UV-B对拟南芥叶片不同来源H2O2的活化和气孔关闭的诱导

在UV-B调控植物许多生理过程中过氧化氢(H2O2)作为第二信使发挥着重要作用,但H2O2来源途径并不清楚。该研究借助气孔开度分析和激光扫描共聚焦显微镜技术,探讨H2O2在介导不同剂量UV-B诱导拟南芥叶片气孔关闭过程中的酶学来源途径。结果发现:0.5W.m-2 UV-B能诱导野生型拟南芥叶片保卫细胞的H2O2产生和气孔关闭,且该效应能被NADPH氧化酶抑制剂二苯基碘(DPI)抑制,而不能被细胞壁过氧化物酶抑制剂水杨基氧肟酸(SHAM)抑制,同时该剂量UV-B也不能诱导NADPH氧化酶功能缺失单突变体AtrbohD和AtrbohF以及双突变体AtrbohD/F保卫细胞的H2O2产生和气孔关闭;相反,0.65 W.m-2 UV-B既能诱导野生型也能诱导NADPH氧化酶突变体保卫细胞的H2O2产生和气孔关闭,且该效应能被SHAM抑制,却不能被DPI抑制。结果表明,不同剂量UV-B通过活化不同生成途径的H2O2来诱导拟南芥叶片气孔关闭,即低剂量UV-B主要诱导NADPH氧化酶AtrbohD和AtrbohF途径来源的H2O2生成,而高剂量UV-B主要活化细胞壁过氧化酶途径来源的H2O2。     

NO对He-Ne激光和增强UV-B辐射小麦幼苗气孔运动的作用机理研究

为探讨NO对He-Ne激光和增强UV-B辐射小麦（Triticum aestivuml）气孔运动的作用机理,采用低剂量（5 mW.mm-2）He-Ne激光和增强（10.08 kJ.m-2.d-1）UV-B辐射并结合药理学实验和激光共聚焦显微技术,对ML7113小麦的叶片及表皮条进行不同的处理,结果显示：（1）UV-B辐射既可诱导小麦叶片气孔关闭,又能够明显增加气孔保卫细胞和叶片的NO水平,且NO清除剂明显抑制了UV-B辐射诱导的小麦叶片气孔关闭,同时气孔保卫细胞和叶片内的NO含量明显减少。（2）一氧化氮合酶（NOS）抑制剂L-NAME对经UV-B辐射诱导的小麦幼苗气孔开度及保卫细胞和叶片内NO含量的抑制程度明显大于硝酸还原酶（NR）抑制剂NaN3对其的抑制程度,说明一氧化氮合酶（NOS）合成途径是小麦叶片经UV-B辐射后NO的主要产生途径。（3）就气孔开度而言,L〉CK〉BL〉B。就小麦叶片及保卫细胞内NO含量而言,B〉BL〉CK〉L。就硝酸还原酶（NR）和一氧化氮合酶（NOS）的活性而言,B组NR活性最低,NOS活性最高,L组NR活性最高,NOS活性最低。表明经He-Ne激光和增强UV-B辐射诱导的小麦气孔开度的变化确实与保卫细胞及叶片中NO含量的多少有关,气孔开度的减小及增大对应于NO含量的增多或减少,同时进一步证实了小麦叶片经He-Ne激光单独辐照后,NO的主要合成途径也来源于NOS途径。     

乙烯对UV-B辐射诱导蚕豆气孔关闭的调控效应(英文)

UV-B辐射作为一种重要的环境信号影响着植物的生长与发育,它能够调控气孔运动和诱导乙烯产生.该试验利用乙烯生物合成抑制剂和乙烯受体抑制剂处理蚕豆叶片表皮条,结合气孔开度分析和乙烯释放量测定,研究乙烯在UV-B辐射调控表皮条气孔运动中的作用.结果发现,将蚕豆叶片表皮条置于0.8 W·m-2的UV-B辐射下1～4 h,乙烯生成和气孔关闭均被显著诱导,且乙烯释放峰先于气孔关闭的起始;乙烯生物合成抑制剂和乙烯受体抑制剂处理均能显著逆转UV-B辐射诱导的气孔关闭;外源乙烯处理也能模拟UV-B辐射的效应诱导可见光下蚕豆表皮条的气孔关闭.可见,乙烯介导了UV-B辐射诱导的蚕豆气孔关闭.     

