bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of a Novel, Defense-Related Arabidopsis Mutant, cir1, Isolated By Luciferase Imaging
Authors: Murray Shane L. , Thomson Catherine , Chini Andrea , Read Nick D. , Loake Gary J.
Published: May 2007

In order to identify components of the defense signaling network engaged following attempted pathogen invasion, we generated a novel PR-1∷luciferase (LUC) transgenic line that was deployed in an imaging-based screen to uncover defense-related mutants. The recessive mutant designated cir1 exhibited constitutive expression of salicylic acid (SA), jasmonic acid (JA)/ethylene, and reactive oxygen intermediate-dependent genes. Moreover, this mutation conferred resistance against the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 and a virulent oomycete pathogen Peronospora parasitica Noco2. Epistasis analyses were undertaken between cir1 and mutants that disrupt the SA (npr1, nahG), JA (jar1), and ethylene (ET) (ein2) signaling pathways. While resistance against both P. syringae pv. tomato DC3000 and Peronospora parasitica Noco2 was partially reduced by npr1, resistance against both of these pathogens was lost in an nahG genetic background. Hence, cir1-mediated resistance is established via NPR1-dependent and -independent signaling pathways and SA accumulation is essential for the function of both pathways. While jar1 and ein2 reduced resistance against P. syringae pv. tomato DC3000, these mutations appeared not to impact cir1-mediated resistance against Peronospora parasitica Noco2. Thus, JA and ET sensitivity are required for cir1-mediated resistance against P. syringae pv. tomato DC3000 but not Peronospora parasitica Noco2. Therefore, the cir1 mutation may define a negative regulator of disease resistance that operates upstream of SA, JA, and ET accumulation.
Overexpression of Pto Induces a Salicylate-Independent Cell Death But Inhibits Necrotic Lesions Caused by Salicylate-Deficiency in Tomato Plants
Authors: Li Jianxiong , Shan Libo , Zhou Jian-Min , Tang Xiaoyan
Published: May 2007

Tomato plants overexpressing the disease resistance gene Pto (35S∷Pto) exhibit spontaneous cell death, accumulation of salicylic acid (SA), elevated expression of pathogenesis-related genes, and enhanced resistance to a broad range of pathogens. Because salicylate plays an important role in the cell death and defense activation in many lesion mimic mutants, we investigated the interaction of SA-mediated processes and the 35S∷Pto-mediated defense pathway by introducing the nahG transgene that encodes salicylate hydroxylase. Here, we show that SA is not required for the 35S∷Pto-activated microscopic cell death and plays a minor role in defense gene activation and general disease resistance in 35S∷Pto plants. In contrast, temperature greatly affects the spontaneous cell death and general resistance in 35S∷Pto plants, and high temperature inhibits the cell death. The NahG tomato plants develop spontaneous, unconstrained necrotic lesions on leaves. These lesions also are initiated by the inoculation of a virulent strain of Pseudomonas syringae pv. tomato. However, the NahG-dependent necrotic lesions are inhibited in the NahG/35S∷Pto plants. This inhibition is most pronounced under conditions favoring the 35S∷Pto-mediated spontaneous cell death development. These results indicate that the signaling pathways activated by Pto overexpression suppress the cellular damage that is caused by SA depletion. We also found that ethylene is dispensable for the 35S∷Pto-mediated general defense.
Characterization of NADH Dehydrogenases of Pseudomonas fluorescens WCS365 and Their Role in Competitive Root Colonization
Authors: Camacho Carvajal Margarita M. , Wijfjes André H. M. , Mulders Ine H. M. , Lugtenberg Ben J. J. , Bloemberg Guido V.
Published: May 2007

The excellent-root-colonizing Pseudomonas fluorescens WCS365 was selected previously as the parental strain for the isolation of mutants impaired in root colonization. Transposon mutagenesis of WCS365 and testing for root colonization resulted in the isolation of mutant strain PCL1201, which is approximately 100-fold impaired in competitive tomato root colonization. In this manuscript, we provide evidence that shows that the lack of NADH dehydrogenase I, an enzyme of the aerobic respiratory chain encoded by the nuo operon, is responsible for the impaired root-colonization ability of PCL1201. The complete sequence of the nuo operon (ranging from nuoA to nuoN) of P. fluorescens WCS365 was identified, including the promoter region and a transcriptional terminator consensus sequence downstream of nuoN. It was shown biochemically that PCL1201 is lacking NADH dehydrogenase I activity. In addition, the presence and activity of a second NADH dehydrogenase, encoded by the ndh gene, was identified to our knowledge for the first time in the genus Pseudomonas. Since it was assumed that low-oxygen conditions were present in the rhizosphere, we analyzed the activity of the nuo and the ndh promoters at different oxygen tensions. The results showed that both promoters are up-regulated by low concentrations of oxygen and that their levels of expression vary during growth. By using lacZ as a marker, it was shown that both the nuo operon and the ndh gene are expressed in the tomato rhizosphere. In contrast to the nuo mutant PCL1201, an ndh mutant of WCS365 appeared not to be impaired in competitive root tip colonization.
Mutation of rpiA in Enterobacter cloacae Decreases Seed and Root Colonization and Biocontrol of Damping-Off Caused by Pythium ultimum on Cucumber
Authors: Lohrke Scott M. , Dery Pierre D. , Li Wei , Reedy Ralph , Kobayashi Donald Y. , Roberts Daniel P.
Published: May 2007

