Biology subjects

Buckley, R. M.

Publications and source records attributed to Buckley, R. M..

10 recordsLinked to original sources

bioRxiv2024

A genotype-first approach identifies variants for orofacial clefts and other phenotypes in dogs

Dog breeding promotes within-group homogeneity through conformation to strict breed standards, and also drives between-group heterogeneity in pursuit of characteristic breed traits. There are over 350 recognized dog breeds that provide the foundation for investigating the genetic basis of phenotypic diversity. Typically, breed standard phenotypes such as stature, fur length, and craniofacial structure are analyzed in genetic association studies. However, such analyses are limited to the assayed phenotypes, leaving difficult to measure phenotypic subtleties potentially overlooked. In this study, the genotype-first approach was adapted to the dog genome to investigate coding variation from over 2000 dogs, leading to discoveries of new mutations related to craniofacial morphology and stature. Breed-enriched variants were prioritized according to gene constraint, which was calculated using a mutation model derived from trinucleotide substitution probabilities in the dog. Among the discovered variants was a splice-acceptor mutation in PDGFRA associated with bifid nose, a characteristic trait of Catalburun dogs, implicating the genes role in midline closure, and a frameshift mutation in LCORL associated with large canine body size, thus highlighting the importance of allelic heterogeneity in selection for breed traits. Most priority variants were not associated with genomic signatures for breed differentiation, as these regions were enriched for constrained genes intolerant to nonsynonymous variation, suggesting a model of breed phenotype diversification based on regulatory changes to essential genes. Identification of trait-associated variants in dogs informs new biological roles for genes. Improved collection of breed disease risk data, along with increased breed representation, will drive further discoveries.

genomics↗

bioRxiv2021

Dog10K_Boxer_Tasha_1.0: A long-read assembly of the dog reference genome

The domestic dog has evolved to be an important biomedical model for studies regarding the genetic basis of disease, morphology and behavior. Genetic studies in the dog have relied on a draft reference genome of a purebred female boxer dog named "Tasha" initially published in 2005. Derived from a Sanger whole genome shotgun sequencing approach coupled with limited clone-based sequencing, the initial assembly and subsequent updates have served as the predominant resource for canine genetics for 15 years. While the initial assembly produced a good quality draft, as with all assemblies produced at the time it contained gaps, assembly errors and missing sequences, particularly in GC-rich regions, which are found at many promoters and in the first exons of protein coding genes. Here we present Dog10K_Boxer_Tasha_1.0, an improved chromosome-level highly contiguous genome assembly of Tasha created with long-read technologies, that increases sequence contiguity >100-fold, closes >23,000 gaps of the Canfam3.1 reference assembly and improves gene annotation by identifying >1200 new protein-coding transcripts. The assembly and annotation are available at NCBI under the accession GCF_000002285.5.

genomics↗

bioRxiv2021

Best practices for analyzing imputed genotypes from low-pass sequencing in dogs

Although DNA array-based approaches for genome wide association studies (GWAS) permit the collection of thousands of low-cost genotypes, it is often at the expense of resolution and completeness, as SNP chip technologies are ultimately limited by SNPs chosen during array development. An alternative low-cost approach is low-pass whole genome sequencing (WGS) followed by imputation. Rather than relying on high levels of genotype confidence at a set of select loci, low-pass WGS and imputation relies on the combined information from millions of randomly sampled low confidence genotypes. To investigate low-pass WGS and imputation in the dog, we assessed accuracy and performance by downsampling 97 high-coverage (>15x) WGS datasets from 51 different breeds to approximately 1x coverage, simulating low-pass WGS. Using a reference panel of 676 dogs from 91 breeds, genotypes were imputed from the downsampled data and compared to a truth set of genotypes generated from high coverage WGS. Using our truth set, we optimized a variant quality filtering strategy that retained approximately 80% of 14M imputed sites and lowered the imputation error rate from 3.0% to 1.5%. Seven million sites remained with a MAF > 5% and an average imputation quality score of 0.95. Finally, we simulated the impact of imputation errors on outcomes for case-control GWAS, where small effect sizes were most impacted and medium to large effect sizes were minorly impacted. These analyses provide best practice guidelines for study design and data post-processing of low-pass WGS imputed genotypes in dogs.

genomics↗

bioRxiv2020

A domestic cat whole exome sequencing resource for trait discovery

Over 94 million domestic cats are considered pets, who, as our companions, are also susceptible to cancers, common and rare diseases. Whole exome sequencing (WES) is a cost-effective strategy to study their putative disease-causing variants. Presented is ~35.8 Mb exome capture design based on the annotated Felis_catus_9.0 genome assembly, covering 201,683 regions of the cat genome. WES was conducted on 41 cats from various breeds with known and unknown diseases and traits, including 10 cats with prior whole genome sequence (WGS) data available, to test WES capture probe performance. A WES and WGS comparison was completed to understand variant discovery capability of each platform. At ~80x exome coverage, the percent of on-target base coverage >20x was 96.4% with an average of 10.4% off-target. For variant discovery, greater than 98% of WGS SNPs were also discovered by WES. Platform specific variants were mainly restricted to a small number of sex chromosome and olfactory receptor genes. Within the 41 cats with ~31 diseases and normal traits, 45 previously known disease or trait causal variants were observed, such as Persian progressive retinal degeneration and hydrocephalus. Novel candidate variants for polycystic kidney disease and atrichia in the Peterbald breed were also identified as well as a new cat patient with a known variant for cystinuria. These results show the discovery potential of deep exome sequencing to validate existing disease gene models and identify novel gene candidate alleles for many common and rare diseases in cats.

