Mutations in MITF and PAX3 Cause “Splashed White” and Other White Spotting Phenotypes in Horses

Abstract

During fetal development neural-crest-derived melanoblasts migrate across the entire body surface and differentiate into melanocytes, the pigment-producing cells. Alterations in this precisely regulated process can lead to white spotting patterns. White spotting patterns in horses are a complex trait with a large phenotypic variance ranging from minimal white markings up to completely white horses. The “splashed white” pattern is primarily characterized by an extremely large blaze, often accompanied by extended white markings at the distal limbs and blue eyes. Some, but not all, splashed white horses are deaf. We analyzed a Quarter Horse family segregating for the splashed white coat color. Genome-wide linkage analysis in 31 horses gave a positive LOD score of 1.6 in a region on chromosome 6 containing the PAX3 gene. However, the linkage data were not in agreement with a monogenic inheritance of a single fully penetrant mutation. We sequenced the PAX3 gene and identified a missense mutation in some, but not all, splashed white Quarter Horses. Genome-wide association analysis indicated a potential second signal near MITF. We therefore sequenced the MITF gene and found a 10 bp insertion in the melanocyte-specific promoter. The MITF promoter variant was present in some splashed white Quarter Horses from the studied family, but also in splashed white horses from other horse breeds. Finally, we identified two additional non-synonymous mutations in the MITF gene in unrelated horses with white spotting phenotypes. Thus, several independent mutations in MITF and PAX3 together with known variants in the EDNRB and KIT genes explain a large proportion of horses with the more extreme white spotting phenotypes.

Author Summary

White spotting coat color phenotypes are the result of aberrations in the development of melanocytes. The analysis of domestic animals with heritable white spotting phenotypes thus helps to better understand the complicated genetic network controlling the proliferation, migration, differentiation, and survival of pigment producing cells. We analyzed the so-called splashed white phenotype in horses, which is characterized by a very distinctive large blaze, extended white markings on the legs, and blue eyes. Splashed white horses are also frequently deaf. However, the phenotype is quite variable and, in some horses with minimal expression, the splashed white phenotype cannot be unambiguously discriminated from the “common” white markings. We studied horses from various breeds and found one mutation in the PAX3 gene and two mutations in the MITF gene that cause the splashed white phenotype. A third mutation in the MITF gene, which we found in a single Franches-Montagnes horse, causes a new coat color phenotype, termed macchiato. Similar mutations in humans cause either Waardenburg or Tietz syndrome, which both are characterized by pigmentation defects and a predisposition for deafness. Our study reveals the molecular basis for a significant proportion of white spotting phenotypes that are intermediate between completely unpigmented horses and common white markings.