ADAM protease inhibitors reduce melanogenesis by regulating PMEL17 processing in human melanocytes

• Melanin content was reduced by the treatment with ADAM protease inhibitor.
• The number of mature melanosomes was reduced by the treatment with ADAM protease inhibitor.
• ADAM protease inhibitor acted on the processing of PMEL17 as γ-secretase and BACE2-independently.

BackgroundADAMs (a disintegrin and metalloprotease) are a family of proteases involved in ectodomain shedding that play a role in various biological processes such as cell adhesion and migration. ADAM10 and ADAM17 are suggested to be involved in pigmentary disorders.ObjectiveWe examined the effect of ADAM protease inhibitors on the modulation of melanogenesis in normal human epidermal melanocytes (NHEM).MethodsNHEMs and B16F10 treated with ADAM protease inhibitors were analyzed. AlamarBlue cell proliferation assay, melanin content assay, tyrosinase activity assay, Western blotting analysis, electron microscopic analysis, and RNA interference were employed.ResultsIn NHEMs, melanin content was reduced by treatment with ADAM protease inhibitors. The inhibitors did not change the protein expression of tyrosinase, TRP-1, and MITF. In B16F10 cells, treatment of the cells with ADAM protease inhibitor diminished the α-MSH-induced increase in melanin content. Electron microscopy showed that the number of fibrillar and mature melanosomes was significantly reduced and that the vacuolar compartments were filled with dense unstructured aggregates after treatment with ADAM protease inhibitors. We therefore focused on the processing of PMEL17, a melanosomal glycoprotein that forms a fibrillar matrix on which melanin gets deposited. Proteolytic processing of PMEL17 is required to form functional fibrils during melanogenesis. Recently, γ-secretase and β-site amyloid precursor protein-cleaving enzyme 2 (BACE2) were found to cleave PMEL17. We found that ADAM protease inhibitors exerted effects on the processing of C-terminal and N-terminal fragments of PMEL17. Using BACE2 siRNA and γ-secretase inhibitor, we showed that ADAM protease inhibitor affected PMEL17 processing in a γ-secretase and BACE2-independent mechanism.ConclusionSeveral proteases, including ADAM proteases, can contribute to the formation of fibrils and their assembly into sheets in melanosomes.