Biology subjects

Bush, S. J.

Publications and source records attributed to Bush, S. J..

8 recordsLinked to original sources

bioRxiv2018

Comprehensive transcriptional profiling of the gastrointestinal tract of ruminants from birth to adulthood reveals strong developmental stage specific gene expression

One of the most significant physiological challenges to neonatal and juvenile ruminants is the development and establishment of the rumen. Using a subset of RNA-Seq data from our high-resolution atlas of gene expression in sheep (Ovis aries) we have provided the first comprehensive characterisation of transcription of the entire the gastrointestinal (GI) tract during the transition from pre-ruminant to ruminant. The dataset comprises 168 tissue samples from sheep at four different time points (birth, one week, 8 weeks and adult). Using network cluster analysis we illustrate how the complexity of the GI tract is reflected in tissue- and developmental stage-specific differences in gene expression. The most significant transcriptional differences between neonatal and adult sheep were observed in the rumen complex. Differences in transcription between neonatal and adult sheep were particularly evident in macrophage specific signatures indicating they might be driving the observed developmental stage-specific differences. Comparative analysis of gene expression in three GI tract tissues from age-matched sheep and goats revealed species-specific differences in genes involved in immunity and metabolism. This study improves our understanding of the transcriptomic mechanisms involved in the transition from pre-ruminant to ruminant. It highlights key genes involved in immunity, microbe recognition, metabolism and cellular differentiation in the GI tract. The results form a basis for future studies linking gene expression with microbial colonisation of the developing GI tract and will contribute towards identifying genes that underlie immunity in early development, which could be utilised to improve ruminant efficiency and productivity.\n\nReference Numbers for Data in the Public RepositoriesThe raw RNA-Sequencing data are deposited in the European Nucleotide Archive (ENA) under study accessions PRJEB19199 (sheep) and PRJEB23196 (goat). Metadata for all samples is deposited in the EBI BioSamples database under group identifiers SAMEG317052 (sheep) and SAMEG330351 (goat).

genomics↗

bioRxiv2018

Arginine to glutamine mutation in olfactomedin-like 3 (OLFML3) is a candidate for severe goniodysgenesis and glaucoma in the Border Collie dog breed.

Goniodysgenesis is a developmental abnormality of the anterior chamber of the eye. It is generally considered to be congenital in dogs (Canis lupus familiaris), and has been associated with glaucoma and blindness. Goniodysgenesis and early-onset glaucoma initially emerged in Border Collies in Australia in the late 1990s and has subsequently been found in Europe and the USA. The objective of the present study was to determine the genetic basis of goniodysgenesis in Border Collies. Clinical diagnosis was based on results of examinations by veterinary ophthalmologists of affected and unaffected dogs from eleven different countries. Genotyping using the Illumina high density canine SNP chip and whole genome sequencing were used to identify candidate genetic regions. Expression profiles and evolutionary conservation of candidate genes were assessed using public databases. Analysis of pedigree information was consistent with an autosomal recessive mode of inheritance for severe goniodysgenesis (potentially leading to glaucoma) in this breed. There was a highly significant peak of association over chromosome 17, with a p-value of 2 x 10-13. Whole genome sequences of three dogs with glaucoma, three severely affected and three unaffected dogs identified a missense variant in the olfactomedin like 3 (OLFML3) gene in all six affected animals. This was homozygous in all nine cases with glaucoma and 12 of 14 other severely affected animals. Of 67 reportedly unaffected animals, only one (offspring of two homozygous affected parents) was homozygous for this variant. The identification of a candidate genetic region and putative causative mutation will aid breeders to reduce the frequency of goniodysgenesis and the risk of glaucoma in the Border Collie population.

genetics↗

bioRxiv2018

Characterisation of the UK honey bee (Apis mellifera) metagenome.

The European honey bee (Apis mellifera) plays a major role in pollination and food production, but is under threat from emerging pathogens and agro-environmental insults. As with other organisms, honey bee health is a complex product of environment, host genetics and associated microbes (commensal, opportunistic and pathogenic). Improved understanding of bee genetics and their molecular ecology can help manage modern challenges to bee health and production. Sampling bee and cobiont genomes, we characterised the metagenome of 19 honey bee colonies across Britain. Low heterozygosity was observed in bees from many Scottish colonies, sharing high similarity to the native dark bee, A. mellifera mellifera. Apiaries exhibited high diversity in the composition and relative abundance of individual microbiome taxa. Most non-bee sequences derived from known honey bee commensal bacteria or known pathogens, e.g. Lotmaria passim (Trypanosomatidae), and Nosema spp. (Microsporidia). However, DNA was also detected from numerous additional bacterial, plant (food source), protozoan and metazoan organisms. To classify sequences from cobionts lacking genomic information, we developed a novel network analysis approach clustering orphan contigs, allowing the identification of a pathogenic gregarine. Our analyses demonstrate the power of high-throughput, directed metagenomics in agroecosystems identifying potential threats to honey bees present in their microbiota.

