A study of the SUMO-conjugating enzyme Ubc9 as a novel regulatory factor of insulin sensitivity

SUMO缀合酶Ubc 9已被证明上调L 6成肌细胞中的GLUT 4，尽管作用机制尚不清楚。在这里，我们研究了Ubc 9在3 T3-L1脂肪细胞中GLUT 4周转和亚细胞靶向腺病毒载体介导的过表达和siRNA介导的Ubc 9基因沉默的生理意义。Ubc 9的过表达导致GLUT 4降解的抑制和其靶向GLUT 4储存室（GSC）的促进，导致GLUT 4水平和胰岛素应答性GLUT 4易位和葡萄糖转运的增加。虽然长期（6小时或更长时间）胰岛素刺激导致GLUT 4下调40- 50%，但溶酶体抑制剂可抑制该下调，并与GSC中GLUT 4的选择性降低相关，Ubc 9的过表达可拮抗胰岛素的这些作用。相比之下，Ubc 9基因沉默与siRNA显着加速胰岛素诱导的GLUT 4下调，而过表达的催化失活突变体Ubc 9-C93 A增加GLUT 4和胰岛素刺激的葡萄糖转运的水平与野生型Ubc 9。这些结果表明，Ubc 9通过抑制溶酶体分选上调GLUT 4，并通过与其催化活性无关的机制促进GLUT 4靶向GSC。因此，Ubc 9在脂肪细胞胰岛素敏感性葡萄糖转运系统的表达和维持中起着不可或缺的作用。

SUMO conjugating enzyme Ubc9 has been shown to upregulate GLUT4 in L6 myoblasts although the mechanism of action is undefined. Here 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 GLUT4 storage compartment (GSC), leading to an increase in GLUT4 level and insulinresponsive GLUT4 translocation and glucose transport. While long-term (6 hours and more) insulin stimulation caused GLUT4 downregulation by 40-50%, which was inhibited with lysosomal inhibitors and was associated with a selective reduction in GLUT4 in GSC, overexpression of Ubc9 antagonized these effects of insulin. By contrast, Ubc9 gene silencing with siRNA markedly accelerated the insulin-induced GLUT4 downregulation, whereas overexpression of the catalytically inactive mutant Ubc9-C93A increased GLUT4 and insulin-stimulated glucose transport to the level comparable to that with wild-type Ubc9. These results suggest that Ubc9 upregulates GLUT4 by inhibition of lysosomal sorting and promotes GLUT4 targeting to GSC by a mechanism unrelated to its catalytic activity. Thus, Ubc9 plays a indispensable role in expression and maintenance of the insulin-sensitive glucose transport system in adipocytes.