Ethylene and jasmonic acid signaling affect the NPR1-independent expression of defense genes without impacting resistance to Pseudomonas syringae and Peronospora parasitica in the Arabidopsis ssi1 mutant

Salicylic acid (SA), ethylene, and jasmonic acid (JA) are important signaling molecules in plant defense to biotic stress. An intricate signaling network involving SA, ethylene, and JA fine tunes plant defense responses. SA-dependent defense responses in Arabidopsis thaliana are mediated through NPR1-dependent and -independent mechanisms. We have previously shown that activation of an NPR1-independent defense mechanism confers enhanced disease resistance and constitutive expression of the pathogenesis-related (PR) genes in the Arabidopsis ssi1 mutant. In addition, the ssi1 mutant constitutively expresses the defensin gene PDF1.2. Moreover, SA is required for the ssi1-conferred constitutive expression of PDF1.2 in addition to PR genes. Hence, the ssi1 mutant appears to target a step common to SA- and ethylene- or JA-regulated defense pathways. In the present study, we show that, in addition to SA, ethylene and JA signaling also are required for the ssi1-conferred constitutive expression of PDF1.2 and the NPR1-independent expression of PR-1. Furthermore, the ethylene-insensitive ein2 and JA-insensitive jar1 mutants enhance susceptibility of ssi1 plants to the necrotrophic fungus Botrytis cinerea. However, defects in either the ethylene- or JA-signaling pathways do not compromise ssi1-conferred resistance to the bacterial pathogen Pseudomonas synringae pv. maculicola and the oomycete pathogen Peronospora parasitica. Interestingly, ssi1 exhibits a marginal increase in the levels of ethylene and JA, suggesting that low endogenous levels of these phytohormones are sufficient to activate expression of defense genes. Taken together, our results indicate that although cross talk in ssi1 renders expression of ethylene- or JA-responsive defense genes sensitive to SA and vice versa, it does not affect downstream signaling leading to resistance.     

Heterotrimeric G proteins-mediated resistance to necrotrophic pathogens includes mechanisms independent of salicylic acid-, jasmonic acid/ethylene- and abscisic acid-mediated defense signaling

Heterotrimeric G proteins are involved in the defense response against necrotrophic fungi in Arabidopsis. In order to elucidate the resistance mechanisms involving heterotrimeric G proteins, we analyzed the effects of the Gβ (subunit deficiency in the mutant agb1-2 on pathogenesis-related gene expression, as well as the genetic interaction between agb1-2 and a number of mutants of established defense pathways. Gβ-mediated signaling suppresses the induction of salicylic acid (SA)-, jasmonic acid (JA)-, ethylene (ET)- and abscisic acid (ABA)-dependent genes during the initial phase of the infection with Fusarium oxysporum (up to 48 h after inoculation). However, at a later phase it enhances JA/ET-dependent genes such as PDF1.2 and PR4 . Quantification of the Fusarium wilt symptoms revealed that Gβ- and SA-deficient mutants were more susceptible than wild-type plants, whereas JA- and ET-insensitive and ABA-deficient mutants demonstrated various levels of resistance. Analysis of the double mutants showed that the Gβ-mediated resistance to F. oxysporum and Alternaria brassicicola was mostly independent of all of the previously mentioned pathways. However, the progressive decay of agb1-2 mutants was compensated by coi1-21 and jin1-9 mutations, suggesting that at this stage of F. oxysporum infection Gβ acts upstream of COI1 and ATMYC2 in JA signaling.     

