杨树（Populus spp.）与茶藨子葡萄座腔菌（Botryosphaeria dothidea）互作中SA和H_2O_2的信号转导特征
|
摘要
本研究以毛白杨(Populus tomentosa)和北京杨(Populus×beijingensis)为试验材料，探讨了杨树与溃疡病菌(Botryosphaeria dothidea)互作过程中的信号转导，重点分析了水杨酸（SA）、水杨酸甲酯（MeSA）和过氧化氢（H_2O_2）作为信号分子的作用及其信号转导特征，主要研究结果如下：
1.检测了B. dothidea侵染后杨树内源SA含量及其相关指标的变化差异，分析了信号分子SA在杨树与B. dothidea互作中的动态特征，并初步揭示了SA与杨树的抗性相关性。接种B. dothidea后，毛白杨中游离态SA含量显著升高，接种后6h开始增加，72h时达到最高值，随后又下降，而未接种对照中游离态SA的含量变化不明显，说明B. dothidea诱导了毛白杨中SA的大量积累，SA参与了毛白杨与B. dothidea的互作过程；相对于结合态SA而言，游离态SA含量的变化总是相对敏感且显著，表明SA主要以游离态的形式参与杨树抗病防卫反应；不同抗性杨树接种B. dothidea后SA含量变化具有显著差异，毛白杨呈现出上述SA作为信号分子的典型功能特征，而北京杨接种B. dothidea后内源SA含量变化不显著，没有出现SA的迅速积累现象，在一定程度上显示了杨树对溃疡病的抗性与杨树体内游离态SA的积累密切相关。
2.针对杨树与B. dothidea互作中SA的合成途径进行研究，发现B. dothidea接种毛白杨后6-12h苯甲酸（benzoic axid，BA）的含量呈下降趋势，同时苯甲酸2-羟化酶（BA2H）活性显著升高，随着接种时间的延长，BA含量增加，而苯丙氨酸解氨酶（PAL）活性显著升高，说明B. dothidea侵染毛白杨后，BA由BA2H催化形成SA，但接种后期BA2H酶活性受到抑制导致BA含量的增加；与毛白杨相比较，北京杨接种后植物体内BA含量变化表现为缓慢上升趋势，整个接种期间没有出现BA含量下降现象；同时研究结果还显示，毛白杨在接种后48-72h，游离态SA与结合态SA的含量变化呈相反趋势，推测存在结合态SA向游离态SA转化的现象。
3.检测了B. dothidea与杨树互作中水杨酸（SA）与水杨酸甲酯（MeSA）的相互转化特征。在不同抗性杨树中SA和MeSA含量水平不同，SA和MeSA的变化具有一定的关系；接种B. dothidea后，毛白杨被诱导导致MeSA含量水平变化显著，并在接种后48h时出现峰值，而北京杨MeSA含量没有显著变化；MeSA和SA含量变化特征揭示，接种后48h MeSA含量达到最大值，伴随着SA含量达到峰值，MeSA含量随之急剧下降，而SA含量却急剧上升，表现出一个在量上的可能转化特征；经对杨树SABP2与SAMT两个基因的克隆及其RT-PCR分析发现，毛白杨接种B. dothidea后，SAMT在12h时表达量达到最大值，SABP2则在接种后48h时表达量达到最大值，但接种前期其表达受到抑制，这与SA和MeSA含量变化相一致，进一步表明在接种点存在MeSA与SA相互转化的现象；同一植株接种后，在未被感染部位，毛白杨中MeSA与SA含量均呈现上升趋势，与接种部位相比较，未感染组织MeSA的含量水平开始上升时间晚，且最高值小，未感染组织中SA的含量比接种点SA的含量低，而北京杨中这两种信号分子并没有显著变化；毛白杨未受感染组织中SABP2基因相对表达量在整个接种期间均处于上调状态，且比接种部位SABP2的表达更活跃，而SAMT的表达量变化不显著，上调幅度不大，这些现象表明，在杨树与溃疡病菌互作体系中接种部位与未受感染部位均存在SA与MeSA通过SABP2与SAMT进行相互转化的现象，MeSA可能作为信号传递分子在杨树与病原菌互作过程中起到关键作用。
4.测定了过氧化氢(H_2O_2)作为第二信使在B. dothidea与杨树互作中的作用，尤其在毛白杨和北京杨上的表现差异。为探讨不同互作体系中H_2O_2及其相关酶活性变化差异与其抗性间的关系，对毛白杨和北京杨接种后H_2O_2含量、CAT和APX酶活性动态变化以及水杨酸结合蛋白SABP基因相对表达进行了研究。结果显示，毛白杨在接种B. dothidea后H_2O_2的含量显著高于北京杨，毛白杨H_2O_2含量于48-72h内迅速积累，较北京杨上升快且含量高，毛白杨接种后72h时，H_2O_2含量达到最大值（737.52mol/g），且对照中H_2O_2含量没有显著变化，表明毛白杨对B. dothidea的抗性可能与植株体内H_2O_2的快速积累有关；为进一步研究H_2O_2的系统产生，用CeCl3染色的细胞化学方法对杨树韧皮部H_2O_2积累情况进行检测，结果表明，毛白杨在接种6h时即有沉淀积累，72h出现一个H_2O_2迸发高峰，大量电子沉积物累积并密集分布于细胞壁，因此细胞壁可能是诱导H_2O_2产生的主要部位；同样，在未受感染部位中H_2O_2变化趋势与接种部位相似，但CeCl3沉淀积累相对较少，初步推测杨树中B. dothidea可以诱导H_2O_2在整株植株中系统性产生；与抗病毛白杨相比，感病北京杨中H_2O_2-CeCl3沉淀颗粒很少，与利用生理学方法检测H_2O_2含量的结果相一致。
5.对毛白杨外施SA的实验表明，SA处理与SA处理后接菌的植株中H_2O_2含量具有相似的变化趋势，且均显著高于对照，外源SA处理使杨树体内H_2O_2含量迅速升高，48h时出现一峰值，SA处理后接菌的杨树体内H_2O_2含量变化趋势与SA单独处理相一致，但含量水平比SA单独处理要高，且SA处理后接菌的植株在接种前期H_2O_2含量明显高于接菌植株，因此表明在毛白杨与B. dothidea的互作过程中，外源SA可明显促进杨树体内H_2O_2的积累；通过对CAT及APX的活性进行测定表明，外源SA可明显抑制CAT活性，而APX活性没有明显变化，可见，在杨树与B. dothidea互作体系中SA可能主要通过对CAT的调节作用，积累H_2O_2产生抗病性；克隆获得编码毛白杨水杨酸结合蛋白PtSABP和北京杨PbSABP蛋白的两个基因，RT-PCR分析表明，接种后毛白杨SABP基因的表达在6-24h时受到抑制，结合前期对SA的研究，推测此时SA与SABP结合，抑制其CAT活性，从而促使24h后H_2O_2含量的显著升高；因此本实验证明H_2O_2的变化差异是由于B. dothidea侵染杨树后引起的，SABP参与了杨树的抗病过程，表明杨树对溃疡病的抗性与H_2O_2的积累情况有关，在杨树的防卫反应中，H_2O_2位于SA下游起作用。
Differences of salicylic acid (SA), methyl malicylate (MeSA) and hydrogen peroxide(H_2O_2) were analyzed in different resistance poplars (Populus tomentosa and Populus×beijingensis) inoculated with Botryosphaeria dothidea. The main results were as follows:
1. Difference of endogenous SA content and its related indicators were analyzed in poplarinoculated with B. dothidea, and the role of signaling molecules SA in poplar canker was clear.The results showed that the SA content of the resistant P. tomentosa had increased significantlyby6h after inoculation with B. dothidea and continued to accumulate to191.42ng/g freshweight (FW) by72h, after which it gradually declined. Levels of SA in control plantsremained low (9.98to14.12ng/g FW) throughout the sampling period. When the susceptible P.×beijingensis was infected with B. dothidea, SA levels were significantly lower than in P.tomentosa. Compared with the combination of SA, the free SA content is always relativelysensitive and significant, indicate that the free SA is involved in the defense response of poplarresistance to disease, indicating that the poplar canker resistance and poplar body of free SAaccumulation is closely related.
