不同低温胁迫对烟草愈伤组织抗氰交替途径诱导和交替氧化酶表达影响的比较

比较了不同低温(14℃和4℃)胁迫对烟草(Nicotiana rusticaL．)愈伤组织抗氰交替途径诱导和交替氧化酶表达的影响。结果显示,不同低温胁迫处理能显著诱导烟草愈伤组织交替途径容量和实际运行的增加,且都呈现出基本相同的变化模式：在胁迫的初期(1—3d)持续增加,在3d时达到最高,而后下降到一个相对恒定的水平。但交替途径容量增加的幅度与温度下降的程度密切相关,而交替途径实际运行量的诱导程度在不同低温胁迫下的差异却很小。表明交替途径容量和实际运行对低温胁迫的响应是不同的。免疫印迹分析结果表明：低温胁迫明显诱导了交替氧化酶总蛋白的增加,且其随低温胁迫进程的变化与交替途径容量的变化基本一致;而对交替氧化酶单体与二聚体在低温胁迫下的含量变化检测结果则显示,烟草愈伤组织中交替氧化酶主要以二聚体形式存在,且这一存在形式并不随低温胁迫程度的加深而发生改变。两种形式的交替氧化酶蛋白含量都能被低温胁迫诱导增加,但其单体水平在两种不同的低温胁迫下并无明显差别,而4℃低温胁迫诱导的二聚体交替氧化酶蛋白含量明显高于14℃。表明不同程度低温对抗氰交替途径发生的不同影响主要是由于对交替氧化酶蛋白二聚体形式的不同诱导程度所致;而高活性的交替氧化酶单体形式则不因低温胁迫程度的加重而被明显诱导升高,使得抗氰交替途径的运行程度在两种不同的低温胁迫处理条件下无显著差异。     

外源~1O_2 和·OH及其清除剂对烟草愈伤组织交替途径发生与运行的影响(英文)

外源1 O2 和·OH处理继代培养 1 4d的烟草愈伤组织 2 4h后 ,愈伤组织内的交替途径的实际运行显著上升 ,但对交替途径容量影响不大 ;而经·OH处理后的愈伤组织交替途径容量和实际运行均明显地受到抑制 ,但交替途径的实际运行对·OH更加敏感。活性氧产生系统中加入相关清除剂进行实验 ,得到与此相对应的结果 :1 O2 的清除剂His处理能明显地降低或抑制1 O2 所增加的交替途径实际运行量 ,但对交替途径容量则几乎均无影响 ,而用·OH的清除剂DMSO和MAN分别处理愈伤组织后 ,DMSO和MAN均能解除·OH对交替途径容量和实际运行的抑制。这些结果表明 ,1 O2 诱导烟草愈伤组织交替途径的实际运行 ,但对其容量的作用不大 ,而·OH则明显地抑制交替途径容量和实际运行。推测1 O2 和·OH对抗氰途径的影响可能是通过对AOX活性的调节     

盐胁迫下烟草愈伤组织内源活性氧及乙烯的产生与交替途径发生、运行的关系

本文初次报告了烟草愈伤组织在不同盐浓度胁迫下交替途径与细胞色素途径的动态变化,同时检测了内源乙烯产生速率和活性氧(O2、H2O2和·OH)的含量及其相关酶(SOD和CAT)活性的变化。结果表明,交替途径实际运行量(ρValt)先逐渐上升,于0.75％盐浓度左右时达到极大值,但交替途径容量(Valt)与ρValt并不同步,且细胞色素途径活性(ρ’Vcyt)与ρValt变化趋势相反。与此同时,愈伤组织体内H2O2和O2迅速积累,分别在0.5％和0.75％左右达到极大值,·OH则一直呈上升趋势。而内源乙烯产生速率先急剧上升,在0.5％盐浓度左右时达到极大值后下降。将上述变化与各呼吸参数变化相比较发现,H2O2含量和乙烯产生速率与Valt变化曲线同步,而O2与ρValt变化曲线一致,但·OH含量与Valt变化曲线呈负相关。据此我们推测抗氰交替途径的发生、运行可能与活性氧的积累以及乙烯的产生密切相关,并讨论了交替途径运行的一些可能的生理作用。     

