THE PMEL-17 SILVER LOCUS PROTEIN - CHARACTERIZATION AND INVESTIGATIONOF ITS MELANOGENIC FUNCTION

The silver mutation in mice causes progressive graying of hair due to the loss of functional follicular melanocytes. Recently the silver loc us gene (called Pmel 17) has been cloned; its encoded product shares h omology with a chick melanosomal matrix protein and a bovine retinal p igment epithelial protein. Although the sequence of the silver gene an d the correlation of its expression with pigment production have been reported, its function in melanogenesis is still unknown. In an effort to characterize that function, we have synthesized the predicted carb oxyl-terminal peptide of the mouse Pmel 17 protein and generated a rab bit polyclonal antibody (alpha PEP13) to it; that antibody recognized the silver protein specifically. The immunoaffinity-purified silver pr otein lacked all of the known melanogenic catalytic activities which o ther tyrosinase-related proteins (TRP) have, nor did it appear to modu late any of those TRP activities. Metabolic labeling experiments demon strated that the silver protein disappears in vivo within a few hours, indicating that it is rapidly degraded, or quickly processed to lose its carboxyl terminus. Crossreactivity experiments showed that a recen tly reported anti-melanosomal matrix protein antibody (alpha MX) also recognizes the silver protein, although at a different epitope from th at of alpha PEP13. Using Western immunoblotting, we analyzed subcellul ar fractions isolated from B16 F10 melanoma cells and found that the s ilver protein was rich in the melanosome fraction but was absent from coated vesicles which deliver TRPs to melanosomes. These results sugge st that the silver locus product is a melanosomal matrix protein which may contribute to melanogenesis as a structural protein, although the possibility remains that it also has a novel catalytic function in me lanogenesis.