title: The GTPase-deficient Rab27A(Q78L) mutant inhibits melanosome transport in melanocytes through trapping of Rab27A effector protein Slac2-a/melanophilin in their cytosol: development of a novel melanosome-targetinG tag.
abstract: The small GTPase Rab27A is a crucial regulator of actin-based melanosome transport in melanocytes, and functionally defective Rab27A causes human Griscelli syndrome type 2, which is characterized by silvery hair. A GTPase-deficient, constitutively active Rab27A(Q78L) mutant has been shown to act as an inhibitor of melanosome transport and to induce perinuclear aggregation of melanosomes, but the molecular mechanism by which Rab27A(Q78L) inhibits melanosome transport remained to be determined. In this study, we attempted to identify the primary cause of the perinuclear melanosome aggregation induced by Rab27A(Q78L). The results showed that Rab27A(Q78L) is unable to localize on mature melanosomes and that its inhibitory activity on melanosome transport is completely dependent on its binding to the Rab27A effector Slac2-a/melanophilin. When we forcibly expressed Rab27A(Q78L) on mature melanosomes by using a novel melanosome-targeting tag that we developed in this study and named the MST tag, the MST-Rab27A(Q78L) fusion protein behaved in the same manner as wild-type Rab27A. It localized on mature melanosomes without inducing melanosome aggregation and restored normal peripheral melanosome distribution in Rab27A-deficient cells. These findings indicate that the GTPase activity of Rab27A is required for its melanosome localization but is not required for melanosome transport.
authors.affiliation: 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.
authors.forename: Morié
authors.initials: M
authors.lastname: Ishida
authors.affiliation: 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.
authors.forename: Saki P
authors.initials: SP
authors.lastname: Arai
authors.affiliation: 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.
authors.forename: Norihiko
authors.initials: N
authors.lastname: Ohbayashi
authors.affiliation: 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. Electronic address: nori@m.tohoku.ac.jp.
authors.forename: Mitsunori
authors.initials: M
authors.lastname: Fukuda
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D006801:Humans; D007153:Immunologic Deficiency Syndromes / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051359:Lymphohistiocytosis, Hemophagocytic; D020460:Melanosomes / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D020125:Mutation, Missense; D016116:Piebaldism / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D000081207:Primary Immunodeficiency Diseases; D021381:Protein Transport / Q000235:genetics; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000075745:rab27 GTP-Binding Proteins
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C492410:MLPH protein, human; C435724:Mlph protein, mouse; D000075745:rab27 GTP-Binding Proteins; C066694:RAB27A protein, human; C512962:Rab27a protein, mouse; D020691:rab GTP-Binding Proteins
keywords: Low Molecular Weight G Proteins; Melanogenesis; Membrane Trafficking; Protein Targeting; Rab; Small GTPases; Subcellular Organelles