Regulation of insulin sensitivity by subcellular GLUT4 targeting

SUMO conjugating enzyme Ubc9 has been shown to upregulate GLUT4 in muscle cells although the mechanism of action is unknown. We investigated the physiological significance of Ubc9 in GLUT4 turnover and subcellular targeting by adenovirus vector-mediated overexpression and by siRNA-mediated gene silencing of Ubc9 in 3T3-L1 adipocytes. Overexpression of Ubc9 resulted in inhibition of GLUT4 degradation and promotion of its targeting to the non-endosomal, non-TGN GLUT4 storage vesicles (GSV), leading to an up-regulation of GLUT4 expression level and insulin-responsive glucose transport. While long-term insulin stimulation caused GLUT4 down-regulation by 40-50%, which was inhibited with lysosomal inhibitors and was associated with a selective reduction in GLUT4 in GSV, overexpression of Ubc9 antagonized these long-term effects of insulin. By contrast, Ubc9 gene silencing with siRNA caused a marked decrease in the GLUT4 level, whereas overexpression of the catalytically inactive mutant of Ubc9 increased GLUT4 and insulin-stimulated glucose transport to the level comparable to that with wild-type Ubc9. These results suggest that Ubc9 up-regulates GLUT4 by inhibition of lysosomal sorting and promotes GLUT4 targeting to GSV by a mechanism unrelated to its catalytic activity. Thus, Ubc9 plays an indispensable role in the expression and maintenance of the insulin-sensitive glucose transport system in adipocytes.