Investigation of the Agouti gene for the identification of useful markers for coat colour association studies in domestic rabbits

Abstract

In wild-type mice, it is well known that Agouti is expressed in skin where it controls the banded-hair Agouti phenotype. Molecular genetics and pharmacological studies show that mutually exclusive binding of the melanocortin 1 receptor (MC1R) by the Agouti protein or by 𰂲-melanocyte-stimulating hormone (a-MSH) signals hair-bulb melanocytes to synthesise preferentially either pheomelanin (yellow-red pigment) or eumelanin (black-brown pigment), respectively. In mice as well as in other species, loss-of-function mutations of the Agouti gene determine only the production of eumelanin while gain-of-function mutations lead to pheomelanin production. A variety of coat colours appear as a result of these alterations that show also epistatic interactions with MC1R mutations. In rabbit, classical studies have suggested the presence of three alleles at the Agouti locus: A (wild type allele), at (black and tan) and a (non-agouti). We recently showed that mutations in the rabbit MC1R gene are associated with coat colours in different breeds. We, furthermore, supposed that the MC1R gene might have partial epistatic actions over the Agouti locus. Here, in order to clarify the interactions between the MC1R and Agouti loci we studied the rabbit Agouti gene with the objective to identify mutations that could be associated or could be useful in association investigations with coat colours in domestic rabbit breeds. Total genomic DNA was extracted from peripheral blood or hair roots of rabbits belonging to breeds having different coat colours. PCR primers were designed to amplify the four exons of the Agouti gene. Sequencing of the fragment encompassing part of intron 2, exon 3 and part of intron 3 showed three single nucleotide polymorphisms (SNPs; A<T, A<G and T<C) in intron 2 and one in intron 3 (G<A). These SNPs produced two haplotypes, A-A-T-G and T-G-C-A. The first haplotype was observed in Giant Grey and Belgian Hare rabbits. The second haplotype was identified in Burgundy Fawn and Checkered Giant animals. Further studies are underway to obtain a complete characterization of the rabbit Agouti gene and to evaluate the identified markers in association studies with coat colours in this species.