STUDIES OF GENETIC VARIATION AT THE KIT LOCUS AND WHITE SPOTTING PATTERNS IN THE HORSE

There are numerous different white spotting patterns in the horse, including two of particular interest tobiano and sabino. In the mouse, genetic variation in the gene KIT causes many white spotting patterns. Due to the phenotypic similarity among white spotting patterns in horses and mice, KIT was investigated as the cause of the tobiano and sabino spotting patterns in horses. Initially, the KIT cDNA sequences from horses with several spotting patterns were compared. Three single nucleotide polymorphisms (SNPs) were identified, though none were associated with a spotting pattern. Three novel splicing variants were also observed: exon 17 skipping, exon 18 skipping and alternative splicing of exon 3. Families segregating for a sabino spotting pattern (designated Sabino 1) and exon 17 skipping were discovered. Sequencing revealed a SNP (KI16+1037) within intron 16 that was completely associated with skipping of exon 17. Using a PCR-RFLP for KI16+1037, linkage was discovered for sabino spotting (LOD=9.02 for =0) and presence of the Sabino 1 allele detected in seven breeds. While all horses with this SNP exhibited the Sabino 1 phenotype, some horses with a sabino phenotype did not possess the SNP. This is most likely due to genetic heterogeneity of the phenotype. Fluorescent in situ hybridization (FISH) was used to investigate the possibility of chromosome inversion in the region of KIT. A chromosomal inversion was discovered spanning ECA3q13 to 3q21 using BAC clones containing KIT and other genes in the same region. The ECA3q inversion was completely associated with Tobiano in the eight horses tested by FISH. This inversion may disrupt regulatory sequences of the KIT gene and thereby cause tobiano spotting. Spotting patterns are important to horse breeders for aesthetic as well as economic reasons. Spotting patterns in the horse may also be an interesting scientific model. The two genetic variants discovered in this work are good examples of genetic diversity due to mechanisms other than SNPs. Study of these variants may be valuable for examining the effects of the KIT gene on health traits. In particular, the KIT gene directs many functions of the mast cell, a cell that is involved in the etiology of inflammation