Use of genomic tools to discover the cause of champagne dilution coat color in horses and to map the genetic cause of extreme lordosis in American Saddlebred horses

Champagne dilution of coat color in horses is caused by dominant gene action. Three sire families were identified as segregating for this trait. Genome wide linkage analysis using 104 microsatellite DNA markers was used to map the gene to ECA14 (LOD > 11.0). Four genes, namely SPARC, SLC36A1, SLC36A2 and SLC36A3, were selected from the region implicated by linkage and their exons sequenced to identify genetic variation. DNA sequences were compared for two homozygotes for Champagne dilution, two heterozygotes and two horses without dilution. A single base change in exon 2 of SLC36A1 was found unique to horses exhibiting Champagne dilution. SLC36A1 encodes a eukaryote specific proton dependent small amino acid transporter also known as LYAAT-1 in rats. This change in base 188 of the cDNA from a C to a G (Genbank REFSEQ: Non-champagne EU432176 and Champagne EU432177), is predicted to cause an amino acid change from threonine to arginine in the first transmembrane region of the protein. This may disrupt placement of the transmembrane portion of the protein in the membrane interrupting function of the ion channel. No exceptions to the association of this mutation with the Champagne dilution (CH) allele were identified based on testing 182 additional horses representing 15 breeds, suggesting that this mutation may be the causative for the dilution phenotype. While this gene is expected to function as a proton/amino acid symporter based on its sequence and gene family, this observation for the horse is the first evidence for a phenotypic effect of mutation of the gene. Identifying this variant also gives breeders a new tool for selecting breeding stock with or without this dilution genotype.;Extreme lordosis is a condition in which the dorsal to ventral curvature of the back is accentuated. The condition is considered a conformation fault by many Saddlebred breeders and is commonly referred to as swayback, softback or lowback. Previous studies suggested the existence of a hereditary component in American Saddlebred horses. A whole genome association study was performed utilizing the Illumina SNP50 beadchip containing over 59,000 SNPs. A 3 Mb region on ECA20 was found associated with extreme lordosis. The distribution of markers indicated that the trait had a simple Mendelian recessive mode of inheritance. Subsequently, the association was confirmed and the region was narrowed to just over 500 kb by a higher density SNP assay using 47 SNPs selected from the equine SNP database. Identification of this associated region will allow future studies a better opportunity to locate the mutation responsible for the lordotic phenotype in young American Saddlebred Horses.;KEYWORDS: horse, champagne, lordosis, swayback, mapping.