Endophytic actinobacteria induce defense pathways in Arabidopsis thaliana

Endophytic actinobacteria, isolated from healthy wheat tissue, which are capable of suppressing a number wheat fungal pathogens both in vitro and in planta, were investigated for the ability to activate key genes in the systemic acquired resistance (SAR) or the jasmonate/ethylene (JA/ET) pathways in Arabidopsis thaliana. Inoculation of A. thaliana (Col-0) with selected endophytic strains induced a low level of SAR and JA/ET gene expression, measured using quantitative polymerase chain reaction. Upon pathogen challenge, endophyte-treated plants demonstrated a higher abundance of defense gene expression compared with the non-endophyte-treated controls. Resistance to the bacterial pathogen Erwinia carotovora subsp. carotovora required the JA/ET pathway. On the other hand, resistance to the fungal pathogen Fusarium oxysporum involved primarily the SAR pathway. The endophytic actinobacteria appear to be able to "prime" both the SAR and JA/ET pathways, upregulating genes in either pathway depending on the infecting pathogen. Culture filtrates of the endophytic actinobacteria were investigated for the ability to also activate defense pathways. The culture filtrate of Micromonospora sp. strain EN43 grown in a minimal medium resulted in the induction of the SAR pathway; however, when grown in a complex medium, the JA/ET pathway was activated. Further analysis using Streptomyces sp. strain EN27 and defense-compromised mutants of A. thaliana indicated that resistance to E. carotovora subsp. carotovora occurred via an NPR1-independent pathway and required salicylic acid whereas the JA/ET signaling molecules were not essential. In contrast, resistance to F. oxysporum mediated by Streptomyces sp. strain EN27 occurred via an NPR1-dependent pathway but also required salicylic acid and was JA/ET independent.     

PDF1.2基因在转草酸氧化酶基因甘蓝型油菜中的表达

采用RT-PCR方法检测茉莉酸（JA）／乙烯（ET）依赖性信号途径中关键基因PDF1。2在转草酸氧化酶基因（OXO）油菜株系与未转化对照中的表达差异。结果表明,在转基因油菜不同株系中PDF1．2都有不同程度的上调表达,预示着转OXO油菜对菌核病的抗性增强可能与激活JA／ET依赖性信号途径有关。     

Environmentally sensitive, SA-dependent defense responses in the cpr22 mutant of Arabidopsis

To investigate the signaling pathways through which defense responses are activated following pathogen infection, we have isolated and characterized the cpr22 mutant. This plant carries a semidominant, conditional lethal mutation that confers constitutive expression of the pathogenesis-related (PR) genes PR-1, PR-2, PR-5 and the defensin gene PDF1.2. cpr22 plants also display spontaneous lesion formation, elevated levels of salicylic acid (SA) and heightened resistance to Peronospora parasitica Emco5. The cpr22 locus was mapped to chromosome 2, approximately 2 cM telomeric to the AthB102 marker. By analyzing the progeny of crosses between cpr22 plants and either NahG transgenic plants or npr1 mutants, all of the cpr22-associated phenotypes except PDF1.2 expression were found to be SA dependent. However, the SA signal transducer NPR1 was required only for constitutive PR-1 expression. A cross between cpr22 and ndr1-1 mutants revealed that enhanced resistance to P. parasitica is mediated by an NDR1-dependent pathway, while the other cpr22-induced defenses are not. Crosses between either coi1-1 or etr1-1 mutants further demonstrated that constitutive PDF1.2 expression is mediated by a JA- and ethylene-dependent pathway. Based on these results, the cpr22 mutation appears to induce its associated phenotypes by activating NPR1-dependent and NPR1-independent branches of the SA pathway, as well as an ethylene/JA signaling pathway. Interestingly, the SA-dependent phenotypes, but not the SA-independent phenotypes, are suppressed when cpr22 mutants are grown under high humidity.     

Interacting signal pathways control defense gene expression in Arabidopsis in response to cell wall-degrading enzymes from Erwinia carotovora

We have characterized the role of salicylic acid (SA)-independent defense signaling in Arabidopsis thaliana in response to the plant pathogen Erwinia carotovora subsp. carotovora. Use of pathway-specific target genes as well as signal mutants allowed us to elucidate the role and interactions of ethylene, jasmonic acid (JA), and SA signal pathways in this response. Gene expression studies suggest a central role for both ethylene and JA pathways in the regulation of defense gene expression triggered by the pathogen or by plant cell wall-degrading enzymes (CF) secreted by the pathogen. Our results suggest that ethylene and JA act in concert in this regulation. In addition, CF triggers another, strictly JA-mediated response inhibited by ethylene and SA. SA does not appear to have a major role in activating defense gene expression in response to CF. However, SA may have a dual role in controlling CF-induced gene expression, by enhancing the expression of genes synergistically induced by ethylene and JA and repressing genes induced by JA alone.     

