Variant in the RFWD3 Gene Associated with PATN1, a Modifier of Leopard Complex Spotting

Leopard Complex spotting (LP), the result of an incompletely dominant mutation in TRPM1, produces a collection of unique pigmentation patterns in the domestic horse. While the LP mutation allows for expression of the various patterns, other loci are responsible for modification of the extent of white. Pedigree analysis of families segregating for high levels of patterning (80-100% white hair coat) indicated a single dominant gene (PATN1) as a major effect modifier for LP. Linkage analysis identified a 16 Mbp region on ECA3p associated with PATN1. Whole transcriptome sequencing of skin samples from horses with and without the PATN1 allele was performed to identify genic SNPs for fine mapping. Two Sequenom assays were utilized to genotype 192 individuals from five LP-containing breeds. The initial panel highlighted a 1.6 Mbp region without a clear candidate gene. In the second round of fine-mapping, a SNP in the 3' UTR of RING finger and WD repeat domain 3 (RFWD3) reached a significance level of p=1.063x10-39. Sequencing of RFWD3 did not identify any coding polymorphisms specific to the PATN1 horses. Genotyping of an additional 52 LP animals increased this association to a p-value of 4.68x10-56. An additional 166 horses of breeds not segregating for LP did not contain the PATN1-associated allele. This variant is a strong candidate for PATN1 and may be used for genotyping lp/lp animals.

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