Browsing by Author "Littlejohn MD"

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    A Capra hircus chromosome 19 locus linked to milk production influences mammary conformation
    (BioMed Central Ltd, 2022-02-11) Jiang A; Ankersmit-Udy A; Turner S-A; Scholtens M; Littlejohn MD; Lopez-Villalobos N; Proser CG; Snell RG; Lehnert K
    Background Economically important milk production traits including milk volume, milk fat and protein yield vary considerably across dairy goats in New Zealand. A significant portion of the variation is attributable to genetic variation. Discovery of genetic markers linked to milk production traits can be utilised to drive selection of high-performance animals. A previously reported genome wide association study across dairy goats in New Zealand identified a quantitative trait locus (QTL) located on chromosome 19. The most significantly associated single nucleotide polymorphism (SNP) marker for this locus is located at position 26,610,610 (SNP marker rs268292132). This locus is associated with multiple milk production traits including fat, protein and volume. The predicted effect of selection for the beneficial haplotype would result in an average production increase of 2.2 kg fat, 1.9 kg protein and 73.6 kg milk yield. An outstanding question was whether selection for the beneficial allele would co-select for any negative pleiotropic effects. An adverse relationship between milk production and udder health traits has been reported at this locus. Therefore, a genome wide association study was undertaken looking for loci associated with udder traits. Results The QTL and production associated marker rs268292132 was identified in this study to also be associated with several goat udder traits including udder depth (UD), fore udder attachment (FUA) and rear udder attachment (RUA). Our study replicates the negative relationship between production and udder traits with the high production allele at position 19:26,610,610 (SNP marker rs268292132) associated with an adverse change in UD, FUA and RUA. Conclusions Our study has confirmed the negative relationship between udder traits and production traits in the NZ goat population. We have found that the frequency of the high production allele is relatively high in the NZ goat population, indicating that its effect on udder conformation is not significantly detrimental on animal health. It will however be important to monitor udder conformation as the chromosome 19 locus is progressively implemented for marker assisted selection. It will also be of interest to determine if the gene underlying the production QTL has a direct effect on mammary gland morphology or whether the changes observed are a consequence of the increased milk volume.
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    A frameshift-deletion mutation in Reelin causes cerebellar hypoplasia in White Swiss Shepherd dogs
    (John Wiley & Sons Ltd on behalf of Stichting International Foundation for Animal Genetics, 2023-10) Littlejohn MD; Sneddon N; Dittmer K; Keehan M; Stephen M; Drögemüller M; Garrick D
    Cerebellar hypoplasia is a heterogeneous neurological condition in which the cerebellum is smaller than usual or not completely developed. The condition can have genetic origins, with Mendelian-effect mutations described in several mammalian species. Here, we describe a genetic investigation of cerebellar hypoplasia in White Swiss Shepherd dogs, where two affected puppies were identified from a litter with a recent common ancestor on both sides of their pedigree. Whole genome sequencing was conducted for 10 dogs in this family, and filtering of these data based on a recessive transmission hypothesis highlighted five protein-altering candidate variants - including a frameshift-deletion of the Reelin (RELN) gene (p.Val947*). Given the status of RELN as a gene responsible for cerebellar hypoplasia in humans, sheep and mice, these data strongly suggest the loss-of-function variant as underlying these effects. This variant has not been found in other dog breeds nor in a cohort of European White Swiss Shepherds, suggesting a recent mutation event. This finding will support the genotyping of a more diverse sample of dogs, and should aid future management of the harmful allele through optimised mating schemes.