2. Study on the SA biosynthetic pathway in the system of poplar inoculated with B.dothidea, The results showed that the levels of benzoic acid was a downward trend at6-12hafter P.tomentosa inoculation, lower than the non-vaccinated control the content, while theactivity of benzeneformic acid2-hydroxylase (BA2H) was significantly elevated, indicatingthat BA change into SA, activity of phenylalanine ammonia lyase (PAL) was significantlyincreased, while the content of BA increased, this may BA2H activity is inhibited which led tothe increase in BA content; compared with P. tomentosa, P.×beijingensis after inoculation, theBA content of the performance of slow upward trend in the entire vaccination time. The resultsalso showed that P. tomentosa inoculated with B. dothidea after48-72h, the change of free SAand conjugated SA, the change was almost the opposite trend, suggesting the existence of bound SA into free SA of poplar inoculated with B. dothidea, the accumulation of the free stateof SA content may come from different channels, such as vaccination of pre-plant biosynthesisand the latter combined with the participation of the SA, BA and poplar anti-ulcer diseaseareclosely related.
3. We analyzed the content of MeSA in resistant P. tomentosa and susceptible P.×beijingensis after inoculation with B. dothidea. The P. tomentosa control plants containedbetween28.65and29.13ng/g FW MeSA, while MeSA levels in infected stems had increasedby6h after inoculation with B. dothidea and remained relatively steady from6to24hpost-inoculation. Then, MeSA content increased significantly in inoculated samples to65.93ng/gFW by48h. No significant induction of MeSA content occurred in the susceptible P.×beijingensis after B. dothidea inoculation. In summary, these quantitative analyses showed thatSA and MeSA production significantly increased in resistant P. tomentosa inoculated with B.dothidea. Moreover, in the susceptible P.×beijingensis, the contents of SA and MeSA were notelevated. Both the basal and induced levels of SA in P. tomentosa plants were increasedcompared with P.×beijingensis. By contrast, the basal level of MeSA in the treated tissue of P.tomentosa plants was similar compared to that in P.×beijingensis. Mock inoculation of thecontrols did not induce SA or MeSA accumulation in treated tissue, indicating that elevatedlevels of these compounds were induced by B. dothidea infection. Here, we identified SAMTand SABP2from P. tomentosa and P.×beijingensis and demonstrated that they shared highsequence similarity to N. tabacum SAMT and P. trichocarpa SABP2, respectively. We alsomeasured the expressions of SABP2and SAMT. We found that SABP2and SAMT were inducedsignificantly and more rapidly in P. tomentosa than in P.×beijingensis after B. dothideainfection, although induction of SABP2occurred in both poplar cultivars after inoculation withthe canker fungus. These results suggested a potential role for SABP2and SAMT in poplardisease resistance responses. Importantly, in P. tomentosa, the expression of SAMT peaked at12h, while SABP2expression peaked at48h, echoing the patterns seen with SA and MeSA. Insummary, SA levels increased in resistant P. tomentosa after inoculation with B. dothidea and peaked at72h after inoculation, SAMT expression was up-regulated, MeSA content increasedby48h after inoculation accompanied by increased expression of SABP2. Although the SAcontent rose in susceptible P.×beijingensis after infection, the increase was not significant andwas not accompanied by decreased MeSA levels and expressions of SAMT and SABP2as in P.tomentosa. Taken together, these data strongly supported the hypothesis that SA and MeSAwere essential parts of the defense response of poplar against pathogen attack. Furthermore, thedata indicated that SA and MeSA in poplar may be converted from one to the other by SAMTand SABP2.
4. To investigate the relation between activity differences of hydrogen peroxide (H_2O_2)and related enzymes in different resistance poplars inoculated with B. dothidea and theresistance of poplar trees，the content of H_2O_2,the activity dynamic changes of CAT and APXand expression of SABP gene relative were studied. The results showed that the content ofH_2O_2of Populus tomentosa was higher than P.×beijingensis at the early stage of inoculation,after72hours of inoculation, H_2O_2content reaches its maximum (737.52mol/g); afterinoculation with B. dothidea, the activity amplitude of CAT and APX of P.tomentosa weresignificantly higher than P.×beijingensis, the activity of CAT and APX were positivelycorrelated with poplar resistance; and associated with the reduction of CAT activity andcontent increased, Although the SA content arise in Populus×beijingensis, it not significant(7.83μg/g FW), and not accompanied by decreased CAT activity, the content of H_2O_2compared with P. tomentosa peak time early and low peak (respectively737.52n mol/g FWand306.99nmol/g FW), Infect the progress of SA give e-to CAT need the SA content attain akey content, P. tomentosa have SA-H_2O_2signal pathway; and detection of H_2O_2in poplarplant phloem by cytochemical method with CeCl3staining, which suggested there were a largenumber of H_2O_2-CeCl3precipitate particles on both sides of cell wall of P.tomentosa phloem at72h after inoculation;
5. SA treatment experiments show that outside of P. tomentosa, H_2O_2content in SA,dealing with SA treatment plants inoculated with a similar trend and were significantly higher than the control, exogenous SA poplar body H_2O_2content increased rapidly,48h a peak, SA,after inoculation of poplar body H_2O_2content trends and SA separate treatment is consistent,but the concentration levels than the SA processed separately to higher SA inoculated plantsinoculated pre-H_2O_2content significantlyhigher than that of inoculated plants, so that duringthe interactions between Populus tomentosa canker exogenous SA can significantly promotethe H_2O_2accumulation of poplar body; determination showed that the CAT and APX activity,and exogenous SA can inhibit the APX activity did not change significantly under the CATactivity, exogenous SA to be seen in the interaction system of poplar canker in SA mayprimarily through the regulation of CAT, the accumulation of H_2O_2disease resistance;Meanwhile, we isolated and identified full-length coding sequence of PtSABP and PbSABPfrom P. tomentosa and P.×beijingensis stems cDNAs, NCBI blast and phylogenetic treeanalysis shows that, proteins of PtSABP and PbSABP belonging to the Catalase_likesuperfamily, their sequences are90%identical to that of tobacco SABP at the amino acid level,RT-PCR analysis showed that gene expression of SABP was inhibited in6-24h after inoculatedwith B. dothidea., suggesting that SA binding with SABP at this time, inhibition of CATactivity, thus contributing increased significantly of the H_2O_2content at24h after inoculation..The experiments show that the difference of content of SA and H_2O_2in poplar is due toinoculated with B. dothidea, SABP involved in the disease resistance of poplar and suggestedthat the resistance of poplar to B. dothidea relevant to the accumulation of SA and H_2O_2.
