Suppressed intrinsic catalytic activity of GLUT1 glucose transporters in insulin-sensitive 3T3-L1 adipocytes.
Previous studies indicated that the erythroidtype (GLUT1) glucose transporter isoform contributes to basal but not insulin-stimulated hexose transport in mouse 3T3-L1 adipocytes. In the present studies it was found that basal hexose uptake in 3T3-L1 adipocytes was about 50% lower than that in 3T3-L1 or CHO-K1 fibroblasts. Intrinsic catalytic activities of GLUT1 transporters in CHO-K1 and 3T3-L1 cells were compared by normalizing these hexose transport rates to GLUT1 content on the cell surface, as measured by two independent methods. Cell surface GLUT1 levels in 3T3-L1 fibroblasts and adipocytes were about 10- and 25-fold higher, respectively, than in CHO-K1 fibroblasts, as assessed with an anti-GLUT1 exofacial domain antiserum, delta. The large excess of cell surface GLUT1 transporters in 3T3-L1 adipocytes relative to CHO-K1 fibroblasts was confirmed by GLUT1 protein immunoblot analysis and by photoaffinity labelling (with 3-[125I]iodo-4-azidophenethylamido-7-O-succinyldeacetylforskoli n) of glucose transporters in isolated plasma membranes. Thus, GLUT1 intrinsic activity is markedly reduced in 3T3-L1 fibroblasts compared with the CHO-K1 fibroblasts, and further reduction occurs upon differentiation to adipocytes. Intrinsic catalytic activities specifically associated with heterologously expressed human GLUT1 protein in transfected CHO-K1 versus 3T3-L1 cells were determined by subtracting appropriate control cell values for hexose transport and delta-antibody binding from those determined in the transfected cells expressing high levels of human GLUT1. The results confirmed a greater than 90% inhibition of the intrinsic catalytic activity of human GLUT1 transporters on the surface of mouse 3T3-L1 adipocytes relative to CHO-K1 fibroblasts. We conclude that a mechanism that markedly suppresses basal hexose transport catalyzed by GLUT1 is a major contributor to the dramatic insulin sensitivity of glucose uptake in 3T3-L1 adipocytes.
Harrison
S A
SA
Program in Molecular Medicine, University of Massachusetts Medical Center, Worcester 01605.
Buxton
J M
JM
Czech
M P
MP
D019325:3-O-Methylglucose; D000273:Adipose Tissue / Q000187:drug effects / Q000378:metabolism*; D000345:Affinity Labels / Q000378:metabolism; D000818:Animals; D000906:Antibodies; D001386:Azides / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005576:Colforsin / Q000031:analogs & derivatives / Q000378:metabolism; D003847:Deoxyglucose / Q000378:metabolism; D004224:Diterpenes; D005347:Fibroblasts / Q000378:metabolism; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008757:Methylglucosides / Q000378:metabolism*; D051379:Mice; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*
D000345:Affinity Labels; D000906:Antibodies; D001386:Azides; D004224:Diterpenes; D007328:Insulin; D008757:Methylglucosides; D009004:Monosaccharide Transport Proteins; C054433:3-iodo-4-azidophenethylamido-7-O-succinyldeacetylforskolin; D019325:3-O-Methylglucose; D005576:Colforsin; D003847:Deoxyglucose