AS160 regulates insulin- and contraction-stimulated glucose uptake in mouse skeletal muscle
AS160 regulates insulin- and contraction-stimulated glucose uptake in mouse skeletal muscle
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DOI：
10.1074/jbc.m605461200
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发表时间：
2006-10-20
影响因子：
4.8
通讯作者：
Goodyear, Laurie J.
中科院分区：
文献类型：
--
作者：
Kramer, Henning F.;Witczak, Carol A.;Goodyear, Laurie J.
胰岛素和收缩是GLUT4易位和增加骨骼肌葡萄糖摄取的有效刺激物。我们最近发现Rab gtpase激活蛋白（GAP） AS160是连接胰岛素和小鼠骨骼肌收缩诱导的不同上游信号级联的推定趋同点。在这里，我们通过体内电穿孔技术研究了这些AS160信号事件的功能意义，在小鼠胫骨前肌中过表达野生型和三种AS160突变体：1)AS160突变阻止四个磷酸化akt底物位点（4P）的磷酸化；2) AS160突变使兔GTPase活性（R/K）丧失；3)含有4P和R/K突变的双突变体AS160 （2M）。注射基因一周后，所有AS160亚型的蛋白表达量升高了7倍以上。为了确定AS160对转染肌肉中胰岛素和收缩刺激的葡萄糖摄取的影响，我们分别在静脉注射葡萄糖和原位肌肉收缩后测量了体内[H-3]2-脱氧葡萄糖摄取。在过表达4P突变体AS160的肌肉中，胰岛素刺激的葡萄糖摄取被显著抑制。然而，这种抑制完全被AS160 Rab GAP活性的破坏所阻止。转染4P突变体AS160也显著损害了收缩刺激的葡萄糖摄取，野生型AS160过表达也是如此。相反，过表达缺乏Rab GAP活性的突变AS160导致假肌和收缩刺激肌的增加。这些数据表明，AS160在体内调节胰岛素和收缩刺激的小鼠骨骼肌葡萄糖代谢，突变体AS160对胰岛素和收缩作用的影响并不相同。我们的研究结果直接暗示AS160是骨骼肌葡萄糖摄取的独立刺激因子的关键收敛因子。
Insulin and contraction are potent stimulators of GLUT4 translocation and increase skeletal muscle glucose uptake. We recently identified the Rab GTPase-activating protein ( GAP) AS160 as a putative point of convergence linking distinct upstream signaling cascades induced by insulin and contraction in mouse skeletal muscle. Here, we studied the functional implications of these AS160 signaling events by using an in vivo electroporation technique to overexpress wild type and three AS160 mutants in mouse tibialis anterior muscles: 1) AS160 mutated to prevent phosphorylation on four regulatory phospho-Akt-substrate sites (4P); 2) AS160 mutated to abolish Rab GTPase activity (R/K); and 3) double mutant AS160 containing both 4P and R/K mutations (2M). One week following gene injection, protein expression for all AS160 isoforms was elevated over 7-fold. To determine the effects of AS160 on insulin- and contraction-stimulated glucose uptake in transfected muscles, we measured [H-3]2-deoxyglucose uptake in vivo following intravenous glucose administration and in situ muscle contraction, respectively. Insulin-stimulated glucose uptake was significantly inhibited in muscles overexpressing 4P mutant AS160. However, this inhibition was completely prevented by concomitant disruption of AS160 Rab GAP activity. Transfection with 4P mutant AS160 also significantly impaired contraction-stimulated glucose uptake, as did overexpression of wild type AS160. In contrast, overexpressing mutant AS160 lacking Rab GAP activity resulted in increases in both sham and contraction-stimulated muscles. These data suggest that AS160 regulates both insulin- and contraction-stimulated glucose metabolism in mouse skeletal muscle in vivo and that the effects of mutant AS160 on the actions of insulin and contraction are not identical. Our findings directly implicate AS160 as a critical convergence factor for independent stimulators of skeletal muscle glucose uptake.