1.检测了B. dothidea侵染后杨树内源SA含量及其相关指标的变化差异，分析了信号分子SA在杨树与B. dothidea互作中的动态特征，并初步揭示了SA与杨树的抗性相关性。接种B. dothidea后，毛白杨中游离态SA含量显著升高，接种后6h开始增加，72h时达到最高值，随后又下降，而未接种对照中游离态SA的含量变化不明显，说明B. dothidea诱导了毛白杨中SA的大量积累，SA参与了毛白杨与B. dothidea的互作过程；相对于结合态SA而言，游离态SA含量的变化总是相对敏感且显著，表明SA主要以游离态的形式参与杨树抗病防卫反应；不同抗性杨树接种B. dothidea后SA含量变化具有显著差异，毛白杨呈现出上述SA作为信号分子的典型功能特征，而北京杨接种B. dothidea后内源SA含量变化不显著，没有出现SA的迅速积累现象，在一定程度上显示了杨树对溃疡病的抗性与杨树体内游离态SA的积累密切相关。
2.针对杨树与B. dothidea互作中SA的合成途径进行研究，发现B. dothidea接种毛白杨后6-12h苯甲酸（benzoic axid，BA）的含量呈下降趋势，同时苯甲酸2-羟化酶（BA2H）活性显著升高，随着接种时间的延长，BA含量增加，而苯丙氨酸解氨酶（PAL）活性显著升高，说明B. dothidea侵染毛白杨后，BA由BA2H催化形成SA，但接种后期BA2H酶活性受到抑制导致BA含量的增加；与毛白杨相比较，北京杨接种后植物体内BA含量变化表现为缓慢上升趋势，整个接种期间没有出现BA含量下降现象；同时研究结果还显示，毛白杨在接种后48-72h，游离态SA与结合态SA的含量变化呈相反趋势，推测存在结合态SA向游离态SA转化的现象。
3.检测了B. dothidea与杨树互作中水杨酸（SA）与水杨酸甲酯（MeSA）的相互转化特征。在不同抗性杨树中SA和MeSA含量水平不同，SA和MeSA的变化具有一定的关系；接种B. dothidea后，毛白杨被诱导导致MeSA含量水平变化显著，并在接种后48h时出现峰值，而北京杨MeSA含量没有显著变化；MeSA和SA含量变化特征揭示，接种后48h MeSA含量达到最大值，伴随着SA含量达到峰值，MeSA含量随之急剧下降，而SA含量却急剧上升，表现出一个在量上的可能转化特征；经对杨树SABP2与SAMT两个基因的克隆及其RT-PCR分析发现，毛白杨接种B. dothidea后，SAMT在12h时表达量达到最大值，SABP2则在接种后48h时表达量达到最大值，但接种前期其表达受到抑制，这与SA和MeSA含量变化相一致，进一步表明在接种点存在MeSA与SA相互转化的现象；同一植株接种后，在未被感染部位，毛白杨中MeSA与SA含量均呈现上升趋势，与接种部位相比较，未感染组织MeSA的含量水平开始上升时间晚，且最高值小，未感染组织中SA的含量比接种点SA的含量低，而北京杨中这两种信号分子并没有显著变化；毛白杨未受感染组织中SABP2基因相对表达量在整个接种期间均处于上调状态，且比接种部位SABP2的表达更活跃，而SAMT的表达量变化不显著，上调幅度不大，这些现象表明，在杨树与溃疡病菌互作体系中接种部位与未受感染部位均存在SA与MeSA通过SABP2与SAMT进行相互转化的现象，MeSA可能作为信号传递分子在杨树与病原菌互作过程中起到关键作用。
4.测定了过氧化氢(H_2O_2)作为第二信使在B. dothidea与杨树互作中的作用，尤其在毛白杨和北京杨上的表现差异。为探讨不同互作体系中H_2O_2及其相关酶活性变化差异与其抗性间的关系，对毛白杨和北京杨接种后H_2O_2含量、CAT和APX酶活性动态变化以及水杨酸结合蛋白SABP基因相对表达进行了研究。结果显示，毛白杨在接种B. dothidea后H_2O_2的含量显著高于北京杨，毛白杨H_2O_2含量于48-72h内迅速积累，较北京杨上升快且含量高，毛白杨接种后72h时，H_2O_2含量达到最大值（737.52mol/g），且对照中H_2O_2含量没有显著变化，表明毛白杨对B. dothidea的抗性可能与植株体内H_2O_2的快速积累有关；为进一步研究H_2O_2的系统产生，用CeCl3染色的细胞化学方法对杨树韧皮部H_2O_2积累情况进行检测，结果表明，毛白杨在接种6h时即有沉淀积累，72h出现一个H_2O_2迸发高峰，大量电子沉积物累积并密集分布于细胞壁，因此细胞壁可能是诱导H_2O_2产生的主要部位；同样，在未受感染部位中H_2O_2变化趋势与接种部位相似，但CeCl3沉淀积累相对较少，初步推测杨树中B. dothidea可以诱导H_2O_2在整株植株中系统性产生；与抗病毛白杨相比，感病北京杨中H_2O_2-CeCl3沉淀颗粒很少，与利用生理学方法检测H_2O_2含量的结果相一致。
5.对毛白杨外施SA的实验表明，SA处理与SA处理后接菌的植株中H_2O_2含量具有相似的变化趋势，且均显著高于对照，外源SA处理使杨树体内H_2O_2含量迅速升高，48h时出现一峰值，SA处理后接菌的杨树体内H_2O_2含量变化趋势与SA单独处理相一致，但含量水平比SA单独处理要高，且SA处理后接菌的植株在接种前期H_2O_2含量明显高于接菌植株，因此表明在毛白杨与B. dothidea的互作过程中，外源SA可明显促进杨树体内H_2O_2的积累；通过对CAT及APX的活性进行测定表明，外源SA可明显抑制CAT活性，而APX活性没有明显变化，可见，在杨树与B. dothidea互作体系中SA可能主要通过对CAT的调节作用，积累H_2O_2产生抗病性；克隆获得编码毛白杨水杨酸结合蛋白PtSABP和北京杨PbSABP蛋白的两个基因，RT-PCR分析表明，接种后毛白杨SABP基因的表达在6-24h时受到抑制，结合前期对SA的研究，推测此时SA与SABP结合，抑制其CAT活性，从而促使24h后H_2O_2含量的显著升高；因此本实验证明H_2O_2的变化差异是由于B. dothidea侵染杨树后引起的，SABP参与了杨树的抗病过程，表明杨树对溃疡病的抗性与H_2O_2的积累情况有关，在杨树的防卫反应中，H_2O_2位于SA下游起作用。
Differences of salicylic acid (SA), methyl malicylate (MeSA) and hydrogen peroxide(H_2O_2) were analyzed in different resistance poplars (Populus tomentosa and Populus×beijingensis) inoculated with Botryosphaeria dothidea. The main results were as follows:
1. Difference of endogenous SA content and its related indicators were analyzed in poplarinoculated with B. dothidea, and the role of signaling molecules SA in poplar canker was clear.The results showed that the SA content of the resistant P. tomentosa had increased significantlyby6h after inoculation with B. dothidea and continued to accumulate to191.42ng/g freshweight (FW) by72h, after which it gradually declined. Levels of SA in control plantsremained low (9.98to14.12ng/g FW) throughout the sampling period. When the susceptible P.×beijingensis was infected with B. dothidea, SA levels were significantly lower than in P.tomentosa. Compared with the combination of SA, the free SA content is always relativelysensitive and significant, indicate that the free SA is involved in the defense response of poplarresistance to disease, indicating that the poplar canker resistance and poplar body of free SAaccumulation is closely related.
2. Study on the SA biosynthetic pathway in the system of poplar inoculated with B.dothidea, The results showed that the levels of benzoic acid was a downward trend at6-12hafter P.tomentosa inoculation, lower than the non-vaccinated control the content, while theactivity of benzeneformic acid2-hydroxylase (BA2H) was significantly elevated, indicatingthat BA change into SA, activity of phenylalanine ammonia lyase (PAL) was significantlyincreased, while the content of BA increased, this may BA2H activity is inhibited which led tothe increase in BA content; compared with P. tomentosa, P.×beijingensis after inoculation, theBA content of the performance of slow upward trend in the entire vaccination time. The resultsalso showed that P. tomentosa inoculated with B. dothidea after48-72h, the change of free SAand conjugated SA, the change was almost the opposite trend, suggesting the existence of bound SA into free SA of poplar inoculated with B. dothidea, the accumulation of the free stateof SA content may come from different channels, such as vaccination of pre-plant biosynthesisand the latter combined with the participation of the SA, BA and poplar anti-ulcer diseaseareclosely related.
3. We analyzed the content of MeSA in resistant P. tomentosa and susceptible P.×beijingensis after inoculation with B. dothidea. The P. tomentosa control plants containedbetween28.65and29.13ng/g FW MeSA, while MeSA levels in infected stems had increasedby6h after inoculation with B. dothidea and remained relatively steady from6to24hpost-inoculation. Then, MeSA content increased significantly in inoculated samples to65.93ng/gFW by48h. No significant induction of MeSA content occurred in the susceptible P.×beijingensis after B. dothidea inoculation. In summary, these quantitative analyses showed thatSA and MeSA production significantly increased in resistant P. tomentosa inoculated with B.dothidea. Moreover, in the susceptible P.×beijingensis, the contents of SA and MeSA were notelevated. Both the basal and induced levels of SA in P. tomentosa plants were increasedcompared with P.×beijingensis. By contrast, the basal level of MeSA in the treated tissue of P.tomentosa plants was similar compared to that in P.×beijingensis. Mock inoculation of thecontrols did not induce SA or MeSA accumulation in treated tissue, indicating that elevatedlevels of these compounds were induced by B. dothidea infection. Here, we identified SAMTand SABP2from P. tomentosa and P.×beijingensis and demonstrated that they shared highsequence similarity to N. tabacum SAMT and P. trichocarpa SABP2, respectively. We alsomeasured the expressions of SABP2and SAMT. We found that SABP2and SAMT were inducedsignificantly and more rapidly in P. tomentosa than in P.×beijingensis after B. dothideainfection, although induction of SABP2occurred in both poplar cultivars after inoculation withthe canker fungus. These results suggested a potential role for SABP2and SAMT in poplardisease resistance responses. Importantly, in P. tomentosa, the expression of SAMT peaked at12h, while SABP2expression peaked at48h, echoing the patterns seen with SA and MeSA. Insummary, SA levels increased in resistant P. tomentosa after inoculation with B. dothidea and peaked at72h after inoculation, SAMT expression was up-regulated, MeSA content increasedby48h after inoculation accompanied by increased expression of SABP2. Although the SAcontent rose in susceptible P.×beijingensis after infection, the increase was not significant andwas not accompanied by decreased MeSA levels and expressions of SAMT and SABP2as in P.tomentosa. Taken together, these data strongly supported the hypothesis that SA and MeSAwere essential parts of the defense response of poplar against pathogen attack. Furthermore, thedata indicated that SA and MeSA in poplar may be converted from one to the other by SAMTand SABP2.
