A missense variant in the coil1A domain of the keratin 25 gene is associated with the dominant curly hair coat trait (Crd) in horse

Abstract

Background: Curly horses present a variety of curl phenotypes that are associated with various degrees of curliness of coat, mane, tail and ear hairs. Their origin is still a matter of debate and several genetic hypotheses have been formulated to explain the diversity in phenotype, including the combination of autosomal dominant and recessive alleles. Our purpose was to map the autosomal dominant curly hair locus and identify the causal variant using genome-wide association study (GWAS) and whole-genome sequencing approaches.

Results: A GWAS was performed using a Bayesian sparse linear mixed model, based on 51 curly and 19 straight-haired French and North American horses from 13 paternal families genotyped on the Illumina EquineSNP50 BeadChip. A single strong signal was observed on equine chromosome 11, in a region that encompasses the type I keratin gene cluster. This region was refined by haplotype analysis to a segment including 36 genes, among which are 10 keratin genes (KRT-10, -12, -20, -23, -24, -25, -26, -27, -28, -222). To comprehensively identify candidate causal variants within all these genes, whole-genome sequences were obtained for one heterozygous curly stallion and its straight-haired son. Among the four non-synonymous candidate variants identified and validated in the curly region, only variant g.21891160G>A in the KRT25 gene (KRT25:p.R89H) was in perfect agreement with haplotype status in the whole pedigree. Genetic association was then confirmed by genotyping a larger population consisting of 353 horses. However, five discordant curly horses were observed, which carried neither the variant nor the main haplotype associated with curliness. Sequencing of KRT25 for two discordant horses did not identify any other deleterious variant, which suggests locus rather than allelic heterogeneity for the curly phenotype.

Conclusions: We identified the KRT25:p.R89H variant as responsible for the dominant curly trait, but a second dominant locus may also be involved in the shape of hairs within North American Curly horses.

Fig. 1

Phenotype of curly horses. Curly horses present curly haired coat (a) and ear (b). Tail (c) and mane (d) present phenotypes ranging from curly to wavy hairs and can resemble dreadlocks. The coat phenotype is associated with various degrees of curliness, ranging from slightly curled (e) and curled (f) up to extremely dense “micro curled” (g) hairs. Pictures were from the ICHO association with permission of Dr. Mitch Wilkinson and Bunny Reveglia (see http://www.ichocurlyhorses.com/ and http://ichophotos.weebly.com)

Fig. 2

Plot of posterior inclusion probabilities showing a strong signal on ECA11. A genome-wide association study based on 46,215 SNPs was performed using a Bayesian sparse linear mixed model (BSLMM). Cumulative inclusion probabilities were computed for sliding windows of 15 SNPs and were plotted along the genome, taking the central SNP as reference. These data represent the probability for a region to have an effect above the polygenic background. A single strong signal encompassing the type I keratin gene cluster was observed on ECA11, with seven windows showing inclusion probabilities above 75% and an additional weak signal on ECA30 was also detected (four windows with inclusion probabilities under 20%)

Fig. 3

Fine-mapping of the Crd locus in horse and whole-genome sequencing identify a candidate variant in the coil1A domain of KRT25 that affects an amino-acid residue conserved among placental mammals. a The IBD segment delineated by haplotype analysis comprises 36 genes, including 10 genes encoding keratin proteins (KRT-10, -12, -20, -23, -24, -25, -26, -27, -28, -222). b Multispecies alignment of KRT25 orthologues shows a perfect conservation of amino acid R89. The p.R89H variant affects the coil1A domain within the α-helical rod domain of KRT25 protein. Ensembl accession numbers for the wild type transcript of each species are ENSTBEP00000001955, ENSETEP00000006824, ENSEEUP00000013767, ENSMUSP00000048439, ENSCAFP00000032235, ENSSSCP00000018507, ENSP00000310573, ENSBTAP00000040707, ENSECAP00000011587, ENSPVAP00000010713, ENSLAFP00000005738 and ENSOCUP00000007766 in order of appearance

Fig. 4

Chromatograms showing the g.21891160G>A variant within the KRT25 gene on ECA11