NO、ROS对SO_2诱导的万寿菊保卫细胞凋亡的调控

一氧化氮(NO)和活性氧(ROS)均是非常重要的信号分子,然而其在二氧化硫(SO2)对观赏植物毒作用过程中可能的信号作用还不清楚.因此,本文以万寿菊叶片下表皮为材料,采用表皮条分析法研究了SO2胁迫引起的保卫细胞死亡和胞内NO、ROS水平变化情况.结果显示:SO2衍生物(终浓度0.4~4.0 mmol·L-1)处理能引起万寿菊叶片下表皮保卫细胞生理活性下降,死亡率增加,且存在剂量效应,细胞出现核固缩、核降解、核拉长等典型核凋亡特征.同时,保卫细胞内NO、ROS和Ca2+水平显著升高(p0.05).用不同浓度的NO干扰剂(NO合酶抑制剂L-NAME、硝酸还原酶抑制剂Na N3及NO清除剂c-PTIO)、ROS清除剂(CAT和As A)、钙离子干扰剂(Ca2+螯合剂EGTA和Ca2+通道抑制剂La Cl3)分别与2.0 mmol·L-1SO2衍生物同时处理后,保卫细胞死亡率及相对应的胞内NO、ROS、Ca2+水平显著低于同期SO2衍生物单独处理组(p0.05).在用As A、L-NAME分别与2.0 mmol·L-1SO2衍生物共同处理后,同时检测ROS、NO及Ca2+含量,发现三者均显著低于SO2衍生物单独处理组;同理,用EGTA和2.0 mmol·L-1SO2衍生物共同处理后,尽管这时Ca2+水平显著下降,但ROS和NO含量却降低不显著.这一结果表明,NO和ROS在Ca2+的上游,且可能通过NO-Ca2+或者NO-ROS-Ca2+信号途径调节SO2诱导的万寿菊保卫细胞凋亡.

Signal regulation of NO and ROS in SO2-induced apoptosis of guard cells in leaves of Tagetes erecta

Both nitric oxide (NO) and reactive oxygen species (ROS) are very important signal molecules, but their roles in signal transduction of SO₂-induced toxicities on ornamental plants are not clear yet. In this study, the epidermal strip experiment was applied to investigate the functions of NO and ROS in the process of SO₂-induced death of lower epidermal guard cells in leaves of Tagetes erecta, one of the ornamental plants. The results showed that SO₂ derivatives (0.4~4.0 mmol·L^-1 of final concentrations) could reduce the guard cells' physiological activities and increase their death rates in a dose-dependent manner. Meanwhile, the significant increase of cellular NO, ROS, and Ca²⁺ levels(p < 0.05)and typical apoptosis features including nucleus condensation, nucleus break and apoptotic bodies were observed. However, exposure to 2.0 mmol·L^-1 of SO₂ derivatives combined with either NO antagonists (c-PTIO, a NO scavenger; NaN₃, a nitrite reductase inhibitor; L-NAME, a NO synthase enzyme inhibitor), ROS scavenger (such as ascorbic acid (AsA) and catalase (CAT)), or Ca²⁺ antagonists (EGTA, a Ca²⁺ chelating agent; LaCl₃, a plasma membrane Ca²⁺ channel blocker) could significantly decline the increased effects of cell death rates and the corresponding NO, ROS and Ca²⁺ levels induced by SO₂. Further studies showed that addition of L-NAME or AsA in 2.0 mmol·L^-1 of SO₂ derivatives led to significant decrease in the levels of NO, ROS and Ca²⁺, whereas addition of EGTA resulted in the decrease of cell death rates and Ca²⁺ levels and had insignificant effects on NO and ROS levels. It was concluded that SO₂-induced apoptosis of leaf guard cells in T. erecta might be mediated by NO-Ca²⁺ or NO-ROS-Ca²⁺ signal pathway.