单克隆抗体和抗合成多肽抗体检测烟草愈伤组织中交替氧化酶表达的比较(英文)

从经过不同温度处理的烟草 (NicotianarusticaL .)愈伤组织中提取并纯化线粒体蛋白 ,分别与交替氧化酶的单克隆抗体和抗合成多肽抗体进行免疫杂交。结果表明 :交替氧化酶的含量随温度的下降而显著上升 ;单克隆抗体的特异性较高于抗合成多肽抗体 ,但后者与交替氧化酶同样有良好的亲和性。因此 ,用合成多肽方法制备的抗体可以用于交替氧化酶的研究中。     

单克隆抗体和抗合成多肽抗体检测烟草愈伤组织中交替氧化酶表达的研究

从经过不同温度处理的烟草（Nicotiana rustica L.)愈伤组织中提取并纯化线粒体蛋白,分别与交替氧化酶的单克隆抗体和抗合成多肽抗体进行免疫杂交。结果表明：交替氧化酶的含量随温度的下降而显上升,单克隆抗体的特异性较高于抗合成多肽抗体,但后与交替氧化酶同样有良好的亲和性。因此,用合成多肽方法制备的抗体可以用于交替氧化酶的研究中。     

低温胁迫下甘肃黄花烟草愈伤组织的抗氰呼吸

继代培养的ｉｃｏｔｉｏｎｏ ｒｕｓｉｔｏａ Ｌ．ｃｖ．Ｇａｎｓｕ Ｙｅｌｌｏｗ Ｆｌｏｗｅｒ）愈伤组织经低温（４℃）处理后,交替途径容量９Ｖａｌｔ）和实际运行量（ρＶａｈ）均被显著地诱导,同时其电子传递途径也在发生交替变化。低温胁迫开始时以细胞色素途径为主,至第２天时则以交替途径为主,并一直持续到第５天,以后则又转为经细胞色素途径为主,而对照却始终以细胞色素途径为主。经放红菌酮或氯霉素处理后,受低     

H2O2和水杨酸对陈化马铃薯切片交替呼吸途径影响的比较

外源5.0 mmol/L H2O2和0.1 mmol/L 水杨酸(salicylic acid, SA)处理均可明显提高陈化24 h的马铃薯切片的交替呼吸途径容量(Valt)及其与总呼吸的比值(Valt/Vt).应用交替氧化酶的单克隆抗体进行Western杂交的结果表明,H2O2和SA处理均可明显提高陈化马铃薯切片中交替氧化酶的表达水平.用氧同位素分辨法研究,结果表明:H2O2处理对陈化马铃薯切片中交替呼吸途径的实际运行没有影响,而SA处理对交替呼吸途径的实际运行具有明显的促进作用.上述结果表明,H2O2和SA对植物组织交替呼吸途径的影响存在差异,二者均可促进交替氧化酶的表达从而诱导交替呼吸途径容量的发生,但H2O2不影响其实际运行,而SA还可同时诱导其实际运行.     

水杨酸对陈化马铃薯切片交替途径活性的诱导作用

在马铃薯切片陈化过程中,总呼吸速率（Ｖｔ）和交替途径容量（Ｖａｌｔ）分别升高了５倍和２８倍,但交替途径在无ＫＣＮ存在下几乎不运行。用２０μｍｏｌ／Ｌ的水杨酸处理陈化切片可引起总呼吸速率和交替途径容量的少量增加,但交替途径的运行却明显加强。水杨酸的这种促进作用在Ｐｅｒｃｏｌｌ纯化的线粒体中也有出现。马铃薯切片中交替途径运行的增加导致了线粒体呼吸控制比及ＡＤＰ／Ｏ的下降。这些都表明水杨酸对马铃薯切片陈化过程中抗氰交替途径的活性具有诱导作用。     

