Identification of novel genetic risk factors of dilated cardiomyopathy: from canine to human.

BACKGROUND: Dilated cardiomyopathy (DCM) is a life-threatening heart disease and a common cause of heart failure due to systolic dysfunction and subsequent left or biventricular dilatation. A significant number of cases have a genetic etiology; however, as a complex disease, the exact genetic risk factors are largely unknown, and many patients remain without a molecular diagnosis. METHODS: We performed GWAS followed by whole-genome, transcriptome, and immunohistochemical analyses in a spontaneously occurring canine model of DCM. Canine gene discovery was followed up in three human DCM cohorts. RESULTS: Our results revealed two independent additive loci associated with the typical DCM phenotype comprising left ventricular systolic dysfunction and dilatation. We highlight two novel candidate genes, RNF207 and PRKAA2, known for their involvement in cardiac action potentials, energy homeostasis, and morphology. We further illustrate the distinct genetic etiologies underlying the typical DCM phenotype and ventricular premature contractions. Finally, we followed up on the canine discoveries in human DCM patients and discovered candidate variants in our two novel genes. CONCLUSIONS: Collectively, our study yields insight into the molecular pathophysiology of DCM and provides a large animal model for preclinical studies.

Genome sequencing of 2000 canids by the Dog10K consortium advances the understanding of demography, genome function and architecture.

BACKGROUND: The international Dog10K project aims to sequence and analyze several thousand canine genomes. Incorporating 20 × data from 1987 individuals, including 1611 dogs (321 breeds), 309 village dogs, 63 wolves, and four coyotes, we identify genomic variation across the canid family, setting the stage for detailed studies of domestication, behavior, morphology, disease susceptibility, and genome architecture and function. RESULTS: We report the analysis of > 48 M single-nucleotide, indel, and structural variants spanning the autosomes, X chromosome, and mitochondria. We discover more than 75% of variation for 239 sampled breeds. Allele sharing analysis indicates that 94.9% of breeds form monophyletic clusters and 25 major clades. German Shepherd Dogs and related breeds show the highest allele sharing with independent breeds from multiple clades. On average, each breed dog differs from the UU_Cfam_GSD_1.0 reference at 26,960 deletions and 14,034 insertions greater than 50 bp, with wolves having 14% more variants. Discovered variants include retrogene insertions from 926 parent genes. To aid functional prioritization, single-nucleotide variants were annotated with SnpEff and Zoonomia phyloP constraint scores. Constrained positions were negatively correlated with allele frequency. Finally, the utility of the Dog10K data as an imputation reference panel is assessed, generating high-confidence calls across varied genotyping platform densities including for breeds not included in the Dog10K collection. CONCLUSIONS: We have developed a dense dataset of 1987 sequenced canids that reveals patterns of allele sharing, identifies likely functional variants, informs breed structure, and enables accurate imputation. Dog10K data are publicly available.

Breed, age, and social environment are associated with personality traits in dogs.

Personality in pets and other domesticated animals is important for their well-being and it can also influence human-animal relationships. Genetic and environmental factors influencing unwanted behavior in dogs are somewhat well known, but the factors influencing dog personality remain understudied. Here we examined environmental and demographic factors associated with seven broad personality traits in a survey of over 11,000 dogs. We utilized linear models and extensive model validation to examine the factors that have the most significant influences on personality and calculated effect sizes to assess the importance of these variables. Breed and age had the strongest associations with dog personality traits. Some environmental factors, especially puppyhood socialization, were also associated with personality. All factors had small effect sizes, highlighting that a lot of variation in personality remains unexplained. Our results indicate that personality traits are complex and strikingly similar in dogs, humans, and other nonhuman animals.

Age, breed, sex and diet influence serum metabolite profiles of 2000 pet dogs.

As an individual's metabolism reflects health and disease states well, metabolomics holds a vast potential in biomedical applications. However, normal physiological factors, such as age, can also influence metabolism, challenging the establishment of disease-specific metabolic aberrations. Here, we examined how physiological and diet-related factors drive variance in the metabolism of healthy pet dogs. We analysed 2068 serum samples using a canine nuclear magnetic resonance (NMR) spectroscopy-based metabolomics platform. With generalized linear models, we discovered that age, breed, sex, sterilization, diet type and fasting time significantly affected the canine metabolite profiles. Especially, breed and age caused considerable variation in the metabolite concentrations, and breeds with very different body conformations systematically differed in several lipid measurands. Our results enhance the understanding how normal physiological factors influence canine metabolism, aid accurate interpretation of the NMR results, and suggest the NMR platform might be applied in identifying aberrations in nutrient absorption and metabolism.

Missense variant in LOXHD1 is associated with canine nonsyndromic hearing loss.

