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New cDNAs and proteins phosphorylated by insulin using proteome analysis

使用蛋白质组分析发现胰岛素磷酸化的新 cDNA 和蛋白质

基本信息

批准号：
13680691
负责人：
OBATA Toshiyuki
金额：
$ 2.69万
依托单位：
The University of Tokushima
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680691/
关键词：
insulin action diabetes Akt proteomics インスリンリン酸化タンパク質プロテオーム解析

项目摘要

Insulin plays a central role in the regulation of glucose homeostasis in part by stimulating glucose uptake and glycogen synthesis. The serine/threonine protein kinase Akt has been proposed to mediate insulin signaling in several processes. However, it is unclear if Akt is involved in insulin-stimulated glucose uptake, and whether which isoforms of Akt are responsible for each insulin action. We confirmed that expression of a constitutively active Akt, using an adenoviral expression vector promoted translocation of glucose transporter 4 (GLUT4) to plasma membrane, 2-deoxyglucose (2-DG) uptake and glycogen synthesis in both Chinese hamster ovary (CHO) cells and 3T3-L1 adipocytes. Inhibition of Akt either by adenoviral expression of a dominant-negative Akt or by the introduction of synthetic 21-mer short-interference RNA (siRNA) against Akt markedly reduced insulin-stimulated GLUT4 translocation, 2-DG uptake and glycogen synthesis. Experiments with isoform-specific siRNA revealed that Akt2, and Akt1 to lesser extent, has an essential role in insulin-stimulated GLUT4 translocation and 2-DG uptake in both cell lines, while Akt1 and Akt2 contribute equally to insulin-stimulated glycogen synthesis. These data suggest a prerequisite role of Akt in insulin-stimulated glucose uptake and distinct functions among Akt isoforms.

胰岛素通过刺激葡萄糖摄取和糖原合成，在调节葡萄糖动态平衡方面发挥核心作用。丝氨酸/苏氨酸蛋白激酶Akt被认为在几个过程中介导胰岛素信号传导。然而，目前尚不清楚Akt是否参与了胰岛素刺激的葡萄糖摄取，以及Akt的哪些亚型负责每种胰岛素的作用。我们证实，在中国仓鼠卵巢(CHO)细胞和3T3-L1脂肪细胞中，使用腺病毒表达载体表达具有组成活性的Akt能促进葡萄糖转运蛋白4(GLUT4)向质膜转位、2-脱氧葡萄糖(2-DG)摄取和糖原合成。通过腺病毒表达显性阴性Akt或引入合成的针对Akt的21聚体短干扰RNA(SiRNA)来抑制Akt，可显著减少胰岛素刺激的GLUT4转位、2-DG摄取和糖原合成。用异构体特异性siRNA进行的实验表明，Akt2和Akt1在胰岛素刺激的GLUT4转位和2-DG摄取方面发挥了重要作用，而Akt1和Akt2对胰岛素刺激的糖原合成起到了同等的作用。这些数据表明Akt在胰岛素刺激的葡萄糖摄取中起着先决条件的作用，并在Akt亚型之间具有不同的功能。