Vigeolas Helene, van Dongen Joost T, Waldeck Peter, Huhn Daniela, Geigenberger Peter
Max Planck Institute of Molecular Plant Physiology, Am Mühlenberg 1, 14476 Golm, Germany.
Plant Physiol. 2003 Dec;133(4):2048-60. doi: 10.1104/pp.103.031963. Epub 2003 Nov 26.
The aim of this study was to investigate whether endogenous restrictions in oxygen supply are limiting for storage metabolism in developing oilseed rape (Brassica napus) seeds. Siliques were studied 30 d after flowering, when rapid lipid accumulation is occurring in the seeds. (a). By using microsensors, oxygen concentrations were measured within seeds and in the silique space between seeds. At ambient external oxygen (21% [v/v]) in the light, oxygen fell to 17% (v/v) between and 0.8% (v/v) within seeds. A step-wise reduction of the external oxygen concentration led within 2 h to a further decrease of internal oxygen concentrations, and a step-wise increase of the external oxygen concentration up to 60% (v/v) resulted in an increase in internal oxygen that rose to 30% (v/v) between and 8% (v/v) within seeds. (b). The increase in oxygen levels in the seeds was accompanied by a progressive increase in the levels of ATP, UTP, and the ATP to ADP and UTP to UDP ratios over the entire range from 0% to 60% (v/v) external oxygen. (c). To investigate metabolic fluxes in planta, 14C-sucrose was injected into seeds, which remained otherwise intact within their siliques. The increase in oxygen in the seeds was accompanied by a progressive increase in the rate of lipid (including triacylglycerol), protein and cell wall synthesis, and an increase in glycolytic flux over a range from sub- to superambient oxygen concentrations. In contrast to lipid synthesis, starch synthesis was not significantly increased at superambient oxygen levels. The levels of fermentation products such as lactate and glycerol-3P increased only at very low (0%-4% [v/v]) external oxygen concentrations. (d). When 14C-acetate or 14C-acetyl-coenzyme A (CoA) was injected into seeds, label incorporation into triacylglycerol progressively increased over the whole range of external oxygen concentrations from 0% to 60% (v/v). (e). Stimulation of lipid synthesis was accompanied by an increase in sugar levels and a decrease in the levels of hexose-phosphates and acetyl-CoA, indicating sucrose unloading and the use of acetyl-CoA as possible regulatory sites. (f). Increased lipid synthesis was also accompanied by an increase in the maximal activities of invertase and diacylglycerol acyltransferase. (g). The developmental shift from starch to lipid storage between 15 and 45 d after flowering was accompanied by an increase in the seed energy state. (h). The results show that at ambient oxygen levels, the oxygen supply is strongly limiting for energy metabolism and biosynthetic fluxes in growing rape seeds, affecting lipid synthesis more strongly than starch synthesis. The underlying mechanisms and implications for strategies to increase yield and storage product composition in oilseed crops are discussed.
本研究的目的是调查内源性氧气供应限制是否会限制发育中的油菜（甘蓝型油菜）种子的储存代谢。在开花后30天对角果进行研究，此时种子中正在快速积累脂质。(a). 通过使用微传感器，测量了种子内部以及种子间角果空间中的氧气浓度。在光照下环境外部氧气含量为21%（体积/体积）时，种子间氧气含量降至17%（体积/体积），种子内部氧气含量降至0.8%（体积/体积）。外部氧气浓度逐步降低在2小时内导致内部氧气浓度进一步下降，而外部氧气浓度逐步增加至60%（体积/体积）导致内部氧气含量增加，种子间氧气含量升至30%（体积/体积），种子内部氧气含量升至8%（体积/体积）。(b). 种子中氧气水平的增加伴随着ATP、UTP水平以及ATP与ADP的比率和UTP与UDP的比率在外部氧气浓度从0%至60%（体积/体积）的整个范围内逐渐增加。(c). 为了研究植物体内的代谢通量，将14C-蔗糖注入种子，种子在其角果内保持完整。种子中氧气的增加伴随着脂质（包括三酰甘油）、蛋白质和细胞壁合成速率的逐渐增加，以及在低于至高于环境氧气浓度范围内糖酵解通量的增加。与脂质合成不同，在高于环境氧气水平时淀粉合成没有显著增加。乳酸和3-磷酸甘油等发酵产物的水平仅在非常低的（0%-4%[体积/体积]）外部氧气浓度下增加。(d). 当将14C-乙酸盐或14C-乙酰辅酶A（CoA）注入种子时，在外部氧气浓度从0%至60%（体积/体积）的整个范围内，标记物掺入三酰甘油的量逐渐增加。(e). 脂质合成的刺激伴随着糖水平的增加以及己糖磷酸和乙酰辅酶A水平的降低，表明蔗糖卸载以及乙酰辅酶A的利用可能是调节位点。(f). 脂质合成增加还伴随着转化酶和二酰甘油酰基转移酶最大活性的增加。(g). 开花后15至45天从淀粉储存到脂质储存的发育转变伴随着种子能量状态的增加。(h). 结果表明，在环境氧气水平下，氧气供应强烈限制了生长中的油菜种子的能量代谢和生物合成通量，对脂质合成的影响比对淀粉合成的影响更强。讨论了潜在机制以及对提高油籽作物产量和储存产物组成策略的影响。
