Two cell type-specific Rab proteins, Rab32 and Rab38 (Rab32/38), have been proposed to regulate the trafficking of melanogenic
enzymes, including tyrosinase and tyrosinase-related protein 1 (Tyrp1), to melanosomes in melanocytes. The same as other GTPases,
Rab32/38 function as switch molecules that cycle between a GDP-bound inactive form and GTP-bound active form, and the cycle
is thought to be regulated by an activating enzyme GEF (guanine nucleotide exchange factor) and an inactivating enzyme GAP
(GTPase-activating protein), which stimulates the GTPase activity of Rab32/38.
Although BLOC-3 has already been identified as a Rab32/38-specific GEF that regulates the trafficking of tyrosinase and Tyrp1,
no physiological GAP for Rab32/38 in melanocytes has never been identified, and it has remained unclear whether Rab32/38 is
involved in the trafficking of dopachrome tautomerase, another melanogenic enzyme, in mouse melanocytes. In this study we
investigated RUTBC1, which was originally characterized as a Rab9-binding protein and GAP for Rab32 and Rab33B in vitro, and
the results demonstrated that RUTBC1 functions as a physiological GAP for Rab32/38 in the trafficking of all three melanogenic
enzymes in mouse melanocytes. The results of this study also demonstrated involvement of Rab9A in the regulation of the RUTBC1
localization and in the trafficking of all three melanogenic enzymes, and we discovered that either excess activation or inactivation
of Rab32/38 achieved by manipulating RUTBC1 inhibits the trafficking of all three melanogenic enzymes. These results collectively
indicated that proper spatiotemporal regulation of Rab32/38 is essential for the trafficking of all three melanogenic enzymes
in mouse melanocytes.