Salicylic acid inhibits jasmonic acid-induced resistance of Arabidopsis thaliana to Spodoptera exigua

The role of salicylic acid (SA) in plant responses to pathogens has been well documented, but its direct and indirect effects on plant responses to insects are not so well understood. We examined the effects of SA, alone and in combination with jasmonic acid (JA), on the performance of the generalist herbivore, Spodoptera exigua, in wild-type and mutant Arabidopsis thaliana genotypes that varied genetically in their ability to mount SA- and JA-mediated defence responses. In one experiment, growth of S. exigua larvae was highest on the Wassilewskija wild-type, intermediate on the Columbia wild-type and the JA-deficient fad mutant, and lowest on the nim1-1 and jar1-mutants, which are defective in the SA and JA pathways, respectively. Activity of guaiacol peroxidase, polyphenoloxidase, n-acetylglucosaminidase, and trypsin inhibitor varied by genotype but did not correlate with insect performance. SA treatment increased growth of S. exigua larvae by approximately 35% over all genotypes, but had no discernable effect on activities of the four defence proteins. In a second experiment, growth of S. exigua was highest across treatments on the cep1 mutant, a constitutive expressor of high SA levels and systemic acquired resistance, and lowest on the fad mutant, which is JA-deficient. JA treatment generally increased activity of all four defence proteins, increased total glucosinolate levels and reduced insect growth by approximately 25% over all genotypes. SA generally inhibited expression of JA-induced resistance to S. exigua when both hormones were applied simultaneously. Across genotypes and treatments, larval mass was negatively correlated with the activity of trypsin inhibitor and polyphenoloxidase and with total glucosinolate levels, and insect damage was negatively correlated with the activity of polyphenoloxidase. SA had little effect on the induction of defence protein activity by JA. However, SA attenuated the induction of glucosinolates by JA and therefore may explain better the interactive effects of SA and JA on insect performance. This study illustrates that direct and indirect cross-effects of SA on resistance to S. exigua can occur in A. thaliana. Effects of SA may be mediated through effects on plant defence chemistry or other aspects of the suitability of foliage for insect feeding and growth.     

Arabidopsis peroxidase AtPRX53 influences cell elongation and susceptibility to Heterodera schachtii

Cyst nematodes establish and maintain feeding sites (syncytia) in the roots of host plants by altering expression of host genes. Among these genes are members of the large gene family of class III peroxidases, which have reported functions in a variety of biological processes. In this study, we used Arabidopsis-Heterodera schachtii as a model system to functionally characterize peroxidase 53 (AtPRX53). Promoter assays showed that under non-infected conditions AtPRX53 is expressed mainly in the root, the hypocotyl and the base of the pistil. Under infected conditions, the AtPRX53 promoter showed upregulation at the nematode penetration sites and in their migration paths. Interestingly, strong GUS activity was observed in H. schachtii-induced syncytia during the early stage of infection and remained strong in the syncytia of third-stage juveniles. Also, AtPRX53 showed upregulation in response to wounding and jasmonic acid treatments. Manipulation of AtPRX53 expression through overexpression and knockout mutation affected both plant morphology and nematode susceptibility. While AtPRX53 overexpression lines exhibited short hypocotyls, aberrant flower development and reduced nematode susceptibility to H. schachtii, the atprx53 mutant showed long hypocotyls and a 3-carpel silique phenotype as well as a non significant increase of nematode susceptibility. Taken together these data, therefore, indicate diverse roles of AtPRX53 in the wound response, flower development and syncytium formation.     

The nematode Neoaplectana carpocapsae and the beet armyworm Spodoptera exigua: Infectivity of prepupae and pupae in soil and of adults during emergence from soil

The nematode, Neoaplectana carpocapsae, infected >90% of the prepupae of Spodoptera exigua in soil even at concentrations as low as five nematodes/cm² of soil surface. Pupae were less susceptible to nematode infection in soil than prepupae, with mortality ranging from 10 to 24% and 10 to 83% for pupae exposed 3–5 days and 6–8 days to the nematode, respectively. Longer exposure (6–8 days) of the pupae to the nematode resulted in higher mortality with a positive relationship with increasing concentrations. Adults of S. exigua were susceptible to nematode infections as they emerged from the soil. The higher nematode concentrations (25 and 50 nematodes/cm²) resulted in higher adult mortality. The majority of nematode-induced mortality occurred within 24 hr after emergence. The susceptibility of emerging S. exigua adults to N. carpocapsae offers a new dimension for insect control.     