拟南芥对细菌性软腐病抗性变异分析

由胡萝卜软腐欧文氏菌胡萝卜软腐亚种 Erwinia carotovora subsp.carotovora （Ecc）引起的细菌性软腐病是世界性的重要流行病害,由于缺乏天然抗源,研究进展缓慢,拟南芥成为软腐病抗性研究的主要试材。本文选用 29 份拟南芥材料,制定了病情分级标准,以接种后 48 小时的病情指数作为软腐病抗性的鉴定指标,筛选出抗软腐病材料 CS906 和感病材料 CS20。通过分析抗、感材料接种 Ecc 后 AOS、ERF1-1、PR1、PDF1.2 和 PAL1 等 5 个基因的表达变化,发现 SA、ET 和 MJ 信号途径都参与了拟南芥对 Ecc 的防卫反应,且基因表达模式在抗、感材料中相似,差异体现在表达量上。本研究对深入探讨拟南芥软腐病抗病机制具有重要意义。     

Signaling pathways that regulate the enhanced disease resistance of Arabidopsis "defense, no death" mutants

Arabidopsis dnd1 and dnd2 mutants lack cyclic nucleotide-gated ion channel proteins and carry out avirulence or resistance gene-mediated defense with a greatly reduced hypersensitive response (HR). They also exhibit elevated broad-spectrum disease resistance and constitutively elevated salicylic acid (SA) levels. We examined the contributions of NPR1, SID2 (EDS16), NDR1, and EIN2 to dnd phenotypes. Mutations that affect SA accumulation or signaling (sid2, npr1, and ndr1) abolished the enhanced resistance of dnd mutants against Pseudomonas syringae pv. tomato and Hyaloperonospora parasitica but not Botrytis cinerea. When SA-associated pathways were disrupted, the constitutive activation of NPR1-dependent and NPR1-independent and SA-dependent pathways was redirected toward PDF1.2-associated pathways. This PDF1.2 overexpression was downregulated after infection by P. syringae. Disruption of ethylene signaling abolished the enhanced resistance to B. cinerea but not P. syringae or H. parasitica. However, loss of NPR1, SID2, NDR1, or EIN2 did not detectably alter the reduced HR in dnd mutants. The susceptibility of dnd ein2 plants to B. cinerea despite their reduced-HR phenotype suggests that cell death repression is not the primary cause of dnd resistance to necrotrophic pathogens. The partial restoration of resistance to B. cinerea in dnd1 npr1 ein2 triple mutants indicated that this resistance is not entirely EIN2 dependent. The above findings indicate that the broad-spectrum resistance of dnd mutants occurs due to activation or sensitization of multiple defense pathways, yet none of the investigated pathways are required for the reduced-HR phenotype.     

beta-Aminobutyric acid-induced protection of Arabidopsis against the necrotrophic fungus Botrytis cinerea

The non-protein amino acid beta-aminobutyric acid (BABA) protects numerous plants against various pathogens. Protection of Arabidopsis plants against virulent pathogens involves the potentiation of pathogen-specific defense responses. To extend the analysis of the mode of action of BABA to necrotrophs we evaluated the effect of this chemical on Arabidopsis plants infected with the gray mold fungus Botrytis cinerea. BABA-treated Arabidopsis were found to be less sensitive to two different strains of this pathogen. BABA protected mutants defective in the jasmonate and ethylene pathways, but was inactive in plants impaired in the systemic acquired resistance transduction pathway. Treatments with benzo-(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester, a functional analog of salicylic acid (SA), also markedly reduced the level of infection. Moreover, BABA potentiated mRNA accumulation of the SA-associated PR-1, but not the jasmonate/ethylene-dependent PDF1.2 gene. Thus, besides jasmonate/ethylene-dependent defense responses, SA-dependent signaling also contributes to restrict B. cinerea infection in Arabidopsis. Our results also suggest that SA-dependent signaling is down-regulated after infection by B. cinerea. The observed up-regulation of the PDF1.2 gene in mutants defective in the SA-dependent signaling pathway points to a cross-talk between SA- and jasmonate/ethylene-dependent signaling pathways during pathogen ingress.     