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    A new mechanism for a familiar mutation - bovine DGAT1 K232A modulates gene expression through multi-junction exon splice enhancement
    (BioMed Central Ltd, 2020-08-26) Fink T; Lopdell TJ; Tiplady K; Handley R; Johnson TJJ; Spelman RJ; Davis SR; Snell RG; Littlejohn MD
    BACKGROUND: The DGAT1 gene encodes an enzyme responsible for catalysing the terminal reaction in mammary triglyceride synthesis, and underpins a well-known pleiotropic quantitative trait locus (QTL) with a large influence on milk composition phenotypes. Since first described over 15 years ago, a protein-coding variant K232A has been assumed as the causative variant underlying these effects, following in-vitro studies that demonstrated differing levels of triglyceride synthesis between the two protein isoforms. RESULTS: We used a large RNAseq dataset to re-examine the underlying mechanisms of this large milk production QTL, and hereby report novel expression-based functions of the chr14 g.1802265AA > GC variant that encodes the DGAT1 K232A substitution. Using expression QTL (eQTL) mapping, we demonstrate a highly-significant mammary eQTL for DGAT1, where the K232A mutation appears as one of the top associated variants for this effect. By conducting in vitro expression and splicing experiments in bovine mammary cell culture, we further show modulation of splicing efficiency by this mutation, likely through disruption of an exon splice enhancer as a consequence of the allele encoding the 232A variant. CONCLUSIONS: The relative contributions of the enzymatic and transcription-based mechanisms now attributed to K232A remain unclear; however, these results suggest that transcriptional impacts contribute to the diversity of lactation effects observed at the DGAT1 locus.
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    Allele-specific binding variants causing ChIP-seq peak height of histone modification are not enriched in expression QTL annotations.
    (BioMed Central Ltd, 2024-06-27) Ghoreishifar M; Chamberlain AJ; Xiang R; Prowse-Wilkins CP; Lopdell TJ; Littlejohn MD; Pryce JE; Goddard ME
    BACKGROUND: Genome sequence variants affecting complex traits (quantitative trait loci, QTL) are enriched in functional regions of the genome, such as those marked by certain histone modifications. These variants are believed to influence gene expression. However, due to the linkage disequilibrium among nearby variants, pinpointing the precise location of QTL is challenging. We aimed to identify allele-specific binding (ASB) QTL (asbQTL) that cause variation in the level of histone modification, as measured by the height of peaks assayed by ChIP-seq (chromatin immunoprecipitation sequencing). We identified DNA sequences that predict the difference between alleles in ChIP-seq peak height in H3K4me3 and H3K27ac histone modifications in the mammary glands of cows. RESULTS: We used a gapped k-mer support vector machine, a novel best linear unbiased prediction model, and a multiple linear regression model that combines the other two approaches to predict variant impacts on peak height. For each method, a subset of 1000 sites with the highest magnitude of predicted ASB was considered as candidate asbQTL. The accuracy of this prediction was measured by the proportion where the predicted direction matched the observed direction. Prediction accuracy ranged between 0.59 and 0.74, suggesting that these 1000 sites are enriched for asbQTL. Using independent data, we investigated functional enrichment in the candidate asbQTL set and three control groups, including non-causal ASB sites, non-ASB variants under a peak, and SNPs (single nucleotide polymorphisms) not under a peak. For H3K4me3, a higher proportion of the candidate asbQTL were confirmed as ASB when compared to the non-causal ASB sites (P < 0.01). However, these candidate asbQTL did not enrich for the other annotations, including expression QTL (eQTL), allele-specific expression QTL (aseQTL) and sites conserved across mammals (P > 0.05). CONCLUSIONS: We identified putatively causal sites for asbQTL using the DNA sequence surrounding these sites. Our results suggest that many sites influencing histone modifications may not directly affect gene expression. However, it is important to acknowledge that distinguishing between putative causal ASB sites and other non-causal ASB sites in high linkage disequilibrium with the causal sites regarding their impact on gene expression may be challenging due to limitations in statistical power.
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    Comparison of the genetic characteristics of directly measured and Fourier-transform mid-infrared-predicted bovine milk fatty acids and proteins.
