Distinct signals in the GLUT4 glucose transporter for internalization and for targeting to an insulin-responsive compartment.

Abstract

Abstract. In adipose and muscle cells, insulin stimulates a rapid and dramatic increase in glucose uptake, primarily by promoting the redistribution of the GLUT4 glucose transporter from its intracellular storage site to the plasma membrane. In contrast, the more ubiquitously expressed isoform GLUT1 is localized at the cell surface in the basal state, and shows a less dramatic translocation in response to insulin. To identify sequences involved in the differential subcellular localization and hormone-responsiveness of these isoforms, chimeric GLUT1/GLUT4 transporters were stably expressed in mouse 3T3-L1 adipocytes. The NH 2 terminus of GLUT4 contains sequences capable of sequestering the transporter inside the cell, although not in an insulin-sensitive pool. In contrast, the COOH-terminal 30 amino acids of GLUT4 are sufficient for its correct localization to an intraceUular storage pool which translocates to the cell surface in response to insulin. The dileucine motif within this domain, which is required for intracellular sequestration of chimeric transporters in fibroblasts, is not critical for targeting to the hormone-responsive compartment in adipocytes. Analysis of rates of internalization of chimeric transporter after the removal of insulin from cells, as well as the subcellular distribution of transporters in cells unexposed to or treated with insulin, leads to a three-pool model which can account for the data. LUCOSE uptake into mammalian cells is accom-plished by a family of proteins, the facilitative glucose transporters, which differ in their tissue distribution and physiological roles. Two of the isoforms identified to date, GLUT1 and GLUT4, are expressed in adipose and muscle, those tissues which exhibit a marked increase in glucose uptake in response to insulin (5). The differential localization of these isoforms within the cell may contribute to their distinct functions. GLUT1, which is expressed in virtually all tissues, is distributed to both the plasma membrane and the interior of the cell in the basal state. Insulin causes a two-to fivefold increase in the amount of GLUT1 present at the cell surface, a recruitment similar to other recycling membrane proteins such as the insulin-like growth factor II and transferrin receptors (39). GLUT4, which is expressed exclusively in insulinresponsive cell types, is excluded from the plasma membrane and instead localizes to an intracellular storage pool in the basal state. Insulin stimulates the specific recruitment of these vesicles to the cell surface, increasing by 10-to 40-fold the amount of GLUT4 present at the cell surface, and thereby dramatically increasing the hexose uptake capacity of the cell (8,9,21,24,34,(36)(37)(38)49 The mechanisms responsible for the insulin-regulated movement of GLUT4 in adipose and muscle may be similar to those utilized by other cell types for regulated secretion. This is supported by the ability of GLUT4 to segregate to large dense core vesicles when expressed in the neuroendocrine cell line PC12 (23). The involvement ...

Immunocytochemical Evidence that GLUT4 Resides in a Specialized Translocation Post-endosomal VAMP2-positive Compartment in Rat Adipose Cells in the Absence of Insulin

“…This route is consistent with the model of two or more intracellular pools in series responsible for GLUT4 sequestration Verhey et al 1995). Studies are now in progress in our laboratory using this immunocytochemical approach to investigate insulin-modulated steps along the GLUT4 trafficking pathway.…”

GLUT1 Glucose Transporter in the Lactating Mammary Gland in the Rat.

“…Insulin-recruitable glucose transporter GLUT4 resides in the intracellular vesicles and is translocated to the plasma membrane upon insulin stimulation in adipocytes and muscle cells [3,16]. Expression experiments of glucose transporters and their mutated molecules in cultured cells suggest that such differential localization of glucose transporters of GLUT family is governed at least by the primary structure of each isoform [8,11,14,29].…”

p115 Interacts with the GLUT4 Vesicle Protein, IRAP, and Plays a Critical Role in Insulin-stimulated GLUT4 Translocation

“…IRAP has a single transmembrane domain with the Nterminus projecting into the cytosol (Keller et al, 1995), whereas GLUT4 has potential sorting and targeting motifs in three cytosolic domains corresponding to N-and C-termini as well as the central loop that connects helices 6 and 7 (Fukumoto et al, 1989). The N-terminus of IRAP has dileucine motifs and an acidic cluster domain similar to that found in the C-terminus of GLUT4, which is thought to be important region for its trafficking (Verhey et al, 1993(Verhey et al, , 1995Corvera et al, 1994;Verhey and Birnbaum, 1994;Haney et al, 1995;Marsh et al, 1995). In this study, we used the N-terminal cytoplasmic domain of IRAP, residues 1-109, conjugated to a chitin-binding protein in order to find cytosolic proteins involved in GSV trafficking, and we identified p115 as one such protein.…”