NO和H2O2在光/暗调控蚕豆气孔运动中的作用及其相互关系

借助表皮条分析和激光扫描共聚焦显微镜技术,对NO和H2O2在光/暗调控蚕豆(Vicia faba L.)气孔运动中的作用及其相互关系进行了探索.结果显示,光下外源NO供体硝普钠(SNP)和H2O2促进气孔关闭的效应明显大于暗中,暗中NO专一性清除剂2,4-羧基苯-4,4,5,5-四甲基咪唑-1-氧-3-氧化物(cPTIO)、一氧化氮合酶(NOS)抑制剂NG-氮-L-精氨酸-甲酯(L-NAME)和H2O2清除剂抗坏血酸(Vc)、过氧化氢酶(CAT)对气孔开度的效应明显大于光下,而且光下蚕豆保卫细胞NO和H2O2水平比暗中明显降低.上述结果表明,光/暗通过影响保卫细胞NO和H2O2的水平调控气孔运动.研究还发现,光下H2O2既诱导NO水平增加,也诱导气孔关闭,cPTIO和L-NAME有效地逆转H2O2的这些效应;光下SNP既诱导H2O2水平增加,也诱导气孔关闭,SNP的上述效应又被Vc和CAT有效逆转.这些结果表明,NO和H2O2在生成及效应上均存在明显的相互作用.另外,L-NAME显著逆转暗和光下H2O2处理对气孔关闭和NO生成的效应表明,蚕豆保卫细胞中可能存在NOS,暗和光下H2O2处理可能通过提高NOS的活性促进NO水平增加,进而诱导气孔关闭.     

外源NO和H2O2对洋葱鳞片外表皮气孔开度的调控

以洋葱(Allium cepa L.)肉质鳞片外表皮为材料,研究不同浓度及不同处理时间的外源NO和H2O2对洋葱鳞片外表皮上气孔开度的调节作用,并结合NO清除剂血红蛋白(Hb)和H2O2清除剂过氧化氢酶(CAT)研究调控过程中NO和H2O2的相互关系.结果显示:单独施用不同浓度的NO和H2O2均可诱导洋葱鳞片外表皮气孔不同程度关闭,并且浓度越大时间越长,其诱导气孔关闭效应越明显;NO和H2O2共同施用所诱导气孔关闭的效应大于其单独施用效应;Hb和CAT能明显减弱NO和H2O2诱导的气孔关闭.研究表明,NO和H2O2能有效诱导洋葱鳞片上气孔关闭,存在明显的浓度效应和时间效应,且两者可能互相依赖,具有协同效应.     

星型孢菌素(STS)对蚕豆气孔运动的调控效应

用激光扫描共聚焦显微技术,初步研究广谱性蛋白激酶抑制剂星型孢菌素(STS)对蚕豆气孔运动的调控效应.结果表明:(1)光下STS对气孔开度无影响但暗中显著促进气孔开放,表明蛋白激酶参与光/暗对气孔运动的调控,光下蛋白激酶活性低而暗中高;(2)与H2O2清除剂抗坏血酸(ASA)和NO清除剂羧基-2-苯-4,4,5,5-四甲基咪唑-1-氧-3-氧化物(cPTIO)一样,STS既降低暗处理和光下外源H2O2、硝普钠(SNP)处理保卫细胞H2O2、NO水平,也促进气孔开放,表明暗中蛋白激酶通过抑制H2O2、NO清除机制提高保卫细胞内源H2O2、NO水平并促进气孔关闭.     