NaCl胁迫下交替呼吸途径对烟草悬浮细胞死亡调节作用的研究

盐分胁迫是植物在自然环境中经常遭遇的环境胁迫因素之一,会引起植物代谢紊乱乃至细胞死亡,这严重限制了植物的生长、繁育和生存。交替呼吸途径是植物较之动物独特的线粒体呼吸途径。该研究在烟草悬浮细胞中调查了交替呼吸途径对Na Cl胁迫引起的植物细胞死亡过程的调节作用及相应的内在机制,以及在200 mmol·L~(-1)Na Cl处理的烟草悬浮细胞中研究了交替呼吸途径和细胞死亡发生及H_2O_2之间的关系。结果表明:(1)随着Na Cl处理浓度的增加,烟草悬浮细胞死亡水平逐渐增加,而交替呼吸途径的容量也逐渐上升。(2)与Na Cl处理相似,外源H_2O_2的处理也能导致烟草悬浮细胞死亡水平的增加。200 mmol·L~(-1)Na Cl的胁迫导致明显的细胞死亡发生和H_2O_2产量的显著性增加;而较之200 mmol·L~(-1)Na Cl胁迫下的细胞,用水杨基氧肟酸(交替呼吸途径的抑制剂)预处理后的细胞再置于200 mmol·L~(-1)Na Cl的胁迫下导致更高水平的细胞死亡和H_2O_2的产生。综上表明,高盐胁迫诱导了烟草悬浮细胞的交替呼吸途径的增加,而交替呼吸途径则可能通过抑制活性氧的产生而起到缓解细胞死亡发生的作用。     

缺磷对烟草悬浮细胞交替呼吸途径的影响及其与活性氧积累的关系

用两个耐低磷能力差异显著的烟草悬浮细胞系,研究了缺磷胁迫对交替呼吸途径的影响及其与活性氧积累的关系.牛津细胞耐缺磷胁迫能力明显高于K326.缺磷降低了K326交替途径的容量及实际运行量,但对牛津的影响不大.无论在缺磷还是正常磷水平下,牛津细胞交替途径容量和实际运行量都比K326高.用分离的线粒体进一步证实了上述结果.缺磷胁迫可以提高AOX的蛋白表达量,但品种间差异不明显.缺磷胁迫下K326细胞中的超氧阴离子积累量远高于牛津细胞.由此认为交替途径可能在细胞耐低磷中起作用.     

The Cyanide-resistant Respiration in Callus of Nicotiana rustica cv. Gansu Yellow Flower Under Low Temperature

The increasement of the operation (ρValt) and the capacity of the alternative pathway (Valt) in tobacco (Nicotiana rustica L. cv. Gansu Yellow Flower) callus that treated by low temperature (4 ℃) were observed. The two electron transport pathways were changed alternatively, the cyanide-resistant respiration was the main respiratory pathway from the 2nd day to the 5th day, while the cytochrome pathway was the main pathway consistently in control callus. Cycloheximide and chloramphenicol treatment which inhibited protein synthesis resulted in dramatic and partly inhibition of Valt induced by low temperature respectively. This further proved above result that low temperature induced Valt. The same result was found when two inhibitors were used together. The analysis of heat production of callus indicated that the operation of the alternative pathway could be regulated to improve the micro-environmental temperature of tobacco callus to adapt to low temperature. The possible participation of the alternative pathway in the response to low temperature and its physiological roles were discussed.     

