Aldolase Mediates the Association of F-actin with the Insulin-responsive Glucose Transporter GLUT4*

To identify potential proteins interacting with the insulin-responsive glucose transporter (GLUT4), we generated fusion proteins of glutathione S-transferase (GST) and the final 30 amino acids from GLUT4 (GST-G4) or GLUT1 (GST-G1). Incubation of these carboxyl-terminal fusion proteins with adipocyte cell extracts revealed a specific interaction of GLUT4 with fructose 1,6-bisphosphate aldolase. In the presence of aldolase, GST-G4 but not GST-G1 was able to co-pellet with filamentous (F)-actin. This interaction was prevented by incubation with the aldolase substrates, fructose 1,6-bisphosphate or glyceraldehyde 3-phosphate. Immunofluorescence confocal microscopy demonstrated a significant co-localization of aldolase and GLUT4 in intact 3T3L1 adipocytes, which decreased following insulin stimulation. Introduction into permeabilized 3T3L1 adipocytes of fructose 1,6-bisphosphate or the metabolic inhibitor 2-deoxyglucose, two agents that disrupt the interaction between aldolase and actin, inhibited insulin-stimulated GLUT4 exocytosis without affecting GLUT4 endocytosis. Furthermore, microinjection of an aldolase-specific antibody also inhibited insulin-stimulated GLUT4 translocation. These data suggest that aldolase functions as a scaffolding protein for GLUT4 and that glucose metabolism may provide a negative feedback signal for the regulation of glucose transport by insulin.

[1]  J. Pessin,et al.  Molecular Basis of Insulin-stimulated GLUT4 Vesicle Trafficking , 1999, The Journal of Biological Chemistry.

[2]  J. Pessin,et al.  Expression of a Dominant Interfering Dynamin Mutant in 3T3L1 Adipocytes Inhibits GLUT4 Endocytosis without Affecting Insulin Signaling* , 1998, The Journal of Biological Chemistry.

[3]  A. Klip,et al.  Actin filaments participate in the relocalization of phosphatidylinositol3-kinase to glucose transporter-containing compartments and in the stimulation of glucose uptake in 3T3-L1 adipocytes. , 1998, The Biochemical journal.

[4]  D. James,et al.  Moving GLUT4: The Biogenesis and Trafficking of GLUT4 Storage Vesicles , 1997, Diabetes.

[5]  D. Chen,et al.  Osmotic Shock Stimulates GLUT4 Translocation in 3T3L1 Adipocytes by a Novel Tyrosine Kinase Pathway* , 1997, The Journal of Biological Chemistry.

[6]  C. Jung,et al.  A Synthetic Peptide Corresponding to the GLUT4 C-terminal Cytoplasmic Domain Causes Insulin-like Glucose Transport Stimulation and GLUT4 Recruitment in Rat Adipocytes* , 1997, The Journal of Biological Chemistry.

[7]  K. Hori,et al.  Mode of interactions of human aldolase isozymes with cytoskeletons. , 1997, Archives of biochemistry and biophysics.

[8]  J. Pessin,et al.  Syntaxin 4, VAMP2, and/or VAMP3/cellubrevin are functional target membrane and vesicle SNAP receptors for insulin-stimulated GLUT4 translocation in adipocytes , 1997, Molecular and cellular biology.

[9]  M. Mueckler,et al.  Insulin unmasks a COOH-terminal Glut4 epitope and increases glucose transport across T-tubules in skeletal muscle , 1996, The Journal of cell biology.

[10]  Amy Johnson,et al.  The Carboxyl Terminus of GLUT4 Contains a Serine-Leucine-Leucine Sequence That Functions as a Potent Internalization Motif in Chinese Hamster Ovary Cells* , 1996, The Journal of Biological Chemistry.

[11]  A. J. Morris,et al.  The Molecular Nature of the F-actin Binding Activity of Aldolase Revealed with Site-directed Mutants (*) , 1996, The Journal of Biological Chemistry.

[12]  A. Klip,et al.  Disassembly of the actin network inhibits insulin-dependent stimulation of glucose transport and prevents recruitment of glucose transporters to the plasma membrane. , 1994, The Journal of biological chemistry.

