过表达番茄GDP-L-半乳糖磷酸酶基因提高烟草抗MV诱导的氧化胁迫能力

为了探讨番茄GDP—L-半乳糖磷酸酶对烟草抗坏血酸（AsA）含量及抗氧化能力的影响,从番茄叶片中分离了GDP-L-半乳糖磷酸酶基因（LeGGP）,并转入到烟草中。以野生型（WT）和转正义LeGGP烟草株系T1-3和T1-15为试材,测定了甲基紫精（MV）处理下AsA、脱氢抗坏血酸（DHA）、H2O2、O2-和叶绿素含量、抗坏血酸过氧化物酶（APX）活性、光合速率和叶绿素荧光参数等。Northem杂交分析表明LeGGP的表达受MV的诱导,在MV处理下,野生型烟草的离体叶圆片发生比转基因烟草更严重的光漂白,转基因烟草的AsA含量及清除H2O2和O2-的能力明显强于野生型,过表达LePGG胀高了烟草的生长量。并且转基因烟草比野生型具有更高的净光合效率（Pn）和光系统Ⅱ（PSII）最大光化学效率（眠）。结果表明,LeGGP的过表达有助于提高烟草AsA含量及抗氧化胁迫能力。     

干旱胁迫下CO2浓度升高对转StAPX番茄植株耐旱能力的影响

在干旱胁迫伴随大气CO2浓度以及升高的CO2浓度（加倍）条件下,以过量表达番茄类囊体膜抗坏血酸过氧化物酶基因（StAPX）的转基因番茄为试材,探明干旱胁迫TCO2浓度升高对转基因及其野生型番茄植株清除活性氧及耐旱能力的影响。结果表明：升高的CO2浓度明显增加了干旱胁迫下植物的光合水平;升高的CO2浓度明显降低了干旱导致的植物体内H2O2．和O2的积累,影响了干旱胁迫下番茄植株的水．水循环系统的活性氧清除酶活性和小分子抗氧化物质含量;干旱胁迫下即使伴随升高的CO2浓度,测试番茄植株体内的渗透调节物质含量变化也不太明显;升高的CO2浓度明显降低了干旱胁迫下的植物细胞膜伤害程度;干旱胁迫下,升高的CO2浓度对转基因番茄株系比对野生型植株的影响更加明显。结果证明干旱逆境下,升高的CO2浓度能够在一定程度上进一步提高转基因番茄植株的耐旱性。     

外源硅对盐胁迫下黄瓜幼苗叶绿体活性氧清除系统的影响

以黄瓜为材料,研究了外源硅（K2SiO3 1．0mmol／L）对NaCl（50mmol/L）胁迫下黄瓜幼苗叶绿体中Na^＋、K^＋向叶绿体分配及活性氧清除系统的影响。结果表明：盐胁迫下硅处理使叶绿体在K^＋与Na^＋之间选择性吸收K^＋,从而降低了叶绿体内Na^＋的含量;同时Si处理可以显著降低盐胁迫下叶绿体中过氧化氢（H2O2）和丙二醛（MDA）含量,提高超氧化物歧化酶（SOD）、抗坏血酸过氧化物酶（APX）、谷胱甘肽还原酶（GR）和脱氢抗坏血酸还原酶（DHAR）的活性及抗坏血酸（ASA）、还原型谷胱甘肽（GSH）含量。说明Si不仅能降低叶绿体对Na^＋的选择吸收,还能增强叶绿体活性氧清除系统清除活性氧的能力,缓解盐胁迫对叶绿体膜的伤害。     

抑制番茄叶绿体DnaJ蛋白基因的表达降低转基因番茄的抗高温能力

DnaJ蛋白是广泛存在于植物细胞内的一种分子伴侣。在胁迫条件下它能够保护细胞内蛋白质等结构的稳定性。本研究克隆到一个番茄叶绿体DnaJ蛋白基（LeCDJI）。半定量PcR分析表明LeCDJ1,的表达受NaCl、高温、PEGTLABA的诱导。利用反义抑制的方法获得了LeCDJl抑制表达的转基因番茄。高温条件下,转基因株系较野生型表现出较严重的光系统II（PsII）光抑制,较低的D1蛋白含量,较高的超氧阴离子（O2-）、过氧化氢（H2O2）含量,及较低的抗坏血酸过氧化物酶（APX）和超氧化物歧化酶（SOD）活性。这些结果表明,抑制LeCDJ1的表达降低了转基因番茄的抗高温能力。     

