IDENTIFICATION OF THE CARBOXY-TERMINUS AS IMPORTANT FOR THE ISOFORM-SPECIFIC SUBCELLULAR TARGETING OF GLUCOSE-TRANSPORTER PROTEINS

Differential trafficking of glucose transporters contributes significa ntly to the establishment of a cell's capacity for hormone-regulatable hexose uptake. In the true insulin-sensitive peripheral target tissue s, muscle and adipose, the transporter isoform GLUT1 residues on the c ell surface and interior of the cell whereas the highly homologous iso form GLUT4 displays virtually exclusive intracellular sequestration, a llowing the latter to redistribute to the cell surface in response to hormone. These patterns are equally pronounced in cells into which the transporters have been introduced by DNA-mediated gene transfer, sugg esting that signals for isoform-specific sorting are recognized in div erse cell types. To determine the primary sequences responsible for th e characteristic distributions, chimeric transporters were constructed in which reciprocal domains were exchanged between GLUT1 and GLUT4. I n addition, a non-disruptive, species-specific epitope ''tag'' was int roduced into a neutral region of the transporter to allow analysis of reciprocal chimeras using a single antibody. These recombinant transpo rters were stably expressed in HIH 3T3 and PC12 cells by retrovirus-me diated gene transfer, and were localized by indirect immunofluorescenc e and laser scanning confocal microscopy, as well as by staining of pl asma membrane sheets prepared from these cells. The results indicate t hat the carboxy-terminal 30 amino acids are primarily responsible for the differential targeting of the glucose transporter isoforms GLUT1 a nd GLUT4, though there is a lesser additional contribution by the amin o-terminal 183 amino acids.