[13]  R. Chakrabarti,et al.  A double leucine within the GLUT4 glucose transporter COOH-terminal domain functions as an endocytosis signal [published erratum appears in J Cell Biol 1994 Sep;126(6):1625] , 1994, The Journal of cell biology.

[14]  D. James,et al.  The amino terminus of GLUT4 functions as an internalization motif but not an intracellular retention signal when substituted for the transferrin receptor cytoplasmic domain , 1994, The Journal of cell biology.

[15]  M. Birnbaum,et al.  A Leu-Leu sequence is essential for COOH-terminal targeting signal of GLUT4 glucose transporter in fibroblasts. , 1994, The Journal of biological chemistry.

[16]  H. Murata,et al.  Domains that confer intracellular sequestration of the Glut4 glucose transporter in Xenopus oocytes. , 1993, The Journal of biological chemistry.

[17]  M. Birnbaum,et al.  Identification of the carboxy terminus as important for the isoform- specific subcellular targeting of glucose transporter proteins , 1993, The Journal of cell biology.

[18]  A. Chawla,et al.  Exofacial epitope-tagged glucose transporter chimeras reveal COOH- terminal sequences governing cellular localization , 1993, The Journal of cell biology.

[19]  J. Slot,et al.  GLUT-4 NH2 terminus contains a phenylalanine-based targeting motif that regulates intracellular sequestration , 1993, The Journal of cell biology.

[20]  M. Czech,et al.  Insulin action on the internalization of the GLUT4 glucose transporter in isolated rat adipocytes. , 1993, The Journal of biological chemistry.

[21]  F. Clarke,et al.  Identification of an actin binding region in aldolase , 1993, FEBS letters.

[22]  G. Holman,et al.  Comparison of GLUT4 and GLUT1 subcellular trafficking in basal and insulin-stimulated 3T3-L1 cells. , 1993, The Journal of biological chemistry.

[23]  B. Jhun,et al.  Effects of insulin on steady state kinetics of GLUT4 subcellular distribution in rat adipocytes. Evidence of constitutive GLUT4 recycling. , 1992, The Journal of biological chemistry.

[24]  D. Taylor,et al.  2-Deoxyglucose and cytochalasin D modulate aldolase mobility in living 3T3 cells , 1992, The Journal of cell biology.

[25]  D. S. Harris,et al.  Translocation of the glucose transporter (GLUT4) to the cell surface in permeabilized 3T3-L1 adipocytes: effects of ATP insulin, and GTP gamma S and localization of GLUT4 to clathrin lattices , 1992, The Journal of cell biology.

[26]  R. Beitner,et al.  Insulin rapidly stimulates binding of phosphofructokinase and aldolase to muscle cytoskeleton. , 1992, The International journal of biochemistry.

[27]  H. Lodish,et al.  Immunoelectron microscopic demonstration of insulin-stimulated translocation of glucose transporters to the plasma membrane of isolated rat adipocytes and masking of the carboxyl-terminal epitope of intracellular GLUT4. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[28]  G I Bell,et al.  Cloning and characterization of the major insulin-responsive glucose transporter expressed in human skeletal muscle and other insulin-responsive tissues. , 1989, The Journal of biological chemistry.

[29]  M. Birnbaum.  Identification of a novel gene encoding an insulin-responsive glucose transporter protein , 1989, Cell.

[30]  H. Lodish,et al.  A glucose transport protein expressed predominately in insulin-responsive tissues. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[31]  D. James,et al.  Molecular cloning and characterization of an insulin-regulatable glucose transporter , 1989, Nature.

[32]  D. Taylor,et al.  Aldolase exists in both the fluid and solid phases of cytoplasm [published erratum appears in J Cell Biol 1988 Dec;107(6 Pt 1):following 2463] , 1988, The Journal of cell biology.

[33]  M. Kasuga,et al.  Phosphatidylinositol 3-kinase acts at an intracellular membrane site to enhance GLUT4 exocytosis in 3T3-L1 cells. , 1996, The Biochemical journal.

[34]  J. D. Pardee,et al.  [18] Purification of muscle actin , 1982 .