激发子α-吡啶羧酸诱发拟南芥和水稻悬浮细胞中依赖于水杨酸、茉莉酸/乙烯和钙离子途径的防卫反应

α-吡啶羧酸(PA)是动物细胞程序化死亡的诱导物.我们前期的研究表明,PA可以激发单子叶模式植物水稻的过敏反应(HR).进一步用双子叶模式植物拟南芥(Arabidopsis thaliana)进行的研究表明,PA是一个广谱的植物HR反应的激发子,包括诱导氧进发和细胞死亡.我们探究了PA诱导的拟南芥防卫反应途径,利用不同信号途径标志基因PR-1,PR-2和PDF1.2受诱导剂量和时间激活的结果,表明PA可以同时激活水杨酸和茉莉酸/乙烯依赖的防卫途径.我们也发现PA诱导水稻悬浮细胞产生活性氧是钙离子依赖性的.综合所有结果,我们认为PA可以作为一个非专化性的植物防卫反应激发子,可望用于系统获得性抗性激发的细胞模型的建立.     

Differential effectiveness of salicylate-dependent and jasmonate/ethylene-dependent induced resistance in Arabidopsis

Salicylic acid (SA), jasmonic acid (JA), and ethylene (ET) are each involved in the regulation of basal resistance against different pathogens. These three signals play important roles in induced resistance as well. SA is a key regulator of pathogen-induced systemic acquired resistance (SAR), whereas JA and ET are required for rhizobacteria-mediated induced systemic resistance (ISR). Both types of induced resistance are effective against a broad spectrum of pathogens. In this study, we compared the spectrum of effectiveness of SAR and ISR using an oomycete, a fungal, a bacterial, and a viral pathogen. In noninduced Arabidopsis plants, these pathogens are primarily resisted through either SA-dependent basal resistance (Peronospora parasitica and Turnip crinkle virus [TCV]), JA/ET-dependent basal resistance responses (Alternaria brassicicola), or a combination of SA-, JA-, and ET-dependent defenses (Xanthomonas campestris pv. armoraciae). Activation of ISR resulted in a significant level of protection against A. brassicicola, whereas SAR was ineffective against this pathogen. Conversely, activation of SAR resulted in a high level of protection against P. parasitica and TCV, whereas ISR conferred only weak and no protection against P. parasitica and TCV, respectively. Induction of SAR and ISR was equally effective against X. campestris pv. armoraciae. These results indicate that SAR is effective against pathogens that in noninduced plants are resisted through SA-dependent defenses, whereas ISR is effective against pathogens that in noninduced plants are resisted through JA/ET-dependent defenses. This suggests that SAR and ISR constitute a reinforcement of extant SA- or JA/ET-dependent basal defense responses, respectively.     

The plant growth-promoting rhizobacterium Bacillus cereus AR156 induces systemic resistance in Arabidopsis thaliana by simultaneously activating salicylate- and jasmonate/ethylene-dependent signaling pathways