    (Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association, 2022-12) Tiplady KM; Lopdell TJ; Sherlock RG; Johnson TJJ; Spelman RJ; Harris BL; Davis SR; Littlejohn MD; Garrick DJ
    Fourier-transform mid-infrared (FT-MIR) spectroscopy is a high-throughput and inexpensive methodology used to evaluate concentrations of fat and protein in dairy cattle milk samples. The objective of this study was to compare the genetic characteristics of FT-MIR predicted fatty acids and individual milk proteins with those that had been measured directly using gas and liquid chromatography methods. The data used in this study was based on 2,005 milk samples collected from 706 Holstein-Friesian × Jersey animals that were managed in a seasonal, pasture-based dairy system, with milk samples collected across 2 consecutive seasons. Concentrations of fatty acids and protein fractions in milk samples were directly determined by gas chromatography and high-performance liquid chromatography, respectively. Models to predict each directly measured trait based on FT-MIR spectra were developed using partial least squares regression, with spectra from a random selection of half the cows used to train the models, and predictions for the remaining cows used as validation. Variance parameters for each trait and genetic correlations for each pair of measured/predicted traits were estimated from pedigree-based bivariate models using REML procedures. A genome-wide association study was undertaken using imputed whole-genome sequence, and quantitative trait loci (QTL) from directly measured traits were compared with QTL from the corresponding FT-MIR predicted traits. Cross-validation prediction accuracies based on partial least squares for individual and grouped fatty acids ranged from 0.18 to 0.65. Trait prediction accuracies in cross-validation for protein fractions were 0.53, 0.19, and 0.48 for α-casein, β-casein, and κ-casein, 0.31 for α-lactalbumin, 0.68 for β-lactoglobulin, and 0.36 for lactoferrin. Heritability estimates for directly measured traits ranged from 0.07 to 0.55 for fatty acids; and from 0.14 to 0.63 for individual milk proteins. For FT-MIR predicted traits, heritability estimates were mostly higher than for the corresponding measured traits, ranging from 0.14 to 0.46 for fatty acids, and from 0.30 to 0.70 for individual proteins. Genetic correlations between directly measured and FT-MIR predicted protein fractions were consistently above 0.75, with the exceptions of C18:0 and C18:3 cis-3, which had genetic correlations of 0.72 and 0.74, respectively. The GWAS identified trait QTL for fatty acids with likely candidates in the DGAT1, CCDC57, SCD, and GPAT4 genes. Notably, QTL for SCD were largely absent in the FT-MIR predicted traits, and QTL for GPAT4 were absent in directly measured traits. Similarly, for directly measured individual proteins, we identified QTL with likely candidates in the CSN1S1, CSN3, PAEP, and LTF genes, but the QTL for CSN3 and LTF were absent in the FT-MIR predicted traits. Our study indicates that genetic correlations between directly measured and FT-MIR predicted fatty acid and protein fractions are typically high, but that phenotypic variation in these traits may be underpinned by differing genetic architecture.
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    Genetic variation in histone modifications and gene expression identifies regulatory variants in the mammary gland of cattle
    (BioMed Central Ltd, 2022-12-08) Prowse-Wilkins CP; Lopdell TJ; Xiang R; Vander Jagt CJ; Littlejohn MD; Chamberlain AJ; Goddard ME
    BACKGROUND: Causal variants for complex traits, such as eQTL are often found in non-coding regions of the genome, where they are hypothesised to influence phenotypes by regulating gene expression. Many regulatory regions are marked by histone modifications, which can be assayed by chromatin immunoprecipitation followed by sequencing (ChIP-seq). Sequence reads from ChIP-seq form peaks at putative regulatory regions, which may reflect the amount of regulatory activity at this region. Therefore, eQTL which are also associated with differences in histone modifications are excellent candidate causal variants. RESULTS: We assayed the histone modifications H3K4Me3, H3K4Me1 and H3K27ac and mRNA in the mammary gland of up to 400 animals. We identified QTL for peak height (histone QTL), exon expression (eeQTL), allele specific expression (aseQTL) and allele specific binding (asbQTL). By intersecting these results, we identify variants which may influence gene expression by altering regulatory regions of the genome, and may be causal variants for other traits. Lastly, we find that these variants are found in putative transcription factor binding sites, identifying a mechanism for the effect of many eQTL. CONCLUSIONS: We find that allele specific and traditional QTL analysis often identify the same genetic variants and provide evidence that many eQTL are regulatory variants which alter activity at regulatory regions of the bovine genome. Our work provides methodological and biological updates on how regulatory mechanisms interplay at multi-omics levels.