外源抗坏血酸对臭氧胁迫下水稻叶片膜保护系统的影响

在田间原位条件下,运用OTCs（open top chamber）装置研究了外源抗坏血酸（exogenous ascorbate acid,ExAsA）对臭氧（O3）胁迫下水稻（Oryza Sativa L.）叶片膜保护系统的影响.研究发现,O3胁迫下的水稻叶片经过ExAsA处理后叶绿素a含量显著升高,而叶绿素b含量变化不明显;相对于对照,经ExAsA处理后的水稻叶片过氧化氢（H2O2）和丙二醛（MDA）含量及相对电导率（REC）均降低,超氧化物岐化酶（SOD）和抗坏血酸过氧化物酶（APX）活性明显提高,抗氧化剂类胡萝卜素（Carotene）含量升高.这表明,ExAsA改善了O3胁迫下水稻叶片的抗氧化系统功能,减少了叶片中活性氧（activity oxygen species,AOS）的积累,抑制了脂质过氧化（lipid peroxidation,LP）,延迟了O3对水稻叶片的老化作用,提高了水稻叶片对O3危害的抗性.     

过表达ZmSKIP基因提高烟草抗低温胁迫的能力

以野生型和过表达ZmSKIP基因烟草为试材, 研究了低温胁迫下过表达ZmSKIP对烟草抗氧化能力的影响。测定了不同低温处理时间下过表达ZmSKIP转基因烟草T3代植株和野生型植株抗氧化酶如超氧化物歧化酶（SOD）、过氧化氢酶（CAT）、过氧化物酶（POD）活性和丙二醛（MDA）含量以及相对电导率, 结果表明, 低温下, 相对于野生型植株, 转基因烟草具有较高的抗氧化酶活性和较低的相对电导率和MDA含量, 说明过表达ZmSKIP提高了转基因植株的耐低温胁迫能力。     

外源H2O2对盐胁迫下小麦幼苗生理指标的影响

以‘郑麦-004’小麦幼苗为供试材料,采用Hoagland营养液培养方法,通过添加H2O2的清除剂过氧化氢酶(CAT)和抗坏血酸(ASA),研究0.05μmol/L外源H2O2处理对150mmol/L NaCl胁迫下小麦幼苗生长和抗氧化系统活性的影响,探讨低浓度外源H2O2对盐胁迫下小麦幼苗伤害的防护作用及其生理机制。结果显示:外源H2O2能缓解盐胁迫对小麦幼苗生长的抑制效应,降低丙二醛(MDA)含量和超氧自由基(O2.-)的产生速率,使小麦幼苗的株高、根长和干重均显著增加,并能提高超氧化物歧化酶(SOD)、过氧化物酶(POD)、CAT、抗坏血酸氧化酶(APX)等保护酶活性和抗氧化物质谷胱甘肽(GSH)的含量;而H2O2清除剂(CAT和AsA)能够逆转外源H2O2对盐胁迫下小麦幼苗生长的促进作用。研究表明,低浓度外源H2O2处理能促进小麦幼苗中的酶类和非酶类抗氧化剂的产生,减少脂质过氧化物的含量,提高小麦幼苗的耐盐性。     

转铜/锌超氧化物歧化酶和抗坏血酸过氧化物酶基因马铃薯的耐氧化和耐盐性研究

高盐等逆境可以加剧植物体内活性氧的产生,进而引起植物细胞死亡。为开发抗逆境作物,以置于氧化诱导型启动子下定位于叶绿体的转铜/锌超氧化物歧化酶（Cu/ZnSOD）和抗坏血酸过氧化物酶基因(APX)马铃薯为材料,研究了其对MV和 NaCl所引起的氧化胁迫的耐受性。结果表明, MV胁迫下,转基因马铃薯叶片膜的相对电导率明显低于对照; NaCl胁迫下,其叶绿素含量高于对照。 在含NaCl 的培养基上,转基因幼苗生根率明显大于对照。另外,NaCl胁迫下转基因马铃薯叶片的SOD和APX酶活性显著高于对照,与其耐盐性的提高相一致。这些研究表明,转入Cu/ZnSOD和APX基因的马铃薯清除活性氧的能力增强,抗逆性得到提高。本实验采用氧化诱导型启动子调控下的SOD和APX两个基因协同作用,使外源基因只有在逆境胁迫时才特异性表达,增强转基因植株的抗逆效果,为培育抗逆经济作物开阔了思路。     

