Rab10 and myosin-Va mediate insulin-stimulated GLUT4 storage vesicle translocation in adipocytes

Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
The Journal of Cell Biology (Impact Factor: 9.69). 08/2012; 198(4):545-560. DOI: 10.1083/jcb.201111091
Source: PubMed

ABSTRACT Rab proteins are important regulators of insulin-stimulated GLUT4 translocation to the plasma membrane (PM), but the precise
steps in GLUT4 trafficking modulated by particular Rab proteins remain unclear. Here, we systematically investigate the involvement
of Rab proteins in GLUT4 trafficking, focusing on Rab proteins directly mediating GLUT4 storage vesicle (GSV) delivery to
the PM. Using dual-color total internal reflection fluorescence (TIRF) microscopy and an insulin-responsive aminopeptidase
(IRAP)-pHluorin fusion assay, we demonstrated that Rab10 directly facilitated GSV translocation to and docking at the PM.
Rab14 mediated GLUT4 delivery to the PM via endosomal compartments containing transferrin receptor (TfR), whereas Rab4A, Rab4B,
and Rab8A recycled GLUT4 through the endosomal system. Myosin-Va associated with GSVs by interacting with Rab10, positioning
peripherally recruited GSVs for ultimate fusion. Thus, multiple Rab proteins regulate the trafficking of GLUT4, with Rab10
coordinating with myosin-Va to mediate the final steps of insulin-stimulated GSV translocation to the PM.

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    07/2014; 1. DOI:10.3389/fmed.2014.00020
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    ABSTRACT: Intracellular transport is largely driven by processive microtubule- and actin-based molecular motors. Nonprocessive motors have also been localized to trafficking cargos, but their roles are not well understood [1-7]. Myosin-Ic (Myo1c), a nonprocessive actin motor, functions in a variety of exocytic events, although the underlying mechanisms are not yet clear. To investigate the interplay between myosin-I and the canonical long-distance transport motor kinesin-1, we attached both motor types to lipid membrane-coated bead cargo, using an attachment strategy that allows motors to actively reorganize within the membrane in response to the local cytoskeletal environment. We compared the motility of kinesin-1-driven cargos in the absence and presence of Myo1c at engineered actin/microtubule intersections. We found that Myo1c significantly increases the frequency of kinesin-1-driven microtubule-based runs that begin at actin/microtubule intersections. Myo1c also regulates the termination of processive runs. Beads with both motors bound have a significantly higher probability of pausing at actin/microtubule intersections, remaining tethered for an average of 20 s, with some pauses lasting longer than 200 s. The actin-binding protein nonmuscle tropomyosin (Tm) provides spatially specific regulation of interactions between myosin motors and actin filaments in vivo [8-12]; in the crossed-filament in vitro assay, we found that Tm2-actin abolishes Myo1c-specific effects on both run initiation and run termination. Together, these observations suggest Myo1c is important for the selective initiation and termination of kinesin-1-driven runs along microtubules at specific actin filament populations within the cell. Copyright © 2015 Elsevier Ltd. All rights reserved.
    Current Biology 02/2015; DOI:10.1016/j.cub.2014.12.008 · 9.92 Impact Factor
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    ABSTRACT: Insulin causes the exocytic translocation of GLUT4 glucose transporters to stimulate glucose uptake in fat and muscle. Previous results support a model in which TUG traps GLUT4 in intracellular, insulin-responsive vesicles termed GLUT4 storage vesicles (GSVs). Insulin triggers TUG cleavage to release the GSVs; GLUT4 then recycles through endosomes during ongoing insulin exposure. The TUG C-terminus binds a GSV anchoring site comprising Golgin-160 and possibly other proteins. Here, we report that the TUG C-terminus is acetylated. The TUG C-terminal peptide bound the Golgin-160-associated protein, ACBD3 (acyl-CoA binding domain containing 3), and acetylation reduced binding of TUG to ACBD3, but not to Golgin-160. Mutation of the acetylated residues impaired insulin-responsive GLUT4 trafficking in 3T3-L1 adipocytes. ACBD3 overexpression enhanced the translocation of GSV cargos, GLUT4 and IRAP, and ACBD3 was required for intracellular retention of these cargos in unstimulated cells. SIRT2, a NAD+-dependent deacetylase, bound TUG and deacetylated the TUG peptide. SIRT2 overexpression reduced TUG acetylation and redistributed GLUT4 and IRAP to the plasma membrane in 3T3-L1 adipocytes. Mutation of the acetylated residues in TUG abrogated these effects. In mice, SIRT2 deletion increased TUG acetylation and proteolytic processing. During glucose tolerance tests, glucose disposal was enhanced in SIRT2 knockout mice, compared to wildtype controls, without any effect on insulin concentrations. Together, these data support a model in which TUG acetylation modulates its interaction with Golgi matrix proteins and is regulated by SIRT2. Moreover, acetylation of TUG enhances its function to trap GSVs within unstimulated cells, and enhances insulin-stimulated glucose uptake. Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
    Journal of Biological Chemistry 01/2015; DOI:10.1074/jbc.M114.603977 · 4.60 Impact Factor

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