genomics↗

bioRxiv2018

Combination of novel and public RNA-seq datasets to generate an mRNA expression atlas for the domestic chicken

BackgroundThe domestic chicken (Gallus gallus) is widely used as a model in developmental biology and is also an important livestock species. We describe a novel approach to data integration to generate an mRNA expression atlas for the chicken spanning major tissue types and developmental stages, using a diverse range of publicly-archived RNA-seq datasets and new data derived from immune cells and tissues.\n\nResultsRandomly down-sampling RNA-seq datasets to a common depth and quantifying expression against a reference transcriptome using the mRNA quantitation tool Kallisto ensured that disparate datasets explored comparable transcriptomic space. The network analysis tool Miru was used to extract clusters of co-expressed genes from the resulting expression atlas, many of which were tissue or cell-type restricted, contained transcription factors that have previously been implicated in their regulation, or were otherwise associated with biological processes, such as the cell cycle. The atlas provides a resource for the functional annotation of genes that currently have only a locus ID. We cross-referenced the RNA-seq atlas to a publicly available embryonic Cap Analysis of Gene Expression (CAGE) dataset to infer the developmental time course of organ systems, and to identify a signature of the expansion of tissue macrophage populations during development.\n\nConclusionExpression profiles obtained from public RNA-seq datasets - despite being generated by different laboratories using different methodologies - can be made comparable to each other. This meta-analytic approach to RNA-seq can be extended with new datasets from novel tissues, and is applicable to any species.

genomics↗

bioRxiv2018

Characterisation of subpopulations of chicken mononuclear phagocytes that express TIM4 and the macrophage colony-stimulating factor receptor (CSF1R)

The phosphatidylserine receptor, TIM4, encoded by TIMD4, mediates the phagocytic uptake of apoptotic cells. We applied anti-chicken TIM4 monoclonal antibodies, in combination with CSF1R reporter transgenes to dissect the function of TIM4 in chick (Gallus gallus). During development in ovo, TIM4 was present on the large majority of macrophages but expression became more heterogeneous post-hatch. Blood monocytes expressed KUL01, class II MHC and CSF1R-mApple uniformly. Around 50% of monocytes were positive for surface TIM4. They also expressed many other monocyte-specific transcripts at a higher level than TIM4- monocytes. In liver, highly-phagocytic TIM4hi cells shared many transcripts with mammalian Kupffer cells and were associated with uptake of apoptotic cells. Although they expressed CSF1R mRNA, Kupffer cells did not express the CSF1R-mApple transgene, suggesting that additional CSF1R transcriptional regulatory elements are required by these cells. By contrast, CSF1R-mApple was detected in liver TIM4lo and TIM4- cells which were not phagocytic and were more abundant than Kupffer cells. These cells expressed CSF1R, alongside high levels of FLT3, MHCII, XCR1 and other markers associated with conventional dendritic cells (cDC) in mice. In bursa, TIM4 was present on the cell surface of two populations. Like Kupffer cells, bursal TIM4hi phagocytes co-expressed many receptors involved in apoptotic cell recognition. TIM4lo cells appear to be a sub-population of bursal B cells. In overview, TIM4 is associated with phagocytes that eliminate apoptotic cells in the chick. In the liver, TIM4 and CSF1R reporters distinguished Kupffer cells from an abundant population of DC-like cells.

immunology↗

bioRxiv2018

Environmentally enriched pigs have transcriptional profiles consistent with neuroprotective effects and microglial depletion

