摘要
以蚕豆叶片下表皮为材料，将荧光探针ＨＰＴＳ导入蚕豆气孔保卫细胞内，利用荧光光谱和激光共聚焦显微技术，检测了ＡＢＡ诱导蚕豆气孔关闭过程中Ｈ２Ｏ２的产生。结果表明，ＡＢＡ和１００μｍｏｌ／Ｌ的Ｈ２Ｏ２可以迅速猝灭ＨＰＴＳ的荧光，ＣＡＴ几乎可完全阻止９μｌ０．２ｍｍｏｌ／Ｌ的ＡＢＡ引起荧光猝灭。因此，ＡＢＡ可以诱导气孔保卫细胞产生Ｈ２Ｏ２，其产生的强度和速度与ＡＢＡ的浓度直接有关，并且推测：这种Ｈ２Ｏ２产生可能是ＡＢＡ诱导气孔关闭过程中信号转导链的一个中间成分。
Changes in H 2O 2 generation in guard cells of Vicia faba induced by ABA were measured by using fluorescence probe, 8 hydroxypyrene 1,3,6 trisulfonic acid (HPTS). Examination of epidermis peel was performed using a laser scanning confocal microscope and spectrofluorometer, set to an excitation light of 405 nm and an emission light of 512 nm. The fluorescence image before stimulation showed dye accumulation in guard cells, distributed homogeneously, with the exception of the chloroplast and nucleus where the fluorescence intensity was stronger, and there was almost no fluorescence in the epidermal cells(PlateⅠ).We also showed that the fluorescence was not significantly fading throughout the experiment. The retention time was long enough for analyses. When guard cells were treated with ABA, fluorescence quenching of HPTS occurred. A does dependent study of the ABA induced oxidative burst was observed, in which both the magnitude and rate of dye quenching was directly related to the concentration of ABA(Fig.2). And the treatment of the guard cells with catalase of 100 units/ml nearly completely obliterated the quenching reaction induced by ABA 10 μmol/L (final concentration). Furthermore, when H 2O 2 10 μmol/L(final concentration) was added to the suspension after above treatment caused little reduction in HPTS fluorescence(Fig.3). The results obtained by laser scanning confocal microscopy suggested that addition of exogenous ABA resulted in a rapid decrease in fluorescence in most cellular compartments of the guard cells(PlateⅠ). Above data support the hypothesis that H 2O 2 is involved in the signal transduction pathway of ABA induced stomatal closure.
基金
国家自然科学基金
国家攀登计划资助项目
关键词
蚕豆
保卫细胞
ABA
H2O2
信号转导
Vicia faba , guard cells, abscisic acid, hydrogen peroxide, signal transduction
