Possible domains responsible for intracellular targeting and insulin-dependent translocation of glucose transporter type 4.
Translocation of the type 4 glucose transporter (GLUT4) to the cell surface from an intracellular pool is the major mechanism of insulin-stimulated glucose uptake in insulin-target cells. We developed a highly sensitive and quantitative method to detect GLUT4 immunologically on the surface of intact cells, using c-myc epitope-tagged GLUT4 (GLUT4myc). We constructed c-myc epitope-tagged glucose transporter type 1 (GLUT1myc) and found that the GLUT1myc was also translocated to the cell surface of Chinese hamster ovary cells, 3T3-L1 fibroblasts and NIH 3T3 cells, in response to insulin, but the degree of translocation was less than that of GLUT4myc. Since GLUT1 and GLUT4 have different intracellular distributions and different degrees of insulin-stimulated translocation, we examined the domains of GLUT4, using c-myc epitope-tagged chimeric glucose transporters between these two isoforms. The results indicated that, (1) all the cytoplasmic N-terminal region, middle intracellular loop and cytoplasmic C-terminal region of GLUT4 have independent intracellular targeting signals, (2) these sequences for intracellular targeting of GLUT4 were not sufficient to determine GLUT4 translocation in response to insulin, and (3) the N-terminal half of GLUT4 devoid both of cytoplasmic N-terminus and of middle intracellular loop seems to be necessary for insulin-stimulated GLUT4 translocation.
Ishii
K
K
Department of Enzyme Genetics, University of Tokushima, Japan.
Hayashi
H
H
Todaka
M
M
Kamohara
S
S
Kanai
F
F
Jinnouchi
H
H
Wang
L
L
Ebina
Y
Y
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism; D000939:Epitopes / Q000235:genetics; D051272:Glucose Transporter Type 1; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000378:metabolism*; D009124:Muscle Proteins; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000276:immunology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012969:Sodium Fluoride / Q000494:pharmacology; D013347:Subcellular Fractions / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000939:Epitopes; D051272:Glucose Transporter Type 1; D051275:Glucose Transporter Type 4; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D016271:Proto-Oncogene Proteins c-myc; D011993:Recombinant Fusion Proteins; C495678:Slc2a1 protein, mouse; C495679:Slc2a1 protein, rat; C495693:Slc2a4 protein, mouse; C495694:Slc2a4 protein, rat; D012969:Sodium Fluoride; D013755:Tetradecanoylphorbol Acetate