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

• Published in: The Journal of biological chemistry Vol. 289; no. 16; pp. 11059 - 11067
• Main Authors: Ishida, Morié, Arai, Saki P, Ohbayashi, Norihiko, Fukuda, Mitsunori
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 18.04.2014
• Subjects: Adaptor Proteins, Signal Transducing - genetics; Adaptor Proteins, Signal Transducing - metabolism; Amino Acid Substitution; Animals; Cell Biology; Chlorocebus aethiops; COS Cells; Humans; Immunologic Deficiency Syndromes - genetics; Immunologic Deficiency Syndromes - metabolism; Immunologic Deficiency Syndromes - pathology; Lymphohistiocytosis, Hemophagocytic; Melanosomes - genetics; Melanosomes - metabolism; Melanosomes - pathology; Mice; Mutation, Missense; Piebaldism - genetics; Piebaldism - metabolism; Piebaldism - pathology; Primary Immunodeficiency Diseases; Protein Transport - genetics; rab GTP-Binding Proteins - genetics; rab GTP-Binding Proteins - metabolism; rab27 GTP-Binding Proteins; Membrane Trafficking; Rab; Subcellular Organelles; Protein Targeting; Small GTPases; Melanogenesis; Low Molecular Weight G Proteins
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M114.552281

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.