4. To investigate the relation between activity differences of hydrogen peroxide (H_2O_2)and related enzymes in different resistance poplars inoculated with B. dothidea and theresistance of poplar trees，the content of H_2O_2,the activity dynamic changes of CAT and APXand expression of SABP gene relative were studied. The results showed that the content ofH_2O_2of Populus tomentosa was higher than P.×beijingensis at the early stage of inoculation,after72hours of inoculation, H_2O_2content reaches its maximum (737.52mol/g); afterinoculation with B. dothidea, the activity amplitude of CAT and APX of P.tomentosa weresignificantly higher than P.×beijingensis, the activity of CAT and APX were positivelycorrelated with poplar resistance; and associated with the reduction of CAT activity andcontent increased, Although the SA content arise in Populus×beijingensis, it not significant(7.83μg/g FW), and not accompanied by decreased CAT activity, the content of H_2O_2compared with P. tomentosa peak time early and low peak (respectively737.52n mol/g FWand306.99nmol/g FW), Infect the progress of SA give e-to CAT need the SA content attain akey content, P. tomentosa have SA-H_2O_2signal pathway; and detection of H_2O_2in poplarplant phloem by cytochemical method with CeCl3staining, which suggested there were a largenumber of H_2O_2-CeCl3precipitate particles on both sides of cell wall of P.tomentosa phloem at72h after inoculation;
5. SA treatment experiments show that outside of P. tomentosa, H_2O_2content in SA,dealing with SA treatment plants inoculated with a similar trend and were significantly higher than the control, exogenous SA poplar body H_2O_2content increased rapidly,48h a peak, SA,after inoculation of poplar body H_2O_2content trends and SA separate treatment is consistent,but the concentration levels than the SA processed separately to higher SA inoculated plantsinoculated pre-H_2O_2content significantlyhigher than that of inoculated plants, so that duringthe interactions between Populus tomentosa canker exogenous SA can significantly promotethe H_2O_2accumulation of poplar body; determination showed that the CAT and APX activity,and exogenous SA can inhibit the APX activity did not change significantly under the CATactivity, exogenous SA to be seen in the interaction system of poplar canker in SA mayprimarily through the regulation of CAT, the accumulation of H_2O_2disease resistance;Meanwhile, we isolated and identified full-length coding sequence of PtSABP and PbSABPfrom P. tomentosa and P.×beijingensis stems cDNAs, NCBI blast and phylogenetic treeanalysis shows that, proteins of PtSABP and PbSABP belonging to the Catalase_likesuperfamily, their sequences are90%identical to that of tobacco SABP at the amino acid level,RT-PCR analysis showed that gene expression of SABP was inhibited in6-24h after inoculatedwith B. dothidea., suggesting that SA binding with SABP at this time, inhibition of CATactivity, thus contributing increased significantly of the H_2O_2content at24h after inoculation..The experiments show that the difference of content of SA and H_2O_2in poplar is due toinoculated with B. dothidea, SABP involved in the disease resistance of poplar and suggestedthat the resistance of poplar to B. dothidea relevant to the accumulation of SA and H_2O_2.
引文
安钰．过氧化氢在合作杨苗木诱导防御性反应中的作用．北京林业大学博士学位论文，2008，7-10．
曹支敏，景耀，周芳．杨树溃疡病流行与土壤条件的关系．西北林学院学报，1991a，6(2)：55-9．
曹支敏，周芳，杨俊秀．杨树溃疡病流行规律与测报研究．森林病虫通讯，1991b，3：5-9．
丁义峰，刘萍．水杨酸诱导植物抗逆性研究进展.生物学教学．2011(11)：42-43．
丁秀英，张军，苏宝林，等．水杨酸在植物抗病中的作用．植物学通报，2001，18（2）：163-168．
杜建玲，刘红霞，于淑平．杨树不同种(品种)间抗溃疡病差异的比较．河北林果研究，2000，15(1)：55-60．
杜建玲，项蔚华，沈瑞祥．诱导杨树抗溃疡病机理的研究．林业科学研究，1995，8(1)：78-81．
高雪．植物苯丙氨酸解氨酶研究进展．现代农业科技，2009，(1)：31-32．
胡景江，朱玮，文建雷．杨树细胞壁HRGP和木质素的诱导积累与其对溃疡病抗性的研究．植物病理学报，1999，29(2)：151-156．
胡景江，朱玮，袁雪丽．溃疡病菌对杨树细胞壁中HRGP和木质素的诱导作用．西北林学院学报，1997，12(3)：14-17．
沈文飚，徐朗莱，叶茂炳，等．水杨酸对小麦叶片抗坏血酸过氧化物酶活性的抑制．南京农业大学学报，1998，21(3)：126-128．
苏晓华，张冰玉，黄秦军，等．我国林木基因工程研究进展和关键领域．林业科学，2003，39(5)：111-118．
理永霞，张星耀，严东辉，等．杨树接种Botryosphaeria dothidea溃疡病菌后蛋白质表达差异分析．南京林业大学学报（自然科学版），2011，35(4)：1-6．
李娜，陆海．植物抗坏血酸过氧化物酶基因家族研究进展．成都大学学报(自然科学版)，2011，30(2)：97-99．
梁军，姜俊清，刘会香，等．我国杨树与溃疡病菌互作的病理学研究．林业科学研究，2005，18(2):214-221.
梁元存，刘爱新．植物系统获得抗性中的信号．山东农业大学学报(自然科学版)，2001(02)：17-18．
苗则彦，赵奎华，刘长远，等．葡萄抗感白腐病品种PAL酶、PPO酶和SOD酶活性比较．沈阳农业大学学报，2003，34(3)：177-180．
钱万强，曹福祥，王猛，等．松材线虫侵染对马尾松体内有机酸及酶的影响．湖南林业科技，2009，36(3)：10-12．
许文力，徐克勤，吴玉斌，等．杨树溃疡病．江苏林业科技，1997(04)：5-6．
肖淑贤，高俊明．番茄诱导抗病性的研究进展．中国西部科技，2011(04)：42-43．
邢会琴，李敏权，徐秉良，等．过氧化物酶和苯丙氨酸解氨酶与苜蓿白粉病抗性的关系．草地学报，2007，l5(4)：37-38．
向玉英，花晓梅，赵经周．杨树溃疡病的发生及病原菌的生物学特性的研究．植物病理学报，1981，11(4)：27-33．
向玉英，魏舜明，侯艳．杨树溃疡病与树皮酚化合物关系的研究．森林病虫通讯，1993，1：5-7．
王颖，胡景江，朱玮，等．杨树溃疡病寄主诱导抗病性的研究．西北林学院学报，1996，11(1)：1-4．
王媛．杨树与溃疡病菌互作的细胞生物学、活性氧代谢及细胞过敏反应,中国林业科学研究院博士学位论文，2007，5-6．
王文天，彭少麟，李冬梅等．薇甘菊水杨酸羧甲基转移酶基因的分离鉴定及表达分析，热带亚热带植物学报，2009，17（5）：445-450．
王春丽，梁宗锁．外源刺激对植物次生代谢的调节及其信号转导途径研究进展．西北植物学报，2009(5)：11-12．
王金生．分子植物病理学.中国农业出版社，1999．
杨俊秀，李武汉，符毓秦，等．抗溃疡病杨树种类的调查研究．西北林学院学报，1990，5(4)：1-10．
阳传和．杨树溃疡病接种方法的比较．北京林学院学报，1985，（2)：76-81．
阳传和．树皮内酚类物质的含量及苯丙氨酸的活性与杨树抗溃疡病的关系．林业科学，1989，25(4)：311-316．
袁嗣令．国内外杨树病害研究概况，四川省永川森林病虫防治试验站，1984．
张星耀，骆有庆．中国森林重大生物灾害．北京：中国林业出版社，2003．
赵嘉平．树木溃疡病菌-葡萄座腔菌属及相关真菌系统分类研究．中国林业科学研究院博士学位论文．2007．5-6．
赵仕光，景耀．杨树对溃疡病的抗性研究(I)对龄及形态结构与抗病性．西北林学院学报，1997，12(3)：35-40．
钟兆康，高雅．不同杨树品种对杨树水泡型溃疡病菌的抗病性测定简报．林业科技通讯，1981，1：25-26．
中科院上海植物生理所主编．现代植物生理学实验指南．北京:科学出版社，1999．
赵仕光，景耀，杨俊秀．杨树树皮内过氧化物酶和多酚氧化酶活性与抗溃疡病的关系．西北林学院学报，1993，8(3)：13-17．
曾大鹏．我国杨树病害的研究现状与防治[J]．中国森林病虫，2002，21(1)：20-26．
郑伟尉，臧运祥．水杨酸(SA)与植物抗病研究进展．河北果树，2005(01)：23-25．
赵仕光，朱玮，岳红艳．溃疡病菌在杨树树皮组织中的扩展和对寄主细胞超微结构的影响．林业科学研究，1999，12(2)：11-26．
张佳娣．活性氧的信号传导途径．安徽农业科学，2010(16)：56-57．
Ament K, Kant MR, Sabelis MW, et al. Jasmonic acid is a key regulator of spider mite-induced volatileterpenoid and methyl salicylate emission in tomato. Plant Physiol,2004,135:2025-2037.
