebred Duroc female pig established using older clone-based sequencing methods was incomplete and unresolved redundancies, short range order and orientation errors and associated misassembled genes limited its utility. We present two annotated highly contiguous chromosome-level genome assemblies created with more recent long read technologies and a whole genome shotgun strategy, one for the same Duroc female (Sscrofa11.1) and one for an outbred, composite breed male (USMARCv1.0). Both assemblies are of substantially higher (>90-fold) continuity and accuracy than Sscrofa10.2. These highly contiguous assemblies plus annotation of a further 11 short read assemblies provide an unprecedented view of the genetic make-up of this important agricultural and biomedical model species. We propose that the improved Duroc assembly (Sscrofa11.1) become the reference genome for genomic research in pigs.

genomics↗

bioRxiv2019

Musashi expression in intestinal stem cells attenuates radiation-induced decline in intestinal homeostasis and survival in Drosophila

Exposure to genotoxic stress by environmental agents or treatments, such as radiation therapy, can diminish health span and accelerate aging. We have developed a Drosophila melanogaster model to study the molecular effects of radiation-induced damage and repair. Utilizing a quantitative intestinal permeability assay, we performed an unbiased GWAS screen (using [~]150 strains from the DGRP reference panel) to search for natural genetic variants that regulate radiation-induced gut permeability. From this screen, we identified the RNA binding protein, Musashi (msi).Msi overexpression promoted intestinal stem cell proliferation, which increased survival after irradiation and rescued radiation-induced gut permeability. We identified that a novel role of Msi in ISC proliferation a novel role for Msi in enhancing ISC function following radiation-induced gut damage, which identifies Msi as a potential therapeutic target.

molecular biology↗

bioRxiv2018

Assignment of virus and antimicrobial resistance genes to microbial hosts in a complex microbial community by combined long-read assembly and proximity ligation

The characterization of microbial communities by metagenomic approaches has been enhanced by recent improvements in short-read sequencing efficiency and assembly algorithms. We describe the results of adding long-read sequencing to the mix of technologies used to assemble a highly complex cattle rumen microbial community, and compare the assembly to current short read-based methods applied to the same sample. Contigs in the long-read assembly were 7-fold longer on average, and contained 7-fold more complete open reading frames (ORF), than the short read assembly, despite having three-fold lower sequence depth. The linkages between long-read contigs, provided by proximity ligation data, supported identification of 188 novel viral-host associations in the rumen microbial community that suggest cross-species infectivity of specific viral strains. The improved contiguity of the long-read assembly also identified 94 antimicrobial resistance genes, compared to only seven alleles identified in the short-read assembly. Overall, we demonstrate a combination of experimental and computational methods that work synergistically to improve characterization of biological features in a highly complex rumen microbial community.

microbiology↗

bioRxiv2018

The genomic and proteomic landscape of the rumen microbiome revealed by comprehensive genome-resolved metagenomics

Ruminants provide essential nutrition for billions of people worldwide. The rumen is a specialised stomach adapted to the breakdown of plant-derived complex polysaccharides, and collectively the rumen microbiota encode the thousands of enzymes responsible. Here we present a comprehensive analysis of over 6.5 terabytes of Illumina and Nanopore sequence data, including assembly of 4941 metagenome-assembled genomes, and several single-contig, whole-chromosome assemblies of novel rumen bacteria. We also present the largest dataset of predicted proteins from the rumen, and provide rich annotation against public datasets. Together these data will form an essential part of future studies of rumen microbiome structure and function.

microbiology↗