Using Regulatory and Epistatic Networks to Extend the Findings of a Genome Scan: Identifying the Gene Drivers of Pigmentation in Merino Sheep

Extending genome wide association analysis by the inclusion of gene expression data may assist in the dissection of complex traits. We examined piebald, a pigmentation phenotype in both human and Merino sheep, by analysing multiple data types using a systems approach. First, a case control analysis of 49,034 ovine SNP was performed which confirmed a multigenic basis for the condition. We combined these results with gene expression data from five tissue types analysed with a skin-specific microarray. Promoter sequence analysis of differentially expressed genes allowed us to reverse-engineer a regulatory network. Likewise, by testing two-loci models derived from all pair-wise comparisons across piebald-associated SNP, we generated an epistatic network. At the intersection of both networks, we identified thirteen genes with insulin-like growth factor binding protein 7 (IGFBP7), platelet-derived growth factor alpha (PDGFRA) and the tetraspanin platelet activator CD9 at the kernel of the intersection. Further, we report a number of differentially expressed genes in regions containing highly associated SNP including ATRN, DOCK7, FGFR1OP, GLI3, SILV and TBX15. The application of network theory facilitated co-analysis of genetic variation with gene expression, recapitulated aspects of the known molecular biology of skin pigmentation and provided insights into the transcription regulation and epistatic interactions involved in piebald Merino sheep

Analysis of the complement and molecular evolution of tRNA genes in cow

<p>Abstract</p> <p>Background</p> <p>Detailed information regarding the number and organization of transfer RNA (tRNA) genes at the genome level is becoming readily available with the increase of DNA sequencing of whole genomes. However the identification of functional tRNA genes is challenging for species that have large numbers of repetitive elements containing tRNA derived sequences, such as <it>Bos taurus</it>. Reliable identification and annotation of entire sets of tRNA genes allows the evolution of tRNA genes to be understood on a genomic scale.</p> <p>Results</p> <p>In this study, we explored the <it>B. taurus </it>genome using bioinformatics and comparative genomics approaches to catalogue and analyze cow tRNA genes. The initial analysis of the cow genome using tRNAscan-SE identified 31,868 putative tRNA genes and 189,183 pseudogenes, where 28,830 of the 31,868 predicted tRNA genes were classified as repetitive elements by the RepeatMasker program. We then used comparative genomics to further discriminate between functional tRNA genes and tRNA-derived sequences for the remaining set of 3,038 putative tRNA genes. For our analysis, we used the human, chimpanzee, mouse, rat, horse, dog, chicken and fugu genomes to predict that the number of active tRNA genes in cow lies in the vicinity of 439. Of this set, 150 tRNA genes were 100% identical in their sequences across all nine vertebrate genomes studied. Using clustering analyses, we identified a new tRNA-Gly^CCCsubfamily present in all analyzed mammalian genomes. We suggest that this subfamily originated from an ancestral tRNA-Gly^GCCgene via a point mutation prior to the radiation of the mammalian lineages. Lastly, in a separate analysis we created phylogenetic profiles for each putative cow tRNA gene using a representative set of genomes to gain an overview of common evolutionary histories of tRNA genes.</p> <p>Conclusion</p> <p>The use of a combination of bioinformatics and comparative genomics approaches has allowed the confident identification of a set of cow tRNA genes that will facilitate further studies in understanding the molecular evolution of cow tRNA genes.</p

Changed Patterns of Genomic Variation Following Recent Domestication: Selection Sweeps in Farmed Atlantic Salmon