The Relationship Between the Activity of Cyanide-resistant Respiration and the Expression of Alternative Oxidase in Different Organs of Mung Bean Seedlings

Twelve peptides, including eight conservative amino acid residues in the amino acid sequence of hydrophilic S helix of the alternative oxidase (AOX), were synthesized by solid-phase method. The polypeptide was coupled to αchymotrypsinogen, and the antibodies against this complex were obtained in rabbit. By using these antibodies, which were raised to immunoreact with total proteins of purified mitochondria from different organs of mung bean (Phaseolous radiatus L.) seedlings, it was found that there were two hybridizable AOX fractions in the mitochondria of mung bean seedlings. Their molecular weight was about 35 kD and 38 kD, respectively. Moreover, among the respiratory parameters obtained in hypocotyl, true leaf and cotyledon of mung bean seedlings true leaf had the highest total respiration (Vt), alternative pathway (AP) capacity(Valt) and the activity of AP (ρValt). Hypocotyl Vt and ρValt were the lowest, but its Vt was higher than that of the cotyledon. The activities of total and cyanide-resistant respiration were consistant with the analysis of Western blotting of AOX expression. The highest Vt and ρValt in true leaf were accompanied by two hybridizable polypeptides of AOX protein. The next was cotyledon Vt and ρValt with only one 38 kD hybridizable polypeptide of AOX protein. Hypocotyl Vt and ρValt were the lowest and its immunobloting band was similar to that of the cotyledon, but the expression amount of 38 kD protein was less than that of the cotyledon. The 35 kD AOX may make the main contribution to the true leaf ρValt.     

小麦幼苗叶片抗氰呼吸对轻度水分胁迫的响应

小麦幼苗经－０．５ＭＰa聚乙二醇（ＰＥＧ－６０００）溶液暗中渗透迫０、６、１２、１８、２４、３０、３６、４２、４８h,叶片的总呼吸速率（Ｖt)呈现先上升后降低的趋势,交替途径呼吸也表现出相同的变化模式。水分胁迫初期（０－１２h),交替途径容量（Ｖalt)、实际运行活性（ρValt)及运行系数（ρ值）均上升,此后（１８－４８h）逐渐下降,水分胁迫也影响了呼吸电子流在２条呼吸途径中的分配比例,胁迫初期的０－１２h内,流经交替途径的电子流增多,而流向细胞色素主路的电子流减少,但随着胁迫时间的延长,交替途径的贡献降低,而细胞色素主路的贡献增加,说明小麦叶片的抗氰呼吸在水分胁迫初期被诱导增加,而随着胁迫进行的延长又表现为下降。     

The alternative oxidase pathway is involved in the BR-induced salt resistance in mustard

In this study, the role of Brassinosteroids (BRs) and the relationship between the mitochondrial alternative oxidase (AOX) and ROS in the BR-induced defence response to salt stress was studied in mustard plants. Salt stress induced a significant activation of AOX. Exogenous BR significantly enhanced the capacity of the cyanide-resistant pathway, and reduced the damage of cell membrane. Pretreatment with brassinazole (Brz, an inhibitor of the BR biosynthesis pathway) significantly blocked the capacity of the cyanide-resistant pathway. BR could partly recover the AOX inactivation under salicylhydroxamic acid (SHAM, an inhibitor of the cyanide-resistant pathway) pretreatment. It was also found that BR could enhance the ROS accumulation and the antioxidant enzyme activities, while the AOX could eliminate the excessive ROS and enhance the antioxidant enzyme activities. Furthermore, the suppression of the cyanide-resistant pathway significantly increased the MDA content and the electrolyte leakage in mustard leaves, and the suppression of the BR biosynthesis had little effect on their recovering. Taken together, the cyanide-resistant pathway was involved in BR-induced salt tolerance and played an important role in maintaining the permeability of the cell membrane.     