An C, Mou Z. Salicylic acid and its function in plant immunity. J Integr Plant Biol,2011,53:412-428.
Antoniw JF, Dunkley AM, White RF, et al. Soluble leaf proteins of virus-infected tobacco (Nicotianatabacum) cultivars [proceedings]. Biochem Soc Trans,1980,8:70-71.
Averyanov A. Oxidative burst and plant disease resistance. Front Biosci (Elite Ed),2009,1:142-152.
Bokszczanin KL, Przybyla AA.[Molecular aspects of allergy to plant products. Part II. Pathogenesis-relatedproteins (PRs), apple allergenicity governed by Mal d1gene]. Pol Merkur Lekarski,2012,32:176-181.
Bolwell BJ, Kalaycio M, Goormastic M, et al. Progressive disease after ABMT for Hodgkin's disease. BoneMarrow Transplant,1997,20:761-765.
Bonifacio A, Martins MO, Ribeiro CW, et al. Role of peroxidases in the compensation of cytosolic ascorbateperoxidase knockdown in rice plants under abiotic stress. Plant Cell Environ,2011,34:1705-1722.
Casano LM, Martin M, Sabater B. Hydrogen peroxide mediates the induction of chloroplastic Ndh complexunder photooxidative stress in barley. Plant Physiol,2001,125:1450-1458.
Charles A, Dawicki DD, Oldmixon E, et al. Studies on the mechanism of short-term regulation of pulmonaryartery endothelial cell Na/K pump activity. J Lab Clin Med,1997,130:157-168.
Chen Z, Ricigliano JW, Klessig DF. Purification and characterization of a soluble salicylic acid-bindingprotein from tobacco. Proc Natl Acad Sci U S A,1993,90:9533-9537.
Chen Z, Silva H, Klessig DF. Active oxygen species in the induction of plant systemic acquired resistance bysalicylic acid. Science,1993,262:1883-1886.
Chern M, Fitzgerald HA, Canlas PE, et al. Overexpression of a rice NPR1homolog leads to constitutiveactivation of defense response and hypersensitivity to light. Mol Plant Microbe Interact,2005,18:511-520.
Clarke JD, Volko SM, Ledford H, et al. Roles of salicylic acid, jasmonic acid, and ethylene in cpr-inducedresistance in arabidopsis. Plant Cell,2000,12:2175-2190.
Colditz F, Niehaus K, Krajinski F. Silencing of PR-10-like proteins in Medicago truncatula results in anantagonistic induction of other PR proteins and in an increased tolerance upon infection with theoomycete Aphanomyces euteiches. Planta,2007,226:57-71.
Dangl JL, Jones JD. Plant pathogens and integrated defence responses to infection. Nature,2001,411:826-833.
Dat JF, Pellinen R, Beeckman T, et al. Changes in hydrogen peroxide homeostasis trigger an active cell deathprocess in tobacco. Plant J,2003,33:621-632.
Davis JM, Wu H, Cooke JE, et al. Pathogen challenge, salicylic acid, and jasmonic acid regulate expressionof chitinase gene homologs in pine. Mol Plant Microbe Interact,2002,15:380-387.
Ding CK, Wang CY, Gross KC, et al. Jasmonate and salicylate induce the expression ofpathogenesis-related-protein genes and increase resistance to chilling injury in tomato fruit. Planta,2002,214:895-901.
Dixon RA, Achnine L, Kota P, et al. The phenylpropanoid pathway and plant defence-a genomicsperspective. Mol Plant Pathol,2002,3:371-390.
Du H, Klessig DF. Identification of a Soluble, High-Affinity Salicylic Acid-Binding Protein in Tobacco.Plant Physiol,1997,113:1319-1327.
Durner J, Klessig DF. Inhibition of ascorbate peroxidase by salicylic acid and2,6-dichloroisonicotinic acid,two inducers of plant defense responses. Proc Natl Acad Sci U S A,1995,92:11312-11316.
Durrant WE, Dong X. Systemic acquired resistance. Annu Rev Phytopathol,2004,42:185-209.
Forouhar F, Yang Y, Kumar D, et al. Structural and biochemical studies identify tobacco SABP2as a methylsalicylate esterase and implicate it in plant innate immunity. Proc Natl Acad Sci U S A,2005,102:1773-1778.
Fragniere C, Serrano M, Abou-Mansour E, et al. Salicylic acid and its location in response to biotic andabiotic stress. FEBS Lett,2011,585:1847-1852.
Fukami H, Asakura T, Hirano H, et al. Salicylic acid carboxyl methyltransferase induced in hairy rootcultures of Atropa belladonna after treatment with exogeneously added salicylic acid. Plant CellPhysiol,2002,43:1054-1058.
Glazebrook J. Genes controlling expression of defense responses in Arabidopsis--2001status. Curr OpinPlant Biol,2001,4:301-308.
Glocova I, Thor K, Roth B, et al. Salicylic acid (SA)-dependent gene activation can be uncoupled from celldeath-mediated gene activation: the SA-inducible NIMIN-1and NIMIN-2promoters, unlike thePR-1a promoter, do not respond to cell death signals in tobacco. Mol Plant Pathol,2005,6:299-314.
Gonzalez-Teuber M, Eilmus S, Muck A, et al. Pathogenesis-related proteins protect extrafloral nectar frommicrobial infestation. Plant J,2009,58:464-473.
Hajduch M, Rakwal R, Agrawal GK, et al. High-resolution two-dimensional electrophoresis separation ofproteins from metal-stressed rice (Oryza sativa L.) leaves: drastic reductions/fragmentation ofribulose-1,5-bisphosphate carboxylase/oxygenase and induction of stress-related proteins.Electrophoresis,2001,22:2824-2831.
Harfouche AL, Rugini E, Mencarelli F, et al. Salicylic acid induces H2O2production and endochitinase geneexpression but not ethylene biosynthesis in Castanea sativa in vitro model system. J Plant Physiol,2008,165:734-744.
Hennig J, Dewey RE, Cutt JR, et al. Pathogen, salicylic acid and developmental dependent expression of abeta-1,3-glucanase/GUS gene fusion in transgenic tobacco plants. Plant J,1993,4:481-493.
Hippauf F, Michalsky E, Huang R, et al. Enzymatic, expression and structural divergences among carboxylO-methyltransferases after gene duplication and speciation in Nicotiana. Plant Mol Biol,2010,72:311-330.
Hiruma K, Nishiuchi T, Kato T, et al. Arabidopsis ENHANCED DISEASE RESISTANCE1is required forpathogen-induced expression of plant defensins in nonhost resistance, and acts through interferenceof MYC2-mediated repressor function. Plant J,2011,67:980-992.
Huang J, Cardoza YJ, Schmelz EA, et al. Differential volatile emissions and salicylic acid levels fromtobacco plants in response to different strains of Pseudomonas syringae. Planta,2003,217:767-775.
Huang J, Gu M, Lai Z, et al. Functional analysis of the Arabidopsis PAL gene family in plant growth,development, and response to environmental stress. Plant Physiol,2010,153:1526-1538.
Iwai T, Seo S, Mitsuhara I, et al. Probenazole-induced accumulation of salicylic acid confers resistance toMagnaporthe grisea in adult rice plants. Plant Cell Physiol,2007,48:915-924.