外源亚精胺对盐胁迫下黄瓜(Cucumis sativus L.)叶绿体活性氧清除系统和结合态多胺含量的影响

采用营养液水培,研究了外源亚精胺(Spd)对NaCl胁迫下抗盐能力不同的两个黄瓜品种幼苗生长、叶绿体中活性氧清除系统、转谷酰胺酶(TGase)活性、结合态多胺含量及植株光合速率的影响.结果表明,外源Spd能提高NaCl胁迫下叶绿体中TGase活性、叶绿体结合态腐胺(Put)、Spd、精胺(Spm)及总多胺含量;提高超氧化物歧化酶(SOD)、抗坏血酸过氧化物酶(APX)和谷胱甘肽还原酶(GR)活性,提高抗坏血酸(AsA)、类胡萝卜素(Car)、还原型谷胱甘肽(GSH)含量及还原型谷胱甘肽/氧化型谷胱甘肽(GSH/ GSSG)比值,降低脱氢抗坏血酸/抗坏血酸(DAsA/AsA)比值;同时显著降低叶绿体过氧化氢(H2O2)和丙二醛(MDA)含量,提高植株净光合速率,缓解NaCl胁迫对幼苗生长的抑制.表明Spd对黄瓜盐害的缓解作用之一可能是通过提高叶绿体结合态多胺含量和叶绿体活性氧清除能力,从而缓解盐胁迫对叶绿体膜的伤害.     

转铜/锌超氧化物歧化酶和抗坏血酸过氧化物酶基因甘薯的耐旱性

水分胁迫使转铜／锌超氧化物歧化酶基因（Cu／Zn SOD）和抗坏血酸过氧化物酶基因（APX）甘薯及未转基因甘薯中超氧阴离子（O2^-）、过氧化氢（H2O2）、丙二醛（MDA）含量和细胞膜相对透性增加,在相同条件下以上指标均为转基因甘薯低于未转基因甘薯;而叶片含水量、净光合速率（Pn）和气孔导度（Gs）均下降,SOD和APX酶活性随胁迫程度的加重先增大后减小,胞间CO2浓度（Ci）则先减小后增大,在相同条件下转基因甘薯中以上指标均高于未转基因甘薯。这些结果表明：转入Cu／Zn SOD和APX基因使转基因甘薯清除活性氧的能力增强,在水分胁迫下能保持较高的叶片含水量和Pn,耐旱性得到提高。     

Thylakoid membrane-bound ascorbate peroxidase is a limiting factor of antioxidative systems under photo-oxidative stress

To evaluate the physiological importance of thylakoid membrane-bound ascorbate peroxidase (tAPX) in the active oxygen species-scavenging system of chloroplasts, the level of tAPX in tobacco plants was altered by expression of the tAPX cDNA in both sense and antisense orientation. The tobacco plants transformed with constructs of antisense tAPXs from spinach and tobacco could not be obtained, suggesting that the suppression of tAPX in higher plants had a severe effect on the growth even under normal conditions. In contrast, the transgenic tobacco plants (TpTAP-12) overexpressing tAPX, which had approximately 37-fold higher activity than that of the wild-type plants, were generated. The TpTAP-12 plants showed increased tolerance to oxidative stress caused by application of methylviologen (MV, 50 microm) under light intensity (300 and 1600 microE m(-2) sec(-1)) and by chilling stress with high light intensity (4 degrees C, 1000 microE m(-2) sec(-1)). At 24 h after the MV treatment under illumination at 300 microE m-2 sec-1, destruction of chlorophyll was observed in the wild-type plants, but not in the TpTAP-12 plants. The activities of thiol-modulated enzymes in the Calvin cycle, the level and redox status of ascorbate (AsA), and the activity of tAPX in the wild-type plants significantly decreased, while those in the TpTAP-12 plants were hardly changed. These observations suggest that tAPX is a limiting factor of antioxidative systems under photo-oxidative stress in chloroplasts, and that the enhanced activity of tAPX functions to maintain the AsA content and the redox status of AsA under stress conditions.     

Thylakoid-bound ascorbate peroxidase mutant exhibits impaired electron transport and photosynthetic activity

In chloroplasts, stromal and thylakoid-bound ascorbate peroxidases (tAPX) play a major role in the removal of H(2)O(2) produced during photosynthesis. Here, we report that hexaploid wheat (Triticum aestivum) expresses three homeologous tAPX genes (TaAPX-6A, TaAPX-6B, and TaAPX-6D) mapping on group-6 chromosomes. The tAPX activity of a mutant line lacking TaAPX-6B was 40% lower than that of the wild type. When grown at high-light intensity photosystem II electron transfer, photosynthetic activity and biomass accumulation were significantly reduced in this mutant, suggesting that tAPX activity is essential for photosynthesis. Despite the reduced tAPX activity, mutant plants did not exhibit oxidative damage probably due to the reduced photochemical activity. This might be the result of a compensating mechanism to prevent oxidative damage having as a consequence a decrease in growth of the tAPX mutant plants.     