Strains of Enterobacter cloacae show promise as biocontrol agents for Pythium ultimum-induced damping-off on cucumber and other crops. E. cloacae A145 is a mini-Tn5 Km transposon mutant of strain 501R3 that was significantly reduced in suppression of damping-off on cucumber caused by P. ultimum. Strain A145 was deficient in colonization of cucumber, sunflower, and wheat seeds and significantly reduced in colonization of corn and cowpea seeds relative to strain 501R3. Populations of strain A145 were also significantly lower than those of strain 501R3 at all sampling times in cucumber, wheat, and sunflower rhizosphere. Populations of strain A145 were not detectable in any rhizosphere after 42 days, while populations of strain 501R3 remained at substantial levels throughout all experiments. Molecular characterization of strain A145 indicated mini-Tn5 Km was inserted in a region of the E. cloacae genome with a high degree of DNA and amino acid sequence similarity to rpiA, which encodes ribose-5-phosphate isomerase. In Escherichia coli, RpiA catalyzes the interconversion of ribose-5-phosphate and ribulose-5-phosphate and is a key enzyme in the pentose phosphate pathway. Ribose-5-phosphate isomerase activity in cell lysates from strain A145 was approximately 3.5% of that from strain 501R3. In addition, strain A145 was a ribose auxotroph, as expected for an rpiA mutant. Introduction of a 1.0-kb DNA fragment containing only the rpiA homologue into strain A145 restored ribose phosphate isomerase activity, prototrophy, seedling colonization, and disease suppression to levels similar to those associated with strain 501R3. Experiments reported here indicate a key role for rpiA and possibly the pentose phosphate pathway in suppression of damping-off and colonization of subterranean portions of plants by E. cloacae.
Regulation of Erwinia carotovora hrpLEcc (sigma-LEcc), Which Encodes an Extracytoplasmic Function Subfamily of Sigma Factor Required for Expression of the HRP Regulon
Authors: Chatterjee Asita , Cui Yaya , Chatterjee Arun K.
Published: May 2007

In Erwinia carotovora subsp. carotovora (Ecc) strain 71 (Ecc71), HrpLEcc, an alternate sigma factor of the extracytoplasmic function subfamily, plays a central role in the expression of the hrp (hypersensitive reaction and pathogenicity) regulon. We document here that sigma-54 (RpoN) is required for full expression of hrpLEcc and that HrpS, in conjunction with sigma-54, activates hrpLEcc transcription. We also made the novel observation that integration host factor is required for the activation of the hrpLEcc promoter. Our findings reveal that the RsmA/rsmB RNA-mediated post-transcriptional system, known to control ex-tracellular enzyme and harpin production, affects hrpLEcc expression as well. For example, hrpLEcc RNA levels are barely detected in an RsmB- strain. Conversely, hrpLEcc mRNA levels are much higher in RsmA- bacteria than in the RsmA+ parent. This effect is due to RsmA-promoted decay of hrpLEcc RNA. Moreover, the following regulators known to control the production of either RsmA, rsmB RNA, or both also affect hrpLEcc expression: GacA (response regulator of a two-component system), KdgR (an IclI type repressor), HexA (a LysR type repressor), RsmC (a putative transcriptional adapter). Based upon the data now available for Ecc and extrapolating from the evidence in other systems, we propose a tentative model that depicts the Hrp regulatory system of Ecc and explains the basis for coregulation of extracellular enzyme production and expression of the Hrp regulon.
Candidate Defense Genes from Rice, Barley, and Maize and Their Association with Qualitative and Quantitative Resistance in Rice
Authors: Ramalingam J. , Vera Cruz C. M. , Kukreja K. , Chittoor J. M. , Wu J.-L. , Lee S. W. , Baraoidan M. , George M. L. , Cohen M. B. , Hulbert S. H. , Leach J. E. , Leung H.
Published: May 2007

Candidate genes involved in both recognition (resistance gene analogs [RGAs]) and general plant defense (putative defense response [DR]) were used as molecular markers to test for association with resistance in rice to blast, bacterial blight (BB), sheath blight, and brown plant-hopper (BPH). The 118 marker loci were either polymerase chain reaction-based RGA markers or restriction fragment length polymorphism (RFLP) markers that included RGAs or putative DR genes from rice, barley, and maize. The markers were placed on an existing RFLP map generated from a mapping population of 116 doubled haploid (DH) lines derived from a cross between an improved indica rice cultivar, IR64, and a traditional japonica cultivar, Azucena. Most of the RGAs and DR genes detected a single locus with variable copy number and mapped on different chromosomes. Clusters of RGAs were observed, most notably on chromosome 11 where many known blast and BB resistance genes and quantitative trait loci (QTL) for blast, BB, sheath blight, and BPH were located. Major resistance genes and QTL for blast and BB resistance located on different chromosomes were associated with several candidate genes. Six putative QTL for BB were located on chromosomes 2, 3, 5, 7, and 8 and nine QTL for BPH resistance were located to chromosomes 3, 4, 6, 11, and 12. The alleles of QTL for BPH resistance were mostly from IR64 and each explained between 11.3 and 20.6% of the phenotypic variance. The alleles for BB resistance were only from the Azucena parent and each explained at least 8.4% of the variation. Several candidate RGA and DR gene markers were associated with QTL from the pathogens and pest. Several RGAs were mapped to BB QTL. Dihydrofolate reductase thymidylate synthase co-localized with two BPH QTL associated with plant response to feeding and also to blast QTL. Blast QTL also were associated with aldose reductase, oxalate oxidase, JAMyb (a jasmonic acid-induced Myb transcription factor), and peroxidase markers. The frame map provides reference points to select candidate genes for cosegregation analysis using other mapping populations, isogenic lines, and mutants.
Agrosuppression: A Bioassay for the Hypersensitive Response Suited to High-Throughput Screening
Authors: Kamoun Sophien , Hamada Walid , Huitema Edgar
Published: May 2007

