Dependence of insulin-stimulated glucose transporter 4 translocation on 3-phosphoinositide-dependent protein kinase-1 and its target threonine-410 in the activation loop of protein kinase C-ζ
Dependence of insulin-stimulated glucose transporter 4 translocation on 3-phosphoinositide-dependent protein kinase-1 and its target threonine-410 in the activation loop of protein kinase C-ζ
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DOI：
10.1210/me.13.10.1766
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发表时间：
1999-10-01
影响因子：
--
通讯作者：
Farese, RV
中科院分区：
文献类型：
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作者：
Bandyopadhyay, G;Standaert, ML;Farese, RV
以前的研究表明：1）非典型蛋白激酶C（PKC）亚型是胰岛素刺激葡萄糖转运所必需的; 2）3-磷酸肌醇依赖性蛋白激酶-1（PDK-1）是非典型PKCa激活所必需的。目前，我们评估了PDK-1在瞬时转染大鼠脂肪细胞中胰岛素作用过程中PKC-ζ活化和表位标记的葡萄糖转运蛋白4（GLUT 4）向质膜易位中的作用。野生型PDK-1的过表达引起共转染的血凝素（HA）标记的PKC-zeta的活性增加，同时增强HA标记的GLUT 4易位。表达激酶失活的PDK-1和在Thr-410（PKC-zeta激活环中PDK-1的直接靶点）突变的PKC-zeta的抗激活形式，抑制胰岛素诱导的HA-PKC-zeta活性和HA-GLUT 4易位增加的程度与激酶失活的PKC-zeta相同。此外，激酶失活的PDK-1的抑制作用被野生型PDK-1的共转染完全逆转，并且被野生型PKC-zeta部分逆转，但不被野生型PKB逆转。与T410 A PKC-zeta突变体相反，对PDK-1活化具有抗性的PKB的类似双突变体（T308 A/S473 A）对胰岛素刺激的HA-GLUT 4易位仅具有很小的影响，并且不抑制由野生型PDK-1过表达诱导的HA-GLUT 4易位。我们的研究结果表明，PDK-1及其下游靶点，PKC-zeta激活环中的Thr-410，是胰岛素刺激的葡萄糖转运所必需的。
Previous studies have suggested that 1) atypical protein kinase C (PKC) isoforms are required for insulin stimulation of glucose transport; and 2) 3-phosphoinositide-dependent protein kinase-1 (PDK-1) is required for activation of atypical PKCa. presently, we evaluated the role of PDK-1, both in the activation of PKC-zeta, and the translocation of epitope-tagged glucose transporter 4 (GLUT4) to the plasma membrane, during insulin action in transiently transfected rat adipocytes. Overexpression of wild-type PDK-1 provoked increases in the activity of cotransfected hemagglutinin (HA)-tagged PKC-zeta and concomitantly enhanced HA-tagged GLUT4 translocation. Expression of both kinase-inactive PDK-1 and an activation-resistant form of PKC-zeta that is mutated at Thr-410, the immediate target of PDK-1 in the activation loop of PKC-zeta, inhibited insulin-induced increases in both HA-PKC-zeta activity and HA-GLUT4 translocation to the same extent as kinase-inactive PKC-zeta. Moreover, the inhibitory effects of kinase-inactive PDK-1 were fully reversed by cotransfection of wild-type PDK-1 and partly reversed by wild-type PKC-zeta but not by wild-type PKB. In contrast to the T410A PKC-zeta mutant, an analogous double mutant of PKB (T308A/S473A) that is resistant to PDK-1 activation had only a small effect on insulin-stimulated HA-GLUT4 translocation and did not inhibit HA-GLUT4 translocation induced by overexpression of wild-type PDK-1. Our findings suggest that both PDK-1 and its downstream target, Thr-410 in the activation loop of PKC-zeta, are required for insulin-stimulated glucose transport.