A developmental response to pathogen infection in Arabidopsis

We present evidence that susceptible Arabidopsis plants accelerate their reproductive development and alter their shoot architecture in response to three different pathogen species. We infected 2-week-old Arabidopsis seedlings with two bacterial pathogens, Pseudomonas syringae and Xanthomonas campestris, and an oomycete, Peronospora parasitica. Infection with each of the three pathogens reduced time to flowering and the number of aerial branches on the primary inflorescence. In the absence of competition, P. syringae and P. parasitica infection also increased basal branch development. Flowering time and branch responses were affected by the amount of pathogen present. Large amounts of pathogen caused the most dramatic changes in the number of branches on the primary inflorescence, but small amounts of P. syringae caused the fastest flowering and the production of the most basal branches. RPS2 resistance prevented large changes in development when it prevented visible disease symptoms but not at high pathogen doses and when substantial visible hypersensitive response occurred. These experiments indicate that phylogenetically disparate pathogens cause similar changes in the development of susceptible Arabidopsis. We propose that these changes in flowering time and branch architecture constitute a general developmental response to pathogen infection that may affect tolerance of and/or resistance to disease.     

A high-throughput chemical screen for resistance to Pseudomonas syringae in Arabidopsis

The study of plant pathogenesis and the development of effective treatments to protect plants from diseases could be greatly facilitated by a high-throughput pathosystem to evaluate small-molecule libraries for inhibitors of pathogen virulence. The interaction between the Gram-negative bacterium Pseudomonas syringae and Arabidopsis thaliana is a model for plant pathogenesis. However, a robust high-throughput assay to score the outcome of this interaction is currently lacking. We demonstrate that Arabidopsis seedlings incubated with P. syringae in liquid culture display a macroscopically visible 'bleaching' symptom within 5 days of infection. Bleaching is associated with a loss of chlorophyll from cotyledonary tissues, and is correlated with bacterial virulence. Gene-for-gene resistance is absent in the liquid environment, possibly because of the suppression of the hypersensitive response under these conditions. Importantly, bleaching can be prevented by treating seedlings with known inducers of plant defence, such as salicylic acid (SA) or a basal defence-inducing peptide of bacterial flagellin (flg22) prior to inoculation. Based on these observations, we have devised a high-throughput liquid assay using standard 96-well plates to investigate the P. syringae-Arabidopsis interaction. An initial screen of small molecules active on Arabidopsis revealed a family of sulfanilamide compounds that afford protection against the bleaching symptom. The most active compound, sulfamethoxazole, also reduced in planta bacterial growth when applied to mature soil-grown plants. The whole-organism liquid assay provides a novel approach to probe chemical libraries in a high-throughput manner for compounds that reduce bacterial virulence in plants.     

拟南芥AtWRKY25、AtWRKY26和AtWRKY33在非生物胁迫条件下的表达分析

WRKY 转录因子家族在调控植物逆境诱导反应、生长发育及其信号转导等方面起着重要的分子生物学功能。文章采用Northern 杂交的方法, 对拟南芥3个WRKY基因进行表达谱分析。结果表明: AtWRKY25、AtWRKY26和AtWRKY33受多种非生物逆境因子(温度因子、高盐、渗透胁迫和激素脱落酸)的影响, 其中低温和高盐对AtWRKY25、AtWRKY26和AtWRKY33的诱导尤为明显, 表明这3个AtWRKY基因可能在响应环境信号方面起着一定的作用。作为序列相似性较高的AtWRKY25、AtWRKY26和AtWRKY33对一些胁迫因子的表达模式呈现一定的相似性; 但AtWRKY33受高温的抑制和低温的快速诱导, 与另外两个基因的表达模式不同, 推测它们对温度胁迫因子的反应存在差异。此外, 对启动子序列的生物信息学分析发现, 3个基因的启动子包含多个与非生物逆境反应相关的顺式作用元件。     

Yeast increases resistance in Arabidopsis against Pseudomonas syringae and Botrytis cinerea by salicylic acid-dependent as well as -independent mechanisms

Cell-wall and glucopeptide components of yeast have been reported to exhibit elicitor activity. The mode of action of defense activation by yeast is not known so far. In this study, we used the model plant Arabidopsis to investigate the activation of defense responses by yeast, the effect on resistance against different pathogens, and the mode of action. Treatment of Arabidopsis plants with an autoclaved yeast suspension induced the expression of systemic acquired resistance-related genes and accumulation of the phytoalexin camalexin. Symptom development and bacterial growth after infection with a virulent strain of the pathogen Pseudomonas syringae was reduced in yeast-pretreated plants. No protection was detectable in mutants affected in the salicylate pathway, while mutants in the jasmonate or camalexin pathway were protected by yeast, indicating that the salicylate pathway is necessary for the yeast-induced resistance against P. syringae. Yeast also reduced symptom development after challenge with Botrytis cinerea. This protection was detectable in all mutants tested, indicating that it is independent of the salicylate, jasmonate, and camalexin pathway.     

