The melanocortin 1 receptor is the principal mediator of the effects of agouti signaling protein on mammalian melanocytes

ABSTRACT The agouti gene codes for agouti signaling protein (ASP), which is temporally expressed in wild-type mouse follicular melanocytes where it induces pheomelanin synthesis. Studies using purified full-length agouti signaling protein has shown that it competes with α-melanocyte stimulating hormone for binding to the melanocortin 1 receptor. We have investigated whether ASP binds exclusively to the melanocortin 1 receptor expressed on mouse melanocytes in primary culture, or additionally activates a receptor that has not been identified yet. We have compared the responses of congenic mouse melanocytes derived from C57 BL/6J-E+/E+, e/e, or Eso/Eso mice to α-MSH and/or ASP. E+/E+ melanocytes express the wild-type melanocortin 1 receptor, e/e melanocytes express a loss-of-function mutation in the melanocortin 1 receptor that results in a yellow coat color, and Eso/Eso is a mutation that causes constitutive activation of the melanocortin 1 receptor and renders melanocytes unresponsive to α-melanocyte stimulating hormone. Mouse E+/E+ melanocytes, but not e/e or Eso/Eso melanocytes, respond to agouti signaling protein with decreased basal tyrosinase activity, and reduction in levels of tyrosinase and tyrosinase-related proteins 1 and 2. Only in E+/E+ melanocytes does agouti signaling protein abrogate the stimulatory effects of α-melanocyte stimulating hormone on cAMP formation and tyrosinase activity. These results indicate that a functional melanocortin 1 receptor is obligatory for the response of mammalian melanocytes to agouti signaling protein.