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    Holstein Friesian dairy cattle edited for diluted coat color as a potential adaptation to climate change
    (BioMed Central Ltd, 2021-11-26) Laible G; Cole S-A; Brophy B; Wei J; Leath S; Jivanji S; Littlejohn MD; Wells DN
    BACKGROUND: High-producing Holstein Friesian dairy cattle have a characteristic black and white coat, often with large proportions of black. Compared to a light coat color, black absorbs more solar radiation which is a contributing factor to heat stress in cattle. To better adapt dairy cattle to rapidly warming climates, we aimed to lighten their coat color by genome editing. RESULTS: Using gRNA/Cas9-mediated editing, we introduced a three bp deletion in the pre-melanosomal protein 17 gene (PMEL) proposed as causative variant for the semi-dominant color dilution phenotype observed in Galloway and Highland cattle. Calves generated from cells with homozygous edits revealed a strong color dilution effect. Instead of the characteristic black and white markings of control calves generated from unedited cells, the edited calves displayed a novel grey and white coat pattern. CONCLUSION: This, for the first time, verified the causative nature of the PMEL mutation for diluting the black coat color in cattle. Although only one of the calves was healthy at birth and later succumbed to a naval infection, the study showed the feasibility of generating such edited animals with the possibility to dissect the effects of the introgressed edit and other interfering allelic variants that might exist in individual cattle and accurately determine the impact of only the three bp change.
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    Non-additive QTL mapping of lactation traits in 124,000 cattle reveals novel recessive loci
    (BioMed Central Ltd, 2022-12) Reynolds EGM; Lopdell T; Wang Y; Tiplady KM; Harland CS; Johnson TJJ; Neeley C; Carnie K; Sherlock RG; Couldrey C; Davis SR; Harris BL; Spelman RJ; Garrick DJ; Littlejohn MD
    BACKGROUND: Deleterious recessive conditions have been primarily studied in the context of Mendelian diseases. Recently, several deleterious recessive mutations with large effects were discovered via non-additive genome-wide association studies (GWAS) of quantitative growth and developmental traits in cattle, which showed that quantitative traits can be used as proxies of genetic disorders when such traits are indicative of whole-animal health status. We reasoned that lactation traits in cattle might also reflect genetic disorders, given the increased energy demands of lactation and the substantial stresses imposed on the animal. In this study, we screened more than 124,000 cows for recessive effects based on lactation traits. RESULTS: We discovered five novel quantitative trait loci (QTL) that are associated with large recessive impacts on three milk yield traits, with these loci presenting missense variants in the DOCK8, IL4R, KIAA0556, and SLC25A4 genes or premature stop variants in the ITGAL, LRCH4, and RBM34 genes, as candidate causal mutations. For two milk composition traits, we identified several previously reported additive QTL that display small dominance effects. By contrasting results from milk yield and milk composition phenotypes, we note differing genetic architectures. Compared to milk composition phenotypes, milk yield phenotypes had lower heritabilities and were associated with fewer additive QTL but had a higher non-additive genetic variance and were associated with a higher proportion of loci exhibiting dominance. CONCLUSIONS: We identified large-effect recessive QTL which are segregating at surprisingly high frequencies in cattle. We speculate that the differences in genetic architecture between milk yield and milk composition phenotypes derive from underlying dissimilarities in the cellular and molecular representation of these traits, with yield phenotypes acting as a better proxy of underlying biological disorders through presentation of a larger number of major recessive impacts.