bioinformatics↗

bioRxiv2020

A deletion in GDF7 is associated with a heritable forebrain commissural malformation concurrent with ventriculomegaly and interhemispheric cysts in cats.

An inherited neurologic syndrome in a family of mixed-breed Oriental cats has been characterized as forebrain commissural malformation concurrent with ventriculomegaly and interhemispheric cysts. However, the genetic basis for this autosomal recessive syndrome in cats is unknown. Forty-three cats were genotyped on the Illumina Infinium Feline 63K iSelect DNA Array and used for analyses. Genome-wide association studies, including a sib-transmission disequilibrium test, a case-control association analysis, and homozygosity mapping, identified a critical region on cat chromosome A3. Short-read whole genome sequencing was completed for a cat trio segregating with the syndrome. A homozygous 7 bp deletion in growth differentiation factor 7 (GDF7) (c.221_227delGCCGCGC [p.Arg74Profs]) was identified in affected cats by comparison to the 99 Lives Cat variant dataset, validated using Sanger sequencing, and genotyped by fragment analyses. This variant was not identified in 192 unaffected cats in the 99 Lives dataset. The variant segregated concordantly in an extended pedigree. Obligate carrier cats were heterozygous. In mice, GDF7 mRNA is expressed within the roof plate when commissural axons initiate ventrally-directed growth. This finding emphasizes the importance of GDF7 in the neurodevelopmental process in the mammalian brain. A genetic test can be developed for use by cat breeders to eradicate this variant.

genetics↗

bioRxiv2020

Werewolf, there wolf: variants in Hairless associated with hypotrichia and roaning in the lykoi cat breed

A variety of cat breeds have been developed via novelty selection on aesthetic, dermatological traits, such as coat colors and fur types. A recently developed breed, the lykoi, was bred from cats with a sparse hair coat with roaning, implying full color and all white hairs. The lykoi phenotype is a form of hypotrichia, presenting as significant reduction in the average numbers of follicles per hair follicle group as compared to domestic shorthair cats, a mild to severe perifollicular to mural lymphocytic infiltration in 77% of observed hair follicle groups, and the follicles are often miniaturized, dilated, and dysplastic. Whole genome sequencing was conducted on a single lykoi cat that was a cross between two independently ascertained lineages. Comparison to the 99 Lives dataset of 194 non-lykoi cats suggested two variants in the cat homolog for Hairless (HR: lysine demethylase and nuclear receptor corepressor) as candidate causal variants. The lykoi cat was a compound heterozygote for two loss of function variants in HR, an exon 3 c.1255_1256dupGT (chrB1:36040783), which should produce a stop codon at amino acid 420 (p.Gln420Serfs*100) and, an exon 18 c.3389insGACA (chrB1:36051555), which should produce a stop codon at amino acid position 1130 (p.Ser1130Argfs*29). Ascertainment of 14 additional cats from founder lineages from Canada, France and different areas of the USA identified four additional loss of function HR variants likely causing the highly similar phenotypic hair coat across the diverse cats. The novel variants in HR for cat hypotrichia can now be established between minor differences in the phenotypic presentations.

genetics↗

bioRxiv2020

A new domestic cat genome assembly based on long sequence reads empowers feline genomic medicine and identifies a novel gene for dwarfism.

