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The Journal of Cell Biology, Vol 108, 1537-1545, Copyright © 1989 by The Rockefeller University Press

ARTICLES

Localization of transferrin receptors and insulin-like growth factor II receptors in vesicles from 3T3-L1 adipocytes that contain intracellular glucose transporters

LI Tanner and GE Lienhard
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03756.

Transferrin receptors in detergent extracts of subcellular membrane fractions prepared from 3T3-L1 adipocytes were measured by a binding assay. There was a small but significant increase (1.2-fold) in the amount of receptor in a crude plasma membrane fraction and a 40% decrease in the number of transferrin receptors in microsomal membranes prepared from insulin-treated cells, when compared with corresponding fractions from control cells. Intracellular vesicles containing insulin- responsive glucose transporters (GT) have been isolated by immunoadsorption from the microsomal fraction (Biber, J. W., and G. E. Lienhard. 1986. J. Biol. Chem. 261:16180-16184). All of the transferrin receptors in this fraction were localized in these vesicles; however, because the GT vesicles contain approximately 30-fold fewer transferrin receptors than GT, on the average only one vesicle in three contains a transferrin receptor. The binding of 125I-pentamannose 6-phosphate BSA to 3T3-L1 adipocytes at 4 degrees C was used to monitor surface insulin- like growth factor II (IGF-II)/mannose 6-phosphate receptors. Exposure of cells to insulin at 37 degrees C for 5 min resulted in a 2.5-4.5- fold increase in surface receptors. There was a corresponding 20% decrease in the amount of IGF-II receptors in the microsomal membranes prepared from insulin-treated cells, as assayed by immunoblotting. Moreover, the IGF-II receptors and GT were located in the same intracellular vesicles, since antibodies to the carboxyterminal peptide of either protein immunoadsorbed vesicles containing 70-95% of both proteins initially present in the microsomal fraction. In conjunction with other studies, these results indicate that in 3T3-L1 adipocytes, three membrane proteins (the GT, the transferrin receptor, and the IGF- II receptor) respond similarly to insulin, by redistributing to the surface from intracellular compartment(s) in which they are colocalized.
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