We describe a novel method, agrosuppression, that addresses the need for an assay of the hypersensitive response (HR) in intact plants that is rapid and adapted to high-throughput functional screening of plant and pathogen genes. The agrosuppression assay is based on inoculation of intact plants with a mixture of Agrobacterium tumefaciens strains carrying (i) a binary plasmid with one or more candidate HR-inducing genes and (ii) a tumor-inducing (oncogenic) T-DNA. In the absence of HR induction, tumor formation is initiated, resulting in a typical crown gall phenotype. However, upon induction of the HR, tumor formation by the oncogenic T-DNA is suppressed, resulting in a phenotype that can be readily scored. We tested and optimized agrosuppression in Nicotiana benthamiana using the inf1 elicitin gene from the oomycete pathogen Phytophthora infestans, which specifically induces the HR in Nicotiana spp., and the gene-for-gene pair Avr9/Cf-9 from the fungal pathogen Cladosporium fulvum and Lycopersicon pimpinellifolium (currant tomato), respectively. Agrosuppression protocols that can be rapidly performed using simple mechanical wounding of petioles of intact N. benthamiana plants were developed and appeared particularly adapted to intensive high-throughput screening. This assay promises to greatly facilitate the cloning of novel plant R genes and pathogen Avr genes and to accelerate functional analyses and structure-function studies of these genes.
Fungal Biology and Agriculture: Revisiting the Field
Authors: Yarden O. , Ebbole D. J. , Freeman S. , Rodriguez R. J. , Dickman M. B.
Published: May 2007

Plant pathology has made significant progress over the years, a process that involved overcoming a variety of conceptual and technological hurdles. Descriptive mycology and the advent of chemical plant-disease management have been followed by biochemical and physiological studies of fungi and their hosts. The later establishment of biochemical genetics along with the introduction of DNA-mediated transformation have set the stage for dissection of gene function and advances in our understanding of fungal cell biology and plant-fungus interactions. Currently, with the advent of high-throughput technologies, we have the capacity to acquire vast data sets that have direct relevance to the numerous subdisciplines within fungal biology and pathology. These data provide unique opportunities for basic research and for engineering solutions to important agricultural problems. However, we also are faced with the challenge of data organization and mining to analyze the relationships between fungal and plant genomes and to elucidate the physiological function of pertinent DNA sequences. We present our perspective of fungal biology and agriculture, including administrative and political challenges to plant protection research.
The Medicago truncatula Sucrose Synthase Gene MtSucS1 Is Activated Both in the Infected Region of Root Nodules and in the Cortex of Roots Colonized by Arbuscular Mycorrhizal Fungi
Authors: Hohnjec Natalija , Perlick Andreas M. , Pühler Alfred , Küster Helge
Published: May 2007

The MtSucS1 gene encodes a sucrose synthase (EC 2.4.1.13) in the model legume Medicago truncatula. To determine the expression pattern of this gene in different organs and in particular during root endosymbioses, we transformed M. truncatula with specific regions of MtSucS1 fused to the gusAint reporter gene. These fusions directed an induction to the vasculature of leaves, stems, and roots as well as to flowers, developing seeds, young pods, and germinating seedlings. In root nodules, strong promoter activity occurred in the infected cells of the nitrogen-fixing zone but was additionally observed in the meristematic region, the prefixing zone, and the inner cortex, including the vasculature. Concerning endomycorrhizal roots, the MtSucS1 promoter mediated strongest expression in cortical cells harboring arbuscules. Specifically in highly colonized root sections, GUS-staining was furthermore detected in the surrounding cortical cells, irrespective of a direct contact with fungal structures. In accordance with the presence of an orthologous PsSus1 gene, we observed a comparable regulation of MtSucS1 expression in the grain legume Pisum sativum in response to microbial symbionts. Unlike other members of the MtSucS gene family, the presence of rhizobial or Glomus microsymbionts significantly altered and enhanced MtSucS1 gene expression, leading us to propose that MtSucS1 is involved in generating sink-strength, not only in root nodules but also in mycorrhizal roots.
Molecular Analysis of the Pathway for the Synthesis of Thiol Tripeptides in the Model Legume Lotus japonicus
Authors: Matamoros Manuel A. , Clemente Maria R. , Sato Shusei , Asamizu Erika , Tabata Satoshi , Ramos Javier , Moran Jose F. , Stiller Jiri , Gresshoff Peter M. , Becana Manuel
Published: May 2007

