title: Selective visualization of GLUT4 storage vesicles and associated Rab proteins using IRAP-pHluorin.
abstract: Fluorescence microscopy and fluorescent protein (FP)-tagged GLUT4 molecule have been great tools to characterize GLUT4 localization and dynamics inside the cell. However, it was difficult to distinguish GLUT4 storage vesicles (GSVs) from other intracellular compartments containing GLUT4 in live cells. Here, we describe the use of IRAP-pHluorin and total internal reflection fluorescence (TIRF) microscopy to selectively visualize GSVs and Rab proteins that associate with GSVs. This assay is also valuable to further defining GSV identity by unraveling other GSV-associated proteins.
authors: Cell Biology and Metabolism Program, NICHD - Eunice Kennedy Shriver National Institute of Child Health and Human Development, NIH - National Institutes of Health, Building 18T, Room 101, 18 Library Drive, Bethesda, MD, 20892, USA.
authors: Yu
authors: Y
authors: Chen
authors: Jennifer
authors: J
authors: Lippincott-Schwartz
mesh_terms: D041721:3T3-L1 Cells; D017667:Adipocytes / Q000166:cytology; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D010122:Cystinyl Aminopeptidase / Q000378:metabolism*; D005089:Exocytosis / Q000187:drug effects; D051275:Glucose Transporter Type 4 / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D007091:Image Processing, Computer-Assisted; D007328:Insulin / Q000494:pharmacology; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D014162:Transfection; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
chemical_list: D051275:Glucose Transporter Type 4; D007328:Insulin; C485187:PHluorin; D049452:Green Fluorescent Proteins; D010122:Cystinyl Aminopeptidase; C105964:leucyl-cystinyl aminopeptidase; D020691:rab GTP-Binding Proteins