Js. Bogan et al., Insulin-responsive compartments containing GLUT4 in 3T3-L1 and CHO cells: Regulation by amino acid concentrations, MOL CELL B, 21(14), 2001, pp. 4785-4806
In fat and muscle, insulin stimulates glucose uptake by rapidly mobilizing
the GLUT4 glucose transporter from a specialized intracellular compartment
to the plasma membrane. We describe a method to quantify the relative propo
rtion of GLUT4 at the plasma membrane, using flow cytometry to measure a ra
tio of fluorescence intensities corresponding to the cell surface and total
amounts of a tagged GLUT4 reporter in individual living cells. Using this
assay, we demonstrate that both 3T3-L1 and CHO cells contain intracellular
compartments from which GLUT4 is rapidly mobilized by insulin and that the
initial magnitude and kinetics of redistribution to the plasma membrane are
similar in these two cell types when they are cultured identically. Target
ing of GLUT4 to a highly insulin responsive compartment in CHO cells is mod
ulated by culture conditions. In particular, we find that amino acids regul
ate distribution of GLUT4 to this kinetically defined compartment through a
rapamycin-sensitive pathway. Amino acids also modulate the magnitude of in
sulin-stimulated translocation in 3T3-L1 adipocytes. Our results indicate a
novel link between glucose and amino acid metabolism.