The thiol tripeptides, glutathione (GSH) and homoglu-tathione (hGSH), perform multiple roles in legumes, including protection against toxicity of free radicals and heavy metals. The three genes involved in the synthesis of GSH and hGSH in the model legume, Lotus japonicus, have been fully characterized and appear to be present as single copies in the genome. The γ-glutamylcysteine synthetase (γecs) gene was mapped on the long arm of chromosome 4 (70.0 centimorgans [cM]) and consists of 15 exons, whereas the glutathione synthetase (gshs) and homoglutathione synthetase (hgshs) genes were mapped on the long arm of chromosome 1 (81.3 cM) and found to be arranged in tandem with a separation of approximately 8 kb. Both genes consist of 12 exons of exactly the same size (except exon 1, which is similar). Two types of transcripts were detected for the gshs gene, which putatively encode proteins localized in the plastids and cytosol. Promoter regions contain cis-acting regulatory elements that may be involved in the plant's response to light, hormones, and stress. Determination of transcript levels, enzyme activities, and thiol contents in nodules, roots, and leaves revealed that γecs and hgshs are expressed in all three plant organs, whereas gshs is significantly functional only in nodules. This strongly suggests an important role of GSH in the rhizobia-legume symbiosis.
The Cre1 and Cre3 Nematode Resistance Genes Are Located at Homeologous Loci in the Wheat Genome
Authors: de Majnik John , Ogbonnaya Francis C. , Moullet Odile , Lagudah Evans S.
Published: May 2007

Differential responses in host-nematode pathotype interactions occur in wheat lines carrying different cereal cyst nematode resistance (Cre) genes. Cre1, located on chromosome 2B, confers resistance to most European nematodes and the sole Australian pathotype, while Cre3, present on chromosome 2D, is highly resistant to the Australian patho-type and susceptible to a number of European pathotypes. Genes encoding nucleotide binding site-leucine rich repeat (NBS-LRR) proteins that cosegregate with the Cre3 locus cross hybridize to homologues whose restriction fragment length polymorphism (RFLP) patterns distinguish near-isogenic Cre1 nematode-resistant wheat lines. Genetic mapping showed that the NBS-LRR gene members that distinguished the Cre1 near-isogenic lines were located on chromosome 2BL at a locus, designated Xcsl107, that cosegregates with the Cre1 locus. A haplotype of NBS-LRR genes from the Xcsl107 locus provides a diagnostic marker for the presence of Cre1 nematode resistance in a wide collection of wheat lines and segregating families. Genetic analysis of NBS-LRR haplo-types that cosegregate with Cre1 and Cre3 resistance, together with flanking cDNA markers and other markers from homoeologous group 2 chromosomes, revealed a conserved gene order that suggests Cre1 and Cre3 are homeoloci.
Depletion of the Photosystem II Core Complex in Mature Tobacco Leaves Infected by the Flavum Strain of Tobacco mosaic virus
Authors: Lehto Kirsi , Tikkanen Mikko , Hiriart Jean-Baptiste , Paakkarinen Virpi , Aro Eva-Mari
Published: May 2007

The flavum strain of Tobacco mosaic virus (TMV) differs from the wild-type (wt) virus by causing strong yellow and green mosaic in the systemically infected developing leaves, yellowing in the fully expanded leaves, and distinct malformations of chloroplasts in both types of infected tissues. Analysis of the thylakoid proteins of flavum strain-infected tobacco leaves indicated that the chlorosis in mature leaves was accompanied by depletion of the entire photosystem II (PSII) core complexes and the 33-kDa protein of the oxygen evolving complex. The only change observed in the thyla-koid proteins of the corresponding wt TMV-infected leaves was a slight reduction of the α and β subunits of the ATP synthase complex. The coat proteins of different yellowing strains of TMV are known to effectively accumulate inside chloroplasts, but in this work, the viral movement protein also was detected in association with the thylakoid membranes of flavum strain-infected leaves. The mRNAs of different enzymes involved in the chlorophyll biosynthesis pathway were not reduced in the mature chlorotic leaves. These results suggest that the chlorosis was not caused by reduction of pigment biosynthesis, but rather, by reduction of specific proteins of the PSII core complexes and by consequent break-down of the pigments.
Transient Expression of Homologous Hairpin RNA Causes Interference with Plant Virus Infection and Is Overcome by a Virus Encoded Suppressor of Gene Silencing
Authors: Tenllado F. , Barajas D. , Vargas M. , Atencio F. A. , González-Jara P. , Díaz-Ruíz J. R.
Published: May 2007

Specific post-transcriptional gene silencing (PTGS) of target genes can be induced in a variety of organisms by providing homologous double-stranded RNA (dsRNA) molecules. In plants, PTGS is part of a defense mechanism against virus infection. We have previously shown and patented that direct delivery to nontransgenic plants of dsRNA derived from viral sequences specifically interfere with virus infection. Here, we show that transient expression of constructs encoding hairpin RNA homologous to a rapidly replicating plant tobamovirus also interferes with virus multiplication in a sequence-dependent manner. A three-day lag period between delivery of hairpin RNA and virus into the same tissues completely block virus infectivity. Several hallmarks characteristic of PTGS were associated with viral interference mediated by hairpin RNA: high level of sequence identity between the hairpin RNA and the target RNA, presence of siRNAs in extracts derived from leaves infiltrated with hairpin RNA, and helper component-proteinase (HC-Pro) of potyviruses, a suppressor of PTGS, overcame interference. No evidence for a mobile silencing suppression signal induced by transient expression of HC-Pro was observed. The approach described here has the potential to be used as a versatile tool for studying the onset of PTGS in cases involving virus infection, in opposition to dsRNA-transgenic plants, which allow primarily for the study of PTGS maintenance.
Expression of Peroxidase-like Genes, H2O2Production, and Peroxidase Activity During the Hypersensitive Response toXanthomonas campestrispv.vesicatoriainCapsicum annuum
Authors: Do Hyun Mee , Hong Jeum Kyu , Jung Ho Won , Kim Sang Hee , Ham Jong Hyun , Hwang Byung Kook
Published: May 2007