The introduction of wild Atlantic salmon into captivity, and their subsequent artificial selection for production traits, has caused phenotypic differences between domesticated fish and their wild counterparts. Identification of regions of the genome underling these changes offers the promise of characterizing the early biological consequences of domestication. In the current study, we sequenced a population of farmed European Atlantic salmon and compared the observed patterns of SNP variation to those found in conspecific wild populations. This identified 139 genomic regions that contained significantly elevated SNP homozygosity in farmed fish when compared to their wild counterparts. The most extreme was adjacent to versican, a gene involved in control of neural crest cell migration. To control for false positive signals, a second and independent dataset of farmed and wild European Atlantic salmon was assessed using the same methodology. A total of 81 outlier regions detected in the first dataset showed significantly reduced homozygosity within the second one, strongly suggesting the genomic regions identified are enriched for true selection sweeps. Examination of the associated genes identified a number previously characterized as targets of selection in other domestic species and that have roles in development, behavior and olfactory system. These include arcvf, sema6, errb4, id2-like, and 6n1-like genes. Finally, we searched for evidence of parallel sweeps using a farmed population of North American origin. This failed to detect a convincing overlap to the putative sweeps present in European populations, suggesting the factors that drive patterns of variation under domestication and early artificial selection were largely independent. This is the first analysis on domestication of aquaculture species exploiting whole-genome sequence data and resulted in the identification of sweeps common to multiple independent populations of farmed European Atlantic salmon

A genomics-informed, SNP association study reveals FBLN1 and FABP4 as contributing to resistance to fleece rot in Australian Merino sheep

<p>Abstract</p> <p>Background</p> <p>Fleece rot (FR) and body-strike of Merino sheep by the sheep blowfly <it>Lucilia cuprina </it>are major problems for the Australian wool industry, causing significant losses as a result of increased management costs coupled with reduced wool productivity and quality. In addition to direct effects on fleece quality, fleece rot is a major predisposing factor to blowfly strike on the body of sheep. In order to investigate the genetic drivers of resistance to fleece rot, we constructed a combined ovine-bovine cDNA microarray of almost 12,000 probes including 6,125 skin expressed sequence tags and 5,760 anonymous clones obtained from skin subtracted libraries derived from fleece rot resistant and susceptible animals. This microarray platform was used to profile the gene expression changes between skin samples of six resistant and six susceptible animals taken immediately before, during and after FR induction. Mixed-model equations were employed to normalize the data and 155 genes were found to be differentially expressed (DE). Ten DE genes were selected for validation using real-time PCR on independent skin samples. The genomic regions of a further 5 DE genes were surveyed to identify single nucleotide polymorphisms (SNP) that were genotyped across three populations for their associations with fleece rot resistance.</p> <p>Results</p> <p>The majority of the DE genes originated from the fleece rot subtracted libraries and over-representing gene ontology terms included defense response to bacterium and epidermis development, indicating a role of these processes in modulating the sheep's response to fleece rot. We focused on genes that contribute to the physical barrier function of skin, including keratins, collagens, fibulin and lipid proteins, to identify SNPs that were associated to fleece rot scores.</p> <p>Conclusions</p> <p>We identified FBLN1 (fibulin) and FABP4 (fatty acid binding protein 4) as key factors in sheep's resistance to fleece rot. Validation of these markers in other populations could lead to vital tests for marker assisted selection that will ultimately increase the natural fleece rot resistance of Merino sheep.</p

Anti-apoptotic seminal vesicle protein IV inhibits cell-mediated immunity.

The in vitro effect of seminal vesicle protein IV (SV-IV) on the cytotoxic activity of human natural or acquired cellular immunity has been investigated by standard immunological procedures, a 51Cr-release cytotoxicity assay, and labeled-ligand binding experiments. The data obtained demonstrate that: (1) fluoresceinated or [125I]-labeled SV-IV binds specifically to the surface of human purified non-adherent monuclear cells (NA-MNC); (2)SV-IV suppresses the cytotoxicity of natural killer (NK) cells against K562 target cells, that of IL-2-stimulated NK (LAK) cells against DAUDI target cells, and that of VEL antigen-sensitized cytotoxic T lymphocytes (CTLs) against VEL target cells; (3) treatment of K562 target cells alone with SV-IV decreases their susceptibility to NK-induced lysis. These findings indicate that the protein SV-IV has a marked in vitro inhibitory effect on NK, LAK and CTL cytotoxicity, providing a better understanding of its immune regulatory functions

Using comparative genomics to reorder the human genome sequence into a virtual sheep genome

Using BAC-end sequences, a sparse marker map and the sequences of the human, dog and cow genomes, an accurate and detailed sub-gene level map of the sheep genome has been constructed

Sheep Genome Functional Annotation Reveals Proximal Regulatory Elements Contributed to The Evolution of Modern Breeds