Bacillus cereus AR156 is a plant growth-promoting rhizobacterium that induces resistance against a broad spectrum of pathogens including Pseudomonas syringae pv. tomato DC3000. This study analyzed AR156-induced systemic resistance (ISR) to DC3000 in Arabidopsis ecotype Col-0 plants. Compared with mock-treated plants, AR156-treated ones showed an increase in biomass and reductions in disease severity and pathogen density in the leaves. The defense-related genes PR1, PR2, PR5, and PDF1.2 were concurrently expressed in the leaves of AR156-treated plants, suggesting simultaneous activation of the salicylic acid (SA)- and the jasmonic acid (JA)- and ethylene (ET)-dependent signaling pathways by AR156. The above gene expression was faster and stronger in plants treated with AR156 and inoculated with DC3000 than that in plants only inoculated with DC3000. Moreover, the cellular defense responses hydrogen peroxide accumulation and callose deposition were induced upon challenge inoculation in the leaves of Col-0 plants primed by AR156. Also, pretreatment with AR156 led to a higher level of induced protection against DC3000 in Col-0 than that in the transgenic NahG, the mutant jar1 or etr1, but the protection was absent in the mutant npr1. Therefore, AR156 triggers ISR in Arabidopsis by simultaneously activating the SA- and JA/ET-signaling pathways in an NPR1-dependent manner that leads to an additive effect on the level of induced protection.     

Induction of systemic resistance in Arabidopsis thaliana in response to a culture filtrate from a plant growth-promoting fungus, Phoma sp. GS8-3

The plant growth-promoting fungus (PGPF), Phoma sp. GS8-3, isolated from a zoysia grass rhizosphere, is capable of protecting cucumber plants against virulent pathogens. This fungus was investigated in terms of the underlying mechanisms and ability to elicit systemic resistance in Arabidopsis thaliana . Root treatment of Arabidopsis plants with a culture filtrate (CF) from Phoma sp. GS8-3 elicited systemic resistance against the bacterial speck pathogen Pseudomonas syringae pv. tomato DC3000 ( Pst ), with restricted disease development and inhibited pathogen proliferation. Pathway-specific mutant plants, such as jar1 (jasmonic acid insensitive) and ein2 (ethylene insensitive), and transgenic NahG plants (impaired in salicylate signalling) were protected after application of the CF, demonstrating that these pathways are dispensable (at least individually) in CF-mediated resistance. Similarly, NPR1 interference in npr1 mutants had no effect on CF-induced resistance. Gene expression studies revealed that CF treatment stimulated the systemic expression of both the SA-inducible PR-1 and JA/ET-inducible PDF1.2 genes. However, pathogenic challenge to CF-treated plants was associated with potentiated expression of the PR-1 gene and down-regulated expression of the PDF1.2 gene. The observed down-regulation of the PDF1.2 gene in CF-treated plants indicates that there may be cross-talk between SA- and JA/ET-dependent signalling pathways during the pathogenic infection process. In conclusion, our data suggest that CF of Phoma sp. GS8-3 induces resistance in Arabidopsis in a manner where SA and JA/ET may play a role in defence signalling.     

Salicylic acid and the plant pathogen Erwinia carotovora induce defense genes via antagonistic pathways

Infection of tobacco plants with the plant pathogenic bacterium Erwinia carotovora subsp. carotovora or treatment of plants with Erwinia -derived elicitor preparations leads to the induction of a number of genes thought to play a role in plant defense response to pathogens. In order to determine the role of salicylic acid (SA) in the induction of the Erwinia responsive genes, the accumulation of mRNAs for these and other genes encoding pathogenesis-related proteins (PR genes) in response to both Erwinia elicitors and SA was determined. PR genes were identified which were preferentially induced by Erwinia elicitor preparations, one gene was induced by SA but not by Erwinia , and another gene was induced by both type of treatments. The differential expression of these genes and the timing of induction suggest that SA is not the signal molecule leading to the early response of plants to Erwinia . This was demonstrated by experiments using transgenic NahG plants that overproduce a salicylate hydroxylase inactivating SA. The elicitation of PR genes by Erwinia was similar in NahG and wild-type plants. Therefore, induction of plant defense genes by Erwinia and SA seems to be by two distinct pathways leading to expression of separate sets of genes. Furthermore, we could demonstrate that Erwinia elicitors antagonize the SA-mediated induction of PR genes. Similarly, SA appeared to inhibit the induction of PR genes elicited by Erwinia . The observed antagonism between the two signal transduction pathways indicates the presence of a common regulatory element in both pathways that acts downstream of SA in the SA-mediated response.