title: A complex of Rab13 with MICAL-L2 and α-actinin-4 is essential for insulin-dependent GLUT4 exocytosis.
abstract: Insulin promotes glucose uptake into skeletal muscle through recruitment of glucose transporter 4 (GLUT4) to the plasma membrane. Rab GTPases are molecular switches mobilizing intracellular vesicles, and Rab13 is necessary for insulin-regulated GLUT4-vesicle exocytic translocation in muscle cells. We show that Rab13 engages the scaffold protein MICAL-L2 in this process. RNA interference-mediated knockdown of MICAL-L2 or truncated MICAL-L2 (MICAL-L2-CT) impaired insulin-stimulated GLUT4 translocation. Insulin increased Rab13 binding to MICAL-L2, assessed by pull down and colocalization under confocal fluorescence and structured illumination microscopies. Association was also visualized at the cell periphery using TIRF microscopy. Insulin further increased binding of MICAL-L2 to α-actinin-4 (ACTN4), a protein involved in GLUT4 translocation. Rab13, MICAL-L2, and ACTN4 formed an insulin-dependent complex assessed by pull down and confocal fluorescence imaging. Of note, GLUT4 associated with the complex in response to insulin, requiring the ACTN4-binding domain in MICAL-L2. This was demonstrated by pull down with distinct fragments of MICAL-L2 and confocal and structured illumination microscopies. Finally, expression of MICAL-L2-CT abrogated the insulin-dependent colocalization of Rab13 with ACTN4 or Rab13 with GLUT4. Our findings suggest that MICAL-L2 is an effector of insulin-activated Rab13, which links to GLUT4 through ACTN4, localizing GLUT4 vesicles at the muscle cell periphery to enable their fusion with the membrane.
authors.affiliation: Program in Cell Biology, Hospital for Sick Children, Toronto, ON M5G 0A4, Canada.
authors.forename: Yi
authors.initials: Y
authors.lastname: Sun
authors.affiliation: Program in Cell Biology, Hospital for Sick Children, Toronto, ON M5G 0A4, Canada.
authors.forename: Javier
authors.initials: J
authors.lastname: Jaldin-Fincati
authors.affiliation: Program in Cell Biology, Hospital for Sick Children, Toronto, ON M5G 0A4, Canada.
authors.forename: Zhi
authors.initials: Z
authors.lastname: Liu
authors.affiliation: Program in Cell Biology, Hospital for Sick Children, Toronto, ON M5G 0A4, Canada.
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Bilan
authors.affiliation: Program in Cell Biology, Hospital for Sick Children, Toronto, ON M5G 0A4, Canada amira@sickkids.ca.
authors.forename: Amira
authors.initials: A
authors.lastname: Klip
mesh_terms: D000185:Actinin / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005089:Exocytosis / Q000502:physiology; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D051275:Glucose Transporter Type 4 / Q000378:metabolism*; D006801:Humans; D007328:Insulin / Q000378:metabolism / Q000494:pharmacology; D008840:Microfilament Proteins / Q000378:metabolism*; D032342:Muscle Cells / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D051381:Rats; D015398:Signal Transduction; D019108:Tight Junctions / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000378:metabolism
chemical_list: C491751:Actn4 protein, rat; D051275:Glucose Transporter Type 4; D007328:Insulin; D008840:Microfilament Proteins; C495694:Slc2a4 protein, rat; D000185:Actinin; D020558:GTP Phosphohydrolases; C541433:Rab13 protein, rat; D020691:rab GTP-Binding Proteins