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.
Kao
A W
AW
Department of Physiology and Biophysics, University of Iowa, Iowa City, Iowa 52242, USA.
Noda
Y
Y
Johnson
J H
JH
Pessin
J E
JE
Saltiel
A R
AR
D016475:3T3 Cells; D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D003847:Deoxyglucose / Q000494:pharmacology; D004705:Endocytosis; D005089:Exocytosis; D005455:Fluorescent Antibody Technique; D005634:Fructose-Bisphosphate Aldolase / Q000276:immunology / Q000378:metabolism*; D005635:Fructosediphosphates / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism; D051275:Glucose Transporter Type 4; D005986:Glyceraldehyde 3-Phosphate / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology*; D051379:Mice; D008845:Microinjections; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D010446:Peptide Fragments / Q000378:metabolism; D011993:Recombinant Fusion Proteins
D000199:Actins; D000906:Antibodies; D005635:Fructosediphosphates; D051275:Glucose Transporter Type 4; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C495693:Slc2a4 protein, mouse; D005986:Glyceraldehyde 3-Phosphate; D003847:Deoxyglucose; D005634:Fructose-Bisphosphate Aldolase; D005947:Glucose; C029063:fructose-1,6-diphosphate