The Elongator complex‐associated protein DRL1 plays a positive role in immune responses against necrotrophic fungal pathogens in Arabidopsis

DEFORMED ROOT AND LEAVES1 (DRL1) is an Arabidopsis homologue of the yeast TOXIN TARGET4 (TOT4)/KILLER TOXIN‐INSENSITIVE12 (KTI12) protein that is physically associated with the RNA polymerase II‐interacting protein complex named Elongator. Mutations in DRL1 and Elongator lead to similar morphological and molecular phenotypes, suggesting that DRL1 and Elongator may functionally overlap in Arabidopsis. We have shown previously that Elongator plays an important role in both salicylic acid (SA)‐ and jasmonic acid (JA)/ethylene (ET)‐mediated defence responses. Here, we tested whether DRL1 also plays a similar role as Elongator in plant immune responses. Our results show that, although DRL1 partially contributes to SA‐induced cytotoxicity, it does not play a significant role in SA‐mediated expression of PATHOGENESIS‐RELATED genes and resistance to the virulent bacterial pathogen Pseudomonas syringae pv. maculicola ES4326. In contrast, DRL1 is required for JA/ET‐ and necrotrophic fungal pathogen Botrytis cinerea‐induced defence gene expression and for resistance to B. cinerea and Alternaria brassicicola. Furthermore, unlike the TOT4/KTI12 gene which, when overexpressed in yeast, confers zymocin resistance, a phenotype of the tot4/kti12 mutant, overexpression of DRL1 does not change B. cinerea‐induced defence gene expression and resistance to this pathogen. Finally, DRL1 contains an N‐terminal P‐loop and a C‐terminal calmodulin (CaM)‐binding domain and is a CaM‐binding protein. We demonstrate that both the P‐loop and the CaM‐binding domain are essential for the function of DRL1 in B. cinerea‐induced expression of PDF1.2 and ORA59, and in resistance to B. cinerea, suggesting that the function of DRL1 in plant immunity may be regulated by ATP/GTP and CaM binding.     

The immune components ENHANCED DISEASE SUSCEPTIBILITY 1 and PHYTOALEXIN DEFICIENT 4 are required for cell death caused by overaccumulation of ceramides in Arabidopsis

Sphingolipids have key functions in plant membrane structure and signaling. Perturbations of plant sphingolipid metabolism often induce cell death and salicylic acid (SA) accumulation; SA accumulation, in turn, promotes sphingolipid metabolism and further cell death. However, the underlying molecular mechanisms remain unclear. Here, we show that the Arabidopsis thaliana lipase-like protein ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1) and its partner PHYTOALEXIN DEFICIENT 4 (PAD4) participate in sphingolipid metabolism and associated cell death. The accelerated cell death 5 (acd5) mutants accumulate ceramides due to a defect in ceramide kinase and show spontaneous cell death. Loss of function of EDS1, PAD4 or SALICYLIC ACID INDUCTION DEFICIENT 2 (SID2) in the acd5 background suppressed the acd5 cell death phenotype and prevented ceramide accumulation. Treatment with the SA analogue benzothiadiazole partially restored sphingolipid accumulation in the acd5 pad4 and acd5 eds1 double mutants, showing that the inhibitory effect of the pad4-1 and eds1-2 mutations on acd5-conferred sphingolipid accumulation partly depends on SA. Moreover, the pad4-1 and eds1-2 mutations substantially rescued the susceptibility of the acd5 mutant to Botrytis cinerea. Consistent with this, B. cinerea-induced ceramide accumulation requires PAD4 or EDS1. Finally, examination of plants overexpressing the ceramide synthase gene LAG1 HOMOLOGUE2 suggested that EDS1, PAD4 and SA are involved in long-chain ceramide metabolism and ceramide-associated cell death. Collectively, our observations reveal that EDS1 and PAD4 mediate ceramide (especially long-chain ceramide) metabolism and associated cell death, by SA-dependent and SA-independent pathways.     

