A single IGF1 allele is a major determinant of small size in dogs

Abstract

The domestic dog exhibits greater diversity in body size than any other terrestrial vertebrate. We used a strategy that exploits the breed structure of dogs to investigate the genetic basis of size. First, through a genome-wide scan, we identified a major quantitative trait locus (QTL) on chromosome 15 influencing size variation within a single breed. Second, we examined genetic variation in the 15-megabase interval surrounding the QTL in small and giant breeds and found marked evidence for a selective sweep spanning a single gene (IGF1), encoding insulin-like growth factor 1. A single IGF1 single-nucleotide polymorphism haplotype is common to all small breeds and nearly absent from giant breeds, suggesting that the same causal sequence variant is a major contributor to body size in all small dogs.

Fig. 1

Relationships of skeletal size, SNP markers, IGF1 haplotype, and serum levels of the IGF1 protein in PWDs. (A) A mixed-model test for association between size and genotype. The association of three genotype categories (A₁A₁, A₁A₂, and A₂A₂) with skeletal size measurements was calculated with the use of all pairwise coefficients of consanguinity for 376 dogs. Each point represents a single SNP position on canine chromosome 15 and negative log P value for the association statistic. (B) PWD IGF1 haplotypes and mean skeletal size. Haplotypes were inferred for 20 markers spanning the IGF1 gene (chromosome 15: 44,212,792 to 44,278,140, CanFam1). Out of the 720 chromosomes with successful inference, 96% carry one of just two haplotypes, B and I, identical to haplotypes inferred for small and giant dogs, respectively (Fig. 3). Data are graphed as a histogram for each genotype: B/B (closed triangle, black line), B/I (open square, dashed line), and I/I (closed circle, gray line). (C) Serum levels of IGF1 protein (ng/ml) as a function of haplotype. Serum levels of IGF1 protein were assayed in 31 PWDs carrying haplotypes B and I. Box plots show the median (center line in box), first and third quartile (box ends), and maximum and minimum values (whiskers) obtained for each category: homozygous B/B (n = 15), heterozygous B/I (n = 7), and homozygous I/I (n = 9).

Fig. 2

Signatures of recent selection on the IGF1 locus across 22 small and giant dog breeds. (A) Heterozygosity ratio (H_R) for small versus giant dogs. (B) Genetic differentiation (F_ST) for small versus giant dogs. For both (A) and (B), a sliding 10-SNP window across IGF1 was used. Dashed lines delimit the 95% confidence intervals based on nonparametric bootstrap resampling. The IGF1 gene interval is indicated above the graphs as a red box drawn to scale. (C) Observed heterozygosity (H_Obs) of SNPs near IGF1 typed in small breeds (<9 kg) and giant breeds (>30 kg). Small breeds have a reduction in observed heterozygosity compared with that of giant breeds. Red and blue points are average observed heterozygosity in small and giant breeds, respectively. Dashed lines are locally weighted scatterplot smoothing (LOWESS) best fit to the data. The IGF1 gene is shown as a black bar with exons indicated by vertical lines.

Fig. 3

Evidence of association and IGF1 haplotypes for 14 small and 9 giant breeds. (A) Mann-Whitney U (MWU) P values for tests of association between individual SNPs and body size (small versus giant) for 116 SNPs on chromosome 15 and 83 SNPs on five control chromosomes. The dashed line indicates Bonferroni correction for multiple tests. Only breeds with data for at least 10 chromosomes were included (14 small and 9 giant breeds). (B) Haplotypes for the 20 markers spanning the small breed sweep interval near IGF1. The haplotypes were inferred independently in each breed. For each individual, fractional chromosome counts were summed for all haplotypes with at least 5% probability according to the haplotype inference software program PHASE. Chromosome sums for each breed were rounded to integer values; several breeds have odd numbers of chromosomes due to rounding error. Only inferred haplotypes carried by at least three dog chromosomes total (i.e., >0.5% frequency overall) are shown. Sequence reads collected from golden jackal (Canis aureus) were used to determine the ancestral allele for each SNP. The haplotypes are rows labeled A to L, and marker alleles are colored yellow for ancestral state (matching the nucleotide observed in the golden jackal) and blue for derived state. SNP positions within IGF1 are shown at the top with IGF1 introns (horizontal line) and exons (vertical bars) indicated. (C) Breed name and the average size of adult males in kilograms are provided. Small breeds less than 9 kg and giant breeds greater than 30 kg are grouped for totals shown at the far right.

Fig. 4

Association of body size and frequency of the SNP 5 A allele. Binomial regression of allele frequency on square root of mean breed mass. Dashed lines indicate the 95% confidence interval on the predicted equation line as estimated from nonparametric bootstrap resampling. Between 5 and 109 (median = 22) dogs were genotyped for each of 143 breeds. The PWD is highlighted in red along with three giant breeds that have larger breed average masses than is predicted by their SNP 5 allele frequency.