Rab32/38-Dependent and -Independent Transport of Tyrosinase to Melanosomes in B16-F1 Melanoma Cells

Int J Mol Sci. 2022 Nov 16;23(22):14144. doi: 10.3390/ijms232214144.

Abstract

B16-F1 melanoma cells have often been used as a model to investigate melanogenesis, but the evidence that melanosome biogenesis and transport occur by the same mechanisms in normal melanocytes and B16-F1 cells is insufficient. In this study, we established knockout B16-F1 cells for each of several key factors in melanogenesis, i.e., tyrosinase (Tyr), Hps4, Rab27A, and Rab32·Rab38 (Rab32/38), and then compared their phenotypes with the phenotypes of corresponding mutant mouse melanocyte cell lines, i.e., melan-c, melan-le, melan-ash, and Rab32-deficient melan-cht cells, respectively. The results showed that Tyr and Rab27A are also indispensable for melanin synthesis and peripheral melanosome distribution, respectively, in B16-F1 cells, but that Hps4 or its downstream targets Rab32/38 are not essential for Tyr transport in B16-F1 cells, suggesting the existence of a Rab32/38-independent Tyr transport mechanism in B16-F1 cells. We then performed comprehensive knockdown screening of Rab small GTPases and identified Rab10 and Rab24, previously uncharacterized Rabs in melanocytes, as being involved in Tyr transport under Rab32/38-null conditions. Our findings indicate a difference between the Tyr transport mechanism in melanocytes and B16-F1 cells in terms of Rab32/38-dependency and a limitation in regard to using melanoma cells as a model for melanocytes, especially when investigating the mechanism of endosomal Tyr transport.

Keywords: Hps4; Rab small GTPase; endosome; melanocyte; melanogenic enzyme; melanoma; melanosome; membrane traffic; tyrosinase; tyrosinase-related protein 1 (Tyrp1).

MeSH terms

  • Animals
  • Melanocytes / metabolism
  • Melanoma* / metabolism
  • Melanosomes* / metabolism
  • Mice
  • Monophenol Monooxygenase* / genetics
  • Monophenol Monooxygenase* / metabolism
  • rab GTP-Binding Proteins* / genetics
  • rab GTP-Binding Proteins* / metabolism

Substances

  • Hps4 protein, mouse
  • Monophenol Monooxygenase
  • rab GTP-Binding Proteins
  • Rab24 protein, mouse
  • Rab32 protein, mouse