Overexpression of anAgouticDNA in the Skin of Transgenic Mice Recapitulates Dominant Coat Color Phenotypes of Spontaneous Mutants

The classical mouse fancyAgoutigene is responsible for the wild-type coat color where hairs are banded black and yellow. TheAgoutigene encodes a 131-amino-acid secreted protein product that regulates phaeomelanin synthesis by melanocytes in mice. Mice with a dominant mutation at this locus,Ay, develop a yellow coat color, obesity, and diabetes, as the result of a deletion that results in ectopic overexpression of theAgoutigene mRNA in all tissues examined. Obesity and diabetes inAymutant mice could be caused by circulation of the protein, or localized action in specific tissues as a paracrine factor acting in cell–cell communication. To test these two possibilities, theAgouticDNA was overexpressed in the skin of transgenic mice using either theTyrosinase-Related Protein-1or the keratin-14 (K14) promoter, the latter with and without an intron. The K14 promoter directed high constitutive levels of expression ofAgoutimRNA in the skin, and several lines of transgenic mice exhibited coat colors resembling dominantAgoutiallele phenotypes. Two highly expressing K14–Agouti transgenic lines, with light-yellow pelage, were analyzed for obesity and hyperglycemia. The transgenic mice were not significantly different from the controls (P> 0.05), indicating that theAgoutiproduct does not act as an endocrine factor. RNase protection assays revealed a correlation between the levels of dorsal and ventral skin expression with pigmentation/phaeomelanin phenotypes. Co-injection experiments with the Agouti transgenes and other transgenes demonstrated co-integration of the two constructs at the same chromosomal site in approximately 95% of F1 progeny, allowing transgene inheritance to be visibly detected.