Iwaoka Ryo, Kataoka Kohsuke
From the Laboratory of Molecular Medical Bioscience, Graduate School of Medical Life Science, Yokohama City University, 1-7-29 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan.
From the Laboratory of Molecular Medical Bioscience, Graduate School of Medical Life Science, Yokohama City University, 1-7-29 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan
J Biol Chem. 2018 Mar 9;293(10):3524-3534. doi: 10.1074/jbc.M117.817932. Epub 2018 Jan 18.
Insulin mRNA expression in pancreatic islet β-cells is up-regulated by extracellular glucose concentration, but the underlying mechanism remains incompletely understood. MafA is a transcriptional activator specifically enriched in β-cells that binds to the insulin gene promoter. Its expression is transcriptionally and posttranscriptionally regulated by glucose. Moreover, AMP-activated protein kinase (AMPK), a regulator of cellular energy homeostasis, is inhibited by high glucose, and this inhibition is essential for the up-regulation of insulin gene expression and glucose-stimulated insulin secretion (GSIS). Here we mutagenized the insulin promoter and found that the MafA-binding element C1/RIPE3b is required for glucose- or AMPK-induced alterations in insulin gene promoter activity. Under high-glucose conditions, pharmacological activation of AMPK in isolated mouse islets or MIN6 cells by metformin or 5-aminoimidazole-4-carboxamide riboside decreased MafA protein levels and mRNA expression of insulin and GSIS-related genes ( and ). Overexpression of constitutively active AMPK also reduced MafA and insulin expression. Conversely, pharmacological AMPK inhibition by dorsomorphin (compound C) or expression of a dominant-negative form of AMPK increased MafA and insulin expression under low-glucose conditions. However, AMPK activation or inhibition did not change the expression levels of the β-cell-enriched transcription factors Pdx1 and Beta2/NeuroD1. AMPK activation accelerated MafA protein degradation, which is not dependent on the proteasome. We also noted that MafA overexpression prevents metformin-induced decreases in insulin and GSIS-related gene expression. These findings indicate that high glucose concentrations inhibit AMPK, thereby increasing MafA protein levels and activating the insulin promoter.
胰岛β细胞中的胰岛素mRNA表达受细胞外葡萄糖浓度上调，但其潜在机制仍未完全阐明。MafA是一种特异性富集于β细胞的转录激活因子，可与胰岛素基因启动子结合。其表达受葡萄糖的转录和转录后调控。此外，细胞能量稳态的调节因子AMP激活的蛋白激酶（AMPK）受高葡萄糖抑制，这种抑制对于胰岛素基因表达上调和葡萄糖刺激的胰岛素分泌（GSIS）至关重要。在此，我们对胰岛素启动子进行诱变，发现MafA结合元件C1/RIPE3b是葡萄糖或AMPK诱导的胰岛素基因启动子活性改变所必需的。在高葡萄糖条件下，通过二甲双胍或5-氨基咪唑-4-甲酰胺核苷对分离的小鼠胰岛或MIN6细胞中的AMPK进行药理激活，可降低MafA蛋白水平以及胰岛素和GSIS相关基因的mRNA表达（以及）。组成型活性AMPK的过表达也降低了MafA和胰岛素表达。相反，在低葡萄糖条件下，通过dorsomorphin（化合物C）对AMPK进行药理抑制或表达AMPK的显性负性形式可增加MafA和胰岛素表达。然而，AMPK的激活或抑制并未改变β细胞富集转录因子Pdx1和Beta2/NeuroD1的表达水平。AMPK激活加速了MafA蛋白降解，这不依赖于蛋白酶体。我们还注意到，MafA过表达可防止二甲双胍诱导的胰岛素和GSIS相关基因表达降低。这些发现表明，高葡萄糖浓度抑制AMPK，从而增加MafA蛋白水平并激活胰岛素启动子。
