Skelly R H, Schuppin G T, Ishihara H, Oka Y, Rhodes C J
E.P. Joslin Research Laboratory, Boston, MA 02215, USA.
Diabetes. 1996 Jan;45(1):37-43. doi: 10.2337/diab.45.1.37.
In the short term (< 2 h), proinsulin biosynthesis is predominately glucose regulated at the translational level; however, the details at the molecular level behind this mechanism are not well defined. One of the major hindrances for gaining a better understanding of the proinsulin biosynthetic mechanism has been a lack of an abundant source of beta-cells that express a phenotype of regulated proinsulin biosynthesis in the appropriate 2.8-16.7 mmol/l glucose range as defined in normal pancreatic islets. In this study, we demonstrate that in the MIN6 cell line, specific glucose-regulated translational control of proinsulin biosynthesis is present in the appropriate glucose concentration range. In addition to that of proinsulin, the biosynthesis of the two proinsulin conversion endopeptidases, PC2 and PC3, was coordinately glucose regulated in MIN6 cells, whereas that of the exopeptidase, carboxypeptidase H, was unaffected by glucose. Proinsulin, PC2 and PC3 biosynthesis was specifically stimulated over that of total MIN6 cell protein synthesis above a threshold of 4 mmol/l glucose that reached a maximum rate between 8 and 10 mmol/l glucose. Glucose-induced proinsulin, PC2, and PC3 biosynthesis was rapid (occurring after a 20-min lag period but reaching a maximum by 60 min), unaffected by the presence of actinomycin D; and in parallel experiments, stimulatory glucose concentrations did not alter MIN6 cell total preproinsulin, PC2, or PC3 mRNA levels. Thus, short-term (< 2 h) glucose stimulation of proinsulin, PC2 and PC3 biosynthesis in MIN6 cells, like that in isolated islets, was mediated at the translational level. Intracellular signals for mediating glucose-stimulated proinsulin PC2 and PC3 biosynthesis translation in MIN6 cells also appeared to be similar to those in pancreatic islets, requiring glucose metabolism and a supporting role for protein kinase A. However, protein kinase C or a Ca(2+)-dependent protein kinase did not appear to be required for glucose-regulated proinsulin biosynthesis in MIN6 cells, as in islets. MIN6 cells are the first beta-cell line that indicate glucose-regulated proinsulin biosynthesis translation essentially identical to that in differentiated islet beta-cells and will be an important experimental model to better define the mechanism of proinsulin biosynthesis in detail.
在短期内（<2小时），胰岛素原生物合成主要在翻译水平受葡萄糖调节；然而，该机制背后分子水平的细节尚未明确界定。深入了解胰岛素原生物合成机制的主要障碍之一是缺乏丰富的β细胞来源，这些β细胞在正常胰岛所定义的2.8 - 16.7 mmol/l葡萄糖范围内表现出受调节的胰岛素原生物合成表型。在本研究中，我们证明在MIN6细胞系中，在合适的葡萄糖浓度范围内存在对胰岛素原生物合成的特定葡萄糖调节的翻译控制。除胰岛素原外，两种胰岛素原转化内肽酶PC2和PC3的生物合成在MIN6细胞中也受葡萄糖协同调节，而外肽酶羧肽酶H的生物合成不受葡萄糖影响。当葡萄糖浓度超过4 mmol/l阈值时，胰岛素原、PC2和PC3的生物合成比MIN6细胞总蛋白合成受到特异性刺激，在8至10 mmol/l葡萄糖之间达到最大速率。葡萄糖诱导的胰岛素原、PC2和PC3生物合成迅速（在20分钟延迟期后发生，但在60分钟时达到最大值），不受放线菌素D存在的影响；并且在平行实验中，刺激葡萄糖浓度未改变MIN6细胞总前胰岛素原、PC2或PC3 mRNA水平。因此，MIN6细胞中胰岛素原、PC2和PC3生物合成的短期（<2小时）葡萄糖刺激，与分离的胰岛一样，是在翻译水平介导的。介导MIN6细胞中葡萄糖刺激的胰岛素原PC2和PC3生物合成翻译的细胞内信号似乎也与胰岛中的相似，需要葡萄糖代谢以及蛋白激酶A的支持作用。然而，与胰岛一样，蛋白激酶C或钙依赖性蛋白激酶似乎并非MIN6细胞中葡萄糖调节的胰岛素原生物合成所必需。MIN6细胞是第一个表明葡萄糖调节的胰岛素原生物合成翻译与分化的胰岛β细胞基本相同的β细胞系，将成为详细界定胰岛素原生物合成机制的重要实验模型。
