280 KERATIN AND KERATINIZATION 



their occurrence as pigment granules, are very widely distributed in nature, 

 occurring in plants, in vertebrates and invertebrates. They are formed by 

 the action of copper-containing oxidases, known as phenolases, that catalyse 

 the oxidation of mono- and di-hydric phenols to o-quinones (Mason, 1953 

 and 1955). Phylogenetically the chemical reactions involved in melani- 

 zation developed before keratinization and quite independently of it. 

 Mason has discussed the wide distribution and varied applications of the 

 " phenolase system " in the different phyla. It constitutes an excellent 

 example of a simple biochemical system, which catalyses essentially the 

 same reactions in plants and highly-organized animals, although with an 

 increased specificity towards substrates as the phylogenetic tree is ascended, 

 and which thus finds expression in widely- different characters at different 

 levels. For example, it is responsible for the browning of plant tissues, the 

 hardening of cuticle of arthropods and the pigmentation of chordates. In 

 the higher animals the site of melanin formation has become limited to the 

 pigment cell or melanocyte. 



The biochemical evidence has been reviewed recently by Mason (1955) 

 and Fitzpatrick, Brunet and Kukita (1958). Briefly, the amino acid 

 tyrosine has been shown to be the precursor of the insoluble pigment. The 

 phenolase, tyrosinase, converts tyrosine to DOPA (3 : 4-dihydroxy- 

 phenylalanine) and to " DOPA quinone", which becomes 5 : 6-dihydroxy- 

 indole, the immediate precursor or monomer of the large polymer molecule 

 melanin, which may then be linked to a protein (Fig. 1 16). 



The darkening of a tissue when treated with DOPA (dihydroxypheny- 

 lalanine), Bloch's DOPA-oxidase reaction, has long been used to demon- 

 strate cytologically the sites of melanin formation (Block, 1921), but it is 

 now less favoured, since it may be non-specifically oxidized to melanin. 

 Tyrosine is preferred as substrate although it may fail to demonstrate 

 tyrosinase when the latter is in low concentration. Histochemically 

 tyrosine activity has been demonstrated in autoradiographs by using 

 radioactive tyrosine and C 14 (see Fitzpatrick et al., 1958). 



The small quantities of melanized material from mammalian sources 

 have hindered biochemical research, but by taking advantage of the larger 

 quantities of material available in pigmented tumours (melanomas) this 

 may be overcome. The granules of the Harding-Passey melanoma appear 

 as aggregates of fine dense granules perhaps incompletely supplied with a 

 protein framework (Mercer, unpublished) (Fig. 115(f)). 



Something of the cytological structure associated with these syntheses 

 has been revealed by the electron microscope and by the histochemical 

 location of tyrosinase activity. The cell is roughly polarized in a manner 

 similar to glandular cells (see p. 110) with a limited basophilic reticulum 

 at the end proximal to the attachment to the basement membrane (Birbeck 

 et al., 1956). Here presumably the protein of the granule is synthesized. 