Nitric oxide modulates ozone-induced cell death, hormone biosynthesis and gene expression in Arabidopsis thaliana

Nitric oxide (NO) is involved together with reactive oxygen species (ROS) in the activation of various stress responses in plants. We have used ozone (O₃) as a tool to elicit ROS-activated stress responses, and to activate cell death in plant leaves. Here, we have investigated the roles and interactions of ROS and NO in the induction and regulation of O₃-induced cell death. Treatment with O₃ induced a rapid accumulation of NO, which started from guard cells, spread to adjacent epidermal cells and eventually moved to mesophyll cells. During the later time points, NO production coincided with the formation of hypersensitive response (HR)-like lesions. The NO donor sodium nitroprusside (SNP) and O₃ individually induced a large set of defence-related genes; however, in a combined treatment SNP attenuated the O₃ induction of salicylic acid (SA) biosynthesis and other defence-related genes. Consistent with this, SNP treatment also decreased O₃-induced SA accumulation. The O₃-sensitive mutant rcd1 was found to be an NO overproducer; in contrast, Atnoa1/rif1 ( Arabidopsis nitric oxide associated 1/resistant to inhibition by FSM1 ), a mutant with decreased production of NO, was also O₃ sensitive. This, together with experiments combining O₃ and the NO donor SNP suggested that NO can modify signalling, hormone biosynthesis and gene expression in plants during O₃ exposure, and that a functional NO production is needed for a proper O₃ response. In summary, NO is an important signalling molecule in the response to O₃.     

Activity and Persistence of Steinernema carpocapsae and Spodoptera exigua Nuclear Polyhedrosis Virus against S. exigua Larvae on Soybean

Experiments were conducted to assess the effects of the entomopathogenic nematode Steinernema carpocapsae and Spodoptera exigua multinucleocapsid nuclear polyhedrosis virus (SeMNPV), alone and in combinations, on mortality of the beet armyworm, S. exigua, larvae on soybean. In 1991 tests, field-grown soybean plants were treated with S. carpocapsae at 0.3 and 0.6 nematodes/cm² of leaflet, SeMNPV at 20 and 40 polyhedral inclusion bodies (PIB)/cm², and all possible combinations. Treated leaflets were collected from plants and bioassayed with 5-day-old larvae. The combination of S. carpocapsae at 0.6 nematodes/cm² + SeMNPV at 40 PIB/cm² produced significantly higher larval mortality (61.7%) compared with either S. carpocapsae (24.8-35.1%) or SeMNPV (26.5-33.7%) alone. In 1992, similar tests were repeated using S. carpocapsae at 0.2 and 0.5 nematodes/cm², and SeMNPV at 14 and 35 PIB/cm². The combination of 0.5 nematodes/cm² + 35 PIB/cm² resulted in significantly higher larval mortality (64.0%) than either pathogen alone (41.5-49.0%). Steinernema carpocapsae and SeMNPV produced additive effects on beet arlnyworm mortality. Persistence of S. carpocapsae was 12-24 hours and SeMNPV was 96-120 hours on soybean.     

Local and systemic wound-induction of RNase and nuclease activities in Arabidopsis: RNS1 as a marker for a JA-independent systemic signaling pathway

Induction of defense-related genes is one way in which plants respond to mechanical injury. We investigated whether RNases are involved in the wound response in Arabidopsis thaliana. As in other plant systems, several activities are induced with various timings in damaged leaves, stems and seedlings in Arabidopsis, including at least three bifunctional nucleases, capable of degrading both RNA and DNA, as well as RNS1, a member of the ubiquitous RNase T(2) family of RNases. The strong induction of RNS1 is particularly interesting because it occurs both locally and systemically following wounding. The systemic induction of this RNase indicates that members of this family may be involved in defense mechanisms in addition to their previously hypothesized functions in nutrient recycling and remobilization. Additionally, the systemic induction appears to be controlled independently of jasmonic acid, and the local induction of RNS1 and the nuclease activities are independent of both JA and oligosaccharide elicitors. Consequently, a novel systemic pathway, likely involving a third signal, appears to exist in Arabidopsis.     

逆境下拟南芥野生型和突变体sex1游离态水杨酸含量及根形态差异

以拟南芥野生型(WT)和突变体sex1为材料,采用HPLC、显微观察等方法,研究了WT在10个不同生长期时游离态水杨酸(SA)含量的变化,比较了在丁香假单胞菌番茄致病变种(Pst.DC3000)、H2O2、甲基紫精(MV)和SA处理环境下WT和sex1中游离态SA含量及幼苗根形态的差异.结果表明:在花序继续开放期(6.30、6.50)以及长角果出现期(8.0),WT中游离态SA含量处在更低水平;2 mmol·L-1 SA处理能够极显著提高WT和突变体sex1中游离态SA的积累,且突变体sex1中游离态SA水平增加更明显,是其他处理游离态SA水平的10倍左右;在MV和H2O2处理下,WT和sex1主根生长受抑制的程度没有明显差异,而突变体sex1幼苗的根毛在低浓度MV环境中比WT更长但密度差异不明显,且WT和sex1根毛在低浓度H2O2环境中差异情况与对照组相似,但外源SA处理使WT和突变体sex1的主根生长均受到明显的抑制,两者的根毛密度随SA增加逐渐降低甚至消失,且突变体sex1比WT受到的抑制作用更明显.研究发现:拟南芥的开花和结果过程与SA依赖途径有一定关系;外源添加SA比Pst.DC3000、H2O2和MV处理更能直接促使拟南芥突变体sex1产生更多的游离态SA;突变体sex1根的生长发育比WT更易受到生长环境条件的影响,且sex1根形态的生长存在一定的SA浓度依赖模式.     