H2S位于H2O2下游参与乙烯诱导拟南芥气孔关闭过程

H2O2和H2S是植物体内重要的信号分子,二者均参与乙烯诱导的拟南芥气孔关闭过程。以拟南芥野生型及其突变体为材料研究了H2O2和H2S在乙烯诱导拟南芥气孔关闭过程中的相互关系。结果表明,乙烯能够诱导野生型拟南芥叶片H2S含量及L-/D-半胱氨酸脱巯基酶(L-/D-CDes)活性显著增加,促进气孔关闭,但对H2O2合成突变体AtrbohD、AtrbohF、Atpao2和Atpao4植株叶片无显著作用;乙烯亦可引起H2S合成突变体Atl-cdes和Atd-cdes气孔保卫细胞H2O2水平的显著增加,但对其气孔运动没有显著作用。此外,H2O2清除剂和合成抑制剂均能抑制乙烯诱导的拟南芥叶片H2S含量和L-/D-CDes活性的增加及气孔开度的减小;而H2S清除剂和合成抑制剂虽能抑制乙烯诱导的气孔关闭,却不能改变乙烯对拟南芥叶片气孔保卫细胞H2O2的作用效应。由此表明H2S位于H2O2下游介导乙烯诱导拟南芥气孔关闭过程。     

NO可能作为H2O2的下游信号介导ABA诱导的蚕豆气孔关闭

ABA、H2O2和硝普钠(SNP)均能诱导蚕豆气孔关闭.NO的清除剂c-PTIO可以减轻由ABA或H2O2所诱导的蚕豆气孔关闭的程度,而过氧化氢酶(CAT)则不能减轻NO诱导的气孔关闭程度.激光共聚焦显微检测结果显示,10μmo1/L的ABA处理后,胞内H2O2的产生速率明显高于NO的产生速率;CAT几乎可完全抑制ABA所诱导的DAF的荧光增加;外源H2O2能显著诱导胞内DAF的荧光增加;c-PTIO对ABA诱导的DCF荧光略有促进作用,但外源SNP不能诱导胞内DCF荧光增加.这些结果表明,在ABA诱导气孔关闭过程中,H2O2可能在NO的上游起作用并受NO的负反馈调节.     

H_2O_2介导H_2S诱导的拟南芥气孔关闭

以拟南芥（Arabidopsis thaliana）为材料,研究了过氧化氢（H2O2）在硫化氢（H2S）调控气孔运动信号转导中的作用。结果表明,光下H2S的供体硫氢化钠（NaHS）能够诱导拟南芥气孔关闭;且能够显著提高叶片和保卫细胞胞质H2O2含量;H2O2的清除剂AsA和H2O2合成酶的抑制剂可不同程度地抑制NaHS诱导的拟南芥气孔关闭及叶片和保卫细胞胞质H2O2水平的升高;NaHS对AtrbohD、AtrbohF、Atpao2和Atpao4突变体气孔关闭、叶片和保卫细胞胞质H2O2水平升高的诱导作用要明显的小于野生型,但对AtPAO2和AtPAO4过表达株系叶片和保卫细胞H2O2水平的升高较野生型显著。据此推测,来源于NADPH氧化酶、细胞壁过氧化物酶和多胺氧化酶途径的H2O2参与H2S诱导的拟南芥气孔关闭。     

ABA诱导蚕豆气孔保卫细胞H2O2的产生

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H     

H_2O_2作为根源信号介导盐胁迫诱导的蚕豆气孔关闭反应

H2O2作为信号分子可被多种胁迫诱导产生并在细胞内积累,进而参与调节植物的抗逆反应。文章通过远红外热成像观察等实验发现,根部NaCl胁迫可诱导蚕豆气孔关闭,叶片温度上升,叶片内Na＋和H2O2含量增加,蒸腾流汁液中H2O2浓度升高。另外,NaCl可直接诱导离体蚕豆根产生H2O2,却不能影响表皮条内H2O2含量。NaCl胁迫条件下产生的蒸腾流汁液可直接诱导表皮条气孔关闭,该过程可被抗氧化剂抗坏血酸（AsA）所逆转。这些结果表明,H2O2作为盐胁迫的根源信号,可能通过维管系统运输参与调节蚕豆气孔的关闭反应。     