Domestication fundamentally reshaped animal morphology, physiology and behaviour, offering the opportunity to investigate the molecular processes driving evolutionary change. Here we assess sheep domestication and artificial selection by comparing genome sequence from 43 modern breeds (Ovis aries) and their Asian mouflon ancestor (O. orientalis) to identify selection sweeps. Next, we provide a comparative functional annotation of the sheep genome, validated using experimental ChIP-Seq of sheep tissue. Using these annotations, we evaluate the impact of selection and domestication on regulatory sequences and find that sweeps are significantly enriched for protein coding genes, proximal regulatory elements of genes and genome features associated with active transcription. Finally, we find individual sites displaying strong allele frequency divergence are enriched for the same regulatory features. Our data demonstrate that remodelling of gene expression is likely to have been one of the evolutionary forces that drove phenotypic diversification of this common livestock species

Skeletal muscle specific genes networks in cattle

While physiological differences across skeletal muscles have been described, the differential gene expression underlying them and the discovery of how they interact to perform specific biological processes are largely to be elucidated. The purpose of the present study was, firstly, to profile by cDNA microarrays the differential gene expression between two skeletal muscle types, Psoas major (PM) and Flexor digitorum (FD), in beef cattle and then to interpret the results in the context of a bovine gene coexpression network, detecting possible changes in connectivity across the skeletal muscle system. Eighty four genes were differentially expressed (DE) between muscles. Approximately 54% encoded metabolic enzymes and structural-contractile proteins. DE genes were involved in similar processes and functions, but the proportion of genes in each category varied within each muscle. A correlation matrix was obtained for 61 out of the 84 DE genes from a gene coexpression network. Different groups of coexpression were observed, the largest one having 28 metabolic and contractile genes, up-regulated in PM, and mainly encoding fast-glycolytic fibre structural components and glycolytic enzymes. In FD, genes related to cell support seemed to constitute its identity feature and did not positively correlate to the rest of DE genes in FD. Moreover, changes in connectivity for some DE genes were observed in the different gene ontologies. Our results confirm the existence of a muscle dependent transcription and coexpression pattern and suggest the necessity of integrating different muscle types to perform comprehensive networks for the transcriptional landscape of bovine skeletal muscle

Bovine proteins containing poly-glutamine repeats are often polymorphic and enriched for components of transcriptional regulatory complexes

peer-reviewedBackground: About forty human diseases are caused by repeat instability mutations. A distinct subset of these diseases is the result of extreme expansions of polymorphic trinucleotide repeats; typically CAG repeats encoding poly-glutamine (poly-Q) tracts in proteins. Polymorphic repeat length variation is also apparent in human poly-Q encoding genes from normal individuals. As these coding sequence repeats are subject to selection in mammals, it has been suggested that normal variations in some of these typically highly conserved genes are implicated in morphological differences between species and phenotypic variations within species. At present, poly-Q encoding genes in non-human mammalian species are poorly documented, as are their functions and propensities for polymorphic variation. Results: The current investigation identified 178 bovine poly-Q encoding genes (Q ≥ 5) and within this group, 26 genes with orthologs in both human and mouse that did not contain poly-Q repeats. The bovine poly-Q encoding genes typically had ubiquitous expression patterns although there was bias towards expression in epithelia, brain and testes. They were also characterised by unusually large sizes. Analysis of gene ontology terms revealed that the encoded proteins were strongly enriched for functions associated with transcriptional regulation and many contributed to physical interaction networks in the nucleus where they presumably act cooperatively in transcriptional regulatory complexes. In addition, the coding sequence CAG repeats in some bovine genes impacted mRNA splicing thereby generating unusual transcriptional diversity, which in at least one instance was tissue-specific. The poly-Q encoding genes were prioritised using multiple criteria for their likelihood of being polymorphic and then the highest ranking group was experimentally tested for polymorphic variation within a cattle diversity panel. Extensive and meiotically stable variation was identified. Conclusions: Transcriptional diversity can potentially be generated in poly-Q encoding genes by the impact of CAG repeat tracts on mRNA alternative splicing. This effect, combined with the physical interactions of the encoded proteins in large transcriptional regulatory complexes suggests that polymorphic variations of proteins in these complexes have strong potential to affect phenotype.Dairy Australia (through the Innovative Dairy Cooperative Research Center