Environmental enrichment (EE) is widely used to study the effects of external factors on brain development, function and health in rodent models, but very little is known of the effects of EE on the brain in a large animal model such as the pig. Twenty-four young pigs (aged 5 weeks at start of study, 1:1 male: female ratio) were housed in environmentally enriched (EE) pens and provided with additional enrichment stimulation (a bag filled with straw) once daily. Litter, weight and sex matched controls n= (24) were housed in barren (B) conditions. Behaviour was recorded on alternate days from study day 10. After 21 days, RNA-sequencing of the frontal cortex of male piglets culled one hour after the enrichment stimulation, but not those at 4 hours after stimulation, showed upregulation of genes involved in neuronal activity and synaptic plasticity in the EE compared to the B condition. This result is mirrored in the behavioural response to the stimulation which showed a peak in activity around the 1 hour time-point. By contrast, EE piglets displayed a signature consistent with a relative decrease in microglial activity compared to those in the B condition. These results confirm those from rodents, suggesting that EE may also confer neuronal health benefits in large mammal models, through a potential relative reduction in neuroinflammatory process and increase in neuroprotection driven by an enrichment-induced increase in behavioural activity.

molecular biology↗

bioRxiv2018

Assembly and validation of conserved long non-coding RNAs in the ruminant transcriptome

mRNA-like long non-coding RNAs (lncRNA) are a significant component of mammalian transcriptomes, although most are expressed only at low levels, with high tissue-specificity and/or at specific developmental stages. In many cases, therefore, lncRNA detection by RNA-sequencing (RNA-seq) is compromised by stochastic sampling. To account for this and create a catalogue of ruminant lncRNA, we compared de novo assembled lncRNA derived from large RNA-seq datasets in transcriptional atlas projects for sheep and goats with previous lncRNA assembled in cattle and human. Few lncRNA could be reproducibly assembled from a single dataset, even with deep sequencing of the same tissues from multiple animals. Furthermore, there was little sequence overlap between lncRNA assembled from pooled RNA-seq data. We combined positional conservation (synteny) with cross-species mapping of candidate lncRNA to identify a consensus set of ruminant lncRNA and then used the RNA-seq data to demonstrate detectable and reproducible expression in each species. The majority of lncRNA were encoded by single exons, and expressed at < 1 TPM. In sheep, 20-30% of lncRNA had expression profiles significantly correlated with neighbouring protein-coding genes, suggesting association with enhancers. Alongside substantially expanding the ruminant lncRNA repertoire, the outcomes of our analysis demonstrate that stochastic sampling can be partly overcome by combining RNA-seq datasets from related species. This has practical implications for the future discovery of lncRNA in other species.

genomics↗

bioRxiv2017

A High Resolution Atlas Of Gene Expression In The Domestic Sheep (Ovis aries)

Sheep are a key source of meat, milk and fibre for the global livestock sector, and an important biomedical model. Global analysis of gene expression across multiple tissues has aided genome annotation and supported functional annotation of mammalian genes. We present a large-scale RNA-Seq dataset representing all the major organ systems from adult sheep and from several juvenile, neonatal and prenatal developmental time points. The Ovis aries reference genome (Oar v3.1) includes 27,504 genes (20,921 protein coding), of which 25,350 (19,921 protein coding) had detectable expression in at least one tissue in the sheep gene expression atlas dataset. Network-based cluster analysis of this dataset grouped genes according to their expression pattern. The principle of guilt by association was used to infer the function of uncharacterised genes from their co-expression with genes of known function. We describe the overall transcriptional signatures present in the sheep gene expression atlas and assign those signatures, where possible, to specific cell populations or pathways. The findings are related to innate immunity by focusing on clusters with an immune signature, and to the advantages of cross-breeding by examining the patterns of genes exhibiting the greatest expression differences between purebred and crossbred animals. This high-resolution gene expression atlas for sheep is, to our knowledge, the largest transcriptomic dataset from any livestock species to date. It provides a resource to improve the annotation of the current reference genome for sheep, presenting a model transcriptome for ruminants and insight into gene, cell and tissue function at multiple developmental stages.\n\nAuthor SummarySheep are ruminant mammals kept as livestock for the production of meat, milk and wool in agricultural industries across the globe. Genetic and genomic information can be used to improve production traits such as disease resiliance. The sheep genome is however missing important information relating to gene function and many genes, which may be important for productivity, have no informative gene name. This can be remedied using RNA-Sequencing to generate a global expression profile of all protein-coding genes, across multiple organ systems and developmental stages. Clustering genes based on their expression profile across tissues and cells allows us to assign function to those genes. If for example a gene with no informative gene name is expressed in macrophages and is found within a cluster of known macrophage related genes it is likely to be involved in macrophage function and play a role in innate immunity. This information improves the quality of the reference genome and provides insight into biological processes underlying the complex traits that influence the productivity of sheep and other livestock species.

genomics↗