Ultraviolet-B radiation induces complex alterations in stomatal behaviour

Both visible and UV wavelengths play an important role in controlling stomatal aperture. We have analysed effects of UV-B radiation on stomatal aperture in Vicia faba , and found them to be complex. Depending on the metabolic state of the guard cell, high fluences of UV-B either stimulate stomatal opening or stomatal closing. Neither of these responses is readily reversed, i.e. once stomata have been exposed to UV-B, they are unable to re-adjust their aperture in response to environmental stimuli like changes in light, humidity or ABA. This lack of responsiveness is unlikely to be due to widespread cellular damage, as UV-induced stomatal closure is largely reverted in response to the H⁺-ATPase activator fusicoccin. It is speculated that UV-B impacts upstream from the plasmalemma based enzyme complexes which facilitate the solute fluxes leading to stomatal opening. Our data may help accommodate seemingly contradictory reports on the effects of UV-B on stomatal aperture and/or conductance.     

Ethylene mediates UV-B-induced stomatal closure via peroxidase-dependent hydrogen peroxide synthesis in Vicia faba L

Ultraviolet B (UV-B) radiation is an important environmental signal for plant growth and development, but its signal transduction mechanism is unclear. UV-B is known to induce stomatal closure via hydrogen peroxide (H(2)O(2)), and to affect ethylene biosynthesis. As ethylene is also known to induce stomatal closure via H(2)O(2) generation, the possibility of UV-B-induced stomatal closure via ethylene-mediated H(2)O(2) generation was investigated in Vicia faba by epidermal strip bioassay, laser-scanning confocal microscopy, and assays of ethylene production. It was found that H(2)O(2) production in guard cells and subsequent stomatal closure induced by UV-B radiation were inhibited by interfering with ethylene biosynthesis as well as ethylene signalling, suggesting that ethylene is epistatic to UV-B radiation in stomatal movement. Ethylene production preceded H(2)O(2) production upon UV-B radiation, while exogenous ethylene induced H(2)O(2) production in guard cells and subsequent stomatal closure, further supporting the conclusion. Inhibitors for peroxidase but not for NADPH oxidase abolished H(2)O(2) production upon UV-B radiation in guard cells, suggesting that peroxidase is the source of UV-B-induced H(2)O(2) production. Taken together, our results strongly support the idea that ethylene mediates UV-B-induced stomatal closure via peroxidase-dependent H(2)O(2) generation.     

The CO(2) response of Vicia guard cells acclimates to growth environment

Stomata of growth chamber-grown Vicia faba leaves have an enhanced CO(2) response, measured as change in stomatal aperture, compared to stomata of greenhouse-grown leaves. Reciprocal transfer experiments showed that the stomatal response to CO(2) acclimated to the growing environment. Stomata of growth chamber-grown leaves transferred to a greenhouse lost their high CO(2) sensitivity within 2-3 d while stomata of greenhouse-grown leaves transferred to a growth chamber acquired a high CO(2) sensitivity within 5-7 d. Experiments measuring the CO(2) responses of stomata in detached epidermis showed that growth chamber and greenhouse-grown stomata have the same contrasting CO(2) sensitivity observed in the intact leaf, indicating that the responses reflect intrinsic guard cell properties. The acclimation properties of the CO(2) response of guard cells have implications for the understanding of stomatal function under the predicted increases in atmospheric CO(2).     