Pepper ascorbate peroxidase-like (CAPOA1), thioredoxin peroxidase-like (CAPOT1), and peroxidase-like (CAPO1) clones were isolated from pepper leaves inoculated with avirulent strain Bv5-4a of Xanthomonas campestris pv. vesicatoria. CAPOA1, CAPOT1, and CAPO1 mRNA disappeared 18 to 30 h after the bacterial infection when the hypersensitive response (HR) was visible. In contrast, peroxidase activity reached a peak at 18 h after infection and then declined at 24 and 30 h when H2O2accumulation level was maximal. These results suggest that the striking accumulation of H2O2and strong decrease in peroxidase activity during the programmed cell death may be due to the strong suppression of CAPOA1, CAPOT1, and CAPO1 gene expression. Infection by Phytophthora capsici or Colletotricum gloeosporioides also induced the expression of the three putative peroxidase genes in pepper tissues. CAPOA1 mRNAs were in situ localized in phloem areas of vascular bundles in pepper tissues infected by Colletotricum. coccodes, P. capsici, or C. gloeosporioides. Exogenous treatment with H2O2strongly induced the CAPOA1 and CAPOT1 transcription 1 h after treatment, while the CAPO1 transcripts accumulated 12 h after H2O2treatment. We suggest that pepper ascorbate peroxidase and thioredoxin peroxidase genes may function as regulators of H2O2level and total peroxidase activity in the oxidative burst during the HR to incompatible pathogen interaction in pepper plant.
The Regulatory Cascade That Activates the Hrp Regulon in Erwinia herbicola pv. gypsophilae
Authors: Nizan-Koren R. , Manulis S. , Mor H. , Iraki N. M. , Barash I.
Published: May 2007

The pathogenicity of Erwinia herbicola pv. gypsophilae (Ehg) is dependent on a plasmid (pPATHEhg) that harbors the hrp gene cluster and additional virulence genes. The hrp regulatory cascade of Ehg comprises an hrpXY operon encoding a two-component system; hrpS encoding a transcriptional factor of the NtrC family and hrpL encoding an alternative sigma factor. Results obtained suggest the following signal transduction model for activating the Hrp regulon: phosphorylated HrpY activates hrpS, HrpS activates hrpL, and HrpL activates genes containing “hrp box” promoter. This model was supported by studies on the effects of mutations in the regulatory genes on pathogenicity and complementation analysis. Nonpolar mutations in hrpX did not affect virulence or transcription of downstream genes. Site-directed mutagenesis of the conserved aspartate 57 in HrpY suggested that its phosphorylation is crucial for activating the hrp regulatory cascade. Studies on the effects of mutations in the hrp regulatory genes on transcriptional activity of downstream genes or of their isolated promoters in planta showed dependency of hrpS expression on active HrpY, of hrpL expression on active HrpS, and of hrpN or hrpJ expression on active HrpL. These results were also partially supported by overexpression of regulatory genes under in vitro conditions. The hrpXY is constitutively expressed with high basal levels under repressive conditions, in contrast to hrpS and hrpL, which exhibit low basal expression levels and are environmentally regulated.
The 2b Protein of Cucumoviruses Has a Role in Promoting the Cell-to-cell Movement of Pseudorecombinant Viruses
Authors: Shi Bu-Jun , Miller Jane , Symons Robert H. , Palukaitis Peter
Published: May 2007

Pseudorecombinant viruses (i.e., those containing a reas-sorted genome of closely related multipartite viruses) are often not as competitive as the parental viruses. The role of the 2b gene in hypervirulence and maintenance of a progressive infection was assessed in a pseudorecombinant virus formed between RNAs 1 plus 2 of Cucumber mosaic virus (CMV) and RNA 3 of Tomato aspermy virus (TAV). The presence of RNA 3 of TAV was found to affect the level of RNA accumulation but not the level of virulence. By contrast, the 2b genes of both TAV and a hypervirulent strain of CMV (WAII-CMV) were found to affect the virulence of the pseudorecombinant viruses but not the levels of viral RNA accumulation. The 2b gene rather than the overlapping open reading frame encoding the C-terminal 41 amino acids of 2a protein of the corresponding virus was found to be essential for promoting infection of the pseudorecombinant viruses in planta. However, the 2b gene was not essential for replication of pseudorecombinant viruses containing CMV RNAs 1 plus 2 and TAV RNA 3. These results indicate that the 2b protein is involved in promoting the cell-to-cell movement of the pseudorecombinant viruses. These data also suggest the existence of specific interaction between the TAV 2b protein and either RNA 3 or its encoded proteins, which may be critical for promoting or maintaining infection or both.
The Arabidopsis Genes RPW8.1 and RPW8.2 Confer Induced Resistance to Powdery Mildew Diseases in Tobacco
Authors: Xiao Shunyuan , Charoenwattana Piyavadee , Holcombe Lucy , Turner John G.
Published: May 2007