Kachroo P, Yoshioka K, Shah J, et al. Resistance to turnip crinkle virus in Arabidopsis is regulated by twohost genes and is salicylic acid dependent but NPR1, ethylene, and jasmonate independent. PlantCell,2000,12:677-690.
Klessig DF, Durner J, Noad R, et al. Nitric oxide and salicylic acid signaling in plant defense. Proc NatlAcad Sci U S A,2000,97:8849-8855.
Leon J, Lawton MA, Raskin I. Hydrogen Peroxide Stimulates Salicylic Acid Biosynthesis in Tobacco. PlantPhysiol,1995,108:1673-1678.
Ma Z, Michailides TJ. Characterization of Botryosphaeria dothidea Isolates Collected from Pistachio andOther Plant Hosts in California. Phytopathology,2002,92:519-526.
Malamy J, Carr JP, Klessig DF, et al. Salicylic Acid: a likely endogenous signal in the resistance response oftobacco to viral infection. Science,1990,250:1002-1004.
Manosalva PM, Park SW, Forouhar F, et al. Methyl esterase1(StMES1) is required for systemic acquiredresistance in potato. Mol Plant Microbe Interact,2010,23:1151-1163.
Park CJ, Kim KJ, Shin R, et al. Pathogenesis-related protein10isolated from hot pepper functions as aribonuclease in an antiviral pathway. Plant J,2004,37:186-198.
Park SW, Kaimoyo E, Kumar D, et al. Methyl salicylate is a critical mobile signal for plant systemicacquired resistance. Science,2007,318:113-116.
Park SW, Liu PP, Forouhar F, et al. Use of a synthetic salicylic acid analog to investigate the roles of methylsalicylate and its esterases in plant disease resistance. J Biol Chem,2009,284:7307-7317.
Pott MB, Effmert U, Piechulla B. Transcriptional and post-translational regulation ofS-adenosyl-L-methionine: salicylic acid carboxyl methyltransferase (SAMT) during Stephanotisfloribunda flower development. J Plant Physiol,2003,160:635-643.
Rasmussen JB, Hammerschmidt R, Zook MN. Systemic Induction of Salicylic Acid Accumulation inCucumber after Inoculation with Pseudomonas syringae pv syringae. Plant Physiol,1991,97:1342-1347.
Reguera M, Bonilla I, Bolanos L. Boron deficiency results in induction of pathogenesis-related proteins fromthe PR-10family during the legume-rhizobia interaction. J Plant Physiol,2010,167:625-632.
Ryals J, Uknes S, Ward E. Systemic Acquired Resistance. Plant Physiol,1994,104:1109-1112.
Sabater-Jara AB, Almagro L, Belchi-Navarro S, et al. Induction of sesquiterpenes, phytoesterols andextracellular pathogenesis-related proteins in elicited cell cultures of Capsicum annuum. J PlantPhysiol,2010,167:1273-1281.
Sels J, Mathys J, De Coninck BM, et al. Plant pathogenesis-related (PR) proteins: a focus on PR peptides.Plant Physiol Biochem,2008,46:941-950.
Shadle GL, Wesley SV, Korth KL, et al. Phenylpropanoid compounds and disease resistance in transgenictobacco with altered expression of L-phenylalanine ammonia-lyase. Phytochemistry,2003,64:153-161.
Shulaev V, Leon J, Raskin I. Is Salicylic Acid a Translocated Signal of Systemic Acquired Resistance inTobacco? Plant Cell,1995,7:1691-1701.
Tripathi D, Jiang YL, Kumar D. SABP2, a methyl salicylate esterase is required for the systemic acquiredresistance induced by acibenzolar-S-methyl in plants. FEBS Lett,2010,584:3458-3463.
Vallet C, Chabbert B, Czaninski Y, et al. Extractibility of structural carbohydrates and lignin deposition inmaturing alfalfa internodes. Int J Biol Macromol,1997,21:201-206.
Verberne MC, Brouwer N, Delbianco F, et al. Method for the extraction of the volatile compound salicylicacid from tobacco leaf material. Phytochem Anal,2002,13:45-50.
Vlot AC, Dempsey DA, Klessig DF. Salicylic Acid, a multifaceted hormone to combat disease. Annu RevPhytopathol,2009,47:177-206.
Vlot AC, Klessig DF, Park SW. Systemic acquired resistance: the elusive signal(s). Curr Opin Plant Biol,2008,11:436-442.
War AR, Paulraj MG, War MY, et al. Role of salicylic acid in induction of plant defense system in chickpea(Cicer arietinum L.). Plant Signal Behav,2011,6.
Wubben MJ, Jin J, Baum TJ. Cyst nematode parasitism of Arabidopsis thaliana is inhibited by salicylic acid(SA) and elicits uncoupled SA-independent pathogenesis-related gene expression in roots. MolPlant Microbe Interact,2008,21:424-432.
Yafei C, Yong Z, Xiaoming Z, et al. Functions of oligochitosan induced protein kinase in tobacco mosaicvirus resistance and pathogenesis related proteins in tobacco. Plant Physiol Biochem,2009,47:724-731.
Yalpani N, Leon J, Lawton MA, et al. Pathway of Salicylic Acid Biosynthesis in Healthy andVirus-Inoculated Tobacco. Plant Physiol,1993,103:315-321.
Yalpani N, Raskin I. Salicylic acid: a systemic signal in induced plant disease resistance. Trends Microbiol,1993,1:88-92.
Yogo K, Tojima H, Ohno JY, et al. Identification of SAMT family proteins as substrates of MARCH11inmouse spermatids. Histochem Cell Biol,2011.
Zhang WP, Jiang B, Lou LN, et al. Impact of salicylic acid on the antioxidant enzyme system and hydrogenperoxide production in Cucumis sativus under chilling stress. Z Naturforsch C,2011,66:413-422.
Zhao N, Guan J, Forouhar F, et al. Two poplar methyl salicylate esterases display comparable biochemicalproperties but divergent expression patterns. Phytochemistry,2009,70:32-39.
Zhu J, Park KC. Methyl salicylate, a soybean aphid-induced plant volatile attractive to the predatorCoccinella septempunctata. J Chem Ecol,2005,31:1733-1746.
曹支敏，景耀，周芳．杨树溃疡病流行与土壤条件的关系．西北林学院学报，1991a，6(2)：55-9．
曹支敏，周芳，杨俊秀．杨树溃疡病流行规律与测报研究．森林病虫通讯，1991b，3：5-9．
丁义峰，刘萍．水杨酸诱导植物抗逆性研究进展.生物学教学．2011(11)：42-43．
丁秀英，张军，苏宝林，等．水杨酸在植物抗病中的作用．植物学通报，2001，18（2）：163-168．
杜建玲，刘红霞，于淑平．杨树不同种(品种)间抗溃疡病差异的比较．河北林果研究，2000，15(1)：55-60．
杜建玲，项蔚华，沈瑞祥．诱导杨树抗溃疡病机理的研究．林业科学研究，1995，8(1)：78-81．
高雪．植物苯丙氨酸解氨酶研究进展．现代农业科技，2009，(1)：31-32．
胡景江，朱玮，文建雷．杨树细胞壁HRGP和木质素的诱导积累与其对溃疡病抗性的研究．植物病理学报，1999，29(2)：151-156．
胡景江，朱玮，袁雪丽．溃疡病菌对杨树细胞壁中HRGP和木质素的诱导作用．西北林学院学报，1997，12(3)：14-17．
沈文飚，徐朗莱，叶茂炳，等．水杨酸对小麦叶片抗坏血酸过氧化物酶活性的抑制．南京农业大学学报，1998，21(3)：126-128．
苏晓华，张冰玉，黄秦军，等．我国林木基因工程研究进展和关键领域．林业科学，2003，39(5)：111-118．
理永霞，张星耀，严东辉，等．杨树接种Botryosphaeria dothidea溃疡病菌后蛋白质表达差异分析．南京林业大学学报（自然科学版），2011，35(4)：1-6．
李娜，陆海．植物抗坏血酸过氧化物酶基因家族研究进展．成都大学学报(自然科学版)，2011，30(2)：97-99．
梁军，姜俊清，刘会香，等．我国杨树与溃疡病菌互作的病理学研究．林业科学研究，2005，18(2):214-221.