过氧化氢对蚕豆气孔运动和质膜K+通道的影响

不同浓度H2 O2 可使蚕豆 (ViciafabaL .)叶片气孔关闭 ,抑制气孔张开 ,10mmol/L的H2 O2 最有效 ,10 μmol/L的H2 O2 仍明显使气孔关闭。且 10 μmol/L的H2 O2 抑制气孔张开作用能被EGTA所消除 ,表明Ca2 参与低浓度H2 O2 使气孔关闭的过程。 2mmol/L的H2 O2 可使质膜内向K 通道电流明显减小 ,而外向K 通道电流显著增加。因此 ,H2 O2 促进蚕豆气孔关闭主要是通过抑制K 通过保卫细胞质膜内向流入 ,或加强K 外向流出实现的     

共聚焦显微技术研究ABA诱导蚕豆气孔保卫细胞H_2O_2的产生(简报)

逆境下,植物细胞内ABA含量急剧增加,同时植物也可通过一些酶代谢反应积累活性氧,如H_2O_2,O_2~-。ABA作为逆境信号对气孔运动的显著调节作用已被诸多实验所证实,但关于其对气孔运动调节的细节还知之甚少。H_2O_2作为氧化信号分子在植物抗病信号转导中已得到广泛研究,但H_2O_2是否介导保卫细胞的气孔运动还缺乏直接的证据。我们已初步发现H_2O_2可参与外源ABA诱     

激光预处理可保护蚕豆细胞免受UV—B辐射的损伤

当蚕豆的胚被 He- Ne激光 (632 .8nm,1 .63J· mm- 2 )照射 5min或被 CO2 激光 (1 0 60nm,2 .53J· mm- 2 )照射 1 min后 ,将其置入 Knop营养液中进行恒温培养。当蚕豆的上胚轴长到大约 3cm时 ,在光背景 (PAR)为 70μmol· m- 2 · s- 1条件下 ,分别用 1 .0 2、3.0 3、4.52 k J· m- 2 的 UV- B辐射蚕豆的上胚轴 7h。根据蚕豆丙二醛 (MDA)、抗坏血酸 (As A)和 UV- B吸收物的含量变化 ,来测试激光对 UV- B照射蚕豆的上胚轴的保护作用。结果显示 :激光预处理可保护蚕豆上胚轴对 UV- B辐射的作用。与对照组 (没有用 UV- B或激光照射 )、UV- B单独照射组比较 ,在激光预处理的条件下 ,MDA的含量明显减少 ,As A和 UV- B吸收化合物的含量增加。如先用激光处理 ,然后再用 UV- B辐射 ,UV- B吸收物的含量将比单独用激光和 UV-B处理获得更好的改善。从而认为 ,激光预处理能增强植物对 UV- B的抵抗力。     

Extracellular calmodulin-induced stomatal closure is mediated by heterotrimeric G protein and H2O2

Extracellular calmodulin (ExtCaM) exerts multiple functions in animals and plants, but the mode of ExtCaM action is not well understood. In this paper, we provide evidence that ExtCaM stimulates a cascade of intracellular signaling events to regulate stomatal movement. Analysis of the changes of cytosolic free Ca2+ ([Ca2+]cyt) and H2O2 in Vicia faba guard cells combined with epidermal strip bioassay suggests that ExtCaM induces an increase in both H2O2 levels and [Ca2+]cyt, leading to a reduction in stomatal aperture. Pharmacological studies implicate heterotrimeric G protein in transmitting the ExtCaM signal, acting upstream of [Ca2+]cyt elevation, and generating H2O2 in guard cell responses. To further test the role of heterotrimeric G protein in ExtCaM signaling in stomatal closure, we checked guard cell responses in the Arabidopsis (Arabidopsis thaliana) Galpha-subunit-null gpa1 mutants and cGalpha overexpression lines. We found that gpa1 mutants were insensitive to ExtCaM stimulation of stomatal closure, whereas cGalpha overexpression enhanced the guard cell response to ExtCaM. Furthermore, gpa1 mutants are impaired in ExtCaM induction of H2O2 generation in guard cells. Taken together, our results strongly suggest that ExtCaM activates an intracellular signaling pathway involving activation of a heterotrimeric G protein, H2O2 generation, and changes in [Ca2+]cyt in the regulation of stomatal movements.