GENETIC REGULATION OF MELANIN BIOSYNTHESIS

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DESCRIPTION: (Adapted from the applicant's abstract) - Melanin pigment is a complex biopolymer found primarily in the skin, hair, and eyes. It functions as a photoprotective pigment of the body surface, provides cosmetic appeal and camouflage, and is involved in the development of the eye and the optic nerves. Melanin synthesis starts with tyrosine and involves a series of steps that lead to black-brown eumelanin or red-yellow pheomelanin. Tyrosinase, as part of a complex with other pigment enzymes and protein factors, catalyzes the first two steps in the melanin synthetic pathway, and the loss of tyrosinase activity is associated with a total loss of melanin in the melanocyte. The investigators propose to study the role of tyrosinase in the regulation of melanin synthesis using human tyrosinase-related oculocutaneous albinism or OCA1 as the model system. Most tyrosinase gene mutations are associated with a total lack of melanin (OCA1A) while a number are associated with the formation of some melanin pigment in the hair, skin, and eyes after birth (OCA1B). The investigators hypothesize that OCA1B mutations produce enzymes with residual acidity, and they feel that the characterization of these mutations and their effect on enzyme structure and function will provide insight into the regulation of melanin synthesis. The investigators propose three specific aims. First, they will characterize the molecular genotype of individuals with OCA1B to extend their initial studies of this type of albinism. They will identify responsible tyrosinase gene mutations by direct DNA sequencing and by mRNA analysis. Second, they will characterize the effects of OCA1B mutations on tyrosinase function. Individual mutations will be recreated and analyzed in expression studies of mutant enzyme. Third, they will perform structure:function studies of recombinant normal and mutant tyrosinase enzyme. Purified Streptomyces antibioticus tyrosinase and human tyrosinase will be used for crystallization and x-ray analysis to determine 3-dimensional structure and functional domains of the enzyme. These studies will provide fundamental information on the role of tyrosinase in the regulation of melanin synthesis and the function of this critical pigment enzyme. Knowledge gained will be important for future development of effective therapy for the protection of the skin from the oncogenic effects of ultraviolet radiation, and for promoting normal ocular development.