盐胁迫下烟草愈伤组织内源活性氧及乙烯的产生与交替途径发生,？…

This paper reported firstly the dynamic changes of the alternative pathway and the cytochrome pathway in tobacco callus under different concentration of salt. The production rate of endogenous ethylene, the contents of active oxygen species (O2-., .OH and H2O2) and the activities of related enzyme (SOD and CAT) were also investigated simultaneously. The results indicated that the actual operation of the alternative pathway (rho Valt) increased gradually and reached the maxima at 0.75% salt concentration, then declined, while the capacity of the alternative pathway (Valt) didn't exhibit the same change pattern with rho Valt and the cytochrome pathway (rho' Vcyt) changed contrary to rho Valt. At the same time, H2O2 and O2-. accumulated rapidly and reached their peak value at about 0.5% and 0.75% salt concentration respectively, while the content of .OH increased consistently. The production rate of endogenous ethylene increased rapidly and reached its maxima at about 0.5% salt concentration. These data demonstrated that changes of Valt in tobacco callus were associated with a parallel production of H2O2 and endogenous ethylene and a opposite production of .OH, while rho Valt changed parally with the production of O2-.. On the basis of above results, it was deduced that the changes of the development and operation of the alternative pathway in tobacco callus under salt stress might be correlated with the production of endogenous ethylene and the active oxygen species. The possible participation of the alternative pathway in response to salt stress was also discussed.     

盐逆境下转基因耐盐小麦与其受体呼吸途径的动态变化

以转基因耐盐品种８９１２２和其受体陇春１３号两种小麦为实验材料,研究科幼苗在不同盐浓度胁迫下呼吸途径动态变化。结果表明,８９１２２出现盐呼吸明显迟于其受体;两听Ｖａｌｔ与ρＶａｌｔ变化并不同步,且Ｖａｌｔ均受高盐浓度的抑制,但低盐浓度能诱导其受体的Ｖａｌｔ;两品种的ρＶａｌｔ与ρ′Ｖｃｙｔ彼此协同调节适应盐境,且ρ′Ｖｃｙｔ仍是盐胁迫过程中线粒体电子传递的主要途径。同时讨论了盐逆境下抗氰呼吸的一些     

The alternative pathway in cucumber seedlings under low temperature stress was enhanced by salicylic acid

The alternative pathway is a cyanide-resistant and non-phosphorylatory electron transport pathway in mitochondria of higher plants. Alternative oxidase (AOX) is the terminal oxidase of this pathway. Our present study investigated the effect of exogenous salicylic acid (SA) on alternative pathway in cucumber (Cucumis sativus L.) seedlings under low temperature stress. Results showed that during the process of low temperature stress, the alternative pathway capacity was enhanced as AOX expression increased in SA pretreated seedlings. Compared with seedlings without SA pretreatment, slower decrease of relative water content and lower levels of electrolyte leakage, H₂O₂ and malonyldialdehyde content were detected in SA pretreated seedlings. These results indicated that SA could alleviate the injury caused by low temperature on cucumber seedlings. Since the special protective functions of alternative pathway and AOX in plants, we suggested that the alternative pathway was related to SA-mediated plant resistance to environmental stresses such as low temperature.     

The Alternative Respiratory Pathway of Euglena Mitochondria

Mitochondria, isolated from heterotrophic Euglena gracilis , have cyanide-resistant alternative oxidase (AOX) in their respiratory chain. Cells cultured under a variety of oxidative stress conditions (exposure to cyanide, cold, or H2O2) increased the AOX capacity in mitochondria and cells, although it was significant only under cold stress; AOX sensitivity to inhibitors was also increased by cold and cyanide stress. The value of AOX maximal activity reached 50% of total respiration below 20 degrees C, whereas AOX full activity was only 10-30% of total respiration above 20 degrees C. The optimum pH for AOX activity was 6.5 and for the cytochrome pathway was 7.3. GMP, AMP, pyruvate, or DTT did not alter AOX activity. The reduction level of the quinone pool was higher in mitochondria from cold-stressed than from control cells; furthermore, the content of reduced glutathione was lower in cold-stressed cells. Growth in the presence of an AOX inhibitor was not affected in control cells, whereas in cold-stressed cells, growth was diminished by 50%. Cyanide diminished growth in control cells by 50%, but in cold-stressed cells this inhibitor was ineffective. The data suggest that AOX activity is part of the cellular response to oxidative stress in Euglena .