The role of calcium/calmodulin-dependent protein kinase cascade in glucose upregulation of insulin gene expression
The role of calcium/calmodulin-dependent protein kinase cascade in glucose upregulation of insulin gene expression
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DOI：
10.2337/diabetes.53.6.1475
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发表时间：
2004-06-01
期刊：
影响因子：
7.7
通讯作者：
Ishida, T
中科院分区：
文献类型：
--
作者：
Yu, X;Murao, K;Ishida, T
据报道，许多因素影响β细胞中的胰岛素合成。虽然葡萄糖是胰腺β细胞中胰岛素基因表达的最重要调节剂，但葡萄糖刺激胰岛素基因转录以响应葡萄糖浓度变化的机制尚未阐明。在这项研究中，我们研究了Ca 2 +/钙调蛋白（CaM）依赖性蛋白激酶（CaM-K）级联反应在胰岛素转录激活中的作用。RT-PCR、Western blotting和免疫组化染色分析显示，CaM-K激酶-α（CaM-KKa）和CaM-KIV定位于大鼠胰腺β细胞及其细胞系INS-1。INS-1细胞暴露于11.2 mmol/l葡萄糖引起胰岛素启动子活性的增加以及刺激后2 min内CaM-KIV活性的上调。我们研究了在转染的INS-1细胞中CaM-KIV的组成型活性形式（CaM-KIVc）或显性阴性突变体（CaM-KIVdn）对胰岛素启动子活性的影响。单独的CaM-KIVc是足够的，并且上游激酶CaM-KK被增强以上调INS-1细胞中的胰岛素启动子活性。此外，CaM-KIVdn的共转染抑制了胰岛素启动子的葡萄糖上调活性的显着程度。综上所述，这些结果表明，CaM-KK/CaM-KIV级联可能在葡萄糖上调的胰岛素基因转录激活中起重要作用。
A number of factors have been reported to affect insulin synthesis in beta-cells. Although glucose is the most important regulator of insulin gene expression in pancreatic beta-cells, the mechanisms whereby glucose stimulates insulin gene transcription in response to changes in glucose concentration have not been clarified yet. In this study, we examined the role of the Ca2+/calmodulin (CaM)-dependent protein kinase (CaM-K) cascade in transcriptional activation of insulin. RT-PCR, Western blotting, and immunohistochemical staining analysis revealed that CaM-K kinase-alpha (CaM-KKalpha) and CaM-KIV were localized in rat pancreatic beta-cells and their cell line, INS-1. Exposure of INS-1 cells to 11.2 mmol/l glucose elicited an increase of insulin promoter activity as well as upregulation of CaM-KIV activity within 2 min after stimulation. We investigated the influence on insulin promoter activity of the constitutively active form (CaM-KIVc) or dominant-negative mutant (CaM-KIVdn) of CaM-KIV in transfected INS-1 cells. CaM-KIVc alone was sufficient, and the upstream kinase, CaM-KK, was enhanced to upregulate the insulin promoter activity in INS-1 cells. Furthermore, cotransfection of CaM-KIVdn suppressed to a significant degree the glucose-upregulated activity of the insulin promoter. Taken together, these results indicated that the CaM-KK/CaM-KIV cascade might play an important role in glucose-upregulated transcriptional activation of the insulin gene.