The efficient intracellular sequestration of the insulin-regulatable glucose transporter (GLUT-4) is conferred by the NH2 terminus.
GLUT-4 is the major facilitative glucose transporter isoform in tissues that exhibit insulin-stimulated glucose transport. Insulin regulates glucose transport by the rapid translocation of GLUT-4 from an intracellular compartment to the plasma membrane. A critical feature of this process is the efficient exclusion of GLUT-4 from the plasma membrane in the absence of insulin. To identify the amino acid domains of GLUT-4 which confer intracellular sequestration, we analyzed the subcellular distribution of chimeric glucose transporters comprised of GLUT-4 and a homologous isoform, GLUT-1, which is found predominantly at the cell surface. These chimeric transporters were transiently expressed in CHO cells using a double subgenomic recombinant Sindbis virus vector. We have found that wild-type GLUT-4 is targeted to an intracellular compartment in CHO cells which is morphologically similar to that observed in adipocytes and muscle cells. Sindbis virus-produced GLUT-1 was predominantly expressed at the cell surface. Substitution of the GLUT-4 amino-terminal region with that of GLUT-1 abolished the efficient intracellular sequestration of GLUT-4. Conversely, substitution of the NH2 terminus of GLUT-1 with that of GLUT-4 resulted in marked intracellular sequestration of GLUT-1. These data indicate that the NH2-terminus of GLUT-4 is both necessary and sufficient for intracellular sequestration.
Piper
R C
RC
Department of Cell Biology and Physiology, Washington University, St. Louis, Missouri 63110.
Tai
C
C
Slot
J W
JW
Hahn
C S
CS
Rice
C M
CM
Huang
H
H
James
D E
DE
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D005455:Fluorescent Antibody Technique; D005822:Genetic Vectors; D006056:Golgi Apparatus / Q000378:metabolism; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D012845:Sindbis Virus / Q000235:genetics; D013329:Structure-Activity Relationship
D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins