title: Perinuclear localization and insulin responsiveness of GLUT4 requires cytoskeletal integrity in 3T3-L1 adipocytes.
abstract: The GLUT4 glucose transporter resides mostly in perinuclear membranes in unstimulated 3T3-L1 adipocytes and is acutely translocated to the cell surface in response to insulin. Using a novel method to purify intracellular GLUT4-enriched membranes, we identified by mass spectrometry the intermediate filament protein vimentin and the microtubule protein alpha-tubulin as components of these membranes. Immunoelectron microscopy of the GLUT4-containing membranes also revealed their association with these cytoskeletal proteins. Disruption of intermediate filaments and microtubules in 3T3-L1 adipocytes by microinjection of a vimentin-derived peptide of the helix initiation 1A domain caused marked dispersion of perinuclear GLUT4 to peripheral regions of the cells. Inhibition of the microtubule-based motor dynein by brief cytoplasmic acidification of cultured adipocytes also dispersed perinuclear GLUT4 and inhibited insulin-stimulated GLUT4 translocation to the cell surface. Insulin sensitivity was restored as GLUT4 was again concentrated near the nucleus upon recovery of cells in physiological buffer. These data suggest that GLUT4 trafficking to perinuclear membranes of cultured adipocytes is directed by dynein and is required for optimal GLUT4 regulation by insulin.
authors.affiliation: Program in Molecular Medicine and Departments of Biochemistry and Molecular Biology and Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA.
authors.forename: A
authors.initials: A
authors.lastname: Guilherme
authors.forename: M
authors.initials: M
authors.lastname: Emoto
authors.forename: J M
authors.initials: JM
authors.lastname: Buxton
authors.forename: S
authors.initials: S
authors.lastname: Bose
authors.forename: R
authors.initials: R
authors.lastname: Sabini
authors.forename: W E
authors.initials: WE
authors.lastname: Theurkauf
authors.forename: J
authors.initials: J
authors.lastname: Leszyk
authors.forename: M P
authors.initials: MP
authors.lastname: Czech
mesh_terms: D016475:3T3 Cells; D017667:Adipocytes / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D000818:Animals; D002458:Cell Fractionation; D003598:Cytoskeletal Proteins / Q000032:analysis / Q000378:metabolism; D003599:Cytoskeleton / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure*; D004398:Dyneins / Q000037:antagonists & inhibitors; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000494:pharmacology*; D007425:Intracellular Membranes / Q000502:physiology* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000032:analysis; D051379:Mice; D009004:Monosaccharide Transport Proteins / Q000032:analysis / Q000378:metabolism*; D009124:Muscle Proteins; D009685:Nuclear Envelope / Q000502:physiology*; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology; D017433:Protein Structure, Secondary; D050683:R-SNARE Proteins; D051381:Rats; D011990:Receptors, Transferrin / Q000032:analysis; D014746:Vimentin / Q000737:chemistry / Q000494:pharmacology
chemical_list: D003598:Cytoskeletal Proteins; D051275:Glucose Transporter Type 4; D007328:Insulin; D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D010446:Peptide Fragments; D050683:R-SNARE Proteins; D011990:Receptors, Transferrin; C495693:Slc2a4 protein, mouse; C495694:Slc2a4 protein, rat; D014746:Vimentin; D004398:Dyneins