Tiessen Axel, Prescha Katrin, Branscheid Anja, Palacios Natalia, McKibbin Rowan, Halford Nigel G, Geigenberger Peter
Max Planck Institute of Molecular Plant Physiology, 14476 Golm, Germany.
Plant J. 2003 Aug;35(4):490-500. doi: 10.1046/j.1365-313x.2003.01823.x.
We recently discovered that post-translational redox modulation of ADP-glucose pyrophosphorylase (AGPase) is a powerful new mechanism to adjust the rate of starch synthesis to the availability of sucrose in growing potato tubers. A strong correlation was observed between the endogenous levels of sucrose and the redox-activation state of AGPase. To identify candidate components linking AGPase redox modulation to sugar supply, we used potato tuber discs as a model system. When the discs were cut from growing wild-type potato tubers and incubated for 2 h in the absence of sugars, redox activation of AGPase decreased because of a decrease in internal sugar levels. The decrease in AGPase redox activation could be prevented when glucose or sucrose was supplied to the discs. Both sucrose uptake and redox activation of AGPase were increased when EDTA was used to prepare the tuber discs. However, EDTA treatment of discs had no effect on glucose uptake. Feeding of different glucose analogues revealed that the phosphorylation of hexoses by hexokinase is an essential component in the glucose-dependent redox activation of AGPase. In contrast to this, feeding of the non-metabolisable sucrose analogue, palatinose, leads to a similar activation as with sucrose, indicating that metabolism of sucrose is not necessary in the sucrose-dependent AGPase activation. The influence of sucrose and glucose on redox activation of AGPase was also investigated in discs cut from tubers of antisense plants with reduced SNF1-related protein kinase activity (SnRK1). Feeding of sucrose to tuber discs prevented AGPase redox inactivation in the wild type but not in SnRK1 antisense lines. However, feeding of glucose leads to a similar activation of AGPase in the wild type and in SnRK1 transformants. AGPase redox activation was also increased in transgenic tubers with ectopic overexpression of invertase, containing high levels of glucose and low sucrose levels. Expression of a bacterial glucokinase in the invertase-expressing background led to a decrease in AGPase activation state and tuber starch content. These results show that both sucrose and glucose lead to post-translational redox activation of AGPase, and that they do this by two different pathways involving SnRK1 and an endogenous hexokinase, respectively.
我们最近发现，ADP-葡萄糖焦磷酸化酶（AGPase）的翻译后氧化还原调节是一种强大的新机制，可根据生长中的马铃薯块茎中蔗糖的可利用量来调节淀粉合成速率。观察到蔗糖的内源水平与AGPase的氧化还原激活状态之间存在很强的相关性。为了确定将AGPase氧化还原调节与糖供应联系起来的候选成分，我们使用马铃薯块茎切片作为模型系统。当从生长中的野生型马铃薯块茎上切下切片并在无糖条件下孵育2小时时，由于内部糖水平的降低，AGPase的氧化还原激活作用减弱。当向切片供应葡萄糖或蔗糖时，可以防止AGPase氧化还原激活作用的降低。当使用EDTA制备块茎切片时，蔗糖摄取和AGPase的氧化还原激活作用均增强。但是，用EDTA处理切片对葡萄糖摄取没有影响。对不同葡萄糖类似物的饲喂表明，己糖激酶对己糖的磷酸化是AGPase葡萄糖依赖性氧化还原激活中的一个重要成分。与此相反，饲喂不可代谢的蔗糖类似物帕拉金糖会导致与蔗糖类似的激活作用，这表明蔗糖代谢在蔗糖依赖性AGPase激活中并非必需。我们还研究了蔗糖和葡萄糖对具有降低的SNF1相关蛋白激酶活性（SnRK1）的反义植物块茎切片中AGPase氧化还原激活的影响。向块茎切片中饲喂蔗糖可防止野生型中AGPase氧化还原失活，但不能防止SnRK1反义株系中AGPase氧化还原失活。但是，饲喂葡萄糖会导致野生型和SnRK1转化体中AGPase产生类似的激活作用。在含有高水平葡萄糖和低蔗糖水平的转化酶异位过表达的转基因块茎中，AGPase氧化还原激活作用也增强。在表达转化酶的背景中表达细菌葡萄糖激酶会导致AGPase激活状态和块茎淀粉含量降低。这些结果表明，蔗糖和葡萄糖均会导致AGPase的翻译后氧化还原激活，并且它们分别通过涉及SnRK1和内源性己糖激酶的两种不同途径来实现这一点。