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    Sequence-based genome-wide association study of individual milk mid-infrared wavenumbers in mixed-breed dairy cattle
    (BioMed Central Ltd, 2021-07-20) Tiplady KM; Lopdell TJ; Reynolds E; Sherlock RG; Keehan M; Johnson TJJ; Pryce JE; Davis SR; Spelman RJ; Harris BL; Garrick DJ; Littlejohn MD
    BACKGROUND: Fourier-transform mid-infrared (FT-MIR) spectroscopy provides a high-throughput and inexpensive method for predicting milk composition and other novel traits from milk samples. While there have been many genome-wide association studies (GWAS) conducted on FT-MIR predicted traits, there have been few GWAS for individual FT-MIR wavenumbers. Using imputed whole-genome sequence for 38,085 mixed-breed New Zealand dairy cattle, we conducted GWAS on 895 individual FT-MIR wavenumber phenotypes, and assessed the value of these direct phenotypes for identifying candidate causal genes and variants, and improving our understanding of the physico-chemical properties of milk. RESULTS: Separate GWAS conducted for each of 895 individual FT-MIR wavenumber phenotypes, identified 450 1-Mbp genomic regions with significant FT-MIR wavenumber QTL, compared to 246 1-Mbp genomic regions with QTL identified for FT-MIR predicted milk composition traits. Use of mammary RNA-seq data and gene annotation information identified 38 co-localized and co-segregating expression QTL (eQTL), and 31 protein-sequence mutations for FT-MIR wavenumber phenotypes, the latter including a null mutation in the ABO gene that has a potential role in changing milk oligosaccharide profiles. For the candidate causative genes implicated in these analyses, we examined the strength of association between relevant loci and each wavenumber across the mid-infrared spectrum. This revealed shared association patterns for groups of genomically-distant loci, highlighting clusters of loci linked through their biological roles in lactation and their presumed impacts on the chemical composition of milk. CONCLUSIONS: This study demonstrates the utility of FT-MIR wavenumber phenotypes for improving our understanding of milk composition, presenting a larger number of QTL and putative causative genes and variants than found from FT-MIR predicted composition traits. Examining patterns of significance across the mid-infrared spectrum for loci of interest further highlighted commonalities of association, which likely reflects the physico-chemical properties of milk constituents.
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    Wool shedding and lamb fleece weights: first-cross and backcross Wiltshire–Romney sheep scored at lamb, hogget and two-tooth ages
    (Taylor and Francis Group, 2024-05-03) Sneddon NW; Handcock RC; Corner-Thomas RA; Kenyon PR; Burnham DL; Garrick DJ; Littlejohn MD; Blair HT; Morris ST
    With decreasing wool values, interest is increasing regarding shedding sheep. To investigate this, two long-term studies introducing Wiltshire genes into Romney flocks were initiated. Data from these two studies provide phenotypic relationships between a range of shedding scores at different ages. The data included shedding scores (on a 0–5 scale) repeated on lambs (∼5 months), hoggets (∼14–18 months) and two-tooths (∼27 months), and lamb fleece weights. Positive relationships between shedding scores on the same animals were observed. Lamb fleece weight was negatively correlated with all shedding scores. Lamb shedding score in February had a correlation of 0.54 (P < 0.001) with the February score as a hogget at Riverside farm. Scoring wool shedding is a laborious activity requiring individual animals to be scored in the shearing position. Therefore, lamb fleece weight was investigated for its relationship with shedding scores, as a potentially easier alternative. Lamb fleece weight had a greater correlation with February hogget shedding score than with the lamb shedding score (−0.76 vs −0.52, P < 0.001). This study indicated that February scores are an accurate predictor of future shedding phenotypes, and when used in conjunction with fleece weight, are a good predictor of phenotypes expressed at later ages.