Effect of temperature on ascorbate peroxidase activity and flowering of Arabidopsis thaliana ecotypes under different light conditions

Four Arabidopsis thaliana ecotypes were grown at 14 degrees C and 22 degrees C under two light conditions (300 microE m-2 s-1 or 150 microE m-2 s-1) and the effect of temperature on their growth and flowering time was studied. Flowering occurred within 31 days (experimental period) at 22 degrees C, whereas a decrease in growth temperature resulted in a delay in flowering (63 days) under both light conditions. At 14 degrees C, membrane-bound APX (tAPX) activity decreased and total chlorophyll (Chl) content increased with growth under both light conditions. However, at 22 degrees C, the tAPX activity increased and total Chl content decreased with growth under both light conditions. These results suggest that at 22 degrees C oxidative stress was high under both light conditions and consequently Chl content decreased under stressful conditions or vice versa for all the four A. thaliana ecotypes studied. Under both the temperature and light conditions, soluble APX activity showed an irregular pattern of growth. The increase in tAPX activity, with growth only at 22 degrees C but not at 14 degrees C, suggests increased H2O2 formation in flowering plants at 22 degrees C for all the four A. thaliana ecotypes studied. Before flowering, the tAPX activity showed a significantly negative correlation with flowering time. Higher oxidative stress in the lower-latitude ecotypes might induce earlier flowering than the higher-latitude ecotypes. From these results, we propose a hypothesis that H2O2 is one of the possible factors in flower induction.     

Hydrogen peroxide-mediated phosphorylation of ERK1/2, Akt/PKB and JNK in cortical neurones: dependence on Ca2+ and PI3-kinase

Primary cortical neurones exposed to an oxidative insult in the form of hydrogen peroxide (H(2)O(2)) for 30 min showed a concentration-dependent increase in oxidative stress followed by a delayed NMDA receptor-dependent cell death measured 24 h later. Extracellular signal-regulated protein kinase (ERK1/2), c-jun N-terminal kinase (JNK) and the kinase Akt/PKB may regulate neuronal viability in response to oxidative insults. Using phospho-specific antibodies, a 15-min stimulation of neurones with H(2)O(2) (100 microm - 1 mm) produced a concentration-dependent phosphorylation of ERK1/2 and Akt/PKB that was partly dependent on extracellular Ca(2+) and phosphatidylinositol 3-kinase (PI3-K). Higher concentrations of H(2)O(2) (1 mm) also stimulated a phosphorylation of JNK which was totally dependent on extracellular Ca(2+) but not PI3-K. H(2)O(2)-induced phosphorylation of ERK1/2, Akt/PKB or JNK were unaffected by the NMDA channel blocker MK801. Blocking ERK1/2 activation with the upstream inhibitor U0126 (10 microm) enhanced H(2)O(2)-induced (100-300 microm range) neurotoxicity and inhibited H(2)O(2)-mediated phosphorylation of the cyclic AMP regulatory binding protein (CREB), suggesting that ERK1/2 signals to survival under these conditions. At higher concentrations (mm), H(2)O(2)-stimulated a phosphorylation of c-jun. It is likely, therefore, that subjecting neurones to moderate oxidative-stress recruits pro-survival signals to CREB but during severe oxidative stress pro-death signals through JNK and c-jun are dominant.     

Photoprotective mechanisms in cold-acclimated and nonacclimated needles of <Emphasis Type="Italic">Picea glehnii</Emphasis>

The response of Picea glehnii, a cold-tolerant species in the boreal zone, to air temperature (T) was investigated for its cold-acclimated needles (i.e. the ones subjected to gradual decrease in T) and nonacclimated needles (i.e. the ones subjected to a sudden decrease in T) were compared under low temperature. Cold-acclimated needles showed a greater increase of zeaxanthin and lutein contents than nonacclimated ones, whereas the nonacclimated needles showed a greater increase of thylakoid-bound ascorbate peroxidase (tAPX) activity than cold-acclimated ones under chilling conditions (after cold acclimation). These results suggest that: (1) low T induces the increase of zeaxanthin and lutein content, and tAPX activity; (2) accumulated zeaxanthin and lutein protect needles from photooxidative stress by dissipating excess energy before the reactive oxygen species (ROS) are formed in response to a gradual decrease in T (with cold acclimation and subsequent chilling condition), and by tAPX scavenging ROS formed in the case of a sudden decrease in T (without cold acclimation and chilling condition).