Copy Number Variation and Missense Mutations of the Agouti Signaling Protein (<i>ASIP)</i> Gene in Goat Breeds with Different Coat Colors

Abstract

In goats, classical genetic studies reported a large number of alleles at the Agouti locus with effects on coat color and pattern distribution. From these early studies, the dominant A^Wt (white/tan) allele was suggested to cause the white color of the Saanen breed. Here, we sequenced the coding region of the goat ASIP gene in 6 goat breeds (Girgentana, Maltese, Derivata di Siria, Murciano-Granadina, Camosciata delle Alpi, and Saanen), with different coat colors and patterns. Five single nucleotide polymorphisms (SNPs) were identified, 3 of which caused missense mutations in conserved positions of the cysteine-rich carboxy-terminal domain of the protein (p.Ala96Gly, p.Cys126Gly, and p.Val128Gly). Allele and genotype frequencies suggested that these mutations are not associated or not completely associated with coat color in the investigated goat breeds. Moreover, genotyping and sequencing results, deviation from Hardy-Weinberg equilibrium, as well as allele copy number evaluation from semiquantitative fluorescent multiplex PCR, indicated the presence of copy number variation (CNV) in all investigated breeds. To confirm the presence of CNV and evaluate its extension, we applied a bovine-goat cross-species array comparative genome hybridization (aCGH) experiment using a custom tiling array based on bovine chromosome 13. aCGH results obtained for 8 goat DNA samples confirmed the presence of CNV affecting a region of less that 100 kb including the ASIP and AHCY genes. In Girgentana and Saanen breeds, this CNV might cause the A^Wt allele, as already suggested for a similar structural mutation in sheep affecting the ASIP and AHCY genes, providing evidence for a recurrent interspecies CNV. However, other mechanisms may also be involved in determining coat color in these 2 breeds.

Sequence characterization of the melanocortin 1 receptor (MC1R) gene in sheep with different coat colours and identification of the putative e allele at the ovine Extension locus

“…However, it seems evident that other loci (e.g. Spotting locus) are involved in determining white coat colour in sheep (Renieri et al, 2008), as also supposed in goats (Fontanesi et al, 2009b). Analysing the ASIP gene in several sheep breeds, we confirmed this hypothesis (Fontanesi et al, in press-a).…”

Coat colours in the Massese sheep breed are associated with mutations in the agouti signalling protein (ASIP) and melanocortin 1 receptor (MC1R) genes

“…Departure from the Hardy-Weinberg equilibrium was highly significant (P 5 0.000018), but this result is clearly biased by the presence of duplicated copies in which one copy (or more copies) could carry the g.5172T allele and the other(s) could carry the g.5172A allele. Similar results are common in loci with CNV (Fontanesi et al, 2009b). Three missense MC1R mutations were genotyped in all Massese sheep, as in part reported by Fontanesi et al (2010a).…”

Molecular genetic basis of pod corn ( Tunicate maize)

“…The combination of gene duplication, promoter rearrangement and deregulated expression, and duplicate mutant copy loss, as exemplified by pod corn (this work) and white sheep and goats (33,34), may be of more general importance in domestication, and possibly also in natural evolution, than has previously been assumed.…”

“…Little is known about the role of tandem gene duplications and loss of duplicate copies in domestication processes, but our findings concerning the Tu locus are not unparalleled. Strikingly similar cases for copy number variation are provided by the Agouti loci controlling coat color in sheep and goats (33,34). The typical coat color of WT sheep is dark-bodied with a pale belly, but artificial selection for white fibers during domestication led to a high frequency of the white coat phenotype in certain breeds of sheep (33).…”