Studying the characteristics of mammalian hoof colors is important for genetic improvements in animals. A deeper black hoof color is the standard for breeding purebred Australian White (AUW) sheep and this phenotype could be used as a phenotypic marker of purebred animals. We conducted a genome-wide association study (GWAS) analysis using restriction site associated DNA sequencing (RAD-seq) data from 577 Australian White sheep (black hoof color = 283, grey hoof color = 106, amber hoof color = 186) and performed association analysis utilizing the mixed linear model in EMMAX. The results of GWAS demonstrated that a specific single-nucleotide polymorphism (SNP; g. 33097911G>A) in intron 14 of the microphthalmia-associated transcription factor (MITF) gene was significantly associated with the hoof color in AUW sheep (p = 9.40 × 10-36). The MITF gene plays a key role in the development, differentiation, and functional regulation of melanocytes. Furthermore, the association between this locus and hoof color was validated in a cohort of 212 individuals (black hoof color = 122, grey hoof color = 38, amber hoof color = 52). The results indicated that the hoof color of AUW sheep with GG, AG, and AA genotypes tended to be black, grey, and amber, respectively. This study provided novel insights into hoof color genetics in AUW sheep, enhancing our comprehension of the genetic mechanisms underlying the diverse range of hoof colors. Our results agree with previous studies and provide molecular markers for marker-assisted selection for hoof color in sheep.

Plumage color is an artificially and naturally selected trait in domestic ducks. Black, white, and spotty are the main feather colors in domestic ducks. Previous studies have shown that black plumage color is caused by MC1R, and white plumage color is caused by MITF. We performed a genome-wide association study (GWAS) to identify candidate genes associated with white, black, and spotty plumage in ducks. Two non-synonymous SNPs in MC1R (c.52G>A and c.376G>A) were significantly related to duck black plumage, and three SNPs in MITF (chr13:15411658A>G, chr13:15412570T>C and chr13:15412592C>G) were associated with white plumage. Additionally, we also identified the epistatic interactions between causing loci. Some ducks with white plumage carry the c.52G>A and c.376G>A in MC1R, which also compensated for black and spotty plumage color phenotypes, suggesting that MC1R and MITF have an epistatic effect. The MITF locus was supposed to be an upstream gene to MC1R underlying the white, black, and spotty colors. Although the specific mechanism remains to be further clarified, these findings support the importance of epistasis in plumage color variation in ducks.

1. Microphthalmia-associated transcription factor (MITF) plays a pivotal role in melanocyte development by regulating the transcription of major pigmentation enzymes (e.g. TYR, TYRP1 and DCT). A single-nucleotide polymorphism (SNP), c.-638T>C, was identified in the MITF promoter, and genotyping of a population (n = 426) revealed that SNP c.-638T>C was associated with skin colour in black-boned chickens. 2. Individuals with genotypes CC and TC exhibited greater MTIF expression than those with genotype TT. Luciferase assays also revealed that genotype CC and TC promoters had higher activity levels than genotype TT. Expression of melanogenesis-related gene (TYR) was higher in the skin of chickens with the CC and CT genotype compared to TT chickens (P < 0.05). 3. Transcription factor-binding site analyses showed that the c.-638C allele contains a putative binding site for transcription factor sterol regulatory element-binding transcription factor 2, aryl hydrocarbon receptor nuclear translocator, transcription factor binding to IGHM enhancer 3 and upstream transcription factor 2. In contrast, the c.-638T allele contains binding sites for Sp3 transcription factor and Krüppel-like factor 1. 4. It was concluded that MITF promoter polymorphisms affected chicken skin colour. SNP c.-638T>C could be used for the marker-assisted selection of skin colour in black-boned chicken breeding.

Waardenburg syndrome (WS) is an autosomal dominant disorder with varying degrees of sensorineural hearing loss, and accumulation of pigmentation in hair, skin and iris. There are four types of WS (WS1-4) with differing characteristics. Mutations in six genes [paired box gene 3 (PAX3), microphthalmia-associated transcription factor (MITF), endothelin 3 (END3), endothelin receptor type B (EDNRB), SRY (sex determining region Y)-box 10 (SOX10) and snail homolog 2 (SNAI2)] have been identified to be associated with the various types. This case report describes the investigation of genetic mutations in three patients with WS2 from a single family. Genomic DNA was extracted, and the six WS-related genes were sequenced using next-generation sequencing technology. In addition to mutations in PAX3, EDNRB and SOX10, a novel heterozygous MITF mutation, p.Δ315Arg (c.944_946delGAA) on exon 8 was identified. This is predicted to be a candidate disease-causing mutation that may affect the structure and function of the enzyme.