Salt I P, Johnson G, Ashcroft S J, Hardie D G
Biochemistry Department, MSI/WTB Complex, Dow Street, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Biochem J. 1998 Nov 1;335 ( Pt 3)(Pt 3):533-9. doi: 10.1042/bj3350533.
The role of the AMP-activated protein kinase (AMPK) cascade in the glucose-sensitive pancreatic beta cell lines HIT-T15 and INS-1 was addressed. In both cell types, removal of glucose leads to a >5-fold activation of AMPK activity. Activation of AMPK was due to phosphorylation, since the effect was reversed by protein phosphatase treatment of the extracts, and was restored by re-addition of MgATP and the purified upstream kinase. When the effects of different concentrations of medium glucose were examined, insulin secretion and AMPK activity were inversely related, and varied over the same concentration range. The activation in response to glucose removal appeared to be due to changes in the concentration of the known regulators of the cascade, i.e. AMP and ATP, since AMPK activation was associated with a large increase in the cellular AMP/ATP ratio, and the two parameters varied over the same range of glucose concentrations. In late-passage HIT-T15 cells that had lost the glucose-dependent insulin secretion response, both AMPK activity and the AMP/ATP ratio also became insensitive to the extracellular glucose concentration. Treatment of INS-1 cells, but not HIT-T15 cells, with AICA riboside (5-aminoimidazole-4-carboxamide riboside) results in accumulation of the ribotide, ZMP (AICA riboside monophosphate), and activation of AMPK. AICA riboside treatment of INS-1 cells, and of isolated rat islets, had both inhibitory and stimulatory effects on insulin secretion. These results show that in beta cell lines the AMP-activated protein kinase, like its yeast homologue the SNF1 complex, can respond to the level of glucose in the medium, and may be involved in regulating insulin release.
研究了AMP激活的蛋白激酶(AMPK)级联在葡萄糖敏感的胰腺β细胞系HIT-T15和INS-1中的作用。在这两种细胞类型中,去除葡萄糖会导致AMPK活性激活超过5倍。AMPK的激活是由于磷酸化,因为用蛋白磷酸酶处理提取物可逆转该效应,并且通过重新添加MgATP和纯化的上游激酶可恢复该效应。当检测不同浓度的培养基葡萄糖的作用时,胰岛素分泌与AMPK活性呈负相关,并且在相同浓度范围内变化。对去除葡萄糖的反应性激活似乎是由于级联反应的已知调节因子即AMP和ATP浓度的变化,因为AMPK激活与细胞内AMP/ATP比值的大幅增加相关,并且这两个参数在相同的葡萄糖浓度范围内变化。在已失去葡萄糖依赖性胰岛素分泌反应的传代后期HIT-T15细胞中,AMPK活性和AMP/ATP比值也对细胞外葡萄糖浓度变得不敏感。用AICA核苷(5-氨基咪唑-4-甲酰胺核苷)处理INS-1细胞而非HIT-T15细胞会导致核苷酸ZMP(AICA核苷单磷酸)积累并激活AMPK。用AICA核苷处理INS-1细胞和分离的大鼠胰岛对胰岛素分泌具有抑制和刺激作用。这些结果表明,在β细胞系中,AMP激活的蛋白激酶与其酵母同源物SNF1复合物一样,可以对培养基中的葡萄糖水平作出反应,并且可能参与调节胰岛素释放。