Functional involvement of Rab1A in microtubule-dependent anterograde melanosome transport in melanocytes

Advance Online Publication August 1, 2012 doi: 10.1242/?jcs.109314 Morié Ishida, Norihiko Ohbayashi, Yuto Maruta, Yuka Ebata and Mitsunori Fukuda*?*Correspondence: 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. Tel.: +81-22-795-7731; Fax: +81-22-795-7733; E-mail: nori{at}m.tohoku.ac.jp. Melanosomes are transported to the cell periphery of melanocytes by coordination between bidirectional microtubule-dependent movements and unidirectional actin-dependent movement. Although both the mechanism of the actin-dependent melanosome transport and the mechanism of the microtubule-dependent retrograde melanosome transport in mammalian skin melanocytes have already been determined, almost nothing is known about the mechanism of the microtubule-dependent anterograde melanosome transport. Small GTPase Rab proteins are common regulators of membrane traffic in all eukaryotes, and in this study we performed genome-wide screening for Rab proteins that are involved in anterograde melanosome transport by expressing 60 different constitutive active (and negative) mutants and succeeded in identifying Rab1A, originally described as a Golgi-resident Rab, as a prime candidate. Endogenous Rab1A protein was found to be localized on mature melanosomes in melanocytes, and its functional ablation either by siRNA-mediated knockdown or by overexpression of a cytosolic form of Rab1A-GTPase-activating protein/TBC1D20 induced perinuclear melanosome aggregation. The results of time-lapse imaging further revealed that long-range anterograde melanosome movements were specifically suppressed in Rab1A-deficient melanocytes, whereas retrograde melanosome transport occurred normally. Taken together, these findings indicate that Rab1A is the first crucial component of the anterograde melanosome transport machinery to be identified in mammalian skin melanocytes.

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