Plant disease resistance (R) gene products recognize pathogen avirulence (Avr) gene products and induce defense responses. It is not known if an R gene can function in different plant families, however. The Arabidopsis thaliana R genes RPW8.1 and RPW8.2 confer resistance to the powdery mildew pathogens Erysiphe orontii, E. cichoracearum, and Oidium lycopersici, which also infect plants from other families. We produced transgenic Nicotiana tabacum, N. benthamiana, and Lycopersicon esculentum plants containing RPW8.1 and RPW8.2. Transgenic N. tabacum plants had increased resistance to E. orontii and O. lycopersici, transgenic N. benthamiana plants had increased resistance to E. cichoracearum, but transgenic L. esculentum plants remained susceptible to these pathogens. The defense responses induced in transgenic N. tabacum and N. benthamiana were similar to those mediated by RPW8.1 and RPW8.2 in Arabidopsis. Apparently, RPW8.1 and RPW8.2 could be used to control powdery mildew diseases of plants from other families.
The Role of Nod Factor Substituents in Actin Cytoskeleton Rearrangements in Phaseolus vulgaris
Authors: Cárdenas Luis , Thomas-Oates Jane E. , Nava Noreide , López-Lara Isabel M. , Hepler Peter K. , Quinto Carmen
Published: May 2007

In order to define the symbiotic role of some of the chemical substituents in the Rhizobium etli Nod factors (NFs), we purified Nod metabolites secreted by the SM25 strain, which carries most of the nodulation genes, and SM17 with an insertion in nodS. These NFs were analyzed for their capabilities to induce root hair curling and cytoskeletal rearrangements. The NFs secreted by strain SM17 lack the carbamoyl and methyl substituents on the nonreducing terminal residue and an acetyl moiety on the fucosyl residue on the reducing-terminal residue as determined by mass spectrometry. We have reported previously that the root hair cell actin cytoskeleton from bean responds with a rapid fragmentation of the actin bundles within 5 min of NF exposure, and also is accompanied by increases in the apical influxes and intracellular calcium levels. In this article, we report that methyl-bearing NFs are more active in inducing root hair curling and actin cytoskeleton rearrangements than nonmethylated NFs. However, the carbamoyl residue on the nonreducing terminal residue and the acetyl group at the fucosyl residue on the reducing terminal residue do not seem to have any effect on root hair curling induction or in actin cytoskeleton rearrangement.
Disruption of Botrytis cinerea Pectin Methylesterase Gene Bcpme1 Reduces Virulence on Several Host Plants
Authors: Valette-Collet Odile , Cimerman Agnès , Reignault Philippe , Levis Caroline , Boccara Martine
Published: May 2007

The pectinolytic enzyme pectin methylesterase (PME) hydrolyses pectin in methanol and polygalacturonic acid. In the expressed sequence tag library of Botrytis cinerea T4, we identified a 1,041 bp Bcpme1 cDNA potentially encoding a 346-amino acid protein of 37 kDa showing 46.8% identity with Aspergillus sp. PMEs. Bcpme1 is a single copy gene and is similarly expressed in glucose and pectin containing media. To evaluate the role of Bcpme1 in Botrytis cinerea virulence, a mutant in Bcpme1 was generated by gene disruption. The Bcpme1 mutant showed similar growth on rich medium but reduced growth on pectin medium. Two isozymes of pI 7.4 and 7.1 were detected in pectin liquid-culture supernatants of wild-type strain Bd90 analyzed by isoelectric focusing-polyacrylamide gel electrophoresis, while those of Bcpme1 mutant possessed only the pI 7.1 isozyme. BCPME1, the pI 7.4 isozyme, is the major PME activity, as PME activity is 75% reduced in Bcpme1 mutant. Moreover, the Bcpme1 mutant was less virulent on apple fruits, grapevine, and Arabidopsis thaliana leaves. Those phenotypes were complemented by reintroducing a Bcpme1 copy in the Bcpme1 mutant. These results showed that B. cinerea possessed more than one PME-encoding gene and that BCPME1 is an important determinant of B. cinerea virulence.
An Investigation into the Involvement of Defense Signaling Pathways in Components of the Nonhost Resistance of Arabidopsis thaliana to Rust Fungi Also Reveals a Model System for Studying Rust Fungal Compatibility
Authors: Mellersh Denny G. , Heath Michèle C.
Published: May 2007

