Human melanocytes from individuals with different skin types, as well as from the skin of the same individual, are heterogeneous in their melanin content. This heterogeneity may be attributed to differences in the activity and expression of the three melanogenic proteins: tyrosinase and tyrosinase-related proteins 1 and 2 (gp75 and DOPAchrome tautomerase, respectively), which in turn are affected by certain regulatory factors. Established melanocyte strains that exhibited intrinsic melanogenic heterogeneity could be separated into subpopulations according to density and melanin content by Percoll density gradient centrifugation. The least melanotic subpopulation consisted of melanocytes that contained an active tyrosinase enzyme and a low amount of melanin. Tyrosinase activity and the quantities of tyrosinase enzyme, tyrosinase-related protein-1 and DOPAchrome tautomerase gradually increased with increased melanin content and Percoll density of the isolated melanocyte subpopulations. We have found a direct correlation between melanin content, tyrosinase activity and the expression of the three melanogenic proteins in melanocyte strains established from different skin types. Addition of the two epidermal cytokines, tumor necrosis factor-alpha or interleukin-1 alpha, to cultures of human melanocytes from different skin types caused decreased proliferation, tyrosinase activity and expression of tyrosinase, tyrosinase-related protein-1 and DOPAchrome tautomerase. Similar results were obtained when Percoll-derived melanocyte subpopulations were treated with tumor necrosis factor-alpha and interleukin-1 alpha. These results indicate that the variation in melanin content in human melanocytes is due to differences in the activity and expression of the melanogenic proteins, which are influenced by autocrine and paracrine factors.