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 degrees 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 degrees C and then stripped with glutathione at 0 degrees 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.
Hare
J F
JF
Department of Biochemistry and Molecular Biology, Oregon Health Sciences University, Portland 97201.
Taylor
K
K
D000273:Adipose Tissue / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D000818:Animals; D001692:Biological Transport; D001710:Biotin; D002352:Carrier Proteins / Q000378:metabolism; D002454:Cell Differentiation; D007328:Insulin / Q000378:metabolism*; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D010766:Phosphorylation; D015398:Signal Transduction
D002352:Carrier Proteins; D007328:Insulin; D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; D001710:Biotin