The domestic cat (Felis catus) numbers over 94 million in the USA alone, occupies households as a companion animal, and, like humans, suffers from cancer and common and rare diseases. However, genome-wide sequence variant information is limited for this species. To empower trait analyses, a new cat genome reference assembly was developed from PacBio long sequence reads that significantly improve sequence representation and assembly contiguity. The whole genome sequences of 54 domestic cats were aligned to the reference to identify single nucleotide variants (SNVs) and structural variants (SVs). Across all cats, 16 SNVs predicted to have deleterious impacts and in a singleton state were identified as high priority candidates for causative mutations. One candidate was a stop gain in the tumor suppressor FBXW7. The SNV is found in cats segregating for feline mediastinal lymphoma and is a candidate for inherited cancer susceptibility. SV analysis revealed a complex deletion coupled with a nearby potential duplication event that was shared privately across three unrelated dwarfism cats and is found within a known dwarfism associated region on cat chromosome B1. This SV interrupted UDP-glucose 6-dehydrogenase (UGDH), a gene involved in the biosynthesis of glycosaminoglycans. Importantly, UGDH has not yet been associated with human dwarfism and should be screened in undiagnosed patients. The new high-quality cat genome reference and the compilation of sequence variation demonstrate the importance of these resources when searching for disease causative alleles in the domestic cat and for identification of feline biomedical models. Author summaryThe practice of genomic medicine is predicated on the availability of a high quality reference genome and an understanding of the impact of genome variation. Such resources have lead to countless discoveries in humans, however by working exclusively within the framework of human genetics, our potential for understanding diseases biology is limited, as similar analyses in other species have often lead to novel insights. The generation of Felis_catus_9.0, a new high quality reference genome for the domestic cat, helps facilitate the expansion of genomic medicine into the felis lineage. Using Felis_catus_9.0 we analyze the landscape of genomic variation from a collection of 54 cats within the context of human gene constraint. The distribution of variant impacts in cats is correlated with patterns of gene constraint in humans, indicating the utility of this reference for identifying novel mutations that cause phenotypes relevant to human and cat health. Moreover, structural variant analysis revealed a novel variant for feline dwarfism in UGDH, a gene that has not been associated with dwarfism in any other species, suggesting a role for UGDH in cases of undiagnosed dwarfism in humans.

genomics↗

bioRxiv2019

Localization of a feline autosomal dominant dwarfism locus: a novel model of chondrodysplasia

Despite the contribution of a few major genes for disproportionate dwarfism in humans, many dwarf patients are yet genetically undiagnosed. In domestic cats, disproportionate dwarfism has led to the development of a defined breed, the Munchkin or Minuet. This study examined the genetic aspects of feline dwarfism to consider cats as a new biomedical model. DNA from dwarf cats was genetically analyzed using parentage, linkage, and genome-wide association studies as well as whole genome sequencing. Each genetic approach localized the dwarfism phenotype to a region on cat chromosome B1. No coding variants suspected as causal for the feline dwarfism were identified but a critical region of [~]5.7 Mb from B1:170,278,183-175,975,857 was defined, which implicates a novel gene controlling disproportionate dwarfism. A yet unidentified but novel gene variant, likely structural or regulatory, produces disproportionate dwarfism in cats, which may define undiagnosed human patients.

genetics↗

bioRxiv2017

Divergent genome evolution caused by regional variation in DNA gain and loss between human and mouse

The forces driving the accumulation and removal of non-coding DNA and ultimately the evolution of genome size in complex organisms are intimately linked to genome structure and organisation. Our analysis provides a novel method for capturing the regional variation of lineage-specific DNA gain and loss events in their respective genomic contexts. To further understand this connection we used comparative genomics to identify genome-wide individual DNA gain and loss events in the human and mouse genomes. Focusing on the distribution of DNA gains and losses, relationships to important structural features and potential impact on biological processes, we found that in autosomes, DNA gains and losses both followed separate lineage-specific accumulation patterns. However, in both species chromosome X was particularly enriched for DNA gain, consistent with its high L1 retrotransposon content required for X inactivation. We found that DNA loss was associated with gene-rich open chromatin regions and DNA gain events with gene-poor closed chromatin regions. Additionally, we found that DNA loss events tended to be smaller than DNA gain events suggesting that they were more tolerated in open chromatin regions. GO term enrichment in human gain hotspots showed terms related to cell cycle/metabolism, human loss hotspots were enriched for terms related to gene silencing, and mouse gain hotspots were enriched for terms related to transcription regulation. Interestingly, mouse loss hotspots were strongly enriched for terms related to developmental processes, suggesting that DNA loss in mouse is associated with phenotypic changes in mouse morphology. This is consistent with a model in which DNA gain and loss results in turnover or \"churning\" of regulatory regions that are then subjected to selection, resulting in the differences we now observe, both genomic and phenotypic/morphological.

evolutionary biology↗

bioRxiv2016

Similar evolutionary trajectories for retrotransposon accumulation in mammals

The factors guiding retrotransposon insertion site preference are not well understood. Different types of retrotransposons share common replication machinery and yet occupy distinct genomic domains. Autonomous long interspersed elements accumulate in gene-poor domains and their non-autonomous short interspersed elements accumulate in gene-rich domains. To determine genomic factors that contribute to this discrepancy we analysed the distribution of retrotransposons within the framework of chromosomal domains and regulatory elements. Using comparative genomics, we identified large-scale conserved patterns of retrotransposon accumulation across several mammalian genomes. Importantly, retrotransposons that were active after our sample-species diverged accumulated in orthologous regions. This suggested a similar evolutionary interaction between retrotransposon activity and conserved genome architecture across our species. In addition, we found that retrotransposons accumulated at regulatory element boundaries in open chromatin, where accumulation of particular retrotransposon types depended on insertion size and local regulatory element density. From our results, we propose a model where density and distribution of genes and regulatory elements canalise retrotransposon accumulation. Through conservation of synteny, gene regulation and nuclear organisation, mammalian genomes with dissimilar retrotransposons follow similar evolutionary trajectories.

evolutionary biology↗