Jasmonic acid and salicylic acid play minor roles in stomatal regulation by CO2, abscisic acid,darkness, vapor pressure deficit and ozone

Jasmonic acid (JA) and salicylic acid (SA) regulate stomatal closure, preventing pathogen invasion into plants. However, to what extent abscisic acid (ABA), SA and JA interact, and what the roles of SA and JA are in stomatal responses to environmental cues, remains unclear. Here, by using intact plant gas-exchange measurements in JA and SA single and double mutants, we show that stomatal responsiveness to CO₂, light intensity, ABA, high vapor pressure deficit and ozone either did not or, for some stimuli only, very slightly depended upon JA and SA biosynthesis and signaling mutants, including dde2, sid2, coi1, jai1, myc2 and npr1 alleles. Although the stomata in the mutants studied clearly responded to ABA, CO₂, light and ozone, ABA-triggered stomatal closure in npr1-1 was slightly accelerated compared with the wild type. Stomatal reopening after ozone pulses was quicker in the coi1-16 mutant than in the wild type. In intact Arabidopsis plants, spraying with methyl-JA led to only a modest reduction in stomatal conductance 80 min after treatment, whereas ABA and CO₂ induced pronounced stomatal closure within minutes. We could not document a reduction of stomatal conductance after spraying with SA. Coronatine-induced stomatal opening was initiated slowly after 1.5–2.0 h, and reached a maximum by 3 h after spraying intact plants. Our results suggest that ABA, CO₂ and light are major regulators of rapid guard cell signaling, whereas JA and SA could play only minor roles in the whole-plant stomatal response to environmental cues in Arabidopsis and Solanum lycopersicum (tomato).     

绿木霉SM1蛋白诱导拟南芥防御反应以及激发活性氧途径研究

SM1蛋白是由绿木霉Trichoderma virens产生的一种富含半胱氨酸的小蛋白,能够作为激发子激发植物防御反应。研究了SM1蛋白对拟南芥Arabidopsis thaliana生长及诱导抗性的作用。结果表明高浓度（>10μg/mL）SM1蛋白液抑制拟南芥的生长,低浓度SM1蛋白液则不影响生长;SM1能诱导拟南芥对细菌性叶斑病Pseudomonas syringae pv. tomato DC3000的抗性,引起拟南芥叶片过氧化氢的积累。SM1蛋白处理后,拟南芥叶片中植物防御反应相关基因PDF1.2、LOX2和活性氧酶基因 SOD、POD等表达显著上升,说明SM1在激活植物的JA/ET和ROS途径中发挥着重要作用。研究为进一步研究SM1诱导植物抗性的机理提供了基础。     

Altered growth and improved resistance of Arabidopsis against Pseudomonas syringae by overexpression of the basic amino acid transporter AtCAT1

Amino acid transporters in plants are crucial for distributing amino acids between plant organs and cellular compartments. The H⁺‐coupled plasma membrane transporter CAT1 (cationic amino acid transporter 1) facilitates the high‐affinity uptake of basic amino acids. The uptake of lysine (Lys) via the roots was not altered in loss‐of‐function mutants, in accordance with the minor expression of CAT1 in roots, but plants ectopically overexpressing CAT1 incorporated Lys at higher rates. Exogenous Lys inhibited the primary root of Arabidopsis, whereas lateral roots were stimulated. These effects were augmented by the presence or absence of CAT1. Furthermore, the total biomass of soil‐grown plants ectopically overexpressing CAT1 was reduced and the time to flowering was accelerated. These effects were accompanied by only minor changes in the overall amino acid profile. Interestingly, CAT1 belongs to a specific small cluster of nitrogen‐containing metabolite transporter genes that are rapidly up‐regulated upon infection with Pseudomonas syringae and that may participate in the systemic response of plants to pathogen attack. The overexpression of CAT1 indeed enhanced the resistance to the hemibiotrophic bacterial pathogen P. syringae via a constitutively activated salicylic acid (SA) pathway, which is consistent with the developmental defects and the resistance phenotype.