梁元存，刘爱新．植物系统获得抗性中的信号．山东农业大学学报(自然科学版)，2001(02)：17-18．
苗则彦，赵奎华，刘长远，等．葡萄抗感白腐病品种PAL酶、PPO酶和SOD酶活性比较．沈阳农业大学学报，2003，34(3)：177-180．
钱万强，曹福祥，王猛，等．松材线虫侵染对马尾松体内有机酸及酶的影响．湖南林业科技，2009，36(3)：10-12．
许文力，徐克勤，吴玉斌，等．杨树溃疡病．江苏林业科技，1997(04)：5-6．
肖淑贤，高俊明．番茄诱导抗病性的研究进展．中国西部科技，2011(04)：42-43．
邢会琴，李敏权，徐秉良，等．过氧化物酶和苯丙氨酸解氨酶与苜蓿白粉病抗性的关系．草地学报，2007，l5(4)：37-38．
向玉英，花晓梅，赵经周．杨树溃疡病的发生及病原菌的生物学特性的研究．植物病理学报，1981，11(4)：27-33．
向玉英，魏舜明，侯艳．杨树溃疡病与树皮酚化合物关系的研究．森林病虫通讯，1993，1：5-7．
王颖，胡景江，朱玮，等．杨树溃疡病寄主诱导抗病性的研究．西北林学院学报，1996，11(1)：1-4．
王媛．杨树与溃疡病菌互作的细胞生物学、活性氧代谢及细胞过敏反应,中国林业科学研究院博士学位论文，2007，5-6．
王文天，彭少麟，李冬梅等．薇甘菊水杨酸羧甲基转移酶基因的分离鉴定及表达分析，热带亚热带植物学报，2009，17（5）：445-450．
王春丽，梁宗锁．外源刺激对植物次生代谢的调节及其信号转导途径研究进展．西北植物学报，2009(5)：11-12．
王金生．分子植物病理学.中国农业出版社，1999．
杨俊秀，李武汉，符毓秦，等．抗溃疡病杨树种类的调查研究．西北林学院学报，1990，5(4)：1-10．
阳传和．杨树溃疡病接种方法的比较．北京林学院学报，1985，（2)：76-81．
阳传和．树皮内酚类物质的含量及苯丙氨酸的活性与杨树抗溃疡病的关系．林业科学，1989，25(4)：311-316．
袁嗣令．国内外杨树病害研究概况，四川省永川森林病虫防治试验站，1984．
张星耀，骆有庆．中国森林重大生物灾害．北京：中国林业出版社，2003．
赵嘉平．树木溃疡病菌-葡萄座腔菌属及相关真菌系统分类研究．中国林业科学研究院博士学位论文．2007．5-6．
赵仕光，景耀．杨树对溃疡病的抗性研究(I)对龄及形态结构与抗病性．西北林学院学报，1997，12(3)：35-40．
钟兆康，高雅．不同杨树品种对杨树水泡型溃疡病菌的抗病性测定简报．林业科技通讯，1981，1：25-26．
中科院上海植物生理所主编．现代植物生理学实验指南．北京:科学出版社，1999．
赵仕光，景耀，杨俊秀．杨树树皮内过氧化物酶和多酚氧化酶活性与抗溃疡病的关系．西北林学院学报，1993，8(3)：13-17．
曾大鹏．我国杨树病害的研究现状与防治[J]．中国森林病虫，2002，21(1)：20-26．
郑伟尉，臧运祥．水杨酸(SA)与植物抗病研究进展．河北果树，2005(01)：23-25．
赵仕光，朱玮，岳红艳．溃疡病菌在杨树树皮组织中的扩展和对寄主细胞超微结构的影响．林业科学研究，1999，12(2)：11-26．
张佳娣．活性氧的信号传导途径．安徽农业科学，2010(16)：56-57．
Ament K, Kant MR, Sabelis MW, et al. Jasmonic acid is a key regulator of spider mite-induced volatileterpenoid and methyl salicylate emission in tomato. Plant Physiol,2004,135:2025-2037.
An C, Mou Z. Salicylic acid and its function in plant immunity. J Integr Plant Biol,2011,53:412-428.
Antoniw JF, Dunkley AM, White RF, et al. Soluble leaf proteins of virus-infected tobacco (Nicotianatabacum) cultivars [proceedings]. Biochem Soc Trans,1980,8:70-71.
Averyanov A. Oxidative burst and plant disease resistance. Front Biosci (Elite Ed),2009,1:142-152.
Bokszczanin KL, Przybyla AA.[Molecular aspects of allergy to plant products. Part II. Pathogenesis-relatedproteins (PRs), apple allergenicity governed by Mal d1gene]. Pol Merkur Lekarski,2012,32:176-181.
Bolwell BJ, Kalaycio M, Goormastic M, et al. Progressive disease after ABMT for Hodgkin's disease. BoneMarrow Transplant,1997,20:761-765.
Bonifacio A, Martins MO, Ribeiro CW, et al. Role of peroxidases in the compensation of cytosolic ascorbateperoxidase knockdown in rice plants under abiotic stress. Plant Cell Environ,2011,34:1705-1722.
Casano LM, Martin M, Sabater B. Hydrogen peroxide mediates the induction of chloroplastic Ndh complexunder photooxidative stress in barley. Plant Physiol,2001,125:1450-1458.
Charles A, Dawicki DD, Oldmixon E, et al. Studies on the mechanism of short-term regulation of pulmonaryartery endothelial cell Na/K pump activity. J Lab Clin Med,1997,130:157-168.
Chen Z, Ricigliano JW, Klessig DF. Purification and characterization of a soluble salicylic acid-bindingprotein from tobacco. Proc Natl Acad Sci U S A,1993,90:9533-9537.
Chen Z, Silva H, Klessig DF. Active oxygen species in the induction of plant systemic acquired resistance bysalicylic acid. Science,1993,262:1883-1886.
Chern M, Fitzgerald HA, Canlas PE, et al. Overexpression of a rice NPR1homolog leads to constitutiveactivation of defense response and hypersensitivity to light. Mol Plant Microbe Interact,2005,18:511-520.
Clarke JD, Volko SM, Ledford H, et al. Roles of salicylic acid, jasmonic acid, and ethylene in cpr-inducedresistance in arabidopsis. Plant Cell,2000,12:2175-2190.
Colditz F, Niehaus K, Krajinski F. Silencing of PR-10-like proteins in Medicago truncatula results in anantagonistic induction of other PR proteins and in an increased tolerance upon infection with theoomycete Aphanomyces euteiches. Planta,2007,226:57-71.
Dangl JL, Jones JD. Plant pathogens and integrated defence responses to infection. Nature,2001,411:826-833.
Dat JF, Pellinen R, Beeckman T, et al. Changes in hydrogen peroxide homeostasis trigger an active cell deathprocess in tobacco. Plant J,2003,33:621-632.
Davis JM, Wu H, Cooke JE, et al. Pathogen challenge, salicylic acid, and jasmonic acid regulate expressionof chitinase gene homologs in pine. Mol Plant Microbe Interact,2002,15:380-387.
Ding CK, Wang CY, Gross KC, et al. Jasmonate and salicylate induce the expression ofpathogenesis-related-protein genes and increase resistance to chilling injury in tomato fruit. Planta,2002,214:895-901.
Dixon RA, Achnine L, Kota P, et al. The phenylpropanoid pathway and plant defence-a genomicsperspective. Mol Plant Pathol,2002,3:371-390.
Du H, Klessig DF. Identification of a Soluble, High-Affinity Salicylic Acid-Binding Protein in Tobacco.Plant Physiol,1997,113:1319-1327.
Durner J, Klessig DF. Inhibition of ascorbate peroxidase by salicylic acid and2,6-dichloroisonicotinic acid,two inducers of plant defense responses. Proc Natl Acad Sci U S A,1995,92:11312-11316.
Durrant WE, Dong X. Systemic acquired resistance. Annu Rev Phytopathol,2004,42:185-209.
Forouhar F, Yang Y, Kumar D, et al. Structural and biochemical studies identify tobacco SABP2as a methylsalicylate esterase and implicate it in plant innate immunity. Proc Natl Acad Sci U S A,2005,102:1773-1778.
Fragniere C, Serrano M, Abou-Mansour E, et al. Salicylic acid and its location in response to biotic andabiotic stress. FEBS Lett,2011,585:1847-1852.
Fukami H, Asakura T, Hirano H, et al. Salicylic acid carboxyl methyltransferase induced in hairy rootcultures of Atropa belladonna after treatment with exogeneously added salicylic acid. Plant CellPhysiol,2002,43:1054-1058.
Glazebrook J. Genes controlling expression of defense responses in Arabidopsis--2001status. Curr OpinPlant Biol,2001,4:301-308.