The Pseudomonas syringae type III effector HopAM1 enhances virulence on water-stressed plants

Pseudomonas syringae strains deliver diverse type III effector proteins into host cells, where they can act as virulence factors. Although the functions of the majority of type III effectors are unknown, several have been shown to interfere with plant basal defense mechanisms. Type III effectors also could contribute to bacterial virulence by enhancing nutrient uptake and pathogen adaptation to the environment of the host plant. We demonstrate that the type III effector HopAM1 (formerly known as AvrPpiB) enhances the virulence of a weak pathogen in plants that are grown under drought stress. This is the first report of a type III effector that aids pathogen adaptation to water availability in the host plant. Expression of HopAM1 makes transgenic Ws-0 Arabidopsis hypersensitive to abscisic acid (ABA) for stomatal closure and germination arrest. Conditional expression of HopAM1 in Arabidopsis also suppresses basal defenses. ABA responses overlap with defense responses and ABA has been shown to suppress defense against P. syringae pathogens. We propose that HopAM1 aids P. syringae virulence by manipulation of ABA responses that suppress defense responses. In addition, host ABA responses enhanced by type III delivery of HopAM1 protect developing bacterial colonies inside leaves from osmotic stress.     

Elicitation of the hypersensitive response by Pseudomonas syringae

The hypersensitive response (HR), a rapid local necrosis accompanied by the production of antimicrobial compounds, is a manifestation of the primary defense mechanisms through which plants prevent colonization by potentially pathogenic microorganisms. The pathogen factors that trigger HR-associated defense mechanisms have yet to be established. Application of transposon mutagenesis and molecular cloning techniques have provided new insights into the bacterial elicitation of the HR. This review focuses on the interaction of Pseudomonas syringae with non-host plants as a model system for the bacterial induction of the HR and discusses (a) the genetics of Pseudomonas syringae pathogenicity and HR elicitation, (b) the environmental conditions controlling expression of Pseudomonas syringae pathogenicity/HR determinants, (c) the nature of the HR elicitor and (d) early plant responses to the pathogen.     

Resistance to Pseudomonas syringae conferred by an Arabidopsis thaliana coronatine-insensitive (coi1) mutation occurs through two distinct mechanisms

A new allele of the coronatine-insensitive locus (COI1) was isolated in a screen for Arabidopsis thaliana mutants with enhanced resistance to the bacterial pathogen Pseudomonas syringae. This mutant, designated coi1-20, exhibits robust resistance to several P. syringae isolates but remains susceptible to the virulent pathogens Erisyphe and cauliflower mosaic virus. Resistance to P. syringae strain PstDC3000 in coi1-20 plants is correlated with hyperactivation of PR-1 expression and accumulation of elevated levels of salicylic acid (SA) following infection, suggesting that the SA-mediated defense response pathway is sensitized in this mutant. Restriction of growth of PstDC3000 in coi1-20 leaves is partially dependent on NPR1 and fully dependent on SA, indicating that SA-mediated defenses are required for restriction of PstDC3000 growth in coi1-20 plants. Surprisingly, despite high levels of PstDC3000 growth in coi1-20 plants carrying the salicylate hydroxylase (nahG) transgene, these plants do not exhibit disease symptoms. Thus resistance to P. syringae in coi1-20 plants is conferred by two different mechanisms: (i) restriction of pathogen growth via activation of the SA-dependent defense pathway; and (ii) an SA-independent inability to develop disease symptoms. These findings are consistent with the hypotheses that the P. syringae phytotoxin coronatine acts to promote virulence by inhibiting host defense responses and by promoting lesion formation.     

Differential volatile emissions and salicylic acid levels from tobacco plants in response to different strains of<Emphasis Type="Italic"> Pseudomonas syringae</Emphasis>

Pathogen-induced plant responses include changes in both volatile and non-volatile secondary metabolites. To characterize the role of bacterial pathogenesis in plant volatile emissions, tobacco plants, Nicotiana tabacum L. K326, were inoculated with virulent, avirulent, and mutant strains of Pseudomonas syringae. Volatile compounds released by pathogen-inoculated tobacco plants were collected, identified, and quantified. Tobacco plants infected with the avirulent strains P. syringae pv. maculicola ES4326 (Psm ES4326) or pv. tomato DC3000 (Pst DC3000), emitted quantitatively different, but qualitatively similar volatile blends of (E)-beta-ocimene, linalool, methyl salicylate (MeSA), indole, caryophyllene, beta-elemene, alpha-farnesene, and two unidentified sesquiterpenes. Plants treated with the hrcC mutant of Pst DC3000 (hrcC, deficient in the type-III secretion system) released low levels of many of the same volatile compounds as in Psm ES4326- or Pst DC3000-infected plants, with the exception of MeSA, which occurred only in trace amounts. Interaction of the virulent pathogen P. syringae pv. tabaci (Pstb), with tobacco plants resulted in a different volatile blend, consisting of MeSA and two unidentified sesquiterpenes. Overall, maximum volatile emissions occurred within 36 h post-inoculation in all the treatments except for the Pstb infection that produced peak volatile emissions about 60 h post-inoculation. (E)-beta-Ocimene was released in a diurnal pattern with the greatest emissions during the day and reduced emissions at night. Both avirulent strains, Psm ES4326 and Pst DC3000, induced accumulation of free salicylic acid (SA) within 6 h after inoculation and conjugated SA within 60 h and 36 h respectively. In contrast, SA inductions by the virulent strain Pstb occurred much later and conjugated SA increased slowly for a longer period of time, while the hrcC mutant strain did not trigger free and conjugated SA accumulations in amounts significantly different from control plants. Jasmonic acid, known to induce plant volatile emissions, was not produced in significantly higher levels in inoculated plants compared to the control plants in any treatments, indicating that induced volatile emissions from tobacco plants in response to P. syringae are not linked to changes in jasmonic acid.     