Seventeen accessions of Arabidopsis thaliana inoculated with the cowpea rust fungus Uromyces vignae exhibited a variety of expressions of nonhost resistance, although infection hypha growth typically ceased before the formation of the first haustorium, except in Ws-0. Compared with wild-type plants, there was no increased fungal growth in ndr1 or eds1 mutants defective in two of the signal cascades regulated by the major class of Arabidopsis host resistance genes. However, in the Col-0 background, infection hyphae of U. vignae and two other rust fungi were longer in sid2 mutants defective in an enzyme that synthesizes salicylic acid (SA), in npr1 mutants deficient in a regulator of the expression of SA-dependent pathogenesis related (PR) genes, and in NahG plants containing a bacterial salicylate hydroxylase. Infection hyphae of U. vignae and U. appendiculatus but not of Puccinia helianthi were also longer in jar1 mutants, which are defective in the jasmonic acid defense signaling pathway. Nevertheless, haustorium formation increased only for the Uromyces spp. and only in sid2 mutants or NahG plants. Rather than the hypersensitive cell death that usually accompanies haustorium formation in nonhost plants, Arabidopsis typically encased haustoria in calloselike material. Growing fungal colonies of both Uromyces spp., indicative of a successful biotrophic relationship between plant and fungus, formed in NahG plants, but only U. vignae formed growing colonies in the sid2 mutants and cycloheximide-treated wild-type plants. Growing colonies did not develop in NahG tobacco or tomato plants. These data suggest that nonhost resistance of Arabidopsis to rust fungi primarily involves the restriction of infection hypha growth as a result of defense gene expression. However, there is a subsequent involvement of SA but not SA-dependent PR genes in preventing the Uromyces spp. from forming the first haustorium and establishing a sufficient biotrophic relationship to support further fungal growth. The U. vignae-Arabidopsis combination could allow the application of the powerful genetic capabilities of this model plant to the study of compatibility as well as nonhost resistance to rust fungi.
Flagellin from an Incompatible Strain of Acidovorax avenae Mediates H2O2 Generation Accompanying Hypersensitive Cell Death and Expression of PAL, Cht-1, and PBZ1, but Not of LOX in Rice
Authors: Tanaka Noriko , Che Fang-Sik , Watanabe Naohide , Fujiwara Satsuki , Takayama Seiji , Isogai Akira
Published: May 2007

Acidovorax avenae causes a brown stripe disease in monocot plants. We recently reported that a rice-incompatible strain of A. avenae caused hypersensitive cell death in rice and that the flagellin of the incompatible strain was involved in this response. The incompatible strain induced the rapid generation of H2O2 accompanying hypersensitive cell death and the expression of defense genes such as PAL, Cht-1, PBZ1, and LOX, whereas the compatible strain did not. The purified incompatible flagellin also induced the expression of PAL, Cht-1, and PBZ1, but LOX expression was not induced by the incompatible flagellin. PAL and LOX enzymatic activities were increased by inoculation with the incompatible strain, whereas only PAL activity was increased by the incompatible flagellin. Interestingly, the flagellin-deficient incompatible strain lost the ability to generate H2O2 and induce hypersensitive cell death, but PAL, Cht-1, and PBZ1 expression still were induced by inoculation with the deficient strain, suggesting that induction of these genes is regulated not only by flagellin but also by some other signal. Thus, the incompatible flagellin of A. avenae is a specific elicitor in rice, but it is not the only factor capable of inducing the rice defense system.
Molecular Evidence that the Extracellular Cutinase Pbc1 Is Required for Pathogenicity of Pyrenopeziza brassicae on Oilseed Rape
Authors: Li Donghui , Ashby Alison M. , Johnstone Keith
Published: May 2007

Recent evidence has suggested that cutinase is required for cuticular penetration and, therefore, is essential for pathogenicity of Pyrenopeziza brassicae, the causal organism of light leaf spot disease of oilseed rape and other brassicas. In order to acquire molecular evidence for the role of cutinase in pathogenesis, the single-copy P. brassicae cutinase gene Pbc1 was disrupted by a transformation-mediated approach. Southern hybridization analysis revealed that in one mutant, NH10-1224, the disruption was due to a tandem insertion of two copies of the disruption vector into the 5′ coding region of Pbc1. In contrast to the wild type, no expression of Pbc1 was detected during in planta growth or in cutin-induced mycelium of NH10-1224 and no cutinase activity was detected in culture supernatants from NH10-1224 using pnitrophenyl butyrate as substrate. Scanning electron microscopy of Brassica napus cotyledons infected with wild-type P. brassicae confirmed that entry into the host is by direct penetration of the cuticle. In contrast, the cutinase-deficient mutant NH10-1224 failed to penetrate the cuticular layer and was unable to develop disease symptoms. This evidence is consistent with the hypothesis that Pbc1 is required for P. brassicae to penetrate the plant cuticle. Demonstration that complementation of NH10-1224 with the Pbc1 wild-type gene restores both cutinase activity and pathogenicity will be required to definitively establish that cutinase is required for successful pathogenesis of brassicas by P. brassicae.
Endophytic Fungal Mutualists: Seed-Borne Stagonospora Spp. Enhance Reed Biomass Production in Axenic Microcosms
Authors: Ernst Michael , Mendgen Kurt W. , Wirsel Stefan G. R.
Published: May 2007

Fungal endophytes mainly belong to the phylum Ascomycota and colonize plants without producing symptoms. We report on the isolation of seed-borne fungal endophytes from Phragmites australis (common reed) that were ascribed to the genus Stagonospora. Nested polymerase chain reaction (PCR) assays revealed that a Stagonospora sp. regularly colonized reed as shown for a period of three years. In spring, it was only detected in roots, whereas in autumn, it could frequently be found in all organs, including seeds. Microcosm experiments revealed that seeds harbored viable propagules of the fungus that colonized the developing germling, indicating vertical transmission. Endophytic growth was confirmed by immunofluorescence microscopy, reisolation of the fungus after surface sterilization, and PCR. Aseptic microcosms were established for studying fungal contributions towards host vitality. Several Stagonospora isolates enhanced reed biomass. Seed-borne endophytic Stagonospora spp. thus can provide improved vigor to common reed, which could be most important when seed-derived germlings establish new reed stands.
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
Authors: Nandi Ashis , Kachroo Pradeep , Fukushige Hirotada , Hildebrand David F. , Klessig Daniel F. , Shah Jyoti
Published: May 2007

