SURFACE-EXPOSED PROTEINS OF 3T3-L1 ADIPOCYTES - IDENTIFICATION OF PHOSPHORYLATED, INSULIN-TRANSLOCATED, AND RECYCLING PROTEINS

Twenty-seven adipocyte-specific, cell surface-exposed proteins were detected by derivatizing undifferentiated and differentiated 3T3-L1 cells with membrane impermeant sulfosuccinimidyl 2-(biotinamido)ethyl-1,3-dithiopropionate at 0 °C. Biotinylated proteins were adsorbed onto streptavidin-agarose, resolved on two-dimensional polyacrylamide gels, and detected by autoradiography or silver staining. Of the surface-exposed proteins specific to adipocytes, three were phosphorylated and seven were glycoproteins that bound to wheat germ agglutinin and eluted with N-acetylglucosamine. Eleven of the adipocyte-specific proteins were bound to streptavidin-agarose after the cells were biotinylated at 20 °C and then stripped with glutathione at 0 °C to isolate plasma membrane proteins that localize to recycling endosomes as well as the cell surface. When insulin-deprived cells were acutely treated with insulin, only a few proteins, including one protein tentatively identified as the GLUT4 glucose transporter, were found to increase in concentration at the cell surface. These latter results imply that up-regulation of glucose transport by the translocation of GLUT4 to the cell surface in response to insulin occurs by exocytic fusion of an intracellular compartment having a limited number of proteins. © 1992.