PDF(Pubmed)
Abstract：
The plastidic 2-C-methylerythritol 4-phosphate (MEP) pathway supplies the precursors of a large variety of essential plant isoprenoids, but its regulation is still not well understood. Using metabolic control analysis (MCA), we examined the first enzyme of this pathway, 1-deoxyxylulose 5-phosphate synthase (DXS), in multiple grey poplar (Populus × canescens) lines modified in their DXS activity. Single leaves were dynamically labeled with 13CO2 in an illuminated, climate-controlled gas exchange cuvette coupled to a proton transfer reaction mass spectrometer, and the carbon flux through the MEP pathway was calculated. Carbon was rapidly assimilated into MEP pathway intermediates and labeled both the isoprene released and the IDP+DMADP pool by up to 90%. DXS activity was increased by 25% in lines overexpressing the DXS gene and reduced by 50% in RNA interference lines, while the carbon flux in the MEP pathway was 25-35% greater in overexpressing lines and unchanged in RNA interference lines. Isoprene emission was also not altered in these different genetic backgrounds. By correlating absolute flux to DXS activity under different conditions of light and temperature, the flux control coefficient was found to be low. Among isoprenoid end products, isoprene itself was unchanged in DXS transgenic lines, but the levels of the chlorophylls and most carotenoids measured were 20-30% less in RNA interference lines than in overexpression lines. Our data thus demonstrate that DXS in the isoprene-emitting grey poplar plays only a minor part in controlling flux through the MEP pathway.
摘要：
质体2-C-甲基赤藓糖醇4-磷酸（MEP）途径提供了多种必需植物类异戊二烯的前体，但是它的规定仍然没有得到很好的理解。使用代谢控制分析(MCA)，我们检查了该途径的第一个酶，1-脱氧木酮糖5-磷酸合成酶（DXS），在多个灰杨树（Populus×canescens）品系中，其DXS活性得到了修饰。在照明中用13CO2动态标记单叶，气候控制的气体交换比色皿与质子转移反应质谱仪耦合，并计算了通过MEP途径的碳通量。碳被快速同化到MEP途径中间体中，并将释放的异戊二烯和IDPDMADP池标记为90％。DXS活性在过表达DXS基因的品系中增加了25%，在RNA干扰品系中减少了50%，而MEP途径中的碳通量在过表达品系中高出25-35％，而在RNA干扰品系中没有变化。在这些不同的遗传背景下，异戊二烯的排放也没有改变。通过将绝对通量与不同光照和温度条件下的DXS活性相关联，发现通量控制系数较低。在类异戊二烯最终产品中，异戊二烯本身在DXS转基因品系中没有变化，但是在RNA干扰系中测得的叶绿素和大多数类胡萝卜素的水平比在过表达系中低20-30％。因此，我们的数据表明，散发异戊二烯的灰杨树中的DXS在控制通过MEP途径的通量中仅起着次要作用。
