Varp is a novel Rab32/38-binding protein that regulates Tyrp1 trafficking in melanocytes.
Two small GTPase Rabs, Rab32 and Rab38, have recently been proposed to regulate trafficking of melanogenic enzymes to melanosomes in mammalian epidermal melanocytes; however, the exact molecular mechanism of Rab32/38-mediated transport of melanogenic enzymes has never been clarified, because no Rab32/38-specific effector has ever been identified. In this study, we screened for a Rab32/38-specific effector by a yeast two-hybrid assay using a guanosine triphosphate (GTP)-locked Rab32/38 as bait and found that VPS9-ankyrin-repeat protein (Varp)/Ankrd27, characterized previously as a guanine nucleotide exchange factor (GEF) for Rab21, functions as a specific Rab32/38-binding protein in mouse melanocyte cell line melan-a. Deletion analysis showed that the first ankyrin-repeat (ANKR1) domain functions as a GTP-dependent Rab32/38-binding domain, but that the N-terminal VPS9 domain (i.e., Rab21-GEF domain) does not. Small interfering RNA-mediated knockdown of endogenous Varp in melan-a cells caused a dramatic reduction in Tyrp1 (tyrosinase-related protein 1) signals from melanosomes but did not cause any reduction in Pmel17 signals. Furthermore, expression of the ANKR1 domain in melan-a cells also caused a dramatic reduction of Tyrp1 signals, whereas the VPS9 domain had no effect. Based on these findings, we propose that Varp functions as the Rab32/38 effector that controls trafficking of Tyrp1 in melanocytes.
Tamura
Kanako
K
Laboratory of Membrane Trafficking Mechanisms, Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University, Aobayama, Aoba-ku, Sendai, Miyagi 980-8578, Japan.
Ohbayashi
Norihiko
N
Maruta
Yuto
Y
Kanno
Eiko
E
Itoh
Takashi
T
Fukuda
Mitsunori
M
D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000172:deficiency / Q000378:metabolism*; D008544:Melanocytes / Q000166:cytology / Q000201:enzymology*; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
D020662:Guanine Nucleotide Exchange Factors; D008562:Membrane Glycoproteins; D034741:RNA, Small Interfering; C512038:VPS9-ankyrin-repeat protein, mouse; D010088:Oxidoreductases; C079621:Tyrp1 protein, mouse; C478766:Rab32 protein, mouse; C498786:Rab38 protein, mouse; D020691:rab GTP-Binding Proteins