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 syringae 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.
cg12 Expression Is Specifically Linked to Infection of Root Hairs and Cortical Cells during Casuarina glauca and Allocasuarina verticillata Actinorhizal Nodule Development
Authors: Svistoonoff Sergio , Laplaze Laurent , Auguy Florence , Runions John , Duponnois Robin , Haseloff Jim , Franche Claudine , Bogusz Didier
Published: May 2007

cg12 is an early actinorhizal nodulin gene from Casuarina glauca encoding a subtilisin-like serine protease. Using transgenic Casuarinaceae plants carrying cg12-gus and cg12-gfp fusions, we have studied the expression pattern conferred by the cg12 promoter region after inoculation with Frankia. cg12 was found to be expressed in root hairs and in root and nodule cortical cells containing Frankia infection threads. cg12 expression was also monitored after inoculation with ineffective Frankia strains, during my-corrhizae formation, and after diverse hormonal treatments. None of these treatments was able to induce its expression, therefore suggesting that cg12 expression is linked to plant cell infection by Frankia strains. Possible roles of cg12 in actinorhizal symbiosis are discussed.
The Root-Knot Nematode Resistance Gene Mi-1.2 of Tomato Is Responsible for Resistance Against the Whitefly Bemisia tabaci
Authors: Nombela Gloria , Williamson Valerie M. , Muñiz Mariano
Published: May 2007

The tomato gene Mi-1.2 confers resistance against root-knot nematodes and some isolates of potato aphid. Resistance to the whitefly Bemisia tabaci previously has been observed in Mi-bearing commercial tomato cultivars, suggesting that Mi, or a closely linked gene, is responsible for the resistance. The response of two biotypes of B. tabaci to tomato carrying the cloned Mi was compared with that of the isogenic untransformed tomato line Moneymaker. Our results indicate that Mi-1.2 is responsible for the resistance in tomato plants to both B- and Q- biotypes. Mi-1.2 is unique among characterized resistance genes in its activity against three very different organisms (root-knot nematodes, aphids, and whiteflies). These pests are among the most important on tomato crops worldwide, making Mi a valuable resource in integrated pest management programs.
Aphid-Induced Defense Responses in Mi-1-Mediated Compatible and Incompatible Tomato Interactions
Authors: de Ilarduya Oscar Martinez , Xie QiGuang , Kaloshian Isgouhi
Published: May 2007

The tomato Mi-1 gene confers resistance to three species of root-knot nematode and potato aphid. We studied changes in expression of jasmonic acid (JA)- and salicylic acid (SA)- dependent defense genes in response to potato and green peach aphids. We determined changes in three PR proteins, lipoxygenase and proteinase inhibitors I and II transcripts, locally and systemically in both compatible and incompatible interactions in tomato. Transcripts for PR-1 were detected earlier and accumulated to higher levels in the incompatible than in the compatible potato aphid/tomato interactions. The transcript profiles of the other genes were similar in compatible compared with incompatible interactions. Pin1 and Pin2 RNAs were detected early and transiently in both compatible and incompatible interactions. In tomato plants containing Mi-1, systemic expression of PR-1 and GluB was detected in both compatible and incompatible interactions at 48 h after infestations with either aphid. These results suggest that aphid feeding involves both SA and JA/ethylene plant defense signaling pathways and that Mi-1-mediated resistance might involve a SA-dependent signaling pathway. Potato aphid feeding generated reactive oxygen species in both compatible and incompatible interactions. However, a hypersensitive response was absent in the Mi-1-mediated resistance response to potato aphids. Reciprocal grafting experiments revealed that resistance is cell autonomous, and local expression of Mi-1 is required for Mi-1-mediated resistance against the potato aphid.
Characterization of Nops, Nodulation Outer Proteins, Secreted Via the Type III Secretion System of NGR234
Authors: Marie Corinne , Deakin William J. , Viprey Virginie , Kopciñska Joanna , Golinowski Wladyslaw , Krishnan Hari B. , Perret Xavier , Broughton William J.
Published: May 2007

The nitrogen-fixing symbiotic bacterium Rhizobium species NGR234 secretes, via a type III secretion system (TTSS), proteins called Nops (nodulation outer proteins). Abolition of TTSS-dependent protein secretion has either no effect or leads to a change in the number of nodules on selected plants. More dramatically, Nops impair nodule development on Crotalaria juncea roots, resulting in the formation of nonfixing pseudonodules. A double mutation of nopX and nopL, which code for two previously identified secreted proteins, leads to a phenotype on Pachyrhizus tuberosus differing from that of a mutant in which the TTSS is not functional. Use of antibodies and a modification of the purification protocol revealed that NGR234 secretes additional proteins in a TTSS-dependent manner. One of them was identified as NopA, a small 7-kDa protein. Single mutations in nopX and nopL were also generated to assess the involvement of each Nop in protein secretion and nodule formation. Mutation of nopX had little effect on NopL and NopA secretion but greatly affected the interaction of NGR234 with many plant hosts tested. NopL was not necessary for the secretion of any Nops but was required for efficient nodulation of some plant species. NopL may thus act as an effector protein whose recognition is dependent upon the hosts' genetic background.