Hearing loss is a common sensory deficit in both humans and dogs. In canines, the genetic basis is largely unknown, as genetic variants have only been identified for a syndromic form of hearing impairment. We observed a congenital or early-onset sensorineural hearing loss in a Rottweiler litter. Assuming an autosomal recessive inheritance, we used a combined approach of homozygosity mapping and genome sequencing to dissect the genetic background of the disorder. We identified a fully segregating missense variant in LOXHD1, a gene that is known to be essential for cochlear hair cell function and associated with nonsyndromic hearing loss in humans and mice. The canine LOXHD1 variant was specific to the Rottweiler breed in our study cohorts of pure-bred dogs. However, it also was present in some mixed-breed dogs, of which the majority showed Rottweiler ancestry. Low allele frequencies in these populations, 2.6% and 0.04%, indicate a rare variant. To summarize, our study describes the first genetic variant for canine nonsyndromic hearing loss, which is clinically and genetically similar to human LOXHD1-related hearing disorder, and therefore, provides a new large animal model for hearing loss. Equally important, the affected breed will benefit from a genetic test to eradicate this LOXHD1-related hearing disorder from the population.

Intronic variant in POU1F1 associated with canine pituitary dwarfism.

The anterior pituitary gland secretes several endocrine hormones, essential for growth, reproduction and other basic physiological functions. Abnormal development or function of the pituitary gland leads to isolated or combined pituitary hormone deficiency (CPHD). At least 30 genes have been associated with human CPHD, including many transcription factors, such as POU1F1. CPHD occurs spontaneously also in mice and dogs. Two affected breeds have been reported in dogs: German Shepherds with a splice defect in the LHX3 gene and Karelian Bear Dogs (KBD) with an unknown genetic cause. We obtained samples from five KBDs presenting dwarfism and abnormal coats. A combined analysis of genome-wide association and next-generation sequencing mapped the disease to a region in chromosome 31 and identified a homozygous intronic variant in the fourth exon of the POU1F1 gene in the affected dogs. The identified variant, c.605-3C>A, resided in the splice region and was predicted to affect splicing. The variant's screening in three new prospective cases, related breeds, and ~ 8000 dogs from 207 breeds indicated complete segregation in KBDs with a carrier frequency of 8%, and high breed-specificity as carriers were found at a low frequency only in Lapponian Herders, a related breed. Our study establishes a novel canine model for CPHD with a candidate POU1F1 defect.

Canine DVL2 variant contributes to brachycephalic phenotype and caudal vertebral anomalies.

A frameshift deletion variant in the Wnt pathway gene dishevelled 2 (DVL2) is associated with a truncated, kinked tail ("screw tail") in English Bulldogs, French Bulldogs and Boston Terriers. These breeds are also characterized by distinctive morphological traits, including a wide head, flat face and short-limbed dwarfism, which are characteristic of Robinow syndrome in humans, caused by defects in genes such as DVL1 and DVL3. Based on these phenotypic and genetic similarities, it has previously been hypothesized that the canine DVL2 variant results in a syndromic phenotype called the Robinow-like syndrome. In our study, we investigated the distribution of the DVL2 variant in 1954 dogs from 15 breeds, identifying breeds with allele variation and enabling the dissection of the genotype-phenotype correlation for the first time. With CT examinations in American Staffordshire Terriers, we confirmed that the DVL2 allele is associated with caudal vertebral malformations and a brachycephalic phenotype. We also hypothesize that the variant may be linked to additional health conditions, including brachycephalic obstructive airway syndrome and congenital heart defects. Altogether, our study strengthens the role of DVL2 as one of the contributors to the "bulldog type" morphology and features on the spectrum of human Robinow syndrome.

Recessive missense LAMP3 variant associated with defect in lamellar body biogenesis and fatal neonatal interstitial lung disease in dogs.

Neonatal interstitial lung diseases due to abnormal surfactant biogenesis are rare in humans and have never been reported as a spontaneous disorder in animals. We describe here a novel lung disorder in Airedale Terrier (AT) dogs with clinical symptoms and pathology similar to the most severe neonatal forms of human surfactant deficiency. Lethal hypoxic respiratory distress and failure occurred within the first days or weeks of life in the affected puppies. Transmission electron microscopy of the affected lungs revealed maturation arrest in the formation of lamellar bodies (LBs) in the alveolar epithelial type II (AECII) cells. The secretory organelles were small and contained fewer lamellae, often in combination with small vesicles surrounded by an occasionally disrupted common limiting membrane. A combined approach of genome-wide association study and whole exome sequencing identified a recessive variant, c.1159G>A, p.(E387K), in LAMP3, a limiting membrane protein of the cytoplasmic surfactant organelles in AECII cells. The substitution resides in the LAMP domain adjacent to a conserved disulfide bond. In summary, this study describes a novel interstitial lung disease in dogs, identifies a new candidate gene for human surfactant dysfunction and brings important insights into the essential role of LAMP3 in the process of the LB formation.