Glocova I, Thor K, Roth B, et al. Salicylic acid (SA)-dependent gene activation can be uncoupled from celldeath-mediated gene activation: the SA-inducible NIMIN-1and NIMIN-2promoters, unlike thePR-1a promoter, do not respond to cell death signals in tobacco. Mol Plant Pathol,2005,6:299-314.
Gonzalez-Teuber M, Eilmus S, Muck A, et al. Pathogenesis-related proteins protect extrafloral nectar frommicrobial infestation. Plant J,2009,58:464-473.
Hajduch M, Rakwal R, Agrawal GK, et al. High-resolution two-dimensional electrophoresis separation ofproteins from metal-stressed rice (Oryza sativa L.) leaves: drastic reductions/fragmentation ofribulose-1,5-bisphosphate carboxylase/oxygenase and induction of stress-related proteins.Electrophoresis,2001,22:2824-2831.
Harfouche AL, Rugini E, Mencarelli F, et al. Salicylic acid induces H2O2production and endochitinase geneexpression but not ethylene biosynthesis in Castanea sativa in vitro model system. J Plant Physiol,2008,165:734-744.
Hennig J, Dewey RE, Cutt JR, et al. Pathogen, salicylic acid and developmental dependent expression of abeta-1,3-glucanase/GUS gene fusion in transgenic tobacco plants. Plant J,1993,4:481-493.
Hippauf F, Michalsky E, Huang R, et al. Enzymatic, expression and structural divergences among carboxylO-methyltransferases after gene duplication and speciation in Nicotiana. Plant Mol Biol,2010,72:311-330.
Hiruma K, Nishiuchi T, Kato T, et al. Arabidopsis ENHANCED DISEASE RESISTANCE1is required forpathogen-induced expression of plant defensins in nonhost resistance, and acts through interferenceof MYC2-mediated repressor function. Plant J,2011,67:980-992.
Huang J, Cardoza YJ, Schmelz EA, et al. Differential volatile emissions and salicylic acid levels fromtobacco plants in response to different strains of Pseudomonas syringae. Planta,2003,217:767-775.
Huang J, Gu M, Lai Z, et al. Functional analysis of the Arabidopsis PAL gene family in plant growth,development, and response to environmental stress. Plant Physiol,2010,153:1526-1538.
Iwai T, Seo S, Mitsuhara I, et al. Probenazole-induced accumulation of salicylic acid confers resistance toMagnaporthe grisea in adult rice plants. Plant Cell Physiol,2007,48:915-924.
Kachroo P, Yoshioka K, Shah J, et al. Resistance to turnip crinkle virus in Arabidopsis is regulated by twohost genes and is salicylic acid dependent but NPR1, ethylene, and jasmonate independent. PlantCell,2000,12:677-690.
Klessig DF, Durner J, Noad R, et al. Nitric oxide and salicylic acid signaling in plant defense. Proc NatlAcad Sci U S A,2000,97:8849-8855.
Leon J, Lawton MA, Raskin I. Hydrogen Peroxide Stimulates Salicylic Acid Biosynthesis in Tobacco. PlantPhysiol,1995,108:1673-1678.
Ma Z, Michailides TJ. Characterization of Botryosphaeria dothidea Isolates Collected from Pistachio andOther Plant Hosts in California. Phytopathology,2002,92:519-526.
Malamy J, Carr JP, Klessig DF, et al. Salicylic Acid: a likely endogenous signal in the resistance response oftobacco to viral infection. Science,1990,250:1002-1004.
Manosalva PM, Park SW, Forouhar F, et al. Methyl esterase1(StMES1) is required for systemic acquiredresistance in potato. Mol Plant Microbe Interact,2010,23:1151-1163.
Park CJ, Kim KJ, Shin R, et al. Pathogenesis-related protein10isolated from hot pepper functions as aribonuclease in an antiviral pathway. Plant J,2004,37:186-198.
Park SW, Kaimoyo E, Kumar D, et al. Methyl salicylate is a critical mobile signal for plant systemicacquired resistance. Science,2007,318:113-116.
Park SW, Liu PP, Forouhar F, et al. Use of a synthetic salicylic acid analog to investigate the roles of methylsalicylate and its esterases in plant disease resistance. J Biol Chem,2009,284:7307-7317.
Pott MB, Effmert U, Piechulla B. Transcriptional and post-translational regulation ofS-adenosyl-L-methionine: salicylic acid carboxyl methyltransferase (SAMT) during Stephanotisfloribunda flower development. J Plant Physiol,2003,160:635-643.
Rasmussen JB, Hammerschmidt R, Zook MN. Systemic Induction of Salicylic Acid Accumulation inCucumber after Inoculation with Pseudomonas syringae pv syringae. Plant Physiol,1991,97:1342-1347.
Reguera M, Bonilla I, Bolanos L. Boron deficiency results in induction of pathogenesis-related proteins fromthe PR-10family during the legume-rhizobia interaction. J Plant Physiol,2010,167:625-632.
Ryals J, Uknes S, Ward E. Systemic Acquired Resistance. Plant Physiol,1994,104:1109-1112.
Sabater-Jara AB, Almagro L, Belchi-Navarro S, et al. Induction of sesquiterpenes, phytoesterols andextracellular pathogenesis-related proteins in elicited cell cultures of Capsicum annuum. J PlantPhysiol,2010,167:1273-1281.
Sels J, Mathys J, De Coninck BM, et al. Plant pathogenesis-related (PR) proteins: a focus on PR peptides.Plant Physiol Biochem,2008,46:941-950.
Shadle GL, Wesley SV, Korth KL, et al. Phenylpropanoid compounds and disease resistance in transgenictobacco with altered expression of L-phenylalanine ammonia-lyase. Phytochemistry,2003,64:153-161.
Shulaev V, Leon J, Raskin I. Is Salicylic Acid a Translocated Signal of Systemic Acquired Resistance inTobacco? Plant Cell,1995,7:1691-1701.
Tripathi D, Jiang YL, Kumar D. SABP2, a methyl salicylate esterase is required for the systemic acquiredresistance induced by acibenzolar-S-methyl in plants. FEBS Lett,2010,584:3458-3463.
Vallet C, Chabbert B, Czaninski Y, et al. Extractibility of structural carbohydrates and lignin deposition inmaturing alfalfa internodes. Int J Biol Macromol,1997,21:201-206.
Verberne MC, Brouwer N, Delbianco F, et al. Method for the extraction of the volatile compound salicylicacid from tobacco leaf material. Phytochem Anal,2002,13:45-50.
Vlot AC, Dempsey DA, Klessig DF. Salicylic Acid, a multifaceted hormone to combat disease. Annu RevPhytopathol,2009,47:177-206.
Vlot AC, Klessig DF, Park SW. Systemic acquired resistance: the elusive signal(s). Curr Opin Plant Biol,2008,11:436-442.
War AR, Paulraj MG, War MY, et al. Role of salicylic acid in induction of plant defense system in chickpea(Cicer arietinum L.). Plant Signal Behav,2011,6.
Wubben MJ, Jin J, Baum TJ. Cyst nematode parasitism of Arabidopsis thaliana is inhibited by salicylic acid(SA) and elicits uncoupled SA-independent pathogenesis-related gene expression in roots. MolPlant Microbe Interact,2008,21:424-432.
Yafei C, Yong Z, Xiaoming Z, et al. Functions of oligochitosan induced protein kinase in tobacco mosaicvirus resistance and pathogenesis related proteins in tobacco. Plant Physiol Biochem,2009,47:724-731.
Yalpani N, Leon J, Lawton MA, et al. Pathway of Salicylic Acid Biosynthesis in Healthy andVirus-Inoculated Tobacco. Plant Physiol,1993,103:315-321.
Yalpani N, Raskin I. Salicylic acid: a systemic signal in induced plant disease resistance. Trends Microbiol,1993,1:88-92.
Yogo K, Tojima H, Ohno JY, et al. Identification of SAMT family proteins as substrates of MARCH11inmouse spermatids. Histochem Cell Biol,2011.
Zhang WP, Jiang B, Lou LN, et al. Impact of salicylic acid on the antioxidant enzyme system and hydrogenperoxide production in Cucumis sativus under chilling stress. Z Naturforsch C,2011,66:413-422.
Zhao N, Guan J, Forouhar F, et al. Two poplar methyl salicylate esterases display comparable biochemicalproperties but divergent expression patterns. Phytochemistry,2009,70:32-39.
Zhu J, Park KC. Methyl salicylate, a soybean aphid-induced plant volatile attractive to the predatorCoccinella septempunctata. J Chem Ecol,2005,31:1733-1746.