Extracellular pyridine nucleotides induce PR gene expression and disease resistance in Arabidopsis

Although it is well known that the pyridine nucleotides NAD and NADP function inside the cell to regulate intracellular signaling processes, recent evidence from animal studies suggests that NAD(P) also functions in the extracellular compartment (ECC). Extracellular NAD(P) [eNAD(P)] can either directly bind to plasma membrane receptors or be metabolized by ecto-enzymes to produce cyclic ADP-ribose and nicotinic acid adenine dinucleotide phosphate, and/or may ADP-ribosylate cell-surface receptors, resulting in activation of transmembrane signaling. In this study, we report that, in plants, exogenous NAD(P) induces the expression of pathogenesis-related ( PR ) genes and resistance to the bacterial pathogen Pseudomonas syringae pv. maculicola ES4326. Chelation of Ca²⁺ by EGTA significantly inhibits the induction of PR genes by exogenous NAD(P), suggesting that exogenous NAD(P) may induce PR genes through a pathway that involves Ca²⁺ signaling. We show that exogenous application of NAD(P) causes accumulation of the defense signal molecule salicylic acid (SA), and induces both SA/NPR1-dependent and -independent PR gene expression and disease resistance. Furthermore, we demonstrate that NAD(P) leaks into the plant ECC after mechanical wounding and pathogen infection, and that the amount of NAD(P) leaking into the ECC after P. syringae pv. tobacco DC3000/ avrRpt2 infection is sufficient for induction of both PR gene expression and disease resistance. We propose that NAD(P) leakage from cells losing membrane integrity upon environmental stress may function as an elicitor to activate plant defense responses. Our data provide evidence that eNAD(P) functions in plant signaling, and illustrate the potential importance of eNAD(P) in plant innate immunity.     

PAD4 functions upstream from salicylic acid to control defense responses in Arabidopsis.

The Arabidopsis PAD4 gene was previously shown to be required for synthesis of camalexin in response to infection by the virulent bacterial pathogen Pseudomonas syringae pv maculicola ES4326 but not in response to challenge by the non-host fungal pathogen Cochliobolus carbonum. In this study, we show that pad4 mutants exhibit defects in defense responses, including camalexin synthesis and pathogenesis-related PR-1 gene expression, when infected by P. s. maculicola ES4 326. No such defects were observed in response to infection by an isogenic avirulent strain carrying the avirulence gene avrRpt2. In P. s. maculicola ES4 326-infected pad4 plants, synthesis of salicylic acid (SA) was found to be reduced and delayed when compared with SA synthesis in wild-type plants. Moreover, treatment of pad4 plants with SA partially reversed the camalexin deficiency and PR-1 gene expression phenotypes of P. s. maculicola ES4 326-infected pad4 plants. These findings support the hypothesis that PAD4 acts upstream from SA accumulation in regulating defense response expression in plants infected with P. s. maculicola ES4 326. A working model of the role of PAD4 in governing expression of defense responses is presented.     

The Pseudomonas syringae type III effector AvrRpt2 functions downstream or independently of SA to promote virulence on Arabidopsis thaliana

AvrRpt2, a Pseudomonas syringae type III effector protein, functions from inside plant cells to promote the virulence of P. syringae pv. tomato strain DC3000 (PstDC3000) on Arabidopsis thaliana plants lacking a functional copy of the corresponding RPS2 resistance gene. In this study, we extended our understanding of AvrRpt2 virulence activity by exploring the hypothesis that AvrRpt2 promotes PstDC3000 virulence by suppressing plant defenses. When delivered by PstDC3000, AvrRpt2 suppresses pathogen-related (PR) gene expression during infection, suggesting that AvrRpt2 suppresses defenses mediated by salicylic acid (SA). However, AvrRpt2 promotes PstDC3000 growth on transgenic plants expressing the SA-degrading enzyme NahG, indicating that AvrRpt2 does not promote bacterial virulence by modulating SA levels during infection. AvrRpt2 general virulence activity does not depend on the RPM1 resistance gene, as mutations in RPM1 had no effect on AvrRpt2-induced phenotypes. Transgenic plants expressing AvrRpt2 displayed enhanced susceptibility to PstDC3000 strains defective in type III secretion, indicating that enhanced susceptibility of these plants is not because of suppression of defense responses elicited by other type III effectors. Additionally, avrRpt2 transgenic plants did not exhibit increased susceptibility to Peronospora parasitica and Erysiphe cichoracearum, suggesting that AvrRpt2 virulence activity is specific to P. syringae.